Transcription Factors For Mitochondrial Biogenesis Research Articles

Anomalous AMPK-regulated angiotensin AT1R expression and SIRT1-mediated mitochondrial biogenesis at RVLM in hypertension programming of offspring to maternal high fructose exposure

BackgroundTissue oxidative stress, sympathetic activation and nutrient sensing signals are closely related to adult hypertension of fetal origin, although their interactions in hypertension programming remain unclear. Based on a maternal high-fructose diet (HFD) model of programmed hypertension, we tested the hypothesis that dysfunction of AMP-activated protein kinase (AMPK)-regulated angiotensin type 1 receptor (AT1R) expression and sirtuin1 (SIRT1)-dependent mitochondrial biogenesis contribute to tissue oxidative stress and sympathoexcitation in programmed hypertension of young offspring.MethodsPregnant female rats were randomly assigned to receive normal diet (ND) or HFD (60% fructose) chow during pregnancy and lactation. Both ND and HFD offspring returned to ND chow after weaning, and blood pressure (BP) was monitored from age 6 to 12 weeks. At age of 8 weeks, ND and HFD offspring received oral administration of simvastatin or metformin; or brain microinfusion of losartan. BP was monitored under conscious condition by the tail-cuff method. Nutrient sensing molecules, AT1R, subunits of NADPH oxidase, mitochondrial biogenesis markers in rostral ventrolateral medulla (RVLM) were measured by Western blot analyses. RVLM oxidative stress was measured by fluorescent probe dihydroethidium and lipid peroxidation by malondialdehyde assay. Mitochondrial DNA copy number was determined by quantitative real-time polymerase chain reaction.ResultsIncreased systolic BP, plasma norepinephrine level and sympathetic vasomotor activity were exhibited by young HFD offspring. Reactive oxygen species (ROS) level was also elevated in RVLM where sympathetic premotor neurons reside, alongside augmented protein expressions of AT1R and pg91phox subunit of NADPH oxidase, decrease in superoxide dismutase 2; and suppression of transcription factors for mitochondrial biogenesis, peroxisome proliferator-activated receptor γ co-activator α (PGC-1α) and mitochondrial transcription factor A (TFAM). Maternal HFD also attenuated AMPK phosphorylation and protein expression of SIRT1 in RVLM of young offspring. Oral administration of a HMG-CoA reductase inhibitor, simvastatin, or an AMPK activator, metformin, to young HFD offspring reversed maternal HFD-programmed increase in AT1R and decreases in SIRT1, PGC-1α and TFAM; alleviated ROS production in RVLM, and attenuated sympathoexcitation and hypertension.ConclusionDysfunction of AMPK-regulated AT1R expression and SIRT1-mediated mitochondrial biogenesis may contribute to tissue oxidative stress in RVLM, which in turn primes increases of sympathetic vasomotor activity and BP in young offspring programmed by excessive maternal fructose consumption.

Anomalous AMPK‐Regulated Angiotensin AT1R Expression and SIRT1‐Mediated Mitochondrial Biogenesis at RVLM in Hypertension Programming of Offspring to Maternal High Fructose Exposure

Tissue oxidative stress, sympathetic activation and nutrient sensing signals are closely related to adult hypertension of fetal origin, although their interactions in hypertension programming remain unclear. Based on a maternal high‐fructose diet (HFD) model of programmed hypertension, we tested the hypothesis that dysfunction of AMP‐activated protein kinase (AMPK)‐regulated angiotensin type 1 receptor (AT1R) expression and sirtuin1 (SIRT1)‐dependent mitochondrial biogenesis contribute to tissue oxidative stress and sympathoexcitation in programmed hypertension of young offspring. Increased systolic and diastolic blood pressure, plasma norepinephrine level and sympathetic vasomotor activity were exhibited by young HFD offspring. Reactive oxygen species (ROS) level was also elevated in rostral ventrolateral medulla (RVLM) where sympathetic premotor neurons reside, alongside augmented protein expressions of AT1R and pg91phox subunit of NADPH oxidase, decrease in superoxide dismutase 2; and suppression of transcription factors for mitochondrial biogenesis, peroxisome proliferator‐activated receptor g co‐activator a (PGC‐1a) and mitochondrial transcription factor A (TFAM). Maternal HFD also attenuated AMPK phosphorylation and protein expression of SIRT1 in RVLM of young offspring. Oral administration of a HMG‐CoA reductase inhibitor, simvastatin, or an AMPK activator, metformin, to young HFD offspring reversed maternal HFD‐programmed increase in AT1R and decreases in SIRT1, PGC‐1a and TFAM; alleviated ROS production in RVLM, and attenuated sympathoexcitation and hypertension. Together these results suggest that dysfunction of AMPK‐regulated AT1R expression and SIRT1‐mediated mitochondrial biogenesis may contribute to tissue oxidative stress in RVLM, which in turn primes increases of sympathetic vasomotor activity and blood pressure in young offspring programmed by excessive maternal fructose consumption.Support or Funding InformationThis work was supported by research grants from the Chang Gung Medical Foundation to J.Y.H. Chan (CMRPG8C0053‐33 and OMRPG8G0011). Y.M. Chao was supported by a Postdoctoral Fellowship (MOST107‐2811‐B‐182A‐501) from the Ministry of Science and Technology, Taiwan.

CHIR-99021 regulates mitochondrial remodelling via β-catenin signalling and miRNA expression during endodermal differentiation.

Mitochondrial remodelling is a central feature of stem cell differentiation. However, little is known about the regulatory mechanisms during these processes. Previously, we found that a pharmacological inhibitor of glycogen synthase kinase-3α and -3β, CHIR-99021, initiates human adipose stem cell differentiation into human definitive endodermal progenitor cells (hEPCs), which were directed to differentiate synchronously into hepatocyte-like cells after further treatment with combinations of soluble factors. In this study, we show that CHIR-99021 promotes mitochondrial biogenesis, the expression of PGC-1α (also known as PPARGC1A), TFAM and NRF1 (also known as NFE2L1), oxidative phosphorylation capacities, and the production of reactive oxygen species in hEPCs. Blocking mitochondrial dynamics using siRNA targeting DRP1 (also known as DNM1L) impaired definitive endodermal differentiation. Downregulation of β-catenin (CTNNB1) expression weakened the effect of CHIR-99021 on the induction of mitochondrial remodelling and the expression of transcription factors for mitochondrial biogenesis. Moreover, CHIR-99021 decreased the expression of miR-19b-2-5p, miR-23a-3p, miR-23c, miR-130a-3p and miR-130a-5p in hEPCs, which target transcription factors for mitochondrial biogenesis. These data demonstrate that CHIR-99021 plays a role in mitochondrial structure and function remodelling via activation of the β-catenin signalling pathway and inhibits the expression of miRNAs during definitive endodermal differentiation.This article has an associated First Person interview with the first author of the paper.

Influence of maternal protein malnutrition on oxidative stress and regulators of mitochondrial biogenesis in female rat hearts over succeeding generations

AimsWe sought to evaluate the effects of maternal protein restriction (LP) on oxidative balance and transcription factors for mitochondrial biogenesis in the hearts of young female rats of both the first (F1) and second (F2) generation. Main methodsWe evaluated oxidative stress biomarkers (lipid peroxidation and protein oxidation), enzymatic antioxidant defense (activity of superoxide dismutase-SOD, catalase, and glutathione-S-transferase-GST), nonenzymatic antioxidant defense (reduced glutathione-GSH and sulfhydryl groups) and gene expression of AMPK, PGC-1α and TFAM. Key findingsInterestingly, lipid peroxidation was decreased (49%, p < 0.001) in the LP-F1 group and 59% (p < 0.001) in LP-F2. In enzymatic defense, we observed increases in SOD activity in the LP-F1 group (79%, p = 0.036) and in CAT activity (approximately 40%, p = 0.041). GSH was increased in F2 in both groups (LP 546%, p < 0.0001 and in NP 491.7%, p < 0.0001). With respect to mitochondrial biogenesis gene transcription, we observed a decrease in AMPK (60%, p < 0. 0001) and an increase in PGC-1α (340%, p < 0.001) in LP compared to NP in the F1 generation. TFAM was decreased in LP-F2L compared to NP-F2L (42%, p = 0.0069) and increased in LP-F2 compared to LP-F1 (160%, p = 0.0037). SignificanceOur study contributes to knowledge of inheritance, showing that despite the potential mitochondrial ‘inheritance’ of cardiovascular damage caused by maternal malnutrition, that damage is not cross-generational and can be eliminated with proper nutrition in the F1 generation.

EBV reduces autophagy, intracellular ROS and mitochondria to impair monocyte survival and differentiation

ABSTRACTEBV has been reported to impair monocyte in vitro differentiation into dendritic cells (DCs) and reduce cell survival. In this study, we added another layer of knowledge to this topic and showed that these effects correlated with macroautophagy/autophagy, ROS and mitochondrial biogenesis reduction. Of note, autophagy and ROS, although strongly interconnected, have been separately reported to be induced by CSF2/GM-CSF (colony stimulating factor 2) and required for CSF2-IL4-driven monocyte in vitro differentiation into DCs. We show that EBV infects monocytes and initiates a feedback loop in which, by inhibiting autophagy, reduces ROS and through ROS reduction negatively influences autophagy. Mechanistically, autophagy reduction correlated with the downregulation of RAB7 and ATG5 expression and STAT3 activation, leading to the accumulation of SQSTM1/p62. The latter activated the SQSTM1-KEAP1- NFE2L2 axis and upregulated the anti-oxidant response, reducing ROS and further inhibiting autophagy. ROS decrease correlated also with the reduction of mitochondria, the main source of intracellular ROS, achieved by the downregulation of NRF1 and TFAM, mitochondrial biogenesis transcription factors. Interestingly, mitochondria supply membranes and ATP required for autophagy execution, thus their reduction may further reduce autophagy in EBV-infected monocytes. In conclusion, this study shows for the first time that the interconnected reduction of autophagy, intracellular ROS and mitochondria mediated by EBV switches monocyte differentiation into apoptosis, giving new insights into the mechanisms through which this virus reduces immune surveillance.Abbreviations: ACTB: actin beta; ATG5: autophagy related 5; BAF: bafilomycin A1; BECN1: beclin 1; CAT: catalase; CSF2: colony stimulating factor 2; CT: control; CYCS (cytochrome C: somatic); DCs: dendritic cells; EBV: Epstein-Barr virus; GSR: glutathione-disulfide reductase; KEAP1: kelch like ECH associated protein 1; IL4: interleukin 4; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MET: metformin; NAC: N-acetylcysteine; NFE2L2/NRF2 nuclear factor: erythroid 2 like 2; NRF1 (nuclear respiratory factor 1); clPARP1: cleaved poly(ADP-ribose) polymerase; Rapa: Rapamycin; ROS: reactive oxygen species; SQSTM1/p62: sequestosome 1; TFAM: (transcription factor A: mitochondrial); TUBA1A: tubulin alpha 1a.

Carbon black suppresses the osteogenesis of mesenchymal stem cells: the role of mitochondria

BackgroundThe rapid increase in carbon black poses threats to human health. We evaluated the effect of CB (Printex 90) on the osteogenesis of bone-marrow-derived mesenchymal stem cells (MSCs). Mitochondria play an important role in the osteogenesis of MSCs and are potential targets of nanomaterials, so we studied the role of mitochondria in the CB Printex 90-induced effects on osteogenesis.ResultsLow doses of Printex 90 (3 ng/mL and 30 ng/mL) that did not cause deleterious effects on MSCs’ viability significantly inhibited osteogenesis of MSCs. Printex 90 caused down-regulation of osteoblastic markers, reduced activity of alkaline phosphatase (ALP), and poor mineralization of osteogenically induced MSCs. Cellular ATP production was decreased, mitochondrial respiration was impaired with reduced expression of ATPase, and the mitochondrial membrane was depolarized. The quantity and quality of mitochondria are tightly controlled by mitochondrial biogenesis, mitochondrial dynamics and mitophagy. The transcriptional co-activator and transcription factors for mitochondrial biogenesis, PGC-1α, Nrf1 and TFAM, were suppressed by Printex 90 treatment, suggesting that decreased biogenesis was caused by Printex 90 treatment during osteogenesis. Mitochondrial fusion and fission were significantly inhibited by Printex 90 treatment. PINK1 accumulated in Printex 90-treated cells, and more Parkin was recruited to mitochondria, indicating that mitophagy increased to remove the damaged mitochondria.ConclusionsThis is the first report of the inhibitory effects of CB on the osteogenesis of MSCs and the involvement of mitochondria in CB Printex 90-induced suppression of MSC osteogenesis.

Standardized Kaempferia parviflora Extract Enhances Exercise Performance Through Activation of Mitochondrial Biogenesis.

Exercise enhances mitochondrial biogenesis in skeletal muscle. Increased mitochondrial function and content can contribute to the improvement in skeletal muscle function and the benefits of exercise by increasing the response to energy demands. The effect of standardized Kaempferia parviflora extract (KPE) on exercise performance was accessed in L6 myotubes and C57BL/6J mice. KPE significantly activated peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) and increased mitochondrial density in L6 myotubes. KPE also upregulated the expression of transcription factors for mitochondrial biogenesis (estrogen-related receptor-α [ERRα], nuclear respiratory factor-1 [NRF-1], and mitochondrial transcription factor A [Tfam]) through activation of PGC-1α in L6 myotubes. In vivo models including normal diet mice and high-fat diet obese mice showed that KPE effectively enhanced running endurance and increased the skeletal muscle weight/body weight ratio. Furthermore, these observations were associated with a significant upregulation of mitochondrial biogenesis regulatory genes in skeletal muscle tissue. KPE enhanced the protein expression of the sirtuin 1 (SIRT1)/adenosine monophosphate (AMP)-activated protein kinase (AMPK)/PGC-1α/peroxisome proliferator-activated receptor-δ (PPARδ) signaling pathway components in vitro and in vivo, acting as an exercise metabolism regulator. These results suggest that KPE has the potential to enhance exercise performance through mitochondrial biogenesis and the SIRT1/AMPK/PGC-1α/PPARδ signaling pathways.

Impaired Nrf2 regulation of mitochondrial biogenesis in rostral ventrolateral medulla on hypertension induced by systemic inflammation

Oxidative stress in rostral ventrolateral medulla (RVLM), where sympathetic premotor neurons reside, is involved in the development of hypertension under systemic inflammation. Mitochondrial dysfunction contributes to tissue oxidative stress. In this study, we sought to investigate whether hypertension developed under systemic inflammation is attributable to impaired mitochondrial biogenesis in RVLM. In normotensive Sprague-Dawley rats, intraperitoneal infusion of a low dose Escherichia coli lipopolysaccharide (LPS) for 7 days promoted a pressor response, alongside a decrease in mitochondrial DNA (mtDNA) copy number, reductions in protein expression of nuclear DNA-encoded transcription factors for mitochondrial biogenesis, including mitochondrial transcription factor A (TFAM) and nuclear factor erythroid-derived 2-like 2 (Nrf2), and suppression of nuclear translocation of the phosphorylated Nrf2 (p-Nrf2) in RVLM neurons; all of which were abrogated by treatment with intracisternal infusion of an interleukin-1β (IL-1β) blocker, IL-1Ra, or a mobile mitochondrial electron carrier, coenzyme Q10 (CoQ10). Microinjection into RVLM of IL-1β suppressed the expressions of p-Nrf2 and TFAM, and evoked a pressor response; conversely, the Nrf2 inducer, tert-butylhydroquinone, lessened the LPS-induced suppression of TFAM expression and pressor response. At cellular level, exposure of neuronal N2a cells to IL-1β decreased mtDNA copy number, increased protein interaction of Nrf2 to its negative regulator, kelch-like ECH-associated protein 1 (Keap1), and reduced DNA binding activity of p-Nrf2 to Tfam gene. Together these results indicate that defect mitochondrial biogenesis in RVLM neurons entailing redox-sensitive and IL-1β-dependent suppression of TFAM because of the increase in the formation of Keap1/Nrf2 complex, reductions in nuclear translocation of the activated Nrf2 and its binding to the Tfam gene promoter may underlie hypertension developed under the LPS-induced systemic inflammation.

Sensing endoplasmic reticulum stress by protein kinase RNA‐like endoplasmic reticulum kinase promotes adaptive mitochondrial DNA biogenesis and cell survival via heme oxygenase‐1/carbon monoxide activity

Endoplasmic reticulum (ER) stress activates the adaptive unfolded protein response, allowing cells to recover folding capacity in the organelle. However, the overwhelming response to severe damage results in apoptotic cell death. Because of the physical proximity between ER and mitochondria, a functional interrelationship between these two organelles, including mitochondrial ATP production and apoptosis, has been suggested. The adaptive response to ER stress includes the maintenance of cellular energetics, which eventually determines cell fate. We previously demonstrated that heme oxygenase-1 (HO-1) activity protects cells against ER stress in a protein kinase RNA-like endoplasmic reticulum kinase (PERK)-dependent pathway. Here, we provide evidence that PERK-mediated induction of HO-1 in murine macrophages, RAW264.7, relays ER stress to mitochondrial DNA (mtDNA) replication and function. ER stress induced by thapsigargin treatments (10-100 nM) resulted in a 2-fold increase in mtDNA contents compared with that in the untreated control. HO-1 activity on ER stress is proven to be critical for mitochondrial integrity because chemical inhibition (zinc protoporphyrin, 5-20 μM) and genetic depletion of HO-1 by small interference RNA transfection suppress the activation of transcription factors for mitochondrial biogenesis. Carbon monoxide (CO), an enzymatic by-product of HO-1 activity is responsible for the function of HO-1. Limited bioavailability of CO by hemoglobin treatment triggers cell death with a concomitant decline in ATP production. Approximately 78.1% of RAW264.7 cells were damaged in the presence of hemoglobin compared with the percentage of injured cells (26.9%) under ER stress alone. Mitochondrial generation of ATP levels significantly declined when CO availability was limited under prolonged ER stress. Taken together, these results suggest that the cellular HO-1/CO system conveys ER stress to cell survival signals from mitochondria via both the activation of transcriptional factors and functional integrity of mtDNA.

Ovariectomy decreases expression of transcription factors for mitochondrial biogenesis in mouse cerebrovascular endothelial cells

Estrogen has protective effects on the cerebrovasculature in physiological as well as pathological states such as stroke. However teasing out effects on individual cell types remains elusive. Thus we adapted Dynabeads (Invitrogen) technology using PECAM1 antibodies to isolate brain vascular endothelial cells. We used appropriate cellular markers to demonstrate enrichment of brain endothelial cells as well as decreased presence of other cell types. We developed a method to correct for variations in endothelial enrichment among different preparations. We found that the level of eNOS mRNA is two‐fold higher in endothelial cells from intact females compared to ovariectomized mice. This agrees with our previous observation of higher eNOS in brain vessels from estrogen‐exposed mice. We also evaluated levels of mitochondrial transcription factors in brain endothelial cells. We observed 61% and 38% higher mRNA levels of NRF1 and TFAM, respectively, in endothelial cells from intact females relative to ovariectomized mice. These two factors are essential in maintenance of respiration as well as mitochondrial biogenesis. The findings extend our previous observations that estrogen impacts cerebrovascular mitochondria. This is the first time that female reproductive competency has been shown to influence endothelial cell master regulators of mitochondrial function.

Endurance Exercise as a Countermeasure for Aging

OBJECTIVE— We determined whether reduced insulin sensitivity, mitochondrial dysfunction, and other age-related dysfunctions are inevitable consequences of aging or secondary to physical inactivity.RESEARCH DESIGN AND METHODS— Insulin sensitivity was measured by hyperinsulinemic-euglycemic clamp and ATP production in mitochondria isolated from vastus lateralis biopsies of 42 healthy sedentary and endurance-trained young (18–30 years old) and older (59–76 years old) subjects. Expression of proteins involved in fuel metabolism was measured by mass spectrometry. Citrate synthase activity, mitochondrial DNA (mtDNA) abundance, and expression of nuclear-encoded transcription factors for mitochondrial biogenesis were measured. SIRT3, a mitochondrial sirtuin linked to lifespan-enhancing effects of caloric restriction, was measured by immunoblot.RESULTS— Insulin-induced glucose disposal and suppression of endogenous glucose production were higher in the trained young and older subjects, but no age effect was noted. Age-related decline in mitochondrial oxidative capacity was absent in endurance-trained individuals. Although endurance-trained individuals exhibited higher expression of mitochondrial proteins, mtDNA, and mitochondrial transcription factors, there were persisting effects of age. SIRT3 expression was lower with age in sedentary but equally elevated regardless of age in endurance-trained individuals.CONCLUSIONS— The results demonstrate that reduced insulin sensitivity is likely related to changes in adiposity and to physical inactivity rather than being an inevitable consequence of aging. The results also show that regular endurance exercise partly normalizes age-related mitochondrial dysfunction, although there are persisting effects of age on mtDNA abundance and expression of nuclear transcription factors and mitochondrial protein. Furthermore, exercise may promote longevity through pathways common to effects of caloric restriction.