Molecular Mechanism Of Astragaloside IV And Andrographolide Promoting Phagocytic And Killing Activities Of Chicken Heterophils Against E.coil O78

Avian colibacillosis is a kind of acute infectious disease induced by Escherichia Coli, which can cause high incidence and mortality rates, decrease the production performance, therefore hinder the development of poultry industry. In the previous study, we have designed an effective formula of traditional Chinese medicine against avian colibacillosis, which consists of Astragalus, Andrographis and Cyperus, named as "Xiang-Qi-Tang"(XQT). Astragaloside IV and Andrographolide are the important ingredients of XQT and play great role in this formula. XQT can significantly resist the infection of E.coli O78, and dramatically alleviate the autopsy changes. Astragaloside IV is an important ingredient of Astragalus, which can improve the function of lung and heart, protect liver, induce interferon and regulate the immunity. Andrographolide is the most effective ingredient of Andrographis, which has many functions, such as clearing Heat and releasing Toxins, diminishing inflammation and alleviating pain. And it also has special efficacy to resist infection caused by bacteria and virus. Many researches showed that at the primary period of infection, these two monomers can also improve the cellular immunity and increase the phagocytosis and cytotoxicity of neutrophils. However, the mechanism of how Astragaloside IV and Andrographolide increase the phagocytosis of neutrophils remains unclear. Heterophils are the most important leukocytes of chicken, with the same biological functions as mammalian neutrophils, which the most important role is to protect chicken from invading by pathogens. At present, the characteristic of heterophils in chicken is little known. Therefore, it is very necessary and important to investigate the phagocytic mechanism of heterophil.The phagocytic and killing process of E. coli O78by heterophils was imitated in vitro. Mix E.coli O78and heterophils at a ratio of10:1, observe the phagocytosis under laser confocal microscopy, and count the colony number of bacteria phagocytized by heterophils on Luria-Bertani plates. We found that Astragaloside IV (5mg/L) and Andrographolide (1mg/L) can significantly improve the phagocytosis and killing ability of heterophils against E.coli O78. Through RT-PCR and ELISA, we also found that Astragaloside IV and Andrographolide can significantly improve the mRNA and protein expression level of TLR4, IL-6, IL-1β and IFN-γ. Heterophils can kill the bacteria through oxygen-dependent and-independent pathways. We found that Astragaloside IV and Andrographolide can significantly improve the mRNA expression level of CATH-2and AvBD-2, and increase the level of respiratory burst. However, Astragaloside IV and Andrographolide can inhibit the production of NO.In summary, we found that Astragaloside IV and Andrographolide can improve the phagocytosis and killing ability of heterophils against E.coli O78through oxygen-dependent and-independent pathway. These results will provide accurate data and reliable theory for the further study of traditional Chinese herbal monomers on improving the avian immunity against infection.