ER stress and ER stress-induced apoptosis are activated in gastric SMCs in diabetic rats

AIM: To investigate the gastric muscle injury caused by endoplasmic reticulum (ER) stress in rats with diabetic gastroparesis. METHODS: Forty rats were randomly divided into two groups: a control group and a diabetic group. Diabetes was induced by intraperitoneal injection of 60 mg/kg of streptozotocin. Gastric emptying was determined at the 4^th and 12^th week. The ultrastructural changes in gastric smooth muscle cells (SMCs) were investigated by transmission electron microscopy. TdT-mediated dUTP nick end labeling (TUNEL) assay was performed to assess apoptosis of SMCs. Expression of the ER stress marker, glucose-regulated protein 78 (GRP78), and the ER-specific apoptosis mediator, caspase-12 protein, was determined by immunohistochemistry. RESULTS: Gastric emptying was significantly lower in the diabetic rats than in the control rats at the 12^th wk (40.71% ± 2.50%, control rats vs 54.65% ± 5.22%, diabetic rats; P < 0.05). Swollen and distended ER with an irregular shape was observed in gastric SMCs in diabetic rats. Apoptosis of gastric SMCs increased in the diabetic rats in addition to increased expression of GRP78 and caspase-12 proteins. CONCLUSION: ER stress and ER stress-mediated apoptosis are activated in gastric SMCs in diabetic rats with gastroparesis.     

Transient Knock Down of Grp78 Reveals Roles in Serum Ferritin Mediated Pro-inflammatory Cytokine Secretion in Rat Primary Activated Hepatic Stellate Cells

Chronic liver diseases, including cancer, are characterized by inflammation and elevated serum ferritin (SF).However, the causal-relationship remains unclear. This study used primary rat hepatic stellate cells (HSC) as amodel to investigate effects of physiological SF concentrations (10, 100 and 1000 pM) because HSCs play a centralrole in the development and progression of liver fibrosis. Physiological concentrations of SF, either horse SF orhuman serum, induced pro-inflammatory cytokine IL1β, IL6 and TNFα secretion in rat activated HSCs (allp<0.05). By contrast, treatment did not alter activation marker αSMA expression. The presence of SF markedlyenhanced expression of Grp78 mRNA (p<0.01). Furthermore, transient knock down of Grp78 by endotoxin EGFSubAabolished SF-induced IL1β and TNFα secretion in activated HSCs (all p<0.05). In conclusion, our resultsshowed that at physiological concentrations SF functions as a pro-inflammatory mediator in primary rat HSCs.We also provide a molecular basis for the action of SF and identified Grp78-associated ER stress pathways as anovel potential therapeutic target for resolution of fibrosis and possible prevention of liver cancer.     

GRP94在HBV感染的人肝细胞肝癌中的表达及意义

目的:探讨葡萄糖调节蛋白(glucose-regulated protein 94,GRP94)在乙型肝炎病毒(hepatitis B virus,HBV)感染的人肝细胞肝癌(human hepatocellular carcinomas,HCC)中的表达及其意义。方法:酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)和实时荧光定量PCR（real-time fluorescence quantitative polymerase chain reaction,RTQF-PCR）方法来检测HCC患者血清。采用免疫组织化学(immunohistochemistry,IHC)方法、蛋白质印迹(Western blot)方法及半定量逆转录聚合酶链反应(reverse transcrition-polymerase chain reaction,RT-PCR),来探讨GRP94在HBV病毒感染的HCC和癌旁组织中的表达。结果:GRP94在癌旁组织中低表达,而在HCC中高表达,两者在Western blot、IHC及RT-PCR中的表达,其差异有统计学意义。同时HCC中GRP94的表达强度与肝癌患者肿瘤TNM分期(P=0.02)、肿瘤大小(P=0.01)、肿瘤结节的数目(P=0.02)、HBV-DNA的拷贝数值(P＜0.01)和血清甲胎蛋白（alpha-fetal protein,AFP）水平(P=0.02)相关,但与患者的性别、年龄差异无关。结论:GRP94与HCC临床病理特征存在一定的相关性,对HCC治疗、预后判定具有一定指导意义。     

GRP78 Secreted by Colon Cancer Cells Facilitates Cell Proliferation via PI3K/Akt Signaling

Glucose regulated protein 78 (GRP78) is usually recognized as a chaperone in the endoplasmic reticulum. However, increasing evidence indicates that GRP78 can be translocated to the cell surface, acting as a signaling receptor for a variety of ligands. Since little is known about the secretion of GRP78 and its role in the progression of colon cancer we here focused on GRP78 from colon cancer cells, and purified GRP78 protein mimicking the secreted GRP78 was able to utilize cell surface GRP78 as its receptor, activating downstream PI3K/Akt and Wnt/β-catenin signaling and promote colon cancer cell proliferation. Our study revealed a new mode of action of autocrine GRP78 in cancer progression: secreted GRP78 binds to cell surface GRP78 as its receptor and activates intracellular proliferation signaling.     

特异性下调GRP78基因表达对卵巢癌细胞耐药性的影响

目的:探讨RNA干扰技术沉默葡萄糖调节蛋白78（GRP78）基因对人卵巢癌细胞耐药性的影响,阐明GRP78基因沉默逆转肿瘤细胞耐药性的生物学机制。方法：构建pSilencerTM3.0-H1-GRP78 siRNA重组质粒,脂质体介导转染至SKOV3/DDP细胞;RT-PCR和Western blotting法检测GRP78基因的蛋白的表达;Western blotting法检测caspase-4和caspase-3蛋白表达;流式细胞术检测细胞凋亡率。结果：转染GRP78 siRNA重组质粒的SKOV3/DDP细胞GRP78基因和蛋白表达较转染空质粒组明显降低(P<0.05);加用顺铂后,转染GRP78 siRNA重组质粒细胞组较未转染GRP78 siRNA重组质粒细胞组caspase-4和caspase-3表达明显增加(P<0.05),细胞凋亡率也明显增加(P<0.05)。结论：抑制GRP78基因表达能通过上调caspase-4和caspase-3表达及增加顺铂诱导的SKOV3/DDP细胞凋亡率,降低SKOV3/DDP细胞对顺铂的耐药性。     

Down‐regulation of BiP/GRP78 sensitizes resistant prostate cancer cells to gene‐therapeutic overexpression of REIC/Dkk‐3

We have recently shown that an adenovirus carrying REIC/Dkk‐3 (Ad‐REIC) exhibits a potent tumor‐specific cell‐killing function for various human cancers. It has also become evident that some human cancers are resistant to Ad‐REIC‐induced apoptosis. The aim of the present study was to determine the molecular mechanisms of resistance to Ad‐REIC. First, we isolated resistant clones from a human prostate cancer cell line, PC3, after repeated exposure to Ad‐REIC. Infection efficiency of the adenovirus vector and expression level of REIC/Dkk‐3 in the resistant clones were similar to those in the parental PC3 cells. By screening for alteration in levels and functional status of proteins involved in Ad‐REIC‐induced apoptosis, we found that BiP/GRP78, an ER‐residing chaperone protein, was expressed at higher levels consistently among resistant cells. Expression levels of BiP and rates of apoptosis induced by Ad‐REIC were inversely correlated. Down‐regulation of BiP with siRNA sensitized the resistant cells to Ad‐REIC in vivo as well as in culture. These results indicate that BiP is a major determinant of resistance to Ad‐REIC‐induced apoptosis. Thus BiP is useful for diagnosis of inherent and acquired resistance of cancers and also as a target molecule to overcome resistance to the gene therapeutic Ad‐REIC.     

Visuospatial perception and navigation in Parkinson’s disease

A shifted field of view, an altered perception of optic flow speed, and gait asymmetries may influence heading direction in Parkinson’s disease (PD). PD participants (left body-side onset, LPD, n = 14; right body-side onset, RPD, n = 9) and Healthy Control participants (n = 17) walked a virtual hallway in which the optic flow speeds of the walls varied. Three-dimensional kinematics showed participants veered away from the faster moving wall. Although veering normally occurs toward the side with smaller step length, in both LPD and RPD this bias was overridden by a shifted field of view, which caused veering in the opposite direction, toward the side of the brain with more basal ganglia damage.     

A new polymorphism in the GRP78 is not associated with HBV invasion

AIM: To examine the association between-86 bp (T > A) in the glucose-regulated protein 78 gene ( GRP78) and hepatitis B virus (HBV) invasion.METHODS: DNA was genotyped for the single-nucleotide polymorphism by polymerase chain reaction followed by sequencing in a sample of 382 unrelated HBV carriers and a total of 350 sex-and age-matched healthy controls. Serological markers for HBV infection were determined by enzyme-linked immunosorbent assay kits or clinical chemistry testing.RESULTS: The distributions of allelotype and genotype in cases were not signi.cantly different from those in controls. In addition, our .ndings suggested that neither alanine aminotransferase/hepatitis B e antigen nor HBV-DNA were associated with the allele/genotype variation in HBV infected individuals.CONCLUSION:-86 bp T > A polymorphism in GRP78 gene is not related to the clinical risk and acute exacerbation of HBV invasion.