Differential TGF-{beta} signaling in retinal vascular cells: a role in diabetic retinopathy?
Purpose. An early hallmark of preclinical diabetic retinopathy is thickening of the capillary basal lamina (BL). TGF-beta, a multipotent cytokine acting through its receptors ALK5 and -1, has been postulated to be involved in this phenomenon. In light of this possible role, TGF-beta signaling and its downstream molecular effects were characterized in cultured vascular endothelial cells and pericytes of the retina. Methods. Bovine retinal endothelial cells and pericytes were stimulated with TGF-beta1 in the presence or absence of SD-208, a specific inhibitor of the TGF-beta type I receptor ALK5, or ALK5 small interfering (si)RNA. TGF-beta-signaling pathways were characterized by analysis of phosphorylated Smad2 or -1/5/8 proteins and TGF-beta target genes (PAI-1, fibronectin, CTGF, Smad7, and Id1) and protein (fibronectin). Results. ALK5 was expressed in both cell types, whereas ALK1 was exclusively expressed in endothelial cells. In endothelial cells, TGF-beta induced Smad2 phosphorylation at high concentrations, which was efficiently blocked by ALK5 inhibition. In contrast, in pericytes, Smad2 phosphorylation was rapidly induced at low concentrations of TGF-beta. The ALK1-Smad1/5/8 pathway was activated by TGF-beta in endothelial cells only. TGF-beta caused ALK5-mediated upregulation of PAI-1, Smad7, and fibronectin and in pericytes at lower TGF-beta concentrations than in endothelial cells. CTGF mRNA expression was induced only in pericytes. Fibronectin protein was confirmed to be regulated by TGF-beta in both cell types. Conclusions. TGF-beta signaling in retinal endothelial cells and pericytes show that these cells, and in particular the pericytes, have the essential characteristics to allow for a role of TGF-beta in BL thickening in preclinical diabetic retinopathy.
Van Geest
Rob J
RJ
Ocular Angiogenesis Group, Departments of Ophthalmology and Cell Biology and Histology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
Klaassen
Ingeborg
I
Vogels
Ilse M C
IM
Van Noorden
Cornelis J F
CJ
Schlingemann
Reinier O
RO
D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D002478:Cells, Cultured; D003930:Diabetic Retinopathy / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D020286:Pericytes / Q000378:metabolism*; D010766:Phosphorylation; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics; D011621:Pteridines / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics; D012171:Retinal Vessels / Q000166:cytology*; D015398:Signal Transduction / Q000502:physiology*; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism; D051906:Smad7 Protein / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism*
D005353:Fibronectins; D017395:Plasminogen Activator Inhibitor 1; D011621:Pteridines; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D018125:Receptors, Transforming Growth Factor beta; C511004:SD-208; D051899:Smad2 Protein; D051906:Smad7 Protein; D016212:Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I