Xinhua Liu

Cardiac fibrosis remains a critical determinant of adverse outcomes in heart disease, yet effective anti-fibrotic therapies are lacking. While multiple semaphorin family members participate in cardiovascular pathophysiology, the role of semaphorin 4f (Sema4f) in cardiac fibrosis remains unexplored. This study investigates the role and mechanisms of Sema4f in fibrotic remodeling post-myocardial infarction (MI). We employed flow cytometry to characterize cell type-specific Sema4f expression patterns in post-MI hearts. Lineage-specific knockout mice (fibroblast vs. myeloid) were subjected to left anterior descending ligation to assess functional consequences. Proteomic analysis of Sema4f-deficient cardiac fibroblasts was conducted to identify downstream effectors. Key pathways were subsequently validated using pharmacological inhibitors. We found that Sema4f expression was markedly upregulated during the fibrotic phase post-MI, primarily due to fibroblast activation. Fibroblast-, but not myeloid-, specific Sema4f deletion significantly reduced fibrosis and improved cardiac function. Proteomic profiling revealed that Sema4f deficiency led to downregulation of pro-fibrotic gene expression, which was associated with impaired actin cytoskeletal remodeling and decreased nuclear translocation of YAP/TAZ. Pharmacological inhibition of either actin remodeling or YAP/TAZ activity attenuated fibrosis, whereas YAP/TAZ activation abolished the anti-fibrotic effects of Sema4f knockout. Our study provides the first evidence demonstrating the functional role of Sema4f in cardiac fibroblast activation and fibrosis progression. We have identified a fibroblast-specific mechanism mediated by the Sema4f-actin cytoskeleton-YAP/TAZ axis, offering novel mechanistic insights into fibrosis regulation and revealing a promising therapeutic target for cardiac fibrosis with potential clinical applications. Graphical abstract

Obesity, a major global health challenge associated with metabolic and cardiovascular disorders, has drawn increasing attention to the therapeutic potential of white adipose tissue (WAT) browning. Although the lysine methyltransferase SETD7 has been implicated in various cardiovascular and metabolic diseases, its role in adipose thermogenesis remains unclear. Here, we reported that SETD7 was upregulated in inguinal WAT (iWAT) of obese mice and was primarily localized to mature adipocytes. Setd7 knockdown ( Setd7 ^⁺/⁻ ) mice exhibited enhanced thermogenic gene expression and iWAT browning upon cold exposure or β3-adrenergic stimulation, whereas thermogenic activity in brown adipose tissue (BAT) remained largely unaffected. In vitro, SETD7 knockdown did not alter adipogenesis but potently augmented thermogenic capacity in beige adipocytes, while SETD7 overexpression exerted the opposite effect. Mechanistically, RNA-Seq analysis revealed that SETD7 deficiency upregulated Adcy7 transcription, leading to increased Sirt1 levels and enhanced Creb1 phosphorylation, thereby activating the thermogenic program. Notably, Setd7 ^{⁺/ –} mice resisted high-fat diet (HFD)-induced obesity, exhibiting reduced weight gain, elevated energy expenditure, and improved metabolic health. Together, these findings identify SETD7 as a negative regulator of iWAT thermogenesis and suggest that targeting SETD7 may represent a promising strategy for combating obesity.

Ligstroside (LIG) is a phenolic secoiridoid glycoside naturally occurring in olives and structurally related to the widely studied olive constituent oleuropein. Although LIG has been reported to exhibit diverse biological activities, it has received comparatively limited attention, and the molecular targets underlying its bioactivity remain largely unknown, thereby limiting exploration of its pharmacological potential. To facilitate target identification and mechanistic investigation, we designed and synthesized a LIG-derived photoaffinity probe (PAL-LIG) through minimal modification of the LIG scaffold. The probe incorporates a photoactivatable diazirine group that enables UVinduced covalent crosslinking with proximal proteins together with an alkyne handle that allows subsequent bioorthogonal conjugation via copper-catalyzed azide–alkyne cycloaddition (CuAAC). Following photo-crosslinking, probe-labeled proteins can be conjugated with azide-functionalized reporter tags, including biotin for streptavidinbased enrichment and pulldown or TAMRA for fluorescence detection and cellular imaging. Using this strategy, PAL-LIG enables enrichment of potential LIG-binding proteins as well as fluorescence-based visualization of probe labeling in cells. Importantly, PAL-LIG retains biological activity comparable to LIG, as demonstrated by its similar ability to promote ATP production in mouse C2C12 myoblasts, indicating that installation of the photoaffinity and alkyne handles minimally perturbs the intrinsic bioactivity of LIG. Collectively, PAL-LIG provides a chemical tool for investigating the direct protein targets and molecular mechanisms of LIG.

Epigenetic regulation plays a pivotal role in adipocyte development and thermogenesis. Ash2l, a key component of the COMPASS (Complex of Proteins Associated with Set1) histone methyltransferase, regulates gene expression through epigenetic mechanisms. This study explored the role of Ash2l in adipose tissue thermogenesis and obesity-related metabolic dysfunction. Ash2l was initially identified through transcriptomic analysis, and its expression was further validated in mouse models of high-fat diet (HFD), cold exposure, and CL316,243 stimulation. In vitro gain- and loss-of-function experiments were conducted to assess the role of Ash2l in adipogenesis and thermogenesis. To knockdown Ash2l in vivo, adeno-associated viruses carrying short hairpin RNA targeting Ash2l (AAV-shAsh2l) were injected into either the brown adipose tissue (BAT) or the inguinal white adipose tissue (iWAT). The functional consequences of Ash2l deficiency were evaluated in mice under room temperature, cold exposure, and HFD conditions. Finally, chromatin immunoprecipitation sequencing (ChIP-seq) was employed as an exploratory analysis to identify genomic regions associated with Ash2l during adipocyte development. Our findings demonstrate that Ash2l modulates the expression of both adipogenic and thermogenic genes in adipocytes. Mice with BAT- or iWAT-knockdown of Ash2l displayed defective cold-induced thermogenesis, aggravated diet-induced obesity, and systemic metabolic dysregulation. Moreover, Ash2l knockdown in BAT under cold exposure or HFD conditions also attenuated thermogenic activity in iWAT, an effect that may be mediated by reduced secretion of FABP4. These findings establish Ash2l as a critical regulator of adipogenesis and thermogenesis. This study provides important insights into the epigenetic role of Ash2l in maintaining metabolic homeostasis under conditions of nutritional excess.

Beige adipocytes play a key role in non-shivering thermogenesis. SEMA3E, a member of the class 3 semaphorin family, is involved in various pathological processes, but its role in adipocyte differentiation and thermogenesis remains unclear. Here, we found SEMA3E expression increased in inguinal white adipose tissue (iWAT) following cold exposure or β-adrenergic agonist CL316,243 stimulation. In vitro, loss- and gain-of-function experiments revealed that SEMA3E promoted beige adipocyte differentiation and enhanced thermogenic genes expression. In vivo, fat transplantation experiments indicated that SEMA3E promoted adipogenesis. Furthermore, adeno-associated virus (AAV)-mediated SEMA3E knockdown in iWAT impaired thermogenesis in mice exposed to cold or CL316,243. RNA-Seq analysis linked SEMA3E to mitochondrial oxidative phosphorylation, and its knockdown reduced mitochondrial respiration by downregulating respiratory chain components expression and lowering mitochondrial oxygen consumption rate. Mechanistically, gene set enrichment analysis suggested SEMA3E regulated beige adipocyte differentiation via the Wnt/β-catenin pathway. SEMA3E knockdown delayed β-catenin degradation, while inhibiting this pathway with IWR-1 rescued the suppressed differentiation and thermogenic genes expression. In conclusion, these findings highlight the crucial role of SEMA3E in beige adipocyte differentiation and thermogenesis.

Background: Skeletal muscle atrophy is a debilitating condition associated with diverse diseases and clinical interventions. Although triggers such as glucocorticoid excess or direct muscle injury disrupt the balance between protein homeostasis and tissue repair, the upstream molecular signals that actively preserve muscle integrity remain largely unknown. Semaphorin 4A (Sema4A) is a Class IV transmembrane semaphorin traditionally recognized for its roles in neural development and immune regulation; however, its function in skeletal muscle maintenance has not been explored. Methods: We first examined Sema4A expression in established models of muscle catabolism. Subsequently, adeno‐associated virus (AAV)‐mediated Sema4A restoration was employed to evaluate its therapeutic potential in both dexamethasone‐induced atrophy and acute injury mouse models. Mechanistic studies were performed in C2C12 myotubes using gain‐ and loss‐of‐function approaches. Results: Sema4A expression was significantly downregulated under atrophic conditions ( p < 0.05). Restoration of Sema4A effectively attenuated dexamethasone‐induced muscle loss (4.766 vs. 5.075 mg/g B.W., p = 0.0414) and accelerated functional recovery (6.219 × 10 ⁻² vs. 7.124 × 10 ⁻² N/g B.W., p = 0.0422). We identified Plexin B2 (Plxnb2) as the functional receptor mediating these effects. At the molecular level, Sema4A signalling suppressed FoxO3a nuclear translocation (FoxO3a positive cells: vector‐Dex 27.99% vs. OE‐Sema4A‐Dex 12.49%, p < 0.05), thereby inhibiting the expression of key atrogenes, while simultaneously reactivating the PI3K‐AKT‐mTOR anabolic pathway. This reprogramming of intracellular metabolic signalling was further associated with the establishment of a reparative immune microenvironment, a process potentially modulated by muscle‐derived factors such as Gdf15. Conclusions: Our study identifies Sema4A as a novel protective regulator that mitigates muscle atrophy and enhances regeneration through a dual mechanism: restoring intracellular anabolic signalling and fostering a proregenerative immune niche. These findings highlight Sema4A as a promising therapeutic target for the treatment of muscle‐wasting disorders.

BACKGROUND: Myocardial infarction (MI)–induced ischemia leads to abnormal ventricular remodeling and cardiac fibrosis, which can ultimately progress to heart failure. Kif23 (kinesin-like protein 23) has been implicated in the progression of various diseases. This study aims to investigate the role of Kif23 in the development of cardiac fibrosis following MI. METHODS: Male C57BL/6J mice received intravenous injections of adeno-associated virus carrying Kif23 shRNA, followed by left anterior descending coronary artery ligation to induce MI. Cardiac function and fibrosis were assessed via echocardiography, histological analysis, and fibrosis marker quantification at 7 and 14 days post-MI. In vitro, adult rat cardiac fibroblasts underwent Kif23 knockdown or overexpression, with subsequent TGF-β1 (Transforming Growth Factor Beta 1) treatment. Proteomic profiling, lipid droplet analysis, and fibrosis/lipid metabolism assessments were performed. RESULTS: Kif23 expression was significantly upregulated in both infarcted myocardium and TGF-β1–stimulated adult rat cardiac fibroblasts. Kif23 knockdown improved cardiac function and attenuated fibrosis post-MI. In vitro, Kif23 silencing suppressed fibroblast proliferation and myofibroblast transdifferentiation, whereas Kif23 overexpression potentiated fibrotic responses. Proteomic profiling identified Ces1d (carboxylesterase 1d) as a key downstream effector upregulated by Kif23 knockdown, concomitant with restored fatty acid β-oxidation. Mechanistically, Kif23 impairs Ces1d-mediated fatty acid β-oxidation via RhoA (Ras Homolog Family Member A)/ROCK1 (Rho-associated protein kinase 1) signaling, driving fibrosis progression. CONCLUSIONS: Our study identifies Kif23 as a novel regulator of post-MI cardiac fibrosis. Mechanistically, Kif23 drives fibroblast proliferation through RhoA activation and exacerbates fibrogenesis by suppressing Ces1d-dependent fatty acid β-oxidation via the RhoA/ROCK1 axis. This metabolic disruption triggers pathological lipid accumulation and myofibroblast transdifferentiation. These findings nominate Kif23 as a promising therapeutic target for antifibrotic interventions.

Objective: High‐altitude environments are characterized by hypobaric hypoxia and pose a significant physiological challenge. While obesity‐related hypoxia is known to impair brown adipose tissue (BAT) function by suppressing lipolysis and thermogenesis, whether hypobaric hypoxia similarly compromises BAT function remains unclear. We hypothesize that, unlike obesity‐associated hypoxia, hypobaric hypoxia perturbs BAT thermogenesis via a distinct mechanism involving the upregulation of Plin2. Methods: Mice were exposed to either normoxia or hypobaric hypoxia (simulating 6000 m, 9.2% O₂) for 30 days. BAT function was assessed by analyzing lipid droplet size, lipid and triglyceride content, and expression of thermogenic protein. Thermogenic capacity was further evaluated in mice subjected to 30 days of normoxia or hypobaric hypoxia, followed by a 2‐day cold exposure or 3‐day treatment with the β3‐adrenergic agonist CL316243. Differential proteomics and in vitro experiments were performed to explore underlying molecular mechanisms. Results: After 30 days of hypobaric hypoxia, mice exhibited reduced body weight (normoxia: 26.64 ± 0.91 vs. hypoxia: 21.94 ± 0.79 g, p = 0.002), yet exhibited enlarged lipid droplets in BAT (142.7 ± 15.52 vs. 387.4 ± 31.91 μm ² , p < 0.001) and increased lipid accumulation. Compared with normoxia, hypoxic mice displayed impaired thermogenic responses to both cold exposure and β3‐adrenergic stimulation, as indicated by suppressed thermogenic gene expression. Mechanistically, hypobaric hypoxia elevated lactate levels and upregulated PPARγ, which subsequently enhanced Plin2 expression, ultimately leading to defective lipolysis and impaired thermogenesis. Conclusions: We identify a novel hypoxia‐lactate/PPARγ‐Plin2 axis that uncouples body weight loss from BAT thermogenesis under hypobaric hypoxia. This pathway represents a previously unrecognized therapeutic target for counteracting metabolic dysfunction induced by high‐altitude exposure.

Diabetic nephropathy (DN) is a leading cause of end-stage renal disease (ESRD), and there has been growing attention towards the role of epigenetics in its pathogenesis. This study aims to investigate the underlying mechanism by which SET domain-containing lysine methyltransferase 7 (SETD7) modulates the progression of DN. SETD7 knockout mice (Setd7−/−) and wild-type controls (Setd7+/+) were intraperitoneally injected with Streptozocin (STZ, 50 mg/kg) to induce DN. The mouse mesangial cell line SV40-MES-13 were stimulated by high glucose in vitro. SETD7 expression was predominantly upregulated in high glucose-induced glomerular mesangial cells (MCs), and in diabetic mice kidney. Notably, SETD7 promoted the transcription of adenosine A2B receptor (A2BR), which mediated the activation and phosphorylation of JAK2 (p-JAK2) via direct interaction. Subsequently, p-JAK2 facilitated the recruitment of signal transducer and activator of transcription 3 (STAT3) to binding sites. This led to nuclear translocation of STAT3 and transcriptional regulation of target genes, ultimately promoting fibrosis and inflammation. Importantly, SETD7 deficiency reduced A2BR transcription, thereby inhibiting fibrosis and inflammation in high glucose-induced MCs. Consistently with these findings, SETD7 knockout in STZ-induced mice conferred significant protection against renal injury and reduced glomerular fibrosis. Our results demonstrate an important role and mechanism of SETD7 in DN by promoting fibrosis and inflammation through the A2BR-mediated JAK2/STAT3 signaling pathway. Targeting SETD7 may represent a promising therapeutic strategy for halting the progression of DN.