Senescent cholangiocytes release extracellular vesicles that alter target cell phenotype via the epidermal growth factor receptor

Abstract

Background & Aims: Primary sclerosing cholangitis (PSC) is a chronic liver disease characterized by peribiliary inflammation and fibrosis. Cholangiocyte senescence is a prominent feature of PSC. Here, we hypothesize that extracellular vesicles (EVs) from senescent cholangiocytes influence the phenotype of target cells. Methods: EVs were isolated from normal human cholangiocytes (NHCs), cholangiocytes from PSC patients and NHCs experimentally induced to senescence. NHCs, malignant human cholangiocytes (MHCs) and monocytes were exposed to 10 8 EVs from each donor cell population and assessed for proliferation, MAPK activation and migration. Additionally, we isolated EVs from plasma of wild-type and Mdr2 −/− mice (a murine model of PSC), and assessed mouse monocyte activation. Results: EVs exhibited the size and protein markers of exosomes. The number of EVs released from senescent human cholangiocytes was increased; similarly, the EVs in plasma from Mdr2 −/− mice were increased. Additionally, EVs from senescent cholangiocytes were enriched in multiple growth factors, including EGF. NHCs exposed to EVs from senescent cholangiocytes showed increased NRAS and ERK1/2 activation. Moreover, EVs from senescent cholangiocytes promoted proliferation of NHCs and MHCs, findings that were blocked by erlotinib, an EGF receptor inhibitor. Furthermore, EVs from senescent cholangiocytes induced EGF-dependent Interleukin 1-beta and Tumour necrosis factor expression and migration of human monocytes; similarly, Mdr2 −/− mouse plasma EVs induced activation of mouse monocytes. Conclusions: The data continue to support the importance of cholangiocyte senescence in PSC pathogenesis, directly implicate EVs in cholangiocyte proliferation, malignant progression and immune cell activation and migration, and identify novel therapeutic approaches for PSC.

Genetic or pharmacological reduction of cholangiocyte senescence improves inflammation and fibrosis in the Mdr2 mouse

“…and contribute to an altered microenvironment through their modified secretome or SASP. 14 Eliminating senescent cells has been shown to reduce tissue dysfunction in age-related pathologies, and prolong life-span. 26 Thus, the aim of the current study was to identify a safe senolytic molecule that selectively targets senescent cholangiocytes, and can improve the PSC-like phenotype in the Mdr2 -/- mouse.…”

“…Hence, it is important to use other markers (i.e., p16, p21, β-gal staining) to detect senescent cells. SASP factors can alter the microenvironment, reinforce the senescent phenotype, and initiate both beneficial and injurious cellular responses (13,14). Senescent cells also highly express the anti-apoptotic B cell lymphoma 2 (BCL2) proteins (BCL2, BCL-XL, MCL1), and therefore are resistant to apoptosis (10,11).…”

“…SASP factors can alter the microenvironment, reinforce the senescent phenotype, and initiate both beneficial and injurious cellular responses. 13 , 14 Senescent cells also highly express antiapoptotic B cell lymphoma 2 (BCL2) proteins [BCL2, B cell lymphoma-extra large (BCL-XL), and myeloid cell leukemia 1 (MCL1)] and, therefore, are resistant to apoptosis. 10 , 11 Thus, cellular senescence has emerged as an important cellular process in health and disease displaying both beneficial ( e.g.…”

“…cartilage, heart, lung, liver etc.) and contribute to an altered microenvironment presumably through their modified secretome or SASP(14). Eliminating senescent cells has shown to reduce tissue dysfunction in age-related pathologies, and prolong life-span(26).…”

Polarized human cholangiocytes release distinct populations of apical and basolateral small extracellular vesicles

“…Abnormal intercellular signaling has emerged as a candidate in these disease processes. An increase in the levels of biliary sEVs has been observed in a cholangiopathy animal model ( Masyuk et al , 2010 ), and cholangiocytes derived from patients with primary sclerosing cholangitis (PSC) exhibit increased EV release ( Al Suraih et al , 2020 ). In addition, EV-associated biomarkers have been identified as potential indicators of disease onset and/or progression ( Arbelaiz et al , 2017 ; Julich-Haertel et al , 2017 ; Olaizola et al , 2018 ).…”