Biomedical subjects

P A Permana

Publications and source records attributed to P A Permana.

14 recordsLinked to original sources

PubMed2002

Microarray profiling of skeletal muscle tissues from equally obese, non-diabetic insulin-sensitive and insulin-resistant Pima Indians.

AIMS/HYPOTHESIS: We carried out global transcript profiling to identify differentially expressed skeletal muscle genes in insulin resistance, a major risk factor for Type II (non-insulin-dependent) diabetes mellitus. This approach also complemented the ongoing genomic linkage analyses to identify genes linked to insulin resistance and diabetes in Pima Indians. METHODS: We compared gene expression profiles of skeletal muscle tissues from 18 insulin-sensitive versus 17 insulin-resistant equally obese, non-diabetic Pima Indians using oligonucleotide arrays consisting of about 40,600 transcripts of known genes and expressed sequence tags, and analysed the results with the Wilcoxon rank sum test. We verified the mRNA expression of ten differentially (best-ranked) and ten similarly (worst-ranked) genes using quantitative Real Time PCR. RESULTS: There were 185 differentially expressed transcripts by the rank sum test. The differential expressions of two out of the ten best-ranked genes were confirmed and the similar expressions of all ten worst-ranked genes were reproduced. CONCLUSION/INTERPRETATION: Of the 185 differentially expressed transcripts, 20 per cent were true positives and some could generate new hypotheses about the aetiology or pathophysiology of insulin resistance. Furthermore, differentially expressed genes in chromosomal regions with linkage to diabetes and insulin resistance serve as new diabetes susceptibility genes.

Adult↗

PubMed2001

Reduced skeletal muscle calpain-10 transcript level is due to a cumulative decrease in major isoforms.

The DNA polymorphism SNP-43 in the calpain-10 gene is associated with insulin resistance and reduced skeletal muscle transcript in Pima Indians. Alternative splicing generates transcript isoforms calpain-10a to -10h. We determined the contribution of calpain-10 mRNA isoforms to the decreased total skeletal muscle calpain-10 mRNA levels observed in the G/G homozygotes. The expression levels of the major isoforms, calpain-10a and -10f, were positively correlated with the total calpain-10 mRNA levels, indicating a cumulative effect.

Calpain↗

PubMed2000

Genomic structure and expression of human KCNJ9 (Kir3.3/GIRK3).

The human KCNJ9 (Kir 3.3, GIRK3) is a member of the G-protein-activated inwardly rectifying potassium (GIRK) channel family. Here we describe the genomic organization of the KCNJ9 locus on chromosome 1q21-23 as a candidate gene for Type II diabetes mellitus in the Pima Indian population. The gene spans approximately 7.6 kb and contains one noncoding and two coding exons separated by approximately 2.2 and approximately 2.6 kb introns, respectively. We identified 14 single nucleotide polymorphisms (SNPs), including one that predicts a Val366Ala substitution, and an 8 base-pair (bp) insertion/deletion. Our expression studies revealed the presence of the transcript in various human tissues including pancreas, and two major insulin-responsive tissues: fat and skeletal muscle. The characterization of the KCNJ9 gene should facilitate further studies on the function of the KCNJ9 protein and allow evaluation of the potential role of the locus in Type II diabetes.

3' Untranslated Regions↗

PubMed2000

Functional analyses of amino acid substitutions Arg883Ser and Asp905Tyr of protein phosphatase-1 G-subunit.

The PPP1R3 gene encoding the G-subunit of protein phosphatase-1 has three polymorphisms in linkage disequilibrium in the Pima Indians: an mRNA-destabilizing element in the 3'-untranslated region (ARE1/ARE2 alleles), Arg883Ser, and Asp905Tyr substitutions. The ARE2 allele, Arg883, and Asp905 variants are associated with insulin resistance and higher prevalence of type 2 diabetes in the Pima Indians. The ARE2 allele is associated with lower PPP1R3 transcript and protein levels in muscle tissue. Here we determined the functional contribution of the amino acid substitutions independent of the ARE alleles to insulin-stimulated glycogen synthesis by adenoviral-mediated gene expression in L6 myotubes. Similar overexpression levels of the G-subunit variants increased glycogen synthase fractional activity in the presence ( approximately 1. 5-fold) of insulin compared to control myotubes transduced with adenovirus encoding beta-galactosidase. The glycogen synthesis rate of myotubes overexpressing the G-subunit variants also increased by approximately 1.7-fold over the control with and without insulin. However, these measures were not significantly different among the variants. This study does not support a role for Arg883 and Asp905 variants independent of the ARE2 allele in the impaired insulin-stimulated glycogen synthesis in the muscle of Pima Indians.

Adenoviridae↗

PubMed2000

A calpain-10 gene polymorphism is associated with reduced muscle mRNA levels and insulin resistance.

Previous linkage studies in Mexican-Americans localized a major susceptibility locus for type 2 diabetes, NIDDM1, to chromosome 2q. This evidence for linkage to type 2 diabetes was recently found to be associated with a common G-->A polymorphism (UCSNP-43) within the CAPN10 gene. The at-risk genotype was homozygous for the UCSNP-43 G allele. In the present study among Pima Indians, the UCSNP-43 G/G genotype was not associated with an increased prevalence of type 2 diabetes. However, Pima Indians with normal glucose tolerance, who have a G/G genotype at UCSNP-43, were found to have decreased rates of postabsorptive and insulin-stimulated glucose turnover that appear to result from decreased rates of glucose oxidation. In addition, G/G homozygotes were found to have reduced CAPN10 mRNA expression in their skeletal muscle. A decreased rate of insulin-mediated glucose turnover, or insulin resistance, is one mechanism by which the polymorphism in CAPN10 may increase susceptibility to type 2 diabetes mellitus in older persons.

Adolescent↗

PubMed2000

Mutations in the genes for hepatocyte nuclear factor (HNF)-1alpha, -4alpha, -1beta, and -3beta; the dimerization cofactor of HNF-1; and insulin promoter factor 1 are not common causes of early-onset type 2 diabetes in Pima Indians.

OBJECTIVE: Maturity-onset diabetes of the young (MODY) is a genetically heterogeneous subtype of type 2 diabetes characterized by an early age at onset and autosomal dominant inheritance. MODY can result from heterozygous mutations in at least five genes. The purpose of this study was to determine whether alterations in known MODY genes and two MODY candidate genes contribute to the development of early-onset type 2 diabetes in Pima Indians. RESEARCH DESIGN AND METHODS: The coding regions of the known MODY genes hepatocyte nuclear factor (HNF)-1alpha, HNF-4alpha, HNF-1beta, and insulin promoter factor 1 and the coding regions of two MODY candidate genes, HNF-3beta and the dimerization cofactor of HNF-1, were sequenced in genomic DNA from Pima Indians. The primary "affected" study population consisted of 46 Pima Indians whose age at onset of type 2 diabetes was < or =20 years. DNA sequence variants identified in the affected group were then analyzed in a group of 80 "unaffected" Pima Indians who were at least 40 years old and had normal glucose tolerance. RESULTS: A total of 11 polymorphisms were detected in these genes. However, none of the polymorphisms differed in frequency among Pima Indians with an early age at onset of diabetes compared with older Pima Indians with normal glucose tolerance. CONCLUSIONS: Mutations in these known MODY or MODY candidate genes are not a common cause of early-onset diabetes in Pima Indians.

Adolescent↗

PubMed1999

Protein targeting to glycogen/PPP1R5: screening of coding and flanking genomic regions for polymorphisms and association analysis with insulin action in Pima Indians.

Insulin resistance, a major predictor of type 2 diabetes mellitus, is genetically inherited in Pima Indians, a population with a high prevalence of the metabolically complex disease. Protein targeting to glycogen/PPP1R5 has recently been identified as a potential regulator of glycogen synthase, the rate-limiting enzyme of the insulin-induced glycogenesis. The gene is located on chromosome 10q23-24, where there is a suggestive linkage to insulin action in this population, establishing it as a functional and positional candidate gene. In this study, we discovered 2 novel polymorphisms upstream of the 5'UTR of the gene, with only one found in Pima Indians, but no polymorphism in the coding sequence. The genotype frequencies of the polymorphism and transcript levels of the gene in skeletal muscle do not correlate with insulin action in the subjects. These results exclude any significant role of protein targeting to glycogen/PPP1R5 in insulin resistance in Pima Indians.

5' Untranslated Regions↗

PubMed1997

Genetic analysis of human type 1 protein phosphatase inhibitor 2 in insulin-resistant Pima Indians.

The rate-limiting enzyme in insulin-mediated nonoxidative glucose disposal, glycogen synthase, has reduced activity in insulin-resistant subjects at risk for developing non-insulin-dependent diabetes mellitus (NIDDM). The synthase-activating enzyme, type 1 protein phosphatase (PP1), also has an abnormally low level of activity. Inhibitor 2 (I-2) reversibly inhibits and facilitates the proper conformation of free catalytic subunits of PP1. This study investigates whether genetic alteration(s) in the I-2 coding locus (PPP1R2) could contribute to insulin resistance in Pima Indians. We determined that the authentic PPP1R2 gene is located on chromosome 3q29 and consists of six exons. The previously reported homologue of PPP1R2 on chromosome 5 is identified as an intronless pseudogene. Comparative sequencing of PPP1R2 exons and splice junctions revealed no mutations in insulin-resistant Pima Indians. The information on the genomic structure of PPP1R2 is important for exploring this gene as a potential candidate contributing to insulin resistance and NIDDM in other populations.

Amino Acid Sequence↗

PubMed1994

Quinobenoxazines: a class of novel antitumor quinolones and potent mammalian DNA topoisomerase II catalytic inhibitors.

The antineoplastic quinobenoxazines A-62176 and A-74932 were shown to be potent inhibitors of mammalian DNA topoisomerase II in vivo. This was demonstrated by their selective inhibition of the SV40 DNA replication stages that require topoisomerase II. Neither drug stabilized a covalent complex of the enzyme with SV40 DNA, which suggests that they are not poisons of DNA topoisomerase II. A-77601, an analog having little antitumor activity, barely inhibited DNA topoisomerase II in vivo, even at high concentrations. These findings were supported by in vitro studies which showed that A-62176 and A-74932, but not A-77601, strongly inhibited the catalytic activity of mammalian DNA topoisomerase II. A-62176 did not cause topoisomerase II-mediated DNA strand breaks in vitro under conditions in which adriamycin produced extensive DNA breakage. The antineoplastic and topoisomerase inhibitory activities of the quinobenoxazines correlate with their ability to unwind DNA. A-62176 antagonized the poisoning of topoisomerase II by VM-26 in vivo and in vitro, but had no effect on DNA breakage induced by camptothecin, a DNA topoisomerase I poison. A-62176 and A-74932 thus inhibit DNA topoisomerase II reactions at a step prior to the formation of the "cleavable complex" intermediate. These findings indicate that stabilization of the DNA topoisomerase II-DNA cleavable complex is not necessary for the antitumor activity of this class of quinolones and that the catalytic inhibition of DNA topoisomerase II may contribute significantly to the anticancer activity of other DNA topoisomerase II inhibitors.

Animals↗

PubMed1994

Inverse relationship between catenation and superhelicity in newly replicated simian virus 40 daughter chromosomes.

High resolution gel electrophoresis was used to demonstrate an inverse relationship between catenation linking number and superhelicity in highly catenated simian virus 40 chromosomes caused by exposure to the topoisomerase II inhibitor ICRF-193. Since ICRF-193 does not unwind DNA, we conclude that the decreased superhelicity in catenated SV40 daughter chromosomes is a direct result of increased catenation. It is likely that catenation decreases superhelicity by interfering with the formation of nucleosomes. The absence of normal chromatin structure in regions of catenation may facilitate access to topoisomerase II under normal conditions. ICRF-193 does not prevent initiation of SV40 DNA replication.

Animals↗

PubMed1994

Mechanism of action of the antileukemic xanthone psorospermin: DNA strand breaks, abasic sites, and protein-DNA cross-links.

Psorospermin, a cytotoxic dihydrofuranoxanthone isolated from Psorospermum febrifugum, produced aberrant simian virus 40 DNA replication intermediates when added to lytically infected CV-1 monkey kidney cells. The aberrant viral intermediates showed dose-dependent DNA strand breaks and protein-DNA cross-links, as well as decreased electrophoretic mobility. Simian virus 40 DNA from psorospermin-treated cells was shown to contain numerous abasic (apyrimidinic/apurinic) sites. The density of abasic sites was a function of the psorospermin dose. We conclude that psorospermin causes extensive loss of DNA bases in vivo. Primary amine groups of cellular proteins are known to react with abasic sites to form covalent protein-DNA cross-links and DNA strand breaks. Cytochrome c cross-linked spontaneously to viral DNA prepared from psorospermin-treated cells but not to DNA from untreated cells. This suggests that the protein-DNA cross-links and many of the DNA strand breaks observed in vivo result from reactions between abasic sites and chromosomal proteins. It is likely that the protein-DNA cross-links and DNA strand breaks contribute to the cytotoxicity and antineoplastic activity of psorospermin.

Aldehydes↗

PubMed1994

Aldehyde-induced protein-DNA crosslinks disrupt specific stages of SV40 DNA replication.

Aldehydes with specific protein-DNA crosslinking ability disrupted simian virus 40 (SV40) DNA replication to cause replication fork failure by the 40S intermediate pathway, in which replicating viral genomes become inactivated and torsionally stressed. In contrast, aldehydes without detectable protein-DNA crosslinking ability had no effect on SV40 DNA replication during the 10 min exposure times employed. This indicates that protein-DNA crosslinks block either DNA polymerase or the entire replication complex. Replication failure by the 40S pathway is known to initiate recombinational events in the damaged SV40 replicons. Similar events in cellular replicons may play a role in the clastogenic effects of formaldehyde. In addition, formaldehyde and acrolein caused accumulation of catenated (topologically linked) SV40 daughter chromosomes--a signature of topoisomerase II inhibition.

Acrolein↗

PubMed1993

SV40 DNA replication intermediates: analysis of drugs which target mammalian DNA replication.

The simian virus 40 chromosome, a model for the mammalian replicon, is a uniquely powerful system for the study of drugs and treatments which target enzymes of the mammalian replication apparatus. High resolution gel electrophoretic analysis of normal and aberrant viral replication intermediates can be used effectively to understand the molecular events of replication failure. These events include breakage of replication forks, aberrant topoisomerase action, failure to separate daughter chromosomes, protein-DNA crosslinking, single and double strand DNA breakage, alterations in topology and inactivation of replication intermediates. The SV40 replication system can also be used to study the recombinational events which often follow drug-induced replication failure.

Animals↗

PubMed1991

Aphidicolin-induced topological and recombinational events in simian virus 40.

Highly compacted (40S) SV40 DNA replication intermediates formed in vivo during aphidicolin exposure and immediately broke down in two stages. In the rapid initial stage, single strand DNA breaks caused loss of superhelicity in the 40S replication intermediates. This DNA breakage was accompanied by the formation of strong, permanent protein-DNA crosslinks which reached a maximum as nicking of the aberrant DNA replication intermediates was completed. These protein-associated DNA strand breaks were not repaired. In the slower second stage of breakdown, the aberrant DNA replication intermediates remained nicked and strongly associated with protein as they underwent DNA replication fork breakage and recombinational changes to produce high molecular weight forms.

Aphidicolin↗