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Biomedical subjects

M Kapoor

Publications and source records attributed to M Kapoor.

At least 55 records · Page 3Linked to original sources
PubMed1997

Evaluation of various diagnostic techniques for Trypanosoma evansi infections in naturally infected camels.

One hundred and eight camels (Camelus dromedarius) from Trypanosoma evansi endemic areas of the Thar Desert of Rajasthan State, India, were evaluated by various diagnostic tests including parasitological tests (wet blood film-WBF, stained thick blood film), chemical test (mercuric chloride), biological test (mouse subinoculation-MSI), and immunodiagnostic tests based on antibody detection (double immunodiffusion test-DID, card agglutination test-CATT), antigen detection (double antibody sandwich enzyme linked immunosorbent assay-Ag-ELISA). Of the tested camels 49 were found infected using the WBF of which nine gave false negative results with the mercuric chloride test. The efficacy of MSI was 87.03 percent, while the mercuric chloride test was 60.18 percent efficient. The diagnostic efficacy of CATT (72.22 percent) was found to be much better than DID (28.70 percent). Ag-ELISA was 86.11 percent efficient in detecting trypanosomal antigens. A good correlation was found between the positive results obtained by wet blood film, CATT and Ag-ELISA. It was inferred that CATT can be used to study the seroprevalence of T. evansi with great ease, however, trypanosome antigen detection may give a more accurate idea of the prevalence of T. evansi in an endemic area.

Agglutination Tests↗
PubMed1997

Epidemiological profile, mineral metabolic pattern and crystallographic analysis of urolithiasis in Kuwait.

This study was conducted to determine the epidemiological profile of urolithiasis in Kuwaiti patients and the associated metabolic abnormalities favouring stone formation in this patient population. Between 1986 and 1994, a total of 421 Kuwaiti patients were studied in Al-Amiri renal stone laboratory. The mineral composition of stones was determined using a combination of stereoscopic microscopy and infrared spectrophotometry. Mineral metabolic screen (MMS) was available in 306 (72.7%) of those patients. The average annual incidence of new stone formation in Kuwaiti patients was 23.9 per 100,000 population. The incidence was only 6.9 per 100,000 population in those from pediatric age group, 33.4 in adults and 73.6 in the elderly. The frequency of new stone formation was higher among males as compared to females at different age groups. Calcium oxalate (CaO) constituted 72.1% of renal stones encountered in adults as compared to only 52.1% and 40.7% in elderly and pediatric age groups. Eighty four patients had recurrent and/or high stone load. Detailed family history was available in 57 of those 84 patients and was suggestive of strong family aggregation of stone disease in 30 (53%) patients. The proportion of urate stones was 15.4% of all stones formed and constituted a major cause of renal stones in children (24.1%), adults (14.4%) and elderly (12.5%). Cystine stones were found in 10 patients (2.4%). Hyperuricemia was detected in 1/3 of patients from pediatric age group. In adults, 89 (38.4%) of 232 patients had abnormal MMS, of which hyperuricosuria with or without hyperuricemia accounted for 79.8% of those abnormalities. In elderly patients, metabolic disorders were detected in 7 (17.5%) of the 40 patients. In the latter group, all except one, had hyperuricosuria with or without hyperuricemia. Cystinuria, distal renal tubular acidosis (d-RTA) and hyperparathyroidism constituted 6.6% of metabolic abnormalities in the patients studied. Primary abnormalities of upper and lower urinary tract was detected in 6% of adult patients as compared to 47% in children and 30% in the elderly. In conclusion; the incidence rate of new stone former in Kuwait is significantly lower from that reported in Europe and USA. This latter finding and the high familial aggregation of urolithiasis in our area constitute a cogent argument against the role of hot climate in the pathogensis of urolithiasis and are in favour of genetic predisposition. Isolated hyperuricosuria was detected in 17% of idiopathic CaO stone formers. The latter finding is of practical importance since red-meat is a common food item in our area and restriction of purine-intake may help to decrease the frequency of urate and CaO stone formation in predisposed patients.

Adolescent↗
PubMed1996

Mutation of phosphoserine 389 affects p53 function in vivo.

To study the importance of phosphorylation for p53 transactivation function, we generated mutations at each of its known phosphorylated serine amino acids. Mutations of murine p53 serine residues individually to either alanine or glutamic acid at positions 7, 9, 12, 18, 37, 312, and 389 resulted in equivalent levels of transcriptional activation in standard transient transfection experiments. However, when p53 transcriptional activity was measured in cells that attain G1 arrest upon contact inhibition, wild-type p53 was inactive, and only alteration at serine 389 to glutamic acid resulted in a functional p53 protein. This Ser --> Glu mutant also has an increased ability to bind DNA. Elimination of the phosphorylation site by substitution of an alanine amino acid resulted in loss of transcriptional activity. We also demonstrated that specific phosphorylation of p53 at serine 389 is induced by cyclin E overexpression in high-density cells. Our data establish for the first time that phosphorylation of p53 at serine 389 is important in activating its function in vivo.

3T3 Cells↗
PubMed1996

The chloroplast chlL gene of the green alga Chlorella vulgaris C-27 contains a self-splicing group I intron.

The chlL gene product is involved in the light-independent synthesis of chlorophyll in photosynthetic bacteria, green algae and non-flowering plants. The chloroplast genome of Chlorella vulgaris strain C-27 contains the first example of a split chlL gene, which is interrupted by 951 bp group I intron in the coding region. In vitro synthesized pre-mRNA containing the entire intron and parts of the flanking exon sequence is able to efficiently self-splice in vitro in the presence of a divalent and a monovalent cation and GTP, to yield the ligated exons and other splicing intermediates characteristic of self-splicing group I introns. The 5' and 3' splice sites were confirmed by cDNA sequencing and the products of the splicing reaction were characterized by primer extension analysis. The absence of a significant ORF in the long P9 region (522 nt), separating the catalytic core from the 3' splice site, makes this intron different from the other known examples of group I introns. Guanosine-mediated attack at the 3' splice site and the presence of G-exchange reaction sites internal to the intron are some other properties demonstrated for the first time by an intron of a protein-coding plastid gene.

Amino Acid Sequence↗
PubMed1996

Disruption of the NAD(+)-specific glutamate dehydrogenase gene of Neurospora crassa by means of the RIP (repeat-induced point mutations) process.

The structural gene for the catabolite-repressed, substrate-induced NAD(+)-specific glutamate dehydrogenase (gdh-1) of Neurospora crassa was disrupted using the process of repeat-induced point mutation (RIP). Plasmids containing incomplete copies of the gene, along with selectable markers, were introduced into germinated conidia by electroporation. The sexual progeny of a transformant containing an ectopically integrated copy of a plasmid, harbouring the 5' flanking region and a part of the coding sequence of gdh-1 DNA, was examined for the occurrence of RIP by (i) Southern blot analysis of the genomic DNA digested with the isoschizomers MboI and Sau3A, (ii) Northern blot analysis of total RNA in cultures subjected to repression and induction conditions for NAD-GDH, (iii) direct assessment of enzymatic activity, and (iv) evaluation of protein levels by Western blot analysis using a polyclonal anti-GDH IgG preparation. Attempts were made at delineating different regions of the gene exhibiting RIP by using 32P-labelled DNA probes, corresponding to (i) the complete gene, (ii) a fragment containing the 5' flanking region plus two-thirds of the coding sequence, and (iii) the 5' flanking segment alone. The extent and relative location of RIP, as revealed by these hybridization probes, appeared to correlate with changes in specific activity under repression and derepression conditions. Mutant progeny, thus recovered, included isolates with altered regulatory features, such as constitutive expression, inability to elicit derepression, higher-than-wildtype GDH levels under derepression and inefficient repression.

Blotting, Northern↗
PubMed1996

Repeat-induced point mutations of HSP80 gene of Neurospora crassa: methylation of duplicated DNA sequences in the vegetative state.

The process of repeat-induced point mutations (RIP) was used to disrupt the gene encoding the 80-kDa heat-inducible protein of Neurospora crassa. Germinated conidia of the wild-type recipient strain were electrotransformed with a plasmid containing a 7-kb fragment harbouring the complete hsp80 gene sequence. Some of the transformants with a duplication of hsp80 gene sequence showed extensive methylation of these sequences even in vegetatively growing cells. The presence of an extra gene copy in transformants of this type resulted in a marked reduction in the expression of this gene. Progeny of a cross of one such transformant, showing methylation of hsp80, was analyzed by Southern blot and Northern blot hybridization to examine the relationship between methylation and the accumulation of hsp80 mRNA under hyperthermia. In addition, HSP80 polypeptide levels were monitored in stressed and unstressed cells by immunoblot analysis using polyclonal anti-HSP80 IgG preparations. A correlation between the extent of RIP and expression of this gene was observed in the progeny isolates.

Blotting, Northern↗
PubMed1995

Synthesis and characterization of an infectious dengue virus type-2 RNA genome (New Guinea C strain).

Dengue virus type 2 (DEN-2), a member of the Flaviviridae family, has a positive-strand RNA genome, 10,723 nucleotides (nt) in length and encoding a single polyprotein precursor consisting of 3391 amino acids (aa). In order to construct a full-length cDNA clone, the viral genome was cloned into 5' (nt 1-2203 under the control of the T7 promoter (pT7)) and 3' (nt 2203-10,723) constructs. A full-length DEN-2 cDNA under pT7 control was assembled in vitro after excising the two cDNA inserts from the 5' and 3' constructs, and joining them with T4 DNA ligase. The RNA produced by in vitro transcription of the cDNA using T7 RNA polymerase was infectious, as shown by transfection of permissive BHK-21 and Vero cells, and propagation of the virus particles released into the culture media. The virus particles stably maintained the conservative mutation introduced into the 5' construct, and the cells infected with the infectious RNA-derived virus synthesized virus-specific DEN-2 antigens, as shown by immunofluorescence and immunoprecipitations. The full-length infectious clone for DEN-2 should be useful for the study of molecular mechanisms involved in viral RNA replication and virus assembly.

Antigens, Viral↗
PubMed1995

Association between NS3 and NS5 proteins of dengue virus type 2 in the putative RNA replicase is linked to differential phosphorylation of NS5.

Dengue virus type 2, a member of the family Flaviviridae, encodes a single polyprotein precursor consisting of 3391 amino acids residues that is processed to at least 10 mature proteins by host and viral proteases. The NS3 protein contains a domain commonly found in cellular serine proteinases that in cooperation with NS2B is involved in polyprotein processing. In addition, NS3 and NS5 proteins contain conserved motifs found in several RNA helicases and RNA-dependent RNA polymerases, respectively. Both enzymatic activities have been suggested to be involved in viral RNA replication. In this report, we demonstrate that the NS3 and NS5 proteins interact in vivo in dengue virus type 2-infected monkey kidney (CV-1) cells and in HeLa cells coinfected with recombinant vaccinia viruses encoding these proteins as shown by coimmunoprecipitations and immunoblotting methods. We also show by immunofluorescence, metabolic labeling, and two-dimensional peptide mapping that NS5 is a nuclear phosphoprotein and that phosphorylation occurs on serine residues at multiple sites. Furthermore, NS5 exists in differentially phosphorylated states in the nuclear and the cytoplasmic fractions, and only the cytoplasmic form of NS5 is found to coimmunoprecipitate with NS3, suggesting that differential phosphorylation may control the interaction between these proteins and its function in the viral RNA replicase.

Amino Acid Sequence↗
PubMed1995

Sequence repeat-induced disruption of the major heat-inducible HSP70 gene of Neurospora crassa.

The process of repeat-induced point mutation (RIP) was used to disrupt hsps-1, the gene encoding the major heat-inducible member of the HSP70 family of Neurospora crassa. A plasmid DNA, containing an incomplete copy of hsps-1 and the selectable marker qa-2+, was introduced into germinated conidia. The sexual progeny of transformants with ectopically integrated hsps-1 DNA was examined for RIP by Southern-blot analysis of MboI- and Sau3A-digested genomic DNA. Progeny strains, showing RIP, were tested for heat shock-responsive expression of hsps-1, by RNA-blot hybridization and Western-blot analysis, as well as for thermotolerance. Isolates with RIP showed low levels of hsps-1 mRNA and a lack of induction of HSP70 protein by heat shock, accompanied by only a marginal decrease in the acquisition of thermotolerance.

Blotting, Northern↗
PubMed1995

The hsp70 gene family of Neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member.

The gene encoding the major heat shock-inducible member of the HSP70 family of Neurospora crassa was cloned and characterized. The 5' nontranscribed region shows the presence of consensus sequence motifs resembling the classical heat shock elements found in many heat shock-responsive eukaryotic promoters, as well as metal-responsive-element sequences. The coding region of the gene contains four introns with boundaries and internal consensus motifs typical of genes of filamentous fungi. None of the other stress-inducible hsp70 genes of fungal origin have, so far, been reported to contain introns. The sequence adjoining the transcriptional initiation zone shows the presence of prominent CT-rich stretches, characteristic of highly expressed fungal genes. The deduced amino acid sequence corresponds to a 646-residue polypeptide, with a calculated molecular mass of 70,561 Da and an average pI of 6.01, exhibiting strong sequence homology with many other eukaryotic HSP70s, with typical HSP70 family signatures 1 and 2 and a bipartite nuclear targeting sequence. Experiments with primer extension revealed the presence of one minor and two major transcriptional start sites. This gene, designated hsps-1, was mapped to a locus on the left arm of linkage group II, in close proximity to the AR-30 translocation breakpoint.

Amino Acid Sequence↗
PubMed1995

Detection of additional restriction fragment length polymorphisms among the weakly virulent (nonaggressive) and highly virulent (aggressive) isolates of Leptosphaeria maculans.

Isolates of Leptosphaeria maculans were analyzed for their genetic relatedness based on DNA restriction fragment length polymorphisms (RFLPs), employing as Southern hybridization probes a combination of heat shock responsive genes (hsp70 and hsp80 from Neurospora crassa), the cutinase gene of Magnaporthe grisea, and cloned genomic DNA sequences from a virulent strain. Southern hybridization analysis revealed a high frequency of DNA polymorphism. Restriction fragments generated by each enzyme-probe combination resulted in distinct banding patterns, clearly separating the isolates into two groups. The cutinase gene probe did not reveal any polymorphisms. Although the majority of the probes used displayed RFLP profiles unique to each group, a nonaggressive isolate, LmA, showed additional genetic characteristics in common with the virulent pathotype.

Ascomycota↗
PubMed1995

Long-term cyclosporin A treatment in adults with refractory nephrotic syndrome.

Patients with refractory nephrotic syndrome (NS) are at risk of infections, thrombosis, renal failure, or inherent side effects of immunosuppressive therapy. In the present study we investigated the efficacy of cyclosporin A (Cy A) in treatment of adult patients with steroid-refractory NS. Fifteen patients were included, 13 of whom were steroid resistant. Initial renal histology showed minimal-change glomerulonephritis (MCGN) in 3 patients, focal segmental glomerulonephritis (FSGN) in 6 patients, and membranous glomerulonephritis (MGN) in 6 patients. Two steroid-dependent patients (one with MCGN and the other with FSGN) were included due to severe steroid side effects. Complete remission (CR) was achieved in those 2 patients, while in the steroid-resistant patients remission was only partial response (PR) and occurred in about half of the patients in each histological subgroup. In patients who responded to Cy A treatment, two attempts were made to taper off the drug, after 6 and 24 months. Unfortunately, both attempts were unsuccessful and NS relapsed. Except for hypertension in those with advanced renal insufficiency, the drug was well tolerated on long-term usage. A second kidney biopsy was carried out in patients in whom renal failure progressed despite Cy A therapy, and who did not show evidence of Cy A toxicity. These findings are in favor of Cy A for treatment of patients with refractory NS. Treatment with Cy A should be maintained for a minimum period of 3 months before considering its failure. The drug was found to be superior to steroid and conventional immunosuppressive drugs in treatment of steroid-refractory NS; however, the response was partial and was limited to only one-half of the patients in the different subgroups.

Administration, Oral↗
PubMed1993

Identification of the heat shock protein of Neurospora crassa corresponding to the stress-inducible peroxidase.

Heat shock and other stress treatments, resulting in thermotolerance in Neurospora crassa cells, stimulate the induction of a peroxidase at a high level. The putative gene encoding this heat shock-induced peroxidase (hspp) has been cloned, using a cDNA clone of the manganese peroxidase of Phanerochaete chrysosporium, as a probe. Northern blot analysis of total RNA from heat-shocked cell showed the stress-dependent accumulation of a approximately 10 kb transcript. The identity of the HSP, corresponding to the heat-induced peroxidase, was established using hybrid-arrested in vitro translation.

Blotting, Northern↗
PubMed1993

The induction of pyruvate kinase synthesis by heat shock in Xenopus laevis embryos.

Heat-shocked Xenopus embryos have an unusually complex heat shock response. The dominant heat shock protein (Hsp) has a relative molecular mass (M(r)) of 62,000 D (Hsp62). Affinity-purified IgGs against the glycolytic enzyme pyruvate kinase (PK; EC 2.7.1.40) specifically immunoprecipitated Hsp62 from extracts of embryos that had been heat-shocked at 37 degrees C for 30 min. Thus, Hsp62 and pyruvate kinase are immunologically cross-reacting. Electrophoretic separation of PK isoforms suggests that heat-shocked Xenopus embryos increase synthesis of an isoform of PK. Thermal denaturation studies suggest that this isoform has enhanced thermal stability. The identification of PK as an Hsp is discussed within the context of a physiological requirement for elevated levels of anaerobic glycolysis in heat-stressed cells as a vital component of the acquisition of thermotolerance.

Animals↗
PubMed1993

Camel trypanosomosis in Rajasthan, India.

Blood samples from 240 camels (Camelus dromedarius) were examined for trypanosome infection. Of these, 18 (7.50%) were found to be infected using the wet blood Giemsa stain technique, while 76 (31.66%) camels were found to be positive for Trypanosoma evansi antigen using the double antibody sandwich enzyme-linked immunosorbent assay (ELISA). The latter was found to be a more useful method for the detection of current infection.

Animals↗
PubMed1993

Acute renal failure in Kuwait--a prospective study.

This prospective study was conducted over a period of 18 months (February 1989 to July 1990) in the State of Kuwait. It covered a population of 1,024,211 and eight multidisciplinary hospitals with an in-patient admission of 118,079 per year. Two hundred and twenty-six adult patients with acute renal failure (ARF) were seen and followed up by nephrologists. This made the calculated annual incidence of ARF 14.7 per 100,000 population, nearly five times that reported by the EDTA registry (Biesenbach et al. 1991). Drugs, sepsis and volume depletion were the most frequent causes, with sepsis resulting in 36% cause specific mortality compared to zero mortality with the other two. The overall mortality rate was only 14% which clearly indicated a markedly improved prognosis in cases of ARF. The prognosis in ARF depended on two major factors, viz. the type of aetiological insult and the presence of predisposing associated medical illnesses. Multiple insults, though common, do not affect the mortality rate. Secondary sepsis or gastrointestinal bleeding as a cause of death in ARF was rarely seen in our study. Those who required dialytic support for renal failure had a 45% patient mortality rate in general. Over 40% of our patients were 60 years or older compared with only 3.5% in the local population. This indicated old age as a major risk factor in the development of ARF. The overall mortality in the elderly did not differ from that in the young, but sepsis in the elderly carried a mortality rate of 60% compared to only 14.8% in the younger age group.

Acute Kidney Injury↗