Transfected Immature Dendritic Cells By Human CD200 Gene Induce Regulatory T Cells In Vitro

The CD200 gene is a new member to be found of the IgSF (immunoglobulin superfamily)and is a molecular of immune regulation.CD200 gene has function of immunological regulation and expresses on a variety of cells generally, including human thymocytes and peripheral B lymphocytes, activated T cells, and follicular dendritic cells. The expressing positions indicate that the distribution of CD200 gene is related to immune tolerance induced. CD200 interacts with the structurally related receptor (CD200R) expressed on myeloid cells and down regulation of myeloid cells activity, regulation of cytokine network, intervention with development and differentiation of help T cell by altering secretion of cytokine and promotion immune tolerance.The DC is the most importantly and has the strongest function of APC in body.It has significance in immunoregulation.Its environment of development, subgroup and phase of maturation determine tendency and end-results of immune response. DCs have multiplicity and plasticity , can induce immune response and immune tolerance. It is thought generally that imDC express costimulation signal molecule lowly and deficient of the second-messenger of T cells immune response , makes T cells disability or low- response.So the imDC can induce immune tolerance.Regulatory T cells are an important part of maintaining self-tolerance. They mainly play a role in suppressive immuno-regulation function. Recently study indicated that CD4~+CD25~+ regulatory T cells express specificially FOXP3. In the function and development of CD4~+CD25~+ regulatory T cells that the FOXP3 is very important. Objective: To establish a cell line of immature dendrtic cell (imDC) which contains CD200 gene, the imDC which contain CD200 gene cultured with cord lymphocytes. To investigate the imDC, which have transfected by CD200 gene, induced to product more quantity of regulatory T cells. The FOXP3 molecule was detected on the surface of CD4~+CD25~+ regulatory T cells .The conclusion is that FOXP3 molecule is a marker of CD4~+CD25~+ regulatory T cells.Methods: The monocytes were isolated from human peripheral blood mononuclear cells which attached to the plastic surface. The acquired monocytes were stimulated by GM-CSF and IL-4 and were induced immature DCs. The molecule of CD83 is a marker of mDC. The plasmid of pcDNA3-CD200 was transfected into imDC with liposome, then checked the express of CD200 and CD83 on the surface of imDC after 48 hours with FCM. Cord lymphocytes cultured with imDCs for 5 days and then regulatory T cells were induced. It is detected that the percentage of CD4~+CD25~+ regulatory T cells and FOXP3 in the lymphocytes with FCM .The imDC was transfected by plasmid of pcDNA3-CD200 as experiment group, was tansfected by plasmid of pcDNA3 as negative group ; and the mutation DC as blank group. The difference of index was contrasted in the three groups.Results: The acquired mononuclear cells were stimulated by GM-CSF and IL-4 , and were induced by immature DCs. The plasmid of pcDNA3-CD200 was transfected into imDC, and after 48 hours the CD200 molecular was detected with FCM. The percentage expression of CD200 molecular and CD83 molecular are significantly enhanced than negative group and blank group. It is not different the percentage expression of CD83 molecular in experiment group and negative control,and the percentage expression of CD83 molecular in experiment group and negative control is lower than blank group. The imDC which included CD200 gene cultured with cord lymphocytes for 5 days, and then detected the percentage express of CD4~+CD25~+ regulatory T cells. The result domenstrated that the percentage of CD4~+CD25~+ regulatory T cells in experiment group is significantly higher than other two groups. FOXP3 molecular was detected in lymphocytes with FCM. The difference in statistics is not significant to contrast the percentage of FOXP3 and CD4~+CD25~+ regulatory T cells.Conclusion: It is feasible that transfect CD200 gene into imDC with liposome, and the expression is good. The imDC include CD200 gene whice can induce and product more regulary T cell.The result of analysis, FOXP3 and CD4~+CD25~+ molecule on the surface of Tr, extrapolate that CD200 gene can induce FOXP3 molecule expression on the surface of Tr.