技术资料

Bruton tyrosine kinase (Btk) is essential for B cell development and function and also appears to be important for myeloid cells. The bone marrow of Btk-deficient mice shows enhanced granulopoiesis compared with that of wild-type mice. In purified granulocyte-monocyte-progenitors (GMP) from Btk-deficient mice,the development of granulocytes is favored at the expense of monocytes. However,Btk-deficient neutrophils are impaired in maturation and function. Using bone marrow chimeras,we show that this defect is cell-intrinsic to neutrophils. In GMP and neutrophils,Btk plays a role in GM-CSF- and Toll-like receptor-induced differentiation. Molecular analyses revealed that expression of the lineage-determining transcription factors C/EBPα,C/EBPβ,and PU.1,depends on Btk. In addition,expression of several granule proteins,including myeloperoxidase,neutrophilic granule protein,gelatinase and neutrophil elastase,is Btk-dependent. In the Arthus reaction,an acute inflammatory response,neutrophil migration into tissues,edema formation,and hemorrhage are significantly reduced in Btk-deficient animals. Together,our findings implicate Btk as an important regulator of neutrophilic granulocyte maturation and function in vivo. View Publication

There is potential interest for combining allogeneic hematopoietic cell transplantation (HCT),and particularly allogeneic HCT with a nonmyeloablative regimen,to the tyrosine kinase inhibitor imatinib (Glivec; Novartis,Basel,Switzerland,http://www.novartis.com) in order to maximize anti-leukemic activity against Philadelphia chromosome-positive leukemias. However,because imatinib inhibits c-kit,the stem cell factor receptor,it could interfere with bone marrow engraftment. In this study,we examined the impact of imatinib on normal progenitor cell function. Imatinib decreased the colony-forming capacity of mobilized peripheral blood human CD133(+) cells but not that of long-term culture-initiating cells. Imatinib also decreased the proliferation of cytokine-stimulated CD133(+) cells but did not induce apoptosis of these cells. Expression of very late antigen (VLA)-4,VLA-5,and CXCR4 of CD133(+) cells was not modified by imatinib,but imatinib decreased the ability of CD133(+) cells to migrate. Finally,imatinib did not decrease engraftment of CD133(+) cells into irradiated nonobese diabetic/severe combined immunodeficient/beta2m(null) mice conditioned with 3 or 1 Gy total body irradiation. In summary,our results suggest that,despite inhibition of hematopoietic progenitor cell growth in vitro,imatinib does not interfere with hematopoietic stem cell engraftment. View Publication

Enforced expression of the HOXB4 transcription factor and downregulation of p21(Cip1/Waf) (p21) can each independently increase proliferation of murine hematopoietic stem cells (HSCs). We asked whether the increase in HSC self-renewal generated by overexpression of HOXB4 is enhanced in p21-deficient HSCs. HOXB4 was overexpressed in hematopoietic cells from wild-type (wt) and p21-/- mice. Bone marrow (BM) cells were transduced with a retroviral vector expressing HOXB4 together with GFP (MIGB4),or a control vector containing GFP alone (MIG) and maintained in liquid culture for up to 11 days. At day 11 of the expansion culture,the number of primary CFU-GM (colony-forming unit granulocyte-macrophage) colonies and the repopulating ability were significantly increased in MIGB4 p21-/- BM (p21B4) cells compared with MIGB4-transduced wt BM (wtB4) cells. To test proliferation of HSCs in vivo,we performed competitive repopulation experiments and obtained significantly higher long-term engraftment of expanded p21B4 cells compared with wtB4 cells. The 5-day expansion of p21B4 HSCs generated 100-fold higher numbers of competitive repopulating units compared with wtMIG and threefold higher numbers compared with wtB4. The findings demonstrate that increased expression of HOXB4,in combination with suppression of p21 expression,could be a useful strategy for effective and robust expansion of HSCs. View Publication