The retroviral transduction of HOXC4 into human CD34(+) cells induces an in vitro expansion of clonogenic and early progenitors.

ABSTRACT

OBJECTIVE: +HOX genes are expressed in the hematopoietic system and increasing data point to their involvement in the control of proliferation and/or differentiation. Genes belonging to the C cluster are preferentially expressed in developing and differentiated lymphoid lineages. However, recent studies demonstrated, by RT-PCR, that the HOXC4 gene is also actively transcribed in the most undifferentiated hematopoietic cells (CD34(+)38(low)) and in more mature myeloid and erythroid progenitors. We evaluated the expression of HOXC4 protein on human CD34(+) cells and the in vitro effect of its overexpression on proliferation and differentiation. MATERIALS AND METHODS: We assessed the expression of HOXC4 on human CD34(+) cells using a polyclonal antibody raised against the C-terminal portion of the protein expressed using the baculovirus system. Overexpression of HOXC4 in human CD34(+) cells was obtained by retroviral gene transfer; its effect on clonogenic (CFU-GM, BFU-E, and CFU-GEMM) and early progenitors (LTC-IC) was evaluated. RESULTS: The HOXC4 protein is indeed expressed in human CD34(+) cells, and its overexpression in human CD34(+) cells increases the proliferation potential of clonogenic and early progenitors. CFU-GM showed a median threefold expansion (range 1.1-19.4; p < 0.002) compared with control transduced with the vector alone. The increment of BFU-E was higher (median ninefold, range 2.5-35; p < 0. 0009) and erythroid colonies presented a larger size with normal morphology. An even more marked effect was observed on LTC-IC (median 13, onefold; range 4.1-102.1, p < 0.0001). CONCLUSION: We demonstrate that HOXC4 is expressed in CD34(+) cells and that its overexpression induces an in vitro expansion of committed as well as very early hematopoietic progenitors. The most striking effect was obtained on LTC-IC with an expansion of 13.1-fold. The enforced expression of HOXC4 induced a significant increase (p < 0.009) in the number of erythroid colonies compared with CFU-GM, although without perturbing, at least in vitro, the maturation program of the cells. On the other hand, the effect of the gene overexpression did not induce any skewing in the colony types derived from the myeloid lineage.