中华糖尿病杂志
中華糖尿病雜誌
중화당뇨병잡지
CHINES JOURNAL OF DLABETES MELLITUS
2013年
9期
536-540
,共5页
姜黄素%受体相互作用蛋白140%炎症%血管内皮细胞
薑黃素%受體相互作用蛋白140%炎癥%血管內皮細胞
강황소%수체상호작용단백140%염증%혈관내피세포
Curcumin%Receptor interaction protein l40%Inflammation%Vascular endothelial cells
目的 明确姜黄素对炎症诱导的血管内皮细胞损伤的保护机制.方法 将人单核/巨噬细胞系(THP-1)分为空白对照组、高脂高糖组、高脂高糖+姜黄素预处理组(高脂高糖浓度为25mmol/L葡萄糖+500 μmol/L棕榈酸),分别给予相应处理24 h,更换培养基继续培养24 h,利用酶联免疫吸附法(ELISA)检测上清及实时定量聚合酶链反应(RT-PCR)分析THP-1细胞内肿瘤坏死因子-α(TNF-α)、白介素-6(IL-6)的表达,同时将上清作用脐静脉血管内皮细胞(HUVECs),HUVECs分为空白对照组、空对照条件培养基组、高脂高糖条件培养基组、姜黄素处理条件培养基组,并行噻唑蓝法(MTT)检测HUVECs增殖、流式细胞仪检测凋亡,Western blotting分析磷酸化细胞外调节蛋白激酶(p-ERK)及B淋巴细胞瘤-2(Bcl-2)蛋白表达.组间比较采用独立样本t检验,多组间比较采用单因素方差分析.结果 高脂高糖组THP-1细胞内受体相互作用蛋白140(RIP140)、TNF-α和IL-6mRNA表达及上清中炎症因子TNF-α和IL-6浓度明显高于空白对照组(t=8.55、9.44、9.73、16.01、19.22,均P＜0.05).相对于高脂高糖组,姜黄素预处理组THP-1细胞内RIP140、TNF-α和IL-6mRNA表达及上清中TNF-α和IL-6浓度明显下降(t=3.59、5.96、5.59、6.95、23.91,均P＜0.05)；高脂高糖条件培养基组HUVECs细胞活力明显低于空对照条件培养基组(24 h,1.22 ±0.07比1.85±0.14,t=6.58,P＜0.05;48 h,1.72±0.02比2.49±0.09,t=10.08,P＜0.05),而姜黄素处理条件培养基组能够改善高脂高糖条件培养基对HUVECs增殖的抑制作用(24 h,1.22±0.07比1.72 ±0.11,t=2.13,P＜0.05;48 h,1.72 ±0.02比2.33 ±0.11,t=6.92,P＜0.05)；与空对照条件培养基组比较,高脂高糖条件培养基组能够明显增加HUVECs凋亡率、p-ERK表达及降低Bcl-2表达(=9.82、9.69、4.61,均P＜0.05),姜黄素处理条件培养基组与高脂高糖条件培养基组比较,下调RIP140表达后能明显降低HUVECs凋亡率、p-ERK表达及增加Bcl-2表达(t=6.35、7.17、3.26,均P＜0.05).结论 姜黄素能够通过抑制高脂高糖诱导的炎症来调控HUVECs的增殖凋亡.
目的 明確薑黃素對炎癥誘導的血管內皮細胞損傷的保護機製.方法 將人單覈/巨噬細胞繫(THP-1)分為空白對照組、高脂高糖組、高脂高糖+薑黃素預處理組(高脂高糖濃度為25mmol/L葡萄糖+500 μmol/L棕櫚痠),分彆給予相應處理24 h,更換培養基繼續培養24 h,利用酶聯免疫吸附法(ELISA)檢測上清及實時定量聚閤酶鏈反應(RT-PCR)分析THP-1細胞內腫瘤壞死因子-α(TNF-α)、白介素-6(IL-6)的錶達,同時將上清作用臍靜脈血管內皮細胞(HUVECs),HUVECs分為空白對照組、空對照條件培養基組、高脂高糖條件培養基組、薑黃素處理條件培養基組,併行噻唑藍法(MTT)檢測HUVECs增殖、流式細胞儀檢測凋亡,Western blotting分析燐痠化細胞外調節蛋白激酶(p-ERK)及B淋巴細胞瘤-2(Bcl-2)蛋白錶達.組間比較採用獨立樣本t檢驗,多組間比較採用單因素方差分析.結果 高脂高糖組THP-1細胞內受體相互作用蛋白140(RIP140)、TNF-α和IL-6mRNA錶達及上清中炎癥因子TNF-α和IL-6濃度明顯高于空白對照組(t=8.55、9.44、9.73、16.01、19.22,均P＜0.05).相對于高脂高糖組,薑黃素預處理組THP-1細胞內RIP140、TNF-α和IL-6mRNA錶達及上清中TNF-α和IL-6濃度明顯下降(t=3.59、5.96、5.59、6.95、23.91,均P＜0.05)；高脂高糖條件培養基組HUVECs細胞活力明顯低于空對照條件培養基組(24 h,1.22 ±0.07比1.85±0.14,t=6.58,P＜0.05;48 h,1.72±0.02比2.49±0.09,t=10.08,P＜0.05),而薑黃素處理條件培養基組能夠改善高脂高糖條件培養基對HUVECs增殖的抑製作用(24 h,1.22±0.07比1.72 ±0.11,t=2.13,P＜0.05;48 h,1.72 ±0.02比2.33 ±0.11,t=6.92,P＜0.05)；與空對照條件培養基組比較,高脂高糖條件培養基組能夠明顯增加HUVECs凋亡率、p-ERK錶達及降低Bcl-2錶達(=9.82、9.69、4.61,均P＜0.05),薑黃素處理條件培養基組與高脂高糖條件培養基組比較,下調RIP140錶達後能明顯降低HUVECs凋亡率、p-ERK錶達及增加Bcl-2錶達(t=6.35、7.17、3.26,均P＜0.05).結論 薑黃素能夠通過抑製高脂高糖誘導的炎癥來調控HUVECs的增殖凋亡.
목적 명학강황소대염증유도적혈관내피세포손상적보호궤제.방법 장인단핵/거서세포계(THP-1)분위공백대조조、고지고당조、고지고당+강황소예처리조(고지고당농도위25mmol/L포도당+500 μmol/L종려산),분별급여상응처리24 h,경환배양기계속배양24 h,이용매련면역흡부법(ELISA)검측상청급실시정량취합매련반응(RT-PCR)분석THP-1세포내종류배사인자-α(TNF-α)、백개소-6(IL-6)적표체,동시장상청작용제정맥혈관내피세포(HUVECs),HUVECs분위공백대조조、공대조조건배양기조、고지고당조건배양기조、강황소처리조건배양기조,병행새서람법(MTT)검측HUVECs증식、류식세포의검측조망,Western blotting분석린산화세포외조절단백격매(p-ERK)급B림파세포류-2(Bcl-2)단백표체.조간비교채용독립양본t검험,다조간비교채용단인소방차분석.결과 고지고당조THP-1세포내수체상호작용단백140(RIP140)、TNF-α화IL-6mRNA표체급상청중염증인자TNF-α화IL-6농도명현고우공백대조조(t=8.55、9.44、9.73、16.01、19.22,균P＜0.05).상대우고지고당조,강황소예처리조THP-1세포내RIP140、TNF-α화IL-6mRNA표체급상청중TNF-α화IL-6농도명현하강(t=3.59、5.96、5.59、6.95、23.91,균P＜0.05)；고지고당조건배양기조HUVECs세포활력명현저우공대조조건배양기조(24 h,1.22 ±0.07비1.85±0.14,t=6.58,P＜0.05;48 h,1.72±0.02비2.49±0.09,t=10.08,P＜0.05),이강황소처리조건배양기조능구개선고지고당조건배양기대HUVECs증식적억제작용(24 h,1.22±0.07비1.72 ±0.11,t=2.13,P＜0.05;48 h,1.72 ±0.02비2.33 ±0.11,t=6.92,P＜0.05)；여공대조조건배양기조비교,고지고당조건배양기조능구명현증가HUVECs조망솔、p-ERK표체급강저Bcl-2표체(=9.82、9.69、4.61,균P＜0.05),강황소처리조건배양기조여고지고당조건배양기조비교,하조RIP140표체후능명현강저HUVECs조망솔、p-ERK표체급증가Bcl-2표체(t=6.35、7.17、3.26,균P＜0.05).결론 강황소능구통과억제고지고당유도적염증래조공HUVECs적증식조망.
Objective To investigate the protective role of curcumin in the dysfunctional vascular endothelial cells induced by inflammation.Methods THP-1 cells were pretreated with 20 μmol/L curcumin under the condition of 25 mmol/L glucose and 500 μmol/L palmitic acid,while other groups received 25 mmol/L glucose and 500 μmoL/L palmitic acid and dimethyl sulfoxide (DMSO),seperately.After 24 h treatment,the real time Polymerase Chain Reaction (PCR) was explored to detect the mRNA expression of receptor interaction protein 140(RIP140),tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6),while the concentration of TNF-α and IL-6 in the supernatant was detected by the enzyme-linked immunosorbent assay (ELISA).Human umbilical vein endothelial cells (HUVECs) were incubated with the supernatant from the THP-1 in each group.The methodology of 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTF) assay,flow cytometry and western-blot was applied to investigate the proliferation,apoptosis and the protein expression of phosphorylated extracellular regulated protein kinases (ERK) and B-cell lymphoma 2 (Bcl-2).Results The mRNA expression of RIP140,TNF-α and IL-6 in the THP-1 cells and the conceutration of TNF-α and IL-6 in the supernatant were significantly higher when treated with 25 mmol/L glucose and 500 μ,mol/L palmitic acid,than those in the control group (t =8.55,9.44,9.73,16.01,19.22 with all P ＜ 0.05).In the groups treated with 25 mmol/L glucose and 500 μmol/L palmitic acid,the pretreatment with curcumin significantly decreased the mRNA expression of RIP140,TNF-α and IL-6 in the THP-1 cells and the concentration of TNF-α and IL-6 in the supernatant (t =3.59,5.96,5.59,6.95,23.91 with all P ＜ 0.05),At 24 h and 48 h treatment,compared with the control group,the supertant from the groups treated with 25 mmol/L glucose and 500 μ mol/L palmitic acid significantly suppressed the proliferation of HUVECs (24 h:1.22 ± 0.07 vs 1.85 ± 0.14,t =6.58,P ＜0.05 ; 48 h:1.72 ± 0.02 vs 2.49 ± 0.09,t =10.08,P ＜ 0.05),while the proliferation of HUVECs could be improved by the pretreatment with 20 μmol/L cureumin (24 h:1.22 ± 0.07 vs 1.72 ± 0.11,t =2.13,P ＜ 0.05 ; 48 h:1.72 ± 0.02 vs 2.33 ± 0.11,t =6.92,P ＜ 0.05).Compared with the control group,the incubation with the supertant from the groups treated with 25 mmol/L glucose and 500 μmol/L palmitic acid significantly increased the apoptosis rate and the level of phosphated ERK,while decreased the Bcl-2 protein expression (t =9.82,9.69,4.61,all P ＜ 0.05).Such effect could be reserved by the pretreatment with 20μmol/L curcumin (t =6.35,7.17,3.26,all P ＜ 0.05).Conclusion Curcumin could protect the vascular endothelial cells from the dysfunction induced by inflammation.
