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Biomedical subjects

S Martin

Publications and source records attributed to S Martin.

At least 253 records · Page 14Linked to original sources

Characteristics of families who attend free vaccine fairs.

OBJECTIVE: To determine the health care resources and perceived barriers to care of families attending free vaccine fairs. DESIGN: A cross-sectional survey. SETTING: Twelve free vaccine fairs in Denver, Colorado, in 1994. PARTICIPANTS: A total of 533 consecutive parents or guardians of children receiving vaccine at the fairs. Interventions. None. MEASUREMENTS/RESULTS: Survey respondents reported that their children received regular health care through a private physician or health maintenance organization (HMO) (47%), a public clinic (20%), or a hospital-based clinic (14%); 18% had no regular site for health care. Twenty-seven percent of the families carried private insurance, although less than half of these plans covered children's vaccines: 9% were enrolled in an HMO or a preferred provider organization and 13% had Medicaid, whereas 50% had no health insurance. Families who received primary care at a private physician's office (OR: 1.7; 95% CI: 1.01-2.7) and those with no regular site for health care (OR: 2.0; 95% CI: 1.01-4.0) were more likely than those who went to a public clinic or hospital clinic to report free vaccine as the most important reason for attending a vaccine fair. Conversely, families who received well-child care at a hospital clinic were more likely to identify no appointment needed as the most important reason (OR: 2.7; 95% CI: 1. 4-5.1). Families with private health insurance (OR: 2.3; 95% CI: 1. 05-4.0) or no health insurance (OR: 2.3; 95% CI: 1.1-4.6) were more likely to identify free vaccine as the most important reason for attending a vaccine fair, whereas those enrolled in an HMO or preferred provider organization identified convenient time as the most important reason (OR: 3.2; 95% CI: 1.2-8.3). Families with Medicaid (OR: 3.2; 95% CI: 1.3-8.3) or with no insurance (OR: 2.1; 95% CI: 1.02-4.6) were more likely than were those with private insurance to identify no appointment needed as the most important reason for attending a vaccine fair. CONCLUSIONS: Most families attending free vaccine fairs have a regular source of health care. For families with private health insurance or with no health insurance, the availability of free vaccine is the major reason to bring their children to a vaccine fair, whereas for families whose insurance routinely covers the cost of childhood vaccine (HMO, Medicaid), convenience is the major determinant.

Colorado↗

Familial defective apolipoprotein B-100 (R3500Q) in Northern Ireland.

Familial defective apolipoprotein B-100 (FDB) R3500Q is an autosomal co-dominant disorder caused by the substitution of glutamine for arginine at amino acid residue 3500 of the apolipoprotein B-100 gene. It is associated with hypercholesterolaemia of varying severity, and with coronary artery disease. Hypercholesterolaemic patients (n = 158) from Northern Ireland were screened for the defect by polymerase chain reaction-mediated, site-directed mutagenesis. Clinical presentation ranged from moderate hypercholesterolaemia with a family history of hypercholesterolaemia or heart disease (n = 104) to those classified as definitely having familial hypercholesterolaemia (FH) (n = 54). Eight (5.1%) unrelated individuals were found to be heterozygous for the FDB R3500Q mutation, including two (3.7%) of those 54 classified clinically as having FH. Treatment with HMG-CoA-reductase-inhibiting drugs (statins) decreased total cholesterol by 22-44% and low-density lipoprotein cholesterol by 34-46% in all but one FDB heterozygote.

Adolescent↗

Interleukin-12 enhances the virus-specific interferon gamma response of pigs to an inactivated pseudorabies virus vaccine.

Cell-mediated immunity is a major component of the host defense system against viral infections. Since interleukin (IL)-12 has been shown to be a potent stimulus for the in vivo generation of interferon-gamma (IFN-gamma)-producing T cells (i.e. Th-1 cells) in laboratory animals, we evaluated the effect of IL-12 on the cellular immune response of pigs to vaccination against pseudorabies virus (PrV), a herpesvirus of swine. The magnitude of the cellular immune response was measured by IFN-gamma ELISPOT analysis of peripheral blood mononuclear cells (PBMC) from pigs which had been immunized twice, at 2-week intervals, with either, modified live virus (MLV) alone or with a commercial inactivated PrV vaccine with or without the coadministration of human recombinant IL-12 (HrIL-12). No significant differences in the titer of virus-neutralizing antibodies or in the intensity of the virus-specific lymphoproliferative response among the different treatment groups was observed. However, the number of virus-specific IFN-gamma-producing cells among PBMC isolated from animals receiving the MLV vaccine was on average 3.5 times more than animals immunized with the inactivated vaccine (P = 0.01). Administration of the inactivated vaccine and IL-12 induced a two-fold higher frequency of virus-specific IFN-gamma-producing cells from that induced by the inactivated vaccine alone (P < 0.05). Despite this enhancement, the level of protection from lethal PrV challenge provided by the inactivated vaccine in combination with IL-12 was the same as that induced by the inactivated vaccine alone. Both of these vaccination regimes provided significantly lower levels of protection than those afforded by the MLV vaccine. This study demonstrates that an inactivated PrV vaccine is a poor inducer of virus-specific IFN-gamma-producing cells and that this response can be enhanced by administration of exogenous IL-12. The data provides evidence of a dichotomy in the humoral and cellular immune responses of pigs to a viral antigen and implies the existence of a Th-1/Th-2 type regulation of the anti-viral immune response in pigs.

Adjuvants, Immunologic↗

Regulatory roles of the P domain of the subtilisin-like prohormone convertases.

A unique feature of the eukaryotic subtilisin-like proprotein convertases (SPCs) is the presence of an additional highly conserved sequence of approximately 150 residues (P domain) located immediately downstream of the catalytic domain. To study the function of this region, which is required for the production of enzymatically active convertases, we have expressed and characterized various P domain-related mutants and chimeras in HEK293 cells and alpha-TC1-6 cells. In a series of C-terminal truncations of PC3 (also known as PC1 or SPC3), PC3-Thr594 was identified as the shortest active form, thereby defining the functional C-terminal boundary of the P domain. Substitutions at Thr594 and nearby sites indicated that residues 592-594 are crucial for activity. Chimeric SPC proteins with interchanged P domains demonstrated dramatic changes in several properties. Compared with truncated wild-type PC3 (PC3-Asp616), both PC3/PC2Pd and PC3/FurPd had elevated activity on several synthetic substrates as well as reduced calcium ion dependence, whereas Fur/PC2Pd was only slightly decreased in activity as compared with truncated furin (Fur-Glu583). Of the three active SPC chimeras tested, all had more alkaline pH optima. When PC3/PC2Pd was expressed in alpha-TC1-6 cells, it accelerated the processing of proglucagon into glicentin and major proglucagon fragment and cleaved major proglucagon fragment to release GLP-1 and tGLP-1, similar to wild-type PC3. Thus, P domain exchanges generated fully active chimeric proteases in several instances but not in all (e.g. PC2/PC3Pd was inactive). The observed property changes indicate a role for the P domain in regulating the stability, calcium dependence, and pH dependence of the convertases.

Animals↗

Intracellular localization of phosphatidylinositide 3-kinase and insulin receptor substrate-1 in adipocytes: potential involvement of a membrane skeleton.

Phosphatidylinositide (PI) 3-kinase binds to tyrosyl-phosphorylated insulin receptor substrate-1 (IRS-1) in insulin-treated adipocytes, and this step plays a central role in the regulated movement of the glucose transporter, GLUT4, from intracellular vesicles to the cell surface. PDGF, which also activates PI 3-kinase in adipocytes, has no significant effect on GLUT4 trafficking in these cells. We propose that this specificity may be mediated by differential localization of PI 3-kinase in response to insulin versus PDGF activation. Using subcellular fractionation in 3T3-L1 adipocytes, we show that insulin- and PDGF-stimulated PI 3-kinase activities are located in an intracellular high speed pellet (HSP) and in the plasma membrane (PM), respectively. The HSP is also enriched in IRS-1, insulin-stimulated tyrosyl-phosphorylated IRS-1 and intracellular GLUT4-containing vesicles. Using sucrose density gradient sedimentation, we have been able to segregate the HSP into two separate subfractions: one enriched in IRS-1, tyrosyl-phosphorylated IRS-1, PI 3-kinase as well as cytoskeletal elements, and another enriched in membranes, including intracellular GLUT4 vesicles. Treatment of the HSP with nonionic detergent, liberates all membrane constituents, whereas IRS-1 and PI 3-kinase remain insoluble. Conversely, at high ionic strength, membranes remain intact, whereas IRS-1 and PI 3-kinase become freely soluble. We further show that this IRS-1-PI 3-kinase complex exists in CHO cells overexpressing IRS-1 and, in these cells, the cytosolic pool of IRS-1 and PI 3-kinase is released subsequent to permeabilization with Streptolysin-O, whereas the particulate fraction of these proteins is retained. These data suggest that IRS-1, PI 3-kinase, as well as other signaling intermediates, may form preassembled complexes that may be associated with the actin cytoskeleton. This complex must be in close apposition to the cell surface, enabling access to the insulin receptor and presumably other signaling molecules that somehow confer the absolute specificity of insulin signaling in these cells.

3T3 Cells↗

Quantitative detection of porcine interferon-gamma in response to mitogen, superantigen and recall viral antigen.

Five monoclonal antibodies (mAbs) specific for porcine interferon-gamma (PoIFN-gamma) were isolated and utilized to develop a PoIFN-gamma sandwich ELISA. Specific reactivity of each mAb with E. coli derived recombinant PoIFN-gamma, but not with rPoIL-2 or rPolL-10, was confirmed in an indirect ELISA and in Western blots. Competitive ELISAs showed that mAbs P2A4 and P2C11 bound an epitope which was not recognized by mAbs P2G10, P1B7 or P2F6. The latter three mAbs were able to neutralize the ability of natural and recombinant PoIFN-gamma to induce the de novo expression of class II MHC antigens on porcine endothelial cells. To simplify the detection of biologically active porcine IFN-gamma, a sandwich ELISA was developed using the mAb P2G10 as a capture antibody and mAb P2C11 as the detecting reagent. The sensitivity of the assay for PolFN-gamma ranged from 1 to 50 ng/ml. Peripheral blood mononuclear cells (PBMC) from all pigs tested produced IFN-gamma when stimulated with either mitogen (PHA) or superantigen (SEB). In contrast, only PBMC obtained from pigs which had previously been vaccinated against PrV produced IFN-gamma in response to stimulation with this virus. Interestingly, cultures with the highest lymphoproliferative response did not necessarily have the highest level of IFN-gamma production.Furthermore, for recall viral antigen, the lymphoproliferative response decreased with time after immunization, whereas the IFN-gamma response increased. Thus, measurement of IFN-gamma production appears to be a good indicator of anti-viral immunological memory.

Animals↗

Incomplete processing of proinsulin to insulin accompanied by elevation of Des-31,32 proinsulin intermediates in islets of mice lacking active PC2.

The prohormone convertases PC2 (SPC2) and PC3/PC1 (SPC3) are the major precursor processing endoproteases in a wide variety of neural and endocrine tissues. Both enzymes are normally expressed in the islet beta cells and participate in proinsulin processing. Recently we generated mice lacking active PC2 due to a disruption of the PC2 gene (Furuta, M., Yano, H., Zhou, A., Rouillé, Y., Holst, J. J., Carroll, R. J., Ravazzola, M., Orci, L., Furuta, H., and Steiner, D. F. (1997) Proc. Natl. Acad. Sci. U. S. A. 94, 6646-6651). Here we report that these PC2 mutant mice have elevated circulating proinsulin, comprising 60% of immunoreactive insulin-like components. Acid ethanol extractable proinsulin from pancreas is also significantly elevated, representing about 35% of total immunoreactive insulin-like components. These increased amounts of proinsulin are mainly stored in secretory granules, giving rise to an altered appearance on electron microscopy. In pulse-chase experiments, the mutant islets incorporate lesser amounts of isotopic amino acids into insulin-related components than normal islets. In both wild-type and mutant islets, proinsulin I was processed more rapidly to insulin, reflecting the preference of both PC2 and PC3 for substrates having a basic amino acid positioned four residues upstream of the cleavage site. The overall half-time for the conversion of proinsulin to insulin is increased approximately 3-fold in the mutant islets and is associated with a 4-5-fold greater elevation of des-31,32 proinsulin, an intermediate that is formed by the preferential cleavage of proinsulin at the B chain-C-peptide junction by PC3 and is C-terminally processed to remove Arg31 and Arg32 by carboxypeptidase E. The constitutive release of newly synthesized proinsulin from both mutant and wild-type islets during the first 1-2 h of chase was normal (<2% of total). These results demonstrate that PC2 plays an essential role in proinsulin processing in vivo, but is quantitatively less important in this regard than PC3, and that its absence does not influence the efficient sorting of proinsulin into the regulated secretory pathway.

Amino Acid Sequence↗

Detection of HLA-specific antibodies by PRA-STAT and their association with transplant outcome.

OBJECTIVE: The aim was to investigate the correlation between renal transplant outcome and the presence of HLA-specific antibodies detected using the ELISA kit PRA-STAT as compared with complement-dependent cytotoxicity (CDC). METHOD: 295 sera from 95 renal transplant recipients (99 transplants) were investigated for the presence of HLA-specific antibodies using both PRA-STAT and CDC. The patients were divided into group I (49 transplants failed within 1 month) and group II (50 successful transplants). RESULTS: The concordance between PRA-STAT and CDC for the detection of HLA class I-specific antibodies was 87.8% (259 of 295). For 19 sera, antibodies were detected only by PRA-STAT; for 17 sera, antibodies were detected only by CDC. No donor-specific antibodies were detected by either technique for patients in group II. For four group I patients (six sera), donor-specific IgG antibodies were detected only by PRA-STAT (one before, three after transplant) and all four transplants failed. For five other group I patients (six sera), donor HLA-specific antibodies were detected only by CDC (one before, four after transplant) and all five transplants failed. The antibodies detected before transplant by CDC were shown to be IgM alloantibodies. CONCLUSION: This study showed that PRA-STAT could detect HLA-specific IgG antibodies relevant to transplant outcome that were not detected by CDC. However, it could not detect IgM alloantibodies that were also shown to be important. PRA-STAT is therefore a useful addition to a histocompatibility laboratory's screening repertoire only when used in conjunction with other techniques.

Adult↗

Transient alteration of T cell fine specificity by a strong primary stimulus correlates with T cell receptor down-regulation.

P14 mice expressing a transgenic TCR specific for the lymphocytic choriomeningitis virus glycoprotein p33 epitope were used to study the induction of CTL effector activity by a variety of ligands. Surprisingly, p33 variants which are weaker agonists for the P14 TCR than the wild-type p33 peptide were able to induce more potent effectors with a broader range of cytolytic specificity. Similarly, low concentrations of p33 were more effective than higher concentrations. These results correlated with no or only moderate TCR down-regulation by variants of p33 and low p33 concentrations. This phenotype observed after 18 h of culture was transient as progressive restoration of reactivity was observed at 42 or 66 h in the cultures stimulated with high p33 concentrations and this correlated with recovery of TCR surface levels. TCR down-regulation was blocked by src family kinase inhibitors. These findings indicate that the specificity of a T cell can be fine-tuned by the nature of the primary stimulus correlating with surface TCR level and imply an important role for src family kinases in the differential regulation of surface TCR levels upon TCR engagement by different ligand/MHC complexes.

Animals↗

Identification of yeast proteins from two-dimensional gels: working out spot cross-contamination.

With the complete sequence of the yeast genome now available, efforts by many laboratories are underway to identify each of the spots on two-dimensional (2-D) gels corresponding to the most abundant yeast proteins. The high mass accuracy now attainable using matrix assisted laser desorption/ionization (MALDI)-mass spectrometry equipped with delayed extraction simplifies the process of identification, such that many spots can be unambiguously identified in a short period of time merely by using peptide mass fingerprinting and generally available database matching programs. Although it is not always possible to match spots between gels run by different laboratories, proteins generally yield the same abundant proteolytic fragments when tryptic digestions are performed. Databases containing these signature peptides not only simplify the task of reidentifying proteins from different gels, but also make it possible to identify small amounts of cross-contaminating proteins from different spots, as well as common extraneous contaminants such as human keratins. In this paper, we present data on the identification of > 20 previously unreported yeast proteins from 2-D gels. Some novel proteins were identified from randomly analyzed spots. Focusing on 14 spots in a narrow-pH-range gel, we demonstrate how organizing peak-table data and peptide match-list data into databases enables the identification of a larger percentage of the peaks.

Acrylic Resins↗

Soluble forms of intercellular adhesion molecule-1 inhibit insulitis and onset of autoimmune diabetes.

Increased concentration of circulating adhesion molecules in human serum have been described in different immune-mediated diseases. Recently, we proposed an immunomodulatory function of soluble forms of the intercellular adhesion molecule-1 (ICAM-1) during the pathogenesis of human Type I (insulin-dependent) diabetes mellitus. To test this hypothesis in nonobese diabetic (NOD) mice, a spontaneous animal model for human Type I diabetes, two recombinant forms of soluble murine ICAM-1 were generated, one monomeric soluble ICAM-1 containing all five extracellular Ig-like domains of ICAM-1 (rICAM-1) and one dimeric protein with the N-terminal extracellular domains fused to the constant regions of murine IgG2a (rICAM-1-Ig). Beginning at age 35 days prediabetic NOD mice received i. p. injections of 5 microg recombinant ICAM-1-proteins three times a week for 4.5 months. At day 170 diabetes development was reduced (p < 0.001) in NOD mice receiving rICAM-1 (8%) or rICAM-1-Ig (8%) treatment in comparison with sham treated animals (45%). After termination of therapy animals treated with multimeric rICAM-1-Ig were protected longer than animals treated with rICAM-1. Prevention of diabetes was associated with decreased infiltration of pancreatic islets by mononuclear cells. A selective downregulation of Th1-type cytokine expression was observed in a second set of experiments in which diabetes development was synchronised by cyclophosphamide. These data support the hypothesis that circulating forms of adhesion molecules have an immunomodulatory function and can intervene in islet inflammation.

Aging↗

A novel strategy for the detection and definition of HLA-specific antibodies in patients awaiting renal transplantation.

Conventional testing for HLA-specific antibodies employs complement-dependent cytotoxicity (CDC) which is labour intensive and dependent on a supply of viable lymphocytes. Our strategy to minimise CDC screening is initially to screen sera by ELISA (Quikscreen) to detect HLA Class I-specific antibodies. Negative sera are then screened by flow cytometry (FCS) using lymphoblastoid cell line pools to detect HLA Class II-specific antibodies. Only Quikscreen- or FCS-positive sera are then tested by CDC and, when indicated, with an ELISA kit (PRA-STAT) for specificity definition. Of 3680 sera, 886 (24.1%) were Quikscreen positive. Of the 2794 Quikscreen-negative sera, 374 (13.4%) were FCS positive. Therefore, only 1265 of the 3680 (34.3%) sera contained HLA-specific antibodies requiring specificity definition. This novel screening strategy has significantly reduced the CDC workload of the laboratory whilst enabling the detection of additional HLA-specific antibodies.

Antibodies↗

Pre-adaptation, adaptation and de-adaptation to high altitude in humans: hormonal and biochemical changes at sea level.

High altitude residence is known to modify body biochemistry and hormone status. However, the effects of such a sojourn on these status observed at sea level both immediately and later after return are not as well established as are the effects of an intermittent acclimation. The aim of this study was therefore to investigate these changes. To achieve our objectives, nine subjects received intermittent acclimation at low pressure in a barometric chamber (8 h daily for 5 days, day 1 at 4500 m, day 5 at 8500 m) before an expedition to the Himalayas. Hormonal and biochemical changes were studied using samples of venous blood taken at sea level before and after acclimation, after return from the expedition and 1 and 2 months after descent. Concentrations of thyroid hormones, adrenaline, noradrenaline (NA), hormones of hydromineral metabolism (aldosterone, renin, arginine vasopressin, atrial natriuretic peptide) as well as prolactin, cortisol, insulin and endothelin 1 were measured. Biochemical measurements made were plasma osmolality, and concentrations of glucose, total cholesterol, total proteins, pre-albumin, transferrin, complement 3C, apolipoproteins A1 and B and serum iron. Acclimation induced no alteration in hormone (except for NA with increases of about 1.5, fold P < 0.05) and biochemistry data. After the expedition, hormone responses were characterized by a higher total triidothyronine concentration (+18%, P < 0.05) while other hormones did not vary. A linear relationship was found between thyroid-stimulating-hormone and body mass changes after the expedition (r = 0.67, P < 0.05). The observed increased concentrations of plasma proteins and total cholesterol (P < 0.05) could be related to the restoration of lean body mass. At 1 and 2 months after return, no changes in hormones were observed but a significant decrease in transferrin concentration was noticed. The higher serum iron concentration reported after 1 month (P < 0.05) could have been the result of a physiological haemolysis. It was concluded that both acclimation and the expedition in the Himalayas affected hormone status and body biochemistry status even though the observed changes were slight and rapidly reversed.

Adaptation, Physiological↗

Tandem and triple high-dose chemotherapy with autologous stem cell rescue in metastatic breast cancer.

The purpose of this phase II study was to evaluate the therapeutic efficacy and toxicity of a tandem or triple high-dose chemotherapy (HDC) with autologous peripheral blood stem cell transplantation (PBSCT) in patients with metastatic breast cancer (MBC) as first line chemotherapy. Conventional chemotherapy consisted of two cycles of epirubicin 120 mg/m2 and ifosfamide 7500 mg/m2 in the case of tandem HDC and one cycle of paclitaxel 135 mg/m2, epirubicin 90 mg/m2 and ifosfamide 6000 mg/m2 in the case of triple HDC. Tandem HDC was composed of two cycles of epirubicin 180 mg/m2, ifosfamide 12000 mg/m2 and carboplatin 900 mg/m2. In the case of triple HDC, paclitaxel 180 mg/m2, etoposide 1500 mg/m2 and thiotepa 600 mg/m2 was added as the third cycle. Patients with tandem HDC (n = 20) were evaluable for both survival and toxicity, and patients with triple HDC (n = 21) only for toxicity because of short-term follow-up. Both tandem and triple HDC were well tolerated and could be safely administered. Non-hematological WHO grade 3 or 4 toxicities were mucositis (8), temporary renal insufficiency (1), myocardial infarction (1), and neuropathy (1). No toxic death occurred. The Kaplan-Meier estimates for 44-months without progression and the overall survival were 12% and 38% respectively. The median survival was 22 months (95% CI: 7.4-51.7 months) and the median progression-free interval 14 months (95% CI: 5.1-43.7 months). In a population with an unfavorable prognosis, tandem HDC showed similar efficacy as to that described in other phase II studies. Triple HDC seems not to improve patient outcome compared to tandem HDC, but a long-term follow up is required.

Adult↗

Heterogeneity of reproductive tract abnormalities in men with absence of the vas deferens: role of cystic fibrosis transmembrane conductance regulator gene mutations.

OBJECTIVE: To determine if the types of reproductive tract abnormalities linked to absence of the vas deferens varies with the cystic fibrosis transmembrane conductance regulator (CFTR) genotype. DESIGN: Prospective data gathering. SETTING: University infertility clinic. PATIENT(S): Forty-six infertile men with absence of the scrotal vas deferens and no signs of cystic fibrosis. INTERVENTION(S): All had blood taken for CFTR gene analysis, 33 had scrotal ultrasounds, and 25 had transrectal ultrasounds. MAIN OUTCOME MEASURE(S): The frequency of testicular, seminal vesicle, and ampullae of the vas deferens malformations was compared between subgroups of men with two, one, or no CFTR gene mutations. RESULT(S): None (0 of 21) of the men with at least one CFTR gene mutations had normal ampullae of the vas or seminal vesicles bilaterally. Two (50%) of 4 men with no CFTR gene mutations had normal ampullae of the vas deferens bilaterally, and 50% had normal bilateral seminal vesicles (statistically significantly different). There was no correlation between testicular malformations and CFTR genotype. CONCLUSION(S): This study indicates that the severity of the malformations in the testis is unrelated to the CFTR genotype, whereas the frequency and severity of wolffian duct malformations are related directly to the CFTR genotype.

Cystic Fibrosis Transmembrane Conductance Regulato↗