中华血液学杂志
中華血液學雜誌
중화혈액학잡지
Chinese Journal of Hematology
2008年
10期
662-666
,共5页
顾玲%李金范%高举%朱易萍%李强%贾苍松%周成燕%马志贵
顧玲%李金範%高舉%硃易萍%李彊%賈蒼鬆%週成燕%馬誌貴
고령%리금범%고거%주역평%리강%가창송%주성연%마지귀
雷帕霉素%哺乳动物雷帕霉素靶蛋白%间变性淋巴瘤激酶
雷帕黴素%哺乳動物雷帕黴素靶蛋白%間變性淋巴瘤激酶
뢰파매소%포유동물뢰파매소파단백%간변성림파류격매
Rapamycin%Mammalian target of rapamycin%Anaplastic lymphoma kinase
目的 探讨哺乳动物雷帕霉素靶蛋白(mTOR)信号通路与间变性淋巴瘤激酶(ALK)阳性淋巴瘤的关系.方法 采用Western blot法分析雷帕霉素作用于Karpas299、BaF3/NPM-ALK及BaF3细胞前后,mTOR相关蛋白表达的改变;MTT法检雷帕霉素对细胞增殖的影响;Annexin V/PI染色法检测雷帕霉素对细胞凋亡的影响;PI染色法检测细胞周期改变.结果 ALK+ Karpos299、BaF3/NPM-ALK和BaF3细胞均高表达mTOR信号通路磷酸化蛋白、磷酸化p70S6K(p-pTOS6K)及4E-BP1(p-4E-BP1)蛋白.去除IL-3 1 h的BaF3细胞p70S6K及4E-BP1蛋白去磷酸化.10 nmol/L雷帕霉素作用48 h,p-p70S6K及p-4E-BP1蛋白表达下降,以p-p70S6K蛋白表达下降为主.相对于对照组各细胞株的抑制率为Karpas299细胞24.4%、BaF3/NPM-ALK细胞37.8%和BaF3细胞61.6%.Karpas299细胞在10 nmol/L雷帕霉素作用后细胞凋亡率从(11.97±0.11)%增至(15.87±0.62)%(P<0.05);BaF3细胞凋亡率由(3.23±0.11)%增至(7.67±0.49)%(P<0.05);BaF3/NPM-ALK细胞凋亡率从(1.90±0.47)%增至(2.80±0.27)%(P>0.05),无诱导调亡作用.三株细胞在10 nmol/L雷帕霉素作用48 h后,G1期细胞比率均有不同程度的增加,以BaF3/NPM-ALK细胞最为显著,从(37.63±1.91)%增加至(69.77±5.44)%;其次为Karpas299细胞,从(31.13±2.51)%增至(40.70±1.47)%;BaF3细胞最少,由(53.57±2.22)%增至(63.70±1.20)%(P值均<0.05).结论 NPM-ALK融合蛋白激酶激活了mTOR信号通路;雷帕霉素通过抑制mTOR活性,使细胞周期停滞于G1期,从而抑制ALK+淋巴细胞肿瘤性生长.
目的 探討哺乳動物雷帕黴素靶蛋白(mTOR)信號通路與間變性淋巴瘤激酶(ALK)暘性淋巴瘤的關繫.方法 採用Western blot法分析雷帕黴素作用于Karpas299、BaF3/NPM-ALK及BaF3細胞前後,mTOR相關蛋白錶達的改變;MTT法檢雷帕黴素對細胞增殖的影響;Annexin V/PI染色法檢測雷帕黴素對細胞凋亡的影響;PI染色法檢測細胞週期改變.結果 ALK+ Karpos299、BaF3/NPM-ALK和BaF3細胞均高錶達mTOR信號通路燐痠化蛋白、燐痠化p70S6K(p-pTOS6K)及4E-BP1(p-4E-BP1)蛋白.去除IL-3 1 h的BaF3細胞p70S6K及4E-BP1蛋白去燐痠化.10 nmol/L雷帕黴素作用48 h,p-p70S6K及p-4E-BP1蛋白錶達下降,以p-p70S6K蛋白錶達下降為主.相對于對照組各細胞株的抑製率為Karpas299細胞24.4%、BaF3/NPM-ALK細胞37.8%和BaF3細胞61.6%.Karpas299細胞在10 nmol/L雷帕黴素作用後細胞凋亡率從(11.97±0.11)%增至(15.87±0.62)%(P<0.05);BaF3細胞凋亡率由(3.23±0.11)%增至(7.67±0.49)%(P<0.05);BaF3/NPM-ALK細胞凋亡率從(1.90±0.47)%增至(2.80±0.27)%(P>0.05),無誘導調亡作用.三株細胞在10 nmol/L雷帕黴素作用48 h後,G1期細胞比率均有不同程度的增加,以BaF3/NPM-ALK細胞最為顯著,從(37.63±1.91)%增加至(69.77±5.44)%;其次為Karpas299細胞,從(31.13±2.51)%增至(40.70±1.47)%;BaF3細胞最少,由(53.57±2.22)%增至(63.70±1.20)%(P值均<0.05).結論 NPM-ALK融閤蛋白激酶激活瞭mTOR信號通路;雷帕黴素通過抑製mTOR活性,使細胞週期停滯于G1期,從而抑製ALK+淋巴細胞腫瘤性生長.
목적 탐토포유동물뢰파매소파단백(mTOR)신호통로여간변성림파류격매(ALK)양성림파류적관계.방법 채용Western blot법분석뢰파매소작용우Karpas299、BaF3/NPM-ALK급BaF3세포전후,mTOR상관단백표체적개변;MTT법검뢰파매소대세포증식적영향;Annexin V/PI염색법검측뢰파매소대세포조망적영향;PI염색법검측세포주기개변.결과 ALK+ Karpos299、BaF3/NPM-ALK화BaF3세포균고표체mTOR신호통로린산화단백、린산화p70S6K(p-pTOS6K)급4E-BP1(p-4E-BP1)단백.거제IL-3 1 h적BaF3세포p70S6K급4E-BP1단백거린산화.10 nmol/L뢰파매소작용48 h,p-p70S6K급p-4E-BP1단백표체하강,이p-p70S6K단백표체하강위주.상대우대조조각세포주적억제솔위Karpas299세포24.4%、BaF3/NPM-ALK세포37.8%화BaF3세포61.6%.Karpas299세포재10 nmol/L뢰파매소작용후세포조망솔종(11.97±0.11)%증지(15.87±0.62)%(P<0.05);BaF3세포조망솔유(3.23±0.11)%증지(7.67±0.49)%(P<0.05);BaF3/NPM-ALK세포조망솔종(1.90±0.47)%증지(2.80±0.27)%(P>0.05),무유도조망작용.삼주세포재10 nmol/L뢰파매소작용48 h후,G1기세포비솔균유불동정도적증가,이BaF3/NPM-ALK세포최위현저,종(37.63±1.91)%증가지(69.77±5.44)%;기차위Karpas299세포,종(31.13±2.51)%증지(40.70±1.47)%;BaF3세포최소,유(53.57±2.22)%증지(63.70±1.20)%(P치균<0.05).결론 NPM-ALK융합단백격매격활료mTOR신호통로;뢰파매소통과억제mTOR활성,사세포주기정체우G1기,종이억제ALK+림파세포종류성생장.
Objective To investigate the relationship between mTOR signaling pathway and ALKpositive lymphoid cell lines. Methods The expression of the downstream effector proteins of mTOR were analyzed by Western blot before and after Karpas 299, BaF3/NPM-ALK and BaF3 cell lines treated with rapamycin. Effect of rapamycin on cell proliferation was detected by MTr assay. FACS was used to analyze apoptosis and cell cycles. Results mTOR signaling phosphoproteins, p-p70S6K and p-4E-BP1 were highly expressed in ALK+ Karpas299, BaF3/NPM-ALK and parental BaF3 cell lines, and they were dephosphorylated after 1 h withdrawal of IL-3 in BaF3 cells. After 48 h exposure to 10 nmoL/L rapamycin, p-p7OS6K and p-4EBP1 proteins expression were decreased, and mainly for the former. The relative inhibitory rate to its control cells was 24.4% in Karpas299, 37.8% in BaF3/NPM-ALK and 61.6% in BaF3. The apoptotic ratio was increased from (11.97±0.11)% to (15.87±0.62)% in Karpas299(P <0.05), from (3.23±0.11)% to (7.67±0.49)% in BaF3 (P<0.05) and from (1.90±0.47)% to (2.80±0.27)% in BaF3/NPMALK (P>0.05). The fraction of G<1> phase cells increased from (37.63±1.91)% to (69.77±5.44)% in BaF3/NPM-ALK, from (31.13±2.51) % to (40.70±1.47) % in Karpas299 and (53.57±2.22) % to (63.70±1.20) % in BaF3 (P < 0.05). Conclusion NPM-ALK kinase can activate mTOR signaling pathway. Rapamycin can inhibit the proliferation of ALK+ lymphoid cells by blocking mTOR signaling pathway and inducing cell cycling arrest at G1 phase.
