Effects Of Long-chain Fatty Acids On Proliferation And Differentiation Of Sheep Preadipocytes

With progressing of society and improving of living standards,increasingly higher dietary structures were required.The contemporary husbandry developed rapidly,but it still hard for the meat production and its quality meet the market.The distribution of adipose tissue and the deposition of triglycerides in the bodies are important factors affecting meat flavor and carcass quality.The body fat deposition is affected by the proliferation and differentiation of preadipocytes,and the proliferation and differentiation of preadipocytes are regulated by different factors.In this paper type I collagenase were used to digest and separate the preadiocytes from tail and subcutaneously of small tail han sheep,primary cultured,subcultured,and induced,treated with different long-chain fatty acids,oil red O stained,and then observed,real-time fluorescence quantitative PCR were used to explore the effects of different long-chain fatty acids on sheep preadipocytes proliferation.The results showed as follows:1.Small tail han sheep tail preadipocytes began to attached in 4h after isolating,and abdominal subcutaneous preadipocytes attached after 6h;the cells showed triangular and irregular in shape.After subcultured,the cell morphology was long-fusiform,and the growth curve showed"S"-like curve.The doubling time of the tail preadipocyte?22.04 h?was significantly shorter than of the subcutaneous?29.56h?.The maximum proliferation density of the tail preadipocyte?58.9×10⁴cells/mL?was significantly larger than that of subcutaneous?29.7×10⁴cells/mL?;the specific growth rates of tail and subcutaneous preadipocytes were 1.25 d^-11 and 1.05 d^-1,with no significant difference?P>0.05?;after 5 days of differentiating with adipogenic induction fluid?DMI?,lipid droplets were detected in the cytoplasm and oil red O stained red.2.The primary cultured tail preadipocytes were treated with different concentration of ALA,LA and PA?concentrations were respectively 100?M,150?M,200?M,250?M,300?M,and 350?M?and compared with the blank control group and the negative control group.The results showed that,treated for 1 day with ALA,the low concentration groups?100?M and 150?M?significantly promoted cell proliferation?P<0.01?.While the high concentration groups?250?M,300?M,and350?M?caused cell death;traeted for 3 days,all groups inhibited cell proliferation,low concentrations?100?M and 150?M?of LA significantly promoted cell proliferation?P<0.05?,high concentrations?200-350?M?inhibited it.150?M PA significantly promoted the proliferation of the preadipocytes?P<0.05?.While it inhibited cell proliferation at 200³50?M concentration.3.In order to study the effects of ALA,LA and PA at differentiation of sheep preadipocytes,the effects of different concentrations?100?M and 150?M?of ALA,LA and PA on lipid droplet formation during differentiation of sheep preadipocytes were studied.Lipid formation was observed under the microscope after staining with red O and real-time fluorescence PCR was used to detect the differentiation-related genes.The results showed that treated with the concentrations of?100?M and 150?M?ALA and LA,the expression of PPAR?,FABP4,LPL,HSL,SREBP-1c and FASN mRNA were up-regulated,LA affected significantly greater than that of same concentration ALA.In addition,150?M ALA up-regulated the expression of PPAR?,FABP4 and LPL more extremely than 100?M?P<0.05?;The expression of PPAR?,FABP4,LPL,HSL,SREBP-1c and FASN were down-regulated by PA?P<0.05?.In summary,in this study,sheep preadipocytes were successfully isolated and cultured in vitro.It showed that low concentrations?100?M and 150?M?of LA and PA promoted preadipocytes proliferation.The expression of preadipocytes differentiation ralated genes were up-regulated by low concentrations?100?M and150?M?of ALA and LA and down-ragulated by PA.