Epothilone은 현재 항암제로 사용되고 있는 탁솔이나 탁소텔 등과 같이 Microtubule(미세소관)을 안정화시켜 세포분열을 억제함으로써 암 세포의 증식을 막아 줄 수 있는 물질이다. 혈관 평활근세포 (vascular smooth muscle cell, VSMC)의 증식은 동맥경화증의 주요 원인으로 알려져 있다. 따라서 본 논문에서는 Epothilone의 과도한 세포증식 억제 가능성을 이용하여 Human colon cancer SW620 세포와 혈관평활근세포(VSMC) 증식에 대한 억제효과와 작용 기작을 규명했다. Epothilone B와 D를 처리한 군의 경우 대장암 세포를 농도의존적으로 저해하였으며 epothilone들에 의해 cell cycle이 arrest (G2/M) 되어 apoptosis가 일어났다. epothilone B 와 D 는 IB kinase (IKKα and IKKβ)를 통해 IkB degradation을 유발시켜 NF-kB를 활성화 시켰으며 결과적으로 nucleus로 p50 과 p65의 translocation을 일으켰다. 또한 epothilone B 와 D은 β-tubulin polymerization과 p50/β-tubulin complex의 형성도 증가시켰다. IKK 저해제인 Sodium salicylic acid을 첨가하거나 IKKα 변이주를 사용하였을 때에 β-tubulin polymerization은 증가되지 않았으나 NF-B dependent apoptotic cell death regulatory genes인 ...
Epothilone은 현재 항암제로 사용되고 있는 탁솔이나 탁소텔 등과 같이 Microtubule(미세소관)을 안정화시켜 세포분열을 억제함으로써 암 세포의 증식을 막아 줄 수 있는 물질이다. 혈관 평활근세포 (vascular smooth muscle cell, VSMC)의 증식은 동맥경화증의 주요 원인으로 알려져 있다. 따라서 본 논문에서는 Epothilone의 과도한 세포증식 억제 가능성을 이용하여 Human colon cancer SW620 세포와 혈관평활근세포(VSMC) 증식에 대한 억제효과와 작용 기작을 규명했다. Epothilone B와 D를 처리한 군의 경우 대장암 세포를 농도의존적으로 저해하였으며 epothilone들에 의해 cell cycle이 arrest (G2/M) 되어 apoptosis가 일어났다. epothilone B 와 D 는 IB kinase (IKKα and IKKβ)를 통해 IkB degradation을 유발시켜 NF-kB를 활성화 시켰으며 결과적으로 nucleus로 p50 과 p65의 translocation을 일으켰다. 또한 epothilone B 와 D은 β-tubulin polymerization과 p50/β-tubulin complex의 형성도 증가시켰다. IKK 저해제인 Sodium salicylic acid을 첨가하거나 IKKα 변이주를 사용하였을 때에 β-tubulin polymerization은 증가되지 않았으나 NF-B dependent apoptotic cell death regulatory genes인 Bax, p53, caspase-3는 증가시켰다. Caspase-3 inhibitor, 사용시 epothilone B-induced cell death와 NF-B 활성을 감소 시켰다. 이상의 결과를 통해 NF-B/IKK 신호가 tubulin polymerization에 비의존적으로 epothilone에 의해 유도되는 대장암세포 SW620 사멸에 중요한 역할을 담당하는 것을 알 수 있었다. 흰쥐경동맥 손상모델에서 Epothilone B와 D의 신생내막형성 억제효과와 흰쥐경동맥 평활근세포(VSMC)의 증식을 촉진 시키는 성장인자의 변화도 조사 하였다. 혈소판유래 성장인자인 PDGF-BB처리 시 apoptosis가 유도되지 않았으나, EPOs를 첨가한 실험결과 epothilone B와 epothilone D는 유의적으로 VSMC의 세포사멸을 유도하였으며 전형적인 세포사멸 염색사의 형태적 변화와 함께 세포의 원형화와 플레이트로부터 탈착이 관찰 되었다. EPOs 처리군의 경우 cell cycle이 arrest (G0/G1) 되어 apoptosis 가 일어나는 것을 알 수 있었다. 또한 EPOs는 세포핵내의 p53발현을 증가시켰으며 이들 p53의 전좌(translocation)가 세포사멸의 주요한 요인임을 확인 하였다. 신호전달 연구를 통해 epothilone B와 D가 p53의 발현과 축적을 유도하였으며 p53조절 caspase-3와 Bax가 활성화 되는 것을 알 수 있었다. 이러한 현상은 Paclitaxel을 양성대조군으로 사용한rat carotid artery balloon-injury model의 동물실험을 통해 epothilone B, D, 그리고 Paclitaxel투여로 신생내막형성이 유의적으로 억제되는 것을 확인하였다. 이상의 결과를 통해 epothilones의 신생내막형성억제 효과는 EPOs에 의해 활성화된p53-dependent caspase-3 경로를 경유한 VSMC 의 세포사멸에 기인함을 알 수 있었다.
Epothilone은 현재 항암제로 사용되고 있는 탁솔이나 탁소텔 등과 같이 Microtubule(미세소관)을 안정화시켜 세포분열을 억제함으로써 암 세포의 증식을 막아 줄 수 있는 물질이다. 혈관 평활근세포 (vascular smooth muscle cell, VSMC)의 증식은 동맥경화증의 주요 원인으로 알려져 있다. 따라서 본 논문에서는 Epothilone의 과도한 세포증식 억제 가능성을 이용하여 Human colon cancer SW620 세포와 혈관평활근세포(VSMC) 증식에 대한 억제효과와 작용 기작을 규명했다. Epothilone B와 D를 처리한 군의 경우 대장암 세포를 농도의존적으로 저해하였으며 epothilone들에 의해 cell cycle이 arrest (G2/M) 되어 apoptosis가 일어났다. epothilone B 와 D 는 IB kinase (IKKα and IKKβ)를 통해 IkB degradation을 유발시켜 NF-kB를 활성화 시켰으며 결과적으로 nucleus로 p50 과 p65의 translocation을 일으켰다. 또한 epothilone B 와 D은 β-tubulin polymerization과 p50/β-tubulin complex의 형성도 증가시켰다. IKK 저해제인 Sodium salicylic acid을 첨가하거나 IKKα 변이주를 사용하였을 때에 β-tubulin polymerization은 증가되지 않았으나 NF-B dependent apoptotic cell death regulatory genes인 Bax, p53, caspase-3는 증가시켰다. Caspase-3 inhibitor, 사용시 epothilone B-induced cell death와 NF-B 활성을 감소 시켰다. 이상의 결과를 통해 NF-B/IKK 신호가 tubulin polymerization에 비의존적으로 epothilone에 의해 유도되는 대장암세포 SW620 사멸에 중요한 역할을 담당하는 것을 알 수 있었다. 흰쥐경동맥 손상모델에서 Epothilone B와 D의 신생내막형성 억제효과와 흰쥐경동맥 평활근세포(VSMC)의 증식을 촉진 시키는 성장인자의 변화도 조사 하였다. 혈소판유래 성장인자인 PDGF-BB처리 시 apoptosis가 유도되지 않았으나, EPOs를 첨가한 실험결과 epothilone B와 epothilone D는 유의적으로 VSMC의 세포사멸을 유도하였으며 전형적인 세포사멸 염색사의 형태적 변화와 함께 세포의 원형화와 플레이트로부터 탈착이 관찰 되었다. EPOs 처리군의 경우 cell cycle이 arrest (G0/G1) 되어 apoptosis 가 일어나는 것을 알 수 있었다. 또한 EPOs는 세포핵내의 p53발현을 증가시켰으며 이들 p53의 전좌(translocation)가 세포사멸의 주요한 요인임을 확인 하였다. 신호전달 연구를 통해 epothilone B와 D가 p53의 발현과 축적을 유도하였으며 p53조절 caspase-3와 Bax가 활성화 되는 것을 알 수 있었다. 이러한 현상은 Paclitaxel을 양성대조군으로 사용한rat carotid artery balloon-injury model의 동물실험을 통해 epothilone B, D, 그리고 Paclitaxel투여로 신생내막형성이 유의적으로 억제되는 것을 확인하였다. 이상의 결과를 통해 epothilones의 신생내막형성억제 효과는 EPOs에 의해 활성화된p53-dependent caspase-3 경로를 경유한 VSMC 의 세포사멸에 기인함을 알 수 있었다.
Epothilones have been reported to trigger apoptotic cell death in hyperproliferative cells such as cancer cells and smooth muscle cells. However their effectiveness on cancer and smooth muscle cell proliferation and molecular mechanisms underlying epothilone-induced apoptotic cell death are limited....
Epothilones have been reported to trigger apoptotic cell death in hyperproliferative cells such as cancer cells and smooth muscle cells. However their effectiveness on cancer and smooth muscle cell proliferation and molecular mechanisms underlying epothilone-induced apoptotic cell death are limited. In the present study, I investigated the effectiveness of epothilones on cancer and smooth muscle cell proliferation, and molecular mechanisms underlying epothilone-induced apoptotic cell death were investigated in SW620 human colon cancer cells and in rat aortic vascular smooth muscle cell (VSMCs). Treatment with epothilone B and D at different concentrations (1-100 nM) dose dependently inhibited cell growth, and caused cell cycle arrest at G2-M, which was followed by apoptosis. Consistent with this induction of apoptotic cell death, epothilone B and D enhanced the constitutional activation of NF-kB via IkB degradation through IB kinase (IKKα and IKKβ) activation and this resulted in p50 and p65 translocation to the nucleus. Moreover, cells treated with sodium salicylic acid, an IKK inhibitor, or transiently transfected with mutant IKKα and β did not show epothilone-induced cell growth inhibition or p50 translocation, though p65 was still translocated to the nucleus. Treatment with epothilone B and D also enhanced β-tubulin polymerization and the formation of p50/β-tubulin complex. However, β-tubulin polymerization was not inhibited in the cells treated by sodium salicylic acid or transiently transfected with mutant IKKα and β. Moreover, epothilone B and D increased the expressions of NF-B dependent apoptotic cell death regulatory genes, i.e., Bax, p53 and the active form of caspase-3, but reduced Bcl-2 expression, and these actions were partially reversed by salicylic acid. In addition, caspase-3 inhibitor, reduced epothilone B-induced cell death and NF-B activation. These findings suggest that the activation of NF-B/IKK signals plays an important role in the epothilone-induced apoptotic cell death of SW620 colon cancer cells in a tubulin polymerization independent manner. It was previously found that epothilone (EPO)-B and EPO-D, novel synthesized microtubule stabilizing agents, inhibited neointimal hyperplasia after in vivo rat carotid artery injury as well as the growth factor activated proliferation of VSMCs. This study investigated whether epothilones can inhibit VSMC proliferation, and prevent neointimal formation through induction of apoptotic cell death, and then subsequently investigated the role of p53 and caspase-3 pathway in apoptotic process. EPO-B and EPO-D significantly induced apoptosis of VSMC in vitro in a dose dependent manner, and balloon-injury-induced neointimal hyperproliferated VSMC of the rat carotid artery resulting in significant inhibition of VSMC proliferation. In further mechanism study, it was found that EPOs potently induced expression and nuclear accumulation of p53. The p53-regulatory apoptotic proteins caspase-3 was concomitantly activated by EPOs treatment in cultured VSMCs and in the neointimal hyperplasic apoptotic areas of EPO-treated rat carotid arteries. These results suggest that the anti-proliferative and anti-neointimal hyperplasia effects of EPOs are due to the apoptotic cell deaths of hyper-proliferated VSMCs via the activation of the p53-dependent caspase-3 pathway. The present results show that epothilones may be developed as a potential antiproliferative agent for treatment of colon cancer and atherosclerosis. Furthermore, epothilones should be examined for further clinical application either as a single agent or in combination with other anti-colon cancer or atherosclerosis agents.
Epothilones have been reported to trigger apoptotic cell death in hyperproliferative cells such as cancer cells and smooth muscle cells. However their effectiveness on cancer and smooth muscle cell proliferation and molecular mechanisms underlying epothilone-induced apoptotic cell death are limited. In the present study, I investigated the effectiveness of epothilones on cancer and smooth muscle cell proliferation, and molecular mechanisms underlying epothilone-induced apoptotic cell death were investigated in SW620 human colon cancer cells and in rat aortic vascular smooth muscle cell (VSMCs). Treatment with epothilone B and D at different concentrations (1-100 nM) dose dependently inhibited cell growth, and caused cell cycle arrest at G2-M, which was followed by apoptosis. Consistent with this induction of apoptotic cell death, epothilone B and D enhanced the constitutional activation of NF-kB via IkB degradation through IB kinase (IKKα and IKKβ) activation and this resulted in p50 and p65 translocation to the nucleus. Moreover, cells treated with sodium salicylic acid, an IKK inhibitor, or transiently transfected with mutant IKKα and β did not show epothilone-induced cell growth inhibition or p50 translocation, though p65 was still translocated to the nucleus. Treatment with epothilone B and D also enhanced β-tubulin polymerization and the formation of p50/β-tubulin complex. However, β-tubulin polymerization was not inhibited in the cells treated by sodium salicylic acid or transiently transfected with mutant IKKα and β. Moreover, epothilone B and D increased the expressions of NF-B dependent apoptotic cell death regulatory genes, i.e., Bax, p53 and the active form of caspase-3, but reduced Bcl-2 expression, and these actions were partially reversed by salicylic acid. In addition, caspase-3 inhibitor, reduced epothilone B-induced cell death and NF-B activation. These findings suggest that the activation of NF-B/IKK signals plays an important role in the epothilone-induced apoptotic cell death of SW620 colon cancer cells in a tubulin polymerization independent manner. It was previously found that epothilone (EPO)-B and EPO-D, novel synthesized microtubule stabilizing agents, inhibited neointimal hyperplasia after in vivo rat carotid artery injury as well as the growth factor activated proliferation of VSMCs. This study investigated whether epothilones can inhibit VSMC proliferation, and prevent neointimal formation through induction of apoptotic cell death, and then subsequently investigated the role of p53 and caspase-3 pathway in apoptotic process. EPO-B and EPO-D significantly induced apoptosis of VSMC in vitro in a dose dependent manner, and balloon-injury-induced neointimal hyperproliferated VSMC of the rat carotid artery resulting in significant inhibition of VSMC proliferation. In further mechanism study, it was found that EPOs potently induced expression and nuclear accumulation of p53. The p53-regulatory apoptotic proteins caspase-3 was concomitantly activated by EPOs treatment in cultured VSMCs and in the neointimal hyperplasic apoptotic areas of EPO-treated rat carotid arteries. These results suggest that the anti-proliferative and anti-neointimal hyperplasia effects of EPOs are due to the apoptotic cell deaths of hyper-proliferated VSMCs via the activation of the p53-dependent caspase-3 pathway. The present results show that epothilones may be developed as a potential antiproliferative agent for treatment of colon cancer and atherosclerosis. Furthermore, epothilones should be examined for further clinical application either as a single agent or in combination with other anti-colon cancer or atherosclerosis agents.
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