Department of Pediatrics, West China Second University Hospital of Sichuan University, Chengdu, Sichuan, China (mainland).
Key Laboratory of Birth Defects and Related Diseases of Women and Children, Sichuan University, Ministry of Education, Chengdu, Sichuan, China (mainland).
Med Sci Monit. 2017 Sep 12;23:4391-4400. doi: 10.12659/msm.906568.
BACKGROUND Previous studies revealed physiological and pathogenetic similarity between vascular smooth muscles cells with severe pulmonary arterial hypertension and tumors. The DNA damage response was found in both pulmonary arterial hypertension (PAH) cells and tumors. The ataxia-telangiectasia mutated proteins (ATM) pathway is considered an important factor in the DNA damage response of tumor formation, but its function in the development of PAH remains unknown. MATERIAL AND METHODS The Sprague-Dawley rat PAH model was established. Three weeks (Group M1), 5 weeks (Group M2), and 7 weeks (Group M3) after drug injection, pulmonary expression of ATM, Checkpoint kinase 2 (Chk2), P53, and P21 were measured. A section of the lungs from Group M2 was used for pulmonary artery vascular smooth muscles cells (PA-SMCs) isolation and culture. The effect of KU60019 in the proliferation and apoptosis of primary cultured rat PA-SMCs was measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and TdT-mediated dUTP nick-end labeling (TUNEL), respectively. RESULTS Immunohistochemistry results show that the expression of ATM, Chk2, and P21 increased in Groups M1 and M2, and decreased in Group M3. Additionally, expression of P53 increased in Group M1, and decreased in Groups M2 and M3. RT-PCR and Western blotting demonstrated that in Groups M1 and M2, the expression of ATM, Chk2, P53, and P21 increased, whereas it decreased in Group M3. In cell culture, 0.3 μM and 0.5 μM KU60019 increased the growth of PA-SMCs, and 0.5 μM KU60019 reduced cell apoptosis. CONCLUSIONS Expression of the ATM-Chk2 pathway increased in early stages of PAH formation, but decreased in late stages. In primary cultured PA-SMCs, KU60019 increased cell proliferation and inhibited cell apoptosis.
先前的研究表明，患有严重肺动脉高压的血管平滑肌细胞与肿瘤在生理和发病机制上具有相似性。在肺动脉高压（PAH）细胞和肿瘤中均发现了 DNA 损伤反应。共济失调毛细血管扩张突变蛋白（ATM）途径被认为是肿瘤形成中 DNA 损伤反应的重要因素，但它在 PAH 发展中的作用尚不清楚。
建立 Sprague-Dawley 大鼠 PAH 模型。在药物注射后 3 周（M1 组）、5 周（M2 组）和 7 周（M3 组），测量肺组织中 ATM、检查点激酶 2（Chk2）、P53 和 P21 的表达。使用 M2 组的一部分肺组织分离和培养肺动脉血管平滑肌细胞（PA-SMCs）。通过 3-(4,5-二甲基-2-噻唑基)-2,5-二苯基-2-H-四唑溴盐（MTT）和末端转移酶介导的 dUTP 缺口末端标记（TUNEL）分别测量 KU60019 对原代培养的大鼠 PA-SMCs 增殖和凋亡的影响。
免疫组织化学结果显示，ATM、Chk2 和 P21 的表达在 M1 组和 M2 组中增加，在 M3 组中减少。此外，P53 的表达在 M1 组中增加，在 M2 组和 M3 组中减少。RT-PCR 和 Western blot 表明，在 M1 组和 M2 组中，ATM、Chk2、P53 和 P21 的表达增加，而在 M3 组中表达减少。在细胞培养中，0.3μM 和 0.5μM KU60019 增加了 PA-SMCs 的生长，而 0.5μM KU60019 减少了细胞凋亡。
在 PAH 形成的早期阶段，ATM-Chk2 途径的表达增加，但在晚期阶段减少。在原代培养的 PA-SMCs 中，KU60019 增加了细胞增殖并抑制了细胞凋亡。
