Division of Developmental and Regenerative Medicine, Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA 15213, USA.
Respir Res. 2013 Nov 5;14(1):119. doi: 10.1186/1465-9921-14-119.
The ability of chemicals to disrupt neonatal development can be studied using embryonic stem cells (ESC). One such chemical is nicotine. Prenatal nicotine exposure is known to affect postnatal lung function, although the mechanisms by which it has this effect are not clear. Since fibroblasts are a critical component of the developing lung, providing structure and secreting paracrine factors that are essential to epithelialization, this study focuses on the differentiation of ESC into fibroblasts using a directed differentiation protocol.
Fibroblasts obtained from non-human primate ESC (nhpESC) differentiation were analyzed by immunohistochemistry, immunostaining, Affymetrix gene expression array, qPCR, and immunoblotting.
Results of these analyses demonstrated that although nhpESCs differentiate into fibroblasts in the presence of nicotine and appear normal by some measures, including H&E and SMA staining, they have an altered gene expression profile. Network analysis of expression changes demonstrated an over-representation of cell-cycle related genes with downregulation of N-myc as a central regulator in the pathway. Further investigation demonstrated that cells differentiated in the presence of nicotine had decreased N-myc mRNA and protein expression and longer doubling times, a biological effect consistent with downregulation of N-myc.
This study is the first to use primate ESC to demonstrate that nicotine can affect cellular differentiation from pluripotency into fibroblasts, and in particular, mediate N-myc expression in differentiating ESCs. Given the crucial role of fibroblasts throughout the body, this has important implications for the effect of cigarette smoke exposure on human development not only in the lung, but in organogenesis in general.
化学物质干扰新生发育的能力可以通过胚胎干细胞（ESC）进行研究。尼古丁就是这样一种化学物质。已知产前接触尼古丁会影响出生后的肺功能，但其作用机制尚不清楚。由于成纤维细胞是发育中肺的关键组成部分，提供结构并分泌旁分泌因子，对上皮化至关重要，因此本研究专注于使用定向分化方案将 ESC 分化为成纤维细胞。
通过免疫组织化学、免疫染色、Affymetrix 基因表达谱、qPCR 和免疫印迹分析从非人类灵长类动物 ESC（nhpESC）分化获得的成纤维细胞。
这些分析的结果表明，尽管 nhpESCs 在尼古丁存在的情况下分化为成纤维细胞，并且在某些方面（包括 H&E 和 SMA 染色）看起来正常，但它们具有改变的基因表达谱。表达变化的网络分析表明，细胞周期相关基因的过表达，以 N-myc 作为该途径的中央调节剂下调。进一步的研究表明，在尼古丁存在下分化的细胞 N-myc mRNA 和蛋白表达减少，倍增时间延长，这是与 N-myc 下调一致的生物学效应。
本研究首次使用灵长类 ESC 证明尼古丁可以影响从多能性到成纤维细胞的细胞分化，特别是调节分化中的 ESC 中的 N-myc 表达。鉴于成纤维细胞在全身的重要作用，这对香烟烟雾暴露对人类发育的影响具有重要意义，不仅在肺部，而且在一般的器官发生中。
