The Presence of Insulin-degrading Enzyme in Human Ileal and Colonic Mucosal Cells

The aim of this research is to characterize the presence of insulin-degrading enzyme in human colon and ileal mucosal cells. Biochemical studies, including the activity-pH profiles, the effects of enzyme inhibitors, immunoprecipitation and western blots, were conducted. The majority of insulin-degrading activity in colon mucosal cells was localized in the cytosol. In both colon and ileum, cytosolic insulin-degrading activities had a pH optimum at pH 7.5, and were extensively inhibited by each of N-ethylmaleimide, p-chloromercuribenzoate, and 1,10-phenanthroline, but were very weakly affected by each of leupeptin, chymostatin, diisopropyl phosphofluoridate and soybean trypsin inhibitor. In the colon and ileum, more than 93% and 96%, respectively, of cytosolic insulin-degrading activities were removed by the mouse monoclonal antibody to human RBC insulin-degrading enzyme, as compared with less than 20% by the normal mouse IgG for both tissues. Further, a western blot analysis revealed that a cytosolic protein of 110 kD, in both human colon and ileum, reacted with the monoclonal antibody to insulin-degrading enzyme. It is concluded that insulin-degrading enzyme is present in the cytosol of human colon and ileal mucosal cells.     

Detection of undiagnosed coagulopathies using routine rapid heparin neutralization.

OBJECTIVE: To determine the most frequent clinical causes of a prolonged activated partial thromboplastin time (APTT) result, and to determine whether a new heparin-removal device (the Hepchek, Pall Biomedical, Glen Cove, NY 11542) is capable of efficiently detecting the causes of these values. DESIGN: A combination of chart review and laboratory testing comparing the criterion standard--the heparin chromogenic substrate assay--with the Hepchek. Laboratory investigations were blinded and controlled. SETTING: Inpatient, acute-care hospital. PATIENTS: A total of 1,000 hospital patients with a variety of hemostatic disorders. MAIN OUTCOME MEASURE: The extent to which the Hepchek accurately identified the etiology of a prolonged APTT result. RESULTS: The APTT was prolonged in 25.2% of samples. The presence of heparin in the sample was confirmed by chromogenic assay or by using the Hepchek heparin-removal filter. The presence of heparin was confirmed in 12.8% of all samples and in more than 50% of all abnormal samples. The cause of the abnormal APTT was often unappreciated by the clinician. Bayesian analysis of the Hepchek's ability to diagnose heparin correctly as the cause of the abnormal APTT showed a sensitivity of 100% and specificity of 99.9%. CONCLUSION: Use of the Hepchek in the routine clinical laboratory is an efficient and rapid method of detecting heparin as a cause of isolated prolonged APTT results, and should reduce demands for unwarranted coagulation analyses and inappropriate treatment with blood products.     

Advances in pharmacotherapy: depression in the elderly– issues and advances in treatment

Depression continues to be a major cause of morbidity and mortality in the elderly. It is estimated that 1–5% of elderly persons who live in the community and 5–43% of nursing–home patients have major depression. Symptoms of depression in the elderly do not differ substantially from younger patients. Tricyclic antidepressants continue to be the drugs of choice in the elderly because of their long record of use with proven efficacy, known adverse effect profile and availability of less expensive generic formulations. The newer secondgeneration antidepressants, including serotonin reuptake inhibitors, appear to offer a major advantage of fewer serious adverse effects in the elderly. This review will highlight recent developments regarding the prevalence and treatment of depression in the elderly.     

Bradykinin-induced release of PGI2 from aortic endothelial cell lines: responses mediated selectively by Ca2+ ions or a staurosporine-sensitive kinase.

1. Bradykinin (100 nM) triggers release of nitric oxide and prostacyclin from both AG07680A and AG04762 bovine cultured aortic endothelial cells. The exposure of these cells to bradykinin is in each case associated with a striking rise in intracellular calcium ion concentration. 2. Exposure of AG07680A cells to 250 nM ionomycin was followed also by a significant release of prostacyclin, whereas 250 nM ionomycin had no capacity to stimulate release of prostacyclin from AG04762 cells. 3. There was a similar concentration-dependent increase in intracellular calcium ion concentration on exposure of AG07680A and AG04762 cells to ionomycin. 4. Exposure of AG04762 cells for 10 min to staurosporine produced a concentration-dependent inhibition (IC50 = 107 +/- 14 nM) in bradykinin-stimulated prostacyclin release. There was no similar inhibitory effect of staurosporine in AG07680A cells. 5. Bradykinin (10 nM) triggered release of nitric oxide from both AG07680A and AG04762 cells, and the effect was not inhibited by 500 nM staurosporine. There was a similar ionomycin-dependent release of nitric oxide from both cell types. 6. These results identify a common pathway for bradykinin-dependent nitric oxide release from both AG07680A and AG04762 cells, involving increases in intracellular calcium ion concentration. In contrast, the bradykinin-dependent release of prostacyclin may involve one of two pathways (involving an increase in intracellular calcium or activation of a staurosporine-sensitive kinase), and the two pathways are selectively exploited in AG07680A and AG04762 cells, respectively.     

Bicycle helmet use by adults: the impact of companionship.

Most of the nearly 1,000 fatal bicycle-related injuries annually could be prevented if riders used safety helmets. Helmet use by adult bicyclists has received relatively little attention because educational campaigns to promote helmet use generally focus on children. Helmet use by adult and child bicyclists at 120 suburban and rural sites in three Maryland counties was observed on two Saturdays in 1990-91 during an evaluation of the impact of a mandatory helmet law. Concordance or discordance of helmet use within various groups of bicyclists--adults only, adults with children, and children only--was recorded. Helmet use among 2,068 adult bicyclists was 49 percent, 51 percent, and 74 percent in the three counties. In two counties combined, 52 percent (365 of 706) of solo adult bicyclists wore helmets compared with only 5 percent (5 of 94) of solo child bicyclists (P < .001). Helmet use or nonuse was concordant among 87 percent of 277 adult-adult pairs, 94 percent of 50 child-child pairs, and 91 percent of 32 adult-child pairs of bicyclists observed. Concordance rates of helmet use or nonuse were similarly high among pairs of adult bicyclists of the same or mixed sexes. These data are consistent with the concept that both adults and children tend to adopt the helmet-wearing behaviors of their companions. Public health efforts focused on adults should encourage helmet use by adult bicyclists both to prevent head injuries and to provide a role model for children.     

A37 REACTIVITY OF PLATELET AUTOANTIBODIES WITH PAPAIN TREATED PLATELETS AND COMPARISON WITH GLYCOPROTEIN SPECIFICITY.

A1 INFLUENCE OF POSTURE ON REACTIONS IN NEW BLOOD DONORS. A2 A CONFIDENTIAL UNIT EXCLUSION SYSTEM IDENTIFIES DONORS WITH A POTENTIAL FOR HIV INFECTION. A3 A STABLE BLOOD SUPPLY FOR THE FUTURE: THE RECRUITMENT OF 16 TO 18 YEAR OLD DONORS TODAY AND THEI CONTRIBUTION AS COMMITTED REGULAR DONORS OF TOMORROW. A4 APPROACH TO A SUPPLY CRISIS OF HYPERIMMUNE RHESUS PLASMA FOR THE PRODUCTION OF RhD IMMUNOGLOBULIN A5 THE INFLUENCE OF AGE, SEX AND ABO BLOOD GROUP ON THE INCIDENCE OF CMV ANTIBODIES IN SYDNEY BLOOD DONORS. A6 THE INCIDENCE OF CATEGORY VI AMONGST WEAK Rh(D) POSITIVE SYDNEY BLOOD DONORS. A7 A MODIFIED METHOD FOR DETECTING HIGH TITRE ANTI-A AND ANTI-B IN GROUP O DONORS A8 IMPROVING THE CLINICAL SPECIFICITY OF ALANINE AMINO TRANSFERASE (ALT) TEST RESULTS WITHIN THE NORMAL BLOOD DONOR POPULATION OF QUEENSLAND. A9 EXTRACTION OF HCV RNA USING A GUANIDINE ISOTHIOCYNATE METHOD. A10 HEPATITIS C VIRUS (HCV) ANTIBODY DETECTION IN TASMANIAN BLOOD DONORS. A11 EFFECTIVE INTERNAL QUALITY CONTROL FOR ENZYME IMMUNOASSAYS A12 DETECTION OF ANTIBODY TO NON-PATHOGENIC RETROVIRUSES (SPUMAVIRUSES) IN HUMAN SERUM A13 DETECTION OF ANTIBODY TO NON-PATHOGENIC RETROVIRUSES (SPUMAVIRUSES) IN HUMAN SERUM A14 A NOVEL BLOOD BAG SYSTEM WITH POTENTIAL, FOR THE ASIA-PACIFIC MARKET. A15 DESIGN OF CONTAINERS SUITABLE FOR THE TRANSPORT OF RED CELL, PLATELET AND FROZEN PLASMA PRODUCTS. A16 EVALUATION OF INDICATOR LABELS FOR QUALITY ASSURANCE OF IRRADIATION PROCEDURE OF BLOOD PRODUCTS. A17 MOLECULAR TYPING FOR UNUSUAL ABO TYPES. A18 AN EXAMPLE OF THE RARE ABO SUBGROUP, A19 RFLP ANALYSIS OF A RH NULL BLOOD DONOR. A20 A RELATIONSHIP BETWEEN LEWIS ERYTHROCYTE PHENOTYPES AND COLORECTAL CANCER. A21 PATERNITY TESTING USING SINGLE LOCUS DNA PROBES: OBSERVATIONS ON THE REFERENCE DATA BASE SIZE A22 USE OF FAMILY AND POPULATION STUDIES TO DETERMINE THE SPECIFICITY AND INHERITANCE OF NEUTROPHIL ANTIGENS DEFINED BY PLANT LECTINS. A23 SAMPLING PLANS: IS THERE RELEVANCE FOR BLOOD COMPONENT QC? A24 QUALITY MANAGEMENT: HOW DO WE DO IT IN A STATE THE SIZE OF QUEENSLAND? A26 THE ENERGY METABOLISM OF CIRCULATING CELLS. A27 ACETATE UTILISATION RATES AND THE EFFECT OF GLUCOSE-FREE PLASMA IN PLATELET CONCENTRATE STORED IN A MIMIMAL MEDIUM (MPM). A28 IMPROVED LEVELS OF 2,3 DJPHOSPHOGLYCERATE IN RED CELL SUSPENSIONS PREPARED FROM BLOOD COLLECTED INTO DEXTROSE-FREE ANTICOAGULANT. A29 EVALUATION OF RED CELL FREEZING METHODS AS A PRELUDE TO ADOPTING -80° C FREEZING IN HIGH GLYCEROL IN ROUTINE PRACTICE. A30 CLUMPING IN PLATELET CONCENTRATES - AN UNSOLVED PROBLEM. A31 AUTOLOGOUS BLOOD: SAFE FOR OTHERS OR NOT? A32 ESTABLISHMENT OF AN AUSTRALIAN HAEMOPHILIA TREATMENT CENTRE DATA BANK. A33 EXPERIENCE IN THE USE OF ROBOTICS AND MICROPLATE TECHNOLOGY TO SEMI-AUTOMATE A ROUTINE HOSPITAL BLOOD BANK. A34 AN ANTI-IgAl/IgA2 ELISA ASSAY FOR THE INVESTIGATION OF HYPESENSITIVITY TRANSFUSION REACTIONS. A35 THE INFLUENCE OF IgG AGGREGATES AND FRESH NORMAL SERUM ON THE MONOCYTE MONOLAYER ASSAY A36 DETECTION OF Rh(D) POSITIVE FETAL CELLS IN PREGNANT Rh(D)-NEGATIVE WOMEN BY FLOW CYTOMETRY. A38 HAEMOSTAT-IX: A HIGH PURITY FACTOR CONCENTRATE FOR THE TREATMENT OF PATIENTS WITH HAEMOPHILIA B. A39 GRAVITY FILTRATION OF PLASMA FROM DONOR BLOOD UTILISING A HOLLOW FIBRE FILTER MEMBRANE DEVICE A40 The Therapeutic Device Problem Reporting Scheme, and the Victorian Red Cross Blood Bank A43 HIGH FREQUENCY ANTIBODIES AND THE ADVANTAGES OF MANUAL POLYBRENE. A44 FACTS AND FANTASY IN THE DEVELOPMENT OF PLATELET ADDITIVE SOLUTIONS. A45 LACK OF EFFECT OF STORAGE CONTAINER ON STORAGE OF PLATELETS PREPARED FROM DEXTROSE-FREE BLOOD, A46 PLATELETS PREPARED FROM DEXTROSE-FREE BLOOD MAY BE STORED WITHOUT AGITATION. A47 QUALITY OF BED CELL CONCENTRATE IN HOSPITALS COMPARED TO THE BLOOD BANK A48FLOW CYTOMETRIC CHARACTERISATION OF LEUCOCYTE - DEPLETED RED CELL CONCEHTRATES. A49 PRODUCTION AND CHARACTERISATION OF HUMAN MONOCLONAL ANTI-D ANTIBODIES. A50 CD55 AND CD59 SUSCEPTIBILITY TO PROTEASE TREATMENT AND THE RESULTANT EFFECT ON COMPLEMENT LYSIS OF RBCs. A51 DIRECT COMPARISON BETWEEN PLATELET STORAGE CONTAINERS - IMPROVEMENT IN STORAGE CHARACTERISTICS OF TUTA PLATELET BAGS OVER THE PAST FOUR YEARS. A52 IMPROVED SOLID-PHASE MIXED PASSIVE HAENAGGLUTININ ASSAY (MPHA) WITH FROZEN PANEL PLATELETS FOR THE DETECTION OF HUMAN PLATELET ANTIBODIES. A53 DEVELOPMENT OF A SOLVENT DETERGENT TREATED THROMBIN CONCENTRATE AS A COMPONENT OF A FIBRIN GLUE KIT. A54 Autologous blood transfusion: a promotional programme A55 AVAILABILITY OF BLOOD PRODUCTS FOR ACUTELY BLEEDING PATIENTS. A56 REMINISCENCES OF 50 YEARS A. A TRANSFUSION ST. A57 A NATIONAL SYSTEM FOR REPORTING TRANSFUSION REACTIONS TO FRACTIONATED BLOOD PRODUCTS. A58 EFFECT OF FLUORESCENT LIGHTING ON THE VISUAL APPEARANCE OF PLATELET CONCENTRATES A59 USING A MICROWAVE OVEN TO THAW FRESH FROZEN PLASMA. A60 COAGULATION CAPACITY OF POOLED PLATELET PLASMA. A61 A COMPARISON OF IMMUNOHAEMATOLOGY SURVEY PERFORMANCE BETWEEN NEK ZEALAND AND AUSTRALIA A62 COMPATIBILITY TESTING: ARE ENZYME TESTS REQUIRED? A63 AN EVALUATION OF THE DIAMED MEROTYPING SYSTEM FOR THE PERFORMANCE OF THE DIRKT ANTIGLOBUDIN TEST. A64 ASSESSMENT OF PERFORMANCE IN BLOOD GROUP ANTIBODY DETECTION. A65 CHARACTERISATION OF MABS TO THROMBIN-HIRUDIN COMPLEXES WITH IMMUNOASSAY POTENTIAL. A66 MONITORING ANTT-HPA-la (P1^A1) PLATELET ANTIBODY LEVELS DURING PREGNANCY USING THE MAIPA TEST. A67 COMPARISON OF PIFT AND MAIPA TEST“ IN THE DETECTION OF ANTI-HPA-la (PI^A1) PLATELET ANTIBODIES. A68 USE OF PLATELET-CROSSMATCHING IN SUPPORT OF A CASE OF MYELODYSPLASIA WTTH A PLATELET SPECIFIC AND B LYMPHOCYTE ANTIBODY A69 The Pattern of Leucocyte Antibody formation in Transfused Patients. A70 DETECTION OF HPA-Ia ANTIBODY IN BREAST MILK A71 ANALYSIS OF PRENATAL SCREENING. A72 DETECTION OF MINOR POPULATIONS OF ERYTHROCYTES A73 MODIFICATIONS TO THE MCNOCYTE-MEDIATED ADCC ASSAY. A74 AN AUTO ANTI-JMH; GAMMA-CLONE POLYSPECIFIC AHG AS A USEFUL TOOL. A75 CLINICALLY SIGNIFICANT ANTI-A1 DERIVED FROM B LYMPHOCYTES FOLLOWING SINGLE LUNG TRANSPLANTATION. A76 CONFIRMATION THAT ANTI-ELO CAUSES HAEMOLYTIC DISEASE OFTHE NEWBORN. A77 ANTI-Do^a STIMULATED BY PREGNANCY. A78 DONOR IgM ANTI-A ASSOCIATED WITH HAEMOLYTIC TRANSFUSION REACTION A79 COLLECTION OF GRANULOCYTES AND PLATELETS USING FENWALL CS 3000 AND HAEMONETICS 30 CELL SEPARATORS - A COMPARISON. A80 APPARENT LYMPHOPENIA IN PLASMAPHERESIS DONORS     

Fos Expression in the Rat Brain Following Vaginal-Cervical Stimulation by Mating and Manual Probing

Vaginal-cervical stimulation (VCS), provided by mating or manual probing, induces many reproductive behavioral and endocrine changes in female rats. These changes include an increase in lordosis duration, heat termination and pseudopregnancy. Electro-physiological and [¹⁴C]2-deoxy-D-glucose studies collectively show that neurons in the medial preoptic area, ventromedial hypothalamus and midbrain central gray respond to manual VCS. In the present study we immunocytochemically labeled brain sections for Fos, the protein product of the immediate early gene c-fos, to detect VCS-responsive neurons in hormone-primed animals receiving VCS by mating or manual probing. In Experiment 1, females receiving mounts and intromissions were compared to: 1) vaginally-masked females receiving mounts but no VCS, 2) females exposed to an intact anesthetized male or 3) females not exposed to males or the testing arena. Those animals receiving VCS showed a dramatic increase in the number of Fos-immunoreactive cells in the medial preoptic area, posterodorsal portion of the medial amygdala and bed nucleus of the stria terminalis, as well as the dorsomedial hypothalamus, ventromedial hypothalamus and midbrain central gray. These effects of VCS were confirmed in Experiment 2 in animals receiving manual vaginal-cervical probing. These findings extend previous electrophysiological and [¹⁴C]2-deoxy-D-glucose studies by providing evidence that additional brain areas respond to VCS by mating, as well as manual probing.     

Serotonin-induced increase in cAMP in ganglia isolated from the myenteric plexus of the guinea pig small intestine: Mediation by a novel 5-HT receptor

Serotonin (5-HT) is a mediator (through 5-HT_1P receptors) of slow EPSPs in myenteric ganglia of the small intestine. The effect of 5-HT can be mimicked by elevating cAMP; therefore, we tested the hypothesis that the slow EPSP-like response to 5-HT is cAMP-mediated. Guinea pig gut was enzymatically dissociated; myenteric ganglia remained intact and were collected by filtration. Neurons in the isolated ganglia retained their ability to manifest the slow EPSP-like response to 5-HT. Exposure to 5-HT raised the ganglionic level of cAMP (ED₅₀ 0.3 μM). This effect was not antagonized by the 5-HT_1P antagonist, N-acetyl-5-hydroxytryptophyl-5-hydroxytryptophan amide (100.0 μM), or mimicked by the 5-HT_1P agonist, 5-hydroxyindalpine (10.0 μM). Increases in cAMP were also evoked by the 5-HT₁ agonist, 5-carboxyamidotryptamine (10.0 μM), the 5-HT₂ agonist, (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI; 1.0–10.0 μM), and by the 5-HT₄ agonists, renzapride (1.0–10.0 μM) and 5-methoxytryptamine (1.0–10.0 μM); however, neither the 5-HT₁/5-HT₂ antagonists, spiperone, methysergide, and methiothepin, nor the 5-HT₄ antagonist, tropisetron (ICS 205–930; 10.0 μM), were able to inhibit the rise in cAMP evoked by these compounds or by 5-HT (0.1–10.0 μM). The 5-HT-evoked elevation of cAMP was antagonized by ketanserin (10.0 μM), which also blocked the effects of 5-methoxytryptamine and DOI, but not those of renzapride. The effective concentration of DOI, however, was higher than that needed for activation of 5-HT₂ receptors, and Northern analysis using a cDNA probe encoding the rat 5-HT₂ receptor failed to reveal the presence of 5-HT₂ mRNA in myenteric ganglia, although it hybridizes with mRNA of the right size in the guinea pig brain. Compounds that failed to change levels of cAMP or to antagonize the action of 5-HT included 8-hydroxy-di-n-propylamino tetralin, R58639, R88226, and sumatriptan. It is concluded that the receptor responsible for the 5-HT-induced rise in cAMP in ganglia isolated from the guinea pig myenteric plexus is not a known subtype of 5-HT receptor. Since the pharmacology of this novel receptor is different from that of the slow EPSP-like response to 5-HT, the receptor probably does not mediate the slow EPSP. © 1993 Wiley-Liss, Inc.     

Insights into Mechanisms of Cisplatin Resistance and Potential for Its Clinical Reversal

Cisplatin in combination with other cytotoxic agents is the backbone for a potential cure of testicular germ cell neoplasms and is a critical factor in the substantial activity observed in the treatment of small cell lung cancer, bladder cancer, and ovarian germ cell tumors. Resistance to cisplatin at the onset of treatment or at relapse limits its curative potential, however. Laboratory studies using both cells selected for cisplatin resistance by exposure to sublethal concentrations and biopsy specimens from patients' tumors provide insights for the potential mechanisms of resistance. The mechanisms identified in vitro include a complex and wide array of related and unrelated pathways such as alterations in cellular drug transport, enhanced DNA repair dependent and independent of signal transduction pathways, and enhanced intracellular detoxification such as glutathione and metallothionein systems. Studies of these mechanisms have identified a number of agents with known potential for administration to humans and that reverse cisplatin resistance in vitro; for example, reversal of cellular accumulation defects by dipyridamole; inhibition of DNA repair by hydroxyurea, pentoxifylline, and novobiocin; inhibition of the glutathione system by ethacrynic acid and buthionine sulfoximine; and inhibition of signal transduction pathways by cyclosporine, tamoxifen, and calcium channel-blocking agents. Current phase I clinical trials are focusing on the most effective doses and schedules to administer these agents in combination with cisplatin. Initial uncontrolled trials in limited numbers of patients suggest that the addition of modulators of cisplatin has the potential to reverse resistance in patients previously failing therapy. Another promising avenue for circumventing cisplatin resistance is the development of noncross-resistant platinum analogs.