目的·初步探究成纤维细胞生长因子2（fibroblast growth factor 2，FGF2）对小鼠结肠CD34⁺CD81⁺间质细胞R-spondin 1（Rspo1）表达的调控作用及机制。方法·利用流式细胞术分选Rspo1-tdTomato荧光报告基因小鼠结肠中CD45^－CD326^－CD31^－GP38⁺CD81⁺Rspo1-tdTomato⁺细胞，并进行体外培养。运用流式细胞术检测培养14 d后该类细胞表面蛋白标志物分子的表达情况。通过实时荧光定量PCR（quantitative real-time PCR，qPCR）检测该细胞分别在FGF2、FGF9、表皮生长因子（epidermal growth factor，EGF）、血小板衍生生长因子-bb（platelet-derived growth factor-bb，PDGFbb）、胰岛素样生长因子1（insulin-like growth factor 1，IGF1）、肝细胞生长因子（hepatocyte growth factor，HGF）刺激条件下Rspo1的表达情况。通过转录组测序和生物信息学方法分析FGF2调控Rspo1表达的信号通路，并用信号通路抑制剂及qPCR进行初步验证。结果·流式分选得到的小鼠结肠间质细胞经过体外培养14 d后，能够表达CD34、CD81和糖蛋白GP38，而不表达CD45、CD326、CD31等其他细胞谱系标志分子。qPCR结果发现，20 ng/mL的FGF2即可显著抑制该细胞Rspo1的表达，其他生长因子则无显著抑制作用。转录组测序和生物信息学分析结果显示，丝裂原活化蛋白激酶（mitogen-activated protein kinase，MAPK）信号通路可能是FGF2调节Rspo1表达的关键信号通路。qPCR结果显示，丝裂原细胞外激酶1/2（mitogen extracellular kinase 1/2，MEK1/2）抑制剂U0216预处理可逆转FGF2对Rspo1表达的抑制作用。结论·FGF2可能通过MEK1/2-细胞外调节蛋白激酶1/2（extracellular regulated protein kinase 1/2，ERK1/2）通路抑制小鼠结肠CD34⁺CD81⁺间质细胞Rspo1的表达。

Objective ·To preliminarily investigate the regulatory effect and underlying mechanism of fibroblast growth factor 2 (FGF2) on R-spondin 1 (Rspo1) expression in CD34⁺CD81⁺ stromal cells from the mouse colon. Methods ·Colonic CD45^-CD326^-CD31^-GP38⁺CD81⁺Rspo1-tdTomato⁺ stromal cells were sorted from Rspo1-tdTomato reporter mice by flow cytometry and subsequently cultured in vitro. The expression of surface protein markers was evaluated by flow cytometry after 14 d of culture. qPCR was employed to quantify Rspo1 expression in response to stimulation with FGF2, FGF9, epidermal growth factor (EGF), platelet-derived growth factor-bb (PDGF-bb), insulin-like growth factor 1 (IGF1), or hepatocyte growth factor (HGF). RNA sequencing and bioinformatic analyses were used to identify the signaling pathways underlying FGF2-mediated regulation of Rspo1, followed by preliminary validation with pathway-specific inhibitors and qPCR. Results ·After 14 d of culture, the sorted colonic stromal cells retained expression of CD34, CD81, and glycoprotein GP38, while remaining negative for other lineages markers CD45, CD326, and CD31. qPCR revealed that 20 ng/mL FGF2 significantly suppressed Rspo1 expression, whereas the other tested growth factors exerted no notable effect. RNA sequencing and bioinformatic analysis indicated that mitogen-activated protein kinase (MAPK) signaling pathway played a key role in the regulatory effect of FGF2 on Rspo1. qPCR further demonstrated that pretreatment with U0126, an inhibitor of mitogen extracellular kinase 1/2 (MEK1/2), reversed FGF2-mediated suppression of Rspo1 expression. Conclusion ·FGF2 may inhibit Rspo1 expression in mouse colonic CD34⁺CD81⁺ stromal cells via the MEK1/2-extracellular regulated protein kinase 1/2 (ERK1/2) signaling pathway.