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Biomedical subjects

M Srivastava

Publications and source records attributed to M Srivastava.

At least 55 records · Page 3Linked to original sources
PubMed1999

Effect of radiation on the xanthine oxidoreductase system in the liver of mice.

The xanthine oxidoreductase system is one of the major sources of free radicals in many pathophysiological conditions. Since ionizing radiations cause cell damage and death, the xanthine oxidoreductase system may contribute to the detrimental effects in irradiated systems. Therefore, modulation of the xanthine oxidoreductase system by radiation has been examined in the present study. Female Swiss albino mice (7-8 weeks old) were irradiated with gamma rays (1-9 Gy) at a dose rate of 0.023 Gy s(-1) and the specific activities of xanthine oxidase (XO) and xanthine dehydrogenase (XDH) were determined in the liver of the animals. The mode and magnitude of change in the specific activities of XO and XDH were found to depend on radiation dose. At doses above 3 Gy, the specific activity of XO increased rapidly and continued to increase with increasing dose. However, the specific activity of XDH was decreased. These findings are suggestive of an inverse relationship between the activity of XO and XDH. The ratio of the activity of XDH to that of XO decreased with radiation dose. However, the total activity (XDH + XO) remained constant at all doses. These results indicate that XDH may be converted into XO. An intermediate form, D/O, appears to be transient in the process of conversion. The enhanced specific activity of XO may cause oxidative stress that contributes to the radiation damage and its persistence in the postirradiation period. Radiation-induced peroxidative damage determined in terms of the formation of TBARS and the change in the specific activity of lactate dehydrogenase support this possibility.

Allopurinol↗
PubMed1999

Fertility outcome after repeat vasoepididymostomy.

PURPOSE: Historically, epididymal obstruction has been treated with surgical reconstruction. We determine whether it is worthwhile for patients to undergo repeat surgical reconstruction after failed vasoepididymostomy or whether they should be advised only to undergo sperm acquisition for assisted reproductive technique. MATERIALS AND METHODS: A total of 18 patients underwent repeat vasoepididymostomy performed by a single urologist (A. J. T.). Cases were divided based on the etiology of obstruction into groups 1--prior vasectomy (4), 2--congenital (7) and 3--inflammatory (7). Data were available regarding time of obstruction between initial and repeat vasoepididymostomy, quality of epididymal fluid, levels of anastomoses, semen analyses at least 12 months after surgery for all 18 men and pregnancy rates based on more than 18 months of followup in 12. RESULTS: Mean patient age at repeat vasoepididymostomy was 40.6 years (50.5, 36 and 39.4 years for groups 1, 2 and 3, respectively). Mean interval between vasectomy and initial vasoepididymostomy was 12.3 years (range 10 to 18). Mean interval between initial and repeat vasoepididymostomy was 19 months (range 12 to 41). Of the patients 10 underwent unilateral and 8 bilateral anastomoses, for a total of 26 repeat anastomoses. Overall patency rate was 66.7% (12 of 18) with sperm in the ejaculate in 75, 85 and 43% of patients in groups 1, 2 and 3, respectively. The patency rates according to the levels of the anastomosis were 66.7, 62.5 and 100% in the caput, corpus and cauda, respectively. Natural conception occurred in 3 of 12 couples (25%, 2 caput and 1 caudal anastomosis) during a mean followup of 23 months (range 13 to 34). All 3 cases had congenital obstruction. Pregnancy was achieved in 2 group 1 cases with cryopreserved sperm extracted at repeat vasoepididymostomy, and in 1 case each in groups 1 and 2 with microsurgical epididymal sperm aspiration and intracytoplasmic sperm injection. CONCLUSIONS: After repeat vasoepididymostomy two-thirds of men have sperm in the semen. Natural conception occurred in 25% of patients (3 of 12) followed for more than 18 months. Inability to establish pregnancy in the remaining 7 of 9 patients with sperm in the semen with a followup longer than 18 months may be due to epididymal dysfunction or partial obstruction and subsequent poor sperm quality. Aspiration of motile sperm and cryopreservation were possible in 11 of 18 cases at repeat vasoepididymostomy and should be recommended in case azoospermia remains or occurs after surgery. It appears worthwhile to offer patients repeat vasoepididymostomy after a failed initial procedure.

Adult↗
PubMed1999

Molecular dissection of nucleolin's role in growth and cell proliferation: new insights.

Cells require optimum protein synthetic activity in order to support cell proliferation, maintain homeostatic and metabolic integrity, and repair damage. Since growth depends on protein synthesis through ribosome biogenesis, the control of biosynthesis of ribosomes is necessarily a key element for control of growth. Nucleolin is a major nucleolar protein of exponentially growing eukaryotic cells, which is directly involved in the regulation of ribosome biogenesis and maturation. The highly conserved nucleolin contains three major domains through which it controls the organization of nucleolar chromatin, packaging of pre-RNA, rDNA transcription, and ribosome assembly. Numerous reports have implicated the involvement of nucleolin either directly or indirectly in the regulation of cell proliferation and growth, cytokinesis, replication, embryogenesis, and nucleogenesis. Nucleolin, an RNA binding protein, is also an autoantigen, a transcriptional repressor, and a switch region targeting factor. In addition, nucleolin exhibits autodegradation, DNA and RNA helicase activities, and DNA-dependent ATPase activity. An interesting aspect of nucleolin action is that it is a target for regulation by proteolysis, methylation, ADP-ribosylation, and phosphorylation by CKII, cdc2, PKC-xi, cyclic AMP-dependent protein kinase, and ecto-protein kinase. For these and other reasons, nucleolin is fundamental to the survival and proliferation of cells. Considerable progress has been made in recent years with the identification of new nucleolin binding proteins that may mediate these many nucleolin-dependent functions. Nucleolin also functions as a cell surface receptor, where it acts as a shuttling protein between cytoplasm and nucleus, and thus can even provide a mechanism for extracellular regulation of nuclear events. Exploration of the regulation of this multifaceted protein in a remarkable number of diverse functions is challenging.

Animals↗
PubMed1999

Pharmacogenomics of the cystic fibrosis transmembrane conductance regulator (CFTR) and the cystic fibrosis drug CPX using genome microarray analysis.

BACKGROUND: Cystic fibrosis (CF) is the most common lethal recessive disease affecting children in the U.S. and Europe. For this reason, a number of ongoing attempts are being made to treat the disease either by gene therapy or pharmacotherapy. Several phase 1 gene therapy trials have been completed, and a phase 2 clinical trial with the xanthine drug CPX is in progress. The protein coded by the principal CFTR mutation, DeltaF508-CFTR, fails to traffic efficiently from the endoplasmic reticulum to the plasma membrane, and is the pathogenic basis for the missing cAMP-activated plasma membrane chloride channel. CPX acts by binding to the mutant DeltaF508-CFTR and correcting the trafficking deficit. CPX also activates mutant CFTR channels. The comparative genomics of wild-type and mutant CFTR has not previously been studied. However, we have hypothesized that the gene expression patterns of human cells expressing mutant or wild-type CFTR might differ, and that a drug such as CPX might convert the mutant gene expression pattern into one more characteristic of wild-type CFTR. To the extent that this is true, a pharmacogenomic profile for such corrective drugs might be deduced that could simplify the process of drug discovery for CF. MATERIALS AND METHODS: To test this hypothesis we used cDNA microarrays to study global gene expression in human cells permanently transfected with either wild-type or mutant CFTR. We also tested the effects of CPX on global gene expression when incubated with cells expressing either mutant or wild-type CFTR. RESULTS: Wild-type and mutant DeltaF508-CFTR induce distinct and differential changes in cDNA microarrays, significantly affecting up to 5% of the total genes in the array. CPX also induces substantial mutation-dependent and -independent changes in gene expression. Some of these changes involve movement of gene expression in mutant cells in a direction resembling expression in wild-type cells. CONCLUSIONS: These data clearly demonstrate that cDNA array analysis of cystic fibrosis cells can yield useful pharmacogenomic information with significant relevance to both gene and pharmacological therapy. We suggest that this approach may provide a paradigm for genome-based surrogate endpoint testing of CF therapeutics prior to human administration.

Cell Line↗
PubMed1998

Diffusion of transferrin receptor clusters.

Two dimensional motion of membrane receptors provides a mechanism for interaction among receptors in the plane of the membrane. In some cases the lateral diffusion leads to formation of clusters which may also be mobile. We have used image cross-correlation (ICCS) spectroscopy technique to measure the translational motion of transferrin receptors in the membrane of 3T3 fibroblasts and HEp2 carcinoma cells. The technique is based on the measurement and analysis of fluctuations in the intensity observed in fluorescence confocal microscope images measured as a function of time. The fluorescence fluctuations arise from stochastic concentration fluctuations about the equilibrium concentration caused by movement of receptors. The amplitude of the fluctuations depend on the number of fluorescent molecules in the observation volume and the dynamics provide the rate of movement. The diffusion observed by this analysis is orders of magnitude slower than that measured by conventional photobleaching techniques. The slower motion corresponds to the diffusion of receptor clusters which provide the more dominant fluctuations.

3T3 Cells↗
PubMed1998

Prognosis of Japanese encephalitis: a multivariate analysis.

To evaluate the role of various clinical neurophysiological laboratory and radiological parameters in the prognosis of Japanese encephalitis (JE), 28 patients were prospectively evaluated. Their age ranged between two and 54 years, with eight of them being female and 11 children. The outcome was defined, at the end of a six-month period, as being poor (bed-ridden), partial (partial dependence for activity for daily life) and complete recovery (independent for activities of daily life). A number of clinical (age, degree of coma, decerebration or decortication and reflex changes), laboratory (CSF cell and protein content, and haemagglutination inhibition titre in the serum), neurophysiological (EEG, EMG, sensory and motor evoked potentials) and the extent of radiological changes were subjected to stepwise multivariate logistic regression analysis. By the end of six months, four patients had died, seven had poor, ten partial and seven had complete recovery. The best set of predictors of outcome included age, Glasgow coma scale and reflex changes.

Adolescent↗
PubMed1998

Failure of macrophage activation in destructive periodontal disease.

Macrophage populations in 22 biopsies of untreated advanced periodontitis were compared with those in 26 biopsies of clinically healthy (minimally inflamed) gingival tissue. The immunohistochemical investigation used high specificity monoclonal antibodies, including a pan-macrophage marker and probes for acute inflammatory, resident histiocytic, and reparative phenotypes. Macrophage expression of the functional activation markers MHC class II, basic fibroblast growth factor (bFGF), acid phosphatase (AP), and tartrate-resistant acid phosphatase (TRAP) was also examined. The study showed that advanced periodontitis and minimally inflamed tissues displayed similar distribution patterns and numbers for the macrophage phenotypic markers: there were, however, regionally-specific differences in the populations. In the advanced periodontitis lesion, there was little evidence of macrophage activation for the expression of HLA-DR, bFGF, and TRAP, although strong expression of HLA-DR and bFGF was observed in association with blood vessels. Macrophages expressing AP showed a distinct regional distribution; this, however, was not associated with foci of degenerate plasma cells. The apparent failure of recruitment and activation of macrophages may in part be both a cause and a consequence of the pathological features of this disease.

Adult↗
PubMed1998

Effect of high linear energy transfer radiation on biological membranes.

Cellular membranes are vital elements, and their integrity is extremely essential for the viability of the cells. We studied the effects of high linear energy transfer (LET) radiation on the membranes. Rabbit erythrocytes (1 x 10(7) cells/ml) and microsomes (0.6 mg protein/ml) prepared from liver of rats were irradiated with 7Li ions of energy 6.42 MeV/u and 16O ions of energy 4.25 MeV/u having maximum LET values of 354 keV/microm and 1130 keV/microm, respectively. 7Li- and 16O-induced microsomal lipid peroxidation was found to increase with fluence. The 16O ions were more effective than 7Li ions, which could be due to the denser energy distribution in the track and the yield of free radicals. These findings suggested that the biological membranes could be peroxidized on exposure to high-LET radiation. Inhibition of the lipid peroxidation was observed in the presence of a membrane-active drug, chlorpromazine (CPZ), which could be due to scavenging of free radicals (mainly HO* and ROO*), electron donation, and hydrogen transfer reactions. The 7Li and 16O ions also induced hemolysis in erythrocytes. The extent of hemolysis was found to be a function of time and fluence, and showed a characteristic sigmoidal pattern. The 16O ions were more effective in the lower fluence range than 7Li ions. These results were compared with lipid peroxidation and hemolysis induced by gamma-radiation.

Animals↗
PubMed1998

Drug-associated acute-onset vanishing bile duct and Stevens-Johnson syndromes in a child.

Acute vanishing bile duct syndrome is a rare but established cause of progressive cholestasis in adults, is most often drug or toxin related, and is of unknown pathogenesis. It has not been reported previously in children. Stevens-Johnson syndrome is a well-recognized immune complex-mediated hypersensitivity reaction that affects all age groups, is drug or infection induced, and has classic systemic, mucosal, and dermatologic manifestations. A previously healthy child who developed acute, severe, rapidly progressive vanishing bile duct syndrome shortly after Stevens-Johnson syndrome is described; this was temporally associated with ibuprofen use. Despite therapy with ursodeoxycholic acid, prednisone, and then tacrolimus, her cholestatic disease was unrelenting, with cirrhosis shown by biopsy 6 months after presentation. This case documents acute drug-related vanishing bile duct syndrome in the pediatric age group and suggests shared immune mechanisms in the pathogenesis of both Stevens-Johnson syndrome and vanishing bile duct syndrome.

Acute Disease↗
PubMed1998

Detection of viruses by electron microscopy: an efficient approach.

Wetting of grids is the most important aspect of the detection of viruses by electron microscopy. Carbon or plastic/carbon coated grids used in most electron microscopy laboratories are negatively charged and hydrophobic (Sogo, Portman, Kaufman and Koller, 1975. J. Microsc. 104, 187-198.). Most viruses are negatively charged at pH 6.0-7.0 which does not allow them to adhere effectively to the support film. Viruses are then washed away or form clumps on the grid making detection more difficult. A suitable wetting agent, 0.1% poly-L-lysine hydrobromide (MW 35000) has been used for the detection of plant viruses (Roberts, personal communication). This chemical was used for processing faecal samples from patients with gastroenteritis for electron microscopy. The result showed a significant increase in terms of virus concentration on the grids and an increase in the general detection rate of positive samples. A study was then undertaken comparing the treatment method of grids using faecal samples. The grids were treated and not treated with poly-L-lysine hydrobromide.

Adenovirus Infections, Human↗
PubMed1998

Analysis of membrane protein cluster densities and sizes in situ by image correlation spectroscopy.

Communication between cells invariably involves interactions of a signalling molecule with a receptor at the surface of the cell. Typically, the receptor is imbedded in the membrane and it is hypothesized that the binding of the signalling molecule causes a change in the state of aggregation of the receptor which, in turn, initiates a biochemical signal within the cell. Subsequently, many of the occupied receptors bind to membrane-associated structures, called coated pits, which invaginate and pinch off to form coated vesicles, thereby removing the receptors from the cell surface. The state of aggregation of membrane receptors is obviously in constant flux. Any useful approach to measuring the state of aggregation must, therefore, allow for dynamic measurements in living cells. It is possible to use fluorescently labelled signalling molecules or antibodies directed at the receptor of interest to visualize the receptor on the cell surface with a fluorescence microscope. By employing a laser confocal microscope, high resolution images can be produced in which the fluorescence intensity is quantitatively imaged as a function of position across the surface of the cell. Calculations of autocorrelation functions of these images provide direct and accurate measures of the density of fluorescent particles on the surface. Combined with the average intensity in the image, which reflects the total average number of molecules, it is possible to estimate the degree of aggregation of the receptor molecules. We refer to this analysis as image correlation spectroscopy (ICS). We show how ICS can be used to measure the density of several receptors on a variety of cells and how it can be used to measure the density of coated pits and the number of molecules per coated pit. We also show how the technique can be used to monitor fusion of virus particles to cell membranes. Further, we illustrate that, by calculating cross-correlation functions between pairs of images, we can extend the analysis to measurements of the distributions as a function of time, on the second timescale, as well as to measurements of the movement of the receptor aggregates on the surface. Finally, we illustrate that, by this approach, we can measure the extent of interaction between two different receptors as a function of time. This represents the most quantitative measurement of the extent of co-localization of receptors available and is independent of the spatial resolution of the confocal microscope. The theory of ICS and image cross-correlation spectroscopy (ICCS), focussing on the interpretation of the data in terms of the biological phenomenon being probed, is discussed.

Animals↗
PubMed1998

Cyanine fluorochrome-labeled antibodies in vivo: assessment of tumor imaging using Cy3, Cy5, Cy5.5, and Cy7.

Monoclonal antibodies to two different targetable antigens were conjugated to each of four commercially available cyanine fluorochromes. Equal amounts of all four antibodies were coinjected into tumor-bearing animals and imaged. Small, superficial tumors were adequately labeled using all four fluorochromes. Large tumors were labeled well only by Cy7, probably due to self-masking and dilution effects. Cy7 was superior to other cyanine fluorochromes for visualizing structures located deep within the animal.

Animals↗
PubMed1998

Effects of ultraviolet-B radiation on phycobilisomes of Synechococcus PCC 7942: alterations in conformation and energy transfer characteristics.

Phycobilisomes (PBS), the major light harvesting antenna of the cyanobacterium Synechococcus contain phycocyanin (PC) and allophycocyanin (APC) as major pigment-protein complexes. PBS also absorb ultraviolet-B (280-320 nm) radiation. Exposure of Synechococcus PBS to low dose of UV-B (approximately 0.28 mw.cm-2) for 90 min induced change in absorption, emission and excitation characteristics of PBS and these changes got enhanced after 3 h of exposure. Room temperature excitation and emission spectra clearly indicated uncoupling of energy transfer from PC to APC on exposure to UV-B. Also, the 77K emission spectra suggested that F682 emission originating from APC decreased by 42% after 3 h of exposure. Circular dichroism (CD) spectra of UV-B exposed PBS indicated changes (14% decrease) in the alpha-helical content after 90 min treatment. SDS-PAGE analysis indicated degradation of a 75 kDa polypeptide (which appear to be a linker polypeptide) on UV-B treatment. The degradation of this polypeptide seems to induce changes in pigment-protein interaction and decoupling of energy transfer within the PBS. Our results for the first time clearly indicate that the PBS of Synechococcus are targets for UV-B damage.

Bacterial Proteins↗
PubMed1998

Mouse cytochrome b561: cDNA cloning and expression in rat brain, mouse embryos, and human glioma cell lines.

A cDNA encoding cytochrome b561 has been isolated from a mouse brain cDNA library, which predicts a protein of 250 amino acids with a deduced Mr of 27,770. Northern blot analysis of different mouse and rat tissues revealed one major mRNA of 3300 bp, which is abundantly distributed in a number of neuroendocrine tissues. In addition, cytochrome mRNA levels in rat brain sections showed the highest distribution of cytochrome b561 in the hypothalamus, hippocampus, thalamus, and striatum, with a moderate level in the cerebral cortex, and the lowest levels in the olfactory bulb and cerebellum. Because non-neuronal cells in the central nervous system contained peptidyl alpha-amidating monooxygenase (PAM), to which cytochrome b561 donates its electrons, we used RT-PCR to document the coexpression of cytochrome b561 with PAM and dopamine beta-hydroxylase (DBH) in glioblastoma cells. Cytochrome b561 expression was detectable in the 11-day-old mouse embryo, and the level of its mRNA increased tenfold by 15 and 17 days of gestation.

Amino Acid Sequence↗
PubMed1998

Nutritional status of rural non-pregnant non-lactating women in reproductive age.

OBJECTIVE: To find out the degree of current undernutrition in rural reproductive age women. SETTING: 49 villages of two adjoining rural blocks of Varanasi. METHODS: 6130 non-pregnant and non-lactating rural women in the age group 18-45 year were studied for sociodemographic characteristics and anthropometry, i.e., weight, height and midarm circumferance. Their percentiles for age and for weight for height were calculated by using cubic spline method. RESULTS: The women in 10th centile weighed < 38 kg and those in > 90th centile weighed 47-48 kg; 74.2% had weight < 45 kg. The 50th centile height ranged between 148-150 cm; 13.5% were < 145 cm. For mid arm circumference 50th and 90th centile values were around 22 and 24 cm, respectively. Mid arm circumference and height had significant linear correlation with weight. CONCLUSION: Around 50% rural UP women in pre-pregnancy state are undernourished. With age these rural women did not change in weight or mid-arm circumference.

Adolescent↗
PubMed1997

Cell cycle and post-transcriptional regulation of annexin expression in IMR-90 human fibroblasts.

Based on the finding that the expression of some annexins varies dramatically as a function of cellular proliferation state [Schlaepfer and Haigler (1990) J. Cell Biol. 111, 229-238], it has been proposed that the cellular level of the annexins might be critical for the regulation of cell growth. To further test this hypothesis, we have studied the expression of various annexins in normal human IMR-90 fibroblasts synchronized by serum deprivation. Using immunoblotting, the cellular content of annexins (Anxs) II, V and VI was found to vary by less than 10% during the cell cycle. However, Anx IV expression increased by 50% during S-phase and the levels of Anxs I and VII were reduced by 40% in early G2/M. However, using RNase protection assays, the mRNAs of Anxs I and VII were found to be uniformly expressed throughout the cell cycle, suggesting that down-regulation of both proteins in G2/M occurred through a post-transcriptional process. In addition, cells transfected with Anx VII cDNA were shown to contain an amount of Anx VII similar to wild-type cells, despite the elevation of Anx VII mRNA content in transfected cells by approx. 2 orders of magnitude. Vector misconstruction or possible secretion of the overexpressed protein were ruled out using appropriate controls. Therefore, as with cell-cycle regulation, Anx VII expression in transfected cells is also controlled by post-transcriptional mechanisms. Furthermore, using pulse-chase analysis, we have determined that annexin VII, and other Anxs, have a slow turnover rate, consistent with the limited changes of expression throughout the cell cycle. Taken together, these results question the hypothesis that cellular expression of Anxs plays a general role in cell growth and support the concept that post-transcriptional mechanisms may control levels of Anxs I and VII.

3T3 Cells↗
PubMed1997

Mucin histochemistry--a simple and effective method for diagnosing premalignant and early malignant lesions of lower gastrointestinal tract.

Mucin histochemistry of colo-rectal mucosa was studied in 124 biopsies which included 28 specimens from normal mucosa, 35 inflammatory lesions, 20 polyps, 26 malignant lesions and 15 specimens from transitional mucosa. The stains used were Haematoxylin and Eosin, Periodic acid schiffs, Alcian blue--Periodic acid schiffs (pH 2.5), Orcein--Alcian blue and Periodic acid sodium borohydride saponification. Normal mucosa, mucosa from inflammatory lesions, juvenile and hyperplastic polyps had neutral mucin in traces along with O-acylated sialomucins and sulphomucins. Predominant changes in adenomatous polyps and malignancy were a significant loss of sulphomucins and O-acylation at position C7, C8 or C9 in sialomucins. The changes in transitional mucosa were more on the pattern of malignant lesions. The alterations in the amount and/or pattern of mucin in colorectal mucosa has been found to predate frank morphological changes of malignancy. A routine use of mucin histochemistry in suspicious colorectal biopsies might be helpful in diagnosing premalignant and early malignant lesions in otherwise apparently normal colorectal mucosa or mucosa with chronic inflammatory changes and thereby reduce mortality.

Histocytochemistry↗