A probable role for trail-induced apoptosis in the pathogenesis of marrow failure: Implications from an in vitro model and from marrow of aplastic anemia patients

Received 4 June 2004; accepted 15 September 2005.

Abstract

Aplastic anemia (AA) is a syndrome of hematopoietic failure involving increased apoptosis of stem cells. In order to investigate the molecular mechanisms participated in the process of marrow failure, we created an in vitro model of hematopoietic cell suppression, by continuous addition of TNF-α and IFN-γ in an vitro long-term bone marrow culture system. An up-regulation of Fas expression was observed in CD34+ cells in cytokine treated cultures, compared to controls. This was accompanied by significant TRAIL and decreased caspase 3 mRNA expression, whereas the expression of Bcl-2 family members was low (Bcl-xl) or absent (Bcl-2, Bax). The expression of these apoptotic genes was also investigated in aplastic anemia patients. Apart from Fas mRNA expression in total marrow and/or CD34+ cells, TRAIL mRNA expression was found only in CD34+ cells in active disease while in total marrow cell compartment this remains a constant finding even in patients in remission. The above data are in agreement with previous studies proposing a major role for the extrinsic apoptosis pathway in the pathogenesis of aplastic anemia and additionally introduce TRAIL as a probable important molecule in the process.

Abbreviations: AA, aplastic anemia, BFU-e, burst forming unit-erythroid, BMMNC, bone marrow mononuclear cells, BSA, bovine serum albumine, CFCs, colony forming cells, CFU-GM, colony forming unit-granulocyte macrophage, CFU-GEMM, colony forming unit-granulocyte, erythroid, macrophage, megakaryocyte, ct, control, CPD-A, CPD-adenine, Epo, erythropoietin, GF, growth factors, GM-CSF, granulocyte macrophage colony stimulating factor, G-CSF, granulocyte-colony stimulating factor, IFN-γ, interferon-gamma, IL-3, interleukine-3, IL-6, interleukine-6, LTBMC, long-term bone marrow cultures, LTC-IC, long-term culture-initiating cells, MAb, monoclonal antibody, MPCR, multiplex PCR, PBS, phosphate buffered saline, PE, phycoerytrin, PI, propidium iodide, RT, reverse transriptase, SCF, stem cell factor, TNF-α, tumor necrosis factor alpha, TRAIL, tumor necrosis factor-related apoptosis-inducing ligand

Keywords: Aplastic anemia, IFN-γ, TNF-α, CD34+, Apoptosis, Fas, TRAIL

Hematology Division, Department of Internal Medicine, University Hospital of Patras, Rion, Patras 26500, Greece

Corresponding author. Tel.: +30 2610 999255; fax: +30 2610 993950.

PII: S0145-2126(05)00395-4

doi:10.1016/j.leukres.2005.09.015

14 of 21