Effect Of Short-chain Fatty Acids On Receptor Gene And Cytokine Expression Of Peripheral Blood Lymphocyte Of Goats

SCFA provides not only to their involvement as the major source of nutrients in ruminants, but also serve as regulator of signal transduction. The experiment was conducted to study the effects of the SCFA on Immune regulation of cell of growing goat in vitro by the teconology of cell biology and molecular biology.Detection of SCFA on lymphocyte prolif after 12h and 24h respectively by MTT method. The results showed that the acetic acid, propionic acid and butyric acid significantly inhibited the proliferation of peripheral blood lymphocytes (P<0.01) at final concentration of 1mmol/L,5mmol/L, 10mmol/L L,and showed time dose-dependent; butyric acid's inhibition is the strongest, followed by propionic acid, then the acetic acid is the smallest role. This affection on lymphocyte proliferation was partly eliminated (P<0.05) after ddA reaction.Dealing goat peripheral blood lymphocytes with SCFA at different concentrations, after 12h or 24h, Detected cytokines by real time RT-PCR,the results showed that:①different concentrations of sodium butyrate on IL-2 and IFN-γmRNA expression was inhibited significantly (P<0.01), and showed time dose-dependent, When processed through the ddA, the inhibitory effect of sodium butyrate was significantly eliminated(P<0.05), The different concentrations of sodium acetate and sodium propionate on IL-2 and IFN-γmRNA had no significant effect on expression (P> 0.05);②After 12h in primary culture, The goat peripheral blood lymphocytes were treated with different concentrations of sodium acetate and sodium propionate,and IL-10 mRNA expression had no significant effects (P>0.05); but the expression was significant enhancement (P<0.05)after 24h,and this promotion reduced after treat with ddA.In contrary, different concentrations of sodium butyrate could significantly inhibited the expression of IL-10 mRNA(P<0.01)after 12h or 24h, ddA could abolish this inhibitory effect of sodium butyrate significantly (P<0.01),③after 40 cycles, we did not detect IL-4 mRNA expression by Real-time quantitative PCR.Treated goat peripheral blood lymphocytes address with different concentrations of SCFA,after 12h or 24h, Examined the effect of cytokine medium by ELISA,the result showed that:①Both acetic acid and propionic acid enhanced the expression of IL-2 and IFN-γcytokine, the difference was not significant (P> 0.05),but butyric acid could significantly inhibit the expression of IL-2 and IFN-γcytokine,ddA could reduce the inhibition(P<0.01)。②The effects of acetic acid and propionic acid on expression of cytokines IL-10 were very significantly enhanced (P<0.01), and showed time dose-dependent. butyric acid was significantly inhibit the expression of IL-10 cytokine (P<0.01). This acid inhibition was eliminated after ddA reaction(P< 0.01).③We had not detected expression of IL-4 in the supernatant of cultured cells.We detected the expression of G protein receptor in goat peripheral blood lymphocytes which deal with different concentrations of SCFA by Real-time quantitative PCR.The results showed that: Sodium acetate significantly enhanced the expression of GPR43 (P<0.05),and showed dose -dependent; Sodium Propionate preference activation GPR41 (P<0.05)and dose-dependent; Sodium butyrate significantly enhanced the expression of GPR41 and GPR43 mRNAs (P<0.05)and dose-dependent. These results suggest that different short-chain fatty acids, respectively corresponding to the expression of G protein receptor enhancement.With different concentrations of sodium butyrate treated goat peripheral blood lymphocytes, Detected its affection on expression of NHE1 and GPR43 by Western Blotting after 24h,The results suggested that:①1mmol/L sodium butyrate increased the expression of NHE1, but the difference was not significant compared with the control group (P>0.05),5mmol/L and 10mmol/L sodium butyrate could significantly enhance the NHE1 protein expression (P<0.05), and 5mM/L enhanced the best. However, ddA can eliminate the affection of butyric acid ignificantly(P<0.05); 1mmol/L and 5mmol/L sodium butyrate can increase GPR43 expression, the difference was not significant (P>0.05), but 10mmol/L sodium butyrate can significantly enhance the expression of GPR43 protein (P<0.05); However, the enhancement role of butyric acid had decreased significantly when dealed with ddA (P<0.05).