Title: Immunohistochemical Localization of Erythropoietin (Epo) and Erythropoietin Receptor (EpoR) in Developing Human Brain ♦ 267
Abstract: We have previously demonstrated the presence of Epo and EpoR mRNA and protein in human fetal spinal cords, and in primary cell cultures of neurons and glial cells derived from fetal spinal cords 5 to 9 weeks postconception. We now describe immunohistochemical localization of both Epo and EpoR in developing human brain. We used a monoclonal antibody raised against the first 26 amino acids of human Epo (Genzyme), and a polyclonal antibody specific to amino acids 221 of the human EpoR (Santa Cruz). Fetal kidney and liver served as positive controls, and absence of the primary antibody as one type of negative control. Antibody specificity for EpoR was further established by elimination of immunoreaction following competative blocking with the specific epitope against which the antibody was raised. CNS tissue obtained at autopsy from adult and fetal brains (10 to 38 wks gestation) were studied. Epo and EpoR reactivity were present at all ages tested, although the patterns of staining varied with developmental stage. Epo: In early fetal life, Epo reactivity was most prominent in the germinal matrix zone (GMZ) and in the subpial granular layer. At later developmental stages neuronal cytoplasmic staining became more marked, particularly in the thalamus, hippocampus, lateral geniculate and cortex. Astrocytes were also positive and displayed a sharp gradient between gray and white matter glia, with gray matter staining positively. EpoR: At 10 wks gestation, EpoR reactivity was moderate in the periventricular zones and subpial regions. At 20 wks, strong immunoreactivity was identified in the periventricular GMZ, in astrocytes originating from the GMZ and in the subpial granular layer, while at 38 wks, EpoR was strongly reactive in astrocytes, particularly in the periventricular and subpial regions. Neurons in both the cortex and deep nuclei were at most weakly reactive. The ependyma and choroid plexus epithelium were strongly immunoreactive at all stages for both Epo and EpoR. We speculate that the function of Epo is not limited to hematopoiesis, and that this important cytokine has previously unrecognized functions within the CNS.