Hexose-6-phosphate Dehydrogenase Research Articles

A fingerprint of 2-[18F]FDG radiometabolites – How tissue-specific metabolism beyond 2-[18F]FDG-6-P could affect tracer accumulation

Studies indicate that the radiotracer 2-[18F]fluoro-2-deoxy-D-glucose (2-[18F]FDG) can be metabolized beyond 2-[18F]FDG-6-phosphate (2-[18F]FDG-6-P), but its metabolism is incompletely understood. Most importantly, it remains unclear whether downstream metabolism affects tracer accumulation invivo. Here we present a fingerprint of 2-[18F]FDG radiometabolites over time in cancer cells, corresponding tumor xenografts and murine organs. Strikingly, radiometabolites representing glycogen metabolism or the oxPPP correlated inversely with tracer accumulation across all examined tissues. Recent studies suggest that not only hexokinase, but also hexose-6-phosphate dehydrogenase (H6PD), an enzyme of the oxidative pentose phosphate pathway (oxPPP), determines 2-[18F]FDG accumulation. However, little is known about the corresponding enzyme glucose-6-phosphate dehydrogenase (G6PD). Our mechanistic invitro experiments on the role of the oxPPP propose that 2-[18F]FDG can be metabolized via both G6PD and H6PD, but data from separate enzyme knockdown suggest diverging roles in downstream tracer metabolism. Overall, we propose that tissue-specific metabolism beyond 2-[18F]FDG-6-P could matter for imaging.

G6PC1 and G6PC2 influence glucose-6-phosphate flux but not 11β-hydroxysteroid dehydrogenase type 1 activity.

In the endoplasmic reticulum (ER) lumen, glucose-6-phosphatase catalytic subunit 1 and 2 (G6PC1; G6PC2) hydrolyze glucose-6-phosphate (G6P) to glucose and inorganic phosphate whereas hexose-6-phosphate dehydrogenase (H6PD) hydrolyses G6P to 6-phosphogluconate (6PG) in a reaction that generates NADPH. 11β-hydroxysteroid dehydrogenase type 1 (HSD11B1) utilizes this NADPH to convert inactive cortisone to cortisol. HSD11B1 inhibitors improve insulin sensitivity whereas G6PC inhibitors are predicted to lower fasting blood glucose (FBG). This study investigated whether G6PC1 and G6PC2 influence G6P flux through H6PD and vice versa. Using a novel transcriptional assay, that utilizes separate fusion genes to quantitate glucocorticoid and glucose signaling, we show that overexpression of H6PD and HSD11B1 in the islet-derived 832/13 cell line activated glucocorticoid-stimulated fusion gene expression. Overexpression of HSD11B1 blunted glucose-stimulated fusion gene expression independently of altered G6P flux. While overexpression of G6PC1 and G6PC2 blunted glucose-stimulated fusion gene expression it had minimal effect on glucocorticoid-stimulated fusion gene expression. In the liver-derived HepG2 cell line, overexpression of H6PD and HSD11B1 activated glucocorticoid-stimulated fusion gene expression but overexpression of G6PC1 and G6PC2 had no effect. In rodents, HSD11B1 converts 11-dehydrocorticosterone (11-DHC) to corticosterone. Studies in wild type (WT) and G6pc2 knockout (KO) mice treated with 11-DHC for 5 weeks reveal metabolic changes unaffected by the absence of G6PC2. These data suggest that HSD11B1 activity is not significantly affected by the presence or absence of G6PC1 or G6PC2. As such, G6PC1 and G6PC2 inhibitors are predicted to have beneficial effects by reducing FBG without causing a deleterious increase in glucocorticoid signaling.

Silencing LINC01234 represses pancreatic cancer progression by inhibiting the malignant phenotypes of pancreatic cancer cells

ObjectivePrevious works have outlined the pivotal involvement of long intergenic non-coding RNA (lincRNA) in cancer progression, while the efficiency of LINC01234 in pancreatic cancer remained obscure. The purpose of this research is to unravel the regulatory mechanism of LINC01234 in pancreatic cancer via modulating microRNA (miR)-513a-3p and hexose 6-phosphate dehydrogenase (H6PD). MethodsPancreatic cancer cells were cultured and clinical tissue specimens were collected. LINC01234, miR-513a-3p and H6PD levels in pancreatic cancer cells and tissues were examined. Plasmids altering LINC01234, miR-513a-3p and H6PD expression were transfected into pancreatic cancer cells to assess the change in biological behaviors of pancreatic cancer cells. The targeting relations among LINC01234, miR-513a-3p and H6PD were validated. ResultsLINC01234 and H6PD levels were elevated while miR-513a-3p level was reduced in pancreatic cancer cells and tissues. LINC01234 deficiency hindered the malignant biological activities of pancreatic cancer cells. MiR-513a-3p depletion or H6PD elevation could abrogate the inhibitory effects of LINC01234 silencing on pancreatic cancer cells. LINC01234 sponged miR-513a-3p that targeted H6PD. ConclusionThe reduced LINC01234 exerts inhibitory impacts on pancreatic cancer cells via targeting miR-513a-3p to restrain H6PD level. The current study broadens the understanding of LINC01234 function and affords novel therapeutic targets for pancreatic cancer treatment.

Elevated Hexose-6-Phosphate Dehydrogenase Regulated by OSMR-AS1/hsa-miR-516b-5p Axis Correlates with Poor Prognosis and Dendritic Cells Infiltration of Glioblastoma.

Objective Glioblastoma (GBM), a type of malignant glioma, is the most aggressive type of brain tumor and is associated with high mortality. Hexose-6-phosphate dehydrogenase (H6PD) has been detected in multiple tumors and is involved in tumor initiation and progression. However, the specific role and mechanism of H6PD in GBM remain unclear. Methods We performed pan-cancer analysis of expression and prognosis of H6PD in GBM using the Genotype-Tissue Expression Project (GTEx) and The Cancer Genome Atlas (TCGA). Subsequently, noncoding RNAs regulating H6PD expression were obtained by comprehensive analysis, including gene expression, prognosis, correlation, and immune infiltration. Finally, tumor immune infiltrates related to H6PD and survival were performed. Results Higher expression of H6PD was statistically significantly associated with an unfavorable outcome in GBM. Downregulation of hsa-miR-124-3p and hsa-miR-516b-5p in GBM was detected from GSE90603. Subsequently, OSMR-AS1 was observed in the regulation of H6PD via hsa-miR-516b-5p. Moreover, higher H6PD expression significantly correlated with immune infiltration of dendritic cells, immune checkpoint expression, and biomarkers of dendritic cells. Conclusions The OSMR-AS1/ miR-516b-5p axis was identified as the highest-potential upstream ncRNA-related pathway of H6PD in GBM. Furthermore, the present findings demonstrated that H6PD blockading might possess antitumor roles via regulating dendritic cell infiltration and immune checkpoint expression.

Progesterone stimulates cortisol production in the maturing bovine cumulus-oocyte complex

In the bovine cumulus oophorus, 11β-hydroxysteroid dehydrogenase type 1 (HSD11B1)-mediated cortisol production dramatically increases during the periovulatory period. This event is closely associated with increased progesterone (P4) production, implying a functional connection between these C21 steroids. In this study, we investigated the mutual regulation of P4 and cortisol production in the bovine cumulus oophorus. Bovine cumulus-oocyte complexes (COCs) were aspirated from follicles 2–5 mm in diameter and subjected to in vitro maturation (IVM) for 24 h in an M199 supplemented with fetal calf serum (FCS) and follicle-stimulating hormone (FSH). COCs were treated with trilostane (0, 0.1, 1, 10 mM), an inhibitor of P4 synthesis, RU486 (0, 0.1, 1, 10 mM), a receptor antagonist for the progesterone receptor (PR) and glucocorticoid receptor (GR), and various concentrations of a synthetic progestogen nomegestrol acetate (NA; 0, 0.001, 0.01, 0.1, 1, 10 mM) to examine effect of P4. The effects of cortisol (0, 0.1, 1, 10 mM) were also examined in the presence or absence of trilostane.Trilostane and RU486 suppressed cumulus expansion, cortisol production, and HSD11B1 but not hexose-6-phosphate dehydrogenase (H6PDH) expression. Concomitant treatment with NA reversed the effects of trilostane. Unlike NA, cortisol did not alter the antagonistic effects of trilostane on cumulus expansion and HSD11B1 expression. Cortisol did not affect P4 production or steroidogenic acute regulatory protein (STAR), cholesterol side-chain cleavage enzyme (CYP11A1), 3β-hydroxysteroid dehydrogenase type 1 (HSD3B1), and HSD11B1 expression. Collectively, these results indicate that locally produced P4 is crucial in regulating the local glucocorticoid environment through PRtg in the maturing bovine cumulus oophorus. Cortisol, however, does not appear to regulate P4 or its production.

H6PD Gene Polymorphisms (R453Q and D151A) and Polycystic Ovary Syndrome: A Case-Control Study in A Population of Iranian Kurdish Women.

Background:Polycystic ovary syndrome (PCOS) is known as the most common endocrine and metabolic disorder in the reproductive-age women. Due to the effects of PCOS on the physical and mental health, the investigation of the factors affecting the development of PCOS is crucial. Hexose-6-phosphate dehydrogenase (H6PD) is a microsomal enzyme that catalyzes the first two reactions of the oxidative chain of the pentose phosphate pathway. The present study examined the polymorphisms of the H6PD gene (R453Q and D151A) in PCOS patients of Iranian Kurdish women.Materials and Methods:In this case-control study, a total, of 200 female volunteers in two equal groups participated in our study. The PCOS patients were selected based on the Rotterdam diagnostic criteria. The association of H6PD gene polymorphisms, D151A and R453Q, with the development of PCOS were investigated. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was used for genotyping. Statistical analysis was applied by the SPSS (version 16) software.Results:Statistically significant lower frequencies of AA+AG genotype (37% vs. 55%, P=0.01) and A allele (22.5% vs. 34%, P=0.01) of R453Q were observed in the patients compared to the controls. In the case of D151A, no significant differences were observed in the frequency of genotypes and alleles between the two groups.Conclusion:The findings of this study suggest that variants of H6PD R453Q affect the risk of PCOS.

Two high-rate pentose-phosphate pathways in cancer cells

The relevant role of pentose phosphate pathway (PPP) in cancer metabolic reprogramming has been usually outlined by studying glucose-6-phosphate dehydrogenase (G6PD). However, recent evidence suggests an unexpected role for a less characterized PPP, triggered by hexose-6-phosphate dehydrogenase (H6PD) within the endoplasmic reticulum (ER). Studying H6PD biological role in breast and lung cancer, here we show that gene silencing of this reticular enzyme decreases cell content of PPP intermediates and d-ribose, to a similar extent as G6PD silencing. Decrease in overall NADPH content and increase in cell oxidative status are also comparable. Finally, either gene silencing impairs at a similar degree cell proliferating activity. This unexpected response occurs despite the absence of any cross-interference between the expression of both G6PD and H6PD. Thus, overall cancer PPP reflects the contribution of two different pathways located in the cytosol and ER, respectively. Disregarding the reticular pathway might hamper our comprehension of PPP role in cancer cell biology.

Reducing NADPH Synthesis Counteracts Diabetic Nephropathy through Restoration of AMPK Activity in Type 1 Diabetic Rats

Diabetic nephropathy (DN) is a major complication of diabetes mellitus and a primary cause of end-stage renal failure. Clinical studies indicate that metabolic surgery improves DN; however, the mechanism remains unclear. Here, we report that Roux-en-Y Gastric Bypass (RYGB) surgery significantly blocked and reversed DN without affecting the insulin signaling pathway. This protective role of RYGB surgery is almost blocked by either inhibition or knockout of 5'AMP-activated protein kinase (AMPK) in podocytes. Furthermore, mRNA microarray data reveal that RYGB surgery obviously reduced the gene expression involved in nicotinamide adenine dinucleotide phosphate (NAPDH) synthesis. The expression of a key NADPH synthase, hexose-6-phosphate dehydrogenase (H6PD), was inhibited by the low plasma corticosterone level after surgery. In addition, blocking NAPDH synthesis by knocking down H6PD mimicked the beneficial role of RYGB surgery through activation of AMPK in podocytes. Therefore, this study demonstrates that reducing NADPH production is critical for renal AMPK activation in response to RYGB surgery.

SAT-591 Lack of Adipose-Specific Hexose-6-Phosphate Dehydrogenase Improves Fat Metabolic Phenotype and Increases Insulin Sensitivity in Mice

Lack of adipose-specific hexose-6-phosphate dehydrogenase improves fat metabolic phenotype and increases insulin sensitivity in mice.ABSTRACTIncreased glucocorticoids (GCs) production in adipose tissue promotes the development of visceral obesity and the MS. The action of GCs in adipose tissue is tightly regulated by 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) coupled with hexose-6-phosphate dehydrogenase (H6PDH). However, the phenotypic consequences of reduction of fat-specific GC action caused by inactivation of H6PDH specifically in adipose tissue have not been explored. To better understand the physiological effects of adipose H6PDH, we generated a tissue-specific animal model of adipocyte-specific H6PDH deletion under the control of the murine adipocyte-specific adiponectin promoter (H6PDHAcKO mice). H6PDHAcKO mice exhibited complete absence of H6PDH expression and decreased GC production with reduction of 11β-HSD1 in adipose tissue. These mice had decreased abdominal fat mass with decreased adipose lipogenic gene expression associated with reduction of their transcription factor C/EBPs mRNA levels. Moreover, H6PDHAcKO mice also decreased adipose lipolysis and reduced plasma FFA levels. In addition, H6PDHAcKO mice decreased fasting glucose levels and increase tolerance to glucose and insulin. These data suggest that H6PDHAcKO mice may provide a good model for study the potential contribution of fat-specific H6PDH inhibition to improve the metabolic phenotype in vivo.

OR03-06 NAD+ Availability Modulates 11β-HSD1-Mediated Glucocorticoid Regeneration in Mouse Skeletal Muscle

11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) is an NADPH-dependant reductase located in the sarcoplasmic reticulum (SR) lumen of skeletal muscle. It generates active glucocorticoids to regulate permissive and adaptive metabolism and contributes to the development of the Cushing’s syndrome phenotype in mice receiving oral corticosterone. The SR enzyme hexose-6-phosphate dehydrogenase (H6PDH) generates NADPH which supports 11β-HSD1 activity. H6PDH depletion disrupts the SR NADPH/NADP ratio leading 11β-HSD1 to assume glucocorticoid-inactivating dehydrogenase activity. Little is understood regarding routes to NAD(P)(H) biosynthesis and metabolism in the SR. Here we asked whether modulating cellular nicotinamide adenine dinucleotide (NAD+) availability (the parent molecule of NAD(P)(H)) would influence muscle 11β-HSD1 activity given its sensitivity to the SR NADPH/NADP ratio. We used FK866 to inhibit nicotinamide phospho-ribosyltransferase (NAMPT, rate-limiting enzyme in NAD+ biosynthesis) to deplete NAD(P)(H) in wild type mouse primary myotubes. FK866 treatment for 48h impaired cellular energetic status, reducing NAD+ (>90%), NADP+ (>50%) and ATP (>30%) without limiting cell viability. 11β-HSD1 reductase activity was decreased to 30% that of untreated cells (152±18 vs. 512±44 pmol/mg protein/h respectively, p<0.005). Employing H6PD knockout myotubes, NADP+-dependent 11β-HSD1 dehydrogenase activity was also impaired following NAMPT inhibition. The NAD+ precursor nicotinamide riboside (NR, 0.5mM), which bypasses NAMPT inhibition through the NR kinase pathway restored NAD+ levels and rapidly rescued 11β-HSD1 reductase activity in wild type and dehydrogenase activity in H6PD knockout myotubes. To assess this in vivo, we examined 11β-HSD1 reductase activity in muscle explants of inducible muscle-specific NAMPT knockout mice in which NAD+ levels are reduced by 90%, and show 40% lower activity compared to wild type explants (114±14 vs. 67±10 pmol/mg protein/h, p=0.04). These data suggest a novel level of redox-regulated 11β-HSD1-mediated glucocorticoid metabolism in skeletal muscle. These data also imply a pathway by which NAD+ status is communicated between the cytosol and the SR, which is contrary to the current belief that the pyridine nucleotide pool in these compartments is separate. NAMPT inhibition is being studied as a potential anti-cancer therapy and these data reveal hitherto unanticipated effects this therapy may have in a range of tissues.

A heretical view: rather than a solely placental protective function, placental 11β hydroxysteroid dehydrogenase 2 also provides substrate for fetal peripheral cortisol synthesis in obese pregnant ewes.

Exposure to glucocorticoid levels higher than appropriate for current developmental stages induces offspring metabolic dysfunction. Overfed/obese (OB) ewes and their fetuses display elevated blood cortisol, while fetal Adrenocorticotropic hormone (ACTH) remains unchanged. We hypothesized that OB pregnancies would show increased placental 11β hydroxysteroid dehydrogenase 2 (11β-HSD2) that converts maternal cortisol to fetal cortisone as it crosses the placenta and increased 11β-HSD system components responsible for peripheral tissue cortisol production, providing a mechanism for ACTH-independent increase in circulating fetal cortisol. Control ewes ate 100% National Research Council recommendations (CON) and OB ewes ate 150% CON diet from 60 days before conception until necropsy at day 135 gestation. At necropsy, maternal jugular and umbilical venous blood, fetal liver, perirenal fat, and cotyledonary tissues were harvested. Maternal plasma cortisol and fetal cortisol and cortisone were measured. Fetal liver, perirenal fat, cotyledonary 11β-HSD1, hexose-6-phosphate dehydrogenase (H6PD), and 11β-HSD2 protein abundance were determined by Western blot. Maternal plasma cortisol, fetal plasma cortisol, and cortisone were higher in OB vs. CON (p < 0.01). 11β-HSD2 protein was greater (p < 0.05) in OB cotyledonary tissue than CON. 11β-HSD1 abundance increased (p < 0.05) in OB vs. CON fetal liver and perirenal fat. Fetal H6PD, an 11β-HSD1 cofactor, also increased (p < 0.05) in OB vs. CON perirenal fat and tended to be elevated in OB liver (p < 0.10). Our data provide evidence for increased 11β-HSD system components responsible for peripheral tissue cortisol production in fetal liver and adipose tissue, thereby providing a mechanism for an ACTH-independent increase in circulating fetal cortisol in OB fetuses.

Lack of adipose-specific hexose-6-phosphate dehydrogenase causes inactivation of adipose glucocorticoids and improves metabolic phenotype in mice.

Excessive glucocorticoid (GC) production in adipose tissue promotes the development of visceral obesity and metabolic syndrome (MS). 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) is critical for controlling intracellular GC production, and this process is tightly regulated by hexose-6-phosphate dehydrogenase (H6PDH). To better understand the integrated molecular physiological effects of adipose H6PDH, we created a tissue-specific knockout of the H6PDH gene mouse model in adipocytes (adipocyte-specific conditional knockout of H6PDH (H6PDHAcKO) mice). H6PDHAcKO mice exhibited almost complete absence of H6PDH expression and decreased intra-adipose corticosterone production with a reduction in 11β-HSD1 activity in adipose tissue. These mice also had decreased abdominal fat mass, which was paralleled by decreased adipose lipogenic acetyl-CoA carboxylase (ACC) and ATP-citrate lyase (ACL) gene expression and reduction in their transcription factor C/EBPα mRNA levels. Moreover, H6PDHAcKO mice also had reduced fasting blood glucose levels, increased glucose tolerance, and increased insulin sensitivity. In addition, plasma free fatty acid (FFA) levels were decreased with a concomitant decrease in the expression of lipase adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) in adipose tissue. These results indicate that inactivation of adipocyte H6PDH expression is sufficient to cause intra-adipose GC inactivation that leads to a favorable pattern of metabolic phenotypes. These data suggest that H6PDHAcKO mice may provide a good model for studying the potential contributions of fat-specific H6PDH inhibition to improve the metabolic phenotype in vivo. Our study suggests that suppression or inactivation of H6PDH expression in adipocytes could be an effective intervention for treating obesity and diabetes.

Interaction of HSD11B1 and H6PD polymorphisms in subjects with type 2 diabetes are protective factors against obesity: a cross-sectional study

BackgroundThe enzyme 11-beta hydroxysteroid dehydrogenase type 1 (HSD11B1) converts inactive cortisone to active cortisol in a process mediated by the enzyme hexose-6-phosphate dehydrogenase (H6PD). The generation of cortisol from this reaction may increase intra-abdominal cortisol levels and contribute to the physiopathogenesis of obesity and metabolic syndrome (MetS). The relationship of HSD11B1 rs45487298 and H6PD rs6688832 polymorphisms with obesity and MetS was studied. We also studied how HSD11B1 abdominal subcutaneous (SAT) and visceral adipose tissue (VAT) gene expression is related to body fat distribution.MethodsRates of obesity and MetS features were cross-sectionally analyzed according to these polymorphisms in 1006 Brazilian white patients with type 2 diabetes (T2DM). Additionally, HSD11B1 expression was analyzed in VAT and SAT in a different cohort of 28 participants with and without obesity who underwent elective abdominal operations.ResultsAlthough polymorphisms of the two genes were not individually associated with MetS features, a synergistic effect was observed between both. Carriers of at least three minor alleles exhibited lower BMI compared to those with two or fewer minor alleles adjusting for gender and age (27.4 ± 4.9 vs. 29.3 ± 5.3 kg/m2; P = 0.005; mean ± SD). Obesity frequency was also lower in the first group (24.4% vs. 41.6%, OR = 0.43, 95% CI 0.21–0.87; P = 0.019). In the second cohort of 28 subjects, HSD11B1 gene expression in VAT was inversely correlated with BMI (r = − 0.435, P = 0.034), waist circumference (r = − 0.584, P = 0.003) and waist-to-height ratio (r = − 0.526, P = 0.010).ConclusionsThese polymorphisms might interact in the protection against obesity in T2DM individuals. Obese individuals may have decreased intra-abdominal VAT HSD11B1 gene expression resulting in decreasing intra-abdominal cortisol levels as a compensatory mechanism against central and general adiposity.

2031-P: Adipose-Specific Lack of Hexose-6-Phosphate Dehydrogenase Causes Inactivation of Adipose Glucocorticoids and Improves Metabolic Phenotype in Male Mice

Excessive glucocorticoids (GCs) production in adipose tissue promotes the development of visceral obesity and the metabolic syndrome. 11β-Hydroxysteroid dehydrogenase type 1 (11 β-HSD1) is critical for controlling intracellular GC production in adipose tissue, and this process is also regulated by hexose-6-phosphate dehydrogenase (H6PDH) that provides cofactor NADPH for 11β-HSD1 amplifying local GC production. To better understand the integrated molecular and physiological role of adipose H6PDH, we created a tissue specific knockout of H6PDH gene mouse model in adipocytes (H6PDHAcKO mice) and characterized the metabolic related phenotypes. H6PDHAcKO mice had decreased levels of intra-adipose corticosterone with reduction of 11β-HSD1 activity in response to H6PDH deletion in adipose tissue. Moreover, H6PDHAcKO mice also exhibited reduced fasting blood glucose levels and increased glucose tolerance with increased insulin sensitivity. In addition, plasma FFA levels were decreased with a concomitant decrease in the expression of lipase ATGL and HSL in the abdominal adipose tissue. These results indicate that inactivation of H6PDH expression in adipose tissue is sufficient to cause intra-adipose GC inactivation leading to the favorable pattern of metabolic phenotype. These data suggest that H6PDHAcKO mice may provide a good model for study the potential contributions of fat-specific H6PDH inhibition in the metabolic phenotype and other pathological disorders in vivo. Disclosure J. Wang: None. X. Wang: None. K. Lutfy: None. T. Friedman: None. M. Jiang: None. Y. Liu: None. Funding National Institute of Diabetes and Digestive and Kidney Diseases (SC1DK104821)

Increased myocardial 18F-FDG uptake as a marker of Doxorubicin-induced oxidative stress

BackgroundOxidative stress and its interference on myocardial metabolism play a major role in Doxorubicin (DXR) cardiotoxic cascade. MethodsMice models of neuroblastoma (NB) were treated with 5 mg DXR/kg, either free (Free-DXR) or encapsulated in untargeted (SL[DXR]) or in NB-targeting Stealth Liposomes (pep-SL[DXR] and TP-pep-SL[DXR]). Control mice received saline. FDG-PET was performed at baseline (PET1) and 7 days after therapy (PET2). At PET2 Troponin-I and NT-proBNP were assessed. Explanted hearts underwent biochemical, histological, and immunohistochemical analyses. Finally, FDG uptake and glucose consumption were simultaneously measured in cultured H9c2 in the presence/absence of Free-DXR (1 μM). ResultsFree-DXR significantly enhanced the myocardial oxidative stress. Myocardial-SUV remained relatively stable in controls and mice treated with liposomal formulations, while it significantly increased at PET2 with respect to baseline in Free-DXR. At this timepoint, myocardial-SUV was directly correlated with both myocardial redox stress and hexose-6-phosphate-dehydrogenase (H6PD) enzymatic activity, which selectively sustain cellular anti-oxidant mechanisms. Intriguingly, in vitro, Free-DXR selectively increased FDG extraction fraction without altering the corresponding value for glucose. ConclusionThe direct correlation between cardiac FDG uptake and oxidative stress indexes supports the potential role of FDG-PET as an early biomarker of DXR oxidative damage.

Absence of hexose-6-phosphate dehydrogenase results in reduced overall glucose consumption but does not prevent 11β-hydroxysteroid dehydrogenase-1-dependent glucocorticoid activation.

Hexose-6-phosphate dehydrogenase (H6PD) is thought to be the major source of NADPH within the endoplasmic reticulum (ER), determining 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) reaction direction to convert inert 11-oxo- to potent 11β-hydroxyglucocorticoids. Here, we tested the hypothesis whether H6pd knock-out (KO) in primary murine bone marrow-derived macrophages results in a switch from 11β-HSD1 oxoreduction to dehydrogenation, thereby inactivating glucocorticoids (GC) and affecting macrophage phenotypic activation as well as causing a more aggressive M1 macrophage phenotype. H6pdKO did not lead to major disturbances of macrophage activation state, although a slightly more pronounced M1 phenotype was observed with enhanced proinflammatory cytokine release, an effect explained by the decreased 11β-HSD1-dependent GC activation. Unexpectedly, ablation of H6pd did not switch 11β-HSD1 reaction direction. A moderately decreased 11β-HSD1 oxoreduction activity by 40-50% was observed in H6pdKO M1 macrophages but dehydrogenation activity was undetectable, providing strong evidence for the existence of an alternative source of NADPH in the ER. H6pdKO M1 activated macrophages showed decreased phagocytic activity, most likely a result of the reduced 11β-HSD1-dependent GC activation. Other general macrophage functions reported to be influenced by GC, such as nitrite production and cholesterol efflux, were altered negligibly or not at all. Importantly, assessment of energy metabolism using an extracellular flux analyzer and lactate measurements revealed reduced overall glucose consumption in H6pdKO M1 activated macrophages, an effect that was GC independent. The GC-independent influence of H6PD on energy metabolism and the characterization of the alternative source of NADPH in the ER warrant further investigations. ENZYMES: 11β-HSD1, EC 1.1.1.146; H6PD, EC 1.1.1.47.

Enhanced hexose-6-phosphate dehydrogenase expression in adipose tissue may contribute to diet-induced visceral adiposity.

BACKGROUNDVisceral fat accumulation increases the risk of developing type 2 diabetes and metabolic syndrome and is associated with excessive glucocorticoids (GCs) Fat depot-specific GC action is tightly controlled by 11β-hydroxysteroid dehydrogenase (11β-HSD1) coupled with the enzyme hexose-6-phosphate dehydrogenase (H6PDH). Mice with inactivation or activation of H6PDH genes show altered adipose 11β-HSD1 activity and lipid storage. We hypothesized that adipose tissue H6PDH activation is a leading cause for the visceral obesity and insulin resistance. Here we explored the role and possible mechanism of enhancing adipose H6PDH in the development of visceral adiposity in vivo.METHODSWe investigated the potential contribution of adipose H6PDH activation to the accumulation of visceral fat by characterization of visceral fat obese gene expression profiles, fat distribution, adipocyte metabolic molecules, and abdominal fat-specific GC signaling mechanisms underlying the diet-induced visceral obesity and insulin resistance in H6PDH transgenic mice fed a standard of high- fat diet (HFD).RESULTSTransgenic H6PDH mice display increased abdominal fat accumulation, which is paralleled by elevated lipid synthesis associated with induction of lipogenic transcriptor C/EBPα and PPARγ mRNA levels within adipose tissue. Transgenic H6PDH mice fed a high- fat diet (HFD) gained more abdominal visceral fat mass coupled with activation of GSK3β and induction of XBP1/IRE1α, but reduced pThr308 Akt/PKB content and browning gene CD137 and GLUT4 mRNA levels within the visceral adipose tissue than WT controls. HFD-fed H6PDH transgenic mice also had impaired insulin sensitivity and exhibited elevated levels of intra-adipose GCs with induction of adipose 11β-HSD1.CONCLUSIONThese data provide the first in vivo mechanistic evidence for the adverse metabolic effects of adipose H6PDH activation on visceral fat distribution, fat metabolism, and adipocyte function through enhancing 11β-HSD1-driven intra-adipose GC action.

Hexose-6-phosphate dehydrogenase controls cancer cell proliferation and migration through pleiotropic effects on the unfolded-protein response, calcium homeostasis, and redox balance.

Hexose-6-phosphate dehydrogenase (H6PD) produces reduced NADPH in the endoplasmic reticulum (ER) lumen. NADPH constitutes a cofactor for many reducing enzymes, and its inability to traverse biologic membranes makes in situ synthesis of NADPH in the ER lumen indispensable. The H6PD gene is amplified in several types of malignancies, and earlier work pointed toward a potential involvement of the enzyme in cancer cell growth. In the present study, we demonstrated a pivotal role of H6PD in proliferation and migratory potential of 3 human breast cancer cell lines. Knockdown of H6PD decreased proliferation and migration in SUM159, MCF7, and MDA-MB-453 cells. To understand the mechanism through which H6PD exerts its effects, we investigated the cellular changes after H6PD silencing in SUM159 cells. Knockdown of H6PD resulted in an increase in ER lumen oxidation, and down-regulation of many components of the unfolded protein response, including the transcription factors activating transcription factor-4, activating transcription factor-6, split X-box binding protein-1, and CCAAT/enhancer binding protein homologous protein. This effect was accompanied by an increase in sarco/endoplasmic reticulum Ca2+-ATPase-2 pump expression and an decrease in inositol trisphosphate receptor-III, which led to augmented levels of calcium in the ER. Further characterization of the molecular pathways involving H6PD could greatly broaden our understanding of how the ER microenvironment sustains malignant cell growth.—Tsachaki, M., Mladenovic, N., Štambergová, H., Birk, J., Odermatt, A. Hexose-6-phosphate dehydrogenase controls cancer cell proliferation and migration through pleiotropic effects on the unfolded protein response, calcium homeostasis, and redox balance.

The Role and Regulation of the 11 Beta-Hydroxysteroid Dehydrogenase Enzyme System in Patients with Inflammatory Bowel Disease.

Glucocorticoids are known to modulate a number of immunological responses including counteracting inflammation. Within tissues expressing the glucocorticoid and mineralocorticoid receptors including the colon, glucocorticoid metabolism is regulated by the isoenzymes of 11ß-hydroxysteroid dehydrogenase (11β-HSD). 11β-HSD1 acts as an oxidoreductase converting inactive cortisone into active cortisol, while 11β-HSD2 acts as a dehydrogenase converting active cortisol to inactive cortisone. Hexose-6 phosphate dehydrogenase (H6PDH) is a key regulator of 11β-HSD1 activity via its generation of NADPH. Variations in the 11β-HSD enzyme system in relation to levels of expression and regulation may have a role in IBD. The aim of this study was to investigate possible abnormalities of 11β-HSD enzyme system in the colon of patients with IBD. By using quantitative real-time PCR, we investigated the transcription levels of 11β-HSD1 and 2 in colonic tissue from IBD patients and healthy controls undergoing a colonoscopy for disease assessment. Disease activity was recorded using clinical (Mayo Score/Harvey-Bradshaw Index), Biochemical (C-reactive protein), histological, and endoscopic parameters. In addition, transcription levels of H6PDH and the glucocorticoid receptor alpha (GR-α) as well as key pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, Rela (subunit for NF Kappa B)) were later examined among this group, and results were correlated with 11β-HSD2 gene expression. Results and patient demographics were expressed as a mean (and SD), and differences between IBD patients and control groups were analyzed using a Student's t test or Mann-Whitney U test as appropriate, with a p value of ≤0.05 considered significant. Results were controlled for disease activity as outlined above. Results have demonstrated a significant downregulation in 11β-HSD2 expression in IBD patients compared with controls (13.8±17.1 au vs. 318.4±521.1au, p=0.01), whereas levels of 11β-HSD1 did not appear to vary across the two groups. Among IBD patients, there was a trend toward higher 11β-HSD1 expression in inflamed tissue compared with matched non-inflamed tissue (422.1±944 au vs. 102.2±103.9, P=0.09). Levels of H6PDH and the GR-α expression did not appear to vary among active inflamed IBD tissue and controls. As a result, we examined the association between pro-inflammatory cytokines and levels of 11β-HSD2 expression. Results showed an upregulation of key pro-inflammatory cytokine mRNA expression (TNF-α, IL-1β, IL-6) during inflammation with an associated downregulation of 11β-HSD2 mRNA expression when compared to controls. Dysregulation in this pathway could have a potential role in IBD pathogenesis and may account for exogenous glucocorticoid resistance in IBD. Further work assessing the role of the 11β-HSD enzyme system in steroid-resistant subjects is warranted.

Interplay between H6PDH and 11β-HSD1 implicated in the pathogenesis of type 2 diabetes mellitus

Extensive studies have been performed on the role of 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) in metabolic diseases. Our previous study reported glucose could directly regulate hexose-6-phosphate dehydrogenase (H6PDH) and 11β-HSD1. Recently, we further investigated the interplay of H6PDH and 11β-HSD1 and their roles in hepatic gluconeogenesis and insulin resistance to elucidate the importance of H6PDH and 11β-HSD1 in pathogenesis of type 2 diabetes mellitus (T2DM). T2DM rats model and H6PDH or 11β-HSD1 siRNA transfected in CBRH-7919 cells were used to explore the effect of H6PDH and 11β-HSD1 in T2DM. The results showed that the expression and activity of H6PDH and 11β-HSD1 in livers of diabetic rats were increased, with the expressions of PEPCK and G6Pase or liver corticosterone increased apparently. It also showed that H6PDH siRNA and 11β-HSD1 siRNA could inhibit the protein expression and enzyme activity by each other. With H6PDH siRNA, the enhancement of gluconeogenesis was blocked and insulin resistance stimulated by corticosterone was reduced. H6PDH and 11β-HSD1 might be the effective and prospective targets for T2DM and metabolic syndromes, based on the interplay between these two enzymes.