■与两栖动物相比,哺乳动物的肢体再生有限。Wnt信号通路在肢体再生中的作用很少被研究。所以,这项研究旨在研究使用化学品CHIR99021和IWP-2的Wnt信号对体外器官培养系统中截肢小鼠手指尖端再生的影响。
■截肢了C57BL/6J小鼠胎儿在E14.5,E16.5和E18.5处的爪子的远端指骨。然后,手培养7天。随后,用1-50µg/ml浓度的CHIR99021和5-10µg/ml浓度的IWP-2处理爪子。最后,通过组织学分析评估新的组织再生,BC的免疫组织化学,TCF1,CAN,K14和P63基因,用RT-qPCR评估β-catenin和Tcf1基因。
■7天后,将E14.5和E16.5天的爪子收缩并压缩,所以选择了活的18.5E的爪子。因此,在25和30μl/ml浓度的CHR99021组中观察到手指尖端处的新生长的质量,但在IWP2治疗中未观察到(*P<0.05;**P<0.01)。qRT-PCR分析证实,与IWP-2组相比,CHIR99021组中β-catenin和Tcf1基因的显着上调(P<0.05)。此外,Alcian-blue染色证明在CHIR组中再生团块处存在软骨样组织。在免疫组织化学分析中β-连环蛋白,ACN,在CHIR处理组中,在指尖中观察到角蛋白-14和P63蛋白表达。
■通过激活Wnt信号通路,在小鼠截肢的指尖的胚层样肿块中形成软骨样组织。
UNASSIGNED: Mammals have limited limb regeneration compared to amphibians. The role of Wnt signaling pathways in limb regeneration has rarely been studied. So, this study aimed to investigate the effect of Wnt-signaling using chemicals CHIR99021 and IWP-2 on amputated mice digit tips regeneration in an in vitro organ culture system.
UNASSIGNED: The distal phalanx of paws from C57BL/6J mouse fetuses at E14.5, E16.5, and E18.5 was amputated. Then, the hands were cultured for 7 days. Subsequently, paws were treated with 1-50 µg/ml concentration of CHIR99021 and 5-10 µg/ml concentration of IWP-2. Finally, the new tissue regrowth was assessed by histological analysis, immunohistochemistry for BC, TCF1, CAN, K14, and P63 genes, and beta-catenin and Tcf1 genes were evaluated with RT-qPCR.
UNASSIGNED: The paws of E14.5 and E16.5 days were shrinkaged and compressed after 7 days, so the paws of 18.5E that were alive were selected. As a result, newly-grown masses at digit tips were observed in 25 and 30 µl/ml concentrations of the CHR99021 group but not in the IWP2 treatment (*P<0.05; **P<0.01). qRT-PCR analysis confirmed the significant up-regulation of beta-catenin and Tcf1 genes in CHIR99021 group in comparison to the IWP-2 group (P<0.05). Moreover, Alcian-blue staining demonstrated the presence of cartilage-like tissue at regenerated mass in the CHIR group. In immunohistochemistry analysis beta-catenin, ACN, Keratin-14, and P63 protein expression were observed in digit tips in the CHIR-treated group.
UNASSIGNED: By activating the Wnt signaling pathway, cartilage-like tissue formed in the blastema-like mass in the mouse\'s amputated digit tips.