Objective:The microcirculation and leukocyte-endothelial cell interaction in synovial tissue of an inflamed joint are known to play a crucial role in the pathogenesis of rheumatoid arthritis. The aim of this study was to characterize the in vivo changes in the microvasculature and in leukocyte-endothelial cell interactions in the mouse synovial tissue using intravital fluorescence microscopy in three stages of antigen-induced arthritis. The expression of E-and P-selectin and ICAM-1 were also studied using immunohistochemistry. Methods: Antigen-induced arthritis (AiA) was produced in Balb/c mice. The severity of arthritis at three different phases was quantified using a clinical and histological score. For the intravital fluorescence microscopy measurements, the patella tendon was partially resected for visualization of the intraarticular synovial tissue of the knee joint. The number of rolling and adherent leukocytes, functional capillary density (FCD) and RBC velocity were quantitatively measured in synovial microvessels. Expression of ICAM-1, E-and P-selectin was assessed by immunohistochemistry. Results: There was a significant increase in the leukocyte rolling fraction in postcapillary venules in the acute phase of AiA (from 0.26 ± 0.05 in controls to 0.45 ± 0.04 8 d after AiA induction). The number of leukocytes adherent to the endothelium was significantly elevated in all phases of arthritis (from 121 ± 27 in controls to 376 ± 62 mm 2 63 d after AiA-induction). Functional capillary density was significantly enhanced in the acute (332 ± 15 cm/cm 2 ) and intermediate phases (320 ± 15 cm/cm 2 ) compared to control values (227 ± 15 cm/cm 2 ). Arthritis resulted in a distinct increase in the expression of ICAM-1 on the synovial endothelium in all phases of AiA. E-and P-selectin expression were detected only in the acute phase. Conclusion: Our model provides new insights into the microcirculatory changes which occur in the synovial tissue of an arthritic joint.
Hemoglobin-based oxygen carriers have been suggested to enhance the formation of oxygen free radicals, especially under conditions of ischemia-reperfusion (I/R), in which activation and adhesion of leukocytes play a pivotal role for propagation of reperfusion injury. This study investigates the effects of the hemoglobin-based oxygen carrier diaspirin-cross-linked hemoglobin (DCLHb) in an I/R model of hamster striated skin muscle. The dorsal skinfold chamber model in the awake Syrian golden hamster was used for analysis of the microcirculation and local tissue [Formula: see text]in striated skin muscle utilizing the technique of intravital fluorescence microscopy and a multiwire platinum surface (Clark type) electrode. Measurements were made before 4 h of pressure-induced ischemia and at 0.5, 2, and 24 h of reperfusion. Animals were treated with 5 ml/kg body wt of either 10% DCLHb ( n = 8), 6% Dextran 60 (Dx-60; 60 kDa, n = 8), or 0.9% NaCl ( n = 7), which was given intravenously 15 min before reperfusion. In animals treated with DCLHb or Dx-60, a significant decrease of leukocytes rolling along and sticking in postcapillary venules, associated with a recovery of functional capillary density and red blood cell velocity, was observed compared with saline-treated controls. In the early reperfusion period (0.5 h), DCLHb and Dx-60 efficiently restored local tissue[Formula: see text], whereas tissue[Formula: see text] decreased from 18.3 ± 1.9 to 15.3 ± 5.3 mmHg in 0.9% NaCl-treated animals. Electron microscopic analysis of the postischemic tissue at 24 h of reperfusion revealed markedly reduced tissue damage in animals treated with DCLHb compared with Dx-60 or isotonic saline. These results indicate that DCLHb attenuates postischemic reperfusion injury of striated skin muscle, presumably through alterations of leukocyte-endothelial cell interactions.
We have recently shown that the purified micronized flavonoid fraction (90% diosmin and 10% hesperidin) Daflon® 500 mg attenuates reperfusion injury in the striated skin muscle of the hamster. Herein, we report on the action of Daflon® 500 mg on postischemic macromolecular leakage of FITC-dextran 150 kD provoked by tourniquet ischemia. Intravital fluorescence microscopy was used for analysis of macromolecular leakage in the microcirculation model of the hamster. A tourniquet ischemia of 4 h duration was induced followed by reperfusion. Animals were treated by gavage of Daflon® 500 mg (n = 6) for 8 days at a daily dose of 30 mg kg-1 body weight. Control animals received equivalent volumes of the vehicle (5% Arabic gum solution, n = 6). Measurements of the microcirculatory parameters were made before induction of ischemia and at 0.5,2 and 24 h of reperfusion. After induction of ischemia, macromolecular leakage from postcapillary venules was significantly enhanced in vehicle-treated animals. Treatment with Daflon® 500 mg significantly attenuated macromolecular leakage of FITC-dextran 150 kD. Preliminary data from a histomorphometric analysis (n = 3/experimental group) indicated that the number of emigrated (extravascular) leukocytes after ischemia reperfusion was markedly reduced in Daflon® 500 mg-treated animals as compared to controls. These data indicate that Daflon® 500 mg prevents leakage of the macromolecular tracer FITC-dextran 150 kD from postcapillary venules after postischemic reperfusion, presumably through an inhibitory action on the emigration of activated leukocytes.
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