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R Singh

Publications and source records attributed to R Singh.

At least 55 records · Page 3Linked to original sources
PubMed2005

Blue native polyacrylamide gel electrophoresis and the monitoring of malate- and oxaloacetate-producing enzymes.

We demonstrate a facile blue native polyacrylamide gel electrophoresis (BN-PAGE) technique to detect two malate-generating enzymes, namely fumarase (FUM), malate synthase (MS) and four oxaloacetate-forming enzymes, namely pyruvate carboxylase (PC), phosphoenolpyruvate carboxykinase (PEPCK), citrate lyase (CL) and aspartate aminotransferase (AST). Malate dehydrogenase (MDH) was utilized as a coupling enzyme to detect either malate or oxaloacetate in the presence of their respective substrates and cofactors. The latter four oxaloacetate-forming enzymes were identified by 2,6-dichloroindophenol (DCIP) and p-iodonitrotetrazolium (INT) while the former two malate-producing enzymes were visualized by INT and phenazine methosulfate (PMS) in the reaction mixtures, respectively. The band formed at the site of enzymatic activity was easily quantified, while Coomassie staining provided information on the protein concentration. Hence, the expression and the activity of these enzymes can be readily evaluated. A two-dimensional (2D) BN-PAGE or SDS-PAGE enabled the rapid purification of the enzyme of interest. This technique also provides a quick and inexpensive means of quantifying these enzymatic activities in normal and stressed biological systems.

Aspartate Aminotransferases↗
PubMed2005

DNA adducts: mass spectrometry methods and future prospects.

Detection of DNA adducts is widely used for the monitoring of exposure to genotoxic carcinogens. Knowledge of the nature and amounts of DNA adducts formed in vivo also gives valuable information regarding the mutational effects that may result from particular exposures. The power of mass spectrometry (MS) to achieve qualitative and quantitative analyses of human DNA adducts has increased greatly in recent years with the development of improved chromatographic interfaces and ionisation sources. Adducts have been detected on nucleic acid bases, 2'-deoxynucleosides or 2'-deoxynucleotides, with LC-MS/MS being the favoured technique for many of these analyses. Our current applications of this technique include the determination of N7-(2-carbamoyl-2-hydroxyethyl)-guanine, which was postulated to be found as a DNA repair product in urine following exposure to acrylamide, and of 8-oxo-7,8-dihydro-2'-deoxyguanosine and 8-oxo-7,8-dihydro-2'-deoxyadenosine, as markers of oxidative damage in human lymphocyte DNA. Higher sensitivity (with a detection limit of 1-10 adducts/10(12) nucleotides) may be achieved by the use of accelerator mass spectrometry (AMS), although this requires the presence of certain isotopes, such as [(14)C], in the material being analysed. In order to make this technique more amenable for studies of human exposure to environmental carcinogens, new postlabelling techniques, incorporating [(14)C] into specific DNA adducts after formation, are being developed. It is expected that combining the use of advanced MS techniques with existing (32)P-postlabelling and immunochemical methodologies will contribute greatly to the understanding of the burden of human exposure to environmental carcinogens.

Animals↗
PubMed2005

Predictive value of arterial ammonia for complications and outcome in acute liver failure.

BACKGROUND AND AIMS: In acute liver failure (ALF), the brain is exposed to high levels of ammonia. Human studies defining the clinical significance of ammonia in ALF are lacking. This prospective study evaluated the relationship of arterial ammonia levels at admission to complications and survival among patients with ALF. METHODS: Eighty consecutive ALF patients admitted from March 2001 to December 2003 were followed up until death or complete recovery. All had arterial ammonia estimation at admission (enzymatic method). Logistic regression analysis was performed to identify independent predictors of mortality. RESULTS: Forty two (52.5%) patients died. Non-survivors had significantly higher median ammonia levels than survivors (174.7 v 105.0 micromol/l; p<0.001). An arterial ammonia level of > or = 124 micromol/l was found to predict mortality with 78.6% sensitivity and 76.3% specificity, and had 77.5% diagnostic accuracy. Patients with higher ammonia levels also developed more complications, including deeper encephalopathy (p = 0.055), cerebral oedema (p = 0.020), need for ventilation (p<0.001), and seizures (p = 0.006). Logistic regression analysis showed that pH, presence of cerebral oedema, and arterial ammonia at admission were independent predictors of mortality (odds ratios 6.6, 12.6, and 10.9, respectively). Incorporating these variables, a score predicting mortality risk at admission was derived: 2.53 + 2.91 ammonia + 2.41 oedema + 1.40 pH, where ammonia is scored as 0 (if <124 micromol/l) or 1 (if > or =124 micromol/l); oedema is scored as 0 (absent) or 1(present); and pH is scored as 1 (if < or =7.40) or 0 (if >7.40). Levels of partial pressure of ammonia were equally correlated with outcome. CONCLUSION: Arterial ammonia at presentation is predictive of outcome and can be used for risk stratification. Ammonia lowering therapies in patients with ALF should be evaluated.

Adolescent↗
PubMed2005

Tetrahydrogestrinone is an androgenic steroid that stimulates androgen receptor-mediated, myogenic differentiation in C3H10T1/2 multipotent mesenchymal cells and promotes muscle accretion in orchidectomized male rats.

The discovery of tetrahydrogestrinone (THG) abuse by several elite athletes led the U.S. Congress to declare it a controlled substance, although conclusive evidence of its anabolic/androgenic activity is lacking. We determined whether THG affects myogenic differentiation and androgen receptor (AR)-mediated signaling, whether it binds to AR, and whether it has androgenic and anabolic effects in vivo. Accordingly, we measured the dissociation constant for THG with a fluorescence anisotropy assay using recombinant AR-ligand binding domain. The AR nuclear translocation and myogenic activity of androstenedione were evaluated in mesenchymal, multipotent C3H10T1/2 cells. We performed molecular modeling of the THG:AR interaction. The androgenic/anabolic activity was evaluated in orchidectomized rats. THG bound to AR with an affinity similar to that of dihydrotestosterone. In multipotent C3H10T1/2 cells, THG upregulated AR expression, induced AR nuclear translocation, dose dependently increased the area of myosin heavy chain type II-positive myotubes, and up-regulated myogenic determination and myosin heavy chain type II protein expression. The interaction between AR and the A ring of THG was similar to that between AR and the A ring of dihydrotestosterone, but the C17 and C18 substituents in THG had a unique stabilizing interaction with AR. THG administration prevented the castration-induced atrophy of levator ani, prostate gland, and seminal vesicles and loss of fat-free mass in orchidectomized rats. We conclude that THG is an anabolic steroid that binds to AR, activates AR-mediated signaling, promotes myogenesis in mesenchymal multipotent cells, and has anabolic and androgenic activity in vivo. This mechanism-based approach should be useful for rapid screening of anabolic/androgenic agents.

Animals↗
PubMed2005

A prospective study of the long-term efficacy of local methyl prednisolone acetate injection in the management of mild carpal tunnel syndrome.

OBJECTIVE: Local glucocorticoid injections are used to treat carpal tunnel syndrome (CTS). However, this treatment is associated with frequent relapses. An important limitation of studies with higher relapse rates is that no attempt has been made to identify patients with mild or severe disease. We evaluated the efficacy of local glucocorticoid injection in patients with mild CTS. METHOD: Mild CTS was defined as intermittent symptoms without absence of sensations, muscle atrophy or weakness of the thenar muscles. Forty-eight patients with idiopathic mild CTS were evaluated before and 3 and 12 months after a single local injection of 40 mg methyl prednisolone acetate. Outcome was assessed by overall satisfaction on a 100 mm visual analogue scale, the Boston self-administered questionnaire for symptom severity and functional scores and improvement in the electrophysiological parameters. RESULTS: At 3 months, 93.7% of the patients reported marked improvement in their symptoms, with significant improvement in the mean values of the nerve conduction parameters distal motor latency at the wrist (DML) (P = 0.00001), distal sensory latency at mid-palm (DSL MP) (P = 0.014) and distal sensory latency at the wrist (DSL W) (P = 0.0003), and symptom severity (P = 4.96 x 10(-8)) and the functional scores (P = 3.56 x 10(-5)). Significant improvement was still present for DML (P = 1.39 x 10(-5)) at 12 months. Almost 50% of the patients achieved normalization in the electrophysiological study. At a median follow-up of 16 months, 79% patients continued to have improvement in their symptoms. Eight patients (16.6%) relapsed following the initial response. CONCLUSIONS: Local glucocorticoid injection results in long-term improvement in nerve conduction parameters, symptom severity and functional scores in patients with mild CTS.

Adult↗
PubMed2005

Microsatellite-based high density linkage map in oil palm (Elaeis guineensis Jacq.).

A microsatellite-based high-density linkage map for oil palm (Elaeis guinensis Jacq.) was constructed from a cross between two heterozygous parents, a tenera palm from the La Me population (LM2T) and a dura palm from the Deli population (DA10D). A set of 390 simple sequence repeat (SSR) markers was developed in oil palm from microsatellite-enriched libraries and evaluated for polymorphism along with 21 coconut SSRs. A dense and genome-wide microsatellite framework as well as saturating amplified fragments length polymorphisms (AFLPs) allowed the construction of a linkage map consisting of 255 microsatellites, 688 AFLPs and the locus of the Sh gene, which controls the presence or absence of a shell in the oil palm fruit. An AFLP marker E-Agg/M-CAA132 was mapped at 4.7 cM from the Sh locus. The 944 genetic markers were distributed on 16 linkage groups (LGs) and covered 1,743 cM. Our linkage map is the first in oil palm to have 16 independent linkage groups corresponding to the plant's 16 homologous chromosome pairs. It is also the only high-density linkage map with as many microsatellite markers in an Arecaceae species and represents an important step towards quantitative trait loci analysis and physical mapping in the E. guineensis species.

Arecaceae↗
PubMed2005

Effect of blood transfusion in combination with Dextran-40 and hypertonic saline solution on cardiopulmonary haemodynamics of endotoxin (lipopolysaccharide) shock in buffalo calves.

The intravenous (i.v.) infusion of lipopolysaccharide (LPS) of E. coli endotoxin in buffalo calves (n = 15) at 5 microg/kg bw per h for 3 h caused a significant (p<0.05) fall in plasma volume, blood volume, haematocrit haemoglobin, and systolic, diastolic and pulse pressure, mean arterial pressure and central venous pressure (CVP), with a marked rise in respiration. Treatment with a combination of i.v. infusion of 7.2% hypertonic saline solution, Plasmex-D-40 (Dextran-40) and blood successfully alleviated hypovolaemia, and raised systolic, diastolic and pulse pressure, mean arterial pressure and central venous pressure. The whole blood was collected from apparently healthy male buffalo calves 24 h prior to infusion and was transfused without cross-matching. No significant fall in haemoglobin, haematocrit and body temperature was observed after transfusion. All these values tended to remain near normal levels. However, this combination of treatment had no effect on high respiratory rate. A one-time blood transfusion did not evoke any cross-reaction and was helpful in raising haematocrit and haemoglobin close to pre-infusion values. The general symptoms of restlessness, respiratory distress, profuse salivation, violent movement of the ears, snoring, intermittent struggle, etc. were markedly reduced. All the treated animals became quiet and lay with eyes open and survived the 7 h of observation.

Animals↗
PubMed2005

Chronomics of circulating plasma lipid peroxides and anti-oxidant enzymes and other related molecules in cirrhosis of liver. In the memory of late Shri Chetan Singh.

BACKGROUND: The chronome (from chronos, time, and nomos, rule; time structure) of lipid peroxidation and anti-oxidant defense mechanisms may relate to the efficacy and management of preventive and curative chronotherapy. PATIENTS AND METHODS: Thirty patients with liver cirrhosis, 25-45 years of age, and 60 age-matched clinically healthy volunteers were synchronized for 1 week with diurnal activity from about 06:00 to about 22:00 and nocturnal rest. Breakfast was around 08:30, lunch around 13:30 and dinner around 20:30. Drugs known to affect the free-radical system were not taken. Blood samples were collected at 6-h intervals for 24 h under standardized, presumably 24-h synchronized conditions. Determinations included plasma lipid peroxides, in the form of malondialdehyde (MDA), blood superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR) activities, and serum total protein, albumin, ascorbic acid, and uric acid concentrations. RESULTS: A marked circadian variation was demonstrated for each variable in each group by population-mean cosinor (P < 0.01). In addition to anticipated differences in overall mean value (MESOR), patients differed from healthy volunteers also in terms of their circadian pattern. CONCLUSION: Mapping the broader time structure (chronome) with age and multifrequency rhythm characteristics of antioxidants and pro-oxidants is needed for exploring their putative role as markers in the treatment and management of liver cirrhosis.

Adult↗
PubMed2005

Occurrence of yellow mosaic geminiviral disease on bitter gourd (Momordica charantia) and its impact on phytochemical contents.

Natural occurrence of yellow mosaic disease was observed on bitter gourd (Momordica charantia). Association of geminivirus with the disease was investigated through polymerase chain reaction using geminivirus-specific primers and Southern hybridization with a probe prepared from the cloned DNA of a known geminivirus. The fruits, leaves and stem of infected and healthy plants were studied for phytochemical composition. The amounts of protein were 49%, 50% and 66% higher, total carotenoids were 36%, 33% and 40% lower, vitamin C were 23%, 48% and 50% lower, total phenols were 28%, 31% and 43% lower, and antioxidant activity were 36%, 48% and 43% lower in the severely virus infected fruits, leaves and stem, respectively, as compared with healthy plants. The loss in the quantity of these phytochemicals was also observed even in mild infected plants, which further increased with the severity of the symptoms. Similarly, ethanol and 50% ethanol soluble extractive were also 25-43% lower in the fruits, leaves and stem of infected plants as compared with the healthy plants. A 45% and 54% lower caffeic acid, and 78% and 59% lower amounts of ferulic acid in the fruits and stem, respectively, and a 25% loss of gallic acid were noticed in the leaves of the severely infected plants.

Antioxidants↗
PubMed2005

Increased expression of Mycobacterium tuberculosis 19 kDa lipoprotein obliterates the protective efficacy of BCG by polarizing host immune responses to the Th2 subtype.

Mycobacterium tuberculosis can not only neutralize immune effector functions, but also has the ability to modulate host-signalling cascades involved in the development of these responses. The 19 kDa antigen (Rv3763), a lipoprotein of M. tuberculosis, elicits high levels of interleukin (IL)-12 from macrophages in addition to its powerful immunomodulatory properties, leading to suppression of antigen-presentation signalling cascades. The present study was aimed at analysing the effect of overexpression of this antigen on the immunostimulatory properties of M. bovis Bacille Calmette-Guerin (BCG). We have constructed a recombinant BCG strain (rBCG19N) producing higher levels of the 19 kDa antigen in both the cytoplasmic (approximately eightfold) and extracellular (approximately fivefold) fractions as compared to the wildtype BCG. Immunization of mice with rBCG19N elicited high levels of interferon-gamma (IFN-gamma) and relatively low levels of IL-10 against the purified 19 kDa antigen. However, in response to total BCG sonicate, mice immunized with rBCG19N produced significantly high levels of IL-10 with relatively very low levels of IFN-gamma. This polarization of the host immune responses towards T-helper 2 subtype resulted in complete abrogation of the protective efficacy of BCG, when rBCG19N was used as a live vaccine against M. tuberculosis challenge in guinea pigs.

Animals↗
PubMed2005

Lifetime risk of symptomatic carpal tunnel syndrome in Type 1 diabetes.

AIMS: To determine the lifetime risk of developing symptomatic carpal tunnel syndrome (CTS) in subjects with Type 1 diabetes and to investigate the effect of glycaemic control, body mass index (BMI), gender and age of onset of diabetes. METHODS: One hundred and twenty-two consecutive subjects with Type 1 diabetes seen by a single investigator, over 1-year, were questioned about previous surgery for, or current symptoms of CTS. Those with current symptoms were referred for nerve conduction studies (NCS). Those with previous surgery and/or diagnostic NCS were classified as having CTS. Lifetime risk was calculated by the Kaplan-Meier method. RESULTS: Twenty six out of 122 patients had CTS, 18 of whom (69%) had undergone median nerve decompression surgery. The predicted lifetime risk of CTS reached 85% after 54 years of Type 1 diabetes (95% confidence interval: 72-97%). The duration of diabetes was greater in those who had developed CTS than in those who had not (29 vs. 19 years, P=0.0001). In those diagnosed with diabetes before the age of 20 there was a lag time of at least 19 years before the development of CTS, whereas patients with later onset of diabetes, began to develop CTS from as early as 5 years diabetes duration. There was no demonstrable effect of glycaemic control, obesity, gender or retinopathy on the appearance of CTS. CONCLUSION: The lifetime risk of developing symptomatic CTS with Type 1 diabetes is high, and is related to age and duration of diabetes, but not to the development of microvascular complications.

Adolescent↗
PubMed2005

Primary alloproliferative TH1 response induced by immature plasmacytoid dendritic cells in collaboration with myeloid DCs.

The role played by dendritic cell (DC) subsets in the immune response to alloantigens is not well defined. In vitro experiments have extensively shown that freshly isolated myeloid (M)DCs induce a strong T lymphocyte proliferation whereas plasmacytoid (P)DCs do not, unless activated by CD40 ligation. The aim of these studies was to explore whether the interplay among PDCs, MDCs and T cells modulates alloresponse. Freshly isolated MDCs and PDCs were merged in different proportions and used as antigen presenting cells (APCs) in mixed lymphocyte cultures (MLC). As described, isolated PDCs only induced a mild alloresponse, while MDCs were potent inducers of alloproliferation. Unexpectedly, when PDCs were merged with even low numbers of MDCs (down to 100 cells) and used as APCs, a potent Th1 cell proliferation was detected. Survival and maturation of PDCs was increased in these MLC conditions, which could partially explain the magnitude of the T-cell response. Interestingly, the proportion of IFNgamma-producing cells generated in such cultures was higher compared to MDC-stimulated cultures. These data suggest that the interaction between both DC subsets is determinant to generate a potent Th1 response, at least in an allogeneic situation, and may be relevant to the outcome of allogeneic stem cell transplantation.

Animals↗
PubMed2005

Impression cytology of the ocular surface.

Impression cytology refers to the application of a cellulose acetate filter to the ocular surface to remove the superficial layers of the ocular surface epithelium. These cells can then be subjected to histological, immunohistological, or molecular analysis. Proper technique is essential as the number of cells sampled can vary considerably. Generally two to three layers of cells are removed in one application but deeper cells can be accessed by repeat application over the same site. Applications for impression cytology include diagnosing a wide range of ocular surface disorders, documenting sequential changes in the conjunctival and corneal surface over time, staging conjunctival squamous metaplasia, and monitoring effects of treatment. It is also a useful investigational tool for analysing ocular surface disease with immunostaining and DNA analysis. It is non-invasive, relatively easy to perform, and yields reliable information about the area sampled with minimal discomfort to the patient. Major ophthalmic centres should develop and introduce this technique into routine clinical practice. This is best achieved with a team approach including the ophthalmologist, pathologist, microbiologist, and the immunologist.

Conjunctiva↗
PubMed2005

Anthropometric and physiological profiles of sepak takraw players.

OBJECTIVES: Anthropometric and physiological profiles of national sepak takraw players were determined. METHODS: Thirty nine players, specialising in the three playing positions (tekong/server, feeder, and killer/spiker) were divided into three age categories of under 15 (U15), under 18 (U18), and under 23 (U23) years of age. Height, weight, percent body fat (%bf), maximum oxygen consumption (Vo(2max)), range of motion (ROM), back and leg strength, and heart rate, for the estimation of oxygen consumption during matches, were recorded. Statistical analysis was performed using one way ANOVA for independent measurements and data are presented as mean+/-standard deviation. RESULTS: The U23 players were significantly taller and heavier with significantly better ROM of the neck, trunk, and ankle joints and back and leg strength than the U15 players. No significant difference was found in %bf between the three age categories. Mean maximum heart rate during exercise was significantly higher in the U15 group when compared to the U18 and U23 groups (p<0.05). Mean Vo2max was similar between the three groups. Estimated oxygen consumption during matches was 69.1%, 68.5%, and 56.4% of Vo2max in the killer, tekong, and the feeder groups, respectively. CONCLUSIONS: The mean height, body weight, and cardiopulmonary capacities of the players were within the Malaysian population norms, but were somewhat lower than those of players of other court games from other countries. %bf was also lower in these players. This study provides the much needed anthropometric and physiological data of sepak takraw players for further development of this sport.

Adolescent↗