Results<?xml:namespace prefix = o ns = "urn:schemas-microsoft-com:office:office" /> We used the Evans blue dye extravasation method to study the lung capillary permeability. Challenge with LPS caused a 2-fold increase in the microvascular permeability index in the lung (Fig. 1). Treatment of the LPS-challenged animals with HES 3.75 and 7.5 mL/kg, but not 15 and 30 mL/kg, significantly reduced (P＜ 0.05) the LPS-induced increase in permeability. HES alone had no effect on permeability index. The wet/dry weight ratio, which represents the percentage of tissue water, is another index of tissue microvascular permeability. The results of this study were in parallel with those of the Evans blue dye study (Fig. 2). LPS significantly increased the lung wet/dry weight ratio. Rats co-treated with HES 3.75 and 7.5 mL/kg, but not 15 and 30 mL/kg, had decreased ratios compared with rats treated only with LPS. HES alone had no significant effect. We studied the functional consequence of HES on neutrophil influx into the lungs by using MPO activity as an index of tissue neutrophil accumulation. As shown in Figure 3, MPO levels were significantly increased from 0.38 ± 0.013 U/g in control rats to 0.67 ± 0.029 U/g in the LPS group of rats and were reduced to 0.51 ± 0.015 U/g, 0.48± 0.026 U/g, 0.56 ± 0.040 U/g, and 0.60± 0.033 U/g in the rats treated with LPS plus HES 3.75, 7.5, 15, or 30 mL/kg. Control rats and rats treated with HES alone had similar lung MPO activities. Increased surface expression of β2 adhesion molecules is important for neutrophil adherence to endothelial cells. As shown in Figure 4, LPS caused a more than twofold increase in surface expression of CD11b. Rats given increasing doses of HES progressively decreased the expression. Treatment of the LPSchallenged animals with HES 3.75, 7.5, 15, and 30 mL/kg reduced the LPS-induced increase of CD11b expression by 28%, 33%, 47%, and 50%, respectively. Treatment of control animals with HES did not affect the CD11b level on neutrophils. CINC plays a pivotal role in neutrophil recruitment and acute lung injury. To address whether HES affects lung CINC expression, we compared CINC protein levels in different groups. ELISA results showed low CINC protein levels in control lungs (Fig. 5); these levels increased by 2.5-fold in lungs of LPS-challenged animals. Treatment with HES 3.75, 7.5, and 15 mL/kg significantly reduced (P ＜ 0.05) the LPS-induced increase in CINC levels. HES alone had no effect on CINC expression. To investigate the possible mechanism of HES action, we performed the EMSA experiment to examine the effect of HES on the activation of NF-kB induced by LPS. We quantitated these NF-kB bands by using densitometry. As shown in Figure 6, the NF-kB activity was low in nuclear protein from control lungs and neutrophils but was markedly increased in specimens from LPS-challenged rats. This increased NF-kB activity was inhibited by treatment with HES in a doserelated manner. The maximal inhibition of LPSinduced increase in NF-kB activity was observed at the HES dose of 7.5 mL/kg in both specimens. This was the same dose at which HES maximally inhibited an LPS-induced increase in lung microvascular permeability(Fig. 1), lung wet/dry weight ratio (Fig. 2), lung accumulation of neutrophils (Fig. 3), and lung CINC protein expression (Fig. 5). |
