Therapeutic effect of exosomes derived from hepatocyte-growth-factor-overexpressing adipose mesenchymal stem cells on liver injury.

Adipose mesenchymal stem cell-derived exosomes (ADMSC-Exo) are a new strategy for the treatment of liver injury. However, mesenchymal stem cells (MSCs) exert therapeutic effects mainly by secreting hepatocyte growth factor (HGF). Therefore, we investigated the role of exosomes derived from ADMSC that overexpress HGF (ADMSCHGF-Exo) on liver injury. MATERIAL AND METHODS: ADMSCs were isolated from young BALB/c female mice. Then exosomes derived from ADMSC transfecting negative control (ADMSCNC-Exo) and HGF overexpression (ADMSCHGF-Exo) were isolated and identified by quantitative polymerase chain reaction (qPCR), flow cytometry, western blot, transmission electron microscope and Nanosight particle tracking analysis. These exosomes were injected into male mice via tail vein after inducing liver injury by administering 40% carbon tetrachloride (CCl4)-olive oil twice a week (3 mL/kg, subcutaneously) for 6 weeks. Liver injury and liver collagen fiber accumulation were determined by histopathological analysis. Then, the levels of serum liver function indexes (alanine aminotransferase, aspartate aminotransferase, albumin, total bilirubin), hepatocyte-specific markers (albumin, cytokeratin-18 and hepatocyte nuclear factor 4α), hepatic fibrosis-related proteins (α-smooth muscle actin and collagen I) and Rho GTPase (cell division cycle 42 and ras-related C3 botulinum toxin substrate 1) were determined by Enzyme-linked immunosorbent assay (ELISA), immunohistochemistry, Western blot and qPCR. RESULTS: ADMSCs were identified by high expression of CD105 and CD44 molecules and low expression of CD45 and CD34. ADMSCs-Exo, ADMSCNC-Exo and ADMSCHGF-Exo transfected cells had similar expression of exosome-specific membrane proteins (CD63, CD81 and CD9). Mice with CCl4-induced liver injury exhibited abnormal serum liver function indexes, altered expression of hepatocyte-specific markers, hepatic fibrosis-related proteins and Rho GTPase protein as well as histopathological changes and collagen fiber accumulation in the liver. These changes were reversed by ADMSC-Exo, ADMSCNC-Exo and ADMSCHGF-Exo administration with ADMSCHGF-Exo displaying the most significant impact. CONCLUSIONS: ADMSCHGF-Exo exerted a hepatoprotective effect in mice with experimental liver injury by alleviating hepatic fibrosis and restoring liver function.

Effect of IL-8 on hepatocellular carcinoma-associated metastasis by targeting MMP9 in mice.

Background: Interleukin-8 (IL-8) and matrix metallopeptidase 9 (MMP9) are overexpressed in hepatocellular carcinoma (HCC), and both are related to tumor metastasis, but whether they regulate HCC metastasis is still unclear. Methods: HCC orthotopic implantation and colonization mice models were established in vivo. Model mice were treated with IL-8 and or MMP9 inhibitors, protein kinase C (PKC) inhibitors, or extracellular regulated protein kinases 1/2 (ERK1/2) inhibitors. Liver metastasis and lung metastasis of model mice were confirmed by hematoxylin and eosin staining. The population of circulating tumor cells (CTCs) was detected by flow cytometry. The expression of MMP9 in tumor tissues was determined by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry. In vitro, HCC LM6 (HCCLM6) cells were treated with IL-8 combined PKC inhibitor or ERK1/2 inhibitor. The expression of MMP9 was confirmed by qRT-PCR and Western blot, and the activation of the PKC/ERK1/2 signaling pathway was confirmed by Western blot. Results: IL-8 promoted liver metastasis and lung metastasis in orthotopic transplantation model mice, increased the proportion of CTCs and promoted the expression of MMP9 in tumor tissues, but these effects were reversed by PKC inhibitor or ERK1/2 inhibitor. In vivo colonization experiments, IL-8 promoted tumor cell metastasis to the liver and lung, but the MMP9 inhibitor reversed the metastasis-promoting effect of IL-8. In cell experiments, IL-8 promoted the expression of p-PKC and p-ERK1/2 and inhibited the expression of PKC and ERK1/2; the promotion of MMP9 expression by IL-8 was reversed by PKC inhibitor or ERK1/2 inhibitor. Conclusions: IL-8 up-regulated the expression of MMP9 by activating the PKC/ERK1/2 signaling pathway, thereby promoting the metastasis and colonization of HCC.