Neudesin Neurotrophic Factor Promotes Bovine Preadipocyte Differentiation and Inhibits Myoblast Myogenesis

Simple Summary Neudesin neurotrophic factor (NENF) is a secreted protein that was significantly inhibited in the fat-muscle co-culture system in our previous study. However, studies on NENF regulation of bovine muscle development and involvement in the cross-talk between adipose tissue and skeletal muscle have not been reported. Hence, the aim of this study was to clarify the roles of NENF in bovine myoblast and preadipocyte differentiation. In this study, we first examined the spatial expression patterns of NENF in different tissues and found that NENF was highly expressed in the muscle of four-day-old and 24-month-old Qinchuan cattle. Compared with 4-day-old Qinchuan cattle, the expression level of NENF was significantly up-regulated in 24-month-old bovine adipose tissue. Then, we detected the expression pattern of the NENF gene in bovine preadipocyte and myoblast differentiation and found that the expression of NENF mRNA peaks at day 6 during preadipocyte differentiation and peaks at day 4 during myoblast differentiation. Furthermore, we found that the endogenous knockdown of NENF inhibited the differentiation of preadipocytes and promoted the differentiation of myoblasts. These findings not only lay the foundation for the construction of regulatory pathways during fat and muscle differentiation but also provide a theoretical basis for molecular breeding of beef cattle. Abstract Neudesin neurotrophic factor (NENF) is a secreted protein that is essential in multiple biological processes, including neural functions, adipogenesis, and tumorigenesis. In our previous study, NENF was significantly inhibited in the bovine adipocytes-myoblasts co-culture system. However, studies on NENF regulation of bovine muscle development and involvement in the cross-talk between adipose tissue and skeletal muscle have not been reported. Hence, the aim of this study was to clarify the functional roles of NENF in bovine preadipocytes and myoblasts. Real-time quantitative PCR (RT-qPCR) was performed to examine the spatial expression patterns of NENF in different tissues, and the results showed that NENF was highly expressed in the muscle of four-day-old and 24-month-old Qinchuan cattle. Compared with four-day-old Qinchuan cattle, the expression level of NENF was significantly up-regulated in 24-month-old bovine adipose tissue. To explore the roles of NENF in bovine myoblast and preadipocyte differentiation, small interfering RNA (siRNA) targeting the NENF gene were transfected into bovine preadipocytes and myoblasts to knock down the expression of the NENF gene. The results showed that the knockdown of NENF in differentiating adipocytes attenuated lipid accumulation, decreased the mRNA expression of adipogenic key marker genes PPARγ, CEBPα, CEBPβ, FASN, and SCD1, and decreased the protein expression of PPARγ, CEBPα, and FASN. The formation of myotubes was significantly accelerated, and the mRNA expression levels of myogenic marker genes MYOD1, MYF5, MYF6, MEF2A, MEF2C, and CKM, and the protein expression levels of MYOD1, MYF6, MEF2A, and CKM were up-regulated in myoblasts where NENF was knocked down. In short, the knockdown of NENF inhibited preadipocyte differentiation and promoted myoblast myogenesis.