hDPSCs (5 × 104 cells) were cultured in an osteogenic differentiation medium containing l -ascorbic acid-2-phosphate (0.2 mM; Fujifilm Wako Pure Chemical), beta-glycerophosphate (5 mM; Fujifilm Wako Pure Chemical), dexamethasone (1 nM; Fujifilm Wako Pure Chemical), and BMP-2 (100 ng/ml; Fujifilm Wako Pure Chemical) for 21 days. Mineralized nodules were stained with Alizarin Red S (Fujifilm Wako Pure Chemical).
Bmp 2
Manufactured by Fujifilm
The BMP-2 is a compact, versatile laboratory equipment designed for a wide range of applications. It functions as a bench-top centrifuge, capable of separating liquids and particulates through centrifugal force. The device is suitable for various sample preparation tasks in research and clinical settings.
Automatically generated - may contain errors
Lab products found in correlation
Dexamethasone (dex), by Fujifilm (2 mentions)
L ascorbic acid 2 phosphate, by Fujifilm (2 mentions)
Beta glycerophosphate, by Fujifilm (2 mentions)
Alizarin red s, by Fujifilm (2 mentions)
Dmem low glucose, by Fujifilm (1 mentions)
Labassay alp kit, by Fujifilm (1 mentions)
Dulbecco modified eagle medium (dmem), by Fujifilm (1 mentions)
5 protocols using bmp 2
1
Osteogenic Differentiation of hDPSCs
2
Mouse Mesenchymal Cell Differentiation
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Mouse mesenchymal ST-2 cells (ATCC) were seeded in multi-well plates (2 × 104 cells/cm2) and cultured in low-glucose DMEM (FUJIFILM Wako) supplemented with 10% FBS and 1% PS at 37 °C in the presence and absence of BMP-2 (FUJIFILM Wako), with and without Myo-EVs for 72 h. Each Myo-EV was diluted with culture medium based on the quantified BCA assay concentration and added at a final concentration of 10 µg/mL. For the control group, equal amounts of culture medium as the added Myo-EVs were used.
3
Osteogenic Differentiation of DPSCs
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DPSCs (2 × 105 cells) were seeded in 48-well cell culture plates and cultured in an osteogenic differentiation medium containing L-ascorbic acid-2-phosphate (0.2 mM; Wako Pure Chemical Industries), beta-glycerophosphate (5 mM; Wako Pure Chemical Industries), dexamethasone (1 nM; Wako Pure Chemical Industries), and bone morphogenic protein-2 (BMP-2, 100 ng/ml; Wako Pure Chemical Industries) for 25 days. Calcified nodules were stained with Alizarin Red S (Wako Pure Chemical Industries) after methanol fixation for 1 min at room temperature. The volume of mineralized nodules stained with Alizarine Red S was measured by ImageJ software (https://imagej.nih.gov/ij/ ).
4
Mouse Osteoblast Differentiation and Isolation
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The mouse bone marrow-derived stromal cell line, ST2 (RIKEN, Tsukuba, Japan) was cultured in RPMI 1640 with 10% FBS and 1% penicillin-streptomycin. ST2 cells were differentiated into osteoblastic cells with 200 ng/mL BMP-2 (FUJIFILM Wako Chemicals) for 3 days. Mouse osteoblasts were isolated from the neonatal calvariae of 3- to 5-day-old C57BL/6J male mice as previously described [29 (link)]. Briefly, the calvariae of neonatal mice were digested 4 times with 1 mg/mL collagenase I and 0.25% trypsin at 37°C for 20 min with gentle agitation. Cells from the second, third, and fourth digestions were collected. The resultant cells were cultured in α-MEM supplemented with 10% FBS and 1% penicillin/streptomycin. Isolated osteoblasts exhibited ALP activity and formed mineralized nodules. Osteoblasts at passage 2 were used in further experiments. Cells were cultured at 37°C in a humidified atmosphere containing 5% CO2.
5
Mesenchymal Cell Line ST2 Osteogenesis
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ST2 cells (RIKEN, Tsukuba, Japan), which represent a mouse mesenchymal cell line, were maintained in lowglucose Dulbecco's modified Eagle's medium (DMEM, FUJIFILM Wako, 5.5 mM glucose) with 10% FBS and 1% penicillin-streptomycin. ST2 cells were cultured until they were confluent and then treated with or without Myo-EVs, BMP-2 (FUJIFILM Wako), advanced glycation end product 3 (AGE3) and high glucose (25 mM). AGE3 was prepared as previously described [25] . Moreover, alkaline phosphatase (ALP) activity was measured with the Lab Assay ALP kit (FUJIFILM Wako) according to the manufacturer's instructions.
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