Bone marrow endothelial cells secrete thymosin beta 4 and AcSDKP

Objective. Bone marrow endothelial cells are the essential component of the bone marrow microenvironment. They produce many kinds of cytokines, includ ing: stimulators and inhibitors, Many researchers have suggested that in th e presence of endothelial cell layer, CD34(+)CD38(-) cells are capable of e xpansion. The ability of the endothelial cell layer to protect hematopoieti c stem cells from extensive differentiation may be related to the inhibitor s derived from endothelial cells, The aim of the present study was to deter mine whether the inhibitors thymosin beta4 and AcSDKP are elaborated by mur ine bone marrow endothelial cells, Materials and Methods. Murine bone marrow endothelial cells (mBMECs) were c ultured in serum-free conditioned medium, Reverse transcriptase polymerase chain reaction (RT-PCR) was used to analyze the differential expression of the thymosin-beta gent, and reverse phase high-performance chromatography ( HPLC and mass spectroscopy were used to determine the concentration of thym osin beta4 (T beta4) and AcSDKP in EC lysate and in the medium (mBMEC-CM). Colony-forming unit granulocyte-macrophage (CFU-GM) colony assays were used to examine the effect of components (mw 3-10 kD, <3 kD) of mBMEC-CM, thymo sin <beta>4, and AcSDKP on the proliferation of hematopoietic cells. Results. mBMECs expressed T beta3 mRNA, In EC lysate and mBMEC-CM, T beta4 and AcSDKP were detected. After adding protease inhibitors, the concentrati on of T beta4 in EC lysate increased significantly, while the concentration of AcSDKP decreased. mBMEC-CM (mw 3-10 kD) had no effect on the formation of CFU-GM, How;ever, mBMEC-CM (mw <3 kD) could inhibit the growth of CFU-GM , T<beta>4 (10(-11)similar to 10(-7)mol/L) and AcSDKP (10(-11)similar to 10 (-5)mol/L) had dose-dependent inhibitory effects on the growth of CFU-GM, A ngiotensin converting enzyme (ACE), the enzyme degrading AcSDKP, could part ially eliminate the inhibitory effect of mBMEC-CM (mw <3 kD) on CFU-GM. Conclusion. BMECs express and secrete T<beta>4 and AcSDKP, T beta4 exists i n the 3-10 kD component of mBMEC-CM, while AcSDKP exists in the <3 kD compo nent of ECCM, Both components exert inhibitory effects on the proliferation of hematopoietic progenitors. (C) 2001 International Society for Experimen tal Hematology. Published by Elsevier Science Inc.