BACKGROUND Diabetic retinopathy(DR)is a major ocular complication of diabetes mellitus,leading to visual impairment.Retinal pigment epithelium(RPE)injury is a key component of the outer blood retinal barrier,and its d...BACKGROUND Diabetic retinopathy(DR)is a major ocular complication of diabetes mellitus,leading to visual impairment.Retinal pigment epithelium(RPE)injury is a key component of the outer blood retinal barrier,and its damage is an important indicator of DR.Receptor for activated C kinase 1(RACK1)activates protein kinase C-ε(PKC-ε)to promote the generation of reactive oxygen species(ROS)in RPE cells,leading to apoptosis.Therefore,we hypothesize that the activation of RACK1 under hypoxic/high-glucose conditions may promote RPE cell apoptosis by modulating PKC-ε/ROS,thereby disrupting the barrier effect of the outer blood retinal barrier and contributing to the progression of DR.AIM To investigate the role and associated underlying mechanisms of RACK1 in the development of early DR.METHODS In this study,Sprague-Dawley rats and adult RPE cell line-19(ARPE-19)cells were used as in vivo and in vitro models,respectively,to explore the role of RACK1 in mediating PKC-εin early DR.Furthermore,the impact of RACK1 on apoptosis and barrier function of RPE cells was also investigated in the former model.RESULTS Streptozotocin-induced diabetic rats showed increased apoptosis and upregulated expression of RACK1 and PKC-εproteins in RPE cells following a prolonged modeling.Similarly,ARPE-19 cells exposed to high glucose and hypoxia displayed elevated mRNA and protein levels of RACK1 and PKC-ε,accompanied by an increases in ROS production,apoptosis rate,and monolayer permeability.However,silencing RACK1 significantly downregulated the expression of PKC-εand ROS,reduced cell apoptosis and permeability,and protected barrier function.CONCLUSION RACK1 plays a significant role in the development of early DR and might serve as a potential therapeutic target for DR by regulating RPE apoptosis and barrier function.展开更多
Objective: To investigate the effect of activated protein C (APC) on inflammatory responses in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). Methods: The second passage of co...Objective: To investigate the effect of activated protein C (APC) on inflammatory responses in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). Methods: The second passage of collagenase digested HUVEC was divided into the following groups: serum free medium control group (SFM control), phosphate buffer solution control group (PBS control), LPS group with final concentration of 1 μg/ml (LPS group), APC group with final concentration of 7 μg/ml, Pre-APC group (APC pretreatment for 30 min prior to LPS challenge), and Post-APC group (APC administration 30 min after LPS challenge). Supernatant was harvested at 0, 4, 8, 12 and 24 h after LPS challenge. Interleukin-6 (IL-6) and Interleukin-8 (IL-8) levels were analyzed with ELISA. Cells were harvested at 24 h after LPS challenge, and total RNA was extracted. Mes-senger RNA levels for IL-6 and IL-8 were semi-quantitatively determined by RT-PCR. Results: Compared with control group, IL-6 and IL-8 levels steadily increased 4 to 24 h after LPS stimulation. APC treatment could increase LPS-induced IL-6 and IL-8 production. The mRNA levels of IL-6 and IL-8 exhibited a similar change. Conclusion: APC can further increase the level of IL-6 and IL-8 induced by LPS. The effect of these elevated cytokines is still under investigation.展开更多
Objective： To investigate the expression of OX40 ligand（OX40L） on C-reactive protein（CRP）-triggered mouse aorta endothelial cells （MAECs） in vitro. Methods： MAECs from aorta were isolated by digestion with col...Objective： To investigate the expression of OX40 ligand（OX40L） on C-reactive protein（CRP）-triggered mouse aorta endothelial cells （MAECs） in vitro. Methods： MAECs from aorta were isolated by digestion with collagenase type II. The cell growth was confirmed by morphological characteristics and the immunological marker, factor Ⅷ（or Willebrand factor, vWF）. The expression of OX40L by MAECs was detected by RT-PCR and western blot after incubating with 100μg/ml CRP for 48 hours. Results： Twenty-day cultures of MAECs formed confluent monolayer of a cobblestone pattern. RT-PCR and western blot assay showed that the level of OX40L expression in MAECs receiving CRP treatment was higher than control. Conclusion： A reliable method is described to isolate and propagate MAECs. CRP upregulates OX40L expression in MAECs.展开更多
Hepatocellular carcinoma(HCC) is the fifth most common cancer, and hepatitis C virus(HCV) infection plays a major role in HCC development. The molecular mechanisms by which HCV infection leads to HCC are varied. HCV c...Hepatocellular carcinoma(HCC) is the fifth most common cancer, and hepatitis C virus(HCV) infection plays a major role in HCC development. The molecular mechanisms by which HCV infection leads to HCC are varied. HCV core protein is an important risk factor in HCV-associated liver pathogenesis and can modulate several signaling pathways involved in cell cycle regulation, cell growth promotion, cell proliferation, apoptosis, oxidative stress and lipid metabolism. The dysregulation of signaling pathways such as transforming growth factor β(TGF-β), vascular endothelial growth factor(VEGF), Wnt/β-catenin(WNT), cyclooxygenase-2(COX-2) and peroxisome proliferator-activated receptor α(PPARα) by HCV core protein is implicated in the development of HCC. Therefore, it has been suggested that this protein be considered a favorable target for further studies in the development of HCC. In addition, considering the axial role of these signaling pathways in HCC, they are considered druggable targets for cancer therapy. Therefore, using strategies to limit the dysregulation effects of core protein on these signaling pathways seems necessary to prevent HCV-related HCC.展开更多
基金Supported by National Natural Science Foundation of China,No.82260211Key Research and Development Project in Jiangxi Province,No.20203BBG73058Chinese Medicine Science and Technology Project in Jiangxi Province,No.2020A0166.
文摘BACKGROUND Diabetic retinopathy(DR)is a major ocular complication of diabetes mellitus,leading to visual impairment.Retinal pigment epithelium(RPE)injury is a key component of the outer blood retinal barrier,and its damage is an important indicator of DR.Receptor for activated C kinase 1(RACK1)activates protein kinase C-ε(PKC-ε)to promote the generation of reactive oxygen species(ROS)in RPE cells,leading to apoptosis.Therefore,we hypothesize that the activation of RACK1 under hypoxic/high-glucose conditions may promote RPE cell apoptosis by modulating PKC-ε/ROS,thereby disrupting the barrier effect of the outer blood retinal barrier and contributing to the progression of DR.AIM To investigate the role and associated underlying mechanisms of RACK1 in the development of early DR.METHODS In this study,Sprague-Dawley rats and adult RPE cell line-19(ARPE-19)cells were used as in vivo and in vitro models,respectively,to explore the role of RACK1 in mediating PKC-εin early DR.Furthermore,the impact of RACK1 on apoptosis and barrier function of RPE cells was also investigated in the former model.RESULTS Streptozotocin-induced diabetic rats showed increased apoptosis and upregulated expression of RACK1 and PKC-εproteins in RPE cells following a prolonged modeling.Similarly,ARPE-19 cells exposed to high glucose and hypoxia displayed elevated mRNA and protein levels of RACK1 and PKC-ε,accompanied by an increases in ROS production,apoptosis rate,and monolayer permeability.However,silencing RACK1 significantly downregulated the expression of PKC-εand ROS,reduced cell apoptosis and permeability,and protected barrier function.CONCLUSION RACK1 plays a significant role in the development of early DR and might serve as a potential therapeutic target for DR by regulating RPE apoptosis and barrier function.
文摘Objective: To investigate the effect of activated protein C (APC) on inflammatory responses in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). Methods: The second passage of collagenase digested HUVEC was divided into the following groups: serum free medium control group (SFM control), phosphate buffer solution control group (PBS control), LPS group with final concentration of 1 μg/ml (LPS group), APC group with final concentration of 7 μg/ml, Pre-APC group (APC pretreatment for 30 min prior to LPS challenge), and Post-APC group (APC administration 30 min after LPS challenge). Supernatant was harvested at 0, 4, 8, 12 and 24 h after LPS challenge. Interleukin-6 (IL-6) and Interleukin-8 (IL-8) levels were analyzed with ELISA. Cells were harvested at 24 h after LPS challenge, and total RNA was extracted. Mes-senger RNA levels for IL-6 and IL-8 were semi-quantitatively determined by RT-PCR. Results: Compared with control group, IL-6 and IL-8 levels steadily increased 4 to 24 h after LPS stimulation. APC treatment could increase LPS-induced IL-6 and IL-8 production. The mRNA levels of IL-6 and IL-8 exhibited a similar change. Conclusion: APC can further increase the level of IL-6 and IL-8 induced by LPS. The effect of these elevated cytokines is still under investigation.
基金supported by National Natural Science Founda-tion of China(30770894)Medical Foundation key scholar of Jiangsu Province(RC2007037)
文摘Objective： To investigate the expression of OX40 ligand（OX40L） on C-reactive protein（CRP）-triggered mouse aorta endothelial cells （MAECs） in vitro. Methods： MAECs from aorta were isolated by digestion with collagenase type II. The cell growth was confirmed by morphological characteristics and the immunological marker, factor Ⅷ（or Willebrand factor, vWF）. The expression of OX40L by MAECs was detected by RT-PCR and western blot after incubating with 100μg/ml CRP for 48 hours. Results： Twenty-day cultures of MAECs formed confluent monolayer of a cobblestone pattern. RT-PCR and western blot assay showed that the level of OX40L expression in MAECs receiving CRP treatment was higher than control. Conclusion： A reliable method is described to isolate and propagate MAECs. CRP upregulates OX40L expression in MAECs.
文摘Hepatocellular carcinoma(HCC) is the fifth most common cancer, and hepatitis C virus(HCV) infection plays a major role in HCC development. The molecular mechanisms by which HCV infection leads to HCC are varied. HCV core protein is an important risk factor in HCV-associated liver pathogenesis and can modulate several signaling pathways involved in cell cycle regulation, cell growth promotion, cell proliferation, apoptosis, oxidative stress and lipid metabolism. The dysregulation of signaling pathways such as transforming growth factor β(TGF-β), vascular endothelial growth factor(VEGF), Wnt/β-catenin(WNT), cyclooxygenase-2(COX-2) and peroxisome proliferator-activated receptor α(PPARα) by HCV core protein is implicated in the development of HCC. Therefore, it has been suggested that this protein be considered a favorable target for further studies in the development of HCC. In addition, considering the axial role of these signaling pathways in HCC, they are considered druggable targets for cancer therapy. Therefore, using strategies to limit the dysregulation effects of core protein on these signaling pathways seems necessary to prevent HCV-related HCC.
基金Supported by the Grant from the Chinese Ministry of Agriculture [No.NYCYTX-43] the Zhejiang Provincial Natural Science Foundation of China [No.R3090332]
