Effects of Leukemia Inhibitory Factor (LIF) on Gene Transfer Efficiency into Murine Hematolymphoid Progenitors

Abstract

We have investigated the effects of the cytokine leukemia inhibitory factor (LIF) on recovery and retroviral infection of murine hematopoietic stem cells maintained in short-term culture. Up to a two-fold increase in CFU-S13 recovery was observed, from 9.7 x 10(-5) cells in untreated controls to 17.6 x 10(-5) cells when 10U/ml LIF is added to the culture medium. Intermediate concentrations of LIF (.1U/ml and 1U/ml) were not significantly different from the control. Histological analysis of spleen colonies harvested thirteen days posttransplant demonstrated that LIF does not cause a detectable alteration in the differentiative potential of CFU-S13. The efficiency of retroviral-vector infection in CFU-S13 is also improved, from 15% (24/158) in untreated controls to 91% (116/127) at a LIF concentration of 10U/ml. LIF concentrations of .1U/ml and 1U/ml increased infection efficiency to 35% (14/40) and 71% (37/51), respectively. Analysis of proviral insertion sites in spleen colonies indicated that some CFU-S13 precursors were infected in the LIF-treated marrows, but no identical pairs were detected in the controls. Finally, long-term expression of provirally-encoded human adenosine deaminase (hADA) was measured in hematopoietic tissues of bone marrow transplant recipients six months posttransplant. In all tissues analyzed (spleen, thymus, bone marrow, splenic B cells, peritoneal macrophages, and blood) differentiated progeny of LIF-treated marrows had higher levels of hADA than untreated controls. Tenfold increases in levels of hADA are detected in some tissues, but levels were variable. These experiments demonstrate that LIF directly or indirectly enhances retroviral infection efficiency of hematopoietic stem cells, and might be used to improved existing gene transfer protocols.