Carbon monoxide alleviates senescence in diabetic nephropathy by improving autophagy

ObjectivesSenescence, characterized by permanent cycle arrest, plays an important role in diabetic nephropathy (DN). However, the mechanism of renal senescence is still unclear, and the treatment targeting it remains to be further explored.Materials and MethodsThe DN mice were induced by HFD and STZ, and 3 types of renal cells were treated with high glucose (HG) to establish in vitro model. Senescence‐related and autophagy‐related markers were detected by qRT‐PCR and Western blot. Further, autophagy inhibitors and co‐immunoprecipitation were used to clarify the mechanism of CO. Additionally, the specific relationship between autophagy and senescence was explored by immunofluorescence triple co‐localization and ELISA.ResultsWe unravelled that senescence occurred in vivo and in vitro, which could be reversed by CO. Mechanistically, we demonstrated that CO inhibited the dysfunction of autophagy in DN mice partly through dissociating Beclin‐1‐Bcl‐2 complex. Further results showed that autophagy inhibitors blocked the improvement of CO on senescence. In addition, the data revealed that autophagy regulated the degradation of senescence‐related secretory phenotype (SASP) including Il‐1β, Il‐6, Tgf‐β and Vegf.ConclusionsThese results suggested that CO protects DN mice from renal senescence and function loss via improving autophagy partly mediated by dissociating Beclin‐1‐Bcl‐2 complex, which is possibly ascribed to the degradation of SASP. These findings bring new ideas for the prevention and treatment of DN and the regulation of senescence.     

p73 regulates autophagy and hepatocellular lipid metabolism through a transcriptional activation of the ATG5 gene

p73, a member of the p53 tumor suppressor family, is involved in neurogenesis, sensory pathways, immunity, inflammation, and tumorigenesis. How p73 is able to participate in such a broad spectrum of different biological processes is still largely unknown. Here, we report a novel role of p73 in regulating lipid metabolism by direct transactivation of the promoter of autophagy-related protein 5 (ATG5), a gene whose product is required for autophagosome formation. Following nutrient deprivation, the livers of p73-deficient mice demonstrate a massive accumulation of lipid droplets, together with a low level of autophagy, suggesting that triglyceride hydrolysis into fatty acids is blocked owing to deficient autophagy (macrolipophagy). Compared with wild-type mice, mice functionally deficient in all the p73 isoforms exhibit decreased ATG5 expression and lower levels of autophagy in multiple organs. We further show that the TAp73α is the critical p73 isoform responsible for inducing ATG5 expression in a p53-independent manner and demonstrate that ATG5 gene transfer can correct autophagy and macrolipophagy defects in p73-deficient hepatocytes. These data strongly suggest that the p73–ATG5 axis represents a novel, key pathway for regulating lipid metabolism through autophagy. The identification of p73 as a major regulator of autophagy suggests that it may have an important role in preventing or delaying disease and aging by maintaining a homeostatic control.     

Autophagy regulates neuronal excitability by controlling cAMP/protein kinase A signaling at the synapse

Autophagy provides nutrients during starvation and eliminates detrimental cellular components. However, accumulating evidence indicates that autophagy is not merely a housekeeping process. Here, by combining mouse models of neuron‐specific ATG5 deficiency in either excitatory or inhibitory neurons with quantitative proteomics, high‐content microscopy, and live‐imaging approaches, we show that autophagy protein ATG5 functions in neurons to regulate cAMP‐dependent protein kinase A (PKA)‐mediated phosphorylation of a synapse‐confined proteome. This function of ATG5 is independent of bulk turnover of synaptic proteins and requires the targeting of PKA inhibitory R1 subunits to autophagosomes. Neuronal loss of ATG5 causes synaptic accumulation of PKA‐R1, which sequesters the PKA catalytic subunit and diminishes cAMP/PKA‐dependent phosphorylation of postsynaptic cytoskeletal proteins that mediate AMPAR trafficking. Furthermore, ATG5 deletion in glutamatergic neurons augments AMPAR‐dependent excitatory neurotransmission and causes the appearance of spontaneous recurrent seizures in mice. Our findings identify a novel role of autophagy in regulating PKA signaling at glutamatergic synapses and suggest the PKA as a target for restoration of synaptic function in neurodegenerative conditions with autophagy dysfunction.     

PAQR3 controls autophagy by integrating AMPK signaling to enhance ATG14L‐associated PI3K activity

The Beclin1–VPS34 complex is recognized as a central node in regulating autophagy via interacting with diverse molecules such as ATG14L for autophagy initiation and UVRAG for autophagosome maturation. However, the underlying molecular mechanism that coordinates the timely activation of VPS34 complex is poorly understood. Here, we identify that PAQR3 governs the preferential formation and activation of ATG14L‐linked VPS34 complex for autophagy initiation via two levels of regulation. Firstly, PAQR3 functions as a scaffold protein that facilitates the formation of ATG14L‐ but not UVRAG‐linked VPS34 complex, leading to elevated capacity of PI(3)P generation ahead of starvation signals. Secondly, AMPK phosphorylates PAQR3 at threonine 32 and switches on PI(3)P production to initiate autophagosome formation swiftly after glucose starvation. Deletion of PAQR3 leads to reduction of exercise‐induced autophagy in mice, accompanied by a certain degree of disaggregation of ATG14L‐associated VPS34 complex. Together, this study uncovers that PAQR3 can not only enhance the capacity of pro‐autophagy class III PI3K due to its scaffold function, but also integrate AMPK signal to activation of ATG14L‐linked VPS34 complex upon glucose starvation.     

Receptor‐mediated clustering of FIP200 bypasses the role of LC3 lipidation in autophagy

Autophagosome formation requires multiple autophagy‐related (ATG) factors. However, we find that a subset of autophagy substrates remains robustly targeted to the lysosome in the absence of several core ATGs, including the LC3 lipidation machinery. To address this unexpected result, we performed genome‐wide CRISPR screens identifying genes required for NBR1 flux in ATG7^KO cells. We find that ATG7‐independent autophagy still requires canonical ATG factors including FIP200. However, in the absence of LC3 lipidation, additional factors are required including TAX1BP1 and TBK1. TAX1BP1''s ability to cluster FIP200 around NBR1 cargo and induce local autophagosome formation enforces cargo specificity and replaces the requirement for lipidated LC3. In support of this model, we define a ubiquitin‐independent mode of TAX1BP1 recruitment to NBR1 puncta, highlighting that TAX1BP1 recruitment and clustering, rather than ubiquitin binding per se, is critical for function. Collectively, our data provide a mechanistic basis for reports of selective autophagy in cells lacking the lipidation machinery, wherein receptor‐mediated clustering of upstream autophagy factors drives continued autophagosome formation.     

Regulation of autophagosome biogenesis by OFD1‐mediated selective autophagy

Autophagy is a lysosome‐dependent degradation pathway essential to maintain cellular homeostasis. Therefore, either defective or excessive autophagy may be detrimental for cells and tissues. The past decade was characterized by significant advances in molecular dissection of stimulatory autophagy inputs; however, our understanding of the mechanisms that restrain autophagy is far from complete. Here, we describe a negative feedback mechanism that limits autophagosome biogenesis based on the selective autophagy‐mediated degradation of ATG13, a component of the ULK1 autophagy initiation complex. We demonstrate that the centrosomal protein OFD1 acts as bona fide autophagy receptor for ATG13 via direct interaction with the Atg8/LC3/GABARAP family of proteins. We also show that patients with Oral‐Facial‐Digital type I syndrome, caused by mutations in the OFD1 gene, display excessive autophagy and that genetic inhibition of autophagy in a mouse model of the disease, significantly ameliorates polycystic kidney, a clinical manifestation of the disorder. Collectively, our data report the discovery of an autophagy self‐regulated mechanism and implicate dysregulated autophagy in the pathogenesis of renal cystic disease in mammals.     

Triptolide promotes autophagy to inhibit mesangial cell proliferation in IgA nephropathy via the CARD9/p38 MAPK pathway

BackgroundMesangial cell proliferation is the most basic pathological feature of immunoglobulin A nephropathy (IgAN); however, the specific underlying mechanism and an appropriate therapeutic strategy are yet to be unearthed. This study aimed to investigate the therapeutic effect of triptolide (TP) on IgAN and the mechanism by which TP regulates autophagy and proliferation of mesangial cells through the CARD9/p38 MAPK pathway.MethodsWe established a TP‐treated IgAN mouse model and produced IgA1‐induced human mesangial cells (HMC) and divided them into control, TP, IgAN, and IgAN+TP groups. The levels of mesangial cell proliferation (PCNA, cyclin D1, cell viability, and cell cycle) and autophagy (P62, LC3 II, and autophagy flux rate) were measured, with the autophagy inhibitor 3‐Methyladenine used to explore the relationship between autophagy and proliferation. We observed CARD9 expression in renal biopsies from patients and analyzed its clinical significance. CARD9 siRNA and overexpression plasmids were constructed to investigate the changes in mesangial cell proliferation and autophagy as well as the expression of CARD9 and p‐p38 MAPK/p38 MAPK following TP treatment.ResultsAdministering TP was safe and effectively alleviated mesangial cell proliferation in IgAN mice. Moreover, TP inhibited IgA1‐induced HMC proliferation by promoting autophagy. The high expression of CARD9 in IgAN patients was positively correlated with the severity of HMC proliferation. CARD9/p38 MAPK was involved in the regulation of HMC autophagy and proliferation, and TP promoted autophagy to inhibit HMC proliferation by downregulating the CARD9/p38 MAPK pathway in IgAN.ConclusionTP promotes autophagy to inhibit mesangial cell proliferation in IgAN via the CARD9/p38 MAPK pathway.

The proliferation of glomerular mesangial cells is the most basic pathological feature for IgA nephropathy (IgAN). Galactose‐deficient IgA1 (Gd‐IgA1) immune complex deposition activates mesangial cells to mediate specific intracellular signal transduction, promotes mesangial cell proliferation, and initiates kidney damage. Autophagy is involved in mesangial cell proliferation. CARD9 is a risk gene for IgAN. The triptolide (TP), purified from Tripterygium wilfordii hook F., promotes autophagy to inhibit mesangial cell proliferation in IgAN via the CARD9/p38 MAPK pathway.     

PTH‐induced EndMT via miR‐29a‐5p/GSAP/Notch1 pathway contributed to valvular calcification in rats with CKD

BackgroundEndothelial‐to‐mesenchymal transition (EndMT) is a common pathophysiology in valvular calcification (VC) among non‐chronic kidney disease (CKD) patients. However, few studies were investigated in CKD‐induced VC. Parathyroid hormone (PTH) was considered to be an important component of EndMT in CKD‐induced cardiovascular diseases. Therefore, determining whether PTH could induce valvular EndMT and elucidating corresponding mechanism involved further study.MethodsPerforming a 5/6 nephrectomy with a high phosphorus diet was done to construct VC models in rats with CKD. miRNA sequencing was used to ascertain changes in microRNA in human umbilical vein endothelial cells (HUVECs) intervened by PTH. VC was observed by Von Kossa staining and scanning electron microscope.ResultsPTH induced valvular EndMT in VC. Global microRNA expression profiling of HUVECs was examined in PTH versus the control in vitro, in which miR‐29a‐5p was most notably decreased and was resumed by PTHrP(7‐34) (PTH‐receptor1 inhibitor). Overexpression of miR‐29a‐5p could inhibit PTH‐induced EndMT in vitro and valvular EndMT in vivo. The dual‐luciferase assay verified that γ‐secretase‐activating protein (GASP) served as the target of miR‐29a‐5p. miR‐29a‐5p‐mimics, si‐GSAP and DAPT (γ‐secretase inhibitor) inhibited PTH‐induced γ‐secretase activation, thus blocking Notch1 pathway activation to inhibit EndMT in vitro. Moreover, Notch1 pathway activation was observed in VC. Blocking Notch1 pathway activation via AAV‐miR‐29a and DAPT inhibited valvular EndMT. In addition, blocking Notch1 pathway activation was also shown to alleviate VC.ConclusionPTH activates valvular EndMT via miR‐29a‐5p/GSAP/Notch1 pathway, which can contribute to VC in CKD rats.     

EZH2‐mediated inhibition of KLF14 expression promotes HSCs activation and liver fibrosis by downregulating PPARγ

ObjectivesInduction of deactivation and apoptosis of hepatic stellate cells (HSCs) are principal therapeutic strategies for liver fibrosis. Krüppel‐like factor 14 (KLF14) regulates various biological processes, however, roles, mechanisms and implications of KLF14 in liver fibrosis are unknown.Materials and MethodsKLF14 expression was detected in human, rat and mouse fibrotic models, and its effects on HSCs were assessed. Chromatin immunoprecipitation assays were utilized to investigate the binding of KLF14 to peroxisome proliferator‐activated receptor γ (PPARγ) promoter, and the binding of enhancer of zeste homolog 2 (EZH2) to KLF14 promoter. In vivo, KLF14‐overexpressing adenovirus was injected via tail vein to thioacetamide (TAA)‐treated rats to investigate the role of KLF14 in liver fibrosis progression. EZH2 inhibitor EPZ‐6438 was utilized to treat TAA‐induced rat liver fibrosis.ResultsKLF14 expression was remarkably decreased in human, rat and mouse fibrotic liver tissues. Overexpression of KLF14 increased LD accumulation, inhibited HSCs activation, proliferation, migration and induced G2/M arrest and apoptosis. Mechanistically, KLF14 transactivated PPARγ promoter activity. Inhibition of PPARγ blocked the suppressive role of KLF14 overexpression in HSCs. Downregulation of KLF14 in activated HSCs was mediated by EZH2‐regulated histone H3 lysine 27 trimethylation. Adenovirus‐mediated KLF14 overexpression ameliorated TAA‐induced rat liver fibrosis in PPARγ‐dependent manner. Furthermore, EPZ‐6438 dramatically alleviated TAA‐induced rat liver fibrosis. Importantly, KLF14 expression was decreased in human with liver fibrosis, which was significantly correlated with EZH2 upregulation and PPARγ downregulation.ConclusionsKLF14 exerts a critical anti‐fibrotic role in liver fibrosis, and targeting the EZH2/KLF14/PPARγ axis might be a novel therapeutic strategy for liver fibrosis.     

Activated mesangial cells induce glomerular endothelial cells proliferation in rat anti‐Thy‐1 nephritis through VEGFA/VEGFR2 and Angpt2/Tie2 pathway

ObjectivesWe aimed to investigate the underlying mechanism of endothelial cells (ECs) proliferation in anti‐Thy‐1 nephritis.Materials and methodsWe established anti‐Thy‐1 nephritis and co‐culture system to explore the underlying mechanism of ECs proliferation in vivo and in vitro. EdU assay kit was used for measuring cell proliferation. Immunohistochemical staining and immunofluorescence staining were used to detect protein expression. ELISA was used to measure the concentration of protein in serum and medium. RT‐qPCR and Western blot were used to qualify the mRNA and protein expression. siRNA was used to knock down specific protein expression.ResultsIn anti‐Thy‐1 nephritis, ECs proliferation was associated with mesangial cells (MCs)‐derived vascular endothelial growth factor A (VEGFA) and ECs‐derived angiopoietin2 (Angpt2). In vitro co‐culture system activated MCs‐expressed VEGFA to promote vascular endothelial growth factor receptor2 (VEGFR2) activation, Angpt2 expression and ECs proliferation, but inhibit TEK tyrosine kinase (Tie2) phosphorylation. MCs‐derived VEGFA stimulated Angpt2 expression in ECs, which inhibited Tie2 phosphorylation and promoted ECs proliferation. And decline of Tie2 phosphorylation induced ECs proliferation. In anti‐Thy‐1 nephritis, promoting Tie2 phosphorylation could alleviate ECs proliferation.ConclusionsOur study showed that activated MCs promoted ECs proliferation through VEGFA/VEGFR2 and Angpt2/Tie2 pathway in experimental mesangial proliferative glomerulonephritis (MPGN) and in vitro co‐culture system. And enhancing Tie2 phosphorylation could alleviate ECs proliferation, which will provide a new idea for MPGN treatment.     

NLRX1/FUNDC1/NIPSNAP1‐2 axis regulates mitophagy and alleviates intestinal ischaemia/reperfusion injury

ObjectivesMitophagy is considered to be a key mechanism in the pathogenesis of intestinal ischaemic reperfusion (IR) injury. NOD‐like receptor X1 (NLRX1) is located in the mitochondria and is highly expressed in the intestine, and is known to modulate ROS production, mitochondrial damage, autophagy and apoptosis. However, the function of NLRX1 in intestinal IR injury is unclear.Materials and methodsNLRX1 in rats with IR injury or in IEC‐6 cells with hypoxia reoxygenation (HR) injury were measured by Western blotting, real‐time PCR and immunohistochemistry. The function of NLRX1‐FUNDC1‐NIPSNAP1/NIPSNAP2 axis in mitochondrial homeostasis and cell apoptosis were assessed in vitro.ResultsNLRX1 is significantly downregulated following intestinal IR injury. In vivo studies showed that rats overexpressing NLRX1 exhibited resistance against intestinal IR injury and mitochondrial dysfunction. These beneficial effects of NLRX1 overexpression were dependent on mitophagy activation. Functional studies showed that HR injury reduced NLRX1 expression, which promoted phosphorylation of FUN14 domain‐containing 1 (FUNDC1). Based on immunoprecipitation studies, it was evident that phosphorylated FUNDC1 could not interact with the mitophagy signalling proteins NIPSNAP1 and NIPSNAP2 on the outer membrane of damaged mitochondria, which failed to launch the mitophagy process, resulting in the accumulation of damaged mitochondria and epithelial apoptosis.ConclusionsNLRX1 regulates mitophagy via FUNDC1‐NIPSNAP1/NIPSNAP2 signalling pathway. Thus, this study provides a potential target for the development of a therapeutic strategy for intestinal IR injury.     

lncRNA XIST-miR137-ATG5调节细胞自噬功能在肠癌细胞5-氟胞嘧啶耐药性中的作用

摘要 目的：本文旨在研究长链非编码RNA XIST-miR137-ATG5的相互作用,同时探讨其调节细胞自噬功能与肠癌细胞5-氟胞嘧啶敏感性的关系。方法：实时聚合酶链反应(real time PCR)检测XIST与miR-137在肠癌细胞中的表达;采用脂质体转染法将si-XIST,miR-137转染入肠癌SW480及HCT116细胞中。采用CCK-8检测瞬时转染si-XIST对肠癌细胞增殖及5-FU敏感性的影响;并利用双荧光素酶报告实验检测miR-137与XIST, miR-137与ATG5相互关系。Western blot方法检测XIST- miR137- ATG5对细胞自噬的影响。结果：与正常结肠细胞FHC比较, XIST在结肠癌细胞系明显高表达,miR-137在结肠癌细胞系明显低表达。与阴性对照组比较,转染si-XIST后,SW480及HCT116细胞增殖能力明显受到抑制,对F-5U的敏感性增强,且抑制自噬蛋白Beclin-1及LC3II/LC3 I的表达。miR-137可与XIST,ATG5 3''UTR结合,抑制XIST和ATG5的表达及功能。在结肠癌SW480细胞中共转染miR-137 inhibitor或过表达ATG5可逆转XIST沉默引起的5-FU耐药,同时可逆转因XIST沉默引起的自噬蛋白表达的抑制。结论：LncRNA XIST或可通过调控mir137-ATG促进结直肠癌细胞SW480自噬从而提高其对5-FU的耐药,针对其这一机制,可为将来针对结肠癌的靶向治疗提供一定的实验基础。     

Autophagy promotes resistance to photodynamic therapy-induced apoptosis selectively in colorectal cancer stem-like cells

Recent studies have indicated that cancer stem-like cells (CSCs) exhibit a high resistance to current therapeutic strategies, including photodynamic therapy (PDT), leading to the recurrence and progression of colorectal cancer (CRC). In cancer, autophagy acts as both a tumor suppressor and a tumor promoter. However, the role of autophagy in the resistance of CSCs to PDT has not been reported. In this study, CSCs were isolated from colorectal cancer cells using PROM1/CD133 (prominin 1) expression, which is a surface marker commonly found on stem cells of various tissues. We demonstrated that PpIX-mediated PDT induced the formation of autophagosomes in PROM1/CD133⁺ cells, accompanied by the upregulation of autophagy-related proteins ATG3, ATG5, ATG7, and ATG12. The inhibition of PDT-induced autophagy by pharmacological inhibitors and silencing of the ATG5 gene substantially triggered apoptosis of PROM1/CD133⁺ cells and decreased the ability of colonosphere formation in vitro and tumorigenicity in vivo. In conclusion, our results revealed a protective role played by autophagy against PDT in CSCs and indicated that targeting autophagy could be used to elevate the PDT sensitivity of CSCs. These findings would aid in the development of novel therapeutic approaches for CSC treatment.     

ERK/Drp1‐dependent mitochondrial fission contributes to HMGB1‐induced autophagy in pulmonary arterial hypertension

ObjectivesHigh‐mobility group box‐1 (HMGB1) and aberrant mitochondrial fission mediated by excessive activation of GTPase dynamin‐related protein 1 (Drp1) have been found to be elevated in patients with pulmonary arterial hypertension (PAH) and critically implicated in PAH pathogenesis. However, it remains unknown whether Drp1‐mediated mitochondrial fission and which downstream targets of mitochondrial fission mediate HMGB1‐induced pulmonary arterial smooth muscle cells (PASMCs) proliferation and migration leading to vascular remodelling in PAH. This study aims to address these issues.MethodsPrimary cultured PASMCs were obtained from male Sprague‐Dawley (SD) rats. We detected RNA levels by qRT‐PCR, protein levels by Western blotting, cell proliferation by Cell Counting Kit‐8 (CCK‐8) and EdU incorporation assays, migration by wound healing and transwell assays. SD rats were injected with monocrotaline (MCT) to establish PAH. Hemodynamic parameters were measured by closed‐chest right heart catheterization.ResultsHMGB1 increased Drp1 phosphorylation and Drp1‐dependent mitochondrial fragmentation through extracellular signal‐regulated kinases 1/2 (ERK1/2) signalling activation, and subsequently triggered autophagy activation, which further led to bone morphogenetic protein receptor 2 (BMPR2) lysosomal degradation and inhibitor of DNA binding 1 (Id1) downregulation, and eventually promoted PASMCs proliferation/migration. Inhibition of ERK1/2 cascade, knockdown of Drp1 or suppression of autophagy restored HMGB1‐induced reductions of BMPR2 and Id1, and diminished HMGB1‐induced PASMCs proliferation/migration. In addition, pharmacological inhibition of HMGB1 by glycyrrhizin, suppression of mitochondrial fission by Mdivi‐1 or blockage of autophagy by chloroquine prevented PAH development in MCT‐induced rats PAH model.ConclusionsHMGB1 promotes PASMCs proliferation/migration and pulmonary vascular remodelling by activating ERK1/2/Drp1/Autophagy/BMPR2/Id1 axis, suggesting that this cascade might be a potential novel target for management of PAH.     

PAK4 suppresses motor neuron degeneration in hSOD1G93A‐linked amyotrophic lateral sclerosis cell and rat models

ObjectivesAmyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons (MN). CREB pathway‐mediated inhibition of apoptosis contributes to neuron protection, and PAK4 activates CREB signalling in diverse cell types. This study aimed to investigate PAK4’s effect and mechanism of action in ALS.MethodsWe analysed RNA levels by qRT‐PCR, protein levels by immunofluorescence and Western blotting, and apoptosis by flow cytometry and TUNEL staining. Cell transfection was performed for in vitro experiment. Mice were injected intraspinally to evaluate PAK4 function in vivo experiment. Rotarod test was performed to measure motor function.ResultsThe expression and activation of PAK4 significantly decreased in the cell and mouse models of ALS as the disease progressed, which was caused by the negative regulation of miR‐9‐5p. Silencing of PAK4 increased the apoptosis of MN by inhibiting CREB‐mediated neuroprotection, whereas overexpression of PAK4 protected MN from hSOD1^G93A‐induced degeneration by activating CREB signalling. The neuroprotective effect of PAK4 was markedly inhibited by CREB inhibitor. In ALS models, the PAK4/CREB pathway was inhibited, and cell apoptosis increased. In vivo experiments revealed that PAK4 overexpression in the spinal neurons of hSOD1^G93A mice suppressed MN degeneration, prolonged survival and promoted the CREB pathway.ConclusionsPAK4 protects MN from degeneration by activating the anti‐apoptotic effects of CREB signalling, suggesting it may be a therapeutic target in ALS.

Schematic representation of the mechanism of PAK4 protecting MN from apoptosis in ALS. PAK4 increases CREB levels and activation, leading to the upregulation of PGC‐1a and Bcl‐2, thereby decreasing cleaved‐caspase3 levels, and inhibiting MN degeneration. miR‐9‐5p is responsible for the decreased expression of PAK4 in ALS.     

JUNB‐FBXO21‐ERK axis promotes cartilage degeneration in osteoarthritis by inhibiting autophagy

Osteoarthritis (OA) is a heterogeneous disease that is extremely hard to cure owing to its complex regulation network of pathogenesis, especially cartilage degeneration. FBXO21 is a subunit of ubiquitin E3 ligases that degrades P‐glycoprotein and EID1 by ubiquitination and activates the JNK and p38 pathways; however, its role in OA remains unknown. Here, the main objective of this study was to evaluate the potential effects and mechanism of FBXO21 in OA degeneration, we revealed that FBXO21 is upregulated in the cartilage of patients with OA, aging, and monosodium iodoacetate‐induced OA rats, and chondrocytes treated with interleukin‐1β, tumor necrosis factor‐α, and lipopolysaccharide. Moreover, the in vivo and in vitro knockdown of FBXO21 suppressed OA‐related cartilage degeneration, as evidenced by activated autophagy, upregulated anabolism, alleviated apoptosis, and downregulated catabolism. In contrast, its overexpression promoted OA‐related cartilage degeneration. In addition, using mass spectrometry and co‐immunoprecipitation assay, we demonstrated that the downstream mechanism of FBXO21 inhibits autophagy by interacting with and phosphorylating ERK. Furthermore, FBXO21 alleviated anabolism and enhanced apoptosis and catabolism by inhibiting autophagy in rat chondrocytes. Interestingly, for its upstream mechanism, JUNB promoted FBXO21 expression by directly targeting the FBXO21 promoter, thus further accelerating cartilage degeneration in SW1353 cells and rat chondrocytes. Overall, our findings reveal that the JUNB‐FBXO21‐ERK axis regulates OA apoptosis and cartilage matrix metabolism by inhibiting autophagy. Therefore, FBXO21 is an attractive target for regulating OA pathogenesis, and its knockdown may provide a novel targeted therapy for OA.