Study On The Relationship Between Polymorphism And Expression Of Key Pro-Inflammatory Factors (IL-1β,IL-6,TNF-α) And Immune Traits In Tibetan Pigs

Tibetan pigs live in high altitude and low oxygen environment for a long time and have excellent characteristics of immune function and disease resistance.Interleukin-1β（IL-1β）,Interleukin-6（IL-6）,and tumor necrosis factor-α（TNF-α）are key pro-inflammatory factors.It plays an important role in the innate immunity of animals.Studies have shown that the infection of endotoxin（LPS）plays a major role in the expression of these key pro-inflammatory factors.In order to explore the characteristics of immune function and strong disease resistance of Tibetan pigs and the related molecular mechanisms,this study compared the blood physiological indexes of Tibetan pigs with stable genetic characteristics and the pigs living in Nyingchi for a long time,combined with the differential expression of key pro-inflammatory factors（IL-1β,IL-6,TNF-α）in immune-related tissues and analyzed the SNP sites.The differential expression of key pro-inflammatory factors（IL-1β,IL-6,TNF-α）in peripheral blood lymphocytes of Tibetan pigs stimulated by endotoxin was discussed.The results are as follows:（1）The inflammatory response ability of Tibetan pigs was significantly stronger than that of approximately gram pigs living in the plateau.The index of white blood cell number（WBC）,red blood cell number（RBC）,hemoglobin number（HGB）,red blood cell volume（HCT）,mean red blood cell volume（MCV）,platelet number（PLT）and procyclidine（PCT）,which are related to immune function in Tibetan pig blood,are relatively high in Tibetan pig.The white blood cell count（WBC）,red blood cell count（RBC）,hemoglobin number（HGB）,erythrocyte specific volume（HCT）and platelet number（PLT）of Tibetan pigs were significantly higher than those of approximately gram pigs（P<0.01）,and the mean red blood cell volume（MCV）and procyclidine（PCT）of Tibetan pigs were significantly higher than those of approximately gram pigs（P<0.05）.（2）IL-1β,IL-6 and TNF-αgenes were highly expressed in Tibetan pig immune tissues.In addition,IL-1β,IL-6 and TNF-αgenes were highly expressed in blood,spleen and liver tissues of Tibetan pigs,and the m RNA expression level of Tibetan pigs was significantly higher than that of approximately gram pigs（P<0.01）.In mandibular lymph nodes,the m RNA expressions of IL-1βand TNF-αgenes in Tibetan pigs were significantly higher than those in Yoog pigs（P<0.05）,and the m RNA expressions of IL-6 genes were extremely significantly higher than those in Yoog pigs（P<0.01）.In the same environment,the immune level of Tibetan pigs is higher than that of gram pigs,and the control of inflammatory response of Tibetan pigs is higher than gram pigs.This difference may be one of the reasons why Tibetan pigs are more resistant to disease.（3）There were five mutation sites in the 3’flanking region of IL-1βgene,which were G⁶⁹⁰T,C¹³⁸³G,C¹⁴⁵⁴T,C¹⁴⁸⁰T and A¹⁴⁹⁷G,and there were significant differences in these sites between Tibetan pigs and approximately gram pigs（P<0.05）.There was one mutation site in the 5’flanker and 3’flanker regions of IL-6 gene:A-72G and C²⁶⁵T were significantly different in Tibetan pigs and Tibetan pigs（P<0.01）,and it was found that transcription factor prediction resulted in the increase of ETS1,the key transcription factor of autoimmunity,and the deletion of RUNX1 and NR2F1,which were easy to cause diseases such as cancer.There was a mutation site in the 5’flanking region of TNF-αgene,G-1017A,but there was no significant difference between Tibetan pigs and abstract II pigs（P>0.05）.（4）Low concentrations（1μg/m L and 10μg/m L）of endotoxin stimulated lymphocytes gradually increased over time,and 10μg/m L of endotoxin stimulated lymphocytes more strongly than 1μg/m L of endotoxin,while high concentrations（100μg/m L）of endotoxin stimulated lymphocytes initially,lymphocytes proliferate.After 48h,apoptosis occurred,and when 1μg/m L concentration of endotoxin stimulated the peripheral blood lymphocytes of Tibetan pigs,the m RNA expression of IL-1βgene increased on the whole,while the m RNA expression of IL-6 and TNF-αgene decreased to a certain extent.After endotoxin stimulation at 10μg/m L,the m RNA expression level of IL-1βgene was increased overall compared with that without endotoxin stimulation,but there was no significant difference compared with that after endotoxin stimulation at 1μg/m L.The m RNA expression levels of IL-6 and TNF-αgenes were increased during 24-36h and reached the peak at36h.Significantly or extremely significantly higher than other time points.The m RNA expressions of IL-1β,IL-6 and TNF-αgenes were the highest at 24h after stimulation with endotoxin concentration of 100μg/m L,and all of them decreased gradually with time.In conclusion,our study indicated that pro-inflammatory factors IL-1β,IL-6 and TNF-αplay an important role in immune function of Tibetan pigs,combined with the differential expression of IL-1β,IL-6 and TNF-αbetween the two pig species,and there are different SNP sites.It is speculated that IL-1β,IL-6and TNF-αmay be the important cytokines causing the stress resistance and disease resistance of Tibetan pigs.To further provide a theoretical basis and scientific support for Tibetan pigs to have stronger resistance.