Characterization of Arnt KO phenotype. (A) Quantitative real-time PCR (qRT-PCR) analysis of ARNT expression in murine HSC subsets. (B) PCR analysis of DNA extracted from bone marrow and peripheral blood, from Arnt KO and heterozygous control (Ctrl) animals, demonstrating effective recombination of Arnt in the hematopoietic lineages. (C) Western blot for ARNT performed on FACS-sorted LSK, MPP, and LT-HSC cells shows loss of ARNT protein in all hematopoietic progenitors; a nonspecific band (NS) illustrates equivalent loading of protein. (D) qRT-PCR analysis of sorted HSCs shows effective deletion of Arnt in all progenitor populations. (E) Histologic analysis of spleen and thymus from 8- to 12-week-old KO mice. Note the change in germinal center architecture in KO spleens (original magnification ×50). (F) Absolute cell counts of hematopoietic tissues from control and KO mice. BM counts are from one femur and tibia (n = 6 ctrl, 6 KO). (G) Complete blood counts (CBC) on peripheral blood shows enhanced white blood cell (WBC) numbers in KO mice (n = 6 ctrl, 6 KO). (H) Representative blood smear illustrating WBC (arrows, upper panels) and spleen showing megakaryocytes (arrows, lower panels), which are quantified from at least 6 independent fields per slide in I and J, respectively (n = 7 ctrl, 6 KO; original magnification ×200). PB, peripheral blood; LN, lymph node; BM, bone marrow. *P < .05, **P < .01.