Abstract Thoracic aortic aneurysm and dissection (TAAD) is a life-threatening disease characterized by progressive medial degeneration, impaired mechanical integrity, and extracellular matrix (ECM) degradation. However, no pharmacological therapy has been proven to halt aneurysm progression or prevent dissection or rupture. Vascular smooth muscle cells (VSMCs) are vital for maintaining medial architecture by sensing and remodeling the surrounding ECM; however, the mechanism by which abnormal ECM mechanics are transmitted to nuclear transcriptional programs that disrupt aortic wall matrix homeostasis remains incompletely understood. Integrative transcriptomic screening of Lysyl oxidase ( LOX )-deficient and β-aminopropionitrile (BAPN)-induced TAAD models identified vestigial-like family member 4 (VGLL4) as a mechanosensitive transcriptional regulator of TAAD. VGLL4 was enriched in VSMCs and markedly increased in aortas from patients with TAAD and BAPN-induced TAAD mice. VSMC-specific deletion of Vgll4 protected mice from BAPN-induced aortic dilation, dissection, rupture-associated mortality, vascular stiffening, ECM degradation, and medial destruction. Mechanistically, pathological matrix remodeling and mechanical stress induced VGLL4 expression in VSMCs, where VGLL4 cooperated with specificity protein 1 (SP1) to activate Wisp1 transcription. In vivo , VSMC-enriched Wnt-inducible signaling pathway protein (WISP1) overexpression exacerbated TAAD progression, whereas Wisp1 knockdown protected against BAPN-induced TAAD and mitigated the severe aortic phenotype driven by VGLL4 overexpression. Secreted WISP1 bound Tissue Inhibitor of Metalloproteinases 3 (TIMP3) through its C-terminal domain and impaired TIMP3-mediated MMP9 inhibition, thereby increasing MMP9 proteolytic activity and accelerating ECM degradation. Consistently, in vivo Wisp1 knockdown protected against BAPN-induced TAAD. Together, these findings define the VGLL4-WISP1-TIMP3/MMP9 axis, which couples pathological ECM mechanics to nuclear transcriptional activation and protease-dependent matrix degradation in VSMCs. This pathway promotes medial structural failure, aortic mechanical stability loss, and TAAD progression, identifying WISP1 as a potential therapeutic target for preserving aortic wall matrix homeostasis.

Hirsutine, a potent drug-like indole alkaloid extracted from Uncaria rhynchophylla, exhibits several biological activities, including cardioprotective effects. However, the underlying regulatory mechanisms remain unclear. Herein, we aimed to examine the therapeutic effects of hirsutine on obesity-related cardiomyopathy and investigate the potential mechanism underlying these effects. An obesity cardiomyopathy mouse model was developed by subjecting mice to a high-fat diet (HFD) for 16 consecutive weeks, followed by an 8-week hirsutine treatment. H9c2 cardiomyocytes treated with palmitate were utilized as an in vitro model. Invasive hemodynamic parameters and left ventricular hypertrophy indices were assessed, and the expression of related signaling molecules was analyzed using western blotting, mass spectrometry, molecular docking, RNA sequencing, immunoprecipitation, histological analysis, and transmission electron microscopy, respectively. Hirsutine significantly alleviated HFD-induced cardiomyopathy in the mouse model. Notably, the therapeutic effect of hirsutine was reversed in Midivi-1-treated mice, indicating that the cardioprotective role of hirsutine is dependent on mitochondrial fission-mediated mitophagy and Parkin. Mechanically, hirsutine maintained Parkin protein stability, and the C-terminal region of 1103-1394 amino acids of leucine-rich pentatricopeptide repeat-containing protein (LRPPRC) functions as a binding motif interacting with Parkin. LRPPRC overexpression significantly enhanced Parkin protein stability, which was attenuated by deletion of the 1103-1394 amino acids of LRPPRC (LRPPRCΔ1103-1394). Collectively, these findings demonstrate that hirsutine ameliorates HFD-induced cardiomyopathy by promoting Parkin protein stability through its interaction with 1103-1394 amino acids of LRPPRC. Therefore, targeting LRPPRC may represent a promising therapeutic strategy underlying the protective effects of hirsutine in HFD-induced cardiomyopathy.

Stem cell-derived extracellular vesicles (EVs) hold therapeutic potential for hypoxia-associated injury, yet the molecular mechanisms underlying their protective effects remain incompletely defined. Herein, we performed comparative proteomic profiling of extracellular vesicles (EVs) secreted by human placenta-derived perivascular stem cells (hPPSCs) and human umbilical cord mesenchymal stem cells (hUCMSCs). We found that hPPSCs-EVs are enriched in proteasome-related proteins. Enzymatic activity assays further confirmed that proteasome activity in hPPSCs-EVs was significantly higher than that in hUCMSCs-EVs. Additionally, we observed that hPPSCs-EVs were efficiently endocytosed by human umbilical vein endothelial cells (hUVECs), leading to marked downregulation of hypoxia-inducible factor 1-alpha (HIF-1α) and intracellular ubiquitinated proteins. Importantly, this HIF-1α degradation persisted even when the host ubiquitin-proteasome system was blocked, indicating that hPPSCs-EVs can function independently of the host proteasomal pathway. These results reveal that hPPSCs-EVs deliver functional proteasomes to endothelial cells, thereby compensating for impaired protein degradation and protecting cells under hypoxic stress. Collectively, our findings provide evidence for an intercellular transfer of proteolytic capacity via stem cell-derived EVs, a mechanism that preserves endothelial proteostasis and highlights the therapeutic potential of proteasome-rich EVs for hypoxia-associated diseases.

A bioengineered kidney using a decellularized kidney scaffold (DKS) offers a promising solution to the kidney shortage. However, the transplantation of bioengineered kidneys using DKS recellularized with human endothelial, renal cells or others has not yet successfully achieved the vascular and renal reconstruction in vivo . In this study, we identified another type of stem cells, designated as human placenta-derived angiogenic stem cells (hPASCs), which serve as seeding cells for the vascularization of DKS. These hPASCs encompass angiogenic subpopulations, exhibit both stem cell properties and the capacity for vascular differentiation. Human fetal kidney organoids (KIO) were established as a source of renal parenchymal cells, comprising renal, immune, and vascular cell populations. We developed a vascularized bioengineered kidney by recellularizing DKS with hPASCs and KIO using a circulation perfusion culture system. The hPASCs bioengineered kidney demonstrated enhanced angiogenesis and reconstructed renal architecture in vivo , by transplantation into rat models of renal subcapsular and partial nephrectomy. Furthermore, immunofluorescence and single-cell analyses revealed that the hPASCs revascularized bioengineered kidney regenerated both vascular and renal parenchymal cells within the host. This study offers another strategy for kidney bioengineering and regeneration. ### Competing Interest Statement The authors have declared no competing interest. National Key Research and Development Program of China, 2023YFC3404301 Zhejiang Provincial Natural Science Foundation of China, LY24H180005 Open Project of State Key Laboratory of Animal Biotech Breeding, 2025SKLAB6-16 The Summit Advancement Disciplines of Zhejiang Province, Wenzhou Medical University–Pharmaceutics Zhejiang Provincial Natural Science Foundation of China, LZ24H120003

Thoracic aortic dissection (TAD) is a life-threatening condition characterized by medial degeneration and vascular smooth muscle cell (VSMC) dysfunction, with no effective medical therapy currently available. The underlying pathological mechanisms of TAD remain incompletely understood. In this study, we used a nonintegrated episomal vector-based reprograming system to generate induced pluripotent stem cells (iPSCs) from TAD patients and healthy controls. Both TAD and normal iPSCs expressed key pluripotency markers and were capable of differentiating into the three germ layers in vitro. These iPSCs were differentiated into vascular smooth muscle cells (VSMCs) through a mesodermal intermediate for disease modeling. VSMCs derived from both TAD and normal iPSCs expressed smooth muscle α-actin (α-SMA), calponin (CNN), and SM22α. However, TAD-iPSC-derived VSMCs exhibited significantly reduced contraction in response to carbachol stimulation compared to their normal counterparts. Whole-exome sequencing identified a mutation in the COL4A2 gene (c.392G > T, p. R131M) in TAD-iPSCs. This mutation was associated with reduced collagen IV expression and increased expression of collagen I and III in TAD-VSMCs, both with and without TGF-β stimulation. Furthermore, noncanonical TGF-β signaling was hyperactivated in TAD-VSMCs, accompanied by elevated MMP9 expression. This patient-specific iPSC model reveals key dysfunctions in VSMC contractility, extracellular matrix (ECM) protein expression, and dysregulated TGF-β signaling, which may contribute to TAD pathogenesis. Our findings provide new insights into the molecular mechanisms driving TAD and offer a platform for future therapeutic development.

Background Marfan syndrome (MFS) is a heritable connective tissue disorder caused by mutations in the Fibrillin‐1 gene, which encodes the extracellular matrix protein fibrillin‐1. Patients with MFS are predisposed to aortic aneurysms and dissections, significantly contributing to mortality. Emerging evidence suggests that endothelial cell (EC) senescence plays a critical role in the pathogenesis of aortic aneurysms in MFS. This study aims to elucidate the role of EC senescence in the development of aortic aneurysms in MFS using a vascular model derived from human induced pluripotent stem cells. Methods and Results We generated human induced pluripotent stem cells lines from 2 patients with MFS carrying specific Fibrillin‐1 mutations and differentiated these into ECs. These MFS–hiPSC‐derived ECs were characterized using immunofluorescence, reverse transcription‐quantitative polymerase chain reaction, and Western blotting. Functional assays including cell proliferation, scratch wound, tube formation, NO content detection, and senescence‐associated β‐galactosidase staining were conducted. RNA sequencing was performed to elucidate underlying signaling pathways, and pharmacological inhibition of the transforming growth factor‐beta pathway was assessed for its therapeutic potential. MFS–hiPSC‐derived ECs recapitulated the pathological features observed in Marfan aortas, particularly pronounced cellular senescence, decreased cell proliferation, and abnormal transforming growth factor‐beta and NF‐κB signaling. These senescent ECs exhibited diminished proliferative and migratory capacities, reduced NO signaling, increased production of inflammatory cytokines, and attenuated responses to inflammatory stimuli. Importantly, senescence and dysfunction in MFS‐hiPSCderived ECs were ameliorated by transforming growth factor‐beta signaling pathway inhibitor, SB‐431542, suggesting a potential therapeutic strategy. Conclusions This study highlights the pivotal role of endothelial cell senescence in the pathogenesis of aortic aneurysms in MFS. Our human induced pluripotent stem cells–based disease model provides new insights into the disease mechanisms and underscores the potential of targeting the transforming growth factor‐beta pathway to mitigate endothelial dysfunction and senescence, offering a promising therapeutic avenue for MFS.

Abstract Background Alzheimer's disease (AD) is a neurodegenerative disorder that affects the central nervous system. Silent information regulator sirtuin 1 (SIRT1) may deacetylate and suppress forkhead box O (FOXO) activities to promote neuronal survival. FOXO1 is involved in the regulation of metabolism, senescence, stress response, and apoptosis. Moreover, endoplasmic reticulum stress (ERS) mediates cell apoptosis. Therefore, this study aimed to determine whether the downregulation of SIRT1 expression exacerbates cognitive dysfunction by activating FOXO1 acetylation and promoting ERS‐mediated apoptosis in amyloid precursor protein/presenilin 1 (APP/PS1) transgenic mice. Methods We used APP/PS1 transgenic mice to construct an in vivo AD model. Additionally, we used β‐amyloid (Aβ)‐incubated HT22 cells and primary neurons (PNs) for in vitro analyses. Cognitive function was assessed using novel object recognition, the Morris water maze, and fear conditioning. Discrepancies between wild‐type (WT) and APP/PS1 transgenic mice were evaluated using an unpaired t test. In addition, one‐way analysis of variance was conducted for behavioral assessments and other tests involving four distinct groups, followed by a Tukey's honestly significant difference test for post hoc pairwise comparisons. Results The expression of SIRT1 was downregulated (in animal experiments, WT mice vs. APP/PS1 mice, n = 3, p = 0.002; in cell experiments, HT22 cells vs. HT22 cells + Aβ1–42, n = 3, p = 0.001; primary neurons vs. primary neurons + Aβ1−42, n = 3, p < 0.001), whereas FOXO1 acetylation was upregulated both in vivo and in vitro (in animal experiments, WT mice vs. APP/PS1 mice, n = 3, p < 0.001; in cell experiments, HT22 cells vs. HT22 cells + Aβ1−42, n = 3, p = 0.004; primary neurons vs. primary neurons + Aβ1–42, n = 3, p < 0.001), leading to cognitive dysfunction, Aβ plaque deposition, and neuronal apoptosis. Quercetin, a SIRT1 agonist, reversed these changes (For SIRT1, APP/PS1 mice vs. Quercetin‐treated APP/PS1 mice, n = 3, p = 0.014; HT22 cells + Aβ1−42 vs. HT22 cells + Aβ1−42 + Quercetin, n = 3, p = 0.003; primary neurons + Aβ1−42 vs. primary neurons + Aβ1−42 + Quercetin, n = 3, p = 0.014. For ac‐FOXO1, APP/PS1 mice vs. Quercetin‐treated APP/PS1 mice, n = 3, p < 0.001; HT22 cells+ Aβ1−42 vs. HT22 cells + Aβ1−42 + Quercetin, n = 3, p = 0.023; primary neurons + Aβ1−42 vs. primary neurons + Aβ1−42 + Quercetin, n = 3, p = 0.003). However, the FOXO1 antagonist AS1842856 invalidated the positive effects of quercetin in APP/PS1 transgenic mice (ac‐FOXO1: Quercetin‐treated APP/PS1 mice vs. AS1842856‐treated APP/PS1 mice, n = 3, p < 0.001). Quercetin counteracted FOXO1 acetylation and ERS‐mediated apoptosis. In contrast, AS1842856 promoted these processes in vivo and in vitro. Conclusion Our findings demonstrate that the downregulation of SIRT1 expression exacerbates cognitive dysfunction by activating FOXO1 acetylation and promoting ERS‐mediated apoptosis.

Bicuspid aortic valve (BAV) is a congenital malformation that predisposes individuals to thoracic aortic aneurysm (TAA), with endothelial dysfunction playing a pivotal role in its pathogenesis. Endothelial cell senescence is a hallmark of endothelial dysfunction, yet direct evidence linking endothelial senescence to BAV-TAA has not been established. In this study, we generated induced pluripotent stem cells (iPSCs) from both BAV-TAA patients and healthy controls, subsequently differentiating them into endothelial cells (iECs). Our findings revealed that BAV-TAA-iECs exhibited senescence phenotype, including impaired proliferation, diminished migratory capacity, upregulated senescence markers (p53, p21, p16), and a pronounced senescence-associated secretory phenotype (SASP). Transcriptomic analysis through RNA sequencing indicated aberrant activation of the FOXO signaling pathway in BAV-TAA-iECs which might contribute to BAV-TAA-iEC senescence. Inhibition of FOXO1 signaling using AS1842856 effectively reversed the senescence phenotype, restored endothelial nitric oxide synthase (eNOS) expression, attenuated SASP cytokine levels, and mitigated inflammation through the p65 and p38 signaling pathways. These findings suggest that endothelial cell senescence plays a critical role in the pathogenesis of BAV-TAA, and targeting FOXO1 signaling may represent a promising therapeutic strategy for BAV-associated aortic diseases.

Endothelial pyroptosis, a pro-inflammatory programmed cell death, promotes endothelial inflammation and is a pivotal process in the initial stage of acute lung injury (ALI). Hydrogen sulfide (H2S), a gasotransmitter primarily dependent on cystathionine γ-lyase (CSE) in the cardiovascular and respiratory systems, plays a protective role during ALI. Nonetheless, the modulatory role and precise molecular mechanism of endothelial endogenous CSE/H2S in the pathogenesis of ALI remain elusive. Herein, we prepared an ALI mouse model using intratracheal administration of LPS (5 mg/kg), and lung injury was assessed by evaluating pulmonary edema, inflammatory response, and endothelial pyroptosis. In this model, H2S production from pulmonary tissues declined in a time-dependent manner, accompanied by a compensatory elevation of CSE protein levels. Treatment with the H2S donor (NaHS) attenuated pulmonary edema, inflammatory cell infiltration, endothelial pyroptosis, and reduced serum levels of tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6). Meanwhile, the inducible deletion of CSE in endothelial cells exacerbated these changes. The blocking effect of CSE/H2S on endothelial pyroptosis (evidenced by caspase-11 activation and GSDMD-NT formation) was also confirmed in cultured pulmonary microvascular endothelial cells (PMECs). Mechanistically, H2S-mediated regulation of sirtuin-1 (SIRT1) expression and activation (via sulfhydration) contributed to the modulatory process. Collectively, we uncovered that endothelial endogenous CSE/H2S alleviates endothelial pyroptosis by activating SIRT1, thereby preventing LPS-induced acute lung injury.

Endothelial pyroptosis, a pro-inflammatory programmed cell death, promotes endothelial inflammation and is a pivotal process in the initial stage of acute lung injury (ALI). Hydrogen sulfide (H2S), a gasotransmitter primarily dependent on cystathionine γ-lyase (CSE) in the cardiovascular and respiratory systems, plays a protective role during ALI. Nonetheless, the modulatory role and precise molecular mechanism of endothelial endogenous CSE/H2S in the pathogenesis of ALI remain elusive. Herein, we prepared an ALI mouse model using intratracheal administration of LPS (5 mg/kg), and lung injury was assessed by evaluating pulmonary edema, inflammatory response, and endothelial pyroptosis. In this model, H2S production from pulmonary tissues declined in a time-dependent manner, accompanied by a compensatory elevation of CSE protein levels. Treatment with the H2S donor (NaHS) attenuated pulmonary edema, inflammatory cell infiltration, endothelial pyroptosis, and reduced serum levels of tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6). Meanwhile, the inducible deletion of CSE in endothelial cells exacerbated these changes. The blocking effect of CSE/H2S on endothelial pyroptosis (evidenced by caspase-11 activation and GSDMD-NT formation) was also confirmed in cultured pulmonary microvascular endothelial cells (PMECs). Mechanistically, H2S-mediated regulation of sirtuin-1 (SIRT1) expression and activation (via sulfhydration) contributed to the modulatory process. Collectively, we uncovered that endothelial endogenous CSE/H2S alleviates endothelial pyroptosis by activating SIRT1, thereby preventing LPS-induced acute lung injury.

BACKGROUND:We have shown that Hippo-YAP signaling pathway plays an important role in endothelial cell differentiation. Vestigial-like family member 4 (VGLL4) has been identified as a YAP inhibitor. However, the exact function of VGLL4 in vascular endothelial cell development remains unclear. In this study, we investigated the role of VGLL4, in human endothelial lineage specification both in 3D vascular organoid and 2D endothelial cell differentiation.METHODS AND RESULTS:In this study, we found that VGLL4 was increased during 3D vascular organoids generation and directed differentiation of human embryonic stem cells H1 towards the endothelial lineage. Using inducible ectopic expression of VGLL4 based on the piggyBac system, we proved that overexpression of VGLL4 in H1 promoted vascular organoids generation and endothelial cells differentiation. In contrast, VGLL4 knockdown (heterozygous knockout) of H1 exhibited inhibitory effects. Using bioinformatics analysis and protein immunoprecipitation, we further found that VGLL4 binds to TEAD1 and facilitates the expression of endothelial master transcription factors, including FLI1, to promote endothelial lineage specification. Moreover, TEAD1 overexpression rescued VGLL4 knockdown-mediated negative effects.CONCLUSIONS:In summary, VGLL4 promotes EC lineage specification both in 3D vascular organoid and 2D EC differentiation from pluripotent stem cell, VGLL4 interacts with TEAD1 and facilitates EC key transcription factor, including FLI1, to enhance EC lineage specification.

本文以西安交通大学、哈尔滨医科大学、温州医科大学为例,探索美育在机能学实验教学课程思政中的应用,提出美育是课程思政建设的重要内涵、目标与切入点.美育融入课程思政建设中是时代要求,也是有效手段.机能学实验教学以课程内容为抓手,以科技发展为载体,以课堂实践为主体,以教学活动为拓展,启迪学生发现科学之美、技术之美、实践之美、生命之美,充分发挥美育在课程思政建设中的重要作用,助力实现立德树人的根本任务.

为了适应移动互联网时代学生的特点,巩固学生的医学机能学知识,训练学生的实验操作技能,培养学生的临床思维能力,该研究通过问卷调查,借助雨课堂,依据赫尔巴特学派的"五段教学法"经典理论,以考一考、听一听、讲一讲、看一看、做一做这五个步骤构建了医学实验机能学课程的精细化教学模式.教学结果显示,雨课堂的混合交互式教学,有助于教师实现医学实验机能学的教学设计由旧有的"粗略定性"向"精准定量"转变,有助于提升学生的课程体验,激发学生的学习兴趣,提高学生的学习效果,促进学生从基础知识到临床思维的转换.

Hydrogen sulfide (H2S), an endogenous gasotransmitter, exhibits the anxiolytic roles through its anti-inflammatory effects, although its underlying mechanisms remain largely elusive. Emerging evidence has documented that cell cycle checkpoint kinase 1 (Chk1)-regulated DNA damage plays an important role in the neurodegenerative diseases; however, there are few relevant reports on the research of Chk1 in neuropsychiatric diseases. Here, we aimed to investigate the regulatory role of H2S on Chk1 in lipopolysaccharide (LPS)-induced anxiety-like behavior focusing on inflammasome activation in the hippocampus. Cystathionine γ-lyase (CSE, a H2S-producing enzyme) knockout (CSE-/-) mice displayed anxiety-like behavior and activation of inflammasome-mediated inflammatory responses, manifesting by the increase levels of interleukin-1β (IL-1β), IL-6, and ionized calcium-binding adaptor molecule-1 (Iba-1, microglia marker) expression in the hippocampus. Importantly, expression of p-Chk1 and γ-H2AX (DNA damage marker) levels were also increased in the hippocampus of CSE-/- mice. LPS treatment decreased the expression of CSE and CBS while increased p-Chk1 and γ-H2AX levels and inflammasome-activated neuroinflammation in the hippocampus of mice. Moreover, p-Chk1 and γ-H2AX protein levels and cellular immunoactivity were significantly increased while CSE and CBS were markedly decreased in cultured BV2 cells followed by LPS treatment. Treatment of mice with GYY4137, a donor of H2S, inhibited LPS-induced increased in p-Chk1 and γ-H2AX levels, mitigated inflammasome activation and inflammatory responses as well as amelioration of anxiety-like behavior. Notably, SB-218078, a selective Chk1 inhibitor treatment attenuated the effect of LPS on inflammasome activation and inflammatory responses and the induction of anxiety-like behavior. Finally, STAT3 knockdown with AAV-STAT3 shRNA alleviated LPS-induced anxiety-like behavior and inhibited inflammasome activation in the hippocampus, and blockade of NLRP3 with MCC950 attenuated neuroinflammation induction and ameliorated LPS-induced anxiety-like behavior. Overall, this study indicates that downregulation of Chk1 activity by H2S activation may be considered as a valid strategy for preventing the progression of LPS-induced anxiety-like behavior.

目的 探讨Pifithrin-α改善异丙肾上腺素(ISO)诱导心肌纤维化的作用及机制.方法 将24只SD雄性大鼠采用信封法随机分为观察组、ISO组和对照组3组,每组8只:观察组和ISO组采用5 mg·kg-1·d-1ISO连续皮下注射1周,观察组再采用2 mg·kg-1·d-1Pifithrin-α腹腔注射1次;对照组连续皮下注射0.9％氯化钠注射液1周.在第4周末时,行超声心动图、血流动力学、心脏重量指数、心肌组织病理学、胶原表达水平和内皮间质转化(EndMT)指标检测.将人微血管内皮细胞分为观察组、转化生长因子-β(TGF-β)组和对照组3组,前两组细胞采用TGF-β刺激,对照组细胞正常培养78 h,检测3组细胞EndMT指标.结果 观察组大鼠左心室舒张末期内径、左心室舒张末期压、左心室重量指数及右心室重量指数低于ISO组,左心室后壁厚度、左心室平均收缩压、左心室压力最大上升速率及左心室压力最大下降速率高于ISO组,差异均有统计学意义(均P＜0.05).观察组大鼠胶原体积分数、Ⅰ型胶原和Ⅲ型胶原表达水平低于ISO组,差异均有统计学意义(均P＜0.05).观察组大鼠心肌组织α-平滑肌肌动蛋白(α-SMA)、波形蛋白和p53表达水平低于ISO组,分化簇31(CD31)表达水平高于ISO组,差异均有统计学意义(均P＜0.05).观察组细胞α-SMA、波形蛋白和p53表达水平低于TGF-β组,CD31表达水平高于TGF-β组,差异均有统计学意义(均P＜0.05).结论 Pifithrin-α可有效减轻ISO诱导的大鼠心肌纤维化和改善心功能,其机制可能与抑制p53介导的EndMT有关.

The Hippo signaling pathway plays a critical role in cardiovascular development and stem cell differentiation. Using microarray profiling, we found that the Hippo pathway components vestigial-like family member 4 (VGLL4) and TEA domain transcription factor 1 (TEAD1) were upregulated during vascular smooth muscle cell (VSMC) differentiation from H1 ESCs (H1 embryonic stem cells). To further explore the role and molecular mechanisms of VGLL4 in regulating VSMC differentiation, we generated a VGLL4-knockdown H1 ESC line (heterozygous knockout) using the CRISPR/Cas9 system and found that VGLL4 knockdown inhibited VSMC specification. In contrast, overexpression of VGLL4 using the PiggyBac transposon system facilitated VSMC differentiation. We confirmed that this effect was mediated via TEAD1 and VGLL4 interaction. In addition, bioinformatics analysis revealed that Ten-eleven-translocation 2 (TET2), a DNA dioxygenase, is a target of TEAD1, and a luciferase assay further verified that TET2 is the target of the VGLL4-TEAD1 complex. Indeed, TET2 overexpression promoted VSMC marker gene expression and countered the VGLL4 knockdown-mediated inhibitory effects on VSMC differentiation. In summary, we revealed a novel role of VGLL4 in promoting VSMC differentiation from hESCs and identified TET2 as a new target of the VGLL4-TEAD1 complex, which may demethylate VSMC marker genes and facilitate VSMC differentiation. This study provides new insights into the VGLL4-TEAD1-TET2 axis in VSMC differentiation and vascular development.

Fibrotic alterations resulting from abnormal tissue repair after lung injury are responsible for the high mortality observed after acute respiratory distress syndrome. Therefore, the prevention and treatment of pulmonary fibrosis has been widely concerned. The Apelin-APJ axis plays an important role in the prevention and treatment of respiratory diseases and organ fibrosis. However, its underlying mechanism remains to be further studied. The aim of this study was to investigate whether the anti-pulmonary fibrosis effect of apelin-APJ axis is related to the activation of angiotensin-converting Enzyme 2 (ACE2). Here, we found that exogenous activation of the Apelin-APJ axis alleviates lipopolysaccharide (LPS)-induced pulmonary fibrosis in mice. In vitro studies revealed that Apelin-13 inhibited LPS-induced endothelial mesenchymal transition in lung microvascular endothelial cells, whereas [Ala13]-Apelin-13 (Apelin-APJ axis inhibitor) accelerated LPS-induced endothelial interstitial transformation in lung microvascular endothelial cells. Notably, angiotensin-converting enzyme 2 (ACE2) inhibitor blocks the beneficial effect of the Apelin-APJ axis activation on LPS-induced pulmonary fibrosis. This finding suggests that the Apelin-APJ axis inhibits pulmonary fibrosis by activating ACE2. Simultaneously, accumulating evidence suggests that ubiquitination may contribute to pulmonary fibrosis. Our study found that LPS increased the ubiquitination of ACE2 protein, whereas Apelin-13 inhibited it. In conclusion, exogenous activation of the Apelin-APJ axis improves LPS-induced pulmonary fibrosis in mice and may be a viable therapeutic target for pulmonary fibrosis.

Recent reports suggested the endoplasmic reticulum stress (ERS)-associated pathway is involved with cognitive impairment in hypoxia condition. ERO1-like protein alpha (Ero1α), an endoplasmic reticulum membrane-bound N-glycoprotein, has been reported to promote oxidative protein folding. However, no studies have reported whether the Ero1α is trapped in hypoxia-induced neuronal loss through the ERS-associated pathways. In our study, this effect of Ero1α was investigated using C57BL/6J mice, the HT22 cells and primary rat neurons. C57BL/6J mice were modeled in a hypoxic chamber for 4 weeks. Behavioral tests were then carried out to test cognitive functions, including the Morris water maze and fear conditioning test. Proteomics showed that Ero1α distinctly upregulated compared with normoxia group and verified using western blotting. Flow cytometry and immunofluorescence were used to analyze the neuroprotective effect of inhibitor EN460 of Ero1α in the HT22 cells. In C57BL/6J mice, hypoxia significantly caused cognitive decline. Brain slice staining results were also used to confirm this effect. Western blot analysis demonstrated that Ero1α, ERS-associated proteins and apoptosis-associated proteins significantly increased in the hypoxia treated groups, further proliferation-related marker protein decreased. EN460, a selective endoplasmic reticulum oxidation 1 (ERO1) inhibitor, counteracted neuronal apoptosis and ameliorated neuronal cell proliferation in the HT22 cells. Taken together, our data indicate that hypoxia induces cognitive impairment, at least in part, by upregulating Ero1α which contributes to neuronal apoptosis through ERS signaling pathway, providing preliminary experimental evidence that the Ero1α is a promising therapeutic target in hypoxia-induced cognitive deficits.

Endothelial cells (ECs) derived from pluripotent stem cells (PSCs) provide great resource for vascular disease modeling and cell-based regeneration therapy. However, the molecular mechanisms of EC differentiation are not completely understood. In this study, we checked transcriptional profile by microarray and found Hippo pathway is changed and the activity of YAP decreased during mesoderm-mediated EC differentiation from human embryonic stem cells (hESCs). Knockdown of YAP in hESCs promoted both mesoderm and EC differentiation indicating by mesodermal- or EC-specific marker gene expression increased both in mRNA and protein level. In contrast, overexpression of YAP inhibited mesoderm and EC differentiation. Microarray data showed that several key transcription factors of EC differentiation, such as FLI1, ERG, SOX17 are upregulated. Interestingly, knockdown YAP enhanced the expression of these master transcription factors. Bioinformation analysis revealed that TEAD, a YAP binds transcription factors, might regulate the expression of EC master TFs, including FLI1. Luciferase assay confirmed that YAP binds to TEAD1, which would inhibit FLI1 expression. Finally, FLI1 overexpression rescued the effects of YAP overexpression-mediated inhibition of EC differentiation. In conclusion, we revealed the inhibitory effects of YAP on EC differentiation from PSCs, and YAP inhibition might promote expression of master TFs FLI1 for EC commitment through interacting with TEAD1, which might provide an idea for EC differentiation and vascular regeneration via manipulating YAP signaling.