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Figure 4. Intravenous (IV) injection of human bone mesenchymal stem cell-derived extracellular vesicles (HBMSC-EV) did not alter hepatic inflammatory status, fibrosis, or proliferation. (A) Hematoxylin and eosin staining, Masson-Trichrome staining, and proliferating cell nuclear antigen (PCNA) immunofluorescence did not demonstrate differences in leukocyte infiltration, fibrosis, and cellular proliferation, respectively. Scale bar = 100 µm. (B) Quantification of % fibrosis on Masson-Trichrome staining was not significantly different (P = 0.8167) between the IV control (IV-C) and IV HBMSC-EV (IV-EV) injection groups. Statistical analysis was performed using the Shapiro-Wilk and unpaired t-test. (C) Immunoblotting showed no differences in major regulators of inflammatory and fibrosis pathways, such as tumor necrosis factor-alpha (TNF-α), transforming growth factor-β (TGF-β), and SMAD 2/3. Cell proliferation regulator mammalian target of rapamycin (mTOR) expression was increased in the IV-EV group, but this finding is difficult to interpret given the complexity of mTOR regulation and the negative PCNA findings. Oxidative protein expression changes are difficult to interpret as well, given the lack of hepatic oxidative injury in this model.