Human HaCaT keratinocytes were maintained at 37 °C and 5% carbon dioxide in MEM medium (Procell, Wuhan, China) supplemented with a 1% antibiotic mixture in the presence of 15% FBS (Gibco, New York, NY, USA). Human SZ95 sebocytes [35 (link)] were maintained at 37 °C and 5% carbon dioxide in DMEM medium (Gibco, New York, NY, USA) supplemented with a 1% antibiotic mixture in the presence of 10% FBS (Gibco, New York, NY, USA).
Mem medium
Manufactured by Procell
Sourced in China
MEM medium is a cell culture medium formulation designed to support the growth and maintenance of various cell types in vitro. It provides essential nutrients and components required for cell survival and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Dmem medium, by Thermo Fisher Scientific (1 mentions)
Dmem medium, by Solarbio (1 mentions)
8305c cell, by Procell (1 mentions)
Grp94, by Abcam (1 mentions)
Tsg101, by Abcam (1 mentions)
Fetal bovine serum (fbs), by Procell (1 mentions)
Bv2 cell line, by Procell (1 mentions)
Myco 5a medium, by Procell (1 mentions)
Rpmi 1640 medium, by Procell (1 mentions)
Caki 1, by Procell (1 mentions)
Penicillin streptomycin liquid, by Thermo Fisher Scientific (1 mentions)
Dmem medium, by Procell (1 mentions)
Dmem f12 medium, by Procell (1 mentions)
Mda mb 231, by Procell (1 mentions)
Mcf 10a, by Procell (1 mentions)
Mcf 7, by Procell (1 mentions)
Leibovitz s l 15 medium, by Procell (1 mentions)
Infinite 200 pro microplate reader, by Tecan (1 mentions)
Cell counting kit 8 cck 8 assay, by AbMole (1 mentions)
8 protocols using mem medium
1
Cultivation of Human Keratinocytes and Sebocytes
2
Cell Line Authentication and Culture
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The 8305C cell, an ATC cell, was purchased from Procell Life Science & Technology Co., Ltd. in September 2021, and was authenticated using STR method. The BCPAP cell, a PTC cell, was obtained from Fuheng Biology in January 2022, and was authenticated using STR method. The 8305C cell and the BCPAP cell were authenticated again using STR method in April and May 2023, respectively. The 8305C cell was cultured in a MEM medium (Procell, China) containing 10% fetal bovine serum (FBS, Gibco, USA) whereas the BCPAP cell was cultured in a DMEM medium (Solarbio, China) containing 10% FBS (Gibco, USA). TC cells were stained using the trypan blue, and the number of viable cells was counted with a hemocytometer.
3
Exosome Isolation from Hyperglycemic HaCaT Cells
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When the HaCaT cells reached 50% confluence, exosome-free serum was used for culture, and normal glucose (5.5 mmol/L) and high glucose (33.3 mmol/L) aliquots were added to the MEM medium (Procell) of the control group and treatment group, respectively. After 48 hours of culture, the collected cell supernatant was centrifuged to remove cell debris (300 × g for 10 min, 3000 × g for 20 min), and the supernatant was filtered through a 0.22-μm pore-size membrane (Guangzhou Jet Bio-Filtration Co., Ltd) for subsequent experiments. Then, the supernatant was centrifuged at 10000 × g for 30 min and transferred to a new ultrafiltration tube. This supernatant was centrifuged at 110000 × g for 70 min, and the pellet was resuspended in PBS (Guangzhou Jet Bio-Filtration Co., Ltd) after the resulting supernatant was removed. After repeated centrifugation at 110000 × g for 70 min, the supernatant was removed, and the pellet was resuspended in an appropriate amount of PBS for subsequent exosome characterization. All operations were carried out at 4°C. The expression of the CD9 (1:1000; Abcam, UK), TSG101 (1:1000; Abcam) and Grp94 (1:1000; Abcam) protein markers was detected by western blotting to confirm the exosome isolations were successful.
4
Collagen Sponge Delivery of Lipid Nanoparticles
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The collagen lyophilized sponge was kindly provided by Tianjin Shiji Kangtai Biomedical Engineering Company, Ltd; BV2 cell line, fetal bovine serum and MEM medium were purchased from Wuhan Procell Life Science & Technology Company; Egg yolk lecithin PL-100M (Injection Grade, Phosphatidylcholine≥98%), purchased from AVT (Shanghai) Pharmaceutical Technology Company; 1,2-propanediol, 99%, AR, purchased from Shanghai Macklin Biochemical Technology Co.; Fluorescein Isothiocyanate (FITC), 97%, Biotech Grade, BR, purchased from Shanghai Macklin Biochemical Technology Co.; Coumarin-6, 98%, HPLC grade, purchased from Shanghai Aladdin Biochemical Technology Co. All other chemicals were of analytical grade and were purchased from the Shanghai Aladdin Biochemical Technology Company. Double distilled water, obtained from a water purification system (Nano-pure Infinity, Barnstead International. Dubuque. IA), was used to prepare all solutions.
5
Clear Cell Renal Cell Carcinoma Cell Lines
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Clear cell renal cell carcinoma cell lines 786-O (male karyotype), 769-P (female karyotype), Caki-1 (male karyotype) and human renal proximal tubule epithelial cells HK2 (male) were obtained from FDCC (Shanghai, China). 786-O and 769-P cells were cultured in RPMI-1640 medium (Procell), Caki-1 was cultured in Myco-5A medium (Procell), while HK2 cells were cultured in MEM medium (Procell). All media was supplemented with 10% fetal bovine serum and 1% antibiotics. Cells were maintained in 5% CO2 at 37°C.
6
Breast Cancer Tissue and Cell Line Characterization
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A total of 66 pairs of BC tissue samples and adjacent normal tissue samples were obtained from BC patients at the Nanyang Second General Hospital, and all tissues were stored at -80℃. The clinical characteristics of these patients were shown in Table 1 . All patients had not undergone any chemotherapy and radiotherapy before surgery. The experiment here got approval from the Ethics Committee of Nanyang Second General Hospital.
Clinical characteristics of patients with breast cancer (n = 66)
| Characteristic | Cases |
|---|---|
| Age | |
| < 50 | 38 |
| ≥ 50 | 28 |
| Histologic grade | |
| I | 3 |
| II | 45 |
| III | 18 |
| Lymph node metastasis | |
| Yes | 35 |
| No | 31 |
| T stage | |
| T1/T2 | 52 |
| T3/T4 | 14 |
| N stage | |
| N0/N1 | 39 |
| N2/N3 | 27 |
7
RvD1 Effects on MG63 Osteosarcoma Viability
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The human osteoblastic osteosarcoma cell line MG63 cells (CL‐0157, Procell, Wuhan, China) were cultured in MEM medium (Procell) supplemented with 10% fetal bovine serum (FBS; Abwbio) at 37°C in a high humidity environment containing 5% CO2. The medium was replaced every 2 days. MG63 cells with 80% confluence were trypsinized from culture flasks.
Cell viability was measured with the Cell Counting Kit‐8 (CCK‐8) assay (AbMole). Briefly, MG63 cells were seeded at a density of 1 × 103 cells/well in a 96‐well plate and cultured in 100 μL of MEM medium containing FBS for 24 h. To examine the effect of RvD1 (Cayman Chemical) on cell viability, MG63 cells were treated with RvD1 (10 ng/ml, 100 ng/ml) for 24 and 48 h. Untreated cells served as control. After treatment, 10 μL CCK‐8 was added to each well, and cells were incubated for another 2 h with 5% CO2 at 37°C. Then, absorbance was read at 450 nm using an infinite 200Pro microplate reader (Tecan). The experiment was performed in sextuplicate. The control group was defined as 100% cell viability, and other groups were normalized to this value.
Cell viability was measured with the Cell Counting Kit‐8 (CCK‐8) assay (AbMole). Briefly, MG63 cells were seeded at a density of 1 × 103 cells/well in a 96‐well plate and cultured in 100 μL of MEM medium containing FBS for 24 h. To examine the effect of RvD1 (Cayman Chemical) on cell viability, MG63 cells were treated with RvD1 (10 ng/ml, 100 ng/ml) for 24 and 48 h. Untreated cells served as control. After treatment, 10 μL CCK‐8 was added to each well, and cells were incubated for another 2 h with 5% CO2 at 37°C. Then, absorbance was read at 450 nm using an infinite 200Pro microplate reader (Tecan). The experiment was performed in sextuplicate. The control group was defined as 100% cell viability, and other groups were normalized to this value.
8
Establishing S100A10 Knockdown HeLa Cells
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The human HeLa cell line (Procell, China) was cultured in MEM medium (Procell) supplemented with 10% fetal bovine serum (FBS) (Gemini, USA) in a humidified incubator at 37°C with 5% CO2. To establish stably transfected cell lines with downregulated S100A10 expression, HeLa cells were infected with lentiviral vectors containing either S100A10 short-hairpin RNA (sh-S100A10) or a corresponding negative control (sh-NC), which were synthesized by Genechem (China). The transfection procedure was conducted in HeLa cells using the Polybrene (Hanbio, China). After the stably transfected cells were selected using puromycin according to the manufacturer's protocol, they were collected and utilized for subsequent experiments. The transfection efficiency was assessed through RT-qPCR analysis.
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