The correlation between YAP nuclear expression and tumor size with prognosis of epithelial ovarian cancer / 安徽医科大学学报

Objective To investigate the correlation between Yes-associated protein(YAP)nuclear expression and tumor size with prognosis of patients with epithelial ovarian cancer(EOC)and to study the role of YAP in EOC.Methods 120 patients with EOC were selected as the experimental group,including 38 patients with early stage(Ⅰ+Ⅱ)EOC and 8 2 patients with advanced stage(Ⅲ+Ⅳ)EOC.3 0 normal ovarian tissues obtained from patients with uterine leiomyoma were enrolled as the control group.Immunohistochemical(IHC)assay was em-ployed to determine YAP expression and sub-location.The relationship between YAP expression and the pathologi-cal parameters of the 120 patients with EOC was analyzed,so as to the prognosis of these patients.EOC cells(C13K and OV2008)were cultured with varying initial cell volumes.Ki67 expression and cell proliferation were tested by immunofluorescence and cloning assay respectively.YAP expression at mRNA and protein levels were de-tected by q-PCR and Western blot respectively when the cell conference of EOC cells reached to low(60％)and high(90％)cell density.Results The YAP nuclear expression was significantly higher in the EOC group com-pared to the control group(P＜0.05).The average diameter of stage Ⅰ+Ⅱ EOC was larger than that of stage Ⅲ+Ⅳ EOC(P＜0.01).The high nuclear expression of YAP was positively associated with pathological grade,clinical stage and the level of Ca125＞1 000 IU/ml,while negatively correlated with tumor size(all P＜0.05).Survival analyses showed that smaller tumor size(＜10 cm)and higher YAP nuclear expression were negatively as-sociated with the 3-year overall survival rate of EOC patients(P＜0.01).C13K and OV2008 cells cultured in the low density group exhibited a high number of clone formation,high Ki67 and YAP expression(P＜0.01).The down-regulation of YAP expression could decrease the cell viability of EOC cells in the low-and high-density groups(P＜0.05).Conclusion Higher level of YAP nuclear expression and smaller tumour size are inversely associated with the clinical prognosis of patients with EOC.Inhibiting YAP nuclear expression leads to a decrease in the prolif-eration capacity of EOC cells.

The mechanism of emodin inhibiting YAP1 and FOXD1 in gastric cancer AGS cells and its related study / 实用医学杂志

Objective To explore the possible mechanism of emodin in inhibiting proliferation,migration,and invasion of AGS cells and in suppressing the expressions of YAP1 and FOXD1.Methods Normal gastric cell GES-1 and gastric cancer cell AGS were cultured with different concentrations of emodin.CCK8 test,scratch test and Transwell assay were used to verify changes in the biological phenotype of AGS cells.TCGA database was applied to analyze expressions of HK2,YAP1 and FOXD1 in gastric cancer tissues and normal gastric tissues.Western blotting method was used to detect the impacts of emodin on HK2,YAP1 and FOXD1 proteins in AGS cells.Exogenous pyruvic acid was added to verify the changes in YAP1 and FOXD1.Results The IC50 of emodin was significantly higher in GES-1 cells than in AGS cells(P<0.05).CCK8 proliferation test,scratch test,and Transwell assay showed that emodin significantly inhibited the biological abilities of AGS(P<0.05 for comparisons).Analysis on the TCGA bioinformatics database found that the expression of key enzymes HK2 in the glycolysis pathway and oncogenes YAP1 and FOXD1 was significantly higher in gastric cancer tissues than in normal gastric tissues(P<0.05 for comparisons).Emodin significantly inhibited the protein expressions of key glycolytic enzymes HK2 and oncogenes YAP1 and FOXD1(P<0.05 for comparisons).With supplement of exogenous glycolytic metabolite pyruvate,the protein expressions of oncogenes YAP1 and FOXD1 significantly increased(P<0.05 for comparisons).Conclusions Emodin has a significant pharmacological inhibitory effect on gastric cancer AGS cells,markedly suppressing their biological phenotype.Emodin not only significantly inhibits the key enzyme HK2 in glycolysis metabolism,but also the protein expressions of oncogenes YAP1 and FOXD1.With the addition of exogenous pyruvate to enhance the glycolytic metabolic pathway,the protein expressions of oncogenes YAP1 and FOXD1 significantly increased.The above results suggest a close association of YAP1 and FOXD1 with glycolytic metabolism.Emodin may inhibit oncogenes YAP1 and FOXD1 through the glycolytic metabolism of gastric cancer AGS cells.

Effects of Polydatin on Proliferation and Apoptosis of Pulmonary Artery Smooth Muscle Cells in Neonatal Rats with Hypoxic Pulmonary Hypertension by Regulating YAP1/TAZ Signaling Pathway / 华中科技大学学报(医学版)

Objective To investigate the effects of polydatin(PD)on the proliferation and apoptosis of pulmonary artery smooth muscle cells(PASMCs)in hypoxic pulmonary hypertension(HPH)neonatal rats,and its mechanism of action.Methods Neonatal rats were randomly separated into six groups:control group,model group,low dose PD group,medium dose PD group,high dose PD group,and high dose PD+Hippo pathway inhibitor(high dose PD+XMU-MP-1)group,with 10 rats in each group.After 2 weeks of hypoxia treatment,the right ventricular systolic blood pressure(RVSP)and right ventricular hypertro-phy index(RVHI)of rats in each group were measured.Hematoxylin-eosin(HE)staining was applied to observe pathological changes in lung tissue,and the percentage of pulmonary artery wall thickness to total thickness(WT)and the percentage of wall area to total area(WA)were calculated.Neonatal rat PASMCs were separated from each group,which were divided into NC group,hypoxia group,low dose PD group,medium dose PD group,high dose PD group,and high dose PD+XMU-MP-1 group.Cell counting kit 8(CCK-8)and 5-ethynyl-2'-deoxyuridine(EdU)were applied to detect cell proliferation.Flow cytometry was applied to detect cell apoptosis.Western blot was applied to detect the expression of Yes-associated protein 1(YAP1),tran-scriptional coactivator with PDZ-binding motif(TAZ),mammalian sterile 20-like kinase 1(MST1),B-cell lymphoma 2(Bcl-2),and Bcl-2 associated protein(Bax)in lung tissue and PASMCs.Results Compared with the control group,the pulmonary artery wall in the model group was significantly thickened,lumen was narrowed,and protein expressions of RVSP,RVHI,WT％,WA％,YAP1,MST1 and TAZ were significantly increased(all P＜0.05).Compared with the model group,pulmonary artery thickening and lumen enlargement were observed in the low,medium and high dose PD groups,and the protein expressions of RVSP,RVHI,WT％,WA％,YAP1,MST1 and TAZ were significantly decreased,which showed a dose-dependent relationship(all P＜0.05).The effect could be reversed by XMU-MP-1.Compared with the NC group,the cell A450nm value,EdU positive rate,the protein expression of YAP1,MST1,TAZ and Bcl-2 in the hydropoxia group were significantly increased.The apoptosis rate and the expression of Bax protein were obviously reduced(all P＜0.05).Compared with the hypoxia group,the cell A450nm value,EdU positive rate,the protein expression of YAP1,MST1,TAZ and Bcl-2 in the low,medium and high dose PD groups were obviously reduced.The apoptosis rate and the expression of Bax were significantly increased,which showed a dose-depend-ent relationship(all P＜0.05).The effect could be reversed by XMU-MP-1.Conclusion PD may inhibit the proliferation of PASMCs in HPH neonatal rats and promote apoptosis by inhibiting YAP1/TAZ signaling pathway.

Metformin:A promising clinical therapeutical approach for BPH treatment via inhibiting dysregulated steroid hormones-induced prostatic epithelial cells proliferation / 药物分析学报

The occurrence of benign prostate hyperplasia(BPH)was related to disrupted sex steroid hormones,and metformin(Met)had a clinical response to sex steroid hormone-related gynaecological disease.How-ever,whether Met exerts an antiproliferative effect on BPH via sex steroid hormones remains unclear.Here,our clinical study showed that along with prostatic epithelial cell(PEC)proliferation,sex steroid hormones were dysregulated in the serum and prostate of BPH patients.As the major contributor to dysregulated sex steroid hormones,elevated dihydrotestosterone(DHT)had a significant positive rela-tionship with the clinical characteristics of BPH patients.Activation of adenosine 5'-monophosphate(AMP)-activated protein kinase(AMPK)by Met restored dysregulated sex steroid hormone homeostasis and exerted antiproliferative effects against DHT-induced proliferation by inhibiting the formation of androgen receptor(AR)-mediated Yes-associated protein(YAP1)-TEA domain transcription factor(TEAD4)heterodimers.Met's anti-proliferative effects were blocked by AMPK inhibitor or YAP1 over-expression in DHT-cultured BPH-1 cells.Our findings indicated that Met would be a promising clinical therapeutic approach for BPH by inhibiting dysregulated steroid hormone-induced PEC proliferation.

Bone morphogenetic protein 7 attenuates renal fibrosis in diabetic kid-ney disease rats by down-regulating Ajuba / 中国病理生理杂志

AIM:Bone morphogenetic protein 7(BMP7)reduces the expression of Yes-related protein 1(YAP1)by down-regulating Ajuba level and decreasing extracellular matrix(ECM)deposition.This study aimed to inves-tigate the influence of these factors on modifying the degree of renal fibrosis in rats with diabetic nephropathy.METH-ODS:Eighteen Sprague-Dawley(SD)rats were randomly divided into three groups:the normal control(NC)group,the diabetes mellitus(DM)group,and the DM group treated with BMP7 overexpressing adeno-associated virus(DM+rAAV-BMP7).Each group consisted of six rats.Diabetic kidney disease(DKD)was established in the DM and DM+rAAV-BMP7 groups by injecting 55 mg/kg streptozotocin(STZ)via the tail vein.NRK-52E cells were divided into three groups:the normal glucose(NG)group,the high glucose(HG)group,and the high glucose group treated with recombinant hu-man BMP7(HG+rhBMP7)group.Pathological changes in renal tissues were observed using hematoxylin and eosin(HE)and Sirius red staining.Immunohistochemical staining was performed to examine the expression sites of Ajuba and YAP1 in the renal cortex.Western blot analysis was conducted to determine the expression levels of BMP7,Ajuba,YAP1,colla-gen type Ⅲ(Col-Ⅲ),and fibronectin(FN)in the rat renal cortex and NRK-52E cells.RT-qPCR was used to measure the mRNA levels of Ajuba and YAP1 in the rat renal cortex.RESULTS:Biochemical indices revealed significantly ele-vated levels of blood glucose,serum creatinine,triglycerides,total cholesterol,and 24-hour urinary protein in the DM group compared to the NC group(P<0.05).In the DM+rAAV-BMP7 group,the levels of serum creatinine,24-hour uri-nary protein,triglycerides,and total cholesterol were lower than those in the DM group(P<0.05).Pathological staining demonstrated that the renal interstitium of the DM group exhibited inflammatory cell infiltration,fibrous tissue,collagen fi-ber deposition,disordered renal tubule arrangement,atrophy,and vacuolar degeneration,which were ameliorated in the DM+rAAV-BMP7 group.Immunohistochemistry revealed that Ajuba and YAP1 were mainly expressed in the cytoplasm and nucleus,with high expression in the cytoplasm of the DM group,which was significantly decreased in the DM+rAAV-BMP7 group.Western blot results indicated that the protein levels of FN,Col-Ⅲ,Ajuba,and YAP1 were up-regulated in the DM and the HG groups(P<0.05),but significantly down-regulated in the DM+rAAV-BMP7 group(P<0.05).RT-qP-CR results demonstrated that the mRNA levels of Ajuba and YAP1 were higher in the DM group and significantly lower in the DM+rAAV-BMP7 group(P<0.05).CONCLUSION:The overexpression of BMP7 can ameliorate renal fibrosis in rats with DKD.This effect is likely mediated by the down-regulation of Ajuba,reduction of YAP1 expression,and subse-quent inhibition of ECM deposition.

Effects of erianin on the apoptosis of ovarian granulosa cells in rats with polycystic ovary syndrome and its mechanism / 中国药房

OBJECTIVE To investigate the effects of erianin （ERI） on the apoptosis of ovarian granulosa cells in rats with polycystic ovary syndrome （PCOS） and its mechanism. METHODS PCOS rat model was constructed by subcutaneous injection of dehydroepiandrosterone， and the successfully constructed rats were randomly divided into PCOS group， ERI low-dose， medium- dose and high-dose groups （10， 20， 40 mg/kg） and ERI high dose + verteporfin group （40 mg/kg ERI + 10 mg/kg verteporfin）， with 10 rats in each group. Another 10 normal rats were selected as the normal group. Rats in each administration group were given corresponding dose of ERI and/or intraperitoneal injection of vitiporfin， and rats in the PCOS group and normal group were orally administered an equal volume of 1% dimethyl sulfoxide， once a day， for 6 consecutive weeks. After administration， the body weight， fasting blood glucose （FPG）， serum levels of estradiol （E2）， testosterone （T）， follicle stimulating hormone （FSH） and luteinizing hormone （LH） were detected in each group； morphological changes in ovarian tissue were observed， and the apoptosis of ovarian tissue cells was analyzed. Apoptosis-associated proteins [B-cell lymphoma-2 （Bcl-2）， Bcl-2 associated X protein （Bax）， Caspase-3] and Hippo-YAP signaling pathway associated proteins [large tumor suppressor kinase 1 （LATS1）， phosphorylated LATS1 （p-LATS1） and Yes associated protein （YAP）， phosphorylated YAP （p-YAP）， transcriptional co-activator with PDZ binding motif （TAZ）] were detected in ovarian tissue. RESULTS Compared with PCOS group， the ovarian polycystic characteristics of the ERI low-dose， medium-dose，and high-dose groups were reduced， the number of atretic follicles was reduced， and the granulosa cell layer was thickened； the body mass， FPG， T， LH， LH/FSH， the number of cystic follicles， cell apoptosis index， protein expressions of Bax， Caspase-3， p-LATS1 and p-YAP were greatly decreased （P＜0.05）； the number of corpus luteum， protein expressions of E2， Bcl-2， LATS1， YAP and TAZ were greatly increased （P＜0.05）. Compared with ERI high-dose group， the above indexes in ERI high-dose + vitiporfin group were inhibited （P＜0.05）. CONCLUSIONS ERI can promote the proliferation of ovarian granulosa cells and improve the level of sex hormones in PCOS rats， and its mechanism of action may be related to the inhibition of the Hippo-YAP signaling pathway.

Effects of erianin on the apoptosis of ovarian granulosa cells in rats with polycystic ovary syndrome and its mechanism / 中国药房

OBJECTIVE To investigate the effects of erianin （ERI） on the apoptosis of ovarian granulosa cells in rats with polycystic ovary syndrome （PCOS） and its mechanism. METHODS PCOS rat model was constructed by subcutaneous injection of dehydroepiandrosterone， and the successfully constructed rats were randomly divided into PCOS group， ERI low-dose， medium- dose and high-dose groups （10， 20， 40 mg/kg） and ERI high dose + verteporfin group （40 mg/kg ERI + 10 mg/kg verteporfin）， with 10 rats in each group. Another 10 normal rats were selected as the normal group. Rats in each administration group were given corresponding dose of ERI and/or intraperitoneal injection of vitiporfin， and rats in the PCOS group and normal group were orally administered an equal volume of 1% dimethyl sulfoxide， once a day， for 6 consecutive weeks. After administration， the body weight， fasting blood glucose （FPG）， serum levels of estradiol （E2）， testosterone （T）， follicle stimulating hormone （FSH） and luteinizing hormone （LH） were detected in each group； morphological changes in ovarian tissue were observed， and the apoptosis of ovarian tissue cells was analyzed. Apoptosis-associated proteins [B-cell lymphoma-2 （Bcl-2）， Bcl-2 associated X protein （Bax）， Caspase-3] and Hippo-YAP signaling pathway associated proteins [large tumor suppressor kinase 1 （LATS1）， phosphorylated LATS1 （p-LATS1） and Yes associated protein （YAP）， phosphorylated YAP （p-YAP）， transcriptional co-activator with PDZ binding motif （TAZ）] were detected in ovarian tissue. RESULTS Compared with PCOS group， the ovarian polycystic characteristics of the ERI low-dose， medium-dose，and high-dose groups were reduced， the number of atretic follicles was reduced， and the granulosa cell layer was thickened； the body mass， FPG， T， LH， LH/FSH， the number of cystic follicles， cell apoptosis index， protein expressions of Bax， Caspase-3， p-LATS1 and p-YAP were greatly decreased （P＜0.05）； the number of corpus luteum， protein expressions of E2， Bcl-2， LATS1， YAP and TAZ were greatly increased （P＜0.05）. Compared with ERI high-dose group， the above indexes in ERI high-dose + vitiporfin group were inhibited （P＜0.05）. CONCLUSIONS ERI can promote the proliferation of ovarian granulosa cells and improve the level of sex hormones in PCOS rats， and its mechanism of action may be related to the inhibition of the Hippo-YAP signaling pathway.

AAMP Promotes Osteosarcoma Cell Metastasis by Regulating YAP Signaling Pathway / 肿瘤防治研究

Objective To determine the role of AAMP in osteosarcoma cells and explore the mechanism of invasion and metastasis of osteosarcoma cells regulated by AAMP through the YAP signaling pathway. Methods Public sequencing data analysis was used to explore the correlation between AAMP and osteosarcoma. q-PCR, Western blot, and immunohistochemistry were used to detect the expression levels of osteosarcoma cell-related molecules. CCK-8 was used to detect cell proliferation ability. Transwell and wound healing assays were used to detect the invasive and metastatic abilities of osteosarcoma cells. Immunofluorescence was used to detect the cell localization and expression levels of related molecules. Results High expression of AAMP was negatively correlated with the prognosis of patients with osteosarcoma (P<0.05), and the expression of AAMP in patients with metastatic osteosarcoma increased (P<0.05). Compared with the control group, the AAMP interference group showed significantly decreased migratory, invasive, and EMT activities (P<0.05). The expression of p-CFL1 reduced after the knockdown of AAMP, and the cell plate pseudopods decreased significantly (P<0.05). A positive correlation was found between the expression levels of AAMP and YAP in osteosarcoma cells (P<0.05). Interfering with YAP expression can affect the migration and invasion of osteosarcoma cells. Conclusion AAMP promotes osteosarcoma cell metastasis by regulating the YAP signaling pathway, suggesting that AAMP may be a key molecule in promoting invasion and metastasis of osteosarcoma.

Effects of formononetin on the apoptosis of intestinal epithelial cells in rats with inflammatory bowel disease by regulating the Hippo/YAP signaling pathway / 中国药房

OBJECTIVE To investigate the effects of formononetin （FMN） on the apoptosis of intestinal epithelial cells in inflammatory bowel disease （IBD） rats and its possible mechanism. METHODS IBD rat model was constructed by using trinitrobenzene sulfonic acid （TNBS） induction. Forty-eight rats with successful modeling were divided into model group （normal saline）， low-dose and high-dose FMN groups （20 and 40 mg/kg FMN）， and high-dose FMN+YAP inhibitor Verteporfin （VTPF） group （40 mg/kg FMN+10 mg/kg VTPF）， with 12 rats in each group. Another 12 rats were set as the normal group （normal saline）. They were given drug/normal saline， once a day， for 7 consecutive days. After the last administration， the disease activity index （DAI） of rats was calculated， and the colon length of rats in each group was measured. The pathological changes in the colon tissue of rats were observed. The levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6） and IL-10 in serum were detected， and the apoptosis of intestinal epithelial cells was detected. The expressions of Yes associated protein （YAP）， cleaved cysteine-containing aspartate proteolytic enzyme 3 （cleaved-caspase-3）， B-cell lymphoma-2 （Bcl-2） and Bcl-2 associated X protein （Bax） were detected in colon tissue of rats. RESULTS Compared with the normal group， DAI score， the levels of TNF-α and IL- 6， the apoptotic rate of intestinal epithelial cells， and the expressions of cleaved-caspase-3 and Bax protein in the model group were increased greatly （P＜0.05）； the length of the colon was greatly decreased （P＜0.05）， and the serum level of IL-10 and the protein expressions of YAP and Bcl-2 were greatly reduced （P＜0.05）. The cell morphology of colon tissue was abnormal， with disordered arrangement and inflammatory cell infiltration. Compared with IBD group， the above indexes of rats were improved significantly in low-dose and high-dose FMN groups （P＜0.05）， in dose-dependent manner （P＜0.05）. VTPF significantly alleviated the effects of FMN on the above indexes of IBD rats （P＜0.05）. CONCLUSIONS FMN may promote the expression of YAP by inhibiting the Hippo/YAP signaling pathway， thereby inhibiting apoptosis of intestinal epithelial cells in IBD rats.

Effects of ligustilide on chemotherapy resistance of cervical cancer cells / 中国药房

OBJECTIVE To investigate the effects of ligustilide on chemotherapy resistance of cervical cancer cells based on Hippo-Yes-associated protein （YAP） signaling pathway. METHODS Human cervical cancer cisplatin-resistant cells HeLa/DDP were divided into control group， cisplatin group （10 μmol/L cisplatin）， cisplatin+ligustilide low-， medium- and high-concentration groups （10 μmol/L cisplatin+25， 50， 100 μmol/L ligustilide）. The proliferation， apoptosis， migration and invasion of HeLa/DDP cells were all detected in each group. The mRNA expressions of YAP and transcriptional coactivator with PDZ binding motif （TAZ） as well as the protein expressions of YAP， TAZ， matrix metalloproteinase 2 （MMP2）， Ki67， cleaved-caspase-3 and caspase-3 were determined in HeLa/DDP cells. RESULTS Compared with control group， the inhibitory rate， apoptotic rate and cleaved- caspase-3/caspase-3 of cisplatin group were increased significantly； scratch healing rate， the number of invasive cells， the mRNA expressions of YAP and TAZ， and the protein expressions of YAP， TAZ， MMP2 and Ki67 were decreased significantly in cisplatin group （P＜0.05）. Compared with cisplatin group， the inhibitory rate of cell proliferation， apoptotic rate and cleaved-caspase-3/ caspase-3 were further increased in cisplatin+ligustilide low-， medium- and high-concentration groups， while scratch healing rate， the number of invasive cells， the mRNA expressions of YAP and TAZ， and the protein expressions of YAP， TAZ， MMP2 and Ki67 were further decreased， in a dose-dependent manner （P＜0.05）. CONCLUSIONS Ligustilide can increase the sensitivity of drug-resistant cervical cancer cells to cisplatin by inhibiting Hippo-YAP signaling pathway.

Study on mechanism of Hippo/YAP pathway inhibition by salvianolic acid B in HuH-7 cells / 中国药理学通报

Aim To investigate whether salvianolic acid B ( Sal B) has inhibitory effect on hepatoma HuH- 7 cells and explore whether it works via Hippo/YAP signaling pathway. Methods HuH-7 cells were induced by TGF-β1 (9 pmol · L

Effect of lidocaine regulating the Hippo-YAP signaling pathway on ischemia-reperfusion injury in orthotopic liver transplantation rats / 中国比较医学杂志

Objective To explore the effect of lidocaine(LID)on ischemia-reperfusion injury in orthotopic liver transplantation(OLT)rats and to analyze its mechanism of action.Methods Sixty rats were randomly divided into Verteporfin group,high-dose LID(High LID),medium-dose LID(Medium LID),low-dose LID(Low LID),Model and Control groups,on average.The rest of the rats except the control rats were used to establish OLT models.Observe the pathological changes in liver tissue were with hematoxylin-eosin staining.Serum aspartate transaminase(AST),total bilirubin(TBIL),lactate dehydrogenase(LDH)activities and alanine transaminase(ALT)were detected.Measure liver tissue levels of proinflammatory factors tumor necrosis factor-α(TNF-α),interleukin(IL)-6,IL-1β,and IL-10 with enzyme-linked immunosorbent assays.Reactive oxygen species(ROS)was detected by a fluorescence probe.Malondialdehyde(MDA)was detected by the thiobarbituric acid colorimetric method.Superoxide dismutase(SOD)was detected by nitrogen blue tetrazole colorimetry.Glutathione peroxidase(GSH-Px)was detected by a spectrophotometry method.Apoptosis of liver histiocytes was detected by in situ end labeling.Detect the expression of mammalian STE20 like protein kinase(MST1),phosphorylation(p)-MST1,large tumor suppressor factor 1(LATS1),p-LATS1,Yes associated protein(YAP),p-YAP,and apoptosis-related proteins B-cell lymphoma 2(Bcl-2)and Bcl-2 related X protein(Bax)with Western blot.Results Compared with the Control group,liver tissue in Model group rats showed injury,liver cell necrosis,and a large degree of inflammatory cell infiltration.Moreover,the cell apoptosis rate;serum AST,ALT,TBIL,and LDH activities;and liver tissue levels of TNF-α,IL-6,IL-1β,MDA,ROS,and Bax were significantly increased.Furthermore,liver tissue levels of IL-10,SOD,GSH-Px,Bcl-2,p-MST1/MST1,p-LATS1/LATS1,and p-YAP/YAP proteins were significantly reduced(P＜0.05).Compared with the Model group,liver tissue injury was reduced in Low LID,Medium LID,and High LID groups.The cell apoptosis rate;serum AST,ALT,TBIL,and LDH activities;and liver tissue levels of TNF-α,IL-6,1L-1β,MDA,ROS,and Bax were significantly reduced.Moreover,liver tissue levels of IL-10,SOD,GSH-Px,Bcl-2,p-MST1/MST1,p-LATS1/LATS1,and p-YAP/YAP proteins were significantly increased(P＜0.05).Hippo-YAP signaling pathway inhibitor verteporfin reversed the improving effect of LID on ischemia-reperfusion injury in OLT rats(P＜0.05).Conclusions LID may activate the Hippo-YAP pathway,which reduces the inflammatory response,oxidative stress,and liver cell apoptosis,and improves liver ischemia-reperfusion injury in OLT rats.

The Potential Mechanism of Hippo Signaling Pathway and Its Related miRNA Intervention in Alzheimer’s Disease and Parkinson’s Disease / 生物化学与生物物理进展

The main characteristics of neurodegenerative diseases represented by Alzheimer’s disease (AD) and Parkinson’s disease (PD) is the progressive irreversible loss of neurons, leading to varying degrees of pathological changes and loss of cognitive function. There is still no effective treatment. With the acceleration of global aging society, the incidence of neurodegenerative diseases is rapidly increasing, becoming a serious global public health concern that urgently requires the development of effective therapeutic strategies. The Hippo signaling pathway, a highly evolutionarily conserved pathway, consists of the core components MST1/2, LATS1/2, and downstream effectors, transcriptional co-activators YAP and TAZ. It plays a crucial role in the regulation of various biological processes such as cell proliferation, differentiation, development, and apoptosis. Dysregulation of the Hippo pathway contributes to the development of many diseases, including cancer, cardiovascular diseases, immune disorders, etc. Therefore, targeting the dysregulated components of the Hippo pathway may be an effective strategy for treating various diseases. Increasing evidence indicates that the Hippo pathway is excessively activated in the development of neurodegenerative diseases, manifested by increased expression of MST1 and downregulation of YAP. Stabilizing the Hippo pathway levels has shown improvements in AD and PD. However, most studies on the Hippo pathway in AD and PD focus on changes in the expression levels of Hippo pathway components, and research in other neurodegenerative diseases is still lacking. Therefore, further investigation is needed to fully understand the mechanistic role of the Hippo pathway in neurodegenerative diseases. Meanwhile, miRNA, similarly dysregulated in neurodegenerative diseases and serving as biomarkers, is a primary target for miRNA therapy in neurodegenerative diseases, including AD and PD. Activating or inhibiting dysregulated miRNAs is the main strategy of miRNA therapy during the neurodegenerative disease development. Evidence suggests that the interaction between the Hippo pathway and miRNA can result in widespread biological effects and crosstalk in the occurrence of different types of diseases. However, studies on the interplay between the Hippo pathway and miRNA in neurodegenerative diseases are relatively scarce. In this paper, we predicted the miRNAs related to Hippo pathway through bioinformatics database, and further screened the miRNAs with crosstalk relationship with Hippo signaling pathway through experiments in combination with PubMed. Then, the mechanism of action of Hippo signaling pathway related miRNAs in AD and PD is further elucidated. It is reported that the Hippo pathway and its related miRNA may exert neuroprotective effects by reducing oxidative stress, improving neuroinflammation, stabilizing autophagy levels, maintaining neuronal mitochondrial function, and ameliorating blood-brain barrier dysfunction, thereby delaying the progression of AD and PD. However, research on miRNA directly regulating the Hippo pathway to improve AD and PD is limited, and observations of the Hippo pathway and its related miRNA in other neurodegenerative diseases are scarce. However, considering the regulatory relationship between the Hippo pathway and miRNA in multiple diseases and their respective roles in key mechanisms of neurodegenerative diseases, such as oxidative stress and neuroinflammation, the crosstalk between miRNA and the Hippo pathway holds a crucial regulatory role in the development of neurodegenerative diseases. Thus, the interaction pathways of the Hippo pathway and its related miRNA may be a pivotal avenue for exploring effective therapeutic strategies for neurodegenerative diseases in the future.

Improvement effects and mechanism of sinomenine on non-alcoholic steatohepatitis in mice / 中国药房

OBJECTIVE To investigate the improvement effects of sinomenine （Sin） on non-alcoholic steatohepatitis （NASH） and its potential mechanism. METHODS Male C57BL/6J mice were randomly divided into standard chow diet （SCD） group， high- fat diet （HFD） group， Sin low-dose group （Sin-L group， 50 mg/kg）， and Sin high-dose group （Sin-H group， 100 mg/kg）， with 6 mice in each group. The mice of SCD groups were fed with SCD， and other groups were given HFD for consecutive 24 weeks to establish NASH model. Since 17th week， the mice in each drug group were given corresponding drug solutions intragastrically， once a day， for 8 consecutive weeks. After the last medication， the body weight and liver weight of mice were determined， and liver indexes were calculated. The contents of total cholesterol （TC） and triglyceride （TG） in liver tissue， the serum contents of aspartate aminotransferase （AST）， alanine aminotransferase （ALT）， interleukin-1β （IL-1β） and IL-18 were all determined. Hepatic steatosis and fibrosis were observed， and hepatic lipid droplets were located. The expressions of IL-18 and IL-1β mRNA， inflammation-related proteins （IL-1β， cleaved-IL-1β）， fibrosis-related proteins [collagen Ⅰ （Col-Ⅰ）， α-smooth muscle actin （α-SMA）， connective tissue growth factor （CTGF）]， and pathway-related protein [adenosine monophosphate-activated protein kinase （AMPK）， phosphorylated AMPK （p-AMPK）， Yes-associated protein （YAP）， phosphorylated YAP （p-YAP）] were all determined. HepG2 human liver cancer cells were selected as subjects and divided into control group， oleic acid （OA） group， Sin 50 μmol/L group， Sin 100 μmol/L group， OA+Sin 50 μmol/L group and OA+Sin 100 μmol/L group. After 24 hours of treatment， the accumulation of lipid droplets was observed， and the expressions of pathway-related proteins were detected.RESULTS Compared to HFD group， hepatic steatosis， zengcheng@gdpu.edu.cn fibrotic lesions and lipid droplet accumulation were all alleviated in Sin groups； body weight， liver weight， liver indexes， the contents of AST， ALT， IL-1β， IL-18 in serum and TG， TC in liver tissue， the mRNA expressions of IL-1β and IL-18， and the expressions of cleaved-IL-1β and fibrosis-related proteins all decreased significantly （P＜0.01）； the protein expression of IL-1β， and the phosphorylation levels of AMPK and YAP proteins significantly increased （P＜0.01）. Compared with OA group， the lipid droplet accumulation of cells in OA+Sin groups significantly decreased， while the phosphorylation levels of AMPK and YAP proteins significantly increased （P＜0.05）. CONCLUSIONS Sin can ameliorate the inflammation， lipid deposition and fibrosis of liver tissue in mice， the mechanism of which may be associated with activating the AMPK signaling pathway and promoting YAP phosphorylation.

Fibroblasts inhibit osteogenesis by regulating nuclear-cytoplasmic shuttling of YAP in mesenchymal stem cells and secreting DKK1

BACKGROUND: Fibrous scars frequently form at the sites of bone nonunion when attempts to repair bone fractures have failed. However, the detailed mechanism by which fibroblasts, which are the main components of fibrous scars, impede osteogenesis remains largely unknown. RESULTS: In this study, we found that fibroblasts compete with osteogenesis in both human bone nonunion tissues and BMP2-induced ectopic osteogenesis in a mouse model. Fibroblasts could inhibit the osteoblastic differentiation of mesenchymal stem cells (MSCs) via direct and indirect cell competition. During this process, fibroblasts modulated the nuclear-cytoplasmic shuttling of YAP in MSCs. Knocking down YAP could inhibit osteoblast differentiation of MSCs, while overexpression of nuclear-localized YAP-5SA could reverse the inhibition of osteoblast differentiation of MSCs caused by fibroblasts. Furthermore, fibroblasts secreted DKK1, which further inhibited the formation of calcium nodules during the late stage of osteogenesis but did not affect the early stage of osteogenesis. Thus, fibroblasts could inhibit osteogenesis by regulating YAP localization in MSCs and secreting DKK1. CONCLUSIONS: Our research revealed that fibroblasts could modulate the nuclear-cytoplasmic shuttling of YAP in MSCs, thereby inhibiting their osteoblast differentiation. Fibroblasts could also secrete DKK1, which inhibited calcium nodule formation at the late stage of osteogenesis.

Hippo-YAP signaling pathway regulates autophagy of human periodontal ligament cells under cyclic tensile stress / 华西口腔医学杂志

OBJECTIVES@#This work aimed to investigate the molecular mechanism of cyclic tensile stress (CTS) stimulating autophagy in human periodontal ligament cells (hPDLCs).@*METHODS@#hPDLCs were isolated and cultured from normal periodontal tissues. hPDLCs were loaded with tensile stress by force four-point bending extender to simulate the autophagy of hPDLCs induced by orthodontic force du-ring orthodontic tooth movement. XMU-MP-1 was used to inhibit the Hippo signaling pathway to explore the role of the Hippo-YAP signaling pathway in activating hPDLC autophagy by tensile stress. The expression levels of autophagy-related genes (Beclin-1, LC3, and p62) in hPDLCs were detected by real-time quantitative polymerase chain reaction. Western blot was used to detect the expression levels of autophagy-related proteins (Beclin-1, LC3-Ⅱ/LC3-Ⅰ, and p62) and Hippo-YAP pathway proteins (active-YAP and p-YAP) in hPDLCs. Immunofluorescence was used to locate autophagy-related proteins (LC3-Ⅱand p62) and Hippo-YAP pathway proteins (active-YAP) of hPDLCs.@*RESULTS@#CTS-activated autophagy in hPDLCs and expression of autophagy-related proteins initially increased and then decreased; it began to increase at 30 min, peaked at 3 h, and decreased (P<0.05). CTS increased the expression of active-YAP protein and decreased the expression of p-YAP protein (P<0.05). When XMU-MP-1 inhibited the Hippo-YAP signaling pathway (P<0.05), active-YAP protein was promoted to enter the nucleus and autophagy expression was enhanced (P<0.05).@*CONCLUSIONS@#The Hippo-YAP signaling pathway is involved in the regulation of autophagy activation in hPDLCs under CTS.

Effects of andrographolide on angiogenesis in diabetic foot rats and its mechanism / 中国药房

OBJECTIVE To investigate the effects of andrographolide （Andro） on angiogenesis in rats with diabetic foot and to explore its mechanism of action based on the Hippo-Yes-associated protein （YAP） signaling pathway. METHODS The rat model of type 2 diabetes was established by using low-dose streptozotocin combined with high-fat and high-glucose diet. On the basis of successful modeling， the rat model of diabetes foot was established by scalding. Model rats were randomly divided into 5 groups with 12 rats in each group： model group， Andro low-dose， medium-dose， and high-dose groups （1， 10， and 20 mg/kg）， as well as inhibitor group （20 mg/kg Andro+100 mg/kg of verteporfin， an specific inhibitor of Hippo-YAP signaling pathway）； other 12 healthy rats were included in the Control group. Rats in each group were intragastrically and intraperitoneally injected with solvents or corresponding drugs， once a day， for 2 consecutive weeks. The wound healing， fasting blood glucose （FBG） and fasting insulin （FINS） were detected in rats after medication. HE staining was performed to observe the tissue damage and capillary number of rat trauma； the number of endothelial progenitor cells （EPCs） in peripheral blood of rats was counted by using flow cytometry； the contents of serum total cholesterol （TC）， triglyceride （TG）， high-density lipoprotein cholesterol （HDL-C） and low-density lipoprotein cholesterol （LDL-C） in rats were determined by fully automatic biochemical analyzer； the expressions of hypoxia- inducible factor 1α （HIF-1α）， vascular endothelial growth factor （VEGF） and Hippo-YAP signaling pathway-related proteins in the traumatic tissues of rats in each group were detected by Western blot. RESULTS Compared with Control group， the wound healing rate， capillary number， the proportion of EPCs， HDL-C content， as well as the protein expression levels of HIF-1α and VEGF and the phosphorylation levels of mammalian Ste20-like kinase 1， large tumor suppressor gene 1 and YAP proteins were significantly reduced in the model group， while the FBG， FINS levels and TC， TG and LDL-C contents were significantly increased （P＜0.05）. Compared with model group， the above indexes were significantly reversed in Andro low-dose， medium-dose and high-dose group， in a dose-dependent manner （P＜ 0.05）； verteporfin attenuated the above reversal effect of Andro （P＜0.05）. CONCLUSIONS Andro has the effects of lowering blood glucose and blood lipids， promoting blood vessel formation and wound healing in rats with diabetic foot， and its mechanism of action may be related to the activation of Hippo-YAP signaling pathway.

Isolated ocular colobomas caused by a novel variant of the YAP1 gene / 中华眼底病杂志

Objective:To identify the causative gene and observe the phenotypic characteristics of a family with isolated microphthalmia-anophthalmia-coloboma (MAC).Methods:A retrospective clinical study. One patient (proband) and 3 family members of a family with MAC visited the Henan Eye Hospital from May 2019 to May 2022 were included in the study. The patient's medical history and family history were inquired in detail, and the best corrected visual acuity (BCVA), slit lamp microscope, fundus photography, optical coherence tomography (OCT), ophthalmological B mode ultrasound and axial length (AL) measurement were performed. The peripheral venous blood of the proband, his parents and brother was collected for Trio whole-exome sequencing and pathogenic gene screening. Fluorescence quantitative Polymerase chain reaction was used to verify the suspicious variations. The clinical features of the patient's ocular and systemic also were observed.Results:The proband, male, was 3 years old at the first visit. The horizontal pendular nystagmus was detected in both eyes. Vertical elliptical microcornea and keyhole-shaped iris colobomas were detected in both eyes. The objective refraction at first visit (3 years old) was -4.00 DS/-0.50 DC×105° (OD) and -3.50 DS/-1.25 DC×80° (OS). Refraction and BCVA at 6 years old: -6.50 DS/-2.00 DC×110°→0.05 (OD) and -6.00 DS/-1.50 DC×80°→0.2 (OS). The AL at 4 years and 10 months old was 24.62 mm (OD) and 23.92 mm (OS), respectively. The AL at 5 years and 7 months old was 25.24 mm (OD) and 24.36 mm (OS), respectively. Ultrasonography shows tissue defects in both eyes. Fundus photography showed the inferior choroidal coloboma involving optic disc. OCT showed the optic disc in both eyes was abnormal with colobomas around, and the retinal neurosensory layer in colobomas area was disordered and thin; the retinoschisis was visible in the left eye. The proband's parents and siblings have normal phenotypes. Whole exome sequencing reveals a denovo heterozygous deletion of YAP1 gene: YAP1, chr11: 10280247-102100671, NM_ 001130145, loss 1 (EXON: 6-9). The results of bioinformatics analysis were pathogenic variants. Parents and siblings were of the wild type. Conclusions:Loss of heterozygosity in exons 6-9 of YAP1 gene is the pathogenic variation in this family. It can cause abnormal development of anterior segment, chorioretinal colobomas, deepening of axial myopia, even severe macular colobomas and retinoschisis.

The role of long-chain non-coding RNA LINC-PINT in drug sensitivity of hypoxia induced in head and neck squamous cell carcinoma through the Hippo/YAP signaling pathway / 实用肿瘤学杂志

Objective The aim of this study was to investigate the role of long chain non-coding RNA LINC-PINT in drug sensitivity of hypoxia induced in head and neck squamous cell carcinoma(HNSCC)through the Hippo/Yes-associated protein(YAP)signaling pathway.Methods The proliferative changes of HNSCC cell lines(AGZY-973 cells,HN4 cells,and HN30 cells)and nor-mal human oral keratinocytes(NHOKs)in hypoxic environment were detected by CCK-8 assay;HN30 cells in good condition were taken and set them as the normal group,hypoxia group,hypoxia+LINC-PINT overexpression group,and hypoxia+overexpression negative control group.The expression of LINC-PINT in HN30 cells was detected by qRT-PCR;CCK-8 assay was applied to de-tect the drug sensitivity of HN30 cells,and the effect of cisplatin on proliferation in HN30 cells;cell apoptosis was detected by flow cy-tometry;and Western blot was applied to detect the expression of hypoxia inducible factor-1α(HIF-1α),p-YAP,and YAP protein in HN30 cells.Results Under hypoxia conditions,the proliferative rates of AGZY-973 cells,HN4 cells and HN30 cells were obvi-ously higher than that of NHOKs cells(P＜0.05).Compared with the normal group,the IC50 value,the expression of HIF-1 α and p-YAP/YAP in the hypoxic group were obviously increased in HN30 cells,the rate of apoptosis,the rates of cell growth inhibition at 24 h and 48 h,and the expression of LINC-PINT protein were obviously decreased(P＜0.05);Compared with the hypoxia+overex-pression negative control group,the IC50 values,the expression of HIF-1α and p-YAP/YAP cells in the hypoxia overexpression of LINC-PINT group was obviously reduced in HN30,the rates of apoptosis,the rates of cell growth inhibition at 24 h and 48 h,and the expression of LINC-PINT protein were significantly increased(P＜0.05).Conclusion Overexpression of LINC-PINT can en-hance the hypoxia-induced cisplatin sensitivity in HNSCC,which may be related to the inhibition of the activation of Hippo/YAP sig-naling pathway.

Hydrogen sulfide inhibits apoptosis of cardiomyocytes and ameliorates myocardial fibrosis in rats after myocardial infarction through downregulating Yes-related protein 1 and a transcriptional co-activator with a PDZ-binding motif / 中华老年医学杂志

Objective:To investigate the effects of exogenous hydrogen sulfide on myocardial fibrosis and apoptosis in rats after myocardial infarction and the underlying mechanisms.Methods:Forty-three Sprague Dawley(SD)rats were divided into 4 groups according to the random number table method: a control group(n=12), a myocardial infarction group(MI group, n=13), an hydrogen sulfide(H 2S)group(n=6)and an MI+ H 2S group(n=12). The rat model of acute myocardial infarction was established by intraperitoneal injections of isoproterenol(50 mg/kg, once a day, for 2 days). Electrocardiogram and troponin changes were recorded 48 h after the last drug administration to determine whether the rat model was successfully constructed.After successful establishment of the model, rats in the MI group and the MI+ H 2S group were intraperitoneally injected with sodium hydrosulfide(56 μmol/kg, once a day, for 6 weeks).6 weeks later, echocardiogram and Masson's trichrome staining were performed to assess changes in cardiac function and collagen volume fraction in each group.Terminal deoxynucleotidyl transferase(TdT)dUTP nick end labeling(TUNEL)was used to detect the myocardial apoptosis rate in each group, and Western-blot was used to detect protein expression of Yes-related protein 1(YAP1), WW domain containing transcriptional regulator1(TAZ), mammalian Ste20-like kinase 2(MST2), Bcl-2-associated X protein(Bax), cysteine protease 3(caspase-3), the ratio of matrix metalloproteinase 3(MMP3)/matrix metalloproteinase inhibitor 2(TIMP2), and B-cell lymphoma factor(Bcl-2). Results:Compared with the control group, myocardial collagen volume fraction was increased( P<0.05), the myocardial cell apoptosis rate was increased( P<0.05), and myocardial YAP1, TAZ, MST2, Bax, caspase-3 protein expression and MMP3/TIMP2 ratio were increased in the MI group(all P<0.05), while the expression of Bcl-2 protein was decreased( P<0.05). Compared with the MI group, collagen volume fraction and the cardiomyocyte apoptosis rate were significantly decreased in the MI+ H 2S group( P<0.05). Also, protein expression of YAP1(2.406±0.024 vs.2.830±0.063), TAZ(0.964±0.090 vs.1.329±0.018), MST2(0.780±0.082 vs.1.788±0.097), Bax(1.500±0.008 vs.0.613±0.003)and caspase-3(0.620±0.024 vs.0.780±0.012)and the MMP3/TIMP2 ratio were decreased(all P<0.05), while protein expression of Bcl-2 was increased( P<0.05)in myocardial tissue. Conclusions:H 2S can mitigate myocardial fibrosis after myocardial infarction, through inhibiting the activation of the YAP1/TAZ signaling pathway, thus reducing apoptosis of cardiomyocytes.