Heat shock protein 90 promotes RNA helicase DDX5 accumulation and exacerbates hepatocellular carcinoma by inhibiting autophagy

Objective:Hepatocellular carcinoma(HCC),the main type of liver cancer,has a high morbidity and mortality,and a poor prognosis.RNA helicase DDX5,which acts as a transcriptional co-regulator,is overexpressed in most malignant tumors and promotes cancer cell growth.Heat shock protein 90(HSP90)is an important molecular chaperone in the conformational maturation and stabilization of numerous proteins involved in cell growth or survival.Methods:DDX5 m RNA and protein expression in surgically resected HCC tissues from 24 Asian patients were detected by quantitative real-time PCR and Western blot,respectively.The interaction of DDX5-HSP90 was determined by molecular docking,immunoprecipitation,and laser scanning confocal microscopy.The autophagy signal was detected by Western blot.The cell functions and signaling pathways of DDX5 were determined in 2 HCC cell lines.Two different murine HCC xenograft models were used to determine the function of DDX5 and the therapeutic effect of an HSP90 inhibitor.Results:HSP90 interacted directly with DDX5 and inhibited DDX5 protein degradation in the AMPK/ULK1-regulated autophagy pathway.The subsequent accumulation of DDX5 protein induced the malignant phenotype of HCC by activating theβ-catenin signaling pathway.The silencing of DDX5 or treatment with HSP90 inhibitor both blocked in vivo tumor growth in a murine HCC xenograft model.High levels of HSP90 and DDX5 protein were associated with poor prognoses.Conclusions:HSP90 interacted with DDX5 protein and subsequently protected DDX5 protein from AMPK/ULK1-regulated autophagic degradation.DDX5 and HSP90 are therefore potential therapeutic targets for HCC.

Expression and significance of heat shock protein 70 and glucose-regulated protein 94 in human esophageal carcinoma

AIM: To investigate the expression and significance of heat shock protein 70 (HSP70) and glucose-regulated protein 94 (grp94) in human esophageal carcinoma and adjacent normal tissues.METHODS: The expression of HSP70 and grp94 in 78human esophageal cancer and adjacent normal tissues was studied by immunohistochemistry and pathology photograph analysis.RESULTS: Both esophageal cancer and adjacent normal tissues could express HSP70 and grp94. Of the 78 cases of esophageal carcinoma, 95.0%(72/78) showed positive HSP70, mainly stained in nuclei, while grp94 was mainly stained in cell plasma, and the positive rate was 71.8%(56/78).There was a significant difference in the expression of HSP70 and grp94 between esophageal cancer and adjacent normal tissues (P＜0.01). Compared with adjacent normal tissues, there was a significant difference between differential types and HSP70 expression (P＜0.01).CONCLUSION: HSP70 and grp94 express differently in cell plasma and nuclei. The expression intensity of HSP70is related to the differentiation of esophageal carcinoma.

Expression of heat shock proteins (HSP27, HSP60, HSP70, HSP90, GRP78, GRP94) in hepatitis B virus-related hepatocellular carcinomas and dysplastic nodules

AIM: Expression of heat shock proteins (HSPs) is frequently up-regulated in hepatocellular carcinoma (HCC), which evolves from dysplastic nodule (DN) and early HCC to advanced HCC. However, little is known about the differential expression of HSPs in multistep hepatocarcinogenesis. It was the purpose of this study to monitor the expression of HSPs in multistep hepatocarcinogenesis and to evaluate their prognostic significance in hepatitis B virus (HBV)related HCC.METHODS: Thirty-eight HCC and 19 DN samples were obtained from 52 hepatitis B surface antigen-positive Korean patients. Immunohistochemical and dot immunoblot analyses of HSP27, HSP60, HSP70, HSP90, glucoseregulated protein (GRP)78, and GRP94 were performed and their expression at different stages of HCC development was statistically analyzed.RESULTS: Expression of HSP27, HSP70, HSP90, GRP78, and GRP94 increased along with the stepwise progression of hepatocarcinogenesis. Strong correlation was found only in GRP78 (Spearman's r= 0.802). There was a positive correlation between the expressions of GRP78, GRP94, HSP90, or HSP70 and prognostic factors of HCC. Specifically, the expression of GRP78, GRP94, or HSP90 was associated significantly with vascular invasion and intrahepatic metastasis.CONCLUSION: The expressions of HSPs are commonly up-regulated in HBV-related HCCs and GRP78 might play an important role in the stepwise progression of HBVrelated hepatocarcinogenesis. GRP78, GRP94, and HSP90 may be important prognostic markers of HBV-related HCC, strongly suggesting vascular invasion and intrahepatic metastasis.

Co-expression of heat shock protein 70 and glucose-regulated protein 94 in human gastric carcinoma cell line BGC-823

AIM: To investigate the co-expression and significance of heat shock protein 70 (HSP70) and glucose-regulatedprotein 94 (grp94) in human gastric carcinoma cell line BGC-823.METHODS: The expression and localization of HSP70 and grp94 in human gastric carcinoma cell line BGC-823 were determined by immunocytochemistry and indirect immunofiuorescence cytochemical staining. Flow cytometry was used to analyze the correlation between expression of HSP70, grpg4 and cell cycle in BGC-823 cell line.RESULTS: Gastric cancer cell line BGC-823 expressed high level of HSP70 and grp94. The positive rate of HSP70 and grp94 was 84.9±4.94% and 79.6±5.16%, respectively. Bothof them were stained in cell plasma. There was a significant difference compared with control group (1.9±0.94%,P＜0.01). During the cell cycle, HSP70 and grp94 were continuously expressed in BGC-823.CONCLUSION: HSP70 and grp94 are highly expressed in human gastric carcinoma BGC-823 cells through the whole cell cycle. There is no relationship between expression of HSP70, grp94 and cell cycle.

Heat shock protein 70 chaperoned alpha-fetoprotein in human hepatocellular carcinoma cell line BEL-7402

AIM: To investigate the interaction between heat shock protein 70 (HSP70) and α-fetoprotein (AFP) in human hepatocellular carcinoma (HCC) cell line BEL7402.METHODS: The expression and localization of HSP70 and AFP in human HCC cell line BEL-7402 were determined by immunocytochemistry and indirect immunofluorescence cytochemical staining. The interaction between HSP70 and AFP in HCC cells was analyzed by immunoprecipitation and Western blot.RESULTS: Immunocytochemical staining detection showed that HCC cell BEL-7402 expressed a high level of HSP70 and AFP synchronously. Both were stained in cell plasma.AFP existed in the immunoprecipitate of anti-HSP70 mAb,while there was HSP70 in the immunoprecipitate of antiAFP mAb.CONCLUSION: HSP70 chaperones AFP in human HCCcell BEL-7402. The interaction between HSP70 and AFP in human HCC cell can be a new route to study the pathogenesis and immunotherapy of HCC.

Expressions and clinicopathologic significance of five heat shock proteins in esophageal squamous cell carcinoma

Objective: To investigate the expressions of heat shock protein (hsp) 10, hsp27, hsp60, hsp70 and hsp90α in esophageal squamous cell carcinoma (ESCC) and normal tissues along the incisal margin (TIM), and discuss the clinico- pathologic features about their expressions. Methods: 120 specimens from ESCC and 36 specimens from TIM were made into tissue chips. The presence and the levels of expression of hsp10, hsp27, hsp60, hsp70 and hsp90α were observed on tissue chips by immunohistochemistry EnVisionTM. Their correlations to clinicopathologic features were analyzed. Results: The positive staining rates of hsp10, hsp27, hsp60, hsp70 and hsp90α in ESCC and TIM were 53.8% and 37.5%, 62.0% and 42.1%, 92.7% and 63.2%, 57.9% and 22.2%, 33.7% and 18.5% respectively. There were no statistical significances between the differential expressions of hsp10, hsp27 and hsp90α in ESCC and TIM (P > 0.05), but there were great statistical signifi- cances about hsp60 and hsp70 (P < 0.01). The level of hsp27 declined with the lower grade of differentiation of ESCC (P < 0.05). Except for hsp27, the positive expressions of the other four HSPs had no correlation to the clinicopathologic features of ESCC. Conclusion: The expressions of hsp10, hsp27, hsp60, hsp70 and hsp90α in ESCC and TIM were a common event. The levels of hsp60 and hsp70 in ESCC were higher than those in TIM. The level of hsp27 declined with the lower grade of differentiation of ESCC showed that it may be play a role in the differentiation of ESCC.

Clinical significance of heat shock protein 10 in large bowel carcinoma

Objective:To investigate the expression of heat shock protein 10(HSP10)during genesis and development of large bowel carcinoma and discuss the clinical significance about its expression.Methods:The expression of HSP10 was observed in specimens from normal colonic mucosa(NC),colorectal adenomas(CA)and colorectal adenocarcinomas(CAC) by immunohistochemistry EnVisionTM.Its correlations to clinicopathologic features,as well as to postoperative survival time of large bowel carcinoma patients were analyzed.Results:The expression of HSP10 was common in normal colonic mucosa, colorectal adenomas and adenocarcinomas and more intensive in colorectal adenomas and adenocarcinomas than that in normal colonic mucosa(P<0.001).The positive expression of HSP10 had no correlation to clinicopathologic features,includ- ing age,gender,primary tumor,infiltrating of regional lymph node,metastasis,clinical stage and histopathology of large bowel carcinoma patients,as well as to their postoperative survival time.Conclusion:HSP10 was overexpressed in the early stage of colorectal adenocarcinoma suggesting that it could serve as an index for early diagnosis of large bowl carcinoma.The positive expression of HSP10 had no correlation to clinicopathologic features or postoperative survival time of large bowel carcinoma patients.

Abrogation of heat-shock protein (HSP)70 expression induced cell growth inhibition and apoptosis in human androgen-independent prostate cancer cell line PC-3m

Aim: To investigate the effect of abrogating heat shock protein (HSP) 70 expression by antisense HSP70 oligonucleotides treatment on human androgen-independent prostate cancer cell line PC-3m growth. Methods: PC-3m cells were treated with 0-16 μmol/L antisense HSP70 oligomers for 0-100 hr. Cell growth inhibition was analyzed using a trypan blue dye exclusion test. Apoptotic cells were detected and confirmed by flow cytometric analysis and DNA fragmentation analysis. The protein expression of HSP70 and bcl-2 affected by antisense HSP70 oligomers were determined using Western blot. Results: Antisense HSP70 oligomer induced apoptosis and then inhibited proliferation of PC-3m cells in a dose- and time-dependent manner. Ladder-like patterns of DNA fragments were observed in PC-3m cells treated with 10 μmol/L antisense HSP70 oligomer for 48 hr or 8 μmol/L for 72 hr on agarose gel electrophoresis. Antisense HSP70 oligomer pretreatment enhanced the subsequent induction of apoptosis by heat shock in PC-3m cells. In addition, undetectable HSP70 expression was observed at a concentration of 10 μmol/L antisense HSP70 oligomer treatment for 48 hr or 8 μmol/L for 72 hr in Western blot, which was paralleled by decreased expression levels of anti-apoptotic protein bcl-2. Conclusion: HSP70 antisense oligomer treatment abrogates the expression of HSP70, which may disrupt HSP70-bcl-2-interactions and further down-regulate bcl-2 expression, in turn inducing apoptosis and inhibiting cell growth in PC-3m cells.

Expression of Heat Shock Protein 70 and 27 in Non-small Cell Lung Cancer and Its Clinical Significance

The heat shock proteins （HSPs） 70 and HSP 27 expression in patients with non-small cell lung cancer （NSCLC） was studied and the relation.ship between HSP 70 and HSP 27 with the clinicopathological features of NSCLC was investigated. The expression of HSP 70 and HSP 27 was detected in tumor tissues from 60 patients with NSCLC by S-P immunohistochemistry. The findings were analyzed in combination with the histological types, histopathological differentiation, lymph node metastasis, patients＇ clinical stages, smoking history and gender. The results showed that of the 60 NSCLC tissue specimens studied, the immunoreactivity of HSP 70 and HSP 27 was detected in 47 （78.3 %） and 43 （71.7 %） specimens, respectively. A positive correlation was found between the overexpression of HSP 70 and HSP 27. The histopathological differentiation, lymph node metastasis, clinical stages and smoking history were correlated to the expression of HSP 70, but not to the expression of HSP 27. No statistical significance was observed in histological types and gender with respect to both HSP 70 and HSP 27 expression. It is suggested that the HSP 70 expression is a powerful and significant prognostic indicator and is related to histopathological differentiation, lymph node metastasis, patients＇ clinical stages, smoking history, whereas HSP 27 expression is not.

Purification of heat shock protein 70-associated tumor peptides and their antitumor immunity to hepatoma in mice

AIM:To purify the heat shock protein (HSP) 70-associated tumor peptides and to observe its non-MHC-I molecule restrictive antitumor effect.METHODS:By ConA-sepharose affinity chromatography,ADP-agarose affinity chromatography, and DEAE anion exchange chromatography, we were able to purify HSP70-associated peptides from mouse hepatoma (HCaF) cells treated in heat shock at 42℃. Specific active immunization and adoptive cellular immunization assay were adopted to observe the immunoprotective effect elicited by HSP70-associated peptide complexes isolated from HcaF.RESULTS: The finally purified HSP-associated peptides had a very high purity and specificity found by SDS-PAGE and Western blot. Mice immunized with HSP70-associated peptide complexes purified from HCaF cells were protected from HCaF living cell challenge. This effect was dose dependent.Adoptive immunization of immune spleen cells of mice immunized with HSP70-associated peptide complexes could elicit immunity against HCaF challenge, and the tumor-free mice could resist repeated challenges. This effect could be continuously enhanced by repeated challenge with HCaF living cells. The tumor-free mice could tolerate the challenge for as high as 1×10^7 HCaF cells. The mice immunized once with spleen cells pulsed with HSP70-associated peptide complexes in vitro could also result in a certain adoptive immunity against HCaF.CONCLUSION:High purity and specificity of HSP70-associated peptides could be achieved from tumor cells by the low-pressure affinity chromatography method used in this study. HSP70-associated peptide complexes derived from the HCaF can elicit non-MHC-I molecule restrictive immunoprotective effect against HCaF.This effect can be transferred by adoptive immunization to mice and enhanced by repeated challenge with HCaF live cells.

Sleep deprivation increase the expression of inducible heat shock protein 70 in rat gastric mucosa

AIM To .investigate if sleep deprivation is able to increase the expression of inducible heat shock protein 70 in gastric mucosa and its possible role in mucosal defense.METHODS Rats for sleep disruption were placed inside a computerized rotating drum, gastric mucosa was taken from rats with 1, 3 and 7 d sleep deprivation. RT-PCR,immunohistochemistry and Western blotting were used to determine the expression of heat shock protein 70.Ethanol (500 mL@ L 1, I.g.) was used to induce gastric muceea damage.RESULTS RT-PCR, Western blotting and immunostaining confirmed that the sleep deprivation as a stress resulted in significantly greater expression of inducible heat shock protein 70 in gastric mucosa of rats. After the 500mL@ L-1 ethanol challenge, the ulcer area found in the rats with 7 d sleep deprivation (19.15 ± 4.2) mm2 was significantly lower (P＜0.01) than the corresponding control (53.7 ± 8.1) mm2.CONCLUSION Sleep deprivation as a stress, in addition to lowering the gastric mucosal barrier, is able to stimulate the expression of inducible heat shock protein 70 in gastric mucosa of rats, the heat shock protein 70 may play an important role in gastric mucosal protection.

Effect of Jianpiyiqi Prescription on the Expression of Heat Shock Proteins in Acetic Acid-induced Chronic Gastric Ulcer Rats

To investigate the effect of Jianpiyiqi prescription on the expression of heat shock proteins and the changes of HSPs in gastric ulcerated rats, the rat model of chronic gastric ulcer was induced by acetic acid. The SABC immunohistochemical method was used to observe HSP70 of mucosa around the gastric ulcer. Imaging analysis was performed. Western dot blot was used to detect HSPs contents in the plasma and gastric mucosal homogenate in each group. The results showed that HSP70 contents of the mucosa around the gastric ulcer in the model group and ranitidine treated group were increased as compared with control group . Jianpiyiqi could increase the expression of HSP70 of the mucosa around the gastric ulcer further as compared with that in the model group and ranitidine treated group . The HSP70 contents in the serum and mucosa in the model group and ranitidine treated group were increased as compared with control group ( P <0.01, P <0 05 respectively). HSP70 of serum and mucosa in the Jianpiyiqi treated group was higher than in the model group and ranitidine treated group ( P <0.05, P <0.01 respectively).It was concluded that HSP might play a role in the process of pathophysiology of gastric ulcer. Jianpiyiqi could enhance gastric ulcer healing through the protective mechanism of HSPs.

Correlation between heat shock protein 70 expression in the brain stem and sudden death after experimental traumatic brain injury

Objective:Theaimof thisstudywas to determinethepatternsof heat-shockprotein70(HSP 70 )biosynthesis followingtraumaticbraininjury,andobservetheeffectof HSP 70 inductionon thefunctionof thevitalcenterinthebrain stem.Methods:Ratmodelsof suddendeathresultedformtraumaticbraininjurywereproduced,andHSP 70 expressionin theratbrainstemwasdeterminedby immunohistochemistry,theinductionof HSP 70 mRNAdetectedby RT-PCR.Re -sults: Thelevelof HSP 70 mRNAwasprominentlyelevatedinthebrainstemas earlyas15minfollowingtheimpactin-jury,whileHSP 70 expressionwasonlyobserved3to6h aftertheinjury.Itwasalsoobservedthatthelevelsof HSP 70 mR-NAbutnottheproteinwereelevatedinthebrainstemof suddendeathrats.Conclusion:Thesynthesisof HSP 70 wassig-nificantlyenhancedinthebrainstemfollowingtraumaticinjury,andtheexpressionof HSP 70 is beneficialto eliminatethe stressagents,andto sustainthecellularproteinhomeostasis.Whentheinjurydisturbsthesynthesisof HSP 70 to disarmthe protectivemechanismof heat-shockproteins,dysfunctionof thevitalcenterinthebrainstem,andconsequentlydeathmay occur.Breachinthesynchronizationof HSP 70 mRNA-proteincanbeindicativeof fataldamageto thenervecells.

Heat shock protein 70 expression in relation to apoptosis in primary bladder transitional cell carcinoma

Bladder carcinoma is the most common tumor in the urinary system. In 1996, a sampleinvestigation showed that bladder carcinoma, in which more than 90% was mainly primary bladder transitional cell carcinoma （BTCC）, was one of the ten highest mortality malignant tumors in China. Bladder carcinoma represented 2% of all malignant tumors and has the fifth most common malignancy in men in Europe and North America.

血清DCP、AFP、AFP-L3和HSP90α在原发性肝细胞癌诊断中的应用价值

目的:探讨血清异常凝血酶原(DCP)、甲胎蛋白(AFP)、甲胎蛋白异质体(AFP-L3)、热休克蛋白90α(HSP90α)检测在原发性肝细胞癌(HCC)诊断中的临床应用价值,为临床提供参考。方法:选取2021年10月—2022年6月安徽医科大学附属巢湖医院收治的64例原发性肝癌(PHC)患者作为研究对象,将其中50例HCC患者与同期就诊的30例失代偿期肝硬化患者、30例乙型病毒性肝炎患者及30例健康人群分为HCC组、肝硬化组、肝炎组及健康对照组,检测并比较分析所有研究对象血清DCP、AFP、AFP-L3、HSP90α水平。结果:HCC组血清AFP、AFP-L3、DCP、HSP90α水平及GALAD评分均较其他三组高,差异有统计学意义(H=39.885、28.618、57.601、46.252、85.734,P<0.05)。绘制ROC曲线,AFP、AFP-L3、DCP、HSP90α及GALAD评分的受试者工作特征曲线下面积(AUC)分别为0.804、0.680、0.883、0.838及0.936。GALAD评分联合HSP90α诊断的AUC最高,为0.940。AFP、DCP与HSP90α三者联合可将诊断敏感度提高至84.0%。HBV组AFP、DCP水平及GALAD评分均高于非HBV组,差异有统计学意义(Z=-2.251、-1.668、-2.288,P<0.05),两组AUC值最高者均为GALAD,分别为0.951、0.903。结论:HCC相关肿瘤标志物单项检测敏感性较低,联合检测可明显提高诊断敏感性和准确性,增加HCC筛查阳性率。

原发性肝癌患者高强度聚焦超声治疗前后HSP70、PTN、PIVKA-Ⅱ水平变化及对早期复发的预测效能

目的探究原发性肝癌(PHC)患者在高强度聚焦超声(HIFU)治疗前后热休克蛋白70(HSP70)、多效生长因子(PTN)、异常凝血酶原Ⅱ(PIVKA-Ⅱ)水平变化及对早期复发的影响和预测效能。方法选取2021年1月—2023年1月收治的PHC 120例,均行HIFU治疗,根据术后6个月是否复发分为复发组(26例)与未复发组(94例)。比较2组临床资料、治疗前后血清HSP70、PTN、PIVKA-Ⅱ水平及变化值(ΔHSP70、ΔPTN、ΔPIVKA-Ⅱ值),分析ΔHSP70、ΔPTN、ΔPIVKA-Ⅱ与HIFU治疗后早期复发的相关性,并评价预测效能。结果2组治疗后血清HSP70、PTN、PIVKA-Ⅱ水平较治疗前下降(P<0.05);复发组治疗前、治疗后血清HSP70、PTN、PIVKA-Ⅱ水平高于未复发组(P<0.01)。复发组血清ΔHSP70、ΔPTN、ΔPIVKA-Ⅱ值低于未复发组(P<0.01)。Spearman相关性分析显示,治疗前血清HSP70、PTN、PIVKA-Ⅱ与肿瘤直径、CNLC分期呈正相关,与分化程度呈负相关(P<0.01)。多因素Logistic回归分析显示,校正肿瘤直径、CNLC分期、分化程度因素前后血清ΔHSP70、ΔPTN、ΔPIVKA-Ⅱ值均与PHC患者HIFU治疗后早期复发独立相关(P<0.01);血清ΔHSP70、ΔPTN、ΔPIVKA-Ⅱ值联合预测PHC患者HIFU治疗后早期复发的曲线下面积为0.916(95%CI:0.851,0.958),敏感度为0.923,特异度为0.872,优于三者单独预测。结论PHC患者HIFU治疗前后HSP70、PTN、PIVKA-Ⅱ水平变化值与早期复发独立相关,且联合预测效能较高。

Abnormal expression of HSP gp96 associated with HBV replication in human hepatocellular carcinoma

BACKGROUND: Heat shock protein (HSP) gp96 is a member of the HSP90 family and presumably overexpresses as a result of stimulation by mutated or abnormal proteins. Its abnormal expression correlates with carcinogenesis, progression and prognosis of hepatocellular carcinoma (HCC). In this study, we investigated the pathological characteristics of liver gp96 expression and its relationship with hepatitis B virus (HBV) replication in HCC patients. METHODS: Tumor specimens were prospectively collected from 30 HCC patients undergoing liver resection. Total RNAs were extracted from HCC or their noncancerous tissues. The distribution of gp96 expression in hepatocytes was investigated by streptavidin peroxidase (S-P) immunohistochemistry and tissue HBV-DNA was detected by the in situ molecular hybridization technique. The association of gp96 expression with HBV replication, and the histopathological characteristics of HCC were analyzed. RESULTS: The gp96 was strongly expressed in HCC (73.3%, 22 of 30) and weakly (46.7%, 14 of 30) in non-cancerous tissues. The gp96 expression in HCC tissues was correlated with degree of tumor differentiation and tumor size, but not with tumor number (P>0.05). Immunohistochemical analysis showed that 17 of 19 HCC patients with HBVDNA -positive were strongly expressed for gp96, whereas only 5 of 11 patients with HBV-DNA-negative were positive for gp96. A significant difference was found between the two groups (89.5% vs. 45.5%, P<0.05). CONCLUSIONS: The abnormal expressions of HSP gp96 in HCC tissues are associated with HBV replication. This finding indicates that HBV infection plays an important role in the development of HCC.

Effects of HSP70 Antisense Oligonucleotide on the Proliferation and Apoptosis of Human Hepatocellular Carcinoma Cells

The study investigated the effects of heat shock protein 70(HSP70) antisense oligonucleotide(ASODN) on the proliferation and apoptosis of a human hepatocellular carcinoma cell line(SMMC-7721 cells) in vitro.HSP70 oligonucleotide was transfected into SMMC-7721 cells by the mediation of SofastTM transfection reagent.Inhibition rate of SMMC-7721 cells was determined by using MTT method.Apoptosis rate and cell cycle distribution were measured by flow cytometry.Immunocytochemistry staining was used to observe the expression of HSP70,Bcl-2 and Bax.The results showed that HSP70 ASODN at various concentrations could significantly inhibit the growth of SMMC-7721 cells,and the inhibition effect peaked 48 h after transfection with 400-nmol/L HSP70 ASODN.Cytometric analysis showed the apoptotic rate was increased in a dose-and time-dependent manner in the HSP70 ASODN-treated cells.The percentage of cells in the G2/M and S phases was significantly decreased and that in the G0/G1 phase increased as the HSP70 ASODN concentration was elevated and the exposure time prolonged.Immunocytochemistry showed that treatment of SMMC-7721 cells with HSP70 ASODN resulted in decreased expressions of HSP70 and Bcl-2 proteins,and an increased expression of Bax protein.It was concluded that the HSP70 ASODN can inhibit the growth of the SMMC-7721 cells and increase cell apoptosis by down-regulating the expression of HSP70.HSP70 ASODN holds promise for the treatment of hepatocellular carcinoma.

A novel intronic circular RNA,circGNG7,inhibits head and neck squamous cell carcinoma progression by blocking the phosphorylation of heat shock protein 27 at Ser78 and Ser82

Background:There is increasing evidence that circular RNAs(circRNAs)play a significant role in pathological processes including tumorigenesis.In contrast to exonic circRNAs,which are the most frequently reported circRNAs in cancer so far,the studies of intronic circRNAs have been greatly lagged behind.Here,we aimed to investigate the regulatory role of intronic circRNAs in head and neck squamous cell carcinoma(HNSCC).Methods:We conducted whole-transcriptome sequencing with four pairs of primary tumor tissues and adjacent normal tissues from HNSCC patients.Then,we characterized circGNG7 expression in HNSCC tissues and cell lines and explored its association with the prognosis of HNSCC patients.We also identified interactions between circGNG7 and functional proteins,which alter downstream signaling that regulate HNSCC progression.Results:In this study,we identified a new intronic circRNA,circGNG7,and validated its functional roles in HNSCC progression.CircGNG7 was predominately localized to the cytoplasm,and its expression was downregulated in both HNSCC tissues andCAL27,CAL33,SCC4,SCC9,HN6,and HN30 cells.Low expression of circGNG7 was significantly correlated with poor prognosis in HNSCC patients.Consistent with this finding,overexpression of circGNG7 strongly inhibited tumor cell proliferation,colony formation,in vitro migration,and in vivo tumor growth.Mechanistically,the expression of circGNG7 in HNSCC cells was regulated by the transcription factor SMAD family member 4(SMAD4).Importantly,we discovered that circGNG7 could bind to serine residues 78 and 82 of the functional heat shock protein 27(HSP27),occupying its phosphorylation sites and hindering its phosphorylation,which reduced HSP27-JNK/P38 mitogen-activated protein kinase(MAPK)oncogenic signaling.Downregulation of circGNG7 expression in HNSCC increased HSP27-JNK/P38 MAPK signaling and promoted tumor progression.Conclusions:Our results revealed that a new intronic circRNA,circGNG7,functions as a strong tumor suppressor and that circGNG7/HSP27-JNK/P38 MAPK signaling is a novel mechanism by which HNSCC progression can be controlled.

Enhanced sensitivity to mitomycin C by abating heat shock protein 70 expression in human bladder cancer cell line of BIU-87

Background Bladder cancer is a relatively common tumor in the urinary system, in which mitomycin C （MMC）-based chemotherapy or combination chemotherapy has been mainly used to treat patients with advanced bladder cancer. The prognosis of patients with advanced bladder cancer is still extremely poor in spite of recent therapeutic advances. To improve the prognosis, the sensitivity of tumor cells to mitomycin C by the induction of apoptosis with the abating heat shock protein 70 （HSP70） expression in human bladder cancer cell lines of BIU-87 was investigated.Methods I-ISPT0 expression was abated in BIU-87 cells by HSV mI^NA anhsense ongomers, lnl i assay at,u the clone-forming test were used for evaluating the sensitivity of cells to MMC. Apoptosis was assessed using both fluorescent microscopy after staining the cells with Hoechst 33258 and DNA fragment ladder agarose electrophoresis. Thirty-two male six-week-old BALB/c nude mice, at the beginning of the experiment, were used to evaluate the effect of antisense oligomers （ASO） on the tumor formation in vivo.Results HSP70 expression in BIU-87 was effectively abated by HSP70 mRNA antisense oligomers. The percentage of apoptotic cells in ASO group was greater than in sense oligomers （SO） [P 〈0. 05, （18.31 ±2. 89）% vs （1.89±0.74）%], nonsense oligomers （NO） [P 〈0.05, （18.31±2.89）% vs （1.78±0.92）%] and blank groups [P〈0.05, （18.31 ±2.89）% vs （1.87±0.84）%], while the sensitivity of tumor cells to mitomycin C was enhanced. The in vivo tumor inhibition rate of ASO plus MMC （ 〉 50% ） was more than that of ASO or MMC group alone （ all P 〈 0. 05 ）.Conclusions The abating level of HSP70 expression can strengthen the sensitivity of BIU-87 to MMC. One of this effect might be related to the induction of apoptosis by abating HSP70 expression.