High-Resolution, 3D Imaging of the Zebrafish Gill-Associated Lymphoid Tissue (GIALT) Reveals a Novel Lymphoid Structure, the Amphibranchial Lymphoid Tissue

Abstract

The zebrafish is extensively used as an animal model for human and fish diseases. However, our understanding of the structural organization of its immune system remains incomplete, especially the mucosa-associated lymphoid tissues (MALTs). Teleost MALTs are commonly perceived as diffuse and scattered populations of immune cells throughout the mucosa. Yet, structured MALTs have been recently discovered in Atlantic salmon (Salmo salar L.), including the interbranchial lymphoid tissue (ILT) in the gills. The existence of the ILT was only recently identified in zebrafish and other fish species, highlighting the need for in-depth characterizations of the gill-associated lymphoid tissue (GIALT) in teleosts. Here, using 3-D high-resolution microscopy, we analyze the GIALT of adult zebrafish with an immuno-histology approach that reveals the organization of lymphoid tissues via the labeling of T/NK cells with an antibody directed to a highly conserved epitope on the kinase ZAP70. We show that the GIALT in zebrafish is distributed over at least five distinct sub-regions, an organization found in all pairs of gill arches. The GIALT is diffuse in the pharyngeal part of the gill arch, the interbranchial septum and the filaments/lamellae, and structured in two sub-regions: the ILT, and a newly discovered lymphoid structure located along each side of the gill arch, which we named the Amphibranchial Lymphoid Tissue (ALT). Based on RAG2 expression, neither the ILT nor the ALT constitute additional thymi. The ALT shares several features with the ILT such as presence of abundant lymphoid cells and myeloid cells embedded in a network of reticulated epithelial cells. Further, the ILT and the ALT are also a site for T/NK cell proliferation. Both ILT and ALT show structural changes after infection with Spring Viraemia of Carp Virus (SVCV). Together, these data suggest that ALT and ILT play an active role in immune responses. Comparative studies show that whereas the ILT seems absent in most neoteleosts (“Percomorphs”), the ALT is widely present in cyprinids, salmonids and neoteleosts, suggesting that it constitutes a conserved tissue involved in the protection of teleosts via the gills.

Antigen-Presenting Cells and T Cells Interact in a Specific Area of the Intestinal Mucosa Defined by the Ccl25-Ccr9 Axis in Medaka

“…In this regard, even though the defined area in the medaka lamina propria where APCs-T cells interaction occur does not appear to be structurally delimited, it shares ILT function in relation to T cells concentration. Recent finding corroborating the existence of Gill-Associated Lymphoid Tissue (GIALT) in zebrafish (46) point to a more complex function of this structure compared to what we observed in the intestine of medaka. The GIALT suffer pronounced structural changes after infection, including a rapid reduction in the number of T/NK cells, followed by a marked repopulation after several days.…”

“…Although there is no report pointing to the existence of an organized mucosal immune response in the gastrointestinal tract of teleost fish, the existence of organized mucosal associated lymphoid tissue in the gills of several fish species, known as interbranchial lymphoid tissue (ILT), has been reported (14,15,46). It is quite intriguing that being both gills and intestine firstline organs for antigens encounter, thus highly immunogenic, in teleost fish organized mucosal lymphoid tissue only evolved in gills.…”

Immunosuppression and apoptosis activation mediated by p53-Bcl2/Bax signaling pathway -The potential mechanism of goldfish (Carassius auratus Linnaeus) gill disease caused by Myxobolus ampullicapsulatus

“…To investigate whether Myxobolus ampullicapsulatus infection could cause immunosuppression of gill tissue in goldfish, immunohistochemical analysis was conducted. T/NK cells, positive signals of ZAP-70, were observed in both the control group ( Figures 9A, B ) and Myx group ( Figures 9D, E ) , mainly distributed in ILT (Interbranchial lymphoid tissue) and ALT (Amphibranchial lymphoid tissue) ( 20 ), and partially dispersed in the gill arch and gill lamella. However, there was no significant difference in the mean optical density ( Figure 9C ) nor IHC score (Immunohistochemistry score) ( Figure 9F ) of gill tissue between the two groups, suggesting that infection with Myxobolus ampullicapsulatus could lead to immunosuppression of gill tissue to a certain extent without the increase in the number of gill lymphocytes.…”

A tessellated lymphoid network provides whole-body T cell surveillance in zebrafish

“…We characterized the TLN by flow cytometry and compared it to other known reservoirs of T cells in fish: the gills, gut, kidney, and spleen (Fig. 2G) (6, 7, 20). The TLN, gills, and gut all had a similar proportion of GFP-positive cells (Fig.…”

“…Further, since diverse lymphocyte repertoires emerged approximately 500 million years before lymph nodes, adaptive immunity necessarily evolved a different surveillance strategy than the one operating in mammals (4, 5). While lymphocytes typically populate and even form lymphoid tissues around the respiratory and digestive organs of lower jawed vertebrates (6, 7), these animals produce robust adaptive immune responses when these barrier sites are bypassed by intraperitoneal or intramuscular immunization (8, 9). These data indicate that non-mammalian jawed vertebrates also perform whole-body antigen surveillance, but how T cells effectively search for antigen in the absence of lymph nodes remains unknown.…”