Chronic kidney disease – a clinical model of premature vascular ageing
Background: Chronic kidney disease (CKD) is a silent and widespread disease that has evolved from subspeciality to a global health burden. Although the population prevalence of CKD in Sweden is unknown, other countries around the world report about 10-15% of the adult population with this condition. However, CKD is present in a much larger proportion (>35%) among elderly and among high-risk diseases, such as hypertension or diabetes. CKD exacerbates by >20-fold the risk of death due to cardiovascular disease (CVD), and vice versa, but the mechanisms and consequences of this interplay are not fully understood. Mortality risk associated with mild reductions in kidney function is exceedingly high: CKD accounts for 10% of all deaths, a similar magnitude as for diabetes and obesity. The presence of albuminuria in a 30-year old man reduces the life expectancy with 10 years. While most CKD patients will die already before they reach dialysis or renal transplantation (RTx), those receiving dialysis have a 20% annual mortality rate; i.e. comparable to colon and ovarian metastatic cancer. Moreover, the quality of life is low and the average dialysis patient is willing to trade an entire life of dialysis for just six years of life in normal health. Despite its considerable health burden, CKD is ranked amongst the most neglected non-communicable diseases. The economic and social impact of CKD is substantial. Loss of plasticity and/or resilience to adaptions to the changing internal uremic environment promote vascular ageing processes in CKD and result in a marked discrepancy between chronological and biological age.
Inclusion: All end-stage renal disease patients eligible for living-donor renal transplantation.
Control subjects: Patients without renal disease undergoing hernia or cholecystectomy at Ersta Hospital. Blood samples as well as skeletal muscle, subcutaneous fat and arterial biopsies are available. A new arterial control material from healthy living donors at Uppsala University Hospital will be collected during fall 2016.
Collaborators in the project:
Paul Shiels, Professor in Gerontology, Glasgow University, Glasgow, UK
Dagmara McGuiness, PhD Gerontology, Glasgow University, Glasgow, UK
Pieter Evenepoel, Professor, University of Leuven, Leuven, Belgium
Louise Nordfors, PhD Genetics, Karolinska Institutet
Karolina Kublickiene, Assoc Professor, Gynecology, Karolinska Institutet
Anna Witasp, Assistant Professor Genetics, Karolinska Institutet
Maria Eriksson, PhD Genetics, BioNut, Karolinska Institutet
Martin Schalling, Professor Genetics, Center for Molecular Medicine, Karolinska Institutet
Magnus Bäck, Professor Cardiology, Karolinska University Hospital
Ulf Hedin, Professor,
Department of Vascular Surgery Karolinska University Hospital,
Anita Aperia, Professor Pediatrics, Karolinska Institutet
Björn Anderstam, PhD, Chemist, Karolinska Institutet
Lars Wennberg, Docent, Transplantation Dept., Karolinska University Hospital
Mathias Haarhaus, MD, PhD Renal Medicine, Karolinska University Hospital
Peter Barany, Associate Professor, Renal Medicine, Karolinska University Hospital,
Tony Qureshi, MD, PhD, Statistician, Karolinska Institutet
Bengt Lindholm, Professor, Baxter Novum, Karolinska Institutet
Annika Wernerson, Professor, Pathology Karolinska University Hospital
Hannes Olauson, PhD Läkare, Pathology, Karolinska University Hospital
Magnus Söderberg, Associate Professor, Pathology Astra Zeneca
Paolo Parini, Professor, Clinical Chemistry, Karolinska University Hospital
Peter Bergman, Associate Professor, Mikrobiology, Karolinska University Hospital,
Per Magnusson, Associate Professor, Clinical Chemistry, Linköping
Eva Hurt-Camejo, Professor, Astra Zeneca, Gothenburg
Torkel Brismar, Associate Professor, Radiology, Karolinska University Hospital
Jonaz Ripsweden MD, PhD Radiology, Karolinska University Hospital
Anders Thorell Professor, Surgery Ersta Hopsital
Biomarker studies: A wide array of putative biomarkers of ageing, calcification, inflammation and oxidative stress are measured to assess their predictive power for presence of arterial VC, CAC score, cardiovascular and all-cause mortality.
Metabolism: Cholesterol, triglycerides, HDL-cholesterol, ApoA1, ApoB, Lp(a), insulin, glucose, TnT, S-creatinine, hemoglobin, HbA1c, TSH, fT3, fT4, Humanin, ApoA truncation, Angiopoietin-2,
Oxidative stress: 8-OH-dG, pentosidine, AGE reader
Inflammation: hsCRP, IL-6, PTX3, TNF, IL-8
Uremic toxins (In collaboration with Leuven University): Cresyl glucoronide, CMPF, phenylalanine, tyrosine, kynuerenic acid, kynuerine, tryptofan, cresylsulphate, inoxyl sulphate, hippuric acid
Vascular calcification: tu-MGP, uc-dp-MGP, PIVKA-II, fetuin-A, Cardiac CT (coronary artery calcification), Sphygmocor (vascular stiffness)
Bone turnover: PTH, Ca, PO4, ALP, Mg, 25(OH) D-vitamin, 1,25(OH) D-vitamin, osteocalcin, bALP, FGF23, sclerostin, TRAcP5A, TRAP5b, cathepsin K, OPG, RANKL , DXA and Osteoprobe
Microbiota: TMAO, betaine, choline
Circulating aging biomarkers: IGF-1, Klotho, PAPP-A, testosterone, GDF11, miRNA
• Endothelial function, miRNA and microvesicle studies: In collaboration with Karolina Kublickiene resistance arteries are isolated from subcutaneous fat from ESRD pts, RTx recipients and donors. Functional and structural properties of the vessels are studied ex vivo using isometric wire and perfusion myography and related to in vivo measurements of endothelial function (EndoPAT) and other novel markers for increased CVD risk. The role of endothelium-derived pathway/s is assessed together with screening of endothelial dysfunctional markers. Next, expression levels of specific miRNA, regulating pathways involved in inflammation (miR-146a), ageing (miR-217) and eNOS biology (miR-155), are quantified by qPCR. Transfection of arteries with specific miRNA mimic and/or antimiR (LifeTechnologies), followed by functional studies (wire myography) and molecular analysis (qPCR, miR in situ hybridisation and immunohistochemistry) allow us to gain insight in the functional relevance and downstream pathways of miR dysregulation, with this identifying potential therapeutic targets. Pressure myography allows studies on long-term effects and evaluates shear stress, pressure-mediated and pharmacological activation, with different agonists and uremic MVs. Vascular wall distensibility and signs of vascular remodeling, calcification and ageing are also assessed. Studies will link endothelial functional properties to degree (%) of media calcification, ApoA proteoforms and expression of osteogenic progenitor cells (CD34+).
• Linking bone health to vascular calcification: In this part Dr Mathias Haarhaus plays an important role. In collaboration with Per Magnusson, Linköping, we will investigate the association of bALP isoforms with VC, bone turnover and bone mineralization. The novel bALP isoform B1x, which has been detected exclusively in serum from patients with CKD, is associated with low bone turnover and up-regulated in calcified vascular smooth muscle. A better characterization of its role in CKD and ageing could identify it as a possible target for future intervention studies for the prevention of VC. We study if reduced bone mechanical strength is associated with VC and thereby explain for the association of fractures with cardiovascular events in CKD. We will also investigate if statin promotes VC in ESRD via induction of vitamin K deficiency or if statin-induced apoptosis serve as a nidus for calcification. Based on existing and future CT heart examinations we will analyze the independent predictive value of the two components (density and volume) of the CAC score. We will also study if high CAC score and/or low bone mass predicts risk of AF.
• Lipid studies: In collaboration with Paolo Parini, Eva Hurt-Camejo and Matteo Pedrelli, KI, we study the impact of CKD on changes in lipoprotein lipid composition, size and selected functionalities: for HDL, the in vitro cholesterol efflux capacity, antioxidant effects and potential atheroprotective mechanisms, and for LDL, the binding to human arterial proteoglycans, a pro-atherogenic mechanism. In a large CKD cohort we plan to relate proteoglycan-binding to CVD mortality, and lipoprotein features to TMAO.
• Progerin studies: In collaboration with Maria Eriksson BioNut KI, epigastric uremic arteries are quantified for progerin expression, and markers for DNA repair deficiency and premature senescence, using IF microscopy. By droplet digital PCR we will screen DNA for the presence of low-degree mosaicism for nucleotide variants that result in increased progerin production.
• Vascular senescence: In collaboration with prof Paul Shiels Gerontology Department at Glasgow University and Pathology Lab at Karolinska we study arterial expression of senescence markers, miRNAs and markers of DNA damage.
Arterial expression: Telomers, SOST, CDKN2A/p16INK4a, miRNA, Progerin, RUNX2, Matrix Gla protein , p16, p21, p53
Muscular expression: Telomers, CDKN2A/p16INK4a, miRNA
Summary and future: The ultimate goal of this project is to find novel treatment strategies for premature vascular aging. Cross-fertilization of data in sub-studies within the project has already led to unexpected findings of high potential interest. The project already include a RCTs of testosterone. In addition, this project will provide background data to determine if novel interventions, such as sirtuins modulators (play a role in mitochondrial biogenesis), thyroid agonists, mTOR inhibitors (rapamycin), senolytics (dasatinib and quercetin), treatments targeting the microbiota (colchicine and resveratrol) and farnesyl transferase inhibitors should be tested in clinical trials. Testing the effects of such drugs on vascular aging processes will be a main target for the next 5 years. The studies within this project will provide translational information directly relevant for improving years of healthy living in an increasingly aged European population. The scope of this cutting edge and ‘beyond state of the art’ approach will provide proof of principle for novel reparative medicine strategies to treat organ dysfunction with real potential for widespread and dramatic impact on health.