Bcl 2

Rat hippocampal tissues were washed 2–3 times with pre-chilled PBS, cut into small pieces and homogenized by adding ten times the volume of tissue in RIPA lysis buffer (Beyotime Institute of Biotechnology, China). Then the supernatant was collected by centrifugation and homogenization (12,000 rpm, 4°C, 10 min), which is the total protein of rat hippocampal tissue. The BCA protein assay kit (AS1086, ASPEN, Wuhan, China) was used for the quantification of total proteins. Samples were separated by SDS PAGE and electro-transferred onto PVDF (0.45 um) membranes (Servicebio,G6015-0.45, Wuhan, China) after activation with methanol. The membranes were then incubated overnight at 4°C with specific primary antibodies:CB1 (Servicebio,GB111214), PI3K(BIOSS,BSM-33219M), P-PI3K (BIOSS,bs5570R), AKT (Servicebio,GB11689), P-AKT (Affinity,AF0908), STAT3 (Servicebio,GB11176), P-STAT3 (RuiyingBiological, RLP0250), Bcl2 (Servicebio, GB113375), BAX (Servicebio, GB11690),ACTIN(Servicebio, GB12001). The membranes were washed three times with TBST for 10 min each. After washing, the membranes were incubated with HRP anti-conjugated secondary antibody (Servicebio, GB23303) for 1 h. The membranes were washed in the same manner as described above. Membranes were scanned using the Fluor Chem FC3 system (Protein Simple, United States).

The piglet SCs was obtained from the cell bank of BLUEFBIO (Shanghai, China). ZEA and DON were obtained from Sigma-Aldrich (St. Louis, MO, USA). 4-PBA was obtained from Target Molecule (Boston, MA, USA). Dulbecco’s modified eagle medium (DMEM) high-glucose medium was purchased from Basalmedia (Shanghai, China). Fetal Bovine Serum (FBS) was purchased from Excell Bio (Suzhou, China). Cell Counting Kit-8 (CCK-8) and bicinchoninic acid kit were purchased from Sparkjade (Shandong, China). Supper RIPA Lysis Buffer was purchased from Coolaber (Beijing, China). The apoptosis detection kit containing Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI) was obtained from Yeasen Biotech Co, Ltd. (Shanghai, China). The mitochondrial membrane potential assay kit with JC-1 and the calcein/PI cell viability/cytotoxicity assay kit were obtained from Beyotime Biotechnology. The digital slide scanner was from 3DHISTECH (Budapest, Hungary). Antibodies to ACTIN, Bcl-2, Bid Caspase-9, Drp1, MFN2, and P53 were purchased from Servicebio (Wuhan, China). The anti-Cyt C antibodies were purchased from HuaBio (Hangzhou, China).

Formalin-fixed paraffin-embedded tissue sections were first baked and deparaffinized. The tissue sections were then placed in citric acid antigen repair buffer (pH 6.0) in a microwave oven for antigen repair. The tissue was placed in 3% hydrogen peroxide solution for 25 min to block endogenous peroxidase and then closed at room temperature for 30 min by adding 3% BSA dropwise to cover the tissue uniformly. primary antibodies (BAX, Servicebio, 50599-2-Ig, 1:300; Bcl-2, Servicebio B13439, 1:200) were added and stored overnight at 4°C and secondary antibodies [HRP IgG (H + L), Servicebio GB23302, 1:200] and incubated at room temperature for 50 min. Then freshly prepared DAB color development solution was added dropwise and the color development time was controlled under the microscope. The nuclei were re-stained with hematoxylin and then sealed by dehydration. The average optical density value was calculated for the relevant area of each section using Image-Pro-PLUS software, and the magnitude of the value was the degree of expression of the relevant subunit protein.

Poly(ε-caprolactone) (PCL; Mn = 100 000 g/mol) was purchased from Changchun SinoBiomaterials Co., Ltd. (Changchun, China). CIS and hexafluoro-isopropyl alcohol (HFIP) were purchased from Aladdin (Shanghai, China). AgNPs (d = 15–20 nm) were provided by Kunming Guiyan Pharmaceutical Co., Ltd. (Kunming, China). Nanjing Microtech Co., Ltd. (Nanjing, China) provided uncovered self-expanding metal stents (SEMS; 20 × 8 mm²). The Shanghai Cell Center of the Chinese Academy of Sciences provides NSCLC cells (A549). Bacterial species (Pseudomonas aeruginosa-ATCC 27853; Staphylococcus aureus-ATCC 25923) and fungi (Candida albicans-ATCC 10231) are provided by the China Type Culture Collection and Shanghai Institutes for Biological Sciences (SIBS). RPMI-1640 medium, fetal bovine serum (FBS), and penicillin/streptomycin were supplied by Sigma-Aldrich, Inc. (China). Thermo Fisher Scientific provided the Live/Dead BacLight bacterial viability kit, the Live/Dead cell viability kit, and the Annexin V-FITC apoptosis detection kit. Servicebio Biotechnology Co., Ltd. (Wuhan, China) provided the Ki67, TUNEL, Bax, caspase-3, Bcl-2, and PCNA antibodies.

The cells of each group were collected and protein was extracted. Protein extracts were prepared in the lysis buffer. The lysis buffer was a combination of RIPA, complete, phosstop, and 10 mM PMSF. A BCA™ protein assay kit (Thermo Fisher Scientific, Rockford, USA) was used to estimate the protein concentration. The protein samples were separated by SDS-PAGE and transferred onto polyvinylidene difluoride membrane (EMD Millipore). The membranes were then blocked with 5% nonfat milk or bovine serum albumin (for detecting phosphorylated protein) in TBST (Tris-buffered saline with 0.1% Tween-20) for 2 h at room temperature. Later, the membranes were incubated overnight with cleaved-caspase-3, Bid, Bcl-2, ERK1/2, p-ERK1/2, and GAPDH (diluted in dilution for primary antibodies, Servicebio, Wuhan, China) at 4°C. After washing with TBST for three times, the membranes were exposed to the corresponding secondary antibodies (1:5,000) at room temperature for 1 h. The target proteins were visualized with an ECL kit.