Buerger`s disease (TAO, thromboangiitis obliterans) is an inflammatory vessel disease affecting small and medium sized arteries and veins [1]. The inflammation induces vessel occlusions by formation of mononuclear cell rich thrombi, leading to acral or limbthreatening ischemia syndromes and/or thrombophlebitis [2,3]. Amputation rates are as high as 75% for minor and 30% for major amputations [4]. Although the ethiology of TAO is still unknown tobacco use is considered to be an indispensible trigger-mechanism [2,3].

Pathohistological and immunhistological findings, the detection of increased autoantibody concentrations, and evidence for cytokine activation finally led to the new paradigm of an immunopathogenesis of TAO, first published by Ketha SS and Cooper LT in 2013 [5].

Based on the hypothesis that TAO is an immunmediated disease with humoral factors may play an important pathogenetic role immunadsorption (IA) was successfully introduced in a pilot study by Baumann et al. and later introduced into clinical routine by the corresponding author [6,7]. In 2014 we first described the detection of G-protein coupled receptor autoantibodies in patients with TAO and their removal by IA [8]. Here we present an update of the results of our ongoing clinical project including the results of a clinical follow-up analysis.

From December 2012 to February 2016 twenty-two patients suffering from Buerger`s disease were treated with IA in our institution and included in this observational cohort-study. Retrospectively, diagnosis of TAO was corrected in two patients due to the finding of an markedly elevated Lp(a) concentration resp. the detection of atherosclerotic coronary lesions. In another patient baseline blood samples got lost on transport to the laboratory, leaving 19 patients for final analysis.

Clinical characteristics are presented in Table 1. All patients had already been treated with best medical treatment including intravenous prostanoid infusions (Iloprost and/or PGE1) prior to IA without clinically satisfactory results. IA was performed during a fiveday course with Fesenius-GlobaffinR-adsorbers (Fesenius Medical Care AG & Co KGaA, Bad Homburg v.d. Höhe, Germany) and an intended clearance of the 2.5-fold plasma volume. G-protein coupled - AAB were analysed by E.R.D.E-AKK-Diagnostik GmbH, Berlin, Germany, using peptides corresponding to the first and/or second extracellular loop of α1, Endothelin A, Angiotensin II Type 1, protease activated receptor (PAR) 1/2. Peptides were coupled to pre-blocked streptavidin-coated 96-well plates (Perbio Science, Bonn, Germany). Patient serum was added in a 1:100 dilution and incubated for 60 min. As detection antibody a horseradish peroxidase conjugated antihuman IgG antibody was used (Dianova, Hamburg, Germany). Antibody binding was visualized by the 1-Step Ultra TMB ELISA (Perbio Science, Bonn, Germany). The absorbance was measured at 450 nm with a SLT Spectra multiplate reader (TECAN, Crailsheim, Germany). Clinical follow-up included regular outpatient visits and/or telephone-contacts for patients living in more remote areas. Descriptive statistics were applied.

Table 1: Clinical characteristics of our cohort.

Overall, G-protein receptor auto-antibodies (AAB) were present in 14 of all patients (74%), with one AAB in 5, 2 AAB in 7, 3 AAB in 1 and 4 AAB in 1 patient(s). The presence of a clustering of AAB directed against the α1-receptor and endothelin A-receptor was seen in 9 out of the 14 AAB-positive patients (64%). AAB directed against ETA- receptors never appeared without AAB directed against the α1- receptor and were exclusively directed against the extracellular receptor-loop 1.

After IA at day 5, 12 out of 14 AAB-positive patients were free of AAB. In two patients AAB persisted, while in two others AAB undulated with negative results immediately after IA and reoccurrence next day before re-starting. Even if these cases were considered as elimination-failures, in the vast majority of our patients (71%) AAB against G-protein-receptors were successfully eliminated by the 5 day course of IA.

Clinical follow-up data was available in 15 patients. There were no disease flares. Median visual analog pain-scale (VAS) values improved from median 7.0 (5-9) before IA to 2.0 (0-5) during last follow-up visit. All patients received pain medication before IA. During last follow-up only 2 patients continued to take opioids, while four patients - including one patient suffering from spondylarthritis- took NSAR. In all but one patient skin lesions heeled. Minor amputations- already anticipated before IA - were performed in two patients. Only one patient underwent unexpected major-amputation after a failed pedal bypass procedure of doubtful indication in case of a wound infection with prior subcritical forefoot-tcpO2-values during the last follow-up visit prior to the operation in our institution. 10 patients returned to their former work or education, 4 retired early, one was still on sick leave. Before IA there were 13 active and 6 recently quitting former smokers, during last follow-up there were 12 non-smokers and only 3 active smokers.

AAB directed against G-protein coupled receptors are increasingly recognized as modulators of various cardiovascular pathologies and diabetes mellitus [9-11]. Their prevalence in these conditions varies grossly von 30 to 60%. With our increasing cohort we were able to reproduce preliminary findings showing almost the same range of positive AAB-analyses. Furthermore, we were able to reproduce the predicted clustering of AAB directed against α1-receptor and endothelin A-receptors, which might be of special pathophysiologic interest as both receptors mediate strong vaso constrictive actions consistently with the clinical finding of a strong tendency towards vasospasms in these patients. Deviant unphysiologic and prolonged activation of post-receptor-actions by G-protein coupled AAB have been published and the pathogenic potential of these circulating AAB has already been elucidated in animal models as well as in clinical studies [12-18]. We were able to confirm, that IA successfully eliminated the vast majority of the AAB in a five-day course, with the majority of the eliminations occurring within the first two to three sessions.

More acute beneficial effects of IA in Buerger`s disease have been published and cover a marked improvement in pain, an improvement of digital pulse curves, an incline in tcpO2 and a decline in tcCO2- levels, and – even more important clinically – ulcer healing [6]. Our results indicate that IA might also improve and stabilize the subsequent clinical course of the disease. However, the beneficial outcome was certainly co-influenced by an unusual high rate of smoking cessation in our cohort, making it impossible to define the exact impact of IA on outcomes. Nevertheless, in many cases clinical improvement after IA was observed before smoking was stopped.