Method for producing dendritic cells
View Patent ↗Disclosed are embryonic stem cell-derived dendritic cells, genetically modified immature dendritic cells capable of maturation, as well as methods for the production of such cells. In one embodiment, the cells made be produced by a method comprising the steps of providing a population of embryonic stem cells; culturing the embryonic stem cells in the presence of a cytokine or combination of cytokines which brings about differentiation of the embryonic stem cells into dendritic cells; and recovering the dendritic cells from the culture. In a further embodiment, the cells may be genetically modified.
1. A process for producing a culture of mouse dendritic cells, comprising:
culturing mouse embryonic stem cells to form embryoid bodies;
culturing said embryoid bodies in the presence of a composition comprising IL-3; and
recovering mouse dendritic cells from said culture, wherein prior to said culturing to form embryoid bodies said mouse embryonic stem cells are maintained in an undifferentiated state in the presence of leukemia inhibitory factor (LIF).
2. The process according to claim 1 , wherein the composition further comprises GM-CSF.
3. The process according to claim 1 , wherein said mouse embryonic stem cells are genetically modified.
4. The process of claim 1 , wherein the mouse dendritic cells express one or more heterologous gene(s).
5. The process of claim 4 , wherein the heterologous gene(s) encodes a cell surface protein.
6. The process of claim 4 , wherein the heterologous gene(s) encodes an antigen.
7. The process of claim 1 , wherein the mouse dendritic cells co-express two or more heterologous genes.
8. The process of claim 1 , wherein one or more endogenous gene(s) has been inactivated in the mouse embryonic stem cells.
9. The process of claim 3 , wherein said mouse embryonic stem cells are transfected with a gene, which gene is expressed in the mouse dendritic cells.
10. The process of claim 9 , wherein the gene is under the control of a promoter which is preferentially active in dendritic cells.
11. The process of claim 9 , wherein the gene is a reporter gene which expresses a detectable gene product in the mouse dendritic cells.
12. The process of claim 11 , wherein the detectable gene product is a fluorescent protein.
13. The process of claim 12 , wherein the fluorescent protein is a green fluorescent protein (GFP).
14. The process of claim 1 , wherein the mouse embryonic stem cells are from a CBA/Ca or a C57B1/6 cell line.
15. The process of claim 1 , wherein the mouse embryonic stem cells are from the ESF116 cell line.
16. A process for producing a culture of mouse dendritic cells, comprising:
culturing mouse embryonic stem cells to form embryoid bodies;
culturing said embryoid bodies in the presence of a composition comprising IL-3; and
recovering mouse dendritic cells from said culture, wherein the mouse embryonic stem cells are transfected with an expression vector comprising;
a) a multiple cloning site designed to facilitate production of fusion protein between a heterologous protein and an enhanced green fluorescent protein, and
b) a promoter operably linked to facilitate expression of the fusion protein.
17. The process of claim 16 , wherein the promoter is an immediate-early promoter of human cytomegalovirus (CMV).
18. The process of claim 16 , wherein the promoter is a CD11c promoter.