中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2011年
4期
503-505,后插1
,共4页
张韬%沈晨阳%李清乐%周冰莹%张小明
張韜%瀋晨暘%李清樂%週冰瑩%張小明
장도%침신양%리청악%주빙형%장소명
趋化素样因子1%核因子-κb%血管平滑肌细胞
趨化素樣因子1%覈因子-κb%血管平滑肌細胞
추화소양인자1%핵인자-κb%혈관평활기세포
Chemokine-like factor 1%Nuclear factor-κB%Vascular smooth muscle cell
目的 应用重组腺病毒载体介导的趋化素样因子1(CKLF1)基因转染大鼠血管平滑肌细胞(SMC),观察CKLF1对SMC内核因子(NF)-κB活性的影响.方法 用腺病毒介导的CKLFl以不同时间梯度转染刺激SMC,采用免疫细胞化学、荧光定量聚合酶链反应(PCR)和Western blot 等方法 分析细胞内CKLF1和NF-κB激活表达.结果 静息状态下SMC中NF-κB有基础表达量,经CKLF1刺激后2～14 d,其活性迅速增加,免疫细胞化学染色细胞质和核呈棕色,CKLF1荧光定量PCR和Western blot结果 分别为1.0500±0.0681、3.3090±0.1523、2.8540±0.1219、1.7050±0.0944、1.4350±0.0858和1.186±0.061、2.264±0.185、2.397±0.149、1.617±0.101、1.381±0.033,与对照组比较差异均有统计学意义(P＜0.05).NF-κB的Western blot结果 为1.184±0.111、2.046±0.221、1.747±0.068、1.498±0.051、1.412±0.087,与对照组比较5～14 d差异有统计学意义(P＜0.05).两组刺激后5～8 d为表达高峰期,且NF-κB活性与CKLF1表达量间明显相关.结论 CKLF1可导致NF-κB表达增加并随刺激时间梯度改变,提示NF-κB信号通路可能参与了CKLF1对SMC的增殖和迁移作用.
目的 應用重組腺病毒載體介導的趨化素樣因子1(CKLF1)基因轉染大鼠血管平滑肌細胞(SMC),觀察CKLF1對SMC內覈因子(NF)-κB活性的影響.方法 用腺病毒介導的CKLFl以不同時間梯度轉染刺激SMC,採用免疫細胞化學、熒光定量聚閤酶鏈反應(PCR)和Western blot 等方法 分析細胞內CKLF1和NF-κB激活錶達.結果 靜息狀態下SMC中NF-κB有基礎錶達量,經CKLF1刺激後2～14 d,其活性迅速增加,免疫細胞化學染色細胞質和覈呈棕色,CKLF1熒光定量PCR和Western blot結果 分彆為1.0500±0.0681、3.3090±0.1523、2.8540±0.1219、1.7050±0.0944、1.4350±0.0858和1.186±0.061、2.264±0.185、2.397±0.149、1.617±0.101、1.381±0.033,與對照組比較差異均有統計學意義(P＜0.05).NF-κB的Western blot結果 為1.184±0.111、2.046±0.221、1.747±0.068、1.498±0.051、1.412±0.087,與對照組比較5～14 d差異有統計學意義(P＜0.05).兩組刺激後5～8 d為錶達高峰期,且NF-κB活性與CKLF1錶達量間明顯相關.結論 CKLF1可導緻NF-κB錶達增加併隨刺激時間梯度改變,提示NF-κB信號通路可能參與瞭CKLF1對SMC的增殖和遷移作用.
목적 응용중조선병독재체개도적추화소양인자1(CKLF1)기인전염대서혈관평활기세포(SMC),관찰CKLF1대SMC내핵인자(NF)-κB활성적영향.방법 용선병독개도적CKLFl이불동시간제도전염자격SMC,채용면역세포화학、형광정량취합매련반응(PCR)화Western blot 등방법 분석세포내CKLF1화NF-κB격활표체.결과 정식상태하SMC중NF-κB유기출표체량,경CKLF1자격후2～14 d,기활성신속증가,면역세포화학염색세포질화핵정종색,CKLF1형광정량PCR화Western blot결과 분별위1.0500±0.0681、3.3090±0.1523、2.8540±0.1219、1.7050±0.0944、1.4350±0.0858화1.186±0.061、2.264±0.185、2.397±0.149、1.617±0.101、1.381±0.033,여대조조비교차이균유통계학의의(P＜0.05).NF-κB적Western blot결과 위1.184±0.111、2.046±0.221、1.747±0.068、1.498±0.051、1.412±0.087,여대조조비교5～14 d차이유통계학의의(P＜0.05).량조자격후5～8 d위표체고봉기,차NF-κB활성여CKLF1표체량간명현상관.결론 CKLF1가도치NF-κB표체증가병수자격시간제도개변,제시NF-κB신호통로가능삼여료CKLF1대SMC적증식화천이작용.
Objective By transfection of adenovirus vector-mediated chemokine-like factor 1 ( CKLF1 ) gene into rat vascular smooth muscle cells ( SMC), to observate the effect of CKLF1 in SMC on nuclear factor-κB (NF-κB) activity. Methods Adenovirus vector-mediated CKLF1 gege was transfected into SMC at different time group. Immunocytochemistry, real-time polymerase chain reaction (PCR) and Western blotting were used to detect the expression of CKLF1 and NF-κB. Results Resting SMC had a low baseline level of NF-κB activity, cells were activated by CKLF1 stimulation from 2 to 14 days, and the activity of NF-κB was rapidly increased. Immunohistochemically, cytoplasma and nucleus were stained as brown. The expression of CKLF1 mRNA and protein was 1. 0500 ± 0. 0681, 3. 3090 ± 0. 1523, 2. 8540 ± 0. 1219, 1. 7050 ±0. 0944, 1. 4350 ±0. 0858 and 1. 186 ±0. 061,2. 264 ±0. 185, 2. 397 ±0. 149, 1. 617±0. 101, 1. 381 ±0. 033 respectively from 2 to 14 days, with the difference being statistically significant (P＜0.05). Western blotting showed that NF-κB activity was 1. 184 ±0. 111, 2.046 ±0.221, 1.747 ±0. 068, 1. 498 ±0. 051, 1. 412 ±0. 087 respectively, with the difference being statistically significant from 5 to 14 days ( P ＜ 0. 05 ). All results suggested that 5 to 8 days after stimulation the expression reached the peak, and the was an obvious correlation between NF-κB activity and CKLF1 expression. Conclusion CKLF1 can enhance the expression of NF-κB in SMC, suggesting that NF-κB signaling pathway might take part in the effect of CKLF1 on SMC proliferation and migration.
