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Biomedical subjects

A K Verma

Publications and source records attributed to A K Verma.

At least 91 records · Page 5Linked to original sources
PubMed1991

Involvement of retinoic acid nuclear receptors in retinoic acid-induced tissue transglutaminase gene expression in rat tracheal 2C5 cells.

The involvement of retinoic acid nuclear receptors (RARs) in the induction of tissue transglutaminase (TG) by retinoic acid in rat tracheal 2C5 cells was determined. The levels of RAR alpha and RAR beta were altered in 2C5 cells by transfection with RAR expression vectors. Increased expression of RAR alpha increased the induction of tissue TG by retinoic acid. In contrast, decreased RAR alpha expression, using an antisense RAR alpha expression vector, diminished the normal level of tissue TG induction caused by retinoic acid. Transfectants overexpressing RAR beta were also more responsive to retinoic acid for the induction of tissue TG, although the magnitude of TG induction was not as great as resulted from RAR alpha overexpression. These results indicate that the levels of the RAR alpha and RAR beta dictate the magnitude of tissue TG induction by retinoic acid.

Animals↗
PubMed1991

Novel heteroarotinoids: synthesis and biological activity.

In this study, 13 heteroarotinoids were synthesized. The key step in each preparation was the condensation of the appropriate chroman-, thiochroman-, or benzothienyl-substituted phosphorus ylide, obtained from the independent synthesis of the corresponding phosphonium salts, with selected polyene-substituted aldehyde esters. Nine of these heterocycles contained a thiochroman group, two had a chroman group, and two others had a benzothienyl system. Screening of the compounds was with one of two assays. One assay measured the ability of a retinoid to inhibit the phorbol ester induced increase of mouse epidermal ornithine decarboxylase (ODC) activity. The other assay measured retinoid-induced differentiation of the human myoloid leukemia cell line HL-60. In the ODC assay, all thirteen compounds were screened. The most active heteroarotinoids were ester 10 [methyl (E)-4-[2-(2,2,4,4-tetramethylthiochroman-6-yl)-1- propenyl]benzoate] and acid 11 [(E)-4-[2-(2,2,4,4-tetramethyl-3,4- dihydro-2H-1- benzothiopyran-6-yl)-1-propenyl]benzoic acid]. Both of these retinoids had ID50 values (dose required for half-maximal inhibition of phorbol ester induced ODC activity) of about 0.3 nmol. In comparison, the ID50 value for trans-retinoic acid (1) was 0.12 nmol while the ID50 values for acids 7 and 9, namely (2Z,4E,6E)-3,7-dimethyl-7-(4,4-dimethyl-thiochroman -6-yl)-2,4,6-heptatrienoic acid and (2E,4E,6E)-3,7-dimethyl-7-(2,2,4,4-tetramethylthiochroman -6-yl)-2,4,6- heptatrienoic acid, respectively, were about 3.5 nmol. Heteroarotinoids 8 and 12-17 had ID50 values of 35 nmol or greater. With a thiochroman unit, the most active acids in decreasing order of activity in the ODC assay were 7 greater than 9 greater than 8. Thus, simple replacement of the terminal propenyl system [C(16,17,18)] in 7 with a cyclopropyl group produced acid 8 [(2E,4E,6E)-7-methyl-7-(4,4-dimethylthiochroman-6-yl)- 2,3-methylene-4,6-heptadienoic acid with markedly reduced activity. With a benzoic acid group as part of the structure attached to the thiochroman unit, the ODC activity was enhanced as shown in 10 and 11. The combination of the 2,2,4,4-tetramethylthiochroman group and the benzoic acid (or ester) terminal group seemed to enhance the biological action which resembles that found with (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)- 1-propenyl]benzoic acid (TTNPB, 6b), a well-known model system.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Human cutaneous myiasis due to mixed infestation in a drug addict.

Soft tissue infestation by maggots of flies (myiasis) is a well-recognized complication of neglected wounds. Mixed infestation by larvae of more than one species of fly is a rarity. We report, for the first time in India, one such case of cutaneous myiasis due to mixed infestation--in this instance by Chrysomya bezziana and Sarcophaga sp. in a drug addict.

Adult↗
PubMed1991

Absence of cognitive impairment after more than 100 lifetime ECT treatments.

The cognitive function scores and subjective memory complaints of eight patients who had each received more than 100 treatments with bilateral modified since wave ECT were equivalent to those of matched patients who had never received ECT. The results suggest that patients given many ECT treatments over several courses do not manifest measurable cognitive impairment at long-term follow-up.

Adult↗
PubMed1990

Expression of human chromosome 2 ornithine decarboxylase gene in ornithine decarboxylase-deficient Chinese hamster ovary cells.

Ornithine decarboxylase (ODC) belongs to a multigene family and some of these may very well be nonfunctional (pseudogenes). We isolated an ODC gene from a human chromosome 2-specific library and transfected the gene into ODC-deficient Chinese hamster ovary cells to directly demonstrate that this ODC gene is functional and ODC is essential for cell proliferation. After screening 2.5 X 10(5) plaques using a human ODC complementary DNA probe, a typical clone with a 5.4-kilobase insert was isolated and then cloned into the HindIII site of the pGem-1 vector. One (phODC 2B1) of these clones containing a 5.4-kilobase ODC gene insert was identified. Restriction enzyme analysis and partial sequencing data revealed that phODC 2B1 contained the full length protein-coding sequences but lacked first exon and 3'-polyadenylation sequences. Primer extension analysis indicated that human ODC mRNA has homologous sequences with the ODC gene from human chromosome 2. To determine that the chromosome 2 ODC gene is functional, ODC-deficient Chinese hamster ovary cells were transfected with the ODC expression vector (phSV2B1-neo) and several G418-resistant transfectants were isolated which expressed 70- to 400-fold more ODC activity than parental or wild-type Chinese hamster ovary cells. Furthermore, these stable transfectants exhibited a higher growth rate than wild-type cells. These results indicate that the ODC gene from human chromosome 2 encodes functional ODC protein, and ODC (and its product putrescine) is required for cell growth.

Animals↗
PubMed1990

The calcium-transporting ATPase and the calcium- or magnesium-dependent nucleotide phosphatase activities of human placental trophoblast basal plasma membrane are separate enzyme activities.

The properties of calcium-stimulated ATP hydrolysis often differ from those of ATP-dependent calcium transport. We have characterized two components of calcium-stimulated ATP hydrolysis in human placental basal plasma membrane. In the absence of magnesium, component 1 apparently has saturable sites for free calcium in both the nanomolar and low micromolar range. It was stimulated by either calcium or magnesium, was unselective for nucleotide substrate, and its activity was very much greater than that of ATP-dependent calcium transport. Component 1 was inhibited by GTP, permitting measurement of component 2 with activity and magnesium stimulation comparable to ATP-dependent calcium transport. Component 2 was inhibited partially by an antibody against purified erythrocyte calcium transporter and completely by sulfhydryl reagents, whereas component 1 was unaffected. A phosphorylated intermediate of the calcium transporter co-migrated with the erythrocyte transporter on acidic sodium dodecyl sulfate-polyacrylamide electrophoresis gels. Immunostaining after transfer to nitrocellulose revealed a doublet. The band of lower molecular weight co-migrated with that of the human erythrocyte membrane transporter. The addition of GTP permits separate measurement of ATP hydrolysis by the calcium transporter of the placental basal plasma membrane and may be useful in defining its properties in other cell membranes under a variety of conditions.

Adenosine Triphosphatases↗
PubMed1990

Absence of effect of supplemental oral calcium on ornithine decarboxylase (ODC) activity in colonic mucosae of healthy individuals with a family history of colorectal cancer.

Tissue activity levels of ornithine decarboxylase (ODC) have been suggested to be a marker of colonic epithelial cell proliferation and risk for colorectal cancer. Supplemental dietary calcium has been hypothesized and reported to suppress colonic epithelial-cell proliferation. We measured sigmoid and rectal mucosal ODC activity levels in 45 healthy, disease-free subjects with strong family histories of colorectal cancer before and after 2 months, during which daily dietary supplementation with calcium carbonate (to provide 600 mg calcium base) was taken. Although the mean ODC activity levels decreased in both sigmoid and rectal specimens, these changes were small in relation to the standard deviation and were not statistically significant. These data suggest that the magnitude of dietary calcium intake does not significantly influence sigmoid-rectal mucosal ODC activity levels and that ODC measurements may not be a useful intermediate endpoint for interventions designed to interrupt the colon carcinogenic sequence.

Administration, Oral↗
PubMed1990

Inhibition of tumor promotion by DL-alpha-difluoromethylornithine, a specific irreversible inhibitor of ornithine decarboxylase.

Knowledge of the mechanisms of carcinogenesis is helpful for planning strategies and in the rational choice of agents for cancer prevention. There is a great potential for intervention at the promotion step of human carcinogenesis. ODC induction is associated with the promotion stage of carcinogenesis. Consequently, DFMO may be a useful drug for cancer prevention in humans. Long-term medication with higher doses (9 gm/m2/da) of DFMO has resulted in several toxic side effects, such as thrombocytopenia and reversible ototoxicity. However, doses of DFMO (less than 1 gm/m2/da), selected by our in vitro human skin punch biopsy assay (16, 47), may be given for a longer period without appreciable toxicity and should be evaluated in human cancer prevention trials.

Animals↗
PubMed1990

Cellular and segmental distribution of Ca2(+)-pump epitopes in rat intestine.

We used a monoclonal antibody (5F10) specific for the human erythrocyte plasma membrane Ca(++)-pump to demonstrate the presence and distribution of Ca(++)-pump epitopes in rat intestine. In paraffin embedded tissue sections, antibody 5F10 binds to epitopes in the basolateral membranes of absorptive cells in rat duodenum and portions of jejunum but not ileum. Western blot analysis of intestinal mucosal proteins with antibody 5F10 shows binding of antibody to major bands of Mr approximately 135,000 and Mr approximately 72,000, and to lesser bands of Mr approximately 125,000 and Mr approximately 27,000. This pattern was seen in mucosal homogenates of rat duodenal and jejunal cells and to a lesser extent in ileal cells. The Mr approximately 135,000 band corresponds to the molecular weight of Ca(++)-pumps in other tissues. The other bands correspond in size to known proteolytic fragments of the Ca(++)-pump. Slot-blot analysis of nitrocellulose immobilized mucosal homogenates shows binding of 5F10 to be greatest in duodenum and least in ileum. Ca(++)-transport studies by the everted gut sac technique show a correlation between vitamin D induction of active Ca(++)-transport and the segmental distribution of Ca(++)-pump epitopes.

Animals↗
PubMed1990

90Y.B72.3 against pancreatic cancer: dosimetric and biological analysis.

Nude mice xenografted with a human pancreatic carcinoma cell line were injected with yttrium-90 (90Y) conjugated to diethylene triaminepenta acetic acid (DTPA) alone, and DTPA covalently linked to a monoclonal antibody, B72.3. The animals were sacrificed in temporal sequence to evaluate isotope distribution. Dosimetry was carried out using the principles outlined in MIRD and ICRU Report 32. Results are expressed as percent uptake per unit mass in organs and tumor and as relative absorbed dose normalized to 90Y uptake in liver at 7 hr. When conjugated to B72.3, an 8-fold increase in isotope localization in the tumor was noted by 24 hr. When the relative absorbed dose is calculated for 90Y and 90Y.B72.3, a 26-fold increase in tumor dose is noted for the 90Y conjugate. Normal tissues show no to modest (less than 5x) enhanced dose with 90Y.B72.3. B72.3, therefore, deserves further investigation as a potential monoclonal antibody for targeting therapeutic radioisotopes and possibly diagnostic radioisotopes to pancreatic cancer. Radiobiological aspects of the low dose rates from radioimmunotherapy are discussed.

Animals↗
PubMed1989

Lack of a role of DNA methylation in tumor promoter 12-O-tetradecanoylphorbol-13-acetate-induced synthesis of ornithine decarboxylase messenger RNA in T24 cells.

Association of alteration in DNA methylation pattern in triggering 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced transcription of ornithine decarboxylase (ODC) gene in T24 cells was determined. In accord with our previous findings (Archiv. Biochem. Biophys., 262: 326-336, 1988), TPA treatment of T24 cells, cultured in serum-free medium, resulted in a dramatic (approximately 15-fold) increase in ODC activity which was accompanied by a proportional increase in hybridizable amount of ODC mRNA. Data from nuclear run-off transcription assay revealed that TPA-induced accumulation of ODC mRNA is the result of increased transcription initiation. Since DNA hypomethylation has been proposed to be a mechanism involved in the regulation of transcription of some gene(s), we examined the changes in the methylation patterns in the ODC gene isolated from the vehicle (ethanol)- and TPA-treated T24 cells. The autoradiograms resulting from the Southern blot analysis of DNA cleaved with several methylation-sensitive restriction endonucleases [e.g., HpaII, MspI, cfoI (HhaI), SalI, XhoI] exhibited no difference in methylation pattern of ODC gene in T24 cells. Also, a single or chronic application of TPA to either noninitiated or 7,12-dimethylbenz(a)anthracene-initiated mouse skin failed to alter DNA methylation pattern of ODC gene. Furthermore, the hypomethylation agent 5-azacytidine failed to induce ODC mRNA in T24 cells. These results indicate that TPA does not affect the methylation status of ODC gene and hypomethylation may not be sufficient for TPA-increased ODC gene transcription in T24 cells.

DNA↗
PubMed1989

Colon ornithine decarboxylase activity following standard endoscopy preparation regimens.

Ornithine decarboxylase (ODC) catalyzes the formation of putrescine from ornithine, which is the first step in the pathway of mammalian polyamine biosynthesis. Tissue activity levels of ODC have been suggested to be a marker of risk for colorectal cancer in hereditary polyposis and in adenoma formers. We analyzed ODC activity in rectal and sigmoid colon mucosal biopsies obtained at 10 cm and at 30 cm in 40 healthy, colon cancer risk factor-free adults following three endoscopic preparation regimens: 1) no special preparation; 2) two phosphate enemas; and 3) "Colyte" lavage preparation 12 hr previously. Levels of ODC, measured in fresh tissue, were approximately twofold higher for enema preparation vs. no preparation (for log-transformed data: sigmoid, P less than 0.0001; rectum, P = 0.0001) and for enema preparation vs. lavage (sigmoid, P = 0.0002; rectum, P = 0.008). Lavage and no preparation ODC levels were not significantly different. ODC activity levels ranged from 0.00 to 352.96 pmol/mg/hr.

Adenomatous Polyposis Coli↗
PubMed1989

The enzyme-activated irreversible inhibitor of ornithine decarboxylase, DL-alpha-difluoromethylornithine: a chemopreventive agent.

DL-alpha-Difluoromethylornithine (DFMO) is an enzyme-activated irreversible inhibitor of mammalian ornithine decarboxylase. DFMO, when administered in drinking water, precludes increases in the levels of intracellular putrescine and spermidine and also inhibits the induction of skin, breast, colon, urinary bladder, and intestinal cancers in experimental animal models. DFMO may be a useful drug for cancer prevention in humans; however, long-term medication with higher doses (9 g/m2/day) of DFMO has resulted in several toxic side effects such as thrombocytopenia and reversible ototoxicity. Smaller doses (less than 1 g/m2/day), selected by our in vitro human skin punch biopsy assay, may be given for a longer period without appreciable toxicity. Further evaluation in human cancer prevention trials is indicated.

Animals↗
PubMed1989

Calcium pump epitopes in placental trophoblast basal plasma membranes.

The syncytiotrophoblast represents the primary cellular barrier between maternal and fetal circulations in the placenta. Large amounts of Ca2+ are transported across this barrier by mechanisms that are not clearly understood. To further understand this phenomenon, we examined rat and human placenta by immunohistochemical and protein blotting techniques with a monoclonal antibody raised against the human erythrocyte plasma membrane Ca2+ pump. Immunohistochemistry with this antibody showed specific staining in the human placenta of the basal (fetal facing) surface of the syncytiotrophoblast. In the rat placenta, immunohistochemistry also showed specific staining of the innermost (fetal facing) layer of the trophoblast and the basal surface of the endoderm of the intraplacental yolk sac. In Western blots of placental homogenates and membranes, the monoclonal antibody bound to a 140,000-mol wt band, characteristic of Ca2+ pumps in other tissues. Western blots of isolated basal membranes showed more intense staining than isolated microvillous membranes, confirming the results of the immunohistochemistry. In addition, Ca2+ transport in basal membrane vesicles from human placenta was inhibited by polyclonal antibodies prepared against the erythrocyte Ca2+ pump. We conclude that basal (fetal facing) layers of human and rat placentas contain a high-affinity Ca2+ pump situated to transport Ca2+ from the maternal to the fetal circulation.

Animals↗
PubMed1989

Plasma membrane calcium pump and 28-kDa calcium binding protein in cells of rat kidney distal tubules.

In an effort to extend our studies on Ca2+ pumps to animal models, we developed a new monoclonal antibody (5F10) prepared against the human erythrocyte Ca2+-Mg2+-adenosinetriphosphatase (ATPase) that recognizes a protein of approximately 140 kDa in rat kidney homogenates. Enzyme-linked immunosorbent assays show that monoclonal antibody 5F10 binds purified Ca2+-Mg2+-ATPase and rat kidney membrane extracts in a concentration-dependent manner. In paraffin-embedded tissue sections, antibody 5F10 binds to an epitope in the basolateral membranes of rat kidney distal convoluted tubule principal cells. The antibody does not bind to intercalated cells. The latter cells were characterized by the presence of large amounts of carbonic anhydrase C. Polyclonal antibodies directed against chick intestinal 28-kDa vitamin D-dependent calcium binding protein (28-kDa CaBP) also bind epitopes in distal convoluted tubule cells, connecting tubules, and portions of collecting duct but not intercalated cells. Western blot and 45Ca blot analysis of renal cytosolic proteins showed that the polyclonal 28-kDa CaBP-directed antibody detects a protein which also binds calcium. Western blot analysis with monoclonal antibody 5F10 shows binding to both the authentic purified erythrocyte Ca2+ pump (approximately 138 kDa) and to tryptic fragments of this pump. Antibody JA3, previously used for staining of human kidney tubules, reacts with a different set of tryptic fragments, showing that the two antibodies are directed against different regions or conformational determinants on the pump molecule. We show that Ca2+-Mg2+-ATPase and 28-kDa CaBP are present in the principal cells of the distal convoluted tubule of the rat and are absent in intercalated cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Ornithine decarboxylase activity as a biochemical marker in individuals predisposed to lung cancer.

Ornithine decarboxylase (ODC) is the first enzyme in the pathway of mammalian polyamine biosynthesis. "Tumor promoters" appear to induce ODC. In addition, increased ODC activity has been observed in normal appearing colonic mucosa of patients with familial polyposis, an autosomal dominant disorder associated with a high incidence of colon cancer. Ornithine decarboxylase activity was measured in bronchoscopic mucosal biopsy specimens from the noninvolved lung in patients with unilateral lung masses. No correlation could be determined in ODC levels from "normal" mucosa between patients with lung cancer compared to those with benign disease, despite elevated ODC activity in the tumors themselves.

Biomarkers↗