Fig. 6: Stimulation by the recombinant human TPO (rhTPO) and phytohemagglutinin-L (PHA-L) leads to the proliferation of MPL-L⁺ VLRA⁺ cells.

a Time points for injection with rhTPO, PHA-L, and the 5-Ethynyl-2’-deoxyuridine (EdU). b Representative sections revealing the number and location of MPL-L⁺ VLRA⁺ cells in the typhlosole after injection, as determined by double-stained immunofluorescence examination. Green fluorescence is associated with anti-VLRA pAb, and red fluorescence is associated with anti-MPL-L pAb. c, d Box plot shows the proportion of MPL-L⁺ VLRA⁺ cells and the mean fluorescence intensity (MFI) of MPL-L within the typhlosoles across varying experimental conditions. The data presented are based on a minimum of three biological replicates per experimental sample: Control (n = 7), PHA-L (n = 9), and rhTPO (n = 7). Statistical significance was determined using one-way ANOVA with a one-sided test approach and Tukey’s adjustment was made for multiple comparisons. e Representative sections revealing the number and location of MPL-L⁺ VLRA⁺ cells in the gill filaments after injection, as determined by double-stained immunofluorescence examination. f-g Box plot shows the proportion of MPL-L⁺ VLRA⁺ cells and the MFI of MPL-L within the gill filaments across varying experimental conditions. The data presented are based on a minimum of three biological replicates per experimental sample: Control (n = 6), PHA-L (n = 4), and rhTPO (n = 4). Statistical significance was determined using one-way ANOVA with a one-sided test approach and Tukey’s adjustment was made for multiple comparisons. h Flow cytometric analysis of MPL-L + VLRA+ cells before and after PHA-L and rhTPO immunostimulation. i Representative sections reveal the number and location of proliferating cells (EdU, brown) in the typhlosoles and gill filaments after the injection of rhTPO with experiments performed at least three times. Arrows indicate the characteristic proliferating cells.