[The role of S100A8/RAGE and Caveolin-1 and the effect of roxithromycin on their expression in a rat model of neutrophilic asthma].

Objective: To explore the role of S100A8, the receptor for advanced glycation endproducts (RAGE) and Caveolin-1 in neutrophilic asthmatic rats, and to further study the intervention of roxithromycin and the possible mechanisms. Methods: Male Brown Norway rats were randomly assigned to a control group, an asthma group and a Roxithromycin group. The asthmatic rat model was established by intraperitoneal injection of ovalbumin (OVA) and Freund's complete adjuvant (FCA) mixture, and aerosol inhalation of OVA. Rats in the Roxithromycin group were given roxithromycin injection 30 mg/kg 30 minutes before each challenge. Rats in the control and the asthma groups were replaced with equal volumes of saline, respectively. Bronchoalveolar lavage fluid (BALF) neutrophil percentage (Neu%) and pathological changes of pulmonary tissue (hematoxylin-eosin, HE staining) were measured to confirm the establishment of asthmatic models. The concentration of inflammatory cytokines and S100A8 were quantified by enzyme-linked immunosorbent assay (ELISA), and the expression of Caveolin-1 and RAGE at protein levels were detected by immunohistochemistry and Western blot. Results: Neu% in BALF of the asthma group was significantly higher than those of the control group, and Neu% in the Roxithromycin group was lower than the asthma group (all P<0.01). Pulmonary histology revealed that there were a large number of inflammatory cells infiltrated in the bronchial and perivascular, pulmonary interstitial and alveolar spaces, and the bronchial wall and smooth muscles were thickened obviously in the asthma group. Rats in the Roxithromycin group showed milder inflammation and airway remodeling change than the asthma group. There was no obvious pathological damage in the control group. The concentration of IL-6 and IL-17 in BALF and serum of rats in the asthma group were significantly higher than those in the control group (P<0.01), and Roxithromycin inhibited the high expression of these cytokines (P<0.05). The expression of S100A8 and RAGE in the asthma group were significantly higher than those in the control group [(20.6±4.4) vs (7.1±2.0) ng/L; (885±118) vs (462±102) ng/L; (14.2±1.7) vs (7.6±1.8) ng/L; (774±166) vs (406±69) ng/L, all P<0.05], and Roxithromycin inhibited the high expression of these proteins [(14.3±3.7) vs (20.6±4.4) ng/L; (650±53) vs (885±118) ng/L; (10.4±1.2) vs (14.2±1.7) ng/L; (560±64) vs (728±72) ng/L] (all P<0.05). Meanwhile, the expression of Caveolin-1 in the asthma group was significantly lower than that in the control group (P<0.01), and Roxithromycin up-regulated its expression (P<0.01). Correlation analysis showed that there was a significantly positive correlation between the expression of S100A8 and RAGE (r=0.706, P<0.01), while there was a significantly negative correlation between the expression of S100A8 and Caveolin-1 (r=-0.775, P<0.01), and between the expression of Caveolin-1 and RAGE (r=-0.919, P<0.01). Conclusion: S100A8 and Caveolin-1 may play an important role in neutrophilic asthma via RAGE, and Roxithromycin may exerts anti-inflammatory effects and inhibition of airway remodeling partly through this signaling pathway.

A novel mechanism of action for salidroside to alleviate diabetic albuminuria: effects on albumin transcytosis across glomerular endothelial cells.

Salidroside (SAL) is a phenylethanoid glycoside isolated from the medicinal plant Rhodiola rosea. R. rosea has been reported to have beneficial effects on diabetic nephropathy (DN) and high-glucose (HG)-induced mesangial cell proliferation. Given the importance of caveolin-1 (Cav-1) in transcytosis of albumin across the endothelial barrier, the present study was designed to elucidate whether SAL could inhibit Cav-1 phosphorylation and reduce the albumin transcytosis across glomerular endothelial cells (GECs) to alleviate diabetic albuminuria as well as to explore its upstream signaling pathway. To assess the therapeutic potential of SAL and the mechanisms involved in DN albuminuria, we orally administered SAL to db/db mice, and the effect of SAL on the albuminuria was measured. The albumin transcytosis across GECs was explored in a newly established in vitro cellular model. The ratio of albumin to creatinine was significantly reduced upon SAL treatment in db/db mice. SAL decreased the albumin transcytosis across GECs in both normoglycemic and hyperglycemic conditions. SAL reversed the HG-induced downregulation of AMP-activated protein kinase and upregulation of Src kinase and blocked the upregulation Cav-1 phosphorylation. Meanwhile, SAL decreased mitochondrial superoxide anion production and moderately depolarized mitochondrial membrane potential. We conclude that SAL exerts its proteinuria-alleviating effects by downregulation of Cav-1 phosphorylation and inhibition of albumin transcytosis across GECs. These studies provide the first evidence of interference with albumin transcytosis across GECs as a novel approach to the treatment of diabetic albuminuria.

Diallyl disulfide and diallyl trisulfide protect endothelial nitric oxide synthase against damage by oxidized low-density lipoprotein.

Garlic is viewed as an effective health food against atherosclerosis. In this study, we examined whether diallyl disulfide (DADS) and diallyl trisulfide (DATS) protect endothelial nitric oxide synthase (eNOS) activation against oxidized LDL (ox-LDL) insult and through what mechanism. We found that DADS and DATS reversed the suppression of eNOS Ser1177 phosphorylation by ox-LDL, and wortmannin abolished the reversal by DADS and DATS. Similarly, the inhibition of cellular cGMP and nitric oxide production by ox-LDL was reversed by DADS and DATS (p<0.05). This increase in nitric oxide bioavailability by the allyl sulfides was attenuated by wortmannin. Immunoprecipitation assay revealed that DADS and DATS preserved the interaction of eNOS with caveolin-1 in the membrane. In addition, DADS and DATS suppressed the reduction of the cellular eNOS protein content by ox-LDL. When cycloheximide was added to block protein synthesis, DADS and DATS suppressed eNOS protein degradation similarly to that noted by MG132. Ox-LDL increased chymotrypsin-like proteasome activity, and this increase was inhibited by the allyl sulfides and MG132 (p<0.05). These results suggest that DADS and DATS protect eNOS activity against ox-LDL insult. This protection can be attributed partly to their mediation of phosphatidylinositol 3-kinase/protein kinase B signaling and prevention of eNOS degradation.

Genistein restores caveolin-1 and AT-1 receptor expression and vascular function in large vessels of ovariectomized hypertensive rats.

OBJECTIVE: The soy-derived phytoestrogen genistein improves endothelial function in postmenopausal women and ovariectomized (OVX) normotensive rats. We hypothesized that genistein would improve vascular reactivity involving changes in endothelial nitric oxide synthase (NOS) expression and its regulatory proteins (caveolin and calmodulin), angiotensin II receptor, and/or oxidative status in OVX spontaneously hypertensive rats (SHRs). DESIGN: After ovariectomy or sham operation, 23-week-old female SHRs received either 17beta-estradiol (2 mg/kg/wk SC), genistein (10 mg/kg/d by gavage), or placebo. RESULTS: In OVX rats, final body weight was increased and uterus weight was decreased, and these values were reduced and increased, respectively, by 17beta-estradiol but unaffected by genistein. Acetylcholine-induced endothelium-dependent vasorelaxation was significantly blunted in aortas from OVX placebo SHRs. The contractions induced by the NOS inhibitor L-NAME and angiotensin II in OVX placebo were lower and higher, respectively, than in sham rats. Estradiol and genistein restored these functional changes. Aortic endothelial NOS and calmodulin-1 expression were unchanged, whereas caveolin-1 and angiotensin II receptor expression was increased in OVX rats. Estradiol and genistein treatment did not modify endothelial NOS, but normalized caveolin-1 and angiotensin II receptor and increased calmodulin-1 expression. Vascular superoxide production was increased in OVX placebo and normalized by both estradiol and genistein. CONCLUSIONS: Genistein prevented all cardiovascular changes induced by estrogen depletion in SHRs to a similar extent as estradiol but had no uterotrophic effect. The present findings may help to explain the potential benefits of genistein as a therapeutic agent for preventing menopausal vascular complications, especially in hypertensive women.