CAJ | 학술논문

目的探讨吡格列酮(Pio)调节树突状细胞(DC)黏附和迁移功能的可能机制.方法体外培养人外周血单核细胞来源的DC和人脐静脉内皮细胞,使用不同浓度的Pio及加过氧化物酶体增殖物活化受体-γ拮抗剂(GW9662)干预DC 24 h,Western印迹和免疫荧光试验检测Pio对表达DC特异性细胞间黏附分子3捕获的非整合蛋白(DC-SIGN)的影响,细胞黏附试验检测Pio对DC黏附功能的作用,细胞迁移试验检测Pio对DC迁移功能的作用.结果 (1)Pio呈浓度依赖性下调DC-SIGN蛋白的表达,在Pio 1.0 μmol/L时DC-SIGN蛋白表达量明显减少(0.96 ±0.09,对照1.25±0.23,均P＜0.05),Pio 10 μmol/L时减少更明显(0.80 ±0.08,均P＜0.01),GW9662干预组(1.10±0.12,均P＜0.01)蛋白表达量增加;(2)在Pio 1.0 μmol/L时DC黏附和迁移降低(10.8%±2.0%和24.6%±0.5%,均P＜0.05),Pio 10 μmol/L时降低更明显(7.6%±1.5%和23.4%±3.0%,均P＜0.01),GW9662干预组DC黏附和迁移增加(12.1%±1.9%和26.8%±0.8%),经抗DC-SIGN抗体作用后DC黏附和迁移较对照降低(5.7%±0.9%和23.2%±2.9%,均P＜0.01).结论 Pio能通过激活过氧化物酶体增殖物活化受体-γ,下调DC-SIGN蛋白表达,从而抑制DC的黏附和迁移功能.
목적 탐토필격렬동(Pio)조절수돌상세포(DC)점부화천이공능적가능궤제.방법 체외배양인외주혈단핵세포래원적DC화인제정맥내피세포,사용불동농도적Pio급가과양화물매체증식물활화수체-γ길항제(GW9662)간예DC 24 h,Western인적화면역형광시험검측Pio대표체DC특이성세포간점부분자3포획적비정합단백(DC-SIGN)적영향,세포점부시험검측Pio대DC점부공능적작용,세포천이시험검측Pio대DC천이공능적작용.결과 (1)Pio정농도의뢰성하조DC-SIGN단백적표체,재Pio 1.0 μmol/L시DC-SIGN단백표체량명현감소(0.96 ±0.09,대조1.25±0.23,균P＜0.05),Pio 10 μmol/L시감소경명현(0.80 ±0.08,균P＜0.01),GW9662간예조(1.10±0.12,균P＜0.01)단백표체량증가;(2)재Pio 1.0 μmol/L시DC점부화천이강저(10.8%±2.0%화24.6%±0.5%,균P＜0.05),Pio 10 μmol/L시강저경명현(7.6%±1.5%화23.4%±3.0%,균P＜0.01),GW9662간예조DC점부화천이증가(12.1%±1.9%화26.8%±0.8%),경항DC-SIGN항체작용후DC점부화천이교대조강저(5.7%±0.9%화23.2%±2.9%,균P＜0.01).결론 Pio능통과격활과양화물매체증식물활화수체-γ,하조DC-SIGN단백표체,종이억제DC적점부화천이공능.
Objective To investigate the effect of pioglitazone(Pio)on dendritic cell-(DC)specific intercellular adhesion molecule-3 grabbing nonintegrin(DC-SIGN)expression in DCs and explore the possible mechanism of Pio inhibiting DC adhesion and transmigration.Methods DCs derived from human peripheral blood mononuclear cells were cultured and divided into 6 groups:blank control group.Pio 0.1 μmol/L group,Pio 1.0 μmol/L group,Pio 10 μmol/L group,GW9662,a peroxisome proliferator activated receptor(PPAR)-γ antagonist,10 μmol/L group,and GW9662 10 μmol/L+Pio 10 μmol/Lgroup.Western blotting was used to detect the protein expression of DC-SIGN 24 h later.Hnman umbilical vein endothelial cells(HUVECs)were obtained and co-cuhured witll the DCs undergoing different treatmdnts.Immunofluorescence test was used to detect the protein expression of DC-SIGN.DCs labeled with 5-ehloromethylfluorescein diacetate(CMFDA)were added into the monocellular layer of fused Ecs.Blank DCs and DCs pretreated witll anti-DC-SIGN antibody were used as blank and experimental groups.Laser confocal microscopy was used to observe the adhesion ability of the DCs.HUVECs were inoculated into the upper chamber of Transwell plate and CMFDA-labeled DCs of above mentioned groups were added to the mono-cellular layer of these Ecs.Serum.free culture mediam witIl monocyte chemoattractant protein-1 was added into the lower chamber.Eight hours later the transmigration ability was observed.Results Westem blotting showed that the DC-SIGN protein expression levels of the DCs of tlle Pio 1.0 μmol/L and Pio 10 μmol/L group were 0.96 ±0.09 and 0.80±0.08 respectively.both significanfly lower than that of the blank control group(1.25±0.23,P＜0.05 and P＜0.01);and tlle DC-SIGN protein expression level of the GW9662 10 μmol/L+Pio 10 μmol/L group was 1.10 ±0.12.significantly higher than that of the Pio 10 μmol/L group(P＜0.05).Immunofluorescence test showed that the DC-SIGN protein expression levels of the DCs of the Pio 1.0 μmol/L and Pio 10 μmol/L group were 22.3 ±5.4 and 14.4±2.3 respectively,both signifieanfly lower than that of the control group(29.5±5.1,P＜0.05 and P＜0.01),and the DC-SIGN protein expression level of the GW9662 10 μmoL/L+Pio 10 μmol/L group was 2,4.9 ±4.3.significantly higher than that of the Pio 10 μmol/L group(P＜0.01),and not significantly different from that of the blank control group(P＞0.05).The adhesion rates of the Pio 1.0 μmol/L and Pio 10 μmol/L groups were 10.8%±2.0% and 7.6%±1.5% respectively.both significantly lower than that of the control group(13.4%±2.1%,P＜0.05 and P＜0.01);and the adhesion rate of the GW9662+Pio 10 μmol/L group was 12.1%±1.9%,significantly higher than that of the Pio 10 μmol/L group(P＜0.01),and not significantly different from that of the blank control group(P＞0.05).The transmigration inhibition rate of DCs of the Pio 0.1,1.0,and 10 μmol/L groups were 4.1%.12.9%,and 17.2% compared with the transmigration rate of the blank control group(P＜0.05,P＜0.05,and P＜0.01).The transmigration rate of the GW9662+Pio 10 μmol/L group was significantly higher than that of the Pi0 10 μmol/L group (P＜0.05),and not significantly different from that of the blank control group(P＞0.05).The transmigration rate of the anti-DC-SIGN intervention group was lower by 17.8%than that of the control group (P＜0.01).Conclusion Pio down-regulates the DC-SIGN protein expression and inhibits DC adhesion and transmigration tllreugh the pathway of PPAR-γ.