Easily-handled method to isolate mesenchymal stem cells from coagulated human bone marrow samples

AIM:To establish an easily-handled method to isolate mesenchymal stem cells(MSCs) from coagulated human bone marrow samples. METHODS: Thrombin was added to aliquots of seven heparinized human bone marrow samples to mimic marrow coagulation. The clots were untreated,treated with urokinase or mechanically cut into pieces before culture for MSCs. The un-coagulated samples and the clots were also stored at 4 ℃ for 8 or 16 h before the treatment. The numbers of colony-forming unit-fibroblast(CFU-F) in the different samples were determined. The adherent cells from different groups were passaged and their surface profile was analyzed with flow cytometry. Their capacities of in vitro osteogenesis and adipogenesis were observed after the cells were exposed to specific inductive agents.RESULTS: The average CFU-F number of urokinasetreated samples(16.85 ± 11.77/106) was comparable to that of un-coagulated control samples(20.22 ± 10.65/106,P = 0.293),which was significantly higher than those of mechanically-cut clots(6.5 ± 5.32/106,P < 0.01) and untreated clots(1.95 ± 1.86/106,P < 0.01). The CFU-F numbers decreased after samples were stored,but those of control and urokinase-treated clots remained higher than the other two groups. Consistently,the numbers of the attached cells at passage 0 were higher in control and urokinase-treated clots than those of mechanically-cut clots and untreated clots.The attached cells were fibroblast-like in morphology and homogenously positive for CD44,CD73 and CD90,and negative for CD31 and CD45. Also,they could be induced to differentiate into osteoblasts and adipocytes in vitro. CONCLUSION: Urokinase pretreatment is an optimal strategy to isolate MSCs from human bone marrow samples that are poorly aspirated and clotted.

Responses of HFR-LWC beams under close-range blast loadings accompanying membrane action

The load-carrying capacities and failure patterns of reinforced concrete components can be significantly changed by membrane effects.However,limited work has been carried out to investigate the blast resistance of Hybrid Fiber Reinforced Lightweight Aggregate Concrete(HFR-LWC)members accompanying membrane action.This paper presents a theoretical approach to quantitatively depicting the membrane behavior and its contribution on the behavior of HFR-LWC beams under close-range blast loadings,and the suitability of the proposed model is validated by a series of field tests.An improved Single-Degree-of-Freedom(SDOF)model was employed to describe the dynamic responses of beam-like members under blast loadings accompanying membrane action,where the mass-load coefficient is determined according to the nonuniformly distributed load induced by close-range explosion,and the membrane action is characterized by an in-plane(longitudinal)force and a resisting moment.The elastoplastic and recovery responses of HFR-LWC beams under the combined action of blast load and membrane force were analyzed by the promoted model.A specially built end-constrain clamp was developed to provide membrane action for the beam member when they are subjected to blast load simultaneously.It is demonstrated that the analytical displacement-time histories are in good agreement with experimental results before peak deflections and that the improved SDOF model is an acceptable tool for predicting the behavior of HFR-LWC beams under blast loadings accompanying membrane action.