Therapeutic Intervention with Anti-Complement Component 5 Antibody Does Not Reduce NASH but Does Attenuate Atherosclerosis and MIF Concentrations in Ldlr-/-.Leiden Mice

Background: Chronic inflammation is an important driver in the progression of non-alcoholic steatohepatitis (NASH) and atherosclerosis. The complement system, one of the first lines of defense in innate immunity, has been implicated in both diseases. However, the potential therapeutic value of complement inhibition in the ongoing disease remains unclear. Methods: After 20 weeks of high-fat diet (HFD) feeding, obese Ldlr-/-.Leiden mice were treated twice a week with an established anti-C5 antibody (BB5.1) or vehicle control. A separate group of mice was kept on a chow diet as a healthy reference. After 12 weeks of treatment, NASH was analyzed histopathologically, and genome-wide hepatic gene expression was analyzed by next-generation sequencing and pathway analysis. Atherosclerotic lesion area and severity were quantified histopathologically in the aortic roots. Results: Anti-C5 treatment considerably reduced complement system activity in plasma and MAC deposition in the liver but did not affect NASH. Anti-C5 did, however, reduce the development of atherosclerosis, limiting the total lesion size and severity independently of an effect on plasma cholesterol but with reductions in oxidized LDL (oxLDL) and macrophage migration inhibitory factor (MIF). Conclusion: We show, for the first time, that treatment with an anti-C5 antibody in advanced stages of NASH is not sufficient to reduce the disease, while therapeutic intervention against established atherosclerosis is beneficial to limit further progression.

[1]  Shelly C. Lu,et al.  Metabolic subtypes of patients with NAFLD exhibit distinctive cardiovascular risk profiles , 2022, Hepatology (Baltimore, Md.).

[2]  Shelly C. Lu,et al.  Metabolic subtypes of patients with NAFLD exhibit distinctive cardiovascular risk profiles , 2022, Hepatology.

[3]  T. Hendrikx,et al.  Oxidized Lipids: Common Immunogenic Drivers of Non-Alcoholic Fatty Liver Disease and Atherosclerosis , 2022, Frontiers in Cardiovascular Medicine.

[4]  Arturo Santos,et al.  Pathophysiological Molecular Mechanisms of Obesity: A Link between MAFLD and NASH with Cardiovascular Diseases , 2021, International journal of molecular sciences.

[5]  Xiaofeng Yang,et al.  Complement Inhibition Targeted to Injury Specific Neoepitopes Attenuates Atherogenesis in Mice , 2021, Frontiers in Cardiovascular Medicine.

[6]  S. Friedman,et al.  Inflammatory and fibrotic mechanisms in NAFLD—Implications for new treatment strategies , 2021, Journal of internal medicine.

[7]  R. Kleemann,et al.  Diet and exercise reduce pre-existing NASH and fibrosis and have additional beneficial effects on the vasculature, adipose tissue and skeletal muscle via organ-crosstalk. , 2021, Metabolism: clinical and experimental.

[8]  A. B. Storsve,et al.  Krill Oil Treatment Increases Distinct PUFAs and Oxylipins in Adipose Tissue and Liver and Attenuates Obesity-Associated Inflammation via Direct and Indirect Mechanisms , 2021, Nutrients.

[9]  Sahmin Lee,et al.  Antibody-Based Therapeutics for Atherosclerosis and Cardiovascular Diseases , 2021, International journal of molecular sciences.

[10]  R. Kleemann,et al.  Cholesterol Accumulation as a Driver of Hepatic Inflammation Under Translational Dietary Conditions Can Be Attenuated by a Multicomponent Medicine , 2021, Frontiers in Endocrinology.

[11]  V. Saini,et al.  Mechanistic Insights into the Oxidized Low-Density Lipoprotein-Induced Atherosclerosis , 2020, Oxidative medicine and cellular longevity.

[12]  R. Kleemann,et al.  A Translational Mouse Model for NASH with Advanced Fibrosis and Atherosclerosis Expressing Key Pathways of Human Pathology , 2020, Cells.

[13]  Jiao Guo,et al.  The role of neutrophils in innate immunity-driven nonalcoholic steatohepatitis: lessons learned and future promise , 2020, Hepatology International.

[14]  B. Stockinger,et al.  Characterizing the original anti‐C5 function‐blocking antibody, BB5.1, for species specificity, mode of action and interactions with C5 , 2020, Immunology.

[15]  V. Fuster,et al.  Complement C5 Protein as a Marker of Subclinical Atherosclerosis. , 2020, Journal of the American College of Cardiology.

[16]  H. Malhi,et al.  Pathogenesis of Nonalcoholic Steatohepatitis: An Overview , 2020, Hepatology communications.

[17]  R. Kleemann,et al.  Combined Treatment with L-Carnitine and Nicotinamide Riboside Improves Hepatic Metabolism and Attenuates Obesity and Liver Steatosis , 2019, International journal of molecular sciences.

[18]  A. Murphy,et al.  Monocytes, Macrophages, and Metabolic Disease in Atherosclerosis , 2019, Front. Pharmacol..

[19]  C. Weber,et al.  ApoE attenuates unresolvable inflammation by complex formation with activated C1q , 2019, Nature Medicine.

[20]  G. Paragh,et al.  The Impact of Obesity on the Cardiovascular System , 2018, Journal of diabetes research.

[21]  R. Kleemann,et al.  Obeticholic Acid Modulates Serum Metabolites and Gene Signatures Characteristic of Human NASH and Attenuates Inflammation and Fibrosis Progression in Ldlr‐/‐.Leiden Mice , 2018, Hepatology communications.

[22]  G. Cao,et al.  Complement Complex C5b-9 Levels Are Associated with the Clinical Outcomes of Acute Ischemic Stroke and Carotid Plaque Stability , 2018, Translational Stroke Research.

[23]  P. Libby,et al.  Atherosclerosis and inflammation: overview and updates. , 2018, Clinical science.

[24]  R. Kleemann,et al.  Key Inflammatory Processes in Human NASH Are Reflected in Ldlr−/−.Leiden Mice: A Translational Gene Profiling Study , 2018, Front. Physiol..

[25]  A. Sanyal,et al.  Preclinical models of non-alcoholic fatty liver disease. , 2018, Journal of hepatology.

[26]  R. Goldschmeding,et al.  Uncovering a Predictive Molecular Signature for the Onset of NASH-Related Fibrosis in a Translational NASH Mouse Model , 2017, Cellular and molecular gastroenterology and hepatology.

[27]  T. Kooistra,et al.  A casein hydrolysate based formulation attenuates obesity and associated non-alcoholic fatty liver disease and atherosclerosis in LDLr-/-.Leiden mice , 2017, PloS one.

[28]  L. Bavia,et al.  The complement component C5 promotes liver steatosis and inflammation in murine non-alcoholic liver disease model. , 2016, Immunology letters.

[29]  Yun Zhang,et al.  Overexpression of complement component C5a accelerates the development of atherosclerosis in ApoE-knockout mice , 2016, Oncotarget.

[30]  T. Kooistra,et al.  Resolvin E1 attenuates atherosclerosis in absence of cholesterol-lowering effects and on top of atorvastatin. , 2016, Atherosclerosis.

[31]  Youming Li,et al.  Serum complement C3 levels are associated with nonalcoholic fatty liver disease independently of metabolic features in Chinese population , 2016, Scientific Reports.

[32]  Armugam P Mekala,et al.  The role of complement activation in atherogenesis: the first 40 years , 2015, Immunologic Research.

[33]  T. Kooistra,et al.  Replacement of Dietary Saturated Fat by PUFA-Rich Pumpkin Seed Oil Attenuates Non-Alcoholic Fatty Liver Disease and Atherosclerosis Development, with Additional Health Effects of Virgin over Refined Oil , 2015, PloS one.

[34]  H. Völzke,et al.  Complement Component 5 Mediates Development of Fibrosis, via Activation of Stellate Cells, in 2 Mouse Models of Chronic Pancreatitis , 2015, Gastroenterology.

[35]  Qing Zhang,et al.  Association between Complement C3 and Prevalence of Fatty Liver Disease in an Adult Population: A Cross-Sectional Study from the Tianjin Chronic Low-Grade Systemic Inflammation and Health (TCLSIHealth) Cohort Study , 2015, PloS one.

[36]  M. Daha,et al.  Functional assessment of mouse complement pathway activities and quantification of C3b/C3c/iC3b in an experimental model of mouse renal ischaemia/reperfusion injury. , 2015, Journal of immunological methods.

[37]  Chang Liu,et al.  Central obesity and nonalcoholic fatty liver disease risk after adjusting for body mass index. , 2015, World journal of gastroenterology.

[38]  R. Kleemann,et al.  Establishment of a General NAFLD Scoring System for Rodent Models and Comparison to Human Liver Pathology , 2014, PloS one.

[39]  W. Huber,et al.  Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2 , 2014, Genome Biology.

[40]  Gongxiong Wu,et al.  Targeted mouse complement inhibitor CR2-Crry protects against the development of atherosclerosis in mice. , 2014, Atherosclerosis.

[41]  Andreas Krämer,et al.  Causal analysis approaches in Ingenuity Pathway Analysis , 2013, Bioinform..

[42]  B. Fromenty,et al.  Mitochondrial adaptations and dysfunctions in nonalcoholic fatty liver disease , 2013, Hepatology.

[43]  E. Feskens,et al.  Activated complement factor 3 is associated with liver fat and liver enzymes: the CODAM study , 2013, European journal of clinical investigation.

[44]  T. Hughes,et al.  The complement membrane attack complex triggers intracellular Ca2+ fluxes leading to NLRP3 inflammasome activation , 2013, Journal of Cell Science.

[45]  L. Gan,et al.  NASH is an Inflammatory Disorder: Pathogenic, Prognostic and Therapeutic Implications , 2012, Gut and liver.

[46]  C. Cook,et al.  The effect of complement C5a on mitochondrial functions of PC12 cells , 2011, Neuroreport.

[47]  A. Zernecke,et al.  Complement C5a inhibition reduces atherosclerosis in ApoE–/– mice , 2011, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[48]  K. Cianflone,et al.  Complement C3 and cleavage products in cardiometabolic risk. , 2011, Clinica chimica acta; international journal of clinical chemistry.

[49]  J. Wojta,et al.  Complement in atherosclerosis: friend or foe? , 2011, Journal of thrombosis and haemostasis : JTH.

[50]  A. Zernecke,et al.  C5a Receptor Targeting in Neointima Formation After Arterial Injury in Atherosclerosis-Prone Mice , 2010, Circulation.

[51]  T. Hughes,et al.  The membrane attack complex of complement drives the progression of atherosclerosis in apolipoprotein E knockout mice , 2010, Molecular immunology.

[52]  R. Steffensen,et al.  Activation of the complement system in human nonalcoholic fatty liver disease , 2009, Hepatology.

[53]  Jichun Yang,et al.  Induction of MIF expression by oxidized LDL via activation of NF-kappaB in vascular smooth muscle cells. , 2009, Atherosclerosis.

[54]  P. Zipfel,et al.  Complement regulators and inhibitory proteins , 2009, Nature Reviews Immunology.

[55]  J. H. van Bockel,et al.  MIF Deficiency Reduces Chronic Inflammation in White Adipose Tissue and Impairs the Development of Insulin Resistance, Glucose Intolerance, and Associated Atherosclerotic Disease , 2009, Circulation research.

[56]  D. Rader,et al.  CD59 but not DAF deficiency accelerates atherosclerosis in female ApoE knockout mice. , 2009, Molecular immunology.

[57]  Gongxiong Wu,et al.  Complement Regulator CD59 Protects Against Atherosclerosis by Restricting the Formation of Complement Membrane Attack Complex , 2009, Circulation research.

[58]  J. Boyle,et al.  Brief Report: Accelerated Atherosclerosis in Low-Density Lipoprotein Receptor–Deficient Mice Lacking the Membrane-Bound Complement Regulator CD59 , 2008, Arteriosclerosis, thrombosis, and vascular biology.

[59]  A. Blom,et al.  Complement regulation in human atherosclerotic coronary lesions. Immunohistochemical evidence that C4b-binding protein negatively regulates the classical complement pathway, and that C5b-9 is formed via the alternative complement pathway. , 2007, Atherosclerosis.

[60]  J. Bernhagen,et al.  MIF is a noncognate ligand of CXC chemokine receptors in inflammatory and atherogenic cell recruitment , 2007, Nature Medicine.

[61]  Jörg Köhl,et al.  Complement factor 5 is a quantitative trait gene that modifies liver fibrogenesis in mice and humans , 2005, Nature Genetics.

[62]  P. Ward,et al.  Role of C5a in inflammatory responses. , 2005, Annual review of immunology.

[63]  P. Libby,et al.  Low-Density Lipoprotein Receptor-Deficient Mice Macrophage Migration Inhibitory Factor Deficiency Impairs Atherosclerosis in , 2022 .

[64]  B. Jaber,et al.  C5a delays apoptosis of human neutrophils via an extracellular signal‐regulated kinase and Bad‐mediated signalling pathway , 2004, European journal of clinical investigation.

[65]  H. Rus,et al.  The role of complement activation in atherosclerosis , 2004, Immunologic research.

[66]  S. Verma,et al.  New Markers of Inflammation and Endothelial Cell Activation: Part I , 2003, Circulation.

[67]  W. Leonard,et al.  Cytokine and Cytokine Receptor Pleiotropy and Redundancy* , 2002, The Journal of Biological Chemistry.

[68]  J. Bernhagen,et al.  Expression of Macrophage Migration Inhibitory Factor in Different Stages of Human Atherosclerosis , 2002, Circulation.

[69]  Y. Chao,et al.  ApoE(-/-) mice develop atherosclerosis in the absence of complement component C5. , 2001, Biochemical and biophysical research communications.

[70]  R. Kinscherf,et al.  Complement C6 deficiency protects against diet-induced atherosclerosis in rabbits. , 1998, Arteriosclerosis, thrombosis, and vascular biology.

[71]  P. Ward,et al.  Sublytic concentrations of the membrane attack complex of complement induce endothelial interleukin-8 and monocyte chemoattractant protein-1 through nuclear factor-kappa B activation. , 1997, The American journal of pathology.

[72]  W D Wagner,et al.  A definition of advanced types of atherosclerotic lesions and a histological classification of atherosclerosis. A report from the Committee on Vascular Lesions of the Council on Arteriosclerosis, American Heart Association. , 1995, Arteriosclerosis, thrombosis, and vascular biology.

[73]  H. Rus,et al.  Quantitative evaluation of the terminal C5b-9 complement complex by ELISA in human atherosclerotic arteries. , 1987, Clinical and experimental immunology.