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S. Yang et al.
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Figure 2. TRPV4 mediates GDF15-induced bone marrow adipocyte remodeling. (A) Bone marrow (BM) adipocytes were co-cultured with leukemia cell lines (THP-1, K562, HL-60) or leukemia cells and anti-GDF15 neutralizing antibody (200 ng/mL) for 4 days. The protein of TRPV4 was detected using western blot analysis. (B) The effect of different concentrations (100 ng, 200 ng, 500 ng) of recombinant human GDF15 (rhGDF15) on the expression of TRPV4 protein for 4 days was analyzed by western blot. (C) Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) was performed to analyze the expression of TRPV4 mRNA after the addi- tion of 200 ng rhGDF15 in BM adipocytes for 2, 4, 6, 8 days. (D) BM adipocytes were treated with dimethylsulfoxide (Ctr), rhGDF15 (200 ng/mL) or rhGDF15 (200 ng/mL) and 4α-phorbol 12,13-didecanoate (4αPDD, 0.25 mg/mL) for 6 days. Adipocytes were stained by oil red O. All images were at a magnification of 200×. (E) The number and average area of adipocytes from the indicated groups were measured using Image-Pro-Plus 5.1. (F) The content of lipid droplets in the indicated groups was detected by optical density values. (G) RT-qPCR was used to analyze HSL and ATGL mRNA in adipocytes from the indicated groups on the fourth day. (H) The content of free fatty acids in the supernatant of BM adipocytes from each group was detected using a colorimetric method. β-actin protein was used as an inter- nal control for the western blot analysis. Three independent experiments were performed. **P<0.01, *P<0.05.
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