Zhou Yong-Zhi, Li Chu-Zhong, Gao Hua, Zhang Ya-Zhuo
Capital Medical University, Beijing Neurosurgical Institute, Beijing Tiantan Hospital, Beijing, China.
Beijing Neurosurgical Institute, Beijing Tiantan Hospital, Beijing Institute for Brain Disorders Brain Tumor Center, China National Clinical Research Center for Neurological Diseases, Key Laboratory of Central Nervous System Injury Research, Capital Medical University, Beijing, China.
World Neurosurg. 2018 Jun;114:e329-e337. doi: 10.1016/j.wneu.2018.02.181. Epub 2018 Mar 8.
Down-regulation of mothers against decapentaplegic homolog 3 (Smad3) results in the formation of tumors both in vivo and in vitro. However, little is known about the effect of Smad3 on adrenocorticotropic hormone-secreting pituitary adenomas (ACTH-PAs). Our objective was to study the expression and effect of Smad3 in ACTH-PAs and its possible mechanisms.
Smad3, COOH-terminally phosphorylated mothers against decapentaplegic homolog 3 (pSmad3), and mothers against decapentaplegic homolog 2 proteins (Smad2) were detected in samples from 5 normal anterior pituitaries and 18 ACTH-PAs by Western blot and immunohistochemical analysis. Then, Smad3 expression was up-regulated by Smad3-CMV plasmid or down-regulated by small interfering RNA in ACTH tumor cells (AtT-20) in vitro. Cell proliferation, apoptosis, ACTH level, and pSmad3, B-cell lymphoma/lewkmia-2 (BCL-2), and pro-opiomelanocortin (POMC) protein expression in the AtT-20 cells were measured to investigate the antitumor effects of Smad3.
Reduced expression of Smad3 and pSmad3 but unchanged Smad2 levels were found in ACTH-PAs compared with normal pituitaries. In vitro, the overexpression of Smad3 inhibited cell proliferation, promoted cell apoptosis, and decreased ACTH secretion; in contrast, Smad3 knockdown increased cell proliferation and decreased cell apoptosis but had no significant effect on ACTH secretion. At the same time, overexpression of Smad3 increased pSmad3 but inhibited BCL-2 and POMC protein expression. On the contrary, underexpression of Smad3 inhibited pSmad3 but promoted BCL-2 and POMC protein expression.
Smad3 is underexpressed in ACTH-PAs. Reversing the expression of Smad3 in AtT-20 cells could suppress cell growth, promote tumor apoptosis, and decrease ACTH secretion. Tumor suppression was possibly mediated by the promotion of pSmad3 and the reduction of BCL-2 and POMC expression.
抑制母亲对脱磷酸化蛋白同源物3（Smad3）会在体内和体外导致肿瘤形成。然而，关于Smad3对促肾上腺皮质激素分泌型垂体腺瘤（ACTH-PAs）的影响知之甚少。我们的目的是研究Smad3在ACTH-PAs中的表达、作用及其可能机制。
通过蛋白质免疫印迹法和免疫组织化学分析，检测5例正常垂体前叶组织和18例ACTH-PAs样本中的Smad3、羧基末端磷酸化的母亲对脱磷酸化蛋白同源物3（pSmad3）和母亲对脱磷酸化蛋白同源物2（Smad2）蛋白。然后，在体外通过Smad3-CMV质粒上调ACTH肿瘤细胞（AtT-20）中Smad3的表达，或通过小干扰RNA下调其表达。检测AtT-20细胞的增殖、凋亡、ACTH水平以及pSmad3、B细胞淋巴瘤/白血病-2（BCL-2）和阿黑皮素原（POMC）蛋白表达，以研究Smad3的抗肿瘤作用。
与正常垂体相比，ACTH-PAs中Smad3和pSmad3表达降低，但Smad2水平无变化。在体外，Smad3过表达抑制细胞增殖，促进细胞凋亡，并降低ACTH分泌；相反，敲低Smad3增加细胞增殖，减少细胞凋亡，但对ACTH分泌无显著影响。同时，Smad3过表达增加pSmad3，但抑制BCL-2和POMC蛋白表达。相反，Smad3表达不足抑制pSmad3，但促进BCL-2和POMC蛋白表达。
Smad3在ACTH-PAs中表达不足。在AtT-20细胞中逆转Smad3的表达可抑制细胞生长，促进肿瘤凋亡，并减少ACTH分泌。肿瘤抑制可能是通过促进pSmad3以及降低BCL-2和POMC表达介导的。
