BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammato...BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.展开更多
Objective： To investigate the mechanisms that Simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A（HMG-CoA） reductase inhibitor, plays an important role in primary prevention of atherosclerosis independently of its...Objective： To investigate the mechanisms that Simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A（HMG-CoA） reductase inhibitor, plays an important role in primary prevention of atherosclerosis independently of its lipid-lowering effect in Apolipoprotein E-deficient mice in the early stage of atherosclerosis. Methods： Twenty-four 6-week old male apoE-deficient mice were randomly divided into two groups：control group（normal saline） and treatment group[simvastatin（5 mg/（kg · d））]. Simvastatin was administered to treatment group mice by gavage and the same volume of normal saline was administered to control group mice by the same method for 2 or 4 weeks. Total cholesterol（TC）, super-oxide dismutase（SOD）, malondialdehyde（MDA） and serum nitric oxide（NO） were measured by biochemical analysis. Results： There was no significant difference in serum TC between control and treatment groups. Compared with the control＇ s, the effects of simvastatin were more significant in decreasing serum MDA level（P 〈 0.01 vs control＇ s at 2-week; P 〈 0.006 vs control＇ s at 4-week）, increasing serum SOD level（P 〈 0.03 vs control＇ s at 2-week; P 〈 0.003 vs control＇ s at 4-week） and NO level （P 〈 0.01 control＇ s at 2-week; P 〈 0.001 vs control＇ s at 4-week） either at 2 or 4 weeks. Conclusion： Simvastatin attenuates oxidative stress and protects endothelial function by the mechanisms of decreasing serum MDA level, increasing serum SOD level and NO level, which were inconsistent with its cholesterol-lowering effect. It may play an important role in primary（if not all） prevention of atherosclerosis and might be independent of lipid-regulation mechanism.展开更多
Mice overexpressing the human apolipoprotein apo CIII are a model of dyslipidemia. They become hypertxiglyceridemic, hypercholesterolemic and have high blood levels of free fatty acids. Blood glucose is normal, but as...Mice overexpressing the human apolipoprotein apo CIII are a model of dyslipidemia. They become hypertxiglyceridemic, hypercholesterolemic and have high blood levels of free fatty acids. Blood glucose is normal, but as the liver integrates lipid and carbohydrate metabolism, conditions of high inter-tissue circulation of energy substxates, such as fasting, may reveal hepatic alterations of glucose metabolism in these （CIII） mice. This hypothesis was explored by in situ liver perfusion in this investigation. The NTG （non-txansgenic） animals showed liver and muscle glycogen content changes compatible with the fed or fasted state. In contrast, glycogen in group CIII was much lower in the fed state. The liver glucose release in group CIII after overnight fasting and adrenaline-stimulated was lower than in group NTG. Total glucose production under gluconeogenic conditions was not different between groups NTG and CIII, but glucose production from alanine was decreased in group CIII. Therefore, dyslipidemia caused by overexpression of apoCIII in mice alters the liver glucose metabolism, particularly compromising glycogen synthesis and degradation. This profile might have adverse outcomes during metabolic challenges that are more severe than fasting.展开更多
Background Restenosis and atherosclerosis are two disorders characterized by abundant proliferation and migration of vascular smooth muscle cells(VSMCs).Previous studies have demonstrated a protective effect of the ce...Background Restenosis and atherosclerosis are two disorders characterized by abundant proliferation and migration of vascular smooth muscle cells(VSMCs).Previous studies have demonstrated a protective effect of the cellular repressor of E1A-stimulated genes(CREG) against restenosis. However,the role of CREG in atherosclerosis is undetermined. The aim of the present study is to examine the impact of CREG on the atherosclerosis.Methods Both immunofluorescence and western blotting were used in this experiment. Results The expression of CREG was decreased markedly in atherosclerotic lesions compared with normal areas of the vessels from both humans and mice species.We furthermore demonstrated that compared with the adenovirus-mediated-GFP control,intravenous administration of adenovirus-mediated CREG to apolipoprotein E deficient mice with six-week high-fat diet significantly reduced the relative area of atherosclerotic lesions in the mice aorta,accompanied by a decreased levels of Tumor necrosis factor(TNF) -αand Interleukin (IL)-1βmeasured by ELISA.Meanwhile,Western analysis revealed that NF-κB activation was also markedly reduced.Studies of cultured human VSMCs identified that overexpression of CREG abrogated the proliferation of human VSMCs stimulated by ox-LDL,along with a significantly decreased releasing of TNF-αand IL-1β.Conversely,down-regulation CREG expression contributed to cells proliferation stimulated by ox-LDL in cultured human VSMCs.Furthermore, overexpression of CREG suppressed the activations of NF-kB and ERK1/2 in cultured cells,while Furthermore, treatment with ERK inhibitor PD98059 reversed the CREG-mediated inhibition of human VSMCs proliferation.Conclusions CREG has a protective effect against atherosclerosis, which is related to inhibiting proliferation and inflammatory response of VSMCs.展开更多
目的探讨高热量饮食和年龄对载脂蛋白E基因敲除(ApoE^(-/-))小鼠脑功能的影响。方法选取8月龄成年ApoE^(-/-)小鼠和18月龄老年ApoE^(-/-)小鼠共20只,随机分为正常饮食成年组、正常饮食老年组、高热量饮食成年组、高热量饮食老年组,每组...目的探讨高热量饮食和年龄对载脂蛋白E基因敲除(ApoE^(-/-))小鼠脑功能的影响。方法选取8月龄成年ApoE^(-/-)小鼠和18月龄老年ApoE^(-/-)小鼠共20只,随机分为正常饮食成年组、正常饮食老年组、高热量饮食成年组、高热量饮食老年组,每组5只。正常饮食成年组、正常饮食老年组小鼠喂食实验室标准饲料,高热量饮食成年组、高热量饮食老年组小鼠喂食高脂饲料,干预8周。用体质量监测和葡萄糖耐量实验测试小鼠体质量、血糖变化,核磁共振波谱检测海马和下丘脑N-乙酰天冬氨酸(NAA)、胆碱(Cho)含量,Y迷宫和旷场实验检测认知功能,Western blot检测脑组织突触体相关蛋白25(SNAP-25)、突触素(synaptophysin)、突触后致密蛋白95(PSD-95)、诱导型一氧化氮合酶(iNOS)、白细胞介素1β(IL-1β)表达。结果与正常饮食成年组比较,高热量饮食成年组海马NAA、下丘脑Cho和NAA、自发交替率、SNAP-25、synaptophysin、PSD-95表达降低,差异有统计学意义(P<0.05,P<0.01),iNOS、IL-1β表达升高,差异有统计学意义(P<0.01);与正常饮食成年组比较,正常饮食老年组海马NAA、下丘脑Cho、SNAP-25、synaptophysin、PSD-95表达降低(P<0.05,P<0.01),iNOS、IL-1β表达升高(P<0.01);与正常饮食老年组比较,高热量饮食老年组海马和下丘脑Cho和NAA、中心路程/总路程、SNAP-25、synaptophysin、PSD-95表达降低(P<0.05,P<0.01),iNOS、IL-1β表达升高(P<0.01);与高热量饮食成年组比较,高热量饮食老年组海马NAA、中心路程/总路程、平均速度、synaptophysin表达降低(P<0.05,P<0.01),iNOS、IL-1β表达升高(1.61±0.10 vs 1.35±0.13,2.04±0.08 vs 1.54±0.11,P<0.05,P<0.01)。结论高热量饮食导致ApoE^(-/-)小鼠代谢障碍和神经炎症,抑制突触蛋白表达引起认知功能障碍;长期高热量饮食和年龄增加促进ApoE^(-/-)小鼠脑功能衰退。展开更多
目的研究C1q/肿瘤坏死因子相关蛋白9(CTRP9)对高脂血症模型载脂蛋白E基因敲除(ApoE^(-/-))小鼠肝脏脂肪病变形成的影响及机制。方法将30只8周龄雄性ApoE^(-/-)小鼠随机分为对照组与观察组,每组15只,对照组小鼠给予尾静脉注射腺病毒空载...目的研究C1q/肿瘤坏死因子相关蛋白9(CTRP9)对高脂血症模型载脂蛋白E基因敲除(ApoE^(-/-))小鼠肝脏脂肪病变形成的影响及机制。方法将30只8周龄雄性ApoE^(-/-)小鼠随机分为对照组与观察组,每组15只,对照组小鼠给予尾静脉注射腺病毒空载绿色荧光蛋白(Ad-GFP),观察组小鼠给予尾静脉注射腺病毒载CTRP9(Ad-CTRP9),高脂高胆固醇饮食(HF/HC)饲喂12周后,采集尾静脉血,处死小鼠收集肝脏组织。采用HE、油红O染色观察CTRP9对ApoE^(-/-)小鼠肝脏脂肪病变形成的影响;采用real time PCR检测CTRP9及脂质合成、炎症相关和线粒体氧化磷酸化相关基因的表达水平;采用western blotting技术检测肝脏组织中CTRP9蛋白表达水平。结果与对照组相比,过表达CTRP9小鼠(观察组)体重和血脂水平差异无统计学意义(P>0.05),CTRP9基因和蛋白表达水平显著升高(P<0.05),肝脏脂肪病变显著减少(P<0.05),肝脏组织中甘油三酯和总胆固醇水平显著下降(P<0.05),脂质合成(FAS、SCD1、SREBP1c)以及炎症相关基因(MCP-1、MIP1α和IL-1β)表达水平显著下调(P<0.01或P<0.001),而脂质代谢(PPARα、PPARγ、LXRα和LXRβ)及线粒体氧化磷酸化相关基因(CPT1、CPT2、COX4、Cytoc、Acadl和Acadm)表达水平显著增加(P<0.05或P<0.01)。结论过表达CTRP9能够抑制肝脏组织中脂肪生成以及炎症反应,同时促进肝细胞脂肪酸氧化磷酸化和能量代谢,增加线粒体能量消耗,从而降低肝脏组织中脂质蓄积。展开更多
文摘BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.
基金This study was supported by Provincial Natural Science Foundation of the Department of Education of Jiangsu(01 KJB320003)Innova-tion Fund of Nanjiing Medical University(CX 003001)
文摘Objective： To investigate the mechanisms that Simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A（HMG-CoA） reductase inhibitor, plays an important role in primary prevention of atherosclerosis independently of its lipid-lowering effect in Apolipoprotein E-deficient mice in the early stage of atherosclerosis. Methods： Twenty-four 6-week old male apoE-deficient mice were randomly divided into two groups：control group（normal saline） and treatment group[simvastatin（5 mg/（kg · d））]. Simvastatin was administered to treatment group mice by gavage and the same volume of normal saline was administered to control group mice by the same method for 2 or 4 weeks. Total cholesterol（TC）, super-oxide dismutase（SOD）, malondialdehyde（MDA） and serum nitric oxide（NO） were measured by biochemical analysis. Results： There was no significant difference in serum TC between control and treatment groups. Compared with the control＇ s, the effects of simvastatin were more significant in decreasing serum MDA level（P 〈 0.01 vs control＇ s at 2-week; P 〈 0.006 vs control＇ s at 4-week）, increasing serum SOD level（P 〈 0.03 vs control＇ s at 2-week; P 〈 0.003 vs control＇ s at 4-week） and NO level （P 〈 0.01 control＇ s at 2-week; P 〈 0.001 vs control＇ s at 4-week） either at 2 or 4 weeks. Conclusion： Simvastatin attenuates oxidative stress and protects endothelial function by the mechanisms of decreasing serum MDA level, increasing serum SOD level and NO level, which were inconsistent with its cholesterol-lowering effect. It may play an important role in primary（if not all） prevention of atherosclerosis and might be independent of lipid-regulation mechanism.
文摘Mice overexpressing the human apolipoprotein apo CIII are a model of dyslipidemia. They become hypertxiglyceridemic, hypercholesterolemic and have high blood levels of free fatty acids. Blood glucose is normal, but as the liver integrates lipid and carbohydrate metabolism, conditions of high inter-tissue circulation of energy substxates, such as fasting, may reveal hepatic alterations of glucose metabolism in these （CIII） mice. This hypothesis was explored by in situ liver perfusion in this investigation. The NTG （non-txansgenic） animals showed liver and muscle glycogen content changes compatible with the fed or fasted state. In contrast, glycogen in group CIII was much lower in the fed state. The liver glucose release in group CIII after overnight fasting and adrenaline-stimulated was lower than in group NTG. Total glucose production under gluconeogenic conditions was not different between groups NTG and CIII, but glucose production from alanine was decreased in group CIII. Therefore, dyslipidemia caused by overexpression of apoCIII in mice alters the liver glucose metabolism, particularly compromising glycogen synthesis and degradation. This profile might have adverse outcomes during metabolic challenges that are more severe than fasting.
文摘Background Restenosis and atherosclerosis are two disorders characterized by abundant proliferation and migration of vascular smooth muscle cells(VSMCs).Previous studies have demonstrated a protective effect of the cellular repressor of E1A-stimulated genes(CREG) against restenosis. However,the role of CREG in atherosclerosis is undetermined. The aim of the present study is to examine the impact of CREG on the atherosclerosis.Methods Both immunofluorescence and western blotting were used in this experiment. Results The expression of CREG was decreased markedly in atherosclerotic lesions compared with normal areas of the vessels from both humans and mice species.We furthermore demonstrated that compared with the adenovirus-mediated-GFP control,intravenous administration of adenovirus-mediated CREG to apolipoprotein E deficient mice with six-week high-fat diet significantly reduced the relative area of atherosclerotic lesions in the mice aorta,accompanied by a decreased levels of Tumor necrosis factor(TNF) -αand Interleukin (IL)-1βmeasured by ELISA.Meanwhile,Western analysis revealed that NF-κB activation was also markedly reduced.Studies of cultured human VSMCs identified that overexpression of CREG abrogated the proliferation of human VSMCs stimulated by ox-LDL,along with a significantly decreased releasing of TNF-αand IL-1β.Conversely,down-regulation CREG expression contributed to cells proliferation stimulated by ox-LDL in cultured human VSMCs.Furthermore, overexpression of CREG suppressed the activations of NF-kB and ERK1/2 in cultured cells,while Furthermore, treatment with ERK inhibitor PD98059 reversed the CREG-mediated inhibition of human VSMCs proliferation.Conclusions CREG has a protective effect against atherosclerosis, which is related to inhibiting proliferation and inflammatory response of VSMCs.
文摘目的探讨高热量饮食和年龄对载脂蛋白E基因敲除(ApoE^(-/-))小鼠脑功能的影响。方法选取8月龄成年ApoE^(-/-)小鼠和18月龄老年ApoE^(-/-)小鼠共20只,随机分为正常饮食成年组、正常饮食老年组、高热量饮食成年组、高热量饮食老年组,每组5只。正常饮食成年组、正常饮食老年组小鼠喂食实验室标准饲料,高热量饮食成年组、高热量饮食老年组小鼠喂食高脂饲料,干预8周。用体质量监测和葡萄糖耐量实验测试小鼠体质量、血糖变化,核磁共振波谱检测海马和下丘脑N-乙酰天冬氨酸(NAA)、胆碱(Cho)含量,Y迷宫和旷场实验检测认知功能,Western blot检测脑组织突触体相关蛋白25(SNAP-25)、突触素(synaptophysin)、突触后致密蛋白95(PSD-95)、诱导型一氧化氮合酶(iNOS)、白细胞介素1β(IL-1β)表达。结果与正常饮食成年组比较,高热量饮食成年组海马NAA、下丘脑Cho和NAA、自发交替率、SNAP-25、synaptophysin、PSD-95表达降低,差异有统计学意义(P<0.05,P<0.01),iNOS、IL-1β表达升高,差异有统计学意义(P<0.01);与正常饮食成年组比较,正常饮食老年组海马NAA、下丘脑Cho、SNAP-25、synaptophysin、PSD-95表达降低(P<0.05,P<0.01),iNOS、IL-1β表达升高(P<0.01);与正常饮食老年组比较,高热量饮食老年组海马和下丘脑Cho和NAA、中心路程/总路程、SNAP-25、synaptophysin、PSD-95表达降低(P<0.05,P<0.01),iNOS、IL-1β表达升高(P<0.01);与高热量饮食成年组比较,高热量饮食老年组海马NAA、中心路程/总路程、平均速度、synaptophysin表达降低(P<0.05,P<0.01),iNOS、IL-1β表达升高(1.61±0.10 vs 1.35±0.13,2.04±0.08 vs 1.54±0.11,P<0.05,P<0.01)。结论高热量饮食导致ApoE^(-/-)小鼠代谢障碍和神经炎症,抑制突触蛋白表达引起认知功能障碍;长期高热量饮食和年龄增加促进ApoE^(-/-)小鼠脑功能衰退。
文摘目的研究C1q/肿瘤坏死因子相关蛋白9(CTRP9)对高脂血症模型载脂蛋白E基因敲除(ApoE^(-/-))小鼠肝脏脂肪病变形成的影响及机制。方法将30只8周龄雄性ApoE^(-/-)小鼠随机分为对照组与观察组,每组15只,对照组小鼠给予尾静脉注射腺病毒空载绿色荧光蛋白(Ad-GFP),观察组小鼠给予尾静脉注射腺病毒载CTRP9(Ad-CTRP9),高脂高胆固醇饮食(HF/HC)饲喂12周后,采集尾静脉血,处死小鼠收集肝脏组织。采用HE、油红O染色观察CTRP9对ApoE^(-/-)小鼠肝脏脂肪病变形成的影响;采用real time PCR检测CTRP9及脂质合成、炎症相关和线粒体氧化磷酸化相关基因的表达水平;采用western blotting技术检测肝脏组织中CTRP9蛋白表达水平。结果与对照组相比,过表达CTRP9小鼠(观察组)体重和血脂水平差异无统计学意义(P>0.05),CTRP9基因和蛋白表达水平显著升高(P<0.05),肝脏脂肪病变显著减少(P<0.05),肝脏组织中甘油三酯和总胆固醇水平显著下降(P<0.05),脂质合成(FAS、SCD1、SREBP1c)以及炎症相关基因(MCP-1、MIP1α和IL-1β)表达水平显著下调(P<0.01或P<0.001),而脂质代谢(PPARα、PPARγ、LXRα和LXRβ)及线粒体氧化磷酸化相关基因(CPT1、CPT2、COX4、Cytoc、Acadl和Acadm)表达水平显著增加(P<0.05或P<0.01)。结论过表达CTRP9能够抑制肝脏组织中脂肪生成以及炎症反应,同时促进肝细胞脂肪酸氧化磷酸化和能量代谢,增加线粒体能量消耗,从而降低肝脏组织中脂质蓄积。
