Senescent human hepatocytes express a unique secretory phenotype and promote macrophage migration

Katharine M Irvine, Richard Skoien, Nilesh J Bokil, Michelle Melino, Gethin P Thomas, Dorothy Loo, Brian Gabrielli, Michelle M Hill, Matthew J Sweet, Andrew D Clouston, Elizabeth E Powell

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    AIM: To develop a model of stress-induced senescence to study the hepatocyte senescence associated secretory phenotype (SASP).
    METHODS: Hydrogen peroxide treatment was used to induce senescence in the human HepG2 hepatocyte cell line. Senescence was confirmed by cytochemical staining for a panel of markers including Ki67, p21, heterochromatin protein 1 beta, and senescence-associated-beta- galactosidase activity. Senescent hepatocytes were characterised by gene expression arrays and quantitative polymerase chain reaction (qPCR), and conditioned media was used in proteomic analyses, a human chemokine protein array, and cell migration assays to characterise the composition and function of the hepatocyte SASP.

    RESULTS: Senescent hepatocytes induced classical markers of senescence (p21, heterochromatin protein 1 beta, and senescence-associated-beta-galactosidase activity); and downregulated the proliferation marker, Ki67. Hepatocyte senescence induced a 4.6-fold increase in total secreted protein (P = 0.06) without major alterations in the protein profile. Senescence-induced genes were identified by microarray (Benjamini Hochbergcorrected P < 0.05); and, consistent with the increase in secreted protein, gene ontology analysis revealed a significant enrichment of secreted proteins among inducible genes. The hepatocyte SASP included characteristic factors such as interleukin (IL)-8 and IL-6, as well as novel components such as SAA4, IL-32 and Fibrinogen, which were validated by qPCR and/or chemokine protein array. Senescent hepatocyteconditioned medium elicited migration of inflammatory (granulocyte-macrophage colony stimulating factor, GM-CSF-derived), but not non-inflammatory (CSF-1-derived) human macrophages (P = 0.022), which could contribute to a pro-inflammatory microenvironment in vivo, or facilitate the clearance of senescent cells. CONCLUSION: Our novel model of hepatocyte senescence provides insights into mechanisms by which senescent hepatocytes may promote chronic liver disease pathogenesis.
    Original languageEnglish
    Pages (from-to)17851–17862
    Number of pages13
    JournalWorld Journal of Gastroenterology
    Issue number47
    Publication statusPublished - 2014


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