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. 2023 Dec 12;300(1):105556. doi: 10.1016/j.jbc.2023.105556

Figure 1.

Figure 1

Differentiation of macrophages from porcine EPSCs.A, schematic overview of macrophages differentiation protocols from pEPSCs. B, representative bright-field images were shown for the different morphologies generated on day 0 (undifferentiated pEPSCs), day 1 (PS), day 2 (ME), day 5 (HE), day 9 (EMPs), and day 11 (pPSCdMs). Scale bars represent 100 μm. C, the expression levels of pluripotent-related genes in pEPSCs, PS, and ME were detected by RT-qPCR. Data indicate mean ± s.d. The p values were calculated by using two-tailed t-tests. n = 3. D, the expression levels of PS-related and mesodermal genes in pEPSCs, PS, and ME were detected by RT-qPCR. Data indicate mean ± s.d. The p values were calculated by using two-tailed t-tests. n = 3. E, the expression levels of HE-related genes in pEPSCs and HE were detected by RT-qPCR. Data indicate mean ± s.d. The p values were calculated by using two-tailed t-tests. n = 3. F, the expression levels of EMP-related genes in pEPSCs, EMPs, and HPCs isolated from porcine bone marrow were detected by RT-qPCR. Data indicate mean ± s.d. The p values were calculated by using two-tailed t-tests. n = 3. G, the expression levels of macrophage-related genes in pEPSCs, pPSCdMs, and PAMs were detected by RT-qPCR. Data indicate mean ± s.d. The p values were calculated by using two-tailed t-tests. n = 3. H, FACS analysis of stage-specific markers on day 0, day 2, day 5, day 9, and day 11 during differentiation from pEPSCs. The percentages of positive populations were shown in red. I, RT-qPCR expression profile analysis of every differentiation stage for pluripotency markers (POU5F1, SOX2, and NANOG), PS markers (TBXT and MIXL1), ME marker (KDR), HE markers (CD31, CD34, RUNX1 and CDH5), EMPs markers (RUNX1, CD45, and ITGA2B), and pPSCdMs markers (CSF1R, CD14, CD163, and PU.1). Data indicate mean ± s.d. n = 3. EMPs, erythromyeloid progenitor cells; EPSCs, expanded potential stem cells; FACS, fluorescence-activated cell sorting; HE, hematopoietic endothelium; HPCs, hematopoietic progenitor cells; ME, mesoderm; PAMs, porcine alveolar macrophages; pEPSCs, porcine expanded potential stem cells; pPSCdMs, pPSC-derived macrophages; PS, primitive streak; RT-qPCR, quantitative reverse transcription PCR.