Fibronectin accumulation in glomerulosclerotic lesions: self-assembly sites and the heparin II binding domain.
BACKGROUND
Glomerulosclerosis is a severe complication of many immunologically-mediated kidney diseases, eventually resulting in loss of renal function. In chronic graft-versus-host disease (GvHD) in mice, a model for human lupus nephritis, the end-stage sclerotic lesions were previously shown to contain large amounts of fibronectin (FN). This study investigated a domain-specific accumulation process of circulating plasma FN (pFN) in sclerotic lesions.


METHODS
GvHD mice were injected with FITC-conjugated pFN or pFN-fragments, with or without heparin pre-incubation. pFN fragments were generated by digestion of pFN by cathepsin D, after which the fragments were separated on a heparin affinity column. Thus, two batches of fragments were obtained with either low or high affinity for heparin.


RESULTS
FN accumulation was accompanied by an up-regulated expression of integrin alpha5beta1, the FN receptor, in the periphery of sclerotic lesions. pFN-FITC injected into GvHD mice was trapped in sclerotic glomeruli within 24 hours. Both heparin and non-anti-coagulant heparin blocked the accumulation of pFN-FITC, indicating that the protective effect of heparin in the trapping of FN is independent of its anticoagulant properties, and probably results from preventing direct binding of FN in the sclerotic lesions. To investigate whether FN binds in the glomerulus via the heparin-binding regions, pFN fragments were generated and injected into GvHD mice. Whereas the fraction with high affinity for heparin did not accumulate in the sclerotic glomeruli, the fraction with low affinity for heparin did. Partial sequencing of the isolated peptides showed that in the glomerulus fibronectin does not bind via the heparin II binding region.


CONCLUSIONS
We hypothesize that the protective effect of heparin treatment may be the result of steric hindrance of the specific binding sites, that is, the I1-5 and/or III1 self-assembly sites of FN.
van Vliet
Anita I
AI
Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands. A.I.van_Vliet@LUMC.nl
van Alderwegen
Isolde E
IE
Baelde
Hans J
HJ
de Heer
Emile
E
Bruijn
Jan A
JA
D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology*; D001665:Binding Sites / Q000187:drug effects; D005109:Extracellular Matrix / Q000378:metabolism; D005260:Female; D005353:Fibronectins / Q000737:chemistry / Q000378:metabolism*; D005923:Glomerulosclerosis, Focal Segmental / Q000188:drug therapy / Q000378:metabolism*; D006086:Graft vs Host Disease / Q000188:drug therapy / Q000378:metabolism*; D006493:Heparin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D010446:Peptide Fragments / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology
D000925:Anticoagulants; D005353:Fibronectins; D010446:Peptide Fragments; C044153:heparin, N-desulfated; D006493:Heparin