Genistein prevents hyperglycemia-induced monocyte adhesion to human aortic endothelial cells through preservation of the cAMP signaling pathway and ameliorates vascular inflammation in obese diabetic mice

Abstract

Hyperglycemia-induced vascular inflammation resulting in the enhanced monocyte-endothelial cell (EC) interaction is the key event in the pathogenesis of atherosclerosis in diabetes. Here, we investigated the effect of isoflavone genistein on hyperglycemia-stimulated vascular inflammation. Human aortic EC (HAEC) were pretreated with genistein before the addition of high glucose (HG; 25 mmol/L) for 48 h. Genistein at a physiological concentration (0.1 μmol/L) significantly inhibited HG-induced adhesion of monocytes to HAEC and suppressed endothelial production of monocyte chemotactic protein-1 (MCP-1) and IL-8. Inhibition of adenylate cyclase or protein kinase A (PKA) significantly attenuated the antiadhesion effect of genistein. Consistently, genistein improved HG-impaired intracellular cAMP production and PKA activity in HAEC. Six-week-old diabetic db/db mice were untreated (db/db) or treated with a diet containing 1 g genistein/kg diet (db/db+G) for 8 wk. Their nondiabetic db/+ mice were used as normal controls. Circulating concentrations of MCP-1/JE and KC were significantly greater, whereas IL-10 concentrations were lower in db/db mice than those in normal mice. Dietary supplementation of genistein did not normalize but significantly suppressed the elevated serum concentrations of MCP-1/JE from 286 ± 30 ng/L to 181 ± 35 ng/L and KC from 321 ± 21 ng/L to 232 ± 20 ng/L while increasing that of IL-10 from 35 ± 4 ng/L to 346 ± 35 ng/L in db/db+G mice. Further, genistein treatment suppressed diabetes-induced adhesion of monocytes to EC by 87% and endothelial secretion of adhesion molecules. We conclude that genistein improves diabetes-caused vascular inflammation, which may be mediated through promoting the cAMP/PKA pathway.

FIGURE 1

Adhesion of monocytes to HAEC cultured with mannitol or various concentrations of glucose for 48 h (A) and monocyte adhesion (B) and IL-8 (C) and MCP-1 (D) production in HAEC cultured with normal glucose (5.5 mmol/L) or HG (25 mmol/L) in the presence or absence of genistein for 48 h. Values are mean ± SEM, n = 3 (means of duplicates). Means without a common letter differ, P < 0.05. G, genistein; HAEC, human aortic endothelial cell; HG, high glucose.

FIGURE 2

Adhesion of monocytes to HAEC cultured with normal glucose or HG with or without ICI 182,780 in the presence or absence of genistein or 17β-estradiol (A). Monocyte adhesion to HAEC cultured with normal glucose or HG with or without SQ22536 or PKI in the presence or absence of genistein (B). Intracellular cAMP production (C) and PKA activity (D) in HAEC cultured with normal glucose or HG in the presence or absence of genistein for 48 h. Values are mean ± SEM, n = 3 (means of duplicates). Means without a common letter differ, P < 0.05. E2, 17β-estradiol (10 nmol/L); G, genistein (5 μmol/L); HAEC, human aortic endothelial cell; HG, high glucose (25 mmol/L); I, ICI 182,780 (1 μmol/L); PKI, protein kinase A inhibitor (2 μmol/L); SQ, adenylate cyclase inhibitor SQ22536 (10 μmol/L).

FIGURE 3

Blood glucose concentration during a glucose tolerance test (A) and an insulin tolerance test (B) in normal, db/db, and db/db+G mice treated for 8 wk. Values are mean ± SEM, n = 10. Means at time without a common letter differ, P < 0.05.