Effects of Toona sinensis leaf extracts on cytokine levels in RAW264.7 Cells induced by LPS and in septic mice

Autor: Chia-Chun Shih, 施佳君
Rok vydání: 2015
Druh dokumentu: 學位論文 ; thesis
Popis: 103
Background: Sepsis remains a major cause of mortality in intensive care. Previous studies have pointed out that sepsis is caused by excessive inflammatory responses and oxidative stress, leading to cell and organ damage in a host. In addition, Toona sinensis is believed to possess anti-oxidant and anti-inflammatory properties. Therefore, we evaluated the effects of Toona sinensis leaf extracts (TS) on the LPS-induced inflammatory responses and oxidative stress in RAW264.7 cells and cytokine levels and liver injury markers in septic mice. Methods: The in vitro study: we divided the cells into four groups, namely, the Control group, TS group, LPS group, and TS+LPS group. The TS group and LPS group were treated with TS (25, 50, 75 microgram/mL) and LPS (1 microgram/mL), respectively. The TS+LPS group was pretreated with TS extracts (25, 50, 75 microgram/mL) prior to the LPS treatment (1 microgram/ml). We analyzed the macrophage activation status, cell viability status, sepsis related cytokines (proinflammatory: IL-17A, IL-1beta, IL-6, IFN-gamma?z?nTNF-alpha and anti-inflammatory: IL-10), transcription related factors (IkappaB-alpha and IKK), oxidative stress, and mitochondrial functions. Furthermore, in the in vivo study, we divided the mice into five groups, namely, the Control group, TS group, sham group, CLP (Cecal ligation and puncture) group, and TS+CLP group. The C57BL/6 mice were supplemented with TS (100 mg/kg B.W./day) in feed prior to the CLP operations. The sepsis related cytokines (IL-17A, IL-10, IL-1beta, IL-6, IFN-gamma?z?nand TNF-alpha) and liver injury markers (AST and ALT) were also assayed. Results: The results of the in vitro study showed that the levels of IL-10 were decreased in the TS group compared with those in the Control group; furthermore, compared with those in the Control group, after the LPS treatment, the activation of macrophages, the levels of IL-6?z?nTNF-alpha?z and IL-10, the protein expression of IkappaB-alpha and IKK, and intracellular oxidative stress were increased; the cell viability and mitochondrial membrane potential were decreased. In addition, the levels of IL-6, ?nTNF-alpha?z?n?nIkappaB-alpha, IKK, and intracellular oxidative stress were significantly decreased in the TS+LPS group compared with those in the LPS group. The results of the in vivo study showed that the levels of IL-17A, IL-1beta, IL-6, and IL-10 were significantly increased in the CLP6hr group compared with those in the Control group; the levels of IL-6 and IL-10 were significantly increased in the CLP12hr group compared with those in the Control group. Besides, our results showed that compared with those in the CLP6hr group, the levels of IL-17A was significantly decreased in the TS+CLP6hr group. On the other hand, the markers of liver injury indicated that the level of ALT was increased in the CLP6hr group compared with that in the sham6hr group; the ALT and AST levels were increased in the CLP12hr group compared with those in the sham12hr group. Besides, the level of ALT was significantly decreased in the TS+CLP12hr group compared with that in the CLP12hr group. Conclusions: The results revealed that TS supplementation would inhibit the levels of oxidative stress and subsequently decrease the activation of transcription related factors to reduce the levels of TNF-alpha and IL-6 in vitro. Simultaneously, TS supplementation would decrease the level of IL-17A and liver injury in septic mice. Therefore, in the future, TS may be an effective adjuvant therapy to antibiotics. Our study may provide the clinical applications of the effects of TS on inflammatory responses of cytokines.
Databáze: Networked Digital Library of Theses & Dissertations
Externí odkaz: