To explore the intervention effect of jiedu dihuang decoction on the inflammatory microenvironment of colon cancer by regulating mitophagy and nucleotide - binding oligomerization domain (NOD) like receptor hot protein domain-related protein 3 (NLRP3) inflammasome.

The animal model was successfully replicated by intraperitoneal injection of azoxymethane (AOM) combined with feeding dextran sodium sulfate (DSS). The animals were randomly divided into A: control group, B: model group, C, D, E: Jiedu Dihuang Decoction low, medium and high dose group, F: positive drug group. Ninety male C57BL/6 mice were randomly divided into 6 groups of 15 mice each, and all groups were treated continuously for 10 weeks. The pathological changes of rectal tissue were evaluated by hematoxylin-eosin (HE) staining. Immunofluorescence staining was used to observe the expression site and intensity of PTEN-induced kinase 1 (PINK1), Parkinson’s disease protein (Parkin) and B-cell lymphoma-2-interacting myosin-like coiled-coil protein 1 (Beclin1) in colon tissue. The expression of NLRP3, apoptosis-associated speck-like protein (ASC) and Caspase-1 mRNA in rectum was detected by polymerase chain reaction (PCR). The protein expression levels of PINK1, Parkin, Beclin1, NLRP3, ASC and Caspase-1 were detected by Western blotting (WB). The ratio of helper T cell 1 (Th1) to helper T cell 2 (Th2) was detected by flow cytometry. The levels of interleukin-4 (IL-4), interleukin-6 (IL-6), interleukin-10 (IL-10), interleukin-12 (IL-12), tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) were detected by enzyme-linked immunosorbent assay (Elisa).

The mRNA expression levels of NLRP3, ASC and Caspase-1 in the model group were 2.06±0.22, 3.03±0.16 and 3.28±0.22, respectively, while those in the control group were 1.01±0.17, 1.01±0.19 and 1.00±0.11, respectively, with statistically significant differences between the two groups (all P<0.01). The mean fluorescence intensities of PINK1, Parkin and Beclin1 in the model group were 76.39±4.89, 83.12±5.01 and 87.43±3.77, respectively, compared with 122.99±6.93, 138.38±3.84 and 143.22±9.42 in the control group, and the differences were statistically significant (all P<0.01). As for the proportion of T helper cells in peritoneal fluid, the model group showed a significantly lower Th1 proportion (0.74±0.09)% and a significantly higher Th2 proportion (2.05±0.26)% compared with the control group [(4.78±0.62)% and (0.26±0.07)%, respectively, all P<0.01]. In terms of peripheral blood cytokine levels, the concentrations of IL-4, IL-6, IL-10 and TNF-α in the model group were (48.98±3.78) pg·mL^-1, (44.42±2.55) pg·mL^-1, (37.67±5.34) pg·mL^-1 and (79.11±6.32) pg·mL^-1, respectively, which were significantly higher than those in the control group [(26.44±5.64) pg·mL^-1, (24.21±2.25) pg·mL^-1, (19.06±3.76) pg·mL^-1 and (35.78±13.17) pg·mL^-1, respectively]. Conversely, the levels of IL-12 and IFN-γ in the model group were (28.28±2.33) ng·mL^-1 and (238.67±35.78) pg·mL^-1, respectively, which were significantly lower than those in the control group [(55.39±3.47) ng·mL^-1 and (517.72±52.15) pg·mL^-1, respectively], with all differences reaching statistical significance (all P<0.01). The mRNA expression levels of NLRP3 in the low, medium, high-dose Jiedu-Dihuang Decoction groups and the positive drug group were 1.75±0.10, 1.59±0.11, 1.31±0.11 and 1.48±0.10, respectively; the mRNA expression levels of ASC were 2.69±0.10, 1.96±0.14, 1.54±0.18 and 1.89±0.09, respectively; and the mRNA expression levels of Caspase-1 were 2.85±0.08, 2.24±0.26, 1.58±0.22 and 1.94±0.16, respectively. Except for the low-dose Jiedu-Dihuang Decoction group, the differences between the other treatment groups and the model group were statistically significant (P<0.05, P<0.01). The mean fluorescence intensities of PINK1 in each treatment group were 87.50±9.38, 98.75±7.13, 107.28±6.60 and 94.75±7.11, respectively; those of Parkin were 94.11±8.17, 103.13±8.59, 111.27±9.66 and 101.27±9.21, respectively; and those of Beclin1 were 100.2±7.11, 107.92±6.84, 122.65±8.36 and 105.44±7.55, respectively. Except for the low-dose Jiedu-Dihuang Decoction group, the differences between the other treatment groups and the model group were statistically significant(P<0.05, P<0.01). The proportions of Th1 in peritoneal fluid of each treatment group were (1.34±0.13)%, (1.78±0.21)%, (2.89±0.23)% and (2.47±0.24)%, respectively; and the proportions of Th2 were (1.65±0.18)%, (1.06±0.11)%, (0.57±0.14)% and (0.68±0.07)%, respectively. Except for the low-dose Jiedu-Dihuang Decoction group, the differences between the other treatment groups and the model group were statistically significant (all P<0.05, P<0.01). The levels of IL-4 in peripheral blood of each treatment group were (41.67±4.31), (36.39±5.93), (33.61±4.30) and (33.94±5.81) pg·mL^-1, respectively; those of IL-6 were (39.10±2.89), (32.85±3.98), (28.91±1.91) and (27.63±1.98) pg·mL^-1, respectively; those of IL-10 were (32.11±7.50), (26.28±4.65), (23.22±4.35) and (26.22±6.15) pg·mL^-1, respectively; those of TNF-α were (64.13±13.12), (57.44±13.62), (47.45±10.06) and (50.78±10.9) pg·mL^-1, respectively; those of IL-12 were (34.09±2.49), (38.53±3.84), (42.67±3.87) and (41.93±4.39) ng·mL^-1, respectively; and those of IFN-γ were (286.57±54.77), (358.59±52.15), (435.28±73.12) and (405.33±39.33) pg·mL^-1, respectively. Except for the low-dose Jiedu-Dihuang Decoction group, the differences between the other treatment groups and the model group were statistically significant(all P<0.05, P<0.01)

Jiedu-Dihuang Decoction can improve the inflammatory microenvironment of colon cancer, which may be related to the regulation of NLRP3 inflammasome by mitochondrial autophagy.