Method of producing erythrocytes without feeder cells
View Patent ↗Provided herein are methods of producing erythrocytes from hematopoietic cells, particularly hematopoietic cells from placental perforate in combination with hematopoietic cells from umbilical cord blood, wherein the method results in accelerated expansion and differentiation of the hematopoietic cells to more efficiently produce administrate erythrocytes. Further provided herein is a bioreactor in which hematopoietic cell expansion and differentiation takes place.
1. A method of producing erythrocytes, comprising:
a) expanding a first population of hematopoietic cells in a first medium in the absence of feeder cells to obtain an expanded population of hematopoietic cells, wherein the first population of hematopoietic cells has an initial cell density of 5×10 4 cells/mL or less and the first medium comprises Epo at a concentration of 1 IU/mL or less and SCF at a concentration of 100 ng/mL or more;
b) differentiating a plurality of hematopoietic cells within said expanded population of hematopoietic cells into erythrocytes by contacting the expanded population of hematopoietic cells with a second medium, wherein the expanded population of hematopoietic cells has an initial cell density greater than 2.75×10 5 cells/mL and the second medium comprises Epo at a concentration of 3 IU/mL or greater and SCF at a concentration of 50 ng/mL or less; and
c) isolating said erythrocytes from said second medium,
wherein said SCF and Ep are not comprised within an undefined component of said first or said second medium.
2. The method of claim 1 , wherein the first medium further comprises pomalidomide.
3. The method of claim 1 , wherein the first medium does not comprise one or more of Flt-3L, IL-11, thrombopoietin (Tpo), homeobox-B4 (HoxB4), or methylcellulose.
4. The method of claim 1 , wherein the second medium does not comprise one or more of Flt-3L, IL-11, thrombopoietin (Tpo), homeobox-B4 (HoxB4), or methylcellulose.
5. The method of claim 1 , wherein the first population of hematopoietic cells has an initial cell density between 2×10 4 and 5×10 4 cells/mL.
6. The method of claim 1 , wherein the first population of hematopoietic cells comprises hematopoietic cells from a plurality of individuals.
7. The method of claim 1 , wherein the first population of hematopoietic cells comprises hematopoietic cells from placenta.
8. The method of claim 1 , wherein the first population of hematopoietic cells comprise cells from two or more tissue sources.
9. The method of claim 1 , wherein the first medium further comprises IGF-1 at a concentration of about 1 to about 1000 ng/mL.
10. The method of claim 1 , wherein the first medium further comprises lipids at a concentration of about 1 to about 1000 μg/mL.