| Cadmium(Cd)poisoning,the human and animal cardiovascular,liver,kidney,reproductive system and immune organs damage.Agaricus blazei polysaccharide(ABP)is a polysaccharide extracted from Agaricus blazei with immunoregulatory,antioxidant and antiinflammatory activities.Based on the establishment of chicken cadmium poisoning model,the effects of ABP on MDA5 signaling pathway and antioxidant function in peripheral blood lymphocytes and spleen of cadmium poisoned chicken were studied in order to clarify the relationship between ABP intervention mechanism of peripheral immune injury in cadmium poisoning chickens.A total of 80 Hyline white chickens(7 days old)were randomly assigned to 4 groups(n=20/group): Control group,Cadmium poisoning model group(Cd),Agaricus blazei polysaccharide group(ABP)and treatment group(Cd+ABP).Control group,fed the normal full price feed,each chicken was supplemented with 0.2 m L saline every day.Cd group,each chicken was administered Cd at a dose of 140 mg/kg Cd Cl2 in its feed every day for 2 months.ABP group,each chicken was supplemented with 0.2 mL ABP(30 mg/mL)in its feed every day for 2 months.Cd+ABP group,each chicken was administered Cd in the same way as those in Cd group,followed by supplementation with ABP in its feed every day for 2 months,samples were taken at 20 th,40 th and 60 th,respectively.First,five chickens were randomly selected for heart blood collection,collected the blood for the separation of lymphocytes and the collection of serum,followed by rapidly separated the spleen,take part of the spleen fixed in 4% paraformaldehyde solution.Separated lymphocytes and the remaining spleen are divided into four parts,the content of cadmium in chicken serum and spleen was detected by inductively coupled plasma mass spectrometry.The extraction of total RNA from lymphocytes and spleen were performed,reverse transcription into c DNA,by real-time quantitative PCR detection of the expression of MDA5 and signal transduction pathways of adaptor protein IPS-1,transcription factors(IRF3,NF-?B)in lymphocytes and spleens.Followed by part of the lymphocytes and spleen protein extraction and the expression of MDA5 in these two proteins was detected by Western Blot.Then,the remaining lymphocytes were cultured in 1640 complete culture medium for 24 h.The supernatant was collected and the remaining spleen was homogenized with physiological saline.The levels of cytokines IL-1?,IL-6,TNF-? and IFN-? were measured by ELISA.Antioxidant markers(SOD,MDA and GSH-Px)were measured with lymphocyte culture supernatant and spleen homogenate.In addition,the chicken spleen was subjected to histopathological examination.The results showed that:(1)the cadmium content in Cd+ABP group was extremely significant redcued(P<0.01).(2)The mRNA levels of MDA5 and the expression of MDA5 protein were increased after cadmium poisoning both in peripheral blood lymphocytes and spleen,with the increase of cadmium poisoning time,the MDA5 expression also increased,while the expression of ABP was significantly reduced;the expression of MDA5 downstream signaling molecules IPS-1,IRF3,NF-?B also increased.(3)The levels of IL-1?,IL-6,TNF-? and IFN-? in peripheral blood lymphocytes and spleen of Cd group increased,and were significantly higher than those in Control group and ABP group(P<0.01),the contents of these cytokines were significantly decreased(P<0.01)after oral administration of ABP.(4)The MDA content of lymphocyte supernatant and spleen were decreased significantly(P<0.01),SOD and GSH-Px activity were significantly increased(P<0.01)in Cd+ABP group.(5)The pathological changes of spleen after cadmium poisoning showed a large number of lymphocytes decreased,red pulp,white pulp boundary are unclear,lymph nodes in the number of lymphocytes decreased,arranged in scattered,and the structure has improved significantly in Cd+ABP group,the numbers of lymphocytes were increase,red pulp,white pulp junction is also clear.The above results showed that the ABP can reduce the accumulation of cadmium in peripheral blood and spleen,reduce the expression of MDA5 and its downstream signal molecules in peripheral blood lymphocytes and spleens after cadmium poisoning,the activity of antioxidant enzymes(SOD,GSH-Px)were significantly increased and the content of MDA was decreased.The ABP had a protective effect on the damage of chicken peripheral blood lymphocytes and spleen of cadmium poisoned.This study provides a scientific basis for further study of ABP intervention cadmium poisoning leading to the mechanism of organism injury. |
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