胡羊,一个独特的中国品种,具有高繁殖性能,在中国也以其罕见的白羊皮而闻名。羊皮的质量受毛囊的影响,毛乳头细胞是毛囊的重要组成部分,在毛囊的生长发育中起着关键作用。本研究有助于阐明miR-148a和miR-10a对毛囊生长发育的影响。
根据基因芯片和高通量测序的结果,以骨形态发生蛋白7(BMP7)为研究对象。生物信息学分析和双荧光素酶报告系统表明,同时用Westernblot和实时定量聚合酶链反应(qRT-PCR)证明miR-148a和miR-10a与BMP7的靶关系。通过双荧光素酶报告系统和蛋白质印迹,BMP7是miR-148a和miR-10a的靶基因。成功分离纯化了湖羊毛乳头细胞,将miR-148a/miR-10a模拟物和抑制剂转染到毛乳头细胞后,细胞计数试剂盒-8(CCK-8)用于确定miR-148a/miR-10a抑制湖羊毛乳头细胞的增殖。此外,miR-148a过表达后,Smad3的表达水平(P<0.05),Smad6(P<0.05),Smad4(P<0.01),Smad5和Smad5显著高于对照组(P<0.01)。抑制miR-148a后,Smad3的表达水平(P<0.05),Smad4(P<0.05),TGF-β和TGF-β明显低于对照组(P<0.01)。miR-10a过表达后,Smad1的表达水平(P<0.01),Smad2(P<0.05),Smad4(P<0.01),Smad5(P<0.01),TGF-β水平明显低于对照组(P<0.05)。抑制miR-10a后,Smad1(P<0.01)和Smad2(P<0.05)的表达水平明显低于对照组。
这些结果揭示了miR-148a,miR-10a和BMP7,以及miR-148a和miR-10a对毛乳头细胞增殖的影响。他们将为miR-148a的后续研究提供基础,miR-10a介导BMP7对毛囊生长发育的调控。
Hu sheep, a unique Chinese breed with high reproductive performance, are also well known for their rare white lambskin in China. The quality of lambskin is affected by hair follicles, and dermal papilla cells are an important component of hair follicles that plays a key role in hair follicle growth and development. This
study helps elucidate the effect of miR-148a and miR-10a on hair follicle growth and development.
Based on the results of gene chip and high-throughput sequencing, bone morphogenetic protein 7 (BMP7) was used as a research object. Bioinformatics analysis and the dual-luciferase reporter system indicated that, along with Western blot and quantitative real-time polymerase chain reaction (qRT-PCR) that miR-148a and miR-10a target relationships with BMP7. BMP7 was the target gene both for miR-148a and miR-10a by the dual-luciferase reporter system and Western blot. Hu sheep dermal papilla cells were successfully isolated and purified, and after transfecting miR-148a/miR-10a mimics and inhibitors into dermal papilla cells, a Cell Counting Kit-8 (CCK-8) was used to determine that miR-148a/miR-10a inhibited the proliferation of Hu sheep dermal papilla cells. In addition, after the overexpression of miR-148a, the expression levels of Smad3 (P < 0.05), Smad6 (P < 0.05), Smad4 (P < 0.01), and Smad5 (P < 0.01) were significantly higher than those of the control groups. After the inhibition of miR-148a, the expression levels of Smad3 (P < 0.05), Smad4 (P < 0.05), and TGF-β (P < 0.01) were significantly lower than those of the control groups. After the overexpression of miR-10a, the expression levels of Smad1 (P < 0.01), Smad2 (P < 0.05), Smad4 (P < 0.01), Smad5 (P < 0.01), and TGF-β (P < 0.05) were significantly lower than those of the control groups. After the inhibition of miR-10a, the expression levels of Smad1 (P < 0.01) and Smad2 (P < 0.05) were significantly lower than those of the control groups.
These results revealed the target relationship between miR-148a, miR-10a and BMP7, and the effect of miR-148a and miR-10a on the proliferation of dermal papilla cells. They will provide the basis for a follow-up
study on how miR-148a, and miR-10a mediate BMP7 regulation of hair follicle growth and development.