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Experimental Studies in Antiinflammatory Effects of Licorice Flavonoids

Author: GuanYan
Tutor: XieQiangMin
School: Zhejiang University
Course: Pharmacology
Keywords: Licorice Flavonoid Anti-inflammatory Antioxidant Lipopolysaccharide Lung injury Cytokines The glycyrrhizin Qin sugar Cigarette smoke extract Lung epithelial cell mucin MUC5AC Interleukin-8 Human small airway epithelial cell line NCI -H292 Cytotoxic
CLC: R285
Type: Master's thesis
Year: 2009
Downloads: 68
Quote: 1
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Abstract

Part One the licorice flavonoids role in the regulation of cytokine expression in mice lung inflammation and oxidation reaction licorice leguminous perennial herb that contains mainly glycyrrhizic acid, glycyrrhetinic acid, flavonoids, alkaloids, and amino acids and other chemical ingredients. Licorice flavonoids with a wide range of pharmacological activities, including anti-oxidation, anti-tumor, anti-HIV, anti-ulcer, liver role. The early studies have shown that licorice flavonoids showed strong inhibition of capsaicin-induced guinea pig cough reflex, we hypothesized that the antitussive mechanism of action and anti-inflammatory effects. The licorice flavonoids experimental study of lung inflammation has not been reported, this study, lipopolysaccharide-induced lung inflammation model in mice to explore the anti-inflammatory effect of licorice flavonoids, and study its anti-inflammatory mechanism. The bronchoalveolar lavage fluid (BALF) in the total number of white blood cells and neutrophils, lymphocytes and macrophages of mice airway instillation of LPS after 24 h, the total number of white blood cells in the model group in BALF neutrophils cells, lymphocytes, and the number of macrophages increased significantly. The total number of white blood cells and neutrophils compared with control group increased by 8.6-fold and 515-fold (P <0.001), lymphocytes and macrophages compared with control group were significantly different (P <0.01). With LPS (model) group, the licorice flavonoids 3,10,30 mg · kg ^{-1 LPS group and DXM 1 mg · kg ^{-1 LPS group significantly inhibited the total number of white blood cells, neutrophils, lymphocytes, and the number of macrophages in the BALF increased significantly. Two pairs of lung water content in the control group lung water content of 32.95 ± 3.27%, the model group was 39.48 ± 2.06%, compared with the control group (P <0.01); licorice flavonoid 3 mg · kg ^{-1 group was 36.25 ± 2.74%, compared with the model group (P <0.05); 10 mg · kg ^{-1 group to 36.77 ± 1.83% (P <0.05); 30 mg · kg ^{-1 group was 35.06 ± 4.61% (P <0.05); DXM 1 mg · kg ^{-1 to 33.47 ± 4.51% (P <0.01). In BALF neutrophil myeloperoxidase enzyme (MPO) and superoxide dismutase (SOD) content of mice airway instillation LPS24h after, the model group, SOD activity in BALF decreased lung tissue in MPO activity unit was significantly higher (P <0.05). With LPS (model) group, the licorice flavonoids 10, 30 mg · kg ^{-1 LPS group and DXM 1 mg · kg ^{-1 LPS group could significantly improve SOD activity in BALF units, a significant reduction in lung tissue MPO activity unit. Tumor necrosis factor-alpha (TNF-alpha) mRNA in lung homogenates and interleukin-1 beta (IL-1beta) mRNA expression in mouse airway instillation of LPS 6h or 24h after lung tissue homogenates after determination of TNF -alpha mRNA express IL-1βmRNA,. Instillation of LPS 6 hours of TNF-alpha mRNA and IL-1βmRNA expression levels higher than 24 h. Instillation of LPS 6 hours of TNF-alpha mRNA compared with control group were significantly different (P <0.01), 24 h of TNF-alpha mRNA and the control group showed no significant difference (P> 0.05). Instillation of LPS 6h or 24h IL-1βmRNA expression compared with the control group were significantly different (P <0.05). The licorice flavonoids 30 mg · kg ^{-1 instillation significantly inhibited the LPS 6 h or 24 h the IL-1βmRNA expression (P <0.05), instillation also markedly inhibited the LPS 6h TNF-alpha mRNA expression ( P <0.01), but 24 h of TNF-alpha mRNA expression. DXM 1mg · kg ^{-1 only 6 h of TNF-alpha mRNA and the expression of IL-1βmRNA plays a significant role (P <0.05), expression of the two cytokines for 24 h had no significant effect. Inhibition on LPS intratracheal instillation of 2 hours after the lung tissue and BALF of TNF-alpha protein levels in mice airway the trickle LPS2h bronchial lavage and lung tissue, centrifugation and homogenized ELIESA law Determination of TNF-alpha protein levels. The results showed that the instillation of LPS 2h of TNF-alpha protein levels with the control group there were significant differences (P <0.001). Licorice flavonoids 10, 30 mg · kg ^{-1 can significantly inhibit the to trickle LPS2h TNF-alpha protein levels, the positive control drug dexamethasone 1 mg · kg ^{-1 there was a marked inhibition. 6 pairs of lung tissue to pathological changes in airway instillation of LPS 24 h after the model mice, lung tissue gap can be seen a large number of red blood cell leakage and neutrophil infiltration, alveolar destruction, alveolar widened gap structure, a transparent film the presence of edema. The licorice flavonoids 10, 30 mg · kg ^{-1 and DXM 1 mg · kg ^{-1 can significantly improve the pathological changes, licorice flavonoids 1 mg · kg ^{-1 < / sup> is not obvious. 7 Conclusion licorice flavonoids can improve the LPS-induced lung inflammation in mice, and its anti-inflammatory effects may be related to inhibition of cytokine expression and regulation of the reaction of the oxidant / antioxidant. Part Two licorice flavonoids, of glycyrrhizin celery sugar CSE induced human small airway epithelial cell line NCI-H292 cells toxic reaction, IL-8 mRNA, and MUC5AC mRNA expression of affect in the first part of the study, we have proved that licorice flavonoids can improve LPS mice caused by inflammation of the lungs, its anti-inflammatory effects may be related to inhibition of cytokine expression and regulation of the reaction of the oxidant / antioxidant. In the second part of the study, we first want to prove the further purified licorice flavonoids and licorice flavonoids liquiritin extract (CSE) against the burning cigarette-induced pulmonary epithelial cells cytotoxicity sugar celery; Second explore energy No adjustment CSE-induced lung epithelial cell mucin MUC5AC mRNA expression and interleukin-8 (IL-8) mRNA expression. Experimental Methods MTT (MTT) colorimetric assay the CSE, drugs and CSE and drugs on cell proliferation of MUC5AC mRNA detected by RT-PCR and IL-8 mRNA of the expression. Induced 1.CSE NCI-H292, A549 and HCC-827 cell toxic reaction CSE was concentration-dependent induction of NCI-H292, A549 and HCC-827 cytotoxic response and reduce cell survival. In the three cell lines, CSE NCI-H292 cell lines are the most sensitive. In addition, the cell line human small airway epithelial cells, rapid proliferation in the passage growth process. Concentration-dependent induction of NCI-H292 cell death induced by 2.CSE NCI-H292 cells toxic reaction time and dose relationship CSE 1.25% to 10.0% was between 24 to 72 hours in culture, cell death increased with time . 3 licorice flavonoids (LF) of NCI-H292 cell survival LF (0.005 to 5μg/ml) no significant effect on survival of NCI-H292 cells. Glycyrrhizin the celery sugar (LA) NCI-H292 cell survival is low concentration of LA (10 ^{-10 ~ 10 ^{-6 M) NCI-H292 cells survival rate had no significant effect. When the LA concentration 10 ^{to 5 to 10 ^{-4 M for 24h, showing obvious cytotoxicity. Cultured for 24h or 48h, this cytotoxic effect is waning, that high concentrations of LA cytotoxic effect is mild and recoverability. NCI-H292 cells induced by toxic protective effect of licorice flavonoids (LF) CSE LF (5μg/ml) CSE-induced NCI-H292 cells poison a significant protective effect, but when the LF concentration of 0.005 to 0.5μg/ml no significant role. 6 the glycyrrhizin celery sugar (LA) of the CSE-induced NCI-H292 cells poison when the LA concentration of 10 ^{-7 to 10 ^{-6 M, can significantly enhance the CSE-induced cytotoxic response (P <0.05 to 0.01). The LA concentration 10 ^{10 to 10 ^{-8 M, the CSE-induced cytotoxic response is not obvious (P> 0.05) 7.CSE NCI-H292 cells induced by IL 8 and MUC5AC mRNA expression in a time and dose relationship between the concentration of 2.5% CSE exposure NCI-H292 cells 6h and 24h could induce the expression of IL-8 mRNA, (P <0.05 to 0.01), and 1.25% CSE exposure NCI-H292 cells 24 h can not induce the expression of IL-8 mRNA, 2.5% CSE concentration exposure of NCI-H292 cells 1h and 3h can not induce the expression of IL-8 mRNA, suggesting that CSE-induced NCI-H292 cells IL-8 mRNA expression was time-and dose-effect relationship. Similarly, 2.5% CSE concentration exposed NCI-H292 cells for 24 h significantly induced MUC5AC mRNA expression (P <0.05), and 1.25% CSE exposure NCI-H292 cells 24h not significantly exposed NCI-H292 cells induced MUC5AC mRNA expression, 2.5% CSE concentration 1h, 3h and 6h can not induce the expression of IL-8 mRNA, prompted CSE induction NCI-H292 cells MUC5AC mRNA expression was time-and dose-effect relationship. 8.LF CSE-induced NCI-H292 cells IL-8 mRNA and MUC5AC mRNA expression 2.5% CSE concentration exposed NCI-H292 cells for 24 hours could induce the expression of IL-8 mRNA, (P <0.01), LF 0.5 and 5μg/ml pre- processing, and does not deal with the sample, it shows a strong inhibitory effect (P <0.05) significantly inhibited (P <0.05), the positive control 1μM dexamethasone processing. In Similarly, LF 0.5μg/ml pretreatment, and does not deal with the sample, exposed to 2.5% CSE concentration NCI-H292 cells for 24 hours induced MUC5AC mRNA expression show significant inhibition (P <0.05). CSE 9.LA NCI-H292 cells induced by IL-8 mRNA and MUC5AC mRNA expression of concentration of 2.5% CSE exposure NCI-H292 cells for 24 hours could induce the expression of IL-8 mRNA, (P <0.05), LA 1 and 10μM pretreatment of and does not deal with the sample, no significant inhibitory effect (P> 0.05). Similarly, LA1 and 10μM pretreatment, and does not deal with the sample exposed to 2.5% CSE concentration NCI-H292 cells for 24 hours induced MUC5AC mRNA expression did not show significant inhibition (P> 0.05). 10. Conclusions These results show that the NCI-H292 is a good research CSE-induced model of epithelial cells in inflammatory mechanisms. Licorice flavonoids (LF) can reduce the CSE-induced epithelial cell cytotoxicity reaction, inhibition of CSE-induced epithelial cell expression of IL-8 and MUC5AC, glycyrrhizin celery sugar no significant role.}}}}}}}}}}}}}}}}}}}}}}}

Experimental Studies in Antiinflammatory Effects of Licorice Flavonoids

Abstract

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