Effects of immunoadsorption on methylglyoxal levels 3 Applied Cardiopulmonary Pathophysiology 15: 3-13, 2011 Effects of protein A immunoadsorption on methylglyoxal levels in patients with chronic dilated cardiomyopathy and diabetes mellitus A. O. Doesch1, S. Mueller1, M. Konstandin1, S. Celik1, L. Frankenstein1, C. Zugck1, T. J. Dengler1, T. Fleming2, A. Bierhaus2, H. A. Katus1 1Department of Cardiology, University of Heidelberg, Germany; 2Department of Medici- ne I and Clinical Chemistry, University of Heidelberg, Germany Abstract Objectives: The objective of this study was to investigate effects of immunoadsorption (IA) on methylglyoxal (MG) levels in patients with chronic non-familial dilated cardiomyopathy (DCM), as well as clinical and humoral markers of heart failure. Background: Previous studies have demonstrated favourable outcomes of IA in DCM patients with diabetes mellitus (DM) and that MG sensitizes cultured cardiomyocytes to injury by post- translational modification of Thioredoxin via glycation. Therapeutic interventions scavenging advanced glycation end-products (AGE) precursors may attenuate myocardial injury in diabet- ic patients. Possible effects of IA on MG levels in patients with DCM and DM have never been analyzed before. Methods: We performed IA using agarose columns on five consecutive days in 10 patients with chronic DCM and DM, congestive heart failure of NYHA class ≥ II, left ventricular ejec- tion fraction (LVEF) ≤ 50 %, and mean time since initial diagnosis of 4.7 ± 3.9 years. Results: Immediately after IA, IgG decreased from 10.7 ± 1.9 to 1.1 ± 0.6 g/L (89.7 %, P = 0.008) and IgG3 from 0.6 ± 0.2 to 0.2 ± 0.2 g/L (66.7 %, P = 0.01). Median NT-pro BNP was reduced from 1665.0 ng/L at baseline to 1163.0 ng/L after 6 months (P = 0.04). Also mean LVEF was significantly improved (25.5 ± 11.7 % to 30.9 ± 11.9 % after 6 months, P = 0.02) and LVEF improved ≥ 5% (absolute) in 7 of 10 (70.0 %) patients. After 6 months, bicycle spiroer- gometry showed a significant increase in exercise capacity from 73.3 ± 15.8 Watts to 93.3 ± 16.4 Watts (P = 0.04) while VO2max rose from 13.0 ± 2.4 to 15.0 ± 2.2 mL/min kg (P = 0.05). No significant changes in MG levels 6 months after IA (169.6 ± 56.9 nM to 208.7 ± 75.2 nM, P = 0.05) were noted. Conclusions: In this study on patients with nonfamilial DCM and DM, IA therapy significantly improved clinical and humoral markers of heart failure severity. However, no significant changes in MG levels were observed. Therefore, these promising results in diabetic DCM pa- tients may not be caused by altering AGE levels. Future blinded prospective multicenter stud- ies are necessary to identify possible underlying mechanisms like polyvalent antibodies in dia- betic patients. Key words: clinical effects, diabetes mellitus, dilated cardiomyopathy, methylglyoxal, protein A immunoadsorption 4 A. O. Doesch et al. Abbreviations protein A immunoadsorption (IA) in patients with chronic dilated cardiomyopathy (DCM) advanced glycation end-products (AGE) and DM [12,13]. This study analyzed possible Angiotensin-Converting Enzyme (ACE) effects of protein A IA on MG levels in pa- angiotensin (AT) tients with chronic DCM and DM. cardiac resynchronization therapy (CRT) C-reactive protein (CRP) diabetes mellitus (DM) Methods dilated cardiomyopathy (DCM) immunoadsorption (IA) Study patients immunoglobulin G (IgG) international normalized ratio (INR) 10 patients (10 male [100.0 percent of total]) left ventricular ejection fraction (LVEF) with DM and chronic non-familial DCM methylglyoxal (MG) (mean symptomatic disease duration 4.7 ± New York Heart Association (NYHA) 3.9 years, no relative diagnosed with DCM, partial thromboplastin time (PTT) conduction defects, arrhythmias, syncope, muscle weakness, or history of first-degree relative with an unexplained sudden death Background under the age of 35 years [14]) were includ- ed in the study. Prior to study entry all pa- DM is a leading metabolic disorder associat- tients had a reduced left ventricular function, ed with severe systemic consequences if i.e. LVEF below 50%, and were in New York poorly managed in the clinical setting. Con- Heart Association functional class II or worse. siderable experimental and clinical studies Detailed patient characteristics and baseline have demonstrated the close association be- laboratory values are summarized in table 1. tween diabetes and significant cardiovascular All patients gave written informed consent morbidity and mortality [1-4]. Methylglyoxal prior to study entry. Preceeding study inclu- (MG), a highly reactive dicarbonyl, is a natu- sion, coronary heart disease was excluded by ral metabolite in glucose metabolism. It is ca- coronary angiography. All patients had been pable of inducing the non-enzymatic reaction taking a stable dose of oral medications (be- glycation, or glycosylation, between reducing ta blockers, Angiotensin-Converting Enzyme sugars and proteins and other biomolecules, (ACE) inhibitors, angiotensin (AT) 1 antago- yielding irreversible advanced glycation end- nists, aldosterone antagonists, diuretics) for at products (AGE). Elevated MG levels are be- least three months before study entry (table lieved to contribute to complications seen in 1) and heart failure medication was held con- poorly controlled diabetic states [5-8]. It has stant during the 6 month follow-up. Exclusion been demonstrated, that MG is capable of criteria were active or chronic infectious dis- sensitizing cultured cardiomyocytes to injury eases, malignancies, chronic alcoholism, by post-translational modification of thiore- pregnancy, or heart failure due to other doxin via glycation [9]. Additionally, recent in- known origins. vestigations have shown that MG induces apoptosis of rat Schwann cells [10] and hu- man vascular endothelial cells [11], indicating Immunoadsorption the significant role MG plays in the etiology of diabetic complications. Therapeutic inter- Immunoadsorption was performed in 10 pa- ventions scavenging AGE precursors may at- tients with DCM, using a central venous tenuate myocardial injury in hyperglycemic catheter to ensure adequate flow rates, with state diseases such as diabetes. Previous stud- one session daily on five consecutive days. ies have demonstrated favourable effects of Immunoglobulin extraction was achieved Effects of immunoadsorption on methylglyoxal levels 5 Table 1: Patient characteristics at baseline (n=10) parameter mean standard deviation patient age [years] 58.0 4.9 time since initial diagnosis [years] 4.7 3.9 NYHA functional class n percent of total NYHA II 8 80.0 NYHA III 2 20.0 cardiac resynchronization therapy device 4 40.0 parameter mean standard deviation LVEF [%] 25.5 11.7 bicycle spiroergometry data: exercise capacity [Watt] 73.3 15.8 VO2max [mL/min*kg] 13.0 2.4 comedication n percent of total beta blocker 10 100.0 ACE inhibitor and/or AT1 antagonist 10 100.0 diuretic 8 80.0 digitalis 6 60.0 aldosterone antagonist 3 30.0 laboratory parameter mean standard deviation sodium [mmol/L] 138.7 2.0 potassium [mmol/L] 4.1 0.6 calcium [mmol/L] 2.4 0.1 urea [mg/dL] 53.2 23.1 creatinine [mg/dL] 1.2 0.4 glucose [mg/dL] 131.1 24.0 haemoglobin [g/dL] 13.3 1.1 haematocrit [%] 39.1 3.0 cholesterol [mg/dL] 172.2 34.1 LDL [mg/dL] 103.3 36.3 HDL [mg/dL] 40.4 12.7 triglycerides [mg/dL] 234.4 297.0 CRP [mg/L] 7.3 5.9 total IgG 10.7 1.9 IgG 3 [g/L] 0.6 0.2 NT-pro BNP [ng/L] 1665.0 (median) with a protein A column (Immunosorba®, Fre- an AT 1 antagonist 7 days prior to IA treat- senius Medical Care, Bad Homburg, Ger- ment. Fresenius Art-Universal was used for many), as described recently [15] and ACE in- plasma separation and the Citem 10 monitor hibitors were discontinued and replaced by (Fresenius Medical Care, Bad Homburg, Ger- 6 A. O. Doesch et al. many) was used to control flow through the the rest of the infusion until the calculated in- columns. Each session was performed to fusion volume has been reached. yield an optimized IgG3 removal (detailed Laboratory tests (including CRP, blood protocol, including plasma flow/volume and count, INR, PTT, and electrolytes) were per- cycle length: table 2 [according to Fresenius formed on day 1, 3, and 5 of IA treatment to Medical Care, Bad Homburg, Germany]). The check electrolyte and coagulation status and 2.5-fold plasma volume was treated every to exclude acute infections. day. Patients’ plasma volume was calculated as previously described [16]. Patients’ body weight was measured daily before and after Follow-up IA treatment and diuretics were administered if patient weight following the IA session had At baseline and 6 months after IA, bicycle increased by more than 1 kg. During the spiroergometry, transthoracic echocardiogra- whole IA session full anticoagulation with un- phy, and routine laboratory tests (including fractionated heparin was performed to avoid electrolytes, blood count, CRP, NT-pro BNP) coagulation in the extracorporeal system (ac- were obtained. cording to manufacturer’s instruction, Frese- nius Medical Care, Bad Homburg, Germany). Patients on warfarin included in the current Echocardiography study were switched to low molecular weight heparin 7 days prior to apheresis treatment. Two-dimensional echocardiography was During the hospital stay full anticoagulation done by experienced operators, unaware of with unfractionated heparin was applied. the investigation. A standardized imaging After completion of five treatment cycles protocol was adopted with cross-sectional im- (day 5), polyclonal IgG (0.5 g/kg, Octagam, aging of