| Objective: To investigate the effects of Tet on LPS-induced febris in SD rats, hyperalgsia in Kunming mice, and inflammatory response in RAW264.7 cell and its possible mechanisms.Methods:1. Rat febris model was constructed by interaperitoneal injection with LPS 100μg·kg-1. Based on changes of the core body temperature of the animals, the antipyretic action of Tet was observed,compared with that of acetaminophen. Westernblotting technology was applied to analyze the expression of cyclooxygenase-2 (COX-2) in liver and brain tissues, and special kits were used to test the levels of myeloperoxidase (MPO) in liver and prostaglandin E2(PGE2) in serum.2.Mouse hyperalgesia model was established by interaperitoneal injection with LPS 100μg·kg-1. Hot-plate and writhing tests were used to evaluate analgesic effects of Tet comparing with acetaminophen. Westernblotting technology was applied to analyze the expression of COX-1, COX-2 and COX-3 in brain, and special ELISA kits were used to determine the levels of IL-6, IL-10,and TNF-αin serum. 3.Cell inflammatory model was set up by LPS( 1μg·ml-1)-stimulating RAW 264.7 cells. P65 translocation kit was used to test effects of Tet on activation of NF-κB; westernblotting technology was applied to analyze its influences on expressions of COX-2 and iNOS; moreover special ELISA and EIA kits were used to test its contribution to the levels of IL-6,IL-10,TNF-α,and PGE2 in culture media.Results:1.That core temperatures of rats increased apparently after LPS injection, and the highest of which reached to 2.5℃above normal indicated that the rat febris model was successfully constructed. Moreover, three peaks of temperature curve appeared at time points of 0.5h, 3h,and 6h,respectively, which was similar to that of previous reports. At the same time, no febrile responses were observed in acetaminophen-treated anilmals. Tet treatment decreased high fever caused by LPS in dose-dependent manner , even made the temperature low to the normal level. Correspondingly, COX-2 protein was highly expressed in liver and brain tissues of model group with highly increasing PGE2 and MPO levels in serum or liver. On the contrary,the various decreases of all these pro-inflammatory molecules were found in Tet-treated animals.2.Hyperalgsia model of mice was established with shortened latency of licking hind paws and increased the number of writhing response after injection of LPS. Acetaminophen treatment showed apparently prolonged latency of licking hind paws and decreased numbers of writhing response. Similar results to that of acetaminophen were obtained when various doses of Tet were used, and it could not only inhibit LPS-induced expression of COX-3 in brain, but also lower contents of IL-6 and TNF-α, meanwhile, increase the level of IL-10 in serum in LPS-treated mice without obvious expression changes of COX-1 and COX-2.3.Being stimulated with LPS 1μg·ml-1, RAW 264.7 cells were observed under LASER confocal microscopy. Erythrine P65 subunit of NF-κB aggregated at nuceluses, which made Erythrine fluorescence of nuceluses much stronger than that of endochylema. Correspondingly, COX-2, iNOS were induced and highly expressed in these cells, with upregulated levels of IL-6, IL-10, TNF-α, and PGE2 in supernatent. On the contrary, in the Tet-treated cells there were weak Erythrine fluorescence in the nuceluses with downregulated expressions of COX-2, iNOS, IL-6, TNF-α, PGE2 and upregulated expression of IL-10.Conclusions: All the resultes indicate that: 1.tet has antipyretic effect,. Its mechanism may be related to inhibiting the express of COX-2 in liver and brain, lowering the content of PGE2 in serum. 2.tet also has analgesic effect, which machanism may be related to inhibiting express of COX-3 in brain, and decreasing generation of IL-6 and TNF-αas well as promoting level of IL-10 in serum. 3.tet has strong anti-inflammatory effect, which may be involved in inhibiting translocation of P65 subunit of NF-κB into the nucleus, downregulating expression of COX-2, iNOS proteins, decreasing contents of IL-6, PGE2 and TNF-αas well as increasing level of IL-10.
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