TNF-α promotes cerebral pericyte remodeling in vitro, via a switch from α1 to α2 integrins

Background: There is increasing evidence to suggest that pericytes play a crucial role in regulating the remodeling state of blood vessels. As cerebral pericytes are embedded within the extracellular matrix (ECM) of the vascular basal lamina, it is important to understand how individual ECM components influence pericyte remodeling behavior, and how cytokines regulate these events. Methods: The influence of different vascular ECM substrates on cerebral pericyte behavior was examined in assays of cell adhesion, migration, and proliferation. Pericyte expression of integrin receptors was examined by flow cytometry. The influence of cytokines on pericyte functions and integrin expression was also examined, and the role of specific integrins in mediating these effects was defined by function-blocking antibodies. Expression of pericyte integrins within remodeling cerebral blood vessels was analyzed using dual immunofluorescence (IF) of brain sections derived from the animal model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). Results: Fibronectin and collagen I promoted pericyte proliferation and migration, but heparan sulfate proteoglycan (HSPG) had an inhibitory influence on pericyte behavior. Flow cytometry showed that cerebral pericytes express high levels of alpha 5 integrin, and lower levels of alpha 1, alpha 2, and alpha 6 integrins. The pro-inflammatory cytokine tumor necrosis factor (TNF)-alpha strongly promoted pericyte proliferation and migration, and concomitantly induced a switch in pericyte integrins, from alpha 1 to alpha 2 integrin, the opposite to the switch seen when pericytes differentiated. Inhibition studies showed that alpha 2 integrin mediates pericyte adhesion to collagens, and significantly, function blockade of alpha 2 integrin abrogated the pro-modeling influence of TNF-alpha. Dual-IF on brain tissue with the pericyte marker NG2 showed that while alpha 1 integrin was expressed by pericytes in both stable and remodeling vessels, pericyte expression of alpha 2 integrin was strongly induced in remodeling vessels in EAE brain. Conclusions: Our results suggest a model in which ECM constituents exert an important influence on pericyte remodeling status. In this model, HSPG restricts pericyte remodeling in stable vessels, but during inflammation, TNF-a triggers a switch in pericyte integrins from alpha 1 to alpha 2, thereby stimulating pericyte proliferation and migration on collagen. These results thus define a fundamental molecular mechanism in which TNF-alpha stimulates pericyte remodeling in an a2 integrin-dependent manner.