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Biomedical subjects

M Sharma

Publications and source records attributed to M Sharma.

At least 127 records · Page 7Linked to original sources
PubMed1994

Fluorescence postlabeling assay of RNA modification.

A new approach has been developed to assay RNA modification by combining enzymatic digestion of RNA to nucleoside monophosphate and fluorescence postlabeling. Nuclease P1 digestion of modified RNA affords both normal and modified nucleotides. The nucleotides are labeled in-situ with dansyl chloride via phosphoramidate derivatives with ethylenediamine following a procedure initially designed to synthesize fluorescence labeled deoxynucleotides. The postlabeled nucleotides are analyzed by HPLC using a fluorescence detector. Fluorescence postlabeling assay of 7-methylGmp in RNA exposed to dimethyl sulfate has validated the technique. In the presence of excess normal nucleotides, the limit of detection (LOD) lies between normal to modified nucleotide ratio of 10(3) to 10(4). HPLC enrichment of the modified nucleotide from the normal nucleotides prior to labeling enhances the LOD by two orders of magnitude.

Chromatography, High Pressure Liquid↗
PubMed1994

The scope of dansyl vs fluorescein label in fluorescence postlabeling assay for DNA damage.

The 5'-phosphate groups of the normal nucleotides of DNA (10 nmol) were labeled with two different fluorescent reagents, namely, dansyl chloride and fluorescein-isothiocyanate, via phosphoramidate derivatives of ethylene and hexamethylenediamines. Using a conventional detector, HPLC analysis showed a linear relationship of fluorescence signal as an integrated peak area and the amounts of nucleotide injected from 2 pmol to 200 fmol (r = 0.996, n = 3) for dansyl-labeled dAmp and from 2 pmol to 90 fmol (r = 0.997) for fluorescein-labeled dAmp, respectively. Although fluorescein-labeled nucleotides are more sensitive than dansylated nucleotides (detection limit < 2 times, S/N = 3), the excess labeling reagent from the fluorescein labeling reaction interferes with HPLC resolution of the labeled nucleotides. The phosphoramidate derivatives of the nucleotides require purification prior to labeling with fluorescein isothiocyanate. As a result, the overall assay procedure becomes complicated. The potential of dansylated nucleotides to assay DNA modifications has been demonstrated by HPLC resolution of the labeled nucleotides of both normal and modified bases without further manipulation of the reaction intermediates.

Chromatography, High Pressure Liquid↗
PubMed1994

Use of micro samples of finger prick blood dried on filter paper for a quick and simple dipstick dot-EIA for diagnosis of amebic liver abscess (ALA).

Filter paper was used as a support to absorb micro samples of finger prick blood for detection of antibodies to Entamoeba histolytica (causative organism of amebiasis) by a rapid dipstick dot-EIA technique; 8 microliters of blood was sufficient to saturate discs (diameter 6 mm) of Whatman paper No. 3. Conditions of elution of blood from the discs were optimized and the best results were obtained when 0.4 ml of buffer was used for elution for 30 minutes at room temperature. The filter paper technique is extremely useful for field use and for diagnosis of amebic liver abscess on a large scale since it does not involve centrifugation of blood and use of sterile vials because blood dried on paper can be stored in polythene bags at room temperature for up to 3 months and for a week at 42 degrees C without significant loss of antibody activity, thereby eliminating the need of refrigeration of samples for storage or transportation to a reference laboratory. Elution of blood from different discs of the same patient was found to be highly reproducible without appreciable loss of sensitivity or specificity. Twenty-four confirmed ALA, 29 non-ALA, and 25 apparently normal healthy controls, with no previous history of amebiasis, were tested. Sensitivity and specificity of the test was found to be 96% and 92%, respectively.

Animals↗
PubMed1994

Androgen treatment decreases estrogen receptor binding in the ventromedial nucleus of the rat brain: a quantitative in vitro autoradiographic analysis.

Androgens oppose the actions of estrogen on a number of neuroendocrine functions in the rat including prolactin and gonadotropin secretion and the activation of the female pattern of sex behavior. Although in nonneural tissues antiestrogenic actions of androgens have been related to actions at the level of the estrogen receptor, previous attempts to demonstrate effects of nonaromatizable androgens on estrogen receptor levels in the brain have been unsuccessful, possibly because of the poor anatomical resolution of the methods used. We have used a new in vitro autoradiographic assay combined with an 125I-labeled estrogen receptor ligand to test the hypothesis that the nonaromatizable androgen, 5 alpha-dihydrotestosterone (5 alpha-DHT), may act to reduce estrogen binding in specific regions of the brain involved in reproductive neuroendocrine and behavioral responses. This in vitro autoradiographic method allows selective measurement of occupied estrogen receptors in tissue sections. Gonadectomized/adrenalectomized rats were divided into two groups per sex. All animals received daily injections of estradiol benzoate (EB: 40 micrograms/kg body wt) for 4 days. Animals in the 5 alpha-DHT treatment group received 5 alpha-DHT (10 mg/kg body wt) every 12 h for 4 days, while animals in the control group received vehicle injections. Animals were killed 4 h after the final EB/5 alpha-DHT injection and their brains processed for in vitro autoradiography. As previously reported, higher levels of estrogen binding were observed in the ventrolateral aspect of the ventromedial nucleus (vIVMN) and the periventricular and medial preoptic area of the female compared to the male.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenalectomy↗
PubMed1994

Nicardipine and verapamil attenuate the pressor response to laryngoscopy and intubation.

In a prospective, double-blind study, we compared the efficacy of iv nicardipine hydrochloride and verapamil hydrochloride in attenuating the cardiovascular responses to laryngoscopy and tracheal intubation, in 45 patients undergoing elective surgery with general anaesthesia. Patients were allocated randomly to one of three groups of 15 patients. Patients in Group I received saline while those in Groups II and III received nicardipine hydrochloride, 0.03 mg.kg-1 or verapamil hydrochloride, 0.1 mg.kg-1 iv three minutes before laryngoscopy and intubation. Patients in Group I showed the greatest increase in SBP 25.4 +/- 2.2 2.2 mmHg and HR 35.7 +/- 3.8 beats.min-1 at one minute after intubation (P < 0.001), and these changes persisted throughout the study period albeit with decreasing magnitude. After drug administration, patients in Groups II and III demonstrated increases in HR of 26 +/- 2.4 and 15.1 +/- 2.2 beats.min-1 and decreases in SBP of 24.8 +/- 2.0 and 18.8 +/- 2.4 mmHg respectively (P < 0.001). It is concluded that nicardipine and verapamil are effective in attenuating pressor responses to laryngoscopy and intubation but did not control the tachycardia.

Adult↗
PubMed1994

Epidemiological determinants of burns in paediatric and adolescent patients from a centre in western India.

A 1-year prospective study of 112 burn patients up to 19 years of age aimed to identify and study the determinants and mortality experiences in these burn patients and found flame burns to be the commonest, followed by scalds and electric burns. All burns other than those caused by electricity were commoner in females, more so between 15 and 19 years of age. Burns were more frequent in winter and 85 per cent of them were domestic. Nearly all burns took place during day time with a higher incidence between 06.00-09.00 h and 14.00-21.00 h. The patient fatality rate (41.1 per cent) was associated with total burn surface area. Referral time-lag was an important determinant of mortality especially in less severe burns. As would be expected, hospital stay was significantly longer in survivors.

Accidents↗
PubMed1994

Neuroimaging of meningeal disease.

In the past two decades, the advent of CT and MRI has made a considerable impact on the evaluation of meningeal diseases, conditions once regulated to cytological, histopathological, or postmortem analyses alone. This article reviews the imaging findings in various meningeal processes with particular attention to the anatomic definition of the meningeal layers and their relationship to the development of meningeal pathology and consequent imaging characteristics.

Central Nervous System Diseases↗
PubMed1994

Characterization of monoclonal antibodies to conserved antigens of Entamoeba histolytica and E. dispar and development of a stool ELISA.

Eight murine monoclonal antibodies (MAbs) were generated against Entamoeba histolytica NIH:200. All MAbs reacted with three axenic strains of E. histolytica, NIH:200, HM-1: IMMS, and SAW 1734R clAR, but not with enteric bacteria or Giardia lamblia. Five of the MAbs reacted with low-molecular-weight, periodate-sensitive antigens of 14-21 kD, while one (AB31) reacted with high-molecular-weight, 90 to 200-kD protein determinants. MAb PC14 appeared to be specific for antigen in its native state. Another MAb (BB12) agglutinated live trophozoites and caused membrane fluorescence in contrast to the five other MAbs tested. Although BB12 reacted with the same 14 to 21-kD band on Western blot as AC55, the latter reacted with different cytoplasmic epitopes. All the MAbs reacted to five pathogenic (E. histolytica) and six nonpathogenic (E. dispar) clinical isolates. These MAbs may be helpful for studying conserved antigens of E. histolytica and were used to develop a sandwich ELISA for the diagnosis of intestinal amoebiasis. The assay was sensitive to 60 ng of E. histolytica antigen. A comparative study of microscopic examination of stool samples and the sandwich ELISA was conducted on 102 samples from patients with gastrointestinal complaints. The ELISA could detect all microscopically positive samples with a sensitivity of 100% and specificity of 93%. A sandwich ELISA using a monoclonal antibody to conserved antigens of E. histolytica has the potential to be a reliable method for the diagnosis of intestinal amoebiasis.

Agglutination Tests↗
PubMed1994

Primary radiation therapy to T1 and T2 breast cancer following conservative surgery. Which patients should be boosted?

We reviewed 199 radiated patients at our institution (201 breasts treated) and its affiliates treated between 1978 and 1989. Of these, 157 were T1 and T2 invasive breast carcinoma. Our intent was to retrospectively compare the results of those who received standard doses of 4,500 to 5,000 cGy to the breast to those that received an additional boost to the surgical bed to a dose totaling at least 5,500 cGy. There were a total of 5 local recurrences in 159 treated breasts. (The mean follow-up time was 36 months.) Of our T1 and T2 patients with clear resection margins that were boosted, there was 1 local recurrence in 28 treated breasts. There was 1 local recurrence in the nonboosted group of 68 patients. Except for one patient, all patients with positive margins were boosted. There were 2 local recurrences in the 23 T1 and T2 breasts with positive margins that were boosted. Of the patients with uncertain margins who were not boosted, there was one local recurrence in 20 treated breasts. Of those with uncertain margins that were boosted, there were no local recurrences in 19 treated breasts. From our results, it would appear that a boost to the primary site is unnecessary if the margins of resection are negative (by either inking or if it is clearly stated in the pathology report). In those patients with uncertain margins, most were done in the years before margins were routinely inked, but generous excisional biopsies were usually done. In this latter group of patients, there also was no added benefit to boosting.

Adult↗
PubMed1994

Purification and characterization of the SegA protein of bacteriophage T4, an endonuclease related to proteins encoded by group I introns.

Although not encoded by an intron, the bacteriophage T4 SegA protein shares common amino acid motifs with a family of proteins found within mobile group I introns present in fungi and phage. Each of these intron-encoded proteins is thought to initiate the homing of its own intron by cleaving the intronless DNA at or near the site of insertion. Previously, we have found that SegA also cleaves DNA. In this report, we have purified the SegA protein and characterized this endonuclease activity extensively. SegA protein cleaved circular and linear plasmids, DNA containing unmodified cytosines, and wild-type T4 DNA containing hydroxymethylated, glucosylated cytosines. In all cases, certain sites on the DNA were highly preferred for cleavage, but with increasing protein concentration or time of incubation, cleavage occurred at many sites. SegA cleaving activity was stimulated by the presence of ATP or ATP gamma S. Sequence analysis of three highly preferred cleavage sites did not reveal a simple consensus sequence, suggesting that even among highly preferred sites, SegA tolerates many different sequences. A T4 segA amber mutant that we constructed had no phenotype, and PCR analyses indicated that several T-even-related phages lack the segA gene. Taken together, our results show that SegA is an endonuclease with a hierarchy of site specificity, and these results are consistent with the insertion of segA DNA into the T4 genome some time after the divergence of the closely consistent with the insertion of segA DNA into the T4 genome some time after the divergence of the closely related T-even phages.

Amino Acid Sequence↗
PubMed1994

MC540 induced photosensitization of glioma & neuroblastoma cells.

Binding and photodynamic action of merocyanine 540 (MC540) has been studied in glioma (U-87MG) and neuroblastoma (Neuro 2A) cells as a function of dye concentration, incubation time of cells with MC540 and growth phase of cells. In the plateau phase, U-87MG cells accumulated more MC540 as compared to exponentially growing cells, whereas in Neuro 2A cells the opposite effect was observed. Exponentially growing U-87MG cells were more photosensitive than plateau phase cells. However, the photosensitivity of Neuro 2A cells was not dependent on the growth phase. Thus, MC540 mediated photosensitization may be useful for photodynamic therapy of brain tumours.

Fluorescent Dyes↗
PubMed1994

A quantitative microbiological study of bacterial vaginosis.

A quantitative microbial study of vaginal secretions from 100 women with bacterial vaginosis (BV) and 50 healthy women matched for age, was undertaken. This was aimed at comparing the bacterial flora colonising the vagina of sexually active asymptomatic women with those with BV and correlating bacterial counts with presence of symptoms. Of the total 475 bacteria isolated anaerobes were predominant among controls and among patients aerobes and anaerobes had equal prevalences. The increased viable counts of all anaerobes (> 10(7) cfu/g) were related with symptoms. Mobiluncus mulieris could be isolated from one patient and Chlamydia trachomatis antigen was absent in all samples tested. Bacteroides, G. vaginalis and Ureaplasma urealyticum were found to be the most commonly occurring group in women with BV. Lactobacillus, and probably Coryneforms have a protective role in health, preventing disease.

Adolescent↗
PubMed1994

Effect of D-penicillamine on lymphocyte subsets: correlation with clinical response in rheumatoid arthritis.

We studied the effects of D-penicillamine (DP) on the clinical response, immunoinflammatory parameters and the lymphocyte subsets in 46 patients with rheumatoid arthritis (RA). Patients were evaluated before the start of the drug and then at 3 and 9 months during the follow up. 38 of 46 (82.6%) patients could continue DP treatment for over 9 months, while in 8 the drug was withdrawn due to adverse effects. Improvement in the various disease activity indices of more than 50% (responders) was seen in 25 of 38 (65.8%) patients. Responders showed a significant decrease in the serum IgA and IgM at 9 months, and in IgM only at 3 months. The serum levels of C3 and C4 did not show any significant change. Serum levels of C-reactive protein and rheumatoid factor (RF) showed a significant decrease at 3 and 9 months. A significant decrease in CD3+ and CD4+ lymphocytes along with a fall in CD4+/CD8+ lymphocyte ratio was also seen in responders at 3 and 9 months, compared to the baseline. Our results suggest that DP may have immunomodulatory action in RA.

Adolescent↗
PubMed1994

Mode, presentation, CT findings and outcome of pediatric head injury.

Twenty five to thirty per cent of all patients admitted with head injuries to any large hospital are children. Head injuries in children differ in several ways from those seen in the adult population. A standard management protocol is used in a service hospital and CT and neurosurgical consultation/transfer is based on specific criteria. The records of 312 patients admitted with head injury to Command Hospital (WC), Chandimandir were retrospectively analysed to compare the mode of injury, initial presentation, CT findings and the outcome of management between those above and those below the age of 10 years. Eighty seven (27.8%) of the 312 patients were children and 71% of them had sustained the injury due to fall from a height. One third of children were brought with vomiting and drowsiness and 9.2% with seizures, but lateralizing signs were found in only 5.7%. Nineteen children underwent a CT of the head and of these 74% had only cerebral edema. Only 4 of the 87 children were transferred to a neurosurgical unit. The mortality rate was lower in children (5.7%) compared to the older age group (11.6%) but the difference was not statistically significant. However, a significantly higher number (80.5%) of children recovered without any residual deficit. Majority of our children can be safely managed in a general surgical unit provided a multidisciplinary approach is used and facilities of CT are readily available, since very few require active neurosurgical intervention.

Accidental Falls↗
PubMed1994

Phytotoxicological evaluation of Tagetes erectes on aquatic stages of Anopheles stephensi.

Petroleum ether (5%) in ethyl acetate fraction of Tagetes erectes showed toxic activity against second and fourth instar larvae of Anopheles stephensi below 100 ppm concentrations. The LC50 values for second and fourth instar larvae were calculated to be 43 and 58 ppm respectively. The second instar larvae were more susceptible than the fourth instar larvae. The extract did not affect the larval developmental period. The growth index of treated mosquitoes was significant in comparison with that of the control and untreated sets (p < 0.01). Treatment with the extract had a significant effect on the mortality and reduction in the adult emergence of the vector.

Animals↗
PubMed1994

Effects of carbon dioxide-induced anesthesia on cholinergic parameters in rat brain.

We report that acetylcholinesterase (AChE) and choline acetyltransferase (ChAT) activities in rat brain were virtually identical whether the rat was anesthetized with carbon dioxide (CO2) before decapitation or decapitated without prior sedation. The AChE and ChAT activities were measured in three brain regions: the hippocampus, cerebral cortex, and cerebellum. Enzyme activities varied significantly by brain region, with the highest values in the hippocampus and the lowest values in the cerebellum. Enzyme activities, however, did not vary with the method of euthanasia, either CO2-induced anesthesia prior to decapitation or decapitation without anesthesia. These data suggest that CO2-induced anesthesia prior to decapitation does not alter activities of these cholinergic markers in rat hippocampus, cerebral cortex, and cerebellum. This method of euthanasia eliminates the need to capture a conscious animal, which reduces stress to the animal and the experimenter.

Acetylcholinesterase↗
PubMed1994

Differential effects on spatial navigation of immunotoxin-induced cholinergic lesions of the medial septal area and nucleus basalis magnocellularis.

The effects on anatomy and behavior of a ribosomal inactivating protein (saporin) coupled to a monoclonal antibody against the low-affinity NGF receptor (NGFr) were examined. In adult rats, NGFr is expressed predominantly in cholinergic neurons of the medial septal area (MSA), diagonal band nuclei, and nucleus basalis magnocellularis (nBM), but also in noncholinergic cerebellar Purkinje cells. Rats with immunotoxin injections to the MSA, nBM, and lateral ventricle were compared to controls on a spatial and cued reference memory task in the Morris maze. Toxin injections to the MSA slightly impaired the initial, but not asymptotic, phase of spatial navigation. Injections to the nBM impaired all phases of spatial navigation. Cued navigation, however, was not affected in either the MSA or nBM group. The ventricular injections severely affected spatial and cued navigation. Acetylcholinesterase (AChE) histochemistry and NGFr and choline acetyltransferase immunohistochemistry revealed a loss of (1) almost all NGFr-positive cholinergic neurons in the MSA and AChE fibers in hippocampus (MSA group); (2) almost all NGFr neurons in the nBM, some in the MSA, most AChE fibers in neocortex and some in the hippocampus (nBM group), and (3) almost all NGFr neurons in the MSA and nBM and their corresponding hippocampal and cortical AChE fibers (ventricular group). Cholinergic nBM projections to the amygdala were largely preserved in all groups. The amount of cholinergic fiber loss in the cortex correlated modestly, but significantly, with the severity of impairment of the asymptotic phase of performance of the spatial task. An unambiguous interpretation of the anatomical locus of behavioral deficits was not possible because of damage to cholinergic striatal interneurons (nBM group) and to noncholinergic cerebellar Purkinje cells (ventricular group). These data suggest that the cholinergic cortical system is critical to the performance of this spatial memory task. Cholinergic denervation of the hippocampus alone, however, is not sufficient to impair markedly performance of this task.

Animals↗