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Lipoprotein (a) and long-term outcome in patients with peripheral artery disease undergoing revascularization

Open AccessPublished:October 12, 2022DOI:https://doi.org/10.1016/j.atherosclerosis.2022.10.002

      Highlights

      • The predictive capability of Lp(a) for cardiovascular mortality was evaluated over a long-term period in two cohorts of symptomatic peripheral artery disease (PAD).
      • Lp(a) was not associated with cardiovascular mortality in both cohorts.
      • No specific pattern of lesion site (iliacal, femoral, below the knee, multivessel) for endovascular repair was detected with elevated Lp(a) levels.
      • This data suggests that elevated Lp(a) does not impose an additional risk to patients with symptomatic PAD and high rates of statin treatment.

      Abstract

      Background and aims

      Despite low LDL-C goals, the residual risk for further cardiovascular (CV) events in patients with peripheral artery disease (PAD) remains high. Lipoprotein (a) (Lp(a)) is a known risk factor for PAD incidence, but little is known regarding the outcome in patients with symptomatic PAD. Thus, this study investigates Lp(a) and CV mortality in PAD after endovascular repair.

      Methods

      A total of 1222 patients with PAD in two cohorts according to Lp(a) assay in nmol/L (n = 964, Lip-LEAD-A) or mg/dl (n = 258, Lip-LEAD-B) were followed up for 4.3 (IQR 3.0–5.6) or 7.6 (IQR 3.2–8.1) years. Lp(a) was measured before endovascular repair for either intermittent claudication (IC) or critical limb ischemia (CLI). Outcome information was obtained from the federal death registry.

      Results

      In Lip-LEAD-A, 141 CV-deaths occurred (annual calculated CV-death rate 3.4%), whereas 64 CV-deaths were registered in Lip-LEAD-B (annual calculated CV-death rate 3.3%). After adjustment for traditional CV risk factors Lp(a) was neither associated with outcome in Lip-LEAD-A (highest tertile HR 1.47, 95%CI [0.96–2.24]) nor in Lip-LEAD-B (highest tertile HR 1.34 [0.70–2.58]). Subanalyses for IC (HR 1.37 [0.74–2.55]; HR 1.10 [0.44–2.80], CLI (HR 1.55 [0.86–2.80], HR 3.01 [0.99–9.10]), or concomitant coronary artery disease (CAD; HR 1.34 [0.71–2.54]; HR 1.21 [0.46–3.17]) failed to show a significant association between Lp(a) and CV-mortality.

      Conclusions

      In this large-scale cohort of symptomatic PAD no association of elevated Lp(a) with CV mortality was found over a median observation period of 5 years. Thus, an even longer study including asymptomatic patients is warranted.

      Graphical abstract

      Keywords

      1. Introduction

      Despite recent advances in secondary prevention of cardiovascular (CV) disease, the residual risk for future CV events remains high [
      • Baigent C.
      • Blackwell L.
      • Emberson J.
      • et al.
      Efficacy and safety of more intensive lowering of LDL cholesterol: a meta-analysis of data from 170,000 participants in 26 randomised trials.
      ]. This particularly extends to patients with peripheral artery disease (PAD), a CV disease that can be seen as a manifestation of systemic and advanced atherosclerotic disease and burden. Lowering of LDL-C, the most relevant objective of secondary prevention in the lipid spectrum, was shown to reduce CV events in statin trials [
      • Baigent C.
      • Keech A.
      • Kearney P.M.
      • et al.
      Efficacy and safety of cholesterol-lowering treatment: prospective meta-analysis of data from 90,056 participants in 14 randomised trials of statins.
      ], as well as in the PCSK9i trial FOURIER [
      • Bonaca M.P.
      • Nault P.
      • Giugliano R.P.
      • et al.
      Low-density lipoprotein cholesterol lowering with evolocumab and outcomes in patients with peripheral artery disease: insights from the FOURIER trial (further cardiovascular outcomes research with PCSK9 inhibition in subjects with elevated risk).
      ], with a distinct PAD cohort. However, in an era of low or even very-low LDL-C treatment goals, according to the most recent ESC guidelines [
      • Mach F.
      • Baigent C.
      • Catapano A.L.
      • et al.
      ESC/EAS Guidelines for the management of dyslipidaemias: lipid modification to reduce cardiovascular risk.
      ] on lipid-lowering treatment, effects of lipoprotein particles heterogeneity on atherosclerotic disease progression, and thus morbidity and mortality, might be more relevant.
      Lipoprotein(a) (Lp(a)), an LDL particle with apolipoprotein(a) attached to apolipoprotein B100, might be one candidate. Lp(a) bears several proatherogenic properties in advance to the LDL particles mediated effects such as selective binding of apo(a) to the extracellular matrix, and the amount of oxidized phospholipids, covalently bound to apo(a) [
      • van der Valk F.M.
      • Bekkering S.
      • Kroon J.
      • et al.
      Oxidized phospholipids on lipoprotein(a) elicit arterial wall inflammation and an inflammatory monocyte response in humans.
      ,
      • Tsimikas S.
      • Brilakis E.S.
      • Miller E.R.
      • et al.
      Oxidized phospholipids, Lp(a) lipoprotein, and coronary artery disease.
      ]. Levels of Lp(a) are predominantly genetically determined [
      • Clarke R.
      • Peden J.F.
      • Hopewell J.C.
      • et al.
      Genetic variants associated with Lp(a) lipoprotein level and coronary disease.
      ,
      • Saleheen D.
      • Haycock P.C.
      • Zhao W.
      • et al.
      Apolipoprotein(a) isoform size, lipoprotein(a) concentration, and coronary artery disease: a mendelian randomisation analysis.
      ]. Epidemiologic, cross-sectional, and prospective data, as well as Mendelian randomization studies, link Lp(a) to both morbidity and mortality in CV disease in primary and secondary prevention [
      • Erqou S.
      • Kaptoge S.
      • Perry P.L.
      • et al.
      Lipoprotein(a) concentration and the risk of coronary heart disease, stroke, and nonvascular mortality.
      ,
      • Kronenberg F.
      • Kronenberg M.F.
      • Kiechl S.
      • et al.
      Role of lipoprotein(a) and apolipoprotein(a) phenotype in atherogenesis: prospective results from the Bruneck study.
      ,
      • O'Donoghue M.L.
      • Morrow D.A.
      • Tsimikas S.
      • et al.
      Lipoprotein(a) for risk assessment in patients with established coronary artery disease.
      ,
      • Patel A.P.
      • Wang M.
      • Pirruccello J.P.
      • et al.
      Lp(a) (Lipoprotein[a]) concentrations and incident atherosclerotic cardiovascular disease: new insights from a large national biobank.
      ].
      For PAD, higher levels of Lp(a) were associated with incidental PAD in a prospective cohort in Hong Kong [
      • Cheng S.W.
      • Ting A.C.
      • Wong J.
      Lipoprotein (a) and its relationship to risk factors and severity of atherosclerotic peripheral vascular disease.
      ] and a cross-sectionally observed cohort in Taiwan [
      • Tseng C.H.
      Lipoprotein(a) is an independent risk factor for peripheral arterial disease in Chinese type 2 diabetic patients in Taiwan.
      ]. In two large European cohorts in the primary setting, the EPIC-Norfolk study and the Edinburgh Artery Study, Lp(a) levels were similarly associated with incidental PAD [
      • Gurdasani D.
      • Sjouke B.
      • Tsimikas S.
      • et al.
      Lipoprotein(a) and risk of coronary, cerebrovascular, and peripheral artery disease: the EPIC-Norfolk prospective population study.
      ,
      • Tzoulaki I.
      • Murray G.D.
      • Lee A.J.
      • et al.
      Inflammatory, haemostatic, and rheological markers for incident peripheral arterial disease: Edinburgh Artery Study.
      ]. On the other side, two different prospective large-scale studies failed to find statistical significance with incidental PAD and Lp(a) [
      • Ridker P.M.
      • Stampfer M.J.
      • Rifai N.
      Novel risk factors for systemic atherosclerosis: a comparison of C-reactive protein, fibrinogen, homocysteine, lipoprotein(a), and standard cholesterol screening as predictors of peripheral arterial disease.
      ,
      • Pradhan A.D.
      • Shrivastava S.
      • Cook N.R.
      • et al.
      Symptomatic peripheral arterial disease in women: nontraditional biomarkers of elevated risk.
      ]. In the secondary prevention setting for patients with PAD, three studies reported a significant association between higher Lp(a) levels and major adverse limb events (MALE) or major adverse cardiac events (MACE) [
      • Hishikari K.
      • Hikita H.
      • Nakamura S.
      • et al.
      Usefulness of lipoprotein(a) for predicting clinical outcomes after endovascular therapy for aortoiliac atherosclerotic lesions.
      ,
      • Golledge J.
      • Rowbotham S.
      • Velu R.
      • et al.
      Association of serum lipoprotein (a) with the requirement for a peripheral artery operation and the incidence of major adverse cardiovascular events in people with peripheral artery disease.
      ,
      • Verwer M.C.
      • Waissi F.
      • Mekke J.M.
      • et al.
      High Lipoprotein(a) Is Associated with Major Adverse Limb Events after Femoral Artery Endarterectomy.
      ]. However, only one study reported data on all-cause mortality and did not find any association [
      • Golledge J.
      • Rowbotham S.
      • Velu R.
      • et al.
      Association of serum lipoprotein (a) with the requirement for a peripheral artery operation and the incidence of major adverse cardiovascular events in people with peripheral artery disease.
      ]. This aforementioned study by Golledge et al. [
      • Golledge J.
      • Rowbotham S.
      • Velu R.
      • et al.
      Association of serum lipoprotein (a) with the requirement for a peripheral artery operation and the incidence of major adverse cardiovascular events in people with peripheral artery disease.
      ] showed data for a combined cohort of patients with abdominal aortic aneurysm and lower extremity artery disease. The other studies that reported MACE or MALE had low event rates and limited total numbers of included patients [
      • Hishikari K.
      • Hikita H.
      • Nakamura S.
      • et al.
      Usefulness of lipoprotein(a) for predicting clinical outcomes after endovascular therapy for aortoiliac atherosclerotic lesions.
      ,
      • Verwer M.C.
      • Waissi F.
      • Mekke J.M.
      • et al.
      High Lipoprotein(a) Is Associated with Major Adverse Limb Events after Femoral Artery Endarterectomy.
      ]. To our knowledge, no further study evaluated CV mortality and Lp(a) in a specific PAD cohort.
      Thus, this study evaluates a possible association of Lp(a) in patients with PAD and long-term CV mortality after endovascular repair either for critical limb ischemia (CLI) or intermittent claudication (IC) in a single-center retrospective cohort.

      2. Patients and methods

      2.1 Patients and study design

      The Vienna Lip-LEAD (LIPids in Lower Extremity Artery Disease) study was designed to evaluate lipid profiles and lipid treatment goals in patients with peripheral artery disease undergoing endovascular repair. The study was approved by the institutional ethics committee and follows the Declaration of Helsinki and contemporary Good Clinical Practice guidelines. All patients treated with endovascular repair of the lower limb arteries at the inpatient department of the Division of Angiology, Medical University of Vienna, were included retrospectively from 1/Jan/2013 to 31/Dec/2018. Only patients with a first admission during this time at the inpatient department were analyzed in this study; further admissions of the same patient for endovascular repair or any other reason were not included. PAD is used as the term for lower extremity artery disease (LEAD) in this study. Patients with missing values of Lp(a) or covariates, an eGFR<15 mg/dl, and prior organ transplantation were excluded from this analysis (flow-chart, Fig. 1). Endovascular repair was indicated either by symptomatic PAD (Fontaine stage II-IV) or in Fontaine stage I in the case of critical stenosis of a previously implanted bypass graft or stent. For diagnostic work-up, all patients received both a hemodynamic assessment (oscillometry and ankle-brachial index measurement) and further imaging (either ultrasonography or contrast-enhanced CT/MR scans) for endovascular repair planning. Ankle-brachial index (ABI) measurement, as defined by the ratio of the brachial and ankle blood pressure according to the TASC II criteria [
      • Norgren L.
      • Hiatt W.R.
      • Dormandy J.A.
      • et al.
      Inter-society consensus for the management of peripheral arterial disease (TASC II).
      ], was performed by experienced and specially trained staff with Doppler sonography (ELCAT, Wolfratshausen, Germany). Ultrasonography was performed by specially trained technicians at the Division of Angiology. CT- or MR-scans were performed at the Department of Radiology, Medical University of Vienna, according to standard examination protocols.
      Routine laboratory parameters were measured on the day of admission, including kidney function, liver function, and a lipid profile (total cholesterol, HDL-C, and Lp(a)) by the laboratory department. LDL-C was calculated according to the Friedewald formula. Lp(a) was included in this routine laboratory testing and measured on a Roche Diagnostics platform. Lp(a) was measured in mg/dL until 17/Feb/2014 and after assay change of the central laboratory in nmol/L. Since mg/dl and nmol/L levels of Lp(a) cannot simply be transformed due to different molar masses according to KIV-2 kringle repeats [
      • Langlois M.R.
      • Chapman M.J.
      • Cobbaert C.
      • et al.
      Quantifying atherogenic lipoproteins: current and future challenges in the era of personalized medicine and very low concentrations of LDL cholesterol. A consensus statement from eas and eflm.
      ], and no assay comparison was performed, patients were separately analyzed in a mg/dL (Lip-LEAD-B) cohort and a nmol/L (Lip-LEAD-A) cohort.
      Medical history including previous myocardial infarction, stroke, PAD, cerebrovascular disease/carotid stenosis, diabetes mellitus, arterial hypertension, and smoking history was recorded from available medical records. Data on medication was registered from the discharge letter for endovascular repair.
      Information on patients’ survival was obtained via queries to the central death registry of the federal department of statistics (Statistik Austria) on 31/Aug/2021. ICD-codes of the I category were categorized as CV-death.
      Medical artwork was used from Servier medical arts under a creative commons license (CCA 3.0 unported license).

      2.2 Definition of parameters

      Peripheral artery disease (PAD), was diagnosed by ABI measurement with a resting ABI <0.9 and clinically graded according to the Fontaine classification. The grade of stenosis was assessed by imaging either with ultrasonography, CT, or MR angiography. Fontaine stage II (both IIa and IIb) was defined as intermittent claudication (IC) and both Fontaine stage III (resting pain) and IV (ulcers) were defined as critical limb ischemia (CLI).
      Diabetes mellitus was defined according to the current ADA guidelines of 2021 [
      2. Classification and diagnosis of diabetes: standards of medical care in diabetes-2021.
      ] by either an HbA1c > 6.5 rel.%, previously known diabetes mellitus, and/or previous antidiabetic medication. In the case of SGLT2i, no other indication besides diabetes mellitus type 2 existed in the country of this study until the end of inclusion (December 2018).
      Coronary artery disease was defined as previously known myocardial infarction, percutaneous coronary intervention, or ischemic cardiomyopathy.
      Hypertension was defined according to medical records (discharge summaries) and verified by the intake of antihypertensive treatment.
      Statin doses were calculated as atorvastatin equivalency doses for better comparison according to Naci et al. [
      • Naci H.
      • Brugts J.J.
      • Fleurence R.
      • et al.
      Dose-comparative effects of different statins on serum lipid levels: a network meta-analysis of 256,827 individuals in 181 randomized controlled trials.
      ].
      The estimated glomerular filtration rate (eGFR) was calculated according to the CKD-EPI formula of 2012 [
      • Inker L.A.
      • Schmid C.H.
      • Tighiouart H.
      • et al.
      Estimating glomerular filtration rate from serum creatinine and cystatin C.
      ].
      Lesion sites were subjected to four categories: 1 iliac region including kissing stenting of the common iliac artery; 2 femoral-popliteal regions; 3 below the knee repair; 4 multiple regions (multisite).

      2.3 Statistical analysis

      The entire statistical analysis was performed with SPSS 27.0 (SPSS Inc. Chicago, IL, USA). Figures were drawn with GraphPad Prism 9.0 (GraphPad Software Inc., San Diego, CA, USA). Data are presented as mean ± standard deviation (SD) or median and percentiles (25th, 75th), as appropriate. A two-sided p-value <0.05 was defined as statistically significant. Student's t-tests or nonparametric equivalents and chi-square tests were applied, as appropriate. Lp(a) levels were divided into tertiles for better comparison. Kaplan-Meier analyses were assessed with a log-rank test and performed with tertiles, respectively. Cox regression analyses were performed categorically with the same grouping. A multivariable-adjusted model was performed with traditional CV risk factors (age, sex, T2DM, LDL-C, HDL-C, arterial hypertension, ezetimibe usage, BMI, Fontaine stage, and CKD-EPI). To evaluate a possible non-linear association of Lp(a) with CV-death, this model was included as a restricted cubic spline with 3 knots in the model. Therefore, R Version 4.0.3 (R Foundation for Statistical Computing, Vienna, Austria) and packages “rms”, “survminer” and “ggplot2” were utilized. Subanalyses were performed with Cox-regression analyses for intermittent claudication, critical limb ischemia, the combination of coronary artery disease and PAD. Subanalyses for LDL-C control <70 mg/dl and <55 mg/dl were only performed in Lip-LEAD-A due to the small sample number as well as overfitting and underpowering issues in Lip-LEAD-B.

      3. Results

      3.1 Baseline characteristics

      3.1.1 LIP-LEAD-A (nmol/L) group

      A total of 964 patients were included in the final analysis and divided into tertiles. Traditional CV risk factors (age, T2DM, hyperlipidemia, smoking) were equally distributed between the tertiles. However, a significantly lower systolic blood pressure was found in the highest vs. the lowest tertile (p = 0.033). Furthermore, a steady significant increase in the percentage of women in higher tertiles was found, ranging from 32.2% in the lowest group to 48.3% in the highest group (overall p<0.001, 1 vs. 3 p<0.001). BMI was higher in the lower tertiles, but only a trend was observed between the highest vs. the lowest tertile overall p = 0.044, 1 vs. 3 p = 0.013). Similar to most traditional CV risk factors, preexisting CV disease, including previously known PAD, stroke, myocardial infarction, CAD, and atrial fibrillation, were equally distributed between the groups. Regarding laboratory values, significantly higher levels of LDL-C (overall p<0.001, 1 vs. 3 p<0.001), HDL-C (overall p = 0.012, 1 vs. 3 p = 0.003), and serum total cholesterol (overall p<0.001, 1 vs. 3 p<0.001) were found in the highest tertile. CV secondary prevention treatment was similar in all groups apart from ezetimibe usage. The highest group had significantly higher prescription rates of ezetimibe while overall the percentage was low (6.6% lowest vs. 11.8% highest tertile, overall p = 0.025, 1 vs. 3 p = 0.021). A detailed overview can be seen in Table 1.
      Table 1Baseline characteristics of Lip-LEAD-A.
      1st Tertile2nd Tertile3rd Tertilep-value1 vs. 3
      n = 320n = 323n = 321
      Lipoprotein (a) nmol/l7 (7, 8)22 (16, 34)180 (108, 230)<0.001<0.001
      Age (yrs)69 ± 1068 ± 1270 ± 110.0970.529
      Female Sex n(%)103 (32.2%)123 (38.1%)155 (48.3%)<0.001<0.001
      Systolic Blood Pressure mmHg148 ± 24145 ± 24139 ± 190.0880.033
      Diastolic Blood Pressure mmHg77 ± 1077 ± 1275 ± 120.3350.516
      BMI kg/m227.4 ± 5.427.0 ± 4.726.4 ± 4.90.0440.013
      Fontaine stage n(%)Fontaine 1/2234 (73.4%)242 (74.9%)229 (71.3%)0.8250.838
      Fontaine 3/485 (26.6%)81 (25.1%)92 (28.7%)
      Previous PAD n(%)97 (30.3%)106 (32.8%)99 (30.8%)0.7700.885
      Ankle-Brachial-Index0.57 ± 0.200.59 ± 0.190.55 ± 0.190.1130.217
      Toe-Brachial-Index0.58 ± 0.230.59 ± 0.220.55 ± 0.200.1080.072
      T2DM n(%)138 (43.1%)135 (41.8%)134 (41.7%)0.9230.724
      Art. Hypertension n(%)281 (87.8%)292 (90.4%)290 (90.3%)0.4740.304
      Hyperlipidemia n(%)269 (84.1%)283 (87.6%)278 (86.6%)0.4070.363
      Active Smoking n(%)103 (32.2%)110 (34.1%)116 (36.1%)0.1350.067
      CAD n(%)94 (29.4%)107 (33.1%)105 (32.7%)0.5350.361
      Stroke n(%)26 (8.1%)24 (7.4%)28 (8.7%)0.8340.785
      Myocardial Infarction n(%)36 (11.3%)56 (17.3%)43 (13.4%)0.0780.409
      Atr. Fibrillation n(%)37 (11.6%)38 (11.8%)34 (10.6%)0.8820.695
      Serum Creatinine mg/dl1.06 ± 0.401.06 ± 0.411.04 ± 0.420.7240.430
      eGFR (CKD-EPI) ml/min/1.73m285.1 ± 25.187.0 ± 27.187.1 ± 26.40.5400.319
      LDL-C mg/dl82.5 ± 34.290.0 ± 37.393.1 ± 39.70.001<0.001
      HDL-C mg/dl48 ± 1650 ± 1753 ± 190.0120.003
      HbA1c rel.%6.3 ± 1.16.4 ± 1.36.2 ± 1.10.1420.655
      Triglycerides mg/dl127 (98, 177)121 (90, 162)125 (89, 172)0.2210.337
      Serum total Cholesterol mg/dl161 ± 41168 ± 45174 ± 460.001<0.001
      C-reactive Protein mg/dl0.28 (0.13, 0.67)0.35 (0.15, 1.05)0.34 (0.12, 0.83)0.1050.338
      ACEi n(%)135 (42.2%)141 (43.7%)107 (33.3%)0.0150.021
      AT2 n(%)118 (36.9%)116 (35.9%)138 (43.0%)0.1360.114
      Beta-Blocker n(%)154 (48.1%)160 (49.5%)159 (49.5%)0.9190.721
      Antihypertensive n(%)266 (83.1%)257 (79.6%)261 (81.3%)0.5120.548
      ASS n(%)271 (84.7%)285 (88.2%)284 (88.5%)0.2760.160
      Statin n(%)275 (85.9%)295 (91.3%)281 (87.5%)0.0920.550
      Statin class usage n(%)Simvastatin149 (46.6%)166 (51.4%)154 (48.0%)0.6860.980
      Rosuvastatin39 (12.2%)39 (12.1%)36 (11.2%)
      Atorvastatin81 (25.3%)84 (26.0%)86 (26.8%)
      Fluvastatin5 (1.6%)3 (0.9%)4 (1.2%)
      Lovastatin1 (0.3%)3 (0.9%)1 (0.3%)
      Ezetimibe n(%)21 (6.6%)22 (6.8%)38 (11.8%)0.0250.021
      Atorvastatin Equivalency Dosages mg33.4 ± 25.734.6 ± 25.733.6 ± 25.80.8070.931
      Patients are presented in tertiles according to their Lp(a) nmol/L level. BMI body mass index. Data are shown as mean ± standard deviation or median and interquartile range, as applicable. p < 0.05 (two-sided) was considered statistically significant.
      PAD peripheral artery disease, T2DM type 2 diabetes mellitus, CAD coronary artery disease, Atr. Fibrillation atrial fibrillation, eGFR estimated glomerular filtration rate, LDL-C low-density lipoprotein cholesterol, HDL-C high-density lipoprotein cholesterol, HbA1c hemoglobin A1c, ACEi angiotensin converting enzyme inhibition treatment, AT2 angiotensin 2 receptor blockage treatment, ASS acetylic salicylic acid treatment. Bold values in the p-value column depict significant results.

      3.1.2 LIP-LEAD-B (mg/dl) group

      A total of 258 patients were included in the final analysis of this group. Patients were divided into tertiles of Lp(a) for better comparison. Similar to the nmol/L group traditional risk factors were equally distributed between the tertiles and no statistically significant difference in sex and BMI was found. Furthermore, previous CV diseases were equally distributed between the tertiles, and percentages of medication for secondary prevention did not differ between the groups. A detailed overview can be seen in Table 2. No significant increase in HDL-C, LDL-C, or TC was seen (in difference from the first group).
      Table 2Baseline characteristics of Lip-LEAD-B.
      1st Tertile2nd Tertile3rd Tertilep-value1 vs. 3
      n = 85n = 89n = 84
      Lipoprotein (a) mg/dl3 (3,5)16 (11, 22)68 (52, 108)<0.001<0.001
      Age (yrs)68 ± 1270 ± 1068 ± 110.5500.927
      Female Sex n(%)29 (34.1%)27 (30.3%)32 (38.1%)0.5610.590
      BMI kg/m226.2 ± 4.626.4 ± 4.626.8 ± 4.40.3860.395
      Fontaine stage n(%)Fontaine 1/264 (75.3%)68 (76.4%)57 (67.9%)0.4230.360
      Fontaine 3/4 CLI21 (24.7%)21 (23.6%)27 (32.1%)
      Previous PAD n(%)37 (43.5%)42 (47.2%)46 (54.8%)0.3290.144
      Ankle-Brachial-Index0.61 ± 0.210.55 ± 0.210.58 ± 0.150.6020.560
      Toe-Brachial-Index0.52 ± 0.170.51 ± 0.130.34 ± 0.20.1070.093
      T2DM n(%)32 (37.6%)41 (46.1%)37 (44%)0.5060.397
      Art. Hypertension n(%)82 (96.5%)78 (87.6%)80 (95.2%)0.0460.688
      Hyperlipidemia n(%)79 (92.9%)79 (88.8%)77 (91.7%)0.6110.756
      Active Smoking n(%)33 (38.8%)35 (39.3%)29 (34.5%)0.7760.562
      CAD n(%)24 (28.2%)31 (34.8%)34 (40.5%)0.2460.094
      Stroke n(%)11 (12.9%)10 (11.2%)9 (10.7%)0.8940.654
      Myocardial Infarction n(%)9 (10.6%)17 (19.1%)19 (22.6%)0.1050.035
      Atr. Fibrillation n(%)6 (7.1%)13 (14.6%)6 (7.1%)0.1530.983
      Serum Creatinine mg/dl1.0 ± 0.41.1 ± 0.521.1 ± 0.50.1640.226
      eGFR (CKD-EPI) ml/min/1.73m289.9 ± 25.181.9 ± 27.185.9 ± 27.40.1460.329
      LDL-C mg/dl88.1 ± 39.188.9 ± 34.793.4 ± 34.20.6000.360
      HDL-C mg/dl51 ± 1650 ± 1750 ± 180.8410.832
      HbA1c rel.%6.2 ± 1.06.3 ± 1.16.3 ± 1.10.5090.243
      Triglycerides mg/dl133 (90, 179)129 (92, 185)124 (94, 171)0.8650.586
      Serum total Cholesterol mg/dl169 ± 44169 ± 46175 ± 420.6420.383
      C-reactive Protein mg/dl0.24 (0.14, 0.61)0.31 (0.12, 0.65)0.32 (0.12, 1.07)0.6750.386
      ACEi n(%)40 (47.1%)33 (37.1%)35 (41.7%)0.4100.143
      AT2 n(%)30 (35.3%)31 (34.8%)32 (38.1%)0.8910.706
      Beta-Blocker n(%)38 (44.7%)44 (49.4%)46 (54.8%)0.4250.191
      Antihypertensive n(%)66 (77.6%)75 (84.3%)71 (84.5%)0.4120.254
      ASS n(%)78 (91.8%)81 (91%)80 (95.2%)0.5290.360
      Statin n(%)70 (82.4%)80 (89.9%)75 (89.3%)0.2600.197
      Statin class usage n(%)Simvastatin29 (34.1%)29 (32.6%)23 (27.4%)0.1260.168
      Rosuvastatin6 (7.1%)8 (9.0%)16 (19.0%)
      Atorvastatin33 (38.8%)36 (40.4%)35 (41.7%)
      Fluvastatin1 (1.2%)4 (4.5%)1 (1.2%)
      Lovastatin1 (1.2%)3 (3.4%)0 (0%)
      Ezetimibe n(%)3 (3.5%)2 (2.2%)8 (9.5%)0.0680.114
      Atorvastatin Equivalency Dosages mg31.82 ± 24.5135.59 ± 27.0228.49 ± 19.880.1530.334
      Patients are presented in tertiles according to their Lp(a) mg/dl level. BMI body mass index. Data are shown as mean ± standard deviation or median and interquartile range, as applicable. p<0.05 (two-sided) was considered statistically significant.
      PAD peripheral artery disease, T2DM type 2 diabetes mellitus, CAD coronary artery disease, Atr. fibrillation atrial fibrillation, eGFR estimated glomerular filtration rate, LDL-C low-density lipoprotein cholesterol, HDL-C high-density lipoprotein cholesterol, HbA1c hemoglobin A1c, ACEi angiotensin converting enzyme inhibition treatment, AT2 angiotensin 2 receptor blockage treatment, ASS acetylic salicylic acid treatment. Bold values in the p-value column depict significant results.

      3.2 Outcome analyses

      3.2.1 LIP-LEAD-A (nmol/L) group

      During the observation time (4.3 years, IQR 3.0–5.6) a total of 326 fatal events occurred (overall death rate 33.8%, calculated annual death rate 7.8%). Furthermore, a total of 141 CV deaths were observed (overall CV-death rate 14.6%, calculated annual death rate 3.4%). In Kaplan-Meier curves no significant difference could be seen between the tertiles for CV-death (log-rank p = 0.244, Fig. 2A) or all-cause death (log-rank p = 0.654).
      Fig. 2
      Fig. 2(A) KM curves for CV-death in Lip-LEAD-A for tertiles according to Lp(a) levels.
      No significant event rate was seen between tertiles (log-rank p = 0.244), (B) KM curves for CV-death in Lip-LEAD-B. for tertiles according to Lp(a) levels. No significant event rate was seen between tertiles (log-rank p = 0.321).
      The functional form between nmol/L Lp(a) and log hazards was further explored using restricted cubic splines with 3 knots. No significant deviation from linearity was observed (Supplemental Fig. 1).
      Log transformed Lp(a) levels were subjected to univariate and multivariable-adjusted Cox-regression analyses (age, sex, T2DM, LDL-C, HDL-C, arterial hypertension, ezetimibe usage, BMI, Fontaine stage, and CKD-EPI). Both the univariate (hazard ratio 1.07, 95% confidence interval 0.95–1.21) and the multivariable-adjusted model (HR 1.10, 95% CI 0.97–1.24) did not show an association for CV-death for an increase of one-unit log transformed Lp(a). Furthermore, patients above the threshold of 175 nmol/L Lp(a) were not associated with CV death in the same continuous analyses (crude HR 1.00, 95% HR 0.99–1.00; multivariable model HR 1.00, 95% HR 0.99–1.01).
      Lp(a) tertiles were further subjected to the same Cox-regression model. Neither in crude (highest tertile HR 1.29, 95% CI 0.86–1.95) nor in multivariable-adjusted models (highest tertile HR 1.47, 95% CI 0.96–2.24) was a significant association seen for CV death (Table 3).
      Table 3Cox-Regression analyses for CV-death in Lip-LEAD-A shown in tertiles. Data are presented as Hazard ratio and 95% confidence interval. Models were adjusted first for age and sex and further multivariable adjusted for traditional CV risk factors (T2DM, LDL-C, HDL-C, arterial hypertension, ezetimibe usage, BMI, Fontaine stage and CKD-EPI).
      Cox Regression Analyses
      Univariate+ Age, sexMultivariable adjusted
      nEventsHR (95% CI)p-valueHR (95% CI)p-valueHR (95% CI)p-value
      OverallTertile 1 (reference)32045
      Tertile 2323410.82 (0.52–1.28)0.3790.97 (0.62–1.52)0.8800.99 (0.63–1.56)0.964
      Tertile 3321551.29 (0.86–1.95)0.2221.37 (0.91–2.08)0.1371.47 (0.96–2.24)0.077
      LDL-C <70Tertile 1 (reference)13220
      Tertile 2101181.17 (0.60–2.27)0.6451.23 (0.63–2.39)0.5501.03 (0.52–2.03)0.940
      Tertile 3101171.32 (0.69–2.54)0.4051.15 (0.59–2.25)0.6790.99 (0.50–1.96)0.968
      LDL-C <55Tertile 1 (reference)6612
      Tertile 249111.32 (0.57–3.05)0.5221.08 (0.46–2.52)0.8571.16 (0.47–2.89)0.750
      Tertile 34160.94 (0.35–2.50)0.8970.91 (0.34–2.44)0.8510.98 (0.33–2.85)0.962
      Intermittent claudicationTertile 1 (reference)23521
      Tertile 2242200.80 (0.42–1.52)0.4970.92 (0.49–1.75)0.8100.96 (0.50–1.86)0.907
      Tertile 3229241.24 (0.69–2.25)0.4761.30 (0.71–2.38)0.3891.37 (0.74–2.55)0.314
      Critical limb ischemiaTertile 1 (reference)8524
      Tertile 281210.92 (0.49–1.73)0.8001.04 (0.55–1.95)0.9130.95 (0.50–1.80)0.876
      Tertile 392311.35 (0.76–2.38)0.3091.44 (0.81–2.57)0.2111.55 (0.86–2.80)0.147
      Concomitant CADTertile 1 (reference)9419
      Tertile 2107261.23 (0.66–2.28)0.5111.44 (0.77–2.70)0.2491.33 (0.69–2.55)0.392
      Tertile 3105251.37 (0.74–2.55)0.3151.38 (0.74–2.58)0.3121.34 (0.71–2.54)0.365
      When stratified according to controlled LDL-C (<70 mg/dl; highest tertile HR 0.99, 95% CI 0.50–1.96) or optimal LDL-C (<55 mg/dl; highest tertile HR 0.98, 95% CI 0.33–2.85) no significant associations for CV-death were found in multivariable adjusted Lp(a) tertile analyses (Table 3).
      In a subanalysis for patients with endovascular repair due to intermittent claudication (Fontaine stage II; highest tertile HR 1.37, 95% CI 0.74–2.55) vs. critical limb ischemia (Fontaine stage III, IV; highest tertile HR 1.55, 95% CI 0.86–2.80) Lp(a) groups were not associated with CV-death as well in multivariable models (Table 3).
      Likewise, in the subcohort of patients with concomitant coronary artery disease (highest tertile 1.34 HR, 95% CI 0.71–2.54) no association was found in multivariable models (Table 3).

      3.2.2 LIP-LEAD-B (mg/dL) group

      During the observation time of the mg/dl group (median 7.6 years, IQR 3.2–8.1) a total of 123 fatal events were registered (overall death rate 47.7%, calculated annual death rate 6.3%). Furthermore, a total of 64 CV-death were registered (overall CV death rate 24.8%, calculated annual death rate 3.3%. In the Kaplan-Meier analysis no association with overall mortality (log-rank p = 0.883) or CV-death was found (log-rank p = 0.321, Fig. 2B).
      The functional form between mg/dl Lp(a) and log hazards was further explored using restricted cubic splines with 3 knots. No major deviation from linearity was observed (Supplemental Fig. 2).
      Both univariate and multivariable Cox-regression analyses did not show an association for CV-death (HR 1.05 (95%CI 0.86–1.30); HR 1.08 (95% CI 0.86–1.34) for an increase of one-unit log-transformed Lp(a) levels. Similar, in categorical analyses for tertiles of Lp(a) no associations both in univariate (highest tertile HR 1.31, 95% CI 0.70–2.43) and multivariable-adjusted analyses (highest tertile HR 1.34, 95% CI 0.70–2.58) were seen (Table 4). Furthermore, in subanalyses for intermittent claudication (Fontaine stage II, highest tertile HR 1.10, 95% CI 0.44–2.80), critical limb ischemia (Fontaine stage III, IV, highest tertile HR 3.01, 95% CI 0.99–9.10), and a subcohort of concomitant CAD (highest tertile HR 1.21, 95% CI 0.46–3.17) no association was found (Table 4).
      Table 4Cox-Regression analyses for CV-death in Lip-LEAD-B shown in tertiles. Data are presented as Hazard ratio and 95% confidence interval. Models were adjusted first for age and sex and further multivariable adjusted for traditional CV risk factors (T2DM, LDL-C, HDL-C, arterial hypertension, ezetimibe usage, BMI, Fontaine stage and CKD-EPI).
      Cox Regression Analyses
      Univariate+Age, sexMultivariable adjusted
      nEventsHR (95% CI)p-valueHR (95% CI)p-valueHR (95% CI)p-value
      OverallTertile 1 (reference)8521
      Tertile 289180.88 (0.45–1.71)0.7100.81 (0.41–1.58)0.5350.82 (0.41–1.62)0.562
      Tertile 384251.31 (0.70–2.43)0.4001.44 (0.77–2.70)0.2571.34 (0.70–2.58)0.381
      Intermittent claudicationTertile 1 (reference)6412
      Tertile 268131.24 (0.56–2.77)0.6001.17 (0.48–2.85)0.7260.85 (0.37–1.99)0.715
      Tertile 357101.01 (0.42–2.43)0.9870.90 (0.42–1.94)0.7951.10 (0.44–2.80)0.828
      Critical limb ischemiaTertile 1 (reference)219
      Tertile 22150.41 (0.13–1.34)0.1390.56 (0.16–1.88)0.3460.70 (0.20–2.50)0.585
      Tertile 327151.30 (0.56–3.01)0.5451.31 (0.56–3.09)0.5313.01 (0.99–9.10)0.051
      Concomitant CADTertile 1 (reference)2411
      Tertile 231110.86 (0.36–2.07)0.7401.01 (0.41–2.46)0.9890.84 (0.33–2.15)0.713
      Tertile 334140.93 (0.40–2.15)0.8601.62 (0.67–3.93)0.2891.21 (0.46–3.17)0.701

      3.3 Lp(a) and stenosis localization

      3.3.1 LIP-LEAD-A (nmol/L) group

      To evaluate a typical pattern of lesions with high Lp(a) levels, locations of endovascular repair were categorized into 4 categories: iliac, femoral including popliteal artery, below the knee, and multiple region repair. Lp(a) levels did not significantly differ between those four sites (p = 0.881, Fig. 3A).
      Fig. 3
      Fig. 3(A) Lp(a) levels in Lip-Lead-A and lesion site. No significant difference of means between iliacal, femoral, below the knee, or multivessel revascularization was seen (overall p = 0.881). (B) Lp(a) in Lip-LEAD-B and lesion site. No significant difference of means between iliacal, femoral, below the knee, or multivessel revascularization was seen (overall p = 0.129).

      3.3.2 LIP-LEAD-B (mg/dL) group

      The endovascular repair site of the mg/dL group was subjected to the same analysis as the nmol/l group. No significant difference was seen (p = 0.129, Fig. 3B).

      4. Discussion

      This study evaluates Lp(a) and its association with all-cause and CV-death in patients with symptomatic PAD after endovascular repair in two separated cohorts according to the assay measurement method of Lp(a). Neither the Lip-LEAD-A (nmol/L) nor the Lip-LEAD-B (mg/dL) cohort showed a significant association with Lp(a). Furthermore, sub-analyses for LDL-C control, only IC, only CLI, or concomitant CAD did not show any significant association with long-term outcome. Likewise, no significant differences in Lp(a) levels and target lesion site (iliac, femoral, BTK, or multisite repair) were found in Lip-LEAD-A or Lip-LEAD-B.
      The evidence of a causal relationship between Lp(a) and a markedly increased risk of CV disease is large. Since the first Mendelian randomization study, linking Lp(a) to CV disease [
      • Sandholzer C.
      • Saha N.
      • Kark J.D.
      • et al.
      Apo(a) isoforms predict risk for coronary heart disease. A study in six populations.
      ], several studies showed an increased risk for CAD [
      • Patel A.P.
      • Wang M.
      • Pirruccello J.P.
      • et al.
      Lp(a) (Lipoprotein[a]) concentrations and incident atherosclerotic cardiovascular disease: new insights from a large national biobank.
      ,
      • Kamstrup P.R.
      • Tybjaerg-Hansen A.
      • Steffensen R.
      • et al.
      Genetically elevated lipoprotein(a) and increased risk of myocardial infarction.
      ,
      • Virani S.S.
      • Brautbar A.
      • Davis B.C.
      • et al.
      Associations between lipoprotein(a) levels and cardiovascular outcomes in black and white subjects: the Atherosclerosis Risk in Communities (ARIC) Study.
      ], stroke [
      • Erqou S.
      • Kaptoge S.
      • Perry P.L.
      • et al.
      Lipoprotein(a) concentration and the risk of coronary heart disease, stroke, and nonvascular mortality.
      ,
      • Patel A.P.
      • Wang M.
      • Pirruccello J.P.
      • et al.
      Lp(a) (Lipoprotein[a]) concentrations and incident atherosclerotic cardiovascular disease: new insights from a large national biobank.
      ], and calcified aortic valve stenosis [
      • Kamstrup P.R.
      • Tybjærg-Hansen A.
      • Nordestgaard B.G.
      Elevated lipoprotein(a) and risk of aortic valve stenosis in the general population.
      ,
      • Vuorio A.
      • Watts G.F.
      • Kovanen P.T.
      Lipoprotein(a) as a risk factor for calcific aortic valvulopathy in heterozygous familial hypercholesterolemia.
      ]. These findings of elevated risk for higher incidences do extend to PAD [
      • Gurdasani D.
      • Sjouke B.
      • Tsimikas S.
      • et al.
      Lipoprotein(a) and risk of coronary, cerebrovascular, and peripheral artery disease: the EPIC-Norfolk prospective population study.
      ,
      • Tzoulaki I.
      • Murray G.D.
      • Lee A.J.
      • et al.
      Inflammatory, haemostatic, and rheological markers for incident peripheral arterial disease: Edinburgh Artery Study.
      ] as well. Furthermore, profound evidence regarding the outcome in primary prevention populations exists [
      • Erqou S.
      • Kaptoge S.
      • Perry P.L.
      • et al.
      Lipoprotein(a) concentration and the risk of coronary heart disease, stroke, and nonvascular mortality.
      ,
      • Kiechl S.
      • Willeit J.
      • Mayr M.
      • et al.
      Oxidized phospholipids, lipoprotein (a), lipoprotein-associated phospholipase A2 activity, and 10-year cardiovascular outcomes: prospective results from the Bruneck study.
      ]. However, results in the secondary prevention setting do not support a clear-cut association. A meta-analysis of O'Donoghue et al. evaluated Lp(a) and outcome as well as MACE in three large RCTs (PEACE, CARE, and PROVE IT-TIMI 22) [
      • O'Donoghue M.L.
      • Morrow D.A.
      • Tsimikas S.
      • et al.
      Lipoprotein(a) for risk assessment in patients with established coronary artery disease.
      ]. This initial analysis failed to show a significant outcome result. A positive outcome was reported only after a second step pooled meta-analysis for four additional RCTs and three observational studies. Yet, the heterogeneity of this meta-analysis was high. Furthermore, all RCTs of this meta-analysis included only CAD disease patients.
      In PAD, to our knowledge, only one study by Golledge et al. evaluated mortality in the secondary prevention [
      • Golledge J.
      • Rowbotham S.
      • Velu R.
      • et al.
      Association of serum lipoprotein (a) with the requirement for a peripheral artery operation and the incidence of major adverse cardiovascular events in people with peripheral artery disease.
      ] but did not report any association. However, the patients of this study had either atherosclerotic PAD or abdominal aortic aneurysms and no specific data for the respective diseases was given. While a recent study linked MALE to high Lp(a) after femoral artery endarterectomy [
      • Verwer M.C.
      • Waissi F.
      • Mekke J.M.
      • et al.
      High Lipoprotein(a) Is Associated with Major Adverse Limb Events after Femoral Artery Endarterectomy.
      ] and the aforementioned study by Golledge et al. [
      • Golledge J.
      • Rowbotham S.
      • Velu R.
      • et al.
      Association of serum lipoprotein (a) with the requirement for a peripheral artery operation and the incidence of major adverse cardiovascular events in people with peripheral artery disease.
      ] linked high Lp(a) need for revascularization in patients with PAD, our study did not find any specific pattern of an affected lesion site.
      A major difference between the profound signal of a relationship between high Lp(a) in the primary versus secondary setting can be seen in LDL-C levels and statin treatment. In the aforementioned meta-analyses of the CARE, PEACE and PROVE-IT TIMI 22 study [
      • O'Donoghue M.L.
      • Morrow D.A.
      • Tsimikas S.
      • et al.
      Lipoprotein(a) for risk assessment in patients with established coronary artery disease.
      ] LDL-C and statin treatment percentages were higher than in the usual primary prevention setting. This might have led to a diminished effect of Lp(a) in these cohorts, which can also be seen in our Lip-LEAD cohort with even higher rates of statin treatment and better LDL-C control. Likewise, in the primary prevention setting, both the Women's Health study [
      • Suk Danik J.
      • Rifai N.
      • Buring J.E.
      • et al.
      Lipoprotein(a), measured with an assay independent of apolipoprotein(a) isoform size, and risk of future cardiovascular events among initially healthy women.
      ] and the Physicians' Health Study [
      • Rifai N.
      • Ma J.
      • Sacks F.M.
      • et al.
      Apolipoprotein (a) size and lipoprotein (a) concentration and future risk of angina pectoris with evidence of severe coronary atherosclerosis in men: the Physicians' Health Study.
      ] only showed an association with Lp(a) and outcome (MACE) in participants with high LDL-C (>121–160 mg/dl). Furthermore, since Lp(a) is genetically determined and thus elevated at birth, the observation period in our cohorts is short in comparison to the lifelong elevated Lp(a).
      However, an “index event bias” as defined by Dahabreh and Kent cannot be ruled out by the null finding of this study [
      • Dahabreh I.J.
      • Kent D.M.
      Index event bias as an explanation for the paradoxes of recurrence risk research.
      ]. Index event bias describes a paradoxical null finding with a risk factor after selection for this risk factor for recurrent events in a specific patient cohort. A similar index event bias can be seen with the “smoker paradox” that describes a beneficial effect of smoking compared to non-smoking after myocardial infarction [
      • Barbash G.I.
      • Reiner J.
      • White H.D.
      • et al.
      Evaluation of paradoxic beneficial effects of smoking in patients receiving thrombolytic therapy for acute myocardial infarction: mechanism of the "smoker's paradox" from the GUSTO-I trial, with angiographic insights. Global Utilization of Streptokinase and Tissue-Plasminogen Activator for Occluded Coronary Arteries.
      ].
      Another major difference of our cohort was that only symptomatic patients either with IC or CLI were analyzed. Previous data show that more than 50% of patients with PAD are, in fact, asymptomatic (Fontaine stage I) [
      • Stoffers H.E.
      • Rinkens P.E.
      • Kester A.D.
      • et al.
      The prevalence of asymptomatic and unrecognized peripheral arterial occlusive disease.
      ,
      • Hooi J.D.
      • Stoffers H.E.
      • Kester A.D.
      • et al.
      Risk factors and cardiovascular diseases associated with asymptomatic peripheral arterial occlusive disease. The Limburg PAOD Study. Peripheral Arterial Occlusive Disease.
      ]. Despite being asymptomatic this major cohort of patients with PAD has a similarly high mortality risk to symptomatic patients [
      • Sigvant B.
      • Lundin F.
      • Wahlberg E.
      The risk of disease progression in peripheral arterial disease is higher than expected: a meta-analysis of mortality and disease progression in peripheral arterial disease.
      ,
      • Diehm C.
      • Schuster A.
      • Allenberg J.R.
      • et al.
      High prevalence of peripheral arterial disease and co-morbidity in 6880 primary care patients: cross-sectional study.
      ]. It seems likely that symptomatic patients feature additive life-limiting risk factors, such as infections and limited physical activity both by IC or amputations aside from higher rates of other CV events [
      • Cambou J.P.
      • Aboyans V.
      • Constans J.
      • et al.
      Characteristics and outcome of patients hospitalised for lower extremity peripheral artery disease in France: the COPART Registry.
      ,
      • Smith G.D.
      • Shipley M.J.
      • Rose G.
      Intermittent claudication, heart disease risk factors, and mortality.
      ]. Latter mentioned problems are associated with reduced life expectancy but are naturally not as prevalent in asymptomatic as in those without symptoms. Thus, a dedicated study on Lp(a) in asymptomatic PAD would be of high interest.
      Nevertheless, our data suggest that Lp(a) levels are not associated with fatal CV outcome in a large cohort of patients with symptomatic PAD. This finding aligns with the aforementioned studies with adequate LDL-C control in other CV diseases. However, the upper limit of confidence intervals for the categorical analyses on CV-death in both cohorts is higher than 1.5 and a trend for higher HR with higher tertiles is present in some analyses. Thus, a moderate association cannot be ruled out by this study. Since outcome RCTs on Lp(a) lowering in the secondary prevention setting are currently conducted, results of these highly anticipated studies might bring more clarity into this field of uncertainty.
      This study has several limitations. Firstly, despite a two-cohort approach, the study was only performed at a single center. Secondly, treatment in a tertiary care hospital might lead to a selection bias. Thirdly, since symptomatic PAD accounts for less than half of the PAD prevalence [
      • Fowkes F.G.R.
      • Housley E.
      • Cawood E.H.H.
      • et al.
      Edinburgh artery study: prevalence of asymptomatic and symptomatic peripheral arterial disease in the general population.
      ] worldwide, an evaluation of only symptomatic patients might present another selection bias. Fourthly, Lp(a) was measured in an “acute-phase” reaction previous to endovascular repair, at least for CLI, which could have obscured baseline Lp(a) levels. Fifthly, no apo(a) isoform size and SNP analyses were performed in this cohort considering Lp(a). Sixthly, no outcome evaluation on MACE or MALE was performed.
      However, this study has several strengths to consider. Firstly, the sample size in comparison to similar PAD studies is large. Secondly, control of traditional risk factors is high in comparison to other cohorts. Thirdly, follow-up time is extensively long and only the hard outcome of fatal CV events was included in this analysis.
      In conclusion, this study does not demonstrate an association with CV-death or a specific pattern of lesion site with Lp(a) in patients with symptomatic PAD. However, while this is in contrast to most studies on the secondary prevention of CAD and a large proportion of patients with PAD remains asymptomatic, further analyses, especially in the latter cohort and extended long-term follow-up studies are warranted.

      CRediT authorship contribution statement

      Bernhard Zierfuss: Conceptualization, Methodology, Validation, Formal analysis, Investigation, Resources, Data curation, Writing – original draft, Visualization. Clemens Höbaus: Data curation, Methodology, Validation, Resources, Data curation, Writing – review & editing, Visualization, Supervision. Anna Feldscher: Investigation, Writing – review & editing. Antonia Hannes: Investigation, Writing – review & editing. Daniel Mrak: Formal analysis, Validation, Writing – review & editing. Renate Koppensteiner: Resources, Writing – review & editing. Herbert Stangl: Validation, Writing – review & editing. Gerit-Holger Schernthaner: Supervision, Resources, Project administration, Writing – review & editing, Supervision.

      Declaration of competing interests

      The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

      Appendix A. Supplementary data

      The following are the Supplementary data to this article:

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