In this study, we found that missense variants in genes encoding components of the telomerase complex occurred at increased frequency in sporadic cirrhosis, suggesting that telomerase deficiency causing accelerated telomere shortening may predispose to cirrhosis and that the clinical spectrum of “telomere diseases”(3
)may be broader and more common than previously suspected. We also confirmed that cirrhotic patients have shorter telomeres of peripheral blood leukocytes than age-matched controls, further implicating telomere dysfunction as a molecular event in the pathophysiology of cirrhosis.
Mutations in telomerase complex genes have been associated with the inherited bone marrow failure syndrome dyskeratosis congenita, apparently acquired aplastic anemia, and familial idiopathic pulmonary fibrosis.(3
) Less than ten percent of patients with dyskeratosis congenita eventually develop severe liver disease with several histopathologic findings, especially after hematopoietic stem-cell transplant. In pedigrees of patients with bone marrow failure and telomerase deficiency, loss-of-function mutations correlate with an unusual high prevalence of severe hepatic disease, mainly represented by cirrhosis and nodular regenerative hyperplasia.(25
) In the present work, we determined that telomerase mutations also are associated with non-familial cirrhosis with an identifiable etiologic factor, and that telomerase mutations might contribute to cirrhosis development in these patients. That mutations may contribute to fibrosis progression is further indicated by the recent observation by others of an absence of telomerase mutations in 200 individuals with chronic hepatitis C virus infection who did not progress to cirrhosis (K.L. Rudolph, personal communication).
Hepatic fibrosis in combination with the formation of regenerative nodules is the pathologic hallmark of cirrhosis.(36
) The most common causes of cirrhosis in the developed world are hepatitis C virus infection and chronic alcohol abuse. However, only a portion of patients with chronic hepatitis C or who abuse alcohol eventually develops cirrhosis, suggesting host factors play a critical role in disease progression.(37
) Numerous attempts to identify genetic risk factors for the development of cirrhosis have had limited success. Most reports have focused on candidate variants that might alter the primary pathologic process, such as oxidant stress and immunologic response, with inconsistent results.(38
) One of the better studied risk factors are mutations in keratins as susceptibility markers for cirrhosis. Mutations in keratins 8 and 18 have been found in patients with cirrhosis due to a variety of causes. A 3.35 fold increase in frequency of mutations in the keratin genes was found relative to controls,(43
)which is somewhat less than the 4.63 fold increase found in TERT
in the current study. It is important to note that in contrast to the studies of keratins and other genes where many of the mutations were of uncertain functional significance, in our study all of the mutations in TERT
were shown to reduce telomerase activity leading to shorten telomeres.
Telomerase-deficient murine models have provided some insights into possible mechanisms that might explain the current observations. Short telomeres rather than telomerase insufficiency causes impairment of regeneration and pathological phenotypes in the mouse.(44
) Also in the telomerase “knockout” model, excessively short and dysfunctional telomeres predispose the mouse to chemically induced cirrhosis, and exogenous telomerase expression in hepatocytes ameliorates hepatic function and fibrosis in response to liver chemical injury, indicating a role of telomeres in pathogenesis of cirrhosis.(26
) In addition, human cirrhosis due to chronic liver injury may improve once liver injury is eliminated (45
) and hepatocyte regenerative capacity and reduced synthesis of collagen are critical in this process. Excessive telomere shorting may impair this repair process.
Telomere length was measured in peripheral blood samples from 50 cirrhotic patients (37% of patients) and was significantly shorter than in healthy controls (). It has been shown previously that telomeres are shorter in cirrhotic than in non-cirrhotic hepatocytes regardless of disease etiology.(27
) Short telomeres in both hepatocytes and peripheral blood leukocytes indicate the constitutive essence of telomere attrition in cirrhosis and implicate short and dysfunctional telomeres as a molecular mechanism for cirrhosis. Excessive telomere shortening (caused by telomerase gene mutations or other factors) may impair the hepatocyte regenerative ability in response to chronic injury, thus facilitating fibrosis progression. For example, as telomeres are eroded with aging (), shorter telomeres in older humans may contribute to the more rapid rate of progression to cirrhosis with hepatitis C virus infection in the more elderly.(50
) In agreement with our findings, in families with idiopathic pulmonary fibrosis and telomerase mutations, short telomeres have been hypothesized to limit pneumocyte proliferation, causing loss of alveolar cells and, secondarily, fibrosis.(12
) Alternatively, in cirrhosis, short telomeres may affect stellate cell differentiation into myofibroblasts upon injury thereby affecting the severity of fibrosis. Additionally, telomere attrition in inflammatory cells may induce a profibrotic response or contribute to the myofibroblast differentiation of cells of bone marrow origin.
Environmental factors may influence disease expression. For example, patients with X-linked dyskeratosis congenita, caused by DKC1
mutations, have extremely short telomeres due to DKC1
gene hemizigosity and present a severe and multi-organ phenotype, including mucocutaneous anomalies, bone marrow failure, and pulmonary and hepatic fibrosis.(3
) In patients with telomerase deficiency due to telomerase mutations,
enzyme function is reduced by haploinsufficiency and telomere shortening may be less intense and clinical phenotype may be less pronounced.(51
) In patients with TERT
mutations, aplastic anemia or pulmonary fibrosis may be the only clinical presentation.(11
) Most patients with telomerase mutations and aplastic anemia do not have respiratory failure, and most patients with pulmonary fibrosis do not have cytopenias, suggesting that environmental factors contribute to disease development in a susceptible patient; for example, most patients with telomerase mutations and pulmonary fibrosis are smokers.(12
) In pedigrees of telomerase mutations, liver disease and aplastic anemia presented alone in different affected individuals, further suggesting a role for environmental factors. In one study, approximately three percent of patients with idiopathic pulmonary fibrosis also had cryptogenic cirrhosis, indicating some overlap between clinical features.(52
In conclusion, telomerase mutations resulting intelomere erosion appear to be a genetic risk factor for human cirrhosis and may predispose affected subjects to disease progression in combination with environmental injury, further supporting telomere attrition as a causal event in cirrhosis pathophysiology. Establishing how shortened telomeres increase the risk of cirrhosis may allow for the design of future therapies to reduce the risk of hepatic fibrosis in susceptible populations. Patients with mutations also may be appropriate targets for more aggressive forms of therapy to treat their primary disease given their increased risk of cirrhosis.