Pancreatic enzymes (Sigma, America) without EDTA were used to digest the third passage of rBMSCs, which was followed by centrifugation at 1000 r/min to dispose of the supernatant and three washes with PBS. PBS was applied to the resuspended cells, and the cell density was adjusted to 6000 cells/μL in the tube. Then, antibody markers for CD105, CD34, CD44, and CD45 were added into the tube prior to incubation in a dark room at ambient temperature for half an hour. The cells were fixed with methanol after PBS washing and centrifugation and then maintained overnight in a refrigerator at 4 °C without light exposure. On the next day, the expression levels of CD105, CD34, CD44, and CD45 were measured using flow cytometry (FCM, Partec, Germany).
Pancreatic enzyme
Manufactured by Merck Group
Sourced in United States
Pancreatic enzymes are a set of enzymes produced by the pancreas that play a crucial role in the digestion of food. These enzymes include amylase, lipase, and protease, which help break down carbohydrates, fats, and proteins, respectively, facilitating the absorption of nutrients.
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Lab products found in correlation
Human inflammasomes pcr array, by Qiagen (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Rneasy mini kit, by Qiagen (1 mentions)
Rnase free dnase set, by Qiagen (1 mentions)
Rt2 first strand kit, by Qiagen (1 mentions)
Mcp 1, by RayBiotech (1 mentions)
Rt2 sybr green rox qpcr mastermix, by Qiagen (1 mentions)
Il 10, by RayBiotech (1 mentions)
Sodium caseinate, by Merck Group (1 mentions)
Resveratrol, by Merck Group (1 mentions)
Methanol, by Sinopharm (1 mentions)
Acetonitrile, by Sinopharm (1 mentions)
Dulbecco s modified eagle s medium f12, by Thermo Fisher Scientific (1 mentions)
Collagenase 2, by Merck Group (1 mentions)
Pepsin, by Merck Group (1 mentions)
α amylase, by Merck Group (1 mentions)
Rpmi 1640 cell culture medium, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Sartorius (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Merck Group (1 mentions)
6 protocols using pancreatic enzyme
1
Characterization of rBMSCs by Flow Cytometry
2
Inflammasome Activation in Immune Cells
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LPS and pancreatic enzymes were purchased from Sigma-Aldrich (St. Louis, MO, USA); lymphocyte separation medium was purchased from Jingyang Biological (Tianjin, China); fetal bovine serum (FBS) was obtained from GIBCO (Australia); RPMI-1640 glucose free medium was purchased from Hyclone (Logan, UT, USA); RNeasy Mini Kit, RNase-Free DNase Set, RT2 First Strand Kit, RT2 SYBR® Green ROX qPCR Mastermix, and Human Inflammasomes PCR Array were obtained from Qiagen (Valencia, CA, USA). ELISA kits of human TNF-α, IL-1β, IL-6, IL-10, and MCP-1 were purchased from RayBiotech (Atlanta, USA). Anti-NLRP3 and anti-Caspase-1 antibodies were purchased from Abcam (Cambridge, United Kingdom). Secondary antibodies were obtained from Sigma-Aldrich (St. Louis, MO, USA).
3
Resveratrol Bioavailability Enhancement Protocol
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Resveratrol and sodium caseinate (analytical grade) were purchased from Sigma-Aldrich China Co., Ltd. (Sigma-Aldrich, Shanghai, China). Polyglycerol fatty acid ester (food grade) was purchased from Shandong Binzhou Jinsheng New Material Technology Co., Ltd. (Binzhou, China). Rice bran oil (Jinlongyu, Shenzhen, China) was purchased from the local market. Acetonitrile and methanol (chromatographic grade) were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shenzhen, China). The enzymes, including porcine gastric mucosal pepsin, pancreatic enzymes, and porcine bile extracts, were bought from Sigma-Aldrich (Sigma-Aldrich, Shanghai, China).
4
Regulation of Pituitary TSH Secretion
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Freshly dissected mouse adenohypophyses were digested with 0.1% collagenase II (Sigma, St. Louis, MO, USA) and 0.25% pancreatic enzyme (Sigma). Anterior pituitary cells were harvested and resuspended in Dulbecco’s modified Eagle’s medium/F12 (GIBCO-Invitrogen, Grand Island, NY, USA), supplemented with 10% fetal bovine serum and antibiotics (100 U/mL penicillin G and 100 µg/mL streptomycin). In total, 10 × 105 cells per well were cultivated in a 24-well plate for 12 h at 37 °C and 5% CO2. For each assay, a minimum of 3–4 solitary trials were conducted, and every trial contained 3–4 replicates per treatment condition.
The profound impact of CK1α on TSH synthesis and secretion from anterior pituitary cells was determined using a CK1α inhibitor, D4476 (0, 5, 10, 20, and 40 μM; MCE, Shanghai, China), CK1α agonist, pyrvinium (0, 0.8, 1.25, and 2.5 μM; MCE), or thyrointegrin receptor antagonist (Tetrac; MCE).
The profound impact of CK1α on TSH synthesis and secretion from anterior pituitary cells was determined using a CK1α inhibitor, D4476 (0, 5, 10, 20, and 40 μM; MCE, Shanghai, China), CK1α agonist, pyrvinium (0, 0.8, 1.25, and 2.5 μM; MCE), or thyrointegrin receptor antagonist (Tetrac; MCE).
5
Enzymatic Hydrolysis of Lyophyllum decastes
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Lyophyllum decastes (Fr.) Singer was collected from an agricultural product processing base in Taixing, Jiangsu Province, China. The enzymes, used in this study, including α-amylase (100 U/mg), pepsin (3,000 U/g), and pancreatic enzyme (4,000 U/g), were purchased from Sigma-Aldrich (St. Louis, United States). SCFA standards including acetic, propionic, butyric, valeric, isobutyric, and isovaleric acids were purchased from Aladdin (Shanghai, China). All other chemicals and solvents used were analytical grade.
6
PDAC Cell Line Maintenance Protocol
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We purchased the human PDAC cell lines ASPC-1 and BXPC-3 from American Type Culture Collection (ATCC) and cultured them in complete growth medium according to the manufacturer’s instructions. Cells were maintained in an incubator at 37°C and 5% CO2 and tested for mycoplasma every 3 months (last verified negative date, May 1, 2020). The experiments in this study used cells within ten passages after thawing.
The following reagents were used. RPMI 1640 cell culture medium was purchased from Gibco. Fetal bovine serum (FBS) was obtained from Biological Industries. Pancreatic enzyme and EDTA were acquired from Sigma. Phosphate-buffered saline (PBS) was from DHeLix.
The following reagents were used. RPMI 1640 cell culture medium was purchased from Gibco. Fetal bovine serum (FBS) was obtained from Biological Industries. Pancreatic enzyme and EDTA were acquired from Sigma. Phosphate-buffered saline (PBS) was from DHeLix.
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