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Biomedical subjects

M Ramanathan

Publications and source records attributed to M Ramanathan.

At least 19 recordsLinked to original sources
PubMed2003

Differential anxiolytic effect of enalapril and losartan in normotensive and renal hypertensive rats.

The effect of angiotensin-converting enzyme (ACE) inhibitor enalapril (EPL) (2 and 4 mg/kg), angiotensin (AT) II receptor antagonist losartan (LRN) (5 and 10 mg/kg), and anxiolytic drug diazepam (DZP) (0.5 mg/kg) on anxiety parameters were evaluated in experimentally induced renal hypertensive rats (RHR). Renal hypertension was induced in Wistar strain male albino rats weighing 200-250 g by following the method of Goldblatt. The animals having systolic blood pressure more than 180-210 mm Hg were subjected to open-field exploratory behaviour, elevated plus maze behaviour, and social interaction tests of anxiety. The RHR showed hyperactivity in open-field behaviour and anxiogenicity in elevated plus maze and social interaction tests. Losartan (5 and 10 mg/kg) and DZP (0.5 mg/kg) significantly attenuated the hyperactivity and anxiogenic behaviour in experimentally induced hypertensive rats and induced anxiolysis in normotensive rats (NTR). Enalapril reversed the hypertension-induced alteration only at higher dose (4 mg/kg) and failed to show any effect in NTR. It can be concluded that renin angiotensin aldosterone system (RAAS) has a significant role on behaviour, and LRN has shown better effect in reversing the hyperactivity and anxiogenicity in the experimentally induced hypertensive rats, indicating a possible role of AT receptor in the mediation of anxiolysis.

Angiotensin II Type 1 Receptor Blockers↗
PubMed2001

In vivo gene expression revealed by cDNA arrays: the pattern in relapsing-remitting multiple sclerosis patients compared with normal subjects.

OBJECTIVES: To use DNA arrays to identify differences in gene expression associated with relapsing-remitting (RR) MS. METHODS: Total RNA was isolated from monocyte depleted peripheral blood mononuclear cells of 15 RR MS patients and 15 age- and sex-matched controls. The RNA was reverse transcribed to radiolabeled cDNA and the resultant cDNA was used to probe a DNA array containing over 4000 named human genes. The binding of radiolabeled cDNA to the probes on the array was measured by phosphorimager. RESULTS: Of more than 4000 genes tested, only 34 were significantly different in RR-MS patients from controls. Of these, 25 were significantly increased and 9 significantly decreased in the RR MS patients. Twelve of these genes have inflammatory and/or immunological functions that could be relevant to the MS disease process. The potentially relevant genes that were elevated (15% to 28%) were P protein, LCK, cAMP responsive element modulator, IL-7 receptor, matrix metalloproteinase-19, M130 antigen, and peptidyl-prolyl isomerase. Those that were significantly decreased (15% to 35%) were SAS transmembrane 4 superfamily protein, STRL22 (C-C chemokine receptor 6), AFX protein, DNA fragmentation factor-45 and immunoglobulin gamma 3 (Gm marker). CONCLUSIONS: The RR-MS disease effect was relatively restricted and most of the mRNAs tested were not different from the normal controls. However, there were significant differences identified in the expression of a subset of mRNAs, including 13 with inflammatory/immune functions that could be relevant to MS. The systematic use of DNA arrays can provide insight into the dynamic cellular pathways involved in MS pathogenesis and its phenotypic heterogeneity.

Adult↗
PubMed2000

Cross-correlation for flow cytometric histogram background subtractions.

Background subtraction is a widely encountered problem in flow cytometry applications for which the currently available analysis techniques are unsatisfactory. The 99% division line method, also referred to as the threshold or marker method, is widely used because it is computationally simple but it has poor accuracy and tends to underestimate the percentage of positive cells when there is overlap between histograms. Model-based approaches are preferred when there are overlapping peaks, but these methods require curve fitting and strong assumptions regarding the shape of the underlying distributions. This report assesses a mathematically rigorous, computationally facile, non-parametric technique called cross correlation for the background subtraction problem. A metric, positivity, derived from cross correlation is shown to overcome the disadvantages of both the 99% division line and model-based methods without compromise.

Flow Cytometry↗
PubMed2000

Introduction of macromolecules into synaptosomes using electroporation.

Synaptic terminals are sites of high metabolic activity and thus are particularly vulnerable to oxidative stress. Oxidative damage to proteins can be toxic to neurons and may cause irreversible cell damage and neurodegeneration. A neuroprotective mechanism used by cells to combat oxidative damage is to selectively degrade damaged proteins. Therefore, it is of interest to study the mechanism of degradation of oxidatively damaged proteins in synaptosomes. One way of oxidizing synaptosomal proteins in vitro is by incubating intact synaptosomes in the presence of an oxidizing agent. A problem with this approach is that it may also cause oxidative damage to the machinery required to recognize and degrade oxidized proteins. We have, therefore, introduced a fluorescent macromolecule into synaptosomes to assess the feasibility of using this technique to study how oxidized proteins are degraded and removed from synaptic terminals. Synaptosomes were subjected to electroporation in the presence of FITC labelled-dextran with an average molecular weight of 70000 (FD-70) and non-specific binding was determined by running parallel experiments in lysed synaptosomes. Following extensive washing, synaptosomes were assayed for the presence of intra-synaptosomal FD-70 by measuring fluorescence in a microplate fluorescence reader. Significant differences in fluorescence were found between intact and lysed synaptosomes with maximal uptake at 100 V/ 1500 microF (approx. 36 pmol/mg protein). To determine if membrane transport was compromised by electroporation, uptake of 3H-arginine was compared in control and electroporated synaptosomes. While untreated electroporated synaptosomes showed a loss of 22% in the ability to transport arginine, preincubation in the presence of 1 mM ATP resulted in a complete restoration of arginine transport. These results show that electroporation is a potentially useful technique for introducing a specific oxidized protein, into synaptic terminals so its metabolic fate can be examined.

Adenosine Triphosphate↗
PubMed2000

Effect of Withania somnifera glycowithanolides on iron-induced hepatotoxicity in rats.

Glycowithanolides, consisting of equimolar concentrations of sitoindosides VII-X and withaferin A, isolated from the roots of Withania somnifera Dunal, have been reported to have an antioxidant effect in the rat brain frontal cortex and striatum. In the present study, the effect of 10 days of oral administration of these active principles, in graded doses (10, 20 and 50 mg/kg), was noted on iron overload (FeSo(4), 30 mg/kg, i.p.) induced hepatotoxicity in rats. Apart from hepatic lipid peroxidation (LPO), the serum enzymes, alanine aminotransferase, aspartate aminotransferase and lactate dehydrogenase, were assessed as indices of hepatotoxicity. Silymarin (20 mg/kg, p.o.) was used for comparison. Iron overload induced marked increase in hepatic LPO and serum levels of the enzymes, which was attenuated by WSG in a dose-related manner, and by silymarin. The results indicate that the reported use of WS in Ayurveda for hepatoprotection against heavy metals and other environmental toxins, may be due the antioxidant action of WSG.

Administration, Oral↗
PubMed2000

Glycosaminoglycans alter the conformation of interferon-gamma.

Interferon-gamma (IFN-gamma) is a potent immunomodulatory cytokine whose physiological roles are modulated by the extracellular matrix. Here, circular dichroism and fluorescence spectroscopic techniques are used to demonstrate that low molecular weight heparin and chondroitin sulfate cause significant changes in secondary and tertiary structure of IFN-gamma. The results suggest that heparin and chondroitin sulfate modulate IFN-gamma activity by causing structural changes in the IFN-gamma dimer.

Animals↗
PubMed2000

Correlates of female sterilization regret in the southern states of India.

This study analyses factors associated with the incidence of sterilization regret in the four south Indian states of Andhra Pradesh, Karnataka, Kerala and Tamil Nadu. Using data from the National Family Health Surveys, in all four states the incidence of regret was found to be less than 10% and the factors significantly associated with it were child loss experience and quality of services. Hence, there is a need to improve the quality of services, both in terms of counselling and service provision, and women need to be counselled about the permanent nature of sterilization in order to avoid future regret.

Adult↗
PubMed2000

What is the signal for the posttranslational arginylation of proteins?

The N-terminal, posttranslational arginylation of proteins is ubiquitous in eukaryotic cells. Previous experiments, using purified components of the reaction incubated in the presence of exogenous substrates, have shown that only those proteins containing acidic residues at their N-terminals are arginylation substrates. However, data from experiments that used crude extracts of brain and nerve as the source of the arginylating molecules, suggest that the in vivo targets for arginylation are more complex than those demonstrated using purified components. One of the proposed functions for arginylation is as a signal for protein degradation and proteins that have undergone oxidative damage have been shown to be rapidly degraded. In the present experiments we have tested the hypothesis that the presence of an oxidatively damaged residue in a protein is a signal for its arginylation. These experiments have been performed by adding synthetic oxidized peptides to crude extracts of rat brain, incubating them with [3H]Arg and ATP and assaying for arginylated peptides using RP-HPLC. Results showed that while the oxidized A-chain of insulin was arginylated in this system, confirming previous experiments, other peptides containing oxidized residues were not. When a peptide containing Glu in the N-terminus was incubated under the same conditions it too was not a substrate for arginylation. These findings show that neither the presence of an N-terminal acidic residue nor an oxidized residue alone are sufficient to signal arginylation. Thus, another feature of the oxidized A-chain of insulin is required for arginylation. That feature remains to be identified.

Adenosine Triphosphate↗
PubMed2000

A rapid spectrofluorimetric technique for determining drug-serum protein binding suitable for high-throughput screening.

PURPOSE: To develop and validate a rapid method for determining the dissociation constants with which pharmaceutical candidates and drugs bind to serum albumin and to alpha1-acid glycoprotein with the goal of deducing the extent of binding. METHODS: The quenching of the intrinsic tryptophan fluorescence of serum albumin and alpha1-acid glycoprotein was monitored by spectrofluorimetry and the data were used to calculate the apparent dissociation constant. Sodium warfarin was used to probe the warfarin-binding site of serum albumin and diazepam was used to probe the benzodiazepine binding site. Additionally, the binding of sodium salicylate, phenylbutazone, sulfinpyrazone, iophenoxic acid, theophylline, chloramphenicol, acetaminophen, lithium chloride and ampicillin were also investigated. Chlorpromazine hydrochloride and imipramine hydrochloride were used as probes for alpha1-acid glycoprotein. The assays were also extended to the multiwell format. The quenching curves were fitted to the quadratic binding equation to determine the dissociation constants. RESULTS: Intrinsic fluorescence measurements are an excellent predictor of the drug binding to human serum albumin and to alpha1-acid glycoprotein. These measurements detect binding to the warfarin and benzodiazepine binding sites of human serum albumin. The dissociation constants estimated using the method compare favorably to the dissociation constants previously reported by Epps et al. using extrinsic fluorescence methodology, and the results correlate well with equilibrium dialysis using drug displacement endpoints. CONCLUSIONS: These measurements can be carried out with small samples and do not require separation of the bound and unbound species. Additionally, the proposed methods eliminate membrane separations, are not compound specific and do not require analytical chromatography or mass spectrometry for quantitation. Spectrofluorimetry may prove to be a useful method for rapidly determining the protein binding of combinatorial libraries.

Anticoagulants↗
PubMed2000

Pharmacokinetic variability and therapeutic drug monitoring actions at steady state.

PURPOSE: To develop a mathematical model for therapeutic drug monitoring and to assess the kinetic relationships between the intensity of corrective action and the approach of drug concentrations to target values. METHODS: A mathematical model that succinctly accounts for the corrective actions and the variability inherent in the pharmacokinetics was used. RESULTS: The validity of the variability term was tested using experimental data for steady state concentrations of the drug procainamide. The approach of the monitored process to the target value followed exponential kinetics and an analytical expression for dependence the variance with time and various dosing parameters was derived. The variance of the drug concentration depends critically on a single non-dimensional parameter containing the rate constant for the therapeutic corrective actions and a coefficient describing the variance rate. When the rate constant for the therapeutic corrective actions was less than this critical value, the variance increased indefinitely. CONCLUSIONS: From a dosing standpoint, large variances in drug concentrations are undesirable because some patients will be overdosed or underdosed. Since deterministic models cannot provide analytical solutions for the moments of drug concentration distribution functions, stochastic models can be used to provide useful insights into the design of therapeutic regimens.

Algorithms↗
PubMed2000

Preventing mediastinal shift after pneumonectomy does not abolish physiological compensation.

To determine the role of mediastinal shift after pneumonectomy (PNX) on compensatory responses, we performed right PNX in adult dogs and replaced the resected lung with a custom-shaped inflatable silicone prosthesis. Prosthesis was inflated (Inf) to prevent mediastinal shift, or deflated (Def), allowing mediastinal shift to occur. Thoracic, lung air, and tissue volumes were measured by computerized tomography scan. Lung diffusing capacities for carbon monoxide (DL(CO)) and its components, membrane diffusing capacity for carbon monoxide (Dm(CO)) and capillary blood volume (Vc), were measured at rest and during exercise by a rebreathing technique. In the Inf group, lung air volume was significantly smaller than in Def group; however, the lung became elongated and expanded by 20% via caudal displacement of the left hemidiaphragm. Consequently, rib cage volume was similar, but total thoracic volume was higher in the Inf group. Extravascular septal tissue volume was not different between groups. At a given pulmonary blood flow, DL(CO) and Dm(CO) were significantly lower in the Inf group, but Vc was similar. In one dog, delayed mediastinal shift occurred 9 mo after PNX; both lung volume and DL(CO) progressively increased over the subsequent 3 mo. We conclude that preventing mediastinal shift after PNX impairs recruitment of diffusing capacity but does not abolish expansion of the remaining lung or the compensatory increase in extravascular septal tissue volume.

Animals↗
PubMed2000

Detection of cytochrome P450 and other drug-metabolizing enzyme mRNAs in peripheral blood mononuclear cells using DNA arrays.

The purpose of this study was to determine whether the expression of cytochrome P450 (CYP) enzyme mRNAs, other drug-metabolizing enzyme mRNAs, and transporter mRNAs can be detected using DNA arrays. Total RNA was isolated from peripheral blood mononuclear cells of 10 multiple sclerosis patients and 10 age- and sex-matched controls. The mRNA was reverse transcribed to radiolabeled cDNA, and the resultant cDNA was used to probe a DNA array containing several thousand known human genes. The signals corresponding to several CYPs, drug-metabolizing, and transporter mRNAs was substantially above background. The results demonstrate that the DNA array technique has the sensitivity and the selectivity for applications in the pharmaceutical sciences. The mean values for mRNAs of specific CYPs and drug-metabolizing enzymes in peripheral blood cells were compared with reported values for liver. The capabilities of DNA arrays may prove useful for characterizing CYP expression in a variety of clinical samples.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Hyperglycaemia in pregnancy: effects on the offspring behaviour with special reference to anxiety paradigms.

Maternal hyperglycemic effect was studied on the offspring behaviour. Offspring were obtained from diabetic rats by mating a normal father with a diabetic mother (NFDM), diabetic father with normal mother (DFNM) and diabetic father with diabetic mother (DFDM). Rats were rendered diabetic by injecting streptozotocin (STZ, 50 mg/kg i.p.) in citrate buffer. Offspring were subjected to various anxiety parameters including open field exploratory behaviour, elevated plus maze and zero maze behaviours, and the social interaction tests at the age of 8 weeks. The results indicate that offspring of NFDM and DFDM showed anxiogenic activity on the elevated plus maze zero maze and the social interaction test. Offspring of NFDM and DFDM exhibited hyper and emotional activity in the open field behaviour test. The behavioural alterations observed in the offspring were comparable to the behavioural alterations noted in STZ diabetic rat as reported earlier. Further offspring of NFDM and DFDM exhibited mild hyperglycaemia. No significant behavioural alterations in the offspring of DFNM were observed. It may be concluded, that exposure of offspring to diabetic environment in their foetal life can lead to anxiogenic/emotional behaviours in adult life.

Animals↗
PubMed1999

Oxidative stress increases ubiquitin--protein conjugates in synaptosomes.

Synaptosomes were incubated in the presence of FeSO4 to test the hypothesis that iron-catalyzed oxidative damage causes an increase in the ubiquitination of synaptosomal proteins. Incubation with 10 or 50 microM FeSO4 caused concentration-dependent increases in carbonyl groups (an indication of protein oxidation) and ubiquitinated proteins (determined by probing Western blots with a monoclonal antibody to ubiquitin). Differences in protein ubiquitination occurred within 5 min of incubation, indicating a rapid response to oxidative stress. Results of experiments with MG-132, an inhibitor of the degradation of ubiquitinated proteins, suggested that oxidative damage stimulated ubiquitination rather than inhibited degradation of ubiquitinated proteins. The data are consistent with the hypothesis that synaptic terminals utilize the ubiquitin/proteasome proteolytic pathway to degrade oxidatively damaged proteins.

Animals↗
PubMed1999

Evaluation of an alternative to the Kolmogorov-Smirnov test for flow cytometric histogram comparisons.

BACKGROUND: The Kolmogorov-Smirnov test is a valid statistical test for comparing distributions that has been recommended for flow cytometric histogram analysis. However, this test is frequently found to be too sensitive for flow cytometric histogram comparisons. Here, a parametric alternative to the Kolmogorov-Smirnov test is proposed that is based on fitting suitable models to flow cytometric data. METHODS: Several flow cytometric histograms derived from cell surface immunophenotyping for intercellular adhesion molecule-1 (ICAM-1) on K562 cells were analyzed using numerical modeling. The prediction intervals derived from the modeling were used for decision making. RESULTS: The residuals after peak fitting flow cytometric data are normally distributed and this permits the use of the prediction limit methodology. The usefulness of the approach for analyzing flow cytometry histograms is examined and the method is shown to avoid the "sensitivity" disadvantages associated with the Kolmogorov-Smirnov test. CONCLUSIONS: The prediction limit method is a viable alternative to the Kolmogorov-Smirnov method.

Flow Cytometry↗
PubMed1999

Flow cytometric analysis of in vitro proinflammatory cytokine secretion in peripheral blood from multiple sclerosis patients.

The cytokines, interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), and interleukin-2 (IL-2) are important endogenous proinflammatory proteins and have been linked to disease activity in multiple sclerosis. In this study, we use flow cytometric methodology to compare the secretion of IFN-gamma, IL-2, and TNF-alpha from peripheral blood-derived T cells of multiple sclerosis patients to the secretion in healthy controls. The percentages of IFN-gamma, IL-2, and TNF-alpha secreting cells are not significantly different between multiple sclerosis patients and controls. However, the TNF-alpha secreting CD3 cell percentage is correlated with the IFN-gamma and IL-2 secreting CD3 cell percentages in multiple sclerosis patients. In the controls, only the TNF-alpha secreting CD3 cell percentage is correlated with IFN-gamma. These findings show that correlated secretion of cytokines occurs in multiple sclerosis and suggest that concerted intercytokine interactions may play an important role in the disease.

Adolescent↗
PubMed1999

Compensatory alveolar growth normalizes gas-exchange function in immature dogs after pneumonectomy.

To determine the extent and sources of adaptive response in gas-exchange to major lung resection during somatic maturation, immature male foxhounds underwent right pneumonectomy (R-Pnx, n = 5) or right thoracotomy without pneumonectomy (Sham, n = 6) at 2 mo of age. One year after surgery, exercise capacity and pulmonary gas-exchange were determined during treadmill exercise. Lung diffusing capacity (DL) and cardiac output were measured by a rebreathing technique. In animals after R-Pnx, maximal O2 uptake, lung volume, arterial blood gases, and DL during exercise were completely normal. Postmortem morphometric analysis 18 mo after R-Pnx (n = 3) showed a vigorous compensatory increase in alveolar septal tissue volume involving all cellular compartments of the septum compared with the control lung; as a result, alveolar-capillary surface areas and DL estimated by morphometry were restored to normal. In both groups, estimates of DL by the morphometric method agreed closely with estimates obtained by the physiological method during peak exercise. These data show that extensive lung resection in immature dogs stimulates a vigorous compensatory growth of alveolar tissue in excess of maturational lung growth, resulting in complete normalization of aerobic capacity and gas-exchange function at maturity.

Animals↗