Study On Expression Pattern Of Acute Phase Protein Genes In Zebrafish Embryo/Larva And Function Of Zebrafish Serum Amyloid A (SAA)

Acute phase response (APR) is a group of rapid physiological responsesoccurring soon after the onset of infection or injury, which contributes to preventionof ongoing tissue damage, trapping and destruction of microorganisms and activationof the repair processes necessary to restore the host’s normal function. Allanimals--from fish to mammals--have demonstrable APR. APR is characterized byincrease in acute phase proteins (APPs) in serum, fever, increased vascularpermeability, and metabolic and pathologic changes. APPs are commonly consideredas plasma proteins primarily synthesized by hepatocytes as part of the acute phaseresponse. Induction of APPs, best characterized in mammals, is mediated by Toll-likereceptors (TLRs), which, upon stimulation, result in the production of thepro-inflammatory cytokines such as IL-1β and TNF-α, that are important inducers ofAPPs. APPs can be classified as positive APPs, if their concentrations in serumincrease, or negative APPs, if their concentrations decrease. In humans, C-ReactiveProtein (CRP), serum amyloid A (SAA), serum amyloid P (SAP), hepatoglobin (HP),fibrinogen, complement component3(C3) and factor B have been shown to bepostive APPs, whereas albumin and transferrin to be negative APPs. Research on APRin several fish species (Bayne and Gerwich,2001; Magnadottir et al.,2011; Russell etal.,2006) has revealed that fish express many homologs to human APPs. CRP, SAA,HP, leukocyte cell-derived chemotaxin2(LECT2) and hepcidin (HAMP) have alsobeen demonstrated as APPs in zebrafish Danio rerio. Acute phase response (APR) indeveloping embryos/larvae has not been systematically studied in zebrafish or anyother fishes, information regarding the expression and role of APPs in developingembryos/larvae is rather limited in teleost.In this study we investigated the APR in zebrafish embryos/larvae challenged with LPS via examining the expression of APP genes encoding CRP, SAA, LECT2,HAMP and HP and APP inducer genes encoding IL-1β and TNF-α. All genes were allup-regulated in embryos/larvae after LPS injection according to qRT-RCR. The resultof WISH demonstrated that both APP and APP-inducing genes displayed maternallylocalized ubiquitously during early normal developmental stages, and then theyexhibited different tissue-specific expression patterns in later developmental stages.LPS challenge resulted in the expression of SAA in the new yolk sac and intestine, theexpression of HAMP in the new site yolk sac, visible up-regulation of LECT2gene inthe yolk sac, and the expression of IL-1β gene in the yolk sac as well was in thedispersed neutrophils of caudal vein, in addition to the tissues they were expressed inthe normal larvae. Collectively, these data suggest that zebrafish embryos are capableof acutely responding to pathogenic challenge, providing protection for themselves.Serum amyloid A (SAA) is one of the most important APP in zebrafish. Theexpression of SAA mRNA was enhanced considerably in zebrafish embryoschallenged with LPS. In this study the recombinant of SAA protein was expressed inE. coli, and its function was analysed in vitro. We demonstrate that recombinant SAAwas not only able to bind to both Gram-positive and negative bacteria such as S.aureus and E. coli but also to LPS and LTA. It was also able to promote thephagocytosis of E. coli and S. aureus by macrophages. These data indicate that SAAcan function as an opsonin in zebrafish.