Yes相关蛋白(YAP)通过激活PI3K/AKT通路促进皮肤鳞状细胞癌细胞侵袭和迁移

目的探讨Yes相关蛋白(YAP)在皮肤鳞状细胞癌(cSCC)中表达及与cSCC侵袭和迁移中的作用。方法通过免疫组织化学染色法检测cSCC、鲍温病(BD)、癌旁正常皮肤组织中YAP的表达水平,并分析与临床病理参数之间的关系;利用慢病毒转染构建YAP基因敲低的A431稳定细胞株,利用四甲基罗丹明标记的鬼笔环肽检测A431细胞微丝分布和数量,Transwell TM实验检测细胞侵袭能力,划痕实验检测A431细胞的迁移能力;免疫荧光细胞化学染色法观察敲低YAP后上皮间质转化(EMT)相关标志物上皮钙黏素(E-cadherin)、锌指转录因子Snail的表达;Western blot法检测E-cadherin、Snail、β-catenin、磷脂酰肌醇3激酶(PI3K)、蛋白激酶B(AKT)、磷酸化的蛋白激酶B(p-AKT)、核糖体蛋白S6(S6)、磷酸化S6(p-S6)、4E结合蛋白1(4EBP1)、磷酸化的4EBP1(p-4EBP1)的表达。结果YAP在cSCC和BD中表达显著高于癌旁正常皮肤组织;cSCC中YAP高表达与肿瘤大小、分化程度、侵袭程度密切相关,与患者的性别、年龄、发病部位、形态类型、是否神经脉管侵犯不相关;敲低A431细胞中YAP后,肿瘤细胞的侵袭、迁移能力降低,细胞微丝变细、伪足变少;E-cadherin表达增加,Snail和β-catenin蛋白表达降低,p-AKT、p-S6及p-4EBP1蛋白表达降低。结论YAP在cSCC中高表达,YAP激活PI3K/AKT信号通路促进cSCC的侵袭、迁移及EMT过程。

LncRNA-ATB promotes autophagy by activating Yes-associated protein and inducing autophagy-related protein 5 expression in hepatocellular carcinoma

BACKGROUND Long non-coding RNAs (lncRNAs) play important roles in many diseases, including hepatocellular carcinoma (HCC). Autophagy is a metabolic pathway that facilitates cancer cell survival in response to stress. The relationship between autophagy and the lncRNA-activated by transforming growth factor beta (lncRNA-ATB) in HCC remains unknown. AIM To explore the influence of lncRNA-ATB in regulating autophagy in HCC cells and the underlying mechanism. METHODS In the present study, we evaluated lncRNA-ATB expression in tumor and adjacent non-tumor tissues from 72 HCC cases by real-time PCR. We evaluated the role of lncRNA-ATB in the proliferation and clonogenicity of HCC cells in vitro. The effect of lncRNA-ATB on autophagy was determined using a LC3-GFP reporter and transmission electron microscopy. Furthermore, the mechanism by which lncRNA-ATB regulates autophagy was explored by immunofluorescence staining, RNA immunoprecipitation (RIP), and Western blot. RESULTS The expression of lncRNA-ATB was higher in HCC tissues than in normal liver tissues, and lncRNA-ATB expression was positively correlated with tumor size, TNM stage, and poorer survival of patients with HCC. Moreover, ectopic overexpression of lncRNA-ATB promoted cell proliferation and clonogenicnity of HCC cells in vitro. LncRNA-ATB promoted autophagy by activating Yesassociated protein (YAP). Moreover, lncRNA-ATB interacted with autophagy-related protein 5 (ATG5) mRNA and increased ATG5 expression. CONCLUSION LncRNA-ATB regulates autophagy by activating YAP and increasing ATG5 expression. Our data demonstrate a novel function for lncRNA-ATB in autophagy and suggest that lncRNA-ATB plays an important role in HCC.

Significance and relationship between Yes-associated protein and survivin expression in gastric carcinoma and precancerous lesions

AIM:To analyze the differences and relevance of Yes-associated protein (YAP) and survivin, and to explore the correlation and signifi cance of their expression in gastric carcinoma and precancerous lesions.METHODS: The PV9000 immunohistochemical method was used to detect the expression of YAP and survivin in 98 cases of normal gastric mucosa, 58 intestinal metaplasia (IM), 32 dysplasia and 98 gastric carcinoma.RESULTS: The positive rates of YAP in dysplasia (37.5%) and gastric carcinoma (48.0%) were significantly higher than that in normal gastric mucosa (13.3%), P<0.01. The positive rates of survivin in IM (53.4%), dysplasia (59.4%) and gastric carcinoma (65.3%) were significantly higher than in normal gastric mucosa (11.2%), P<0.01. Survivin expression gradually increased from 41.7% in well differentiated adenocarcinoma through 58.3% in moderately differentiated adenocarcinoma to 75.6% in poorly differentiated adenocarcinoma, with significant Rank correlation, rk=0.279, P<0.01. The positive rate of survivin in gastric carcinoma of diffused type (74.6%) was significantly higher than that in intestinal type (51.3%), P<0.05. In gastric carcinoma with lymph node metastasis (76.9%), the positive rate of survivin was signifi cantly higher than that in the group without lymph node metastasis (41.2%), P<0.01. In 98 cases of gastric carcinoma, the expression of YAP and of survivin were positively correlated, rk=0.246, P<0.01.CONCLUSION: YAP may play an important role as a carcinogenic factor and may induce survivin expression. Detecting both markers together may help in early diagnosis of gastric carcinoma.

Central role of Yes-associated protein and WW-domain-containing transcriptional co-activator with PDZ-binding motif in pancreatic cancer development

Pancreatic ductal adenocarcinoma(PDAC) remains a deadly disease with no efficacious treatment options. PDAC incidence is projected to increase, which may be caused at least partially by the obesity epidemic. Significantly enhanced efforts to prevent or intercept this cancer are clearly warranted. Oncogenic KRAS mutations are recognized initiating events in PDAC development, however, they are not entirely sufficient for the development of fully invasive PDAC.Additional genetic alterations and/or environmental, nutritional, and metabolic signals, as present in obesity, type-2 diabetes mellitus, and inflammation, are required for full PDAC formation. We hypothesize that oncogenic KRAS increases the intensity and duration of the growth-promoting signaling network.Recent exciting studies from different laboratories indicate that the activity of the transcriptional co-activators Yes-associated protein(YAP) and WW-domaincontaining transcriptional co-activator with PDZ-binding motif(TAZ) play a critical role in the promotion and maintenance of PDAC operating as key downstream target of KRAS signaling. While initially thought to be primarily an effector of the tumor-suppressive Hippo pathway, more recent studies revealed that YAP/TAZ subcellular localization and co-transcriptional activity is regulated by multiple upstream signals. Overall, YAP has emerged as a central node of transcriptional convergence in growth-promoting signaling in PDAC cells. Indeed, YAP expression is an independent unfavorable prognostic marker for overall survival of PDAC. In what follows, we will review studies implicating YAP/TAZ in pancreatic cancer development and consider different approaches to target these transcriptional regulators.

Yes-associated protein promotes endothelial-tomesenchymal transition of endothelial cells in choroidal neovascularization fibrosis

AIM:To reveal whether and how Yes-associated protein(YAP)promotes the occurrence of subretinal fibrosis in agerelated macular degeneration(AMD).METHODS:Cobalt chloride(Co Cl2)was used in primary human umbilical vein endothelial cells(HUVECs)to induce hypoxia in vitro.Eight-week-old male C57 BL/6 J mice weighing 19-25 g were used for a choroidal neovascularization(CNV)model induced by laser photocoagulation in vivo.Expression levels of YAP,phosphorylated YAP,mesenchymal markers[αsmooth muscle actin(α-SMA),vimentin,and Snail],and endothelial cell markers(CD31 and zonula occludens 1)were measured by Western blotting,quantitative real-time PCR,and immunofluorescence microscopy.Small molecules YC-1(Lificiguat,a specific inhibitor of hypoxia-inducible factor 1α),CA3(CIL56,an inhibitor of YAP),and XMU-MP-1(an inhibitor of Hippo kinase MST1/2,which activates YAP)were used to explore the underlying mechanism.RESULTS:Co Cl2 increased expression of mesenchymal markers,decreased expression of endothelial cell markers,and enhanced the ability of primary HUVECs to proliferate and migrate.YC-1 suppressed hypoxia-induced endothelialto-mesenchymal transition(End MT).Moreover,hypoxia promoted total expression,inhibited phosphorylation,and enhanced the transcriptional activity of YAP.XMU-MP-1 enhanced hypoxia-induced End MT,whereas CA3 elicited the opposite effect.Expression of YAP,α-SMA,and vimentin were upregulated in the laser-induced CNV model.However,silencing of YAP by vitreous injection of small interfering RNA targeting YAP could reverse these changes.CONCLUSION:The findings reveal a critical role of the hypoxia-inducible factor-1α(HIF-1α)/YAP signaling axis in End MT and provide a new therapeutic target for treatment of subretinal fibrosis in AMD.

Transcription factor glucocorticoid modulatory element-binding protein 1 promotes hepatocellular carcinoma progression by activating Yes-associate protein 1

BACKGROUND Glucocorticoid modulatory element-binding protein 1(GMEB1),which has been identified as a transcription factor,is a protein widely expressed in various tissues.Reportedly,the dysregulation of GMEB1 is linked to the genesis and development of multiple cancers.AIM To explore GMEB1’s biological functions in hepatocellular carcinoma(HCC)and figuring out the molecular mechanism.METHODS GMEB1 expression in HCC tissues was analyzed employing the StarBase database.Immunohistochemical staining,Western blotting and quantitative realtime PCR were conducted to examine GMEB1 and Yes-associate protein 1(YAP1)expression in HCC cells and tissues.Cell counting kit-8 assay,Transwell assay and flow cytometry were utilized to examine HCC cell proliferation,migration,invasion and apoptosis,respectively.The JASPAR database was employed for predicting the binding site of GMEB1 with YAP1 promoter.Dual-luciferase reporter gene assay and chromatin immunoprecipitation-qPCR were conducted to verify the binding relationship of GMEB1 with YAP1 promoter region.RESULTS GMEB1 was up-regulated in HCC cells and tissues,and GMEB1 expression was correlated to the tumor size and TNM stage of HCC patients.GMEB1 overexpression facilitated HCC cell multiplication,migration,and invasion,and suppressed the apoptosis,whereas GMEB1 knockdown had the opposite effects.GMEB1 bound to YAP1 promoter region and positively regulated YAP1 expression in HCC cells.CONCLUSION GMEB1 facilitates HCC malignant proliferation and metastasis by promoting the transcription of the YAP1 promoter region.

基于Hippo-YAP信号通路探讨黄芩苷干预膝骨关节炎大鼠的疗效和作用机制

目的:探讨黄芩苷干预膝骨关节炎(knee osteoarthritis,KOA)大鼠的疗效和作用机制。方法:将50只大鼠随机分为假手术组、模型组、低剂量黄芩苷组、高剂量黄芩苷组、高剂量黄芩苷联合抑制剂组,每组10只。将假手术组以外的大鼠采用切断前交叉韧带和切除内侧半月板的方法建立右膝关节KOA模型,假手术组大鼠于右膝关节内侧做一切口后缝合。造模成功后,低剂量黄芩苷组、高剂量黄芩苷组大鼠分别按照50 mg·kg^(-1)、100 mg·kg^(-1)的剂量给予黄芩苷生理盐水溶液灌胃;高剂量黄芩苷联合抑制剂组大鼠按照100 mg·kg^(-1)的剂量给予黄芩苷生理盐水溶液灌胃,并按照1 mg·kg^(-1)的剂量给予XMU-MP-1溶液腹腔注射;假手术组和模型组大鼠均给予等量生理盐水灌胃。每天给药1次,连续给药30 d。分别于给药前、给药15 d时、给药30 d时测量大鼠双侧膝关节肿胀程度。给药结束后处死大鼠,取大鼠右侧膝关节软骨组织,采用苏木精-伊红染色和番红O-固绿染色观察膝关节软骨组织病理变化,采用Mankin评分标准评价软骨退变情况,采用ELISA试剂盒检测肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素(interleukin,IL)-1β、IL-10、基质金属蛋白酶(matrix metalloproteinase,MMP)-13表达水平,采用Western blotting检测滑膜组织中切割活化的半胱氨酸天冬氨酸蛋白酶(Cleaved-cysteine aspartic acid specific protease,Cleaved-Caspase)-3、B淋巴细胞瘤-2相关X蛋白(Bcl2 associated X protein,Bax)、B淋巴细胞瘤-2(B-cell lymphoma-2,Bcl-2)蛋白、Yes相关蛋白(Yes-associated protein,YAP)、Tafazzin(TAZ)蛋白的表达水平。结果:①大鼠右膝关节肿胀程度。给药15 d时和给药30 d时,高剂量黄芩苷组大鼠右膝关节肿胀程度均低于模型组、低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组(P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000);模型组、低剂量黄芩苷组、高剂量黄芩苷联合抑制剂组大鼠右膝关节肿胀程度两两比较,差异均无统计学意义(P=0.063,P=0.215,P=0.399;P=0.052,P=0.261,P=0.240)。②大鼠右膝关节软骨组织病理变化。干预结束后,模型组大鼠右膝关节软骨萎缩、排列混乱;低剂量黄芩苷组、高剂量黄芩苷组、高剂量黄芩苷联合抑制剂组大鼠右膝关节软骨萎缩、细胞排序情况均较模型组改善,且高剂量黄芩苷组大鼠的改善情况优于低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组。③大鼠右膝关节软骨Mankin评分。干预结束后,低剂量黄芩苷组、高剂量黄芩苷组、高剂量黄芩苷联合抑制剂组大鼠右膝关节软骨Mankin评分均低于模型组(P=0.000,P=0.000,P=0.000),高剂量黄芩苷组右膝关节软骨Mankin评分低于低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组(P=0.000,P=0.000),低剂量黄芩苷组大鼠右膝关节软骨Mankin评分低于高剂量黄芩苷联合抑制剂组(P=0.000)。④大鼠右膝关节软骨组织中TNF-α、IL-1β、IL-10、MMP-13表达水平。低剂量黄芩苷组、高剂量黄芩苷组、高剂量黄芩苷联合抑制剂组大鼠右膝关节软骨组织中TNF-α、IL-1β、MMP-13表达水平均低于模型组(P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000),IL-10表达水平均高于模型组(P=0.000,P=0.000,P=0.000);高剂量黄芩苷组大鼠右膝关节软骨组织中TNF-α、IL-1β、MMP-13表达水平均低于低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组(P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000),IL-10表达水平高于低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组(P=0.000,P=0.000)。⑤大鼠右膝关节软骨组织细胞凋亡及Hippo-YAP信号通路相关蛋白表达水平。低剂量黄芩苷组、高剂量黄芩苷组、高剂量黄芩苷联合抑制剂组大鼠右膝关节软骨组织中Cleaved-Caspase-3、Bax表达水平均低于模型组(P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000),BCL-2、YAP、TAZ表达水平均高于模型组(P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000;P=0.000,P=0.000,P=0.000);高剂量黄芩苷组大鼠右膝关节软骨组织中Cleaved-Caspase-3、Bax表达水平均低于低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组(P=0.000,P=0.000;P=0.000,P=0.000),BCL-2、YAP、TAZ表达水平均高于低剂量黄芩苷组和高剂量黄芩苷联合抑制剂组(P=0.000,P=0.000;P=0.000,P=0.000;P=0.000,P=0.000)。结论:黄芩苷干预KOA大鼠,能够缓解膝关节肿胀,抑制炎症反应和细胞凋亡,延缓关节软骨退变,其作用机制可能与激活Hippo-YAP信号通路有关。

Reactive oxygen species-induced activation of Yes-associated protein-1 through the c-Myc pathway is a therapeutic target in hepatocellular carcinoma

BACKGROUND The Hippo signaling pathway regulates organ size by regulating cell proliferation and apoptosis with terminal effectors including Yes-associated protein-1(YAP-1).Dysregulation in Hippo pathway has been proposed as one of the therapeutic targets in hepatocarcinogenesis.The levels of reactive oxygen species(ROS)increase during the progression from early to advanced hepatocellular carcinoma(HCC).AIM To study the activation of YAP-1 by ROS-induced damage in HCC and the involved signaling pathway.METHODS The expression of YAP-1 in HCC cells(Huh-7,HepG2,and SNU-761)was quantified using real-time polymerase chain reaction and immunoblotting.Human HCC cells were treated with H2O2,which is a major component of ROS in living organisms,and with either YAP-1 small interfering RNA(siRNA)or control siRNA.To investigate the role of YAP-1 in HCC cells under oxidative stress,MTS assays were performed.Immunoblotting was performed to evaluate the signaling pathway responsible for the activation of YAP-1.Eighty-eight surgically resected frozen HCC tissue samples and 88 nontumor liver tissue samples were used for gene expression analyses.RESULTS H2O2 treatment increased the mRNA and protein expression of YAP-1 in HCC cells(Huh-7,HepG2,and SNU-761).Suppression of YAP-1 using siRNA transfection resulted in a significant decrease in tumor proliferation during H2O2 treatment both in vitro and in vivo(both P<0.05).The oncogenic action of YAP-1 occurred via the activation of the c-Myc pathway,leading to the upregulation of components of the unfolded protein response(UPR),including 78-kDa glucoseregulated protein and activating transcription factor-6(ATF-6).The YAP-1 mRNA levels in human HCC tissues were upregulated by 2.6-fold compared with those in nontumor tissues(P<0.05)and were positively correlated with the ATF-6 Levels(Pearson’s coefficient=0.299;P<0.05).CONCLUSION This study shows a novel connection between YAP-1 and the UPR through the c-Myc pathway during oxidative stress in HCC.The ROS-induced activation of YAP-1 via the c-Myc pathway,which leads to the activation of the UPR pathway,might be a therapeutic target in HCC.

Yes-associated protein at the intersection of liver cell fate determination

A recent publication highlights the importance of high yes-associated protein(YAP) expressing cells in liver regeneration following partial hepatectomy.Although the names of the cell populations described in these articles [hybrid periportal hepatocytes(HybHP) or epithelial-mesenchymal transition(EMT)-reprogrammed hepatocytes] are not identical, they all express high levels of YAP.We hypothesize that the HybHP and EMT-reprogrammed hepatocytes might be a similar cell population. Hippo signaling is the primary pathway that regulates YAP activity. According to the contribution of these two types of cells to liver regeneration and the high YAP expression, Hippo-YAP signaling activation may be a common regulatory pathway experienced by cells undergoing dedifferentiation and reactivating proliferative activity during liver regeneration.Although no evidence has shown that HybHP cells contribute to hepatocellular carcinoma in mouse models, we can not rule out the possibility that these highly regenerative cells can further develop into tumor cells when they acquire mutations caused by viral infection or other risk factors like alcohol. The detailed mechanistic insight of the regulation of YAP expression and activity in HybHP(or other types of cells contributing to liver regeneration) is unknown. We hypothesize that liver regeneration under various conditions will eventually lead to divergent consequences, likely due to the duration of YAP activation regulated by Hippo-large tumor suppressor 1 and 2 pathway in a context-and cell typedependent manner.

Association of Elevated Yes-Associated Protein Expression with Gastric Cancer and Its Clinicopathological Features: A Meta-Analysis

Objectives: To evaluate the difference of YAP-positive expression between GC and adjacent tissues, as well as the association of elevated YAP expression with clinicopathological features of GC. Methods: PubMed, Embase, Web of Science databases and the Chinese National Knowledge Infrastructure (CNKI) were searched from inception up to December 2018. The pooled ORs and corresponding 95% CIs were used to assess the strength of association. The heterogeneity among eligible studies was evaluated by the Q-test and I2 values. The sensitivity analysis was performed by sequential omission of individual studies. Moreover, Begg’s test and Egger’s test were used to evaluate publication bias. Results: A total of 2229 patients from 16 studies were included in this meta-analysis. The results showed that positive YAP expression was closely correlated with GC but not adjacent non-tumor tissue (OR = 8.08, 95% CI = 4.41 - 14.80). Additionally, YAP overexpression was found to be associated with more advanced TNM stage (OR = 2.68, 95% CI = 1.61 - 4.48), deeper invasion depth (OR = 2.05, 95% CI = 1.32 - 3.19), and lymph node metastasis (OR = 1.95, 95% CI = 1.29 - 2.96). No significant correlation was observed between YAP overexpression and degree of differentiation (OR = 1.17, 95% CI = 0.63 - 2.16), as well as gender of patients (OR = 1.12, 95% CI = 0.91 - 1.37) or tumor size (OR = 1.11, 95% CI = 0.82 - 1.49) of gastric cancer. Conclusions: This meta-analysis demonstrated that YAP might be a promising diagnostic marker and even a therapeutic target for gastric cancer.

长链基因间非编码RNA-p21通过调控YAP表达促进胃癌细胞凋亡

目的:探讨lincRNA-p21在胃癌细胞凋亡中的作用及其潜在机制。方法:在胃癌MGC-803细胞系中分别转染siRNA以及过表达质粒实现下调/上调lincRNA-p21表达。lincRNA-p21和YAP的mRNA水平由qRT-PCR方法测定。之后通过流式细胞术和TUNEL测定对细胞凋亡进行检测,并用Western blot检测细胞凋亡相关蛋白。结果:上调lincRNA-p21促进胃癌细胞凋亡,反之亦然。研究还发现下调lincRNA-p21会引起YAP mRNA及蛋白表达量升高。最重要的是,由lincRNA-p21下调引起的抑制凋亡作用可以通过敲低YAP表达来解除。结论:lincRNA-p21可以通过调控YAP表达促进胃癌细胞凋亡,这可能为胃癌治疗提供新思路。

Iduna调控Hippo/YAP通路抑制炎症反应从而减轻围术期小鼠的神经认知障碍

目的:探讨Iduna对老龄小鼠围术期神经认知障碍(PND)过程中炎症因子的影响并分析Hippo/YAP信号通路在此发挥的作用。方法:取20只C57BL/6老龄小鼠分为对照(control)组和模型(model)组(行剖腹探查术诱导建立PND小鼠模型),每组10只,用荧光定量PCR检测PND小鼠脑组织中Iduna的mRNA水平。在PND造模前,通过侧脑室注射Iduna过表达腺相关病毒(AAV)载体、sh-Iduna AAV载体和(或)Hippo/YAP信号通路抑制剂CA3。将40只C57BL/6老龄小鼠分为control+AAV9-Con组、control+AAV9-Iduna组、model+AAV9-Con组和model+AAV9-Iduna组,每组10只,在造模后7 d,用水迷宫实验检测动物学行为,用Western blot检测脑组织中YAP和TAZ表达,用ELISA法检测脑组织中IL-6、TNF-α、TGF-β1和IL-10水平。将40只C57BL/6老龄小鼠分为control+DMSO组、control+CA3组、model+DMSO组和model+CA3组,每组10只,在造模后7 d,用水迷宫实验检测动物学行为,用ELISA法检测脑组织中炎症因子水平。将60只C57BL/6老龄小鼠分为control+AAV9-Con组、control+AAV9-sh-Iduna、control+AAV9-sh-Iduna+CA3组、model+AAV9-Con组、model+AAV9-sh-Iduna组和model+AAV9-sh-Iduna+CA3组,每组10只,在造模后7 d,用水迷宫实验检测动物学行为,用ELISA法检测脑组织中炎症因子水平。结果:与control比较,model组小鼠脑组织中Iduna的mRNA表达下调(P<0.05)。与model+AAV9-Con组比较,model+AAV9-Iduna组小鼠神经认知障碍显著减轻(P<0.05),脑组织中IL-6和TNF-α表达显著减少(P<0.05),TGF-β1和IL-10表达显著增加(P<0.05),而model+AAV9-sh-Iduna组中的上述情况与model+AAV9-Iduna组相反。Iduna可负向调控Hippo/YAP信号通路相关蛋白YAP和TAZ表达。Hippo/YAP信号通路抑制剂CA3的作用与AAV9-Iduna相似,可抑制PND小鼠围术期神经认知障碍(P<0.05),降低脑组织中IL-6和TNF-α水平(P<0.05)。结论:Iduna通过抑制Hippo/YAP信号通路减轻老龄小鼠PND,降低脑组织中IL-6和TNF-α水平。

激活Yes相关蛋白(YAP)抑制铁死亡减轻小鼠急性肝损伤

目的探索Yes相关蛋白(YAP)可否通过调控铁死亡影响急性肝衰竭的发生发展。方法将8周龄C57BL/6小鼠20只随机分为对照组、急性肝衰竭模型组、YAP激动剂XMU-MP-1干预组和YAP抑制剂维替泊芬(verteporfin)干预组。肝组织HE染色、肝脏生化学检测观察小鼠肝损伤表现;试剂盒检测小鼠肝组织中铁(Fe)、丙二醛(MDA)、谷胱甘肽(GSH)含量;透射电镜观察小鼠肝细胞线粒体改变;荧光定量PCR和Western blot法检测YAP及铁死亡关键基因谷胱甘肽过氧化物酶4(GPX4)、5-脂氧合酶(5-LOX)的表达情况。结果与对照组相比,急性肝衰竭小鼠肝组织严重淤血,可见炎细胞浸润伴肝小叶结构破坏,XMU-MP-1干预组肝损伤减轻。随肝衰竭发生,血浆丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)水平显著升高,XMU-MP-1干预组肝功能改善。此外,电镜观察到肝衰竭小鼠肝细胞内线粒体变小,双层膜密度增高,XMU-MP-1减轻线粒体改变。肝衰竭小鼠肝组织内Fe、MDA水平增加,及GPX4蛋白表达降低、5-LOX表达升高均提示铁死亡参与小鼠急性肝衰竭发生,而活化YAP可抑制铁死亡表现。结论活化YAP可通过抑制铁死亡减轻急性肝衰竭小鼠肝损伤。

Hippo信号通路中的核心因子YAP/TAZ参与骨形成的作用与机制

背景:随着机械信号在骨骼中的作用研究逐渐增多,作为力学敏感因子的YAP/TAZ也逐渐受到大众关注。现有研究发现YAP/TAZ可能在骨形成的过程中作为一个重要介质参与其中,但具体机制尚不清楚。目的:对YAP/TAZ参与骨形成的最新研究进展进行综述。方法:第一作者以“Hippo pathway,YAP/TAZ,Bone,Osteogenesis”为英文检索词,以“Hippo信号通路、YAP/TAZ、骨骼、成骨”为中文检索词,检索PubMed数据库和中国知网2016年至2022年3月发表的相关文献,对筛选出的文献进行归纳分析。最终纳入56篇相关文献进行综述。结果与结论:①YAP/TAZ作为Hippo通路的核心因子,在其功能调控上具有特殊性,除了在分子水平上可以调控其活性,细胞微环境的改变,如细胞浓度、细胞形状和细胞外基质的刚度也可以促使其活性改变从而发挥不同功能作用。②YAP/TAZ参与成骨信号传导,并且调控也受细胞微环境影响。过表达或沉默细胞中的YAP/TAZ会促使或抑制成骨信号的传导,从而改变细胞分化结局。③YAP/TAZ参与骨组织细胞的成骨过程,并对不同类型的骨组织细胞有不同功能。但对于过表达或沉默YAP/TAZ方法不同,选择观察细胞种属类型差异较大,结果难以统一量化。④动物实验中证实YAP/TAZ对正常骨骼形态及颌面部形态的正常发育有着至关重要的作用,并且在不同成骨分化阶段有着不同的功能。然而对于实验动物敲除YAP/TAZ条件基因不统一、观察时机不一致导致实验成果难以相互比较。YAP/TAZ在骨形成中有着重要地位,但对于其作用机制还有待深入研究,期待后续有学者深入探索其作用机制,并利用其特性在骨性疾病中发挥作用。

Hippo-YAP通路参与NaAsO 2对PC12细胞周期及细胞凋亡的影响

目的探讨NaAsO 2对PC12细胞YAP蛋白核质分布的影响及Hippo-YAP通路在NaAsO 2影响细胞周期及细胞凋亡中的作用。方法以PC12细胞作为研究对象,利用核质分离及免疫荧光观察NaAsO 2对YAP蛋白核质分布的影响。再将细胞分为control组、NaAsO 2组和NaAsO 2+XMU-MP-1(MST1/2抑制剂)组、XMU-MP-1组,利用流式细胞术、Western blot验证Hippo-YAP通路在NaAsO 2影响细胞周期及细胞凋亡中的作用。结果相较于control组,核质分离与免疫荧光结果都表明NaAsO 2组细胞质中YAP蛋白明显增加(P<0.05),细胞核中YAP蛋白明显下调(P<0.01);并且NaAsO 2组p-MST1、p-LATS1、p-YAP以及凋亡相关蛋白P53和BAX表达明显上调(P<0.05),BCL-2蛋白表达明显下调(P<0.01);G 0/G 1期细胞比例降低(P<0.01),S期比例上升(P<0.05);细胞凋亡率增加(P<0.01)。与NaAsO 2组相比,NaAsO 2+XMU-MP-1组上述指标均有逆转(P<0.05)。结论Hippo-YAP通路可能参与NaAsO 2对PC12细胞周期及细胞凋亡的影响。

YAP蛋白与肿瘤

Hippo-YAP信号通路是新发现的生长控制信号通路,能够限制调结细胞的生长增殖,细胞凋亡,信号通路异常有利于肿瘤的发生。YAP蛋白还参与调控AKT-mTORC、WNT和MAPK等信号通路,进而影响肿瘤细胞的生长和侵润转移。YAP蛋白在多种类型的人类肿瘤中表达异常增加,或许成为肿瘤新的治疗靶点。

FOXA1和YAP在胃癌中的表达及临床意义

目的探讨胃癌组织中FOXA1蛋白和YAP蛋白的表达及其与胃癌患者临床病理特征的关系。方法收集西安交通大学医学部第二附属医院2009年1月至2013年12月间临床资料保存完整的胃癌组织80例和相应癌旁正常组织80例。应用免疫组化染色测定FOXA1和YAP1在胃癌及癌旁组织中的表达,分析两者表达与胃癌临床病理特征的关系;并进一步检测两者在胃癌中表达水平的相关性。结果 FOXA1蛋白和YAP蛋白在胃癌组织中的免疫组化评分显著高于其在癌旁组织中的免疫组化评分(FOXA1:3.50±0.20 vs.1.13±0.11,P<0.01;YAP:3.40±0.20vs.1.55±0.14,P<0.01);FOXA1蛋白的阳性表达与肿瘤大小、浸润深度、有无淋巴结转移及TNM分期有显著相关性(P<0.01);YAP蛋白的阳性表达与胃癌的分化程度以及淋巴结转移有显著相关性(P<0.01);FOXA1和YAP蛋白在胃癌组织中的表达具有显著相关性(P<0.01)。结论FOXA1和YAP在胃癌组织中存在显著高表达;FOXA1和YAP的阳性表达与胃癌患者的临床病理特征密切相关,两者在胃癌的发生发展可能有协同促进作用,联合检测这两种蛋白的表达对胃癌的临床诊断及预后判断有一定的指导意义。

YAP蛋白在胃腺癌中的表达情况及其临床意义

目的探讨胃腺癌中YAP蛋白的表达情况及其与胃腺癌患者临床病理特征及预后之间的关系。方法应用免疫组织化学法检测人胃腺癌组织中YAP蛋白的表达情况,并用Kaplan-Meier单因素生存分析法和Cox模型多因素回归分析法分析YAP蛋白表达与胃腺癌患者的临床病理特征及预后之间的关系。结果胃腺癌组织中YAP蛋白的细胞质阳性表达(31.6%vs 1.0%)、细胞核阳性表达(22.4%vs 0.0%)均明显高于正常胃黏膜组织(P﹤0.01)。YAP蛋白细胞质阳性表达与细胞核阳性表达呈正相关(Spearman相关系数=0.5818,P﹤0.0001)。Kaplan-Meier单因素生存分析结果显示,YAP蛋白细胞核阳性组患者的5年生存率较YAP蛋白细胞核阴性组低(P﹤0.05)。Cox模型多因素回归分析显示,YAP蛋白的表达未能作为胃腺癌患者预后的独立预测因子(P﹥0.05)。结论部分胃腺癌组织中存在YAP蛋白的高表达,且YAP蛋白细胞核表达的胃腺癌患者的预后较差,可以作为此病的早期诊断指标。

心肌肥大相关蛋白YAP在大鼠糖尿病心肌病中的表达

目的:观察Hippo信号通路效应蛋白YAP在糖尿病大鼠心肌肥大中的改变,分析可能的作用机制。方法:健康雄性SD大鼠随机分为2组,每组6只。正常对照组:大鼠正常饮水饮食喂养12周;糖尿病心肌病组:大鼠腹腔注射STZ(55 mg/kg)复制糖尿病模型,以血糖≥16.7 mmo1/L,持续1周确定造模成功,继续饲养12周。测定大鼠空腹血糖水平和体质量;心室内插管检测左心室动力学变化,称量心脏质量计算心体比;荧光定量PCR检测心肌肥大相关基因心房钠尿肽和脑利钠肽mRNA表达,Western blot法检测YAP和p-YAP蛋白表达改变。结果:与正常对照组相比,糖尿病组大鼠心体比增加,左心室舒张末压增高,左心室发展压、左心室内压最大上升和下降速率及左心室做功均降低;心肌肥大相关基因心房钠尿肽和脑利钠肽mRNA表达增加;YAP蛋白表达增加,p-YAP/YAP比值降低(P<0.05~P<0.01)。结论:糖尿病可诱导心肌肥大,其机制可能与增加Hippo信号通路效应蛋白YAP的表达,降低其磷酸化水平有关。

长链非编码RNA AGAP2-AS1通过YAP通路影响结肠癌细胞的生物学行为

目的:探究AGAP2-AS1在结肠癌组织中的表达及对结肠癌恶性生物学行为和Yes相关蛋白(Yes-associated protein,YAP)信号通路的影响。方法:通过TCGA数据库分析457例结肠癌和42例健康样本的AGAP2-AS1表达水平。收集于2018年1月至2019年3月在义乌市中心医院就诊且通过病理科确诊的结肠癌组织和癌旁组织20例,通过实时荧光定量PCR检测其AGAP2-AS1的表达,之后进一步检测SW480、HCT-116和NCM460细胞中的AGAP2-AS1表达。利用CCK-8试剂盒、克隆形成、细胞划痕和流式细胞实验分析其细胞增殖、迁移、侵袭及凋亡的改变。Western blot检测YAP、p-YAP以及基质金属蛋白酶(matrix metallopeptidase,MMP)2、MMP9的表达。免疫荧光检测YAP在细胞内的定位情况。共转染pcDNA3.1-AGAP2-AS1和si-YAP质粒,然后检测YAP、p-YAP、MMP2、MMP9蛋白的表达。结果:TCGA数据库显示AGAP2-AS1在结肠癌组织显著高表达,并且AGAP2-AS1在结肠癌组织样本和细胞中表达也显著增加。敲低/过表达AGAP2-AS1后HCT-116细胞迁移,增殖能力显著降低/增加,凋亡显著增加/抑制;同时敲低AGAP2-AS1后会诱导YAP磷酸化,减少YAP入核调控,且MMP2、MMP9的表达也显著降低(P<0.05)。共转染结果显示AGAP2-AS1通过激活YAP通路上调MMP2、MMP9表达(P<0.05)。结论:AGAP2-AS1在结肠癌组织和细胞系中均高表达,且诱导结肠癌细胞增殖、迁移并抑制细胞凋亡。此外,AGAP2-AS1通过激活YAP通路调控MMP2、MMP9的表达影响结肠癌侵袭和转移。