| Cryptosporidum parvum is an important zoonotic parasite that infects intestinal epithelialcells of humans and various other mammals，and causes protracted diarrheal in young andimmunodeficient individuals and can lead to death for AIDS patients.The antimicrobial peptides（AMPs） is an critical part of innate immune system and secreted by enterocytes andimmnocytes. Th17, charactered by IL-17A and IL-17F, is another important immune systembesides Th1and Th2, and be important in host defense pathogen infection. Recently, people havefocused on the alteration of antimicrobial peptides and Th17cytokines expressions in the intestineepithelial cell which is infected by bacteria. However, the alteration of AMPs，Th17cytokines andanti-inflammatory cytokines, such as IL-10, response to C. parvum infection is little known.Therefore, the aim of the study was to investigate the intestine immune response in BALB/c miceinfected with C. parvum, including the alteration of antimicrobial peptides, Th17cytokines andIFN-γ-induced chemokines etc.3—4week BALB/c mice were gavaged with1×106oocysts of C. parvum per mouse andkilled on day7and14post infection. Jejunum and colon samples were collected for tissuefragment culture in vitro. Spontaneous IL-12and IL-17proteins in gut fragment cultures wereassayed by ELISA. The results showed that the secretion of IL-12was highly increased in jejunalframents cultures at14days post infection compared to uninfected control（124.02±2.24pg/ml vs14.34±8.60pg/ml, P<0.001）, while in the colon, the secretion of IL-12on day7and14postinfection were both increased significantly （307.93±1.98pg/ml vs24.95±0.24pg/ml, p<0.001;77.32±0.88pg/ml vs24.95±0.24pg/ml, P<0.001）. In jejunal fragment cultures, the expression ofIL-17was increased makedly on day7and14post infection（153.68±37.02pg/ml vs12.52±9.58pg/ml, P<0.001；77.69±10.91pg/ml vs12.52±9.58pg/ml, P<0.001）. However, the production ofIL-17in colon was down-regulated on day7post infection compared to uninfected samples（133.66±32.16pg/ml vs245.86±36.37pg/ml, P<0.001）AMPs are inducible small molecule polypeptides and take important part in the inflammatoryreaction, act as barrier in the host defense pathogenic microorganism. The small intestine is consisted of enterocyte, goblet cell, enteroendocrine cell and paneth cell. Goblet cell secretesmucin and intelectin, while lysozyme, defensin and most intelectin are secreated by paneth cells,which located in the base of crypt. In the study, we collected jejunum and colon samples,embedded in paraffin and stained with H.E and AB-PAS. Through the histological observations,the number of goblet cells and paneth cells were increased significantly response to C. parvuminfection. And the RT-PCR assay showed the expression of mucin1and mucin2were up-regulatedsignificantly in jejunum compared to uninfected samples, while the production of mucin2decreasein colon compared to uninfected samples. The mRNA expression of intelectin1and intelectin2were decreased on day7post infection in jejunum, but elevated production was observed at14days post infection as3-4folds in jejunum. However, little expression of intelectin1andintelectin2were found in colon. We also found the production of lysozyme secreted by panethcells was down-regulated makedly on day7post infection compared to control, while nodifference was observed between the samples infected for14days and control. It wasdemonstrated that AMPs was important for the immune response during C. parvum infection.Th17is another important adaptive immune response system, and IL-17A and IL-17F havebeen demonstrated be critical in defense inflammation and microbe infection. The results ofRT-PCR showed that the mRNA level of IL-17A and IL-17F were increased significantlycompared to uninfected controls, and can be up to be28-fold and46.44-fold respectively at7dayspost infection in jejunum. IFN-γ and the chemokines it induced CXCL-9, CXCL-10and CXCL-11all response to C. parvum actively as reported before. Besides, we also observed the alteration ofIL-10and Tgfb3, indicated that C. parvum infection can induce complicated immune response.In a word, the most severe response to C. parvum infection in mucosal immune systemlocated in small intestine, and important alteration of humoral immune response were alsoobserved in both jejunum and colon. It suggests that AMPs, Th17cytokines and IFN-γ-inducedchemokines may take important parts in mediating host defense C. parvum infections, and theresults may also establish foundation for study the mechanism of intestinal immune response ofmice infected by C. parvum. |
