PGC-1α Protein Expression Research Articles

Adiponectin treatment improves insulin resistance in mice by regulating the expression of the mitochondrial-derived peptide MOTS-c and its response to exercise via APPL1-SIRT1-PGC-1α.

Adiponectin stimulates mitochondrial biogenesis through peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α), a major regulator of mitochondrial biogenesis. MOTS-c (mitochondrial open reading frame of the 12S rRNA) is a biologically active mitochondrial-derived peptide encoded by mitochondrial DNA. It influences the mechanisms of obesity and diabetes. We hypothesised that the adiponectin pathway may regulate the production and/or secretion of MOTS-c in skeletal muscle. We aimed to determine whether exercise and adiponectin affect MOTS-c to improve insulin resistance in mice. To investigate this hypothesis, we used wild-type C57BL/6 mice subjected to high-fat diet, an exercise regimen, and i.p. injection of recombinant mouse adiponectin (Acrp30) or MOTS-c, and adiponectin knockout (Adipoq-/-) mice (C57BL/6 background) subjected to i.p. injection of Acrp30. C2C12 myotubes were also treated with sirtuin 1 (SIRT1) inhibitor, PGC-1α inhibitor, SIRT1 activator, plasmid-expressed active APPL1 (adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper), pcDNA-SIRT1, or siRNA against APPL1, SIRT1 or PGC-1α. In Adipoq-/- mice, MOTS-c levels in the plasma and skeletal muscle were downregulated. In C2C12 myotubes, adiponectin increased the mRNA expression of MOTS-c. APPL1 protein level following adiponectin treatment positively correlated with MOTS-c protein and mRNA levels in C2C12 myotubes. SIRT1 overexpression increased the adiponectin-induced mRNA and protein expression of MOTS-c, SIRT1 and PGC-1α. Pharmacologic and genetic inhibition of PGC-1α suppressed the increases in MOTS-c mRNA and protein levels induced by SIRT1 overexpression. In mice, plasma and skeletal muscle MOTS-c levels were significantly downregulated following high-fat-diet. Exercise and i.p. Acrp30 or MOTS-c increased MOTS-c levels and adiponectin mRNA and protein expression in the plasma and skeletal muscle. Our findings showed that the APPL1-SIRT1-PGC-1α pathway regulates the production and/or secretion of skeletal muscle MOTS-c by mediating adiponectin signalling. Our study provides an insight into the cellular and molecular pathways underlying the pathogenesis of diabetes and shows that MOTS-c is a potential novel therapeutic target in the treatment of diabetes. Graphical abstract.

Fermented Soy Paste Alleviates Lipid Accumulation in the Liver by Regulating the AMPK Pathway and Modulating Gut Microbiota in High-Fat-Diet-Fed Rats.

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of liver disease due to lipid accumulation in the hepatocyte. Diet, especially a high-fat diet, is one risk factor that leads to NAFLD. Many natural compounds such as isoflavones have antiobesity effects. Therefore, intake of these functional compounds through daily dietary choices is a method of improving health. Miso is a kind of fermented soy paste, which is rich in isoflavones and has a different biological activity. In this study, we investigated the effects of different concentrations of fermented soy paste on NAFLD in high-fat-diet (HFD)-fed Sprague-Dawley (SD) rats. The results showed that 2% fermented soy paste decreased serum triacylglycerol (TG) and alanine aminotransferase (ALT) and reduced lipid accumulation in the liver through induced fatty acid oxidation by activating the adenosine 5'-monophosphate -activated protein kinase (AMPK) pathway and increasing PGC1α and CPT1α protein expression. Furthermore, we found that 2% fermented soy paste increased the abundance of Prevotellaceae NK3B31 and Desulfovibrio. Taken together, fermented soy paste improved HFD-induced lipid accumulation in the liver by activating fatty acid oxidation and modulating gut microbiota.

Testosterone Decreases Placental Mitochondrial Content and Cellular Bioenergetics.

Placental mitochondrial dysfunction plays a central role in the pathogenesis of preeclampsia. Since preeclampsia is a hyperandrogenic state, we hypothesized that elevated maternal testosterone levels induce damage to placental mitochondria and decrease bioenergetic profiles. To test this hypothesis, pregnant Sprague–Dawley rats were injected with vehicle or testosterone propionate (0.5 mg/kg/day) from gestation day (GD) 15 to 19. On GD20, the placentas were isolated to assess mitochondrial structure, copy number, ATP/ADP ratio, and biogenesis (Pgc-1α and Nrf1). In addition, in vitro cultures of human trophoblasts (HTR-8/SVneo) were treated with dihydrotestosterone (0.3, 1.0, and 3.0 nM), and bioenergetic profiles using seahorse analyzer were assessed. Testosterone exposure in pregnant rats led to a 2-fold increase in plasma testosterone levels with an associated decrease in placental and fetal weights compared with controls. Elevated maternal testosterone levels induced structural damage to the placental mitochondria and decreased mitochondrial copy number. The ATP/ADP ratio was reduced with a parallel decrease in the mRNA and protein expression of Pgc-1α and Nrf1 in the placenta of testosterone-treated rats compared with controls. In cultured trophoblasts, dihydrotestosterone decreased the mitochondrial copy number and reduced PGC-1α, NRF1 mRNA, and protein levels without altering the expression of mitochondrial fission/fusion genes. Dihydrotestosterone exposure induced significant mitochondrial energy deficits with a dose-dependent decrease in basal respiration, ATP-linked respiration, maximal respiration, and spare respiratory capacity. In summary, our study suggests that the placental mitochondrial dysfunction induced by elevated maternal testosterone might be a potential mechanism linking preeclampsia to feto-placental growth restriction.

Sitagliptin mitigates oxidative stress and up-regulates mitochondrial biogenesis markers in Brown adipose tissues of high-fat diet fed obese mice through AMPK phosphorylation

AimsThe present study was designed to investigate the effects of Sitagliptin (SGN) on oxidative stress, 5' adenosine monophosphate-activated protein kinase (AMPK) phosphorylation, together with mitochondrial biogenesis and thermogenic markers in intercapsular brown adipose tissue (iBAT) of Swiss albino mice fed high-fat diet for sixteen weeks. MethodsMice were distributed into five groups with last eight weeks of treatment: Normal control (NC): control diet with regular drinking water; Obese control (OC): 60% high-fat diet with 20% fructose drinking water (HFFR); Treatment-1: (OC + MET): HFFR with metformin (MET) 100 mg/kg/day; Treatment-2 (OC + SGN30): HFFR with SGN 30 mg/kg/day; Treatment-3 (OC + SGN20): HFFR with SGN 20 mg/kg/day. Fasting blood glucose, serum free fatty acid and adiponectin levels were measured. Oxidative stress, P-AMPK protein expression and mRNA expression of PPARα, PGC1-α, NRF-1, TFAM, and UCP-1 in iBAT were studied. ResultsSGN treatment for eight weeks ameliorated oxidative stress, improved P-AMPK protein expression and enhanced mitochondrial biogenesis and thermogenic mediators in iBAT. ConclusionThe findings from the present study suggest the beneficial role of SGN in ameliorating obesity associated oxidative stress and up-regulated mitochondrial biogenesis and thermogenic markers in iBAT of obese mice, possibly through AMPK activation.

Does Heat Acclimation Upregulate Skeletal Muscle Markers Of Oxidative Metabolism And Mitochondrial Biogenesis?

Heat acclimation (HA) increases tolerance to exercise performed in the heat and may improve aerobic fitness and performance in highly trained individuals. However, it is unknown if HA effects the expression of proteins related to mitochondrial biogenesis and oxidative capacity in skeletal muscle in active young individuals. PURPOSE: To investigate the effect of HA on the expression of oxidative and mitochondrial proteins in the skeletal muscle of healthy, active young-adult males and females. METHODS: In a quasi-experimental design, thirteen (7 males, 6 females) aerobically fit (VO2max > 75th percentile) individuals underwent 10-days of HA over 14-days. Participants performed two 45-minute bouts of exercise (walking at 30-40% maximal velocity) with 10 minutes rest per session in a hot environment (dry temperature ~42°C, relative humidity 30-50%). Pre- and post-HA VO2max (room temperature), second ventilatory threshold (VT), protein expression of PGC-1α, TFAM, CaMK, Cytochromes I-IV, and HSP72 were measured pre-HA and post-HA. Heat acclimation was determined comparing heart rate (HR), core temperature (CT), sweat rate, and ratings of perceived exertion (RPE) on days 1 and 10. RESULTS: Participants were acclimated as indicated by pre- and post-HA resting-CT (37.3 ± 0.38 vs. 37.1 ± 0.28°C; p = 0.04), maximal-CT (38.8 ± 0.35 vs. 38.2 ± 0.38°C; p<0.0001), mean-HR (138.5 ± 17.7 vs. 125.9 ± 14.0 bpm; p=0.03), maximal-HR (170.2 ± 26.4 vs. 149.4 ± 24.5 bpm; p=0.003), mean-RPE (12.4 ± 1.2 vs. 9.8 ± 1.1; p< 0.0001), and maximal-RPE (15.0 ± 1.6 vs. 12.2 ± 1.9; p=0.002). Pre- to Post-HA VO2max (50.2 ± 8.6 vs. 51.7 ± 11.2 mL/kg/min; p=0.08) and VT (31.2 ± 7.0 vs. 33.9 ± 8.7; p=0.19) were unchanged. Protein expression was unchanged pre- to post-HA; (PGC-1α p=0.21, TFAM p=0.34, CaMK 0-0.19, Cytochromes I p=0.73; II p=0.85, III p=0.62, IV p=0.70, Hsp72 p=0.27). CONCLUSIONS: 10-days of low intensity exercise in the heat resulted in HA, but did not increase aerobic fitness or markers of mitochondrial biogenesis and oxidative metabolism in physically active individuals. Supported by a research grant from the College of Education, University of New Mexico.

AMPK-SIRT1-PGC1α Signal Pathway Influences the Cognitive Function of Aged Rats in Sevoflurane-Induced Anesthesia.

To understand the effect of AMP-activated protein kinase (AMPK)-SIRT1 (silent information regulator 1)-PPARγ coactivator-1α (PGC1α) signaling pathway on the cognitive function of sevoflurane-anesthetized aged rats. Aged rats were divided into Normal group, Sevo group (Sevoflurane anesthesia), Sevo + AICAR (the AMPK activator) group, Sevo + EX527 group (the AMPK inhibitor), and Sevo + AICAR + EX527 group. The cognitive function of rats was determined by the Morris water maze. Hippocampal neuronal apoptosis was evaluated by TUNEL and Fluoro-Jade C (FJC) staining, and the expression of cleaved caspase-3 was detected by immunohistochemistry. ROS, SOD, and MDA levels and the fluorescence intensity of GFAP in the hippocampus were assayed. The mitochondrial membrane potential (MMP), mitochondrial mass, ATP level, and the expression of AMPK-SIRT1-PGC1α were determined by the corresponding methods. Rats in the Sevo group manifested significant extension in the escape latency, with fewer platform crossings; and meanwhile, the apoptotic rate, the number of FJC-positive cells, and the fluorescence intensity of GFAP of neurons were elevated, with up-regulation of cleaved caspase-3. Moreover, the level of MDA and ROS was increased evidently, with significant down-regulation of SOD activity, ATP, mitochondrial mass and MMP levels, and AMPK, SIRT1 and PGC-1α protein expressions. However, sevoflurane-induced changes above were improved after the administration of AICAR, and EX527 could reverse AICAR-induced improvements in Sevo-anesthetized aged rats. Activating AMPK-SIRT1-PGC1α pathway can improve the cognitive function and mitigate the neuronal injury in Sevo-anesthetized aged rats by antagonizing the oxidative stress and maintaining the mitochondrial function.

1945-P: Pioglitazone Promotes the Function of White and Brown Adipose Tissue in Diet-Induced Obese Mice

PPARγ agonist pioglitazone is an insulin sensitizer in the treatment of type 2 diabetes. Here, we investigate the effects of pioglitazone on white and brown fat in diet-induced obese mice. After 16-week diet intervention, C57BL/6 mice were divided randomly into three groups: normal chow diet group (NC group), high-fat diet group (HFD group) and high-fat diet + pioglitazone group (HFDP group). Pioglitazone (30-40mg/kg/day) was administered in drinking water for 6 weeks. After 6-week pioglitazone treatment, there was no significant difference in both body weight and fat mass between the HFDP group and the HFD group. However, fasting blood glucose, glucose tolerance and insulin sensitivity were significantly improved in HFDP group. What’s more, the weight of liver and epididymal adipose tissue (eWAT) were not reduced by pioglitazone treatment. However, the weight of brown adipose tissue (BAT) in the HFDP group was significantly higher than HFD group. Histological staining showed that the hepatic steatosis was ameliorated by pioglitazone. Quantitative real-time PCR revealed that mRNA levels of heat-producing genes (PRDM16, FGF-21, UCP1) and mitochondrial biosynthesis and functional genes (Tfam, cidea, cox7a1, Fabp4) were upregulated in eWAT by pioglitazone. Western blotting results also showed that the protein expression of UCP1 and PGC1α was upregulated by pioglitazone both in eWAT and in BAT when compared with the HFD group. To sum up, our study revealed that pioglitazone could promote the browning of WAT and activate the function of BAT with improved hepatic steatosis in diet-induced obese mice. Disclosure F. Xu: None. W. Wang: None. X. Zheng: None. W. Guo: None. Y. Shen: None.

1498-P: A Feedback Loop between miRNA Processing Complex and miR-3975 in Western Diet–Induced Brown Adipose Tissue Dysfunction

Brown adipose tissue (BAT) is considered as promising therapeutic targets for obesity and metabolic disorder through modulation of energy homeostasis. microRNAs (miRNAs) are established as essential modulators of biological processes in many tissues, including the differentiation and function of BAT. Here we demonstrated that high-fat/high fructose associated dysfunction of BAT coincides with impaired miRNA-processing pathway in a feedback loop with miR-3975 activation. Eight-week-old male C57BL/6 mice were fed a low-fat diet (10% fat and 17% sucrose; LFD) and high fat-high fructose (45% fat, 17% sucrose, and 17% fructose; HFHFD) for 4, 12, and 20 weeks (n=6-9 per group). We evaluated expression levels of apoptosis markers, thermogenesis markers, miRNA biogenesis regulators, and miRNAs in BAT. Mice fed an HFHFD showed a gradually significance increased body weight and brown adipose tissue weight when compared to the mice fed with LFD. BAT of HFHFD fed mice has a pale appearance versus feeding period matched LFD fed mice. The protein expression of thermogenesis markers, UCP1 (P &amp;lt;0.05), and PGC1α (P &amp;lt;0.05) were shown to decrease after 12 weeks of HFHFD diet feeding. The apoptosis marker, the Bax/Bcl-2 ratio, was upregulated at 12 weeks (P &amp;lt; 0.01). Mice fed HFHFD for 12 and 20 weeks showed a significant decrease in Dicer (P &amp;lt; 0.01 for 12 and 20 weeks) and TRBP2 (P &amp;lt; 0.01 for 12 weeks and P &amp;lt; 0.0001 for 20 weeks). miR-3975, which is predicted miRNA to regulate both Dicer and TRBP2 by IPA was activated at 12 weeks (P &amp;lt; 0.01), while inactivated at 20 weeks (P &amp;lt; 0.001). Our results suggest that miR-3975 acts as a component of a negative feedback loop that blocks miRNA biogenesis machinery through the inhibition of Dicer and TRBP2 expression. Therefore, long-term HFHFD may disturb miRNA processing and contribute to brown adipose tissue dysfunction and apoptosis by modulation of miRNA expression. We are currently investigating the novel feedback loop in brown adipocyte primary cells. Disclosure M. Park: None. S. Meruvu: None. J. Zhang: None. M. Choudhury: None.

Peripherally Restricted Dual Cannabinoid Receptor Agonist Modifies Mitochondrial Signaling and Inhibits Atrial Electrical Remodeling in a Tachypaced Ex Vivo Rat Model

Atrial fibrillation (AF) leads to rate‐dependent changes, which progressively increase the risk for recurrence and persistence of the arrhythmia (collectively defined as atrial remodelling). Atrial effective refractory period (AERP) shortening and conduction velocity prolongation are hallmarks of atrial remodelling markedly promoting AF substrate. The molecular mechanisms underlying atrial remodelling are not fully elucidated and therapeutic strategies to reduce AF burden by preventing atrial remodelling are not yet available. Cannabinoid (CB) receptor ligands exert cardioprotective effects. We hypothesized that CB13, a peripherally restricted dual CB receptor agonist with limited brain penetration, would reduce atrial remodelling.Sprague Dawley (SD) rat hearts were excised and hung on a Langendorff set‐up, and supplemented with CB13 (1μM) or without (control) (paced and non‐paced). Coronary pressure of 60–80 mmHg was maintained. Electrophysiological (EP) parameters were measured by placing a mini‐hook platinum‐iridium quadrupole electrode on the right atrium (RA) for pacing and atrial recording (CB13 n=8; control n=8). RA were tachypaced for 90 min at double diastolic threshold and a cycle length of 5 ms higher than failed 1:1 atrial capture. S1S2 stimulation protocol was used to measure AERP, defined as the longest interval with failed 1:1 capture. The effects of CB13 on atrial refractoriness and AF substrate were assessed by analysis of high resolution electrogram recordings and protein expression via western blot.Post‐tachypacing EP recordings showed reduced AERP in control compared to pre‐tachypacing (54.38±3.05 vs. 44.75±3.67 ms; p&lt;0.01). AERP post‐tachypacing in CB13‐treated hearts remained unchanged compared to pre‐tachypacing (55.25±3.46 vs. 55.50±2.67 ms; ns). Thus, CB13‐treatment resulted in preservation of AERP compared to control (103.6±9.2 vs. 82.3±4.4 % of baseline; p&lt;0.05). Protein expression of PGC1α was significantly increased by CB13 compared to controls (paced and non‐paced). Upregulation of mitochondrial biogenesis may be related to increased phosphorylation of AMPKα at residue threonine 172 (compared to total AMPKα, p=0.12, CB13 vs. control paced). Furthermore, gap junction connexin 43 was significantly downregulated in paced controls vs. non‐paced controls, while CB13 remained upregulated (no significance vs. controls).CB13 exposure during tachypacing prevents AERP reduction and gap junction downregulation probably through mechanisms involving mitochondrial biogenesis. Thus, by inhibiting atrial remodeling CB13 and the subsequent activation of CB receptors may be a viable treatment strategy for AF.Support or Funding InformationThis work was supported by studentships from the Canadian Institutes of Health Research and Mitacs (D.L.)

Milk fat globule membrane and its component phosphatidylcholine induce adipose browning both in vivo and in vitro

The functional induction of brown-like adipocytes in white adipose tissue (WAT) provides a defense against obesity. The aim of this study was to analyze the effects of milk fat globule membrane (MFGM) and its component phosphatidylcholine (PC) on the brown remodeling of WAT. Male C57BL/6 J mice were fed a high-fat diet (HFD) for 8 weeks and then fed HFD for another 8 weeks with MFGM. In vitro studies were performed in C3H10T1/2 pluripotent stem cells, 3T3-L1 pre-adipocytes and differentiated inguinal WAT stromal vascular cells (SVCs) to determine the role of MFGM and PC on the formation of brown-like adipocytes. MFGM decreased fasting glucose and serum insulin levels in HFD-fed mice. MFGM improved glucose tolerance and insulin sensitivity, and induced browning of inguinal WAT. MFGM and its component PC stimulated transformation of brown-like adipocytes in C3H10T1/2 pluripotent stem cells, 3T3-L1 adipocytes and SVCs by increasing the protein expression of UCP1, PGC-1α, PRDM16 as well as the mRNA expression of other thermogenic genes and beige cell markers. MFGM and PC also increased mitochondrial DNA (mtDNA) copy number, mitochondrial density and oxygen consumption rate and up-regulated the mRNA expression of mitochondria-biogenesis-related genes in vitro. PPARα inhibitor GW6471 treatment or knockdown of PPARα using lentivirus-expressing shRNA inhibited the PC-induced increase in the protein expression of UCP1, PGC-1α and PRDM16 in C3H10T1/2 pluripotent stem cells and 3T3-L1 adipocytes, indicating the potential role of PPARα in PC-mediated brown-like adipocyte formation. In conclusion, MFGM and milk PC induced adipose browning, which has major protective effects against obesity and metabolic dysfunction.

Oxidative Phosphorylation Promotes Primary Melanoma Invasion

Dermal invasion is a hallmark of malignant melanoma. Although the molecular alterations that drive the progression of primary melanoma to metastatic disease have been studied extensively, the early progression of noninvasive primary melanoma to an invasive state is poorly understood. To elucidate the mechanisms underlying the transition from radial to vertical growth, the first step in melanoma invasion, we developed a zebrafish melanoma model in which constitutive activation of ribosomal protein S6 kinase A1 drives tumor invasion. Transcriptomic analysis of ribosomal protein S6 kinase A1-activated tumors identified metabolic changes, including up-regulation of genes associated with oxidative phosphorylation. Vertical growth phase human melanoma cells show higher oxygen consumption and preferential utilization of glutamine compared to radial growth phase melanoma cells. Peroxisome proliferator activated receptor γ coactivator (PGC)-1α, has been proposed as a master regulator of tumor oxidative phosphorylation. In human primary melanoma specimens, PGC1α protein expression was found to be positively associated with increased tumor thickness and expression of the proliferative marker Ki-67 and the reactive oxygen species scavenger receptor class A member 3. PGC1α depletion modulated cellular processes associated with primary melanoma growth and invasion, including oxidative stress. These results support a role for PGC1α in mediating glutamine-driven oxidative phosphorylation to facilitate the invasive growth of primary melanoma.

Impairment of PGC-1α-mediated mitochondrial biogenesis precedes mitochondrial dysfunction and Alzheimer's pathology in the 3xTg mouse model of Alzheimer's disease.

Impairment of mitochondrial biogenesis and mitochondrial dysfunction is a prominent feature of Alzheimer's disease (AD). However, the extent to which the impairment of mitochondrial biogenesis influences mitochondrial dysfunction at the onset and during progression of AD is still unclear. Our study demonstrated that the protein expression pattern of the transcription factor pCREB/CREB, together with the protein expression of PGC-1α, NRF1 and TFAM are all significantly reduced in early ages of 3xTg-AD mice. We also found reduced mRNA expression levels of PKAC-α, CREB, PGC-1α, NRF1, NRF2 and TFAM as early as 1month-of-age, an age at which there was no significant Aβ oligomer deposition, suggesting that mitochondrial biogenesis is likely impaired in ages preceding the development of the AD pathology. In addition, there was a decrease in VDAC2 expression, which is related to mitochondrial content and mitochondrial function, as demonstrated by protein expression of complex IV, as well as complex II+III, and complex IV activities, at later ages in 3xTg-AD mice. These results suggest that the impairment in mitochondrial biogenesis signaling mediated by PGC-1α at early ages of the AD mice model likely resulted in mitochondrial dysfunction and manifestation of the AD pathology at later ages. Taken together, enhancing mitochondrial biogenesis may represent a potential pharmacological approach for the treatment of AD.

The impact of DRP1 on myocardial fibrosis in the obese minipig.

The heart is a highly oxidative tissue, thus mitochondria play a major role in maintaining optimal cardiac function. Our previous study established a dietary-induced obese minipig with cardiac fibrosis. The aim of this study was to elucidate the role of mitochondrial dynamics in cardiac fibrosis of obese minipigs. Four-month-old Lee-Sung minipigs were randomly divided into two groups: a control group (C) and an obese group (O) by feeding a control diet or a high-fat diet (HFD) for 6months. Exposure of H9c2 cardiomyoblasts to palmitate was used to explore the effects of high-fat on induction of myocardial injury in vitro. The O pigs displayed greater heart weight and cardiac collagen accumulation. Obese pigs exhibited a lower antioxidant capacity, ATP concentration, and higher oxidative stress in the left ventricle (LV). The HFD caused downregulation in protein expression of PGC-1α and OPA1, and upregulation of DRP1, FIS1, and PINK1 in the LV of O compared to C pigs. Furthermore, palmitate induced apoptosis and decreased ATP content in H9c2 cells. Palmitate elevated the protein expression of DRP1 and PINK1 in these cells. Inhibition of DRP1 protein expression by siDRP1 in H9c2 cells resulted in enhanced ATP and decreased palmitate-induced apoptosis. These results suggest that mitochondrial dynamics were linked to the progression of obesity-related cardiac injury. Inhibition of DRP1 after palmitate exposure in H9c2 cells resulted in improved ATP level and decreased apoptosis in vitro suggesting that mitochondrial fission serves a key role in progression of obesity-induced cardiac fibrosis.

Canagliflozin ameliorates obesity by improving mitochondrial function and fatty acid oxidation via PPARα in vivo and in vitro

AimsSodium-glucose cotransporter 2 (SGLT2) inhibitors have been reported to significantly reduce body weight. This study investigated whether SGLT2 inhibitors directly affect adipose tissues and the underlying mechanisms in vivo and in vitro. Main methodsMale C57BL/6 mice were fed a normal diet, high-fat diet (HFD), or HFD with canagliflozin for 14 weeks. 3T3-L1 adipocytes were treated with canagliflozin. Metabolic parameters were measured. Key findingsCanagliflozin reduced body weight, fat mass, and white adipose tissue (WAT) weight and inhibited adipocyte hypertrophy. Canagliflozin improved glucose and lipid metabolic disorders induced by HFD. Furthermore, canagliflozin treatment reversed the suppressed mRNA and protein expression of PGC-1α, NRF1, tfam and CPT1b, which are markers of mitochondrial biogenesis, function and fatty acid oxidation in mice with obesity. In vitro, canagliflozin increased mitochondrial DNA to nuclear DNA and upregulated the expression of PGC-1α, NRF1, tfam, COX5b, COX8b, Atp5o, and CPT1b mRNA and PGC-1α, NRF1, tfam, COX5b, CPT1b protein in 3T3-L1 adipocytes in a dose-dependent manner, while these increases were inhibited by GW6471, a PPARα antagonist. SignificanceOur study showed that canagliflozin protected against HFD-induced obesity and obesity-related metabolic disorders by improving mitochondrial function and fatty acid oxidation in adipose tissue and adipocytes. Such energy-dissipating effects of canagliflozin may be mediated by PPARα.

Inhibition of Lipolysis With Acipimox Attenuates Postburn White Adipose Tissue Browning and Hepatic Fat Infiltration.

Extensive burn injuries promote an increase in the lipolysis of white adipose tissue (WAT), a complication that enhances postburn hypermetabolism contributing to hyperlipidemia and hepatic steatosis. The systemic increase of free fatty acids (FFAs) due to burn-induced lipolysis and subsequent organ fatty infiltration may culminate in multiple organ dysfunction and, ultimately, death. Thus, reducing WAT lipolysis to diminish the mobilization of FFAs may render an effective means to improve outcomes postburn. Here, we investigated the metabolic effects of Acipimox, a clinically approved drug that suppresses lipolysis via inhibition of hormone-sensitive lipase (HSL). Using a murine model of thermal injury, we show that specific inhibition of HSL with Acipimox effectively suppresses burn-induced lipolysis in the inguinal WAT leading to lower levels of circulating FFAs at 7 days postburn (P < 0.05). The FFA substrate shortage indirectly repressed the thermogenic activation of adipose tissue after injury, reflected by the decrease in protein expression of key browning markers, UCP-1 (P < 0.001) and PGC-1α (P < 0.01). Importantly, reduction of FFA mobilization by Acipimox significantly decreased liver weight and intracellular fat accumulation (P < 0.05), suggesting that it may also improve organ function postburn. Our data validate the pharmacological inhibition of lipolysis as a potentially powerful therapeutic strategy to counteract the detrimental metabolic effects induced by burn.

Dietary thymol supplementation promotes skeletal muscle fibre type switch in longissimus dorsi of finishing pigs.

As one of the key points related to meat quality, skeletal muscle fibre type is determined by energy metabolism and genetic factors, but its transformation could be also greatly influenced by many factors. Thymol, the primary effective ingredients of thyme, is well known for its anti-oxidation and anti-inflammatory, while little is known about its effect on skeletal muscle oxidative metabolism and fibre type switch. Therefore, in order to investigate its effects and possibility to be applied in livestock production, 36 150-day-old fattening Pigs were fed with different diet for six-week experiment. As a result, the drip loss ratio of longissimus dorsi (LD) was significantly reduced (p<.05). Oxidative metabolism-related enzyme activity, the mRNA levels and protein expression of COX5B and PGC1α, mRNA level of myosin heavy chain I (MyHC I) and protein level of MyHC IIa were significantly upregulated (p<.05). While compared with control group, the protein expression of MyHC IIb was significantly decreased (p<.05). The result revealed that thymol could promote the oxidative metabolism in the muscle of pigs and improve the meat quality to a certain extent.

The Ability of Carnitine to Act as a Type 1Histone Deacetylase Inhibitor May Explain the Favorable Impact of Carnitine Supplementation on Mitochondrial Biogenesis in the Elderly

A number of studies have shown that carnitine supplementation – alone or in conjunction with supplemental lipoic acid - promotes mitochondrial biogenesis (MB) in skeletal, muscle, and brain of aging rodents; no such effect is seen in younger animals. These findings parallel clinical studies in which supplemental carnitine improves physical and mental energy in elderly humans, while decreasing body fat and increasing lean mass – effects that have not been achieved with carnitine in younger people. The age dependence of these phenomena appears to reflect the fact that tissue carnitine levels, especially those in muscle, decline during aging; carnitine supplementation restores higher, more youthful tissues carnitine levels in aging animals, but has relatively little impact in this regard on younger ones. The effect of supplemental carnitine on MB in aging animals appears to be mediated, in whole or in part, by increased expression of PPARγ-coactivator-1α (PGC-1α), a key driver of MB. There is recent evidence that, in low millimolar intracellular concentrations such as those seen in skeletal muscle, carnitine functions as an inhibitor of type 1 histone deacetylases (HDACs). Moreover, it has been reported that drug inhibitors of these deacetylases boost mRNA and protein expression of PGC-1α, presumably by promoting transcription of the PGC-1a gene; these drugs also amplify MB. It is therefore proposed that intracellular carnitine provides a moderate tonic inhibition of type 1 HDACs that supports PGC1α transcription and that diminishes with age as tissue carnitine levels decline; hence, carnitine supplementation in the elderly restores youthful expression of PGC-1α and promotes MB. The complementary impact of lipoic acid on MB may reflect the fact that the promoter of the gene coding for nuclear respiratory factor-1 (NRF-1) contains antioxidant response elements; hence, NRF-1 transcription is promoted by phase 2 inducers such as lipoic acid. PGC-1α and NRF-1 collaborate in driving the expression of mitochondrial proteins. Additional nutraceutical measures which may likewise support MB – citrulline, taurine, N-acetylcysteine, high-dose biotin, and astaxanthin – are discussed. The adverse impact of metabolic syndrome on MB in skeletal muscle may be mediated by toll-like receptor 4 (TLR4) signaling stimulated by saturated fatty acids; antagonists of TLR4 signaling, possibly including ferulic acid and phycocyanobilin, may therefore promote MB in the context of metabolic syndrome. Restoration of youthful MB in the elderly may have favorable impacts on physical capacity and cognitive function, body composition, insulin sensitivity, and oxidative stress.

Pro-renin receptor suppresses mitochondrial biogenesis and function via AMPK/SIRT-1/ PGC-1α pathway in diabetic kidney.

Abnormal mitochondrial biogenesis and function has been linked to multiple diseases including diabetes. Recently, we demonstrated the role of renal (Pro)renin receptor (PRR) in the dysregulation of mitochondria. We hypothesized that PRR contributes to the reduction of mitochondrial biogenesis and function in diabetic kidney via PGC-1α/AMPK/SIRT-1 signaling pathway. In vivo and in vitro studies were conducted in C57BL/6 mouse and mouse renal mesangial cells (mRMCs). Control and streptozotocin-induced diabetic mice were injected with scramble or PRR shRNA and followed for a period of eight weeks. PRR mRNA and protein expression increased by 44% and 39% respectively (P<0.05) in kidneys of diabetic mice, and in mRMCs exposed to high glucose by 43 and 61% respectively compared to their respective controls. These results were accompanied by reduced mRNA and protein expressions of PGC-1α (67% and 75%), nuclear respiratory factors (NRF-1, 48% and 53%), mitochondrial transcriptional factor A (mtTFA, 56% and 40%), mitochondrial DNA copy number by 75% (all, P<0.05), and ATP production by 54%, respectively in diabetic kidneys and in mRMCs exposed to high glucose. Compared to non-diabetic control mice, PRR knockdown in diabetic mice and in mRMCs, not only attenuated the PRR mRNA and protein expression but also normalized mRNA and protein expressions of PGC-1α, NRF-1, mtTFA, mitochondrial DNA copy number, and ATP production. Treatment with AMPK inhibitor, Compound C, or SIRT-1 inhibitor, EX-527, alone, or combined with PRR siRNA caused marked reduction of mRNA expression of PGC-1α, NRF-1 and mtTFA, and ATP production in mRMCs exposed to high glucose. In conclusion, our study demonstrated the contribution of the PRR to the reduction of mitochondrial biogenesis and function in diabetic kidney disease via decreasing AMPK/SIRT-1/ PGC-1α signaling pathway.

Changes in Mitochondria-Associated Protein Expression and Mitochondrial Function in Response to 2 Weeks of Enriched Environment Training After Cerebral Ischaemia-Reperfusion Injury.

An enriched environment (EE) can stimulate the recovery of neurological function following a cerebral ischaemia-reperfusion injury; however, the impact of EE's on mitochondrial function has been insufficiently studied. Our research aimed to assess whether EE's therapeutic impact involved the enhancement of mitochondrial dysfunction. Following 2 weeks of EE training, we tested both mitochondrial function and mitochondria-associated protein expression within the cerebral cortex following cerebral ischaemia-reperfusion injury. We subjected Sprague-Dawley rats to transient focal cerebral ischaemia and categorized the rats into three separate groups, i.e. an enriched environment (EE) group, a standard condition (SC) group and a sham control group (no middle cerebral artery embolization). The rats within the EE group were raised in enriched conditions for 2 weeks, while the rats within the SC group, in comparison, were reared in standard conditions for 2 weeks. After 2 weeks, the cerebral cortices of the rats were removed. We then measured a series of indices, i.e. the protein expression of peroxisome proliferator-activated receptor gamma coactivator (PGC-1α), nuclear respiratory factor-1 (NRF-1), mitochondrial transcription factor A (TFAM) and mitochondrial protein cytochrome C oxidase subunit IV (COX IV). Furthermore, the number of mitochondria was evaluated through electron microscopy.EE upregulated the protein expression of PGC-1α, NRF-1 as well as TFAM, which function as the master regulators of mitochondrial biogenesis, in comparison with the SC group. The EE group's COX IV protein expression also exhibited an increase. Moreover, the amount of mitochondria in the peri-infarct region of the cortex increased as result of EE training. Over 2 weeks, EE training significantly increased mitochondrial biogenesis-associated protein expression and mitochondrial function. A possible mechanism of the EE leading to the improvement of neurological function is that it increases brain mitochondrial biogenesis after the rats' cerebral ischaemia-reperfusion injury. Mitochondrial biogenesis stimulation or enhancement could become an innovative strategy for neuroprotection in future treatment.

Mitochondrial damage are involved in Aflatoxin B1-induced testicular damage and spermatogenesis disorder in mice

Aflatoxin B1 (AFB1) is an unavoidable environmental pollutants, which seriously endangers human and animal health. AFB1 has male reproductive toxicity, yet the underlying mechanisms remain inconclusive. Mitochondra are a kind of crucial organelle for maintaining spermatogenesis in testis. Thus, we hypothesized that AFB1 can impair mitochondria to aggravate testicular damage and spermatogenesis disorder. To verify this hypothesis, 48 male mice were intragastrically administered with 0, 0.375, 0.75 or 1.5 mg/kg body weight AFB1 for 30 days, respectively. In this study, we found AFB1 caused testicular histopathological lesions and spermatogenesis abnormalities, with the elevation of oxidative stress (increased H2O2, whereas decreased SOD and GSH). Significant mitochondria structure damage of germ cells and Leydig cells, MMP loss, ATP contents reduction, and inhibited activities of mitochondrial complexes I-IV in mice testis were found in AFB1 treatment groups. Besides, AFB1 inhibited mitochondrial biogenesis and mitochondrial dynamics, presenting as the decreased mRNA and protein expressions of PGC-1α, Nrf1, Tfam, Drp1, Fis1, Mfn1 and Opa1. The results revealed that the mitochondrial damage were involved in AFB1-induced testicular damage and spermatogenesis disorder, providing a considerable direction to clarify potential mechanisms of AFB1 reproductive toxicity.