Hypoxia-induced phenotypic switch of fibroblasts to myofibroblasts through a matrix metalloproteinase 2/tissue inhibitor of metalloproteinase-mediated pathway: implications for venous neointimal hyperplasia in hemodialysis access.
OBJECTIVE
Hemodialysis grafts fail because of venous neointimal hyperplasia formation caused by adventitial fibroblasts that have become myofibroblasts (ie, alpha-smooth muscle actin [SMA]-positive cells) and migrate to the neointima. There is increased expression of hypoxia-inducible factor (HIF)-1alpha in venous neointimal hyperplasia formation in experimental animal models and clinical samples. It was hypothesized that, under hypoxic stimulus (ie, HIF-1alpha), fibroblasts will convert to myofibroblasts through a matrix metalloproteinase (MMP)-2-mediated pathway.


METHODS
Murine AKR-2B fibroblasts were made hypoxic or normoxic for 24, 48, and 72 hours. Protein expression for HIF-1alpha, alpha-SMA, MMP-2, MMP-9, tissue inhibitor of metalloproteinase (TIMP)-1, and TIMP-2 was performed to determine the kinetic changes of these proteins. Immunostaining for alpha-SMA, collagen, and fibronectin was performed.


RESULTS
At all time points, there was significantly increased expression of HIF-1alpha in the hypoxic fibroblasts compared with normoxic fibroblasts (P < .05). There was significantly increased expression of alpha-SMA at all time points, which peaked by 48 hours in hypoxic fibroblasts compared with normoxic fibroblasts (P < .05). There was a significant increase in the expression of active MMP-2 by 48-72 hours and a significant increase in TIMP-1 by 48-72 hours by hypoxic fibroblasts (P < .05). By 72 hours, there was significant increase in TIMP-2 expression (P < .05). Immunohistochemical analysis demonstrated increased expression of alpha-SMA, collagen, and fibronectin as the duration of hypoxia increased.


CONCLUSIONS
Under hypoxic conditions, fibroblasts will convert to myofibroblasts through an MMP-2-mediated pathway, which may provide insight into the mechanism of venous neointimal hyperplasia.
Misra
Sanjay
S
Department of Radiology, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA. misra.sanjay@mayo.edu
Fu
Alex A
AA
Misra
Khamal D
KD
Shergill
Uday M
UM
Leof
Edward B
EB
Mukhopadhyay
Debabrata
D
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D015687:Cell Hypoxia / Q000502:physiology; D002460:Cell Line; D005347:Fibroblasts / Q000166:cytology* / Q000502:physiology*; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D051379:Mice; D010100:Oxygen / Q000378:metabolism*; D006435:Renal Dialysis / Q000379:methods; D015398:Signal Transduction / Q000502:physiology*; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism*; D017539:Tunica Intima / Q000503:physiopathology
D019715:Tissue Inhibitor of Metalloproteinase-1; D020778:Matrix Metalloproteinase 2; D010100:Oxygen