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OBJECTIVE To investigate the effect of 717 Jiedu decoction on neutrophil extracellular traps (NETs) and CYR61/CCN1 expression in local tissue of rats after Agkistrodon halys bite. METHODS SD rats were randomly divided into normal control group, Agkistrodon halys bite group and high-dose and low-dose 717 Jiedu decoction groups. The serum and thigh gastrocnemius were collected, and the pathological changes of gastrocnemius in each group were observed by Hematoxylin-Eosin staining(HE) and Masson staining, the ultrastructure of gastrocnemius in each group was observed by transmission electron microscopy, the apoptosis of gastrocnemius cells in each group was detected by TUNEL method, and the serum circulating-free cell (cf-DNA) level was detected by PicoGreen fluorescence. The expression levels of myeloperoxidase (MPO) in serum and gastrocnemius were detected by enzyme-linked immunosorbent assay(ELISA), the change of neutrophil elastase (NE) in gastrocnemius was detected by immunohistochemistry, and the level of citcitinic histone H3 (CitH3) in gastrocnemius was detected by immunofluorescence. The expressions of CYR61/CCN1 protein and its mRNA in gastrocnemius were detected by Western blot and PCR. RESULTS Compared with the normal group, the model group showed significant inflammatory infiltration and bleeding reaction, and the degree of collagen fibrosis increased. Ultrastructure showed that the structure of gastrocnemius cells in the model group was seriously damaged, and the distribution of muscle segments was asymmetrical. The contents of cf-DNA and MPO in serum were significantly increased (P<0.01), the protein expressions of MPO and CitH3 in gastrocnemius were increased (P<0.05 or P<0.01), and the protein and mRNA expressions of CYR61/CCN1 in gastrocnemius were up-regulated. All the administration groups of 717 Jiedu decoction significantly improved the local inflammatory response and bleeding symptoms of rats, reduced the degree of collagen fibrosis, effectively inhibited the apoptosis rate of gastrocnemius cells of pit viper bitten rats, and reduced the contents of cf-DNA and MPO in serum (P<0.05 or P<0.01). The expressions of NE and MPO in gastrocnemius were inhibited in the high-dose 717 Jiedu decoction group, and the expression of CitH3 in gastrocnemius muscle was decreased in all administration groups of 717 Jiedu decoction (P<0.05 or P<0.01), and the high expression of CYR61/CCN1 protein and its mRNA was inhibited. CONCLUSION The venom of Agkistrodon halys can induce the formation of neutrophil NETs in local tissue and the overexpression of CYR61/CCN1. The 717 Jiedu decoction could improve the ultrastructure of gastrocnemius muscle and inhibit cell apoptosis, which may also improve snitch symptoms by regulating the NETs-Cyr61/CCN1 pathway. The signaling molecules of NETs and CYR61/CCN1 are expected to be new targets for the prevention and treatment of local tissue damage caused by Agkistrodon halys venom.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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