Fig. 5: TLR8 regulates erythropoiesis in erythroid cell-intrinsic manner.

a Schematic illustrating the inducer-incubator assay workflow. Numbers of colonies generated from incubator cells collected on days 4 (b) and 8 (c) of erythroid differentiation (n = 4 biologically independent experiments). d Representative flow cytometry (left) showing the percentage of CD71⁺CD235a⁺ erythroid precursor cells of incubator cells on day 11 of erythroid differentiation. Statistics of the fold changes are shown on the right (n = 3 biologically independent experiments). e Cell cycle analysis of incubator cells on day 11 of erythroid differentiation (n = 3 biologically independent experiments). f Apoptosis of incubator cells (annexin-V⁺ cells) on day 11 of culture (n = 3 biologically independent experiments). g Schematic illustrating experimental design. After 8 days of erythroid differentiation, FACS-sorted CFU-E cells were harvested and subjected to colony-forming assays with 10 μM VTX2337 or 10 μM CUCPT8m treatment. h Numbers and i percentages of colonies generated from FACS-sorted CFU-E cells under the indicated conditions (n = 4 biologically independent experiments). j Schematic illustrating experimental design. Single CFU-E was sorted and seeded into 96-well plates on day 5 of erythroid differentiation. These purified CFU-E cells were further induced differentiation toward erythroid lineage for 5 days with 10 μM VTX32337 or 10 μM CUCPT8m treatment. k Cell number on day 5 of erythroid differentiation upon vehicle, 10 μM VTX2337 or 10 μM CUCPT8m treatment (n = 88 for vehicle, n = 65 for VTX2337, n = 75 for CUCPT8m from biologically triplicate). The representative flow cytometry (l) showing percentages of CD71⁺CD235a⁺ erythroid cells after treatment with vehicle or 10 μM CUCPT8m for 5 days during erythroid differentiation and its statistics (m) (n = 3 biologically independent experiments). Data are mean ± SEM; statistical significance was determined using the unpaired two-tailed Student’s t-test. *P < 0.05, **P < 0.01, ****P < 0.0001, ns = not significant. Source data are provided as a Source Data file.