Elevated glucose and fatty acid levels impair substance P-induced dermal microvascular endothelial cell migration and proliferation in an agarose gel model system.
Substance P (SP), a sensory nerve derived neuropeptide, has been implicated in wound repair. Our hypothesis was that oxidative effects of elevated glucose and fatty acid levels as seen with diabetes mellitus inhibit SP-mediated endothelial cell directional migration and proliferation. Using a 2% agarose gel, immortalized human microvascular endothelial cells (HMEC-1) were plated into a 1.5-mm well, and agonist (SP; 10(-4) mol/L) was loaded into a 3-mm well; controls included NaCl, albumin (bovine serum albumin), and vascular endothelial cell growth factor. The SP receptor antagonist spantide 1 was used to confirm SP specificity. Elevated glucose (40 mmol/L) and fatty acids (40 micromol/L) were added to the medium with and without vitamin E and vitamin C treatment to determine whether endothelial cell responses to SP were altered by metabolic perturbations and whether they could be recovered with antioxidant treatment. Using computer-assisted image analysis, migration distance was measured. Cells were counted using a hemocytometer. Human microvascular endothelial cell 1 migration toward the SP exceeded NaCl or bovine serum albumin; vascular endothelial cell growth factor had similar effects. The SP receptor antagonist, spantide, inhibited SP-induced HMEC-1 migration. Substance P treatment was associated with increased cell number. Ki-67 staining was increased in SP-treated cells compared with controls. Elevated glucose and fatty acid levels diminished cell migration toward SP. The antioxidants vitamins C and E significantly improved proliferation but only marginally improved migration. Our data suggest that glucose and fatty acids perturb SP-induced HMEC-1 migration and proliferation in an agarose gel migration model.
Wang
Qiang
Q
Department of Surgery, University of Washington, Seattle, Washington 98104, USA.
Muffley
Lara A
LA
Hall
Kyla
K
Chase
Marie
M
Gibran
Nicole S
NS
D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology*; D005227:Fatty Acids / Q000494:pharmacology*; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D064729:Neurokinin-1 Receptor Antagonists; D012685:Sepharose / Q000737:chemistry; D013373:Substance P / Q000031:analogs & derivatives / Q000494:pharmacology
D005227:Fatty Acids; D064729:Neurokinin-1 Receptor Antagonists; D013373:Substance P; D012685:Sepharose; C042867:spantide; D005947:Glucose