Lower extremity peripheral arterial disease (PAD) is one of the most common manifestations of atherosclerosis1. Atherosclerosis is a chronic inflammatory disease of the vessel wall2 and inflammation is deeply involved in the initiation and progression of atherosclerosis3. This study aims to determine the differences in the inflammatory parameters of patients with claudication and with chronical limb threatening ischaemia (CLTI). The second goal is to analyze the evolution of inflammatory parameters after revascularization in patients with CLTI.

An observational, prospective, longitudinal study, including patients with PAD, was conducted from january 2018 to december 2021. Inclusion criteria: patients with PAD suggested by the clinical history and objective examination and confirmed with ankle-brachial index. Exclusion criteria: medical conditions responsible for body composition changes or pro-inflammatory state in the last past 3 months, including active foot infection. An informed written consent was obtained. The study was approved by the ethical commission of the Local Hospital (75/2017). The patients were observed at admission or immediately prior revascularization and after three months. The following data was registered at this two time-points: clinical characteristics; cardiovascular risk factors, usual medication, routine analytic evaluation and inflammatory parameters. The inflammatory parameters registered were: positive acute phase proteins (C-reactive Protein- CRP- and fibrinogen), negative acute phase proteins (albumin, total cholesterol and high-density lipoprotein- HDL). A panel of cytokines were measured at admission (Bio-Plex Pro Human Cytokine 27-plex Assay #M500KCAF0Y- Bio-Rad Laboratories). Continuous variables were expressed as the mean ± standard deviation and as the percentage for categorical variables. The Shapiro-Wilk test was used to assess all continuous variables for normality. Continuous variables between two groups were compared with Student's t-test or with Mann-Whitney. Categorical variables between two groups were compared with Chi-square. A p-value of less than .05 was considered significant. Statistical evaluation was performed using SPSS software, version 20.0 (SPSS, Inc., Chicago, IL, USA).

116 patients (mean age: 67.65±9.53 years-old) 64% with claudication and 46% with CLTI were enrolled in the study. No differences were registered between patients with claudication and CLTI on age, cardiovascular risk factors (hypertension, dyslipidemia, diabetes mellitus, smoking habits) and medication. There was a higher number of men in the claudication group (88.89% versus 71.70%, p=0.019). Patients with CLTI when compared to claudicants have a decrease in haemoglobin/ hematocrit and lymphocytes and an increase in platelets. Analyzing the inflammatory parameters, we noted that patients with CLTI, compared with claudicants had increased serum levels of positive acute phase proteins: CRP (35.73±46.61mg/L versus 9.18±26.12mg/L, p=0.000), and fibrinogen (466.18±208.07mg/dL versus 317.37±79.42mg/dL, p=0.000). CLTI patients had decreased negative acute phase proteins: albumin (3.53±0.85g/dL versus 3.91±0.72g/dL, p=0.001), total cholesterol (145.41±38.59mg/dL versus 161.84±34.94mg/dL, p=0.013) and HDL (38.70±12.19mg/dL versus 51.31±15.85mg/dL, p=0.000). CLTI patients also had significantly higher cytokines serum levels (TNF-α; IL-1ra; IL-17a; FGF basic; IP-10)(Fig. 1). When analyzing the data at the third month of follow-up, it became evident that after resolution of CLTI there is a reversion in the inflammatory parameters (Fig.2). In CLTI patients there is an increase in negative acute phase proteins: albumin (at admission: 3.53±0.85g/dL versus three months: 3.70±0.73g/dL, p=0.043), HDL (at admission: 38.70±12.19mg/dL versus three months: 50.00±13.77mg/dL, p=0.034) and total cholesterol (at admission 145.41±38.59mg/dL versus three months: 153.89±37.71mg/dL, p=0.524) (Fig.2). There is a decrease in positive acute phase proteins in CLTI patients at three months CRP (at admission: 35.73±46.61mg/L versus three months: 6.27±0.95g/dL, p=0.009) and fibrinogen (at admission: 466.18±208.07mg/dL versus three months: 346.00±153.65mg/dL, p=0.022) (Fig.2). Analyzing these data, in patients with claudication, no significant changes were noted at three months of follow up(Fig.2). Despite this "inflammatory improvement" in patients with CLTI, their "inflammatory burden" remains higher than in claudicants (Fig. 2).

Patients with CLTI have an inflammatory state, with potential deleterious consequences, that can be at least be partially reversed after revascularization. These results are explained by the fact that atherosclerosis lesions are associated with an increased synthesis of proinflammatory cytokines4.These cytokines cause a decrease in synthesis of negative acute phase protein, an increased in acute phase protein, a generation of proatherogenic lipoprotein profile, a reduction in muscle mass and cardiovascular injury4. Recognizing that patients with CLTI have an inflammatory state with lethal consequences, that can be partially reversed is an opportunity to implement a timely revascularization and to optimize medical treatment.

1- S. S. Signorelli, F. Valerio, and G. Malaponte, "Inflammation and peripheral arterial disease:The value of circulating biomarkers (review)," International Journal of Molecular Medicine, vol. 33, no. 4, pp. 777-783, 2014, doi: 10.3892/ijmm.2014.1657. 2- D. Wolf and K. Ley, "Immunity and Inflammation in Atherosclerosis," Circulation Research, vol. 124, no. 2, pp. 315-327, Jan. 2019, doi: 10.1161/CIRCRESAHA.118.313591. 3- P. Libby, P. M. Ridker, and A. Maseri, "Inflammation and atherosclerosis," Circulation, vol.105, no. 9, pp. 1135-1143, Mar. 2002, doi: 10.1161/hc0902.104353. 4- G. A. Kaysen, "Biochemistry and biomarkers of inflamed patients: Why look, what to assess," in Clinical Journal of the American Society of Nephrology, 2009, vol. 4, no. SUPPL. 1. doi: 10.2215/CJN.03090509.