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Biomedical subjects

A A Kumar

Publications and source records attributed to A A Kumar.

At least 37 records · Page 2Linked to original sources
PubMed1995

Genotoxicity testing of two red dyes in the somatic and germ line cells of Drosophila.

Two red dyes, rhodamine B and amaranth, were tested for their genotoxic effects in the somatic (wing primordia) and germ line cells of Drosophila melanogaster following the wing spot and the sex-linked recessive lethal tests. Second- and third-instar larvae, carrying suitable genetic markers, were subjected to chronic exposure to different concentrations of the test dyes. The results indicate that rhodamine is genotoxic in both somatic and germ line cells and amaranth is non-genotoxic.

Abnormalities, Drug-Induced↗
PubMed1993

Genotoxicity of 2,4-dichlorophenoxyacetic acid tested in somatic and germ-line cells of Drosophila.

The genotoxic potential of 2,4-dichlorophenoxyacetic acid, a commonly used chlorophenoxy herbicide, was tested in Drosophila somatic and germ-line cells following the protocols of the wing spot test and the sex-linked recessive lethal test. In the wing spot test second- and third-instar larvae, carrying genetic markers mwh and flr3, were exposed to different concentrations of the herbicide so that induced genetic changes would be phenotypically expressed as mosaic spots on the wings of eclosing adults. The Basc (Muller-5) standard technique but with larval exposure was followed for the sex-linked recessive lethal test. The results obtained indicate that the test compound is genotoxic both in the somatic and germ-line cells of Drosophila.

2,4-Dichlorophenoxyacetic Acid↗
PubMed1991

Expression of recombinant human plasminogen in mammalian cells is augmented by suppression of plasmin activity.

We present evidence that over-expression of human plasminogen, the precursor to the serine protease plasmin, can be cytotoxic to mammalian cells. When an expression vector containing plasminogen cDNA is transfected into baby hamster kidney cells, the number of drug-resistant colonies as well as the levels of plasminogen secreted by those colonies is lower than observed in similar transfections of other protease precursor genes. The recombinant plasminogen accumulates intracellularly as degraded NH2-terminal fragments. In contrast, a mutant of plasminogen that produces inactive plasmin (active site Ser740 changed to Ala) is synthesized by these cells as a full-length plasminogen molecule, and the colony numbers and expression levels are normal. Thus, the generation of plasmin activity is responsible for the cytotoxic phenomena and the degradation associated with plasminogen expression. In addition, experiments using a plasminogen mutant that cannot be activated to plasmin (activation cleavage site Arg560 to Gly) or using coexpression of antisense urokinase RNA indicate that an endogenous plasminogen activator is responsible for converting newly synthesized plasminogen to plasmin. Finally, coexpression of plasminogen with alpha 2-plasmin inhibitor, a serpin which is the physiologic inhibitor of plasmin, prevents the toxic effects of intracellular plasmin activity and allows the synthesis and secretion of native human plasminogen.

Animals↗
PubMed1991

Endoproteolytic processing of the human protein C precursor by the yeast Kex2 endopeptidase coexpressed in mammalian cells.

The human protein C precursor undergoes extensive co- and posttranslational modification during its biosynthesis in the liver. These modifications include glycosylation, gamma-carboxylation, and beta-hydroxylation of specific amino acids and endoproteolytic processing to remove the pre- and propeptides as well as the pair of basic amino acids which connect the light and heavy chains in the precursor. Previous studies with a recombinant mammalian expression system have indicated that the endopeptidase in several mammalian cell types which recognizes and cleaves this dibasic site has a substrate specificity for sites which also include a basic amino acid in the -4 position (Foster et al., 1990). Since the human protein C precursor has His154 in the -4 position, it is poorly and incompletely cleaved in BHK and several other mammalian cell lines and also apparently secreted from the liver as a mixed population of mature two-chain and precursor one-chain molecules. In the present study, a mammalian expression system has been used to study the effect of coexpressing the protein C precursor together with the yeast Kex2 endopeptidase which is known to recognize and process dibasic pairs within peptide precursors in yeast. Coexpression of the KEX2 gene resulted in complete conversion of the protein C precursor to the mature two-chain form. Amino-terminal sequencing of the cleavage products has indicated that the cleavage occurs in the correct location and that this site is preferentially recognized by the yeast endopeptidase within the context of the mammalian cell secretory pathway.

Amino Acid Sequence↗
PubMed1991

Stereotactic brain biopsy in the diagnosis of malignant lymphoma.

Fifteen patients with cerebral involvement by malignant non-Hodgkin's lymphoma were identified, among more than 200 patients who underwent stereotactic biopsy at The Johns Hopkins Hospital. All but one of these cases were diagnosed accurately by the stereotactic biopsy procedure. In 12 of 14 patients, the material was adequate to classify the lymphoma according to the Working Formulation. Because all but one of the lesions were intermediate or high-grade neoplasms, a diagnosis of lymphoma was often possible by conventional light microscopic examination alone. Monotypic light chain expression was demonstrated by immunohistochemical techniques in six patients, and positivity for B-cell markers was observed in an additional case. In one instance, two stereotactic biopsy specimens were interpreted as being suggestive of lymphoma, but necrosis and inflammation prevented a definitive diagnosis. Nine patients had no known risk factors for cerebral lymphoma, and the diagnosis often was unsuspected clinically.

Adolescent↗
PubMed1990

Endoproteolytic processing of the dibasic cleavage site in the human protein C precursor in transfected mammalian cells: effects of sequence alterations on efficiency of cleavage.

The human protein C precursor undergoes extensive co- and posttranslational modification during its biosynthesis in the liver. These modifications include glycosylation, gamma-carboxylation and beta-hydroxylation of specific amino acids, and endoproteolytic processing to remove the pre- and propeptides and also to remove the pair of basic amino acids that connect the light and heavy chains in the precursor. Specific molecular signals have been elucidated which direct several of these modifications; however, the mechanism for cleavage and removal of the basic amino acid pair has not been established. In the present study, a recombinant mammalian expression system has been used to study the molecular signals that direct removal of this basic amino acid pair. Mutations were introduced by site-directed mutagenesis either to insert additional basic amino acids or to alter the sequence adjacent to the basic pair by point mutations. The mutant protein precursors were expressed and analyzed for the degree of processing to 2-chain form and also for the location of the cleavage site (by N-terminal sequencing) and subsequent removal of the basic amino acids from the newly formed C terminus of the light chain. These experiments have shown that human protein C can be readily synthesized and secreted in several mammalian cell lines. However, cell lines vary considerably in their capacity to remove the dibasic pair in the protein C precursor and, like the liver, secrete a mixed population of 1-chain and 2-chain forms of the protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1990

Artificial intelligence for medical decision making.

Artificial intelligence techniques find extensive applications in medical decision making and other aspects of health care. A number of successful expert systems have been developed in various disciplines of medicine. This paper gives an overview of expert system techniques, describes some practical systems, and discusses the relevance of such systems in clinical diagnosis and management of diseases.

Decision Making, Computer-Assisted↗
PubMed1989

Intraoperative CT-guided serial microbiopsy of brain lesions and concomitant use of CT air and angiographic studies.

The value of step-by-step confirmation of the location of the brain biopsy probe is demonstrated, using serial CT scanning to confirm the exact source of the sample tissue. Concomitant use of CT angiography and a CT air study makes possible the biopsy of small areas and of areas near major cerebral vessels, as well as of lesions near critical areas like the optic chiasma, basal ganglia, midbrain, and other complex lesions. The pathologist on-line examination of the CT-confirmed serial samples enables the surgeon to obtain the most informative tissue for the ultimate histopathological diagnosis. Our experience with the technique is illustrated by examples of its application in some complex cases.

Biopsy↗
PubMed1987

Propeptide of human protein C is necessary for gamma-carboxylation.

Protein C is one of a family of vitamin K dependent proteins, including blood coagulation factors and bone proteins, that contains gamma-carboxyglutamic acid. Sequence analysis of the cDNAs for these proteins has revealed the presence of a prepro leader sequence that contains a pre sequence or hydrophobic signal sequence and a propeptide containing a number of highly conserved amino acids. The pre region is removed from the growing polypeptide chain by signal peptidase, while the pro region is subsequently removed from the protein prior to secretion. In the present study, deletion mutants have been constructed in the propeptide region of the cDNA for human protein C, and the cDNAs were then expressed in mammalian cell culture. These deletions included the removal of 4, 9, 12, 15, 16, or 17 amino acids comprising the carboxyl end of the leader sequence of 42 amino acids. The mutant proteins were then examined by Western blotting, barium citrate adsorption and precipitation, amino acid sequence analysis, and biological activity and compared with the native protein present in normal plasma. These experiments have shown that protein C is readily synthesized in mammalian cell cultures, processed, and secreted as a two-chain molecule with biological activity. Furthermore, the pre portion or signal sequence in human protein C is 18 amino acids in length, and the pro portion of the leader sequence is 24 amino acids in length. Also, during biosynthesis and secretion, the amino-terminal region of the propeptide (residues from about -12 through -17) is important for gamma-carboxylation of protein C, while the present data and those of others indicate that the carboxyl-terminal portion of the propeptide (residues -1 through -4) is important for the removal of the pro leader sequence by proteolytic processing.

Amino Acid Sequence↗
PubMed1987

Sequence of the Ampullariella sp. strain 3876 gene coding for xylose isomerase.

The nucleotide sequence of the gene coding for xylose isomerase from Ampullariella sp. strain 3876, a gram-positive bacterium, has been determined. A clone of a fragment of strain 3876 DNA coding for a xylose isomerase activity was identified by its ability to complement a xylose isomerase-defective Escherichia coli strain. One such complementation positive fragment, 2,922 nucleotides in length, was sequenced in its entirety. There are two open reading frames 1,182 and 1,242 nucleotides in length, on opposite strands of this fragment, each of which could code for a protein the expected size of xylose isomerase. The 1,182-nucleotide open reading frame was identified as the coding sequence for the protein from the sequence analysis of the amino-terminal region and selected internal peptides. The gene initiates with GTG and has a high guanine and cytosine content (70%) and an exceptionally strong preference (97%) for guanine or cytosine in the third position of the codons. The gene codes for a 43,210-dalton polypeptide composed of 393 amino acids. The xylose isomerase from Ampullariella sp. strain 3876 is similar in size to other bacterial xylose isomerases and has limited amino acid sequence homology to the available sequences from E. coli, Bacillus subtilis, and Streptomyces violaceus-ruber. In all cases yet studied, the bacterial gene for xylulose kinase is downstream from the gene for xylose isomerase. We present evidence suggesting that in Ampullariella sp. strain 3876 these genes are similarly arranged.

Actinomycetales↗
PubMed1984

Bone marrow transplant in adrenoleukodystrophy.

An allogeneic bone marrow transplant (BMT) from a normal HLA identical sibling donor was performed in a 13-year-old boy with rapidly progressive adrenoleukodystrophy (ALD). Engraftment and complete hematologic recovery occurred within 4 weeks, but neurologic deterioration continued. The patient died of an adenovirus infection 141 days after BMT. ALD is characterized by abnormally high plasma levels of very long chain fatty acids (VLCFA) as a result of impaired capacity to degrade them. Ten days after BMT, the white blood cell VLCFA levels and enzyme activity became normal; after 3 months, there was progressive reduction of plasma VLCFA to levels only slightly above normal.

Adolescent↗
PubMed1983

Fluorescent analogues of methotrexate: characterization and interaction with dihydrofolate reductase.

The dansylated derivatives of lysine and ornithine analogues of methotrexate exhibit fluorescence properties characteristic of the dansyl moiety with an excitation at 328 nm and an emission maximum at 580 nm in aqueous media. As in the case of dansyl amino acids, the fluorescence emission is dependent upon the polarity of the medium. In solvents of low dielectric constant there is an enhancement of the dansyl fluorescence intensity as well as a shift to shorter wavelengths. The dansylated analogues show a reduction in the quantum yields as compared to N epsilon-dansyl-L-lysine and 5-(N,N-dimethylamino)-1-naphthalenesulfonic acid. The absorption spectra of the two dansyl analogues are similar to the spectra of the parent basic amino acid precursors but with reduced molar extinction values. The two fluorescent analogues of methotrexate were found to be potent inhibitors of purified dihydrofolate reductases from Lactobacillus casei and from chicken liver. The binding of these fluorescent analogues to either dihydrofolate reductase resulted in 10-15-nm blue shift of the ligand emission maxima and a 2-5-fold enhancement of the emission. These fluorescent properties of the bound ligands indicate a possible interaction of the dansyl moiety with a region on the enzyme molecule which is more hydrophobic relative to the surrounding solvent.

Animals↗
PubMed1983

High-performance liquid chromatography of methotrexate analogs containing terminal lysine or ornithine and their dansyl derivatives.

A procedure utilizing a reverse-phase semipreparative high-performance liquid chromatography column and a binary solvent system consisting of trifluoroacetic acid and 1-propanol has been developed for the semipreparative scale purification and analytical identification of four newly synthesized analogs of methotrexate. The methotrexate analogs containing a lysine or an ornithine residue in place of a terminal glutamate residue together with their respective dansyl derivatives were purified in milligram quantities by the procedures described.

Chromatography, High Pressure Liquid↗