Effects of lethal irradiation in zebrafish and rescue by hematopoietic cell transplantation

Abstract

The study of hematopoiesis has been greatly facilitated by transplantation of blood cell populations into recipient animals. Efficient engraftment of donor cells generally requires ablation of the host hematopoietic system. The zebrafish has recently emerged as a developmental and genetic system to study hematopoiesis. To enable the study of hematopoietic stem cell ( IntroductionThe study of the biologic effects of ␥ irradiation began over a century ago, 1 but it was not until deployment of the atomic bomb in 1945 that the clinical manifestations of lethal total body irradiation (TBI) were fully realized. Acute irradiation injury was replicated using animal models, and seminal experiments by Jacobsen et al 2,3 and Lorenz et al 4 showed that shielding or transplantation of hematopoietic tissues was sufficient for radioprotection of otherwise lethal doses. Although humoral factors produced from these tissues were initially proposed to be the radioprotective elements, 2 subsequent studies showed that survival correlated with the level of hematopoietic repopulation by donor-derived cells. [5][6][7][8] Pioneering studies by Till and McCulloch and colleagues showed that donor bone marrow contained rare cells that generated clonal, macroscopic spleen colonies, 9,10 a fraction of which regenerated spleen colonies 11 or repopulated multilineage hematopoiesis 12 in hosts receiving serial transplants. These experiments led to the concept of the hematopoietic stem cell (HSC).TBI has since been used to condition hosts prior to transplantation of donor bone marrow, both therapeutically in humans and as a means of testing murine blood cell subsets for the presence of HSCs. The development of monoclonal antibodies and multiparameter flow cytometry, combined with hematopoietic cell transplantation (HCT), has led to the prospective isolation of murine HSCs, [13][14][15][16] committed lymphoid 17 and myeloerythroid 18 progenitors, and their downstream progeny by cell-surface phenotypes. These powerful techniques have allowed individual components of the immune system to be tested functionally, which has led to the dissection of the hematopoietic hierarchy and the resolution of effector cell function. HCT also provides the means to study the effects of genetic mutations in isolated cell types when introduced into a normal cellular environment.The zebrafish has recently emerged as a unique vertebrate model in which forward genetic screens can uncover novel genes involved in blood cell development and function. 19 Many zebrafish blood mutants identified in these screens are embryonic lethal, and we have recently described embryonic HCT to address issues of cell autonomous mutant gene function. 20 Similar transplantation of whole kidney marrow (WKM) cells into unconditioned adult recipients resulted in the disappearance of donor-derived cells within several weeks of transplantation. This suggests that donorderived cells are either rejected by the host immune system or that engraftment of donor cells requires the creation of niche spa...

Isolation and characterization of hematopoietic stem cells in teleost fish

“…Transplantation of thymocytes and kidney marrow cells from lck:GFP animals resulted in the transient and permanent repopulation of the thymus in the recipients, respectively. These results indicate that the thymus contains only short-term thymic repopulating progenitors, whereas long-term repopulating HSCs are present in the kidney (Langenau et al, 2004).…”

“…Transplantation of kidney hematopoietic cells from normal donors rescued the 40 Gy-irradiated recipients (Traver et al, 2004). Thus, transplantation of hematopoietic cells from the teleost kidney recapitulates the radioprotective effect of comparable transplants in the mouse model (Lorenz et al, 1951).…”

CD41+ cmyb+ precursors colonize the zebrafish pronephros by a novel migration route to initiate adult hematopoiesis

“…In addition to many transplantation experiments from embryo to embryo, we also transplanted AGM CD41 + cells into conditioned adult recipients. Whereas we routinely achieve LTR using adult kidney cells transplanted into sublethally irradiated adult hosts (Traver et al, 2004), we have not observed LTR when using any cell population from embryonic donors. In the mouse, it has been demonstrated that embryonic cells do not express class I MHC molecules before E10.5 (Ozato et al, 1985).…”

Co-injection strategies to modify radiation sensitivity and tumor initiation in transgenic Zebrafish

“…Animals were scored for tumors at 25 days of life, killed and fixed in 4% PFA. Subsequently, animals were sectioned and stained by TUNEL or hematoxylin/eosin as described previously (Traver et al, 2004). Tumor sections were photographed under  1000 oil emersion and apoptotic cell number and total cell number were counted per field (n ¼ 3 fields per sample, Supplementary Figure 3).…”