Synergistic Effects Of Cyclosporin A And Hematopoietic Growth Factor On Human Bone Marrow Cell Multiplication In Longterm Bone Marrow Culture And Colonal Culture In Semi-Solid Ayar Medium

Cyclosporin A (CYA) is a potent immunosuppressive drug widely used in transplant settings due to its specific inhibition of T–cell activation. Several Studies have reported the synergistic effect of CyA with hematopoietic growth factor in colony culture of mice bone marrow. We Studied the effect of IL-3, IL-6, SCF,CyA and nutrition medium ( condition medium - PHA) on the development of hematopoietic cells of human bone marrow because this unique cyclic endecapeptide has been successfully utilized in organ transplantation for graft rejection and graft versus host disease and also since CYA stimulates colony formation by hematopoietic cells in vitro.

Human bone marrow cells were taken from the posterior illiac crest of volunteer donors aged 5-35 years. Cells were cultured in complete culture medium (Containing 12.5% FCS 12.5% horse serum and 50 µmol Hydrocortisone with IL-3, IL-6, SCF and CY-A) on 24 –well microplates at 37 degree centigrade with 5% co2 for 4 week and colony culture was performed for 14 days in semisolid Agar medium .The cell count by light microscope and colony assay by inverted microscope were performed weekly for four weeks.

Long term bone marrow culture and semisolid agar culture stimulation with IL-3, IL-6 , SCF and CyA had two as much cloning efficiencies than that of parallel cultures without CyA and IL-3, IL-6 ,SCF in inducing proliferation and conservation of CFU – GM in Bone marrow culture. Culture inactive PHA- condition medium showed higher stimulation than active condition medium. Cloning efficiencies were calculated by mean mean colony number per medium and data were presented as comparison of mean colonies in CyA group to control group.

Statistical analysis using the t-student test was performed to determine significance of differences between cultures in presence or absence of CyA. Consequently,these results show a direct positive effect of Cy A on the signal transduction pathways in hematopoietic stem and progenitor cells.