| Mesenchymal stem cell (MSC), originating from mesoblast, resides mainly in post-natal bone marrow as well as human umbilical cord blood, and coexists with hematopoietic stem cell (HSC). Functionally MSC not only exerts the crucial role of supporting hemopoiesis by participating in the formation of hematopoietic microenvironment, but also maintains its capacity for renewal and its potential of multilineage differentiation when induced properly. So, MSC is referred to as adult multipotent stem cell.The existence of non-hematopoietic stem cell in bone marrow was first proposed by Cohnheim in 1867. In 1976, Friedenstein isolated roughly from bone marrow the cell component through cell adherenting method. Whereafter more and more cultures of bone marrow-derived MSC were established. In 2000, Erices separated and established the culture of human umbilical cord blood-derived MSC.Several recent reports suggest that MSC, whatever it is intro- or extro- bone morrow derived, can be induced to differentiate into diverse cell types. The formal differentiation results in connective tissue cells, such as osteoblast, chondrocyte, adipocyte, tendon cell, and myogenic cell. The untraditional differentiation leads to acquire properties of cells of other lineages. This kind of transgermic differentiation is called transdifferentiation (TD). For example, MSC may transdifferetiate into neurons. At present, neuron-oriented differentiation of MSC are mostly based on bothSanchez-Ramos protocol, whose induction includes mainly trophic factors/growth factors; and Woodbury protocol, which emphasizes the utility of antioxidants. After induction, special protein markers of neuron/ neural stem cell, such as nestin, trkA, NSE, NeuN, NF-M, tau, etc, are expressed in all levels in MSC.In this study, MSC was isolated respectively from rabbit femeral bone marrow and human umbilical cord blood sterilely, and cultivated in vitro to observe the regularities of cell growth, recovery, adherenting, and proliferation. Growth curves of different passges (P=5 and P=10) of rabbit MSC were drawn according to the MTT test. Modified Woodbury protocol was adopted to induce the neuron-oriented differentiation of rabbit bone marrow-derived MSC, that is, BME was applied to MSC in low dose and a relatively short span (D=3mM, and T=2h). Then, induced MSC was cultivated subsequently and protein markers NSE and nestin were identified with immunocytochemistric method. At the same time, MSC pre- and post- induction was tested for the existence and change of K+ ionic current by whole-cell recording method of patch clamp, in order to observe the electrophysiological properties. All data were presented in the format "means" or "means + S", and compared statistically by analysis of variance (a =0.05) .Result: 1 Morphologically diverse cells were obtained from the cultures of human umbilical cord blood, but formed no fusion and no steady proliferation. However, expanding-MSC cultures were established from the samples of rabbit bone marrow in about 3w, which were pure and well-organized cell groups, and steadily passaged more than 16 passages in vitro. 2 The growth feature was changless after passaging approved by MTT test. Although cells grew slower when recovery, turn back to normal after passaging. 3 NSE and nestin were identified in induced rabbit bone marrow derived-MSC irnmunocytochernistrically. The average positive rates were about 71%~73% and 37%~38% respectively. And the positive rates kept changless in subsequent culture in neural medium. 4 Giga-sealing can be formed in rabbit bone marrow-derived MSC pre- and post- induction in dual time span, which suggests the feasibility of further whole-cell recording test for the ionic currents. But no ionic current is recorded.Conclusion: 1 MSC derived from rabbit bone marrow can establish expanding-culture in vitro, and no change was observed in either morphology or characteristic after passaging and recovery. Human umbilical cord blood contains MSC, which may possibly form the same culture in vitro. 2 After BME...
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