Regulating Effects Of Allicin On The Immune Function Of Reticuloendotheliosis Virus Infected SPF Chickens

Reticuloendotheliosis virus(REV)is classified as a member of the viruses in the family Retroviridae,specifically gammaretroviruses in the same genus as mammalian C-type retroviruses,causes immunosuppressive,oncogenic and runting-stunting syndrome in multiple avian hosts.Immunosuppression plays an important role in the occurrence and development of the disease,but the underlying molecular mechanism remains unclear.In this study,we inoculated SPF chickens with REV SNV-C5 strain,and the factors that might be involved in the process of immunosuppression were detected.Allicin,the main responsible component of garlic,has a broad spectrum of pharmacological properties such as anti-bacterial and anti-viral effects.There is still a lack of safe and effective vaccines or drugs for prevention and control of REV.In order to provide a new way for the reasonable and effective prevention and control of REV,we aimed to investigate the effects of allicin on REV infected chickens by feeding diets supplenmented with different doses of allicin on experimental chickens.1.Allicin alleviated the growth inhibition and immunosuppression induced by REV infectionThe experimental chickens were fed with a diet supplemented with different doses of allicin(0,150,300,600 mg/kg)and subjected to REV inoculation(100TCID50).Samples of immune organs including thymus,spleen and bursa were collected at 2W,3W,4W,5W post REV infection.The results showed that REV infection caused dysplasia of chickens(P < 0.0001),and atrophy of thymuses(P < 0.05)as well as bursas(P < 0.05)and intumesce of spleen(P <0.05)from 2 weeks post REV infection compared to control chickens,which was effectively alleviated by 300mg/kg allicin(P < 0.0001),indicated that REV-infection caused immune dysfunction,allicin ameliorated the growth inhibition and immunosuppression.2.Allicin attenuated REV-induced inflammatory reactionThe effect of allicin and REV on the release of cutokines were analyzed.Compared to untreated chickens,REV infection promoted the expression of IL-1(P < 0.01),TNF-(P < 0.01),IFN-?(P < 0.01),IL-4(P < 0.01)and IL-10(P < 0.05).Moreover,REV infection induced a significant increase of TLR-3(P < 0.05),TLR-4(P < 0.01)as well as TLR-7(P < 0.05),and dramatically induced the mRNA expression of MDA5(P < 0.05).We also detected REVup-regulated the level of STAT6(P < 0.01)and subsequently promoted the expression of ISGs(P <0.05)and IRF-7(P < 0.01)to regulate the expression of IFN,but allicin altered these changes.Additionally,REV significantly activated the NF-?B p65(P < 0.01)and MAPK pathways(P <0.05),whereas treatment with allicin depressed this phenomenon.And we found that inhition of ERK inhibited the expression of REV genes(P < 0.0001).These results suggested that REV was primarily recognized by TLRs and MDA5,and then STAT,MAPK,IRF and Nrf2 signaling pathways were activated,and finally the nuclear translocation of NF-?B were regulated to exert antiviral responses.Allicin alleviated the changes of signaling pathways mentioned above,which explained the potential anti-REV effect of allicin.3.Allicin reduced the oxidative damage caused by REVThe effect of allicin and REV on the antioxidant system were analyzed.High expression of malondialdehyde(MDA)(P < 0.05),protein carbonyls(P < 0.01),translocation of Nrf2(P <0.05)and decreased activity of antioxidant enzymes(P < 0.05)were observed in REV-administrated chickens,suggested REV infection broke the balance of oxidation and antioxidation,resulting oxidative damage of the body.Allicin treatment effectively improved the antioxidant capacity of REV-infected chicks,decreased the nuclear translocation of Nrf2 and improved the oxidation resistance.4.Effect of allicin and REV infection on adipocytesIn the first study,we found that REV infection significantly lowered abdominal fat contents than control group.In this study,we investigated the effect of REV infection on adipocytes and the effect of allicin.We isolated the pre-adipocyte from 15 days 'embryo and induced pre-adipocytes differentiationed into mature adipocytes with IBMX,rosiglitazone,dexamethasone and insulin.We treated the mature adipocytes with 0 TCID50,5 TCID50,10TCID50,20 TCID50,40 TCID50 REV for 0h,12 h,24h,48 h,72h respectively.The results showed that 20 TCID50,40 TCID50 REV treated adipocytes for 1h promoted the expression of TNF-?,and increased the phosphorylation of p38(P < 0.01),ERK(P < 0.0001),JNK(P <0.0001)and NF-?B(P < 0.0001),indicated that REV triggered the inflammatory reaction in adipocyte.In addition,REV promoted the phosphorylation of AMPK(P < 0.0001),promoted energy metabolism in adipocytes,and decreased the synthesis of TG(P < 0.1)and differentiation(P < 0.0001).5?g/mL,10?g/mL,25?g/mL,50?g/mL allicin were treated REV infected adipocytes for 24 h,48h,72 h,which showed that 25?g/mL,50?g/mL allicin treated adipocytes for 24 h decreased the expression of TNF-?(P < 0.01)and phosphorylation of MAPK(P < 0.01),NF-?B(P < 0.0001)and AMPK(P < 0.0001),suggested that allicinalleviated intracellular inflammatory stress.In summary,REV infection resulted in growth inhibition and immnosupression by induced inflammatory and oxidative damage in some extent.Allicin significantly alleviated the release of inflammatory cytokines induced by REV infection,increased the activity of antioxidant enzymes,anti-inflammatory and antioxidant capacity of experimental chickens,indicating that allicin has immunoregulation effect on REV infection.