Development of a method of thymocyte differentiation of bone marrow-enriched CD34+CD38- cells in postnatal allogeneic cultured thymic epithelia to evaluate immunodeficiency disorders.
An in vitro model of CD34+CD38- stem cell (SC) differentiation in postnatal cultured thymic epithelia fragment (CTEF) cocultures is described. Sequential phenotypic analysis of the progeny of the SC-CTEF demonstrated predominantly thymocytes and minor populations of promyelocytes, monocytes and natural killer cells. Triple-positive CD3+CD4+CD8+, double-positive CD4+CD8+, and mature single-positive CD4+ and CD8+ T cells, which were TCR alpha beta+, were identified indicating normal thymocyte maturation. In kinetic studies, mature single-positive CD4+ T cells increased from 29% of total cells at one week to 54% at four weeks of coculture. These findings demonstrate that coculture of bone marrow-derived SC and allogeneic cultured thymic epithelia in vitro results in continuous normal predominantly thymocyte differentiation. The SC-CTEF cocultures were then infected with two different strains of human immunodeficiency virus. CD4+ thymocytes were markedly decreased. However, inhibition of early thymocyte maturation steps was also suggested by the presence of increased triple-negative and CD44+CD25-CD3-thymocytes and decreased CD44+CD25+ thymocytes. This model system of thymocyte maturation will be useful in the evaluation of primary T cell immunodeficiency disorders, gene therapy of SC and pharmacological augmentation of thymic function.
Knutsen
A P
AP
Pediatric Research Institute, St. Louis University Health Sciences Center, Missouri 63110, USA.
Roodman
S T
ST
Ruiz
M E
ME
Mueller
K R
KR
Bouhasin
J D
JD
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D015703:Antigens, CD; D018952:Antigens, CD34 / Q000032:analysis*; D000943:Antigens, Differentiation / Q000032:analysis*; D001854:Bone Marrow Cells; D002454:Cell Differentiation; D004847:Epithelial Cells; D004848:Epithelium / Q000737:chemistry; D005434:Flow Cytometry; D015658:HIV Infections / Q000473:pathology; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis; D008562:Membrane Glycoproteins; D009699:N-Glycosyl Hydrolases / Q000032:analysis*; D010641:Phenotype; D015375:Receptors, Interleukin-2 / Q000032:analysis; D013950:Thymus Gland / Q000166:cytology*
D015703:Antigens, CD; D018952:Antigens, CD34; D000943:Antigens, Differentiation; D018960:Hyaluronan Receptors; D008562:Membrane Glycoproteins; D015375:Receptors, Interleukin-2; D009699:N-Glycosyl Hydrolases; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1