title: Genetically Engineered In Vitro Erythropoiesis.
abstract: BACKGROUND
  Engineered blood has the greatest potential to combat a predicted future shortfall in the US blood supply for transfusion treatments. Engineered blood produced from hematopoietic stem cell (HSC) derived red blood cells in a laboratory is possible, but critical barriers exist to the production of clinically relevant quantities of red blood cells required to create a unit of blood. Erythroblasts have a finite expansion capacity and there are many negative regulatory mechanisms that inhibit in vitro erythropoiesis. In order to overcome these barriers and enable mass production, the expansion capacity of erythroblasts in culture will need to be exponentially improved over the current state of art. This work focused on the hypothesis that genetic engineering of HSC derived erythroblasts can overcome these obstacles.
  
  
  OBJECTIVE
  The objective of this research effort was to improve in vitro erythropoiesis efficiency from human adult stem cell derived erythroblasts utilizing genetic engineering. The ultimate goal is to enable the mass production of engineered blood.
  
  
  METHODS
  HSCs were isolated from blood samples and cultured in a liquid media containing growth factors. Cells were transfected using a Piggybac plasmid transposon.
  
  
  RESULTS
  Cells transfected with SPI-1 continued to proliferate in a liquid culture media. Fluorescence-activated cell sorting (FACS) analysis on culture day 45 revealed a single population of CD71⁺CD117⁺ proerythroblast cells. The results of this study suggest that genetically modified erythroblasts could be immortalized in vitro by way of a system modeling murine erythroleukemia.
  
  
  CONCLUSIONS
  Genetic modification can increase erythroblast expansion capacity and potentially enable mass production of red blood cells.
authors.affiliation: Department of Basic Science Research, Cellologi, LLC, California, USA.|Santa Barbara Cottage Hospital, Santa Barbara, California, USA.
authors.forename: Cristopher
authors.initials: C
authors.lastname: Geiler
authors.affiliation: Department of Basic Science Research, Cellologi, LLC, California, USA.
authors.forename: Inez
authors.initials: I
authors.lastname: Andrade
authors.affiliation: Department of Basic Science Research, Cellologi, LLC, California, USA.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Clayton
authors.affiliation: Department of Basic Science Research, Cellologi, LLC, California, USA.|Santa Barbara Cottage Hospital, Santa Barbara, California, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Greenwald
keywords: Adult Stem Cells; Erythroblasts; Erythroleukemia; Erythropoiesis; Hematopoietic Stem Cells