ABSTRACT
The Wilms' tumor protein Wt1 is required for embryonic development and has been implicated in hematologic disorders. Since Wt1 deficiency may compromise the proliferation and differentiation of erythroid progenitor cells, we analyzed the possible role of the transcriptionally active Wt1 isoform, Wt1(-KTS), in regulating the expression of the erythropoietin receptor (EpoR). Wt1 and EpoR were coexpressed in CD117(+) hematopoietic progenitor cells and in several hematopoietic cell lines. CD117(+) cells of Wt1-deficient murine embryos (Wt1(-/-)) exhibited a significantly lower proliferation response to recombinant erythropoietin than CD117(+) cells of heterozygous (Wt1(+/-)) and wild-type littermates (Wt1(+/+)). EpoR expression was significantly diminished in hematopoietic progenitors (CD117(+)) that lacked Wt1, and the erythroid colony-forming capacity was reduced by more than 50% in fetal liver cells of Wt1-deficient embryonic mice. Wt1(-KTS) significantly increased endogenous EpoR transcripts in transfected cells. The proximal EpoR promoter of human and mouse was stimulated more than 10-fold by Wt1(-KTS) in transiently cotransfeced K562 erythroleukemia cells. A responsible cis-element, which is highly conserved in the EpoR promoter of human and mouse, was identified by mutation analysis, electrophoretic mobility shift assay, and chromatin immunoprecipitation assay. In conclusion, activation of the EpoR gene by Wt1 may represent an important mechanism in normal hematopoiesis.
Subject(s)
Genes, Wilms Tumor , Hematopoietic Stem Cells/metabolism , Receptors, Erythropoietin/genetics , Animals , Base Sequence , Binding Sites/genetics , Cell Line , DNA/genetics , DNA/metabolism , DNA Primers/genetics , Embryonic Stem Cells/metabolism , Erythropoiesis/genetics , Erythropoietin/pharmacology , Genes, Reporter , Hematopoiesis/drug effects , Hematopoiesis/genetics , Hematopoietic Stem Cells/cytology , Hematopoietic Stem Cells/drug effects , Heterozygote , Humans , Mice , Mice, Knockout , Mutagenesis, Site-Directed , Promoter Regions, Genetic , Proto-Oncogene Proteins c-kit/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Erythropoietin/deficiency , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Transcriptional Activation , Transfection , WT1 Proteins/deficiency , WT1 Proteins/genetics , WT1 Proteins/metabolismABSTRACT
Three years ago, the Working Party on Facial Transplantation concluded that until there was more information available about risks any potential patient would be exposed to, it would be unwise to proceed with transplantation of the human face. Over the last three years, there has been a deepening understanding of the potential psychological problems of facial transplantation as well as a very considerable debate on the ethical aspects of the procedure. Further data on experimental work in animal models of facial transplantation as well as medium-term follow-up data from 24 hand and forearm transplants in 18 patients has now become available. Furthermore, a partial facial transplantation has been performed in France and a second one in China. In this second edition of the report, the technical, immunological, psychological, and ethical issues are discussed again in the light of this developing knowledge. In particular, there has been a major expansion of the sections on the psychological and societal issues, as well as the ethical and legal problems of facial transplantation. The working party still has considerable reservations about facial transplantation. Although it accepts that on balance the risks cannot be precisely quantified, they remain substantial. Therefore, if patients are allowed to make an informed choice to proceed, they must be very carefully selected and protected in the process, along with the families of both the donors and the recipients. To achieve this, the working party insists that 15 minimum requirements, described at the end of this report, must be fulfilled before it would be appropriate for a research ethics committee/institutional review board to approve of a proposal to undertake facial transplantation.
