Search PubMed⌕ Search

Biomedical subjects

A K Roy

Publications and source records attributed to A K Roy.

At least 55 records · Page 3Linked to original sources
PubMed1995

Molecular cloning and sequence analysis of the rat liver carnitine octanoyltransferase cDNA, its natural gene and the gene promoter.

The full-length cDNA and the natural gene for rat peroxisomal carnitine octanoyltransferase (COT) have been isolated and sequenced. The 2681 bp long cDNA contains an open reading frame for 613 amino acids, resulting in a protein with a deduced molecular weight of 70,301, and a C-terminal peroxisomal targeting sequence (Ala-His-Leu). The isolated COT cDNA has 51 bp of the 5' untranslated region (UTR), 791 bp of 3' UTR, two putative polyadenylation sites, and a poly(A19-23) tail. Screening of a rat genomic DNA library in the lambda phage with the COT cDNA probe resulted in the isolation of seven overlapping clones, together containing the complete COT gene with seventeen exons. All of the exon-intron boundary sequences conform to the GT-AG rule. The COT gene appears to spread over 40 to 60 kbp region of the rat genome. The transcription initiation site of the COT gene was determined through primer extension, and the promoter sequence up to the position -1140 was established. The promoter lacks the canonical TATA box and a promoter-reporter construct containing the sequence encompassing -1140 to +84 base positions and the firefly luciferase reporter cDNA yielded about 100-fold increase in promoter activity in transfected hepatoma cells. Some of the consensus sequences for putative cis elements present in the promoter sequence are: the two CCAAT motifs for CTF/NF1/CBP binding (at -284 and -93), two GC boxes for Sp1 binding (at -160 and -68), two AP2 sites (at -359 and -25), a half site (TGACCT) for the peroxisome proliferator activated receptor (PPAR) binding at -737 within a partial palindromic sequence region. Potential regulatory elements, such as several palindromes and repeat motifs for five different sequence segments, are also identified.

Amino Acid Sequence↗
PubMed1995

Production and characterization of the extracellular domain of human CD7 antigen: further evidence that CD7 has a role in T cell signaling.

CD7 is a T cell-associated antigen which appears early in ontogeny and persists on circulating T cells. It appears to have a significant role in T cell development and function. The precise mechanism by which this molecule mediates its effect is not known. In this paper, we expressed the extracellular domain of CD7 in the baculovirus system and used this product to study the function CD7 might have in T cell activation. The recombinant protein was found to be structurally similar to the native CD7 and recognized by monoclonal and polyclonal antibodies to CD7. This protein inhibited T cell proliferation induced by anti-CD3/anti-CD7 costimulation. It also inhibited the augmentation effect of anti-CD7 on suboptimal PHA stimulation. However, it did not block T cell proliferation induced by optimal doses of PHA, staphylococcal entertoxin A or B. Interestingly, the recombinant protein inhibited antigenic- and alloantigenic-induced T cell proliferation. The latter finding strongly suggests that a ligand for CD7 exists and crosslinking CD7 by this ligand may be responsible for the costimulatory role it plays in T cell activation.

Adult↗
PubMed1995

Regulation of androgen action by receptor gene inhibition.

Regulated functions of hormonal agents play a critical role in health and disease. Target cell responsiveness to a hormonal signal is a product of both cellular concentrations of the hormone ligand and the corresponding receptor protein. The major thrust of the drug design for treatment of endocrine-related problems, so far, has been directed to ligand derivatives. In certain cases, receptor regulation through antigene technology has much to offer with improvements in both target cell and hormonal specificity. Three different antigene approaches are currently being explored. The first approach is to inhibit the expression of the receptor gene by disrupting the DNA protein interaction at critical cis-elements by short triple helix-forming oligonucleotides. The second approach is to sequester and inactivate the receptor mRNA by the antisense mRNA produced in the target tissue directed by a heterologous tissue-specific promoter. The third approach is the tissue-specific expression of a catalytic ribozyme that binds to the specific receptor mRNA and selectively degrades it before its translation into the protein. In this study, we have characterized the promoter of the rat androgen receptor, and by progressive deletion from its 5' end have identified two critical cis-regulatory elements, one at the -960 to -940 region and the other at the -554 to -574 positions. The former is an activator while the latter is an inhibitor domain. The inhibitory domain is the binding site for the nuclear factor kappa B (NF-kappa B) and more specifically, the p50/p50 homodimer of this transcription factor family. We have also provided correlative data to show that under normal physiological conditions, the NF-kappa B functions as an antiandrogen during the age-dependent desensitization of the liver. In addition to the naturally functioning antiandrogenic influence of NF-kappa B, we have designed an artificial antiandrogenic agent, a triplex-forming oligonucleotide (TFO) directed to the -960/-940 activator domain of the rat androgen receptor gene promoter. This oligonucleotide at a TFO-to-promoter ratio of 500 is able to cause about 60% inhibition of rAR promoter function in transfected COS-1 cells. These results clearly demonstrate the feasibility of the antigene approach for effective inhibition of steroid hormone action.

Androgens↗
PubMed1995

Nuclear factor kappa B functions as a negative regulator for the rat androgen receptor gene and NF-kappa B activity increases during the age-dependent desensitization of the liver.

Transcriptional regulation of the steroid hormone receptor genes plays a central role in temporal changes of target cell sensitivity during development, maturation, and aging. Sequence-specific DNA-protein interactions mediate these regulatory functions. Progressive 5' deletion of the rat androgen receptor (rAR) gene immediately beyond the -572 base pair (bp) region causes a marked increase in its promoter activity. DNase I footprinting with nuclear proteins revealed a protected area encompassing -574- to -554-bp positions that begins with a perfectly palindromic nuclear factor kappa B (NF-kappa B) motif. Electrophoretic mobility shift analyses (EMSA) showed that the decameric rAR NF-kappa B site at positions -574 to -565 cross-competes with the authentic kappa immunoglobulin light chain enhancer for specific protein binding. Supershift with specific antibodies to NF-kappa B subunits confirmed that the two retarded bands observed in the EMSA with the labeled rAR probe are due to p50/p65 and p50/p50 dimers of the NF-kappa B/Rel proteins. Fragments of rAR promoter with either deletion or point mutation of the NF-kappa B site are found to be about 2- to 3-fold more effective as compared to the wild type control in driving a heterologous reporter gene in cellulo. Thus, unlike most other known cases, NF-kappa B acts as a negative regulator for the rAR gene. The physiological relevance of this repressor function is evident from a 10-fold increase in the p50/p50 form of the NF-kappa B activity in the liver of aged rats exhibiting hepatic androgen desensitization. The newly identified repressor element is a rare example of a naturally occurring perfect palindromic binding motif for the NF-kappa B/Rel family of transcription factors. This repressor factor and the positively acting age-dependent factor, ADF, described earlier (Supakar, P. C., Song, C. S., Jung, M. H., Slomczynska, M. A., Kim, J.-M., Vellanoweth, R. L., Chatterjee, B. & Roy, A. K. (1993) J. Biol. Chem. 268, 26400-26408) function to coordinate the tissue-specific down-regulation of the rAR gene during aging.

Aging↗
PubMed1995

Tubular hypertrophy due to work load induced by furosemide is associated with increases of IGF-1 and IGFBP-1.

We have examined the expression of insulin-like growth factor 1 (IGF-1), IGF binding protein-1 (IGFBP-1), and IGF binding protein-3 (IGFBP-3) in the rat distal nephron during increased cell work load and hypertrophy, induced by the diuretic, furosemide. Furosemide was given for six days to increase distal sodium delivery and uptake. To mitigate salt loss, the animals drank 0.8% NaCl and 0.1% KCl. Control rats were infused with vehicle (0.9% saline) and drank tap water. Furosemide increased urinary volume (13-fold) and sodium excretion (eightfold), and decreased urine osmolarity (fourfold). By immunocytochemistry, staining for IGF-1 and IGFBP-1 was markedly increased in distal convoluted tubules and cortical collecting ducts; both segments also underwent hypertrophy. Increased staining for the peptides was evident early (1 hr, 18 hr) after furosemide, prior to hypertrophy of cells. Whereas transcripts of IGF-1 and IGFBP-3 mRNA showed little or no increase in extracts from furosemide-treated kidney cortices, IGFBP-1 mRNA was increased threefold 18 hours after furosemide. Alterations of IGF-1 and IGFBP-1 were independent of changes in plasma aldosterone, glucocorticoids or arginine vasopressin. That IGFBP-1 mRNA increased threefold without significant changes in IGF-1 mRNA suggests that hypertrophic stimuli might initially induce the synthesis of IGF binding protein followed by the trapping of extracellular IGF-1. The present study raises the possibility of IGF-1 and IGFBP-1 being involved in processes that lead to tubular hypertrophy. IGFBP-1 may regulate these effects by binding to and interaction with IGF-1.

Animals↗
PubMed1995

Endoscopic US of the gastrointestinal tract with endoscopic, radiographic, and pathologic correlation.

Endoscopic ultrasonography (US) makes it possible to evaluate the layers of the wall of the gastrointestinal tract, as well as surrounding structures. The authors have used endoscopic US to evaluate the gastrointestinal tract in 160 patients. In the esophagus, endoscopic US is useful in staging esophageal carcinoma, planning radiation therapy ports, and locating and characterizing esophageal masses. Endoscopic US of the esophagus is also used to identify varices and evaluate the results of therapy. Applications of endoscopic US in the stomach include staging of gastric carcinoma, localization and characterization of nonmucosal gastric masses, detection and evaluation of gastric varices, and evaluation of gastric lymphoma. In the pancreas, endoscopic US is used for detecting pancreatic masses; staging pancreatic, distal common bile duct, and ampullary carcinoma; and evaluation biliary tract obstruction. Applications of endoscopic US in the rectum include localization and characterization of nonmucosal lesions and local staging of rectal carcinoma.

Digestive System Diseases↗
PubMed1995

Androgen action.

Androgens are C-19 steroids that provide major regulatory influences on male reproductive function. Testosterone, the principal androgenic steroid, is secreted by the Leydig cells of the testes. Both testosterone and its 5 alpha reduced derivative 5 alpha-dihydrotestosterone (DHT) are physiological ligands for the androgen receptor (AR). Ligand-activated AR acts as a nuclear transcription factor and mediates androgen action. AR, along with receptors for a number of C-21 steroids such as glucocorticoid, mineralocorticoid, and progesterone, share the same 15 base pair consensus element composed of 5'-GGA/TACAnnnTGTTCT-3'. Despite this cross-reactivity at the level of the DNA, physiologically, androgens regulate their target genes with a high degree of receptor specificity. Such a regulatory specificity appears to be due to multiphasic interactions involving enzymatic activation/inactivation of the steroid ligand, interaction with specific receptor-associated nuclear factors on or around the hormone response element, and differential regulation of the receptor gene expression. Conversion of testosterone to 5 alpha-dihydrotestosterone in target cells is a widespread activation mechanism that amplifies the androgenic signal. Unlike the testosterone-AR complex, DHT-activated AR has a longer half-life, and thus prolongs androgen action. Oxido-reduction of androgens by 17 beta-hydroxysteroid dehydrogenase and sulfurylation by androgen sulfotransferase are two major pathways of androgen inactivation in target cells. Prenatal deprivation of androgen action, due to mutations in either the AR or the 5 alpha-reductase gene, results in developmental abnormalities of male reproductive tissues and also cause partial or complete androgen-insensitivity syndromes. Elucidation of various molecular steps in androgen action is allowing development of improved therapeutic agents for the management of disorders of androgen action such as the prostatic hypertrophy and neoplasia.

Androgens↗
PubMed1995

Cytogenetic study of primary amenorrhoea.

A study of chromosomal pattern was done in 60 cases of primary amenorrhoea of different age groups to determine the incidences of chromosomal abnormalities in them and to detect those cases of classical 45, X Turner's syndrome and Turner mosaics that do not bear the Turner stigmata. Buccal smears were examined for sex chromatin followed by karyotype using leucocyte culture method. Majority (63.3%) of cases were found chromosomally incompetent of which the major abnormality was 45, X/46, XX mosaicism (33.3%) followed by 45, X Turner's syndrome (26.6%). But only 43.7% of these Turner's syndrome had classical Turner stigmata. Two cases of complete testicular feminisation syndrome with male genotype (46,XY) and inguinal testis were also detected.

Adult↗
PubMed1994

Androgen and estrogen sulfotransferases of the rat liver: physiological function, molecular cloning, and in vitro expression.

Cloning of the androgen and estrogen sulfotransferases has allowed studies on their spatiotemporal regulation and physiological function. These two enzymes appear to be involved in regulating target cell sensitivity for the androgenic and estrogenic steroids. Recombinant androgen sulfotransferase produced in insect Sf9 cells through a baculoviral expression vector is capable of transferring the sulfate from PAPS (phosphoadenosine phosphosulfate) to 3 alpha, 3 beta, 17 alpha, and 17 beta hydroxyl groups of a number of steroid substrates indicating a broad range of substrate specificity.

Aging↗
PubMed1994

Reentry monitoring in the treatment of physicians with substance dependence.

A total of 37 physicians and other health care professionals have been accepted into a monitoring and advocacy program for an average of 24 months. Thirty-five are working in their profession, one is in treatment, and one has repeatedly relapsed and is no longer practicing medicine (an overall improvement rate of 97%). The elements of "reentry monitoring" are identified and proposed as a standard for the treatment of physicians with substance dependence.

Alcoholism↗
PubMed1994

In vitro analysis of the interaction between sucralfate and ketoconazole.

In healthy volunteers, the bioavailability of ketoconazole is significantly decreased during simultaneous administration with sucralfate. In an effort to address this problem, we examined the interaction between sucralfate and ketoconazole in aqueous solutions and in simulated gastric fluid (SGF) at various initial pHs (1, 2, 3, and 6) in the presence or absence of glutamic acid hydrochloride (GA). Samples from each solution were taken 30 min and 2 h after the addition of ketoconazole to evaluate the solubility of ketoconazole over the usual time period of maximal absorption of ketoconazole in humans. The addition of GA to SGF leads to an increase in solution acidity, while the pHs of SGF at a pH of 1, 2, or 3 are markedly increased by the addition of sucralfate. There is a net decrease in acidity from initial pHs for the pH 1, 2, and 3 solutions when GA and sucralfate are combined. The concentration of ketoconazole in SGF at pHs of 1, 2, 3, 4, and 6 was evaluated in order to assess the pH-dependent solubility properties of the drug in the absence of other interacting species. Regardless of the initial pH, combinations of GA plus ketoconazole showed high concentrations of ketoconazole (approximately 100%) in solution. In contrast, significant decreases in the concentration of soluble ketoconazole were observed when sucralfate was mixed with ketoconazole, and, in some cases, soluble ketoconazole was not detectable. The addition of GA to a mixture of sucralfate and ketoconazole leads to a significant increase in the concentration of solubilized ketoconazole. Nonetheless, important sucralfate-ketoconazole interactions are still observed. After 2 h, approximately 35% of the maximal ketoconazole concentration remained in solution. Comparison of the ketoconazole concentrations at different pHs with the predicted concentrations of the three protonation species of ketoconazole [H2(ketoconazole)(2+), H(ketoconazole)(+), or ketoconazole] showed no correlation. Therefore, the decrease in ketoconazole solubility is not simply a reflection of pH perturbation associated with the dissolution of sucralfate. The observed data are most consistent with a model that has H2(ketoconazole)(2+) or H(ketoconazole)(+) forming an electrostatic interaction with the sucralfate polyanion. The findings of this study suggest that the coadministration of sucralfate with other azole antifungal agents should be investigated.

Chemistry, Pharmaceutical↗
PubMed1994

Development of methods to monitor exposure to 1-nitropyrene.

On the basis of 32P-postlabeling analysis, treatment of rats with 1-nitropyrene (1-NP) resulted in the formation of multiple DNA adducts in the liver, mammary glands, and peripheral lymphocytes. The one adduct resulting from nitroreduction, N-(deoxyguanosin-8-yl)-1-aminopyrene, constitutes only a minor component among the adducts. In the present study, incubation of calf thymus DNA with mutagenic ring-oxidized metabolites of 1-NP in vitro in the presence and absence of xanthine oxidase also resulted in the formation of multiple adducts. On the basis of their chromatographic behavior, it appears that DNA adducts derived from such metabolites may have been formed in vivo; however, this needs to be confirmed. [3H]1-NP was given to male and female F344 rats and Sprague-Dawley rats by gavage at five dose levels in the range of 0.1 to 1000 micrograms/kg bw. This led to stable hemoglobin adducts accounting for 0.08 +/- 0.05% of the dose (n = 3 rats). The radioactivity associated with hemoglobin following administration of [3H]1-NP was cleared with a half-life of about 14 days, which is faster than that of unmodified erythrocytes in the rat (t1/2 = 30 days). Treatment of the hemoglobin with 1% HCl in acetone, to precipitate the globin, released the radioactivity; it was all bound to the heme moiety. The structures of the heme adducts have not been elucidated; yet, because of their stability, they may be useful as dosimeters for human exposure to 1-NP.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗
PubMed1994

Albendazole therapy for neurocysticercosis.

Based on clinical evaluation and computed tomography (CT) of the brain, 30 cases of neurocysticercosis were diagnosed. Diagnosis was supported by presence of histopathologically proven subcutaneous cysticerci in 12 cases. Three primary neurological syndromes were established i.e. epilepsy in 22 cases, increased intracranial tension in 6 cases and meningoencephalitis in 2 cases. Albendazole was administered orally in a dose of 15 mg/kg bodyweight/day for 30 days without prophylactic steroids. Follow up CT study at 3 months and 12 months revealed complete regression of all lesions in 2 cases, partial regressions in 14 cases and change in morphology in 4 cases. Transient appearence of fresh subcutaneous cysticerci as a side effect of therapy was noted in 4 cases. Albendazole, though acting slow, is considered a suitable alternative to praziquantel in medical management of parenchymal neurocysticercosis.

Adolescent↗
PubMed1994

Biomonitoring of nitropolynuclear aromatic hydrocarbons via protein and DNA adducts.

Nitropolynuclear aromatic hydrocarbons (nitro-PAHs) are widely distributed in the environment. For several chemicals in this class of compounds, mutagenic activity in bacterial and mammalian systems and tumorigenic activity in laboratory animals have been clearly documented. Procedures for assessing the risk to humans from exposure to nitro-PAHs have not been clearly defined, despite the wide-spread occurrence of such agents in the environment and their possible involvement in the etiology of some human cancers. Several methods are available for determining exposure, uptake, and metabolic activation of genotoxic carcinogens in humans. DNA adducts currently are regarded as the most direct markers of genotoxicity. However, several proteins are equally capable of forming adducts with electrophiles derived from xenobiotics. We focused on developing methods to detect and quantify adducts of 1-nitropyrene and 1,6-dinitropyrene with proteins and with DNA. 1-Nitropyrene is the most abundant nitro-PAH in emissions from combustion sources such as diesel engines. Although 1,6-dinitropyrene is far more mutagenic and more tumorigenic than 1-nitropyrene, it is present in the environment at lower levels. Seeking a highly sensitive method, we have utilized the 32P-postlabeling technique to establish the pattern of the DNA adducts formed in rat tissues, as well as in peripheral blood lymphocytes, following administration of both 1-nitropyrene and 1,6-dinitropyrene. We also present results on hemoglobin and albumin adducts formed after administration of these nitro-PAHs. [3H]1-Nitropyrene was given to male or female Fischer-344 or Sprague-Dawley rats by gavage at five dose levels ranging from 0.1 to 1,000 micrograms/kg of body weight. This led to stable hemoglobin adducts, which accounted for 0.08% +/- 0.05% of the dose. The radioactivity associated with hemoglobin following administration of [3H]1-nitropyrene was cleared with a half-life of 13.6 days. This is faster than the clearance of unmodified erythrocytes in the rat (half-life of 30 days). Treating the hemoglobin with 1% hydrochloric acid in acetone, to precipitate the globin, released the radioactivity so that none remained bound to the globin. Rather, the radioactivity remained bound to the heme moiety. To obtain structural information about the heme adducts, we incubated [3H]1-nitrosopyrene and [3H]4,5-epoxy-4,5-dihydro-1-nitropyrene with rat hemoglobin. In each case, [3H] was bound mainly to globin and, to a lesser extent, to the heme moiety.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylation↗
PubMed1993

A novel regulatory element associated with age-dependent expression of the rat androgen receptor gene.

A large body of evidence indicates that the genetic program of aging has co-evolved with the sexual mode of reproduction (Partridge, L., and Barton, N. H. (1993) Nature 362, 305-311). Age-dependent changes in target cell sensitivity to reproductive hormones can be considered part of this evolutionary linkage. Here we describe a novel regulatory element in the rat androgen receptor (AR) gene promoter associated with its age-dependent expression in the liver. This element consists of two (19 and 25 base pairs) contiguous sites, one specifically binding an Age-dependent Factor (ADF) and the other an Associated Factor (AF). Both deletion and point mutations of the ADF site result in about a 5-fold decline in the AR promoter function. Unlike AF, which is relatively tissue specific, ADF appears to be ubiquitous. The ubiquitous and evolutionarily conserved nature of ADF suggests a fundamental role of this novel transcription factor in programmed gene expression.

Aging↗