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J K Reddy

Publications and source records attributed to J K Reddy.

At least 91 records · Page 5Linked to original sources
PubMed1992

Identification of a peroxisome proliferator-responsive element upstream of the gene encoding rat peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase.

Ciprofibrate, a hypolipidemic drug that acts as a peroxisome proliferator, induces the transcription of genes encoding peroxisomal beta-oxidation enzymes. To identify cis-acting promoter elements involved in this induction, 5.8 kilobase pairs of promoter sequence from the gene encoding rat peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase (EC 4.2.1.17/EC 1.1.1.35) was inserted upstream of a luciferase reporter gene. Transfection of this expression vector into rat hepatoma H4IIEC3 cells in the presence of ciprofibrate resulted in a 5- to 10-fold, cell type-specific increase in luciferase activity as compared to cells transfected in the absence of drug. A peroxisome proliferator-responsive element (PPRE) was localized to a 196-nucleotide region centered at position -2943 from the transcription start site. This PPRE conferred ciprofibrate responsiveness on a heterologous promoter and functioned independently of orientation or position. Gel retardation analysis with nuclear extracts demonstrated that ciprofibrate-treated or untreated H4IIEC3 cells, but not HeLa cells or monkey kidney cells, contained sequence-specific DNA binding factors that interact with the PPRE. These results have implications for understanding the mechanisms of coordinated transcriptional induction of genes encoding peroxisomal proteins by hypolipidemic agents and other peroxisome proliferators.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1992

Inhibition of spontaneous testicular Leydig cell tumor development in F-344 rats by dehydroepiandrosterone.

The incidence of spontaneous Leydig cell tumors of testis is very high in old F-344 rats. We have examined the effect of dehydroepiandrosterone (DHEA), a steroid hormone with antimitotic and anticarcinogenic properties, on spontaneous Leydig cell tumorigenesis. Fifteen-week-old male F-344 rats were fed a diet containing DHEA (0.45% w/w) for 84 weeks. At the termination of experiment none of the 13 rats had Leydig cell hyperplasia or Leydig cell tumors. All the eight control rats of comparable age had Leydig cell tumors. These findings suggest that DHEA is a potent inhibitor of spontaneous Leydig cell tumors of testis in aged rats.

Aging↗
PubMed1992

Rat urate oxidase produced by recombinant baculovirus expression: formation of peroxisome crystalloid core-like structures.

Urate oxidase (EC 1.7.3.3), which catalyzes the oxidation of uric acid to allantoin, is present in most mammals but absent in humans and hominoid primates. In rats and most other mammals that catabolize uric acid to allantoin, this enzyme is localized within the crystalloid cores of peroxisomes present in liver parenchymal cells. To determine whether urate oxidase forms these crystalloid cores or whether core-forming protein(s) exist in association with urate oxidase, a baculovirus expression vector system was used to overproduce the full-length rat urate oxidase in Spodoptera frugiperda cells. Urate oxidase was expressed to a level of approximately 30% of the total protein in this system. Immunoblot analysis demonstrated that the baculovirus-generated protein had electrophoretic and immunologic properties similar to those of urate oxidase expressed in rat liver. Immunofluorescence and electron microscopic examination revealed that the overexpressed recombinant urate oxidase is present in both the cytoplasm and the nucleus of infected insect cells as numerous 1- to 3-microns discrete particles. These insoluble protein aggregates, which were positively stained for urate oxidase by protein A-gold immunocytochemical approach, did not appear to be delimited by a single membrane. They revealed a crystalloid structure reminiscent of rat peroxisomal core consisting of bundles of tubules with an inner diameter of approximately 50 A. The recombinant urate oxidase particles, isolated by a single-step procedure, were composed entirely of 35-kDa urate oxidase subunit. These studies indicate that rat urate oxidase is capable of forming insoluble crystalloid core-like structures.

Animals↗
PubMed1992

Hepatocarcinogenicity of dehydroepiandrosterone in the rat.

Dehydroepiandrosterone, a major secretory steroid hormone of the human adrenal gland, possesses mitoinhibitory and anticarcinogenic properties. It also induces peroxisome proliferation in the livers of rats and mice. Because peroxisome proliferators exhibit hepatocarcinogenic potential, it is necessary to examine the long term hepatic effects of dehydroepiandrosterone since this hormone is contemplated for use as a potential cancer chemopreventive agent in humans. Dehydroepiandrosterone was administered in the diet at a concentration of 0.45% to F-344 rats for up to 84 weeks. At the termination of the experiment, 14 of 16 rats developed hepatocellular carcinomas. Liver tumors induced by dehydroepiandrosterone lacked gamma-glutamyl transpeptidase and glutathione S-transferase (placental form); these phenotypic properties are identical to the features exhibited by liver tumors induced by other peroxisome proliferators. Dehydroepiandrosterone was also shown to markedly inhibit liver cell [3H]thymidine labeling indices, suggesting that cell proliferation is not a critical feature in liver tumor development with this agent. These results show that although dehydroepiandrosterone exerts anticarcinogenic effects in a variety of tissues, the peroxisome-proliferative property makes it a hepatocarcinogen.

Animals↗
PubMed1992

Loss of peroxisomal enzyme expression in preneoplastic and neoplastic lesions induced by peroxisome proliferators in rat livers.

Immunohistochemical staining of enoyl CoA hydratase (ECH), a key peroxisomal enzyme, revealed that the putative preneoplastic lesions induced in livers by administration of the peroxisome proliferator (PP) clofibrate (0.3% in diet) to rats for 60 weeks or more, lacked this enzyme so that they could be detected as ECH-negative foci. ECH and other peroxisomal enzymes such as acyl CoA oxidase, catalase and carnitine-dependent acetyltransferase were also either not or only weakly expressed in most hepatic hyperplastic nodules and hepatomas induced by ciprofibrate (0.025% in diet), Wy-14,643 (0.1%) or BR-931 (0.2%), while being strongly induced in surrounding hepatocytes. These results indicate that the expression of ECH and other peroxisomal enzymes is repressed in putative preneoplastic and neoplastic lesions induced by PPs in rat livers and that these peroxisomal enzymes might therefore be used as negative markers.

Animals↗
PubMed1992

Phenotypic properties of liver tumors induced by dehydroepiandrosterone in F-344 rats.

Dehydroepiandrosterone (DHEA), a C19 adrenal steroid hormone, induces peroxisome proliferation in liver cells and is hepatocarcinogenic in the rat. The present study deals with the phenotypic properties of DHEA-induced liver lesions. A majority of the altered areas (80-87%), neoplastic nodules (> 94%) and hepatocellular carcinomas (HCC, 80-100%) lacked the marker enzymes gamma-glutamyltranspeptidase and placental form of glutathione S-transferase (GSTP). Northern blot analysis of HCC from 4 rats revealed no detectable GSTP mRNA. These HCC, however, showed a marked decrease in the staining of glucose-6-phosphatase and adenosine triphosphatase. These results indicate that the phenotypic properties of liver tumors induced by DHEA and amphipathic carboxylate peroxisome proliferators are similar.

Animals↗
PubMed1992

Dehydroepiandrosterone-induced peroxisome proliferation in the rat liver.

We have investigated the effect of dehydroepiandrosterone (DHEA), a naturally occurring steroid hormone, on hepatic peroxisomes in the rat. Dietary administration of DHEA at a concentration of 0.45% for 2 weeks resulted in a 200% increase in the liver weight and a more than 5.0-fold increase in the volume density of peroxisomes. SDS-polyacrylamide gel electrophoresis of the postnuclear fraction of livers showed a marked increase in the 80,000-molecular weight polypeptide. Northern blot analysis of total RNA showed 2- and 12-fold increases, respectively, in the levels of catalase and peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase bifunctional enzyme (PBE) mRNA. Administration of a single large dose of DHEA led to a 10- to 40-fold increase in PBE mRNA levels by 24 h. These changes are qualitatively similar to those caused by a variety of structurally different peroxisome proliferators.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1991

Molecular evolution of the urate oxidase-encoding gene in hominoid primates: nonsense mutations.

Nucleotide sequences of portions of second and fifth exons of urate oxidase encoding gene (UOX) of chimpanzee, gorilla, orangutan, rhesus monkey and squirrel monkey obtained following amplification by polymerase chain reaction have been compared with corresponding sequences of human, baboon and rat UOX. Two or more nonsense mutations are found in the coding regions of this UOX gene thus far analyzed in human, chimpanzee, gorilla and orangutan, but not in the baboon, rhesus monkey and squirrel monkey. Of these nonsense mutations, the stop codon at amino acid position 33 is constant in the human and the three great apes suggesting that this may be the original mutation responsible for the inactivation of the UOX gene during hominoid evolution.

Amino Acid Sequence↗
PubMed1991

Rat urate oxidase: cloning and structural analysis of the gene and 5'-flanking region.

The structural gene (UOX) encoding rat urate oxidase (UOX) spans at least 23 kb and is composed of eight exons and seven introns. All of the exon-intron splice junction sequences conformed to the GT/AG consensus established for eukaryotic genes. The transcription start point (tsp) was determined using S1-type nuclease protection riboprobe, and assigned to an adenine 54 nucleotides (nt) upstream of the ATG start codon. A 456-bp 5'-terminal fragment, starting at the ATG codon, carries a putative TATA (ATAAAA) sequence at -32, and two putative 'CAAT box' sequences at -62 and -71 bp upstream from the tsp. No sequence resembling 'GC' box hexanucleotides (GGGCGG or CCGCCC) was found. The structural features of the 5'-flanking region of the UOX gene are distinct from the 5'-flanking sequences of peroxisomal beta-oxidation system genes which contain one or more 'GC' box elements but lack TATA- and CAAT-like features [Osumi et al., J. Biol. Chem. 262 (1987) 8138-8143; Ishii et al., J. Biol. Chem. 262 (1987) 8144-8150]. The 5'-flanking region of the UOX gene reveals a sequence, TTAGTAATT at nt -276 from the tsp, which appears to be complementary to the underlined part of the liver-specific LF-B1/HNF-1 consensus sequence, GTTAATNATTAAC (where N = A, C, T, G or no nt).

Amino Acid Sequence↗
PubMed1991

Pancreatic hepatocytes. An in vivo model for cell lineage in pancreas of adult rat.

Multiple foci of hepatocytes differentiate in the pancreas of adult rats subjected to a copper depletion-repletion regimen. Copper deficiency for seven to nine weeks causes an irreversible depletion of over 80% of the acinar cells in the pancreas. When transferred to a normal diet, these rats exhibit only a minimal and spotty acinar cell recovery. This disruption of tissue organization appears to trigger a profound change in cellular commitment, which leads to hepatocyte differentiation in the "oval cells" in the periductal interstitium and the epithelial cells lining the small pancreatic ductules. Pancreatic hepatocytes express several liver-specific genes including albumin, a2u-globulin, carbamoylphosphate synthetase-I, and urate oxidase. Both carbamoylphosphate synthetase-I and glutamine synthetase, the ammonia-metabolizing enzymes, are expressed by all pancreatic hepatocytes; in liver, these are expressed by different populations of hepatocytes. The magnitude of hepatocyte differentiation in this model should facilitate studies on the molecular events regulating changes in cell lineage or differentiation commitment within the pancreas.

Animals↗
PubMed1991

Induction of rat liver DNA alterations by chronic administration of peroxisome proliferators as detected by 32P-postlabeling.

The mechanisms of the hepatocarcinogenicity of non-mutagenic peroxisome proliferators, i.e. compounds used as hypolipidemic drugs and industrial plasticizers, are not sufficiently understood. To gain more information on the mechanism of their action, the chronic effects of two structurally diverse peroxisome proliferators on rat-liver DNA were investigated by the 32P-postlabeling assay. Male F-344 rats (1.5 month old) were fed ciprofibrate (0.025%) in the diet for 2, 5, 8, and 16 months or Wy-14643 (0.1%) for 18 months. Liver DNA from individual treated animals (3-4 per group) and age-matched controls was analyzed by the nuclease P1/bisphosphate version of the 32P-postlabeling assay. Three distinct types of exposure-related DNA alterations were observed: (i) A significant reduction of the age-dependent accumulation of I-compounds (putative indigenous DNA modifications) (type 1), (ii) adduct-like DNA derivatives induced by the treatments (type 2), and (iii) as yet structurally uncharacterized radiolabeled material occupying substantial areas of DNA adduct maps and accumulating in an exposure time-dependent manner (type 3). DNA from liver tumors generated by these agents displayed only traces of I-compounds, lacked all but one adduct-like derivatives, and had no type 3 alterations. Thus, in contrast to the non-mutagenicity of peroxisome proliferators in short-term assays, chronic administration of these compounds led to DNA alterations that were detectable by 32P-postlabeling assay.

Affinity Labels↗
PubMed1991

Expression and compartmentalization of integral plasma membrane proteins by hepatocytes and their progenitors in the rat pancreas.

A combination of Western blotting, Northern blotting and immunofluorescence was used to examine the expression and compartmentalization of plasma membrane proteins by those hepatocyte-like cells that arise in the pancreases of rats subjected to sequential dietary copper depletion and repletion. The pancreatic hepatocytes were found to: (1) express several integral membrane proteins known to be concentrated within the apical, lateral or basolateral domains of the plasma membranes of hepatocytes in liver; and (2) compartmentalize the membrane proteins to equivalent plasma membrane domains, despite the organization of these cells into clusters instead of highly vascularized plates. The apical plasma membrane proteins dipeptidylpeptidase IV and HA 4 were found to line bile canaliculus-like openings between adjacent pancreatic hepatocytes; these openings were shown to be continuous with the pancreatic exocrine duct by India ink infusion. In contrast, the basolateral plasma membrane protein rat hepatic lectin-1 and lateral plasma membrane protein HA 321 were detected elsewhere about the surfaces of the pancreatic hepatocytes: by analogy to their respective localizations on hepatocytes in liver, rat hepatic lectin-1 was concentrated on those surfaces exposed to the pancreatic matrix at the periphery of the hepatocyte clusters (the basal surface equivalent), whereas HA 321 was concentrated on those surfaces exposed to adjacent hepatocytes within the clusters. The hepatocyte plasma membrane proteins were found to be expressed in the pancreas at different times during the copper depletion/repletion protocol: for example, rat hepatic lectin-1 and the bulk of the HA 4 were expressed relatively late in the protocol, only after large numbers of pancreatic hepatocytes had appeared; whereas dipeptidylpeptidase IV was induced greater than 10-fold early in the protocol and proved to be an apical-specific marker for those ductular epithelial cells that are believed to be the progenitors of the pancreatic hepatocytes.

Animals↗
PubMed1991

An overview of peroxisome proliferator-induced hepatocarcinogenesis.

Peroxisome proliferators are hepatocarcinogens in rats and mice. Chronic administration of these compounds results in the development of altered areas and neoplastic nodules followed by hepatocellular carcinomas. All three types of hepatic lesions do not express gamma-glutamyltranspeptidase, glutathione 8-transferase-P, and alpha-fetoprotein and are resistant to iron accumulation after overload. The mechanism by which nongenotoxic peroxisome proliferators induce hepatic tumors is not well understood. It has been proposed that with continuous administration of peroxisome proliferators, liver cells are subjected to persistent oxidative stress resulting from marked proliferation of peroxisomes and a differential increase in the levels of H2O2 producing (20- to 30-fold) and degrading (2-fold) enzymes. Free oxygen radicals lead to DNA damage (both directly and through lipid peroxidation) and thus may cause initiation and promotion of the carcinogenic process.

Animals↗
PubMed1991

A comparative study of the effects of clofibrate, ciprofibrate, WY-14,643, and di-(2-ethylhexyl)-phthalate on liver protein expression in mice.

Chemical, physical, or mechanical insults to a cellular system cause changes in the rates of protein synthesis or degradation. To define patterns of quantitative protein changes and identify the biochemical basis of those changes, we are studying fluctuations in mouse liver protein abundance in response to factors such as stages of maturation, changes in diet, and exposure to chemicals. Two-dimensional gel electrophoresis (2DE) together with computer-assisted data analysis is used to collect quantitative data from groups of 2DE patterns. Our analysis of liver protein expression in mice treated with four different chemicals known to cause peroxisome proliferation serves as an example of the changes in protein expression that can be monitored by using this approach. The database we are building indicates that changes specific to a particular treatment as well as generalized effects can be identified, providing the opportunity to identify patterns of protein expression that are indicators of biological response to different types of cellular insults.

Animals↗