Selective intestinal decontamination in the prevention of bacterial infection in patients with acute liver failure.

To investigate whether selective intestinal decontamination from oral administration of poorly absorbable antibiotics is effective in preventing bacterial infection in patients with acute liver failure, the incidence of nosocomial infection in 34 patients consecutively admitted to hospital between 1985-1990 and treated with either neomycin + colistin + nystatin or norfloxacin + nystatin (group I) was compared to the incidence of infection in 57 patients who did not receive oral, poorly absorbable antibiotics and who were consecutively admitted between 1972-1984 (group II). Patients from groups I and II had similar clinical and laboratory data at hospital admission. Twelve patients from group I and 33 from group II developed bacterial infection during the study period. The probability of infection was significantly different (p = 0.0083) in the two groups: 19% vs. 39% at the 3rd day of admission, 33% vs. 74% at the 7th day, and 48% vs. 78% at the 14th day, respectively. This difference was due to a different rate of infection from enterobacteria. Enterobacteria caused one infectious episode in group I and 24 in group II (p less than 0.001). The incidence of infections caused by other organisms, however, was similar in both groups (15 and 19 episodes, respectively). These results suggest that selective intestinal decontamination is useful in reducing the risk of infection from enterobacteria in patients with acute liver failure.


Hepatitis B x antigen and polymerase antibodies in the serum of hepatitis B carriers with or without hepatitis delta virus infection. Effects of interferon treatment.

Previous work has shown that the hepatitis B x antigen (HBxAg) and antibodies directed against the polymerase of hepatitis B virus (anti-pol) are early markers of hepatitis B virus (HBV) replication in natural infections. The present study was carried out to test the hypothesis that the appearance of one or both of these markers signaled reactivation in chronic carriers with liver disease who were treated with alpha-interferon (IFN). The results show that HBV DNA decreased among the patients who responded to therapy, and that among these responders, neither HBxAg nor anti-pol became detectable in serum for 12 months after treatment, in contrast to controls. Hence, the loss of HBxAg and anti-pol correlate with decreased levels of HBV DNA in response to IFN therapy. However, different patterns of HBxAg and anti-pol were observed among alpha-IFN-treated HBV carrier patients who were also chronically infected with the hepatitis delta virus (HDV). The treatment of such patients often resulted in the loss of HDV RNA from serum and delta antigen from liver. Most of these patients had increased levels of HBV DNA in serum. HBxAg and/or anti-pol also became detectable in patients who lost markers of HDV, implying that the suppression of HDV by IFN is accompanied by the appearance of early markers of HBV reactivation in some of the treated patients.


Failure of acyclovir to enhance the antiviral effect of alpha lymphoblastoid interferon on HBe-seroconversion in chronic hepatitis B. A multi-centre randomized controlled trial.

Serum HBeAg levels and HBe-seroconversion were investigated in patients with chronic HBeAg-positive hepatitis who were randomized to receive either alpha lymphoblastoid interferon (5 megaunits subcutaneously daily for 16 weeks) plus acyclovir (2 g intravenously daily during weeks 1 and 2 and weeks 9 and 10) (n = 49) or no treatment (n = 48). HBeAg levels in serial dilutions of patient serum were assessed quantitatively by radioimmunoassay and compared with the values found for negative control serum. One year after the start of therapy 44 treated patients and 43 control patients were available for follow-up. A complete response (HBe-seroconversion) occurred in 11 treated patients (25%) and six controls (14%) (difference: 11%, 95% CI-5-28%). A partial response (HBeAg less than 50% of initial level) was found significantly more often for treated patients (n = 13, 30%) than for controls (n = 2, 5%) (difference: 25%, 95% CI 10-40%). During acyclovir-interferon combination therapy the decrease in HBeAg level was similar to that achieved during therapy with interferon alone. We conclude that acyclovir does not enhance the effect of interferon on serum HBeAg levels. Since HBeAg levels continue to decline during interferon treatment and rebound thereafter to pretreatment levels, prolongation of therapy may yield a higher response rate.


Steatosis associated with immotile cilia syndrome: an unrecognized relationship?

The present study deals with a case of hepatic parenchymal steatosis in a child with primary ciliary dyskinesia (immotile cilia syndrome) well documented by functional and ultrastructural evaluation of the ciliary epithelia. Hepatic steatosis was associated with ultrastructural evidence of retention of material either in the cisternae of the endoplasmic reticulum or in proximity of the Golgi apparatus of hepatocytes. It is suggested that the absence of dynein in the axoneme is probably part of a diffuse genetic defect which may extend to cytoplasmic, non axonemal, dynein and lead to a disturbance of various microtubule-dependent cell activities.


Frequency and significance of chronic ulcerative colitis in severe corticosteroid-treated autoimmune hepatitis.

To assess the prevalence and significance of chronic ulcerative colitis in patients with severe autoimmune hepatitis and to determine the frequency of cholangiographic and histologic features of primary sclerosing cholangitis in those with colitis, 105 patients who had been screened by annual proctoscopic examination were studied. Patients with features of colitis were compared to counterparts without colitis who had been matched by age, sex, disease severity and treatment regimen. Seventeen patients (16%) had findings of chronic ulcerative colitis. Twelve of these underwent cholangiography and five (42%) had features of primary sclerosing cholangitis. Patients with and without cholangiographic abnormalities were indistinguishable by clinical, laboratory, immunoserologic, and histologic features. Fibrous obliterative cholangitis was present in only two patients, including one with normal cholangiography. Patients with colitis entered remission less frequently (59 vs. 94%, p less than 0.05), failed treatment more commonly (41 vs. 6%, p less than 0.05) and progressed to cirrhosis more frequently (75 vs. 25%, p less than 0.05) than counterparts without colitis. Patients with colitis but normal cholangiography, however, responded satisfactorily to therapy. We conclude that chronic ulcerative colitis can coexist with severe autoimmune hepatitis in the absence of primary sclerosing cholangitis or hepatitis C infection. Under such circumstances its presence does not adversely influence treatment outcome. Primary sclerosing cholangitis cannot be excluded by routine examinations and its presence is associated with a poor treatment response. Cholangiography should be considered in all patients with autoimmune hepatitis and colitis, especially in those recalcitrant to therapy.


Hepatitis D virus superinfection remains a rare occurrence in non-drug abusers in Hong Kong.

Recent studies from Taiwan reported an increased prevalence of hepatitis D virus (HDV) superinfection as well as a spread of HDV infection to non-drug abusers. We examined the current prevalence of HDV superinfection in Hong Kong by testing sera from a large cohort of patients with acute and chronic hepatitis B virus (HBV) infection, collected between January 1988 and December 1990, for IgG antibody to HDV (anti-HDV). Anti-HDV was detected in 13 (93%) of 14 parenteral drug abusers who had HBV-related chronic liver disease. However, anti-HDV was detected in only one (0.15%) of 664 non-drug abusers who had chronic HBV infection and none of 31% patients with acute hepatitis B. We concluded that HDV superinfection remains a rare occurrence in non-drug abusers in Hong Kong.


Uridine catabolism by the isolated perfused rat liver.

A new approach in the treatment of gastrointestinal tumors with 5-fluorouracil involves the infusion of high doses of uridine to improve the chemotherapeutic efficiency of the former. High amounts of uracil formed from uridine can interfere with the hepatic catabolism of 5-fluorouracil and thus increase its bioavailability and toxicity. In our study, we analysed the metabolite pattern of uridine in the effluent of isolated perfused rat livers in relation to portal uridine levels. The livers were perfused hemoglobin-free without recirculation at a constant flow. In the perfusate, uridine was changed from 0.5 to 100 mumol/l. The complete degradation of [2-14C]uridine and [2-14C]uracil was monitored via the release of labeled CO2. Radioactive catabolites of uridine including uracil and the sum of dihydrouracil and beta-ureidopropionate were separated by high-performance liquid chromatography and counted using a radioactivity flow monitor. Portal uridine concentrations were increased from 0.5 to 100 mumol/l and were accompanied by a rise in the relative amount of non-metabolized uridine in the effluent from 13 to 78%. At uridine concentrations above 50 mumol/l, there was a constant release of uracil into the effluent, indicating saturation of uridine phosphorolysis or transport. The amount of 14CO2 formed by the liver reflecting complete uridine breakdown was higher than any other uridine metabolite when uridine concentration varied from 0.5 to 15 mumol/l. Saturation of 14CO2 formation was achieved at a uridine concentration of 25 mumol/l. Higher peak values of 14CO2 release were observed after direct infusion of equivalent amounts of uracil into the portal vein.(ABSTRACT TRUNCATED AT 250 WORDS)


Detection of serum HBV-DNA by polymerase chain reaction (PCR) in patients before reactivation of chronic hepatitis B.

Reactivation of chronic hepatitis B is characterized by the reappearance of HBV-DNA in serum. The purpose of the study was to determine whether, before reactivation, HBV-DNA would be detectable in serum, using a sensitive procedure of detection, namely polymerase chain reaction (PCR). We studied 17 patients with chronic hepatitis B who experienced an episode of reactivation, defined by the reappearance of HBV-DNA in serum. None of these 17 sera had HBV-DNA demonstrable by dot-blot hybridization nor liquid hybridization in sera collected before reactivation. Using PCR, HBV-DNA was detected, before reactivation, in 13 of the 17 episodes of reactivation tested with Southern-blot and hybridization. HBV-DNA was not detectable with PCR in the serum of four patients who subsequently experienced an episode of reactivation. In conclusion, our results show low level HBV replication before reactivation in most, but not all, HBs-positive, HBV-DNA-negative patients. This suggests that reactivation may occur even in patients with no HBV-DNA demonstrable in serum with PCR prior to reactivation.


Peptide-induced modulation of target cell sensitivity to natural killing.

We have previously shown that the capacity of class I molecules to confer resistance to NK in transfected target cells maps to the Ag-binding site (ABS) of the HLA class I structure. Here we examine the effect of peptide (reagents specific for the ABS) pretreatment on the NK sensitivity of class I+ target cells. Synthetic peptides (10-17 amino acids in length) were used to pretreat C1R target cells expressing either no serologically detectable HLA-A, B class I molecules, or C1R transfectants expressing individual HLA-A or -B locus class I molecules. In each case in which the class I allele had previously been shown to directly bind a given peptide, peptide-pulsing of target cells resulted in increased sensitivity to NK-mediated conjugation and cytolysis. The NK susceptibility of C1R target cells expressing no HLA-A, B class I molecules or the nonprotective HLA-A2.1 or HLA-A2M70 mutant class I molecules was unaffected by pretreatment with HLA-A2-binding peptides. These results support the intimate involvement of the HLA class I ABS and potentially ABS-bound peptides in determining target cell sensitivity to NK. Furthermore, these findings form the basis of an effective screening procedure for discerning peptide class I allele-specific interactions.


Activation of resting peripheral blood lymphocytes through the T cell receptor induces rapid phosphorylation of Op18.

Op18 is a highly conserved major cytosolic phosphoprotein that has been implicated in signal transduction in a wide variety of cell types. Freshly isolated peripheral blood lymphocytes (PBL) constitutively express low levels of mostly unphosphorylated Op18. After mitogenic stimulation of PBL, Op18 synthesis is induced at a time when cells are entering S-phase. In this study, we have examined the phosphorylation of Op18 in freshly isolated PBL after activation of the T cell receptor by OKT3. Quantitative analysis of Op18 phosphorylation was undertaken by metabolic labeling with 32Pi and PhosphorImager analysis of two-dimensional gels. After 10 or 15 min of activation by OKT3, one of the three major phosphorylated forms of Op18, designated Op18c, increased approximately 10-fold, which represented a most pronounced change among a large number of phosphoproteins analyzed. In time course experiments, increased Op18 phosphorylation to yield Op18c was observed as early as 2 min. Continued OKT3-induced activation for 20 to 72 h resulted in a further increase in phosphorylated Op18 forms, which paralleled new Op18 synthesis and occurred at a time when cells were entering S-phase, as determined by [3H]-thymidine incorporation. Inhibitors of lymphoid proliferation, cyclosporin A and RPM, had no effect on early (less than 15 min) phosphorylation. Addition of calphostin C, a specific inhibitor of protein kinase C, 1 min prior to stimulation of resting T cells with OKT3 completely inhibited further phosphorylation of Op18. Incubation of PBL with calphostin C for 75 min decreased constitutive levels of phosphorylated Op18. In contrast, inhibition of cyclic nucleotide-dependent protein kinases with HA1004 had no effect on Op18 phosphorylation. Activation of cAMP-dependent protein kinase with Forskolin or 8Br-cAMP did not increase Op18 phosphorylation. Our results suggest that Op18 phosphorylation is mediated by protein kinase C activation as an early event in T cell activation through the T cell receptor.


Mapping of the rabbit MHC reveals that class I genes are adjacent to the DR subregion and defines an insertion/deletion-related polymorphism in the class II region.

Molecular analyses of genes in the rabbit MHC (RLA) by pulsed field gel electrophoresis have shown that the relative order of class II genes (DP, DO, DQ, DR) is identical to that in humans and similar to that in the mouse. However, a major difference from either HLA or H-2 was observed at the DR end of the RLA class II complex: class I genes are located in close proximity to DR with no interposed class III sequences. A MluI fragment of 180 kb and a 210-kb SalI fragment both hybridized with the DR probe as well as with different class I probes including that for pR27, a class I gene with T cell-limited pattern of expression. Comparison of two different RLA haplotypes, A and B, indicated that the distance between the DQ and DR subregions differs by approximately 700 kb in the two haplotypes. Testing other unrelated rabbits suggested that this difference segregates within the rabbit population and presumably derives from an insertion/deletion event in different haplotypes. A further difference between the A and B haplotypes included variable distance between genes encoding DO beta and DP; the DR end of the complex and the class I genes linkage was conserved in the two haplotypes.


VH genes in tandem array comprise a repeated germline motif.

In a study of human VH gene heterogeneity, we have previously used sequence-specific oligonucleotide probes to demonstrate polymorphism of 56pl and three highly homologous VH3 germline elements. We now extend these findings with VH nucleotide sequences obtained from a person who possesses restriction fragments corresponding to each of these four VH3 genes. From a lambda-phage library of genomic DNA, distinct phage clones containing putative 56pl, hv3005, 1.9III, and hv3019b9 genes were selected by screening with oligonucleotide probes. PCR amplification, subcloning, and sequencing from the respective clones 3d216, 3d24, 3d28, and 3d277, yielded exact 56p1, hv3005, 1.9III, and hv3019b9 nucleotide sequences. A panel of oligonucleotide probes was shown to hybridize to these cloned VH3 genes with exact specificity, demonstrating the ability of the probes to predict the sequence of detected target DNA. Based on their chromosomal organization and their previously determined distribution in the population, these VH3 genes represent at least three distinct loci. From each of the VH3-containing phage clones, a VH4 element was also identified and sequenced. Linked to 3d24 and 3d28, respectively, were VH4 sequences identical to hv4005 and 1.9II, corroborating previous reports. The VH4 elements linked to 3d216 and 3d277 were distinct from published VH4 sequences. Nucleotide sequence homology was 97 to 99% among the VH3 sequences, and 93 to 99% among the VH4. These findings indicate that the VH3-VH4 gene pairs we have identified are a repeated germline motif, apparently resulting from multiple duplications of tandemly arrayed VH genes.


VH-related idiotopes detected by site-directed mutagenesis. A study induced by the failure to find CD4 anti-idiotypic antibodies mimicking the cellular receptor of HIV.

The function of the CD4 cell surface protein as coreceptor on T helper lymphocytes and as receptor for HIV makes this glycoprotein a prime target for an immune intervention with mAb. A detailed understanding of the structural determinants on the therapeutic CD4 mAb that are involved in Ag binding or are recognized by anti-idiotypic mAb (anti-Id) may be important for designing antibodies with optimal therapeutic efficacy. Seven anti-Id raised against the CD4 mAb M-T310 were selected from a large panel with the intention to obtain CD4 mimicking structures with specificity for HIV gp120. The selected anti-Id did not react with other CD4-specific mAb cross-blocking M-T310. Among these, mAb M-T404, although having the same L chain as M-T310 and a VH region sequence differing only at 14 amino acid positions, was not recognized by the anti-Id. M-T310 H chain complexed with the J558L L chain reacted with all anti-Id, thus demonstrating that the recognized idiotopes are located within the VH region. To identify the idiotopes of M-T310 seen by the anti-Id, variants of M-T404 containing one or more of the M-T310-derived substitutions were generated by oligonucleotide-directed mutagenesis. The reactivity pattern of the mutant proteins with the anti-Id demonstrated that the idiotopes reside within the complementarity determining region (CDR) 2 and CDR3 loops of the VH region. A major idiotope was defined by a single amino acid in CDR2 that was recognized by three anti-Id, whereas the four other anti-Id reacted with determinants of CDR3. Although the performed amino acid substitutions did influence the Id recognition, Ag binding was not significantly affected, suggesting that none of the anti-Id can be considered as a mimicry of the CD4 Ag.


IL-4 receptor signal transduction in human monocytes is associated with protein kinase C translocation.

IL-4 regulates B cell differentiation and monocyte functions. Protein kinases, such as kinase C (PKC), transduce receptor signals. The involvement of PKC in IL-4R signaling was investigated in human monocytes. Treatment with IL-4 (10 ng/ml) for 10 min resulted in a significant redistribution of the PKC activity from cytosol to nuclear fraction. Total PKC activity localized in the nuclear fraction of IL-4-treated and control monocytes was, respectively, 68 and 19%. In contrast, similar PKC activity was found in membrane fraction of IL-4-treated and control cells. The kinetics of IL-4-mediated redistribution of PKC activity to the nuclear fraction were rapid. Within 30 s of IL-4 exposure, 29% of the total PKC activity localized in the nuclear fraction as compared to 15% in control monocytes and increased to 69% at 10 min. The PKC activity in the nuclear fraction appears to be a sequestered form. Extraction with Triton X-100 and additional sonication were required for functional assay of PKC activity. Additional support for PKC involvement in IL-4R signaling is provided by the dose-dependent effect of IL-4 on PKC activity and the abrogation of this effect after heat denature and immunoabsorption of IL-4. Furthermore, electron microscope examination and subcellular marker enzyme assays excluded significant contamination of the nuclear fraction by plasma membranes or subcellular organelles and IL-4 altering membrane disruption. The data presented indicate that IL-4R signaling in human monocytes involves PKC translocation to a nuclear fraction.


Differences between ATP-mediated cytotoxicity and cell-mediated cytotoxicity.

There has been considerable recent interest in the possible role of extracellular ATP in cell-mediated cytotoxicity. To investigate the similarities and differences between ATP-mediated lysis and CTL-mediated lysis, we studied in detail the ATP-mediated lysis of EL-4 cells, which are highly susceptible to lysis by extracellular ATP. ATP-mediated lysis was time and concentration dependent; most lysis occurred within 4 to 6 h of incubation. The kinetics of ATP- and cell-mediated lysis were similar. However, in contrast to CTL-mediated lysis, ATP-mediated lysis of EL-4 cells did not appear to be accompanied by characteristic chromosomal degradation (apoptosis). In order to compare these cytotoxic processes in greater detail, ATP-resistant clones were derived by growing EL-4 cells continuously in the presence of high concentrations of ATP. These cloned EL-4 lines showed marked resistance to ATP-mediated cytotoxicity across a wide range of concentrations but were as susceptible as the parent EL-4 cells to cell-mediated cytotoxicity by an alloreactive effector population from a MLC. Thus, there appear to be a number of differences between ATP-mediated and cell-mediated cytotoxicity in this system; most notable is the identification of cell lines that are resistant to ATP but susceptible to CTL-mediated lysis.


Administration of human recombinant IL-7 to normal and irradiated mice increases the numbers of lymphocytes and some immature cells of the myeloid lineage.

In vitro experiments performed by several investigators have demonstrated that IL-7 is a growth factor for immature B lymphocytes, thymocytes, and mature T lymphocytes. To evaluate the potential therapeutic use for human rIL-7 (rhuIL-7) as a hematopoietin, we have studied the in vivo hematopoietic effects of rhuIL-7 in mice. In these experiments, sublethally irradiated and normal mice were treated with or without rhuIL-7 for up to 26 days. Administration of rhuIL-7 significantly increased the white blood cell count in the peripheral blood and spleen in both normal and irradiated mice. Treatment with rhuIL-7 also accelerated lymphocytic recovery in irradiated mice. Precursor and mature B lymphocytes showed the greatest expansion in response to rhuIL-7 administration, with smaller increases in T lymphocytes being observed. In mice recovering from high dose irradiation, rhuIL-7 treatment resulted in preferential expansion of CD8+ T lymphocytes and more rapid normalization of the CD4/CD8 ratios. Differential analysis of peripheral blood smears demonstrated that rhuIL-7 also increased the numbers of immature granulocytes in both normal and irradiated mice. Moreover, administration of rhuIL-7 to normal, irradiated, cyclophosphamide-pretreated, or 5-fluorouracil-pretreated mice increased the number of acetylcholinesterase-positive megakaryocytes in the spleen, but not the bone marrow. Therefore, although the major in vivo effects of rhuIL-7 were on cells of the lymphocytic lineage, rhuIL-7 also increased the numbers of some immature cells of the myeloid lineage.


IL-1 beta maturation: evidence that mature cytokine formation can be induced specifically by nigericin.

Mouse peritoneal macrophages stimulated with LPS produce large amounts of pro-IL-1 beta. When these cells were pulse-labeled with [35S]methionine, however, little labeled cytokine appeared in the medium after a chase, and that which was externalized was not processed to its mature biologically active form. In an effort to promote proteolytic maturation of IL-1 beta, macrophages were treated with agents that were expected to compromise their viability. The calcium ionophore A23187 and the detergent saponin caused complete release of nonprocessed 35-kDa pro-IL-1 beta and liberation into the extracellular medium of the cytoplasmic marker enzyme LDH and the lysosomal enzyme beta-N-acetylglucosaminidase. Hypotonic lysis resulted in the release of a 20-kDa IL-1 beta species that was distinct from the 17-kDa mature species. Importantly, incubation of the murine macrophages with the potassium/proton ionophore nigericin led to a quantitative conversion of pro-IL-1 beta to a 17-kDa species. The N-terminus of this nigericin-derived product possessed the amino acid sequence expected for mature biologically active IL-1 beta. Monensin, an ionophore similar to nigericin, did not induce release or proteolysis of IL-1 beta. Complete release of mature IL-1 beta required concentrations of nigericin in excess of 2 microM and a minimum of 10 min of treatment. Mature 17-kDa IL-1 beta was observed within the nigericin-treated cells before their lysis. Nigericin's effect was not limited to mouse peritoneal macrophages, inasmuch as the ionophore also induced release and proteolytic maturation of IL-1 beta produced by LPS-stimulated human peripheral blood monocytes. Treatment of macrophages with LPS and nigericin, therefore, results in a unique series of intracellular events that promote formation of mature 17-kDa IL-1 beta.


CD11b (Mac-1): a marker for CD8+ cytotoxic T cell activation and memory in virus infection.

We have found that CD11b, a cell surface integrin of macrophages, granulocytes, and NK cells, is expressed by a subset of CD8+ T cells that include both the active virus-specific CTL and the virus-specific memory CTL populations. CD8+CD11b+ cells comprise less than 3% of naive mouse splenocytes, but after lymphocytic choriomeningitis virus (LCMV) infection increase by 9- to 12-fold by the peak (day 8) of the virus-specific CTL response. Depletion of day-8 splenocytes with anti-Mac-1 and C' or enrichment by sorting for CD11b+ or CD8+CD11b+ spleen cells demonstrated that LCMV-specific CTL are CD11b+. The CD11b+ subpopulation also contained the bulk of the IL-2-responsive CD8+ cells. MEL-14, a homing marker down-regulated on activated T cells, was down-regulated on the majority of CD8+ cells that became CD11b+. Less than 1% of LCMV-immune splenic lymphocytes expressed CD11b. Antibody and C' depletion of this population severely impaired the ability of immune splenocytes to respond to in vitro secondary stimulation with LCMV-infected peritoneal macrophages, but did not affect the generation of a primary allospecific CTL response in MLC. Mixing of CD8-depleted and CD11b-depleted LCMV-immune splenocytes failed to restore the ability of these cells to mount a virus-specific memory CTL response, indicating that a cell coexpressing CD8 and CD11b is essential for this response. As determined by limiting dilution analysis, the precursors for the LCMV-specific memory CTL response were enriched in the CD11b+ population of LCMV-immune splenocytes. CD11b stained far fewer CD8+ splenocytes from naive mice than did CD44 (Pgp-1), and among immune splenocytes it identified a small subpopulation of CD44hi cells, indicating that CD11b may be the best single marker available for discriminating between naive and memory CD8+ T cells.


Biosynthesis and secretion of complement component (C3) by activated human polymorphonuclear leukocytes.

We tested the hypothesis that human polymorphonuclear leukocytes (PMN), bearing complement receptors CR1 and CR3, might also synthesize C3, particularly when activated by LPS or cytokines. Northern blot analysis of total RNA, obtained from purified PMN stimulated overnight with LPS or cytokines (IFN-gamma, TNF-alpha, and IL-1) showed the 5.3-kb RNA transcript reported for C3 in hepatocytes and monocytes. No transcripts for C4 and factor B were detected. Time course studies of C3 mRNA expression in PMN treated with LPS or TNF-alpha demonstrated a steady increase with a plateau at 24 h that correlated with secretion of C3, determined by ELISA. In contrast, IFN-gamma and IL-1 induced a transient increase in C3 transcript with a peak around 8 h after stimulation, which was not reflected in an increased rate of C3 secretion. The content of C3 protein in PMN culture media, measured by ELISA, was about 4 ng/ml/10(7) cells after overnight stimulation with LPS or TNF-alpha. A very small amount of C3 (about 0.7 ng/ml/10(7) cells) was detected in supernatants from unstimulated and IFN-gamma- or IL-1-induced PMN. Immunoprecipitation with a polyclonal anti-human C3, followed by SDS-PAGE analysis, from [35S]methionine labeled PMN, revealed the presence in culture supernatants of three major bands at 185, 115 and 70 kDa, corresponding to pro-C3, alpha and beta chains, respectively. Analysis of [14C]methylamine incorporation and of autolytic cleavage showed that the C3 produced in tissue culture by PMN contained an intact thiolester bond. The capacity of PMN to secrete functional C3 in response to LPS and TNF-alpha might be an important mechanism of host defense at sites of inflammation.


Inhibition of phospholipase A2 activity in guinea pig eosinophils by human recombinant IL-1 beta.

The effect of human rIL-1 beta on the release of arachidonic acid (AA) and on the phospholipase A2 (PLA2) activity in guinea pig eosinophils was investigated. Stimulation of [3H]AA-labeled eosinophils with the ionophore A23187 resulted in a time and concentration-dependent release of AA in parallel to hydrolysis of endogenous phosphatidylcholine (PC). Both events were abrogated by the chelation of intracellular free calcium, but not by its depletion from the medium, suggesting that the ionophore-induced AA release involves a PLA2 activity dependent on the mobilization of intracellular calcium. Addition of human rIL-1 beta (0.01 to 100 ng/ml) to eosinophils for 15 min had no effect on the release of AA induced by the ionophore. However, prolonged incubation with human rIL-1 beta (30 to 180 min) inhibited in a concentration- and time-dependent manner the release of AA and the hydrolysis of phosphatidylcholine in ionophore-stimulated eosinophils. Our results also showed that eosinophil homogenates contain a calcium-dependent PLA2 whose activity was markedly reduced when eosinophils were pretreated with human rIL-1 beta. The inhibition was time and concentration dependent and was observed in the presence of calcium and phospholipid excess. Finally, studies with Fura-2-loaded eosinophils showed that the ionophore A23187 stimulated an increase in intracellular calcium concentration that was not altered by pretreating the eosinophils with human rIL-1 beta. These results suggest that human rIL-1 beta inhibits the release of AA by eosinophils via the inhibition of a PLA2 activity and through a calcium-independent mechanism. Inhibition by human rIL-1 beta required a prolonged incubation (30 to 180 min) and was observed after its removal from the medium, suggesting that human rIL-1 beta did not interact directly with the PLA2 itself, but with a metabolic process involved with the regulation of its activity in eosinophils.


Comparison of actin changes and calcium metabolism in plastic- and fibronectin-adherent human neutrophils.

Human neutrophils adherent to a polystyrene plastic surface are vigorously activated, whereas those adherent to fibronectin manifest only a priming response. The basis of these metabolic differences was further characterized; polystyrene-adherent cells, which were shown to spread quickly upon adhesion, exhibited an increase of cytoskeleton-associated actin (F-actin) (measured by a nitrobenzoxadiazole-phallacidin fluorescent staining assay) and a decrease of monomeric G-actin concentration (measured by a DNase inhibition assay); in contrast, fibronectin-adherent cells exhibited little spreading and decreased their F-actin, after 1.5 min of adhesion, to 33.49 +/- 6.9% (mean +/- SD, n = 5) of initial levels found in suspended cells before plating. Actin depolymerization in fibronectin-adherent cells was confirmed by measuring G-actin, which sharply increased during the first minute of adhesion, rising from 0.065 +/- 0.007 to 0.20 +/- 0.035 microgram/microgram of protein (mean +/- SEM, p less than 0.05), and then remained elevated during 5 min of observation. In contrast, soluble fibronectin induced a decrease of G-actin in suspended cells. Cells pretreated with 1 microM cytochalasin D and allowed to adhere to a plastic surface did not spread, failed to generate O2-, and exhibited elevated concentrations of G-actin (0.1 to 0.2 microgram/microgram of protein) during the 5 min of observation. Actin changes, as well as respiratory burst, in adherent cells were shown to proceed through a pertussis toxin-insensitive pathway. Fluo-3 measurements of intracellular Ca2+ concentrations ([Ca2+]i) showed a fourfold and twofold [Ca2+]i increase in polystyrene- and fibronectin-adherent cells, respectively, after 2 min. The small rise in [Ca2+]i in fibronectin-adherent cells corresponds to a primed response of these cells to subsequent activation with FMLP. Ionomycin (1 microM) added to neutrophils just before adhesion on fibronectin induced full activation, i.e., O2- production and actin polymerization. The metabolic events controlling metabolic priming and actin depolymerization are as yet uncharacterized, but fibronectin receptor-linked responses beyond the mediation of cell adhesion have now been identified, suggesting complex metabolic functions of integrin receptors.


Neutralization of HIV-1 by anti-idiotypes to monoclonal anti-CD4. Potential for idiotype immunization against HIV.

Anti-idiotypic antibodies were raised in rabbits against a panel of 11 murine mAb directed to the human CD4 receptor. Selection of mAb for vaccination was based on inhibition studies demonstrating that these mAb recognized CD4/V1 epitopes implicated in HIV-1-gp120 binding. Purified antisera showed high titer anti-Id activity and reacted specifically with Ag-combining site-related Id of the mAb used for their generation. Anti-Id either detected a private Id of the immunizing mAb or displayed a partial cross-reactivity with Id of other mAb to CD4. Eight anti-Id to six different mAb were shown to recognize determinants of recombinant HIV-1-gp120 or of HIV-1-gp160 as shown by ELISA and radioimmunoprecipitation assay. These anti-Id were capable of inhibiting HIV infection up to 100% in a MT-4 cell assay in vitro. In addition to neutralizing infectivity of cell-free virus, anti-Id to two mAb--the mAb IOT4a and 7.3F11--were also shown to inhibit HIV-induced syncytia formation up to 100%. Anti-Id to the mAb IOT4a, 7.3F11, and to the mAb anti-Leu3a interfered with rgp120 binding to cellular CD4 as assessed by flow cytometry. These results demonstrated that mAb specific for both CDR2- and CDR3-like regions of CD4 were capable of inducing HIV-1-gp120 cross-reacting anti-Id neutralizing HIV-1 in vitro. These studies may have implications for the development of a gp120 internal image based vaccine against HIV.


Role of cytokines in alveolar macrophage accessory cell function in HIV-infected individuals.

Mononuclear phagocytes, including alveolar macrophages (AM), can be chronically infected with HIV and thus serve as a reservoir for the virus. Acting as AC during the generation of an immune response, HIV-infected mononuclear phagocytes can facilitate viral T cell infection by several mechanisms, including direct contact of T cells with HIV-infected macrophages as well as cytokine-induced up-regulation of latent T cell infection. Our laboratory has shown that AM from HIV-infected individuals have enhanced AC function compared to normal AM. In this study we explored AM production and secretion of IL-1 beta and IL-6, two cytokines critical for optimal AC function, in normal volunteers and HIV-infected patients. Cultured AM supernatants and lysates were generated in the presence and absence of LPS and standard mitogens. In initial mixing experiments HIV AM supernatants enhanced mitogen-induced T cell proliferation using normal AM as AC significantly more than normal AM supernatants, suggesting that HIV AM secreted more T cell stimulatory factors than normal AM. Neither group could enhance T cell proliferation induced by HIV AM suggesting these cells already secreted optimal levels of these factors. AM from HIV+ individuals produced and secreted more IL-1 beta (measured by ELISA) and IL-6 (measured in a B9 bioassay and by immunoprecipitation) than normal AM both spontaneously and in the presence of low LPS concentrations and mitogens. In some cases depleting HIV AM supernatants of IL-1 beta and IL-6 on immunoaffinity columns abrogated their enhancement properties indicating that these cytokines were important in the observed enhancement. However, in other patients different factors must also be involved as depletion of IL-1 beta and IL-6 in their AM supernatants had no effect on enhancement function. These results show that HIV AM secretory products are important in the enhanced AC function demonstrated by these cells. However, although augmented IL-1 beta and IL-6 secretion likely contribute significantly to this enhancement, other AC secretory factors and/or functions must also be involved.


A flow-cytometric method to study DNA fragmentation in lymphocytes.

A method to measure DNA fragmentation cell by cell in a cell population was implemented based on acridine orange procedure to determine DNA content of single cells by flow cytometry. Using this method it can be observed that the fragmentation process induced by irradiation in thymic cells occurs in a fraction of the population, thus indicating that this process is not evenly distributed over the total population, and that it corresponds to a fast phenomenon in which the cells suddenly lose DNA material.


Cytokine patterns of immunologically mediated tissue damage.

Reactional states in leprosy are produced by different immunologic mechanisms and are responsible for a major component of tissue damage of the disease. Reversal reactions exhibit increased CD4 T cell infiltration in lesions and augmented cell-mediated immune reactivity to Ag of Mycobacterium leprae that can rapidly produce nerve damage. Erythema nodosum leprosum (ENL) reactions also have CD4 T cell infiltration but appear to be associated with the formation of immune complexes that are responsible for panniculitis, arthritis, vasculitis, and nerve injury. Because these reactional states may serve as paradigms for other types of human immunologically mediated tissue damage, this study sought to characterize the dynamic changes in cytokines associated with these reactions. Expression of cytokine mRNA in lesions of leprosy reactional states were measured by PCR. In reversal reactions, IL-1 beta, TNF-alpha, IL-2, and IFN-gamma mRNA were prominent and found to increase during the reaction, concomitant with decreases in expression of mRNA for IL-4, IL-5, and IL-10. In ENL, selective increases in the expression of IL-6, IL-8, and IL-10 mRNA was observed, with persistent expression of IL-4 and IL-5 mRNA. Reversal reactions represent naturally occurring delayed-type hypersensitivity reactions that favor macrophage activation and protective immunity, but which can engender concomitant cell injury. In contrast, ENL lesions represent immediate-type hypersensitivity reactions reflecting the selective stimulation of cytokines that attract neutrophils, stimulate antibody production, and down-regulate macrophage activation. The analysis of cytokine dynamics within different inflammatory responses can provide insights into immune mechanisms of tissue damage, and provide a useful framework for developing strategies for therapeutic intervention.


Biological activities of polyethylene-glycol immunoglobulin conjugates. Resistance to enzymatic degradation.

Serum IgG has been covalently bonded to polyethylene glycols of either 2000 or 8000 molecular weight to produce immunoglobulin conjugates with 4.4-27.2% of primary amines bonded to polyethylene glycol. Polyethylene glycol immunoglobulin conjugates retain the ability, comparable to native IgG, to bind to a range of protein and microbial antigens, but have a reduced ability to bind to Fc receptors or to fix complement C3. When 6.8% or more of available primary amines are conjugated, IgG-PEG conjugates are impervious to trypsin, and at 14% or more conjugation, more resistant than native IgG to pepsin and chymotrypsin. We suggest that PEG-Ig conjugates may be useful for the oral treatment of various gastrointestinal diseases in which secretory humoral immunity is insufficient.


Characterization of monoclonal antibodies physically adsorbed onto polystyrene latex particles.

The physicochemical and immunochemical properties of monoclonal antibodies (MAbs), adsorbed onto polystyrene latex particles, have been investigated. Both native and pH 2 pretreated MAbs were compared before and after immobilization. It was found that the antigen binding capacity of the immobilized, acidic pretreated MAbs was significantly higher than for the immobilized, native IgG molecules. This enhanced antigen binding capacity appeared to be due to an improved molecular orientation following adsorption of the monomeric, pH 2 treated IgG fraction. Additionally, experiments using F(ab')2 fragments demonstrated that the Fc portion of the MAb molecule is of major importance for achieving the enhanced binding capacity. Binding studies showed that the (apparent) affinity of native and pH 2 pretreated MAbs were similar; the K(a) values of the immobilized MAbs were higher than those of MAbs in solution.


U937 cells stimulated with opsonised zymozan particles provide a convenient laboratory source of tumour necrosis factor alpha.

The U937 cell line has been shown to generate tumour necrosis factor alpha (TNF-alpha) in response to soluble stimuli such as PMA and LPS, but only after treatment with GM-CSF. We report here the generation of TNF-alpha from U937 cells following phagocytosis of opsonised zymozan particles without the need for pre-treatment with GM-CSF. The release of TNF-alpha from U937 cells was demonstrated by a specific radioimmunoassay, L929 cell killing and neutrophil 'priming'. The biological activities in the cell supernatant were inhibited by TNF-alpha antiserum. Phagocytosis was required for TNF-alpha production. Non-opsonised zymozan or latex particles which were not phagocytosed or pretreatment with cytochalasin B, which inhibited phagocytosis of opsonised zymozan particles, all failed to trigger TNF-alpha production. Phagocytosis failed to trigger detectable IL-1 generation, and production of IL-6 was insufficient to produce biological effects on neutrophils. The U937 supernatant thus provides a source of human TNF-alpha which can be generated conveniently and cheaply for experimental investigations.


Evaluation of biological response modifiers in the enhancement of tumor uptake of technetium-99m labeled macromolecules. A preliminary report.

Imaging tumors with radioactive monoclonal antibodies remains attractive but continues to be challenging. With the hypothesis that the use of biological response modifiers (BRMs) may augment the tumor uptake, technetium-99m(99mTc)-labeled tumor necrosis factor (TNF) and nuclear histone specific TNT-1-F(ab')2 were evaluated in tumor bearing mice given a single dose of interferon (IFN). Ukrain or pokeweed mitogen as BRMs. As early as 1.5 h post injection (p.i.) of the radioactive macromolecules, the absolute tumor uptake (% administered dose/g) of each agent was enhanced (e.g., TNF, control = 1.8 +/- 0.4, Ukrain = 3.2 +/- 0.5, P = 0.006) and tumor to muscle ratios were elevated (e.g., TNF, control a 4.1 +/- 2.2, interferon 8.3 +/- 2.7, P = 0.01). The absolute tumor uptake remained practically unchanged at 4 h p.i. Generally with BRMs, the blood clearance was rapid and tumor/blood ratios and tumor/muscle ratios were higher than in the control group, increasing to greater than 200% for IFN as a BRM. The early enhancement in tumor uptake of macromolecules, leading to excellent delineation of tumors by scintigraphy is highly encouraging and warrants further studies to explore the full potential of BRMs.


Quantitative dot-blot assay for proteins using enhanced chemiluminescence.

A sensitive non-radioactive method for detection of specific proteins on Western blots is commercially available. The protein is immobilized on nitrocellulose membrane and immunolabelled with HRP-conjugated secondary antibody. HRP catalyzes the oxidation of luminol, a cyclic diacylhydrazide, resulting in the emission of light which is recorded on film. Using dot blot, we have shown that the signal generated by this system is proportional to the amount of protein loaded onto the membrane. Standard curves were linear (r2 greater than 0.95) over a 10-50-fold range. Linearity was also achieved with tissue extracts probed for a specific antigen. The sensitivity of the method is such that less than 10 fmol protein can be measured. The sensitivity and range are comparable to a previously reported dot-blotting assay based on the use of 125I-protein A, but the method does not require the handling of radioactive compounds. This method was used to estimate the level of chromogranin A in a mixture of proteins extracted from human brain.


Isolation of functionally active intraepithelial lymphocytes and enterocytes from human small and large intestine.

A mild purification method has been developed for the isolation of human intraepithelial lymphocytes (IEL) and enterocytes from the same individual. The isolation procedure includes mechanical disruption of the mucosal layer, treatment with reducing agent and sedimentation followed by Percoll gradient centrifugation. Finally, epithelial cells are removed from the IEL fraction using magnetic beads coated with the anti-epithelial antigen monoclonal antibody (mAb) BerEP4. Leucocytes are removed from the enterocyte fraction using magnetic beads coated with mAbs directed against common leucocyte antigen (CD45). Using this procedure IEL and enterocytes have been isolated from apparently normal jejunal, ileal and colonic tissue specimens. Recoveries of IEL were 7 x 10(5), 4 x 10(5) and 1 x 10(5)/cm2 mucosa from jejunum, ileum and colon respectively. 1-2 x 10(6) enterocytes/cm2 mucosa were recovered from small intestine while the corresponding value for colonic biopsies was approximately 2 x 10(5) enterocytes/cm2. The IEL fraction was pure as judged by the low percentages of B cells, macrophages and BerEP4 positive cells (less than 4%) present in the purified fraction. The enterocyte fraction contained less than 2% CD45+ cells. The two cell fractions were viable and expanded in vitro. Enterocytes expanded spontaneously while IEL required initial stimulation with mitogens. The isolation procedure described here will make it possible to study the function of human IEL, interactions between IEL and enterocytes and the role of both cell types in local immunity.


Immunology of Giardia and Cryptosporidium infections.

Giardiasis and cryptosporidiosis occur in immunocompetent persons, and as opportunistic infections in immunocompromised individuals. The causative organisms of giardiasis (Giardia lamblia) and cryptosporidiosis (Cryptosporidium parvum) have markedly different life cycles, but currently available evidence suggests that intestinal antibody plays a major part in host protection against both parasites. In the case of G. lamblia, the evidence points to a role for intestinal antitrophozoite antibody in blocking adherence of trophozoites to intestinal epithelium, with consequent removal of trophozoites from the intestine by peristalsis. Similarly, recent work supports the view that C. parvum infection can be prevented or interrupted by antibody that binds to Cryptosporidium life cycle stages present in the intestinal lumen (sporozoites and merozoites). Molecular characterization of G. lamblia and C. parvum immunodominant antigens would facilitate future studies aimed at determining whether vaccination against giardiasis or cryptosporidiosis is feasible.


Deficient polymorphonuclear cell and mononuclear cell antibody-dependent cellular cytotoxicity in pediatric and adult human immunodeficiency virus infection.

Children with human immunodeficiency virus (HIV) infection have impaired polymorphonuclear leukocyte (PMNL) function leading to secondary bacterial infections and possible acceleration of underlying viral disease. The chief antiviral defense mechanism of PMNL is antibody-dependent cellular cytotoxicity (ADCC). Accordingly, the ADCC of PMNL and mononuclear cells from HIV-positive children was compared with that of HIV-positive adults, healthy adults, and age-matched healthy children. PMNL and mononuclear cells from HIV-positive children incubated with hyperimmune HIV immune globulin (HIVIG) gave significantly lower ADCC compared with PMNL or mononuclear cells of healthy age-matched children incubated with HIVIG (P less than .05). The ADCC of mononuclear cells of healthy adults in the presence of plasma from HIV-infected children was significantly less than that of the same cells in the presence of plasma from HIV-positive symptomatic or asymptomatic adults. Augmentation of ADCC of the PMNL from HIV-positive children with interferon-gamma or granulocyte-macrophage colony-stimulating factor did not occur. Thus, the defect in ADCC of HIV-positive children is due to defects of both effector cells and antibody function.


Antibody response to rubella virus antigen and structural proteins in retinitis pigmentosa.

Elevated serum ELISA IgG antibodies to rubella virus (RV) were found by three independent determinations in 41 (72%) of 57 adults with the retinal degeneration retinitis pigmentosa, while antibody responses to five other common neurotropic viruses were normal. However, these patients lacked clinical signs of active RV infection or known recent RV exposure, and 56 lacked IgM anti-RV antibody. Unusual relative percentages of IgG antibody to RV structural proteins compared with those of controls were found in patients' sera by radioimmunoprecipitation assay. For retinitis pigmentosa patients, percentage of RV envelope glycoprotein E1 antibody was similar to, of RV envelope glycoprotein E2 antibody was greater than, and of antibody to RV nucleocapsid C protein was lower than control percentages. Abnormal immunity to RV was also suggested by a lack of increased proliferation of lymphocytes to RV antigen despite elevated anti-RV antibody in patients with retinitis pigmentosa. Not associated with age or particular genetic pattern, these divergences from normal immunity suggest an unusual association between RV proteins and retinitis pigmentosa.


Meningococcal disease and influenza-like syndrome: a new approach to an old question.

The temporal and spatial association of meningococcal disease and influenza-like syndrome (ILS) was assessed from surveillance data on these diseases in France for a 6-year period (1985-1990). Using time series methods to account for the usual seasonal pattern meningococcal disease, the incidence of meningococcal disease in a given week was found to be linked to incidence of ILS in the 5 previous weeks but not to that in subsequent weeks. Geographic spread of meningococcal disease correlated with spread of ILS. This study also suggests that meningococcal disease is more severe for a 2-month period during and after an ILS epidemic: The proportion of cases with purpura fulminans increased by 24% and those resulting in death by 26% during this period. No shift in the age distribution was observed. When an ILS epidemic is identified, medical practitioners should be informed of the likelihood of an increased incidence and severity of meningococcal disease.


Lactate and glucose concentrations in brain interstitial fluid, cerebrospinal fluid, and serum during experimental pneumococcal meningitis.

Metabolic abnormalities during bacterial meningitis include hypoglycorrhachia and cerebrospinal fluid (CSF) lactate accumulation. The mechanisms by which these alterations occur within the central nervous system (CNS) are still incompletely delineated. To determine the evolution of these changes and establish the locus of abnormal metabolism during meningitis, glucose and lactate concentrations in brain interstitial fluid, CSF, and serum were measured simultaneously and sequentially during experimental pneumococcal meningitis in rabbits. Interstitial fluid samples were obtained from the frontal cortex and hippocampus by using in situ brain microdialysis, and serum and CSF were directly sampled. There was an increase of CSF lactate concentration, accompanied by increased local production of lactate in the brain, and a decrease of CSF-to-serum glucose ratio that was paralleled by a decrease in cortical glucose concentration. Brain microdialysate lactate concentration was not affected by either systemic lactic acidosis or artificially elevated CSF lactate concentration. These data support the hypothesis that the brain is a locus for anaerobic glycolysis during meningitis, resulting in increased lactate production and perhaps contributing to decreased tissue glucose concentration.


Prophylaxis against wound infection following herniorrhaphy or breast surgery.

The effect of perioperative antibiotic prophylaxis on definite wound infections was assessed for 3202 herniorrhaphies or selected breast surgery procedures. Patients were identified preoperatively and monitored for greater than or equal to 4 weeks. Thirty-four percent of patients (1077/3202) received prophylaxis at the discretion of the surgeon; 86 definite wound infections (2.7%) were identified. Prophylaxis recipients were at higher risk for infection, with a higher proportion of mastectomies, longer procedures, and other factors. Patients who received prophylaxis experienced 41% fewer definite wound infections (odds ratio [OR], 0.59; 95% confidence interval [CI], 0.35-0.99; P = .04) and 65% fewer definite wound infections requiring parenteral antibiotic therapy (OR, 0.35; 95% CI, 0.15-0.88; P = .02) after adjustment for duration of surgery and type of procedure. Additional adjustment for age, body mass index, the presence of drains, diabetes, and exposure to corticosteroids did not change the magnitude of this effect meaningfully. The effect of prophylaxis was similar for all procedures studied. In the absence of formal guidelines, surgeons at these institutions administered prophylaxis preferentially to patients at highest risk.


Participation of complement in the phagocytosis of the conidia of Aspergillus fumigatus by human polymorphonuclear cells.

Nonspecific immunity plays a major role in the clearance of the opportunistic fungal pathogen, Aspergillus fumigatus. However, the mode of recognition of Aspergillus conidia by the different elements of an intact phagocyte system remains poorly understood. This study concentrated on the initial interaction between infective conidia and human polymorphonuclear cells (PMNL) using a double-label immunofluorescent method. At 37 degrees C, association was rapid. Although ingestion rates were slower than association, both association and ingestion plateaued at 90 min and were considerably reduced in the presence of fetal calf serum. In the presence of autologous plasma, association increased as the conidia-to-cell ratio increased, but the percentage of associated conidia that were ingested decreased. At 4 degrees C, phagocytosis was negligible. Phagocytosis experiments in the presence of plasma treated in various ways to inhibit the complement pathways demonstrated that optimal association was dependent on an active alternative complement pathway.


Adherence of Candida albicans to tissues from mice with drug- or radiation-induced immunodeficiencies.

Host factors that influence binding of Candida albicans to murine spleen, lymph node, and kidney were studied. Organs were harvested from BALB/cByJ and AKR/J mice immunocompromised by irradiation, cyclophosphamide, and cortisone acetate alone or in combination. Tissues from treated mice and untreated littermates were compared for their ability to bind C. albicans in ex vivo assays. Immunosuppressive regimens decreased yeast binding to splenic marginal zones, but when mice recovered for 5 days after treatment, adherence to spleen was similar to adherence in untreated littermates. Adherence to lymph node and kidney in treated mice was not different from binding to these tissues in untreated mice. Total serum immunoglobulin titers correlated with binding of yeast cells to mouse spleen. Blocking studies ruled out a mannosyl-fucosyl receptor-mediated binding. These results suggest that ex vivo adherence of C. albicans represents a host immune defense mechanism by which the immunocompetent host binds blood-borne yeast cells to host immune cells in reticuloendothelial organs to prevent dissemination to other organs.


Cytokine gene expression of endothelial cells infected with Trypanosoma cruzi.

Coronary microvascular spasm and platelet hyperreactivity have been implicated in the pathogenesis of Chagas' cardiomyopathy. To clarify further the role of the microvasculature in this disease, alterations in cytokine gene expression due to Trypanosoma cruzi infection of human umbilical vein endothelial cells were examined. Northern blot analysis of total RNA from endothelial cells demonstrated that interleukin (IL)-1 beta, IL-6, and colony-stimulating factor 1 (CSF-1) mRNA expression was absent or minimal in uninfected cells but significantly increased in infected cells. c-sis mRNA levels diminished with increased time of infection. In situ hybridization studies also demonstrated high levels of IL-6 mRNA in individual infected cells. Significant levels of IL-6 and IL-1 beta protein were detected in the supernatants of infected endothelial cells. The serum of an acutely infected individual contained high levels of IL-6 protein, suggesting the potential importance of cytokines secreted by the vascular endothelium in the pathogenesis of Chagas' cardiomyopathy.


Monitoring of antibodies against human immunodeficiency virus type 1 p25 core protein as prognostic marker.

Anti-p25 antibodies were evaluated by cross-sectional analysis of sera from 130 human immunodeficiency virus type 1-infected patients and in a longitudinal study of 56 patients by retrospective analysis of sequentially collected sera. High and stable antibody levels were found in Centers for Disease Control stage II or III patients, 78% of whom had levels greater than 10 arbitrary units/mL. Patients with AIDS-related complex displayed heterogeneous levels. Patients with AIDS had the lowest values: less than or equal to 10 units/mL in 96% of cases. In patients whose CD4+ cell counts eventually fell below 200/mm3 or who developed AIDS (or both), antibodies were initially less than 40 units/mL and/or they declined with a rate greater than 1 log unit/5 years, beginning at least 4 years before the index symptom. Because the only point at which CD4+ cell counts significantly differed between progressors and nonprogressors was 1 year before the disease, both initial anti-p25 values and antibody decline seemed to be better long-term prognostic markers than CD4+ cell counts.


Early diagnosis of human immunodeficiency virus infection in children less than 6 months of age: comparison of polymerase chain reaction, culture, and plasma antigen capture techniques.

Three techniques were evaluated for their ability to detect human immunodeficiency virus (HIV) in infants from birth to 6 months of age. Polymerase chain reaction (PCR) and HIV cocultivation were of comparable sensitivity, detecting 90% of all positive specimens. Both techniques found positive results in approximately 5% of samples from seroreverting children. Both assays detected HIV in only half of infected newborns, suggesting that this fraction of children was infected during gestation. Plasma p24 antigen was detected in three-fourths of all samples tested but in only half of infected children during the first 2 months of life and 88% of samples from children during the next 4 months. The specificity of p24 antigen detection was 100%.


Concordance of human immunodeficiency virus detection by polymerase chain reaction and by serologic assays in a Dutch cohort of seronegative homosexual men.

In three subgroups of a clinically and socially well defined group of Dutch homosexual men, the prevalence of human immunodeficiency virus type 1 (HIV-1) sequences in seronegative blood samples was studied using the polymerase chain reaction (PCR). In 19 seronegative partners of seropositive persons, no HIV-1 sequences were found by PCR in either early (1984/1985) or more recent (1987) samples. In 42 seronegative persons selected by their high risk for HIV-1 infection, none harbored HIV-1 sequences in either early (1985/1986) or late (1989) samples. In 15 people who seroconverted for HIV-1, only 2 samples collected 3 months before seroconversion were PCR-positive. These persons were also HIV antigen-positive at this time. These data suggest that a latent infection greater than 6 months does not occur and that the combination of HIV antibody and HIV antigen tests is appropriate and conclusive in most cases of HIV-1 infection.


Immunosuppression with cyclosporine during the incubation period of experimental woodchuck hepatitis virus infection increases the frequency of chronic infection in adult woodchucks.

The immunosuppressive agent cyclosporine was given to adult woodchucks during acute experimental infection with woodchuck hepatitis virus (WHV). All 17 woodchucks given WHV alone or with a vehicle resolved the infection (i.e., zero chronicity), but when cyclosporine was given throughout the incubation and acute phases of infection (0-12 or 14 weeks; n = 12), the rate of chronic infection increased to 92%. When cyclosporine was given only during the incubation period (0-4 weeks; n = 10) or only during the acute phase of infection (2-12 weeks; n = 9), the rates increased to 50% and 55%, respectively. However, when the drug was given after the acute phase (8-18 weeks; n = 9), the chronic infection rate (11%) did not differ from that in untreated and vehicle controls. Immune responses inhibited by cyclosporine are important in resolution of acute WHV infection and occur mainly during the first 8 weeks. Immunosuppression of these responses for even short intervals during incubation (e.g., 0-4 weeks) increases the risk of chronicity.


Role of cell wall polysaccharide in the assessment of IgG antibodies to the capsular polysaccharides of Streptococcus pneumoniae in childhood.

The interference of antibodies to pneumococcal cell wall polysaccharide (CWPS) in the measurement of antibodies to capsular polysaccharides in children was assessed after vaccination with pneumococcal polysaccharide vaccine. ELISAs were developed to measure IgG subclasses specific for pneumococcal types 3, 6, 19, and 23 and CWPS. Analysis of antibody levels to all four capsular polysaccharides was affected by the presence of antibodies to CWPS, and their removal altered both anti-capsular polysaccharide antibody levels and the interpretation of responses to the vaccine. Thus, it is likely that CWPS contaminating pure capsular polysaccharide reagents used in most standard immunoassays is responsible for falsely elevated measurements of antibodies to capsular polysaccharide and the incorrect assessment of anti-pneumococcal antibody status in childhood.


Neonatal mouse model of group B streptococcal infection.

Neonatal mice were infected with type III group B streptococcal (GBS) strain M781 by the intraperitoneal route. Age-related susceptibility to challenge was seen within the first 5 days of life. Quantitative blood cultures demonstrated a rapid increase in bacterial numbers during the first 30 h after challenge. Infected pups showed clinical signs of septicemia, and most succumbed within 48 h of challenge. Histopathologic evaluation of the neonates showed bacterial infection within 1 day after challenge. Pregnant adult mice were given a single inoculation of serum raised in rabbits against a tetanus toxoid-conjugated type III GBS polysaccharide vaccine. This serum passively protected 100% of the offspring. This neonatal mouse model of GBS infection and protection may be suitable for study of various forms of intervention.


Salmonella typhimurium bacteremia: association with the virulence plasmid.

Virulence plasmids, which are found in a small number of Salmonella serotypes, greatly enhance the extraintestinal growth of salmonellae and lower the LD50 by 2-5 logs in experimental murine infections. To determine if virulence plasmids are important in the pathogenesis of Salmonella bacteremia in humans, blood and fecal isolates of Salmonella typhimurium from California were examined for the presence of a virulence plasmid. Colony blots were done using a labeled probe made from the highly conserved EcoRI fragment of the Salmonella dublin virulence plasmid. A total of 42% of the fecal and 76% of the blood isolates hybridized with the probe. This is the first evidence that the virulence plasmid is important in the pathogenesis of Salmonella bacteremia in humans.


Anti-tetanus toxoid antibodies in intravenous gamma globulin: an alternative to tetanus immune globulin.

The levels of anti-tetanus toxoid IgG antibodies were measured in 29 lots of intravenous gamma globulin (IVIG). The antibody levels varied from 4 to 90 IU/mL (geometric mean, 18.6; 90% confidence interval, 9.7-35.7). The variation from manufacturer to manufacturer accounted for most of the observed differences among lots; there was relatively little variability among multiple lots from a single manufacturer. IVIG may be an acceptable alternative to horse or human tetanus immune globulin.


Seroepidemiology of Chlamydia pneumoniae TWAR infection in Seattle families, 1966-1979.

Incidence rates of Chlamydia pneumoniae infection and information on reinfection and transmission within families were obtained by serologic study of serum samples from prospective family studies conducted 1966-1979. Specimens (n = 3671) from 343 subjects in 68 families were tested for TWAR antibody using the microimmunofluorescence assay. Acute infection was defined as a fourfold rise in antibody titer between consecutive specimens. Sixty-four episodes of infection were identified in 58 persons; 4 had 2 infections and 1 had 3. From late 1975 until early 1979, when 3 serum specimens were collected yearly, rates of infection by age groups 0-4, 5-9, 10-14, 15-19, and greater than or equal to 20 years were 0, 9.2, 6.2, 2.2, and 1.5/100 person-years, respectively. Reinfections, defined as infections in persons with previous antibody, constituted most acute infections among adults. Acute infections more often affected a single family member than multiple members, but 2 or 3 family members were infected during the same period 12 times.


Compartmentalization of lipopolysaccharide production correlates with clinical presentation in meningococcal disease.

Lipopolysaccharide (LPS, endotoxin) was quantified in plasma and cerebrospinal fluid (CSF) collected simultaneously from patients with systemic meningococcal disease. High levels (median, 3800 ng/L; range, 750-14,000) were present in plasma and low levels (median, 40 ng/L; range, less than 25-165) in CSF of patients with fulminant septicemia. Conversely, high levels (median, 2500 ng/L; range, less than 25-500,000) in CSF and low or undetectable levels (median, less than 25 ng/L; range, less than 25-210) in plasma were associated with meningitis without septic shock. Levels of LPS were significantly correlated with protein levels in CSF (r = .50, P = .01) and inversely correlated with the ratio of glucose in CSF to that in blood (r = -.62, P = .0005). LPS level in CSF greater than 800 ng/L was significantly associated with greater than or equal to 10(9) leukocytes/L, protein levels greater than 0.5 g/L, and a glucose ratio less than 0.5. Thus, quantification of LPS levels in the plasma and CSF in systemic meningococcal disease is a better predictor of pathophysiologic events than is demonstrating the presence of live bacteria as in conventional culture.


Comparison of urine and serum concentrations of interleukin-6 in women with acute pyelonephritis or asymptomatic bacteriuria.

The mucosal and systemic interleukin-6 (IL-6) response to urinary tract infection was analyzed in women with acute pyelonephritis or asymptomatic bacteriuria. Urine and serum samples were obtained at diagnosis and after treatment. IL-6 activity was elevated in urine samples from most bacteriuric women, regardless of the severity of infection. Urinary levels greater than 20 units/mL occurred in 25 of 29 women with acute pyelonephritis and in 36 of 42 women with asymptomatic bacteriuria. Elevated serum IL-6 levels were found mainly in patients with acute pyelonephritis: Levels greater than 20 units/mL occurred in 14 of 28 women with acute pyelonephritis compared with 0 of 28 women with asymptomatic bacteriuria. These results suggest that bacteriuria is accompanied by elevated urinary IL-6 levels and that this IL-6 is locally produced. The spread of IL-6 to the circulation in patients with acute pyelonephritis may contribute to the elevation of fever and C-reactive protein characteristic of the disease.


Absence of intrathecal synthesis of some interferon-alpha subtypes in bacterial meningitis.

Sixty-five cerebrospinal fluid (CSF) samples from 54 infants and children with bacterial meningitis were analyzed for the presence of interferon (IFN)-alpha with a biologic assay. Of the 65 samples, 3 were positive (2-4 IU/mL) and only 1 of 56 collected early was at 4 IU/mL. These results suggest that some subtypes of IFN-alpha already reported as present in viral infections of the central nervous system are not detected in bacterial meningitis by our IFN assay. This difference may be helpful in differentiating bacterial from viral infections and also in evaluating the quality of the virologic investigation; moreover, the rarity of IFN-gamma in CSF in bacterial meningitis needs further investigation to understand its role in the pathogenesis of the disease.


Diagnosis of Mediterranean spotted fever by indirect immunofluorescence of Rickettsia conorii in circulating endothelial cells isolated with monoclonal antibody-coated immunomagnetic beads.

Rickettsia conorii, an obligate intracellular bacterium that infects vascular endothelial cells, is the etiologic agent of Mediterranean spotted fever (MSF). A new procedure using indirect immunofluorescence was used to directly detect R. conorii in circulating endothelial cells (CEC). CEC were separated from other blood components by using anti-endothelial cell monoclonal antibody-coated magnetic beads. An anti-R. conorii polyclonal rabbit antiserum was used to stain rickettsiae. The entire procedure took 3 h. R. conorii was detected in CEC from 9 of 12 patients ultimately confirmed as having MSF. Among the patients, 5 with R. conorii isolated by centrifugation-shell vial assay were also positive by the new technique. None of 3 patients whose diagnosis was other than MSF had R. conorii detected in CEC. The procedure may be helpful for rapid diagnosis of MSF and may lead to new technical approaches for the diagnosis of infectious diseases caused by intracellular pathogenic microorganisms.


Cytokine-induced human multinucleated giant cells have enhanced candidacidal activity and oxidative capacity compared with macrophages.

The granulomatous response to infection is characterized by formation of multinucleated giant cells (MGC). A model has been developed for the study of MGC using fresh human peripheral blood monocytes cultured in medium supplemented with autologous serum and a combination of recombinant human interferon-gamma and interleukin-3 (100 units/mL each). Differential Giemsa staining demonstrated a 53% increase in candidacidal activity of MGC (35.1% +/- 2.0% of ingested organisms were killed by MGC) compared with identically cultured mononuclear macrophages (which killed 22.9% +/- 1.8% of organisms ingested; P less than .05). There was no significant difference in the number of organisms ingested. MGC stimulated with phorbol myristate acetate produced 2.2 times as much superoxide anion per unit of cytoplasmic protein as identically cultured and stimulated macrophages (34.3 vs. 16.2 nmol of superoxide/microgram of cell protein; P less than .01). This was corroborated with single-cell measurements of oxidative activity using digital image analysis. These observations support the hypothesis that MGC have an advantage in microbicidal activity over macrophages that may be due, at least in part, to enhanced oxidative capacity.


Restaurant-associated outbreak of giardiasis.

An outbreak of giardiasis occurred among staff of a job training center after a meeting at a restaurant. Twenty-seven (75%) of 36 attendees became ill compared with 1 (3%) of 31 staff members not attending (relative risk, 23.3; 95% confidence interval, 3.4-161.4). Because most attendees ate all items on a fixed menu, no individual food item could be conclusively associated with illness. Circumstantial evidence suggests, however, that ice contaminated by a food handler may have been the vehicle. The restaurant had multiple sanitary violations, and 2 employees, 1 with asymptomatic giardiasis and the other with a Giardia-infected, diapered child, served ice to the attendees. This outbreak demonstrates the potential for restaurant-based giardiasis outbreaks.


Fine structure of spermiogenesis in the coccid insect Aspidiotus perniciosus: late stages of differentiation and structure of the mature sperm bundles.

Spermatids of A. perniciousus are organized into bundles each containing 32 cells. During the process of differentiation, nuclear elongation and chromatin condensation take place. The mature sperm bundles are finally surrounded by the secondary sheath which is composed of a trilaminar membrane. The mature sperm is not differentiated into head, mid piece or tail but has two tapering ends and an electron dense central core. Centrioles, the nuclear membrane, mitochondria and a flagellum appear to be lacking. Microtubules are arranged in a spiral pattern forming the enveloping microtubular sheath.


Ultrastructure of symbiont-like microorganisms associated with the sperm of ixodid ticks.

The endospermatophores found in the seminal receptacles of three species of Ixodid ticks: Hyalomma marginatum, H. dromedarii and Amblyomma hebraeum were examined. The seminal receptacles showed the presence of symbiont-like microorganisms in association with the sperm cells. These yeast-like microorganisms represent extracellular symbionts, produced by the male and introduced in the genital system of the female through copulation. The symbionts frequently occur freely in large agglomerations surrounded by a large amount of alpha-glycogen rosettes. They do not alter their shape nor do they attach to the sperm cells. Another kind of Rickettsia-like microorganisms is present in tissues of some tick organs. This could be a form of disease agents or naturally occurring microorganisms in the tissue of tick organs.


Intestinal parasitic infection among sewage workers in Alexandria, Egypt.

This research was conducted to study the risk of intestinal parasitic infections among sewage workers. A hundred sewage workers in Alexandria and another hundred as control group not exposed to sewage were selected for this research. The results indicated that sewage workers were at a higher risk of developing intestinal parasitic infection especially Ascariasis and Amoebiasis. Nail examination revealed that sewage workers were at a higher risk of infection by Entrobius vermicularis.


Profile of portal hypertension in hepatosplenic schistosomiasis.

Intrasplenic pressure changes versus ova count in stool of schistosomal portal hypertension in cases of bilharzial hepatic fibrosis were studied before and after 40 mg/kg body weight praziquantel therapy. Praziquantel acts on the vascular level in which it decreases the portal hypertension caused by bilharziasis as detected by decreased size of portal and splenic veins diameter as well as a significant decrease of portal pressure by the decrease of intrasplenic pressure (P less than 0.05). On the other hand, praziquantel decrease bilharzial stool egg count after 3 months of therapy (P less than 0.05).


Cutaneous leishmaniasis in outpatients of dermatology and venereology clinic, Benha Faculty of Medicine, Zagazig University, Egypt.

Zoonotic cutaneous leishmaniasis (ZCL) is a disease of an ancient history and more or less encountered in several Egyptian Governorates. However, no data is available concerning CL. in Qualyobia Governorate. In the present study, a number of 10920 patients who attended the outpatient's clinic of Dermatology and Venereology, during the year 1991, were clinically examined for any manifestations suggesting CL. Lesion when present was examined by Giemsa stained smear and N.N.N. medium culture. Serum samples were separated from blood of suspected CL. cases for detection of antileishmanial antibodies by the indirect haemagglutination tests. The results showed that six patients had parasitologically proven lesions and that IHAT was less specific in diagnosing CL. It was concluded that CL exists as endogenous cases in Qualyobia Governorate and that the presence of CL and VL within nearly the same focus is a point of medical importance.


Alternate chemotherapy in experimental schistosomiasis.

Schistosoma mansoni infected mice were treated with a single oral dose of either praziquantel (400 mg/kg) or oxamniquine (100 mg/kg). The uncured mice, after the first dose, were given a second dose of the same drug or alternatively with the other drug. The drug efficacy was monitored by stool examination and egg count using Kato-thick smear technique. Praziquantel was found to be more effective, it gave 60% cure rate after the first dose, increased to 100% after the second dose, than oxamniquine which had 37% and 74% cure rates respectively. Furthermore, in uncured mice praziquantel resulted in more egg count reduction than oxamniquine. Treatment with the alternate drug of mice not cured with the first treatment gave 100% cure rate with praziquantel and 83% with oxamniquine. It was concluded that praziquantel is generally more effective than oxamniquine and cases not cured with a single dose of either drugs may be effectively treated with a second dose of the other.


Effect of Crytocotyle lingua on the ionic concentration of Littorina obstusata (L.).

The concentration of Sodium, Potassium and Calcium ions (the most important inorganic ions in the body fluid) were determined in both parasitized and uninfected Littorina obtusata (L.). The ionic concentrations were determined using a Pye Unicam SP atomic absorption spectrophotometer. Sodium and calcium were determined using the absorption mode while, potassium concentration was measured in the emission mode using special Potassium filter. The results reveals, no significant difference in the concentration of Na+, K+ and Ca++ between parasitized and uninfected L. obtusata. It seems that although the presence of the parasite within the molluscan hosts caused dramatic effects in the tissue and physiology of the snail, the blood ionic concentration of parasitized snail remains isosmotic to that of the surrounding medium.


Myopathic changes in schistosomal hepatosplenomegaly; histopathological, histochemical and immunopathological studies.

Eighteen patients suffering mainly from schistosomal hepatosplenomegaly (B.H.S.M.) were selected & classified into 3 groups according to the degree of B.H.S.M. Muscle biopsy for histopathological, histochemical and immunopathological studies were taken. The results showed that fatiguability as a symptom was present in all cases but motor weakness was present in four cases. Twelve cases had peripheral neuropathy in the form of glove and stocking hypothesia, while pyramidal tract lesion was present in four cases. Two cases, however, showed both peripheral neuropathy and pyramidal tract lesions. Histopathological and histochemical assessment revealed frank myopathic changes in 14 cases while only one case showed definite neuropathic changes. The remainder cases were inconclusive. Immunopathological examination revealed marked deposition of IgG in 50% of cases, while IgM and IgA showed mild to moderate reactions. It was concluded that muscular changes in schistosomal patients are myopathic in nature and that immunological mechanisms could be considered as a factor in the pathogenesis of this muscular disorder.


The efficacy of IHAT, IFAT and Dot-ELISA in sero-diagnosis of toxoplasmosis in complicated pregnancies.

Studying of 72 pregnant women having congenital anomalies, stillbirth and repeated abortion of first and second trimesters showed Toxoplasma antibodies with high titre by IFAT and Dot-ELISA. Analysis of the results showed a positive correlation between IFAT and Dot-ELISA but not with IHAT. However, Dot-ELISA is simple, sensitive and specific.


Scanning electron microscopy of the small intestine of rats experimentally infected with Cynodiplostomum azimi.

Scanning electron micrographs of the proximal intestine of rats infected with C. azimi showed mild villous changes and excessive mucus secretion as early as the first day after infection. On the second day the regular leaf-like pattern of the villi was not seen, the epithelial lining of the mucosa was damaged with large amounts of mucus. Goblet cell openings appeared either empty or filled with secretions. The mucosal damage persisted for three months. Four months after infection, villi regained part of their normal pattern. Their epithelial lining although less damaged, appeared delicate. Villi away from the worm were less affected. Pathological changes at different intervals of infection were discussed in relation to the surface structure of the parasite.


Leishmania infection sought in rodents in north Sinai, Egypt.

Zoonotic cutaneous leishmaniasis (ZCL) is endemic in certain foci in North Sinai. In the present study, eight species of commensal and wild rodents were trapped from seven areas. Suggestive ZCL lesions were seen on R. rattus, R. norvegicus, G. pyramidum, G. ander soni, G. gerbillus and Meriones sacramenti. Positive Giemsa stained smears were identified in G. pyramidum. The results were discussed. It was concluded that the ZCL transmission cycle, animal reservoirs, sand fly vector and man, is documented.


Immune response of gametocytes of Plasmodium falciparum in exogenous cases in Dakahlia Governorate, Egypt.

Following chloroquine treatment gametocytes numbers fell with a constant rate of loss over a period from 15-21 days. The gametocytes had a half-life of 2-4 days. The sex ratio was constant 4 female: 1 male. Antibodies to the gametocytes were detected in some patients. It was concluded that the gametocytes of P. falciparum are long-lived and can stimulate antibody production but the immune response appears to play no part in their elimination, which probably takes place in the spleen as a part of the normal process of removing old, damaged and malformed red cells.


A freeze etching study on the distribution of nuclear pores during spermatogenesis in ticks.

In this study the freeze-fracturing and freeze-etching techniques was used to study pore distribution during spermatogenesis in ticks. Three species of Ixodid ticks were studied: Hyalomma dromedarii, H. marginatum and Amblyomma hebraeum. In all three species of the ticks examined, the nuclei of the early stages of male germ cells showed varying degrees of aggregation into pore-rich and pore-poor areas. Spermatocytes demonstrated significant pore clustering in pore-rich areas, leaving areas almost devoid of pores. Spermatids on the other hand displayed a random distribution of nuclear pores. This random distribution was soon modified in the course of nuclear differentiation. Nuclear pores disappeared in the nuclei of mature spermatozoa of the ticks examined.


Tonsilar toxoplasmosis.

One hundred cases were suffering from chronic tonsillitis, aged between 4-12 years, of both sexes (60 males and 40 females), from outpatient clinic of E.N.T. unit at Benha Faculty of Medicine. They were examined serologically by indirect haemagglutination--antibody technique (IHAT) for toxoplasmosis. After tonsillectomy, histopathological and electronmicroscopical studies were done. Ten percent of cases were positive serologically, among males (11.71%) and females (7.5%). The positivity rate was high among age group 10-12 years than others. Two cases showed histopathological and ultrastructural changes indicating toxoplasmosis.


Heterophyids: experimental chemotherapy and immunotherapy.

Heavy infections with the tiny flukes heterophyids can cause intestinal pain and mucous diarrhea, thus the study of praziquantel (Pzq) and the new drug cyclosporin A (CsA) as antiparasitic drugs were undertaken in this work, as well as the effect of matecercarial and adult antigens as immunizing agents. To assess the result of our work, the number and length of the recovered heterophyids were studied. Description of the surface by SEM was carried out for the groups: treated by cyclosporin A and immunized by adult heterophyid antigen (third fraction) which gave the highest percentage reduction.


A contribution to the anatomy and histology of the female reproductive system of Amblyomma cajennense (Acarina: Ixodidae).

The genital system of the tick Amblyomma cajennense is composed of several parts: a single tubular ovary where a great number of ova develop except for a longitudinal groove on its dorsal surface, with feeding the ova develop multiplying the size and weight of the ovary, as they reach maturity they are ovulated into the lumen. Two oviducts are connected to the ovary on each side, these thicken at a certain region forming the knee shaped ampulla. This significant region is where great numbers of mature spermatozoa are found. The vagina is composed of a vestibular and a cervical part and leads to the exterior by the genital aperture. A large seminal receptacle opens into the cervical vagina, the function of the receptable is to receive and store endospermatophores.


A general survey of ciliates associated with Periplaneta americana (L.) in the Central Region of Saudi Arabia.

The aim of this work is to identify and classify different species of ciliates which live in the alimentary canal of Periplaneta americana (L.) in the Central Region of Saudi Arabia. This insect was chosen because of the direct and indirect relationship to man. Identification of ciliates that live in the alimentary canal of this insect may be useful in the future for eradication of insects probably by making an epidemic among such insects.


Seroepidemiology of Toxoplasma infection in Riyadh, Saudi Arabia.

A total of 362 apparently healthy Saudis of both sexes living in Riyadh and aged 4-60 years, were investigated for Toxoplasma antibodies using the indirect haemagglutination test (IHT). Eighty one (22.4%) were positive with titres of 1/64 or greater. A significantly higher prevalence of Toxoplasma antibodies was correlated with age and association with cats. Economic status showed an apparent, but not significant, negative correlation with antibody prevalence rates. No significant association was found between Toxoplasma infection and sex, consumption of raw or undercooked meat, and urban or rural residence. The results provide an evidence that cats may be the main source of human infection with Toxoplasma in Riyadh.


Gastro-vascular and micro-vascular changes in chronic murine schistosomiasis mansoni-response to propranolol.

Histopathologic study of the gastric wall of mice infected with Schistosoma mansoni was carried out at 8, 12, 16 and 20 weeks post-infection (p.i.). The gastric vascular structures were more sensitive to injury with the progression of liver disease than the glandular cells, with marked submucosal edema and thickening of submucosal and subserosal vessels. By transmission electron microscopy, the mucosal microvessels had thickened basement membrane and conspicuous endothelial cells with granular cytoplasm thrown into numerous intraluminal microprojections and with increased number and size of pinocytotic vesicles. When propranolol was administered orally as 20 mg/kg daily doses for two weeks at 6, 10, 14 and 18 weeks p.i., a striking regression of gastro vascular and micro-vascular changes has been observed, particularly with early drug therapy. Based on the present study, it can be concluded that "structural autoregulation" of gastric vasculature to the emerging portal hypertension in chronic murine schistosomiasis mansoni, is a dynamic process that could be reversed by propranolol therapy. This response could, possibly, improve gastric mucosal perfusion and functional abnormalities which may predispose the gastric mucosa to severe damage or hemorrhage.


Effect of Capillaria hepatica infection on Schistosoma mansoni challenge in mice.

The combined infection between Capillaria hepatica and Schistosoma mansoni was studied. The results of this work revealed that C. hepatica infection induced significant reduction of S. mansoni worm load in the two groups infected with C. hepatica and challenged with S. mansoni either during worm maturation period of C. hepatica, or at the time of presence of C. hepatica eggs in the liver Reduction in total and tissue egg count was also reported, but eggs excreted in the stool showed no difference in count from that of S. mansoni-infected controls. Oogram pattern of the experimental groups revealed a higher percentage of dead eggs and absence of mature and some developing stages. Scanning electron microscopy demonstrated severe destruction of adult worm of both groups. All these data showed the vigorous destructive effect of C. hepatica infection on the challenged S. mansoni.


A brief note on phlebotomine sandflies in Riyadh, Saudi Arabia.

A total of 294 specimens were collected and identified as three species of the genus Phlebotomus and four species of the genus Sergentomyia. Of interest was the demonstration of promastigotes in the foregut of two females P. papatasi.


Identification of the Schistosoma haematobium soluble egg antigens inducing antibody production and/or T cell proliferation in humans.

Soluble antigens were prepared from Schistosoma haematobium eggs collected from urine of 6-16 year-old children with urinary schistosomiasis. The electrophoretic profile of the soluble egg antigen (SEAH) preparation was almost identical to that (SEAh) obtained from UNDP/World Bank/WHO, Switzerland and prepared from S. haematobium eggs retrieved from intestines of infected hamsters. Reactivity of 50 individual patients with S. haematobium in Western blots led to the identification of the SEA protein bands carrying human B cell epitopes. Some, but not all, of these SEA proteins initiated peripheral blood T lymphocyte proliferation in T cell Western assays. These antigens are probably the ones inducing granulomatous response in vivo, and that are responsible for the immunopathology of the disease.


Some ultrastructural studies on the excretory bladder of the adult Cynodiplostomum azimi (Trematoda: Digenea) in infected rats.

Transmission electron microscope was used to reveal the reserve bladder system of the adult Cynodiplostomum azimi in experimentally infected rats. It was shown that this system consisted of a number of lacunae. The lacunar lining appeared as syncytial epithelium containing many nuclei, mitochondria, dense secretory bodies, Golgi complex and bundles of smooth muscles. The outer surface of the excretory epithelium was highly folded and lamellated. These lamellae were continuous with the excretory epithelium. Small lipid droplets were observed within the excretory epithelium, while larger ones were associated with the lamellae. The large lipid droplets were released in the lacunal lumen after the rupture of the lamellae surrounding them.


Melanization response of two mosquito species against Wuchereria bancrofti.

The melanization response of two mosquito against Wuchereria bancrofti was studied under different conditions of temperatures, adult age and feeding regimes. The two mosquito species were Culex pipiens, a susceptible species for W. bancrofti transmission and Aedes Caspius, a refractory one.


Ultrastructural aspects of intramolluscan developing cercariae of Echinostoma liei.

The ultrastructure of the daughter redia of Echinostoma liei inside their host Biomphalaria alexandrina is described. The tegument consists of one layer, bearing numerous microvilli on its outer surface. Besides, the description of the ultrastructure of the developing cercariae within the daughter rediae is also included in the present study. The young cercariae are covered with a thin nucleated primitive temporary epithelium, which is lost when the true tegument is formed beneath it. The mechanism of spine formation was not apparent, but there is some indication that the spines are associated with a thickened part of the basal membrane of the tegument. Section through these cercariae showed that the tissues lying below the tegument are disposed in consecutive layers, suggestively representing developing muscle tissues.


The effect of cyclosporin A (CSA) on murine visceral toxocariasis canis.

Swiss mice experimentally infected with T. canis were treated by Cyclosporin A (CSA). The direct antiparasitic effect was noticed when administered on the same day of infection and was more effective when repeated for eight weeks. Although immunosuppression of CSA increased the susceptibility of mice to T. canis larvae, its direct antiparasitic effect was more effective and led to diminution of living T. canis larvae. Cellular immunity was inhibited as revealed by diminution of T. lymphocytes by decreased lymphocytes migration inhibition percentage on repeated administration of CSA for four weeks.


Cyclosporin A in experimental trichinosis: histopathological study.

Cyclosporin A (CsA), a relatively new drug, was previously used in a series of studies on experimental trichinosis. This work was conducted to study the histopathological changes of CsA treatment at different time intervals from T. spiralis infection in mice. The work included study of the effect of the drug on the larvae treated either in vivo or in vitro. The drug was most effective when it was administered on the same day of infection, then when given one week before. The effect was evidenced by diminution both in the number of larvae and in the cellular reactions.


Evaluation of a direct agglutination test and methylene blue dye test in diagnosis of cutaneous leishmaniasis.

The usefulness and sensitivity of a direct agglutination test (DAT) in the diagnosis of cutaneous leishmaniasis infection has been investigated. Trypsin treated, formalin fixed and coomassie blue stained Leishmania promastigotes were used as antigens: L. infantum, L. donovani, L. aethiopica. Although the titres of sera from patients with localized cutaneous leishmaniasis were low, sera from lepromatous, tuberculous and toxoplasmic patients gave high titres indicating cross reactivity. Comparable results were obtained when the same sera were tested using freshly prepared antigen or antigens stored for 5 months at 4 degrees C, and with addition of 0.78% 2-mercaptoethanol to the diluent.


Identification and characterization of specific hydatid antigen fraction(s).

A specific hydatid antigen was prepared in this study from Echinococcus granulosus cyst in livers and lungs of camels. Elimination of host "camel" protein from crude hydatid fluid was achieved by two methods: Salting out using ammonium sulfate precipitation method and immunoaffinity purification using coupled anticamel antibody to cyanogenbromide activated sepharose 4B gel. Testing the prepared hydatid antigen against anticamel serum, using immunodiffusion method, indicated that the affinity purified hydatid antigen was almost completely purified from camel protein. Characterization of the affinity purified hydatid antigen, using immunoelectrophoresis, showed positive arc 5 precipitation when tested against known positive antihydatid sera. Further characterization with gradient gel electrophoresis, showed with silver stain that the dominant and most consistently demonstrable proteins occurred as a complex in the 52/62 KDa region. Strong reaction with the 52/62 KDa complex was consistently observed when the affinity purified hydatid antigen was probed with known positive reference antihydatid sera. The identified hydatid antigen fraction(s) with 52/62 KDa complex can provide promising non-invasive parameter for diagnosis of Hydatidosis.


Effect of larval population density on dynamics of growth in Chrysomyia albiceps (Wiedemann). I. Effect of feeding behaviour and growth rate of larvae.

The feeding behaviour of uncrowded and crowded larvae differed as soon as feeding began. In case of high density populations, larvae tend to form dense clusters and this gregariousness is responsible for much of the injury and resulting predation. Larval growth rate had the same pattern up to 200 larvae per 20 grams meat, then this pattern greatly changed with the increase of densities. Larval survivorship remained high at densities up to 160 larvae per 20 gm meat, then decreased rapidly as larval density increased. The duration of larval development was extended and synchronization of larval pupal ecdysis was affected. Percentage of pupation and pupal weight were greatly decreased at high densities.


Lymphatic leishmaniasis. An adult case without visceral involvement from Sharkia Governorate, Egypt.

Lymphatic leishmaniasis was reported as an indigenous infection in a 30 year old male from Abo Hamad, Sharkia, Egypt. Cervical lymphadenopathy was the only clinical sign and no visceral involvement. Diagnosis was based on histopathological examination of needle biopsied material. The patient was successfully treated with sodium stibogluconate as shown clinically and parasitologically. It was concluded that lymphatic leishmaniasis should be in mind when dealing with lymphanopathy of unknown aetiology.