Effect of 3-isobutyl-1-methylxanthine and zaprinast on non-adrenergic non-cholinergic relaxation in the rat gastric fundus. Vasoactive intestinal polypeptide (VIP) and nitric oxide (NO) have been proposed as inhibitory non-adrenergic non-cholinergic (NANC) neurotransmitters in the rat gastric fundus. The smooth muscle relaxant actions of VIP and NO are medaited by cAMP and cGMP, respectively; therefore the effect of inhibitors of phosphodiesterases responsible for cyclic nucleotide breakdown on relaxation induced by VIP, NO and electrical field stimulation was investigated. The non-specific phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX), the cGMP-specific phosphodiesterase inhibitor, zaprinast, the adenylate cyclase activator, forskolin, and the cyclic nucleotide analog, 8-bromo cGMP, produced concentration-dependent relaxation of rat gastric fundus strips precontracted by PGF2 alpha. IBMX potentiated isoprenaline-induced relaxation but not relaxation induced by sodium nitroprusside, VIP, NO or electrical field stimulation. Zaprinast potentiated the relaxation induced by sodium nitroprusside, while having no influence on relaxation due to any other stimulus. The combination of both phosphodiesterase inhibitors did not significantly affect the electrically induced relaxation. It is concluded that both cAMP and cGMP mediate relaxation in the rat gastric fundus. Further research is needed to investigate the role of the cyclic nucleotides during NANC relaxation of this tissue.


Phorbol esters induce transient changes in the accessibility of the carboxy-terminal domain of nuclear lamin A. Treatment of human epithelial cells in culture with phorbol esters (TPA) gives rise to a transient and reversible loss of accessibility to antibodies of the nonhelical carboxy-terminal domain of nuclear lamin A that distinguishes it from lamin C. No change in the accessibility of epitopes present in the common domain of lamins A and C was observed. Loss of accessibility of lamin A was not due to proteolytic degradation nor to modification of the isoelectric point of lamin A and did not depend upon protein kinase C activation nor protein synthesis. Perturbation of desmosome organization by growth in low calcium blocked the effect of TPA on lamin A. Prolonged exposure to nocodazole, one of the effects of which is a perinuclear collapse of intermediate filaments, also blocked the effect of TPA on lamin A. These results suggest that the initial target of TPA may be at the level of cell-cell contacts and that the perturbation induced by TPA may be propagated via the structural link formed by intermediate filaments between the cell surface and the nucleus, giving rise to a change in conformation of the carboxy-terminal domain of lamin A or to an interaction of this domain with another nuclear component. These results form the basis for the hypothesis that the interphase nuclear lamina may play an active role in the process of mechanochemical signal transduction.


Alpha IIb beta 3 integrin expression and function in subpopulations of murine tumors. Subpopulations of B16 amelanotic melanoma (B16a) cells, isolated by centrifugal elutriation from enzymatically dispersed solid tumors, demonstrated different abilities to form lung colonies when injected intravenously. In contrast, no differences in experimental metastasis were observed among subpopulations obtained from Lewis lung (3LL) tumors. Lung colonization by B16a and 3LL subpopulations correlated positively with observed differences (B16a) or lack of differences (3LL) in tumor cell ability to induce aggregation of homologous platelets, to adhere to subendothelial matrix or fibronectin, and with the percentage of cells in the G2/M phase of the cell cycle. Both B16a and 3LL cells express alpha IIb beta 3 integrin receptors; however, differences in the receptor expression level were found only among B16a subpopulations. Comparison of the amount of alpha IIb beta 3 receptor expressed on cell surface with tumor cell ability to induce platelet aggregation (TCIPA) and to adhere to fibronectin or subendothelial matrix revealed a positive correlation. Pretreatment of tumor cells with alpha IIb beta 3-specific antibodies inhibited tumor cell matrix adhesion, TCIPA, and lung colony formation. We propose that alpha IIb beta 3 integrin receptor expression, tumor cell matrix adhesion, and tumor cell-induced platelet aggregation can be important parameters to indicate the metastatic potential of some tumor cells and that the alpha IIb beta 3 is a multifunctional receptor involved in both tumor cell-matrix and tumor cell-platelet interactions. Further, the correlation among cell cycle phase, metastatic ability, and receptor expression suggests that metastatic propensity may be transiently expressed and/or increased in some tumor cell subpopulations.


Immunocytochemical analysis reveals differences between the subcellular localization of normal and delta Phe508 recombinant cystic fibrosis transmembrane conductance regulator. Cystic fibrosis (CF) is caused by mutations in the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR). The most common mutation responsible for CF is the deletion of amino acid residue Phe508, with an average allelic frequency of 70%. We have isolated an anti-CFTR monoclonal antibody which specifically recognizes recombinant normal and delta Phe508-CFTR produced by a vaccinia virus expression system. Immunocytochemical analysis of L cells expressing either normal or delta Phe508-CFTR showed a marked difference in subcellular distribution. Normal CFTR had a distinct localization in the perinuclear area and was also associated with the plasma membrane. delta Phe508-CFTR essentially lacked the membrane-associated distribution and was present throughout the cytoplasm. This heterologous expression system thus provides a model system for studying the subcellular localization of different mutant forms of CFTR.


Increased expression of cytokeratin and ferritin-H genes in tumorigenic clones of the SW 613-S human colon carcinoma cell line. Subclones of the SW 613-S human colon carcinoma cell line differ by their ability to induce tumors in nude mice and by their level of amplification of the c-myc gene. Clones with a high level of amplification are tumorigenic in nude mice whereas those with a low level are not. Genes overexpressed in the tumorigenic clones as compared to the nontumorigenic ones were searched by differential screening of a cDNA library. Two cDNA clones corresponding to cytokeratin K18 and ferritin-H chain were isolated. The steady state level of the corresponding mRNAs is higher in cells of all tumorigenic clones. The level of cytokeratin K8 mRNA, the specific partner of cytokeratin K18 in intermediate filaments of epithelial cells, is also elevated in these cells. For all three genes, this is mainly due to an increase in the transcription rate, as shown by a nuclear run-on assay. Immunoblotting experiments showed that cytokeratins K8, K18, and K19 are more abundant in cells of tumorigenic clones. The mRNA of the other subunit of apo-ferritin (ferritin-L chain) is expressed at the same level in both types of clones. The mRNAs of cytokeratins K18 and K8 and of ferritin-H chain are also overexpressed in cells of nontumorigenic clones which have acquired a tumorigenic phenotype after transfection of c-myc gene copies.


The Ca2+ release channel in junctional sarcoplasmic reticulum: gating and blockade by cations. 1. By using a sarcoplasmic reticulum preparation containing feet structures and the 45Ca2+/filtration technique, the opening and closing response of the Ca(2+)-channel was studied. 2. Extravesicular Sr2+ can activate the channel even though this cation is less efficient than Ca2+ in stimulating the Ca2+ release. Higher Sr2+ concentrations display inhibitory action. 3. By studying the closing response high- and low-affinity cations can be distinguished, according to the concentration range required to exert their effect. 4. The synergistic behavior observed by combining high- and low-affinity blocking cations suggest that they interact through the same binding site. 5. The high-and low-affinity cations are noncompetitive blockers of the activating Ca2+ suggesting the existence of an inhibitory site which is different to the activating site.


Nutrition and somatomedin XXIX. Molecular regulation of IGFBP-1 in hepatocyte primary culture. The insulinlike growth factors (IGFs) circulate in association with insulinlike growth factor binding proteins (IGFBPs) that modulate IGF action, but mechanisms of IGFBP regulation are poorly understood. We investigated the regulation of IGFBPs in primary cultures of rat hepatocytes, measuring the appearance of export proteins by ligand blotting after separation via SDS/PAGE, and evaluating mRNA with cDNA probes. Northern blotting studies revealed that IGFBP-1 was expressed at high levels in cultured hepatocytes, in which sustained release of both insulinlike growth factor I and albumin marks preservation of differentiated status. In contrast, transcripts of IGFBP-3 and IGFBP-2 were not detected. Release of IGFBP-1 was unaffected by exposure to glucose (20-500 mg/dl) or to provision of amino acids (0.25-6.25 times normal rat arterial plasma levels). Hormonal studies revealed little effect of glucagon, inhibition by insulin, stimulation by dexamethasone, and blunting of dexamethasone effects by added insulin. Adding dexamethasone provided progressive stimulation: 5-, 11-, and 26-fold at 10(-9), 10(-8), and 10(-7) M, all P less than 0.01; increases in IGFBP-1 protein (ligand blot) and IGFBP-1 mRNA (Northern blot) were highly correlated (r = 0.62, P less than 0.001). In contrast, adding insulin resulted in progressive suppression of both IGFBP-1 protein and IGFBP-1 mRNA, 43% at 10(-10) M, 74% at 10(-9) M, and 83% (maximal) at 10(-8) M; ED50 of approximately 10(-10) M is within the physiological range of insulin concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)


Experimental diabetic neuropathy. Effect of ganglioside treatment on axonal transport of cytoskeletal proteins. Abnormalities in axonal transport of proteins are thought to play an important role in the pathogenesis of diabetic neuropathy. Gangliosides exert a positive action on numerous alterations in biochemistry and physiology of diabetic nerves. This study was undertaken to assess the effects of exogenous gangliosides on the axonal transport of structural proteins such as actin and tubulin in the sensory fibers of short-term (9-wk) and long-term (6-mo) diabetic rats. Adult Sprague-Dawley rats were made diabetic with a single injection of 70 mg/kg streptozocin i.p. Subgroups were injected daily with either highly purified ganglioside mixture (10 mg/kg i.p.) or saline for 1 mo, beginning either 2 or 17 wk after streptozocin injection. Age-matched rats were used as controls. Axonal transport was studied by the pulse-labeling technique. Three weeks after labeling, sciatic nerves were dissected out and processed for sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography. In diabetic rats of both experimental designs, the transport rate of tubulin and actin was decreased by approximately 30% compared with control rats. Ganglioside treatment counteracted such alterations in both 9-wk and 6-mo diabetic rats. These data suggest a pharmacological effect that could be correlated with molecular interactions between integral membrane glycolipids and cytoskeletal elements.


Non-verbal communication skills of surgically treated children with infantile spasms. The authors present preliminary findings on the effects of surgery on the development of early non-verbal social communication skills in eight children with intractable infantile spasms. After a mean follow-up of 15.2 months, there was no statistically significant change in the post-surgical non-verbal communication behavior of these children beyond the expected developmental change. Implications of these findings for the developmental impairment associated with infantile spasms are discussed.


Transient increase in renal insulin-like growth factor binding proteins during initial kidney hypertrophy in experimental diabetes in rats. The insulin-like growth factors, insulin-like growth factor I and insulin-like growth factor II are bound to six distinct classes of insulin-like growth factor binding proteins (IGFBPs) in the circulation and in extracellular fluids. Diabetic renal hypertrophy is preceded by a transient increase in kidney insulin-like growth factor I suggestive of a renotropic function for insulin-like growth factor I. In order to examine a possible involvement of IGFBPs in initial diabetic kidney growth and in kidney insulin-like growth factor I accumulation, we studied rat kidney IGFBPs by ligand blotting during the first 4 days after induction of diabetes. Six distinct bands were identified in kidney and liver tissue with apparent molecular weight values of 38-47 (doublet), 34, 30, 24 and 20 kDa. The 38-47 kDa doublet band probably corresponds to the insulin-like growth factor binding subunit of IGFBP-3, the 24 kDa band to IGFBP-4 and the 30 kDa band to IGFBP-1 and/or IGFBP-2, as these IGFBPs in rats have similar molecular weight. In untreated diabetic rats a transient increase in the kidney 30 kDa band was demonstrable 24 h after induction of diabetes with a maximal rise (two-fold) after 48 h, followed by a decrease to baseline values after 4 days. In untreated diabetic rats the 38-47 kDa doublet band also increased (two-fold) in kidney during the first 2 days after induction of diabetes, followed by a subsequent decrease. Insulin-treatment prevented both the increase in the 30 kDa and in the 38-47 kDa bands.(ABSTRACT TRUNCATED AT 250 WORDS)


ATP-dependent bacterial transporters and cystic fibrosis: analogy between channels and transporters. The traffic ATPases superfamily includes known transporters, both prokaryotic and eukaryotic, including the medically important proteins, P-glycoprotein, and the cystic fibrosis gene product (CFTR), which is known to be a Cl- channel. The structure and mechanism of action of the best-studied members of the superfamily, the periplasmic permeases, are described and related to that of CFTR and eukaryotic traffic ATPases in general. The contention is put forward that the distinction between the architecture and mechanisms of action of channels and transporters is blurred.


Long-term culture of normal human colonic epithelial cells in vitro. Studies of normal cellular function as well as the understanding of cellular mechanisms of carcinogenesis and other diseases of the large intestine have been limited, particularly due to the lack of long-term culture of normal human large intestinal epithelial cells (NHLIEC). Using the epithelia from surgically resected human colon, we have dissociated a sufficient number of viable NHLIEC and maintained them in in vitro culture for up to 5 months. Normal-appearing human large intestinal mucosal fragments (1 mm2) were treated with 0.01 mg/ml trypsin, 0.2 mg/ml collagenase + 0.1 mM EGTA or 0.1 mg/ml trypsin + 0.1 mM EGTA in a Stomacher laboratory blender to isolate the cells. Compared with other methods, the use of the Stomacher blender combined with low concentrations of proteolytic enzymes yielded greater numbers of cells per gram of tissue, with up to 84% viable cells. Primary and serially passaged NHLIEC were cultured in CMRL-1066, MEM with 5% serum, and serum-free KGM. These media were all supplemented with insulin, hydrocortisone, epithelial growth factor, and bovine pituitary extract. CMRL-1066 was found to be the best medium for NHLIEC. Contaminating fibroblasts were selectively removed by briefly allowing the cells to adhere to the culture vessel and adding 25 U/ml collagenase to the culture media at the first subculture treatment. The epithelial nature and secretory function of the established cells were confirmed by morphological criteria (light microscopy, phase contrast microscopy and electron microscopy), immunoreactivity to cytokeratin, and positive mucin cytochemistry. We propose that using this methodology for the culture and maintenance of NHLIEC for an extended period of time would serve as a valuable model for a variety of investigations.


Fine specificity of the human T-cell response to the hepatitis B virus preS1 antigen. The T-cell response to hepatitis B virus envelope antigens was studied in 11 hepatitis B vaccine recipients; 7 were selected to analyze the fine specificity of the T-cell response to the preS1 antigen. Four distinct T-cell epitopes were identified by peripheral blood lymphomononuclear cell stimulation with a panel of short synthetic peptides covering the preS1 sequence. The immunodominance of the preS1 epitopes included within peptides 21-30 and 29-48 was shown by their capacity to restimulate an HLA class II restricted proliferative response of T cells primed with the whole preS1 antigen. Conversely, peptide-specific T cells selected by peripheral blood lymphomononuclear cell stimulation with peptides 21-30 and 29-48 were able to recognize the native preS1 molecule, confirming that these epitopes are actually generated by the intracellular processing of preS1. Finally, amino acid residues essential for T-cell activation by peptide 21-30 were identified using 10 analogues of the stimulatory peptide containing single alanine substitutions. These results may be relevant to the design of efficient synthetic vaccines against hepatitis B virus infection.


Recurrent and acquired hepatitis C viral infection in liver transplant recipients. To examine the postliver transplant recurrence of hepatitis C virus (HCV) infection in patients with pretransplant infection, as well as its acquisition in patients without prior infection, we used the polymerase chain reaction to amplify HCV RNA in serum and/or liver samples of 89 patients with alcoholic and cryptogenic cirrhosis undergoing liver transplantation. Results were correlated with histologic findings from posttransplant liver biopsies. Ninety-five percent of patients with pretransplant infection had posttransplant viremia. In contrast, 35% of patients without pretransplant infection acquired the virus (P less than 0.0001). Pretransplant HCV infection predisposed patients to hepatitis in the new graft. HCV RNA was present in serum of 96% of patients with posttransplant hepatitis. Fifty-six percent of patients with posttransplant HCV infection had no evidence of liver damage at least 1 year posttransplant. However, of those patients with histologic hepatitis, chronic active hepatitis was common. It is concluded that although HCV infection recurs posttransplant in almost all infected patients, acquisition of the HCV infection with transplant is common. Pretransplant HCV infection is an independent risk factor for the development of posttransplant hepatitis. HCV infection accounts for the majority of posttransplant hepatitis not due to cytomegalovirus, and although many patients with posttransplant viremia have little evidence of histologic hepatitis, significant hepatic damage may occur.


[Continuous ambulatory intravenous morphine infusion for pain therapy in advanced ovarian cancer]. We report on a female outpatient with cancer of the ovary, who received continuous intravenous morphine infusion for terminal pain control. The patient was treated over a period of 48 days with a morphine dosage ranging from 10 to 60 mg/h, which kept her free of pain. With treatment, she was alert, communicative with her relatives and moved freely. At a later stage, we complemented the treatment with Diazepam and Haloperidol. No side-effects were observed over the whole period of morphine infusion.


Streptomycin biosynthesis and its regulation in Streptomycetes. New insights into the gene orders, structures, evolution, and functions of streptomycin (Sm) biosynthetic genes (str) were gained via hybridization studies, determination of nucleotide sequences, and measurement of expression in the str gene clusters of Streptomyces griseus and S. glaucescens. Both str clusters showed considerable divergence in macro and micro structure. Genes putatively involved in pathways leading to the (dihydro-)streptose and N-methyl-L-glucosamine moieties of Sm were identified. Additional regulatory elements, such as gene strS and conserved TTA codons in the N-terminal sections of reading frames, are reported. Evidences for the involvement of physiological state, signal transduction, and activators in the control of Sm production are presented.


Effect of Helicobacter pylori infection on colloidal bismuth subcitrate concentration in gastric mucus. Necropsy gastric mucus infected with Helicobacter pylori has a reduced capacity to concentrate colloidal bismuth subcitrate when compared with non-infected mucus. Mucus mounted in a modified in vitro diffusion chamber was bathed with colloidal bismuth subcitrate solutions at different concentrations and pH levels. Bismuth was measured by atomic absorption spectrophotometry to assess intramucus colloidal bismuth subcitrate concentrations. Bismuth concentrations in non-infected mucus were higher than in Helicobacter pylori infected mucus at all experimental colloidal bismuth subcitrate concentrations and pH levels. Regardless of the infection status, the intramucus concentration of colloidal bismuth subcitrate was dependent upon the concentration of the bathing solution and independent of the pH and the mucus thickness. Colloidal bismuth subcitrate solubility in saline solution varied with pH, and was least soluble in the pH range 1.1 to 3.25 and more soluble above and below this pH range. This study suggests that Helicobacter pylori infection is associated with physicochemical changes in the gastric mucus with a reduction in its capacity to concentrate colloidal bismuth subcitrate. Such a reduction may compromise the attainment of optimum colloidal bismuth subcitrate concentrations necessary for its bactericidal activity.


Differences in the mode of the extension of gastric cancer classified by histological type: new histological classification of gastric carcinoma. By combining two of the morphological characteristics of gastric cancer, the degree of differentiation of the glandular tubules and the amount of mucus in the cytoplasm, the histological type of the gastric carcinoma was categorised into four groups. Group I: tubular differentiation--well; mucus in cytoplasm--poor; group II: tubular differentiation--well; mucus in cytoplasm--rich; group III: tubular differentiation--poor; mucus in cytoplasm--poor; group IV: tubular differentiation--poor; mucus in cytoplasm--rich. A study of the relation between the types of primary lesion and the mode of extension and recurrence of gastric carcinoma in 200 autopsy cases was then undertaken. In group I, the frequency and extent of haematogenous metastasis such as in the liver was high, while in group IV, that of lymph node metastasis, direct invasion into surrounding organ, and peritoneal dissemination were higher. In group III, which showed the intermediate mode of extension in nature to those of group I and IV, although the frequency and severity of the bone marrow metastasis was the highest. There were significant differences in the modes of development and the extent of infiltration in all groups.


Treatment of malignant esophageal obstruction with silicone-coated metallic self-expanding stents. Six patients with high-grade malignant esophageal obstruction were treated with silicone-coated metallic self-expanding esophageal stents (Z stents). Endoscopic placement of stents was well tolerated. All patients achieved excellent palliation, defined by a decrease of at least two dysphagia grades, which was sustained. Complications occurred during follow-up in four patients and included stent migration, silicone disruption with tumor ingrowth, food impaction, and perforation (discovered at autopsy) at the distal stent site. Three of the four complications were promptly treated by endoscopic or radiologic intervention. Recent modification in stent design and placement technique will hopefully reduce complications. The self-expanding stent has several theoretical advantages over the rigid plastic stent and Nd:YAG laser for palliation of obstructing esophageal malignancy.


Effective peritoneal therapy of acute pancreatitis in the rat with glutaryl-trialanin-ethylamide: a novel inhibitor of pancreatic elastase. The six hour peritoneal lavage with glutaryl-trialanin-ethylamide, a low molecular competitive inhibitor of pancreatic elastase (IC50-8 mumol/l), effectively suppresses the evolution of taurocholate induced acute pancreatitis in the rat. The lavage alone is followed by a marked decrease of fat necrosis and amylase and lipase activity in serum. The area of pancreatic haemorrhage was significantly reduced only after the lavage solution was supplemented with Glt-Ala3-NHEt. The effect was not enhanced by a bolus injection of the inhibitor before starting the lavage. The combination of Glt-Ala3-NHEt with aprotinin or nafamstate mesilate produced only marginal greater benefit. The effect of Glt-Ala3-NHEt on pancreatic haemorrhage is time and dose related even with delayed onset of the lavage. Animals treated with peritoneal lavage without Get-Ala3-NHEt lived longer than controls (p less than 0.05), but by 60 hours the survival rate of both groups was almost the same (76 v 74%). All animals lavaged with Glt-Ala3-NHEt survived 120 hours and the difference in the survival rate between this and both remaining groups was significant (100% v 76% v 74% - p less than 0.05). The results were considered favourable and preliminary clinical trials of Glt-Ala3-NHEt in subjects with acute pancreatitis justified.


Dual sequential non-cross-resistant chemotherapy for advanced stage squamous cell carcinoma of the cervix. Forty-three patients with recurrent and metastatic squamous cell carcinoma of the cervix received a program of sequential combination chemotherapy incorporating induction with cisplatin, vinblastine, and bleomycin (PVB) and subsequent consolidation with 5-fluorouracil, doxorubicin, cyclophosphamide, and vincristine (FACV). The overall response rate was 62% and 10 patients (23%) achieved complete remission. Response status was improved in 11 patients at the completion of FACV after initial PVB therapy, including 9 complete remissions. The median survival for all patients in the study was 38 weeks and 50 weeks for the responding patients. Myelosuppression was the principal toxicity encountered and 10 episodes of neutropenic sepsis occurred, including one septic death. However, the only cumulative toxicity was peripheral neuropathy, although this was only moderate to severe in 2 patients. These results are encouraging, but will require confirmation in randomized comparison to cisplatin, presently accepted as standard single-agent therapy.


Conservative treatment of ectopic pregnancy and its effect on corpus luteum activity. Corpus luteum activity was monitored in 15 women undergoing nonsurgical management of ectopic pregnancy with local methotrexate injection followed by alternating oral methotrexate and citrovorum factor (group A, n = 8) or local methotrexate injection alone (group B, n = 7). All patients initially demonstrated a viable corpus luteum (plasma progesterone ranged from 1.4 to 19 ng/ml). The treatment was successful in 14, with the exception of one whose tube ruptured 11 days after local administration of methotrexate, despite a continuous decrease in beta human chorionic gonadotropin, 17 beta-estradiol and plasma progesterone levels. There seems to be no correlation between the success of the treatment and the behavior of beta human chorionic gonadotropin, 17 beta-estradiol and plasma progesterone. Three patients from group A and two from group B displayed an initial rise in beta human chorionic gonadotropin following the initiation of the therapy, but the corpus luteum response differed. In group B patients, 17 beta-estradiol and plasma progesterone levels increased in parallel with beta human chorionic gonadotropin. Group A patients displayed a continuous decrease in 17 beta-estradiol and plasma progesterone levels despite the elevation of beta human chorionic gonadotropin, suggesting a possible effect of the systemic methotrexate on corpus luteum activity.


Epitope specific antibody response against human type II collagen induced by anti-idiotypic antibody. A species specific epitope on human type II collagen (CII) recognized with mAb, 2-60, was found to be localized on a cyanogen bromide-cleaved peptide (CB10) of human CII. To characterize the antibody response to the 2-60 epitope, we raised rabbit anti-idiotypic (Id) antibody against 2-60. The rabbit anti-2-60 Id antibody reacted not only with 2-60 mAb but also with 2-56 and 3-11 mAbs which were reactive with the epitopes related to the 2-60 epitope on CB10. The anti-Id antibody inhibited the binding of these three mAbs to human CII. Thus, rabbit anti-2-60 Id antibody recognized the cross-reactive idiotype expressed on 2-60, 2-56 and 3-11. The anti-2-60 Id antibody inhibited about thirty percent of the binding of polyclonal anti-human CII antibody derived from DBA/1J mice, thereby suggesting that the 2-60 idiotype might be expressed on a major fraction of the anti-human CII antibody. Immunization with the rabbit anti-2-60 Id antibody elicited antibody response to the 2-60 epitope, in DBA/1J (H-2q, Ighc), BALB/c (H-2d, Igha) and DBA/2 (H-2d, Ighc) mice. On the other hand, an epitope-specific antibody response induced by rabbit anti-Id antibody to 1-5 mAb reactive with a putative arthritogenic epitope on human CII was shown to be influenced by a single dominant gene, possibly the Igh gene. Our findings suggest that antibody responses against two distinct epitopes on human CII are probably regulated by different mechanisms.


Microvessel quantitation and prognosis in invasive breast carcinoma. The prognostic significance of microvessel quantitation in invasive breast carcinoma was analyzed in a study group that comprised 88 patients with axillary node-negative carcinoma and 32 patients with axillary node-positive carcinoma who had a minimum follow-up period of 9 years. Microvessels were identified by immunohistochemistry using antibodies to endothelial markers, including factor VIII-related antigen and blood group isoantigens (ABH). Factor VIII-related antigen staining provided more consistent results for microvessel quantitation than did staining for ABH isoantigens. The three most vascular areas within a tumor were selected, and the microvessels within a x200 microscopic field of each area were counted by two investigators simultaneously. Node-positive carcinomas demonstrated significantly higher microvessel counts than did node-negative carcinomas (mean +/- SD, 99 +/- 42 and 73 +/- 22, respectively; P less than .001). In node-negative carcinomas, tumors from patients who experienced distant recurrence had higher microvessel counts than did tumors from patients who were disease-free (84 +/- 19 and 70 +/- 22; P = .01). Similarly, in patients with node-positive carcinoma, microvessel counts were considerably higher in tumors from patients who experienced distant recurrence than in patients who did not, although the difference did not reach statistical significance (113 +/- 44 and 93 +/- 34, respectively). Among patients with node-negative carcinoma, those with a microvessel count of less than 84 had a recurrence rate of 20% compared with 57% in patients with counts greater than 84 (P = .003). Microvessel counts were independent of histologic parameters, ploidy status, and S-phase fraction but correlated with peritumoral vascular invasion. Both microvessel counts and vascular invasion were independent prognostic parameters by multivariate analysis. High vessel counts may represent increased tumor angiogenesis and are correlated with tumor aggressiveness. Microvessel quantitation may be an additional prognostic factor that, when used in conjunction with more established parameters, can help in appropriate patient management.


CD56 (NKH1)-positive hematolymphoid malignancies: an aggressive neoplasm featuring frequent cutaneous/mucosal involvement, cytoplasmic azurophilic granules, and angiocentricity. CD56 (NKH1) expression is a rare phenomenon in malignant lymphomas, mostly confined to those occurring in the nasal or nasopharyngeal region. In this study we provide a detailed clinicopathologic analysis of nine patients with CD56-positive hematolymphoid malignancies occurring in sites other than the upper aerodigestive tract. The disease occurred predominantly in young and middle-aged men (mean age, 40 years) who often presented with swinging fever, skin rash, and/or hepatosplenomegaly, usually in the absence of peripheral lymphadenopathy. There was frequent involvement of the skin and mucosal sites, such as the salivary gland, lungs, and small intestine. The disease pursued a highly aggressive course, with most patients dying within weeks despite cytotoxic therapy. Although the cytologic appearances and immunophenotypic profile varied from case to case, the group of tumors could be unified by two morphologic features, namely, the presence of azurophilic granules in the cytoplasm of the neoplastic cells and the frequent occurrence of angiocentric and angiodestructive infiltrates. Since CD56 reactivity appears to confer a poor prognosis in hematolymphoid malignancies, we recommended inclusion of CD56 antibody in the routine panel for immunophenotypic analysis.


Alpha-fetoprotein-producing acinar cell carcinoma of the pancreas. A pancreatic carcinoma and liver metastases associated with marked elevation of the serum alpha-fetoprotein (AFP) level were resected from a 57-year-old man. On microscopic examination, the tumor cells showed a predominantly acinar arrangement, with tubular and trabecular structures; in some foci it had features of a medullary pattern. Alpha-fetoprotein, lipase, trypsin, chymotrypsin, and alpha 1-antitrypsin were strongly demonstrated in tumor tissue by immunohistochemical techniques. A biochemical analysis of AFP on affinity sepharose columns revealed that the AFP derived from the tumor tissues was similar to that of hepatocellular carcinoma. Ultrastructural study showed that most of the tumor cells had abundant rough endoplastic reticulum and numerous zymogen granules. No squamoid corpuscles, neuroendocrine granules, bile production, or bile canaliculi were recognized. These findings suggest that this unique tumor originated from acinar cells.


Expression and chromosome localization of the murine cystic fibrosis transmembrane conductance regulator. A 13.5-kb genomic fragment of the mouse cystic fibrosis transmembrane conductance regulator (CFTR) gene was isolated from a C57BL/6J liver DNA library, using a human CFTR exon 10 probe. This region of the human gene includes the most common cystic fibrosis mutation (deletion of the Phe508 residue) in the first nucleotide binding domain of CFTR. Sequence analysis demonstrated 87% identity between the predicted mouse and the normal human CFTR exon 10 sequences, including conservation of the Phe508 residue. Northern analysis revealed that the mouse gene is expressed in intestine, lung, stomach, kidney, and salivary gland. In contrast to human CFTR, murine CFTR transcripts were not detectable by Northern analysis in the liver or pancreas. More sensitive PCR analysis, however, revealed that the mouse CFTR gene is weakly expressed in other tissues, including liver and pancreas. During development, mouse CFTR transcripts were observed as early as Embryonic Day 13. Southern analysis of mouse x Chinese hamster somatic cell hybrid DNAs mapped the mouse CFTR locus (Cftr) to Chromosome 6 (Chr 6). Subsequent typing of the progeny of an interspecies backcross revealed that Cftr is closely linked to the proto-oncogene c-met locus (Met) in the centromeric region of mouse Chr 6, consistent with the observation that there is a conserved chromosomal segment on human chromosome 7 and mouse Chr 6.


Regional assignment of the human keratin 5 (KRT5) gene to chromosome 12q near D12S14 by PCR analysis of somatic cell hybrids and multicolor in situ hybridization. Keratin 5 is the major type II keratin of the basal cells of epidermis and of other stratified epithelia. With its type I partner, keratin 14, it constitutes a major fraction of the cytoskeleton of the basal cells. Because the inheritance of epidermolysis bullosa simplex, a disease of epidermal basal cell fragility, was mapped in one family to chromosome 12q close to D12S14, we undertook to localize the gene for keratin 5. Polymerase chain reaction analysis of somatic cell hybrids mapped the keratin 5 gene to chromosome 12, and multicolor fluorescence in situ hybridization localized it to 12q very near D12S14. This sublocalization exemplifies the utility of in situ physical localization in assessing the candidacy of genes thought to underlie inherited disorders.


The interferon- and virus-inducible IFI-56K and IFI-54K genes are located on human chromosome 10 at bands q23-q24. IFI-56K and IFI-54K are two human genes that are strongly induced by interferon and viruses. These genes are closely related at the protein, RNA, and promoter levels. By means of the somatic cell hybrid technique, the two genes have been previously located on chromosome 10. Using in situ hybridization, we show here that both IFI-54K and IFI-56K genes map to 10q23-q24. This result does not confirm the previous localization of the IFI-56K gene at the junction of the 10q25 and 10q26 bands.


Assignment of the gene coding for the alpha 2-macroglobulin receptor to mouse chromosome 15 and to human chromosome 12q13-q14 by isotopic and nonisotopic in situ hybridization. The assignment of the gene encoding the alpha 2-macroglobulin receptor (A2MR), which was first described as the low-density lipoprotein receptor-related protein, was confirmed by nonisotopic and isotopic in situ hybridizations on normal human metaphases to the region 12q13-q14. The same human cDNA, which has 95% sequence identity with the mouse A2mr, was hybridized to metaphases containing the Robertsonian translocation Rb(6;15)1Ald. The mouse A2mr gene was assigned to chromosome 15 in the region B2-D1. This locus and other loci on mouse chromosome 15 have been shown to be homologous with loci on human chromosome 12q.


[A new, highly synergistic drug combination for the treatment of infections with multiresistant mycobacteria, especially the mycobacterium avium complex]. Rationally designed combinations of rifampicin/thiacetazone plus isoniazid and/or ethambutol are highly effective in the treatment of patients (including HIV-pos.) infected with multiply resistant mycobacteria of the M. avium complex (MAC). Clinical results are very promising. The high efficacy of these combinations is due to the synergistic potentiation of single drug activities. As soon as rifabutin is marketed it should replace rifampicin in the combination treatment of patients with highly rifampicin-resistant MAC bacteria.


Direct demonstration of binding of aggregated mouse IgG1 to the 40-kDa Fc receptor for IgG (Fc gamma RII) in both high and low responders. Several directly fluorochrome-conjugated murine monoclonal antibodies (mAb) of the IgG1 subclass and directed against B or T cell antigens were found to bind to monocytes via the 40-kDa Fc receptor for IgG (Fc gamma RII). As expected from the established polymorphism of Fc gamma RII, strong staining was observed in about 75% of individuals. In the remaining 25% staining was clearly weaker, but could be definitely demonstrated with a mAb against the B cell-specific CD19 differentiation antigen. Specificity of binding to Fc gamma RII was confirmed by the ability to block the binding of the CD19 mAb by pre-incubation with aggregated IgG1 or with mAb against Fc gamma RII. The extent of T cell proliferation induced with a CD3 mAb of the IgG1 isotype (a-Leu 4), which is dependent on the interaction of monocyte Fc gamma RII with the Fc portion of the CD3 mAb, exactly correlated with the amount of binding to Fc gamma RII in all individuals. Proliferation was dose-dependent for both high and low responders; cells of low responders did not proliferate at concentrations below 16 ng/ml of mAb, whereas there was a small but unequivocal proliferation at higher concentrations. These results confirm that monocytes from previously characterized "non-responders" are able to bind aggregated murine IgG1 via Fc gamma RII. They also demonstrate that directly labeled mAb can cause extensive nonspecific staining which may not be excluded by the use of control antibodies of the same isotype.


A new approach to the cloning of genes encoding T-cell epitopes. The molecular structure of antigens recognized exclusively by T cells, such as minor histocompatibility antigens and some antigens that provoke autoimmune responses, has proved difficult to determine. Recently, several antigens induced on tumor cells by mutagen treatment have been cloned by transfection of genomic DNA libraries into P1.HTR cells, screening for antigen expression using T-cell clones, and subsequent recovery of the integrated DNA by cosmid rescue. We have modified this technique and have stably transfected P1.HTR cell lines with polyoma T antigen, which allows episomal replication of the shuttle vector, pCDM8. Using pCDM8-CAT constructs, we have determined the frequency of transfection and plasmid copies taken up per cell under optimal transfection conditions. Using a pCDM8 construct which expresses the tumor-specific antigen, P91A (pCDM8-tum-), that is recognized by a T-cell clone, we have found that cells transfected with this antigen can be recognized by the T-cell clone when they are present at only 1%-3% of a mixed population. Progeny of a single cell transfected with pCDM8-tum-: pCDM8-CAT at proportions of 1:10, 1:25, and 1:50 are recognized by the T-cell clone. Furthermore, Hirt extracted plasmid DNA from transfectants expressing the tum- antigen can be amplified in bacteria, transfected back into P1.HTR recipients, and recognized by the T-cell clone. This approach should enable reasonably rapid screening of cDNA libraries for even relatively low abundance messages encoding, for example, minor histocompatibility and alloantigens, and allow their subsequent cloning.


Genomic organization and chromosomal location of the human gene encoding the B-lymphocyte activation antigen B7. The human B lymphocyte activation antigen B7 provides regulatory signals for T lymphocytes as a consequence of binding to its ligands CD28 and CTLA-4. The cDNA for B7 has previously been isolated and predicted to encode a type I membrane protein. The predicted polypeptide has a secretory signal peptide followed by two contiguous Ig-like domains, a hydrophobic transmembrane region and a short cytoplasmic tail. Here we report the exon-intron genomic organization of human B7 and the chromosomal location. The gene has six exons that span approximately 32 kilobases of DNA. Exon 1 is not translated and the second exon contains the initiation ATG codon and encodes a predicted signal peptide. This gene structure is characteristic for several eukaryotic genes with tissue-specific expression. The third and fourth exons correspond to two Ig-like domains whereas the fifth and sixth exons encode respectively the trans-membrane portion and the cytoplasmic tail. This close relationship between exons and functional domains is a characteristic feature of genes of the Ig superfamily. Cell surface expression of the B7 gene product has previously been mapped to human chromosome 12 by antibody reactivity with the B7-specific monoclonal antibody BB-1. We here demonstrate that the B7 gene is located to the q21-qter region of chromosome 3 by DNA blot analysis of human x rodent somatic cell hybrids.


Cytologic diagnosis of tuberculous lymphadenitis in children by fine needle aspiration. One hundred consecutively diagnosed cases of tuberculous lymphadenitis in children have been analysed retrospectively. All cases were stained by May Grunwald Giemsa for cytomorphology and Ziehl Neelsen stain for acidfast bacilli (AFB). In 52 cases the material was sufficient and AFB cultures were possible. A diagnosis of tuberculosis was made when smears showed epithelioid cell granulomas or AFB on either smears or culture. M. tuberculosis and atypical mucobacteria were cultured in 26 and 3 cases respectively. In 6 cases the diagnosis of tuberculosis would have been missed but for culture studies, the cytologic smears were necrotic and stains for AFB negative.


Play therapy with young children.

Play therapy with young children. This article sets out to describe and evaluate the use of Play-Therapy with young handicapped children. The children, mostly of pre-school age, have been seen at the Wolfson Centre in London, over the last five years. The work is on-going. The developmental programme adapted and followed, was designed at the Centre in the 1970s, by Cooper, Moodley & Reynell. It was used to teach children with specific language delays. It has proved ideal to use with children with a variety of problems in their learning, also those with developmental delay. The teaching/play sessions have taken place in the children's homes and in the Wolfson Centre. Parental involvement has been important for success of the programme. Other professionals beside the teacher have been involved at the Centre, for monitoring the children's progress. The results shown in the annual reviews, or when the child attends for formal assessment, are encouraging. They point to the conclusion that this form of therapy can not only help the child at home, but also helps when he/she starts school.


Mapping of a visceral leishmaniasis-specific immunodominant B-cell epitope of Leishmania donovani Hsp70. We have shown that a member of the 70-kDa heat shock protein (Hsp70) family is a major target of the humoral immune response during Leishmania donovani infection. A recombinant fusion protein was recognized by sera from 92% (35 of 38) of patients with visceral leishmaniasis, including representatives from each of the major regions where it is endemic. Serological analysis of recombinant Hsp70, expressed by a series of deletion constructs, identified the carboxy-terminal region as the immunodominant site. This region, which is the most evolutionarily divergent part of the molecule, was recognized by all sera from patients with visceral leishmaniasis which exhibited an anti-Hsp70 response. Purified recombinant L. donovani Hsp70 was not recognized by sera from patients with cutaneous leishmaniasis, Chagas' disease, leprosy, malaria, or schistosomiasis. To determine the regions involved in antibody recognition, a series of overlapping peptides were synthesized on polyethylene pins by the Pepscan method, and a hexamer, EADDRA, was identified by the visceral leishmaniasis serum samples as an immunodominant B-cell epitope.


Immunological characterization of the lipooligosaccharide B band of Bordetella pertussis. Two structurally and immunologically different components of Bordetella pertussis endotoxin can be visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining: a major A band and a faster-migrating minor B band. Certain mutant strains of B. pertussis express only the B band, while the wild-type strains produce both lipooligosaccharides (LOS). Two monoclonal antibodies (MAbs) directed against the minor LOS B band were generated, allowing the study of this surface molecule on different strains of Bordetella. These two MAbs, designated BL-8 and BL-9, reacted strongly with phenol-water-purified LOS obtained from a B. pertussis LOS B mutant strain. Sodium periodate treatment of the purified LOS prevented binding of the MAbs, indicating the carbohydrate nature of the epitope(s). Western immunoblotting experiments revealed that the epitope(s) recognized by these MAbs is conserved on all B. pertussis and Bordetella bronchiseptica Vir- (avirulent) variant strains tested but is not present on Bordetella parapertussis and B. bronchiseptica Vir+ (virulent) wild-type strains. Further studies showed that although present in the lipopolysaccharide B band expressed by Vir- strains, the epitope(s) recognized by the MAbs is not accessible on the surface of intact B. bronchiseptica cells. For B. pertussis, the density and accessibility of this epitope(s) are dependent on the virulence-associated or LOS phenotype expressed by the strain. Our data demonstrate that the expression and accessibility of the epitope(s) are significantly greater on the LOS B variant strains and LOS AB Vir- strains compared with fresh B. pertussis clinical isolates. For these latter strains, which are Vir+, this epitope(s) was barely detectable on the surface of intact bacteria, despite Western blot analyses that revealed specific reactions between the MAbs and the LOS B band. The two LOS B-specific MAbs had no bacteriolytic activity against a LOS AB wild-type strain, while the control MAb BL-2, which is specific for the B. pertussis LOS A band, significantly reduced the number of living bacteria in the same assay. Moderate lytic activity against a mutant strain expressing only the LOS B band was observed for MAb BL-8 but not for MAb BL-9 or BL-2. These data demonstrate that the type, amount, and surface exposure of the LOS are related to the phenotype expressed by a specific B. pertussis strain. In addition, the LOS B MAbs also reveal the antigenic conservation of carbohydrate epitopes among B. pertussis and B. bronchiseptica strains.


Epitope mapping of hemagglutinating adhesion HA-Ag2 of Bacteroides (Porphyromonas) gingivalis. Thirteen monoclonal antibodies (MAbs) directed against hemagglutinating adhesion HA-Ag2 of Bacteroides (Porphyromonas) gingivalis were produced by immunizing mice with the relevant immunoprecipitate from crossed immunoelectrophoresis (CIE). Crossed immuno-affinoelectrophoresis and hemagglutination experiments confirmed that our MAbs recognized a molecule able to bind erythrocytes and involved in the hemagglutination process. In immunoelectron microscopy, these MAbs labelled amorphous material as novel cell-bound appendages distinct from fimbriae. CIE experiments allowed differentiation of the MAbs according to reactivity with immunoprecipitates Ag2, Ag8a, and Ag8c, which define HA-Ag2. The epitopes recognized by nine MAbs were mapped on three main antigenic domains (I, II, and III) by competition experiments and further grouped according to chemical composition and distribution on CIE immunoprecipitate. Domain I, defined by two MAbs, comprises an epitope with protein and carbohydrate determinants and distributed on Ag2 only. Epitopes of domain IIA, defined by four MAbs, are distributed on Ag8a, Ag8c, and Ag2 and are essentially composed of protein determinants but also have carbohydrate determinants that enhance the binding of the MAbs but are not essential. Epitopes of domain IIB, defined by two MAbs, and of domain III, defined by a single MAb, have a composition similar to that of domain IIA epitopes but are distributed on Ag8a and Ag8c only. A competition enzyme-linked immunosorbent assay with serum from normal subjects and patients with periodontitis suggested that domain I is more immunogenic than domain II.


Development in severely visually handicapped children and visual therapy remediation. The motor and cognitive development of a visually handicapped child is delayed due to various constraints placed on learning by the handicap. The child requires a functional visual assessment, and assessment of mental & motor development. Based on these findings a total remediation programme needs to be worked out. This is individualised, integrated & includes stimulation of residual vision, optional use of residual vision and special measures to promote general development.


Amelioration of clinical severity through raised fetal hemoglobin in sickle cell anaemia. In the present study, the levels of fetal hemoglobin (HbF) in sickle cell anemia patients were compared with sickle cell trait, beta thalassemia major and control. The mean HbF levels in beta thalassemia major and sickle cell anemia were 51.62 and 19.63% respectively. However, when the amount of HbF was expressed in terms of gram hemoglobin per deciliter whole blood, the mean values were 2.88 and 1.81 respectively between the two groups, suggesting that the genetic mechanism controlling the different threshold levels of increased HbF in these disorders could probably be similar. The elevated. HbF level in sickle cell anemia along with moderate hematologic profile observed in the present study is suggested to provide amelioration of the clinical severity unlike in beta thalassemia major where despite raised HbF levels, the severe clinical implications are attributed to marked imbalance in the globin chain synthesis.


Expression of Streptococcus mutans gtf genes in Streptococcus milleri. The Streptococcus mutans glucosyltransferase (GTF) genes gtfB and gtfC were ligated into Escherichia coli-streptococcus shuttle plasmids and introduced into Streptococcus milleri. gtfB transformant KSB8 formed an S. mutans-like rough colony on mitis salivarius agar and expressed an extracellular GTF-I, of 158 kDa, and two cell-bound GTF-Is, of 158 and 135 kDa. gtfC transformant KSC43 formed a semirough colony on mitis salivarius agar and expressed primarily an extracellular GTF-SI, of 146 kDa, and two cell-bound GTF-SIs, of 146 and 152 kDa. The extracellular GTFs from KSB8 and KSC43 were purified and characterized. The two types of GTF also reacted specifically with monoclonal antibodies directed against each enzyme. Both enzymes synthesized significant amounts of oligosaccharides, consisting primarily of alpha-1,6-glucosidic linkages, as well as water-insoluble glucans, containing alpha-1,3-glucosidic linkages. Insoluble-glucan-synthesizing activities of both enzymes were stimulated (three- to sixfold) by the addition of dextran T10 and were inhibited in the presence of 1.5 M ammonium sulfate. The Km(s) for sucrose and the optimal pHs were also similar for both enzymes. However, when the transformants were grown in Todd-Hewitt broth supplemented with sucrose, KSC43 cells, expressing GTF-SI activity, adhered to glass surfaces in vitro, while KSB8 cells, expressing GTF-I activity, did not. These results are discussed relative to the potential role of the gtfB and gftC genes in S. mutans cariogenicity.


Inhibition of adhesion of S-fimbriated Escherichia coli to buccal epithelial cells by human milk fat globule membrane components: a novel aspect of the protective function of mucins in the nonimmunoglobulin fraction. We investigated the presence of factors in human milk that inhibit invasion of pathogenic bacteria. The effect of human milk fat globule membrane (HMFGM) components on adhesion of cloned S-fimbriated Escherichia coli to human buccal epithelial cells was analyzed. S fimbriae are a common feature of E. coli strains causing sepsis and meningitis in newborns and are bound to epithelia via sialyl-(alpha-2-3)galactoside structures. Human milk fat globules (HMFG) could be agglutinated by the above-mentioned bacteria. Agglutination could be inhibited by fetuin, human glycophorin, and alpha 1-acid glycoprotein. In addition, pretreatment of HMFG with Vibrio cholerae neuraminidase markedly reduced bacterium-induced agglutinations, indicating the involvement of neuraminic acid-containing glycoproteins. In contrast, lipid droplets of infant formula or artificial lipid emulsions (Intralipid) could not be agglutinated. HMFG were present in stools of breast-fed neonates as shown by indirect immunofluorescence staining with a monoclonal antibody directed against carbohydrate residues present on HMFGM. These HMFG could be agglutinated by bacteria. HMFG inhibited E. coli adhesion to buccal epithelial cells. To further characterize relevant E. coli binding structures, HMFGM components were separated by gel chromatography. The mucin fraction showed the most pronounced inhibitory effect on adhesion of S-fimbriated E. coli to human buccal epithelial cells. Our data suggest that HMFG inhibit bacterial adhesion in the entire intestine and thereby may provide protection against bacterial infection.


Effects of staphylococcal enterotoxin B on rodent mast cells. Staphylococcal enterotoxin B (SEB) was tested in rodent mast cell cultures for the release of serotonin. Both rat RBL-2H3 mast cells and murine peritoneal cells released serotonin after SEB stimulation in culture. Release of serotonin in RBL-2H3 cells depended on the concentration of SEB; an appreciable release was seen at 50 micrograms/ml. The release of serotonin was not due to cell death. Serotonin release could be enhanced by bradykinin but not by vasoactive intestinal peptide, substance P, lipopolysaccharide from Salmonella typhimurium, the calcium ionophore A23187, acetylcholine, adenosine, 5-hydroxyeicosatetraenoic acid, indomethacin, or phorbol myristate acetate. SEB bound directly to the membrane of RBL-2H3 mast cells, and the SEB-binding site, the presumptive receptor, appeared to be a protein. The SEB receptor could not be capped under membrane-capping conditions, and serotonin release could not be enhanced by attempts to cross-link the receptor. These results suggest that mast cells may be an important cell type involved in SEB toxicosis and that release of serotonin may be enhanced by activation of the kinin-kallikrein system.


Potentiation by novobiocin of the cytotoxic activity of etoposide (VP-16) and teniposide (VM-26). The coumermycin antibiotic novobiocin, which interacts with the nuclear enzyme topoisomerase II, produced supra-additive toxicity to WEHI-3B D+ leukemia cells at clinically achievable concentrations, when combined with teniposide (VM-26) or etoposide (VP-16). Simultaneous exposure of cells to both agents was required for maximum efficacy of the combination. Novobiocin also produced supra-additive toxicity to A549 human lung carcinoma cells when combined with VM-26 or VP-16. At concentrations above the peak plasma levels achievable in patients, novobiocin lost its potentiating activity. Exposure of WEHI-3B D+ cells to novobiocin did not modify the cytotoxicity produced by the topoisomerase II inhibitor m-AMSA, whereas, in contrast, novobiocin antagonized the cytotoxicity of m-AMSA in A549 cells. Although it has been suggested that inhibitors of the syntheses of DNA and RNA interfere with the cytotoxic activity of the epipodophyllotoxins, maximum potentiation of the cytotoxicities of VP-16 and VM-26 occurred at novobiocin concentrations that decreased the rates of synthesis of both DNA and RNA in WEHI-3B D+ cells by about 50%. The number of DNA-topoisomerase-II covalent complexes stabilized by VM-26 in WEHI-3B D+ cells was greatly increased when cells were exposed simultaneously to VM-26 and novobiocin for 1 hr, but not when cells were treated with m-AMSA and novobiocin for the same period of time. Novobiocin did not affect the amount of covalent complexes produced by VM-26 in isolated nuclei, suggesting that the potentiating activity of novobiocin was not due to its direct interaction with the nuclear topoisomerase II enzyme. Our findings suggest that therapeutic levels of novobiocin may be capable of enhancing the clinical activities of VP-16 and VM-26.


Insulin-like growth-factor-binding protein gene expression and protein production by human tumour cell lines. The secretion of insulin-like growth-factor-binding proteins (IGFBPs) and expression of the genes encoding IGFBP-1, IGFBP-2 and IGFBP-3 have been studied in a panel of cell lines derived from breast carcinomas, Wilms' tumour, neuroblastoma, retinoblastoma, colon carcinoma, liver adenocarcinoma, Burkitt's lymphoma and a non-small-cell lung carcinoma. All cell lines, with the exception of the Burkitt's lymphoma cell line, secreted IGFBPs, as detected by affinity labelling. A 34-kDa BP was present in the conditioned media of all IGFBP-secreting cell lines, whereas BPs ranging from 18 kDa to 53 kDa were variably secreted. All IGFBP-secreting cell lines expressed the IGFBP-2 gene as determined by Northern blot analysis. The Wilms' tumour, the neuroblastoma and the retinoblastoma cell line expressed the IGFBP-2 gene only. All other cell lines, with the exception of the Burkitt's lymphoma, expressed the IGFBP-2 gene and, in addition, either the IGFBP-1 gene and/or the IGFBP-3 gene. IGFBP-1 gene expression could be detected by reverse transcriptase polymerase chain reaction only. IGFBP-3 gene expression was detected by Northern blot analysis, but transcripts were less abundant than IGFBP-2 mRNAs. These findings indicate that the expression of multiple BP genes and the secretion of BPs may be a common property of tumour cells.


Capillary occlusion and secondary angiogenesis: case report. The first follow-up control 5 days later (14.1.1990) showed a fresh capillary occlusion with massive red blood cell diapedesis. The arteriolar limb of the capillary is filled with erythrocytes up to the occlusion. Since capillary occlusion probably allowed some plasma flow at first, blood cells distal to the occlusion were washed away. After 53 days (8.3.1990) extravasal cells could no longer be made out. At the upper margin of the image a newly formed capillary loop can be seen. The channel of the old capillary seemed to have disappeared. Seventy-three days after occlusion, on 28th of March, the tip of the capillary, already demonstrated on 8th of March, had further grown distally. The capillary had crossed the neighbouring diagonal capillary. Twelve days later (9.4.1990) a remarkable growth connected with formation of a collateral was observed. The new capillary grew in direction of the former capillary channel. On 22nd of April the new capillary had almost got back to its old form. The collateral vessel formed on 9th of April had also grown and could be well distinguished. The different branches were immediately well perfused (mean erythrocyte velocity is v = 0.74 mm/s). The capillary were growing at different speeds. The new capillary had visibly grown by 220 microns over a period of 47 days which corresponds to an average growth of 4.7 microns a day. The capillary was growing fastest between the 9th and 22nd of April. A minimum of growth of 2 microns was observed within the first 11 days of the observation period.


Macromolecular transport across endothelial monolayers. We studied the macromolecular size-selective transport characteristics of polycarbonate (PC) filters with defined pore radius (rp; 15,000 to 400 A) as well as 2,000 A rp PC filter-bovine pulmonary artery endothelial cell (EC) monolayer sandwich under zero hydrostatic pressure conditions using fluorescein isothiocyanate-hydroxyethyl starch (FITC-HES, 16 A less than ae less than 100 A), and 2-methoxy-2,4-diphenyl-3(2H) furanone-bovine serum albumin (MDPF-BSA, ae = 35.5 A). We surprised to find substantial convective solute transport (solute drag) across the filter-endothelial sandwich. This effect was increased by large rp (15,000 A) filters and prevented by 400 A rp filters. Positive hydrostatic pressure across 2,000 A rp filters increased convective solute transport and negative pressure prevented this effect. High, medium and low permeability monolayers on 2,000 A filters progressively attenuated the solute drag effect seen across these filters without cells. The decline in monolayer permeability was associated with an increased filter area covered by cells; approximately 50 and 95% as well as greater than 99%, respectively. Although significant restricted diffusion was seen across low permeability monolayers, this pattern was distinct from that measured in single frog capillaries. Restricted diffusion by low permeability monolayers under conditions that produce solute drag document the significant barrier effects of high confluence endothelial monolayers, in vitro. These data show that solute transport across endothelial monolayers is due to diffusion+convective solute drag. The degree of the solute drag effect across the filter-endothelial sandwich is a direct function of monolayer confluence.


[Nevus spongiosus et albus mucosae].

[Nevus spongiosus et albus mucosae]. An 8-year-old girl with a non-familial case of white sponge nevus (WSN) is presented. The differential diagnosis is discussed with reference to anamnestic, clinical and histopathological data. In keratin expression, WSN resembles the epithelia of the hard palate and the tongue.


[Immunohistologic characterization of skin metastases in a patient with simultaneous cancers of the rectum and cervix]. A patient with skin metastases several years after surgical treatment of carcinoma of the cervix and the rectum is presented. Comparative histology and immunohistochemical analysis with anti-cytokeratin antibodies implicated the carcinoma of the cervix as the source of the skin infiltrates. Based on this patient's case record, the use of anti-cytokeratin antibodies for identification and subtyping of epithelial or carcinoma cells is discussed with special reference to cytokeratin 7.


Effect of osteopathic medical management on neurologic development in children. For 3 years, children between 18 months and 12 years of age with and without recognized neurologic deficits were studied at the Osteopathic Center for Children. Their response to 6 to 12 osteopathic manipulative treatments directed to all areas of impaired inherent physiologic motion was estimated from changes in three sensory and three motor areas of performance. Houle's Profile of Development was used to compare neurologic with chronologic age and rate of development, and scores were age-adjusted. Results in children after treatment were compared with those following a waiting period without treatment. Neurologic performance significantly improved after treatment in children with diagnosed neurologic problems and to a lesser degree in children with medical or structural diagnoses. The advances in neurologic development continued over a several months' interval. The results support the use of osteopathic manipulative treatment as part of pediatric healthcare based on osteopathic medical philosophy and principles.


DNA ligands as radiomodifiers: studies with minor-groove binding bibenzimidazoles. An iodinated bibenzimidazole, iodoHoechst 33258, was previously reported to markedly sensitize DNA and cells to UV-A, exemplifying the potential of iodinated DNA ligands as radiosensitizers, a rational extension of sensitization by halogenated pyrimidines. However, unlike the latter sensitizers, iodoHoechst 33258 is not a sensitizer of ionizing radiation, presumably due to the innate radioprotective properties of the uniodinated ligand. Experiments with purified DNA show that both Hoechst 33258 and Hoechst 33342 decrease the yield the radiation-induced DNA strand breakage. The ligands bind at discrete sites in the minor groove of DNA, and analysis on DNA sequencing gels show pronounced protection at the ligand binding sites, as well as more generalized protection. The extent of protection of strand breakage on plasmid DNA and the fact that it persists in the presence of 0.5 M NaCl (which prevents low affinity ionic binding between the high affinity sites) suggests that the protective effects of bound ligand are not confined to the high affinity binding sites in the minor groove. The mechanisms of this generalized protection is unknown, but there is some evidence indicating that the H-atom donation from the ligand may account for the site-specific protection. The extent of protection is much diminished, but still evident, in the presence of 100 mM mannitol, a known hydroxyl radical scavenger, indicating that some of the protective effects might relate to DNA damage mediated by direct action. Further evaluation of the mechanisms of protection should enable development of both more active radioprotectors and, by elimination of the radioprotective features from halogenated DNA ligands, more effective radiosensitizers.


Treatment of myelodysplastic syndromes with hematopoietic growth factors. Clinical trials with hematopoietic growth factors (granulocyte-macrophage colony-stimulating factor [GM-CSF], granulocyte colony-stimulating factor [G-CSF], interleukin-3, or erythropoietin) have been performed on patients with myelodysplastic syndromes. Absolute neutrophil counts can be readily raised to within the normal range by treatment with GM-CSF or G-CSF. Increases in platelets and hemoglobin have been reported after treatment with interleukin-3 and erythropoietin, respectively. Cytogenetic and molecular genetic analyses have demonstrated that both normal and malignant precursor cells are stimulated by cytokine therapy.


Differentiation therapy.

Differentiation therapy. A variety of compounds are able to induce myeloid leukemia cells to differentiate into morphologically and functionally mature cells. Vitamin D derivatives, retinoids, interferons, and polar-planar compounds are agents that demonstrate such activity in vitro and have been employed as therapy for the myelodysplastic syndromes.


Thin-layer electrophoresis of hydroxyethyl starches on a modified silica gel support. The recent developed thin-layer electrophoresis on modified silanized silica gel was applied to the separation of hydroxyethyl starches (HES) and glycogen and of HES with different degrees of substitution. This method permits a rapid qualitative and semi-quantitative determination of HES in animal tissues such as liver, lung, heart and kidney after their disintegration with alkali and precipitation with ethanol.


Bayley's infant behavior record ratings of infants with recurrent apnea: behavioral profile and correlates with apnea, age, and developmental status. The goal of this study was to provide a behavioral profile of apneic infants and to examine relationships between behavioral ratings, apnea frequency, age, and concurrent developmental status. Two samples of previously apneic infants were included in this exploratory analysis using the Infant Behavior Record (IBR) as a measure of temperament. Frequency of IBR suspect ratings was more than twice those reported for the Bayley standardization sample for tension, object orientation, attention, and reactivity. Apnea frequency, age, and developmental status were significantly related to a number of IBR ratings with relationships remaining significant for apnea and decreased responsiveness to persons and sensory interest after controlling for age and developmental status in the combined samples. These findings are consistent with a behavioral profile described as a lethargic temperament but may reflect early physiologic instability rather than inherent behavioral style.


Comparing the effects of neonatal intensive care unit intervention on premature infants at different weights. This study investigated the efficacy of an intervention program in the neonatal intensive care unit (NICU) on the development of premature infants from low socioeconomic status (SES) backgrounds. Sixty premature infants born at a county hospital over an 8-month period and their mothers were the focus of this study. The infants were assigned to either a control group or an experimental group. The experimental group received teaching and reinforcement about their babies' behavior from a child development specialist when they visited the NICU. Of the initial 60 infants, 49 completed the study at 8 months, but because data from only the first born of twin pairs were used, 41 infants were included in the final analysis. Infants were divided into two weight groups: one below and one above 1500 g. There was a significant effect of the intervention on the infants' mental and motor scores at 4 and 8 months. Significant differences on the total Home Observation for Measurement of the Environment (HOME) were noted between the groups. There were some significant differences between intervention and control groups on the interaction of the mother with the infant at 4 months but not in the play situation at 8 months. The lighter premature infants had a greater boost from the intervention than did the heavier premature infants. The intervention had no effect on the mother's perception of her infant's temperament or on her confidence skills. In line with other research findings, the infants who weighed less than 1500 g seemed to have benefited most from the intervention, and the effect of the intervention was most notable at 4 months.(ABSTRACT TRUNCATED AT 250 WORDS)


Parvalbumin and calbindin immunocytochemistry reveal functionally distinct cell groups and vibrissa-related patterns in the trigeminal brainstem complex of the adult rat. Immunocytochemistry for calbindin (CA) and parvalbumin (PA) was combined with retrograde tracing from the thalamus, superior colliculus (SC), and cerebellum to define the ascending projections of neurons in the rat's trigeminal (V) brainstem complex that express immunoreactivity for these calcium binding proteins. Many PA-immunoreactive neurons were observed in trigeminal nucleus principalis (PrV). Many of these cells projected to thalamus and a few sent axons to SC. In ventral PrV, PA-immunoreactive neurons were arranged in a vibrissa-related pattern. A very small number of large CA-immunoreactive neurons were observed in dorsomedial PrV. None of these cells were labeled by our tracer deposits. Small neurons in V subnucleus oralis (SpO) were also immunoreactive for PA, but none were retrogradely labeled. A small percentage of the large neurons in SpO were CA-immunoreactive; many of these were retrogradely labeled by tracer injections in the thalamus and/or SC. In V subnucleus interpolaris (SpI), many small to medium sized cells were PA-positive and they were arrayed in a vibrissae-like pattern. None of these neurons were retrogradely labeled from any of the above-listed targets, but many were retrogradely labeled by tracer injections into ipsilateral PrV. SpI also contained many large CA-immunoreactive cells. Many of these projected to the thalamus and/or SC and some were also retrogradely labeled by tracer injections into ipsilateral PrV. In V subnucleus caudalis (SpC), very dark PA-immunoreactive neurons were located in the inner part of lamina II and less often in laminae I. Lightly labeled cells were located in the magnocellular laminae and formed vibrissa-related aggregates. None of these neurons were retrogradely labeled by our tracer injections. CA-immunoreactive cells were located throughout the depth of lamina II in SpC and smaller numbers were also visible in lamina I and layers III-V. A small percentage of the CA-positive cells in lamina I and in the magnocellular layers were retrogradely labeled from the thalamus. These data indicate that PA and CA antisera identify two cell populations in whisker-related regions of the V brainstem complex and that PA cells are somatotopically patterned in PrV, SpI, and SpC. These markers also distinguish two cell groups in superficial laminae of the medullary dorsal horn.


Changes of chemoarchitectural organization of the rat spinal cord following ventral and dorsal root transection. Time-related changes in the distribution of chemical messengers in the rat spinal cord following the transection of dorsal and ventral roots were observed by using immunohistochemistry for the following antigens: microtubule-associated protein 2 (MAP2), calcitonin gene-related peptide (CGRP), substance P (SP), galanin (Gal), Met-enkephalin (Enk), neuropeptide Y (NPY), and serotonin (5-HT). To investigate dendrocytoarchitectonic organizational changes, morphometric analyses were performed on both the gray and the white matter of tissue samples stained with MAP2 antiserum. A significant reduction in the area of gray matter on the lesioned side was seen from 1 to 24 weeks postoperation, and progressive changes in the shape of the gray matter were also observed. CGRP-immunoreactive fibers were reduced in number in the posterior horn after root transection, except in the lateral part of lamina I. In contrast, CGRP immunoreactivity in the anterior horn cells of the ipsilateral side was increased early after transection, but later it progressively decreased. Root transection also caused significant reduction in the number of SP-immunoreactive fibers in the posterior horn, but no changes were seen in the anterior horn. Gal immunoreactivity was also affected by root transection, and it changed in a similar way to CGRP immunoreactivity. 5-HT-immunoreactive fibers were increased in the posterior horn after transection, and later decreased. In the anterior horn, there were no changes in the intensity or distribution pattern of 5-HT-immunoreactive nerve fibers following root transection. Enk and NPY immunoreactivity in the anterior and posterior horns was not affected by root transection up to 24 weeks postoperative. These results show that spinal root transection caused significant changes in the chemoarchitectural organization of nerve fibers containing certain types of chemical messengers, such as CGRP, SP, Gal, and 5-HT, in addition to altering dendritic geometry in the spinal cord.


Immunoreactivity to GABAA receptor in the outer plexiform layer of the cat retina. The distribution of GABAA receptor in the outer plexiform layer of cat retina was studied by immunocytochemistry with monoclonal antibodies. Staining was observed at the base of the cone pedicle, extracellularly, in association with the "triad" synaptic complex. Some bipolar dendrites and the basal processes that interconnect the cone pedicles were also stained. Rod spherules and horizontal cells were negative. The findings support the idea that the cone horizontal cells are GABAergic.


The tectorecipient zone in the inferior olivary nucleus in the rat. Tecto-olivary and olivocerebellar projections in the rat were investigated in order to identify the tectorecipient zone in the inferior olivary nucleus and to determine whether inferior olivary neurons projecting to the cerebellar tecto-olivo-recipient zones (lobule VII, crus II, lobulus simplex, and paramedian lobule) originate in the different regions within the tectorecipient zone. An electrophysiological method and an axonal transport technique of wheat-germ agglutinin-conjugated horseradish peroxidase were used. The tectorecipient zone was identified in the caudomedial region of the medial accessory olive. Neurons projecting to lobule VII originated in the caudomedial region of the tectorecipient zone, but those to crus II, lobulus simplex, and paramedian lobule originated in its rostrolateral region. These observations suggest that there are two independent tecto-olivo-cerebellar systems: 1) superior colliculus--the medial region of the tectorecipient zone--lobule VII--the caudomedial region of the fastigial nucleus; and 2) superior colliculus--the rostralateral region of the tectorecipient region--crus II, lobulus simplex, and paramedian lobule--the dorsolateral protuberance of the fastigial nucleus.


Two types of motoneurons supplying dorsal fin muscles in lamprey and their activity during fictive locomotion. The location and dendritic morphology of motoneurons supplying the dorsal fin muscles were studied in the lamprey spinal cord (Ichthyomyzon unicuspis). Motoneurons were retrogradely labelled after injection of HRP into the fin muscles or after its application on the cut ends of the ventral roots. HRP-labelled cells were subsequently reconstructed, in the horizontal and/or transverse planes. Fin motoneurons were also injected intracellularly with Lucifer Yellow and their detailed three-dimensional structure was analysed by confocal laser-scanning microscopy. Unlike myotomal motoneurons, which are closely spaced in the lateral cell column, fin motoneurons were distributed along the spinal cord separately or in pairs. They could be distinguished from motoneurons supplying trunk muscles by having a limited number of dendrites in the lateral part of the spinal cord. In addition, some fin motoneurons extend their dendrites into the dorsal column. The motor cells innervating fin muscles were divided into two types based on their dendritic morphology. Type I have a widespread dendritic tree in the rostrocaudal direction and, with few exceptions, completely restricted to the ipsilateral side. A proportion (25%) of these cells have dendrites extending into the dorsal column. Type II fin motoneurons extended their dendrites both ipsi- and contra-laterally. The contra-lateral dendrites pass below and above the central canal. The dendrites send off branches into the dorsal columns on both the ipsi- and the contra-lateral sides. Electron microscopic analysis showed that both type I and type II fin motoneurons receive numerous synaptic contacts from dorsal column axons. During fictive locomotion both types of motoneurons are active in antiphase in relation to myotomal motoneurons and to the main locomotor burst.


Callosal and superior temporal sulcus contributions to receptive field properties in the macaque monkey's nucleus of the optic tract and dorsal terminal nucleus of the accessory optic tract. To assess the functional contribution of the cortical input to the receptive field properties of nucleus of the optic tract (NOT) and dorsal terminal nucleus (DTN) neurons, a first set of experiments evaluated the response properties of NOT-DTN cells in monkeys with split corpus callosum. With respect to visual latency, direction specificity, directional tuning width, velocity tuning, ocular dominance, and binocular interaction, they were indistinguishable from NOT-DTN neurons in normal monkeys. However, a clear difference was found regarding the extent of the receptive fields. Whereas, in normal monkeys, NOT-DTN receptive fields include the contralateral hemifield and the fovea as well as substantial parts of the ipsilateral visual field, receptive fields in callosum-split monkeys stop abruptly at, or close to, the vertical 0-meridian and do not extend into the ipsilateral visual field. In addition, the location of the highest sensitivity within the receptive fields in callosum-split monkeys is shifted away from the vertical 0-meridian in comparison to normal animals. In a second set of experiments, we antidromically identified cortical neurons within the superior temporal sulcus that project to the NOT-DTN. These neurons were found in area MT mostly near the border of MTp or MSTl. All of them are direction selective for ipsiversive stimulus movement, and their receptive fields extend substantially into the ipsilateral visual hemifield. Neurons with other preferred directions did not project to the NOT-DTN. These results contribute to the explanation of the ipsiversive directional deficits in slow eye movements after cortical lesions, as well as the asymmetries in optokinetic nystagmus with hemifield stimulation after transection of the corpus callosum. The more general implication of the results is that a particular function of a cortical area can only be understood by knowing its subcortical connections.


Anatomy and fine structure of neurons in the deutocerebral projection pathway of the crayfish olfactory system. Golgi impregnation and neurobiotin injection were used to examine details of the neural pathways in the olfactory system of the freshwater crayfish, Procambarus clarkii. Deutocerebral projection neurons (globuli cells) were directly injected with neurobiotin. These neurons have dendritic arborizations in the ipsilateral olfactory and accessory lobes, and they project axons to the lateral protocerebrum, where they terminate in microglomeruli of the hemi-ellipsoid body. The axons of the deutocerebral projection neurons are readily impregnated by Golgi procedures, and they terminate as an expanded membranous knot about 5 microns in diameter. Electron microscopy on Golgi-stained terminals has revealed that each knot makes several hundred synapses with small spine-like or shaft-like processes of postsynaptic neurons. Injection of neurobiotin into local interneurons of the hemi-ellipsoid body and subsequent examination of stained preparations with the electron microscope reveals that these cells are a major postsynaptic target of the deutocerebral projection neurons. Furthermore, the local interneurons make extensive efferent synaptic connections with unidentified neurons in the terminal medulla.


Eccrine syringofibroadenomatosis: a clinical and histologic study and review of the literature. A 56-year-old man had an 11-year history of a psoriasiform eruption of the palms, soles, and shins. An examination revealed well-demarcated patches and plaques of erythematous, fissured, and hyperkeratotic skin with focal erosions. There was no clinical evidence of ectodermal dysplasia. On histologic examination these lesions proved to be eccrine hamartomas that consisted of anastomosing cords and strands of cuboidal epithelial cells with well-formed ducts and a fibrovascular mucinous stroma. Eccrine ductal origin was indicated by histopathologic, histochemical, immunopathologic, and electron microscopic evaluation. These multiple palmoplantar eccrine hamartomas, unassociated with ectodermal dysplasia, represent a sporadic hamartomatous condition that is best designated as "eccrine syringofibroadenomatosis."


Multiple linear cylindromas.

Multiple linear cylindromas. We report a case of multiple cylindromas with a linear arrangement on the lower right extremity. The patient had more than 100 tumors, but the scalp was spared. Some tumors showed overlapping features with eccrine spiradenoma. We believe this is the first report of multiple linear cylindromas.


Generalized papular xanthomatosis in mycosis fungoides. Xanthomas can occur in association with underlying lymphoproliferative disease, or they can result from lipid deposition in damaged or altered skin. We report a case of generalized papular xanthomas that developed in a patient with Sézary syndrome. The xanthomas were composed of foamy histiocytes that were shown by immunoperoxidase staining to be of the monocyte/macrophage lineage. Electron microscopic studies revealed lipid vacuoles, lysosomes, and myelin figures but no Birbeck granules, features that are consistent with a non-X histiocytosis. Generalized papular xanthomatosis has not been previously described in a patient with cutaneous T-cell lymphoma.


Correlations of unique clinical, immunotypic, and histologic findings in cutaneous gamma/delta T-cell lymphoma. Cutaneous T-cell lymphoma is a malignancy of T cells that express a clone-specific heterodimer T-cell receptor for antigen. The second recognized case of an epidermotropic malignancy of T-cells expressing gamma/delta T-cell receptor-expressing cells is reported. The immunophenotype of the malignant T-cells was CD3+, CD2+, CD7+, gamma/delta T-cell receptor+, CD4-, CD8-, and alpha/beta T-cell receptor-. The clinical features were remarkable for extreme epidermotropism with a scant dermal lymphomatous infiltrate. Profound keratinocyte necrosis occurred in areas of malignant skin infiltrates. Despite cutaneous lesions covering more than 50% of the skin surface of the patient, no adenopathy or splenomegaly was detected. The intense epidermotropism of this patient's gamma/delta T-cell receptor-expressing cells and the putative cytolytic properties of CD4- CD8- gamma/delta contributed to the destruction of epidermis. Remission was induced with a combination of electron beam and extracorporeal photochemotherapy.


The effect of short-term fasting and a single meal on protein synthesis and oxygen consumption in cod, Gadus morhua. Rates of protein synthesis and oxygen consumption (MO2) in cod were compared in both fasted and refed animals. During a 14-day fast both protein synthesis and respiration rates fell to stable values after 6 days. When a meal of whole sandeel at 6% body weight was fed to fish fasted for 6 days, protein synthesis and MO2 increased to a maximum at between 12 and 18 h after feeding. Peak MO2 was about twice the pre-feeding values, while whole animal protein synthesis increased four-fold. There were differences between tissues in the timing of maximum protein synthesis; the liver and stomach responded faster than the remainder of the body. Maximum protein synthesis rates in the liver and stomach occurred at 6 h after feeding, at which time their calculated contribution to total MO2 was 11%. Similar calculations suggested that the integrated increment in whole animal protein synthesis contributed between 23% and 44% of the post-prandial increase in MO2. It was concluded that protein synthesis is an important contributor to increased MO2 after feeding in cod.


The effects of decentralization on substance P-immunoreactivity in the anterior major pelvic ganglion of the male guinea pig. The anterior major pelvic ganglion (AMPG) of the male guinea pig possesses a substance P (SP)-immunoreactive peri-neuronal plexus. Selective nerve transections involving the principal inputs of the AMPG, the hypogastric and pelvic nerves, indicate that the SP-immunoreactive peri-neuronal plexus is derived from multiple sources: an extrinsic source involving both the hypogastric and pelvic nerves, and another source (possibly the projections of small intensely fluorescent cells). SP-immunoreactivity (IR) is not normally present in the neuronal perikarya of the AMPG. Evidence is presented that suggests the absence of SP-IR is due to an active suppression of SP-synthesis. This seems to be achieved by a trans-synaptic mechanism involving the hypogastric nerve which, after transection, leads to the appearance of neuronal perikarya exhibiting SP-IR (less than 1% of the total neuronal population of the AMPG). Up to 65% of the neuronal perikarya of the AMPG have the ability to synthesize SP, as demonstrated by SP-IR after 24 h in vitro. A more potent factor in the down-regulation of SP synthesis seems to be exerted by the pelvic genito-urinary organs, especially the prostate.


Modern histochemical methods using enzymes as markers. In the 25 years since the idea that histochemically detectable enzymes could serve as a marker of ligands, the world of histochemistry has undergone a dramatic change. Today slides of immunostained tissue sections may be filed together with hematoxylin- and eosin-stained slides and may be examined at one's leisure. Antigens are now localized at the ultrastructural level as well as at the light microscopic level permitting one to wonder about the intricacies and beauties of living creatures. Genes which mandate our body since the day of conception to the day of death can now easily be probed with enzyme markers. Even with all these success stories, some dreams which I had hoped to accomplish with the method are not yet realized. Unlike the chemical and physical properties of inert objects, living organisms are provided with a fixed genetic program. By recognizing the point in the program which is being executed, one can deduce the potential FUTURE activities of the cells and tissues. I hypothesize that this is not an impossible task since (1) histochemistry and immunohistochemistry already provide us information on the PAST activities of cells and tissues, (2) in situ hybridization describes PRESENT gene activities, and (3) newer methods allow for the determination of potential gene function. Thus it is possible to peek at the future.


Neutralization of the anticoagulant effects of glycosaminoglycans by serum amyloid P component: comparison with other plasma and platelet proteins. Serum amyloid P protein (SAP) is a heparin-binding protein that is found in blood and connective tissues including some types of vascular basement membrane. In this article we present evidence that SAP is capable of blocking the anticoagulant effects of glycosaminoglycans. SAP neutralized the catalytic effect of heparin on the thrombin-antithrombin III reaction more effectively than vitronectin, histidine-rich glycoprotein, fibronectin, and high-molecular-weight kininogen and almost as effectively as platelet factor 4. SAP also blocked the effects of heparin and dermatan sulfate on the inhibition of thrombin by heparin cofactor II. We found evidence for the formation of a high-affinity 1:1 complex between SAP and heparin and for inhibition of binding of both thrombin and antithrombin III to heparin-Sepharose by SAP. We conclude that SAP may account for much of the heparin-neutralizing capacity of plasma under some conditions and that basement-membrane-bound SAP may modulate extravascular coagulation by blocking the anticoagulant effects of basement membrane glycosaminoglycans.


Coordinated appearance of beta 1 integrins and fibronectin during corneal wound healing. Fibronectin, which appears at wound sites, serves as a temporary matrix for the epithelial migration involved in healing. Cellular responses to fibronectin have not yet been elucidated. We visualized beta 1 integrins, fibronectin, laminin, and collagen type IV by immunofluorescence techniques at various intervals after single, nonpenetrating incision in the corneas of rats to investigate the chronologic changes in the localization of these proteins during corneal epithelial wound healing. In unwounded corneas beta 1 integrins were found on basal cells of the corneal epithelium and fibronectin was found in the corneal subepithelial region, at Descemet's membrane, and in streaks between collagen fibers of the stroma. Both laminin and collagen type IV were found in the subepithelial region and Descemet's membrane. Immediately after the incision, fibronectin was visible on the surface of the V-shaped defect of the stroma; epithelial cell expressed beta 1 integrins began to migrate over the defect and to fill it. When healing was almost completed, beta 1 integrin staining of epithelial cells diminished, except for in basal cells. Staining of fibronectin diminished at the interface between the new epithelium and the stroma. Laminin and collagen type IV were not seen between the migrating epithelial cells and the underlying stroma until epithelial cells completely covered the defect. Appearance and disappearance of beta 1 integrins and fibronectin are thus well coordinated during corneal epithelial wound healing process, suggesting an active involvement of beta 1 integrins and fibronectin.


Short amino acid sequences derived from C1q receptor (C1q-R) show homology with the alpha chains of fibronectin and vitronectin receptors and collagen type IV. The human C1q receptor (C1q-R) is a 65-70-kd, highly acidic, hydrophobic glycoprotein that is expressed on a wide variety of cell surfaces. Although the C1q-R itself appears to bind preferentially to C1q, the region of the ligand to which C1q-R binds is the primary binding site for several other molecules, including fibronectin, laminin, and C1q inhibitor (chondroitin 4-sulfate proteoglycan) as well as the complement C1r2C1s2 tetramer. In order to further characterize the C1q-R molecule with regard to its structure and function, highly purified C1q-R was obtained from Raji cells using DEAE-Sephacel and C1q-Sepharose CL-4B chromatography. Studies performed with 125I-labeled C1q-R demonstrated that whereas the C1q-R molecule binds poorly to a variety of human collagens including types II, III, and V, markedly enhanced binding is observed with type IV collagen and moderately enhanced binding with type I collagen. Amino acid composition studies show that the C1q-R molecule contains approximately 44% hydrophobic and 12.6% hydrophilic residues with a ratio of negatively charged to positively charged residues of about 2:1. Treatment of 125I-labeled C1q-R with endoglycosidase F lowers the apparent molecular size from 70 to 58 kd, whereas endoglycosidase H lowered the size to 64 kd. Treatment with neuraminidase, on the other hand, shifted the size of C1q-R to 60 kd. These results suggest the presence of several highly sialylated complex-type or high mannose-type N-linked oligosaccharide side chains. Because purified C1q-R has a blocked amino terminus, amino acid sequences representing internal fragments of the molecule were generated by electroblotting and in situ enzymatic digestion. When these short sequences were searched against the National Biomedical Research Foundation computer data base, a seven-amino-acid sequence, VSWQGQI, showed significant homology (100% and 80% in a five-amino-acid overlap, respectively) with the alpha chains of the human fibronectin (alpha 5 beta 1) and vitronectin (alpha v beta 3) receptors, and to a lesser degree with epidermal growth factor receptor and T cell receptor. A second sequence, ISEDNIR, showed homology with mouse collagen type IV (86% in a six-amino-acid overlap), calmodulin (60% in a seven-amino-acid overlap), and a Leishmania major surface antigen, gp63. These observations seem to predict that C1q-R has pockets of conserved sequences that are similar to those not only present in its ligand(s) but also in other cell surface receptors that may, in part, fulfill similar functions.


[Aggregation of erythrocyte suspensions of diabetic patients in dextran 70 solutions]. Our previous studies have shown an erythrocyte hyperaggregation during diabetes. This hyperaggregation phenomenon seems usually be promoted by quantitative plasma protein's changes mainly fibrinogen. The aim of the present work is to appreciate the possible effect of only diabetic red cells on aggregation induced by dextran 70 (Dx 70). Aggregation measurements were performed with Sefam aggregometer. Patients were insulin-dependent (IDD) and non insulin-dependent diabetics (NIDD), divided in two groups of 11 well controlled diabetics (HbA1C 6.8 +/- 0.5%) (7 IDD and 4 NIDD) and 11 poorly controlled diabetics (HbA1C 10.8 +/- 2.7%) (7 IDD and 4 NIDD). They were compared with a control group consisting of 22 healthy subjects. Results can be summarized as follows: red cell aggregation induced by Dx 70 was not significantly different between the well controlled diabetics and controls. Similar results were obtained for the poorly controlled diabetics. By contrast, when studying red cell aggregation in autologous plasma (H = 40%), aggregation was found to be significantly more important in both diabetic groups than that of the controls. Thus, results emphasize the importance of suspending medium in erythrocyte hyper-aggregation phenomenon during diabetes. Cellular factors do not seem to interfere on aggregation phenomenon for diabetics included in this study.