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Biomedical subjects

S K Gupta

Publications and source records attributed to S K Gupta.

At least 271 records · Page 15Linked to original sources
PubMed1998

Dog zona pellucida glycoprotein-3 (ZP3): expression in Escherichia coli and immunological characterization.

An internal fragment (978 bp) corresponding to the dog zona pellucida glycoprotein-3 (DZP3), excluding the N-terminal signal sequence and the C-terminal transmembrane-like domain, was amplified by polymerase chain reaction from a full-length cDNA clone. The amplified SacI and PstI restricted fragment was cloned in-frame downstream of the T5 promoter under lac operator control for expression in the pQE-30 vector. Recombinant DZP3 (rec-DZP3) was expressed as a polyhistidine fusion protein in Escherichia coli. Optimum expression of rec-DZP3 was observed at 1.0 mM isopropyl-beta-D-thiogalactopyronoside. Immunoblots with a murine monoclonal antibody, MA-451 (raised against porcine ZP3beta-a homologue of DZP3 and cross-reactive with dog zona pellucida), revealed a major band of 42 kDa. Localization studies revealed that the recombinant protein was present only in an insoluble intracellular fraction. Further optimization studies revealed that the level of expression of rec-DZP3 was significantly higher in Luria broth medium containing glycerol rather than glucose and maximum expression was observed when cultures were induced during the mid-log phase of growth. Batch fermentation with glycerol as the carbon source yielded 30 mg/L of rec-DZP3 compared to 4 mg/L from a shake flask culture. Immunization of two male rabbits with Ni-NTA-purified rec-DZP3 and two female dogs with the rec-DZP3 conjugated to diphtheria toxoid generated high antibody titers against rec-DZP3 as determined by enzyme-linked immunosorbent assay. Rabbit immune serum reacted with porcine ZP3beta but failed to react with porcine ZP3alpha in a Western blot. Moreover, antisera when tested by indirect immunofluorescence on dog ovarian sections showed positive fluorescence with zona pellucida. The availability of rec-DZP3 will help in evaluating its efficacy for fertility regulation in stray dogs.

Animals↗
PubMed1998

Analgesic efficacy and pharmacokinetics of topical nimesulide gel in healthy human volunteers: double-blind comparison with piroxicam, diclofenac and placebo.

OBJECTIVE: The present study was conducted to compare the analgesic efficacy of a new topical gel formulation of nimesulide (10 mg of pure drug) with that of placebo, diclofenac and piroxicam gels (10 mg of pure drug) in three parallel groups in a double-blinded, randomized fashion with vehicle placebo. The analgesic activity of nimesulide was subsequently correlated with its pharmacokinetic profile. METHODS: The drugs were applied on a fixed marked area on the skin of the right forearm. Pain stimulus was administered using a modification of the Hollander method, before and at 15, 30, 60, 120 min and 240 min post-treatment. The pain experienced by the subjects was ranked separately on the visual analogue scale (VAS) and the ten-point category scale. Antinociception induced by the treatments was evaluated through the placebo-related ratings (PRR) and total pain relief (TOTPAR) analysis. The plasma concentration of nimesulide was estimated using high-performance liquid chromatography (HPLC). RESULTS: Nimesulide exhibited better efficacy than diclofenac, piroxicam and placebo. It demonstrated faster onset of action in concordance with earlier studies. Peak analgesic effect was observed at 120 min post-treatment, which correlated with the pharmacokinetic profile of the drug in gel formulation. In this study, diclofenac was found to be superior to piroxicam though both drugs exhibited peak analgesic effect at 60 min post-treatment. In the modified Hollander method, a good correlation was found between the ten point category scale and the VAS, indicating that it may serve as a sensitive and reliable method for the screening of analgesic drugs. CONCLUSION: The superior analgesic activity of nimesulide (as gel formulation), correlating with its pharmacokinetic profile, indicates that the topical route of administration may be a safe and effective alternative to the presently used oral and rectal routes.

Administration, Topical↗
PubMed1998

Polymerase chain reaction-based risk assessment for Wilms tumor in sporadic aniridia.

PURPOSE: Sporadic cases of aniridia have a 30% risk for the development of Wilms tumor. Current guidelines for sporadic aniridia recommend screening by renal ultrasonography for the presence of tumors every 6 months until age 5 years. Deletions of chromosome 11p13 that affect both PAX6 (aniridia) and WT1 (Wilms tumor) loci are the basis for the association of these two uncommon disorders. We sought to develop a rapid polymerase chain reaction-based test that could rule out a chromosome 11p13 deletion covering the PAX6-WT1 region in sporadic aniridia. METHODS: Five patients with sporadic aniridia were recruited. Polymerase chain reaction-based genotyping was carried out for six highly informative marker loci across the PAX6-WT1 region to determine whether these patients had one or two haplotypes. The results were compared with those obtained from two cell lines with known deletions in the PAX6-WT1 region. RESULTS: All five patients were heterozygous at least at one of the four marker loci in the PAX6-WT1 region, indicating that there were no cases of gross chromosomal deletion. The cell lines showed hemizygosity in the four marker loci within the PAX6-WT1 region and in one of the two flanking marker loci. CONCLUSIONS: We have developed a rapid DNA test with an estimated sensitivity of 94.0% to 99.2%, using standard DNA diagnostic techniques and equipment, to rule out chromosomal deletion in sporadic aniridia. Patients in whom a chromosome 11p13 deletion has been ruled out do not require repeated renal imaging to screen for Wilms tumor.

Aniridia↗
PubMed1998

Effects of PAX6 mutations on retinal function: an electroretinographic study.

PURPOSE: To investigate the retinal function in aniridic patients with documented PAX6 mutations to determine the range of electroretinogram abnormalities in aniridic patients and to relate electroretinogram findings with specific PAX6 mutations. METHODS: Eleven patients with typical aniridia and fully characterized PAX6 mutations underwent electroretinography. RESULTS: In all 11 patients, electroretinogram recordings were abnormal, ranging from mild to severe. Rod-related and cone-related activities were equally affected. The amplitude of the oscillatory potentials was the most reduced, followed by the b-wave, then to a milder degree the a-wave. Mutations affecting the paired domain of the PAX6 protein had the biggest impact on the electroretinogram amplitudes. Implicit times were increased in a subgroup with mutations affecting only the homeodomain. CONCLUSION: Patients with aniridia have varying degree of retinal dysfunction, ranging from severely abnormal to almost normal. The paired domain appears to have more impact on retinal function than other regions of the PAX6 protein. It is unclear whether mutations affecting the homeodomain lead to alteration of the photoreceptor function.

Adolescent↗
PubMed1998

Genotype/phenotype correlations in aniridia.

PURPOSE: To detect and characterize mutations in cases of familial and sporadic aniridia in Maritime Canada, and to look for indications of genotype/phenotype correlation within the cohort. METHODS: Twelve consecutive and unrelated patients (probands) who had total or nearly complete absence of irides, and four affected relatives, were recruited from Maritime Canada. Clinical data were obtained by chart review and electroretinogram testing. Mutations in the PAX6 gene were detected by single-strand conformation polymorphism and characterized by sequence analysis. RESULTS: Eleven different PAX6 mutations, 10 of which are novel, were found. The four patients with congenital cataracts all had mutations in the C-terminal proline-serine-threonine (PST)-rich domain of the PAX6 protein. Electroretinograms of nine of 11 patients displayed depressed scotopic maximum response b-wave amplitudes. The greatest decrease in b-wave amplitudes was seen in patients in whom the paired domain was disrupted by mutation. CONCLUSION: Some aspects of the phenotype of aniridia appear to correlate with the predicted effect of point mutations on the paired and PST domains of the PAX6 protein.

Adolescent↗
PubMed1998

Lattice corneal dystrophy type 1 in a Canadian kindred is associated with the Arg124 --> Cys mutation in the kerato-epithelin gene. sgupta@ogh.on.ca.

PURPOSE: To identify the mutation responsible for lattice corneal dystrophy type 1 in an extended Canadian kindred. METHODS: A search for a mutation in the candidate gene, kerato-epithelin, was carried out by single-strand conformation polymorphism and sequencing analyses. RESULTS: AC --> T mutation at position 417 was detected in exon 4 of the kerato-epithelin gene, which is expected to cause an Arg124 --> Cys change. This is the same nucleotide change described previously in two Swiss families with lattice corneal dystrophy type 1. CONCLUSION: Although the possibility that the three families (two previously described Swiss families and this Canadian kindred) are related has not been excluded, it appears that the unique phenotype of lattice corneal dystrophy type 1 is caused by this particular amino acid change.

Arginine↗
PubMed1998

Passive versus electrotransport-facilitated transdermal absorption of ketorolac.

OBJECTIVE: To investigate the bioavailability (extent and rate of absorption) of ketorolac from two cutaneous absorption sources, active electrotransport and passive transdermal, and to examine the enantiomeric selectivity of bioavailability for each source. METHODS: Based on a crossover study in 12 healthy volunteers, the extent and rate of absorption of ketorolac, delivered by a patch, were found by estimating the input rate function of the drug. For that purpose, deconvolution was used in two steps. First, intravenous data were analyzed to estimate the ketorolac disposition function, and second, postpatch data were deconvolved to estimate the unknown patch input profile given the disposition function estimated in the first step. Because the input rate function curves to be estimated for the patches may be of arbitrary shape, a spline was used to model the patch input function, whereas intravenous data were modeled with use of a sum of exponentials. Differences in the extent of absorption (F) for the four treatment-enantiomer combinations were further examined with a mixed-effect regression model, based on the sets of four individual estimates of bioavailability. RESULTS: On average, the F value for the active electrotransport treatment, which exhibited the faster absorption rate, was four times greater than the F for the passive transdermal treatment. Further, during the passive treatment, R-ketorolac yielded an average F that is 42% greater than that for S-ketorolac and also exhibited a smaller absorption lag-time. During the active treatment, there was no important enantiomeric difference in either extent or rate of absorption.

Administration, Cutaneous↗
PubMed1998

Effects of isolation and culture of turkey primary follicular oocytes on morphology and germinal vesicle integrity.

A novel approach to the production of transgenic poultry is to use primary follicular oocytes (PFOs). However, fundamental information regarding the impact of isolation and culture procedures on PFO integrity is lacking. This study describes the isolation and culture of PFOs from mature turkeys and the effects of these procedures on PFO morphology and germinal vesicle (GV) integrity. To isolate PFOs, ovarian cortex was incubated in trypsin-EDTA alone or further incubated in collagenase plus hyaluronidase (CH). About 200 to 500 PFOs, ranging in size from less than 100 microns in diameter to 1,000 microns, were recovered from each ovary. The culture of PFOs less than 100 microns in diameter for 4 h resulted in blebbing of the oolemma followed by extrusion of ooplasm. Primary follicular oocytes 100 to 250 microns in diameter survived culture for 24 h whereas larger PFOs survived for up to 7 d. Those PFOs with intact granulosa cell investments survived longer than those fully or partially denuded of granulosa cells with CH. Co-culture of PFOs (100 to 250 microns in diameter) on a monolayer of granulosa cells derived from mature, yellow-yolk follicles augmented PFO survival rates. The rate of GV breakdown was not influenced by the isolation or culture of the PFO. These data provide the basis for developing procedures for the in vitro maturation and in vitro fertilization of isolated PFOs.

Animals↗
PubMed1998

Effect of current density on pharmacokinetics following continuous or intermittent input from a fentanyl electrotransport system.

The pharmacokinetics of fentanyl were determined in two open-label crossover studies following 24-h periods of delivery by an electrotransport transdermal system (E-TRANS [fentanyl] system) in young healthy male volunteers. A direct current was applied continuously in study 1 (at 50, 100, and 200 microA; surface area = 5 cm2; n = 8), but in study 2 it was limited to the first 20 min of each hour (at 150, 200, and 250 microA; surface area = 2 cm2; n = 12). The opioid effects of fentanyl were blocked with naltrexone administered every 12 h. With increasing electrical current, the increase in serum fentanyl concentration, amount absorbed, and AUC values were proportional in study 2 but not in study 1. It is hypothesized that the lack of proportionality in study 1 is due to lower current density (microA/cm2) in this study. It appears that for fentanyl, the current density should be about 75 microA/cm2 or greater for a linear relation between current and amount absorbed as seen in study 2. Compared with intravenously infused fentanyl, the serum concentrations resulting from E-TRANS (fentanyl) system application revealed a slightly dampened rate of increase (stratum-corneum barrier effect) and decrease in serum concentrations, and a similar intersubject variability in fentanyl AUC values. Fentanyl pharmacokinetics with either E-TRANS (fentanyl) or intravenous infusion were time-invariant over a 24-h application period, with similar mean half-life values (about 15-18 h). E-TRANS (fentanyl) administration (either continuous or intermittent input) was safe and well tolerated. Adverse effects were mild to moderate; they consisted mainly of local erythema and pruritus (which resolved in most patients within 24 h after system removal) and occasional opioid effects.

Adult↗
PubMed1998

Topical pefloxacin in bacterial keratitis.

PROBLEM: To evaluate the clinical efficacy and safety of topical pefloxacin 0.3% drops as the sole antibiotic used to treat culture positive bacterial corneal ulcers. METHODS: Forty two consecutive Gram's smear-positive cases of bacterial corneal ulcers were enrolled for this prospective open labelled clinical trial. All patients underwent a complete clinical and microbiological work up and were put on topical 0.3% pefloxacin drops with supportive cycloplegic, vitamins and antiglaucoma therapy. Of 42 cases, 4 cases of mycotic keratitis and 6 culture negative cases were excluded from the study. RESULTS: Positive microbiologic cultures were obtained in 84.2% (32 of 38) cases. Staphylococcus aureus (14/32; 43.7%) and coagulase negative Staphylococci (12/32; 37.5%) were the two most common organisms isolated. Resolution of the corneal ulcer was achieved in 31 out of 32 cases (96.9%) with a mean duration of 9.3+/-5.3 days (range 3-21 days). Best corrected visual acuity of 20/200 or better was achieved in 65.6% of cases at 4 weeks post resolution. Corneal deposits were observed in one case which disappeared 8 days following discontinuation of therapy. CONCLUSIONS: Topical pefloxacin is effective as a single antibiotic agent for the treatment of bacterial keratitis.

Adolescent↗
PubMed1998

Morphological and oil content variation in seeds of Azadirachta indica A. Juss. (Neem) from northern and western provenances of India.

Seed morphology (seed length and 20 seed weight) and oil content was studied in Azadirachta indica A. Juss. (Neem) of five provenances of northern and western India. Trees with wide ranges of girths were considered for study. Maximum average oil content was observed in trees from Hisar provenance. Seed oil content in most of the provenances was not consistently and significantly correlated with morphological parameters of seeds. Age of the tree had no significant effect on the oil yield.

Genotype↗
PubMed1998

Coat protein-mediated resistance to pea enation mosaic virus in transgenic Pisum sativum L.

Pea (Pisum sativum L.) plants were transformed in planta by injection/electroporation of axillary meristems with a chimeric pea enation mosaic virus (PEMV) coat protein gene contruct. R1 progenies of these plants were shown to harbor the transgene by polymerase chain reaction (PCR) and genomic Southern analysis, while transgene expression was demonstrated by western blot analysis. Transgenic R2, R3 and R4 plants displayed delayed or transient PEMV multiplication and attenuated symptoms as compared to control inoculated individuals.

Base Sequence↗
PubMed1998

Use of a pharmacokinetic/pharmacodynamic model to design an optimal dose input profile.

A model for the pharmacodynamic effect of a drug (designated only X), and the use of the model to explore optimal input is described. The data analyzed here are from a crossover comparison study of the effect of 4 active treatments, yielding distinct concentration vs. time curves, plus placebo in 32 subjects. The model expresses total effect as the sum of placebo effect and (pure) drug effect. The latter allows for possible tolerance (found) and time effects (not found). Random effects allow interindividual differences to be expressed. Conditioning on the fitted model, a population optimal input profile is designed that obeys certain protocol constraints. The profile minimizes the expectation of a sum of squared differences between target effect and the resulting response over a given time interval. The target is a fixed constant, chosen to be either the individuals' maximum effect level in response to a baseline input regimen used in the study or the maximum effect level for the typical individual in the population in response to this regimen, as predicted from the model. The expectation is taken over the estimated nonparametric distribution of the 32 subjects' random effects. As one goal of early clinical studies of drugs may be to provide a basis for designing an optimal delivery profile (with respect to a specified loss function), we suggest this report as an example of a reasonable way to go about finding such a profile.

Analgesics↗
PubMed1998

Leptin gene therapy and daily protein administration: a comparative study in the ob/ob mouse.

We have compared the efficacy of daily injection of recombinant leptin protein (rh-leptin) with adenovirus-mediated delivery of the murine or human leptin gene (Ad-leptin) for treatment of obesity in the obese (ob/ob) mouse model. We demonstrate an improved correction profile for obesity and associated surrogate markers using the adenovirus delivery method. Rate of weight loss and percentage satiety were significantly greater in the mice treated with Adleptin. These findings were associated with lower peak serum leptin levels with Ad-leptin (22.9 +/- 2.6 ng/ml for the human gene, and 48.9 +/- 11.5 ng/ml for the murine gene) compared to rh-leptin (385.2 +/- 36.0 ng/ml). (Values are given as mean +/- standard error of the mean.) Importantly rh-leptin and ex vivo-expressed Ad-leptin were equivalently active in a functional cell-based assay. The primary difference in the two therapeutic approaches is the continuous chronic secretion of leptin mediated by gene delivery, versus the intermittent bolus delivery and rapid clearance of the daily injection of rh-leptin protein. Thus, in vivo findings suggest that leptin effects are better achieved at lower steady-state levels, a pharmacological feature attained here by gene therapy. These findings may have implications for the potential use of leptin in the treatment of obesity.

Adenoviridae↗
PubMed1998

Cysticercosis of the oral cavity: report of five cases and a review of literature.

This paper reports on five cases of cysticercosis of tongue and buccal mucosa, diagnosed on fine-needle aspiration cytology (FNAC), affecting child patients who presented between January 1994 and October 1997. Four cases presented with gradually increasing nodular swelling of the dorsum of tongue and in the fifth case the swelling was situated on the buccal mucosa of the left side. A clinical diagnosis of cysticercosis was not entertained in any of these patients, who each presented with a solitary lesion; instead, it was considered to be a benign cyst or benign tumour of salivary gland or mesenchymal tissue, before FNAC diagnosis. These lesions of the oral cavity may present first to a dentist and, in endemic areas, cysticercosis should be included in the differential diagnosis of solitary nodular lesions of the oral cavity, particularly in young individuals.

Cheek↗
PubMed1998

Molecular cloning of a novel chemokine receptor-like gene from early stage chick embryos.

Proliferation, differentiation and regulated trafficking of cells are the hallmarks of development and embryogenesis. This led us to speculate a role for chemokines and their receptors in this process. Here, we report the molecular cloning of AvCRL1, a novel member of the G-protein coupled receptor family from early stage 3 days old chick embryos. While the function and ligand for this receptor remain unknown, its sequence and gene structure indicates that it is most related to the family of chemokine receptors, with highest homology to the virally induced human BLR-1 and the CXCR3 or gammaIP-10/Mig-1 receptors.

Amino Acid Sequence↗
PubMed1998

Metabolism of remifentanil during liver transplantation.

We have investigated the pharmacokinetics of remifentanil and its less potent metabolite, GR90291, in six adult patients undergoing orthotopic liver transplantation (OLT). A single bolus infusion of remifentanil 10 micrograms kg-1 min-1 was given at the beginning of the dissection and anhepatic phases of OLT. Remifentanil and GR90291 concentrations were measured in subsequent serial arterial and mixed venous blood samples. Mean arterial clearance of remifentanil was significantly greater (P = 0.02) in the dissection phase (79.54 ml min-1 kg-1) than in the anhepatic phase (39.57 ml min-1 kg-1). Steady state volumes of distribution were not significantly different. Clearance of remifentanil during the anhepatic phase was similar to that of healthy adult patients. Mean maximum concentration (Cpmax) of GR90291 was lower in the dissection phase than in the anhepatic phase (P = 0.026). There was no significant pulmonary metabolism of remifentanil.

Adolescent↗