Identifying nonalcoholic fatty liver disease patients with active fibrosis by measuring extracellular matrix remodeling rates in tissue and blood.

Excess collagen synthesis (fibrogenesis) in the liver plays a causal role in the progression of nonalcoholic fatty liver disease (NAFLD). Methods are needed to identify patients with more rapidly progressing disease and to demonstrate early response to treatment. We describe here a novel method to quantify hepatic fibrogenesis flux rates both directly in liver tissue and noninvasively in blood. Twenty-one patients with suspected NAFLD ingested heavy water (2 H2 O, 50-mL aliquots) two to three times daily for 3-5 weeks prior to a clinically indicated liver biopsy. Liver collagen fractional synthesis rate (FSR) and plasma lumican FSR were measured based on 2 H labeling using tandem mass spectrometry. Patients were classified by histology for fibrosis stage (F0-F4) and as having nonalcoholic fatty liver or nonalcoholic steatohepatitis (NASH). Magnetic resonance elastography measurements of liver stiffness were also performed. Hepatic collagen FSR in NAFLD increased with advancing disease stage (e.g., higher in NASH than nonalcoholic fatty liver, positive correlation with fibrosis score and liver stiffness) and correlated with hemoglobin A1C. In addition, plasma lumican FSR demonstrated a significant correlation with hepatic collagen FSR.ConclusionUsing a well-characterized cohort of patients with biopsy-proven NAFLD, this study demonstrates that hepatic scar in NASH is actively remodeled even in advanced fibrosis, a disease that is generally regarded as static and slowly progressive. Moreover, hepatic collagen FSR correlates with established risks for fibrotic disease progression in NASH, and plasma lumican FSR correlates with hepatic collagen FSR, suggesting applications as direct or surrogate markers, respectively, of hepatic fibrogenesis in humans. (Hepatology 2017;65:78-88)

Turnover rates of hepatic collagen and circulating collagen-associated proteins in humans with chronic liver disease.

Accumulation and degradation of scar tissue in fibrotic liver disease occur slowly, typically over many years. Direct measurement of fibrogenesis, the rate of scar tissue deposition, may provide valuable therapeutic and prognostic information. We describe here results from a pilot study utilizing in vivo metabolic labeling to measure the turnover rate of hepatic collagen and collagen-associated proteins in plasma for the first time in human subjects. Eight subjects with chronic liver disease were labeled with daily oral doses of 2H2O for up to 8 weeks prior to diagnostic liver biopsy and plasma collection. Tandem mass spectrometry was used to measure the abundance and fractional synthesis rate (FSR) of proteins in liver and blood. Relative protein abundance and FSR data in liver revealed marked differences among subjects. FSRs of hepatic type I and III collagen ranged from 0.2-0.6% per day (half-lives of 4 months to a year) and correlated significantly with worsening histologic fibrosis. Analysis of plasma protein turnover revealed two collagen-associated proteins, lumican and transforming growth factor beta-induced-protein (TGFBI), exhibiting FSRs that correlated significantly with FSRs of hepatic collagen. In summary, this is the first direct measurement of liver collagen turnover in vivo in humans and suggests a high rate of collagen remodeling in advanced fibrosis. In addition, the FSRs of collagen-associated proteins in plasma are measurable and may provide a novel strategy for monitoring hepatic fibrogenesis rates

Subject Demographics.

<p>*both scores 0 to 4 (Batts Ludwig, 1995)</p><p>Details regarding diagnosis, heavy water labeling duration, pathology scoring and tissue samples analyzed from each clinical subject. Abbreviations: autoimmune hepatitis (AIH); hepatitis C virus (HCV); human immunodeficiency virus (HIV); orthotopic liver transplantation (OLT).</p><p>Subject Demographics.</p

List of 20 hepatic proteins with relative abundances observed to most significantly positively correlate with histological fibrosis score.

<p>List of 20 hepatic proteins with relative abundances observed to most significantly positively correlate with histological fibrosis score.</p

Comparison of Hepatic Collagen Kinetics and Histologic Fibrosis Score.

<p>(A) Linear regression of hepatic type I collagen FSR (% new per day) and histopathologic fibrosis score in human subjects with chonic liver disease. (B,C) Bar graphs depicting the relative abundance of unlabeled (old) and labeled (new) hepatic type I collagen (B) and type III collagen (C) per unit mass of liver protein, normalized to 30 days of labeling <i>in vivo</i>. Collagen FSR values (% new per 30 days) are displayed above each bar. Values shown represent the subject or the mean of subjects with each histopathology score. Abbreviation: fractional synthesis rate (FSR).</p

List of 10 hepatic proteins with fractional synthesis rates (FSRs) that most significantly positively correlate with histological fibrosis score.

<p>List of 10 hepatic proteins with fractional synthesis rates (FSRs) that most significantly positively correlate with histological fibrosis score.</p

Comparison of Plasma Lumican FSR to Hepatic Collagen Kinetics and Histopathologic Fibrosis Score.

<p>(A) Linear regression of FSR of plasma lumican vs FSR of hepatic collagen in human subjects with chronic liver disease. (B) Linear regression of plasma lumican FSR vs histopathologic fibrosis score in human subjects with chronic liver disease. (C) Comparison of FSRs of liver and plasma lumican in individual subjects in whom liver values were measurable. Abbreviation: fractional synthesis rate (FSR).</p

Antibody profiles in COVID-19 convalescent plasma prepared with amotosalen/UVA pathogen reduction treatment.

BackgroundCOVID-19 convalescent plasma (CCP), from donors recovered from severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection, is one of the limited therapeutic options currently available for the treatment of critically ill patients with COVID-19. There is growing evidence that CCP may reduce viral loads and disease severity; and reduce mortality. However, concerns about the risk of transfusion-transmitted infections (TTI) and other complications associated with transfusion of plasma, remain. Amotosalen/UVA pathogen reduction treatment (A/UVA-PRT) of plasma offers a mitigation of TTI risk, and when combined with pooling has the potential to increase the diversity of the polyclonal SARS-CoV-2 neutralizing antibodies.Study design and methodsThis study assessed the impact of A/UVA-PRT on SARS-CoV-2 antibodies in 42 CCP using multiple complimentary assays including antigen binding, neutralizing, and epitope microarrays. Other mediators of CCP efficacy were also assessed.ResultsA/UVA-PRT did not negatively impact antibodies to SARS-CoV-2 and other viral epitopes, had no impact on neutralizing activity or other potential mediators of CCP efficacy. Finally, immune cross-reactivity with other coronavirus antigens was observed raising the potential for neutralizing activity against other emergent coronaviruses.ConclusionThe findings of this study support the selection of effective CCP combined with the use of A/UVA-PRT in the production of CCP for patients with COVID-19