Angiogenic sprouting from the aortic vascular wall is impaired in the BB rat model of autoimmune diabetes.
BACKGROUND
Diabetes is associated with impaired neovascularization leading to reduced revascularization of ischemic tissue and impaired wound healing. Endothelial progenitor cells in diabetes were previously shown to be numerically reduced and functionally impaired. We hypothesize that diabetes also has a long-term effect on angiogenic cells residing in the vessel wall. To test this hypothesis, angiogenic sprout formation from ex vitro cultured aortic rings isolated from diabetic and non-diabetic BioBreeding (BB) rats was assessed.


METHODS
Diabetes prone BB (BBDP) rats spontaneously develop autoimmune diabetes were suboptimally treated with insulin by subcutaneous implantation of slow-release insulin-pellets. Neonatally thymectomized BBDP rats, pre-diabetic BBDP rats and diabetes resistant BBDR rats served as non-diabetic controls. After follow-up thoracic aortas were harvested and cultured in vitro in Matrigel to induce sprout formation. Sprout length was quantified after 4, 7, 10 and 14 days of culture. The total number of sprout-derived cells was measured and in vitro proliferative capacity of sprout cells was quantified. Finally, expression of Flk-1, CD31 and smooth muscle alpha-actin on sprout cells was determined.


RESULTS
Mean blood glucose levels in diabetics were significantly elevated compared with non-diabetics. Both long-term and short-term diabetes significantly reduced sprout formation (p<0.05 vs. non-diabetics). Reduced sprout length in diabetics was reflected by significantly reduced numbers of sprout cells that could be isolated (p<0.05 vs. non-diabetics). Isolated sprout cells from diabetics revealed significantly reduced proliferative capacity after in vitro culture (p<0.05 vs. non-diabetics). Immunofluorescent staining indicated an endothelial phenotype of both freshly isolated and in vitro cultured sprout cells as indicated by CD31 and Flk-1 expression and absence of smooth muscle alpha-actin expression.


CONCLUSIONS
Diabetes in BB rats impairs angiogenic sprouting from cells residing in the vascular wall, independent of effects on circulating cells or circulating angiogenic/anti-angiogenic factors. The angiogenic impairment of diabetic sprout cells is, to some extent, imprinted upon the cells.
Onuta
Geanina
G
Department of Cell Biology - Immunology section, University Medical Center Groningen, University of Groningen, The Netherlands.
Westerweel
Peter E
PE
Zandvoort
André
A
van Riezen
Manon
M
Rozing
Jan
J
Hillebrands
Jan-Luuk
JL
Verhaar
Marianne C
MC
D000818:Animals; D001013:Aorta, Thoracic / Q000473:pathology*; D001327:Autoimmune Diseases / Q000276:immunology / Q000473:pathology*; D015415:Biomarkers / Q000378:metabolism; D002196:Capillaries / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism; D002452:Cell Count; D049109:Cell Proliferation; D003920:Diabetes Mellitus / Q000276:immunology / Q000473:pathology*; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000166:cytology / Q000254:growth & development / Q000378:metabolism; D005260:Female; D008297:Male; D018919:Neovascularization, Physiologic; D009924:Organ Culture Techniques; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D051381:Rats; D011913:Rats, Inbred BB; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
D015415:Biomarkers; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D040301:Vascular Endothelial Growth Factor Receptor-2