Sevoflurane Postconditioning Upregulates HIF-1α Pathway Enhances BNIP3 Mediated Mitochondrial Autophagy in Myocardium of Aged Mice

BACKGROUND: Diminished mitochondrial autophagy in aged myocardium may be due to impaired HIF-1α protein expression. Previous studies confirmed that upregulation of HIF-1α expression protects myocardial tissue from ischemia/reperfusion (I/R) injury and found that sevoflurane post-conditioning (SpostC) mediated mitochondrial autophagy plays a significant role in myocardial protection. However, the protective mechanism of SpostC in aged myocardium is unclear. This study aimed to investigate whether SpostC regulates BNIP3 - mediated mitochondrial autophagy by upregulating HIF-1α expression, thus alleviating myocardial I/R injury in aged mice.Methods: An in vivo mouse model of myocardial I/R injury was established and treated with sevoflurane at the time of reperfusion, and at the end of reperfusion, echocardiographic changes, myocardial infarct size, mitochondrial ultrastructure, and autophagosomes were measured, mitochondrial respiratory function and enzyme activity were detected, serum LDH, CKM, CK-MB, TNNT2, IL-6 levels were determined, and Western blot was used to examine the expression levels of phosphorylated HIF-1α, LC3-II, BNIP3, Beclin1, TLR9, and IL-6 protein in myocardial tissue.RESULTS: In young, healthy myocardium, SpostC upregulated the expression of HIF-1α, activated the downstream target gene BNIP3 protein, and upregulated the expression levels of autophagy essential proteins LC3-II, Beclin-1, and TLR9, attenuated myocardial oxidative stress injury, stabilized mitochondrial ultrastructure, inhibited cardiomyocyte apoptosis, and ultimately reduced myocardial infarct size. In aged myocardium, SpostC also played an excellent myocardial protective role.CONCLUSION: SpostC was able to upregulate HIF-1α expression, promote BNIP3-mediated mitochondrial autophagy, reduce myocardial infarct size, and alleviate myocardial I/R injury in aged mice.

Effects of Bushen-Tiaojing-Fang on the pregnancy outcomes of infertile patients with repeated controlled ovarian stimulation

Bushen-Tiaojing-Fang (BSTJF) is commonly used to treat infertility. This study investigated the effects of BSTJF on the pregnancy outcomes of patients with repeated controlled ovarian stimulation (COS), on mitochondrial function, and on oxidative stress in ovarian granulosa cells (GCs) and follicular fluid (FF). The samples and clinical data of 97 patients, including 35 in the control group, 29 in the placebo group and 33 in the BSTJF group, were collected for this study. The mitochondrial ultrastructure, ATP content, mitochondrial DNA (mtDNA) number, 8-hydroxy-2-deoxyguanosine (8-OHdG), Mn-superoxide dismutase (Mn-SOD), glutathione peroxidase (GSH-Px) activity levels, and mRNA expression levels of Mn-SOD, GSH-Px, and nuclear factor erythroid-derived factor 2-related factor 2 (Nrf2) were analyzed. The high-grade embryo (P < 0.001), implantation (P = 0.033), and clinical pregnancy (P = 0.031) rates, as well as the ATP content (P = 0.014), mtDNA number (P = 0.035), GSH-Px activity (P = 0.004 in GCs and P = 0.008 in FF) and mRNA expression levels (P = 0.019), were significantly lower in the placebo group than in the control group, whereas the 8-OHdG content was significantly (P = 0.006 in FF) higher in the placebo group than in the control group. Compared with those in the placebo group, the high-grade embryo rate (P = 0.007), antioxidant enzyme activity (P = 0.037 and 0.036 in Mn-SOD; P = 0.047 and 0.030 in GSH-Px) and mRNA level (P < 0.001 in Nrf2, P = 0.039 in Mn-SOD and P = 0.002 in GSH-Px) were significantly higher in the BSTJF group, as were changes in mitochondrial ultrastructure, ATP (P = 0.040) and mtDNA number (P = 0.013). In conclusion, BSTJF can improve oxidative stress in patients with repeated COS and pregnancy outcomes.

Extract of Cynomorium songaricum ameliorates mitochondrial ultrastructure impairments and dysfunction in two different in vitro models of Alzheimer’s disease

Background Alzheimer’s disease (AD) is one of the most common neurodegenerative disorders, but there is still no effective way to stop or slow its progression. Our previous studies demonstrated that extract of Cynomorium songaricum (ECS), a Chinese herbal medicine, had neuroprotective effects in AD models in vivo. However, the pharmacological mechanism of ECS in AD is still unclear. Methods To study the mechanisms of action of the effects of ECS on AD, we used Aβ25–35- and H2O2-exposed HT22 cells to mimic specific stages of AD in vitro. The mitochondrial membrane potential (MMP), intracellular ATP, intracellular reactive oxygen species (ROS), and expression levels of mitochondrial dynamics-related proteins in each group were examined. Furthermore, we explored the mechanisms by which ECS reduces the phosphorylation of Drp1 at Ser637 and the changes in the concentrations of intracellular calcium ions in the two models after FK506 intervention. Results The results showed that ECS significantly enhanced the MMP (P < 0.05), increased intracellular ATP levels (P < 0.05) and decreased intracellular ROS levels in the Aβ- and H2O2-induced cell models (P < 0.05). Additionally, ECS regulated the expression levels of mitochondrial dynamics-related proteins by reducing the phosphorylation of Drp1 at Ser637 (P < 0.05) and decreasing the expression of Fis1 in the H2O2-induced models (P < 0.05). Further study indicated that ECS reduced the overload of intracellular calcium (P < 0.05). Conclusion Our study results suggest that ECS protects the mitochondrial ultrastructure, ameliorates mitochondrial dysfunction, and maintains mitochondrial dynamics in AD models.

Pentoxifylline Enhances Antioxidative Capability and Promotes Mitochondrial Biogenesis in D-Galactose-Induced Aging Mice by Increasing Nrf2 and PGC-1α through the cAMP-CREB Pathway

Aging is a complex phenomenon associated with oxidative stress and mitochondrial dysfunction. The objective of this study was to investigate the potential ameliorative effects of the phosphodiesterase inhibitor pentoxifylline (PTX) on the aging process and its underlying mechanisms. We treated D-galactose- (D-gal-) induced aging mice with PTX and measured the changes in behavior, degree of oxidative damage, and mitochondrial ultrastructure and content as well as the expression of nuclear factor erythroid 2-related factor 2- (Nrf2-) mediated antioxidant genes and peroxisome proliferator-activated receptor-gamma coactivator 1-alpha- (PGC-1α-) dependent mitochondrial biogenesis genes. The results demonstrated that PTX improved cognitive deficits, reduced oxidative damage, ameliorated abnormal mitochondrial ultrastructure, increased mitochondrial content and Nrf2 activation, and upregulated antioxidant and mitochondrial biogenesis gene expression in the hippocampus of wild-type aging mice. However, the above antiaging effects of PTX were obviously decreased in the brains of Nrf2-deficient D-gal-induced aging mice. Moreover, in hydrogen peroxide-treated SH-SY5Y cells, we found that cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) and Nrf2/PGC-1α act in a linear way by CREB siRNA transfection. Thus, PTX administration improved the aging-related decline in brain function by enhancing antioxidative capability and promoting mitochondrial biogenesis, which might depend on increasing Nrf2 and PGC-1α by activating the cAMP-CREB pathway.

Qiangji Jianli Decoction Alleviates Hydrogen Peroxide-Induced Mitochondrial Dysfunction via Regulating Mitochondrial Dynamics and Biogenesis in L6 Myoblasts

Oxidative stress can cause the excessive generation of reactive oxygen species (ROS) and has various adverse effects on muscular mitochondria. Qiangji Jianli decoction (QJJLD) is an effective traditional Chinese medicine (TCM) that is widely applied to improve muscle weakness, and it has active constituents that prevent mitochondrial dysfunction. To investigate the protective mechanism of QJJLD against hydrogen peroxide- (H2O2-) mediated mitochondrial dysfunction in L6 myoblasts. Cell viability was determined with MTT assay. Mitochondrial ultrastructure was detected by transmission electron microscope (TEM). ROS and mitochondrial membrane potential (MMP) were analyzed by fluorescence microscope and flow cytometry. The superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) activity, and malondialdehyde (MDA) level were determined by WST-1, TBA, and DTNB methods, respectively. The mRNA and protein levels were measured by quantitative real-time PCR (qRT-PCR) and Western blot. The cell viability was decreased, and the cellular ROS level was increased when L6 myoblasts were exposed to H2O2. After treatment with QJJLD-containing serum, the SOD and GSH-Px activities were increased. MDA level was decreased concurrently. ROS level was decreased while respiratory chain complex activity and ATP content were increased in L6 myoblasts. MMP loss was attenuated. Mitochondrial ultrastructure was also improved. Simultaneously, the protein expressions of p-AMPK, PGC-1α, NRF1, and TFAM were upregulated. The mRNA and protein expressions of Mfn1/2 and Opa1 were also upregulated while Drp1 and Fis1 were downregulated. These results suggest that QJJLD may alleviate mitochondrial dysfunction through the regulation of mitochondrial dynamics and biogenesis, the inhibition of ROS generation, and the promotion of mitochondrial energy metabolism.

Small mitochondrial protein NERCLIN regulates cardiolipin homeostasis and mitochondrial ultrastructure

ABSTRACTCardiolipin (CL) is an essential phospholipid for mitochondrial structure and function. Here we present a small mitochondrial protein, NERCLIN, as a negative regulator of CL homeostasis and mitochondrial ultrastructure. Primate-specific NERCLIN is expressed ubiquitously from GRPEL2 locus on a tightly regulated low level, but induced by heat stress. NERCLIN overexpression severely disrupts mitochondrial cristae structure and induces mitochondrial fragmentation. Proximity labeling suggested interactions of NERCLIN with CL synthesis and prohibitin complexes on the matrix side of the inner mitochondrial membrane. Lipid analysis indicated that NERCLIN regulates mitochondrial CL content. The regulation may occur directly through interaction with PTPMT1, a proximal partner on the CL synthesis pathway, as its product phosphatidylglycerol was also reduced by NERCLIN. We propose that NERCLIN contributes to stress-induced adaptation of mitochondrial dynamics and turnover by regulating the mitochondrial CL content. Our findings add NERCLIN to the group of recently identified small mitochondrial proteins with important regulatory functions.

Protease OMA1 Modulates Mitochondrial Bioenergetics and Ultrastructure through Dynamic Association with MICOS Complex

ABSTRACTRemodeling of mitochondrial ultrastructure is a complex dynamic process that is critical for a variety of mitochondrial functions and apoptosis. Although the key regulators of this process - mitochondrial contact site and cristae junction organizing system (MICOS) and GTPase Optic Atrophy 1 (OPA1) have been characterized, the mechanisms behind this regulation remain incompletely defined. Here, we found that in addition to its role in mitochondrial division, metallopeptidase OMA1 is required for maintenance of contacts between the inner and outer membranes through a dynamic association with MICOS. This association is independent of OPA1, appears to be mediated via the MICOS subunit MIC60, and is important for stability of MICOS machinery and the inner-outer mitochondrial membrane contacts. We find that OMA1-MICOS relay is required for stability of respiratory supercomplexes, optimal bioenergetic output in response to cellular insults, and apoptosis. Loss of OMA1 affects these activities; remarkably it can be partially compensated for by an artificial MICOS-emulating tether protein that bridges the inner and outer mitochondrial membranes. Our data show that OMA1-mediated support of mitochondrial ultrastructure is required for maintenance of mitochondrial architecture and bioenergetics under both basal and homeostasis-challenging conditions, and suggest a previously unrecognized role for this protease in mitochondrial physiology.

Role of IRE1α in podocyte proteostasis and mitochondrial health

Glomerular epithelial cell (GEC)/podocyte proteostasis is dysregulated in glomerular diseases. The unfolded protein response (UPR) is an adaptive pathway in the endoplasmic reticulum (ER) that upregulates proteostasis resources. This study characterizes mechanisms by which inositol requiring enzyme-1α (IRE1α), a UPR transducer, regulates proteostasis in GECs. Mice with podocyte-specific deletion of IRE1α (IRE1α KO) were produced and nephrosis was induced with adriamycin. Compared with control, IRE1α KO mice had greater albuminuria. Adriamycin increased glomerular ER chaperones in control mice, but this upregulation was impaired in IRE1α KO mice. Likewise, autophagy was blunted in adriamycin-treated IRE1α KO animals, evidenced by reduced LC3-II and increased p62. Mitochondrial ultrastructure was markedly disrupted in podocytes of adriamycin-treated IRE1α KO mice. To pursue mechanistic studies, GECs were cultured from glomeruli of IRE1α flox/flox mice and IRE1α was deleted by Cre–lox recombination. In GECs incubated with tunicamycin, deletion of IRE1α attenuated upregulation of ER chaperones, LC3 lipidation, and LC3 transcription, compared with control GECs. Deletion of IRE1α decreased maximal and ATP-linked oxygen consumption, as well as mitochondrial membrane potential. In summary, stress-induced chaperone production, autophagy, and mitochondrial health are compromised by deletion of IRE1α. The IRE1α pathway is cytoprotective in glomerular disease associated with podocyte injury and ER stress.