Dehydrogenase Gene Expression Research Articles

Inhibitory effect of novel red ginseng treated with high temperature and high pressure on oxidative damage

Ginseng is one of the most commonly used traditional herbal medicines. This study was to investigate the inhibitory effect of novel red ginseng treated with high temperature and high pressure in oxidative stress‐induced mouse by high intensity exercise. Experimental groups were no exercise group(CON), high‐intensity exercise group(E), common red ginseng extract supplementation with high‐intensity exercise group(CGE), novel ginseng red extract supplementation with high‐intensity exercise group(NGE). High‐intensity exercise was performed by treadmill running for two weeks. After the experimental period, changes of glutamic pyruvic transaminase(GPT) and glutamic oxaloacetic transaminase(GOT) in serum was measured, and MDA levels, glycogen contents, glucose‐6‐phosphate dehydrogenase(G6PDH), lactate dehydrogenase(LDH) activities and gene expressions of Cu/Zn SOD and myogenic factor4(MRF4) on tissue were analyzed. Our results show that the levels of GOT, GPT and MDA in NGE group were significantly lower than those of other groups. Moreover, the levels of muscular glycogen, G6PDH activity, LDH activity of NGE group were markedly higher than those of other groups. In the case of gene expression on muscle tissue, CGE or NGE groups led to up regulation of SOD and MRF4 gene expression. These results indicate that NGE can inhibit high‐intensity induced oxidative stress and enhance exercise capacity.

Gene expression and function involved in polyol biosynthesis of Trichosporonoides megachiliensis under hyper-osmotic stress

Among three erythritol reductase isogenes (er1, er2, and er3) in Trichosporonoides megachiliensis SN-124A, er1 and er2 each had one stress response element (STRE) approximately 2kbp upstream of their respective initiator codon; in contrast, er3 had two STREs, 148 and 40bp upstream from the initiator codon. Based on intracellular erythritol accumulation and gene expression profiles, er3 seemed to be highly responsive to stress than er1 or er2. Under hyper-osmotic conditions, intracellular glycerol production, increased significantly within 1.5h together with glycerol-3-phosphate dehydrogenase gene (gpd1) expression; in contrast, neither er gene expression nor the corresponding production of intracellular erythritol increased significantly within the first 1.5h of hyper-osmotic culture. However, within 24h of hyper-osmotic culture, erythritol production and er3 gene expression increased significantly and in parallel. Thus, we concluded that, as an initial response to hyper-osmotic growth conditions, T.megachiliensis produces glycerol as an osmoregulatory compatible solute via GPD; however, within 24h, it begins to produce erythritol, mainly via ER3, as the preferred compatible solute. Heterologous expression of ers in a Saccharomyces cerevisiae mutant indicated that any of three ers might not function in S.cerevisiae for erythritol biosynthesis in spite of ers and corresponding ERs expression. Hence, although er is annotated as a galactose-inducible crystalline-like yeast protein gene (gcy1) homolog, er may be functionally different from gcy1 in glycolytic metabolism. Otherwise, S.cerevisiae is not likely to produce erythrose, the substrate of erythrose reductase due to metabolic characteristics.

Monitoring of Inosine Monophosphate Dehydrogenase Activity and Expression during the Early Period of Mycophenolate Mofetil Therapy in De Novo Renal Transplant Patients

Measurement of inosine-monophosphate dehydrogenase (IMPDH) activity or gene expression was used as a further approach in pharmacokinetics (PK)/pharmacodynamic (PD)-guided mycophenolate mofetil (MMF) therapy. Forty-four de novo kidney transplant patients were enrolled; 35 of these completed the study, and were followed for 24 weeks for clinical status, PK parameters, IMPDH activity and IMPDH1/2 gene expression. IMPDH activity and expression were measured in peripheral blood mononuclear cells before transplant and at week 2,4,12 and 24, drawn before (t0) and 2 h (t2 h) after MMF administration. No significant correlation was found between IMPDH activity/expression and PK parameters. For both genes, significant enhancement in t2 h expression was observed, then decreases towards week 24 with a trend following steroid dosages. Seven patients experienced acute rejection (AR) and exhibited significantly higher pre-transplant expression of both IMPDH1 (median 3.42 vs. 0.84; p=0.0025), and IMPDH2 genes (135 vs. 104; p=0.0218) with respect to non-rejecting patients. A significant association was also found between pre-transplant IMPDH1 mRNA and haematological complications (p=0.032). This study suggests that high steroid dosages may influence IMPDH1/2 expression, hampering their use as a PD biomarker, particularly during the early post-transplant period. The measurement of pre-transplant levels of IMPDH1/2 may contribute to prediction of individual drug responsiveness to improve the clinical management of patients in MMF therapy.

Association Between Cytosolic Glycerol 3-Phosphate Dehydrogenase Gene Expression in Human Subcutaneous Adipose Tissue and BMI

Background/Aims: Cytosolic glycerol 3-phosphate dehydrogenase (cGPDH) is a key enzyme providing glycerol 3-phosphate for triacylglycerol synthesis in adipose tissue and is regarded as a marker for adipocyte differentiation. The aim of this study was to test the hypothesis that an increase in cGPDH gene expression in subcutaneous adipose tissue is associated with obesity. Methods: mRNA levels in human subcutaneous adipose tissue were analysed by Real-Time PCR. Results: We found that human subcutaneous adipose tissue cGPDH activity and cGPDH mRNA level were greater in obese patients than in lean subjects and were positively correlated with BMI and fat mass. Moreover, a strong positive correlation between subcutaneous adipose tissue cGPDH mRNA level and cGPDH activity was found. The data presented here indicates also that PPARγ mRNA level is positively correlated with body mass index and fat mass as well as with adipose tissue cGPDH mRNA level. Moreover, the association between subcutaneous adipose tissue cGPDH mRNA level and fatty acid translocase (FAT/CD36) mRNA level was also observed. Conclusion: The obtained results suggest that in comparison to lean subjects the increase in subcutaneous adipose tissue cGPDH gene expression in the obese, is probably the result of adipose tissue expansion during obesity.

Interspecies Electron Transfer via Hydrogen and Formate Rather than Direct Electrical Connections in Cocultures of Pelobacter carbinolicus and Geobacter sulfurreducens

Direct interspecies electron transfer (DIET) is an alternative to interspecies H(2)/formate transfer as a mechanism for microbial species to cooperatively exchange electrons during syntrophic metabolism. To understand what specific properties contribute to DIET, studies were conducted with Pelobacter carbinolicus, a close relative of Geobacter metallireducens, which is capable of DIET. P. carbinolicus grew in coculture with Geobacter sulfurreducens with ethanol as the electron donor and fumarate as the electron acceptor, conditions under which G. sulfurreducens formed direct electrical connections with G. metallireducens. In contrast to the cell aggregation associated with DIET, P. carbinolicus and G. sulfurreducens did not aggregate. Attempts to initiate cocultures with a genetically modified strain of G. sulfurreducens incapable of both H(2) and formate utilization were unsuccessful, whereas cocultures readily grew with mutant strains capable of formate but not H(2) uptake or vice versa. The hydrogenase mutant of G. sulfurreducens compensated, in cocultures, with significantly increased formate dehydrogenase gene expression. In contrast, the transcript abundance of a hydrogenase gene was comparable in cocultures with that for the formate dehydrogenase mutant of G. sulfurreducens or the wild type, suggesting that H(2) was the primary electron carrier in the wild-type cocultures. Cocultures were also initiated with strains of G. sulfurreducens that could not produce pili or OmcS, two essential components for DIET. The finding that P. carbinolicus exchanged electrons with G. sulfurreducens via interspecies transfer of H(2)/formate rather than DIET demonstrates that not all microorganisms that can grow syntrophically are capable of DIET and that closely related microorganisms may use significantly different strategies for interspecies electron exchange.

Increased Pheromone cCF10 Expression in Enterococcus faecalis Biofilm Formed by Isolates from Renal Transplant Patients

Renal transplant recipients are at a high risk of developing infectious complications even caused by commensal bacteria. This is because of various physiological non-immunological, and immunological protective mechanisms are not fully efficient in RTx patients. Therefore, rapid and precise diagnostic tools are essential in this particular group of patients. We aimed to develop simple and sensitive protocol Flow-Fish for the study of gene expression in enterococci and to compare expression of genes involved in virulence regulation in biofilm and planktonic form of Enterococcus faecalis. Proper optimization of the method was demonstrated with analysis of dehydrogenase gene expression. According to expectation reduction of the dehydrogenase gene expression was observed in biofilm. Furthermore, expression of studied gene was higher in clinical than in commensal strains. We have also found that in contrast to dehydrogenase gene, pheromone cCF10 gene expression increasing then clinical strains formed biofilm.

Vitamin D3 enhances ATRA-mediated neurosteroid biosynthesis in human glioma GI-1 cells

Emerging evidence indicates that vitamin D (VD) is an important modulator of brain development and function. To investigate whether VD modulates neurosteroid biosynthesis in neural cells, we investigated the effect of VD(3) on steroidogenic gene expression in human glioma GI-1 cells. We found that VD(3) enhanced CYP11A1 and 3β-hydroxysteroid dehydrogenase gene expression. The induction of CYP11A1 gene expression by VD(3) was dose- and incubation time-dependent. Calcipotriol, a VD(3) receptor (VDR) agonist, also induced CYP11A1 gene expression in GI-1 cells, indicating that VDR is involved in this induction. The induction of progesterone (PROG) de novo synthesis was observed along with the induction of steroidogenic genes by VD(3). Furthermore, VD(3) enhanced all-trans retinoic acid (ATRA)-induced CYP11A1 gene expression and PROG production. This suggests cooperative regulation of steroidogenic gene expression by the two fat-soluble vitamins, A and D. In addition, a mixed culture of neuronal IMR-32 cells and GI-1 cells treated with ATRA and VD(3) resulted in the induction of PROG-responsive gene expression in the IMR-32 cells. This result shows a paracrine action of PROG that is induced in and released by the GI-1 cells. The relationship between neurological dysfunction associated with VD deficiency and neurosteroid induction by VD is discussed.

Taraxerone enhances alcohol oxidation via increases of alcohol dehyderogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activities and gene expressions

The present study, taraxerone (d-friedoolean-14-en-3-one) was isolated from Sedum sarmentosum with purity 96.383%, and its enhancing effects on alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activities were determined: EC50 values were 512.42±3.12 and 500.16±3.23μM for ADH and ALDH, respectively. In order to obtain more information on taraxerone related with the alcohol metabolism, 40% ethanol (5mL/kg body weight) with 0.5–1mM of taraxerone were administered to mice. The plasma alcohol and acetaldehyde concentrations of taraxerone-treated groups were significantly lowered than those of the control group (p<0.01): approximately 20–67% and 7–57% lowered for plasma alcohol and acetaldehyde, respectively. Compare to the control group, the ADH and ALDH expressions in the liver tissues were abruptly increased in the taraxerone-treated groups after ethanol exposure. In addition, taraxerone prevented catalase, superoxide dismutase, and reduced glutathione concentrations from the decrease induced by ethanol administration with the concentration dependent manner.

123 A FUNCTIONAL LINK BETWEEN VITAMIN D RECEPTOR AND JAK-STAT SIGNALING RESULTS IN SYNERGISTIC HEPATOCELLULAR RESPONSE TO CALCITRIOL AND INTERFERON-α

Background and Aims: The Hepatitis C Virus (HCV) interacts with lipid particles (such as LDL receptor) to entry the cell, circulating as lipo-viral particles and being secreted as VLDL. On the other hand, serum LDL is a predictive response factor in triple theraphy (Telaprevir+peginterferon+ribavirin). LDLr is a glycoprotein membrane that controls the entry of cholesterol into the cells. The JFH-1 transfection of Huh7.5 cells modulates LDLr gene expression. The aim of this study was to investigate the interaction between HCV infection (genotype 1) and LDLr gene expression in vivo. Methods: Blood samples from patients with HCV genotype 1 (n =17) were obtained and peripheral blood mononuclear cells (PBMC) isolated. Patients included were: RNA-HCV (−) after treatment with peginterferon plus ribavirin (P+R) (SVR, n = 8) and patients with RNA-VHC(+)(Non-responders (NR), n = 9). Total RNA was isolated from PBMC and gene expression diacylglycerol acyl transferase 1(DGAT1), SREBF1, FASN, LDLr and DAGT2 genes were studied. Gene expression was analyzed by real time semiquantitative PCR using the qRT-PCR kit from Quantance (Bioline, London, UK) usin the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene expression as housekeeping. The relationship between gene expression, treatment outcome after P+R therapy and lipid profile was also analyzed. Results: DGAT1, DGAT2 genes are induced in RNA-HCV(+) patients (1.5±0.4 and 1.6±0.3 fold, respectively) compared to expression levels in RNA-HCV(−) patients(SVR). LDLr gene expression showed a marked decrease(1.6±0.4 fold) gene expression in RNA-HCV(+) patients compared to patients RNA-HCV(−). In RNA-HCV(SVR) patients, we found over-expression of Fatty Acid Synthase(FASN) and Sterol Regulatory Element-binding transcription factor 1(SREBF1) genes, both related to lipid metabolism (1.4±0.3 and 2.4±0.5 fold, respectively). LDLr gene expression was increased in patients with SVR (2.72±0.74) in comparison with NR (0.65±0.15); p < 0.0001. Conclusion: HCV infection modulates the expression levels of genes related to lipid metabolism in vivo, especially for LDLr. This fact, together with the evidence that LDL levels correlates with treatment response, strongly support the role of lipid metabolism in hepatitis C infection.

Influence of maternal metabolism and parental genetics on fetal maldevelopment in diabetic rat pregnancy

The purpose of this study was to investigate the influence of parental transgenerational genetics and maternal metabolic state on fetal maldevelopment in diabetic rat pregnancy. Rats from an inbred malformation-resistant (W) strain, and an inbred malformation-prone (L) strain, were cross-mated to produce two different F(1) hybrids, WL and LW. Normal (N) and manifestly diabetic (MD) WL and LW females were mated with normal males of the same F(1) generation to obtain WLWL and LWLW F(2) hybrids. Maternal diabetes increased malformation and resorption rates in both F(2) generations. MD-WLWL offspring had higher resorption rate but similar malformation rate compared with the MD-LWLW offspring. Malformed MD-WLWL offspring presented with 100% agnathia/micrognathia, whereas malformed MD-LWL offspring had 60% agnathia/micrognathia and 40% cleft lip and palate. The MD-WL dams showed increased β-hydroxybutyrate levels and alterations in concentrations of several amino acids (taurine, asparagine, citrulline, cystine, glutamic acid, leucine, tyrosine, and tryptophan) compared with MD-LW dams. Fetal glyceraldehyde-3-phosphate dehydrogenase (Gapdh) activity and gene expression were more altered in MD-WLWL than MD-LWLW. Fetal gene expression of reactive oxygen species (ROS) scavenger enzymes was diminished in MD-WLWL compared with MD-LWLW. Glial cell line-derived neurotrophic factor and Ret proto-oncogene gene expression was decreased in both MD-WLWL and MD-LWLW fetuses, whereas increased bone morphogenetic protein 4 and decreased Sonic hedgehog homolog expression was found only in MD-LWLW fetuses. Despite identical autosomal genotypes, the WL and LW dams gave birth to offspring with markedly different malformation patterns. Together with fetal differences in enzymatic activity and expression of Gapdh, ROS scavengers, and developmental genes, these results may suggest a teratological mechanism in diabetic pregnancy influenced by maternal metabolism and parental strain epigenetics.

11-Hydroxy-β-steroid dehydrogenase gene expression in canine adipose tissue and adipocytes: Stimulation by lipopolysaccharide and tumor necrosis factor α

The enzyme 11β-hydroxysteroid dehydrogenase 1 (11β-HSD-1) is expressed in a number of tissues in rodents and humans and is responsible for the reactivation of inert cortisone into cortisol. Its gene expression and activity are increased in white adipose tissue (WAT) from obese humans and may contribute to the adverse metabolic consequences of obesity and the metabolic syndrome. The extent to which 11β-HSD-1 contributes to adipose tissue function in dogs is unknown; the aim of the present study was to examine 11β-HSD-1 gene expression and its regulation by proinflammatory and anti-inflammatory agents in canine adipocytes. Real-time PCR was used to examine the expression of 11β-HSD-1 in canine adipose tissue and canine adipocytes differentiated in culture. The mRNA encoding 11β-HSD-1 was identified in all the major WAT depots in dogs and also in liver, kidney, and spleen. Quantification by real-time PCR showed that 11β-HSD-1 mRNA was least in perirenal and falciform depots and greatest in subcutaneous, omental, and gonadal depots. Greater expression was seen in the omental depot in female than in male dogs ( P = 0.05). Gene expression for 11β-HSD-1 was also seen in adipocytes, from both subcutaneous and visceral depots, differentiated in culture; expression was evident throughout differentiation but was generally greatest in preadipocytes and during early differentiation, declining as cells progressed to maturity. The inflammatory mediators lipopolysaccharide and tumor necrosis factor α had a main stimulatory effect on 11β-HSD-1 gene expression in canine subcutaneous adipocytes, but IL-6 had no significant effect. Treatment with dexamethasone resulted in a significant time- and dose-dependent increase in 11β-HSD-1 gene expression, with greatest effects seen at 24 h (2nM: approximately 4-fold; 20nM: approximately 14-fold; P = 0.010 for both). When subcutaneous adipocytes were treated with the peroxisome proliferator activated receptor γ agonist rosiglitazone, similar dose- and time-dependent effects were noted. However, no effects were seen when adipocytes from the gonadal WAT depot were treated with rosiglitazone. The induction of 11β-HSD-1 expression, by the pro-inflammatory cytokine tumor necrosis factor α and by lipopolysaccharide may have implications for the pathogenesis of obesity and its associated diseases in the dog.

Up-regulation of glyceraldehyde-3-phosphate dehydrogenase gene expression by HIF-1 activity depending on Sp1 in hypoxic breast cancer cells

Hypoxia up-regulates the expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in a cell type-specific manner. It is unknown whether this occurs in breast cancer. Here, we report that hypoxia up-regulates the GAPDH gene expression through breast cancer-specific molecular mechanisms in MCF-7 cells. Mutation analysis identified a novel hypoxia response element (HRE), in addition to the HRE found previously in prostate cancer LNCaP cells. Knockdown and overexpression of hypoxia-inducible factor (HIF)-1α indicated that HIF-1 contributed to the up-regulation of GAPDH gene expression by hypoxia. Although chromatin immunoprecipitation (ChIP) and plasmid immunoprecipitation analyses revealed the presence of HIF-1α on the novel HRE in both hypoxic cell lines, a mutation in either the novel HRE or its 3′-flanking GC-box resulted in a reduction of hypoxia-increased GAPDH promoter activity only in MCF-7 cells. ChIP analysis showed that Sp1 bound to the GC-box in MCF-7 cells, but not in LNCaP cells, in normoxia and hypoxia. Knockdown of Sp1 reduced hypoxia-increased promoter activity and expression level of GAPDH in MCF-7 cells. These results indicate that in MCF-7 cells, the activation of HIF-1 on the novel HRE contributes to the breast cancer-specific hypoxic induction of GAPDH gene expression and absolutely depends on the presence of Sp1 on the GC-box.

Oxidative stress‐induced attenuation of thrombospondin‐1 expression in primary rat astrocytes

Astrocytes, the major glial population in the central nervous system (CNS), can secrete thrombospondin (TSP)-1 that plays the role in synaptogenesis and axonal sprouting during CNS development and tissue repair. However, little is known about the regulation of TSP-1 expression in astrocytes under oxidative stress condition. Here, a hypoxic mimetic reagent, cobalt chloride (CoCl(2)), was used to initiate hypoxia-induced oxidative stress in primary rat astrocytes. CoCl(2) at the concentration range of 0.1-0.5 mM was found to cause no significant cell death in primary rat astrocytes. However, CoCl(2) at 0.2-0.5 mM increased intracellular reactive oxygen species (ROS) levels and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene expression that is known as a hallmark for oxidative damage. We further found that TSP-1 mRNA expression in astrocytes was inhibited dose- and time-dependently by CoCl(2). TSP-1 mRNA levels were increased in CoCl(2)-exposed astrocytes in the presence of the inhibitors (U0126 and PD98059) of mitogen-activated protein kinase/extracellular signal-regulated kinases (MAPK/ERK), when compared to that detected in the culture only exposed to CoCl(2). Moreover, the inhibition in TSP-1 mRNA expression by CoCl(2) was blocked by the addition of the potent antioxidant, N-acetylcysteine (NAC). Thus, we conclude that CoCl(2) inhibits TSP-1 mRNA expression in astrocytes via a ROS mechanism possibly involving MAPK/ERK. This inhibition may occur after CNS injury and impair the supportive function of astrocytes on neurite growth in the injured CNS tissues.

Sorbitol Dehydrogenase Gene Expression and Enzyme Activity in Apple: Tissue Specificity during Bud Development and Response to Rootstock Vigor and Growth Manipulation

Sorbitol is the primary photosynthate and translocated carbohydrate in apple (Malus ×domestica), and most of it is converted to fructose by sorbitol dehydrogenase (SDH) in sink tissues. We studied the expression of nine SDH genes, SDH activity, and sorbitol content of apple 1) in buds and floral tissues from dormancy to bloom, 2) in leaves and shoot tips of trees on two rootstocks, the moderately vigorous ‘Malling Merton 111’ (MM.111) and the dwarfing ‘Malling 9’ (M.9), and 3) in shoot tips in response to application of prohexadione-Ca to suppress shoot growth and defoliation and girdling (D/G) to deprive the shoot tip of sorbitol. In mature, orchard-grown trees, sorbitol was the main soluble carbohydrate in expressed xylem sap from dormancy to bloom at levels over 3- to 6-fold those of glucose and fructose, the other major sugars present. Sorbitol levels there increased from dormancy to its highest concentration at the half inch green stage and declined by bloom, while those of the other sugars increased. SDH activity per milligram of protein increased over 4-fold from dormancy to flowering. Three of the nine known SDH genes (SDH1, SDH2, and SDH3) were expressed in immature and mature leaves and all buds from dormancy to bloom, as well as in all floral organs, except that only SDH3 transcript was found in stamen tissue. Two genes, SDH6 and SDH9, were floral-tissue specific; SDH6 transcript was detected in all floral organs except stamens at full bloom, and SDH9 was only expressed in anthers with pollen. In buds and leaves of young, container-grown trees, SDH1 and SDH2 generally accounted for the majority of total SDH expression. There were generally no effects of rootstock on SDH expression, SDH activity, or sorbitol concentration in leaves, while apical shoot tips on M.9 rootstock exhibited greater SDH activity than those on MM.111 or lateral shoot tips on either rootstock, though SDH expression of apical and lateral shoot tips on M.9 was lower than on MM.111. Prohexadione-Ca reduced apical but not lateral shoot growth, increased apical but not lateral shoot tip sorbitol content, had no effect on SDH activity, and increased SDH1 expression of all shoot tips. D/G treatment reduced shoot growth, sorbitol content, and SDH activity, but increased SDH1 expression of apical shoot tips only and SDH2 expression of lateral shoot tips only. This work indicates that sorbitol and other sugars are abundantly available from dormancy to bloom, that SDH activity increased during this period, and that SDH expression is at least in part developmentally regulated within the individual floral and leaf tissues. In shoot tips and leaves of young trees, SDH transcript level was not correlated with sorbitol availability or SDH activity, suggesting that other factors have significant regulatory effects after SDH expression on SDH activity.

Molecular study of ovine glucose 6-phosphate dehydrogenase gene expression in respect to different energy intake

Glucose 6-phosphate dehydrogenase (G6PD) plays an important role in a ruminant's metabolism catalyzing the first committed reaction in the pentose phosphate pathway as it provides necessary compounds of NADPH for the synthesis of fatty acids. The cloning of ovine ( Ovis aries) G6PD gene revealed the presence of two cDNA transcripts (oG6PD A and oG6PD B), with oG6PD B being a product of alternative splicing and with no similarity to any other previously reported G6PD transcript. Here, we attempt to study the effect of energy balance in ovine G6PD transcript expression, trying simultaneously to find out any potential physiological role of the oG6PD B transcript. Changes of energy balance that lead to synergistic changes in the expression of both transcripts, but in opposite directions and not in a proportional way. Negative energy balance favours the presence of the oG6PD B transcript leading to a significant increase of its expression, compared to oG6PD A expression ( P < 0.05). In contrast, positive energy balance leads to a significant increase of oG6PD A compared to oG6PD B expression ( P < 0.05). In either condition oG6PD B expression is unchanged. Regression analysis showed that there is an energy balance threshold where the expression of both transcripts shows no change.

Adipocyte Differentiation, Mitochondrial Gene Expression and Fat Distribution: Differences between Zidovudine and Tenofovir after 6 Months

Abnormal lipid metabolism and cell oxidative mechanisms are reported in patients on antiretroviral treatment. We compared the expression of several key adipocyte genes in HIV-infected patients randomized to antiretroviral regimens containing zidovudine (AZT) or tenofovir disoproxil fumarate (TDF). Subcutaneous fat was sampled from 32 HIV-positive treatment-naive patients before and 6 months after randomization to AZT/lamivudine/efavirenz (n=15) or TDF/emtricitabine/efavirenz (n=17) plus 15 HIV-negative matched controls. Expression of genes involved in adipocyte differentiation, lipid metabolism, mitochondrial function and glucocorticoid generation were profiled using real-time PCR. Lipoprotein lipase and hepatic lipase activity were assessed. Before treatment, 11beta-hydroxysteroid dehydrogenase expression was down-regulated compared with controls. Following 6 months treatment with AZT, there was a significant increase in visceral adipose tissue (VAT; P=0.02) and the ratio of VAT to subcutaneous adipose tissue (P=0.008), down-regulation of cytochrome B (P=0.003) and cytochrome oxidase (COX)-3 gene expression (P=0.03), up-regulation of NADH dehydrogenase (P=0.008) and nuclear-encoded COX-4 (complex IV) gene expression (P=0.012). Genes involved with adipocyte cortisol generation, fatty acid metabolism and the tricarboxylic acid cycle were up-regulated. In the TDF-treated patients, there was no significant change in regional body fat or mitochondrial genes compared with pretreatment values. Changes in the expression of genes involved with cortisol and fatty acid metabolism were less marked with TDF. Interference with the mitochondrial electron transport chain appears to occur early in an AZT-containing regimen and occurs at a time when there is increased visceral fat and up-regulation of genes involved with adipocyte differentiation and fatty acid flux.

Independent Confirmation of Association between Metabolic Phenotypes of Polycystic Ovary Syndrome and Variation in the Type 6 17β-Hydroxysteroid Dehydrogenase Gene

Few candidate genes for polycystic ovary syndrome (PCOS) are widely agreed upon largely due to lack of replication. Type 6 17beta-hydroxysteroid dehydrogenase (HSD17B6) gene expression is increased in PCOS ovarian theca. Previous genetic study of HSD17B6 reported significant association of rs898611 with PCOS risk and metabolic phenotypes. Our objective was to replicate association between polymorphisms in HSD17B6 and PCOS in a well-characterized replication cohort. We conducted a case-control association study. Subjects were recruited from reproductive endocrinology clinics; controls were recruited from the surrounding communities of the University of Alabama at Birmingham and Cedars-Sinai Medical Center in Los Angeles. Genotyping occurred at Cedars-Sinai Medical Center. Participants included 335 White women with PCOS and 198 White controls. We assessed HSD17B6 genotype, PCOS status, and metabolic traits. The minor allele of rs898611 was not associated with PCOS; however, it was associated with increased body mass index (P = 0.031), increased fasting insulin (P = 0.008), decreased fasting glucose/insulin ratio (P = 0.038), and increased homeostasis model assessment of insulin resistance (HOMA-IR) (P = 0.021). rs10459247 and rs10876920 were associated with increased fasting insulin (P = 0.031 and 0.019, respectively), and rs10876920 was also associated with increased HOMA-IR (P = 0.046). Haplotype T-A-T-C was associated with reduced fasting insulin (P = 0.046), and haplotype C-A-C-T was associated with increased body mass index (P = 0.032). Although we did not replicate association between PCOS and rs898611, we replicated associations of this variant and others in HSD17B6 with metabolic traits. These replication data suggest a role for HSD17B6 in PCOS. How HSD17B6, an enzyme involved in steroid metabolism, may influence BMI and insulin resistance in PCOS remains to be determined.

What Is the Metabolic Role of Phosphoenolpyruvate Carboxykinase?

The enzyme phosphoenolpyruvate carboxykinase (GTP; EC 4.1.1.32) (PEPCK)2 has the unusual distinction of being very well studied but metabolically misunderstood. As we will document in this minireview, the enzyme has been almost exclusively linked to gluconeogenesis to the point that changes in the levels of PEPCK mRNA or its activity are associated with the control of hepatic glucose output and, more recently, with alterations in life span. That a tissue such as brown adipose tissue, which does not make glucose, has more PEPCK activity on a protein basis than is present in the liver is largely ignored. In addition, all eukaryotes have a gene for both a mitochondrial (PEPCK-M) and cytosolic (PEPCK-C) form of the enzyme. In the livers of most mammals studied to date (including humans), 50% of the total PEPCK activity is PEPCK-M. However, for reasons to be discussed, only PEPCK-C has been studied in any detail in mammals. Thus, the “strange case of PEPCK-M” deserves our attention. This minireview is an attempt to broaden our prospective on the metabolic role of this enzyme by reviewing the body of literature that has accumulated demonstrating that PEPCK plays a key role in a several metabolic processes associated with cataplerosis.

Investigation of associations of Yarrowia lipolytica, Staphylococcus xylosus, and Lactococcus lactis in culture as a first step in microbial interaction analysis.

The interactions that may occur between microorganisms in different ecosystems have not been adequately studied yet. We investigated yeast-bacterium interactions in a synthetic medium using different culture associations involving the yeast Yarrowia lipolytica 1E07 and two bacteria, Staphylococcus xylosus C2a and Lactococcus lactis LD61. The growth and biochemical characteristics of each microorganism in the different culture associations were studied. The expression of genes related to glucose, lactate, and amino acid catabolism was analyzed by reverse transcription followed by quantitative PCR. Our results show that the growth of Y. lipolytica 1E07 is dramatically reduced by the presence of S. xylosus C2a. As a result of a low amino acid concentration in the medium, the expression of Y. lipolytica genes involved in amino acid catabolism was downregulated in the presence of S. xylosus C2a, even when L. lactis was present in the culture. Furthermore, the production of lactate by both bacteria had an impact on the lactate dehydrogenase gene expression of the yeast, which increased up to 30-fold in the three-species culture compared to the Y. lipolytica 1E07 pure culture. S. xylosus C2a growth dramatically decreased in the presence of Y. lipolytica 1E07. The growth of lactic acid bacteria was not affected by the presence of S. xylosus C2a or Y. lipolytica 1E07, although the study of gene expression showed significant variations.

Alcohol dehydrogenase gene expression in Chironomusriparius exposed to di(2-ethylhexyl) phthalate

Di(2-ethylhexyl) phthalate (DEHP) is an industrial additive that is widely used as a plasticizer. Due to its widespread use, DEHP is often found in freshwater ecosystems and many freshwater species have been exposed to various levels of DEHP in natural aquatic systems. Alcohol dehydrogenase (ADH) is a metabolizing enzyme produced in response to exposure to DEHP. To evaluate the effects of DEHP exposure on the ADH metabolizing process of Chironomus, the full-length cDNA of ADH from Chironomus riparius was determined through molecular cloning and rapid amplification of cDNA ends (RACE). The expression of ADH was then analyzed during different life-cycle developmental stages and under various DEHP concentrations. In addition, a comparative and phylogenetic study among different orders of insects and vertebrates was conducted through analysis of sequence databases. The complete cDNA sequence of the ADH gene was 1134 bp in length. The amino acid sequence of C. riparius ADH was found to have a low degree of homology (around 70%) with other insects available in the databases. ADH mRNA was highly expressed during various developmental stages. ADH gene expression by C. riparius increased significantly after short-term exposure (24 h) to DEHP, regardless of the exposure concentration. ADH gene expression also increased in C. riparius following exposure to DEHP for 7 days. These results suggest that DEHP affects the metabolism associated with ADH in Chironomus species.