Background & Aims
Binge drinking is associated with an increased risk of liver disease. Morbidity and mortality of alcohol‐related liver disease (ALD) is associated with collagen deposition in the ...hepatic extracellular matrix (ECM). However, the acute effects of binge drinking on ECM turnover are unknown. We aimed to investigate the effects on hepatic ECM turnover following a binge drinking episode.
Methods
We performed a pathophysiological intervention study with 15 non‐alcoholic fatty liver disease (NAFLD) patients, 15 ALD patients and 10 healthy controls. We used 40% ethanol in 9 mg/mL NaCl administered through a nasogastric tube to simulate binge drinking. Hepatic vein catheterisation allowed simultaneous hepatic‐ and systemic vein sampling. Markers of ECM formation and degradation were measured with competitive ELISA.
Results
The interstitial matrix formation marker PRO‐C3 increased by 1.2 ng/mL (10%, P < .001) 24 hours after binge drinking. In participants with existing liver fibrosis determined by elevated baseline PRO‐C3, hepatic levels increased by 0.09 ng/mL (95% CI: 0.03‐0.15, P = .005) while systemic PRO‐C3 decreased 0.11 ng/mL (95% CI: −0.15 to −0.06, P < .001) in 3 hours. PRO‐C8 increased by 30% (+0.9 ng/mL, P = .014) in liver‐diseased patients with F0‐F1 but not in any other group. Twenty‐four‐hour changes in systemic C3M and PRO‐C3 were not associated (P = .911).
Conclusions
Binge drinking induced an acute burst of PRO‐C3 in healthy individuals and patients with liver disease. Markers of ECM degradation were not correlated to markers of ECM formation, suggesting that even a single episode of binge drinking promotes excessive hepatic fibrogenesis.
Overview of study design and main finding where PRO‐C3 increased in all study groups following a single binge drinking episode. Permission to reproduce material from other sources: Graphical created with BioRender.com reproduced with permission.
Aims/hypothesis
The treatment of type 2 diabetes with full peroxisome proliferator-activated receptor gamma (PPARγ) agonists improves insulin sensitivity, but is associated with weight gain, heart ...failure, peripheral oedema and bone loss. Endotrophin, the C-terminal fragment of the α3 chain of procollagen type VI (also called Pro-C6), is involved in both adipose tissue matrix remodelling and metabolic control. We established a serum assay for endotrophin to assess if this novel adipokine could identify type 2 diabetic patients who respond optimally to PPARγ agonists, improving the risk-to-benefit ratio.
Methods
The BALLET trial (NCT00515632) compared the glucose-lowering effects and safety of the partial PPARγ agonist balaglitazone with those of pioglitazone in individuals with type 2 diabetes on stable insulin therapy. The per protocol population (
n
= 297) was stratified into tertiles based on baseline endotrophin levels. Participants were followed-up after 26 weeks, after which correlational analysis was carried out between endotrophin levels and measures of glucose control. This is a secondary post hoc analysis.
Results
Endotrophin was significantly associated with therapeutic response to balaglitazone and pioglitazone. At week 26, only individuals in the upper two tertiles showed significant reductions in HbA
1c
and fasting serum glucose compared with baseline. The OR for a 1% and a 0.5% reduction in HbA
1c
for individuals in the upper two tertiles were 3.83 (95% CI 1.62, 9.04)
p
< 0.01, and 3.85 (95% CI 1.94, 7.61)
p
< 0.001, respectively. Endotrophin levels correlated with adipose tissue mass, insulin resistance and fatty liver index. Notably, PPARγ-associated adverse effects, such as moderate-to-severe lower extremity oedema, only occurred in the lower tertile.
Conclusions/interpretation
Elevated endotrophin serum levels predict response to two insulin sensitisers and reduce the risk of associated adverse effects, thereby, identifying patients with type 2 diabetes who may profit from PPARγ agonist treatment.
Background & Aims
Fibrogenesis results in release of certain extracellular matrix protein fragments into the circulation. We evaluated the diagnostic and prognostic performance of two novel ...serological markers, the precisely cleaved N‐terminal propeptide of type III collagen (Pro‐C3) and a peptide of helical collagen type III degradation (C3M), in chronic hepatitis C (CHC) patients.
Method
Pro‐C3 and C3M were measured by ELISA in plasma from CHC patients (n = 194) from a prior phase II antifibrotic trial (NCT00244751). Plasma samples and paired liver biopsies were obtained at baseline and after 1‐year. Patients were stratified according to Ishak stages 2‐4. Internal cross‐validation was performed by bootstrap analysis.
Results
Pro‐C3 levels were significantly higher in CHC patients in Ishak stage 4 compared to stage 2 (P < 0.001) or 3 (P < 0.01). Pro‐C3 could significantly distinguish moderate (stage 4) from mild fibrosis (stage 2/3) (AUC = 0.72, P < 0.001). Importantly, an overall significance in Pro‐C3 (P = 0.007) levels was observed between the groups of −1, 0, +1 and +2 change in Ishak stage at 12 months. Pro‐C3 was significantly increased in group +1 (P = 0.030) and +2 (P = 0.021) compared to group 0. No significant differences were observed for C3M. In multivariate analysis, only baseline Pro‐C3, but not FibroTest, had an independent association with fibrosis progression.
Conclusions
Pro‐C3 is a useful test to predict fibrogenesis and monitor disease progression. Moreover, it could differentiate mild from moderate disease. Pro‐C3 may become a promising blood parameter be included in future studies for monitoring disease progression and eventually for evaluation of potential antifibrotic therapies.
Summary
Background
Alcohol is a main cause of preventable deaths and frequently leads to the development of alcohol‐related liver disease. Due to the lack of diagnostics, patients are commonly ...diagnosed after developing clinical manifestations. Recently, the biomarker PRO‐C3 was shown to accurately identify fibrosis due to non‐alcoholic fatty liver disease.
Aim
To assess the diagnostic accuracy of PRO‐C3, the ADAPT score and best‐performing non‐patented serological test to detect advanced alcohol‐related liver fibrosis.
Methods
We enrolled 426 patients with alcohol overuse in a prospective biopsy‐controlled study. We evaluated the accuracy of PRO‐C3 and the PRO‐C3‐based algorithm ADAPT to detect advanced liver fibrosis.
Results
The accuracy of PRO‐C3 was good with an AUROC of 0.85 (95% CI 0.79‐0.90). The best‐performing non‐patented test was the Forns index with an AUROC of 0.83 (95% CI 0.78‐0.89). The ADAPT algorithm performed better as compared to both the Forns index and PRO‐C3 alone with an AUROC = 0.88 (95% CI 0.83‐0.93).
Conclusion
PRO‐C3 is a new marker with high accuracy to detect advanced alcohol‐related liver fibrosis. The diagnostic accuracy of PRO‐C3 can be further improved by using the ADAPT algorithm in which the test outperforms currently available non‐patented serological fibrosis markers. The study is registered in the Odense Patient Data Exploratory Network (OPEN) under study identification numbers OP_040 (https://open.rsyd.dk/OpenProjects/da/openProject.jsp?openNo=40) and OP_239 (https://open.rsyd.dk/OpenProjects/openProject.jsp?openNo=239&lang=da).
PRO‐C3 and ADAPT algorithm accurately identify patients with advanced fibrosis due to alcohol related liver disease
Liver-associated complications still frequently lead to mortality in people with HIV (PWH), even though combined antiretroviral treatment (cART) has significantly improved overall survival. The ...quantification of circulating collagen fragments released during collagen formation and degradation correlate with the turnover of extracellular matrix (ECM) in liver disease. Here, we analysed the levels of ECM turnover markers PC3X, PRO-C5, and PRO-C6 in PWH and correlated these with hepatic fibrosis and steatosis.
This monocentre, retrospective study included 141 PWH. Liver stiffness and liver fat content were determined using transient elastography (Fibroscan) with integrated CAP function. Serum levels of formation of cross-linked type III collagen (PC3X), formation of type V collagen (PRO-C5) and formation type VI collagen (PRO-C6), also known as the hormone endotrophin, were measured with ELISA.
Twenty-five (17.7%) of 141 PWH had clinical significant fibrosis with liver stiffness ≥ 7.1 kPa, and 62 PWH (44.0%) had steatosis with a CAP value > 238 dB/m. Study participants with fibrosis were older (p = 0.004) and had higher levels of AST (p = 0.037) and lower number of thrombocytes compared to individuals without fibrosis (p = 0.0001). PC3X and PRO-C6 were markedly elevated in PWH with fibrosis. Multivariable cox regression analysis confirmed PC3X as independently associated with hepatic fibrosis. PRO-C5 was significantly elevated in participants with presence of hepatic steatosis.
Serological levels of cross-linked type III collagen formation and endotrophin were significantly associated with liver fibrosis in PWH receiving cART and thus may be suitable as a non-invasive evaluation of liver fibrosis in HIV disease.
Increased attention is paid to the structural components of tissues. These components are mostly collagens and various proteoglycans. Emerging evidence suggests that altered components and noncoded ...modifications of the matrix may be both initiators and drivers of disease, exemplified by excessive tissue remodeling leading to tissue stiffness, as well as by changes in the signaling potential of both intact matrix and fragments thereof. Although tissue structure until recently was viewed as a simple architecture anchoring cells and proteins, this complex grid may contain essential information enabling the maintenance of the structure and normal functioning of tissue. The aims of this review are to (1) discuss the structural components of the matrix and the relevance of their mutations to the pathology of diseases such as fibrosis and cancer, (2) introduce the possibility that post-translational modifications (PTMs), such as protease cleavage, citrullination, cross-linking, nitrosylation, glycosylation, and isomerization, generated during pathology, may be unique, disease-specific biochemical markers, (3) list and review the range of simple enzyme-linked immunosorbent assays (ELISAs) that have been developed for assessing the extracellular matrix (ECM) and detecting abnormal ECM remodeling, and (4) discuss whether some PTMs are the cause or consequence of disease. New evidence clearly suggests that the ECM at some point in the pathogenesis becomes a driver of disease. These pathological modified ECM proteins may allow insights into complicated pathologies in which the end stage is excessive tissue remodeling, and provide unique and more pathology-specific biochemical markers.
Background and Aims
Risk prediction in alcohol‐related liver disease (ArLD) is an unmet need. We aimed to assess PRO‐C3 models to predict liver‐related events (LRE) in patients with a history of ...excessive alcohol use without an established diagnosis of chronic liver disease.
Methods
A prospective cohort study of 462 patients with ArLD, split into a derivation cohort of 221 secondary care patients and a validation cohort of 241 primary care patients. Baseline variables, including fibrogenesis marker PRO‐C3, were used to develop a prediction model. Prognostic accuracy was compared to enhanced liver fibrosis (ELF), fibrosis‐4‐index (FIB‐4), transient elastography (TE) and ADAPT.
Results
In the derivation and validation cohorts, 67 (30%) and 19 (8%) experienced an LRE during a median follow‐up of 5.2 years (IQR: 3.2‐6.8) and 4.0 years (IQR: 2.7‐5.6). On top of PRO‐C3 and ADAPT score, we generated a model (ALPACA) of independent predictors of LREs (PRO‐C3, AST/ALT, platelets). ALPACA had high prognostic accuracy with a C‐statistic of 0.85 in the derivation cohort, comparable to ELF (0.83) and TE (0.84) and significantly higher than FIB‐4 (0.78), PRO‐C3 (0.80) and ADAPT (0.81). In the validation cohort, all tests had comparable C‐statistics. Compared to low‐risk patients (ALPACA ≤11), high‐risk patients (>11) had a subhazard ratio for LREs of 12.6 (95% CI 5.9‐26.8, p < .001) and higher cumulative incidence (57% vs. 7%, p < .001; derivation cohort). We observed similar subhazard ratio in the validation cohort.
Conclusions
PRO‐C3‐based scores are reliable tools to predict LREs in ArLD patients and are suitable for risk stratification in primary and secondary care.
Background and Aims
Liver fibrosis results from a prolonged wound healing response to continued injury with excessive production of extracellular proteins. In patients with chronic liver disease, the ...monitoring of liver fibrosis dynamics is of high interest. Whilst markers of fibrogenesis exist, markers of hepatic fibrosis resolution remain an unmet clinical need. Thus, we sought to develop an assay quantifying a circulating proteolytic fragment of cross‐linked type III collagen as a biomarker of fibrolysis, testing its utility in two clinical cohorts of liver fibrosis of distinct aetiology and regressing endotype
Methods
We used a monoclonal antibody targeting the C‐telopeptide of type III collagen following C‐proteinase cleavage to develop and validate a neo‐epitope‐specific enzyme‐linked immunosorbent assay (CTX‐III). A potential fibrosis resolution marker, CTX‐III, was measured in two clinical cohorts of patients with obesity‐associated non‐alcoholic fatty liver disease undergoing bariatric surgery or hepatitis C virus infection from a clinical trial study evaluating the anti‐fibrotic effect of farglitazar.
Results
CTX‐III was robust and specific for the targeted neo‐epitope with good reproducibility in EDTA plasma. We assessed type III collagen remodelling using a panel of biomarkers, including a type III collagen formation marker (PRO‐C3), degradation (C3M), and CTX‐III (fibrolysis). Net fibrolysis was increased in patients with non‐alcoholic fatty liver disease following bariatric surgery (p < .001). Moreover, net fibrolysis identified spontaneous fibrotic regressors from stable and progressors (p < .05 and p < .001) among hepatitis C virus infection patients.
Conclusion
Circulating CTX‐III as a marker of fibrolysis indicates the biomarker's beneficial use in assessing hepatic fibrosis resolution.
The present study describes the assessment of true formation of type III collagen in different pathologies using a neo-epitope specific competitive Enzyme-linked immunosorbent assay (ELISA) towards ...the N-terminal propeptide of type III collagen (PRO-C3).
The monoclonal antibody was raised against the N-protease mediated cleavage site of the N-terminal propeptide of type III collagen and a competitive ELISA was developed using the selected antibody. The assay was evaluated in relation to neo-epitope specificity, technical performance, and as a marker for liver fibrosis and muscle mass using the rat carbon tetrachloride (CCl4) model and a study of immobilization induced muscle loss in humans, respectively.
The ELISA was neo-epitope specific, technically stable and can be assessed in serum and plasma samples. In the CCl4 liver fibrosis model it was observed that serum PRO-C3 were significantly elevated in rats with liver fibrosis as seen by histology (56% elevated in the highest quartile of total hepatic collagen compared to control rats, p<0.001) and correlated significantly to total hepatic collagen in the diseased rats (r=0.46, p<0.01) and not in control rats, suggesting the pathological origin of the epitope. Human plasma PRO-C3 correlated significantly to muscle mass at baseline (R(2)=0.44, p=0.036).
The developed neo-epitope specific serum ELISA for type III procollagen (PRO-C3) reflects true formation as it is specific for the propeptide cleaved off the intact collagen molecule. In a clinical and in a rodent study we showed that this marker was highly related to liver fibrosis and muscle mass.
Acute-on-chronic liver failure (ACLF) is characterised by organ failure(s), high short-term mortality, and, pathophysiologically, deranged inflammatory responses. The extracellular matrix (ECM) is ...critically involved in regulating the inflammatory response. This study aimed to determine alterations in biomarkers of ECM turnover in ACLF and their association with inflammation, organ failures, and mortality.
We studied 283 patients with cirrhosis admitted for acute decompensation (AD) with or without ACLF, 64 patients with stable cirrhosis, and 30 healthy controls. A validation cohort (25 ACLF, 9 healthy controls) was included. Plasma PRO-C3, PRO-C4, PRO-C5, PRO-C6, and PRO-C8 (i.e. collagen type III–VI and VIII formation) and C4M and C6M (i.e. collagen type IV and VI degradation) were measured. Immunohistochemistry of PRO-C6 was performed on liver biopsies (AD n = 7, ACLF n = 5). A competing-risk regression analysis was performed to explore the prognostic value of biomarkers of ECM turnover with 28- and 90-day mortality.
PRO-C3 and PRO-C6 were increased in ACLF compared to AD (p = 0.089 and p <0.001, respectively), whereas collagen degradation markers C4M and C6M were similar. Both PRO-C3 and PRO-C6 were strongly associated with liver function and inflammatory markers. Only PRO-C6 was associated with extrahepatic organ failures and 28- and 90-day mortality (hazard ratio HR; on log-scale 6.168, 95% CI 2.366–16.080, p <0.001, and 3.495, 95% CI 1.509–8.093, p = 0.003, respectively). These findings were consistent in the validation cohort. High PRO-C6 expression was observed in liver biopsies of patients with ACLF.
This study shows, for the first time, evidence of severe net interstitial collagen deposition in ACLF and makes the novel observation of the association between PRO-C6 and (extrahepatic) organ failures and mortality. Further studies are needed to define the pathogenic significance of these observations.
This study describes a disrupted turnover of collagen type III and VI in Acute-on-chronic liver failure (ACLF). Plasma biomarkers of these collagens (PRO-C3 and PRO-C6) are associated with the severity of liver dysfunction and inflammation. PRO-C6, also known as the hormone endotrophin, has also been found to be associated with multi-organ failure and prognosis in acute decompensation and ACLF.
Display omitted
•Collagen type III and VI formation is increased in ACLF compared to AD.•PRO-C3 and PRO-C6 correlate with the severity of liver dysfunction and inflammation in AD and ACLF.•High PRO-C6 levels were found to be indicative for the presence of multi-organ failure and worse survival.
PDF
