Nicotine exposure during differentiation causes inhibition of N-myc expression.

BACKGROUND: 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. METHODS: Fibroblasts obtained from non-human primate ESC (nhpESC) differentiation were analyzed by immunohistochemistry, immunostaining, Affymetrix gene expression array, qPCR, and immunoblotting. RESULTS: 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. CONCLUSIONS: 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.

Effects of exogenous estrogen on uterine leukocyte recruitment.

OBJECTIVE: To determine the role of estrogen in leukocyte recruitment to the human endometrium. DESIGN: Prospective, controlled in vivo study. SETTING: Academic research laboratory. PATIENT(S): Ten patients presenting for donor oocytes. INTERVENTION(S): Endometrial biopsies for the evaluation of leukocyte populations were collected from perimenopausal women in two consecutive regulated cycles who were given two different regimens of estrogen with identical progesterone treatment. MAIN OUTCOME MEASURE(S): Immunohistochemical identification of endometrial leukocyte populations and relative levels of expression of three chemokine genes. RESULT(S): The total uterine leukocyte population increased significantly when the women received oral estrogen, which resulted in higher serum estrogen levels. This rise in leukocytes was due to a significant increase in both the uterine natural killer cells and the macrophage populations. T-cell numbers did not change relative to circulating estrogen levels. The relative abundance of mRNA from three chemokines was also determined. No changes were found in the expression of M-CSF or MCP-1. Interleukin 8 decreased in glands relative to estrogen levels. CONCLUSION(S): These data demonstrate that changes in circulating levels of estrogen can regulate the recruitment of bone marrow-derived cells to the uterine endometrium; however, the mechanism whereby that occurs remains elusive.