IMMUNOHISTOCHEMICAL COLOCALIZATION OF THE ALPHA-SUBUNIT OF NEUTROPHILNADPH OXIDASE AND ECTO-5'-NUCLEOTIDASE IN KIDNEY AND LIVER

Immunohistochemical colocalization of the cu-subunit of neutrophil NAD PH oxidase and ecto-5'-nucleotidase in kidney and liver. In kidney and liver, fibroblasts and fibroblast-like cells, respectively, are sourc es of erythropoietin (Epo) formation, and these cells also bear a numb er of other similarities. Renal Epo expression is localized in peritub ular type I fibroblasts of the cortical labyrinth, and in the liver, a part from parenchymal cells, transcription is found in Ito cells. Both the renal peritubular cells and Ito cells contain ecto-5'-nucleotidas e (5'NT). It had been suggested that 5'NT is involved in the oxygen se nsing mechanism via a hydrolysis of AMP to adenosine, which in turn ma y stimulate EPO synthesis. However, the molecular mechanism of the cel lular response to hypoxia is currently not well understood. Based on t he notion that a heme protein probably acts as the oxygen sensor, it h as recently been proposed that a b-type cytochrome as part of the neut rophil NADPH oxidase may influence intracellular superoxide levels dep ending on local oxygen tension. Superoxide levels were otherwise shown to determine the EPO production in hepatoma cell lines. By double imm unofluorescence labelling the ol-subunit of cytochrome b(558) (alpha-S U) and 5'NT were simultaneously localized in rat kidney and liver, and in the kidney Epo mRNA and alpha-SU were double-labelled. Positive si gnal for alpha-SU was found in the majority of renal peritubular fibro blasts in the cortex and outer medulla, and in Ito cells. In both orga ns, the cells that coexpress 5'NT and Epo mRNA also contain an immunor eactivity for alpha-SU. In these cells, cytochrome b(558) as part of a n NADPH oxidase may be involved in a presumptive oxygen sensing mechan ism using H2O2 as a possible second messenger for EPO gene regulation.