Podocyte injury or podocyte loss in the renal glomerulus has been proposed as the crucial mechanism in the development of
focal segmental glomerulosclerosis. However, it is poorly understood how podocytes respond to injury. In this study, glomerular expression of
connexin43 (
Cx43)
gap junction protein was examined at both
protein and transcript levels in an experimental model of podocyte injury,
puromycin aminonucleoside (PAN)
nephrosis. A striking increase in the number of immunoreactive dots with anti-Cx43 antibody was demonstrated along the glomerular capillary wall in the early to nephrotic stage of PAN
nephrosis. The conspicuous change was not detected in the other areas including the mesangium and Bowman's capsule. Immunoelectron microscopy showed that the immunogold particles for
Cx43 along the capillary wall were localized predominantly at the cell-cell contact sites of podocytes. Consistently, Western blotting and
ribonuclease protection assay revealed a distinct increase of
Cx43 protein, phosphorylation, and transcript in glomeruli during PAN
nephrosis. The changes were detected by 6 hours after PAN injection. These findings indicate that the increase of
Cx43 expression is one of the earliest responses that have ever been reported in podocyte injury. To show the presence of functional gap junctional intercellular communication (GJIC) in podocytes, GJIC was assessed in podocytes in the primary culture by transfer of
fluorescent dye,
Lucifer yellow, after a single-cell microinjection. Diffusion of the
dye into adjacent cells was observed frequently in the cultured podocytes, but scarcely in cultured parietal epithelial cells of Bowman's capsule, which was compatible with their
Cx43 staining. Thus, it is concluded that Cx43-mediated GJIC is present between podocytes, suggesting that podocytes may respond to injury as an integrated epithelium on a glomerulus rather than individually as a separate cell.