Calcium ions (Ca2+) play pivotal roles in regulating numerous cellular functions, including metabolism and growth, in normal and cancerous cells. Consequently, Ca2+ signaling is a vital determinant of cell fate and influences both cell survival and death. These intracellular signals are susceptible to modulation by various factors, including changes in the extracellular environment, which leads to mechanical alterations. However, the effect of extracellular matrix (ECM) stiffness variations on intracellular Ca2+ signaling remains underexplored. In this study, we aimed to elucidate the mechanisms of Ca2+ regulation through the mitochondria, which are crucial to Ca2+ homeostasis. We investigated how Ca2+ regulatory mechanisms adapt to different levels of ECM stiffness by simultaneously imaging the mitochondria and endoplasmic reticulum (ER) in live cells using genetically encoded biosensors. Our findings revealed that the uptake of mitochondrial Ca2+ through Voltage-Dependent Anion Channel 1 (VDAC1), facilitated by intracellular tubulin, is influenced by ECM stiffness. Unraveling these Ca2+ regulatory mechanisms under various conditions offers a novel perspective for advancing biomedical studies involving Ca2+ signaling.

Circular RNAs (circRNAs) are a subclass of non-coding RNAs that are important for the regulation of gene expression in eukaryotic organisms. CircRNAs exert various regulatory roles in cancer progression. However, the role of hsa_circ_0064636 in osteosarcoma (OS) remains poorly understood. In the present study, the expression of hsa_circ_0064636 in OS cell lines was measured by reverse transcription-quantitative PCR (RT-qPCR). Differentially expressed mRNAs and microRNAs (miRNA or miRs) were screened using mRNA(GSE16088) and miRNA(GSE65071) expression datasets for OS. miRNAs that can potentially interact with hsa_circ_0064636 were predicted using RNAhybrid, TargetScan and miRanda. Subsequently, RNAhybrid, TargetScan, miRanda, miRWalk, miRMap and miRNAMap were used for target gene prediction based on the overlapping miRNAs to construct a circ/miRNA/mRNA interaction network. Target genes were subjected to survival analysis using PROGgeneV2, resulting in a circRNA/miRNA/mRNA interaction sub-network with prognostic significance. miRNA and circRNA in the subnetwork may also have survival significance, but relevant data are lacking and needs to be further proved. RT-qPCR demonstrated that hsa_circ_0064636 expression was significantly increased in OS cell lines. miR-326 and miR-503-5p were identified to be target miRNAs of hsa_circ_0064636. Among the target genes obtained from the miR-326 and miR-503-5p screens, ubiquitination factor E4A (UBE4A) and voltage dependent anion channel 1 (VDAC1) were respectively identified to significantly affect prognosis; only miR-326 targets UBE4A and only miR-503 targets VDAC1. To conclude, these aforementioned findings suggest that hsa_circ_0064636 may be involved in the development of OS by sponging miR-503-5p and miR-326to inhibit their effects, thereby regulating the expression of VDAC1 and UBE4A.

BACKGROUND: Zishenhuoxue decoction (ZSHX), a Chinese herbal medicine, exhibits myocardial and vascular endothelial protective properties. The intricate regulatory mechanisms underlying myocardial ischemic injury and its association with dysfunctional mitochondrial quality surveillance (MQS) remain elusive.
OBJECTIVE: To study the protective effect of ZSHX on ischemic myocardial injury in mice using a TMBIM6 gene-modified animal model and mitochondrial quality control-related experiments.
METHODS: Using model animals and myocardial infarction surgery-induced ischemic myocardial injury TMBIM6 gene-modified mouse models, the pharmacological activity of ZSHX in inhibiting ischemic myocardial injury and mitochondrial homeostasis disorder in vivo was tested.
METHODS: Our focal point entailed scrutinizing the impact of ZSHX on ischemic myocardial impairment through the prism of TMBIM6. This endeavor was undertaken utilizing mice characterized by heart-specific TMBIM6 knockout (TMBIM6CKO) and their counterparts, the TMBIM6 transgenic (TMBIM6TG) and VDAC1 transgenic (VDAC1TG) mice.
RESULTS: ZSHX demonstrated dose-dependent effectiveness in mitigating ischemic myocardial injury and enhancing mitochondrial integrity. TMBIM6CKO hindered ZSHX\'s cardio-therapeutic and mitochondrial protective effects, while ZSHX\'s benefits persisted in TMBIM6TG mice. TMBIM6CKO also blocked ZSHX\'s regulation of mitochondrial function in HR-treated cardiomyocytes. Hypoxia disrupted the MQS in cardiomyocytes, including calcium overload, excessive fission, mitophagy issues, and disrupted biosynthesis. ZSHX counteracted these effects, thereby normalizing MQS and inhibiting calcium overload and cardiomyocyte necroptosis. Our results also showed that hypoxia-induced TMBIM6 blockade resulted in the over-activation of VDAC1, a major mitochondrial calcium uptake pathway, while ZSHX could increase the expression of TMBIM6 and inhibit VDAC1-mediated calcium overload and MQS abnormalities.
CONCLUSIONS: Our findings suggest that ZSHX regulates mitochondrial calcium homeostasis and MQS abnormalities through a TMBIM6-VDAC1 interaction mechanism, which helps to treat ischemic myocardial injury and provides myocardial protection. This study also offers insights for the clinical translation and application of mitochondrial-targeted drugs in cardiomyocytess.

An increase in α-synuclein (α-syn) levels and mutations in proteins associated with mitochondria contribute to the development of familial Parkinson\'s disease (PD); however, the involvement of α-syn and mitochondria in idiopathic PD remains incompletely understood. The voltage-dependent anion channel I (VDAC1) protein, which serves as a crucial regulator of mitochondrial function and a gatekeeper, plays a pivotal role in governing cellular destiny through the control of ion and respiratory metabolite flux. The ability of resveratrol (RES), which is a potent phytoalexin with antioxidant and anti-inflammatory properties, to regulate VDAC1 in PD is unknown. The objective of this study was to evaluate the role of VDAC1 in the pathological process of PD and to explore the mechanism by which resveratrol protects dopaminergic neurons by regulating VDAC1 to maintain the mitochondrial permeability transition pore (mPTP) and calcium ion balance. The effects of RES on the motor and cognitive abilities of A53T mice were evaluated by using small animal behavioral tests. Various techniques, including immunofluorescence staining, transmission electron microscopy, enzyme-linked immunoadsorption, quantitative polymerase chain reaction (PCR), and Western blotting, among others, were employed to assess the therapeutic impact of RES on neuropathy associated with PD and its potential in regulating mitochondrial VDAC1. The findings showed that RES significantly improved motor and cognitive dysfunction and restored mitochondrial function, thus reducing oxidative stress levels in A53T mice. A significant positive correlation was observed between the protein expression level of VDAC1 and mitochondrial α-syn expression, as well as disease progression, whereas no such correlation was found in VDAC2 and VDAC3. Administration of RES resulted in a significant decrease in the protein expression of VDAC1 and in the protein expression of α-syn both in vivo and in vitro. In addition, we found that RES prevents excessive opening of the mPTP in dopaminergic neurons. This may prevent the abnormal aggregation of α-syn in mitochondria and the release of mitochondrial apoptosis signals. Furthermore, the activation of VDAC1 reversed the resveratrol-induced decrease in the accumulation of α-syn in the mitochondria. These findings highlight the potential of VDAC1 as a therapeutic target for PD and identify the mechanism by which resveratrol alleviates PD-related pathology by modulating mitochondrial VDAC1.

Inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) are high-conductance channels that allow the regulated redistribution of Ca2+ from the endoplasmic reticulum (ER) to the cytosol and, at specialized membrane contact sites (MCSs), to other organelles. Only a subset of IP3Rs release Ca2+ to the cytosol in response to IP3. These \'licensed\' IP3Rs are associated with Kras-induced actin-interacting protein (KRAP, also known as ITPRID2) beneath the plasma membrane. It is unclear whether KRAP regulates IP3Rs at MCSs. We show, using simultaneous measurements of Ca2+ concentration in the cytosol and mitochondrial matrix, that KRAP also licenses IP3Rs to release Ca2+ to mitochondria. Loss of KRAP abolishes cytosolic and mitochondrial Ca2+ signals evoked by stimulation of IP3Rs via endogenous receptors. KRAP is located at ER-mitochondrial membrane contact sites (ERMCSs) populated by IP3R clusters. Using a proximity ligation assay between IP3R and voltage-dependent anion channel 1 (VDAC1), we show that loss of KRAP reduces the number of ERMCSs. We conclude that KRAP regulates Ca2+ transfer from IP3Rs to mitochondria by both licensing IP3R activity and stabilizing ERMCSs.

The strategy for treating bladder cancer (BC) depends on whether there is muscle invasion or not, with the latter mostly treated with intravesical therapy, such as with bacillus Calmette-Guérin (BCG). However, BCG treatment is unsuccessful in 70% of patients, who are then subjected to radical cystectomy. Although immune-checkpoint inhibitors have been approved as a second-line therapy for a subset of BC patients, these have failed to meet primary endpoints in clinical trials. Thus, it is crucial to find a new treatment. The mitochondrial gatekeeper protein, the voltage-dependent anion channel 1 (VDAC1), mediates metabolic crosstalk between the mitochondria and cytosol and is involved in apoptosis. It is overexpressed in many cancer types, as shown here for BC, pointing to its significance in high-energy-demanding cancer cells. The BC cell lines UM-UC3 and HTB-5 express high VDAC1 levels compared to other cancer cell lines. VDAC1 silencing in these cells using siRNA that recognizes both human and mouse VDAC1 (si-m/hVDAC1-B) reduces cell viability, mitochondria membrane potential, and cellular ATP levels. Here, we used two BC mouse models: subcutaneous UM-UC3 cells and chemically induced BC using the carcinogen N-Butyl-N-(4-hydroxybutyl) nitrosamine (BBN). Subcutaneous UM-UC3-derived tumors treated with si-m/hVDAC1 showed inhibited tumor growth and reprogrammed metabolism, as reflected in the reduced expression of metabolism-related proteins, including Glut1, hexokinase, citrate synthase, complex-IV, and ATP synthase, suggesting reduced metabolic activity. Furthermore, si-m/hVDAC1-B reduced the expression levels of cancer-stem-cell-related proteins (cytokeratin-14, ALDH1a), modifying the tumor microenvironment, including decreased angiogenesis, extracellular matrix, tumor-associated macrophages, and inhibited epithelial-mesenchymal transition. The BBN-induced BC mouse model showed a clear carcinoma, with damaged bladder morphology and muscle-invasive tumors. Treatment with si-m/hVDAC1-B encapsulated in PLGA-PEI nanoparticles that were administered intravesically directly to the bladder showed a decreased tumor area and less bladder morphology destruction and muscle invasion. Overall, the obtained results point to the potential of si-m/hVDAC1-B as a possible therapeutic tool for treating bladder cancer.

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) plays an important role in vascular smooth muscle cell (VSMC) phenotypic switching, which is an early pathogenic event in various vascular remodeling diseases (VRDs). However, the underlying mechanism is not fully understood.
METHODS: An IP‒LC‒MS/MS assay was conducted to identify new binding partners of G6PD involved in the regulation of VSMC phenotypic switching under platelet-derived growth factor-BB (PDGF-BB) stimulation. Co-IP, GST pull-down, and immunofluorescence colocalization were employed to clarify the interaction between G6PD and voltage-dependent anion-selective channel protein 1 (VDAC1). The molecular mechanisms involved were elucidated by examining the interaction between VDAC1 and apoptosis-related biomarkers, as well as the oligomerization state of VDAC1.
RESULTS: The G6PD level was significantly elevated and positively correlated with the synthetic characteristics of VSMCs induced by PDGF-BB. We identified VDAC1 as a novel G6PD-interacting molecule essential for apoptosis. Specifically, the G6PD-NTD region was found to predominantly contribute to this interaction. G6PD promotes VSMC survival and accelerates vascular neointimal hyperplasia by inhibiting VSMC apoptosis. Mechanistically, G6PD interacts with VDAC1 upon stimulation with PDGF-BB. By competing with Bax for VDAC1 binding, G6PD reduces VDAC1 oligomerization and counteracts VDAC1-Bax-mediated apoptosis, thereby accelerating neointimal hyperplasia.
CONCLUSIONS: Our study showed that the G6PD-VDAC1-Bax axis is a vital switch in VSMC apoptosis and is essential for VSMC phenotypic switching and neointimal hyperplasia, providing mechanistic insight into early VRDs.

UNASSIGNED: Alzheimer\'s disease (AD) is a widespread neurodegenerative disorder with a significant global impact, affecting approximately 50 million individuals, and projections estimate that up to 152 million people will be affected by 2050. AD is characterized by beta-amyloid plaques and tau tangles in the brain, leading to cognitive decline.
UNASSIGNED: Recent research on AD has made significant strides, including the development of an \"amyloid clock\" biomarker that tracks AD progression through positron emission tomography (PET) scans. Surf4 and other genes have been discovered to play a role in regulating beta-amyloid toxicity, while inhibiting the enzyme hexokinase-2 has shown positive results in preclinical studies. New brain mapping techniques have identified early brain-based causes of cognitive changes in AD, and biomarkers such as neuronal pentraxin protein Nptx2 and astrocytic 7-subunit of the nicotinic acetylcholine receptors (7nAChRs) show potential for early detection. Other approaches, such as replenishing the enzyme Tip60, selectively degrading the modified protein p-p38 with PRZ-18002, and targeting the protein voltage-dependent anion channel-1 (VDAC1), have shown promise in enhancing cognitive function and preventing pathophysiological alterations linked to AD. Baseline blood samples and other biomarkers such as urine formic acid, p-tau 198, microRNAs, and glial fibrillary acidic protein (GFAP) have also been discovered for early detection and intervention of AD. Additionally, recent FDA approvals for medications such as aducanumab and lecanemab provide options for reducing AD symptoms and improving function, while clinical trials for dementia vaccines show promise for the nasal and beta-amyloid 40 vaccines as well as vaccinations targeting tau.
UNASSIGNED: These advancements in AD research, including biomarker discovery and the development of disease-modifying treatments, are crucial steps towards improving the lives of those affected by AD and finding a cure for this debilitating disease.

The present study aimed to explore how resveratrol (Res) confers myocardial protection by attenuating ferroptosis. In vivo and in vitro myocardial ischemia/reperfusion injury (MIRI) models were established, with or without Res pretreatment. The results showed that Res pretreatment effectively attenuated MIRI, as evidenced by increased cell viability, reduced lactate dehydrogenase activity, decreased infarct size, and maintained cardiac function. Moreover, Res pretreatment inhibited MIRI-induced ferroptosis, as shown by improved mitochondrial integrity, increased glutathione level, decreased prostaglandin-endoperoxide synthase 2 level, inhibited iron overload, and abnormal lipid peroxidation. Of note, Res pretreatment decreased or increased voltage-dependent anion channel 1/glutathione peroxidase 4 (VDAC1/GPX4) expression, which was increased or decreased via anoxia/reoxygenation (A/R) treatment, respectively. However, the overexpression of VDAC1 via pAd/VDAC1 and knockdown of GPX4 through Si-GPX4 reversed the protective effect of Res in A/R-induced H9c2 cells, whereas the inhibition of GPX4 with RSL3 abolished the protective effect of Res on mice treated with ischemia/reperfusion.Interestingly, knockdown of VDAC1 by Si-VDAC1 promoted the protective effect of Res on A/R-induced H9c2 cells and the regulation of GPX4. Finally, the direct interaction between VDAC1 and GPX4 was determined using co-immunoprecipitation. In conclusion, Res pretreatment could protect the myocardium against MIRI-induced ferroptosis via the VDAC1/GPX4 signaling pathway.

BACKGROUND: The occurrence of hyperlipidemia is significantly influenced by lipid synthesis, which is regulated by sterol regulatory element binding proteins (SREBPs), thus the development of drugs that inhibit lipid synthesis has become a popular treatment strategy for hyperlipidemia. Alisol B (ALB), a triterpenoid compound extracted from Alisma, has been reported to ameliorate no-nalcoholic steatohepatitis (NASH) and slow obesity. However, the effect of ALB on hyperlipidemia and mechanism are unclear.
OBJECTIVE: To examine the therapeutic impact of ALB on hyperlipidemia whether it inhibits SREBPs to reduce lipid synthesis.
METHODS: HepG2, HL7702 cells, and C57BL/6J mice were used to explore the effect of ALB on hyperlipidemia and the molecular mechanism in vivo and in vitro.
METHODS: Hyperlipidemia models were established using western diet (WD)-fed mice in vivo and oleic acid (OA)-induced hepatocytes in vitro. Western blot, real-time PCR and other biological methods verified that ALB regulated AMPK/mTOR/SREBPs to inhibit lipid synthesis. Cellular thermal shift assay (CETSA), molecular dynamics (MD), and ultrafiltration-LC/MS analysis were used to evaluate the binding of ALB to voltage-dependent anion channel protein-1 (VDAC1).
RESULTS: ALB decreased TC, TG, LDL-c, and increased HDL-c in blood, thereby ameliorating liver damage. Gene set enrichment analysis (GSEA) indicated that ALB inhibited the biosynthesis of cholesterol and fatty acids. Consistently, ALB inhibited the protein expression of n-SREBPs and downstream genes. Mechanistically, the impact of ALB on SREBPs was dependent on the regulation of AMPK/mTOR, thereby impeding the transportation of SREBPs from endoplasmic reticulum (ER) to golgi apparatus (GA). Further investigations indicated that the activation of AMPK by ALB was independent on classical upstream CAMKK2 and LKB1. Instead, ALB resulted in a decrease in ATP levels and an increase in the ratios of ADP/ATP and AMP/ATP. CETSA, MD, and ultrafiltration-LC/MS analysis indicated that ALB interacted with VDAC1. Molecular docking revealed that ALB directly bound to VDAC1 by forming hydrogen bonds at the amino acid sites S196 and H184 in the ATP-binding region. Importantly, the thermal stabilization of ALB on VDAC1 was compromised when VDAC1 was mutated at S196 and H184, suggesting that these amino acids played a crucial role in the interaction.
CONCLUSIONS: Our findings reveal that VDAC1 serves as the target of ALB, leading to the inhibition of lipid synthesis, presents potential target and candidate drugs for hyperlipidemia.