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Biomedical subjects

B Roy

Publications and source records attributed to B Roy.

At least 37 records · Page 2Linked to original sources
PubMed2000

Increases in submaximal cycling efficiency mediated by altitude acclimatization.

To investigate the hypothesis that respiratory gas exchange and, in particular, the O(2) consumption (VO(2)) response to exercise is altered after a 21-day expedition to 6,194 m, five male climbers (age 28.2 +/- 2 yr; weight 76.9 +/- 4.3 kg; means +/- SE) performed a progressive and prolonged two-step cycle test both before and 3-4 days after return to sea level. During both exercise tests, a depression (P < 0.05) in VO(2) (l/min) and an increase (P < 0.05) in minute ventilation (VE BTPS; l/min) and respiratory exchange ratio were observed after the expedition. These changes occurred in the absence of changes in CO(2) production (l/min). During steady-state submaximal exercise, net efficiency, calculated from the rates of the mechanical power output to the energy expended (VO(2)) above that measured at rest, increased (P < 0.05) from 25.9 +/- 1.6 to 31. 3 +/- 1.3% at the lighter power output and from 24.4 +/- 1.3 to 29.5 +/- 1.5% at the heavy power output. These changes were accompanied by a 4.5% reduction (P < 0.05) in peak VO(2) (3.99 +/- 0.17 vs. 3.81 +/- 0.18 l/min). After the expedition, an increase (P < 0.05) in hemoglobin concentration (15.0 +/- 0.49 vs. 15.8 +/- 0.41 g/100 ml) was found. It is concluded that, because resting VO(2) was unchanged, net efficiency is enhanced during submaximal exercise after a mountaineering expedition when the exercise is performed soon after return to sea level conditions.

Acclimatization↗
PubMed2000

Assessment of androclonal variation in an indica rice PTB 28.

Anther culture (AC) derived 160 doubled haploid (DH) families of an indica rice PTB 28 were field evaluated at Port Blair (lat 11 degrees.41'13.04"N; long 92 degrees 43'30.16"E), to assess the extent and range of induced variability among the androclones at segregating A2 generation and identify promising selectants for genetic improvement. Though the coefficient of variation (CV) for plant height was least (2.65%), majority of the androclones were shorter (81.7%). CV was high in case of panicle number per plant (24.59%) and tiller number per plant (28.28%), while it was maximum in respect of yield per plant (37.0%). Tiller and panicle number per plant registered maximum reduction compared to parent, however, displayed larger variations in double haploid (DH) lines. Out of 160 DH families, only 10 (6.3%) lines out-yielded the parent. Extreme types with wide variations were encountered which reaffirms the possibility of employing anther culture in rice improvement by restoring yield advantage in advanced generations or by tailoring them through recombination breeding.

Evaluation Studies as Topic↗
PubMed1999

Successful myoblast transplantation in fibrotic muscles: no increased impairment by the connective tissue.

BACKGROUND: Implantation of normal myoblasts may eventually be a treatment for inherited myopathies such as Duchenne muscular dystrophy. METHODS: We report a comparative study of the effectiveness on myoblast implantation: (1) into the muscles of young (2 months) mdx mice nonirradiated and noninjected with notexin (group 1), (2) into muscles of old mdx mice (15 months) nonirradiated and noninjected with notexin (group 2), and (3) into muscles of 5 months mdx mice irradiated 3 months before the transplantation (group 3). Roughly 3 million cells were injected with bFGF in the Tibialis anterior. RESULTS: Although mice of groups 2 and 3 had significantly more (P<0.05) fibrotic tissue in their muscles than those of group 1, the transplantation success was not significantly different among the three groups. CONCLUSION: Therefore these results demonstrated that myoblast transplantation can be successful even when there is abundant fibrosis.

Aging↗
PubMed1999

Simultaneous determination of pyrimidine or purine deoxyribonucleoside triphosphates using a polymerase assay.

In this paper, we describe an improved enzymatic assay for the determination of deoxyribonucleoside triphosphates (dNTPs). This is based on the elongation of 32P 5'-end-labeled oligonucleotide primers annealed to complementary oligonucleotide templates. Incorporation within the primer/template (p/t) was catalyzed by the Klenow fragment of Escherichia coli DNA polymerase I under conditions where the concentration of the dNTP to be analyzed is limiting. Using a combination of two different sized p/t pairs, dCTP and dTTP (or dATP and dGTP) were assayed together. Since the elongated products were clearly separated after electrophoresis on a denaturing 10% polyacrylamide gel, the two dNTPs could be quantified in a single lane. This method allows for the first time the simultaneous determination of two pyrimidine or two purine deoxyribonucleoside triphosphates. Consequently, a large number of biological samples can be tested in a single experiment. The high sensitivity of this method enables the quantification of low concentrations of dNTPs, such as those found in resting nondividing cells. Furthermore, this new protocol is well suited for the determination of dNTPs in cells treated with the antiretroviral ddI, since the Klenow fragment has a low affinity for ddATP, the active form of ddI.

Anti-HIV Agents↗
PubMed1999

Downregulation of JAK3 protein levels in T lymphocytes by prostaglandin E2 and other cyclic adenosine monophosphate-elevating agents: impact on interleukin-2 receptor signaling pathway.

The Janus kinase, JAK3 plays an important role in interleukin-2 (IL-2)-dependent signal transduction and proliferation of T lymphocytes. Our findings show that prostaglandin E2 (PGE2) can inhibit upregulation of JAK3 protein in naive T cells and can downregulate its expression in primed cells. Reduction in JAK3 was selective because expression of other tyrosine kinases (JAK1, p56(lck), and p59(fyn)) and signal transducer and activator of transcription (STAT)5, which are linked to IL-2 receptor (IL-2R) signaling pathway, were not affected. Inhibition of JAK3 may be controlled by intracellular cyclic adenosine monophosphate (cAMP) levels, as forskolin, a direct activator of adenylate cyclase and dibutyryl cAMP (dbcAMP), a membrane permeable analogue of cAMP suppressed JAK3 expression. Moreover, 3-isobutyl-1-methylxanthine (IBMX), an inhibitor of cAMP phosphodiesterase, potentiated PGE2-induced suppression of JAK3. In naive T cells, but not primed T cells, PGE2 and other cAMP elevating agents also caused a modest reduction in surface expression of the common gamma chain (gammac) that associates with JAK3. The absence of JAK3, but not IL-2R in T cells correlated with impaired IL-2-dependent signal transduction and proliferation. The alteration in IL-2 signaling included decreased tyrosine phosphorylation and DNA binding activity of STAT5 and poor induction of the c-Myc and c-Jun pathways. In contrast, IL-2-dependent induction of Bcl-2 was unaffected. These findings suggest that suppression of JAK3 levels may represent one mechanism by which PGE2 and other cAMP elevating agents can inhibit T-cell proliferation.

1-Methyl-3-isobutylxanthine↗
PubMed1999

The mammalian endoplasmic reticulum stress response element consists of an evolutionarily conserved tripartite structure and interacts with a novel stress-inducible complex.

When mammalian cells are subjected to calcium depletion stress or protein glycosylation block, the transcription of a family of glucose-regulated protein (GRP) genes encoding endoplasmic reticulum (ER) chaperones is induced to high levels. The consensus mammalian ER stress response element (ERSE) conserved among grp promoters consists of a tripartite structure CCAAT(N9)CCACG, with N being a strikingly GC-rich region of 9 bp. The ERSE, in duplicate copies, can confer full stress inducibility to a heterologous promoter in a sequence-specific but orientation-independent manner. In addition to CBF/NF-Y and YY1 binding to the CCAAT and CCACG motifs, respectively, we further discovered that an ER stress-inducible complex (ERSF) from HeLa nuclear extract binds specifically to the ERSE. Strikingly, the interaction of the ERSF with the ERSE requires a conserved GGC motif within the 9 bp region. Since mutation of the GGC triplet sequence also results in loss of stress inducibility, specific sequence within the 9 bp region is an integral part of the tripartite structure. Finally, correlation of factor binding with stress inducibility reveals that ERSF binding to the ERSE alone is not sufficient; full stress inducibility requires integrity of the CCAAT, GGC and CCACG sequence motifs, as well as precise spacing among these sites.

Adaptation, Biological↗
PubMed1999

Successful myoblast transplantation in primates depends on appropriate cell delivery and induction of regeneration in the host muscle.

Myoblast transplantation (MT) may be a potential treatment for severe recessive hereditary myopathies. The limited results of MT in clinical trials led us to improve this technique in monkeys, an animal model phylogenetically similar to humans. Three Macaca mulata monkeys were used as donors and six as receivers for MT. Myoblasts were grown in culture from muscle biopsies of adult monkeys and infected with a retroviral vector encoding the LacZ gene. Different numbers of cells (i.e., 4 x 10(6), 8 x 10(6), and 24 x 10(6) cells) were transplanted into different muscles and 8 x 10(6) cells (resuspended in a notexin solution) were injected in one muscle of four monkeys. For these transplantations, the cell suspension (in a volume of about 100 microl) was injected at 35 sites less than 1 mm apart. Two other monkeys received 100 x 10(6) myoblasts resuspended in 1 ml of HBSS or 1 ml of notexin. For these two monkeys, the myoblasts were injected at 200-250 sites within a small portion of the muscle. All monkeys were immunosuppressed with daily injections of FK506. Four weeks after MT, the transplanted muscle portions were biopsied and the presence of beta-galactosidase-positive (beta-Gal+) muscle fibers was investigated. The number of beta-Gal+ fibers was 822 +/- 150 (site grafted with 4 x 10(6) cells), 1253 +/- 515 (8 x 10(6) cells), 1084 +/- 278 (24 x 10(6)), and 2852 +/- 1211 (notexin). In the monkeys grafted with 100 x 10(6) myoblasts, the number of beta-Gal+ fibers was 4850 (site without notexin) and 9600 (site with notexin). We demonstrated that a precise mechanical distribution of myoblasts into the tissue improves substantially MT in primates. The presence of notexin with the transplanted cells further increased the success of their transplantation. These are the best results obtained either with MT or gene therapy in primates and they encourage the possibility to human MT trials.

Animals↗
PubMed1999

Nitric oxide: a radical molecule in quest of free radicals in proteins.

A number of enzymes use an amino acid free radical cofactor. Tyrosyl and tryptophanyl radicals react with nitric oxide (NO) with an almost diffusion-limited rate. The catalytically competent tyrosyl radical in ribonucleotide reductase (RR) and prostaglandin H synthase (PGHS) recombines with NO in a radical-radical reaction. The unstable adduct formed can dissociate to regenerate the tyrosyl radical. However, upon prolonged incubation with NO, the diiron center of mouse RR leaks out, while the adduct is successively oxidized into an iminoxyl radical and a nitrotyrosine in PGHS. These data provide a plausible mechanism for the physiological inactivation of RR observed in various models, and may help in understanding the inhibition of PGHS reported in some cases. Reversible combination with NO is an intrinsic property of tyrosyl radicals, which also occurs with Y(D) and Y(Z) in photosystem II, where NO has been useful in the analysis of the oxygen-evolving complex.

Amino Acids↗
PubMed1999

Modulation of calcium mobilization in aortic rings of pregnant rats: Contribution of extracellular calcium and of voltage-operated calcium channels.

Pregnancy is associated with decreased vascular responsiveness to vasopressor stimuli. We have tested the involvement of Ca2+ mobilization in myotropic responses of aortic rings obtained from pregnant and virgin rats. Contractions of the rings to phenylephrine, in the absence of calcium in the bathing medium, were lower in tissues from virgin than from pregnant rats. Concentration-response curves to CaCl2 that were measured after stimulation by phenylephrine in the absence of Ca2+ were shifted to higher levels of contraction. This was not observed when KCl was used to prestimulate the aorta. D-600, a phenylalkylamine calcium channel blocker, similarly inhibited these responses to CaCl2 in tissues from both pregnant and virgin animals. D-600 exerted a concentration-dependent inhibition of responses to phenylephrine and KCl. However, the calcium antagonist was less effective in aortic rings of pregnant than of virgin rats. Basal 45Ca2+ uptake was lower in aortic rings from pregnant than from virgin rats, and Bay K 8644 was unable to reverse this difference. The time course of basal and stimulated (KCl) 45Ca2+ influx was lower in aorta of pregnant rats at all times studied. Moreover, when the intracellular calcium pools were emptied with phenylephrine, the refilling of these pools was delayed in aortic rings of pregnant rats. These results indicate an altered extracellular calcium mobilization of aortic rings from pregnant rats. These changes may be due to a functional alteration of the voltage-operated calcium channels during pregnancy.

Animals↗
PubMed1999

Transplantation of human myoblasts in SCID mice as a potential muscular model for myotonic dystrophy.

Myotonic dystrophy (DM), the most frequent hereditary myopathy in adults, is characterized clinically by muscle weakness, myotonia, and systemic symptoms. Although the specific genetic basis for DM has been established, less is known about the cellular defects responsible for its pleiotropic manifestations. DM pathogenesis studies are presently limited due to the absence of animal models. In the present study, we transplanted myoblasts of DM patients into the Tibialis anterior of Severe Combined Immunodeficient (SCID) mice to determine whether this approach could reproduce the muscular characteristics of DM. One to 4 months after transplantation, a variable number of innervated human muscle fibers, recognized by an antibody specific for the human dystrophin, were found in the transplanted muscles. The CTG expansion was retained in human muscle fibers as determined by Southern blot analysis. Although the histological characteristics of DM were absent in these fibers, electromyographic recording showed typical myotonic discharges in muscles transplanted with DM myoblasts. The specificity of the myotonic runs was demonstrated by its inhibition by apamin, a drug that specifically blocks DM myotonia. We conclude that transplantation of myoblasts from DM patients into SCID mice represents a potential in vivo model for basic studies of this disease.

Animals↗
PubMed1999

Variation at 4 short tandem repeat loci in 8 population groups of India.

We have determined the nature and extent of variation at 4 STR loci (CSF1P0, TPOX, TH01, VWA) in 8 caste and tribal population groups of eastern and northern India. Large differences in allele frequencies among the groups were found. Average heterozygosities in all populations were high (approximately 80%). The overall extent of gene differentiation among the 8 groups was high (GST = 0.04). The nature of genomic affinities based on these 4 STR loci does not completely agree with our earlier finding based on classical genetic markers that geographic proximity of habitat has a greater influence on genetic similarity between populations than sociocultural proximity does.

Developing Countries↗
PubMed1999

Negligible male gene flow across ethnic boundaries in India, revealed by analysis of Y-chromosomal DNA polymorphisms.

From the historically prevalent social structure of Indian populations it may be predicted that there has been very little male gene flow across ethnic boundaries. To test this finding, we have analyzed DNA samples of individuals belonging to 10 ethnic groups, speaking Indo-European or Austroasiatic languages and inhabiting the eastern and northern regions of India. Eight Y-chromosomal markers, two biallelic and six microsatellite, were studied. All populations were monomorphic for the deletion allele at the YAP (DYS287) locus and for the 119-bp allele at the DYS288 locus. Y-chromosomal haplotypes were constructed on the basis of one RFLP locus and five microsatellite loci. The haplotype distribution among the groups showed that different ethnic groups harbor nearly disjoint sets of haplotypes. This indicates that there has been virtually no male gene flow among ethnic groups. Analysis of molecular variance revealed that there was significant haplotypic variation between castes and tribes, but nonsignificant variation among ranked caste clusters. Haplotypic variation attributable to differences in geographical regions of habitat was also nonsignificant.

DNA↗
PubMed1998

Induction of 15-lipoxygenase expression by IL-13 requires tyrosine phosphorylation of Jak2 and Tyk2 in human monocytes.

The enzyme 15-lipoxygenase (15-LO) participates in the dioxygenation of polyenoic fatty acids. This activity leads to the degradation of mitochondrial membranes during reticulocyte differentiation, the production of pro- and anti-inflammatory mediators by a variety of cell types, and the oxidation of lipids in atherosclerotic lesions. The cytokines, IL-4 and IL-13, are reported to induce the expression of 15-LO in human peripheral blood monocytes. In this report we explore the signaling mechanisms involved in the IL-13-mediated induction of 15-LO expression. First we demonstrate that the delayed induction of 15-LO requires continuous stimulation of monocytes for a minimum period of 12 h. We also found that tyrosine kinase inhibitors blocked the induction of 15-LO in a dose-dependent manner. By immunoprecipitation and antiphosphotyrosine blotting experiments, IL-13 was shown to induce tyrosine phosphorylation of Jak2 and Tyk2, but not Jak1 or Jak3, within 5 min of treatment in human monocytes. To investigate whether the early induction of tyrosine phosphorylation of both Jak2 and Tyk2 was ultimately involved in 15-LO expression, we generated antisense oligodeoxyribonucleotides (ODNs) against Tyk2 and Jak2. We employed a cationic lipid-mediated delivery technique to transfect the monocytes and found that both antisense ODNs inhibited expression of their target proteins by 75-85%. The treatments were specific and did not affect the expression of each other. Furthermore, the antisense ODNs to Jak2 and Tyk2 both inhibited the induction of expression of 15-LO in monocytes treated with IL-13. Parallel experiments with sense ODNs to Jak2 and Tyk2 did not affect their protein levels or the induction of 15-LO by IL-13, and down-regulation of Jak1 also did not affect expression of 15-LO. Our results suggest the novel finding that IL-13 can induce tyrosine phosphorylation of both Jak2 and Tyk2 in primary human monocytes. This occurs as an early and essential signal transduction event for the IL-13-mediated induction of 15-LO expression. These data represent the first characterization of upstream kinases involved in the induced expression of 15-LO.

Arachidonate 15-Lipoxygenase↗
PubMed1998

Successful transplantation of genetically corrected DMD myoblasts following ex vivo transduction with the dystrophin minigene.

Myoblast transplantation and gene therapy are two promising therapeutical approaches for the treatment of Duchenne Muscular Dystrophy (DMD). So far, both strategies have met many hurdles, mainly because of immune reactions. In this study, we investigated a third and novel strategy based on the combination of these two basic ones, i.e., transplantation of genetically modified myoblasts. We first derived a primary culture from a muscle biopsy of a young DMD patient (3 years old). Adenoviral-mediated dystrophin gene transfer into these DMD cultures and expression of the dystrophin transgene were achieved in vitro. The transduced cultures were then transplanted the same day in immunodeficient SCID mouse muscles. Three weeks following the graft, many human dystrophin-positive fibers were observed throughout sections of the injected muscles. However, many fibers expressed human MHC antigens without expressing human dystrophin due to the low percentage of infected primary muscle cells in vitro (even when a high MOI [400] was used) and to a reduction and even to a complete loss of transgene copy number during myoblast replication. From our results, we conclude that, although not at a high proportion, (1) DMD primary myoblast cultures are infectable by adenoviruses; (2) they can be efficiently transplanted back in a muscle, leading to normal fusion of infected myoblasts with the host fibers; and (3) they can correct the dystrophin deficiency in the host fibers by the expression of a mini-dystrophin transgene.

Adenoviridae↗
PubMed1998

Application of least squares method in matrix form: simultaneous determination of ibuprofen and paracetamol in tablets.

Least squares method in matrix form which is K-matrix representation of Beer's law is presented for simultaneous determination of ibuprofen and paracetamol in tablets without prior separation from each other. The concentration of each component in the mixture was determined spectrophotometrically from absorbances of the mixture measured at 225, 226, 228, 232, 230, 234, and 235 nm. Mixtures of known composition were used as standards to minimise errors due to presence of both compounds in the same solution. Excellent results were obtained by this method.

Acetaminophen↗
PubMed1998

[Diagnostic value of ERTL4: a screening test of language disorders in 4-year-old children].

BACKGROUND: Language disorders in children are frequent and may sometimes severely affect their development. ERTL4 (screening test for language disorders of children aged 4) is the first test for screening language disorders to be used by doctors. It has not yet been evaluated. AIM: The objective of this study is to assess and optimize the diagnostic value of the ERTL4 test and to evaluate its potential interest as a screening test. MATERIAL AND METHODS: A sample of 370 children was randomly constituted among children aged between 3 years 9 months and 4 years 6 months from schools in Nancy, France and surrounding areas. These children undertook both the ERTL4 test and a standardized examination of language and speech as a reference. RESULTS: Among the 325 children included in the analysis, 34.1% presented with disorders requiring speech and language therapy. ERTL4 sensitivity and specificity were 88.6% and 66.8%, respectively. Varying thresholds for disorder diagnoses and proposals for a modified test led to a sensitivity of 72.9% and a specificity of 91.0%. The proportion of well-classified subjects improved from 73.8% to 85.3% (P < 10(-3)). The positive predictive value was 78.7% and the negative predictive value was 88.1% (with the observed frequency in the sample). CONCLUSION: ERTL4 is a good test for doctors to screening of children's language disorders and their management.

Adolescent↗
PubMed1998

Myoblast transplantation in non-dystrophic dog.

Dog myoblasts obtained from muscle biopsies were infected in vitro with a defective retroviral vector containing a cytoplasmic beta-galactosidase (beta-Gal) gene. These myoblasts were initially transplanted in the irradiated muscles of SCID mice and beta-Gal positive muscle fibers were observed. beta-Gal myoblasts were also transplanted back either in the donor dogs (autotransplantation model) or in unrelated recipient dogs (allotransplantation model). Following these myoblast injections, a rapid inflammatory reaction developed within the muscle as indicated by an expression of P-selectin and of pro-inflammatory cytokine mRNAs (interleukin 6 (IL-6) and transforming growth factor beta (TGF-beta), and by a neutrophil infiltration. Following either auto- or allotransplantation in inadequately or non-immunosuppressed dogs, a specific immune reaction also developed within 2 weeks as indicated by the infiltration of CD4+ and of CD8+ lymphocytes, the increased expression of IL-10 and granzyme B mRNAs and the presence of antibodies reacting with the injected cells. Some dogs were immunosuppressed with several combinations of FK506, cyclosporine (CsA) and RS-61443. In dogs immunosuppressed with CsA combined with RS-61443, only a few myoblasts and myotubes expressing beta-Gal were observed 1-2 weeks after the transplantation, but no muscle fibers expressing beta-Gal were observed after 4 weeks, and antibodies against the injected cells were formed. In dogs immunosuppressed with FK506 alone, although no antibodies against the injected cells were produced, there were no small cells and no muscle fibers expressing beta-Gal 1 month after the transplantation. However, FK506 triggered diarrhea and vomiting in dogs. When the dogs were immunosuppressed with FK506 combined with CsA and RS-61443, muscle fibers expressing beta-Gal were present 4 weeks after the transplantation and no antibodies reacting with donor myoblasts were detected. These results indicate that the combination of three immunosuppressive agents (i.e., FK506, CsA and RS-61443) is effective in controlling the specific immune reactions following myoblast transplantation in dogs and they underline that the outcome of myoblast transplantation is dependent in part on an adequate immunosuppression. These results obtained here in normal dogs may justify myoblast transplantation in dystrophic dogs despite the side effects of FK506.

Animals↗