The effects of a clinically used purified micronized flavonoid fraction (S5682) containing 90% diosmin and 10% hesperidin on increased microvascular permeability induced by histamine, bradykinin and leukotriene B-4 (LTB(4)) were investigated by intravital microscopy in the cheek pouch preparation of diabetic hamsters. We also investigated the effects of S 5682 on macromolecular permeability increase and leukocyte adhesion during ischemia-reperfusion using the same preparation. Diabetes was induced by an intraperitoneal injection of streptozotocin (50 mg/kg). S 5682, suspended in 10% lactose solution, or vehicle (10% lactose) was administered orally for 25 days at 20 mg/kg/day (10 mg/kg twice a day), starting 5 days after the streptozotocin injection. Fluorescein isothiocyanate-labelled dextran (molecular weight 150,000) was given intravenously, 30 min after completion of the cheek pouch preparation. The leukocytes were stained by continuous intravenous infusion of acridine orange (0.5 mg/kg/min). Histamine (2 mu M), bradykinin (1 mu M), and LTB(4) (0.01 mu M), applied topically for 5 min, increased the number of fluorescent vascular leakage sites in postcapillary venules. A temporary ischemia (duration: 30 min) with total circulatory arrest of the cheek pouch was obtained by clamping the neck of the everted pouch. The maximum number of leaky sites (per cm(2) in the prepared area) which occurs either at 5 min after the beginning of each topical application or 10 min after the onset of reperfusion was quantified in UV light microscopy. The results from 60 animals divided into ten groups of 6 animals each are presented as means +/- SEM. In comparison with vehicle, S 5682 significantly inhibited the macromolecular permeability increasing the effect of histamine (343.8 +/- 18.5 vs. 91.0 +/- 8.2 leaks/cm(2); p < 0.001), bradykinin (347.0 +/- 14.6 vs. 110.3 +/- 8.5 leaks/cm(2); p < 0.001) and LTB(4) (323.0 +/- 15.5 vs. 161.3 +/- 13.8 leaks/cm(2); p < 0.001). At reperfusion, after 30 min ischemia, S 5682 significantly decreased the observed macromolecular permeability(168.5 +/- 19.7 vs. 52.7 +/- 6.3 leaks/cm(2); p < 0.01). Flavonoid-treated animals also tended to have a lower number of leukocytes adhering to the venular endothelium(104.8 +/- 11.0 vs. 75.8 +/- 9.7/6 mm(2); p > 0.05). These results demonstrate that oral administration of S 5682 for 25 days at 20 mg/kg body weight/day has a protective effect on leakage of macromolecules after application of permeability-increasing substances and during ischemia-reperfusion in the cheek pouch microvasculature of diabetic hamsters. In conclusion, the present data illustrating the inhibitory effect of a clinically relevant dose of S 5682 on the inflammatory processes induced in this in vivo model of microcirculation may serve as a rational basis to explain its clinical efficacy.
