Stigmasterol alleviates allergic airway inflammation and airway hyperresponsiveness in asthma mice through inhibiting substance-P receptor.

CONTEXT: Stigmasterol has significant anti-arthritis and anti-inflammatory effects, but its role in immune and inflammatory diseases is still unclear. OBJECTIVE: The potential advantages of stigmasterol in asthma were explored in IL-13-induced BEAS-2B cells and asthmatic mice. MATERIALS AND METHODS: The optimal target of stigmasterol was confirmed in asthma. After detecting the cytotoxicity of stigmasterol in BEAS-2B cells, 10 µg/mL and 20 µg/mL stigmasterol were incubated with the BEAS-2B cell model for 48 h, and anti-inflammation and antioxidative stress were verified. Asthmatic mice were induced by OVA and received 100 mg/kg stigmasterol for 7 consecutive days. After 28 days, lung tissues and BAL fluid were collected for the following study. To further verify the role of NK1-R, 0.1 µM WIN62577 (NK1-R specific antagonist), and 1 µM recombinant human NK1-R protein were applied. RESULTS: NK1-R was the potential target of stigmasterol. When the concentration of stigmasterol is 20 µg/mL, the survival rate of BEAS-2B cells is about 98.4%, which is non-toxic. Stigmasterol exerted anti-inflammation and antioxidant stress in a dose-dependent manner and decreased NK1-R expression in IL-13-induced BEAS-2B. Meanwhile, in vivo assay also indicated the anti-inflammation and antioxidant stress of stigmasterol after OVA challenge. Stigmasterol inhibited inflammation infiltration and mucus hypersecretion, and NK1-R expression. DISCUSSION AND CONCLUSIONS: The protective effect of stigmaterol on asthma and its underlying mechanism have been discussed in depth, providing a theoretical basis and more possibilities for its treatment of asthma.

Magnetic Charge Model for Leakage Signals from Surface Defects in Ferromagnetic Material.

A novel three-dimensional theoretical model of magnetic flux leakage (MFL) is proposed in this paper based on the magnetic dipole model. The magnetic dipole model assumes that a ferromagnetic specimen with defects is exposed to a uniform external magnetic field that causes a uniform magnetization around the defect surface. Under this assumption, the MFL can be regarded as arising from magnetic charges on the defect surface. Previous theoretical models were mostly used to analyze simple crack defects such as cylindrical and rectangular cracks. In this paper, we developed a magnetic dipole model for more complex defect shapes such as circular truncated holes, conical holes, elliptical holes, and double-curve-shaped crack holes to complement the existing defect shapes. Experimental results and comparisons with previous models demonstrate that the proposed model provides a better approximation of complex defect shapes.

Interleukin-8 gene polymorphism is associated with acute coronary syndrome in the Chinese Han population.

BACKGROUND: Acute coronary syndrome (ACS) is one of the most common forms of heart disease. Recent studies have shown that interleukin (IL)-8 plays a key role in the development of atherosclerotic plaques, but the relationship between the common genetic variants of IL-8 and ACS has not been extensively studied. METHODS: This case-control study in the Chinese Han population included 675 patients with ACS and 636 age- and sex-matched controls. We investigated IL-8 polymorphisms and their association with susceptibility to ACS. The investigation was replicated in the second study comprising 360 cases and 360 control subjects. The plasma concentration of IL-8 was measured by enzyme-linked immunosorbent assay. RESULTS: IL-8 -251 A/T polymorphism was associated with increased susceptibility to ACS (P=0.004; odds ratio=1.30; 95% confidence interval: 1.12-1.53). The second study yielded similar results. An increased IL-8 level was found in the plasma of acute myocardial infarction patients, suggesting that IL-8 -251 A/T may affect the expression of IL-8. CONCLUSION: IL-8 -251 A/T polymorphism is associated with ACS risk in the Chinese Han population and the A allele of IL-8 -251 A/T may be an independent predictive factor for ACS.

[Prevention of rupture of atherosclerotic plaque by Candesartan in rabbit model].

OBJECTIVE: To evaluate Candesartan therapeutic effect against atherosclerotic plaque rupture and to explore the related mechanisms. METHODS: Thirty-four New Zealand White male rabbits were randomly divided into three groups: the control group, the model control group and the Candesartan intervention group. The control group rabbits were fed with a normal diet. Rabbits of the latter two groups were fed with a 1% high-cholesterol diet and received a balloon catheter injury respectively one week after the cholesterol feeding. Candesartan (0.5 mgⁱkg⁻¹â±d⁻¹) was given to the Candesartan group rabbits 2 days before the performance of the balloon catheter injury. By the end of 12(th) week of the experiment, Russell's viper venom was used for rabbits of both the model control and the Candesartan groups in order to induce rupture of the plaques developed and followed by sacrifice of all the rabbits of the 3 groups. The aortas were removed and fixed for histological evaluation. Immunohistochemistry of MMP-9, macrophage markers and collagen were performed. The protein expression of MMP-9 was determined using Western blot analysis. RESULTS: In the model control group, 7 of 9 rabbits with a total of 12 plaques developed rupture and thrombosis of the plaques after the induction. In contrast, only 2 of 10 rabbits with a total of 3 plaques demonstrated rupture and thrombosis in the Candesartan group (P < 0.05). The control group rabbits did not have plaque rupture and thrombosis. Compared with the model group, both the percentage area of MMP-9 and macrophages in the plaques were significantly decreased in the Candesartan group (12.35% ± 4.28% vs 32.58% ± 9.16%, P < 0.05; 13.87% ± 4.91% vs 23.8% ± 7.45%, P < 0.05). There was an increased percentage of collagen content in total plaques of the Candesartan group (30.27% ± 11.36% vs 4.18% ± 1.28%, P < 0.01). Compared with the model group, the protein expression of MMP-9 was significantly decreased in the Candesartan group (P < 0.01). CONCLUSION: Candesartan has a preventive value against atherosclerotic plaque rupture in hypercholesterolemic rabbits, likely through its reduction of MMP-9 expression, inhibition of macrophage accumulation and increase of collagen content within the plaques.

Pioglitazone suppresses inflammation and fibrosis in nonalcoholic fatty liver disease by down-regulating PDGF and TIMP-2: Evidence from in vitro study.

BACKGROUND: The prevalence of nonalcoholic fatty liver disease (NAFLD) has been increasing worldwide. Pioglitazone is a pharmacologic agonist of peroxisome proliferators-activated receptor-γ (PPAR-γ) that was reported to ameliorate hepatic steatosis and inflammatory changes. OBJECTIVE: We aimed to evaluate the effects of pioglitazone in NAFLD and investigate the underlying mechanism by testing platelet derived growth factor (PDGF) and tissue inhibitory of metalloproteinase-2 (TIMP-2). METHODS: A total of C57BL/6 wild-type mice were randomized to three groups, control group (NC, n= 60), high-fat control group (HF, n= 60), and pioglitazone treatment group (L,n= 60). Mice were administrated with high-fat diet to construct NAFLD model. Enzyme-linked immunosorbent assay (ELISA) was used to measure protein expression of PDGF and TIMP-2. Liver histology samples were stained with hematoxylin and eosin (H&E). RESULTS: Upon pioglitazone treatment, the PDGF and TIMP-2 expression levels were decreased compared with high-fat diet-fed mice devoid of drug stimulation. Analysis of liver histology showed pioglitazone treatment could reduce steatosis and inflammatory changes, which was helpful to inhibit hepatic fibrosis in NAFLD mice. CONCLUSIONS: The study showed pioglitazone might exert an inhibitory effect on hepatic inflammation and fibrosis in NAFLD. Moreover, this study provided novel evidence for the promising clinical application of pioglitazone in intervening NAFLD.

[Rac1 accelerates endothelial cell senescence induced by hypoxia in vitro].

To investigate the role and mechanism of Rac1 protein in the process of the human umbilical vein endothelial cell (HUVEC) senescence, we used hypoxia as a model for modulating HUVECs entering replicative senescence in vitro. Premature senescence of HUVECs was evidenced by detecting the SA-beta-Gal activity and PAI-1 expression. Meanwhile, cell cycle distribution and cell proliferation rate were investigated by flow cytometry assay and BrdU staining. The results indicated that the HUVECs became enlarged and flattened, both SA-beta-Gal activity and PAI-1 expression increased obviously, while cell proliferation was inhibited and G(1) phase cell cycle arresting occurred when HUVECs were treated with continued hypoxia for 96 h. Accompanied with these changes, the expression of activated Rac1 increased obviously in cells after hypoxia. All these observations suggested that endothelial senescence could be induced by continued hypoxia and it might correlate with the activity of Rac1. To further define the relationship between Rac1 and HUVEC senescence, HUVECs were transiently infected with the constitutively active form of Rac1 (V12Rac1) or dominant negative form of Rac1 (N17Rac1) using retrovirus vector pLNCX-V12Rac1 or pLNCX-N17Rac1. We observed the changes of these three kinds of HUVECs (HUVECs, N17Rac1-HUVECs, V12Rac1-HUVECs) after hypoxia for 48 h and 96 h, the expression and localization of serum response factor (SRF), which is one of the downstream signal molecules of Rac1, were also investigated. The results obtained indicated that after continued hypoxia for 48 h, HUVECs infected by V12Rac1 showed obvious senescence accompanied with SA-beta-Gal activation, PAI-1 expression increase, G(1) phase arrest and cell proliferation inhibition which were similar to HUVECs after continued 96-hour hypoxia treatment, while the senescence of HUVECs infected by N17Rac1 was significantly inhibited even if the cells were exposed to hypoxia for more than 96 h. All the results identified that the activation of Rac1 might accelerate HUVEC senescence induced by hypoxia and that inactivation of Rac1 could partly block the cell senescence. To further investigate the mechanism of HUVEC senescence induced by Rac1, we detected the expression of total SRF (tSRF) and nuclear SRF (nSRF) in these three kinds of HUVECs by immunofluorescent analysis and Western blot assay after hypoxia. The results showed that the expression of nSRF decreased obviously and the nuclear translocation of SRF was inhibited in HUVECs infected by V12Rac1 compared with those in the normal HUVECs. In contrast, the expression of nSRF increased obviously in the HUVECs infected by N17Rac1. These results suggest that activation of Rac1 accelerates endothelial cell senescence and inhibition of Rac1 activity prevents HUVECs from entering senescence induced by hypoxia, while the nuclear translocation of SRF regulated by Rac1 might play an important role in the process of senescence.

[Serum response factor participates in RhoA-induced endothelial cell F-actin rearrangements].

RhoA is one of the main members of RhoGTPase family involved in cell morphology, smooth muscle contraction, cytoskeletal microfilaments and stress fiber formation. It has been demonstrated that RhoA modulates endothelial cell permeability by its effect on F-actin rearrangement, but the molecular mechanism of rearrangement of actin cytoskeleton remains unclear. Recent studies prove that RhoA/Rho kinase regulates smooth muscle specific actin dynamics by activating serum response factor (SRF)-dependent transcription. To further investigate the molecular mechanism of the rearrangement of vascular endothelial cell actin cytoskeleton, we explored the relationship between the activation of SRF and F-actin rearrangement induced by RhoA in human umbilical vein endothelial cells (HUVECs). HUVECs were infected with the constitutively active forms of RhoA (Q63LRhoA) or the dominant negative forms of RhoA(T19NRhoA) using retrovirus vector pLNCX-Q63LRhoA or pLNCX-T19NRhoA, the positive clone was obtained by G418 selection. The expression and distribution of SRF in normal and infected cells were evaluated by immunohistochemistry and Western blot in complete medium and in serum-free medium. The effect of F-actin polymerization was detected by Rhodamine-Phalloidine staining. Infection of PLNCX-Q63LRhoA induced F-actin rearrangement and stress fiber formation in HUVECs, as well as enhanced the expression of SRF in the nuclei. In contrast, the cells infected with T19NRhoA showed no distinct changes. With serum deprivation, the expression of SRF increased obviously in both normal and infected HUVECs, but the subcellular localization of SRF was evidently different. In HUVECs, the localization of SRF was in the nuclei after 3 d with serum deprivation, but it was redistributed outside the nuclei after 5 d with serum deprivation. In cells infected with Q63LRhoA, the immunolocalization of SRF was always in the nuclei compared with HUVECs infected with T19NRhoA, which was almost always localized in the cytoplasm. In HUVECs, the rearrangement of F-actin and formation of stress fiber increased after 3 d with serum deprivation, but appeared decreased and unpolymerized after 5 d with serum deprivation. The polymerization of F-actin and the formation of stress fiber in HUVECs infected with Q63LRhoA kept during the period of serum-free culture, whereas the rearrangement of F-actin in cells infected with T19NRhoA was not found. These results suggest that RhoA influences endothelial F-actin rearrangement in part by regulating the expression and subcellular localization of SRF.