| Part1Effects of Low-Dose-X-Irradiation for rabbit mesenchymal stemcells: an in vitro studyObjective: Rabbit bone marrow stromal cells (MSCs) were cultured in vitro foramplification and identification. Cell cycle distribution and apoptosis phase was detectedby flow cytometry and proliferative activity was detected by CCK-8after low-dose X-rayirradiation. Furthermore, explore the proliferation of MSCs after low-dose irradiation andits relationship with the MAPK/ERK signaling pathway, providing an experimental basisfor further research.Methods:5mL bone marrow was extracted from the rabbit tubercle of tibia. Theoriginal states and passages were observed under an inverted microscope. Then the cellsurface markers (i.e., CD34, CD44and CD90) were identified by flow cytometry. Afterirradiation with CCK-8assay to detect the proliferation of cells, taking Western-blot forthe best dose group, relationships observed X-ray irradiation on proliferation of MSCs andMAPK/ERK signaling pathway.Results: The MSCs expressed CD44and CD90, however, did not express CD34. Inaddition, CCK-8assay test results indicate that low-dose irradiation of75mGy cansignificantly stimulate the proliferation of MSCs, and had been statistically significantcompared with other irradiated groups. Conclusions: The cultured cells have the essential phenotypes and characteristics ofthe MSCs. X-ray irradiation affect on the cell cycle and regulate the MAPK/ERKsignaling pathway to stimulate the proliferation of MSCs. Part2Mesenchymal stem cells of Low-Dose-X-Irradiation for discregeneration in a rabbit modelObjective: To investigate the effectiveness of MSCs after low-dose-X-irradiation forthe repair of rabbit intervertebral disc degeneration.Methods:20white New Zealand rabbits are randomly divided into three groups:1)normal group (n=4);2) control group (n=8); and3) experimental group (n=8). Induce andestablish the degeneration merely by means of puncturing the annulus fibrosus withneedles of defined gauges and aspirating some nucleus pulposus tissue from theintervertebral discs,2weeks after inducing degeneration model, in control group, onlywere induced and established the degeneration; but in experimental group, the MSCs oflow-dose-X-irradiation were transplanted. Transplanted into the intervertebral discs afterfour and six weeks, each time point select normal group2, control group4, theexperimental group4for examing the lumbar lateral X-ray, and calculating the disc heightindex (DHI); measuring disc T2-weighted signal intensity changes by lumbar MRI;detecting the gene expression of disc type II collagen and aggrecan by RT-PCR andevaluating the disc degeneration and grading by HE staining.Results: the disc height index (DHI) moderate decreases in experimental controlgroup and experimental group (P＜0.05). MRI results showed that the control group andthe experimental group disc signal intensity varying degrees of decline, but theexperimental group was significantly stronger than the signal intensity of experimentalcontrol group(P＜0.05), In the control group, histological analysis showed significantly disordered annulus fibrosus; but slightly disordered annulus layered structure with freshnucleus pulposus in the experimental group. RT-PCR results showed that the experimentalgroup of type II collagen and aggrecan mRNA expression levels have a significantdifference, comparing with the control group (P＜0.05).Conclusion: After low-dose X-ray irradiation of bone marrow mesenchymal stemcells transplanted into the intervertebral disc degeneration, X-ray, MRI, pathologydetection, RT-PCR confirmed retard disc degeneration, promoting disc type II collagenand aggrecan gene expression and effectively repairing disc degeneration with a brightfuture. |
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