IFNτ 通过 STAT1 和 MAPK42/44 信号通路介导体外牛滋养层细胞的趋化性、运动性、代谢和 CK18 下调。

IFNτ mediates chemotaxis, motility, metabolism and CK18 downregulation in bovine trophoblast cells in vitro via STAT1 and MAPK42/44 signaling.

机构信息

Department of Anatomy, University of Veterinary Medicine Hannover, Germany.

出版信息

Placenta. 2018 Apr;64:17-26. doi: 10.1016/j.placenta.2018.02.004. Epub 2018 Feb 21.

DOI:10.1016/j.placenta.2018.02.004

PMID:29626977

Abstract

INTRODUCTION

IFNτ is the ruminant pregnancy recognition signal. In the study we investigated the autocrine influence of IFNτ on bovine F3 trophoblast cells. In detail chemotaxis, motility, metabolism, cell polarisation (CK18; ezrin) and the underlying classical (STAT1) and non-classical (MAPK42/44) signaling pathways were examined.

METHODS

Cellular signaling was analysed by densitometric Western blot (STAT1, MAPK42/44, proteinkinase B) and RT-PCR (IFNAR1, -2). Cellular assays were carried out for chemotaxis (agarose spot assay), cell motility (live cell imaging), metabolism (MTT) and cell polarisation (CK18; ezrin). In vivo-produced conceptuses of gestational days (GD) 20-39 underwent immunohistochemistry (CK18; ezrin) to set the in vitro findings (cell polarisation) in proportion to the in vivo situation.

RESULTS

IFNτ (10-1000 ng/ml) mediated dose-dependent effects. 10 ng/ml IFNτ induced chemotaxis and motility, whilst 1000 ng/ml led to reduced chemotaxis, motility and a 92-fold activation of MAPK44. Stimulation of cells with 10-1000 ng/ml IFNτ promoted metabolism (1.4-fold), increased the gene expression of IFNAR1/2 (24 h) and downregulated CK18 but not ezrin. All described in vitro effects were significant. Signaling, motility and metabolism could be blocked by specific inhibitors (PD98059, LY294002). CK18 and ezrin expression patterns in the trophoblast of in vivo conceptuses differed depending on GD.

DISCUSSION

IFNτ is a major factor for bovine F3 trophoblast cells and mediates a variety of cellular actions ranging from chemotaxis to polarisation. IFNτ exerts its effects via classical (STAT1) and non-classical (MAPK42/44) signaling pathways in a dose-dependent way. We hypothesize that (dose-dependent) IFNτ regulation of the cellular effects could also be essential for bovine elongation and implantation.

摘要

简介

IFNτ 是反刍动物妊娠识别信号。本研究探讨了 IFNτ 对牛 F3 滋养层细胞的自分泌影响。详细研究了趋化性、运动性、代谢、细胞极化（CK18；ezrin）以及潜在的经典（STAT1）和非经典（MAPK42/44）信号通路。

方法

通过密度计量 Western blot（STAT1、MAPK42/44、蛋白激酶 B）和 RT-PCR（IFNAR1、-2）分析细胞信号转导。进行趋化性（琼脂斑点分析）、细胞运动性（活细胞成像）、代谢（MTT）和细胞极化（CK18；ezrin）细胞检测。对妊娠第 20-39 天的体内生产的胚胎进行免疫组织化学（CK18；ezrin），使体外研究结果（细胞极化）与体内情况成比例。

结果

IFNτ（10-1000ng/ml）介导了剂量依赖性作用。10ng/ml IFNτ诱导趋化性和运动性，而 1000ng/ml IFNτ导致趋化性、运动性降低和 MAPK44 激活 92 倍。用 10-1000ng/ml IFNτ刺激细胞促进代谢（1.4 倍），增加 IFNAR1/2 基因表达（24 小时）并下调 CK18 但不影响 ezrin。所有描述的体外作用均有统计学意义。特定抑制剂（PD98059、LY294002）可阻断信号转导、运动性和代谢。体内胚胎滋养层的 CK18 和 ezrin 表达模式随妊娠天数而变化。