Study on the experimental mechanism of Warm Purgative and Strengthening Spleen therapy combined with antibiotics in the treatment of sepsis / 中国中西医结合急救杂志

ABSTRACT

Objective To explore the synergism efficacy and mechanism of Warm Purgative and Strengthening Spleen (WPSS) therapy combined with antibiotics in the treatment of sepsis. Methods Thirty-two SPF Spargue-Dawley (SD) rats were used to replicate the rat sepsis model by cecum ligation perforation (CLP) method and equally divided into model control (MC) group, ceftriaxone group, Chinese herbal medicine (CHM) group and ceftriaxone +CHM group. Eight SD rats underwent sham surgery were used as a sham operation (Sham) group. Rats in Sham and MC groups were administered with 0.9% normal saline (NS) by intraperitoneal injection and gavage. Rats in CHM group were administered with modified Dahuang Fuzi Decoction (DFD, 8 mg/kg) by gavage + 0.9% NS by intraperitoneal injection, Bid. Rats in ceftriaxone group were administered with 0.9% NS by gavage and ceftriaxone (120 mg/kg) by intraperitoneal injection, Bid. Rats in ceftriaxone + CHM group were administered with modified DFD (8 mg/kg) by gavage and ceftriaxone (120 mg/kg) intraperitoneal injection, Bid. The drugs were given for 2 days. The mortality of rats in each group was observed after treatment. The intestinal flora changes and intestinal permeability [intestinal mucosa injury index (IMII), intestinal mucosa secretory immunoglobulin (sIgA), serum D-lactic acid, diamine oxidase (DAO) and sIgA] were detected. Meanwhile, the levels of serum inflammation indexes [lipopolysaccharide (LPS), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6)] were detected. Results ① Mortality ceftriaxone+CHM group (25.0%) < CHM group (37.5%) and ceftriaxone group (37.5%) < MC group (50.0%), the differences between groups were statistically significant (all P < 0.05). ② 16S rDNA sequencing analysis: the ratio of Bacteroidetesin in MC group was lower than that in the Sham group [(24.36±7.15)% vs. (45.20±6.05)%], and the ratio of Proteobacteria in MC group was higher than that in Sham group [(10.03±7.55)% vs. (0.41±0.21)%]. The diversity of intestinal flora in ceftriaxone group was significantly lower than that in Sham and CHM groups (404.60±17.09 vs. 470.80±16.97, 469.20±14.59), the differences between groups were statistically significant (all P < 0.05). The principal component analysis (PCA) suggested that the composition of CHM group was closer to that of Sham group, which indicated that WPSS therapy could reduce intestinal flora disorders in rats with sepsis. ③The pathological changes of intestinal mucosa light microscope showed the intestinal mucosa of Sham group was intact; the intestinal mucosa became thinner, and local inflammatory cells had infiltration in MC group. The thickness of intestinal mucosa in CHM, ceftriaxone and CHM+ceftriaxone groups was slightly thicker, and the infiltration of local inflammatory cells was less than that in MC group. The thickness of intestinal mucosa in CHM group and ceftriaxone+CHM group was slightly thicker than that in the ceftriaxone group, and the arrangement was more regular than that in MC group and ceftriaxone group.④Intestinal mucosa permeability and inflammatory state IMII, D-lactic acid, DAO, LPS, TNF-α and IL-6 of rats in MC group were higher than those of rats in Sham group [IMII 4.37±0.56 vs. 0.26±0.29, D-lactic acid (mg/L)12.35±0.83 vs. 7.30±1.29, DAO (kU/L) 2.16±0.43 vs. 0.32±0.06, LPS (kU/L) 0.663±0.012 vs. 0.095±0.003, TNF-α (μg/L) 251.03±82.69 vs. 52.15±6.25, IL-6 (μg/L) 160.50±4.77 vs. 54.30±3.36], while sIgA in MC group was lower than that in Sham group (mg/L 11.57±0.17 vs. 26.76±1.99). IMII, D-lactic acid, DAO, LPS, TNF-α and IL-6 of rats in CHM, ceftriaxone and CHM+ ceftriaxone groups were significantly lower than those of rats in MC group, while sIgA in CHM, ceftriaxone and CHM+ceftriaxone groups were significantly higher than that of rats in MC group. The change of CHM+ceftriaxone group was more significant than those of CHM group and ceftriaxone group [IMII1.78±0.23 vs. 1.96±0.62, 3.39±0.43, D-lactic acid (mg/L) 8.56±0.37 vs. 9.62±0.57,11.42±0.39, DAO (kU/L)1.14±0.12 vs. 1.72±0.24, 2.01±0.32, sIgA (mg/L) 25.34±1.49 vs. 23.99±7.85, 17.46±1.20, LPS (kU/L)0.302±0.007 vs. 0.387±0.004, 0.715±0.013, TNF-α (μg/L) 57.10±3.98 vs. 101.49±21.49, 141.91±20.20, IL-6 (μg/L) 93.71±2.39 vs. 87.12±7.31, 104.27±1.84]. Conclusion WPSS therapy may improve the efficacy of antibiotics in the treatment of sepsis by regulating the intestinal flora and reducing the intestinal mucosa permeability and inflammation level.