Objective To observe the effects of cryopreservation and resuscitation on the biological characteristics of mesenchymal stem cells (MSCs) derived f rom human umbilical cord blood. Methods MSCs were isolated and cultured f rom human umbilical cord blood in vitro. The cells were passaged , and the third generation of MSCs were cryopreserved in－196 ℃ liquid nitrogen for 4 weeks with cryopreservation medium , which contained 10 % dimethyl sulfoxide (DMSO) and 90 % fetal calf serum ( FCS) . The morphology , proliferation and differentiation of MSCs were investigated and compared with those of MSCs before cryopreservation. Results There was no significant difference of morphology between pre-cryopreserved MSCs and the ones af ter resuscitation. It was observed that all MSCs were spindle-shaped and showed adherence growth characteristic before and af ter cryopreservation. The cell growth curves of MSCs were also similar before and af ter cryopreservation. Even though the curve of resuscitated MSCs descended a little as compared with that of pre-cryopreserved MSCs , there was no significant difference ( Pgt; 0. 05) . After 2-week adipocytic differentiation induction , fat drops could be found in the kytoplasm of MSCs and they were red when stained with oil-red O staining , which suggested that MSCs could be induced and differentiated into adipocytes. Af ter 4-week osteoblastic differentiation induction , MSCs could be induced and differentiated into osteoblasts , and calcium node showed black when stained with Von Kossa staining. There were no significant changes of the differentiating ability of MSCs into adipocyte and osteoblast before and after cryopreservation. Conclusion MSCs derived from human umbilical cord blood maintains their biological characteristics af ter cryopreservation and resuscitation.
Objective To investigate the optimal dosage of bone marrow mesenchymal stem cells （BMSCs） transplantations for treatment of hepatic ischemia-reperfusion injury in rats， and to provide prophase experimental basis for it. Methods BMSCs of Wistar rats were isolated and cultivated by bone marrow adherent culture method. BMSCs of the fourth generation were prepared for cell transplantation. Thrity hepatic ischemia-reperfusion injury models of maleWistar rats were successfully established， and then were randomly divided into blank control group， 5×105 group， 1×106group， 2×106 group， and 3×106 group， each group enrolled 6 rats. The 200 μL cell suspension of BMSCs were transfusedinto the portal vein with number of 5×105， 1×106， 2×106， and 3×106 separately in rats of later 4 groups， and rats of blank control group were injected with phosphate buffered saline of equal volume. At 24 hours after cell transplantation， blood samples were collected to test aspartate aminotransferase （AST） and alanine aminotransferase （ALT）， liver tissueswere obtained to test malonaldehyde （MDA）， superoxide dismutase （SOD）， and nuclear factor-κB （NF-κB） p65 protein.Liver tissues were also used to perform HE staining to observe the pathological changes. Results Compared with blank control group， 5×105 group， and 3×106 group， the levels of AST， ALT， and MDA were lower （P＜0.05） while activity levels of SOD were higher （P＜0.05） in 1×106 group and 2×106 group， and expression levels of NF-κB p65 protein were lower with the pathological injury of liver tissue improved， but there were no significant differences on levels of AST， ALT， MDA， and SOD （P＞0.05）， and both of the 2 groups had the similar pathological change. Conclusion The optimal dosage of the BMSCs transplantations after hepatic ischemia-reperfusion injury is 1×106.
Objective To explore repair role of allogeneic bone marrow mesenchymal stem cells （BM-MSCs） transplantation on treating hepatic ischemia reperfusion injury （HIRI） in rats. Methods Ten rats were executed to get BM-MSCs， then BM-MSCs were cultured in vitro and dyed by 4，6-diamidino-2-phenylindole （DAPI）. Models of 70% hepatic ischemia reperfusion injury were eatablished. Thirty two rats were randomly divided into sham operation group （Sham group）， ischemia reperfusion group （I/R group）， Vitamin C group （VC group）， and BM-MSCs group. Serum samples were analyzed for ALT and AST， and hepatic tissue were for superoxide dismutase （SOD） and malondialdehyde （MDA）. Liver sections were stain with hematoxylin and eosin （HE） for histological analysis， TUNEL staining was applied to detect hepatic apoptosis. Serum and tissues were both collected at 24 h after reperfusion. Results The isolated BM-MSCs maintained vigorous growth in vitro. Specific markers for MSCs antigens CD29 and CD44 were detected by flow cytometry， but antigens CD34 and CD45 were not be detected. Models of HIRI were stable， and BM-MSCs were detected around the periportal area by DAPI staining. Compared with I/R group， levels of ALT， AST， MDA， and AI in the VC group and BM-MSCs group decreased at 24 h after reperfusion （P＜0.05）， meanwhile SOD level increased （P＜0.05）. Compared with VC group， levels of ALT， AST， MDA， and AI in the BM-MSC group decreased at 24 h after reperfusion （P＜0.05）， meanwhile SOD level increased （P＜0.05）. Conclusion BM-MSCs could protect HIRI by alleviating oxidative stress and inhibiting cellular apoptosis.