3D-confocal structural analysis of bone marrow-derived renal tubular cells during renal ischemia/reperfusion injury.

Bone marrow cells (BMC) have been shown to migrate into injured sites for parenchymal repair. However, the extent of BMC involvement is controversial. To determine whether and to what extent BMC contribute to renal parenchymal repair, we employed three-dimensional (3D) fluorescent confocal microscopy/video in renal warm and cold ischemia/reperfusion (I/R) injury using enhanced green fluorescent protein transgenic rats and their radiation chimeras. After induction of renal warm I/R injury in chimeras, BM-derived renal tubular cells were found in 2D microscopy as isolated single cells or clusters of 2-3 cells. Likewise, cold I/R injury resulted in host-derived tubular cells with frequencies approximately 0.2%. However, stringent confocal microscopic analysis and 3D image construction revealed that BM-derived tubules identified in 2D images were frequently artifacts of overlapping cells separately stained with different markers. The actual frequency in 3D analysis was approximately one-fourth of that seen in 2D analysis. 3D confocal imaging precisely detected BM-derived tubular epithelial cells and could be useful to study BMC contribution to tissue repair.