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S Vasudevan

Publications and source records attributed to S Vasudevan.

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

Detection of viral ribonucleic acid and histologic analysis of inflamed synovium in Ross River virus infection.

OBJECTIVE: To document the histology of Ross River virus (RRV) arthritis and to examine inflamed synovium for viral RNA. METHODS: Biopsy tissue from the inflamed knees of 12 patients with RRV infection was studied using conventional and immunostaining techniques. Reverse transcriptase-polymerase chain reaction technology was used to probe for the presence of viral RNA in the synovial biopsy samples and in serum. RESULTS: Hyperplasia of the synovial lining layer, vascular proliferation, and mononuclear cell infiltration were the main histologic changes. RRV RNA was found in knee biopsy tissue that was obtained from 2 patients at 5 weeks after the onset of symptoms. CONCLUSION: RRV RNA was identified in inflamed synovium more than a month after symptoms began. Inflammation was apparent in the absence of detectable virus in the majority of patients.

Adult↗
PubMed2000

The yeast hnRNP-like protein Hrp1/Nab4 marks a transcript for nonsense-mediated mRNA decay.

The nonsense-mediated mRNA decay (NMD) pathway monitors premature translation termination and degrades aberrant mRNAs. In yeast, it has been proposed that a surveillance complex searches 3' of a nonsense codon for a downstream sequence element (DSE) associated with RNA-binding proteins. An interaction between the complex and the DSE-binding protein(s) triggers NMD. Here we describe the identification and characterization of the Hrp1/Nab4 protein as a DSE-binding factor that activates NMD. Mutations in HRP1 stabilize nonsense-containing transcripts without affecting the decay of wild-type mRNAs. Hrp1p binds specifically to a DSE-containing RNA and interacts with Upf1p, a component of the surveillance complex. A mutation in HRP1 that stabilizes nonsense-containing mRNAs abolishes its affinity for the DSE and fails to interact with Upf1p. We present a model describing how Hrp1p marks a transcript for rapid decay.

Biological Transport↗
PubMed1999

Plasma and urinary selenium in Saudi Arabian patients with dilated cardiomyopathy.

We measured selenium (Se) levels in the urine and blood plasma samples of 72 Saudi Arabian patients with dilated cardiomyopathy (DCM) and 70 control subjects of the same origin. To correct for differences in the hydration state of the subjects, the selenium concentration for each urine sample was normalized by dividing it by the concentration of creatinine (CREAT) in the same sample. The median (and range) of the values found for the concentration of Se in plasma, urine, and normalized concentration in urine for the control subjects was 1.306 (0.66-2.50) microM, 0.478 (0.05-2.00) microM, and 56.7 (10.6-426.5) microM Se/M CREAT, respectively, whereas, for the patients, it was 1.246 (0.53-2.45) microM, 0.39 (0.05-1.90) microM, and 75.1 (4.9-656.2) microM Se/M CREAT, respectively. Additionally, the patients were separated into three subgroups according to the severity of their disease state as judged by NYHA procedure, and were then compared to the control group. Only group 4 (the most severe state of the disease) had a significantly lower concentration of urinary Se than the control group. However, the difference became nonsignificant when normalized for CREAT levels. There was no significant difference in the plasma Se levels between the controls and any of the patient groups. As the plasma Se in the control group and in the DCM patients both fell on the low end of the "normal" range, with the patients being marginally lower than the controls, there is no firm evidence from this study to suggest that Se is related to the high incidence rate of DCM found in Saudi Arabia.

Adolescent↗
PubMed1998

Mode of binding of anti-P-glycoprotein antibody MRK-16 to its antigen. A crystallographic and molecular modeling study.

Monoclonal antibody MRK-16 recognizes a discontinuous extracellular epitope on the multidrug resistance-associated ATP-binding cassette transporter, P-glycoprotein. The atomic basis for specificity of this antibody is of interest because of its potential as a modulator of P-glycoprotein activity. The crystal structure of Fab MRK-16 is reported to a resolution of 2.8 A. A structure for a portion of the epitope was derived by comparison to regions of solved structures with similar primary sequence. This has permitted a proposal for the mode of binding of the peptide epitope to the antibody, in which the peptide makes specific contacts with complementarity-determining regions H1, H2, and H3 from the heavy chain and L3 from the light chain. These interactions are consistent with epitope mapping studies and with the observation that MRK-16 is specific for human class I P-glycoprotein. This result identifies side chains in MRK-16 that would be amenable to alteration in antibody engineering experiments to derive improved multidrug resistance inhibitors for clinical use during chemotherapy. In particular, Arg-H97 contacts both Glu-746 and Asp-744 of the peptide, Arg-L96 contacts Asp-743, and Thr-H33 interacts with Thr-747. All of these epitope residues were implicated in mediating specificity by epitope mapping studies.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Cycloheximide sensitivity of orotic acid biosynthesis induced by ammonia and glycine administration.

Administration of either ammonia or glycine to both rats and mice results in an increased synthesis in the liver and urinary excretion of orotic acid. The two most relevant observations obtained are that carbamoyl phosphate synthesized inside the mitochondria is involved in the increased synthesis of orotic acid and that this latter process is almost completely abolished by cycloheximide and actinomycin D, inhibitors of protein and RNA synthesis. Orotic acid synthesis could be controlled by an induction-suppression mechanism. Inhibition of synthesis of excess orotic acid brought about by N-(phosphonacetyl)-L-aspartic acid but not by acivicin, suggests that glutamine-dependent cytosolic synthesis of carbamoyl phosphate, is not involved. Administration of ornithine together with glycine completely suppressed the synthesis of orotic acid, but promoted a twofold increase of urea excretion. The concentration of ornithine rather than that of carbamoyl phosphate or the activity of the enzymes involved, may represent a limiting factor controlling both the flux of ammonia in the urea cycle and the availability of mitochondrial carbamoyl phosphate for orotic acid synthesis. Two enzymes have been found to be induced by glycine: ornithine decarboxylase and aspartate transcarbamoylase (aspartate carbamoyltransferase). Both enzymes may contribute to the increase in orotic acid synthesis, aspartate transcarbamoylase more directly and ornithine decarboxylase by lowering the ornithine concentration. Ornithine decarboxylase activity was completely suppressed but that of aspartate transcarbamoylase was further increased by cycloheximide treatment. Inhibition of orotic acid biosynthesis by cycloheximide appears to be the result of a decreased availability in the cytosol of carbamoyl phosphate synthesized inside the mitochondria.

Ammonium Chloride↗
PubMed1995

Characterization of the rat m3 muscarinic acetylcholine receptor produced in insect cells infected with recombinant baculovirus.

The m3 muscarinic acetylcholine receptor from rat heterologously produced in insect cells after infection with a recombinant baculovirus has an apparent molecular mass of approximately 75 kDa. Polyclonal antibodies raised against a carboxy-terminal nonapeptide that is unique to the m3 subtype can detect the receptors produced in the insect cells by Western blot and can also immunoprecipitate solubilized receptor. Immunofluorescence microscopy as well as electron microscopy revealed that the receptor was located intracellularly, visualized as a ring around the nucleus of the infected insect cells. Solubilization of the receptor was accomplished with digitonin which was added in increments (over 10 min) to a final concentration of 0.8% (mass/vol). The solubilized receptor is unstable when the ligand-binding site is not protected by a ligand. Here the low-affinity ligand propylbenzilylcholine (approximately 10 nM) has demonstrable protective ability during solubilization, but the usefulness of this ligand is limited by a very slow off rate. From the behaviour of the solubilized receptor during DEAE-Sephacel chromatography and lectin-affinity chromatography it can be deduced that the receptor produced in insect cells is heterogeneously glycosylated in the producing insect cells.

Affinity Labels↗
PubMed1995

Nucleotide pool imbalances in the livers of patients with urea cycle disorders associated with increased levels of orotic aciduria.

Liver samples obtained at autopsy from patients with ornithine transcarbamylase (OTC) deficiency, a urea cycle disorder that is associated with high levels of orotic acid biosynthesis and excretion were analysed for nucleotide pools. As a control, liver samples from patients with a deficiency of mitochondrial carbamyl phosphate synthetase (CPS-I) which is not associated with increased levels of orotic acidurias were also analysed. The results show that liver tissue from OTC deficiency patients exhibited an increased ratio of uridine nucleotides to adenosine nucleotides, while in CPS-I deficiency patients, no such increase was noted. This study indicates that genetic disorders that are associated with increased loads of orotic acid exhibit abnormally high ratios of uridine to adenosine nucleotides in the liver. This type of imbalance is analogous to that seen in the liver of rats and mice exposed to an orotic acid supplemented or an arginine-deficient diet under liver tumor promoting conditions. It is likely that an imbalance in nucleotide pools may have a significant role in the pathophysiology associated with these disorders.

Adenine Nucleotides↗
PubMed1994

Preliminary crystallographic analysis of a Fab specific for P-glycoprotein with and without bound peptide.

The antigen-binding fragment (Fab) of anti-peptide monoclonal antibody C219 raised against the multidrug resistance associated P-glycoprotein has been crystallized with and without bound peptide. The crystals of the Fab in the absence and presence of peptide belong to space groups P2(1) and P2(1)2(1)2(1), respectively. The volumes of both crystal forms are consistent with the presence of four Fab molecules per asymmetric unit. Diffraction data to 3.2 A resolution have been collected on a San Diego Multiwire Area Detector system from both crystal forms. Determination of the molecular replacement solutions is underway.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1994

An earlier proliferative response of hepatocytes in gamma-glutamyl transferase positive foci to partial hepatectomy.

One of the hallmarks of initiated hepatocytes is their resistance to several hepatotoxins. This property forms the basis for their selective growth under conditions which are inhibitory to the non-initiated hepatocytes. Selective growth of initiated hepatocytes also occurs, albeit at a low level, in initiated rat liver without exposure to any known promoting regimen and/or in the absence of any known selective pressure to which initiated hepatocytes can possibly be resistant. This latter phenotypic property of initiated hepatocytes was further characterized by comparing the kinetics of response of hepatocytes in gamma-glutamyl transferase positive foci and in the surrounding liver to 2/3 partial hepatectomy both in the presence and in the absence of a promoting regimen. Male Fischer 344 rats (130-150 g) were initiated with a single dose of diethylnitrosamine and 1 week later they were placed on either a semi-synthetic basal diet or a promoting diet containing 1% orotic acid. Partial hepatectomy was performed 15 weeks after initiation and animals from both groups were killed at 12, 16, 20, 24, 30, 36, 48, 72 or 96 h after operation. Each animal received a pulse of 3H-labelled thymidine 1 h prior to killing. Autoradiographic studies revealed that hepatocytes in gamma-glutamyl transferase positive foci in the livers of rats fed the basal diet were significantly labelled at 16 h post-partial hepatectomy while surrounding hepatocytes were still virtually quiescent (LI 12.7 +/- 4.7 versus 1.2 +/- 0.5%, respectively). Higher labelling index in foci compared to the surrounding liver was also seen at 20 h post-PH (36.9 +/- 2.6 versus 21.5 +/- 2.4). Similar earlier response of hepatocytes in gamma-glutamyl transferase positive foci was also seen in initiated rats exposed to dietary orotic acid. In addition, orotic acid treatment appears to have imposed a slight delay on the entry of hepatocytes in the surrounding liver into 'S' phase and thereby enhancing the differential of growth response between these two populations.

Animals↗
PubMed1994

Perturbations of endogenous levels of orotic acid and carcinogenesis: effect of an arginine-deficient diet and carbamyl aspartate on hepatocarcinogenesis in the rat and the mouse.

Feeding excess orotic acid (OA) in the diet promotes the carcinogenic process in different organs including the liver. A number of metabolic and genetic disorders are associated with increased synthesis of endogenous OA and some of these disorders appear to pose an increased risk of liver cancer development. This study therefore examines whether excess OA of endogenous origin also exerts a promoting effect on hepatocarcinogenesis in the mouse and the rat. Increased endogenous synthesis of OA was achieved by (i) feeding a diet deficient in arginine (AD) and (ii) feeding excess dietary carbamylaspartate (CA), a precursor for the synthesis of OA. A single dose of diethylnitrosamine (DENA) was given i.p. to male Fischer 344 rats (200 mg/kg) or to male DBA/2 mice (90 mg/kg). One week later they were placed on either AD diet or the same diet supplemented with 1.35% arginine (AS) for a total of 4 weeks. Two-thirds partial hepatectomy (PH) was performed at the end of the second week. All animals were then transferred to a control semisynthetic basal diet for a total of 20 weeks before they were killed. The results indicated that AD diet increased the incidence of hepatic nodules in both rats (percentage area occupied by nodules was 4.7 +/- 0.4 in the AD group compared to a control value of 0.7 +/- 0.5) and mice (4/10 mice had nodules > 5 mm diameter in the AD group while none in the AS group had such large nodules). In another experiment male Fischer 344 rats similarly initiated with DENA were exposed to either basal diet or basal diet containing 2% CA for 4 weeks coupled with PH performed at the end of the second week. This regimen was followed by 20 weeks of feeding basal diet to both groups. Rats given CA developed larger hepatic foci and nodules (0.84 +/- 0.56 mm3) compared to the control group, which was fed basal diet throughout the experiment (0.07 +/- 0.03 mm3). Further, both AD diet and dietary CA, like dietary OA, induced an increase in hepatic uridine nucleotides. Taken together, these results suggest that increased levels of endogenously synthesized OA, like exogenously supplied excess OA, can induce an imbalance in hepatic nucleotide pools and can exert a promoting effect on hepatocarcinogenesis.

Animals↗
PubMed1993

Human beta 2-adrenergic receptor produced in stably transformed insect cells is functionally coupled via endogenous GTP-binding protein to adenylyl cyclase.

Spodoptera frugiperda insect cells (Sf9) containing the stably integrated human beta 2-adrenergic receptor gene under the control of the baculovirus IE1 promoter expressed up to 350,000 human receptors/cell. The number of receptors did not change with cell density or age of culture. The adrenergic receptors overexpressed in the insect cells were functional with respect to their ligand binding and signalling properties. Coupling of the receptors to endogenous GTP-binding proteins is demonstrated by hormone-dependent stimulation of GTPase and adenylyl cyclase activity in the transformed insect cells. Western-blot analysis revealed that the endogenous GTP-binding protein appears to be of the heterotrimeric type. Antibodies raised against the mammalian alpha subunit of stimulatory GTP-binding proteins cross-react with the insect alpha subunit of GTP-binding proteins, which also exhibits the same apparent molecular mass as its mammalian counterpart. The beta subunit of GTP-binding proteins from insect cells reacts with anti-peptide serum directed against the C-terminal amino acids of the mammalian beta subunit of GTP-binding proteins, but is about approximately 2 kDa larger than that of the beta subunit of GTP-binding proteins from bovine brain. Exposure of the transformed insect cells to L-isoproterenol rapidly induces uncoupling and internalization of 30% of the heterologously expressed receptors. In contrast to the situation in mammalian cells, prolonged exposure of the agonist (24 h) does not result in down regulation of the remaining 70% of the receptors.

Adenylyl Cyclases↗
PubMed1993

The effects of various inhibitors on the regulation of orotic acid excretion in sparse-fur mutant mice (spf/Y) deficient in ornithine transcarbamylase.

Experiments were conducted to determine whether the excessive orotic aciduria, induced in sparse-fur male mice (spf/Y) deficient in ornithine transcarbamylase (OTC), may be regulated by some inhibitors, such as acivicin (0.014 mmol/100 g body weight, i.p.), N-(phosphonoacetyl)-L-aspartate (PALA, 2.5 mg/100 g body weight, i.p.), adenine (3 g/kg diet) and cycloheximide (0.35 mmol/kg body weight, i.p.). We also administered ornithine (1 mmol/100 g body weight, i.p.), a substrate of the urea cycle, to alleviate the metabolic deficiency of arginine in spf/Y mice which may also be responsible for excessive orotic aciduria. The orotic aciduria remained insensitive to acivicin, indicating mitochondria as the source of carbamyl phosphate. However, orotate excretion was significantly decreased by PALA (P < 0.01), due to its effect on the aspartate transcarbamylase activity. The ingestion of adenine resulted in an increase (P < 0.05) of urinary orotate, suggesting the blockage of the utilization of orotate for nucleotide biosynthesis. Ornithine administration led to a reduction (P < 0.01) of the excretion of orotate induced by the OTC deficiency in these mice, indicating that one of the regulatory steps in its synthesis may be the availability of ornithine. There were no changes in urinary orotate excretion in spf/Y mice when treated with cycloheximide. On the other hand, pretreatment with cycloheximide in an artificial model of OTC deficiency (Swiss-ICR normal mice on an arginine-deficient diet treated thereafter with norvaline, an inhibitor of OTC), caused a significant decrease in urinary orotate. These results suggest that spf/Y mice are unique in that the increased synthesis of orotate is not sensitive to cycloheximide. Perhaps this may reflect an adaptive phenomenon developed by the mutant mice to handle excess mitochondrial carbamyl phosphate and orotic acid.

Adenine↗
PubMed1993

The development of hepatocellular carcinoma in initiated rat liver after a brief exposure to orotic acid coupled with partial hepatectomy.

Previous work from this laboratory has revealed that a minimum of 10-20 weeks of continuous exposure to 1% dietary orotic acid (OA) is necessary for this regimen to exert a significant promoting effect on the carcinogenic process in rat liver. The present study investigates the effect of partial hepatectomy (PH), given during a short-term exposure (4 weeks) to OA, on the development of hepatocyte nodules (HN) and hepatocellular carcinoma (HCC) initiated by diethylnitrosamine (DEN). Male Fischer 344 rats (130-150 g) were given a single dose of DEN (200 mg/kg body wt i.p.). Starting a week later they were fed either a semisynthetic basal diet (BD) or the same diet containing 1% OA for 2 weeks; two-thirds PH was then performed followed by another 2 weeks of BD or OA diet respectively. At the end of this treatment some animals from both groups were killed while the rest were continued on BD and killed at 20 or 56 weeks thereafter. The results showed no difference between the two groups in the incidence of gamma-glutamyltransferase-positive foci when rats were killed at 2 weeks after PH. However, 4 week exposure to OA coupled with PH significantly enhanced the incidence of HN and HCC when this protocol was followed by 20 or 56 weeks of BD feeding respectively, leading to 63% incidence of HCC in the OA-fed group, while no HCC was observed in control animals. It is concluded that a type of stable or permanent change(s) ('imprinting' or 'memory effect') is induced in the initiated rat liver by this treatment, which imposes a promoting environment in the liver even after withdrawal of the promoter.

Animals↗
PubMed1993

Increasing the interval between initiation and the onset of exposure to orotic acid decreases its promoting effect on rat liver carcinogenesis.

The present study was designed to determine whether a delay in the start of the promoting regimen after the administration of a carcinogen would influence the promoting efficacy of orotic acid on the development of hepatocellular carcinoma in rats. Male Fischer 344 rats weighing 130-150 g were injected with a single dose of diethylnitrosamine (200 mg/kg body wt i.p.) then divided into 3 groups: groups 1 and 2 were given semi-synthetic basal diet or the same diet containing 1% orotic acid (OA) respectively starting 1 week after the carcinogen; group 3 received the OA diet starting 5 weeks after the administration of diethylnitrosamine. Animals from these 3 groups were sacrificed after 25, 32, 42 and 60 weeks of being fed their respective diets. The results indicated that delaying the start of the OA diet after the carcinogen resulted in about a 50% decrease in the incidence of hepatic nodules and/or hepatocellular carcinomas at various time points during the experiment. This decrease in promoting efficacy of OA was not apparently explainable by lack of metabolic effects of OA, at least in terms of induction of nucleotide pool imbalance, a condition that appears to be important for OA to exert its tumor promoting effects.

Animals↗