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Biomedical subjects

B Singh

Publications and source records attributed to B Singh.

At least 613 records · Page 34Linked to original sources
PubMed1991

Induction of mutations at the thymidine kinase locus in CHO cells by restriction endonucleases.

Induced mutation frequencies were measured at the tk locus (encoding for the enzyme thymidine kinase) following treatment of Chinese hamster ovary cells (CHO KI) with two restriction endonucleases (REs), PvuII and EcoRI, which generate 'blunt-ended' and 'cohesive-ended' DNA double-strand breaks (dsb), respectively. Electroporation was used to introduce these enzymes into the cells. Restriction endonucleases generating blunt-ended dsb have been shown to mimic the action of ionising radiation in causing chromosome aberrations, cell killing, mutations and oncogenic transformation. Here we show that the tk locus, PvuII induced an approximately 11-fold higher mutation frequency than EcoRI at the same enzyme concentrations. There are four PvuII and six EcoRI restriction sites in the Chinese hamster thymidine kinase gene. Hence the higher mutation induction by PvuII, despite the lower number of restriction sites than EcoRI in the tk gene, suggests that blunt-ended dsb represent more effective and critical mutagenic DNA lesions than the cohesive-ended type. In this respect, our results are similar to those we obtained previously for chromosomal aberrations and for cell killing. Results from the present study suggest that mutations could arise from unrepaired or misrepaired dsb possibly via induction of chromosomal deletions or stable exchanges between chromosomes.

Animals↗
PubMed1991

A community screening test for high alcohol consumption using biochemical and haematological measures.

A discriminant function based on a number of biochemical and haematological tests from an extended multiple biochemical analysis and full blood count, together with weight, smoking status and systolic blood pressure is developed. The function was far more effective at detecting high alcohol use (greater than 40 g ethanol per day) than serum gamma-glutamyl transpeptidase (GGT) or the Short Michigan Alcoholism Screening Test (SMAST) in a community sample of adult males. When classifying high alcohol consumption by GGT only, several division criteria were considered, the most effective being at 40 i.u./l. In terms of identifying high alcohol consumers, rather than alcoholics, the SMAST was no better than GGT, and both had unacceptably low sensitivity (49%, 51%) and poor performance on other measures, thus limiting their use as community screening tools. The discriminant function, however, had an estimated community sensitivity of 78%, was similarly high on other performance measures, and would perform satisfactorily as a community screening tool, particularly in situations where there was a tendency for individuals to under-report their alcohol consumption.

Adult↗
PubMed1991

Overexpression of Acetohydroxyacid Synthase from Arabidopsis as an Inducible Fusion Protein in Escherichia coli: Production of Polyclonal Antibodies, and Immunological Characterization of the Enzyme.

Acetohydroxyacid synthase (AHAS, EC 4.1.3.18) is the first enzyme unique to the biosynthesis of valine, leucine, and isoleucine. This enzyme is the target site of several classes of structurally unrelated herbicides. The conventional method of antibody production using purified protein has not been successful with this enzyme. Two separate fragments of a gene encoding a portion of the mature region of AHAS from Arabidopsis were fused with the trpE gene from Escherichia coli using the pATH1 vector. E. coli cells transformed with each respective plasmid expressed a fusion protein at levels greater than 10% of the total cell protein. The fusion protein was purified and used to immunize rabbits. Antisera obtained from the immunized rabbits immunoprecipitated AHAS activity from Arabidopsis cell free extracts. The anti-AHAS antisera reacted with a 65 kilodalton protein band in electrophoretically resolved extracts of Arabidopsis. In cross-reactivity tests, this antibody was able to immunoprecipitate AHAS activity from various plant species. Furthermore, a protein band with a molecular mass of 65 kilodaltons was detected in the crude extracts of all plant species tested on a Western blot. These results indicate that the 65 kilodalton protein represents AHAS in various plant species. The wide spectrum of cross-reactivity for the antisera supports the view that the AHAS enzyme is highly conserved across all plant species.

Journal Article↗
PubMed1991

Clinical correlates of in vitro Plasmodium falciparum cytoadherence.

To determine whether isolates of Plasmodium falciparum have intrinsically different cytoadherent properties and whether these differences contribute to the clinical severity of human falciparum malaria, we studied the cytoadherence to C32 melanoma cells in vitro of 59 parasite isolates from patients with naturally acquired infections in Thailand. Parasitized erythrocytes adhere to these melanoma cells principally via the glycoprotein CD36, which is also expressed on most vascular endothelium. In vitro cytoadherence was significantly greater for isolates from patients with biochemical evidence of severe malaria. The cytoadherent properties of P. falciparum parasites may thus be a virulence factor in human falciparum malaria. However, there was no correlation between the degree of in vitro cytoadherence and cerebral symptoms, which suggests that other receptors and/or host factors may be important in the adherence of malaria parasites to cerebral vascular endothelium. The cytokines tumor necrosis factor, interleukin-1, and gamma interferon, which have been implicated in the pathogenesis of cerebral malaria and are known to promote intercellular adhesion in other systems, did not enhance the cytoadherence of P. falciparum isolates to C32 melanoma cells.

Adult↗
PubMed1991

Hidden severe psychiatric morbidity in sentenced prisoners: an Australian study.

OBJECTIVE: The aim of this survey was to estimate the prevalence of severe mental disorders in a representative sample of sentenced prisoners. METHOD: The subjects were selected as a random sample of sentenced prisoners in Melbourne's three metropolitan prisons. Interviews were conducted with 158 men and 31 women. Clinicians used the Structured Clinical Interview for DSM-III-R (SCID) to diagnose psychotic, affective, and substance use disorders. RESULTS: Six prisoners (3%) received current diagnoses of psychotic disorders, and 23 (12%) were diagnosed as having current mood disorders, mainly major depression. A lifetime diagnosis of at least one mental disorder each was made for 82% of the respondents, and in 26% more than one lifetime disorder was diagnosed. Sixty-nine percent received lifetime diagnoses of dependence on or abuse of alcohol, other psychoactive substances, or a combination of these. CONCLUSIONS: These findings do not indicate a large-scale shift of deinstitutionalized psychotically ill people from mental hospitals to prisons. They do, however, highlight the diversion into the corrections system of substance-dependent people and the apparent pool of prisoners with largely untreated major depression.

Adolescent↗
PubMed1991

An educational approach to leprosy control: an evaluation of knowledge, attitudes and practice in two poor localities in Bombay, India.

Based on the hypothesis that a systematic, carefully planned educational approach to leprosy would yield results in terms of knowledge, attitudes and case presentation superior to those of the established and traditional mass survey method, ALERT-India launched a programme in S ward of Bombay in February 1985, to compare the two. An intensive programme of health education, using trained teams, was carried out in one zone of this ward over a period of 12 months. Eight months later, mass survey work (as used routinely in previous years and on a country-wide basis) was carried out in an adjacent zone. In 1987, the Centre for Social and Technological Change in Bombay, in association with the School of Oriental and African Studies, University of London, was requested to evaluate the effect of the above educational approach in terms of knowledge, attitudes and practice in both the trial and control zones. Other aspects of this experimental approach, including its cost and effectiveness in identifying cases of leprosy, will be published separately. The design of the 'KAP' evaluation and the social and environmental controls introduced in the statistical analysis are described. The results pointed to a considerable degree of ignorance about leprosy as a disease (and its treatment) in both the study and the control zones. Knowledge about early symptoms was particularly weak and on all aspects scores for women were invariably lower than men. General education enhanced the absorption of specific knowledge, and the education of children compensated adequately for lack of parental education in this respect. Overall the evaluation indicated that the intensive educational approach was superior to the survey approach in terms of improving knowledge, attitudes and practice.

Educational Status↗
PubMed1991

Temperature induced changes in microsomal (Na+,K+)-ATPase and lipid composition of Candida kefyr.

Growth temperature affected both the membrane lipid composition and microsoma (Na+, K+)-ATPase activity of Candida kefyr. Higher growth, temperature (37 degrees C) increased the amount of total lipids, phospholipids and free sterol. Ratios of phosphatidylcholine to phosphatidylethanolamine as well saturated to unsaturated fatty acids increased with a rise in growth temperature. Km of the ATPase isolated from the yeast grown at 27 degrees C was minimum, suggesting that the membranes of C. kefyr grown at optimal growth temperature provide the most suitable environment for the activity of ATPase.

Candida↗
PubMed1991

Evaluation of a new hydrocolloid occlusive dressing for central catheters used in total parenteral nutrition.

Catheter-related sepsis still remains one of the most frequent and serious complication of total parenteral nutrition. Strategies for preventing contamination of central venous lines have focused on decreasing the number of micro-organisms around the exit site and inhibiting their entry into the catheter wound. This prospective study compares a new occlusive hydrocolloid dressing (Visiband; Convatec Squibb) with that of a polyurethane film dressing for nutritional catheters. Dressings were changed either on day 3 or day 5 after application. Swab smears of the catheter exit site at each dressing change were stained by Gram's method before inoculation onto a blood agar plate, a chocolate agar plate and a MacConkey agar plate. Significantly less colonisation occurred under the former dressing at day 3 and day 5 dressing changes. In addition, the polyurethane film dressing was associated with a significant increase in skin colonisation (P = 0.04) and the number of positive Gram-stain microbes if left unchanged for 5 days (P = 0.0018). Staphylococcus aureus catheter-related sepsis occurred in 1 patient on day 18 in the polyurethane film dressing group. In addition, Candida albicans colonisation was confined to patients with the polyurethane film dressing. While the type of dressing applied to the catheter exit site may influence the incidence of catheter colonisation and infection, it must be emphasised that strict adherence to aseptic technique during catheter insertion and manipulation of the dressing is vital in the prevention of catheter-related sepsis during total parenteral nutrition.

Adult↗
PubMed1991

The role of c-mos proto-oncoprotein in mammalian meiotic maturation.

The developmentally regulated expression of c-mos oncogene has led to the speculation that this gene may be involved in gametogenesis and early development. To directly test this possibility, we have used an electric field mediated transfer method to introduce an antibody against the c-mos gene product into living immature mouse oocytes. Control oocytes exposed to the electric field without antibody, non-immune IgG, or c-mos antibody pre-absorbed with the mos peptide underwent normal germinal vesicle breakdown (GVBD) (90%) and formed a polar body by 8 h. Oocytes transferred with antibody against c-mos product underwent GVBD, and chromosome condensation as judged by Hoechst 33258 staining. However, antibody transferred oocytes did not form a polar body. Confocal fluorescence microscopy using antibodies against tubulin demonstrated that 90% of the oocytes that received antibody against c-mos did not assemble a meiotic spindle. In a few instances an abnormal spindle-like structure did form. Electron microscopy confirmed that the nuclear envelope disassembled and revealed many microtubules in a disorganized manner. Western blot analysis showed the presence of p39c-mos in mouse mature oocytes, spermatocytes and granulosa cells. These results suggest a role for c-mos in regulating the assembly and/or function of the spindle during meiotic division in murine.

Animals↗
PubMed1991

Inhibition of c-mos protein kinase blocks mouse zygotes at the pronuclei stage.

The c-mos gene product is required for activation of the maturation-promoting factor (MPF) during oocyte maturation. The c-mos protein also acts as a cytostatic factor which is responsible for meiotic metaphase arrest of vertebrate eggs via stabilization of MPF. Here we show that mouse zygotes contain the c-mos protein. Introduction of a kinase-inhibitory anti-mos antibody into mouse zygotes 12 h after fertilization prevented the first cleavage of zygotes at the pronuclei stage. A second anti-mos antibody, known to allow the mos kinase to function, did not interfere with the formation of two-cell embryos. In addition to its known role in MPF activation in oocyte maturation and meiotic metaphase arrest, our findings indicate that the c-mos protein kinase is also required for completion of pronuclei breakdown following fertilization of mouse eggs.

Amino Acid Sequence↗
PubMed1991

Phosphorylation of v-mos Ser 47 by the mitotic form of p34cdc2.

P85gag-mos is hyperphosphorylated during mitosis in normal rat kidney (NRK) cells transformed by Moloney murine sarcoma virus ts110. We now report that P85gag-mos is phosphorylated in vitro by the mitotic form of the cdc2 kinase (p34cdc2, known as M-phase kinase) derived from virus-transformed cells. The major site of P85gag-mos phosphorylation by the M-phase kinase in vitro lies within the amino-terminal portion of the viral mos protein sequence spanning residues 45-53, as determined by tryptic peptide mapping. A synthetic peptide corresponding to amino acids 37-55 of v-mos was specifically phosphorylated by the M-phase kinase, whereas v-mos peptides either lacking Ser 47 or substituted with Ala at residue 47 were not phosphorylated. Protein sequencing analyses established that the M-phase kinase specifically phosphorylates Ser 47. Tryptic phosphopeptide mapping of the in vivo-phosphorylated gag-mos protein from mitotic cells indicated that the 45-53 v-mos region was also phosphorylated within mitotic cells. These findings demonstrate that the M-phase kinase phosphorylates the viral mos protein at Ser 47. These results were unexpected in view of earlier reports regarding cdc2 kinase activation/stabilization by the c-mos kinase in maturing oocytes.

Animals↗
PubMed1991

Plasma renin activity in diabetics with and without microangiopathy.

Plasma renin activity was measured in ten diabetics with nephropathy, 10 uncomplicated diabetics and 10 normal healthy controls. All the groups were comparable for age, sex, and duration of diabetes. Plasma renin activity was found to be significantly lower in patients with nephropathy in comparison to uncomplicated diabetics and normal healthy controls. There was no correlation between plasma renin activity and mean blood pressure or degrees of renal failure. These findings raise the possibility that microangiopathic changes occurring in the glomeruli and juxta-glomerular apparatus may alter the renin-angiotensin system. Also the finding of low renin in diabetics with nephropathy may modify management policies.

Adult↗
PubMed1990

Regulatory function for murine intraepithelial lymphocytes. Two subsets of CD3+, T cell receptor-1+ intraepithelial lymphocyte T cells abrogate oral tolerance.

The murine intraepithelial lymphocyte (IEL) population is enriched in T cells that express the gamma delta-TCR, however, the biologic function served by these T cells remains obscure. IEL are considered to be major effector cells in mucosal immunity, and we have investigated whether IEL subsets could reverse orally induced systemic unresponsiveness (oral tolerance; OT) and support secondary type responses when adoptively transferred to mice orally tolerized with SRBC. When purified CD3+ IEL from mice orally primed with SRBC were transferred to adoptive hosts and challenged with SRBC, splenic IgM, IgG1, IgG2b, and IgA anti-SRBC plaque-forming cell responses were observed. However, CD3+ IEL from HRBC orally primed mice did not abrogate SRBC induced OT. Further, HRBC-primed CD3+, IEL converted HRBC-specific OT but not SRBC-specific OT. CD3+ IEL could be separated into four subsets based on expression of CD4 and CD8. CD3+, CD4-, 8+ T cells were the major subset (74.5%), with smaller numbers of CD4- and CD8- (double negatives, DN) (7.8%), CD4+, 8- (7.6%) and CD4+, CD8+ (double positives) (10.1%) T cells. Interestingly, both the CD3+, CD8+, and the CD3+, DN IEL subsets abrogated OT, resulting in significant IgM, IgG1, IgG2b, and IgA anti-SRBC plaque-forming cell responses when adoptively transferred to mice with OT. However, neither CD3+, CD4+, CD8-, nor double positive T cells affected OT when studied in this system. The CD3+, CD8+ IEL subset could be further separated into Thy-1+ (16.6%) and Thy-1- (83.4%) cells; adoptive transfer of Thy-1- cells abrogated oral tolerance whereas the Thy-1+ subset was without effect. When the expression of TCR on IEL with this biologic function was determined by use of monoclonal anti-alpha beta TCR (H57.597), TCR2-, CD3+ IEL possessed immunoregulatory function whereas the alpha beta-TCR+ (TCR2+) fraction did not abrogate OT. Immunoprecipitation of membrane fractions obtained from purified CD3+, CD4-, CD8+, Thy-1- IEL with polyclonal anti-delta peptide (Tyr-Ala-Asn-Ser-Phe-Asn-Asn-Glu-Lys-Leu) antibody revealed bands of 45 and 35 kDa, corresponding to the delta- and gamma-chains, respectively. These results suggest that gamma delta-TCR+ IEL possess a regulatory function, namely the restoration of immune responses in a state of oral tolerance. Further, both CD3+, CD4-, CD8+, Thy-1-, and CD3+, DN IEL T cells exhibit this effector contrasuppressor function.

Administration, Oral↗