Clinical Significance of High Expression of Endoplasmic Reticulum Stress-related Proteins Glucose-regulating Protein 78 and X-box Binding Protein 1 induced by Fusobacterium Nucleatum in Esophageal Squamous Cell Carcinoma / 中山大学学报(医学科学版)

ObjectiveTo analyze the induction effect of Fusobacterium nucleatum （Fn） on endoplasmic reticulum stress-related proteins Glucose-regulating protein 78（GRP78） and X-box binding protein 1（XBP1） in esophageal squamous cell carcinoma （ESCC）， and to explore its potential mechanism and clinical significance. MethodsESCC cells KYSE150 and KYSE140 were infected with Fn for 12 h， 24 h and 48 h. The oxidative stress indexes （ROS， MDA and SOD） and the expression of GRP78 and XBP1 in each group were detected by oxidative stress index kit and Western blot. The experiment was divided into Fn groups， Fn+siNC1 groups， Fn+siGRP78 groups， Fn+siNC2 groups and Fn+siXBP1 groups； the oxidative stress indexes， paclitaxel （PTX） response efficacy， abilities of proliferation， invasion and metastasis in each group were compared. The infection of Fn and the expression of GRP78 and XBP1 in 234 ESCC and paracancerous tissues were detected by RNA scope and immunohistochemistry. The correlation between each factor and clinicopathological characteristics of patients was analyzed by Chi-square test. The influence of each factor on the survival of patients was compared by Kaplan-meier survival estimate. ResultsCompared with Fn uninfected KYSE150 and KYSE140 cells， the content of ROS and MDA was gradually increased， the activity of SOD was gradually decreased， and the expression of GRP78 and XBP1 was gradually increased in Fn infected groups （12 h， 24 h and 48 h） （P < 0.05）. Compared with Fn groups， Fn+siNC1 groups， and Fn+siNC2 groups， ROS and MDA contents were decreased， SOD activity was increased， PTX response efficacy was enhanced， and abilities of proliferation， invasion and metastasis were decreased in Fn+siGRP78 and Fn+siXBP1 groups （P < 0.05）. The rates of Fn， GRP78 and XBP1 in ESCC tissues were 43.16%， 69.66% and 60.68%， respectively. And the three indexes were significantly consistent （P < 0.05）. The patients with positive Fn infection and high expression of GRP78 and XBP1 were mostly males with a history of smoking and drinking， and the tumor differentiation degree was low， the invasion degree was deep， the lymph node metastasis rate was high， and the clinical stage was mostly stage Ⅲ/Ⅳ. The 5-year survival time of patients with above positive indexes was shortened （P < 0.05）. ConclusionsFn could induce endoplasmic reticulum stress by inducing the high expression of GRP78 and XBP1， and promote the malignant evolution of ESCC.

Effect of wheat-grain moxibustion on the expression of Beclin-1/GRP78 in spinal dorsal horn in rats with cervical spondylotic radiculopathy / 中国针灸

OBJECTIVE@#To observe the effect of wheat-grain moxibustion at "Dazhui" (GV 14) on the expressions of Beclin-1 and GRP78 in spinal dorsal horn in rats with cervical spondylotic radiculopathy (CSR), and to explore the possible analgesic mechanism of wheat-grain moxibustion for CSR.@*METHODS@#A total of 48 SD rats were randomly divided into a sham operation group, a model group, a wheat-grain moxibustion group and a wheat-grain moxibustion+3-MA group, 12 rats in each group. The CSR model was prepared by spinal cord insertion method. Three days after modeling, the rats in the model group were intraperitoneally injected with 1 mL of 0.9% sodium chloride solution; the rats in the wheat-grain moxibustion group were treated with wheat-grain moxibustion at "Dazhui" (GV 14, 6 cones per time) on the basis of the model group; the rats in the wheat-grain moxibustion+3-MA group were intraperitoneally injected with 3-MA solution and wheat-grain moxibustion at "Dazhui" (GV 14, 6 cones per time). The three groups were intervened for 7 days, once a day. The gait score and mechanical pain threshold were observed before treatment and 7 days into treatment; after the treatment, the expressions of mRNA and protein of Beclin-1 in spinal dorsal horn were detected by real-time fluorescence quantitative PCR and immunohistochemistry; the expression of GRP78 protein in spinal dorsal horn was detected by Western blot method; the autophagosomes and ultrastructure in spinal dorsal horn neurons were observed by electron microscope.@*RESULTS@#After the treatment, compared with the sham operation group, in the model group, the gait score was increased and the mechanical pain threshold was decreased (P<0.01), and the expression of GRP78 protein in spinal dorsal horn was increased (P<0.01). Compared with the model group and the wheat-grain moxibustion+3-MA group, in the wheat-grain moxibustion group, the gait score was decreased and mechanical pain threshold was increased (P<0.01), and the expression of GRP78 protein in spinal dorsal horn was decreased, and the expressions of mRNA and protein of Beclin-1 were increased (P<0.01). Under electron microscope, the ultrastructure of spinal dorsal horn neurons in the wheat-grain moxibustion group was not significantly damaged, and its structure was basically close to normal, and the number of autophagosomes was more than the other three groups.@*CONCLUSION@#Wheat-grain moxibustion at "Dazhui" (GV 14) has analgesic effect on CSR rats. The mechanism may be related to moderately up-regulate the expression of Beclin-1, enhance autophagy and reduce endoplasmic reticulum stress.

Research progress in the role of glucose-regulated protein 78 in mucormycosis / 中华微生物学和免疫学杂志

Mucormycosis is an infectious disease characterized by rapidly progressive vascular invasiveness, thrombosis and tissue necrosis, which could be potentially responsible for blocking the exudation of leukocytes to infection sites and affecting drug distribution. Glucose-regulated protein 78 (GRP78) is a key protein involved in regulating the invasiveness of Mucorales. Endoplasmic reticulum GRP78 is overexpressed under various stress conditions and transported to the cell membrane to become a cell surface receptor for Mucorales entering into vascular endothelial cells. This article reviewed the mechanisms and pathogenesis of GRP78-mediated host cell invasion and summarized the progress in related targeted drugs, aiming to provide reference for developing multi-target intervention against mucormycosis.

Effect of GSH and niacin combination on protein oxidation, ER stress, glycation and aggregation in HLE cells under high glucose condition / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM:To evaluate the effects of reduced glutathione(GSH)and niacin combination on protein oxidative stress, endoplasmic reticulum(ER)stress, glycation, and aggregation of the αβ crystalline in human lens epithelial(HLE)cells treated with high glucose levels. <p>METHODS:HLE cells were cultured and exposed to 25 mmol/L glucose to promote high glucose conditions. Groups of cells were co-treated with three different combinations of dosages: 10 μmol/L GSH+2 μmol/L niacin, 30 μmol/L GSH+25 μmol/L niacin, and 100 μmol/L GSH+25 μmol/L niacin. After 72h incubation, protein carbonyl content(PCC)and glucose reactive protein(GRP78)content were assessed using ELISA examinations. After two-week incubation, advanced glycation end products(AGEs)were also assessed and the expression of αβ crystalline was measured using Western Blot examination. The SPSS 18.0 statistical package was used for all data analyses.<p>RESULTS:PCC and GRP78 levels in the co-treated groups were not significantly reduced compared to control(<i>P</i>>0.05). In contrast, there was a significant decrease of the AGEs levels in all groups co-treated with GSH and niacin when compared with the control group(<i>P</i><0.05). In addition, the αβ crystalline expression increased after high dose glucose administration, but decreased in all groups co-treated with GSH and combinations of GSH and niacin.<p>CONCLUSION:The results suggest that combinations of GSH and niacin inhibit the aggregation of proteins and prevent glycation in hyperglycemic human lens epithelial cells. This study shows that this combination may play an active role in preventing diabetic cataract mainly from the AGEs pathway.

Immunohistochemical study of amelogenin binding proteins in an amelogenin point mutation mouse / Estudio inmunohistoquímico de proteínas de unión de la amelogenina en un ratón con mutación puntual de amelogenina

Amelogenin is one of the enamel matrices secreted by ameloblasts. A mutation of the amelogenin gene can cause hereditary dental enamel defects known as amelogenesis imperfecta (AI). Since lysosome-associated membrane protein-1 (LAMP-1), -3 (LAMP-3), and 78kDa glucose-related protein (Grp78) were identified as binding proteins of amelogenin, several studies have suggested the involvement of these binding proteins with the cell kinetics of ameloblasts in normal or abnormal conditions. The purpose of this study is to investigate the distribution of these amelogenin binding proteins in the ameloblast cell differentiation of mice with a point mutation of the amelogenin gene (Amelx*). The incisors of Amelx* mice had a white opaque color and the tooth surface was observed to be rough under a scanning electron microscope. Among the sequential ameloblast cell differentiation in the Amelx* mice, the shape of ameloblasts at the transition stage was irregular in comparison to those in wild-type (WT) mice. Immunostaining of Grp78 revealed that the whole cytoplasm of the transition stage ameloblasts was immunopositive for Grp78 antibody, while only the distal part of cell was positive in the WT mice. Furthermore, in the Amelx* mice, the cytoplasm of the transition stage ameloblasts was immunopositive for LAMP-1 and LAMP-3. These results suggest that Amelx* may cause the abnormal distribution of amelogenin binding proteins in the cytoplasm of ameloblasts.

La amelogenina es una de las matrices de esmalte secretadas por los ameloblastos. Una mutación del gen de amelogenina puede causar defectos hereditarios del esmalte dental conocidos como amelogénesis imperfecta (AI). Dado que la proteína de membrana asociada a lisosoma-1 (LAMP-1), -3 (LAMP-3) y la proteína relacionada con la glucosa de 78 kDa (Grp78) se identificaron como proteína de unión a amelogenina, varios estudios han sugerido la participación de estas proteínas con la cinética celular de los ameloblastos en condiciones normales o anormales. El objetivo del estudio fue investigar la distribución de LAMP-1, LAM-3 y Grp78 durante la diferenciación celular de ameloblastos de ratones con una mutación puntual del gen de amelogenina (Amelx*). Los incisivos de los ratones Amelx* presentaron un color blanco opaco y se observó en microscopio electrónico de barrido que la superficie del diente era áspera. La diferenciación celular secuencial y la forma de los ameloblastos en la etapa de transición en los ratones Amelx* fue irregular en comparación con los ratones silvestres (RS). La inmunotinción de Grp78 reveló que todo el citoplasma de los ameloblastos en etapa de transición fue inmunopositivo para el anticuerpo Grp78, mientras que solo la parte distal de la célula fue positiva en los ratones RS. Además, en ratones Amelx*, el citoplasma de los ameloblastos en etapa de transición fue inmunopositivo para LAMP-1 y LAMP-3. Estos resultados sugieren que Amelx* puede causar distribución anormal de proteínas de unión a amelogenina en el citoplasma de los ameloblastos.

Icariside II improves left ventricular function by improving endoplasmic reticulum stress and caspase 12 signaling in SHR / 中国药理学通报

Aim: To explore the mechanism of icariside II (ICS II) on improving left ventricular function based on endoplasmic reticulum stress and caspase-12 signaling in spontaneously hypertensive rats (SHRs). Methods: Thirty 14-week-old male SHRs were divided into model group, ICS II low, middle and high dose groups and positive drug group (n = 6). WKY was used as control group (n =6). ICS II groups were respectively given ICS 114, 8, 16 mg · kg-1(ig, qd), and positive drug group was given losartan (20 mg · kg-1). At the end of 26th weeks, anesthetized rats were measured by ultrasound for detection of the left ventricular function, RT-PCR was used to determine the level of GRP78 mR- NA in the left ventricle tissue, and Western blot was used to assess the levels of GRP78 and cleaved-caspase- 12/9/3 protein in the left ventricle tissues. Results: Compared with WKY group, the internal diameter and posterior wall thickness of the left ventricular end diastolic increased, while the ejection fraction and fractional shortening decreased in SHR group. GRP78 mRNA and protein levels were up-regulated, and the levels of cleaved-caspase-12/9/3 protein were raised in left ventricle (P < 0.05). Compared with SHR group, the internal diameter and posterior wall thickness of the left ventricular end diastolic increased in ICS II medium and high dose groups and positive drug group (P <0. 05), while the ejection fraction and fractional shortening decreased (P<0.05). GRP78 mRNA and protein levels were down-regulated, the levels of cleaved-caspase-12/ 9/3 protein declined in left ventricle (P <0.05). Conclusions: ICS II could improve left ventricular function in SHRs, and its mechanism may be related to improving left ventricular endoplasmic reticulum stress and down-regulating the elevated caspase-12 signaling.

Effects of difluoromethylalanine on cardiac hypertrophy and endoplasmic reticulum stress in type 2 diabetic rats / 中国药理学通报

Aim: To investigate the protective effects of difluoromethylornithine (DFMO) on cardiac hypertrophy and to explore its influence on the protein expression of GRP78 and CHOP in type 2 diabetic rats. Methods: The type 2 diabetic mellitus (T2DM) rat model was induced by 4 weeks' high sucrose-high fat diet feeding and one-time intraperitoneal injection of streptozotocin. Rats were randomly divided into three groups:control group, T2DM model group and DFMO treatment group. Rats of treatment group were treated with 2% DFMO drinking for 12 weeks. The changes in blood glucose and cardiac index were calculated. Myocardium superoxide dismutase (SOD), malondialdehyde (MDA) and the total antioxidant capacity (T- A0C) were determined. The morphological changes were observed by Masson staining, and the protein expressions of GRP78 and CHOP were measured by Western blot. Results: Compared with T2DM model group, the blood glucose, cardiac index and the degree of myocardial fibrosis significantly decreased (P < 0. 05), the content of MDA decreased and the activity of SOD and T-AOC increased. Meanwhile, the protein expressions of GRP78 and CHOP were down-regulated in treatment group (P < 0. 05). Conclusions: DFMO exerts protective effects on cardiac hypertrophy in T2DM rats, which may be related to its function of the down-regulation of protein expression of GRP78 and CHOP and inhibition of endoplasmic reticulum stress.

Naringin pretreatment alleviating myocardial cell hypoxia/reoxygenation injury by suppressing endoplasmic reticulum stress-induced apoptosis / 中国药理学通报

Aim: To study the effect of naringin (Nar) on endoplasmic reticulum stress (ERS) -induced apoptosis in H9c2 myocardial cell hypoxia/reoxygenation (H/R) injury and its molecular mechanism. Methods: H9c2 cells were cultured in vitro and randomly classified into five groups: normal control (group C), H/R group, H/R with Nar 10 mg · L-1 (group L), H/R with Nar20 mg · L-1 (group M) and H/R with Nar40 mg · L-1 (group H). Myocardial cells were normally cultured until the end of the experiment in group C. The myocardial cells were treated by hypoxia for 4 h before reoxygenation for 24 h in group H/R. The myocardial cells were cultured with Nar(10, 20, 40 mg · L-1) respectively 6 h before hypoxia, and after 6 h they were treated by hypoxia for 4 h before reoxygenation for 24 h in group L, group M and group H. The survival rate of cells was determined by MTT method after experiment. The apoptosis of cardiomyocytes was detected by TUNEL. CCAAT/enhancer-binding protein-homologous protein(CHOP), activating transcription factor-4(ATF4), eukaryotic initiation factor 2α (eIF2α), p-eIF2α, double-stranded RNA like endoplasmic reticulum kinase (PERK) and p-PERK were all assessed by Western blot. Results: Compared with group C, H9c2 cell viability decreased, induced apoptosis and the protein expression of CHOP, ATF4, peIF2α/eIF2α and p-PERK/PERK in group H/R were significantly increased (P < 0. 05). Compared with group H/R, H9c2 cell viability increased, the apoptosis and the protein expression of CHOP, ATF4, peIF2α/eIF2α and p-PERK/PERK were reduced in group L, group M and group H. Of the three groups, group M and group H showed the most significant effect (P < 0. 05). Conclusions: The Nar pretreatment can reduce myocardial cell apoptosis caused by H/R injury, suggesting that Nar can help to relieve the ERS-associated apoptosis through the PERK-eIF2α-ATF4- CHOP pathway.

Cripto Enhances Proliferation and Survival of Mesenchymal Stem Cells by Up-Regulating JAK2/STAT3 Pathway in a GRP78-Dependent Manner

Cripto is a small glycosylphosphatidylinositol-anchored signaling protein that can detach from the anchored membrane and stimulate proliferation, migration, differentiation, vascularization, and angiogenesis. In the present study, we demonstrated that Cripto positively affected proliferation and survival of mesenchymal stem cells (MSCs) without affecting multipotency. Cripto also increased expression of phosphorylated janus kinase 2 (p-JAK2), phosphorylated signal transducer and activator of transcription 3 (p-STAT3), 78 kDa glucose-regulated protein (GRP78), c-Myc, and cyclin D1. Notably, treatment with an anti-GRP78 antibody blocked these effects. In addition, pretreatment with STAT3 short interfering RNA (siRNA) inhibited the increase in p-JAK2, c-Myc, cyclin D1, and BCL3 levels caused by Cripto and attenuated the pro-survival action of Cripto on MSCs. We also found that incubation with Cripto protected MSCs from apoptosis caused by hypoxia or H₂O₂ exposure, and the level of caspase-3 decreased by the Cripto-induced expression of B-cell lymphoma 3-encoded protein (BCL3). These effects were sensitive to down-regulation of BCL3 expression by BCL3 siRNA. Finally, we showed that Cripto enhanced expression levels of vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), and hepatocyte growth factor (HGF). In summary, our results demonstrated that Cripto activated a novel biochemical cascade that potentiated MSC proliferation and survival. This cascade relied on phosphorylation of JAK2 and STAT3 and was regulated by GRP78. Our findings may facilitate clinical applications of MSCs, as these cells may benefit from positive effects of Cripto on their survival and biological properties.

Effects of resveratrol on expression of renal fibrosis proteins ORP150, GRP78, and GRP94 in rat kidney tissue / 中草药

Objective To assess the effect of resveratrol on the expression of endoplasmic reticulum stress molecular partners150-KD oxygen-regulated protein (ORP150) in renal tissues of rats with unilateral ureteral obstruction (UUO). Methods The UUO rat model of renal interstitial fibrosis was established. The rats were randomly divided into sham operation group, model group, enalapril group, the high-dose, medium-dose and low-dose groups of resveratrol, each group of 10. After 14 d of surgery, rats were sacrificed to collect serum and kidney tissue to detect the level of serum Scratinine (Scr) and blood urea nitrogen (BUN); Pathological changes were stained by HE to evaluate of renal tubular damage index, renal interstitial collagen deposition area was detected by Masson staining, apoptosis of in situ cell in renal tissue was determined by TUNEL assay, and using the western blot for the detection of protein expression of ORP150, GRP78, and GRP94 in renal tissue. Results After comparison of the treatment groups with model group, BUN and Scr levels in serum were significantly reduced in high-dose resveratrol group, the damage degree of renal tubules was significantly reduced, the relative area of the renal interstitial collagen decreased, In addition, the expression of ORP150 was increased (P < 0.05, 0.01), GRP78 and GRP94 proteins expression were significantly reduced, (P < 0.05, 0.01). Moreover, the resveratrol high-dose group seems more effective than enalapril groups in reversing the phenotype (P < 0.05). Conclusion Resveratrol was able to protect renal function, alleviate hydronephrosis, reduce interstitial injury of renal tubular, induce the expression of the key endoplasmic reticulum stress signal molecule ORP150 and the down-regulated molecular partner GRP78 and GRP94, delay the apoptosis of renal tubular epithelial cell of rats after UUO and inhibit renal interstitial fibrosis caused by apoptosis deposition.

Effects of isoflurane on learning and memory and the expression of GRP78,ATF4 and CHOP in frontal lobe neurons of aged rats / 中华行为医学与脑科学杂志

Objective To observe the changes of learning and memory ability and the expression of GRP8,ATF4 and CHOP in the frontal lobe neurons of rats after isoflurane anesthesia.Methods Male SD aged rats were randomly divided into 4 groups with 15 in each group.Rats in ISO group received 1.5% isoflurane 2 h,SAL group received intraperitoneal injection Salubrinal(1 mg/kg),ISO+SAL group received 1.5% 2 h after intraperitoneal injection Salubrinal(1 mg/kg),C group only inhaled 30% air and oxygen mixture.Morris water maze (MWM) test was performed 24 hours after isoflurane anesthesia,and then the left frontal lobe of rats was collected,gene transcription and protein expression changes of 78-kDa glucose-regulated protein(GRP78),activating transcription factor 4(ATF4),C/EBP homologous protein(CHOP) were evaluated by real-time fluorescence quantitative PCR and immunofluorescence.Results Compared with the C group,the latent time of ISO group was significantly prolonged(ISO group(19.10±2.98)s vs C group (10.54±2.05)s,P<0.05);the number of times passing through the target platform of ISO group was decreased significantly(ISO group (6.78±1.47) vs Cgroup (9.03±1.69),P<0.05);protein expression level of GRP78 was significantly increased in group ISO (ISO group (965.8±86.5) vs C group(247.5±46.3),P<0.05);protein expression level of ATF4 was significantly increased in group ISO(ISO group (470±69.4) vs C group (275.4±56.3),P<0.05) protein expression level of CHOP was significantly increased in group ISO(ISO group (618.7±83.3) vs C group(174.5±71.2),P<0.05).The transcription trends of GRP78,ATF4,CHOP were consistent with their protein expression.Conclusion The decrease of short-term memory ability after isoflurane anesthesia may be related to the activation of endoplasmic reticulum stress pathway in frontal lobe neurons.

Resveratrol alleviates ISO-induced cardiomyocyte hypertrophy in rat / 基础医学与临床

Objective To explore the role of resveratrol (RES) on cardiomyocyte hypertrophy of rat induced by isoproternol (ISO) and the effect of Res on the expression of GRP78 and GRP94 in endoplasmic reticulum stress of cardiomyocyte hypertrophy.Methods Hypertrophic model of cardiomyocytes was induced by ISO.Cardiomyocytes was divided into four group: control group, the model group, RES+ISO group and RES group.Hypertrophy status of cardiomyocytes was determined by Leica 2Q500 image analysis system measuring the cell surface area and the gene expression of ANP.The content of LDH and MDA was measured in different groups, and the protein expressions of GRP78 and GRP94 were detected by Western blot.Results Compared with control group, ISO induced cardiomyocytes hypertrophy, endoplasmic reticulum stress related factors GRP78 and GRP94 protein expression were increased, compared with ISO group, RES intervention effectively suppressed the cardiomyocytes hyper-trophy induced by ISO, reduced the protein expression of GRP78 and GRP94, at the same time, reduced lactate dehydrogenase (LDH) and malondialdehyde (MDA) release in cell medium.Conclusions Treatment of RES may protect cardiomyocytes hypertrophy, which is partially mediated by inhibiting the expression of ERS factors GRP78 and GRP94.

Effects of leptin on endoplasmic reticulum stress related proteins after focal cerebral ischemia / 中国神经精神疾病杂志

Objective To investigate the effect of leptin on endoplasmic reticulum stress related protein after focal cerebral ischemia in rats. Methods Ischemia was induced by occluding the middle cerebral artery in rats brain using the filament occlusion method. Forty SD rats were randomly divided into sham operation group, cerebral is-chemia group and leptin-preconditioning group. Leptin was injected subcutaneously before occlusion of blood vessel. Longa 5 score neurological function scale, body weight and brain edema changes were measured 6 hours after MCAO, and the brain was removed to detect the endoplasmic reticulum marker protein: glucose-regulated protein 78 (GRP78) and C/EBP-homologous protein (CHOP) by immunohistochemical method. Results There was no difference in the body weight changes between leptin-preconditioning group and ischemic group. In the leptin-preconditioning group, the neurological function score (1.90±0.31 vs. 2.50±0.52, P<0.05) and the degree of brain edema (3.60±0.52 vs. 7.70±0.94, P<0.001) were significantly lower than those in the cerebral ischemia group. Moreover, the expression of GRP78 in leptin-preconditioning group was significantly higher than that in ischemia group (48.69 ±5.06 vs. 35.78± 4.35, P<0.01), and the expression of CHOP was significantly lower than that of ischemia group (60.24 ±4.11 vs. 38.81±5.34, P<0.01). Conclusion Leptin can reduce the neurological deficit and may be associated with the up-reg-ulation of GRP78 protein, and down regulation of CHOP protein to weaken the endoplasmic reticulum stress caused by cerebral ischemia

Effect of PI3K/Akt signaling pathway and endoplasmicreticulum stress on apoptosis of hepatocytes in rats with hepatic fibrosis / 基础医学与临床

Objective To observe the changes of PI3K/Akt signaling pathway and endoplasmic reticulum stress related protein GRP78 and CHOP in CCl4 induced liver fibrosis and to explore their effects on hepatic fibrosis.Methods Thirty SD rats were randomly divided into normal control group, 4 and 8 weeks liver fibrosis group (hypodermic injection of 40% CCl4).Pathological changes of liver tissue was observed by HE staining.The techniques of real-time PCR was applied to detect mRNA of GRP78 and CHOP in liver.Detected expression of Akt1, phospho-Akt1, GRP78 and CHOP protein by western blot.Meanwhile, the cell apoptosis in liver was detected by TUNEL.Results Compared with the normal control group, GRP78 and CHOP mRNA and protein in 4 and 8 weeks liver fibrosis group was increased(P<0.05), while expression of Akt1, phospho-Akt1 in 4 and 8 weeks liver fibrosis group was lower than that in normal control group(P<0.05).Compared with normal control group, the apoptosis of hepatocytes in 4 and 8 weeks liver fibrosis group was elevated (P<0.05).Conclusions PI3K/Akt signaling pathway and endoplasmic reticulum stress may play important roles in the induction of apoptosis of hepatocytes in rats with hepatic fibrosis.

Protective effect of earthworm active ingredients against endoplasmic reticulum stress-induced acute liver injury in mice / 中国中药杂志

To study the protective effect of earthworm active ingredients(EWAs) against endoplasmic reticulum stress(ERS)-induced acute liver injury in mice. The model of liver injury was induced through intraperitoneal injection of 10%CCl4. Serum glutamic-pyruvic transaminase(ALT), glutamic-oxaloacetic transaminase(AST), superoxide dismutase(SOD) and glutathione peroxidase(GSH-PX) activity and malondialdehyde(MDA) concentration were detected by colorimetric method. Histological examination was performed through hematoxylin-eosin staining and light microscopy, and apoptosis was detected using terminal transferase dUTP nick end labeling. The expressions of ERS related proteins, including glucose regulated protein 78(GRP78), protein kinase R-like ER kinase(PERK), eukaryotic transcription initiation factor 2α(eIF2α), active transcription factor-4(ATF4) and CCAAT/enhancer binding homologous protein(CHOP), were measured by immunohistochemistry and Western blot. According to the results, compared with the model group,serological indexes in the high, middle and low doses of EWAs were significantly improved (P<0.05 or P<0.01), the extent of liver lesion was decreased and the degree of injury was significantly reduced, and that the liver index and the spleen index of mice were significantly changed(P<0.05 or P<0.01). In liver tissue, the expressions of GRP78 and CHOP were significantly decreased(P<0.05 or P<0.01). The protein expressions of GRP78, CHOP and its upstream signaling pathway PERK-eIF2-ATF4 were significantly decreased in each dose group(P<0.05 or P<0.01). In summary, EWAs has a significant protective effect on ERS-induced acute liver injury, and its mechanism may be correlated with the inhibition of oxidative stress and ERS, and down-regulation of ERS marker protein CHOP expression, andinhibition of apoptosis.

Expression of endoplasmic reticulum chaperone GRP78 in breast cancer and its prognosis significance / 国际外科学杂志

Objective To investigate the expression of GRP78 in breast cancer,the relationships among the expression and the clinicopathological characteristics,prognosis were also investigated.Methods The expression of GRP78 was detected in 172 paraffin-embedded breast cancer samples with immunohistochemistry Envision method,the relationships among the expression and the clinicopathological characteristics,prognosis were investigated.Results Positive expression of GRP78 is common in breast cancer.Strong expression of GRP78 was detected in 71 cases (41.28％),and weak expression was detected in 101 cases (58.72％).GRP78 expression wasn't associated with the clinicopathological characterristics except T stage and Her-2 status.Univariate analysis (log-rank test) showed that GRP78 expression correlated with disease free survival and overall survival significantly.Patients with strong GRP78 expression had poorer prognosis compared to those with weak GRP78 expression(P ＜ 0.05).Multivariate analysis utilizing Cox regression analysis showed that GRP78 is an independent biomarker of disease free survival (P ＜ 0.05),but not an independent biomarker of overall survival (P ＞ 0.05).Conclusions Positive expression of GRP78 is common in breast cancer and strong expression is associated with poorer survival.

Altered expression of Chop and Grp78 in neural tube of rat with spina bifida / 国际儿科学杂志

Chop and Grp78 genes are key genes in endoplasmic reticulum stress(ERS).To explore the gene alteration of Chop and Grp78 in embryo with spina bifida,we studied Chop and Grp78 gene expression by qRT-PCR in neural tube of rat with spina bifida in E12 and primary cultured neuroepithelial stem cells(NEP).We found higher expression of both Chop and Grp78 in neural tube of rat with spina bifida than normal rats.While Chop and Grp78 genes were up-regulated after NEP differentiated into neuron.The up-regulation of Chop and Grp78 genes in neural rube of rat and NEP might be involved in the abnormal apoptosis in spina bifida related with ERS.And we found the primary culture of NEP provided a good model for embryo early development research.

Role of ERS in Astragaloside Ⅳ-induced cardioprotection against ischemia/reperfusion injury in rats / 中国药理学通报

Aim To explore the role of endoplasmic re-ticulum stress( ERS) in Astragaloside Ⅳ-induced car-dioprotection against ischemia/reperfusion injury in rats. Methods A model of myocardial ischemia 30 min followed by 120 min reperfusion was made by liga-ting coronary artery in male Wistar rats. Rats were di-vided randomly into 4 groups: sham group, ischemia/reperfusion group, ERS inhibitor TUDCA group, As-tragaloside Ⅳgroup. Myocardial samples were collect-ed from the risk zones during ischemia and reperfu-sion, ERS was determined by measuring levels of glu-cose regulated protein 78 ( GRP78 ) , an established marker of ERS with Western blot. Immunofluorescence study was used to test GRP78 intensity with laser scan-ning confocal microscopy, TTC method was used to measure the infarct size,hematoxylin-eosin staining was used to observe the changes of morphological changes of myocardium. Results There was no statistical difference in GRP78 expression during ischemia com-pared to the sham group, but was markedly increased upon reperfusion. Astragaloside Ⅳ could mimic TUD-CA and significantly decreased the GRP78 expression, reduced infarct size and improved the morphology of myocardial tissue with a significant statistical difference compared with the control group ( P<0. 05 ) . Conclu-sions ERS is induced upon reperfusion but not during ischemia in isolated rat hearts. Astragaloside Ⅳ pre-vents myocardial reperfusion injury presumably by the inhibition of ERS.

Expression of endoplasmic reticulum stress-associated molecule GRP78 in testicular tissue of rats in differ-ent phases of morphine-dependence / 实用医学杂志

Objective To investigate expression level of endoplasmic reticulum stressmarker GRP78 in the testicular tissue in rats with different phases of morphine-dependence. To explore the role of ERS in morphine-de-pendence. Methods SD rats were divided into 6 groups: morphine (mor) -withdrawal group, mor-extinct group, mor-kindling group and their control groups, normal saline (NS)-withdrawal group, NS-extinct group, NS-kindling group. The experimental rats were injected with morphine subcutaneously on increasing dosage to establish the con-ditioned place preference (CPP) model. The rats in control groups were injected NS. Then the rats were suffered from withdrawal for 48 h, extinction and kindling by morphine, separately. The GRP78 expression level in testicular tissues of rats in the time point mentioned above were measured using Western Blot. Results The time of rats in the paired-box was (528.0 ± 81.0) s, which was significantly higher than that in the NS control group (P<0.001). It was (396.8 ± 116.9) s after extinctive phase, which was significantly higher than that the withdrawal phase of rats (P < 0.001). Also it was (396.8 ± 116.9) s after kindling with morphine which was significantly higher than that the extinctive phase of rats (P < 0.001). These changes of the time indicated that the animal models of extinction and kindling were established in the study. The GRP78 levels were down-regulated in 48 h after withdrawal (P <0.05), and increased a bit afterextinctive phase, but up-regulated highly after kindling with morphine (P < 0.01). Conclusion ERS may be related in the morphine dependence and it might play an important role of testicular dys-function in male drugabuser.

Myocardial expression of Caspase-12 and GRP78 in cardiac arrest and beating heart mitral valve replacement / 实用医学杂志

Objective To observe the expression of Caspase-12 and GRP78 of endoplasmic reticulum stress (ERS) in cardiac arrest and beating heart mitral valve replacement Methods Thirty patients with rheumatic heart disease mitral stenosis were randomly divided into beating heart group (BH,n=15) and cardiac arrest group(CA, n = 15). Both groups accepted MVR by beating heart surgery and cardiac arrest surgery under cardiopulmonary bypass (CPB) respectively. Right atrial myocardial tissues were collected at prior the start of CPB (T0), after aortic cross-clamping 30 minutes (BH group 30 minutes after CPB, T1) and stitched right atrium (T2) respectively. The method of reverse transcriptase polymerase chain reaction (RT-PCR) was applied to detect the expression level of Caspase-12 and GRP78 in two groups and positive staining of Caspase-12 and GRP78 of myocardial tissue slices in both groups was observed by immunohistochemical method. Results The expression of Caspase-12 in CA group heightened at T1and significantly increased at T2 (P < 0.05) but the expression of Caspase-12 in BH group had increased in T2 only (P < 0.05). Caspase-12 in CA group expressed higher than that in BH group at T1 and T2. The expression of GRP78 had increased at T1 in two groups but it in CA group expressed higher than that inBH group at T2. The number of positive staining of Caspase-12 and GRP78 in CA group was higher than that in BH group at T2. Conclusion MVR of beating heart can reduce the reaction of ERS to enhance the myocardial protection under CPB.