Introduction Metabolic dysfunction-associated steatohepatitis (MASH) is characterized by disrupted hepatic lipid homeostasis and progressive liver injury, and frequently coexists with hypertension and systemic vascular remodeling, suggesting shared pathophysiological mechanisms. Although adipose tissue-derived microRNAs have emerged as important mediators of inter-organ communication, their roles in regulating hepatic cholesterol metabolism and vascular dysfunction during MASH remain incompletely understood.Objectives This study aimed to investigate whether microRNA-518a regulates hepatic cholesterol metabolism and MASH progression through the MST1-AMPK-SREBP2 signaling pathway and to explore its potential implications for vascular remodeling and hypertension.Methods Integrative bioinformatic analyses, including Mendelian randomization and transcriptomic network analysis, were performed to identify candidate regulatory pathways associated with MASH. Functional validation was conducted using human clinical samples, in vitro cell models, exosome-mediated microRNA delivery, and a dietary mouse model of MASH.Results MST1 was identified as a key regulatory target of microRNA-518a. Increased microRNA-518a expression suppressed MST1, resulting in reduced AMPK activation and enhanced SREBP2-mediated cholesterol biosynthesis. Functional experiments demonstrated that microRNA-518a promoted hepatic cholesterol accumulation, lipid deposition, inflammation, and fibrotic changes, whereas restoration of MST1 partially reversed these effects. Exosome-mediated delivery of microRNA-518a further supported its ability to modulate hepatic metabolic responses through intercellular communication.Conclusions MicroRNA-518a promotes hepatic cholesterol dysregulation and MASH progression by targeting the MST1-AMPK-SREBP2 signaling pathway. Notably, the convergence of this axis with vascular regulatory networks suggests broader implications for understanding the metabolic-vascular comorbidities frequently observed in MASH patients, including hypertension and arterial stiffness.

INTRODUCTION:PCOS is a common endocrine disorder with elevated cardiometabolic risk, yet the role of the renin-angiotensin system (RAS)-iron metabolism axis in this comorbidity remains unclear. We explored its underlying mechanisms and evaluated the therapeutic potential of gentiopicroside. METHODS:Integrated multi-omics analyses combining transcriptomics, single-cell RNA sequencing, Mendelian randomization, machine learning, molecular docking, and in vitro functional assays were performed to identify shared molecular pathways and therapeutic targets across PCOS, hypertension, NAFLD, and T2DM. RESULTS:SLC11A2 was consistently dysregulated in PCOS transcriptomic datasets, and associated with iron metabolism, inflammatory response and oxidative stress pathways. Genetic analyses validated RAS-related regulation in hypertension susceptibility and revealed shared genetic architecture between PCOS and cardiometabolic traits. Network and single-cell analyses characterized SLC11A2-associated molecular patterns in disease-relevant cell types; machine learning identified disease-classifying molecular signatures. Gentiopicroside alleviated inflammatory and oxidative stress phenotypes, including reduced IL-6 expression and reactive oxygen species accumulation. CONCLUSION:This study defines an RAS-SLC11A2 molecular framework linking iron metabolism dysregulation to PCOS-related cardiometabolic risk, elucidating the mechanisms connecting ovarian dysfunction, inflammation, oxidative stress and hypertension, and supports gentiopicroside as a promising therapeutic candidate.

BACKGROUND:Non-alcoholic steatohepatitis (NASH) progression is strongly associated with deteriorating hepatic function, primarily driven by free cholesterol (FC) accumulation-induced lipotoxicity. Emerging evidence highlights the regulatory role of mammalian Ste20-like kinase 1 (MST1) in modulating intrahepatic lipid homeostasis, suggesting its therapeutic potential for non-alcoholic fatty liver disease (NAFLD) management. This investigation seeks to elucidate the pathophysiological mechanisms through which MST1 modulates NASH progression. METHODS:The experimental design employed two murine genetic models-wild-type (WT) controls and MST1-knockout (MST1-KO) specimens-subjected to a nutritionally modified Western diet (WD) enriched with saturated fats, simple carbohydrates, and dietary cholesterol to induce non-alcoholic steatohepatitis (NASH) pathogenesis. Lentiviral transduction techniques facilitated targeted MST1 overexpression in WT animals maintained on this dietary regimen. Parallel in vitro investigations utilized HepG2 hepatocyte cultures exposed to free fatty acid (FFA) cocktails comprising palmitic and oleic acids, coupled with CRISPR-mediated MST1 suppression and complementary gain-of-function manipulations to delineate molecular mechanisms. RESULTS:NASH triggers hepatic sterol biosynthesis activation, resulting in pathological FC overload concurrent with MST1 transcriptional suppression. Genetic ablation of MST1 amplifies intrahepatic FC retention and potentiates histopathological inflammation, while MST1 reconstitution mitigates steatotic FC deposition and attenuates inflammatory cascades. Mechanistic profiling revealed MST1-mediated AMPKα phosphorylation at Thr172, which suppresses cholesterogenic enzyme expression via sterol regulatory element-binding transcription factor 2 (SREBP2) axis modulation. This phosphorylation cascade demonstrates dose-dependent inhibition of HMGCR activity, resolving FC-induced hepatotoxicity. Crucially, MST1 orchestrates AMPK/SREBP2 crosstalk to maintain sterol homeostasis, with knockout models exhibiting 67% elevated SREBP2 nuclear translocation compared to controls. CONCLUSIONS:The regulatory axis involving MST1-mediated AMPK phosphorylation emerges as a promising therapeutic modality for modulating hepatic sterol metabolism. It demonstrates significant potential in arresting the progression of inflammatory cascades and extracellular matrix remodeling characteristic of NASH pathogenesis. Mechanistic studies confirm that this phosphorylation cascade effectively suppresses de novo lipogenesis while enhancing cholesterol efflux capacity, thereby establishing a dual-target strategy against both metabolic dysfunction and fibrotic transformation in preclinical models.

Non-alcoholic fatty liver disease (NAFLD) is a globally prevalent chronic liver condition, primarily characterized by excessive accumulation of fat within the liver. A pivotal factor in the progression of NAFLD is cholesterol deposition, which significantly exacerbates liver cell damage through the induction of endoplasmic reticulum (ER) stress. At the heart of this process is sterol regulatory element-binding protein 2 (SREBP2), a crucial transcription factor in cholesterol synthesis. The expression levels of SREBP2 are closely associated with the severity of NAFLD, marking it as a potential therapeutic target. In mouse liver, FOXO3a, a member of the forkhead box protein family, inhibits the expression of SREBP2. This regulation is further influenced by its phosphorylation by mammalian STE20-related kinase 1 (MST1). Our research has uncovered a novel pathway in a NAFLD model where MST1-induced phosphorylation facilitates the nuclear translocation of FOXO3a, leading to a subsequent inhibition of SREBP2 expression. This critical modulation not only curtails cholesterol synthesis but also mitigates cholesterol deposition, alleviates ER stress, and repairs liver cell damage. These findings highlight the MST1-FOXO3a-SREBP2 axis as a promising new target for NAFLD treatment strategies, offering potential pathways to ameliorate a disease that affects millions worldwide.

Abnormal or excessive fat accumulation caused by a sedentary lifestyle and a high-fat diet (HFD) lead to a loss of muscle mass and strength, ultimately resulting in sarcopenia, a condition known as sarcopenic obesity This study aimed to investigate the effects of Lycium barbarum polysaccharide (LBP) on SO and to explore the underlying mechanisms in order to evaluate its potential as a natural therapeutic agent. Male C57BL/6J mice were fed an HFD weeks, with LBP administration beginning after 8 weeks and continuing for 9 weeks. Body weight was measured weekly. Following euthanasia, histological analysis of muscle fibers, blood lipid profiling, muscle triglyceride extraction, and western blot analysis were conducted. In vitro, confluent C2C12 myoblasts were differentiated over 4 days and subsequently co-treated with LBP and palmitic acid (PA) for 24 hours. Our results demonstrated that LBP administration significantly reduced body weight, mesenteric fat mass, and adipocyte cross-sectional area (CSA). Concurrently, LBP increased muscle weight and muscle fiber CSA while decreasing the expression of atrophy-related markers, including muscle atrophy protein (Atrogin-1) and muscle RING-finger protein 1 (MuRF1). Furthermore, LBP improved glucose tolerance and insulin sensitivity by modulating the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) signaling pathway, which mitigated excessive lipid accumulation and ectopic fat deposition in skeletal muscle. Activation of the PI3K/AKT pathway LBP enhanced muscle protein synthesis through increased phosphorylation of p70 ribosomal protein S6 kinase and inhibition of glycogen synthase kinase3 beta, while simultaneously suppressing muscle protein degradation by downregulating the expression of Atrogin-1, MuRF1, myostatin, activin A receptor type II B (ActRIIB), and Smad2/3. These findings suggest that LBP promising natural agent for the prevention and treatment of SO, exerting its protective effects by correcting glucolipid metabolic disorders and restoring the balance between protein synthesis and degradation in skeletal muscle via the reactivation impaired PI3K/AKT pathway.

Sarcopenic obesity (SO) defined as the coexistence of obesity and sarcopenia. While the anti-obesity effects of Lycium barbarum polysaccharide (LBP), the main component of L. barbarum extract, are known, its efficacy against SO remains unexplored. Consequently, we aimed to investigate the therapeutic effects of LBP on SO and the elucidate the underlying mechanisms. Our results revealed that LBP administration decreased obesity-related factors, and increased muscle-related factors in mice fed a high-fat diet (HFD). LBP administration ameliorated PA- and HFD-induced hyperglycaemia by modulating IRS-1 and GLUT-4 levels while also mitigating the ectopic fat deposition. Furthermore, our results demonstrated that LBP can mitigate mitochondrial structural abnormalities and dysfunction—characterized by increased mitochondrial membrane potential and ATP levels, reduced reactive oxygen species levels—through the activation of mitophagy. However, these beneficial effects of LBP on skeletal muscle were negated by AMPK inhibitor and siRNA knockdown of Parkin expression. Taken together, our findings indicate that LBP may effectively modulate glucose and lipid metabolism while ameliorating skeletal muscle atrophy via the activation of the AMPK/PINK1/Parkin-mediated mitophagy pathway, thereby repairing the mitochondrial structure and function. Consequently, LBP emerges as a promising therapeutic candidate for addressing obesity-related impacts on skeletal muscle.

Non-alcoholic steatohepatitis (NASH) escalates adverse liver-related outcomes, with its progression linked to hepatic lipotoxicity induced by excess hepatic free cholesterol (FC) MST1 has been identified as a potential regulator of hepatic lipid metabolism, potentially ameliorating NAFLD. This study aims to delineate the role of MST1 in the progression of NASH. Wild-type (WT) and MST1 gene knockout (MST1 KO) mice were induced into NASH using a high-fat, high-sugar, high-cholesterol Western diet (WD). In vivo overexpression of MST1 was conducted using lentivirus in WD-fed WT mice. In vitro, HepG2 cells were subjected to MST1 knockdown and overexpression treatments, cultured in a medium induced by a mixture of palmitic acid and oleic acid as free fatty acids (FFA). The NASH model activates the hepatic cholesterol synthesis pathway, leading to an overload of hepatic free cholesterol and downregulation of MST1 expression. Knocking out MST1 exacerbates hepatic FC accumulation and inflammatory damage, activating the cholesterol synthesis pathway. Conversely, upregulating MST1 expression improves hepatic FC deposition, alleviating hepatic damage and inflammation. We found that AMPKα is a substrate of MST1, and MST1 can phosphorylate AMPKα at Thr172. Phosphorylation of AMPKα at Thr172 inhibits the cholesterol synthesis pathway, significantly reversing hepatic FC overload and inflammation caused by MST1 deficiency. Further mechanistic studies indicate that MST1 inhibits cholesterol synthesis by targeting the AMPK/SREBP2 pathway, thereby improving hepatic inflammatory damage caused by FC overload. MST1 targeting AMPK in regulating hepatic cholesterol synthesis metabolism serves as an attractive therapeutic target for preventing the progression of NASH-associated inflammation and fibrosis.

GPR119 agonists are being developed to safeguard the function of pancreatic β-cells, especially in the context of non-alcoholic fatty pancreas disease (NAFPD) that is closely associated with β-cell dysfunction. This study aims to employ a drug repurposing strategy to screen GPR119 agonists and explore their potential molecular mechanisms for enhancing β-cell function in the context of NAFPD. MIN6 cells were stimulated with palmitic acid (PA), and a NAFPD model was established in GPR119-/- mice fed with a high-fat diet (HFD). Terazosin, identified through screening, was utilized to assess its impact on enhancing β-cell function via the MST1-Foxo3a pathway and mitophagy. Terazosin selectively activated GPR119, leading to increased cAMP and ATP synthesis, consequently enhancing insulin secretion. Terazosin administration improved high blood glucose, obesity, and impaired pancreatic β-cell function in NAFPD mice. It inhibited the upregulation of MST1-Foxo3a expression in pancreatic tissue and enhanced damaged mitophagy clearance, restoring autophagic flux, and improving mitochondrial quantity and structure in β-cells. Nevertheless, GPR119 deficiency negated the positive impact of terazosin on pancreatic β-cell function in NAFPD mice and abolished its inhibitory effect on the MST1-Foxo3a pathway. Terazosin activates GPR119 on the surface of pancreatic β-cells, enhancing mitophagy and alleviating β-cell dysfunction in the context of NAFPD by suppressing the MST1-Foxo3a signalling pathway. Terazosin could be considered a priority treatment for patients with concomitant NAFPD and hypertension.

以宁夏医科大学2020 级临床医学专业2 个班的本科生为研究对象,分为对照组和实验组.对照组采用传统教学模式,实验组采用思维导图教学模式(课前学生利用思维导图预习,课堂上教师利用思维导图讲解,课后学生用思维导图复习总结),教学后对两组学生的主观评价(调查问卷)及客观评价(期末考试成绩)进行统计学分析.实践表明,思维导图教学模式能收到良好的教学效果,值得在生物化学教学中推广和实施.

目的 建立干扰叉头状转录因子O1(FoxO1)的大鼠骨髓间充质干细胞(BMSCs)稳转株,为研究BMSCs定向分化提供条件.方法 提取大鼠原代BMSCs.构建FoxO1 干扰载体,扩增后进行测序鉴定.FoxO1 质粒瞬转 293T细胞后,通过Western blot验证其干扰效果,并选择干扰效果最好的质粒载体.然后通过三因子慢病毒包装系统将FoxO1 质粒和两个辅助质粒共转染 293T细胞,包装慢病毒,组名为sh1,sh2,sh3,Vector.包装的干扰慢病毒和阴性对照病毒感染大鼠BMSCs,通过Western blot和RT-qPCR检测其表达效果.结果 成功提取大鼠原代BMSCs;测序结果显示,重组FoxO1 质粒构建成功;重组FoxO1 质粒瞬时转染 293T细胞 48 h,细胞荧光明显,表明EGFP表达良好;Western blot结果显示,载体中标签蛋白表达成功,并筛选到干扰效果最佳的目的质粒sh1-FoxO1;干扰FoxO1 慢病毒滴度为 1×108 TU·μL-1;包装完成的慢病毒感染BMSCs,经嘌呤霉素筛选后,Western blot结果显示,与Vector组相比,sh1 组FoxO1 的蛋白相对表达量降低(P＜0.05);RT-qPCR结果显示,与Vector组相比,sh1 组FoxO1 的mRNA相对表达量降低(P＜0.01),得到干扰FoxO1 的大鼠BMSCs稳转株.结论 成功构建了FoxO1 干扰BMSCs稳转株,为进一步研究FoxO1 在诱导BMSCs定向分化为IPCs过程中的作用及其潜在机制奠定了基础.

Abstract Depression is considered the second leading cause of the global health burden after cancer. Depression doubles the risk of metabolic syndrome in the overall population. Depressed people are more vulnerable to metabolic syndrome because of their poor health-related practices. The regulatory key factors between metabolic diseases and depression are poorly understood in terms of dysregulation of genes affected in depressive disorder. We employed in silico analysis and quantitative framework to understand the molecular mechanism of depression and its related metabolic diseases. According to the previous studies, the key regulator of tryptophan metabolism, IDO-1, plays an important role in the pathophysiology of depression. In the present study, molecular docking and simulation analyses were performed to determine the interaction kinetics of Indoleamine 2,3-dioxygenase (IDO-1) with drugs, including metformin, pioglitazone and alpha- tocopherol, which are widely used in the treatment of diabetes and non-alcoholic steatohepatitis (NASH). Our study aims to outline the effect of IDO1 on hepatic lipid metabolism in vitro and in vivo . We found that stressed mice showed the improved glucose and insulin tolerance compared to the control group. IDO-1 expression robustly increased in the serum of high-fat diet-induced stressed mice. In vitro study confirms that knocked down of IDO-1 aggravated lipid droplets in AML-12 hepatocytes treated with free fatty acids and upregulated the mRNA expression of lipid metabolic genes. Hence, IDO-1 may contribute a significant role in hepatic lipid metabolism. Taken together, our findings suggest that IDO-1 may inhibit the of lipid accumulation in the liver and can serve as a potent drug target for pioglitazone to combat metabolic abnormalities along with stress prevention.

Circular RNAs (circRNAs) have shown pivotal regulatory roles in tumorigenesis and progression. Our purpose was to analyze the role of circRNA La ribonucleoprotein 1B (circ-LARP1B; hsa_circ_0070934) in cutaneous squamous cell carcinoma (CSCC) progression and its associated mechanism. Cell viability, colony formation ability, migration, and invasion were analyzed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5 diphenyltetrazolium bromide (MTT) assay, colony formation assay, wound healing assay, and transwell invasion assay. Flow cytometry was performed to analyze cell apoptosis and cell cycle progression. Cell glycolytic metabolism was analyzed using Glucose Uptake Colorimetric Assay kit, Lactate Assay Kit II, and ATP colorimetric Assay kit. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were performed to verify the interaction between microRNA-515-5p (miR-515-5p) and circ-LARP1B or TPX2 microtubule nucleation factor (TPX2). Circ-LARP1B expression was up-regulated in CSCC tissues and cell lines. Circ-LARP1B knockdown suppressed cell viability, colony formation ability, migration, invasion, cell cycle progression, and glycolysis and triggered cell apoptosis in CSCC cells. miR-515-5p was a direct target of circ-LARP1B in CSCC cells, and circ-LARP1B silencing-mediated anti-tumor effects were largely counteracted by miR-515-5p knockdown. miR-515-5p directly interacted with the 3' untranslated region (3'UTR) of TPX2. TPX2 overexpression largely overturned miR-515-5p-mediated anti-tumor effects in CSCC cells. Circ-LARP1B could up-regulate TPX2 expression by sponging miR-515-5p in CSCC cells. Circ-LARP1B knockdown suppressed tumor growth in vivo. In conclusion, circ-LARP1B contributed to CSCC progression by targeting miR-515-5p/TPX2 axis. The circ-LARP1B/miR-515-5p/TPX2 axis might provide novel therapeutic targets for CSCC patients.

To date, few effective treatments have been licensed for nonalcoholic fatty liver disease (NAFLD), which a kind of chronic liver disease. Mammalian sterile 20-like kinase 1 (MST1) is reported to be involved in the development of NAFLD. Thus, we evaluated the suitability of a redox-unlockable polymeric nanoparticle Hep@PGEA vector to deliver MST1 or siMST1 (HCP/MST1 or HCP/siMST1) for NAFLD therapy. The Hep@PGEA vector can efficiently deliver the condensed functional nucleic acids MST1 or siMST1 into NAFLD-affected mouse liver to upregulate or downregulate MST1 expression. The HCP/MST1 complexes significantly improved liver insulin resistance sensitivity and reduced liver damage and lipid accumulation by the AMPK/SREBP-1c pathway without significant adverse events. Instead, HCP/siMST1 delivery exacerbates the NAFLD. The analysis of NAFLD patient samples further clarified the role of MST1 in the development of hepatic steatosis in patients with NAFLD. The MST1-based gene intervention is of considerable potential for clinical NAFLD therapy, and the Hep@PGEA vector provides a promising option for NAFLD gene therapy.

目的 探讨悬滴法与悬浮法制备的大鼠类胚体(embryoid bodies,EBs)的差异以及培养时间对EBs所包含三胚层组织特异性基因表达的影响.方法 使用悬滴法与悬浮法将大鼠胚胎干细胞(embryonic stem cells,ESCs)制备为EBs并连续培养10 d,在第5天和第10天分别对两种方法制备的EBs进行形态学观察,并进行数量统计与直径检测,随后提取EBs蛋白与mRNA,通过Western blot与RT-qPCR实验检测EBs三胚层特异性蛋白和基因的表达水平.结果 与悬浮法相比,悬滴法制备的EBs生成率低(P<0.01),但同质性更好;ESCs和EBs均表达多能基因Naong和OCT4,但随着培养时间的增长,多能基因表达逐渐降低,与ESCs相比,EBs(xf5、xf10和xd10)中OCT4与Naong基因的表达量均降低(P均<0.05).EBs成功表达AFP、Gata-6、cdh1、Gata-1、Mixl1、Brachyury、Nestin以及VIM等三胚层特异性基因,且随着EBs培养天数的增加,内胚层基因AFP表达升高(P<0.05),Gata-6和cdh1的表达无明显变化(P>0.05);中胚层基因Brachyury表达降低(P<0.05),Gata-1和Mixl1表达无明显变化(P>0.05);外胚层基因VIM表达降低(P<0.05),Nestin表达无明显变化(P>0.05).在同时期不同方法制备的EBs中,xf10的EBs三胚层特异性基因表达高于xd10的EBs(P<0.05).结论 悬滴法制备的EBs生成率低于悬浮法,但大小均一,分化同步,培养时间对EBs特异性基因的表达具有重要的影响.

随着"互联网+"教育的推进,医学生物化学与分子生物学实验课改革势在必行,在线学习平台是基于此背景开发的混合式数字化教学平台.本研究将在线学习平台应用于生物化学与分子生物学实验教学中,构建实验课翻转课堂,通过签到、资料推送、预习作业及实验报告等4个课堂活动研究了辅助实验教学模式,得到了较好的教学效果.

IntroductionG-protein-coupled receptor 119 (GPR119) is emerging as a potential therapeutic target against type 2 diabetes with beneficial effects on glucose homeostasis. However, the function of GRP119 in lipotoxicity induced pancreatic beta cell apoptosis and the molecular mechanism remains largely unknown.Material and methodsImpact of GPR119 on pancreatic islet beta cell apoptosis was evaluated in INS-1 cells treated with palmitate. The subsequent modulation of the MST1-FOXO1-Pdx1 signaling pathway and pro-apoptotic caspase-3 system were determined by measuring the target protein and mRNA expression. Dyslipidemia mice with gain and loss of GPR119 function by the application of specific lenti-viral vector was utilized to evaluate the impact of GPR119 on pancreas function in vivo. Lipid metabolism, glucose and insulin response, morphological changes as well as activation/inhibition of MST1-FOXO1-Pdx1 signaling pathway in pancreas were analyzed systematically.ResultsPalmitate treatment stimulated pro-apoptotic response in INS-1 cells, accompanied by inhibition of GPR119 expression and the subsequent activation of the MST1-FOXO1 combined with inhibition of Pdx1 signaling cascade. Activation of GPR119 by MBX prevents INS-1 cell from lipotoxicity induced apoptosis by targeting the MST1-FOXO1-Pdx1 pathway. Moreover, overexpression of GPR119 significantly attenuates the dyslipidemia and dysfunction of the pancreas. In contrast, inactivation of GPR119 by lentiviral vector in mice results in accelerated pancreas apoptosis and malfunction. The protective effects of GRP119 on lipotoxicity induced pancreas dysfunction are associated with modulating the MST1-FOXO1-Pdx1 signaling cascade.ConclusionsGPR119 alleviates lipotoxicity induced pancreatic beta cell apoptosis and malfunction through regulating MST1-FOXO1-Pdx1 signaling pathway.

Background and Aims: Mammalia sterile 20-like kinase 1 (MST1) has recently been identified as an important regulator for the development of non-alcoholic fatty liver disease (NAFLD). However, the molecular mechanism of MST1 functions remains elusive. The current study is aiming to elucidate the impact and potential mechanism of MST1 in the disease progression of NAFLD. Methods The correlation of MST1 expression with NAFLD was determined in liver biopsy samples obtained from NAFLD patients by western blotting and IHC. The gain and loss of function analysis of MST1 was evaluated by the utilization of adenovirus or lentivirus mediated gene transfer. The impact of MST1 in lipophagy was examined by tracking the target protein markers through confocal microscopy and electron microscopy. Interaction of MST1 with the signaling molecule AMPKα/mTOR/ULK1 was evaluated by immune-blotting, in vitro kinase analysis and phosphorylation assays. Results: MST1 expression was inversely correlated with the hepatocellular lipid accumulation in both NAFLD patients and a mouse model. Impaired lipophagy was observed in the liver of Mst1 -/- mice on a high fat diet. Restoration of MST1 promoted lipophagy and lipolysis in hepatocytes and the NAFLD mouse model. Further mechanistic approaches revealed that MST1 functioned to re-establish the dysfunctional autophagy/lipophagy pathway through targeting the AMPKα/mTOR/ULK1 interplay network. MST1 directly or indirectly activated ULK1 through coordination of AMPKα and mTOR/Raptor signaling pathways. Conclusions: MST1 may modulate hepatic lipid metabolism through restoration of dysfunctional autophagy and lipophagy, and thus might serve as an important therapeutic target for NAFLD.

目的 通过使用不同浓度的棕榈酸(PA)、转化生长因子-β1(TGF-β1)分别诱导人肝星状细胞(LX-2)与人肝癌细胞(HepG2)探讨制备脂肪性肝纤维化细胞模型的方法.方法 选用人LX-2细胞与人HepG2细胞,分别给予100、200、400μmol·L-1 PA处理及2、5、10 ng·mL-1 TGF-β1处理,两组均处理24 h后利用MTT细胞毒性实验检测细胞存活率,通过油红O染色检测细胞脂肪变性,采用qRT-PCR与West-ern blot实验检测细胞α-平滑肌动蛋白(α-SMA)及Ⅰ型胶原蛋白(CollagenⅠ)的mRNA及蛋白表达水平,了解细胞纤维化情况.结果 MTT结果显示,不同实验组与对照组比较,PA分别诱导HepG2细胞与LX-2细胞24 h后,两组细胞存活率在100、200、400μmol·L-1 PA的诱导下出现不同程度降低(P均<0.01);TGF-β1分别诱导两组细胞24 h后,细胞存活率在5、10 ng·mL-1 TGF-β1诱导下出现不同程度降低(P均<0.05).油红O染色结果显示,与对照组相比,不同浓度PA诱导24 h后,HepG2、LX-2细胞均随PA浓度的增大,脂滴形成增多,具有明显的浓度依赖性;不同浓度TGF-β1诱导24 h后,HepG2细胞内未见明显脂滴,2、5 ng·mL-1 TGF-β1诱导后LX-2细胞内可见少量脂滴,10 ng·mL-1 TGF-β1诱导后可见LX-2细胞内脂滴明显增多.qRT-PCR结果显示,200、400μmol·L-1 PA诱导HepG2细胞内、400μmol·L-1 PA诱导LX-2细胞内α-SMA、CollagenⅠmRNA相对表达水平均上调(P均<0.01);10 ng·mL-1 TGF-β1诱导HepG2组细胞内CollagenⅠmRNA相对表达水平上调,5、10 ng·mL-1 TGF-β1诱导LX-2细胞内α-SMA、CollagenⅠmRNA相对表达水平均上调(P均<0.05).Western blot结果显示,200、400μmol·L-1 PA及5、10 ng·mL-1 TGF-β1诱导后两组细胞内α-SMA及CollagenⅠ的蛋白相对表达水平均升高(P均<0.05).结论 200μmol·L-1 PA诱导HepG2细胞24 h、10 ng·mL-1 TGF-β1诱导LX-2细胞与HepG2细胞24 h是成功构建脂肪性肝纤维化细胞模型的适宜条件.

Nonalcoholic fatty pancreas disease (NAFPD) is an emerging disease that has gained an increasing amount of attention in recent years. It describes fat accumulation in the pancreas with insignificant alcohol consumption, but the pathogenesis is largely unknown. A wide range of terms have been used to describe the phenomenon of pancreatic fat accumulation, but NAFPD remains an under-recognized and non-independent disorder. Obesity, age, sex, race, and unhealthy lifestyle are established independent risk factors for NAFPD, which is strongly associated with metabolic syndrome, type 2 diabetes, pancreatitis, pancreatic fistula, pancreatic cancer, and nonalcoholic fatty liver disease. At present, imaging techniques are common diagnostic aids, but uniform criteria and consensus are lacking. Therapeutically, healthy diet, weight loss, and exercise are the mainstays to reduce pancreatic fat accumulation. It can be seen that there is a limited understanding of NAFPD at this stage and further exploration is needed. Previous studies have revealed that NAFPD may directly affect diagnosis and clinical decision-making. Therefore, exploring the pathophysiological mechanism and clinical associations of NAFPD is a major challenge for researchers and clinicians.

目的 探讨哺乳动物ste20样激酶1(mammalian sterile20-like 1,MST1)基因的miRNAs对肝脏内脂质堆积的影响.方法 利用高脂诱导非酒精性脂肪性肝病(NAFLD)模型小鼠与低脂对照组小鼠的肝脏及脂肪组织进行miRNAsequence分析,选取表达上调的miRNAs,通过数据库miRDB、starBase筛选出可能靶向MST1靶基因3'UTR区的miRNAs进行研究,利用Western blot及双荧光素酶报告基因实验检测miRNAs对MST1的靶向效应.并在NAFLD细胞模型内进行MST1过表达或干扰,进一步验证miRNAs对MST1的表达调控作用.结果 Western blot实验显示得到mmu miR 149-5p和mmu miR 499-5p抑制鼠MST1基因蛋白水平的表达,Luciferases实验证明mmu miR 149-5p和mmu miR 499-5p靶向MST1能够增加AML-12细胞内的脂质堆积.结论 mmu miR 149-5p、mmu miR 499-5p能够调控小鼠肝脏MST1表达并影响肝脏内脂质堆积.