Fig. 2

Suppression of tRF-3022b inhibits subcutaneous tumor growth and induces M2 polarization via LGALS1 and MIF. a Representative images of subcutaneous xenograft tumors (n = 4 mice per group). b Tumor volume at indicated time points was calculated and plotted (n = 4 mice per group). c Final weight of subcutaneous xenograft tumors was measured and quantified (n = 4 mice per group). d Transmission electron micrograph (TEM) of exosomes isolated from HCT116 cell (scale bar = 200 nm). e Western blot analysis of TSG101, CD9, and E-cadherin (negative control) in exosomes. Cellular lysates were used as positive loading controls. f mRNA levels of various cytokines and markers in the M0 macrophages were measured by qRT-PCR analysis. g mRNA levels of M2 macrophage markers were measured by qRT-PCR. h Flow cytometry was used to quantify the expression of CD206, an M2 macrophage marker. i Cluster heat map of the DEGs between tRF-3022b-knockdown and negative control HCT116 cells. j Silver SDS-PAGE gel image shows proteins immunoprecipitated by the tRF-3022b probe in HCT116 and RKO cells. k–l Western blot analysis of products from RNA pull-down assays using the tRF-3022b probe and a control probe suggested LGALS1 and MIF in HCT116 and RKO cells. m Protein levels of LGALS1 in CRC cells with tRF-3022b knockdown. n Protein levels of MIF in CRC cells with tRF-3022b knockdown. o Immunofluorescence images of tRF-3022b with LGALS1 and MIF co-staining. Cell nuclei were counterstained with Hoechst. Scale bar: 20 μm. p Diagram outlining discussed hypotheses about the roles of tRF-3022b in the CRC tumor microenvironment. (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001)