Second harmonic microscopy to quantify renal interstitial fibrosis and arterial remodeling

Interstitial fibrosis is a powerful pejorative predictor of pro- gression of nephropathies in a variety of chronic renal diseases. It is characterized by the depletion of kidney cells and their replacement by extracellular matrix, in particular, type-I fibrillar collagen, a protein scarce in normal interstitium. However, assessment of fibrosis remains a challenge in research and clinical pathology. We develop a novel methodology based on second harmonic generation SHG micros- copy, and we image collagen fibers in human and mouse unstained kidneys. We take into account the variability in renal shape, and we develop automated image processing for quantitative scoring of thick murine tissues. This approach allows quantitative 3-D imaging of in- terstitial fibrosis and arterial remodeling with high accuracy. More- over, SHG microscopy helps to raise pathophysiological questions. First, imaging of a large volume within a mouse kidney shows that progression of fibrosis is a heterogeneous process throughout the dif- ferent renal compartments. Second, SHG from fibrillar collagens does not overlap with the glomerular tuft, despite patent clinical and ex- perimental glomerulosclerosis. Since glomerulosclerosis involves SHG-silent nonfibrillar collagens, our work supports pathophysiologi- cal differences between interstitial fibrosis and glomerulosclerosis, a clearly nonfibrotic process. © 2008 Society of Photo-Optical Instrumentation Engi-