Stress-induced hematopoietic failure in the absence of immediate early response gene X-1 (IEX-1, IER3)

Abstract

© F e r r a t a S t o r t i F o u n d a t i o ntributes significantly to MDS pathogenesis in these patients. The present study shows that null mutation of IEX-1 perturbs HSC quiescence and impairs maturation of platelets and red blood cells (RBCs). Upon BM transplantation or non-myeloablative radiation, changes resembling MDS occurred, including thrombocytopenia, a trend toward anemia, and granulocyte dysplasia. These observations underscore an indispensable role for IEX-1 in maintenance of HSC quiescence as well as in multiple differentiation steps along thrombopoiesis and erythropoiesis. IEX-1-deficient mice confer a novel model for investigating how mitochondrial dysfunction may cause MDS and how to prevent or slow down the disease. Methods MiceIEX-1 knockout (KO) mice on mixed 129Sv/C57BL/6 background were generated by gene-targeting deletion in our laboratory, as previously described. 17 The KO mice were bred with wild type (WT) C57BL/6 (B6) for 10 generations to obtain IEX-1 KO B6 mice. WT B6 mice and mice on 129Sv/B6 background were derived from breeding as above and used as controls. Cognate BL/6.SJL-Ptprc a Pep 3 /BoyJ (CD45.1) or PepJ mice were obtained from Jackson Laboratory. All experimental mice and BM donor mice were used at 6-8 weeks of age. Animals were maintained in the pathogen-free animal facilities of Massachusetts General Hospital in compliance with institutional guidelines. All animal studies were approved by the Subcommittee on Research Animal Care of the institute. Extended methods can be found in the Online Supplementary Appendix. Results Perturbation of HSC quiescence in the absence of IEX-1IEX-1 KO mice display no gross developmental defect except for modest hypertension at 2-months of age or older, 17,18 in agreement with its function required primarily in response to stress. Given the significant loss of IEX-1 reported in patients with early stage MDS, 14,16 we examined the status of LSK (Lineage-Sca-1 + c-Kit + ) stem cells in IEX-1 KO mice. Loss of IEX-1 led to a significant decline in the proportion of LSK cells, in comparison with WT mice (0.10±0.07% vs. 0.24±0.06 %; P<0.01) ( Figure 1A). Apparently, the decline contradicted increased cycling of the cells, as shown in Figure 1B and C, in which relatively higher proportions of KO LSK cells entered G1 and G2/S/M cycling phases than WT counterparts, concurrent with a reduced percentage of the cells at a Go or quiescent phase. We thus examined whether the decline was ascribed to increased apoptosis of the cells, because apoptosis in HSCs was abnormally high in many MDS patients. 19 Increased apoptosis of IEX-1 KO over WT BM cells was confirmed by apoptosis-specific terminal deoxynucleotydyl transferase-mediated dUTP nick end labeling or TUNEL (11.6±0.7% vs. 2.2±0.4%; P<0.01), which detected apoptosis at both early and late stages. Early apoptotic marker Annexin V staining further revealed that apoptosis was increased by more than 50% in the BM (6.9±0.5% vs. 4.5±0.3%; P<0.01) ( Figure 1D In spite of a reduced percentage of L...

Noninvasive low-level laser therapy for thrombocytopenia

“…1G). To recapitulate this finding in vivo, mature MKs were sorted, treated with LLL or sham light, labeled with vital fluorescent dye carboxyfluorescein diacetate succinimidyl ester (CFSE), and intravenously infused into recipient mice (11). LLLtreated MKs generated higher levels of platelets than control counterparts from day 2 to day 5 after infusion in recipients (Fig.…”

“…Point mutations in mitochondrial cytochrome c cause dysregulated platelet formation and thrombocytopenia in humans (10), suggesting that platelet biogenesis is sensitive to mitochondrial activity. We recently showed that inadequate mitochondrial function predisposed mice lacking immediate early responsive gene X-1 (IEX-1) to thrombocytopenia upon stress (11). One of the major functions of IEX-1 is to enhance ATP synthase activity at the mitochondrial respiratory chain (12), and its null mutation compromises ATP generation and increases the production of reactive oxygen species (ROS) in mitochondria (13).…”

Extramedullary Hematopoiesis (EMH) in Laboratory Animals: Offering an Insight into Stem Cell Research

“…IEX-1 −/− mice developed MDS after total body irradiation (93). The BM erythroid dysplasia occurred in parallel to increasing EMH, with a relatively higher level of CD71 + /Ter-119 + erythroblasts and internuclear bridge and binucleated erythroid precursors in irradiated IEX-1 −/− spleens than WT counterparts.…”

“…IEX-1 −/− HSCs from BM produced significantly fewer circulating platelets and red blood cells when transplanted into WT mice, despite their enhanced repopulation capability. These findings highlight a role for this early response gene in multiple differentiation steps within hematopoiesis, including erythropoiesis and in the regulation of HSC quiescence (93).…”

Low-level light treatment ameliorates immune thrombocytopenia

“…5b). Since platelet apoptosis is correlated with platelet lifespan and clearance23, we monitored circulating platelets in these mice after injection of a biotinylation agent as previously described24, which is a common assay for monitoring the lifespan of platelets in the circulation. In comparison with control mice, the percentage of biotin-labeled platelets was decreased precipitously on day 2 and day 3 and remained below a 50% level on day 3 in ITP + sham mice (Fig.…”