Regulation Of Mitophagy Research Articles

IRF1-mediated upregulation of PARP12 promotes cartilage degradation by inhibiting PINK1/Parkin dependent mitophagy through ISG15 attenuating ubiquitylation and SUMOylation of MFN1/2

Osteoarthritis (OA) is an age-related cartilage-degenerating joint disease. Mitochondrial dysfunction has been reported to promote the development of OA. Poly (ADP-ribose) polymerase family member 12 (PARP12) is a key regulator of mitochondrial function, protein translation, and inflammation. However, the role of PARP12 in OA-based cartilage degradation and the underlying mechanisms are relatively unknown. Here, we first demonstrated that PARP12 inhibits mitophagy and promotes OA progression in human OA cartilage and a monosodium iodoacetate-induced rat OA model. Using mass spectrometry and co-immunoprecipitation assay, PARP12 was shown to interact with ISG15, upregulate mitofusin 1 and 2 (MFN1/2) ISGylation, which downregulated MFN1/2 ubiquitination and SUMOylation, thereby inhibiting PINK1/Parkin-dependent chondrocyte mitophagy and promoting cartilage degradation. Moreover, inflammatory cytokine-induced interferon regulatory factor 1 (IRF1) activation was required for the upregulation of PARP12 expression, and it directly bound to the PARP12 promoter to activate transcription. XAV-939 inhibited PARP12 expression and suppressed OA pathogenesis in vitro and in vivo. Clinically, PARP12 can be used to predict the severity of OA; thus, it represents a new target for the study of mitophagy and OA progression. In brief, the IRF1-mediated upregulation of PARP12 promoted cartilage degradation by inhibiting PINK1/Parkin-dependent mitophagy via ISG15-based attenuation of MFN1/2 ubiquitylation and SUMOylation. Our data provide new insights into the molecular mechanisms underlying PARP12-based regulation of mitophagy and can facilitate the development of therapeutic strategies for the treatment of OA.

RABIF promotes hepatocellular carcinoma progression through regulation of mitophagy and glycolysis

The RAB interacting factor (RABIF) is a putative guanine nucleotide exchange factor that also functions as a RAB-stabilizing holdase chaperone. It has been implicated in pathogenesis of several cancers. However, the functional role and molecular mechanism of RABIF in hepatocellular carcinoma (HCC) are not entirely known. Here, we demonstrate an upregulation of RABIF in patients with HCC, correlating with a poor prognosis. RABIF inhibition results in decreased HCC cell growth both in vitro and in vivo. Our study reveals that depleting RABIF attenuates the STOML2-PARL-PGAM5 axis-mediated mitophagy. Consequently, this reduction in mitophagy results in diminished mitochondrial reactive oxygen species (mitoROS) production, thereby alleviating the HIF1α-mediated downregulation of glycolytic genes HK1, HKDC1, and LDHB. Additionally, we illustrate that RABIF regulates glucose uptake by controlling RAB10 expression. Importantly, the knockout of RABIF or blockade of mitophagy sensitizes HCC cells to sorafenib. This study uncovers a previously unrecognized role of RABIF crucial for HCC growth and identifies it as a potential therapeutic target.

UCF101 Rescues against Diabetes-Evoked Cardiac Remodeling and Contractile Anomalies through AMP-Activated Protein Kinase-Mediated Induction of Mitophagy

Introduction: Diabetes mellitus is known to provoke devastating anomalies in myocardial structure and function, while effective therapeutic regimen is still lacking. The selective protease inhibitor UCF101 (5-[5-(2-nitrophenyl) furfuryl iodine]-1,3-diphenyl-2-thiobarbituric acid) has been shown to fend off ischemic heart injury, although its impact on diabetic cardiomyopathy remains elusive. Methods: Our present work was conducted to examine the effect of UCF101 on experimental diabetes-evoked cardiac geometric and functional abnormalities as well as mechanisms involved. Adult mice were made diabetic using streptozotocin (STZ, 50 mg/kg, i.p., for 5 days) while receiving UCF101 (7.15 mg/kg, i.p.). Results: STZ evoked cardiac hypertrophy, interstitial fibrosis, mitochondrial ultrastructural damage, oxidative stress, dampened autophagy (LC3B, Beclin 1, elevated p62), mitophagy (FUNDC1 and Parkin with upregulated TOM20), increased left ventricular end systolic diameter, reduced fractional shortening, ejection fraction, cardiomyocyte shortening capacity, velocities of shortening/re-lengthening, and rise in intracellular Ca2+ in conjunction with elongated diastole and intracellular Ca2+ removal, the responses were overtly reconciled by UCF101 with little effects from UCF101 itself. Levels of cell injury markers Omi/HtrA2, TNFα, and stress signaling (JNK, ERK, p38) were overtly enhanced along with compromised phosphorylation of cellular fuel AMP-activated protein kinase (AMPK) (Thr172) and cell survival molecule GSK3β, as well as downregulated SERCA2a and elevated phospholamban, the effect was reversed by UCF101 (except for SERCA2a). AMPK knockout, pharmacological inhibition, the mitophagy inhibitor liensinine, and parkin knockout nullified UCF101-offered cardioprotection in diabetes. UCF101 reversed STZ-induced upregulation in the AMPK degrading enzymes PP2A and PP2C. Conclusion: These findings suggest that UCF101 rescues diabetes-mediated alterations in cardiac structure and function, likely through AMPK-mediated regulation of mitophagy.

Mitophagy Defects Exacerbate Inflammation and Aberrant Proliferation in Lymphocytic Thyroiditis.

Background: Mitochondrial dysfunction in the thyroid due to defective mitophagy has been observed in lymphocytic thyroiditis (LT). However, the effect of impaired mitophagy on the pathogenesis of LT is not well understood. The aim of this study is to investigate the role of mitophagy dysregulation in the thyroid gland. Methods: We analyzed RNA sequencing data of human thyroid glands with/without LT from Genotype-Tissue Expression (GTEx; n = 653) and performed RNA sequencing in thyroid glands of phosphatase and tensin homolog-induced putative protein kinase 1 (Pink1) knock-out and wild-type mice. We evaluated the phenotypic and histopathologic characteristics of the human (n = 16) and mouse thyroids. Additionally, we assessed cell proliferation, reactive oxygen species (ROS) production, and cytokine secretion of human thyroid epithelial cells (HTori-3) treated with PINK1 siRNA or a mitophagy inhibitor. Results: We found that expression of PINK1, a key regulator of mitophagy, was compromised in human thyroids with LT. Thyroid glands of Pink1-deficient mice exhibited increased inflammatory responses and nodular hyperplasia. Furthermore, mitophagy defects led to the production of pro-inflammatory cytokines and ROS in thyroid cells, resulting in immune cell recruitment. Notably, these mitophagy defects upregulated both the RNA expression and protein secretion of amphiregulin (AREG), an epidermal growth factor receptor (EGFR) ligand, in thyroid cells, while decreasing the protein expression of cAMP response element-binding protein (CREB), a transcription factor that suppresses AREG transcription. Finally, we demonstrated that aberrant cell proliferation in thyroid cells, driven by mitophagy defects, was mitigated after treatment with cetuximab, an EGFR inhibitor. Conclusions: In this study, we observed that mitophagy defects in the thyroid not only intensify inflammation through the accumulation of ROS, cytokine production, and immune cell recruitment but also contribute to hyperplasia via the EGFR pathway, facilitated by increased secretion of AREG from thyroid cells.

Copper aggravated synaptic damage after traumatic brain injury by downregulating BNIP3-mediated mitophagy

ABSTRACT Synaptic damage is a crucial pathological process in traumatic brain injury. However, the mechanisms driving this process remain poorly understood. In this report, we demonstrate that the accumulation of damaged mitochondria, resulting from impaired mitphagy, plays a significant role in causing synaptic damage. Moreover, copper induced downregulation of BNIP3 is a key player in regulating mitophagy. DMSA alleviates synaptic damage and mitochondrial dysfunction by promoting urinary excretion of copper. Mechanistically, we find that copper downregulate BNIP3 by increasing the nuclear translocation of NFKB, which is triggered by TRIM25-mediated ubiquitination-dependent degradation of NFKBIA. Our study underscores the importance of copper accumulation in the regulation of BNIP3-mediated mitophagy and suggests that therapeutic targeting of the copper-TRIM25-NFKB-BNIP3 axis holds promise to attenuate synaptic damage after traumatic brain injury.

Coordinating BNIP3/NIX-mediated mitophagy in space and time.

Mitochondria maintain organellar homeostasis through multiple quality control pathways, including the clearance of defective or unwanted mitochondria by selective autophagy. This removal of mitochondria, mitophagy, is controlled in large part by the outer mitochondrial membrane mitophagy receptors BNIP3 and NIX. While it has long been appreciated that BNIP3 and NIX mediate mitophagy by controlling the recruitment of autophagic machinery to the mitochondrial surface, the requirement for the carefully controlled spatiotemporal regulation of receptor-mediated mitophagy has only recently come to light. Several new factors that regulate the BNIP3/NIX-mediated mitophagy pathway have emerged, and various loss-of-function cell and animal models have revealed the dire consequences of their dysregulation. In this mini-review, we discuss new insights into the mechanisms and roles of the regulation of BNIP3 and NIX and highlight questions that have emerged from the identification of these new regulators.

Shaoyao-Gancao Decoction, a famous Chinese medicine formula, protects against APAP-induced liver injury by promoting autophagy/mitophagy

BackgroundAcetaminophen (APAP)-induced hepatotoxicity is a major cause of acute liver failure (ALF), during which autophagy is triggered as a cellular defense mechanism. Shaoyao-Gancao Decoction (SGD), a traditional prescription in Chinese Medicine, is renowned for its therapeutic effects on liver diseases. However, the efficacy and mechanisms of SGD in treating APAP-induced liver injury remain underexplored. PurposeThis study aims to provide robust evidence regarding the protective effects of SGD against APAP overdose in vitro and in vivo, as well as to elucidate its hepatoprotective mechanisms and active components. Study designThe hepatoprotective mechanisms and active components of SGD were investigated through a combination of in vivo and in vitro experiments. MethodsThe protective effects of SGD on APAP-induced liver injury were assessed using a murine model and primary hepatocytes. RNA sequencing and subsequent experimental validations were conducted to uncover the underlying mechanisms of SGD's hepatoprotective actions. Comprehensive chemical profiling of SGD was performed using UHPLC-Q-Exactive Orbitrap HRMS to identify potential active ingredients. Immunohistochemistry, immunofluorescence, quantitative real-time PCR (qPCR), western blotting, enzyme-linked immunosorbent assay (ELISA), and flow cytometry were utilized to investigate the specific cellular changes in liver tissues and hepatocytes influenced by SGD. ResultsSGD was observed to mitigate APAP-induced mitochondrial damage, inflammation, and necrosis by promoting mitochondrial autophagy. The inhibition of autophagy negated the hepatoprotective effects of SGD. Additionally, a detailed characterization of SGD's chemical composition revealed that Licoisoflavone B, Liquiritin, Liquiritin apioside, Licorice saponin G2 and Paeoniflorin Sulfit were potentially critical compounds in the regulation of autophagy and mitophagy. ConclusionOur findings demonstrate that SGD promotes autophagy/mitophagy, which effectively mitigates APAP-induced hepatotoxicity, suggesting SGD's potential as a promising therapeutic agent for APAP-induced liver injury.

Dioscin improves fatty liver hemorrhagic syndrome by promoting ERα-AMPK mediated mitophagy in laying hens

BackgroundMitochondria play a crucial role in upholding metabolic homeostasis. Mitochondrial damage closely associated with the pathogenesis of fatty liver hemorrhagic syndrome (FLHS), while mitophagy being among the most effective methods for eliminating the damaged mitochondria. Dioscin, a natural extract, can activate autophagy; however, its effects on FLHS regarding mitophagy regulation remain unelucidated. PurposeWe explored the impact of dioscin on FLHS induced by a high-energy and low-protein (HELP) diet in laying hens, mainly focused the protective effects of dioscin on mitochondrial injury. MethodTo investigate the impact of dioscin on fatty liver syndrome in laying hens, we first induced the condition by feeding them a high-energy and low-protein diet. Then, we assessed lipid metabolism-related markers using oil red staining and a commercial detection kit. In addition, the role of dioscin on fatty liver syndrome in laying hens was confirmed by assessing the activation of hepatocyte fat deposition and hepatocyte apoptosis; and the mechanism of dioscin in FLHS was investigated through LMH cell experiment in vitro. Furthermore, CETSA and molecular docking were conducted for additional confirmation. ResultThe results showed that dioscin alleviated mitochondrial damage, relieved the excessive deposition of hepatic lipid droplets and oxidative stress induced by HELP diet in laying hens. Furthermore, dioscin regulated the mitophagy by activating the estrogen receptor α (ERα)/adenosine 5′-monophosphate-activated protein kinase (AMPK) signaling pathway, thus mitigating mitochondria injury and apoptosis in hepatocytes. In addition, we found that dioscin promoted the translocation of nuclear transcription factor into nucleus by activating ERα-AMPK signaling, facilitating autophagic flux in the liver of laying hens and LMH cells. Furthermore, cells pretreated with the lysosomal acidification inhibitor bafilomycin A1 blocked the inhibitory effect of dioscin on the apoptosis induced by palmitic acid (PA)-stimulation in LMH cells, suggesting that dioscin reduces PA-induced apoptosis by activating mitophagy. Moreover, dioscin-induced lysosomal acidification and mitochondrial biogenesis were reversed in PA-induced LMH cells pretreated with ERα-specific inhibitor methylpiperidino pyrazole. ConclusionThis study firstly demonstrated that dioscin alleviates fatty liver syndrome induced by HELP diet in laying hens. The findings from this study illustrated that dioscin plays a significant role in reducing mitochondrial damage and apoptosis, and these beneficial effects mainly achieve through promotion of ERα-AMPK signaling, which mediates autophagy within the liver of laying hens fed a HELP-diets. These findings provide a theoretical basis for considering dioscin as a possible treatment option for mitigating FLHS in egg-laying hens.

Mitophagy Regulates Kidney Diseases

Background: Mitophagy is a crucial process involved in maintaining cellular homeostasis by selectively eliminating damaged or surplus mitochondria. As the kidney is an organ with a high dynamic metabolic rate and abundant mitochondria, it is particularly crucial to control mitochondrial quality through mitophagy. Dysregulation of mitophagy has been associated with various renal diseases, including acute and chronic kidney diseases, and therefore a better understanding of the links between mitophagy and these diseases may present new opportunities for therapeutic interventions. Summary: Mitophagy plays a pivotal role in the development of kidney diseases. Upregulation and downregulation of mitophagy have been observed in various kidney diseases, such as renal ischemia-reperfusion injury, contrast-induced acute kidney injury, diabetic nephropathy, kidney fibrosis, and several inherited renal diseases. A growing body of research has suggested that PINK1 and Parkin, the main mitophagy regulatory proteins, represent promising potential therapeutic targets for kidney diseases. In this review, we summarize the latest insights into how the progression of renal diseases can be mitigated through the regulation of mitophagy, while highlighting their performance in clinical trials. Key Message: This review comprehensively outlines the mechanisms of mitophagy and its role in numerous kidney diseases. While early research holds promise, most mitophagy-centered therapeutic approaches have yet to reach the clinical application stage.

The tumor suppressor Parkin exerts anticancer effects through regulating mitochondrial GAPDH activity.

Cancer cells preferentially utilize glycolysis for energy production, and GAPDH is a critical enzyme in glycolysis. Parkin is a tumor suppressor and a key protein involved in mitophagy regulation. However, the tumor suppression mechanism of Parkin has still not been elucidated. In this study, we identified mitochondrial GAPDH as a new substrate of the E3 ubiquitin ligase Parkin, which mediated GAPDH ubiquitination in human cervical cancer. The translocation of GAPDH into mitochondria was driven by the PINK1 kinase, and either PINK1 or GAPDH mutation prevented the accumulation of GAPDH in mitochondria. Parkin caused the ubiquitination of GAPDH at multiple sites (K186, K215, and K219) located within the enzyme-catalyzed binding domain of the GAPDH protein. GAPDH ubiquitination was required for mitophagy, and stimulation of mitophagy suppressed cervical cancer cell growth, indicating that mitophagy serves as a type of cell death. Mechanistically, PHB2 served as a key mediator in GAPDH ubiquitination-induced mitophagy through stabilizing PINK1 protein and GAPDH mutation resulted in the reduced distribution of PHB2 in mitophagic vacuole. In addition, ubiquitination of GAPDH decreased its phosphorylation level and enzyme activity and inhibited the glycolytic pathway in cervical cancer cells. The results of in vivo experiments also showed that the GAPDH mutation increased glycolysis in cervical cancer cells and accelerated tumorigenesis. Thus, we concluded that Parkin may exert its anticancer function by ubiquitinating GAPDH in mitochondria. Taken together, our study further clarified the molecular mechanism of tumor suppression by Parkin through the regulation of energy metabolism, which provides an experimental basis for the development of new drugs for the treatment of human cervical cancer.

Unraveling the mechanism of quercetin alleviating BHPF-induced apoptosis in epithelioma papulosum cyprini cells: SIRT3-mediated mitophagy

Fluorene-9-bisphenol (BHPF), as an alternative to bisphenol A, is now increasingly used in plastic products. The accumulation of BHPF in the water environment has posed potential safety risks to aquatic organisms. Unfortunately, the toxicity of BHPF on the physiological metabolism of aquatic animals remains unclear, especially on the molecular mechanisms of BHPF kidney toxicity and antagonizing BHPF toxicity. Quercetin (QCT), a naturally occurring flavonoid, has been reported to mitigate the toxic effects on aquatic organisms induced by a variety of environmental contaminants. It is unclear whether QCT can be a candidate for mitigating BHPF toxicity. In this study, we investigated the protective effect of QCT on BHPF-induced apoptosis and elucidated the possible mechanism of the protective effect mediated by QCT. We treated epithelioma papulosum cyprini cells (EPCs) with 20 μM of BHPF and/or 20 μM of QCT, and the results showed that BHPF significantly increased the release of reactive oxygen species (ROS) from EPCs, decreased the expression of SIRT3, and initiated endogenous apoptosis. Molecular docking provides evidence for the interaction of QCT and SIRT3. Our intervention with Honokiol (HKL) showed that QCT or HKL treatment significantly attenuated BHPF-induced mitochondrial dysfunction and mitochondrial apoptosis (mtApoptosis) in EPCs, and activated mitophagy, restoring autophagy flux. To further investigate the specific mechanism of the protective effect of QCT, we intervened with Cyclosporin A (CsA), and our results suggest that QCT activation of SIRT3-promoted regulation of mitophagy may be a therapeutic strategy to attenuate the toxic effects of BHPF on EPCs. In conclusion, our findings suggest that BHPF induces oxidative damage and mtApoptosis in EPCs and that QCT activates mitophagy and improves autophagic flux through activation of SIRT3, thereby alleviating apoptosis mediated by mitochondrial dysfunction in EPCs. Our study provides a theoretical basis for reassessing the safety of BHPF for aquatic organisms and reveals a novel detoxification mechanism against the toxic effects of BHPF.

Cardioprotective effect of Vitamin D on cardiac hypertrophy through improvement of mitophagy and apoptosis in an experimental rat model of levothyroxine -induced hyperthyroidism.

Mitochondria are known to be involved in mediating the calorigenic effects of thyroid hormones. With an abundance of these hormones, alterations in energy metabolism and cellular respiration take place, leading to the development of cardiac hypertrophy. Vitamin D has recently gained attention due to its involvement in the regulation of mitochondrial function, demonstrating promising potential in preserving the integrity and functionality of the mitochondrial network. The present study aimed to investigate the therapeutic potential of Vitamin D on cardiac hypertrophy induced by hyperthyroidism, with a focus on the contributions of mitophagy and apoptosis as possible underlying molecular mechanisms. The rats were divided into three groups: control; hyperthyroid; hyperthyroid + Vitamin D. Hyperthyroidism was induced by Levothyroxine administration for four weeks. Serum thyroid hormones levels, myocardial damage markers, cardiac hypertrophy indices, and histological examination were assessed. The assessment of Malondialdehyde (MDA) levels and the expression of the related genes were conducted using heart tissue samples. Vitamin D pretreatment exhibited a significant improvement in the hyperthyroidism-induced decline in markers indicative of myocardial damage, oxidative stress, and indices of cardiac hypertrophy. Vitamin D pretreatment also improved the downregulation observed in myocardial expression levels of genes involved in the regulation of mitophagy and apoptosis, including PTEN putative kinase 1 (PINK1), Mitofusin-2 (MFN2), Dynamin-related Protein 1 (DRP1), and B cell lymphoma-2 (Bcl-2), induced by hyperthyroidism. These results suggest that supplementation with Vitamin D could be advantageous in preventing the progression of cardiac hypertrophy and myocardial damage.

Melatonin regulates mitochondrial dynamics and mitophagy: Cardiovascular protection.

Despite extensive progress in the knowledge and understanding of cardiovascular diseases and significant advances in pharmacological treatments and procedural interventions, cardiovascular diseases (CVD) remain the leading cause of death globally. Mitochondrial dynamics refers to the repetitive cycle of fission and fusion of the mitochondrial network. Fission and fusion balance regulate mitochondrial shape and influence physiology, quality and homeostasis. Mitophagy is a process that eliminates aberrant mitochondria. Melatonin (Mel) is a pineal-synthesized hormone with a range of pharmacological properties. Numerous nonclinical trials have demonstrated that Mel provides cardioprotection against ischemia/reperfusion, cardiomyopathies, atherosclerosis and cardiotoxicity. Recently, interest has grown in how mitochondrial dynamics contribute to melatonin cardioprotective effects. This review assesses the literature on the protective effects of Mel against CVD via the regulation of mitochondrial dynamics and mitophagy in both in-vivo and in-vitro studies. The signalling pathways underlying its cardioprotective effects were reviewed. Mel modulated mitochondrial dynamics and mitophagy proteins by upregulation of mitofusin, inhibition of DRP1 and regulation of mitophagy-related proteins. The evidence supports a significant role of Mel in mitochondrial dynamics and mitophagy quality control in CVD.

Enhancing terminal erythroid differentiation in human embryonic stem cells through TRIB3 overexpression

Tribbles pseudokinase 3 (TRIB3) expression significantly increases during terminal erythropoiesis in vivo. However, we found that TRIB3 expression remained relatively low during human embryonic stem cell (hESC) erythropoiesis, particularly in the late stage, where it is typically active. TRIB3 was expressed in megakaryocyte-erythrocyte progenitor cells and its low expression was necessary for megakaryocyte differentiation. Thus, we proposed that the high expression during late stage of erythropoiesis could be the clue for promotion of maturation of hESC-derived erythroid cells. To our knowledge, the role of TRIB3 in the late stage of erythropoiesis remains ambiguous. To address this, we generated inducible TRIB3 overexpression hESCs, named TRIB3tet-on OE H9, based on a Tet-On system. Then, we analyzed hemoglobin expression, condensed chromosomes, organelle clearance, and enucleation with or without doxycycline treatment. TRIB3tet-on OE H9 cells generated erythrocytes with a high proportion of orthochromatic erythroblast in flow cytometry, enhanced hemoglobin and related protein expression in Western blot, decreased nuclear area size, promoted enucleation rate, decreased lysosome and mitochondria number, more colocalization of LC3 with LAMP1 (lysosome marker) and TOM20 (mitochondria marker) and up-regulated mitophagy-related protein expression after treatment with 2 μg/mL doxycycline. Our results showed that TRIB3 overexpression during terminal erythropoiesis may promote the maturation of erythroid cells. Therefore, our study delineates the role of TRIB3 in terminal erythropoiesis, and reveals TRIB3 as a key regulator of UPS and downstream mitophagy by ensuring appropriate mitochondrial clearance during the compaction of chromatin.

Identification of novel mitophagy-related biomarkers for Kawasaki disease by integrated bioinformatics and machine-learning algorithms.

Kawasaki disease (KD) is a systemic vasculitis primarily affecting the coronary arteries in children. Despite growing attention to its symptoms and pathogenesis, the exact mechanisms of KD remain unclear. Mitophagy plays a critical role in inflammation regulation, however, its significance in KD has only been minimally explored. This study sought to identify crucial mitophagy-related biomarkers and their mechanisms in KD, focusing on their association with immune cells in peripheral blood. This research used four datasets from the Gene Expression Omnibus (GEO) database that were categorized as the merged and validation datasets. Screening for differentially expressed mitophagy-related genes (DE-MRGs) was conducted, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. A weighted gene co-expression network analysis (WGCNA) identified the hub module, while machine-learning algorithms [random forest-recursive feature elimination (RF-RFE) and support vector machine-recursive feature elimination (SVM-RFE)] pinpointed the hub genes. Receiver operating characteristic (ROC) curves were generated for these genes. Additionally, the CIBERSORT algorithm was used to assess the infiltration of 22 immune cell types to explore their correlations with hub genes. Interactions between transcription factors (TFs), genes, and Gene-microRNAs (miRNAs) of hub genes were mapped using the NetworkAnalyst platform. The expression difference of the hub genes was validated using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Initially, 306 DE-MRGs were identified between the KD patients and healthy controls. The enrichment analysis linked these MRGs to autophagy, mitochondrial function, and inflammation. The WGCNA revealed a hub module of 47 KD-associated DE-MRGs. The machine-learning algorithms identified cytoskeleton-associated protein 4 (CKAP4) and serine-arginine protein kinase 1 (SRPK1) as critical hub genes. In the merged dataset, the area under the curve (AUC) values for CKAP4 and SRPK1 were 0.933 [95% confidence interval (CI): 0.901 to 0.964] and 0.936 (95% CI: 0.906 to 0.966), respectively, indicating high diagnostic potential. The validation dataset results corroborated these findings with AUC values of 0.872 (95% CI: 0.741 to 1.000) for CKAP4 and 0.878 (95% CI: 0.750 to 1.000) for SRPK1. The CIBERSORT analysis connected CKAP4 and SRPK1 with specific immune cells, including activated cluster of differentiation 4 (CD4) memory T cells. TFs such as MAZ, SAP30, PHF8, KDM5B, miRNAs like hsa-mir-7-5p play essential roles in regulating these hub genes. The qRT-PCR results confirmed the differential expression of these genes between the KD patients and healthy controls. CKAP4 and SRPK1 emerged as promising diagnostic biomarkers for KD. These genes potentially influence the progression of KD through mitophagy regulation.

Ablation of Akt2 rescues chronic caloric restriction-provoked myocardial remodeling and dysfunction through a CDK1-mediated regulation of mitophagy

Chronic caloric restriction triggers unfavorable alterations in cardiac function albeit responsible scenarios remain unclear. This work evaluated the possible involvement of Akt2 in caloric restriction-evoked cardiac geometric and functional changes and responsible processes focusing on autophagy and mitophagy. Akt2 knockout and WT mice were subjected to caloric restriction for 30 weeks prior to assessment of myocardial homeostasis. Caloric restriction compromised echocardiographic parameters (decreased LV wall thickness, LVEDD, stroke volume, cardiac output, ejection fraction, fractional shortening, and LV mass), cardiomyocyte contractile and intracellular Ca2+ capacity, myocardial atrophy, interstitial fibrosis and mitochondrial injury associated with elevated blood glucocorticoids, autophagy (LC3B, p62, Atg7, Beclin-1), and mitophagy (Pink1, Parkin, TOM20), dampened cardiac ATP levels, mitochondrial protein PGC1α and UCP2, anti-apoptotic protein Bcl2, intracellular Ca2+ governing components Na+-Ca2+ exchanger, phosphorylation of SERCA2a, mTOR (Ser2481) and ULK1 (Ser757), and upregulated Bax, phospholamban, phosphorylation of Akt2, AMPK, and ULK1 (Ser555), the responses except autophagy markers (Beclin-1, Atg7), phosphorylation of AMPK, mTOR and ULK1 were negated by Akt2 ablation. Levels of CDK1 and DRP1 phosphorylation were overtly upregulated with caloric restriction, the response was reversed by Akt2 knockout. Caloric restriction-evoked changes in cardiac remodeling and cardiomyocyte function were alleviated by glucocorticoid receptor antagonism, Parkin ablation and Mdivi-1. In vitro experiment indicated that serum deprivation or glucocorticoids evoked GFP-LC3B accumulation and cardiomyocyte dysfunction, which was negated by inhibition of Akt2, CDK1 or DRP1, whereas mitophagy induction reversed Akt2 ablation-evoked cardioprotection. These observations favor a protective role of Akt2 ablation in sustained caloric restriction-evoked cardiac pathological changes via correction of glucocorticoid-induced mitophagy defect in a CDK1-DRP1-dependent manner.

MANF facilitates breast cancer cell survival under glucose-starvation conditions via prkn-mediated mitophagy regulation

ABSTRACT During tumor expansion, breast cancer (BC) cells often experience reactive oxygen species accumulation and mitochondrial damage because of glucose shortage. However, the mechanism by which BC cells deal with the glucose-shortage-induced oxidative stress remains unclear. Here, we showed that MANF (mesencephalic astrocyte derived neurotrophic factor)-mediated mitophagy facilitates BC cell survival under glucose-starvation conditions. MANF-mediated mitophagy also promotes fatty acid oxidation in glucose-starved BC cells. Moreover, during glucose starvation, SENP1-mediated de-SUMOylation of MANF increases cytoplasmic MANF expression through the inhibition of MANF’s nuclear translocation and hence renders mitochondrial distribution of MANF. MANF mediates mitophagy by binding to PRKN (parkin RBR E3 ubiquitin protein ligase), a key mitophagy regulator, in the mitochondria. Under conditions of glucose starvation, protein oxidation inhibits PRKN activity; nevertheless, the CXXC motif of MANF alleviates protein oxidation in RING II-domain of PRKN and restores its E3 ligase activity. Furthermore, MANF-PRKN interactions are essential for BC tumor growth and metastasis. High MANF expression predicts poor outcomes in patients with BC. Our results highlight the prosurvival role of MANF-mediated mitophagy in BC cells during glucose starvation, suggesting MANF as a potential therapeutic target.

Linagliptin mitigates cisplatin-induced kidney impairment via mitophagy regulation in rats, with emphasis on SIRT-3/PGC-1α, PINK-1 and Parkin-2

Cisplatin (CDDP) often leads to kidney impairment, limiting its effectiveness in cancer treatment. The lack of mitophagy in proximal tubules exacerbates this issue. Hence, targeting SIRT-3 and PGC1-α shows promise in mitigating CDDP-induced kidney damage. The potential renoprotective effects of linagliptin, however, remain poorly understood. This study represents the first exploration of linagliptin's impact on CDDP-induced kidney impairment in rats, emphasizing its potential role in mitophagic pathways. The experiment involved four rat groups: Group (I) received saline only, Group (II) received a single intraperitoneal injection of CDDP at 6 mg/kg. Groups (III) and (IV) received linagliptin at 6 and 10 mg/kg p.o., respectively, seven days before CDDP administration, continuing for an additional four days. Various parameters, including renal function tests, oxidative stress, TNF-α, IL-1β, IL-6, PGC-1α, FOXO-3a, p-ERK1, and the gene expression of SIRT-3 and P62 in renal tissue, were assessed. Linagliptin improved renal function, increased antioxidant enzyme activity, and decreased IL-1β, TNF-α, and IL-6 expression. Additionally, linagliptin significantly upregulated PGC-1α and PINK-1/Parkin-2 expression while downregulating P62 expression. Moreover, linagliptin activated FOXO-3a and SIRT-3, suggesting a potential enhancement of mitophagy. Linagliptin demonstrated a positive impact on various factors related to kidney health in the context of CDDP-induced impairment. These findings suggest a potential role for linagliptin in improving cancer treatment outcomes. Clinical trials are warranted to further investigate and validate its efficacy in a clinical setting.

Abstract Tu090: P21-activated kinase 1 safeguards cardiac function from impairment during fasting

Excessive calorie intake poses a heightened risk for cardiovascular disease, while caloric restriction and intermittent fasting have demonstrated benefits in enhancing cardiovascular health and improving cardiometabolic risk profiles. Despite this knowledge, the precise mechanisms underlying the cardioprotective effects of these dietary regimens remain inadequately understood. P21-activated kinase 1 (PAK1) is a serine/threonine protein kinase known to play a crucial role in promoting cardiomyocyte survival in response to various stressors. In this study, we aimed to elucidate the role of PAK1 in maintaining cardiac function during fasting, utilizing mice with modified PAK1 activity in the heart. Under normal feeding conditions, heart function exhibited no discernible differences among wild type (WT), transgenic (TG), and knockout (KO) mice, as evidenced by consistent fractional shortening (FS), ejection fraction (EF), E/A ratio, and isovolumic relaxation time (IVRT). After a one-day fast, systolic contractile function remained robust across all groups of mice. However, the E/A ratio decreased in KO mice, and IVRT was prolonged in KO mice, while shortened in TG mice, highlighting the crucial role of PAK1 in maintaining diastolic cardiac function. Upon a two-day fast, systolic function was compromised in both WT and KO mice, whereas TG mice exhibited preserved function, as evidenced by FS and EF measurements, emphasizing PAK1's equal significance in sustaining systolic cardiac function. Furthermore, fasting accelerated mitophagy, a process eliminating dysfunctional mitochondria through lysosomal degradation, as observed by mitochondria-associated LC3-II and a dual fluorescent mitophagy reporter. Additionally, we established PAK1 as a positive regulator of mitophagy, suggesting its potential protective role in the heart during fasting via mitophagy modulation. In summary, PAK1 emerges as a critical mediator of cardiac function during fasting, presenting promising avenues for therapeutic interventions aimed at improving cardiovascular health.

Inhibition of mitophagy via the EIF2S1-ATF4-PRKN pathway contributes to viral encephalitis

IntroductionMitophagy, a selective form of autophagy responsible for maintaining mitochondrial homeostasis, regulates the antiviral immune response and acts as viral replication platforms to facilitate infection with various viruses. However, its precise role in herpes simplex virus 1 (HSV-1) infection and herpes simplex encephalitis (HSE) remains largely unknown. ObjectivesWe aimed to investigate the regulation of mitophagy by HSV-1 neurotropic infection and its role in viral encephalitis, and to identify small compounds that regulate mitophagy to affect HSV-1 infection. MethodsThe antiviral effects of compounds were investigated by Western blot, RT-PCR and plaque assay. The changes of Parkin (PRKN)-mediated mitophagy and Nuclear Factor kappa B (NFKB)-mediated neuroinflammation were examined by TEM, RT-qPCR, Western blot and ELISA. The therapeutic effect of taurine or PRKN-overexpression was confirmed in the HSE mouse model by evaluating survival rate, eye damage, neurodegenerative symptoms, immunohistochemistry analysis and histopathology. ResultsHSV-1 infection caused the accumulation of damaged mitochondria in neuronal cells and in the brain tissue of HSE mice. Early HSV-1 infection led to mitophagy activation, followed by inhibition in the later viral infection. The HSV-1 proteins ICP34.5 or US11 deregulated the EIF2S1-ATF4 axis to suppress PRKN/Parkin mRNA expression, thereby impeding PRKN-dependent mitophagy. Consequently, inhibition of mitophagy by specific inhibitor midiv-1 promoted HSV-1 infection, whereas mitophagy activation by PRKN overexpression or agonists (CCCP and rotenone) attenuated HSV-1 infection and reduced the NF-κB-mediated neuroinflammation. Moreover, PRKN-overexpressing mice showed enhanced resistance to HSV-1 infection and ameliorated HSE pathogenesis. Furthermore, taurine, a differentially regulated gut microbial metabolite upon HSV-1 infection, acted as a mitophagy activator that transcriptionally promotes PRKN expression to stimulate mitophagy and to limit HSV-1 infection both in vitro and in vivo. ConclusionThese results reveal the protective function of mitophagy in HSE pathogenesis and highlight mitophagy activation as a potential antiviral therapeutic strategy for HSV-1-related diseases.