Wnt signaling pathway inhibitor promotes mesenchymal stem cells differentiation into cardiac progenitor cells in vitro and improves cardiomyopathy in vivo

BACKGROUND Cardiovascular diseases particularly myocardial infarction(MI)are the leading cause of mortality and morbidity around the globe.As cardiac tissue possesses very limited regeneration potential,therefore use of a potent small molecule,inhibitor Wnt production-4(IWP-4)for stem cell differentiation into cardiomyocytes could be a promising approach for cardiac regeneration.Wnt pathway inhibitors may help stem cells in their fate determination towards cardiomyogenic lineage and provide better homing and survival of cells in vivo.Mesenchymal stem cells(MSCs)derived from the human umbilical cord have the potential to regenerate cardiac tissue,as they are easy to isolate and possess multilineage differentiation capability.IWP-4 may promote the differentiation of MSCs into the cardiac lineage.AIM To evaluate the cardiac differentiation ability of IWP-4 and its subsequent in vivo effects.METHODS Umbilical cord tissue of human origin was utilized to isolate the MSCs which were characterized by their morphology,immunophenotyping of surface markers specific to MSCs,as well as by tri-lineage differentiation capability.Cytotoxicity analysis was performed to identify the optimal concentration of IWP-4.MSCs were treated with 5μM IWP-4 at two different time intervals.Differentiation of MSCs into cardiomyocytes was evaluated at DNA and protein levels.The MI ratmodel was developed.IWP-4 treated as well as untreated MSCs were implanted in the MI model,then the cardiac function was analyzed via echocardiography.MSCs were labeled with 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(DiI)dye for tracking,while the regeneration of infarcted myocardium was examined by histology and immunohistochemistry.RESULTS MSCs were isolated and characterized.Cytotoxicity analysis showed that IWP-4 was non-cytotoxic at 5μM concentration.Cardiac specific gene and protein expression analyses exhibited more remarkable results in fourteen days treated group that was eventually selected for in vivo transplantation.Cardiac function was restored in the IWP-4 treated group in comparison to the MI group.Immunohistochemical analysis confirmed the homing of pre-differentiated MSCs that were labeled with DiI cell labeling dye.Histological analysis confirmed the significant reduction in fibrotic area,and improved left ventricular wall thickness in IWP-4 treated MSC group.CONCLUSION Treatment of MSCs with IWP-4 inhibits Wnt pathway and promotes cardiac differentiation.These pre-conditioned MSCs transplanted in vivo improved cardiac function by cell homing,survival,and differentiation at the infarcted region,increased left ventricular wall thickness,and reduced infarct size.

血清胸苷激酶1、细胞角质蛋白19片段抗原21-1及dickkopfWNT信号通路抑制剂1对食管癌的诊断价值

目的探讨血清胸苷激酶1(TK1)、细胞角质蛋白19片段抗原21-1(CYFRA21-1)及dickkopf WNT信号通路抑制剂1(DKK1)对食管癌的诊断价值。方法选取62例食管癌患者作为食管癌组,59例健康体检者作为对照组。对比两组受试者TK1、CYFRA21-1、DKK1水平,对比不同分期食管癌患者TK1、CYFRA21-1、DKK1水平,分析TK1、CYFRA21-1、DKK1单独及联合检测对食管癌的诊断价值。结果食管癌组患者TK1、CYFRA21-1、DKK1水平均明显高于对照组,差异均有统计学意义(P﹤0.01)。Ⅲ~Ⅳ期食管癌患者TK1、CYFRA21-1、DKK1水平均明显高于Ⅰ~Ⅱ期患者,差异均有统计学意义(P﹤0.01)。TK1+CYFRA21-1+DKK1联合检测诊断食管癌的灵敏度和特异度分别为88.50%、79.10%,曲线下面积为0.779(95%CI:0.695~0.864),均高于三者单独检测。结论TK1、CYFRA21-1、DKK1联合检测有利于提高食管癌的诊断效能,防止误诊漏诊,三者对其病情评估具有一定参考价值。

Inhibition of Melanoma Cell Proliferation by Targeting Wnt/β-catenin Pathway through Sox4 RNA Interference

The effect of siRNA-mediated Sox4 gene silencing on Wnt/β-catenin signaling pathway of human malignant melanoma cell line A375 was investigated.Two types of dsRNA targeting Sox4 were constructed and transfected into A375 cells,and untreated cells and cells transfected with scramble RNA were used as blank control and negative control respectively.The expression levels of mRNA and protein of Sox4,Wnt3a,β-catenin and Wnt/β-catenin signaling target gene Survivin were detected after real-time PCR and Western blot respectively.MTT assay was used to measure cell proliferation after Sox4 knockdown.β-catenin/TCF transcription reporter assay was used for assessing Wnt/β-catenin signaling pathway activity.Our results showed that the two types of Sox4 siRNA were transfected into A375 cells successfully.As compared with untreated cells,Sox4 siRNAs had no significant influence on Wnt3a expression,and Sox4 siRNAs led to the decrease of β-catenin at protein level.Wnt/β-catenin signaling pathway activity was inhibited significantly.As a target of Wnt/β-catenin signaling,Survivin was decreased at both mRNA and protein levels,and cell proliferation was attenuated.Our study suggests that Sox4 may play an important role in Wnt/β-catenin signaling pathway in human malignant melanoma cells by regulating β-catenin protein level,indicating that Sox4 is involved in the progression of malignant melanoma through Wnt/β-catenin signaling pathway.

DKK4通过负调控Wnt/β-catenin信号通路抑制乳腺癌细胞的增殖、迁移和侵袭

Wnt信号通路在机体多种生理过程中均具有重要作用,已证明有多种蛋白质参与其调节,包括DKK(Dickkopf)家族成员。然而,DKK4在乳腺癌中的生物学功能和分子机制尚不清楚。本研究通过在线数据库TCGA(the cancer genome atlas)和UALCAN(ualcan.path.uab.edu/)分析发现,DKK4在乳腺癌中的表达与其甲基化程度负相关,并影响其肿瘤分期和患者生存率。MethHC数据库分析显示,DKK4在多数乳腺癌组织中的表达低于癌旁组织。RT-PCR检测发现,DKK4在8种人乳腺癌细胞株中不表达或低表达,在2种人正常乳腺细胞中表达;qPCR分析发现,13对组织标本中有11对癌组织的表达低于癌旁组织(P<0.0001)。构建过表达DKK4的人乳腺癌细胞株MCF7和YCCB1,克隆形成的结果显示,MCF7和YCCB1稳定细胞株形成的细胞集落数量分别为对照组的28.66%和37.26%,增殖能力明显低于对照组细胞(P<0.001);Transwell实验结果显示,细胞迁移能力减弱(590 vs. 2 052;1 310 vs. 5 137,P<0.001);侵袭能力也明显减弱(220 vs. 872;2 532 vs. 5 089;P<0.001)。流式细胞仪分析发现,DKK4可将乳腺癌细胞周期阻滞于G0/G1期((57.06±0.64)%vs.(50.13±1.08)%;(51.94±0.93)%vs.(31.00±1.03)%,P<0.001),细胞凋亡率明显增高((31.55±0.77)%vs.(9.85±0.58)%;(28.19±0.99)%vs.(17.92±0.58)%,P<0.001)。进一步采用Western印迹检测结果显示,DKK4能够使细胞周期调控蛋白P53、P27、P21和细胞凋亡因子胱天蛋白酶-3、胱天蛋白酶-7、胱天蛋白酶-9表达上调。Western印迹检测过表达DKK4对Wnt/β-catenin信号通路的影响。结果表明,细胞周期蛋白D1(cyclin D1)、C-Myc蛋白、环氧合酶2 (cyclooxygenase 2,Cox 2)、C-Jun蛋白、磷酸化应激活化蛋白激酶(p-JNK)和活化的β-联蛋白(activeβ-catenin)等的表达下调,而上皮型钙黏着蛋白(E-Cadherin)表达上调。综上所述,DKK4通过负调控Wnt/β-catenin信号通路抑制乳腺癌细胞的增殖、迁移和侵袭,阻滞细胞周期,并促进其凋亡。

多发性骨髓瘤患者的血清PINP、DKK1和SFRP3水平及其临床意义

目的探讨多发性骨髓瘤(MM)患者的血清I型原胶原氨基端前肽(PINP)、dickkopf Wnt信号通路抑制因子1(DKK1)和分泌型卷曲相关蛋白3(SFRP3)水平及其临床意义。方法纳入150例MM患者(MM组)及150健康体检者(对照组)。比较两组研究对象之间、不同临床分期MM患者之间、不同临床疗效MM患者之间的血清DKK1、PINP、SFRP3水平。采用Logistic回归模型分析治疗前血清DKK1、PINP和SFRP3水平与MM患者临床疗效的关系。采用受试者工作特征(ROC)曲线分析治疗前血清DKK1、PINP和SFRP3水平对MM患者临床疗效的预测效能。结果MM组患者治疗前血清DKK1、PINP和SFRP3水平高于对照组(P<0.05);Ⅰ期、Ⅱ期、Ⅲ期MM患者治疗前血清DKK1、PINP、SFRP3水平依次升高(P<0.05);有效组患者治疗前血清DKK1、PINP、SFRP3水平低于无效组(P<0.05)。治疗前血清DKK1、SFRP3、PINP水平升高是影响MM患者临床疗效的独立危险因素(P<0.05)。治疗前血清DKK1和SFRP3水平对MM患者临床疗效无预测价值(曲线下面积<0.5,P>0.05),而治疗前血清PINP水平对MM患者的临床疗效有一定的预测价值(曲线下面积=0.663,P<0.05)。结论MM患者治疗前的血清DKK1、PINP和SFRP3水平高于健康人群,且与疾病严重程度有关,是MM患者临床疗效的影响因素。治疗前血清PINP水平对预测MM患者的临床疗效有一定的价值。

Wnt/β-catenin通路参与脂肪间充质干细胞对大鼠肾小球系膜细胞增殖的调控

目的探讨脂肪间充质干细胞(ADSC)经Wnt/β联蛋白(β-catenin)通路对肾小球系膜细胞增殖的影响。方法以大鼠ADSC的条件培养基作用于HBZY-1大鼠肾小球系膜细胞,通过流式细胞术分析细胞周期、RNA干扰、实时定量PCR和Western blot等技术分析HBZY-1细胞增殖能力、Wnt信号通路中相关基因和蛋白表达水平的变化。结果在ADSC条件培养基作用下,HBZY-1细胞增殖受到明显抑制,dickkopf WNT信号通路抑制物1(DKK1)mRNA表达水平明显升高,而纤连蛋白(fibronectin)和转化生长因子β1(TGF-β1)的mRNA表达显著降低,同时系膜细胞的β-catenin和Bcl-2蛋白的表达受到明显抑制。ADSC中DKK1 mRNA表达水平显著高于HBZY-1细胞,利用小干涉RNA敲低ADSC的DKK1表达后,其β-catenin蛋白的表达水平显著升高。将干扰后的ADSC条件培养基作用于HBZY-1细胞,可重新增加HBZY-1细胞Wnt信号通路中β-catenin和Bcl-2的蛋白水平。结论 Wnt/β-catenin信号通路可能参与ADSC对HBZY-1肾小球系膜细胞增殖的调控。

Dickkopfs分泌蛋白调控肿瘤细胞机制

Dickkopfs是Wnt/β-catenin信号通路的抑制因子,具有抑制肿瘤生长的作用。Dickkopfs分泌蛋白可用作临床诊断和治疗的靶点,因此,有关Dickkopfs分泌蛋白精准有效调控肿瘤细胞机制及靶向抑制剂的研究,是癌症研究中重要的研究对象之一。目前的研究对Dickkopfs蛋白家族缺乏系统性归纳,因此对Dickkopfs在肿瘤细胞中的调控机制进行分析。结果发现:DKK1、DKK2、DKK4主要通过引起蛋白酶体β-catenin降解、诱导细胞凋亡和阻止细胞增殖来抑制Wnt/β-catenin通路;DKK3是通过对β-catenin的下游信号通路因子进行负调控的详细机制。阐述了Dickkopfs分泌蛋白在临床诊断、治疗及预后的应用,并对其可能存在抑制肿瘤血管生成机制进行了展望。

Aberrant expression and function of TCF4 in the proliferation of hepatocellular carcinoma cell line BEL-7402

Wnt signaling pathway is essential for development and tumorigenesis,however,this signaling pathway in the progress of hepatocellular carcinoma (HCC) remains unclear. In this paper,we studied the function of human T-cell transcription factor-4 (TCF4),a key factor of Wnt signaling pathway,on the proliferation of HCC cell line. We showed that the expression of TCF4 mRNA in HCC cell line BEL-7402 was higher than that in immortalized normal liver cell line L02. Blockage of Wnt pathway by △NTCF4,a dominant negative TCF4,could suppress BEL-7402 cells growth and decrease the expression of cyclin D1 and c-myc,two of target genes of Wnt pathway. On the other hand,stimulating Wnt pathway by introducing a degradation-resistant β-catenin S37A could increase BEL-7402 cells proliferation. But all the treatments had no effect on L02 cells. Our data indicated that TCF4 might be another key factor in Wnt pathway involved in HCC cells proliferation and TCF4 could be an effective therapeutic target for suppressing the growth of hepatocellular cancers.

甲胎蛋白、胸苷激酶1、Dickkopf相关蛋白1联合检测在原发性肝癌中的诊断价值

目的探讨血清甲胎蛋白(AFP)、胸苷激酶1(TK1)、Dickkopf相关蛋白1(DKK1)联合检测在原发性肝癌中的诊断价值。方法选取120例原发性肝癌患者(原发性肝癌组)、90例良性肝病患者(良性肝病组)和80例健康体检者(健康对照组)作为研究对象,比较3组受试者血清中AFP、TK1、DKK1的水平及阳性表达情况,计算AFP、TK1、DKK1单独及联合检测诊断原发性肝癌的灵敏度、特异度和准确度。结果原发性肝癌组患者血清中AFP、TK1和DKK1的水平均高于良性肝病组和健康对照组(P﹤0.05)。原发性肝癌组患者血清中AFP、TK1和DKK1的阳性表达率均高于良性肝病组和健康对照组(P﹤0.05)。AFP、TK1、DKK1联合检测诊断原发性肝癌的灵敏度、特异度和准确度均高于AFP、TK1、DKK1单独检测的结果。结论血清AFP、TK1、DKK1联合检测对原发性肝癌的诊断具有较高的灵敏度、特异度和准确度,对于临床原发性肝癌的诊断具有重要的参考价值,值得临床推广和应用。

加味桂枝附子汤联合艾灸治疗类风湿关节炎疗效研究

目的:探讨加味桂枝附子汤联合艾灸治疗类风湿关节炎(RA)疗效及对血清白细胞介素-21(IL-21)、Wnt信号通路关键抑制因子Dickkopf-1(DKK-1)的影响。方法:选取80例RA患者随机分为对照组、联合组,各40例。对照组给予加味桂枝附子汤,联合组给予加味桂枝附子汤联合艾灸治疗。比较两组中医证候积分、临床疗效、晨僵时间、压痛关节数(TJC)、肿胀关节数(SJC)、关节功能评分、视觉模拟评分(VAS)、风湿性关节炎活动度评分(DAS28)、血清IL-21、血清DKK-1、不良反应。结果:与治疗前比较,两组治疗后晨僵、关节疼痛、关节肿胀评分、晨僵时间、TJC、SJC、WOMAC评分、VAS评分、DAS28评分、血清IL-21、DKK-1水平均下降,且治疗后联合组上述指标均低于对照组,差异有统计学意义(均P<0.05)。联合组总有效率92.50%高于对照组72.50%,差异有统计学意义(P<0.05)。两组患者便秘、失眠、皮肤瘙痒等不良反应发生率比较,差异无统计学意义(均P>0.05)。结论:加味桂枝附子汤联合艾灸治疗RA可有效改善患者临床症状,提高关节功能,减轻疼痛,降低血清IL-21、DKK-1水平,安全性较高。

Wnt4 increases the thickness of the epidermis in burn wounds by activating canonical Wnt signalling and decreasing the cell junctions between epidermal cells

Background:Burn wound healing is a complex process and the role of Wnt ligands varies in this process.Whether and how Wnt4 functions in burn wound healing is not well understood.In this study,we aim to reveal the effects and potential mechanisms of Wnt4 in burn wound healing.Methods:First,the expression of Wnt4 during burn wound healing was determined by immunoflu-orescence,Western blotting and qPCR.Then,Wnt4 was overexpressed in burn wounds.The healing rate and healing quality were analysed by gross photography and haematoxyline and eosin staining.Collagen secretion was observed by Masson staining.Vessel formation and fibroblast distribution were observed by immunostaining.Next,Wnt4 was knocked down in HaCaT cells.The migration of HaCaT cells was analysed by scratch healing and transwell assays.Next,the expression ofβ-catenin was detected by Western blotting and immunofluorescence.The binding of Frizzled2 and Wnt4 was detected by coimmunoprecipitation and immunofluorescence.Finally,the molecular changes induced by Wnt4 were analysed by RNA sequencing,immunofluorescence,Western blotting and qPCR in HaCaT cells and burn wound healing tissues.Results:The expression of Wnt4 was enhanced in burn wound skin.Overexpression of Wnt4 in burn wound skin increased the thickness of epidermis.Collagen secretion,vessel formation and fibroblast distribution were not significantly impacted by Wnt4 overexpression.When Wnt4 was knocked down in HaCaT cells,the ratio of proliferating cells decreased,the ratio of apoptotic cells increased and the ratio of the healing area in the scratch healing assay to the number of migrated cells in the transwell assay decreased.The nuclear translocation ofβ-catenin decreased in shRNA of Wnt4 mediated by lentivirus-treated HaCaT cells and increased in Wnt4-overexpressing epidermal cells.RNA-sequencing analysis revealed that cell junction-related signalling pathways were significantly impacted by Wnt4 knockdown.The expression of the cell junction proteins was decreased by the overexpression of Wnt4.Conclusions:Wnt4 promoted the migration of epidermal cells.Overexpression of Wnt4 increased the thickness of the burn wound.A potential mechanism for this effect is that Wnt4 binds with Frizzled2 and increases the nuclear translocation ofβ-catenin,thus activating the canonical Wnt signalling pathway and decreasing the cell junction between epidermal cells.

二肽基肽酶4抑制剂抑制Wnt/β-catenin信号通路改善慢性心力衰竭大鼠心肌纤维化

目的探讨二肽基肽酶4(DPP-4)抑制剂对慢性心力衰竭大鼠心肌纤维化的影响及与Wnt/β-catenin信号通路的关系。方法将50只SPF级SD大鼠,体重(200±20)g,随机分为对照组(Control group)、模型组(Model group)、二肽基肽酶4抑制剂组(DPP-4inhibitor group)、氯化锂组(LiClgroup)及氯化锂+DPP-4组(LiCl+DPP-4inhibitor group),每组10只。利用腹腔注射阿霉素(2.5mg/kg)的方法构建慢性心力衰竭大鼠模型。利用超声检测各组大鼠心功能;HE及Masson染色检测各组大鼠心脏病理学改变;Western blot检测各组大鼠心脏组织中TGF-β1,α-SMA,GSK-3β,p-GSK-3β及β-catenin蛋白的表达。结果与模型组相比,DPP-4inhibitor组大鼠心功能显著改善,心脏组织病理损伤及心肌纤维化减轻,心脏组织中TGF-β1和α-SMA蛋白表达明显减少,p-GSK-3β及β-catenin蛋白表达也显著降低(P<0.05);LiCl加重慢性心力衰竭大鼠模型的心脏功能、病理损伤及心肌纤维化程度,增加TGF-β1和α-SMA蛋白表达,并显著增加p-GSK-3β及β-catenin蛋白表达;但是DPP-4抑制剂能够减轻甚至部分逆转LiCl对慢性心力衰竭大鼠的影响。结论二肽基肽酶4抑制剂改善慢性心力衰竭大鼠心肌纤维化,与抑制Wnt/β-catenin信号通路相关蛋白表达相关。

多西他赛、雷替曲塞治疗无法手术胃癌患者的疗效及作用机制

目的探讨多西他赛联合雷替曲塞治疗无法手术的晚期胃癌患者的疗效及对血清dickkopf WNT信号通路抑制剂1(DKK1)、三叶因子3(TFF3)和内皮抑制蛋白(endostatin)水平的影响。方法采用随机数字表法将92例晚期无法接受手术治疗的胃癌患者分为研究组和对照组,每组46例。研究组采用多西他赛联合雷替曲塞治疗,对照组采用多西他赛联合5-氟尿嘧啶治疗。比较两组患者的疗效,化疗前后血清endostatin、DKK1、TFF3水平,不良反应发生情况及生存情况。结果化疗后,研究组患者总有效率为75.56%,高于对照组的52.27%,差异有统计学意义(P﹤0.05)。化疗前,两组患者血清endostatin、DKK1、TFF3水平比较,差异均无统计学意义(P﹥0.05);化疗后,研究组患者血清DKK1、TFF3水平均低于对照组,endostatin水平高于对照组,差异均有统计学意义(P﹤0.05)。研究组患者恶心呕吐、骨髓抑制发生率均低于对照组,差异均有统计学意义(P﹤0.05)。研究组患者中位生存时间为22.0个月,长于对照组的17.0个月,差异有统计学意义(P﹤0.05)。结论多西他赛联合雷替曲塞治疗无法手术的晚期胃癌患者较多西他赛联合5-氟尿嘧啶更具有优势。

多囊卵巢综合征患者子宫内膜病变的基因表达谱

目的:通过基因芯片技术筛选出多囊卵巢综合征(PCOS)患者子宫内膜正常组(PCOS-E)和子宫内膜病变组(PCOS-EH)的差异表达基因,探讨PCOS合并子宫内膜病变的相关致病基因。方法:选取2018年3月~2019年11月于广东省妇幼保健院行宫腔镜检查+诊刮术的PCOS患者,根据病理检查结果:分为两组PCOS-E和PCOS-EH。两组中各随机选取10例子宫内膜标本,利用芯片差异显著性分析(SAM)软件筛选两组差异表达基因,并进行Gene Ontology(GO)及Pathway富集分析,建立基因表达谱。结果:基因芯片差异显著性分析结果:共筛选出有表达意义的基因29871个,PCOS-EH组子宫内膜组织中存在表达差异的基因共810个,其中上调基因408个,下调基因402个。GO生物学过程富集类别128个,分子功能富集类别23个,细胞组成富集类别19个,发现多个基因功能簇的富集。共筛选出23条代谢通路的差异基因,涉及Wnt信号通路、细胞因子-细胞因子受体相互作用、PI3K-Akt信号通路、MAPK信号通路,其中Wnt信号通路的负调控(q-value=4.96E-05)具有显著差异。利用KEGG通路分析发现,Wnt信号通路中涉及8个基因,其中2个差异基因表达下调(SFRP1=-2.70、SFRP4=-4.55),6个差异基因表达上调(WIF1=11.88、DKK4=9.40、NKD1=8.93、WNT6=4.65、BAMBI=3.79、PRKCG=2.99),表达均显著差异。结论:PCOS患者子宫内膜病变的基因表达谱与正常增殖期子宫内膜存在明显差异,累及Wnt信号传导通路,这些改变可能与PCOS患者子宫内膜病变相关,相关基因表达(产物)可作为早期预测的生物标记物。

Transplantation of Mesenchymal Stem Cells Attenuates Acute Liver Failure in Mice via an Interleukin-4-dependent Switch to the M2 Macrophage Anti-inflammatory Phenotype

Background and Aims:Transplantation of mesenchymal stem cells(MSCs)derived from bone marrow(BM)is an alternative treatment of acute liver failure(ALF)mainly be-cause of the resulting anti-inflammatory activity.It is not known how MSCs regulate local immune responses and liver regeneration.This study explored the effects of MSCs on hepatic macrophages and the Wnt signaling pathway in ALF.Methods:MSCs were isolated from BM aspirates of C57BL/6J mice,and transplanted in mice with ALF induced by D-galactosamine(D-Gal).The proliferation of hepato-cytes was assayed by immunohistochemical(IHC)staining of Ki-67 and proliferating cell nuclear antigen(PCNA).The levels of key proteins in the Wnt signaling pathway were assayed by western blotting and cytokines were determined enzyme-linked immunosorbent assays(ELISAs).A mac-rophage polarization assay characterized the M1/M2 ratio.The potential role of interleukin-4(IL-4)in the biological ac-tivity of MSCs was determined by silencing of IL-4.Results:Transplantation of allogeneic MSCs significantly attenuated D-Gal-induced hepatic inflammation and promoted liver re-generation.MSC transplantation significantly promoted a phenotypic switch from proinflamatory M1 macrophages to anti-inflammatory M2 macrophages,leading to significant Wnt-3a induction and activation of the Wnt signaling path-way in mice with D-Gal-induced ALF.Of the paracrine fac-tors secreted by MSCs(G-CSF,IL-6,IL-1 beta,IL-4,and IL-17A),IL-4 was specifically induced following transplantation in the ALF model mice.The silencing of IL-4 significantly ab-rogated the phenotypic switch to M2 macrophages and the protective effects of MSCs in both the ALF model mice and a co-culture model in an IL-4 dependent manner.Conclu-sions:In vivo and in vitro studies showed that MSCs ame-liorated ALF through an IL-4-dependent macrophage switch toward the M2 anti-inflammatory phenotype.The findings may have clinical implications in that overexpression of IL-4 may enhance the therapeutic effects of allogeneic MSC transplantation in the treatment of ALF.

Drug discovery of sclerostin inhibitors

Sclerostin, a protein secreted from osteocytes, negatively regulates the WNT signaling pathway by binding to the LRP5/6 co-receptors and further inhibits bone formation and promotes bone resorption. Sclerostin contributes to musculoskeletal system-related diseases, making it a promising therapeutic target for the treatment of WNT-related bone diseases. Additionally, emerging evidence indicates that sclerostin contributes to the development of cancers, obesity, and diabetes, suggesting that it may be a promising therapeutic target for these diseases. Notably, cardiovascular diseases are related to the protective role of sclerostin. In this review, we summarize three distinct types of inhibitors targeting sclerostin, monoclonal antibodies, aptamers, and small-molecule inhibitors, from which monoclonal antibodies have been developed. As the first-in-class sclerostin inhibitor approved by the U.S. FDA,the monoclonal antibody romosozumab has demonstrated excellent effectiveness in the treatment of postmenopausal osteoporosis;however, it conferred high cardiovascular risk in clinical trials. Furthermore,romosozumab could only be administered by injection, which may cause compliance issues for patients who prefer oral therapy. Considering these above safety and compliance concerns, we therefore present relevant discussion and offer perspectives on the development of next-generation sclerostin inhibitors by following several ways, such as concomitant medication, artificial intelligence-based strategy, druggable modification, and bispecific inhibitors strategy.