| High energy diet can improve the performance of animal fattening and production,and promote the fat deposition of subcutaneous and visceral in different degrees.Excessive fat deposition not only can increase the incidence of lipid metabolism diseases,but al so affect the carcass quality of animals,affect the utilization rate of diet,and cause the waste of energy in the diet to some extent.The adipocytes of subcutaneous and visceral are differentiated from the former adipocytes.Because of their different growth microenvironment and types,the former adipocytes in different parts have different differentiation and lipid accumulation.In this study,primary pre-adipocytes from subcutaneous and visceral tissues of mice were used as experimental models to study the effects of different energy levels on autophagy,differentiation and lipid metabolism of preadipocytes,The effects of autophagy activator rapamycin and inhibitor 3-methyladenine(3-MA)on lipid accumulation of subcutaneous and visceral pre-adipocytes in mice were also studied.The results are as follows:Experiment 1: extraction and culture of primary pre-adipocytes from subcutaneous and visceral organs of mice.The results showed that both subcutaneous and visceral pre-adipocytes showed fibrous or irregular shape,and there was no significant difference between them in appearance,and the cell composition after subculture was more uniform,and the cell morphology was not significantly different from that of primary culture.The growth curves of subcutaneous and visceral pre-adipocytes were typical "S" type,and the proliferation ability of subcutaneous pre-adipocytes was stronger than that of visceral pre-adipocytes from the logarithmic phase,the results showed that the proliferation ability of subcutaneous pre-adipocytes was higher than that of visceral pre-adipocytes in the later stage of growth.Experiment 2: effects of different energy levels on autophagy and adipogenic differentiation of subcutaneous and visceral pre-adipocytes in mice.Adipogenic differentiation of subcutaneous and visceral pre-adipocytes was induced by high glucose(4500 mg/L glucose)and low glucose(1000 mg/L glucose),respectively,and the differences in autophagy,differentiation and lipid metabolism were measured.The results showed that the accumulation of triglyceride in subcutaneous pre-adipocytes was more than that in visceral pre-adipocytes in both high glucose group and low glucose group,and the total triglyceride content in high glucose group was lower than that in low glucose group(P < 0.05).Compared with the high glucose group,the low glucose group promoted the expression of LC-3 protein in subcutaneous and visceral pre-adipocytes during differentiation,decreased the protein expression of p62,SREBP-1c and PPAR-γ,and had no significant effect on FAS and HSL protein,but showed low expression of HSL.The expression of SREBP-1c protein in subcutaneous pre-adipocytes induced by high glucose group was higher than that in visceral pre-adipocytes(P < 0.05).The expression of HSL protein in subcutaneous pre-adipocytes was lower than that in visceral pre-adipocytes(P <0.05).In low glucose group,the expression of p62 and SREBP-1c protein in subcutaneous pre-adipocytes was higher than that in visceral pre-adipocytes(P < 0.05),while the expression of LC-3 and HSL was lower than that in visceral pre-adipocytes(P < 0.05).The effect of energy level on LC-3,p62 and FAS proteins of visceral pre-adipocytes was greater than that of subcutaneous pre-adipocytes,but the effects of SREBP-1c,PPAR-γand HSL proteins on visceral pre-adipocytes were lower than those of subcutaneous pre-adipocytes.Experiment 3: to explore the intervention effect of autophagy on adipogenic differentiation of subcutaneous and visceral pre-adipocytes induced by different energy levels.In experiment 2,pre-adipocytes were induced by rapamycin in high glucose group and 3-MA in low glucose group.The results showed that rapamycin group increased the triglyceride content of visceral pre-adipocytes(P < 0.05).3-MA group decreased the lipid content of subcutaneous pre-adipocytes(P < 0.05).Rapamycin group increased the expression of LC-3 protein in subcutaneous and visceral pre-adipocytes(P < 0.05),while3-MA group decreased the expression of LC-3 protein in subcutaneous and visceral pre-adipocytes during induction(P < 0.05).Both rapamycin group and 3-MA group promoted the expression of PPAR-γ in subcutaneous pre-adipocytes during induction,and3-MA promoted the expression of SREBP-1c and PPAR-γ in visceral pre-adipocytes during induction(P < 0.05).Rapamycin group promoted the expression of FAS and HSL protein in subcutaneous pre-adipocytes during induction(P < 0.05).3-MA decreased the expression of FAS protein in subcutaneous and visceral pre-adipocytes,but promoted the direct HSL expression in visceral pre-adipocytes during induction(P < 0.05).In conclusion,the proliferation and lipid accumulation of subcutaneous pre-adipocytes were stronger than those of visceral pre-adipocytes;the differentiation ability of subcutaneous pre-adipocytes was stronger than that of visceral pre-adipocytes,and the degree of autophagy and lipolysis was lower than that of visceral pre-adipocytes.Compared with high energy,low energy level increased the degree of autophagy of pre-adipocytes,but accompanied by the decrease of differentiation and lipolysis.The effect of energy level on autophagy and lipid synthesis of visceral pre-adipocytes was greater than that of subcutaneous pre-adipocytes,but the effect on differentiation and lipolysis of subcutaneous pre-adipocytes was greater.Rapamycin increased the lipid accumulation of visceral pre-adipocytes by promoting the differentiation of visceral pre-adipocytes and reducing lipolysis;3-m A reduced lipid accumulation of subcutaneous pre-adipocytes by inhibiting differentiation and lipid synthesis of subcutaneous pre-adipocytes.Therefore,autophagy plays different roles in adipogenic differentiation of subcutaneous and visceral pre-adipocytes,and the differentiation degree and lipid metabolism state of subcutaneous and visceral pre-adipocytes are also different at the same time;The energy environment can affect the level of autophagy,which may be the way of different fat deposition in different parts of livestock and poultry under the same energy environment. |
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