DEMONSTRATION OF PDGF B-CHAIN MESSENGER-RNA IN GLOMERULI IN MESANGIAL PROLIFERATIVE NEPHRITIS BY INSITU HYBRIDIZATION

We used the technique of in situ hybridization to determine if cells expressing PDGF B-chain mRNA can be detected in a model of mesangial proliferative nephritis in the rat induced with antibody directed against the Thy 1 antigen present on the mesangial cell membrane. The method involved hybridization with a digoxigenin-labeled cRNA probe for the murine PDGF B-chain followed by defection with an anti-digoxigenin-alkaline phosphatase conjugate and subsequent colorimetric reaction. In normal rats (N = 4). the majority of glomeruli (74%) were negative for PDGF B-chain mRNA, whereas 65% of glomeruli from rats with mesangial proliferative nephritis (N = 4) had segmental or diffuse staining for PDGF B-chain mRNA in a mesangial pattern. The difference. as measured using a semiquantitative scale, was significant (mean scores 0.4 +/- 0.2 vs. 1.9 +/- 0.2; scale 0 to 3+; P < 0.001). The increase in PDGF B-chain mRNA positive cells localized to areas of hypercellularity and was associated with a significant increase in cells positive for PDGF B-chain by immunostaining with a specific monoclonal antibody (0.8 +/-0.1 vs. 1.7 +/- 0.4. scale 0 to 3+, normal vs. diseased rats. P < 0.005). Complement depletion, which prevents the mesangial cell proliferation. also prevented the increase in cells expressing PDGF B-chain mRNA and protein. Thus. this method of in situ hybridization can successfully detect cells expressing PDGF mRNA in active glomerulonephritis, and may be useful for detecting cells expressing genes for other growth factors and cytokines in both human and experimental models of glomerular injury.