Expression of alternatively spliced fibronectin variants during remodeling in proliferative glomerulonephritis

Fibronectin (Fn) plays an important role in tissue remodeling during embryogenesis, wound repair, and vascular disease, and is thought to regulate cellular processes such as cell adhesion, migration, proliferation, and differentiation through specialized domains within the molecule. In addition, Fn can be alternatively spliced at three regions: extradomains EIIIA, EIIIB, and a variable segment V, potentially giving rise to functionally distinct variants of the molecule. We have previously shown a sequential expression of cellular Fn first by platelets, followed by macrophages, then mesangial cells in habu snake venom-induced proliferative glomerulonephritis (Am J Pathol 145: 585-597, 1994). These studies examined the cellular sources and glomerular localization of Fn in general but did not distinguish between the various alternatively spliced isoforms. In this study, we examine by in situ hybridization and immunohistochemistry the temporal expression and cellular sources of EIIIA, EIIIB, and V in a model of proliferation glomerulonephritis that has cell migration, proliferation, and extracellular matrix synthesis as features of tissue remodeling. Macrophages were the first cells to express Fn mRNA showing an EIIIA+, EIIIB-, and V95+ pattern beginning at 8 hours after habu snake venom injection. Migrating mesangial cells at the margins of early lesions (8 and 24 hours) did not overexpress mRNA encoding these Fn variants, but immunofluorescence microscopy revealed V95 and EIIIA protein at the margins of lesions. EIIIB was absent in lesions at this time. At 48 hours and peaking at 72 hours after habu snake venom injection, mesangial cells in central aspects of glomerular lesions expressed abundant mRNA and protein for V95 and EIIIA. EIIIB mRNA and protein was slight in the mesangium at these times. Parietal epithelial cells, particularly adjacent to glomerular lesions, also expressed abundant mRNA and protein for all three variants throughout the course of the disease, beginning at 24 hours after habu snake venom injection. Expression of mRNA and protein for all three isoforms declined by 2 weeks after habu snake venom injection. These studies show that migrating mesangial cells do not require their own synthesis of Fn and suggest that they might rely on exogenous sources of Fn, particularly V95+ and EIIIA+ forms. Commencement of enhanced expression of EIIIA and EIIIB mRNA and protein by resident glomerular cells coincided with the temporal course of cell proliferation, acquisition of alpha-smooth muscle cell actin phenotype, and matrix synthesis, suggesting that Fn isoforms have specific functions during the course of glomerular remodeling.