Characterization Of Lymphocytes During Humoral Immune Response In Nile Tilapia

The immunity is made up of humoral immunity and cellular immunity.In vertebrates,with both innate immunity and adaptive immunity,the humoral immunity is mainly mediated via B cells.Moveover,the humoral immunological memory were existed in mammals and birds.However,the specific mechanism and the characteristic of immunological memory during teleost humoral immunity remains unclear while their humoral immune system were complete and in result the understanding of adaptive immunity evolution and the progress of research focus on fish vaccine were slowed.Thus,the characterization of lymphocytes as well as the characteristic of immunological memory during primary and secondary humoral immunities in typical teleost Nile tilapia were explored as well as the expression patterns and potential roles of spleen B cells were analyzed.The main results are as follows:The primary and secondary humoral immunities of Nile tilapia were induced with recombinant proteins Sip and/or α-enolase and then the serum Ig M were analyzed.The expression levels of total Ig M were rapidly increased at 12-15 d post primary immunization,reached peak at 30-45 d,then declined and calmed at 90 d ultimately.Besides,an affinity maturation was detected since 60 d post injection.However,the rapidly and unenhanced responses with the longer-term antigen-specific antibodies post secondary immunization was observed.Moreover,no obvious difference was detected between two secondary humoral immune responses induced by distinct immunogens.These results indicate that there was no antigen-specific immunological memory in Nile tilapia humoral immunity,instead of leading a more rapidly secondary humoral immunity.Meanwhile,the expression profiles of Nile tilapia head kidney lymphocytes RNASeq during primary and secondary humoral immunities were analysed.The main response progress during primary humoral immunity were biosynthesis and cell cycles while a few immune related players were also identified,such as the key genes of lymphocytes development and differentiation.However,the main immune related responders against different antigens during secondary humoral immunity were both involved in T cells development process.The results of trend analysis and the evaluation of lymphocyte subpopulation marker genes both indicated that there was no clear and specific responder among the whole process.That is to say,the Nile tilapia head kidney is not the key player.Besides,in order to discover the crucial organ of Nile tilapia humoral immunity,the expression profiles of a series of lymphocyte subpopulation marker genes were evaluated via q RT-PCR.The results showed that the low-density lymphocytes were detected from each organs.HSCs,CLPs,and B cells were mainly located in head kidney,spleen and peripheral blood,T cells and NCC were concentrated distributed in specific lymphocyte layers of each organ while Mo/Mφ mainly located in kidney.The HSCs and T/B cells in posterior kidney and spleen were sharply induced during the primary humoral immunity while the T/B cells in head kidney and spleen were promoted during the secondary immunity with a widespread apoptosis.These results suggest the spleen might been the pivotal place for B cells development and differentiation.Furthermore,the expression profiles of Nile tilapia spleen sc RNA-Seq during humoral immunity were evaluated.Compared to the head kidney sc RNA-Seq,more T/B cells were identified in spleen while the HSCs and GC marker genes were lost.Besides,the proportions of B cells were reduced post immunization but the BCR levels were promoted.However,no distinct Bm was observed.In summary,the spleen B cells suffered distinct differentiation along with the primary and secondary humoral immunities and in result the BCR sequences were changed with affinity maturation.