Inactivation of the human immunodeficiency virus type 1 (HIV-1) by ultraviolet and X irradiation. Here we report the kinetics of inactivation of HIV-1 by ultraviolet (UV) or X irradiation. Inactivation of HIV-1 by UV irradiation followed quasi first-order, i.e., single-hit, kinetics. The LD37 for inactivation of syncytia formation in SupT1 cells by limiting dilutions was approximately 780 J/m2. The LD37 for inactivation of HIV-1-induced syncytia by UV irradiation was nearly identical when measured in UV repair-proficient and -deficient lymphoblastoid cell lines (LCLs), demonstrating that immediate host cell reactivation (repair) of UV damage in the HIV-1 genome does not occur. The ability of HIV-1 to induce the accumulation of reverse transcriptase activity showed similar dose-dependent inhibition by UV irradiation (LD37, 845 J/m2). Inactivation of HIV-1-induced syncytia formation by X rays also approached first-order kinetics. The LD37 for syncytia formation as measured by limiting dilutions was approximately 3 x 10(3) Gy. HIV-1-induced accumulation of reverse transcriptase activity was slightly more resistant to inactivation by X rays, with an LD37 of approximately 4.5 x 10(3) Gy. Syncytia-forming ability was still present in HIV-1 preparations X-irradiated with 1.6 x 10(4) Gy. For the first time, we utilized the polymerase chain reaction (PCR) to measure the effects of radiation on virus replication. A decrease in the presence of the HIV-1 DNA could be detected by PCR in PBL cultures infected with UV- and X-irradiated virus. It required 12.96 J/m2 to eliminate the signal specific for HIV-1 DNA completely. The ability of HIV-1 to establish long-term infection in LCLs was also resistant to UV and X irradiation. Only linear and circular forms of HIV-1 DNA could be detected in LCLs established from PBL cultures infected with UV-irradiated virus. The significance of the relative resistance of HIV-1 to inactivation by UV and X irradiation is discussed.


Disturbances in acute phase plasma proteins during melancholia: additional evidence for the presence of an inflammatory process during that illness. 1. Leukocyte enumeration through flow cytometry has revealed that severe depression may be accompanied by a systemic immune activation, indicative of an inflammatory response. The latter condition allegedly involves an important modification of acute phase plasma protein (APP) equilibrium. 2. In order to elucidate whether the state of severe depression is represented by alterations in APPs, the authors measured: alpha 1 antitrypsin (alpha 1 AT), alpha 2 macroglobulin (alpha 2 M), haptoglobin (Hp), alpha 1 acid glycoprotein (alpha 1 S), transferrin (Tf), complement component 4 (C4) and C-reactive protein (CRP). Interleukin-1-beta (II-1 beta) and interleukin-6 (II-6) circulating levels were determined. 3. Hyperhaptoglobinemia and hypotransferrinemia are hallmarks for major depression and depression per se, respectively. The disorders in Hp and Tf circulating levels are highly sensitive to (83%) and specific for (100%) melancholia as opposed to the healthy state. 4. Disorders in both APPs are significantly related to the absolute number of blood monocytes. 5. The authors observed a trend towards lower alpha 2M and higher alpha 1S values in severely depressed subjects. Severity of depression was significantly related to Hp and alpha 1S (both positively) and to alpha 2M and Tf (both negatively) values. 6. No significant intercategory differences in C4 could be established, whilst only a few subjects exhibited measurable CRP, II-1 beta and II-6 circulating levels. 7. Our findings may support the hypothesis that depression is accompanied by an inflammatory response.


Serum pre-S1 and pre-S2 antigens as prognostic markers in interferon therapy for chronic hepatitis B. We investigated the changes of serum pre-S1 and pre-S2 antigens before and after interferon therapy in 35 carriers with HBeAg, to examine their clinical significance in the therapy. Both antigens were measured quantitatively by solid-phase enzyme immunoassays using monoclonal antibodies specific to each antigen. The titers of these antigens of responders before therapy were significantly lower (p less than 0.001) than those of non-responders (6.23 +/- 1.09 versus 8.65 +/- 2.09 and 4.46 +/- 1.27 versus 6.50 +/- 1.50, respectively). The titers decreased significantly 6 months after interferon therapy in the responders (p less than 0.05), whereas they did not change in the non-responders. Our findings indicate that pre-S1 and pre-S2 antigens have a close correlation with hepatitis B virus replication and are valuable markers for predicting the responsiveness to interferon therapy.


Hydroxyurea induction of fetal hemoglobin synthesis in sickle-cell disease. In the past 8 years, it has become apparent that some cytotoxic drugs that interfere with DNA replication can reprogram erythroid progenitors to switch from adult hemoglobin to fetal hemoglobin (HbF) production. Hydroxyurea has now been shown to substantially increase HbF in patients with sickle cell anemia. Since HbF interferes with sickle hemoglobin polymerization, hydroxyurea may become an important therapeutic agent for patients with sickle cell anemia.


Spectrum of fetal hemoglobin responses in sickle cell patients treated with hydroxyurea: the National Institutes of Health experience. Hydroxyurea is one of several cytostatic agents that increase fetal hemoglobin (HbF) production in some patients with sickle-cell disease, although their mechanisms of action remain to be defined. We have studied the effects of hydroxyurea in several hospitalized patients with sickle-cell disease treated for 3 months, who were then maintained on outpatient therapy. Among hydroxyurea-treated patients, we found that about 75% respond with at least a twofold increase in the percentages of F reticulocytes and HbF. Among the responders, HbF levels increased twofold to 10-fold, with three patients achieving levels of 10% to 15%. Statistical analysis of the three cellular variables that determine HbF levels in patients with sickle-cell disease--namely, increased F-cell production, F-cell survival, and HbF/F cells--disclosed that HbF production, as measured by an increase in F reticulocytes, accounted for about 70% of the HbF elevation, with smaller contributions coming from augmentation of HbF/F cells and preferential survival of F cells. Four responders were re-treated with their optimal weekly hydroxyurea dose, given either in daily fractions or over 4 consecutive days, after a 1- to 3-month washout period. Greater HbF responses were attained with the optimal hydroxyurea dose than with the dose-regimen escalation, and usually occurred after a lag period. Furthermore, increases in HbF and F-cell levels were more rapid in patients receiving therapy on 4 out of 7 days rather than on a daily schedule. Our calculations show that the increases in HbF/F and F cells and the decrease in the fraction of dense cells during limited hydroxyurea administration should cause a significant improvement in intracellular sickle hemoglobin polymerization tendency. Controlled prospective trials are necessary to establish whether these effects lead to clinical benefit. Alternate schedules of hydroxyurea administration, or its use in conjunction with other means to elevate HbF or reduce mean cell hemoglobin concentration, may achieve greater inhibition of polymerization and thus be more likely to result in unequivocal amelioration of disease manifestations.


Laser and photodynamic therapy of esophageal cancer. With proper selection of patients with non-operative esophageal cancer, palliative therapy with Nd:YAG laser enhances the quality of life by improving the patient's ability to swallow soft or solid food. Also, the use of non-thermal laser light to initiate the process of photodynamic therapy (PDT) is an additional laser technique that appears effective, not just in palliative treatment, but also in the cure of early esophageal cancer. For PDT, considerable work will be necessary to provide standard techniques in tumor staging, light and drug delivery and in light dosimetry. Ultimately, palliative and curative therapy of esophageal cancer will most likely incorporate multiple modalities, including surgery, radiation, chemotherapy, stents, laser and other thermal modalities, photodynamic therapy, and combinations of these methods.


Hydroxyurea and erythropoietin therapy in sickle cell anemia. Hydroxyurea has been shown to increase fetal hemoglobin (Hb F) production in patients with sickle cell disease and therefore has the potential to alleviate both the hemolytic and vaso-occlusive manifestations of the disease. Preliminary evidence indicates that recombinant human erythropoietin (rhEpo) may also induce Hb F. Three sickle cell anemia patients were treated with escalating doses of intravenous rhEpo and, subsequently, with daily oral hydroxyurea. After the optimal hydroxyurea dose was attained, rhEpo was added again. Two additional patients were treated with hydroxyurea alone. Treatment with rhEp, either alone or in combination with hydroxyurea, had no significant effect on the percentage of F reticulocytes or F cells. In contrast, hydroxyurea treatment was associated with a 1.5-fold to sevenfold increase in F cells and a 2.3- to 27-fold increase in the percentage of Hb F. In the three patients whose response reached a plateau, hydroxyurea treatment was associated with lessened hemolysis, decreased serum bilirubin and lactate dehydrogenase levels, and prolonged 51chromium-labeled RBC survival. Hydroxyurea treatment also resulted in decreased numbers of irreversibly sickled cells and in decreased sickling at partial oxygen saturation, increased oxygen affinity, increased total RBC cation content, and diminished potassium:chloride co-transport. All five patients treated with hydroxyurea experienced a decrease in severity and frequency of painful sickle crises. This study confirms that hydroxyurea therapy increases Hb F production and provides objective evidence of a significant reduction in hemolytic rate and intracellular polymerization. In contrast, rhEpo, either alone or in combination with hydroxyurea, offered no measurable benefit. Based on these encouraging preliminary data, large-scale, controlled clinical trials are warranted to study the safety and efficacy of hydroxyurea in the treatment of sickle cell disease.


Provision of food and fluids in terminal care: a sociological analysis. The artificial provision of nutrition and hydration to those with end-stage malignant disease is addressed. The physiology of cancer is investigated and is found to render such treatment futile in many instances. An analysis of the sociology of food, and the role of gender in its provision is discussed, and placed in the social milieu of the acute hospital, where there is to be found a cultural replication of the family. It is hypothesized that it is the synthesis of the powerful symbols of food and family that is at the root of behaviour in this area. Implications for patients and caregivers are explored.


Current role of laser and photodynamic therapy in gastrointestinal tumors and analysis of a 10-year experience. Laser energy can be used for a variety of neoplastic diseases including benign tumors, early stage malignancies, and advanced carcinomas, either with curative intent or for palliation. Nd:YAG laser photocoagulation of 168 colorectal adenomas allowed a complete eradication in 70% of cases, after a mean follow-up of 22 months. Advanced and obstructing tumors were treated with Nd:YAG laser to recanalize the lumen. In the upper gastrointestinal tract the recanalization of the lumen by means of laser photocoagulation improved the quality of life and survival. In fact, in our series of 308 patients treated, 1-year survival was 23% in recanalized patients and 7% in nonrecanalized patients. In the lower gastrointestinal tract, 289 cancer patients were treated and an amelioration of symptoms related to the obstruction was obtained in 93%. The current indication for photodynamic therapy is mainly the treatment of flat or ulcerative early stage tumors in the esophagus and stomach of high risk patients. Out of 17 patients treated, 14 were locally cured.


Galanin inhibition of enteric cholinergic neurotransmission: guanosine triphosphate-binding protein interactions with adenylate cyclase. The ability of galanin, a unique 29 amino acid peptide, to affect cholinergic neurotransmission was examined in the guinea pig myenteric plexus. The effects of galanin on tritiated acetylcholine ([3H]ACh) release were studied with cultured guinea pig myenteric plexus neurons. Functional correlations were made with longitudinal strips of ileal smooth muscle with attached myenteric plexus for examination of isometric contraction. Galanin abolished potassium-stimulated [3H]ACh release (170% +/- 18% of basal vs 109% +/- 16%). Galanin inhibited [3H]ACh release stimulated by forskolin or cholera toxin. [3H]ACh release stimulated by cholecystokin octapeptide, calcitonin gene-related peptide, substance P, or vasoactive intestinal peptide was also suppressed by galanin (10(-8) mol/L). Inhibitory effects were reversed by pertussis toxin preexposure, indicating involvement of guanosine triphosphate-binding proteins. Galanin inhibited contractility of longitudinal smooth muscle strips exposed to cholecystokinin-8 (EC50 7.0 X 10(-9) mol/L for cholecystokinin alone vs 1.3 X 10(-8) mol/L for cholecystokinin-8 plus galanin) and abolished contractile responses to calcitonin gene-related peptide. Galanin inhibits cholinergic neurotransmission in myenteric plexus neurons. Inhibitory effects involve guanosine triphosphate-binding protein mediation.


Heat-shock gene expression excludes hepatic acute-phase gene expression after resuscitation from hemorrhagic shock. To determine whether the gene expression of both acute-phase reactants (APR) and the major heat-shock protein (hsp-72) can occur simultaneously, transcriptional rates were measured during shock and resuscitation. A nuclear runoff technique was applied to hepatic biopsy specimens obtained from pigs before shock, during 40% blood volume hemorrhagic shock (1 and 2 hours), and after resuscitation (4 and 6 hours). Shock-induced transcription of hsp-72 was elevated elevenfold over sham operation at 2 hours (p less than 0.02, Mann-Whitney rank test). Individually shocked animals did not transcribe both classes of stress genes but segregated into two groups: (1) strong APR transcriptional responders and (2) hsp-72 transcriptional responders. In group 2, APR transcription was significantly suppressed. Antichymotrypsin transcription was an average of eighteenfold lower in group 2 versus group 1 (p less than 0.05 at 1,2, and 6 hours). Different classes of stress protein genes are not transcribed simultaneously. We infer that their increased accumulation at the mRNA level is the result of sequential transcription. Hsp-72 transcription excludes that of the APR genes that may be critical to survival after stress.


Protein kinase C is a mediator of the adaptation of vascular endothelial cells to cyclic strain in vitro. The mechanism by which hemodynamic forces influence the function of the endothelium lining a blood vessel are unknown. The aim of this study was to determine the effect of in vitro cyclic strain on endothelial cell (EC) activation of protein kinase C (PKC). Confluent bovine aortic ECs grown on flexible-bottomed culture plates were subjected to 24% maximum strain at a frequency of 60 cycles/min for 24 hours. Changes in PKC activity and evidence of translocation from cytosol to membrane fractions were assessed by immunocytochemical staining of ECs with antibodies specific to PKC and direct measurement of PKC activity in cytosol and membrane. To determine whether activation of PKC was responsible for some effects of cyclic stretch on ECs, a specific PKC inhibitor, calphostin C, was added to ECs subjected to cyclic stretch for 5 days and control ECs grown under static conditions. Immunocytochemical staining of ECs demonstrated translocation of PKC alpha- and beta-antibody fluorescence from the cytosol to the perinuclear and nuclear regions in ECs subjected to cyclic strain. This was confirmed by direct measurements of PKC activity, which demonstrated an early transient translocation of PKC activity from cytosol to membrane fraction at 10 seconds followed by a sustained elevation in PKC activity in the membrane at 100 seconds. Calphostin C abrogated the increase in EC proliferation that occurs in response to stretch. We conclude that cyclic stretch of ECs results in activation of PKC, which may be responsible for mediating the effects of cyclic stretch on EC growth.


Influence of hypercortisolemia on the acute-phase protein response to endotoxin in humans. The response to systemic infection includes the coordinated appearance of hepatic acute-phase proteins, the production of which may be influenced by a counterregulatory hormonal background. This study sought to assess the potential for hypercortisolemic conditions to influence fibrinogen kinetics and other acute-phase protein responses in humans with endotoxemia. Eleven hospitalized healthy male volunteers underwent two separate determinations of fibrinogen kinetics, one baseline and one after administration of endotoxin (2 ng/kg intravenously; lot EC-5). Seven volunteers were studied without hormonal manipulation and four in the presence of a hypercortisolemic background (hydrocortisone infusion, 3 micrograms/kg/min). Fibrinogen fractional synthetic rates were estimated from the incorporation of orally administered 15N-glycine, and fibrinogen, C-reactive protein, cortisol, tumor necrosis factor-alpha, and interleukin-6 levels were also determined. The presence of an antecedent hypercortisolemic background resulted in an attenuated interleukin-6 response, as well as decreased fibrinogen synthesis and C-reactive protein appearance. The current data suggest that glucocorticoid hormonal influences are of importance in the regulation of endotoxin-induced cytokine and acute-phase protein responses.


Parenteral anticoagulation with the heparinoid Lomoparan (Org 10172) in patients with heparin induced thrombocytopenia and thrombosis. Progressive thrombocytopenia may develop in as many as 5% of patients receiving heparin anticoagulation. In these patients, the risk of thromboembolic complications as well as continued thrombocytopenia necessitates discontinuation of heparin and initiation of an alternative anticoagulant when indicated. The heparinoid Lomoparan (Org 10172) is a mixture of several non-heparin low molecular weight glycosaminoglycans with proven anticoagulant efficacy that is generally non-reactive with platelets in the presence of plasma from patients with heparin induced thrombocytopenia, whereas standard heparin will induce platelet aggregation. We evaluated the role of heparinoid as a potential alternative anticoagulant in patients with heparin induced thrombocytopenia. During a 6 month period, we identified six patients with heparin induced thrombocytopenia who required an alternative parenteral anticoagulant, four as primary treatment for specific medical problem, and two as anticoagulation during a necessary surgical procedure. Heparinoid was used successfully in both medical and surgical patients requiring parenteral anticoagulation. In no case was there an exacerbation of the thrombocytopenia nor thromboembolic complications while on heparinoid therapy. Three of our patients sustained hemorrhagic complications, predominantly in the post-surgical setting in association with elevated anti-factor Xa levels and additional anticoagulant agents. We feel that these results confirm the utility of heparinoid anticoagulation in a select subset of patients with heparin induced thrombocytopenia who require continued parenteral anticoagulation.


Availability of the B beta(15-21) epitope on cross-linked human fibrin and its plasmic degradation products. The binding of radiolabeled monoclonal antifibrin antibody 59D8 (specific for fibrin but not fibrinogen) to a series of degraded fibrin clots showed that the availability of the B beta(15-21) epitope (against which 59D8 had been raised) was inversely proportional to the extent of clot lysis. Examination of digest supernatants revealed that the B beta(15-21) epitope was released from clots as a high molecular weight degradation product in the presence of calcium ions but that the generation of low molecular weight peptides occurred in the absence of calcium ions. To address the question of epitope accessibility, we compared levels of fibrin clot binding among four radioactively labeled antibodies: antifibrin monoclonal antibody 59D8, two antifibrinogen monoclonal antibodies that cross-reacted with fibrin, and an affinity-purified polyclonal antifibrinogen antibody. We expected that the antifibrinogen antibodies would show enhanced binding to clots in comparison with the antifibrin antibody. However, the epitope accessibility experiments showed that all four antibody preparations bound fibrin clots at comparable levels. Taken together, these studies demonstrated that one fibrin-specific epitope, B beta(15-21), remains available on clots as they undergo degradation by plasmin and, importantly, that the epitope is not solubilized at a rate faster than the rate at which the clot is itself solubilized. The availability of the B beta(15-21) epitope during the course of plasminolysis assures the potential utility of antifibrin antibodies such as 59D8 for detecting thrombi and targeting plasminogen activators.


The metabolism of cyclamate to cyclohexylamine and its cardiovascular consequences in human volunteers. A group of 194 diabetic patients were given calcium cyclamate (1 g/day as cyclamic acid equivalents) for a period of 7 days. Blood and urine samples were collected to determine the formation of cyclohexylamine, which is an indirectly acting sympathomimetic amine. Blood pressure and heart rate were recorded before and after treatment. Urine samples were collected each day and analyzed for cyclamate (to check compliance) and cyclohexylamine (to monitor the development of metabolizing activity). After 7 days intake most individuals (78%) did not excrete significant amounts of cyclohexylamine (less than 0.1% of the daily dose of cyclamate) but a small number (8; 4% of the group) excreted more than 20% of the daily dose as cyclohexylamine in the urine. Similar interindividual variations were found in the plasma concentrations of cyclohexylamine after 7 days intake of cyclamate, with 8 individuals having concentrations of 300-1942 ng/ml. The changes in cardiovascular parameters in these 8 subjects between pre- and postdosing were similar to those found in 150 subjects with plasma cyclohexylamine concentrations less than 10 ng/ml. Twenty of the subjects were restudied after receiving calcium cyclamate for 2 weeks at a daily dose equivalent to 2 g of cyclamic acid (0.66 g tds). Plasma concentrations of cyclohexylamine, heart rate, and blood pressure were measured every 30 min for a period of 8 hr (one dose interval) after the final dose. Twelve patients had plasma concentrations of cyclohexylamine greater than 10 ng/ml (89-2043 ng/ml) at the start of the dose-interval investigations. There were no transient increases or decreases in plasma concentrations of cyclohexylamine which might have resulted in a transient change in blood pressure or heart rate. These data indicate that the metabolism of cyclamate (2 g/day) to cyclohexylamine would not affect blood pressure or heart rate even in individuals with high metabolizing ability.


Repeated lindane exposure in the rat results in changes in spontaneous motor activity at 2 weeks post-exposure. Male Wistar rats were given 25 doses of lindane, 10 mg/kg per day, 1 ml/kg p.o. in olive oil, or control vehicle. Two weeks after the last dose, the animals were assessed for modifications in spontaneous motor activity, plus-maze behavior, shuttle-box active avoidance acquisition, brain regional concentrations of biogenic amines and metabolites, and regional [35S]TBPS binding. Rats treated with lindane showed an increase in spontaneous motor activity. Although no additional behavioral or neurochemical modifications were found, the changes in activity observed at 2 weeks post-exposure further demonstrate the need to assess for long lasting neurobehavioral sequelae of repeated lindane exposure.


[Damage to human endothelial cells by cholestane-3 beta,5 alpha,6 beta-triol and the protective effects of preparations that raise the intracellular level of cAMP]. Effect of drugs, which are able to elevate the intracellular level of cAMP, on resistance of human umbilical vein endothelial cells (HUVEC) to cholestane-3 beta,5 alpha, 6 beta-triol (Triol)-induced injury was studied. Triol at a concentration of 62 microM caused death of 50% of cells after a 24 hour incubation. Addition of forskolin (10 microM), methylisobutylxantine (100 microM), or 8-Br-cAMP (100 microM) into the incubation medium prevented injury of HUVEC under these conditions. These findings indicate that endothelial resistance to the injury can be regulated by the adenylate cyclase system. A comparative study on Triol-induced injury of adult human aortic endothelial cells isolated separately from zones of low (LP) and high (HP) probability of atherosclerosis was also performed. In 7 cases endothelial cells isolated from the LP zones were more resistant to Triol-induced injury, in 2 cases the differences were not significant. The development of atherosclerotic lesion in HP zones is likely to be associated with a higher sensitivity of endothelial cells from these zones to different injuring agents.


[Catecholamine blood levels in sheep during superovulation in estrus]. The effects of a serum gonadotropin (SG) superovulation hormonal preparation were investigated on catecholamine levels (norepinephrine, dopamine and epinephrine) in the blood plasma of ewes with synchronized oestrus in the oestrus period. In this trial the blood plasma of eleven ewes of the Slovak Merino breed was analyzed to detect catecholamines in the oestrus period. Superovulation was induced by an i.m. administration of 1500 IU SG as soon as oestrus synchronization with Agelin vaginal tampons finished (20 mg chlorsuperlutin) which lasted for 10 days. Catecholamines were detected in the blood plasma before synchronization, on the day of Agelin vaginal tampons application, and in 48 and 72 hours after the hormone administration--on the days of the expected ovulation. Catecholamine concentrations in the blood plasma were determined by a radioenzymatic assay using a Catechola test (Praha). The results indicate that synchronization and hormonal stimulation influence plasma catecholamine levels. The norepinephrine (NE) concentration in the blood plasma of the control samples has the value of 8.31 +/- 0.732 pmol/ml. An insignificant increase in the NE levels (13.12 +/- 0.120 pmol/ml; Fig. 1) was recorded on day 1 of the trial, after the start of synchronization. On the day of the expected ovulation the NE concentrations rose to 17.12 +/- 1.289 pmol/ml (P less than 0.001) and they remained significantly increased (P less than 0.01) at the level of 12.89 +/- 1.020 pmol/ml on the following day of the experiment. The dopamine level (DA) in the plasma of a control sample is 6.42 +/- 0.350 pmol/ml (Fig. 2).(ABSTRACT TRUNCATED AT 250 WORDS)


Localization of the virus neutralizing and hemagglutinin epitopes of E1 glycoprotein of rubella virus. Current serological assays using whole rubella virus (RV) as a target antigen for detecting RV-specific antibodies fail to define specific RV proteins and antigenic determinants such as hemagglutinin (HA) and virus-neutralizing (VN) epitopes of rubella virus. A panel of E1 deletion mutants and a subset of E1-specific monoclonal antibodies (MAb) were used for the initial analysis of HA and VN epitopes of E1 glycoprotein. A peptide region (E1(193) to E1(269)) was found to contain HA and VN epitopes. Using both overlapping synthetic peptides and truncated fusion proteins within this region, the HA epitope defined by MAb 3D9F mapped to amino acid residues E1(214) to E1(240), while two VN epitopes defined by MAb 21B9H and MAb 16A10E mapped to amino acid residues E1(214) to E1(233) and E1(219) to E1(233), respectively. The epitopes defined in this study are recognized by antibody whether or not the epitopes are glycosylated.


Definition of linear antigenic regions of the HPV16 L1 capsid protein using synthetic virion-like particles. Mice of three haplotypes (H-2d, H-2b, and H-2d/b) were immunized with synthetic HPV16 virus-like particles (VLPs), produced using a vaccinia virus doubly recombinant for the L1 and L2 proteins of HPV16. The resultant anti-VLP antisera recognized HPV16 capsids by ELISA assay and baculovirus recombinant HPV16 L1 and L2 protein on immunoblot. Overlapping peptides corresponding to the HPV16 L1 amino acid sequence were used to define the immunoreactive regions of the L1 protein. The majority of the L1 peptides were reactive with IgG from the mice immunized with the synthetic HPV16 capsids. A computer algorithm predicted seven B epitopes in HPV16 L1, five of which lay within peptides strongly reactive with the murine antisera. The murine anti-VLP antisera failed to react with the two peptides recognized by anti-HPV16L1 monoclonal antibodies raised by others against recombinant L1 fusion protein. We conclude that the immunoreactive epitopes of HPV16 defined using virus-like particles differ significantly from those defined using recombinant HPV16 L1 fusion proteins, which implies that such fusion proteins may not be the antigens to look for HPV16L1 specific immune responses in HPV-infected patients.


Epitope mapping and conformational analysis of SV40 T-antigen deletion mutants. Deletion mutants of SV40 T-antigen were stably expressed in BALB/c 3T3E cells and characterized in immunoprecipitation assays using T-antigen-specific monoclonal antibodies. The epitope recognized by antibody PAb602 was narrowed to T-antigen residues 230-362. Coprecipitation results were compatible with a p53-binding region between T-antigen amino acids 347 and 517. Amino terminal deletions (1-108 or 98-229) of T-antigen appeared to have pronounced effects on the conformation of distal regions of the molecule, based both on antibody binding and on association with p53.


Enhanced sensitivity of a second generation ELISA for antibody to hepatitis C virus. A second generation ELISA for combined detection of antibodies to three hepatitis C virus (HCV) recombinant proteins, i.e. C100, C33c and core, was compared with a first generation anti-HCV ELISA in which only antibodies to C100 are detected. The results of the ELISAs were evaluated in 225 haemophilia patients (panel A) and 44 patients with non-A, non-B (NANB) hepatitis (panel B). HCV infection was established by cDNA-polymerase chain reaction (PCR) (in panel B only) and by studying the anti-HCV reaction patterns in 4 separate ELISAs for detection of antibodies to the recombinant proteins C100, C33c, core and a combination of two synthetic peptides sp67/65 derived from the C100 region. The sensitivity for the detection of HCV infection had increased from 0.92[95% confidence interval (CI): 0.87-0.95] to 1.00 (95% CI: 0.89-1.00) in haemophiliacs and from 0.84 (95% CI: 0.66-0.95) to 1.00 (95% CI: 0.89-1.00) in NANB hepatitis patients when the second generation ELISA was used instead of the first generation ELISA. Concurrently the chance of a false negative result was reduced in panel A and B from 0.37 to 0 and from 0.28 to 0, respectively. Analysis of anti-HCV reaction patterns revealed that 172 of 206 (83.5%) anti-HCV ELISA-reactive haemophilia patients had antibodies to all 4 antigens tested. In the NANB hepatitis patients 18 of 31 (58.1%) anti-HCV ELISA-reactive subjects reacted with 4 antigens. In the PCR tested panel of NANB hepatitis patients 2 subjects who showed antibody reactivity to only one antigen and 5 patients with reactivity to 2 antigens were PCR-positive.(ABSTRACT TRUNCATED AT 250 WORDS)


Antiglobulin testing for CR1-related (Knops/McCoy/Swain-Langley/York) blood group antigens: negative and weak reactions are caused by variable expression of CR1. The Knops, McCoy, Swain-Langley and York antigens have recently been identified as being on complement receptor type 1 (CR1, CD35, C3b/C4b receptor). We examined the relationship between CR1 expression and the reactivity of the CR1-related blood group antigens with their specific antibodies. RBC from donors of selected phenotypes were tested by hemagglutination using two monoclonal antibodies to CR1, as well as anti-Kna, -McCa, -S1a, -'Kn/McC' and -Yka. Monoclonal antibodies 3D9 and E11 required approximately 250 and approximately 400 CR1/RBC to obtain a positive reaction. Agglutination of antigen-positive cells by human polyclonal antisera was related to the CR1/RBC: thus, cells expressing 20-100 CR1/RBC were negative and included the previously designated null phenotypes for this collection, 100-150 were weak or negative, and greater than 200 were usually positive. One RBC sample carried Yka on the 190,000 dalton (A or F allele), but not the 220,000 dalton (B or S allele) variant of CR1, and gave inconsistent reactions with Yka antisera. These data provide an explanation for certain of the serologic characteristics of the CR1-related blood group antigen system.


[Primary choriocarcinoma of the ovary--a case report]. A report is given on a rare case of nongestational primary chorionepithelioma of the ovary. A 30 years old woman was treated by salpingo-oophorectomy and methotrexate-monotherapy. She is showing long-term remission.


Immunohistochemistry with keratin and smooth muscle actin monoclonal antibodies in canine digestive tract and extramural glands. The canine digestive system and its extramural glands (parotid gland, liver, pancreas) were immunohistochemically studied using a panel of twelve monoclonal antibodies (MoAbs) specific for human keratin proteins and for alpha-smooth muscle actin. Various epithelial tissues and cells were characterized by different keratin staining patterns. So, the epithelial lining of the upper alimentary tract was characterized by staining with the MoAb 6B10, specific for keratin-type (K) 4, and the absence of staining with the MoAbs directed against K 8 and 18 (CAM 5.2 and RGE 53, DE-K18 respectively), whereas the lower alimentary tract epithelium was not labeled by 6B10, but stained by the latter MoAbs. In the salivary glands the luminal and basal cells of the adenomeres as well as the different ductal structures could be immunohistochemically differentiated. The duct epithelium in liver and pancreas showed next to keratin staining characteristics in common with hepatocytes and exocrine pancreatic cells, additional staining by several keratin MoAbs. The keratin staining patterns in the canine tissues showed, in addition to similarities also distinct discrepancies when compared to the staining patterns in corresponding human tissues. Myoepithelial cells in salivary and oesophageal glands could be differentiated from other basally located epithelial cells by their exclusive immunoreactivity for alpha-smooth muscle actin. Canine pancreatic endocrine cells were not labeled by any of the keratin MoAbs. It is concluded that immunohistochemistry with polypeptide specific MoAbs specific for human keratin-types can be used to differentiate between different types of canine epithelial tissues and epithelial cells in the digestive tract. As a result such reagents may find their application in developmental biology and pathology of this species.


Foot-and-mouth disease virus C3 Resende subtype analysed by means of competition RIA using neutralizing monoclonal antibodies. Foot-and-mouth disease virus (FMDV) was analysed using 30 monoclonal antibodies (MAbs) obtained from Balb/c mice immunized with FMDV C3 Resende (C3R) subtype 7 and 14 days before fusion No. 15 and 16 respectively. Fourteen MAbs were neutralizing and by means of competition radioimmuno assay it was possible to classify them into four groups. The first group consisted of MAbs specific for three sequential and three conformational epitopes. The second group consisted of MAbs specific for two conformational and for one sequential epitope. The third and the fourth groups consisted only of one MAb each, being specific for conformationally and one sequentially dependent epitope, respectively.


2-Mercaptoethanol influences the in vitro function of bovine peripheral blood mononuclear cells. The effects of 2-mercaptoethanol (2-ME) on some in vitro functions of bovine peripheral blood mononuclear cells (PBMC) were examined. It was shown that 2-ME enhanced, in a dose-dependent manner, the production of antibodies to bovine coronavirus. In this test the optimal concentration of 2-ME was 50 microM. This molarity of 2-ME was also optimal for the pokeweed mitogen (PWM)-induced proliferation of PBMC obtained from the 7 cattle tested. Similarly, the spontaneous proliferation of PBMC from 4 out of these cattle was enhanced. Thus, 2-ME evoked an increase (up to 2.5 times) or a decrease (at most 10 times) of the quota between the PWM-induced and the spontaneous proliferation (stimulation index). In general, the presence of 50 microM 2-ME enhanced the in vitro production of interferon by bovine PBMC. On the contrary, the highest proliferative response of PBMC to stimulation with bovine virus diarrhoea virus was achieved in cultures without 2-ME or in cultures with 0.5 or 5 microM 2-ME. Since the effects of 2-ME varied, for different tests as well as for cattle tested, attention should be paid to the use of 2-ME in cultures of bovine PBMC.


Discontinuous and continuous stimulation of FRTL-5 thyroid cells with bTSH cause different cAMP and nuclear proliferation antigen responses. Pulsatile TSH secretion has been described in man. We investigated the effect of discontinuous TSH stimulation on FRTL-5 thyroid cells. FRTL-5 monolayers were pulsed with TSH in 4 h incubation periods with alternate 4 h TSH-free intervals, or continuously incubated with TSH. The cAMP production of cells was measured in the supernatant of monolayers. Expression of a nuclear proliferation antigen in FRTL-5 monolayers was determined by a monoclonal antibody (Ki-67) using the alkaline phosphatase-anti-alkaline-phosphatase staining method. The TSH concentration in the stimulation series ranged from 0.01 to 1.0 U/l medium. Rhythmic cAMP production was observed in both discontinuous and continuous stimulation. With discontinuous stimulation cAMP production peaked after about 24 and 48 h, while in the continuous presence of TSH peaks were observed at 32-40 and 48 h. At all TSH concentrations the effect of discontinuous stimulation was higher than in continuously stimulated cultures. The discontinuous incubation stimulated nuclear proliferation antigen expression of FRTL-5 more intensely and there was a positive correlation with TSH concentration. We conclude that the rhythmic pattern of cAMP production after TSH stimulation is independent of the TSH pulse. The amplitude of stimulation and proliferation of FRTL-5, however, is increased by discontinuous TSH application.


Immunoreactive somatostatin in the rat ovary. Immunoreactive SRIH was detected in the rat ovary (15.6 pg/mg wet weight, 520 pg/mg protein) and was localized to the granulosa cells (168 +/- 6 pg/10(6) cells). Serial dilution studies showed parallelism of the inhibition curve for synthetic SRIH-14 and those of extracts of whole ovary and media conditioned by granulosa cells. The quantity of immunoreactive SRIH released into granulosa cell conditioned media decreased with time, while the intracellular content remained relatively constant. Gel chromatography showed peaks of immunoreactivity co-eluting with SRIH-14 (38%), SRIH-28 (31%) and a high molecular weight component. The addition of synthetic SRIH-14 stimulated meiotic maturation in cumulus-enclosed rat oocytes, with dose dependency being observed at SRIH-14 concentrations between 1 and 1000 pmol/l. No evidence of pre-pro-SRIH gene expression could be demonstrated in either rat ovary or testis using both Northern analysis and reverse transcriptase/polymerase chain reaction amplification of polyadenylated RNA. SRIH may be produced in the ovary during a specific stage of ontogeny or by an alternative gene. It is also possible that SRIH is actively taken up and stored by granulosa cells without being produced locally.


CD4+, CD33-, CD13-, CD14- acute monoblastic leukaemia. We report a case of acute monoblastic leukaemia in which the expression of the CD4 antigen occurred in the absence of myeloid and monocytic lineage specific markers. Unexpected marker profiles have biological and diagnostic implications and we also suggest that the inappropriate expression of the CD4 antigen may be implicated in the poor prognosis of this case.


Granulocyte colony-stimulating factor-producing malignant lymphoma. A 71-year-old woman with leukocytosis was admitted for treatment of malignant lymphoma. During the clinical course, neutrophilia of unknown origin occurred in parallel with the progression of the malignant lymphoma. The supernatant of lymphoma tissue culture contained a high titer of granulocyte colony-stimulating factor (G-CSF), and lymphoma cells were positive when immunohistochemically stained by anti-G-CSF antibody. Western blot analysis and mouse colony assay of the supernatant also confirmed that the lymphoma produced G-CSF.


Hyaline-vascular type Castleman's disease with concomitant malignant B-cell lymphoma. This case report describes a patient with localized hyaline-vascular (H-V) type Castleman's disease with concomitant malignant B-cell lymphoma. Malignant lymphoma has been described in association with multicentric type Castleman's disease, but not in association with the localized H-V type. Evidence for a relation between the two lesions in this patient by means of histologic, flow-cytometric, cytogenetic and gene rearrangement studies was not found.


Histochemistry of the human hair follicle with consideration of anagen phases I to VI. The hair follicle is composed of different epithelial layers under participation of mesenchymal and nerval factors. The present study is an attempt to localize differentiation and functional markers in the human hair follicle during anagen phases I to VI. Monoclonal antibody K 8.12 against keratins 13 and 16 showed an increasing reactivity with certain types of the follicle epithelia during anagen I to VI. Ki67 was expressed within the innermost layer of the outer hair root sheath. Scattered Ki67-positive matrix cells could be additionally identified during anagen V and VI but were absent in anagen I to IV. During anagen Merkel cells became more abundant in the bulbar area. Neuropeptide-like immunoreactivity expressed by bulbar (especially matrix) cells were evenly seen during the early anagen I and II. The findings are in favour of a neurohumoral modulatory role during anagen phases accompanied by an increase of expression of certain proliferation-associated antigens like keratin 16 and Ki67 among the complex epithelia of human hair follicles.


Immunohistochemical evidence for the presence of oxytocin in the opossum corpus luteum. Corpora lutea from opossums late in pregnancy were examined by immunohistochemistry for the presence of oxytocin. Oxytocin-immunoreactivity was observed in all corpora lutea examined but not elsewhere in ovarian tissue. The immunoreactive staining observed was confined primarily to the perinuclear cytoplasm of reactive luteal cells. Not all luteal cells showed oxytocin-immunoreactivity. The immunohistochemical localization of oxytocin in the pregnant opossum corpus luteum demonstrates for the first time this peptide in a metatherian ovary. Its presence in this primitive species suggests that oxytocin has a fundamental role in the physiology of the mammalian ovary.


Glycoconjugate composition of mammalian parotid glands elucidated in situ by lectins and glycosidases. Sugar specific lectins (PNA, RCA I, LPA, SBA, DBA, GSA IB4, GSA II, WGA, LTA, UEA I, Con A, LCA) with and without prior selective glycosidase digestion (sialidase, alpha-fucosidase, alpha-mannosidase, beta-N-acetylglucosaminidase, alpha- and beta-galactosidase, beta-glucosidase) were used in order to investigate the distribution of native accessible carbohydrates and obtain information dealing with the composition of terminal disaccharides within glycoconjugates present in acinar compartments and ductal segments of mammalian (mouse, rat, hare, and rabbit) parotid glands. Glycoconjugates containing variable amounts of mannose, glucose, N-acetylgalactosamine and N-acetylglucosamine were present in the parotid glands of all species. However, these carbohydrate chains exhibited a different composition of terminal sequences within each type of gland. For example, sialylated components having the terminal dimers sialic acid-galactose and sialic acid-N-acetylgalactosamine were found in all acinar cells, whereas fucoglycoconjugates with terminal disaccharide fucose-galactose were localized in the rat striated ducts and hare acinar cells. The terminal sequence alpha-galactose-beta-galactose was demonstrated in the mouse acinar cells. Finally, glycoconjugates characterized by the terminal dimer beta-galactose-N-acetylgalactosamine were demonstrated in the mouse acinar and ductal cells and the rat ductal ones. Thus, present findings outlined and further confirmed the possibility to elucidate the oligosaccharide structure in situ using lectin histochemistry combined with enzymatic degradation.


Histochemical analysis of sialomucin in Paget cells of mammary and extramammary Paget's disease. The cytoplasmic sialomucin in Paget cells of both mammary and extramammary Paget's diseases was examined, using a new method proposed by Volz et al. (1987a, b). The staining methods used involved an electrolyte-Alcian Blue (pH = 5.8) and periodic acid Schiff. Oxidation was performed with 0.4 mmol/l periodate in 1 mol/l HCl at 4 degrees C or 50 mmol/l periodate in distilled water at room temperature for 1 h. Methylation, saponification, borohydride reduction, and digestion with diastase, neuraminidase (Vibrio cholerae) or chondroitinase ABC, were also employed. The cytoplasmic mucin was found to exhibit positive reaction for the above staining which were variously altered by the chemical modification procedures and diminished in intensity or abolished by digestion with neuraminidase. These results suggest that the cytoplasmic mucin in Paget cells is sialomucin without side-chain substituent in genital Paget's disease, and that with a substituent at C7 in mammary Paget's disease.


Peripheral neuropathy in monoclonal gammopathy of undetermined significance: prevalence and immunopathogenetic studies. In an unselected series of patients with monoclonal gammopathy of undetermined significance (MGUS) we found neuropathy in 2 of 34 patients with IgG (6%), 2 of 14 with IgA (14%), and 8 of 26 with IgM MGUS (31%). The neuropathy was subclinical in 6 patients (1 IgG, 1 IgA, and 4 IgM). Patients with IgG or IgA MGUS had a prominent motor impairment with electrophysiologic and morphologic findings suggestive of predominant axonal degeneration. No deposit of the M-protein in sural nerve and no reactivity of the M-protein with nerve was detected in these patients. Patients with IgM MGUS had a prominent sensory impairment with evidence of predominant demyelination. In 6 of these patients the M-protein reacted with the myelin-associated glycoprotein (MAG). The higher prevalence of neuropathy in patients with IgM MGUS may be related to the frequent reactivity of IgM M-proteins with MAG.


A study of right unilateral spatial neglect in left hemispheric lesions: the difference between right-handed and non-right-handed post-stroke patients. We report 20 cases of right unilateral spatial neglect caused by lesions in the left cerebral hemisphere. Differences in neuropsychological symptoms and lesions sites are discussed in connection with handedness. Of the right-handed patients, 6 had severe aphasia, 4 had Gerstmann's syndrome, and 1 had pure agraphia, but unilateral spatial neglect in these cases disappeared after a number of months. Six of the non-right-handed patients had moderate-to-severe aphasia, while the other 3 cases had no aphasia at all. Eight of the 9 cases in this group continued to have right unilateral spatial neglect for more than 6 months. Lesion site as determined by CT differed as to hemisphere, but all fell into the common area previously mentioned in connection with such disorders: i.e., the temporal, parietal and occipital lobes.


Prevalence of anti-HCV in cryptogenic cirrhosis in a suburban Detroit community. To assess the role of the hepatitis C virus in patients with unexplained chronic liver disease, we tested for the presence of anti-hepatitis C antibody (anti-HCV) in the stored serum of patients with cryptogenic cirrhosis and a variety of other chronic liver diseases. The anti-HCV assay was performed by both the enzyme-linked and recombinant immunoblot methods in 16 patients with cryptogenic cirrhosis. Eight of these 16 patients (50%) were seropositive. Six of these eight patients were born outside of the United States, compared with only one of eight seronegative patients (p = 0.021). Of the anti-HCV-positive cryptogenic cirrhotic patients, 50% also had markers of previous hepatitis B infection, compared with only 12.5% of seronegative patients. Evidence of anti-HCV positivity was found in 10%, 19%, 0%, and 0% in patients with alcoholic cirrhosis, autoimmune hepatitis, primary biliary cirrhosis, and primary sclerosing cholangitis, respectively. We conclude that in a suburban American population, hepatitis C accounts for a significant percentage of patients with presumed cryptogenic cirrhosis. Unrecognized risk factors may account for a higher prevalence of HCV in foreign-born patients with cryptogenic cirrhosis. A low prevalence of anti-HCV positivity is found in other forms of chronic liver disease.


Cystic fibrosis patients bearing both the common missense mutation Gly----Asp at codon 551 and the delta F508 mutation are clinically indistinguishable from delta F508 homozygotes, except for decreased risk of meconium ileus. The glycine-to-aspartic acid missense mutation at codon 551 (G551D), which is within the first nucleotide-binding fold of the cystic fibrosis transmembrane conductance regulator (CFTR), is the third most common cystic fibrosis (CF) mutation, with a worldwide frequency of 3.1% among CF chromosomes. Regions with a high frequency correspond to areas with large populations of Celtic descent. To determine whether G551D confers a different phenotype than does delta F508, the most common CF mutation, we studied 79 compound heterozygotes for G551D/delta F508, from nine centers in Europe and North America. Each subject was matched, by age and sex, with a delta F508 homozygote from the same center. A retrospective cohort analysis was performed on the following outcome parameters: age at diagnosis, sweat chloride, meconium ileus at birth, height, weight, weight for height, FVC, FEV1, chest X-ray score, pseudomonas colonization, pancreatic sufficiency, and Shwachman clinical score. There was less meconium ileus among the G551D/delta F508 compound heterozygotes (relative risk 0.33; 95% confidence interval .13-.86), as well as a trend toward later age at diagnosis of pancreatic insufficiency. No statistically significant difference was found between the groups for any other parameter. These results suggest that the CF genotype can be a predictor of pancreatic and intestinal phenotype. Prenatal counseling for the two genotype groups should differ only with respect to probability of meconium ileus. Clinical outcome (after survival of meconium ileus) for G551D/delta F508 compound heterozygotes and delta F508 homozygotes is indistinguishable; therefore, prognostic counseling should not differ.


Development, multiplexing, and application of ARMS tests for common mutations in the CFTR gene. The amplification refractory mutation system (ARMS) is a simple, rapid and reliable method for the detection of any mutation involving single base changes or small deletions. We have applied ARMS methodology to the detection of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Single ARMS tests have been developed for 11 CFTR mutations found in the northwest of England. ARMS reactions for the most common mutations have been multiplexed to give a test which will detect the presence of the delta F508, G551D, G542X, and 621 + 1G----T mutations in a DNA sample. The multiplex test has been validated by the analysis of over 500 previously genotyped samples and has been found to be completely accurate. The rapid detection of the most common mutations has enabled early molecular confirmation of suspected cystic fibrosis in neonates, rapid typing of cystic fibrosis patients and their relatives, and testing of sperm and egg donors.


Life-threatening anaphylactoid reactions to propofol (Diprivan). Fourteen patients who had had a life-threatening reaction within a few minutes after receiving propofol (Diprivan) were investigated for anaphylaxis 4-6 weeks after the incident. Three kinds of immunologic tests were carried out: skin tests (prick tests and intradermal tests with the drugs used and Intralipid, the solvent for propofol), a leukocyte histamine release test, and a radioimmunoassay (RIA) of immunoglobulin E (IgE) against propofol and muscle relaxants, when they had been given with propofol. It had been previously shown that these were always negative in patients anesthetized with propofol without any complications. Thirteen of the 14 patients had at least one positive test supporting hypersensitivity to propofol; 2 patients had three tests positive; 4 had two tests positive; and 7 had one test positive. The skin tests with Intralipid were negative in 4 patients whose tests with propofol were positive. Two patients who had been given muscle relaxants at the same time as the propofol had positive IgE-RIA to both drugs. In one patient, results of all the tests remained negative, and the mechanism involved in the reaction remained unidentified. It is note-worthy that 9 patients of 14 had allergic histories that were known before the anesthetic (atopy; allergy to antibiotics, muscle relaxants, lidocaine, colloids) and that none of the patients had ever received propofol or Intralipid before. It is possible that the IgE that linked abnormally with the propofol had specific binding sites for the phenyl nucleus and the isopropyl groups, which are present in propofol and many other drugs.(ABSTRACT TRUNCATED AT 250 WORDS)


The role of radiotherapy in the primary management of cutaneous melanoma. The historical role of radiotherapy in the management of cutaneous melanoma has largely been for palliation. Fractionation schedules using infrequent large doses have proved convenient and effective. Response rates range from 50% to 85% with the likelihood of complete response strongly dependent on tumor volume. These observations, along with radiobiological data showing a wide range of cellular radiosensitivity within the same histotype, suggest that radiotherapy would be most reliably effective against microscopic or subclinical disease. A few investigators have evaluated the role of radiotherapy combined with surgery as a curative approach in early-stage, high-risk disease. Patterns of failure after surgery alone indicate a substantial risk of local and regional recurrence that can be reduced by radiotherapy. Strategies for integrating radiotherapy into primary management are reviewed and additional studies are suggested.


Dual Analyte immunoassay--a new approach to neural tube defect and Down's syndrome screening. A microtiter plate based Dual Analyte enzyme-immunoassay method for the simultaneous measurement of alpha-fetoprotein (AFP) and Free-beta human chorionic gonadotrophin (hCG) was evaluated. This rapid assay, which has application in both Neural Tube Defect screening and Down's screening, shows good precision with between assay coefficients of variation between 5 and 7.5% for AFP and 3.7 to 5.8% for Free-beta(hCG). Correlation with single analyte procedures is good, with correlation coefficients being greater than 0.91 in both cases. Clinical discrimination in detecting both types of abnormalities is not compromised by this new simultaneous Dual Analyte assay. We conclude that the Dual Analyte approach, which combines analytes achieving the highest known detection efficiency, will bring about improvements in the efficiency of screening, reduce costs and improve report turnaround, all leading to better quality of patient care.


Urinary albumin and low molecular weight protein excretion in the nephrotic syndrome--sequential studies during corticosteroid treatment. In a preliminary investigation into the behaviour of low molecular weight proteins in the nephrotic syndrome, we have measured urinary concentrations of albumin, alpha-1-microglobulin (alpha 1-m) and retinol-binding protein (RBP) in six children for up to 11 days during the course of steroid therapy for nephrotic syndrome. The results in part support the concept of independent proximal tubular absorption of albumin and low molecular weight proteins, and indicate that in the nephrotic syndrome the excretion of RBP and alpha 1-m, two generally accepted markers of tubular proteinuria, is anomalous.


Correlation between multi-headed spermatozoa in the ejaculate and histological findings in the testis. Semen smears of 163 patients as well as bioptic material of 163 patients (93 cases of paraffin histology and 70 cases of cryostat sections) were grouped by their content of multi-headed spermatozoa or multi-nucleated spermatids, respectively. It could be demonstrated that findings in the ejaculate concerning multi-headed spermatozoa are in good agreement with those in the testis in respect of the presence of double- and multi-nucleated spermatids. Furthermore, it could be shown that the frequency of multi-headed spermatozoa is positively correlated to inflammatory processes either of immunologic origin or caused by micro-organisms.


Ganglioside GD1b is the target antigen for a biclonal IgM in a case of sensory-motor axonal polyneuropathy: involvement of N-acetylneuraminic acid in the epitope. We report on a 54-year-old man with a sensory-motor polyneuropathy associated with a biclonal IgM-kappa gammopathy, which reacted with the ganglioside GD1b. Examination of nerve biopsy specimens showed some reduction in the density of myelinated fibers and axonal degeneration with a loss of large fibers and a relative increase in the density of small fibers. Immunodetection on thin-layer chromatography of the glycolipid antigens showed strong reactivity of the patient's serum IgM-kappa with GD1b ganglioside and weak binding to GD1a. biclonal IgM antibodies did not react with GM1, asialo-GM1, GT1b, GD2, or GD3. Indirect immunofluorescence staining showed binding of IgM-kappa mainly in a crescent-like pattern on the internal side of myelin sheaths, which could correspond either to an enlarged periaxonal (adaxonal) space or to the internal mesaxon or to both. The immunostaining was abolished after absorption of the serum with GD1b.


Immunogenicity of free synthetic peptides corresponding to T helper epitopes of the influenza HA 1 subunit. Induction of virus cross reacting CD4+ T lymphocytes in mice. Four linear synthetic peptides corresponding to residues 12-29, 50-67, 121-138 and 131-147 of the HA 1 subunit of H3 subtype influenza virus (NT/60/68) were tested for their capacity to elicit in vivo peptide-specific CD4+ T cells cross reacting with whole virus. By studying the in vitro peptide proliferative response of lymph node cells from mice sensitized in vivo with free peptides emulsified in complete or incomplete Freund adjuvant, it was found that region 12-29 could be recognized by CD4+ T lymphocytes in the context of H-2k and H-2b, region 50-67 in association with H-2b and region 121-138 in the context of H-2d MHC molecules. Outbred OF 1 mice could recognize regions 50-67 and 121-138. Peptides 50-67 and 121-138 are of potential interest for synthetic vaccine design since they induced in BALB/c (peptide 121-138) and OF 1 (both peptides) mice a CD4+ T cell population that cross reacted with whole virus. The region 50-67 is of particular interest since only few substitutions have been found in this area in natural variants of the H3 virus subtype.


Epitope specificities of human serum antibodies reactive with respiratory syncytial virus fusion protein. Respiratory syncytial (RS) virus continues to cause serious human respiratory disease and no prophylactic vaccine is yet available. Serum antibodies to RS virus fusion protein (F) that have the appropriate specificities and activities could confer protection against severe RS virus infections. To explore human serum antibody responses to RS virus F we first characterised four epitopes on F and then measured the concentrations of human serum antibodies to these sites for 389 sera. Individuals varied in serum antibody concentration to the epitopes. The distribution patterns of the concentrations of antibodies reactive to each epitope were different. Antigenic variation of F at these epitopes in Southampton RS virus isolates was examined by immunofluorescence. The F proteins from different isolates varied within and between RS virus subtypes which co-circulated in the outbreak of winter 1985-1986. Variations in F detected by immunofluorescence were consistent with differences between the strains' susceptibilities to monoclonal antibody antiviral action.


Successful treatment of mycoplasmosis in layer chickens with single dose therapy. The efficacy of treatment with single dose administration of 5 drugs at different dosages to layer hens naturally infected with Mycoplasma gallisepticum was studied. The drugs were tiamulin, which was administered orally, tylosin (parenterally and orally), spiramycin (orally), long-acting oxytetracycline (parenterally) and tylosindihydrostreptomycin (parenterally). Cure was assessed by the absence of nasal discharge. The cure rate was significantly higher (P less than 0.05) in treated hens than in untreated hens, as early as 1 day after treatment. Remission for 33 days was achieved in 60% of hens treated with 100 mg oxytetracycline, in 100% of hens treated with 100 mg or 200 mg spiramycin, in 92% and 85% of hens treated with 100 mg tylosin, parenterally and orally, and in 89% and 88% of birds given 100 mg tiamulin and tylosin-dihydrostreptomycin, respectively.


Effect of albumin on adenylate cyclase receptor-related signal transduction of human peripheral blood mononuclear cells. In the present study we investigated in vitro the effect of human serum albumin (HSA) on receptor-stimulated cAMP production in isolated human peripheral blood mononuclear cells (PBMC). The cAMP production is strongly correlated with the pH of the medium during long incubations with albumin. Adenylate cyclase is stimulated by receptor agonists like histamine, forskolin, prostaglandin E2 and the beta-adrenergic agonist isoprenaline, in the presence or absence of HSA. This protein, at concentrations above 0.1%, dose-dependently inhibits both basal and agonist-stimulated cAMP levels in PBMC. In the presence of 0.5% HSA a significant reduction of 30-60% (cell batch dependent) is induced, a reduction which is not incubation time dependent. Washing the cells after a period of incubation with 2% HSA does not reverse the HSA-induced cAMP inhibition. Oleic acid-evoked conformational changes in HSA were not capable of influencing the inhibition processes of HSA on the isoprenaline-stimulated cAMP production. Structure-controlled interactions between HSA and membrane or adenylate cyclase are therefore unlikely. Bovine serum albumin and chicken albumin had different effects upon the agonist-stimulated cAMP production as compared with HSA. At this moment no explanation for this behavior can be provided. The findings indicate that albumin may inhibit non-specifically cAMP production in PBMC and possibly influences membrane-controlled processes.


High percentage of CD8+, Leu-7+ cells in rheumatoid arthritis synovial fluid. While analyzing the phenotype of the synovial fluid mononuclear cells (SFMC) clustered about dendritic cells in rheumatoid arthritis (RA) joint effusions, it was noted that most of the clustering cells were CD8+ and coexpressed Leu-7. Therefore, the present study was conducted to investigate the frequency of CD8+, Leu-7+ cells in RA SF: SFMC from 13 patients with RA and from 12 patients with non-RA inflammatory arthritides were examined for CD8 and Leu-7 expression using 2-color immunofluorescence flow cytometry. RA SFMC had statistically significantly greater percentages of total CD8+ cells, total Leu-7+ cells, and CD8+, Leu-7+ cells, compared with SFMC from the non-RA patients. These RA CD8+, Leu-7+ SFMC had a distinctive electron microscopic appearance compared with CD8+, Leu-7- SFMC. When peripheral blood mononuclear cells (PBMC) from 31 RA patients (including 7 from the SFMC group) were compared with PBMC from 15 normal controls, the percentage of CD8+, Leu-7+ cells was not significantly greater in the RA patients. However, the combination of a modest increase in CD8+, Leu-7+ cells and a decrease in total CD8 cells in RA PBMC altered the composition of the RA CD8 population compared with normal PBMC, such that over 40% of RA peripheral blood CD8 cells coexpressed Leu-7. The increased frequency of CD8+, Leu-7+ cells in RA SFMC may arise from the fact that a high percentage of the CD8+ PBMC in RA patients are also Leu-7+. This altered composition of CD8 cells in RA SF may have a role in the pathogenesis of the disease.


Analysis of intraarticular fibrinolytic pathways in patients with inflammatory and noninflammatory joint diseases. Intraarticular activation of the fibrinolytic system has been suspected to occur in patients with arthritis. We undertook the present study to investigate the relation of this activation to clinical symptoms, and the molecular pathways involved. We quantitatively assessed levels of plasmin-alpha 2-antiplasmin (PAP) complexes in synovial fluid (SF) from 25 patients with rheumatoid arthritis (RA), 7 with seronegative spondylarthropathy (SSA), and 10 with osteoarthritis (OA), and conducted an analysis to determine the plasminogen-activating pathway via which these complexes were generated. In addition, we studied the relationship of intraarticular fibrinolysis to clinical and biochemical parameters. All patients studied had increased SF levels of PAP complexes. Levels in patients with RA and SSA were slightly higher than those in patients with OA. These complexes were probably formed by activation of urokinase-type plasminogen activator (u-PA), and not tissue-type plasminogen activator (t-PA), since SF levels of both u-PA antigen and u-PA-plasminogen activator inhibitor (PAI) complexes were increased in 27 of the 42 patients. Conversely, SF levels of t-PA were below normal in all but 1 patient. In some patients, activation of factor XII presumably also contributed to plasminogen activation in SF, since levels of factor XIIa-C1 inhibitor in SF were increased in 8 of the 42 patients and correlated, as did u-PA-PAI levels, with levels of PAP complexes. Several of the parameters of fibrinolysis in SF, particularly u-PA antigen and u-PA-PAI-1 complexes, were found to correlate with clinical and biochemical parameters. Our results suggest that plasminogen is frequently activated in the joints of patients with inflammatory or noninflammatory arthropathy and that this activation mainly occurs via a u-PA-, and in some cases also via a factor XII-, dependent pathway. The possible relation of this activation process to stimulation of synovial cells by cytokines is discussed.


Immunopathology of rheumatoid arthritis. Antikeratin antibodies precede the clinical disease. We sought to determine whether circulating antikeratin antibodies (AKA) precede the onset of rheumatoid arthritis (RA). By matching the registers of 2 previous population studies with the registry of patients receiving antirheumatic drugs several years later, pre-illness serum specimens could be obtained from 39 individuals who subsequently developed RA. AKA were assayed with the standard indirect immunofluorescence technique. Ten of 39 serum specimens from individuals who subsequently developed seropositive RA, and 1 of 15 sera from individuals who developed seronegative RA, were positive for IgG-class AKA by immunofluorescence assay. The AKA-positive sera were also positive for rheumatoid factors. The findings focus attention on the role of pre-illness immunologic events in the pathogenesis of RA.


T cell receptor V beta repertoire of double-negative alpha/beta T cells in patients with systemic sclerosis. To analyze the T cell receptor V beta gene on double-negative (DN) alpha/beta T cells, which are increased in number, on peripheral blood lymphocytes (PBL) from patients with systemic sclerosis (SSc). The DN alpha/beta T cells were sorted by flow cytometry from PBL obtained from 3 patients with SSc. The V beta repertoire was analyzed by polymerase chain reaction. Only 1 or 2 V beta genes (V beta 5/7, 5, or 17) were predominantly expressed on DN alpha/beta T cells from these 3 patients. The V beta repertoire on DN alpha/beta T cells in PBL from patients with SSc is rather restricted.


Autoantigenic epitopes of the B polypeptide of Sm small nuclear RNP particles. Identification of regions accessible only within the U1 small nuclear RNP. To analyze the autoantigenic epitopes of the Sm B polypeptide of the U1 small nuclear RNP (snRNP) using complementary DNA (cDNA) clones. Expression of Sm B fusion proteins in lambda phage vectors, immunoblots, immunoprecipitations, and affinity purification of antibodies. Immunoblots using antibodies affinity-purified from B fusion proteins demonstrated that there were cross-reactive epitopes between the B'/B and A polypeptides of the U1 snRNP. Immunoprecipitation assays suggested that there were at least 3 different autoantigenic epitopes on the B polypeptide that could be classified into 2 general groups based upon autoantibody reactivity. The first group of autoantibodies, which bound 2 separate autoantigenic epitopes (BU1-1, BU1-2), participated in immunoprecipitation of the U1 snRNP alone. The second group, which bound the third type of autoantigenic epitope (BSm-1), immunoprecipitated all the abundant Sm snRNPs. There is at least 1 region on the B proteins that is accessible to antibodies only within the structure of the U1 snRNP, as well as a region that is accessible on all Sm snRNPs. These data support the notion that the native U1 RNP, perhaps containing B proteins in a different conformation than those found on other Sm snRNPs, may drive the humoral immune response in systemic lupus erythematosus.


Methemoglobinemia secondary to automobile exhaust fumes. Methemoglobinemia is an uncommon cause of cyanosis. A 28-year-old male presented to the emergency department cyanotic and short of breath after exposure to noxious automobile fumes. He did not improve with the administration of 100% oxygen therapy. The initial arterial blood gas with cooximetry was: pH of 7.38, PaCO2 of 43 mm Hg, PaO2 of 118 mm Hg, measured oxygen saturation of 70%, and a methemoglobin level of 24.8%. Methylene blue was given (2 mg/kg intravenously) and the patient's symptoms resolved. On the following day he was discharged home without complication. A comprehensive review of the literature revealed no reported cases of methemoglobinemia secondary to accidental exposure to exhaust fumes.


Nitric oxide: a pathogenetic factor in autoimmunity. Nitric oxide (NO) has been identified recently as a multifunctional mediator, produced by, and acting on, most cells of the body. Besides its function as endothelium-derived relaxing factor, as a neurotransmitter and as an immune defence molecule, evidence is accumulating that NO participates in inflammatory- and autoimmune-mediated tissue destruction. Modulation of NO synthesis and action represents a new approach to the treatment of inflammatory and autoimmune conditions.


Airway macrophages releasability in bronchial asthma. Asthma is a multifactorial disease on genetic basis. Its development is influenced by maternal and environmental factors, i.e. allergens and adjuvants. Early identification of candidates at high risk for development of asthma will enable giving recommendations on preventive measures focussing on exposure to tobacco smoke and other pollutants, indoor and outdoor allergens and possibly viral infections during infancy.


Function and specificity of human gamma/delta-positive T cells. gamma/delta+ T-cells are a recently identified subpopulation of T-lymphocytes expressing an "alternative" T-cell receptor (TCR) molecule consisting of disulfate-linked or nonlinked gamma and delta chains. Despite a limited number of V gamma and V delta genes in the germ line, there is a large and diverse gamma delta TCR repertoire due to extensive N region variability. Recently developed monoclonal antibodies against V gamma and V delta gene products are useful reagents for the identification and isolation of gamma/delta+ T-cell subpopulations. The physiological significance of gamma/delta+ T-cells is still unknown. However, accumulating evidence indicates that human gamma/delta+ T-cells frequently recognize bacterial ligands as well as certain tumor cells. Interestingly, reactivity towards microbial antigens is usually restricted to a subpopulation of gamma/delta+ T-cells expressing a V gamma 9/V delta 2 TCR. However, different bacteria-reactive V gamma 9+/V delta 2+ gamma/delta+ T-cells display extensive N region variability, suggesting the involvement of a gamma/delta-specific superantigen in these responses. Little is known about the role of gamma/delta+ T-cells under pathological conditions. Rare cases of gamma/delta+ T-cell leukemias and lymphomas have been described. In addition, discrete changes in the distribution of gamma/delta+ T-cell subpopulations have been observed during HIV infection. Current thinking favors the interpretation that gamma/delta+ T-cells play a role in the immune reaction during infection and in the regulation of physiological or pathophysiological autoimmune responses.


Structural analysis of class II major histocompatibility complex proteins. Numerous studies have contributed much data that, taken together, provide considerable insight into the structural features of class II molecules. The putative peptide-binding region has been modeled from the crystal structure of the class I molecule HLA-A2. Mutational analyses and peptide-binding studies support this model. Direct structural studies have established features of the noncovalent association of the alpha and beta chains of class II molecules, and detergents and phospholipids have been shown to affect class II protein structure and/or peptide-binding capacity. Spectroscopic studies have revealed dynamic features of class II molecules in solution, and suggest differences in the overall structures of class I and II molecules that may reflect differences in function.


A technique for demonstrating the nerve supply of whole larynges. At present there is a large gap in our understanding of the exact distribution of sensory and motor nerves within the larynx. Gross dissection is only accurate for large-caliber nerves, whereas microscopic sections show only isolated examples of terminal branching. To fill this gap, we modified an old technique called Sihler's stain to process five whole canine larynges. The technique requires eight steps that take approximately 3 months. The result is an entire larynx in which muscle and cartilage are rendered translucent, while nerve is counterstained. Nerve branching can be continuously traced from the large-caliber nerves to the terminal branches. We found that the nerve patterns are surprisingly complex and contain frequent anastomotic networks. This technique is a powerful tool for studying certain neurolaryngological problems.


Antigenic change in a human IgG4-specific CH3 epitope upon binding of a monoclonal antibody against a neighboring IgG4-specific epitope. Two sets of monoclonal antibodies (mAbs) probably reacting with two different epitopes in the CH3 domain of the human IgG4 molecule were studied. We observed that the commercially available mAb HP 6011 inhibited the antigen binding of the three mutually inhibitable mAbs, 40-A2, 41-E8 and 43-F11 (40-series), made by us. However, the 40-series mAbs, including those with similar affinity such as mAb HP6011, were not able to inhibit mAb HP 6011. When the 40-series mAbs were preincubated with IgG4, the mAb HP 6011 could partially displace these antibodies. This one-way inhibition indicates that upon binding mAb HP 6011 changes the antigenic structure of the IgG4 molecule by disrupting the epitope for the 40-series mAbs. A steric hindrance of this epitope by mAb HP 6011 is more unlikely, since the small Fab fragment of mAb HP 6011 also inhibited the reaction of the 40-series mAbs.


Squamous cell metaplasia of the bladder urothelium. A retrospective study of 36 patients. In a retrospective study of 28 women and eight men with squamous cell metaplasia in different parts of the bladder, including the trigone, no histopathological differences were observed among the regions. All the five (female) patients with parakeratosis had a concomitant invasive bladder tumour. Thirty-eight% of all the patients had a simultaneous neoplastic tumour. The metaplastic lesions were investigated for keratin in 13 patients, and all were positive. In seven out of eight patients, the urothelium adjacent to the squamous cell metaplasia was also positive for keratin, indicating a direct transformation of the urothelium to squamous cell epithelium. The metaplastic cells were investigated for oestrogen receptors in five men and five women, and all were negative, suggesting no relationship between estrogens and squamous cell metaplasia of the bladder. Squamous cell metaplasia in the bladder is not considered a premalignant condition. However, metaplasia and neoplastic tumours are often associated with chronic tissue damage, and the presence of metaplasia may give a warning of conditions that can also cause cancer.


Using a cationic carbocyanine dye to assess RNA loading in Northern gel analysis. In Northern blotting, one must have a means of assessing the uniformity of RNA loaded into each lane of a gel. As an alternative to "common gene" controls and previously published nucleic acid dyes (ethidium bromide, acridine orange, methylene blue), we have utilized a cationic carbocyanine dye (Stains All) for the assessment of RNA gel loading uniformity over the range of 5-25 micrograms RNA/lane. The following protocol is suitable for messages of well-characterized mobility and utilizes xylene cyanol as a 4-kb marker; as such, it will migrate between 28S and 18S rRNA over a wide range of agarose concentrations. Optimally, it is best that the message(s) of interest should migrate either as a smaller species than 18S or as a larger species than 28S; this allows either the 28S or 18S ribosomal band to be separated from the message(s) of interest by severing the gel transversely at the xylene cyanol front. Severing the gel in such a manner makes it possible to simultaneously submit that portion of the gel containing either the 28S or 18S rRNA band to Stains All staining while immediately continuing with the transfer of that portion of the gel containing the mRNA of interest. We have found the dye to interact linearly with rRNA whether data were gathered by densitometrically scanning the gels themselves or photographs of the gels.


CD59: a molecule involved in antigen presentation as well as downregulation of membrane attack complex. CD59 is a recently discovered complement (C) regulatory protein. Three activities of CD59 have been recognized so far. It can downregulate the activation of the C cascade at membrane attack complex formation stage, it participates in T-cell rosette formation with erythrocytes and appears to be necessary for T-cell activation. CD59 is broadly distributed on cells of various organs and this is compatible with its function of protecting cells and tissues from incidentally activated autologous C. CD59 is encoded by a single gene residing on chromosome 11. The entire sequence of CD59 cDNA is known, from which the amino acid structure of CD59 has been deduced. Mature CD59 is made up of 103 amino acids. It has two potential N-glycosylation sites. Carbohydrate constitutes about 20% of the molecular mass of CD59. The protein part of the molecule is covalently linked to an oligosaccharide which, in turn, is glycosydically linked to phosphatidylinositol (PI). CD59 is anchored to the cells via PI moiety. The fine structure of the PI anchor of CD59 has not yet been established. Decreased expression of CD59 has been shown in two diseases. In paroxysmal nocturnal hemoglobinuria, erythrocytes (type III) lacking CD59 (and other PI proteins) have susceptibility to lysis by membrane attack complex. In psoriasis, expression of CD59 (and CD55) is drastically decreased in psoriatic lesions, presumably due to PI cleavage involving signal transduction mechanism(s) leading to increased proliferation of various cell types in lesional skin.


Comparative aspects of basal forebrain organization in vertebrates. The present survey shows that basal ganglia organization in the three classes of amniotes has many features in common. Yet, there are several differences that are important to note since they contribute to a better understanding of the evolution of those brain structures. They also underscore once again that it is questionable to generalize the results obtained in a certain species. This question is the more important in view of the general practice in mammalian research. Comparative studies may be of great help to solve this problem. The reported differences in acetylcholine release after pharmacological treatment in the striatum of rats and reptiles have stimulated further research that has led to the conclusion that the nucleus accumbens in the rat is a very heterogeneous structure with respect to the regulation of the release of acetylcholine by D2 dopamine and NMDA receptor activation. In this regard, the striatum of reptiles may offer a model for studying the functioning of the caudomedial part of the rat nucleus accumbens.


Etiologic factors and pathologic alterations in selenium-vitamin E deficiency and excess in animals and humans. The etiology of selenium-vitamin E (Se-E) deficiency diseases may be complex. Many of the syndromes involve combined deficiency of selenium and vitamin E. Selenium moves into the animal and human food chain from soil and plants, which may contain inadequate amounts of the nutrient in many areas of the world. Vitamin E may be in low concentration in many animal feeds unless supplements are added. Some syndromes, such as steatitis in cats, result from an increased requirement of vitamin E in diets that contain large amounts of polyunsaturated fatty acids, and these diseases will only respond to vitamin E administration. Deficiency syndromes in animals owing to pure Se deficiency are infrequent and have been produced mainly by laboratory studies utilizing extreme deficiency conditions. Other factors that may affect the occurrence of these deficiency diseases are concurrent dietary deficiency of S-containing amino acids, bioavailability of different forms of dietary Se, intake of compounds that antagonize Se (e.g., silver salts), and exposure to various prooxidant substances (e.g., iron compounds, oxygen, ozone, and various drugs).


Effects of glutathione and of cysteine on intestinal absorption of selenium from selenite. The influence of glutathione (1 mmol/L) (GSH) on in vitro mucosal uptake and in vivo absorption of 75Se-labeled selenite (10 mumol/L) was investigated in rat jejunum. For comparison, the effect of L-cysteine (1 mmol/L) on in vivo absorption of 75Se-labeled selenite was also studied. In the in vitro uptake experiments, only the mucosal surface was exposed to the incubation medium for 3 min. For the in vivo experiments, a luminal perfusion technique was employed. GSH inhibited in vitro mucosal Se uptake, whereas absorption in vivo was stimulated by GSH. L-Cysteine also stimulated in vivo Se absorption, confirming former in vitro mucosal uptake experiments. Thus, unlike L-cysteine, GSH affected in vitro and in vivo absorption of Se from selenite differently. Enzymatic cleavage of products of the reaction of selenite with GSH occurring more efficiently under in vivo than in vitro conditions may be a prerequisite for the stimulatory effect of GSH on Se absorption. This apparently does not apply to the stimulatory effect of cysteine. Since GSH occurs in the intestinal lumen under physiological conditions, it may contribute to the high bioavailability of Se from selenite.


Stimulation of mucosal uptake of selenium from selenite by some thiols at various sites of rat intestine. The influence of several thiols (conc. 1 mmol/L) on mucosal uptake of 75Se from 75Se-labeled selenite (conc. 10 mumol/L) across the brush border of rat jejunum and cecum was investigated in vitro using a short-term uptake technique. L-Cysteine (L-Cys) stimulated 75Se uptake in the mid- and distal jejunum and cecum, but not in the proximal jejunum. The effect was maximal in the distal jejunum. D-Cys was less effective in the jejunum and similarly effective in the cecum. L-Leucine (L-Leu) and L-glutamic acid significantly reduced the stimulatory effect of L-Cys on Se uptake in the distal jejunum, whereas the respective effect of D-Cys was not diminished by L-Leu. Cysteamine stimulated mucosal 75Se uptake at all intestinal sites tested, whereas the effect of mercaptopyruvate was restricted to the distal jejunum. Thioglycolate also enhanced 75Se uptake in the distal jejunum. The stimulatory effects of L-Cys, mercaptopyruvate, and thioglycolate were Na(+)-dependent, whereas the effect of cysteamine also occurred in the absence of Na+. Mercaptosuccinate, D-penicillamine, ergothioneine, and thiosulfate did not enhance mucosal 75Se uptake. It is concluded from these findings that the reaction of some thiols with selenite results in Se compounds that are rapidly absorbed by the intestinal epithelium through various Na(+)-dependent and Na(+)-independent mechanisms. The high bioavailability of Se from selenite found by others might thus be the result of the presence of thiols in the gastrointestinal tract.


Glycoalkaloid and nitrate content of potatoes as affected by method of selenium application. A comparison of two methods of selenium application, banding and foliar spray, of sodium selenite (Na2SeO3) on total glycoalkaloid (TGA) and nitrate nitrogen (NO3-N) was studied during each of two consecutive years. The levels of application used were 0.0, 1.6 (0.75), 3.36 (1.5), and 5.6 (2.5) kg/ha (ppm soil). Both TGA and NO3-N were significantly reduced by application of 1.5 and 2.5 ppm of sodium selenite. Tuber selenium levels were significantly increased at all levels of application, using either banding or foliar spray, but were well below the toxic range for human consumption. Banding resulted in greater uptake of Se, and greater decreases in TGA and NO3-N as compared to foliar spray.


Selenium and blood pressure studies in young and adult normotensive, renal, and spontaneously hypertensive animals. With reports of either no change or reduction of blood pressure, the relationship between selenium and blood pressure has not been clear. Normal Se values are not available for the Sprague Dawley (SD) rat or in the young and adult rat with various models of experimental hypertension. This study measured serum Se levels in the young and adult normotensive (NT), Grollman renal hypertensive (RH), and Okamoto-Aoki spontaneous hypertensive rats (SHR). The young animals have statistically significant (P less than 0.001) lower Se values as measured by the fluorometric method than those found at adulthood. Selenium levels were found to be altered in the adult SHR animals when compared with the RH and NT animals. The serum Se value for the normotensive SD rat was found to be 65.0 +/- 3.5 micrograms/dL, and for the two experimental models, 63.7 +/- 4.6 micrograms/dL for the RH, whereas the SHR level was elevated to 75.04 +/- 4.8 micrograms/dL (P less than 0.001). Elevated serum Se values in the adult SHR animals suggests an altered metabolism in SHR animals.


Lymphocyte glutathione peroxidase activity during exacerbations in multiple sclerosis. Glutathione peroxidase, one of the major antioxidants in the human brain, has been found to have decreased activity in patients suffering from multiple sclerosis (MS). This study compares the activity of lymphocyte glutathione peroxidase (L-GSH-px) in MS patients suffering from acute relapses with clinically stable MS patients and with control patients referred with nondemyelinating neurological diseases. All three groups showed an increase of mean enzymatic activity (MEA) during the observation period. The highest MEA in this study was observed in the MS groups. However, there were no significant differences in the L-GSH-px activity in the three groups. These results are not in accordance with previous investigations, and the need for further research in this field is emphasized.


Some properties of selenoprotein P.

Some properties of selenoprotein P. Selenoprotein P is a newly characterized selenoprotein. It is the first protein described to contain multiple selenocysteines. It is secreted by the liver into the plasma and turns over rapidly. Its concentration is sensitive to the selenium status of the animal. Its function is unknown.


Metabolic differences and similarities of selenium in blood and brain of the rat following the administration of different selenium compounds. A common intermediate, i.e., selenite, was found in the serum of the rat; the maximum levels occurred 3 h after administration independent of chemical forms. This indicates that both the reduction of selenate to selenite, and oxidation of seleno-dl-methionine to selenite existed in the metabolic pathways of the rat. We found that water-soluble selenium compounds led to a similar maximum content in blood and serum, but seleno-dl-methionine had a higher affinity for the brain and, by gel filtration chromatography, for the higher mol-wt (25-100 K Da) fractions of serum protein, when compared with inorganic forms.


Biogeochemical regioning problems and the biogeochemical selenium provinces in the former USSR. Current trends in biogeochemical research in the former USSR are exemplified for the trace element selenium (Se). Vast regions of the former USSR are low in Se, giving rise to selenium deficiency diseases in animals and to Kaschin-Beck disease in humans, whereas isolated high-Se regions are comparatively rare. The Se content of plants depends on geological soil-type and secondary processes such as weathering and leaching. In general, a direct correlation between the Se content of feedstock and of the blood in animals is observed, whereas corresponding data for humans remain to be accumulated.


Selenium content and distribution in rat tissues irradiated with gamma rays. The effects of supplementation with selenous yeast and ionizing radiation on selenium (Se) content and distribution were evaluated in rat tissues (liver, kidney, spleen, heart, muscle, blood, front brain, hind brain, hypothalamus, pituitary, adrenal glands, testes, and hair). This study had 16 Se-supplemented (0.5 micrograms Se/d) and 16 placebo adult male Wistar rats. One half of the animals (eight Se-supplemented and eight placebos) were irradiated with a single dose of 4.2 Gy from a Co-60 source and sacrificed 7 d after irradiation along with nonirradiated animals and analyzed for Se content determination. The data obtained showed that selenous yeast supplementation increased Se levels in rat tissues (highest increases in hypothalamus, 161%; hind brain, 126%; spleen, 110%; and adrenal gland, 105%). Ionizing radiation induced significant changes in Se content and distribution (decrease in liver, blood, hair, femoral muscle, spleen, and hypothalamus; increase in kidney, testes, adrenal glands, and brain of placebo group). Supplementation with selenous yeast reduces changes in Se content and distribution after irradiation. It seems that the animal tissue susceptibility to oxidative damage may be correlated to their ability to retain Se in tissues.


Regulation of cellular immune responses by selenium. Selenium (Se) is an essential nutritional factor that affects the development and expression of cell-mediated immune responses directed toward malignant cells. These studies have shown that dietary (2 ppm for 8 wk) or in in vitro (1 x 10(-7)M) supplementation with Se (as sodium selenite) results in a significant enhancement of the proliferative responses of spleen lymphocytes from C57Bl/6J mice in response to stimulation with mitogen or antigen. Se deficiency (0.02 ppm for 8 wk) had the opposite effect. The alterations in the ability of the cells to proliferate, which occurred in the absence of changes in the endogenous levels of interleukin-2 (Il2) or interleukin 1, were apparently related to the ability of Se to alter the kinetics of expression of high-affinity Il2 receptors on the surface of activated lymphocytes. This resulted in an enhanced or delayed clonal expansion of the cells, and in an increased or decreased frequency of cytotoxic cells within a given cell population. The changes in tumor cytotoxicity were paralleled by changes in the amounts of lymphotoxin produced by the activated cells. Dietary Se modulations had a comparable effect on macrophage-mediated tumor cytodestruction. The results also suggested that Se exerts its effect 8-24 h after stimulation, and that it most likely affects processes in the cytoplasmic and/or nuclear compartments of activated lymphocytes.


Selenium deficiency in Yugoslavia.

Selenium deficiency in Yugoslavia. Data on selenium (Se) deficiency in Yugoslavia are presented. The results include Se content of soil, cereal crops, and garlic grown in these soils, and human serum and scalp hair from several towns and regions. All data indicate a serious Se deficiency: soil (n = 140), the mean value of 200 +/- 69.6 micrograms/kg Se; wheat, (58) mean = 20.5 +/- 12.4 micrograms/kg; corn, (79) mean = 13.7 +/- 13.6 micrograms/kg; and garlic, (66) mean = 13.7 +/- 17.1 micrograms/kg Se. Analyses of human tissue show a very low Se status of the Yugoslav population: serum, (n = 875) mean = 50.0 +/- 18.0 micrograms/L and scalp hair, (388) mean = 94 +/- 16 micrograms/kg Se. In some regions, Se contents of grain, garlic, and human serum and hair are approaching those in the low-Se belt in China. It is assumed that very low Se status of a human population could be a risk factor in the development of Balkan Endemic Nephropathy (BEN) and in a high incidence of urinary tract tumors (UTT) in endemic areas.


The role of selenium in thyroid hormone metabolism and effects of selenium deficiency on thyroid hormone and iodine metabolism. Selenium deficiency impairs thyroid hormone metabolism by inhibiting the synthesis and activity of the iodothyronine deiodinases, which convert thyroxine (T4) to the more metabolically active 3,3'-5 triiodothyronine (T3). Hepatic type I iodothyronine deiodinase, identified in partially purified cell fractions using affinity labeling with [125I]N-bromoacetyl reverse triiodothyronine, is also labeled with 75Se by in vivo treatment of rats with 75Se-Na2SeO3. Thus, the type I iodothyronine 5'-deiodinase is a selenoenzyme. In rats, concurrent selenium and iodine deficiency produces greater increases in thyroid weight and plasma thyrotrophin than iodine deficiency alone. These results indicate that a concurrent selenium deficiency could be a major determinant of the severity of iodine deficiency.


A contribution to the world selenium map. Atomic absorption spectrophotometric method was used to determine the serum selenium levels of 86 healthy individuals. Variations in age, sex, and geographically different urban regions of Yugoslavia were investigated. A group of 63 healthy children, ages 8-15 yr, were examined. Mean +/- standard deviation of the serum selenium concentration was 57 +/- 9 micrograms/L; age and geographic area had no effect on the Se status of children, but the difference between boys and girls was significant (P less than 0.05). A group of 23 men from Zagreb, ages 22-37 yr, were examined. The group was divided into three age subgroups and no difference was found among these groups. The mean Se concentration was 69 +/- 18 micrograms/L, and a statistically significant difference was found only between the group of adults and the group of children (P less than 0.05).


Monitoring circulating B cells in patients with multiple myeloma at diagnosis or in plateau phase: how prevalent is light chain isotype suppression? Peripheral blood lymphocytes from 25 patients with multiple myeloma at diagnosis or in plateau phase, and from 18 healthy subjects, were analysed for the surface expression of CD19 and immunoglobulin (Ig) light chain isotypes by dual parameter flow cytometry. The aim was to test for the existence of light chain isotype suppression (LCIS). LCIS is a hypothetical suppressor mechanism which supposedly operates in myeloma patients with stable disease, causing a drop in the number of circulating B cells bearing the same Ig light chain isotype as the myeloma paraprotein. Our experiments emphasized that the use of a non-Ig, pan-B cell marker, combined with dual antigen analysis, is necessary to distinguish Ig on B cells form (cytophilic) Ig on some non-B cells. We found good correlation between results obtained using polyclonal versus monoclonal anti-light chain antibodies, and washed blood versus peripheral blood mononuclear cells. There was no evidence of an abnormal ratio of CD19+, K+ to CD19+, lambda + cells in any of the patients. Thus, these patients did not manifest LCIS, and these experiments raise doubts about the existence of this phenomenon. The results also illustrate the problem that may arise in the interpretation of one parameter flow cytometry.