Advanced glycation of fibronectin impairs vascular repair by endothelial progenitor cells: implications for vasodegeneration in diabetic retinopathy.
OBJECTIVE
Vascular repair by marrow-derived endothelial progenitor cells (EPCs) is impaired during diabetes, although the precise mechanism of this dysfunction remains unknown. The hypothesis for the study was that progressive basement membrane (BM) modification by advanced glycation end products (AGEs) contributes to impairment of EPC reparative function after diabetes-related endothelial injury.


METHODS
EPCs isolated from peripheral blood were characterized by immunocytochemistry and flow cytometry. EPC interactions on native or AGE-modified fibronectin (AGE-FN) were studied for attachment and spreading, whereas chemotaxis to SDF-1 was assessed with the Dunn chamber assay. In addition, photoreactive agent-treated monolayers of retinal microvascular endothelial cells (RMECs) produced circumscribed areas of apoptosis and the ability of EPCs to "endothelialize" these wounds was evaluated.


RESULTS
EPC attachment and spreading on AGE-FN was reduced compared with control cells (P < 0.05-0.01) but was significantly restored by pretreatment with Arg-Gly-Asp (RGD). Chemotaxis of EPCs was abolished on AGE-FN but was reversed by treatment with exogenous RGD. On wounded RMEC monolayers, EPCs showed clustering at the wound site, compared with untreated regions (P < 0.001); AGE-FN significantly reduced this targeting response (P < 0.05). RGD supplementation enhanced EPC incorporation in the monolayer, as determined by EPC participation in tight junction formation and restoration of transendothelial electric resistance (TEER).


CONCLUSIONS
AGE-modification of vascular substrates impairs EPC adhesion, spreading, and migration; and alteration of the RGD integrin recognition motif plays a key role in these responses. The presence of AGE adducts on BM compromises repair by EPC with implications for vasodegeneration during diabetic microvasculopathy.
Bhatwadekar
Ashay D
AD
Centre for Vision Science, Queen's University Belfast, Royal Victoria Hospital, Northern Ireland, UK.
Glenn
Josephine V
JV
Li
Gang
G
Curtis
Tim M
TM
Gardiner
Tom A
TA
Stitt
Alan W
AW
D000818:Animals; D002417:Cattle; D002448:Cell Adhesion / Q000502:physiology; D002465:Cell Movement; D054377:Chemokine CXCL12 / Q000494:pharmacology; D002633:Chemotaxis / Q000187:drug effects; D003930:Diabetic Retinopathy / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D005353:Fibronectins / Q000378:metabolism*; D005434:Flow Cytometry; D017127:Glycation End Products, Advanced / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D009842:Oligopeptides / Q000494:pharmacology; D011765:Pyruvaldehyde / Q000633:toxicity; D012171:Retinal Vessels / Q000166:cytology; D013234:Stem Cells / Q000378:metabolism*; D014945:Wound Healing
D054377:Chemokine CXCL12; D005353:Fibronectins; D017127:Glycation End Products, Advanced; D009842:Oligopeptides; D011765:Pyruvaldehyde; C047981:arginyl-glycyl-aspartic acid