Journal: Toxics
Article Title: Mercury Chloride Impacts on the Development of Erythrocytes and Megakaryocytes in Mice
doi: 10.3390/toxics9100252
Figure Lengend Snippet: HgCl 2 does not impact the production of EPO, but increases the expression of EPOR in EMPs in B10.S mice. EPO protein in the serum and BM, EPO mRNA in the kidney, and EPOR in EMPs in the BM in B10.S mice treated with 50 μM HgCl 2 or the control for 4 w were measured. FACS-purified EMPs from the BM of B10.S mice were treated with HgCl 2 or vehicle or EPO in vitro, and the Jak2/STAT5 signaling pathway and differentiation of EMPs were measured thereafter. ( A ) EPO protein (IU/L) in the serum of B10.S mice treated with 50 μM HgCl 2 or water for 4 w. ( B ) EPO protein (IU/g protein) in the BM of B10.S mice treated with 50 μM HgCl 2 or water for 4 w. ( C ) EPO mRNA (fold change) in the kidney of B10.S mice treated with 50 μM HgCl 2 or water for 4 w. ( D ) The expression of EPOR (MFI) in EMPs in the BM of B10.S mice treated with 50 μM HgCl 2 or water for 4 w. ( E ) The expression of EPOR mRNA (fold change) in FACS-purified EMPs from the BM of B10.S mice treated with 50 μM HgCl 2 or water for 4 w. ( F ) EMPs were purified from the BM of control B10.S mice, and the FACS-purified EMPs were treated thereafter with EPO or vehicle in vitro for 16 h. After that, the expression of p-Jak2 (MFI) in EMPs was measured. ( G ) EMPs were purified from the BM of control B10.S mice, and the FACS-purified EMPs were treated thereafter with EPO or vehicle in vitro for 16 h. After that, the expression of p-STAT5 (MFI) in EMPs was measured. ( H ) EMPs were purified from the BM of B10.S mice treated with 50μM HgCl 2 or water for 4 w, and the EMPs were treated thereafter with EPO or vehicle in the presence or absence of the Jak2 inhibitor fedratinib for 16 h in vitro. After that, the expression of p-STAT5 (MFI) in the EMPs was measured. ( I ) EMPs were purified from the BM of control B10.S mice, and the FACS-purified EMPs were treated thereafter with HgCl 2 and/or EPO or vehicle in vitro for 16 h. After that, the expression of p-Jak2 (MFI) in EMPs was measured. Each dot represents one mouse or one sample, and a total of 5 to 12 mice or samples were used for each group. Asterisk indicates a significant difference compared to the counterpart control group. p < 0.05 was considered as the level of significant difference.
Article Snippet: Ab (clone) and fluorescein used to stain progenitors, erythroblasts and leukocytes included lineage cocktail (Lin, CD3 (145-2C11), CD11b (M1/70), B220 (RA3-6B2), Gr-1 (RB6-8C5) and Ter119 (TER-119))-FITC or biotin, c-Kit-APC (2B8), Scal-1-PerCP-Cy5.5 (D7), CD150-PE-Cy7 (TC15-12F12.2), CD150-APC (TC15-12F12.2), CD41-FITC (MWReg30), CD41-PE (MWReg30), CD105-PB (MJ7/18); CD105-PE-Cy7 (MJ7/18), CD71-PE-Cy7 (RI7217), CD45-PE-Cy7 (30-F11), CD11b-PerCP-Cy5.5 (M1/70), Ly6G-PE (1/A8), F4/80-PE-Cy7 (BM8), Ki67-PE (16A8), p-STAT1-PE (pY701); p-STAT3-PE (Tyr705), streptavidin (SA)-PB, SA-APC-Cy7, anti-rabbit Ab-PE, anti-rabbit Ab-FITC and anti-rabbit Ab-APC (Biolegend, San Diego, CA, USA), and rabbit anti-mouse p-Jak1 (Tyr1022) (Nanjing Jiancheng, China), rabbit anti-mouse p-Jak2 (Tyr1007/1008), rabbit anti-mouse p-STAT5 (Tyr694), and rabbit anti-mouse EPO receptor (EPOR) (Beyotime Biotechnology, China), and rat anti-mouse CD16/32 (Fc block, 2.4G2) (BD Biosciences, San Diego, CA), and 4’6-diamidino-2-phenylindole (DAPI) (Sigma, St Louis, MO, USA).
Techniques: Expressing, Purification, In Vitro
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