Cellular senescence, neuropeptides and hepatic fibrosis: Additional insights into increasing complexity
Authors: Steven P. O'Hara, Nicholas F. La Russo
Journal: Hepatology (2017)

Potential conflict of interest: Nothing to report. See Article On Page 528 Cells respond to stress through initiation of programs that promote their differentiation to terminal fates, including cellular senescence, a process described over 50 years ago when it was observed that normal human cells had a limited ability to proliferate in culture.1 The senescent phenotype is now recognized as a stable form of cell cycle arrest, mediated predominantly by two proteins, p21 and p16INK4a. This pathway is triggered by a variety of stimuli including telomere erosion, mitogenic or oncogenic signaling, oxidative stress, and DNA damage. Until recently, the physiological relevance of this process was unclear. Based on multiple studies from multiple disciplines, it is now generally accepted that the senescent cellular phenotype prevents continued proliferation of aged (e.g., shortened telomeres) or damaged (e.g., DNA double‐strand breaks) cells, supporting the notion that senescence is a barrier to proliferation of potentially neoplastic cells. In addition, senescence is likely important in tissue repair/remodeling, preservation of tissue architecture, and, ultimately, protective mechanisms for overall organ function. In contrast, accumulation of senescent cells has been implicated in both aging itself and multiple age‐associated diseases, as well as in diseases not limited to old age including liver diseases such as primary biliary cholangitis and primary sclerosing cholangitis (PSC).2 In PSC, a cholangiopathy characterized by concentric fibrosis and inflammation leading to biliary cirrhosis, cholangiocyte senescence is a prominent feature.3 Moreover, in livers of patients with PSC, senescent cholangiocytes exhibit characteristics of the senescence‐associated secretory phenotype (SASP),3 a potentially pathologic state of proinflammatory cytokine, chemokine, and growth factor hypersecretion.4 Indeed, data from in vitro and in vivo models suggest that persistent cholangiocyte stress initiates signaling cascades, including N‐Ras/mitogen‐activated protein kinase activation, that culminate in increased p16INK4a expression dependent on ETS1, a transcription factor driving cholangiocyte senescence and influencing the SASP.3 This working model further suggests that the cholangiocyte SASP primes the peribiliary microenvironment for inflammation and fibrosis. In contrast to the proposed inciting role of senescent cholangiocytes in biliary injury, previous work using the carbon tetrachloride model of hepatic fibrosis suggested that, in hepatic stellate cells (HSCs), senescence functions to limit fibrosis.6 Using this model, Krizhanovsky et al.6 demonstrated that activated HSCs (i.e., myofibroblasts) both transition to a senescent state, thus limiting their own numbers, and, through the SASP, direct their own removal by natural killer cells. Moreover, senescent HSCs up‐regulate matrix metalloproteinase‐9, consistent with the SASP, which further promotes fibrosis resolution.7 Taken together, these experiments reveal an additional layer of complexity for the functional and pathological consequences of cellular senescence, both in general and in liver disease; that is, while accumulation of senescent cholangiocytes may be detrimental to bile ducts, likely through SASP, HSC senescence may function to limit hepatic fibrosis and impede disease progression. These considerations emphasize the importance of continued exploration of the cellular targets, molecular mechanisms, and pathophysiological consequences of cellular senescence in liver disease. In that context, Wan et al. report on the intersection of the neuropeptide substance P (SP) and both cholangiocyte and myofibroblast senescence.8 SP, encoded by the TAC1 gene, is expressed in the central and peripheral nervous systems. In addition to functioning there as a proinflammatory peptide, SP is expressed in a variety of peripheral organs, including the liver. Upon binding to its receptor, neurokinin‐1 receptor (NK‐1R), which is expressed on several resident liver cells including cholangiocytes and HSCs, SP promotes a variety of physiological and potentially pathophysiological effects including inflammation and cell survival.9 In this issue, Wan et al. report experiments on the role of the SP/NK‐1R signaling axis in the development of liver fibrosis in the Abcb4–/– (Mdr2–/–) and bile duct–ligated (BDL) mouse models of biliary injury. They observed that both models exhibited increased levels of TAC1 and NK‐1R mRNA in the liver as well as elevated circulating SP. Increased expression of hepatic TAC1 and NK‐1R mRNA and increased circulating SP were also observed in PSC patient samples compared to healthy controls. Importantly, they used both genetic and pharmacologic approaches to determine whether the SP/NK‐1R signaling axis contributed to fibrosis in the rodent models. First, BDL was performed in mice deficient in NK‐1R; when compared to control wild‐type BDL mice, NK‐1R‐deficient mice had improved serum profiles of liver injury, decreased fibrosis, and diminished numbers of activated myofibroblasts. Second, when Mdr2–/– mice were treated with the NK‐1R antagonist L‐733,060, they also exhibited improved hepatic phenotype and diminished myofibroblasts compared to untreated Mdr2–/– mice. Importantly, the improved phenotype of the BDL NK‐1R–/– mice and the L‐733,060‐treated Mdr2–/– mice correlated with decreased total liver senescence‐associated beta‐galactosidase staining and diminished expression of the senescence‐associated genes p16INK4A and p21. Based on these data, they reasonably concluded that the SP/NK‐1R signaling axis likely contributes to the increased senescence and fibrosis observed in these hepatobiliary injury models. To pursue this further, they assessed senescence markers in cells isolated from mouse liver. Surprisingly, the profibrogenic effects of SP were associated with increased cholangiocyte, but decreased myofibroblast, expression of the senescence‐associated genes p21 and P16INK4a. Additionally, BDL performed in NK‐1R‐deficient animals or administration of L‐733,060 to Mdr2–/– mice partially reversed these cell‐specific changes in p16INK4a and p21. In additional in vitro experiments, and consistent with the in vivo data, they found that SP treatment of cultured human HSCs or murine cholangiocytes either reduced (HSC) or increased (cholangiocyte) p16INK4a and p21 expression. These results further suggest that the SP/NK‐1R signaling axis has divergent effects on cellular senescence depending on cell type but may contribute to a common physiological/pathophysiological outcome, i.e., increased fibrosis. A working model of this concept is depicted in Fig. 1.Figure 1: Working model of SP‐associated hepatobiliary fibrosis. In this issue, the role of the SP/NK‐1R signaling axis in the development of liver fibrosis in the Abcb4 –/– (Mdr2 –/–) and BDL mouse models of biliary injury is explored. Several cell types in the liver express the SP cognate receptor NK‐1R, including cholangiocytes and HSCs. Senescence of both of these cell types has been reported in models of liver injury. The cholangiocyte senescence phenotype is proinflammatory/profibrotic, while myofibroblast senescence limits fibrosis. In this model, SP promotes cholangiocyte senescence and limits myofibroblast senescence, hence promoting fibrosis. This paradigm is supported by the observation that BDL in NK‐1R‐deficient mice and treatment of the Mdr2 –/– mouse with the NK‐1R inhibitor L‐733,060 limit cholangiocyte but promote myofibroblast senescence. The work by Wan et al. provides additional justification for continued investigation of the pathogenic importance and therapeutic target potential of cellular senescence and SASP in liver disease.Thus, their data not only broaden the scope and implications of cellular senescence and its role in hepatic fibrosis but also provide additional support to the concept that the senescent phenotype is an integral component of fundamental physiological/pathophysiological aspects of tissue repair. As with any good science, their work generates many questions. For example, how does the SP/NK‐1R signaling axis cause dramatic phenotypic differences in different cell types; could this be a cell type–specific, epigenetic phenomenon? While it is known that senescent myofibroblasts are removed by natural killer cells,6 what is the mechanism of senescent cholangiocyte removal? Moreover, why do senescent cholangiocytes accumulate in PSC livers? Are PSC cholangiocytes genetically or epigenetically more likely to undergo senescence when challenged, or are they resistant to senescence surveillance and removal? How can these observations advance us toward a therapeutic target for PSC? Can we target senescence‐associated processes, such as autophagy and the SASP, for therapeutic benefit? Most senescent cells share with cancer cells the feature of apoptosis resistance, identified as a molecular target for which pharmaceuticals are already in use. In addition, a new, exciting pharmacological approach to both aging and senescence‐associated pathologies (e.g., lung fibrosis) is targeted killing of senescent cells with drugs called senolytics.10 With the use of senolytics, the prediction is that the targeted removal of senescent cells will limit fibrotic and inflammatory damage to tissues and promote healing; proof of concept of this hypothesis has recently been demonstrated.11 While it remains to be seen whether targeted removal of excessive senescent cells may promote healing in liver disease, the work by Wan et al. provides additional justification for continued investigation of the pathogenic importance and therapeutic target potential of cellular senescence and SASP in liver disease. Author names in bold designate shared co‐first authorship.