Fig. 3

Exosomes derived from colon cancer cells facilitated the expression of NGF in Schwann cells via miR-21-5p. A. The expression of NGF in Schwann cells was increased when added colon cancer-CM was by Western blot and qRT-PCR. B. ELISA assay showed that the expression of NGF in Schwann cells was augmented when colon cancer-CM was added. C. Exosomes were labeled with PKH26, and sNF96.2 cells were transfected lentivirus with stable expression of GFP. Incubation of sNF96.2 cells with PKH26-labeled exosomes for 6 h. D. Western blot and qRT-PCR showed that the expression of NGF in Schwann cells was increased when added colon cancer-Ex. E. ELISA assay showed the expression of NGF in Schwann cells was augmented when added colon cancer-Ex. F. Venn diagram indicated that five miRNAs were highly expressed in exosomes of colon cancer serum and tissues from the GEO and EVmiRNA database. G. The qRT-PCR showed the expression of five miRNAs in the exosomes of FHC, SW480, and HCT116 cells. H. The qRT-PCR showed the expression of miR-21-5p in sNF96.2 cells treated with exosomes. I. The qRT-PCR analysis of miR-21-5p in the CM of FHC, SW480, and HCT116 cells was treated with RNase R (3 U/μg) alone or combined with Triton X-100 (0.1%) for 20 min. J. Western blot and qRT-PCR showed that inhibition of miR-21-5p in SW480 cells significantly blocked the increased expression of NGF in Schwann cells incubated with the exosome of the SW480 cells. K. Western blot and qRT-PCR showed that the expression of NGF was conspicuously increased or decreased in Schwann cells upon miR-21-5p overexpression or knockdown in Schwann cells, respectively. CM: conditioned medium. Ex: Exosomes. Inh: inhibitor. All data were revealed as means ± standard deviation (SD) for no less than three independent experiments. Significant P values showed as *P < 0.05, **P < 0.01, ***P < 0.001. n.s means the difference was not significant