| Acting as the fist line of specific immune defense, the mucosal immune system participates in protecting organisms from a variety of pathogens. Secretory immunologlobulins (slgs) are the dominating defensive factors of mucosal immune system. Polymeric immunoglobulin receptor (plgR) plays an indispensable role in mucosal immunity, mainly medicating the transportation of polymeric immunoglobulins (pig) through the epithelial cells to protect the organisms. The efficient of secretion of plgR is the necessary condition for pig playing the mucosal defense function, and it plays an important role in mucosal immunity. As the research for teleost plgR has been increasingly active, the immune response of plgR and slgs attract much concern.This study researched the transcription levels of pIgR and IgM in different tissues after intraperitoneal injected and immersed with inactivated Vibrio anguillarum, investigated the protein levels of plgR and IgM in skin nucus, gill mucus, intestinal mucus and bile, moreover, detected the plgR-IgM complex in three kinds of mucus and bile, finally, demonstrated the location of pIand IgM in the mucosa-associated tissues, it will lay the foundation for the further research of teleost mucosal immune mechanism. The following are the details:Following intraperitoneal injection and immersion immunization with inactivated Vibrio anguillarum, the transcription levels of pIgR and IgM was detected by quantitative real-time PCR. Post immunization, the transcriptional levels of IgM gene in tissues of flounder were presented a similar tendency to pIgR gene that increased firstly, then decreased, however, IgM gene mRNA levels increased more slowly and the peaks appeared more lately than pIgR gene. In skin, gill and hindgut, the peaks of immersed group pIgR were higher than injected group and they were lower than injected group in other tissues; in skin and gill, the peaks of immersed group IgM were higher than injected group and they were lower than injected group in other tissues.The protein levels of pIgR and IgM were detected by ELISA following intraperitoneal injection and immersion of inactivated V. anguillarum. The results of ELISA showed that protein levels of pIand IgM indicated a similar variation trends that increased firstly and then decreased to the control levels over a period of time in external secretions including skin mucus, gill mucus, intestinal mucus and bile of flounder, the protein expression of pIgR were increased more quickly than IgM and the peaks appeared more early than IgM in both of immunized groups. In skin mucus and intestinal mucus, the peaks of immersed group pIgR were higher than injected group and they were lower than injected group in gill mucus and bile; in skin mucus and gill mucus, the peaks of immersed group IgM were higher than injected group, and they were lower than injected group in intestinal mucus and bile.Under non-reducing conditions, the anti-serum Ig mAb 2D8 and anti-mucus Ig mAb 1A2 reacted with serum, gill mucus, skin mucus, intestinal mucus and bile at molecular weight of-800kDa. The polyclonal antibodies against rpIgR reacted with gill mucus, skin mucus, intestinal mucus and bile of flounder at molecular weight of-800kDa, but not reacted with serum of flounder.Immunohistochemistry located the pIgR and IgM in mainly mucosa-associated lymphoid tissues of flounder using mouse-anti-pIgR and mouse-anti-IgM antibodies, and the immuno-staining of the two antibodies were substantially accordant. In mucosa-associated tissues of flounder, the obvious positive signals appeared in experimental group, and the control group was negative. The pIgR positive signals mainly located in epithelial cells, goblet cells and surface of skin, gill and hindgut, the base of epithelial cells in gill and hindgut and epithelial cells of hepatic duct. The staining of IgM primarily distributed in epithelial cells and goblet cells in skin, gill and hindgut, subcutaneous lamina propria of hindgut and epithelial cells of hepatic duct. |
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