Interleukin 6 (il 6)

COP (purity >98%) was purchased from Shanghai Yuan Ye Biotechnology Co., Ltd (Shanghai, China). Dextra sulfate sodium was obtained from MP Biomedical (Irvine, CA, USA). Cefadroxil (purity >99%), terramycin (purity >98%) and erythromycin (purity >99%) were purchased from Dalian Meilun Biotechnology Co., Ltd (Dalian, China). Mesalazine (MSZ) was obtained from Losan Pharma GmbH, Germany. ELISA kits (TNF-α, IL-6, IL-10, IL-18, IL-1β, TGF-β, and IFN-γ) were obtained from Shanghai Enzyme-linked Biotechnology Co., Ltd (Shanghai, China). BCA kit and myeloperoxidase (MPO) assay kit were obtained from the Jiancheng Bioengineering Institute of Nanjing (Nanjing, Jiangsu, China). Primary antibodies against p65, IκBα, p-IκBα, NLRP3, ASC, Capase-1, Caspase-1 p10, GAPDH and Histone H3 were purchased from Affinity Biosciences (OH, USA). All other reagents and chemicals were at least of analytical grade.

The Fas L, TGF β1, IL-1, and IL-6 in the transfected cells and supernatant were analyzed using commercial ELISA kits for humans (catalog numbers: Fas L, ml000971; TGF β1, ml013583; IL-1, ml001554; IL-6, ml001532; Shanghai Enzyme-linked Biotechnology Co., Ltd., Shanghai, China) following the manufacturer’s instructions.

The serum of mice was collected, and the serum TNF-α, IL-6, and IL-1β (Shanghai Enzyme-linked Biotechnology Co., Ltd.) levels were measured according to the manufacturer’s instructions.

Sodium butyrate (Figure 1(a); HPLC ≥ 98.5% purity) was obtained from Sigma-Aldrich (Shanghai) Trading Co. Ltd. (China). TNF-α, IL-1β, IL-6, IL-10, TGF-β, and IL-17 assay kits were purchased from Shanghai Enzyme-linked Biotechnology Co. Ltd. (Shanghai, China). Primary antibodies, including anti-iNOS (Cell Signaling, 13120S), anti-Arg-1 (Cell Signaling, 93668S), anti-ROR gamma(t) (Invitrogen, 14-6988-82), anti-FoxP3 (Beijing Biosynthesis Biotechnology Co. Ltd., Beijing, China, bs-10211R), anti-β-actin (Proteintech, 20536-1-AP), and anti-insulin (Abcam, ab181547). Goat anti-rabbit secondary antibodies (cat. no. 10285-1-AP) were obtained from ProteinTech Group, Inc. (Chicago, IL, USA).

The ELISA kits of TNF-α (1102589), IL-1β (1102590), and IL-6 (1099199) were obtained from Shanghai Enzyme-Linked Biotechnology Co., Ltd. (Shanghai, China). The enzyme plate was sealed after adding the sample and incubated at 37°C for 30 min, washed 5 times with detergent, and incubated at 37°C for 30 min with the HRP labeled enzyme labeled reagent. The plate was washed 5 times again, and the color reagent was applied 20 min later and the reaction was stopped by adding the termination solution. OD at 450 nm was analyzed.

Naringenin (NRG, purity > 98%) was purchased from the Aladdin Bio-Chem Technology Co., Ltd. (Shanghai, China), and D-α-tocopherol polyethylene glycol succinate (TPGS; batch number: 20121203) was purchased from Xi′an Healthful Biotechnology Co., Ltd. (Xi’an, China). Pepsin and pancreatin were purchased from Shanghai Macklin Biochemical Co., Ltd. (Shanghai, China). The KH₂PO₄ was purchased from Sinopharm Chemical Reagent Co., Ltd. (Beijing, China). The montelukast sodium tablets were purchased from Merck Sharp and Dohme Ltd. (Cramlington, Northumberland, UK). The lipopolysaccharide (LPS) and capsaicin were obtained from the Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China), and thyroxine was purchased from Shenzhen Wenle Biological Technology Co., Ltd. (Shenzhen, China). The IL-6, CRP, MDA, and SOD enzyme-linked immunosorbent assay (ELISA) kits were obtained from Shanghai Enzyme-linked Biotechnology Co., Ltd. (Shanghai, China). All reagents and solvents were used without further purification.

ELISA kit detected the concentration of IL-1β, IL-6, MPO and TNF-α (Shanghai Enzyme-linked Biotechnology Co., Ltd., China). According to the instruction of ELISA kit, the standard (50 μL) and sample (40 μL Sample diluent and 10 μL sample) were added to the standard holes and sample holes of microtiter plate, respectively. The culture plates were incubated at 37°C in 5% CO₂ for 30 min and then rinsed with washing solution. Then 50 μL enzyme-labeled reagent was added into each well, and the samples were incubated for 30 min and washed. Then 50 μL color developer A and color developer B were added, mixed, and stained in the dark for 10 min. After the reaction was stopped by adding stop solution, the absorbance density (OD) was measured at 450 nm by enzyme-labeled instrument (Molecular, CA, USA).