An Experimental Study Of The Cultivation In Vitro And Tissue Compatibility In Vivo Of Reticulated Vitreous Carbon Combined Canine Bone Marrow Stromal Stem Cells

Objective: Making sure the reticulated vitreous carbon is non-toxic and conducive to cell growth in vitro. In vivo experiment, the reticulated vitreous carbon is implanted into femoral head and then tissue slices were deal with by Masson staining to observe the histocompatibility of the bone tissue and reticulated vitreous carbon.Methods: The experiment in vitro: the marrow stromal stem cells were extracted on distal of the canine′s femur. The method of differential sticking wall and medium selection was used to extract the bone marrow stromal stem cells. Then, the cells were subcultured in vitro. The third-generation cells were identified by Flow Cytometry and detected cytotoxicity of reticulated vitreous carbon by MTT. The third-generation cells were implanted on the surface of reticulated vitreous carbon at the cell concentration of 2.0 x 109-1 L. The cells of dual culture were observed the cell adhesion and proliferation after 14 days. The experiment in vivo: femoral greater trochanter is exposed by surgery. Then a hole was drilled into the 4 to 5 mm to subchondral bone with along the direction of the femoral neck from the intersection point of the vertical axis of the femoral neck and the lateral femoral, in where reticulated vitreous carbon was implanted. The canines were killed 6 weeks and 12 weeks later. The femoral greater trochanter with implant materials was cut down, decalcified and observed by Masson staining.Results:(1) The less cells were discovered nearby the material after 7 days by dual culture, but the more cells were discovered around the material under inverted microscope. The cells gradually moved closer to support material 14 days later, and there were a large number of cells adhere to the material of surface, with a small amount of cell matrix secreted. After 21 days, a large number of cells were proliferated and connected together, filled in the material surface and the gap, with a large amount of cell matrix secreted, some of the cells showed contact inhibition.(2) After 14 days cells were dual cultured with the material, the cells mostly appeared fusiform with good ductility, at the same time, cells were surrounded by a large amount of extracellular matrix, a small amount of cells extended pseudopodia and adhered to material surface under scanning electron microscopy.(3) The experiment in vivo showed that a lot of loose connective tissue and bone tissue were filled in the gap of reticulated vitreous carbon.Conclusion: Experiment shows that reticulated vitreous carbon is non-toxic and conducive to adhesion and growth of bone marrow stromal stem cells in vitro. Experiment shows that reticulated vitreous carbon have good histocompatibility in animals in vivo.