Differentiation Potential Of Bone Marrow Mesenchymal Stem Cells Into Lymphatic Endothelial Cells

Introduction:Mesenchymal stem cells(MSCs)are a type of non-hematopoietic stem cells which reside in the bone marrow. These cells possess the capacity of self-renew and multipotential,according to recent studies,it can also inducing immune suppression,Moreover,MSCs are easy to isolate and replicate extensively in vitro. MSCs have already been used in treatment of ischemic and injury diseases, At present, replacement therapies of MSCs are carried out in many field.It is agreed that breast cancer related lymphedema is a chronic, debilitating condition that has traditionally been seen as refractory or incurable diease. we acknowledged that the main cause of breast cancer related lymphedema is the disruption of lymphatic vessels when conduct axillary dissection and (or)sentinel lymph node dissection.Lymphangiogenesis is undoubtedly the key point to cure lymohedema. We hypothesized that if we can find a method to enhance lymphangiogenesis to reconstitute the collecting lymphatic vessels and to cure the condition.Objectives:To culture and explore the differentiation potential of bone marrow mesenchymal stem cells (MSCs) into lymphatic endothelial cells in vitro.Methods:MSCs were isolated from human bone marrow use adherence culture method. After amplification and cultured to3-4passage,MSCs were detected by flow cytometry for CD90、CD105、CD14、CD34and HLA-DR. Than MSCs were divided into two groups._experimental group as compared with control group. The experimental group were cultured in endothelial cell growth medium-2with VEGF-C (156s),the control group were cultured in endothelial cell growth medium-2only.After7days,we observed the morphological changes of MSCs under the phase contrast microscope. Podoplanin VEGFR-2and VEGFR-3were detected by Flow cytometry and their gene were examined by RT-PCR.Tubulogenesis formation assay was performed to verified the fuction of the new lymphatic endothlieal cells.Results:adherence cultured MSCs showed a fibroblast-like morphology. After cultured for3passages,It expressed high levels of CD90(90.03%), CD105(97.86%) and only a very low level of CD34(2.00%), CD14(2.18%) and HLA-DR(1.56%). After induction, The experimental group exhibited a monolayer with cobblestone appearance, the expression rate of podoplanin, VEGFR-2and VEGFR-3are26.53%、2.53%and8.55%_respectively.In the control group, the expression rate of podoplanin VRGFR-2and VEGFR-3are2.84%、1.61%and2.30%_respectively only.podoplanin and VRGFR-2have statistical significance(P<0.01),but VEGFR-3has no significant difference compared with control group.RT-PCR analysis also showed that podoplanin and vegfr-3mRNA expressed in the experimental group selectively, without vegfr-2.In the control group,we can see the expression of podoplanin. Tubulogenesis formation assay demonstrted that differentiational MSCs have the fuction of forming capillary-like structures in vitro.Conclusions:1. The findings provide evidence that it is possible to isolate mesenchymal stem cell from human bone marrow.2. MSCs were capable of expressing a lymphatic phenotype when exposed to endothelial cell growth medium-2with purified VEGF-C (156s)3. Induced MSCs can form Lymphatic Vessel chondrocyte.4. MSCs have mmuch more Application Prospect in Lymphagiogenesis.