Bone marrow stromal cells of bcl-2 transgenic mice express widespread Bcl-2 protein and reduce apoptosis in a serum-free medium
Stem cell transplantation is a vibrant area of research with the goal of treatment of uncurable diseases. Bone marrow stromal cells (MSCs, including marrow stem and progenitor cells) have been employed to improve neurological functional recovery after stoke and injury. Grafted cell death is a common feature of intracerebral transplantation. Bcl-2 has been shown to protect a variety of cell types from apoptoic cell death. We tested the hypothesis that MSCs harvested from bcl-2 transgenic mice promote survival and decrease apoptosis of cells in vitro. MSCs were harvested from adult male C57BL/6 transgenic mice (n=4) overexpressing bcl-2, and from normal control C57BL/6 mice (n=4). Cells were cultured in regular medium, Iscove s Modified Dulbecco s medium (IMDM) supplemented with 10% fetal bovine serum (FBS); and then isolated by their adherence to the plastic dishes from non-adherent hematopoietic cells at 72 h of incubation. To examine the effect of bcl-2 on cell death induced by serum deprivation. MSCs were washed and incubated with the serum-free medium. Immunostaining and Western blot were used to identify Bcl-2 protein expression, and TUNEL staining was used to identify in situ DNA fragmentation of apoptotic cells. Immunocytochemical staining indicated that 100% of MSCs harvested from bcl-2 transgenic mice express Bcl-2 protein. Western blot showed that Bcl-2 protein was 8 times more intense in the MSCs harvested from transgenic mice than that from normal control mice at 12 days cultured in regular medium. After 48 h cultured in the serum-free medium, 5.8% of MSCs from bcl-2 transgenic mice were presented apoptoic morphology, compared with 39% of MSCs from normal controls. Our finding demonstrates that the overexpression of bcl-2 gene in the MSCs reduces apoptotic cell death.