Abstract: The aim of this study was to establish an in vitro study model of subcutaneous and intramuscular preadipocytes in yak, and to detect the expression differences of key genes in the differentiation process of preadipocytes in the two parts, so as to provide experimental materials and theoretical basis for the study of the molecular mechanism of fat deposition in different parts of yaks. The subcutaneous adipose tissue and longissimus dorsi muscle tissue of 5 healthy male Maiawa yaks aged 18 to 22 months were collected, and the subcutaneous and intramuscular preadipocytes were separated by collagenase digestion. The cells were then divided into intramuscular group and subcutaneous group according to the cell sources. Immunofluorescence identification, growth curve mapping, Oil red O identification of adipocytes and real-time fluorescence quantitative technology were used to detect the expression of key differentiation genes PPARγ, C/EBPα, FASN and HSL. The results showed that yak subcutaneous and intramuscular adipocytes on the first day of cultivation were mostly circular, with the increase of incubation time, formed a spindle, the normal "S" growth curve model for CCK-8 detection, and growth rate of subcutaneous preadipocytes on day 4 was significantly higher than that of intramuscular preadipocytes. After identification by immunofluorescence, the expression of PREF-1 was positive, and they were identified as preadipocytes. After differentiation, mature adipocytes with large and round fat droplets were formed, and red fat droplets were detected by Oil red O staining, and the key genes were expressed. The expression levels of PPARγ, FASN and HSL during subcutaneous and intramuscular preadipocyte differentiation were significantly increased with the increase of differentiation time (P<0.05), the expression of C/EBPα in intramuscular and subcutaneous preadipocytes increased gradually at the prophase of differentiation (P<0.05), and the expression level was decreased at the later stage of differentiation. The expression of FASN and HSL in intramuscular preadipocytes showed significant increase on day 3, while in subcutaneous preadipocytes showed significant increase on day 6. In conclusion, subcutaneous and intramuscular preadipocytes of yak were successfully isolated and differentiated into mature adipocytes. Preliminary detection showed that the expression levels of key adipogenic differentiation genes PPARγ, HSL and FASN all showed an increasing trend during the differentiation period for 9 days, while the expression levels of C/EBPα decreased significantly after 6 days of differentiation. This study provided a reference for further studying the molecular rules of fat deposition in subcutaneous and intramuscular adipocytes in yaks.

Key words: yak, preadipocytes, different parts, differentiation of key genes