Beclin 1 Recombinant Rabbit Monoclonal Antibody [47F02]

Skeletal muscle atrophy, a debilitating complication of COPD, is closely linked to cigarette smoke (CS) exposure. The epigenetic regulator HDAC2 has been implicated, but the upstream regulatory mechanisms and precise downstream pathways are unclear. Using a CS-induced mouse atrophy model and C2C12 myotubes treated with cigarette smoke extract (CSE), we systematically investigated the role of USP47/HDAC2/CYP1A1/ROS axis through gain/loss-of-function studies, RNA-seq, ChIP-qPCR, co-immunoprecipitation, and ubiquitination assays. HDAC2 was downregulated in atrophic muscle, and its overexpression mitigated CS-induced atrophy, improved grip strength, and enhanced muscle regeneration. HDAC2 acted as a transcriptional repressor of CYP1A1 by deacetylating H3K9 and H3K27 at the promoter, thus curtailing ROS-driven excessive autophagy. We further discovered that the deubiquitinase USP47 is the key upstream regulator of HDAC2. USP47 directly interacted with HDAC2, promoted its deubiquitination, and enhanced its protein stability. Consequently, USP47 overexpression phenocopied the benefits of HDAC2 overexpression, which were effectively nullified by restoring CYP1A1 expression. In conclusion, we delineate a previously unrecognized signaling axis wherein USP47 stabilizes HDAC2 to inhibit the CYP1A1/ROS/autophagy cascade, ultimately protecting against CS-induced skeletal muscle atrophy. Targeting the USP47-HDAC2 interface presents a novel therapeutic strategy for combating muscle wasting in COPD.

Ethnopharmacological relevance Guanxin Ⅱ (GXⅡ) is a traditional Chinese herbal formula, which has the functions of activating blood to resolve stasis and move qi to relieve pain. Qi and blood stagnant is regarded as the main pathogenesis of atherosclerosis (AS) and depression, but the specific mechanism of GXⅡ in the treatment of AS comorbid depression (co-depression) remains unclear. Aim of the study This study aimed to investigate the therapeutic mechanisms of GXⅡ in ameliorating AS co-depression through multi-target network regulation and ATG3-mediated autophagy pathway, integrating pharmacochemical profiling, network pharmacology, and experimental validation. Materials and methods The absorbed components of GXⅡ in mouse plasma were identified by UPLC-Q-TOF/MS. ApoE −/− mice were fed with a high fat diet (HFD) combined with restraint stress for 16 weeks, and GXⅡ was administered orally. Anti-AS effects were evaluated by serum lipid panel, aortic Oil Red O staining and HE staining. Antidepressant effects were assessed by open field test (OFT), sucrose preference test (SPT), and tail suspension test (TST). Network pharmacology was employed to predict core targets, which were validated by molecular docking and molecular dynamics simulation. The regulatory effects of GXⅡ on autophagy were confirmed by Western blotting and transmission electron microscopy in mouse hippocampus, and further verified in LPS-stimulated primary hippocampal neurons focusing on ATG3-mediated pathway. Results GXⅡ treatment ameliorated dyslipidemia and reduced aortic plaque formation in mice with AS co-depression. It also demonstrated antidepressant effects in the OFT, SPT and TST. Network pharmacology analysis identified 77 overlapping targets among GXⅡ, AS and depression, with 15 core genes selected via PPI network topology. Molecular docking and dynamics simulation revealed robust binding interactions between GXⅡ constituents and AKT1, STAT3, and TLR4. Notably, GXⅡ restored the suppressed autophagy in the hippocampus of AS co-depressed mice, as evidenced by upregulated LC3, Beclin1 and ATG3 expression. These autophagy-regulating effects were consistently observed in primary hippocampal neurons and were reduced upon lentiviral knockdown of ATG3, confirming its essential role in mediating GXⅡ's therapeutic action. Conclusions GXⅡ ameliorates atherosclerosis and depression-like behaviors in comorbid mice via modulating the AKT1-STAT3-TLR4 signaling axis and activating ATG3-mediated autophagy. These findings provide pharmacological evidence to support the application of GXⅡ in the treatment of AS co-depression, and highlight ATG3 as a potential therapeutic target.

Mycobacterium tuberculosis infection of the trachea causes tracheobronchial tuberculosis (TBTB), a chronic inflammatory illness, and its pathological features include macrophage pyroptosis. RanBP3, as a potential regulator, participates in cell stress, but its specific mechanism of action in TBTB has not been clarified. The mouse and bone marrow-derived macrophages (BMDMs) models were constructed by infecting with M. smegmatis . RanBP3 protein level and the levels of pyroptosis-related proteins were measured by Western blotting (WB). The levels of inflammatory factors were measured by ELISA. Pearson’s correlation analysis was employed to evaluate their correlation with RanBP3, and the interaction of RanBP3/CCAR2/SIRT1 was verified by Co-immunoprecipitation (Co-IP). The detection of macrophage pyroptosis and mitophagy was done using flow cytometry, and mitophagy was further assessed by transmission electron microscopy. Bacterial survival in BMDMs was evaluated by colony-forming unit assays. Hematoxylin and eosin (H&E) and Masson staining were used to assess lung tissue damage. RanBP3 was down-regulated in TBTB mucosa. At the cellular level, RanBP3 overexpression alleviated BMDM’s pyroptosis by promoting mitophagy and reduced intracellular bacterial survival, whereas RanBP3 knockdown exacerbated pyroptosis and increased bacterial survival. Mechanistically, RanBP3 competitively binds to CCAR2, disrupts the CCAR2-SIRT1 interaction, and promotes SIRT1 release, thereby activating mitophagy. At the animal level, RanBP3 overexpression promoted mitophagy, alleviated macrophage pyroptosis, and alleviated TBTB, but the SIRT1 inhibitor reversed these effects. RanBP3 mediates the release of SIRT1 through the CCAR2-SIRT1 axis, thereby activating mitophagy and inhibiting pyroptosis, providing a new target for TBTB treatment.

Distal hereditary motor neuropathy (dHMN) is a progressive neurological disease characterized by distal limb muscle weakness and amyotrophy. Sigma 1 receptor (σ1R), a gene product of SIGMAR1, mutations have been reported to induce dHMN, but its mechanism remains unknown. This study aims to explore the effect of C238T and 31_50del mutations in σ1R on neuronal SH-SY5Y cell functions. The SH-SY5Y cells that overexpressed σ1R, C238T mutant σ1R (σ1R C238T ) or 31_50del mutant σ1R (σ1R 31_50del ) were constructed by pEGFPN1 vectors. We used Western blot (WB) and immunofluorescence (IF) staining to detect the expression of σ1R and green fluorescent proteins (GFP). Then, we evaluated the impact of σ1R mutation on apoptosis, autophagy, endoplasmic reticulum stress, and the involvement of the unfolded protein response (UPR) pathway in SH-SY5Y cells. We found that σ1R C238T and σ1R 31_50del downregulated σ1R and promoted the apoptosis of SH-SY5Y cells. σ1R C238T and σ1R 31_50del increased p-PERK, p-eIF2α, p-JNK, BIP, ATF4, CHOP, ATF6, XBP1, Caspase3, Caspase12 expressions and Ca 2+ concentration, whereas decreased ATP content in SH-SY5Y cells. Besides, the expressions of LC3B, Lamp1, ATG7, Beclin-1 and phosphorylation of AMPK and ULK1 were increased, while the p62 level decreased after C238T or 31_50del mutation of σ1R. Additionally, AMPK knockdown abolished the apoptosis mediated by σ1R C238T or σ1R 31_50del in SH-SY5Y cells. Our results indicated that C238T or 31_50del mutation in σ1R promoted motor neuron apoptosis through the AMPK/ULK1 pathway in dHMN. This study shed light on a better understanding of the neurons pathological mechanisms mediated by σ1R C238T and σ1R 31-50del in dHMN.