Biomedical subjects

B Singh

Publications and source records attributed to B Singh.

At least 631 records · Page 35Linked to original sources

PubMed1990

Mitochondrial import of the human chaperonin (HSP60) protein.

The mitochondrial import of a member of the "chaperonin" group of proteins which play an essential role in the import of protein into organelles and their subsequent proper folding has been examined. The cDNA for human hsp60 (synonyms: GroEL homolog, P1) was transcribed and translated in vitro and its import into isolated rat heart mitochondria examined. The protein was converted into a mature form of lower molecular mass (= 58 kDa) which was resistant to trypsin treatment. The import of human hsp60 into mitochondria was inhibited in the presence of an uncoupler and also no import occurred when the N-terminal presequence was lacking. These results indicate that the chaperonin protein(s) are transported into mitochondria by a process similar to other imported mitochondrial proteins. Our results also indicate that although the P1 protein precursor was efficiently imported into mitochondria, in comparison to precursors of other mitochondrial proteins (viz. ornithine carbamoyltransferase and uncoupling protein) much less binding of pre P1 to mitochondria was observed. The significance of this latter observation at present is unclear.

Amino Acid Sequence↗

PubMed1990

The cyclin B2 component of MPF is a substrate for the c-mos(xe) proto-oncogene product.

Previous studies from this laboratory have shown that purified MPF from Xenopus eggs contains cyclin B2 complexed with cdc2 kinase. The activation of MPF during oocyte maturation is known to require expression of the c-mos(xe) proto-oncogene. We show here that immunoprecipitates of either v-mos from Moloney murine sarcoma virus-transformed NIH 3T3 cells or c-mos from Xenopus eggs phosphorylate cyclin B2 in vitro. Phosphopeptide analysis reveals a pattern similar to that observed with cdc2 kinase. Moreover, ablation of c-mos(xe) from oocytes by antisense oligonucleotide injection reduces the rate of cyclin B2 phosphorylation in oocyte extracts by 40%. These results suggest that the mechanism of activation of MPF by c-mos(xe) involves phosphorylation of the cyclin component.

Animals↗

PubMed1990

Activation-induced cell death in T cell hybridomas is due to apoptosis. Morphologic aspects and DNA fragmentation.

Some T cell hybridomas, upon activation via the TCR, rapidly undergo cell death. In this paper, we demonstrate that this activation-induced cell death (AICD) is accompanied by morphologic changes seen at the electron and light microscopy levels. The most striking changes are an extensive condensation of the chromatin and formation of membrane blebs. In addition to the morphologic changes, a significant portion of genomic DNA is broken at an interval of approximately 200 bp, producing a ladder of oligonucleosome-sized fragments after gel electrophoresis. Taken together, these observations indicate that AICD proceeds via apoptosis, or programmed cell death. This is additionally supported by the observation that AICD-associated phenomena are at least partially inhibited by cycloheximide or actinomycin D. Curiously, AICD and its associated DNA fragmentation are completely inhibited by aurintricarboxylic acid, a known nuclease inhibitor. The possible relationship between AICD in vitro, and the negative selection process (wherein selection may proceed via AICD of developing, autoreactive thymocytes) is discussed.

Aurintricarboxylic Acid↗

PubMed1990

Heterogeneity of presenting features and their relation to treatment outcome in 120 children with T-cell acute lymphoblastic leukemia.

Presenting features of 120 consecutive children with T-cell acute lymphoblastic leukemia (ALL), representing 15% of all patients diagnosed as having ALL during the study period, were analyzed to determine relationships with treatment outcome. Patients' ages ranged from 1.7 to 18.8 years (median, 10.3 years) and their leukocyte counts from 1.7 to 1,070 x 10(9)/L (median, 100 x 10(9)/L). Central nervous system (CNS) leukemia was present in 12.5% of the cases, a mediastinal mass in 61%, and L2 lymphoblast morphology in 32%. A relatively high proportion of cases, 26%, had normal karyotypes at presentation. Of the cases tested, membrane CD1 expression was found in 38% of cases, CD3 in 33%, CD4 in 50%, CD5 in 94%, CD8 in 55%, and CD10 in 35%. Four presenting features were found to confer an increased risk of treatment failure: age greater than or equal to 15 years, L2 lymphoblast morphology, abnormal karyotype, and membrane CD3 expression. This study illustrates the heterogeneity of presentations of childhood T-cell ALL and suggests that the relative importance of risk factors in ALL differs according to immunophenotype and treatment strategy.

Age Factors↗

PubMed1990

Myeloid-associated antigen expression lacks prognostic value in childhood acute lymphoblastic leukemia treated with intensive multiagent chemotherapy.

Frequency and clinical significance of myeloid-associated antigen expression in blast cells were assessed in 372 consecutive children with acute lymphoblastic leukemia (ALL). A comprehensive panel of myeloid monoclonal antibodies representing seven cluster groups showed myeloid-associated antigen expression in 61 cases (16.4%), 18 of which expressed two or more antigens. The antigens expressed comprised CD11b (8.9% of the total series), CD13 (6.5%), CD33 (3.2%), CD36 (1.9%), CD15 (1.6%), CD14 (1.3%), and CDw12 (1.1%). No significant associations were found between myeloid-associated antigen expression and the presence of known adverse prognostic features (eg, higher leukocyte count, nonwhite race, older age). Myeloid-associated antigen expression had no effect on remission induction or event-free survival for the 267 children who had been treated with the same combination of chemotherapeutic agents (P = .34). Thus, blast cell expression of myeloid-associated antigens in childhood ALL appears to lack prognostic value in the context of contemporary intensive chemotherapy.

Antibodies, Monoclonal↗

PubMed1990

Critical role of an amino acid residue in a T cell determinant is due to its interaction with a neighboring non-critical residue.

Several lines of evidence support the concept of two functionally distinct sites on antigen: the epitope, involved in interaction with the T cell receptor and the agretope, interacting with Ia. We investigated the Ia and T cell receptor interaction sites on the synthetic polypeptide antigen poly-18 [poly-EYK(EYA)5] using T cell hybridoma clones specific for this antigen in the context of I-Ad. Peptides with amino acid sequences related to poly-18 were synthesized. These were used to identify the critical residues in the minimum peptide sequence required for activation. Clone A.1.1 responds to the minimal peptide EYK(EYA)4 but not to (EYA)5. This identifies Lys3 as a critical amino acid for this hybridoma. Surprisingly, the substituted peptide EYAEAA(EYA)3 could activate A.1.1, indicating that an Ala at position 5 instead of a Tyr obviates the critical requirement for Lys3. This demonstrates that the function of critical residues may extend beyond contacting the T cell receptor or Ia, to include a third role: that of interacting with other amino acids of the T cell epitope, thus influencing the antigen's recognition by T cells.

Amino Acid Sequence↗

PubMed1990

Prediction of the secondary structure and functional sites of major histocompatibility complex molecules.

A method for the theoretical prediction of the antigenic determinants and the antigen-interactive receptor sites of immunological proteins from their primary structure would constitute a useful tool for their study. Such a method developed in this laboratory uses hydrophilicity, accessibility, flexibility, and recognition profiles, together with the predicted secondary structure (alpha-helices, beta-sheets, and turns). The secondary structure is determined by a modification of the method of Lim (1974), as described below. A study of human and mouse class I and class II major histocompatibility complex (MHC) antigens, central to the regulation of immune responses and to the phenomenon of graft rejection, was carried out using the above method. Comparison of the predictions with some of the available experimental and theoretical information supports the validity and usefulness of the approach.

Animals↗

PubMed1990

Volatile fraction of neem oil as a spermicide.

The volatile, odorous fraction of neem oil coded as NIM-76 obtained by steam distillation was investigated for in vitro spermicidal activity. The data showed that the minimum concentration which inhibited spermatozoal motility was 0.25 mg/ml for rat and 25 mg/ml for human spermatozoa. The effect of the drug on spermatozoal motility was found to be dose-dependent. The activity of this drug was not altered in the presence of vaginal or cervical mucus. Intra-vaginal application of NIM-76 in rabbits showed no irritation to the vaginal mucosa.

Animals↗

PubMed1990

Open pelvic fractures.

We reviewed 32 patients with open pelvic fractures treated over a 5-year period. Eleven patients had isolated fractures of the ilium and in 21 patients pelvic ring disruption was associated with a perineal laceration. Three patients died from massive haemorrhage soon after admission and one patient from severe sepsis. Early pelvic stabilization and meticulous attention to the soft tissue injury proved invaluable in the management of open unstable pelvic fractures.

Adolescent↗

PubMed1990

Mood changes, obstetric experience and alterations in plasma cortisol, beta-endorphin and corticotrophin releasing hormone during pregnancy and the puerperium.

The relationships between mood change, obstetric experience and alterations in plasma cortisol, beta-endorphin (beta-EP) and corticotrophin-releasing hormone (CRH) were examined in a prospective study of 97 primiparous Australian women. Psychological measures were administered between the 28th week of pregnancy and the 3rd postnatal month, including the Profile of Mood States (POMS) and the Montgomery Asberg Depressive Rating Scale (MADRS). Blood samples were collected for cortisol, beta-EP and CRH assay on most of these occasions and during labour. Factor analysis was used to identify key subsets of psychological variables for use in the subsequent analyses. 'Mood disturbance' and 'tiredness' factors peaked at 38 weeks' gestation, while 'difficulty falling asleep' was greatest around the time of birth. Cortisol, beta-EP and CRH concentrations rose significantly as pregnancy advanced and peaked at birth; plasma CRH correlated with plasma cortisol (r = 0.54) and beta-EP (r = 0.32). Women with the highest 'mood disturbance' and MADRS depression scores at 28 weeks' gestation received significantly more pain relief during labour. Those women whose mood deteriorated from 38 weeks' gestation to postnatal day 2 had larger falls in plasma beta-EP after delivery (p less than 0.01) than those women whose mood improved or remained constant. Women in this mood-deteriorated subgroup also had significantly higher MADRS depression scores at 3 months (p less than 0.01). Mild antenatal depression (MADRS greater than 13) occurred in 5.2% of women and mild postnatal depression in 4.7%. Overall, these data suggest a role for circulating CRH in the regulation of maternal cortisol secretion and significant relationships between maternal postnatal mood states and beta-EP and between antenatal mood states and obstetric events.

Adult↗

PubMed1990

Enhanced mutability at the tk locus in the radiosensitive double-strand break repair mutant xrs5.

The thymidine kinase locus (tk) has been utilised as the target locus to measure the induced mutation frequency following X-irradiation in the X-ray-sensitive xrs5 mutant and its parent CHO K1 line of Chinese hamster cells. Mutations to tk- cells were measured by plating cells in selective medium containing trifluorothymidine after a post-irradiation expression time of 4 days. Our results show that the mutation frequency was 3-4 times higher in the xrs5 mutant than in the CHO K1 cell line. This enhanced mutation frequency in xrs5 is though to result from the deficiency in DNA double-strand break repair in this cell line which also results in the enhanced cell killing and higher frequencies of chromosomal aberrations in response to X-irradiation. The findings of the present study suggest that DNA double-strand break is a critical lesion leading to mutations in irradiated cells.

Animals↗

PubMed1990

Use of antipeptide antibodies to probe the catalytic activity of p37v-mos.

Several site-directed antipeptide antibodies were generated to probe the kinase function of p37v-mos. Anti-mos(37-55) antibodies allowed autophosphorylation of p37v-mos, as well as transphosphorylation of exogenously added purified bovine vimentin. Rabbit antipeptide antibodies against v-mos residues 158-70, 194-206, 260-271, and 363-374 and a mouse monoclonal antibody against residues 344-359 completely inhibited p37v-mos protein kinase activity in vitro. p37v-mos autophosphorylation and vimentin transphosphorylation were affected similarly. These results suggest important roles for insert and the carboxy-terminal domains in the catalytic activity of p37v-mos.

Amino Acid Sequence↗

PubMed1990

Characterization of agretopes and epitopes involved in the presentation of beef insulin to T cells.

Beef insulin-specific I-Ad-restricted T cell hybridomas were derived from the fusion of antigen-primed (BALB/c X B6)F1 T cells with BW5147 thymoma. Specificity analysis revealed that the A-chain loop region is involved in antigen recognition. Hybridoma A20.2.15 is specific for beef insulin and cross-reacted with sheep insulin, but not with pork insulin. Using synthetic peptides we showed that the A-chain loop containing peptide A1-A14 jointed to the B7-B15 peptide by a disulfide bond can activate this hybridoma. Fragments generated by enzyme digest further suggest that the peptide recognized on beef insulin appears to involve A-chain loop residues A5-A12 and B-chain residues B7-B13 that are linked by the A7-B7 disulfide bridge. We found that beef insulin needs to be processed prior to T cell activation. Glutaraldehyde fixation and chloroquine treatment of presenting cells abolished their capacity to present insulin. Beef insulin denatured by pH changes cannot activate, thus suggesting that simple denaturation is not sufficient for presentation by antigen presenting cells. Finally, the agretope on beef insulin is comprised of two functional regions B7-B13 on the B chain and the A-chain loop in the A-chain, while residues A8 and A10 are probably involved in interaction with the T cell receptor.

Amino Acid Sequence↗

PubMed1990

A numerical model for the oxygenation of blood in lung capillaries--effect of nth order one-step kinetics of oxygen uptake by haemoglobin.

A numerical model is described for the oxygenation of blood in lung capillaries by considering the transport mechanisms of molecular diffusion, convection and the facilitated diffusion due to the presence of haemoglobin. In order to represent the oxygen dissociation curve accurately in the model, the nth order one-step kinetics of oxygen uptake by haemoglobin has been used. The resulting system of coupled, non-linear partial differential equations is solved numerically. It is shown that the blood is required to traverse a larger distance in the capillary before becoming fully oxygenated with nth order one-step kinetics in comparison to first-order one-step kinetics.

Capillaries↗

PubMed1990

Role of the first external domain of I-A beta chain in immune responses and diabetes in non-obese diabetic (NOD) mice.

Diabetes in the non-obese diabetic (NOD) mouse is a multigenic autoimmune disease and is possibly controlled by three recessive loci, including one that is linked to the major histocompatibility complex (MHC). The first external domain of the Class II MHC I-A beta chain in these mice is unique and has been suggested as being responsible for autoimmunity. The I-A alpha chain in these mice is I-A alpha d, and they lack the expression of I-E molecules. We have investigated immune responses to various Ir gene control antigens in NOD mice to determine the influence of the NOD Ia and particularly the I-A beta chain. We find that sheep insulin is highly immunogenic while other insulins are weakly immunogenic in these mice. Hen egg lysozyme, pigeon cytochrome C and the synthetic polypeptide Poly 18, Poly EYK(EYA)5 antigen produce good antibody responses. Apart from H-2d, NOD are the only mice where Poly 18 antigen is immunogenic. In these mice Poly 18 induced good T-cell proliferative response, which was inhibited by anti-Ia antibody, and the mice were able to respond to tyrosine-containing polypeptide Poly EYA but not to the phenylalanine-containing antigen Poly EFA. We also found that synthetic peptide 48-60 of the NOD I-A beta chain is highly immunogenic in syngeneic NOD mice both for T cells and B cells. Using an I-A beta chain-specific monoclonal antibody, we are able to prevent induction of diabetes when the antibody was administrated in prediabetic, young mice. Our results suggest that the immune response to various antigens and autoimmune diabetes in NOD mice is directly influenced by the I-A beta chain.

Amino Acid Sequence↗

PubMed1990

Prevention of diabetes in the BB rat by early immunotherapy using Freund's adjuvant.

The BB rat spontaneously develops an insulin-dependent diabetes mellitus (IDDM) that closely resembles this disease in man. The pathogenesis involves autoimmune destruction of pancreatic islet beta-cells. In the present study, a single intraperitoneal injection of complete Freund's adjuvant (CFA) in diabetes-prone (DP) BB/Wor rats between 9 and 28 days of age reduced the incidence of diabetes at 120 days from 89% to 10-28%, whereas injection of CFA after 40 days of age was ineffective. The CFA-injected, diabetes-free DP rats had normal levels of pancreatic insulin and little or no mononuclear leukocyte infiltration in the islets. These protective effects of CFA were not associated with any changes in peripheral blood leukocyte counts or monoclonal antibody-defined T cell, B cell, or macrophage subsets in the spleen of the DP rats. These results suggest that it is possible to prevent diabetes by adjuvant treatment in early life without general immunosuppression or sustained therapy.

Adjuvants, Immunologic↗

PubMed1990

Bilateral choanal atresia: key to success with the transnasal approach.

The intranasal approach is generally accepted as the route of choice in the correction of bilateral choanal atresia but unfortunately it is associated with a significant incidence of stenosis. Over a five-year period, we have diagnosed bilateral choanal atresia in 30 neonates. Trans-nasal surgery was performed on 27 of them. All belonged to the Black race group. The incidence was 1:8,300 live births. There was a female predominance of 5:1. Four (13 per cent) had other associated anomalies. One was a twin. Various factors were analysed to determine the cause of the stenosis. These were: 1. Drilling compared with curettage of the bony atretic plate. 2. Stenting versus non-stenting. 3. Soft versus hard Portex tube stenting. 4. Variation of stenting period--six, eight and twelve weeks. 5. The use of prophylactic broad spectrum antibiotic for the whole of the stenting period. The conclusion derived from this study is that curettage of bony atresia, stenting with soft Portex tube for a period of six weeks and the use of broad spectrum antibiotic for the entire stenting period eliminates the problem of stenosis associated with the trans-nasal approach for the correction of bilateral choanal atresia.

Choanal Atresia↗

PubMed1990

A novel, nonsteroidal inhibitor of androgen binding to the rat androgen binding protein: diethyl [[[3-(2,6-dimethyl-4-pyridinyl)-4-fluorophenyl]amino]methylene] propanedioate.

In vitro binding studies demonstrate the binding specificity of a series of 4-aryl-2,6-dimethylpyridines for the rat epididymal androgen binding protein (rABP). The compounds bound competitively to rABP but have very weak or no demonstrable affinity for rat ventral prostate androgen receptor and human sex hormone binding globulin. In particular, compound 11, diethyl [[[3-(2,6-dimethyl-4-pyridinyl)-4-fluorphenyl]amino]methylene] propanedioate, bound with high affinity to rABP (binding affinity about 1/3 that of the endogenous ligand 5 alpha-dihydrotestosterone). However, additional in vitro binding studies indicated that 11 did not bind to testicular or epididymal ABP from rabbit, rhesus monkey, and human. Nevertheless, the specificity and relatively high affinity of these nonsteroidal compounds make them unique and potentially ideal agents for the study of the role of ABP in spermatogenesis and sperm maturation in the rat.

Androgen-Binding Protein↗