目的探讨细胞周期依赖性蛋白激酶抑制剂roscovitine对C型尼曼-皮克病(NPC)小鼠(npc-/-)球状神经轴突形成的影响。方法采用不同浓度的roscovitine(72,144,300 and 600nmoles/day)对5.5周龄的npc-/-小鼠进行两周侧脑室灌注,以DMSO灌注的同...目的探讨细胞周期依赖性蛋白激酶抑制剂roscovitine对C型尼曼-皮克病(NPC)小鼠(npc-/-)球状神经轴突形成的影响。方法采用不同浓度的roscovitine(72,144,300 and 600nmoles/day)对5.5周龄的npc-/-小鼠进行两周侧脑室灌注,以DMSO灌注的同龄npc-/-小鼠为对照。抗非磷酸化神经细丝蛋白抗体SMI32和抗有丝分裂期磷酸化表位抗体MPM-2免疫染色检测小鼠脑内球状神经轴突的数量,Western blot检测细胞骨架蛋白磷酸化程度的变化。结果Roscovitine组球状神经轴突数量较DMSO组显著减少,神经细丝和tau蛋白的过度磷酸化程度较对照组明显下降(P<0.05)。结论细胞周期依赖性蛋白激酶抑制剂能明显改善NPC的神经元细胞骨架损害。展开更多
The tumor necrosis factor （TNF）-related apoptosis-inducing ligand （TRAIL/APO2L） is a member of the TNF gene superfamily that induces apoptosis upon engagement of cognate death receptors. While TRAIL is relatively ...The tumor necrosis factor （TNF）-related apoptosis-inducing ligand （TRAIL/APO2L） is a member of the TNF gene superfamily that induces apoptosis upon engagement of cognate death receptors. While TRAIL is relatively non-toxic to normal cells, it selectively induces apoptosis in many transformed cells. Nevertheless, breast tumor cells are particularly resistant to the effects of TRAIL. Here we report that, in combination with the cyclin-dependent kinase inhibitor roscovitine, exposure to TRAIL induced marked apoptosis in the majority of TRAIL-resistant breast cancer cell lines examined. Roscovitine facilitated TRAIL death-inducing signaling complex formation and the activation of caspase-8. The cFLIPL and cFLIPs FLICE-inhibitory proteins were significantly down-regulated following exposure to roscovitine and, indeed, the knockdown of cFLIP isoforms by siRNA sensitized breast tumor cells to TRAIL-induced apoptosis. In addition, we demonstrate that roscovitine strongly suppressed Mcl-1 expression and up-regulated E2F1 protein levels in breast tumor ceils. Significantly, the silencing of Mci-1 by siRNA sensitized breast tumor cells to TRAIL-induced apoptosis. Furthermore, the knockdown of E2F1 protein by siRNA reduced the sensitizing effect of roscovitine in TRAIL-induced apoptosis. In summary, our results reveal a pleitropic mechanism for the pro-apoptotic influence of roscovitine, highlighting its potential as an antitumor agent in breast cancer in combination with TRAIL.展开更多
为了明确蓝藻中丝氨酸/苏氨酸激酶的功能是否与调控细胞的生长分裂相关,以丝状鱼腥藻7120、单细胞集胞藻6803和聚球藻7002为对象,利用OD750光吸收测定和MTT方法研究了不同浓度丝氨酸苏氨酸激酶抑制剂roscovitine对其生长和脱氢酶活性的...为了明确蓝藻中丝氨酸/苏氨酸激酶的功能是否与调控细胞的生长分裂相关,以丝状鱼腥藻7120、单细胞集胞藻6803和聚球藻7002为对象,利用OD750光吸收测定和MTT方法研究了不同浓度丝氨酸苏氨酸激酶抑制剂roscovitine对其生长和脱氢酶活性的影响。结果表明:4 h roscovitine处理后对鱼腥藻7120和集胞藻6803生长量影响不大,对聚球藻7002的生长有促进作用。4 h roscovitine的处理对鱼腥藻7120有浓度依赖的显著抑制活性,对集胞藻6803的活性无影响,但是却促进聚球藻7002的活性。药物作用4 d后,7120的生长和活性均显著降低,并有浓度效应;6803的生长量较对照减少,但活性变化不明显;聚球藻7002的生长和活性均未受影响。显微观察结果显示,roscovitine对3种细胞形态没有影响,但药物作用4 d后的7120藻丝体较短。结果表明丝氨酸/苏氨酸抑制剂roscovitine影响丝状藻7120的生长和活性。展开更多
Abnormal proliferation and migration of vascular smooth muscle cells （VSMCs） are the major cause of in-stent restenosis （ISR）. Intervention proliferation and migration of VSMCs is an im- portant strategy for antir...Abnormal proliferation and migration of vascular smooth muscle cells （VSMCs） are the major cause of in-stent restenosis （ISR）. Intervention proliferation and migration of VSMCs is an im- portant strategy for antirestenotic therapy. Roscovitine, a second-generation cyclin-dependent kinase in- hibitor, can inhibit cell cycle of multiple cell types. We studied the effects of roscovitine on cell cycle distribution, proliferation and migration of VSMCs in vitro by flow cytometry, BrdU incorporation and wound healing assay, respectively. Our results showed that roscovitine increased the proportion of Go/G1 phase cells after 12 h （69.57±3.65 vs. 92.50±1.68, P=0.000）, 24 h （80.87±2.24 vs. 90.25±0.79, P=0.000） and 48 h （88.08±3.86 vs. 88.87±2.43, P=-0.427） as compared with control group. Roscovifine inhibited proliferation and migration of VSMCs in a concentration-dependent way. With the increase of concen- tration, roscovitine showed increased capacity for growth and migration inhibition. Roscovitine （30 μmol/L） led to an almost complete VSMCs growth and migration arrest. Combined with its low toxicity and selective inhibition to ISR-VSMCs, roscovitine may be a potential drug in the treatment of vascular stenosis diseases and particularly useful in the prevention and treatment of ISR.展开更多
文摘目的探讨细胞周期依赖性蛋白激酶抑制剂roscovitine对C型尼曼-皮克病(NPC)小鼠(npc-/-)球状神经轴突形成的影响。方法采用不同浓度的roscovitine(72,144,300 and 600nmoles/day)对5.5周龄的npc-/-小鼠进行两周侧脑室灌注,以DMSO灌注的同龄npc-/-小鼠为对照。抗非磷酸化神经细丝蛋白抗体SMI32和抗有丝分裂期磷酸化表位抗体MPM-2免疫染色检测小鼠脑内球状神经轴突的数量,Western blot检测细胞骨架蛋白磷酸化程度的变化。结果Roscovitine组球状神经轴突数量较DMSO组显著减少,神经细丝和tau蛋白的过度磷酸化程度较对照组明显下降(P<0.05)。结论细胞周期依赖性蛋白激酶抑制剂能明显改善NPC的神经元细胞骨架损害。
文摘The tumor necrosis factor （TNF）-related apoptosis-inducing ligand （TRAIL/APO2L） is a member of the TNF gene superfamily that induces apoptosis upon engagement of cognate death receptors. While TRAIL is relatively non-toxic to normal cells, it selectively induces apoptosis in many transformed cells. Nevertheless, breast tumor cells are particularly resistant to the effects of TRAIL. Here we report that, in combination with the cyclin-dependent kinase inhibitor roscovitine, exposure to TRAIL induced marked apoptosis in the majority of TRAIL-resistant breast cancer cell lines examined. Roscovitine facilitated TRAIL death-inducing signaling complex formation and the activation of caspase-8. The cFLIPL and cFLIPs FLICE-inhibitory proteins were significantly down-regulated following exposure to roscovitine and, indeed, the knockdown of cFLIP isoforms by siRNA sensitized breast tumor cells to TRAIL-induced apoptosis. In addition, we demonstrate that roscovitine strongly suppressed Mcl-1 expression and up-regulated E2F1 protein levels in breast tumor ceils. Significantly, the silencing of Mci-1 by siRNA sensitized breast tumor cells to TRAIL-induced apoptosis. Furthermore, the knockdown of E2F1 protein by siRNA reduced the sensitizing effect of roscovitine in TRAIL-induced apoptosis. In summary, our results reveal a pleitropic mechanism for the pro-apoptotic influence of roscovitine, highlighting its potential as an antitumor agent in breast cancer in combination with TRAIL.
文摘为了明确蓝藻中丝氨酸/苏氨酸激酶的功能是否与调控细胞的生长分裂相关,以丝状鱼腥藻7120、单细胞集胞藻6803和聚球藻7002为对象,利用OD750光吸收测定和MTT方法研究了不同浓度丝氨酸苏氨酸激酶抑制剂roscovitine对其生长和脱氢酶活性的影响。结果表明:4 h roscovitine处理后对鱼腥藻7120和集胞藻6803生长量影响不大,对聚球藻7002的生长有促进作用。4 h roscovitine的处理对鱼腥藻7120有浓度依赖的显著抑制活性,对集胞藻6803的活性无影响,但是却促进聚球藻7002的活性。药物作用4 d后,7120的生长和活性均显著降低,并有浓度效应;6803的生长量较对照减少,但活性变化不明显;聚球藻7002的生长和活性均未受影响。显微观察结果显示,roscovitine对3种细胞形态没有影响,但药物作用4 d后的7120藻丝体较短。结果表明丝氨酸/苏氨酸抑制剂roscovitine影响丝状藻7120的生长和活性。
基金supported by grants from the National Natural Science Foundation of China(Nos.30870641 and 81030021)the National Basic Research of China "973" Program(No.2011CB504403)
文摘Abnormal proliferation and migration of vascular smooth muscle cells （VSMCs） are the major cause of in-stent restenosis （ISR）. Intervention proliferation and migration of VSMCs is an im- portant strategy for antirestenotic therapy. Roscovitine, a second-generation cyclin-dependent kinase in- hibitor, can inhibit cell cycle of multiple cell types. We studied the effects of roscovitine on cell cycle distribution, proliferation and migration of VSMCs in vitro by flow cytometry, BrdU incorporation and wound healing assay, respectively. Our results showed that roscovitine increased the proportion of Go/G1 phase cells after 12 h （69.57±3.65 vs. 92.50±1.68, P=0.000）, 24 h （80.87±2.24 vs. 90.25±0.79, P=0.000） and 48 h （88.08±3.86 vs. 88.87±2.43, P=-0.427） as compared with control group. Roscovifine inhibited proliferation and migration of VSMCs in a concentration-dependent way. With the increase of concen- tration, roscovitine showed increased capacity for growth and migration inhibition. Roscovitine （30 μmol/L） led to an almost complete VSMCs growth and migration arrest. Combined with its low toxicity and selective inhibition to ISR-VSMCs, roscovitine may be a potential drug in the treatment of vascular stenosis diseases and particularly useful in the prevention and treatment of ISR.
