Skip to main content
Download PDF
Download ePub

Plasma fibrin clot properties and cardiovascular mortality in patients with type 2 diabetes: a long‐term follow‐up study

Cardiovascular Diabetology volume 20, Article number: 47 (2021) Cite this article

Abstract

Background

Patients with type 2 diabetes mellitus (T2DM) are at high risk of cardiovascular mortality, but the mechanisms behind this remain unclear. Prothrombotic fibrin clot properties have been shown in T2DM and cardiovascular disease. We hypothesized that formation of denser clots, which are resistant to fibrinolysis, has a negative impact on cardiovascular mortality in T2DM.

Methods

We studied 133 T2DM patients aged 43–83 years. Plasma fibrin clot turbidity, permeation, compaction, and efficiency of clot lysis using 3 assays including the determination of maximum concentration (D-Dmax) and rate of increase in D-dimer concentration (D-Drate) released during tissue plasminogen activator-induced degradation, were evaluated at the time of enrollment, along with thrombin generation and fibrinolytic proteins. During a median follow-up period of 72 months, cardiovascular mortality was recorded.

Results

Cardiovascular deaths (n = 16, 12%) occurred more frequently in patients with increased D-Dmax (> 4.26 mg/l, hazard ratio [HR] 5.43, 95% confidence interval [CI] 1.99–14.79), or decreased D-Drate (< 0.07 mg/l/min, HR 2.97, 95% CI 1.07–8.23), or increased peak thrombin (> 283.5 nM, HR 5.65, 95% CI 2.07–15.51). These predictors had an even more potent impact on cardiovascular mortality in patients with prior cardiovascular disease (64.7%) and with corresponding risks as follows: HR 6.18, 95% CI 2.02–18.96; HR 8.98, 95% CI 2.99–26.96; and HR 5.35, 95% CI 1.62–17.72, respectively. Other investigated fibrin variables and fibrinolytic proteins did not associate with cardiovascular mortality. In multivariable analysis, cardiovascular mortality was predicted by D-Dmax > 4.26 mg/l, age > 65 years, prior cardiovascular disease, and C-reactive protein > 3 mg/l.

Conclusions

This study is the first to show that formation of denser fibrin clots resistant to fibrinolysis could be a risk factor for long-term cardiovascular mortality in T2DM.

Background

Type 2 diabetes mellitus (T2DM) is considered to be an epidemic of the twenty-first century, having a global prevalence of 9.3% in 2019 [1]. Over the last 15 years, the mortality and incidence of cardiovascular events has declined both in patients with and without T2DM, though at a slower rate among diabetic patients [2]. Cardiovascular disease affects 32.2% of T2DM patients and leads to mortality in 9.9% of patients having T2DM of more than 10 years duration [3]. Coronary artery disease (CAD) and stroke are major contributors to overall mortality in T2DM patients [3, 4].

There is growing evidence which shows that prothrombotic fibrin clot properties, including formation of more compact clots which are more resistant to lysis, are involved in atherothrombotic events such as myocardial infarction and ischemic stroke [5,6,7]. Several groups have also shown an association of T2DM with a prothrombotic clot phenotype [8,9,10,11,12]. Of note, T2DM has been reported to unfavorably affect plasma fibrin clot characteristics, such as leading to hypolysability, in patients with concomitant advanced CAD [13, 14]. The mechanisms behind impaired clot susceptibility to lysis observed in T2DM are complex. It has been suggested that hypofibrinolysis in T2DM is associated with elevated levels of plasminogen activator inhibitor-1 (PAI-1) [15] and thrombin-activatable fibrinolysis inhibitor (TAFI) [16], along with increased cross-linking of α2-antiplasmin into fibrin networks [8, 17], and glycation of fibrinogen [18] and plasminogen [19], while the mechanical properties of fibrin are independent from fibrinogen glycation [20]. Prolonged T2DM duration might also contribute to prothrombotic fibrin clot alterations including impaired fibrinolysis [12], with enhanced oxidative stress playing a substantial role [21]. Patient sex is also an important modulator of fibrin clot properties, both in T2DM and CAD patients [14, 22]. Asymptomatic women with coronary plaques, as visualized by coronary computed tomography angiography, have reduced fibrin clot lysability compared to men with and without coronary plaques, and women without coronary plaques, suggesting a sex-dependent link between coronary atherosclerosis and fibrin clot lysability [22].

Fibrinogen, the main component of fibrin, has been reported to be predictive of major adverse cardiovascular events, especially among diabetic patients, both following acute coronary syndrome [23] and in those with stable coronary disease [24]. Moreover, fibrin clots that are resistant to lysis have predicted adverse outcome in diabetic patients following acute coronary syndrome [25]. The same was true for maximum turbidity. In another study [26], area under the turbidity curve, but not the turbidity lysis time, associated with cardiovascular events in CAD patients, also after adjustment for T2DM, which was diagnosed in 29% of the study participants. To the best of our knowledge, there have not been any published reports investigating the long-term impact of fibrin clot phenotype on mortality in T2DM patients. We hypothesized that formation of denser clots, which are more resistant to fibrinolysis, leads to increased long-term cardiovascular mortality in T2DM.

Methods

Patients

Patients were recruited from March 2011 to November 2011 from the John Paul II Hospital, Krakow, Poland. The study population was previously described in detail [12]. Briefly, 156 patients with T2DM aged 18 years or greater who fulfilled the American Diabetes criteria for diagnosis of T2DM were enrolled. Patients were excluded if they presented with signs of acute infection, suffered from arterial or venous thromboembolic events within the six months prior to enrollment, were treated with anticoagulants at the time of enrollment, or were diagnosed with cancer, chronic inflammatory disease, liver injury, or stage 4 or 5 chronic kidney disease. Pregnant women were also excluded.

Data collected in the questionnaire included demographic data, time since T2DM diagnosis, comorbidities, and medications used. Obesity was defined as body-mass index > 30 kg/m2. Hypercholesterolemia was defined as low-density lipoprotein (LDL) cholesterol > 2.5 mmol/l or ongoing lipid-lowering treatment [27]. Other comorbidities were defined as previously described [12]. The study was approved by the local bioethics committee. All patients provided informed written consent.

Laboratory investigations

At baseline, fasting blood samples were obtained from the antecubital vein between 8:00 and 10:00 am. Glycated hemoglobin (HbA1c), fasting glucose, creatinine, lipid profile, and high sensitivity C-reactive protein (CRP) were assayed by routine laboratory techniques. Fibrinogen was determined with the von Clauss method. Plasminogen and α2-antiplasmin were measured using chromogenic assays (STA Stachrom α2-antiplasmin and plasminogen, Diagnostica Stago, Asnieres, France). Commercially available immunoenzymatic assays were used to measure tissue plasminogen activator (tPA) antigen (TintElize, Umea, Sweden), PAI-1 antigen (American Diagnostica, Stamford, CT, USA), TAFI antigen (Chromogenix, Lexington, MA, USA), and soluble thrombomodulin (Diagnostica Stago).

Plasma thrombogenic potential was assessed using calibrated automated thrombography (Thrombinoscope BV, Maastricht, the Netherlands) in a 96-well plate fluorometer (Ascent Reader, Thermolab systems OY, Helsinki, Finland) at 37 °C according to the manufacturer’s instructions. A volume of 80 µl of platelet-poor plasma was diluted with 20 µl of a commercially available tissue factor-based activator (Diagnostica Stago) containing 5 pmol/l recombinant tissue factor, 4 µmol/l phosphatidylserine/phosphatidylcholine/phosphatidylethanolamine vesicles, and 20 µl of FluCa solution (Hepes, pH 7.35, 100 nmol/l CaCl2, 60 mg/ml bovine albumin, and 2.5 mmol/l Z-Gly-Gly-Arg-amidometylcoumarin). The peak thrombin level was analysed [28]. Plasma samples were analyzed in duplicate, and the intra-assay variability was 6%.

Fibrin clot parameters, assayed at the time of enrollment, included: turbidity, permeation, compaction, lysability, maximum concentration (D-Dmax), and rate of D-dimer (D-Drate) released during tPA-induced clot lysis, and were previously described in detail [12]. Two parameters were assayed during turbidity measurements: lag phase, which reflects the time required for fibrin protofibrils to grow to a sufficient length to allow for lateral aggregation, and maximum absorbance at plateau (ΔAbs), which reflects fiber cross-sectional area. Plasma was diluted 2:3 with 0.05 mol/l Tris buffer and was incubated with 1 U/ml human thrombin (Sigma) and 16 mmol/l calcium chloride. Absorbency was read every 7 s at 350 nm for 15 min with a Dynex MRX2 plate reader. Lag phase was recorded as the time needed for absorbency to change by 0.01 from baseline [29]. Fibrin clot permeation was determined by measuring flow rates of buffer through the fibrin clots obtained from the 120 µl of citrated plasma, by adding 20 mmol/l calcium chloride and 1 U/ml human thrombin, as previously described [30, 31]. The mass of consecutive drops through each tube during a period of 60 min was recorded for exact volume. A permeation coefficient (Ks), indicating the average size of fibrin clot pores, was calculated from the following equation: Ks = \(\frac{{\text{Q}}\times {\text{L}}\times\upeta }{{\text{t}}\times {\text{A}}\times \Delta {\text{p}}}\), where Q is the flow rate in time (t); L is the length of a fibrin gel; η is the viscosity of liquid (in poise); A is the cross-sectional area (in square centimeters), and Δp is the differential pressure (in dynes per square centimeter). To assess compaction, the citrated plasma was mixed (3:2) with 0.7 IU/ml thrombin, 0.1% Tween 80, and 20 mmol/l calcium chloride in TBS, and then clots were formed in tubes prepared as in the permeation experiments [31, 32]. After centrifugation at 6000 g for 60 s, the volume of supernatant evacuated from the tubes was assessed based on the difference in mass of the tube. Compaction was expressed as this volume divided by the initial plasma volume used to form the fibrin clot. Two plasma clot lysis assays were performed. In the modified assay by Lisman et al. [33], citrated plasma was mixed (1:1) with HEPES buffer containing calcium chloride (final concentration, 17 mmol/l), diluted recombinant tissue factor (final dilution 105 times; Innovin, Dade Behring), phospholipid vesicles (final concentration, 10 µmol/l), and recombinant tPA (rtPA, final concentration, 30 U/mL, 56 ng/mL; Boehringer Ingelheim, Ingelheim, Germany). Clot lysis time (CLT) was defined as time from the midpoint of the baseline to maximum turbid transition, to the final plateau phase. In the second assay, using the modified approach by Williams et al. [34], 1 µg/ml rtPA (Boehringer Ingelheim) was added simultaneously with human thrombin. This lysis time was defined as the time required for a 50% decrease in clot turbidity measured at 405 nm (t50%) at 37 °C in a Spectra Max 340 kinetic microplate reader (Molecular Devices, Sunnyvale, CA, USA). Finally, plasma lysis was assessed using a modified version of the method described by Collet et al. [35, 36]. The plasma samples were recalcified with calcium chloride (final concentration, 20 mmol/l) and then 1 U/ml human thrombin (Sigma-Aldrich, St Louis, MO, USA) was added. After 2 h of incubation at room temperature, tubes containing the clots were connected via plastic tubing to a reservoir with buffer (0.05 M Tris–HCl, 0.15 M NaCl, pH 7.5), containing 0.2 µM rtPA. The lysis rate was determined by measuring the concentration of released D-dimers (American Diagnostica), a marker of plasmin-mediated fibrin degradation, in the eluate every 20 min. Two measures of fibrin clot architecture, namely the high maximum concentration of D-dimer concentration (D-Dmax) and the rate of increase in D-dimer concentrations released from the clot (D-Drate) were determined in each subject. The experiment was usually stopped after 80–120 min when the fibrin gel collapsed under the pressure.

Follow-up

The primary endpoint was cardiovascular death. The length of follow-up was censored at the time of death. Follow-up by telephone was conducted between July 2017 and January 2018. The family of the deceased patient was asked about the cause of death. In the case of incomplete medical record data or loss to clinical follow-up, patient status was assessed in March 2019 using the National Mortality Registry maintained by the State Systems Department of Ministry of Digital Affairs. The International Statistical Classification of Diseases 10 (ICD-10) codes I70.9, I25.0, I25.9, and I11.0 were classified to be cardiovascular death. In the case of ICD-10 code R99, the correct classification of death was based on additional clinical data. The secondary endpoint was all-cause death.

Statistical analysis

Continuous data were presented as medians (interquartile range, IQR) and were compared using the Mann–Whitney U test. Categorial data were presented as numbers (percentages). To test the differences in proportions between two groups, Fisher’s exact test was used. Continuous variables (peak thrombin, ΔAbsmax, Ks, compaction, CLT, t50%, D-Dmax, and D-Drate) were dichotomized using the cut-off values found in the analysis of receiver operating curves (ROC), which optimally classified the clinical endpoint, as described previously [37]. Multivariable Cox proportional hazards survival regression, after adjustment for age and sex, was used to investigate the effect of high peak thrombin, high ΔAbsmax, low Ks, high compaction, prolonged CLT, prolonged t50%, high D-Dmax, and low D-Drate, on clinical outcomes. The results were presented as hazard ratios (HR) with 95% confidence interval (CI). The HR were additionally adjusted for possible confounding factors, identified in the comparison analysis of two groups. Kaplan–Meier survival curves were drawn. Six factors (age > 65 years, previous cardiovascular event, CRP level > 3 mg/l, hypercholesterolemia defined as LDL > 1.8 mmol/l or treatment with statins, D-Dmax and peak thrombin) were analyzed for the association with the cardiovascular mortality in the univariate logistic regression model. The Nagelkerke R2 represented the contribution of D-Dmax and peak thrombin to the variation in cardiovascular mortality. Factors, which significantly associated with cardiovascular mortality in the univariate regression model, were included in the multivariable logistic regression model. Results of the regression model were presented as odds ratio (OR) with 95% CI. To address the problem of multiple comparisons, the results were controlled with the Benjamini–Hochberg procedure. If continuous variables correlated with Spearman’s correlation coefficient > 0.5, only one of them was included in the multivariable regression model. A two-sided p-value of < 0.05 was considered statistically significant. No formal sample size calculation was performed, as the number of patients included in the original study was fixed [12]. We performed a post hoc power calculation based on a 12% event rate, HR of 5.4, with the distribution of patients in the subgroups 20%/80%. Given these estimates, and with a significance level set at 5%, a post hoc power of 78% was found for D-Dmax. Statistical analyses were performed using IBM SPSS Statistics for Windows, Version 25.0. (Armonk, NY: IBM Corp.). The Benjamini–Hochberg procedure was applied in R [38]. Figures were prepared using GraphPad Prism (version 8.4.2 for Windows, GraphPad Software, San Diego, CA, USA).

Results

Patient characteristics at baseline

A total of 156 patients with T2DM (87 men and 69 women) were enrolled. At the time of enrollment, median period since diagnosis of T2DM was 5.0 (IQR, 2.0–10.0) years, while the HbA1C levels ranged from 5.2 to 12.6 (median, 6.5) %. The patients were mostly (60.3%) treated with biguanides, while 26.3% of patients received insulin. A history of cardiovascular events was recorded in 67.3% of patients, including 64.1% with CAD.

Patient characteristics during follow-up with respect to cardiovascular death in the entire study group

The median length of follow-up was 72 (68–74) months. Twenty-three (14.7%) patients were lost to follow-up. Final analysis involved 133 patients, 79 men and 54 women, aged 43.0–83.0 (median 66.0) years. During follow-up, 16 patients (12.0%) died from cardiovascular causes (2.18 per 100 patient-years). Patients who died from cardiovascular causes did not differ from the survivors in terms of demographic data and time since T2DM diagnosis, but they more commonly had nephropathy at baseline and less commonly were treated with metformin when compared to the survivors (Table 1). Despite similar HbA1c, patients who died from cardiovascular causes tended to have lower fasting glucose levels when compared with the remaining patients (Table 2). Patients who died from cardiovascular causes had lower total and LDL-cholesterol and higher CRP when compared with the remaining patients (Table 2). Patients with LDL-cholesterol above the a median, i.e. ≥ 2.3 mmol/l, did not differ from the remaining subjects in regard to the parameters of clot structure and function. The baseline fibrinogen concentration, tPA and PAI-1 antigen, α2-antiplasmin and TAFI activity, as well as thrombomodulin antigen did not differ between patients who died from cardiovascular causes and the survivors (Table 2). Analysis of several variables describing plasma fibrin clot structure and function showed that the clot turbidity, permeability, compaction, and lysability, represented by CLT, were comparable in patients who died from cardiovascular causes and the survivors (Table 2). Patients who died from cardiovascular causes had higher D-Dmax and lower D-Drate when compared with the survivors (Table 2). The absolute difference in D-Drate between those groups corresponds with the standardized effect size of 0.68, and it is statistically significant in the Mann–Whitney’s test.

Table 1 Baseline characteristics of type 2 diabetes mellitus (T2DM) patients divided according to the cardiovascular death
Full size table
Table 2 Baseline laboratory investigations of type 2 diabetes mellitus patients divided according to the cardiovascular death
Full size table

Correlations between the investigated parameters in the entire study group

The D-Dmax correlated with ∆Abs (r = 0.432, p < 0.001), Ks (r = − 0.496, p < 0.001), compaction (r = − 0.446, p < 0.001), CLT (r = − 0.463, p < 0.001), and t50% (r = 0.518, p < 0.001), but not with peak thrombin (r = 0.14, p = 0.10). The correlations between D-Dmax and CRP, and between D-Drate and CRP, were weak (r = 0.208, p = 0.020, and r = − 0.208, p = 0.020, respectively).

Predictors of cardiovascular mortality in the entire study group

Based on the ROC curves, D-Dmax > 4.26 mg/l was regarded as a decreased value and D-Drate < 0.07 mg/l/min was considered as decreased. At baseline, increased D-Dmax was noted in 25 patients (18.8%), while decreased D-Drate was observed in 54 (40.6%) patients. Cardiovascular deaths occurred more frequently in patients with increased maximum concentration of D-dimer released from the clot during lysis induced by tPA (D-Dmax) and decreased rate of D-dimer release (D-Drate, Table 3, Fig. 1a, b). Another predictor of cardiovascular death was increased peak thrombin (Table 3, Fig. 1c). Based on the ROC curves, increased peak thrombin > 283.5 nM was noted in 23 (17.3%) patients at baseline. Adjustment for cardiovascular disease history prior to the study enrollment, nephropathy, or treatment with metformin did not influence the hazard ratios (Table 3). In univariate regression model analysis, the high D-Dmax was responsible for 13% variation in cardiovascular mortality (OR 5.88, 95% CI 1.94–17.79), while high peak thrombin accounting for 10% of that variation (OR 4.91, 95% CI 1.60–15.04). Since D-Drate correlated with D-Dmax (r = − 0.536, p < 0.001), only D-Dmax was included in the regression model. The multivariable model including increased D-Dmax, along with traditional risk factors for cardiovascular mortality such as age > 65 years, cardiovascular events prior to study enrollment (myocardial infarction or stroke), and inflammatory state represented by CRP > 3 mg/l, accounted for 37% variation in cardiovascular death (Fig. 2a). When increased D-Dmax was replaced in the model by increased peak thrombin, the explained variation in cardiovascular mortality was 42% (Fig. 2b). The Benjamini–Hochberg procedure controls did not affect results obtained in the multivatiate regression model. Despite the fact that D-Dmax correlated with many of the investigated parameters as presented above, only D-Dmax was found to be a predictor of cardiovascular death.

Table 3 Predictors of cardiovascular and all-cause mortality in type 2 diabetes mellitus patients
Full size table
Fig. 1
figure 1

Kaplan–Meier curves showing probability of survival in diabetic patients (a–c) and in diabetic patients with cardiovascular disease (d–f) with increased maximum D-dimer concentration (D-Dmax), or decreased rate of D-dimer (D-Drate) released during clot lysis, or increased peak thrombin generated, as compared with the remainder. Cut-offs were estimated based on the receiver operating curves. The D-Dmax > 4.26 mg/l was regarded as increased in all diabetic patients, while D-Dmax > 4.25 mg/l was regarded as increased in diabetic patients with cardiovascular disease; in both groups D-Drate < 0.07 mg/l/min was considered to be decreased, and peak thrombin > 283.5 nM was regarded as increased. P-values were calculated in the Mantel-Cox test

Full size image
Fig. 2
figure 2

Associations of increased maximum concentration of D-dimer released during clot lysis (D-Dmax), peak thrombin and cardiovascular risk factors with cardiovascular mortality. Cardiovascular risk factors included: age > 65 years, prior cardiovascular disease, C-reactive protein > 3 mg/l. The D-Dmax and peak thrombin were dichotomized using the cut-off value found in the receiver operating curves which optimally classified the cardiovascular death. A D-Dmax > 4.26 mg/l and peak thrombin > 283.5 nM were regarded as increased. Squares represent odds ratio (OR) estimates, and horizontal lines represent 95% confidence intervals (CI), found in the multivariable regression analysis. CV denotes cardiovascular; CRP, C-reactive protein

Full size image

Predictors of cardiovascular mortality in the subgroup of patients with prior cardiovascular disease

When T2DM patients with a history of cardiovascular disease prior to enrollment were analyzed separately (n = 86, 64.7%), 13 (15.1%) deaths were recorded. The predictors of cardiovascular mortality were D-Dmax (HR 6.18, 95% CI 2.02–18.96, Fig. 1d) and D-Drate (HR 8.98, 95% CI 2.99–26.96, Fig. 1e), as well as peak thrombin (HR 5.35, 95% CI 1.62–17.72, Fig. 1f).

Predictors of all-cause mortality in the entire study group

During follow-up, all-cause death was recorded in 22 patients (16.5%, 2.99 per 100 patient-years), indicating 6 non-cardiovascular fatal events. Non-survivors were older (72.0 [66.0–77.0] vs. 65.0 [59.0–71.0] years, p = 0.004) when compared with survivors. Other demographic data, time since T2DM diagnosis, comorbidities, medications, and laboratory investigations, including fibrin properties, did not differ between the two groups. Similar to cardiovascular mortality, Kaplan–Meier curves showed that all-cause mortality was predicted by high D-Dmax, low D-Drate, and high peak thrombin with similar cut-offs and HR with 95% CI as for cardiovascular mortality, before and after adjustment for sex, age, cardiovascular disease history, nephropathy, and metformin treatment (Table 3).

Comparison of patients who died from non-cardiovascular vs. cardiovascular causes

When compared with patients who died from cardiovascular causes (n = 16, 12%), patients who died from non-cardiovascular causes (n = 6, 4.5%) were less frequently treated with β-blockers (2 [33.3] vs. 14 [87.5], p = 0.03) and had a higher concentration of LDL-cholesterol by 0.9 mmol/l (p = 0.03), although the frequency of treatment with statin was comparable between groups. Moreover, patients who died from non-cardiovascular causes had 11.4% increased clot compaction (p = 0.008), 7.4% shorter t50% (p = 0.049), and tended to have lower D-Dmax (p = 0.08) when compared with patients who died from cardiovascular causes.

Discussion

To the best of our knowledge, this study is the first to evaluate the association between plasma fibrin clot properties and cardiovascular mortality during long-term follow-up of T2DM patients. Among several fibrin variables tested at enrollment, only D-Dmax and D-Drate were identified to be independent predictors of cardiovascular mortality in T2DM patients. These fibrin-related parameters (not associated with plasma fibrinogen concentration), in addition to advanced age, prior cardiovascular disease, and elevated CRP levels, helped to identify the risk of cardiovascular death. These associations were even more pronounced when patients with concomitant T2DM and prior cardiovascular disease were analyzed.

Assay utilizing D-dimer concentrations: clinical relevance, possible mechanisms, and perspectives for future studies

Two measures of fibrin clot architecture determined in the assay introduced by Collet et al. in 1999 [35] indicate slightly different features of the clot. The D-Dmax indicates the fibrin mass subjected to lysis by rtPA, while the D-Drate reflects the ability of rtPA to penetrate into the fibrin clots. To understand the clinical relevance of these two parameters, it should be clear that opposite results associate with unfavorable fibrin clot characteristics: increased D-Dmax and decreased D-Drate. In contrast to turbidity, where fibrin clot formation and degradation take place simultaneously [39], D-Drate is the measure of tPA-induced degradation of the previously prepared plasma clots, generated in the presence of thrombin and calcium. The assay in which D-Drate or D-Dmax are evaluated may reflect the situation in which accumulating intravascular fibrin is exposed to exogenous tPA. For example, damage to the atherosclerotic plaque, which is rich in fibrin [40], results in the formation of fibrin on the surface of the exposed site [41]. A modified version of the method described by Collet et al. [35, 36] might be considered as a method to measure fibrin removal from the atherosclerotic plaques. We observed the association of D-Drate and/or D-Dmax with cardiovascular events also in our previous studies [36, 42]. Cardiovascular mortality was not predicted by other fibrinolytic tests (CLT and t50%) in which much lower concentrations of rtPA were applied, similar to those encountered during thrombolysis [43]. Results from our study support the use of a comprehensive set of plasma fibrin clot assays over the single test to better characterize fibrin clots in a given clinical setting.

The mechanisms behind impaired fibrinolysis in T2DM patients, including non-survivors during long-term follow-up, remain unclear despite extensive experimental efforts over the last decade [44]. Adjustment for potential clinical confounders, such as metformin treatment [45, 46], did not influence the results. Higher cardiovascular mortality was not attributed to the differences in fibrinogen concentration, the most important modulator of fibrin clot properties [8, 47], or PAI-1, or TAFI. It is hypothesized that post-translational modifications of fibrinogen (i.e. glycation and oxidation) are involved [48, 49].

Our study opens the discussion about the fibrin-targeted pharmacotherapy, which could improve the clinical outcomes in T2DM patients. Antithrombotic agents, such as aspirin monotherapy or in combination with low-dose rivaroxaban, have a favorable impact on cardiovascular mortality in T2DM [50]. The same agents can improve fibrin clot properties, including the acceleration of fibrinolysis [51, 52]. Prevention of mortality among T2DM patients receiving antithrombotic medications might be partly attributed to improved fibrin clot phenotype. Larger studies, including patients receiving antithrombotic agents, are needed to validate this intriguing hypothesis. Furthermore, it remains unclear whether the cardiovascular protective role of sodium-glucose cotransporter inhibitors or glucagon-like peptide-1 receptor agonists in T2DM is related to an improvement in fibrin clot properties.

Other predictors of cardiovascular death: thrombin generation and CRP

Thrombin generation, as a potential predictor of cardiovascular mortality in T2DM, deserves a comment, given the contradictory results between the Ludwigshafen Risk and Cardiovascular Health (LURIC) [53] and Gutenberg Health Studies [54]. Our study provides data in favor of the positive association of thrombin generation and cardiovascular mortality in T2DM. Elevated CRP was another predictor of cardiovascular death in our study. Previously, elevated CRP level has been reported to be a predictor of cardiovascular and all-cause mortality in T2DM patients [55], although it has not been found to independently predict fibrin clot lysis in T2DM [9]. We speculate that elevated CRP increases the negative impact of dense fibrin clot formation, which are resistant to lysis in T2DM, although the mechanisms of this link remain elusive.

Study limitations

Our study has several limitations. Firstly, the number of study participants was limited; however, the study was sufficiently powered to detect associations between plasma fibrin clot and cardiovascular events. The small sample size may have impeded adequate identification of possible confounding factors on the results. Non-survivors may have been a group with more advanced co-morbidities which could be reflected by a higher percentage of patients with nephropathy and prior stroke, when compared with survivors. Nevertheless, adjustment for previous cardiovascular events and nephropathy did not significantly influence the hazard ratios of investigated parameters for cardiovascular death. Larger studies in this area could allow for better distribution of comorbidities between the studied groups.

Secondly, antidiabetic medications being used and glycemic control, as reflected by HbA1c, were not assessed during follow-up. Because the median time since T2DM diagnosis at baseline was 5 years, we suspect that the impact of disease duration outweighed the effects of possible periods of inadequate glycemic control during that time [12].

Thirdly, use of the modified assay by Collet et al. in clinical practice is limited due to lack of standardization and the fact that it is a time-consuming protocol, requiring hands-on laboratory expertise to obtain reproducible results [35]. Our findings clearly show that further efforts to optimize the determination method of fibrin clot density and lysability in real-life patients are worthwhile as they might be clinically relevant in formulating a prognosis related to cardiovascular mortality. Another limitation of the study was that our analysis included only one of the thrombin generation parameters, i.e. peak thrombin. Although endogenous thrombin potential could add some extra information, both parameters are well correlated with each other [56].

Finally, we did not analyze specific cardiovascular events during follow-up since we focused on death as the single hard endpoint.

Conclusions

In summary, our current study shows that the formation of denser clots, which are resistant to fibrinolysis, may predict cardiovascular mortality in T2DM.

Availability of data and materials

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Abbreviations

ΔAbs:

Maximum absorbance at plateau

CAD:

Coronary artery disease

CI:

Confidence interval

CLT:

Clot lysis time

CRP:

C-reactive protein

D-Dmax :

Maximum concentration of D-dimer released during tissue plasminogen activator induced clot lysis

D-Drate :

Rate of increase in D-dimer concentration released during tissue plasminogen activator-induced clot lysis

HbA1c:

Glycated hemoglobin

HR:

Hazard ratio

Ks :

Darcy’s constant or clot permeability

LDL:

Low-density lipoprotein

OR:

Odds ratio

PAI-1:

Plasminogen activator inhibitor 1

ROC:

Receiver operating curve

rtPA:

Recombinant tissue plasminogen activator

t50% :

Time required for a 50% decrease in clot turbidity

TAFI:

Thrombin-activatable fibrinolysis inhibitor

tPA:

Tissue plasminogen activator

References

  1. Saeedi P, Petersohn I, Salpea P, Malanda B, Karuranga S, Unwin N, et al. Global and regional diabetes prevalence estimates for 2019 and projections for 2030 and 2045: Results from the International Diabetes Federation Diabetes Atlas, 9th edition. Diabetes Res Clin Pract. 2019;157:107843. https://doi.org/10.1016/j.diabres.2019.107843.

    Article  PubMed  Google Scholar 

  2. Rawshani A, Rawshani A, Franzén S, Eliasson B, Svensson AM, Miftaraj M, et al. Mortality and cardiovascular disease in type 1 and type 2 diabetes. N Engl J Med. 2017;376:1407–18. https://doi.org/10.1056/NEJMoa1608664.

    Article  PubMed  Google Scholar 

  3. Einarson TR, Acs A, Ludwig C, Panton UH. Prevalence of cardiovascular disease in type 2 diabetes: a systematic literature review of scientific evidence from across the world in 2007–2017. Cardiovasc Diabetol. 2018;17:83. https://doi.org/10.1186/s12933-018-0728-6.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Niedziela J, Hiczkiewicz J, Kleinrok A, Pączek P, Leszek P, Lelonek M, et al. Prevalence, characteristics, and prognostic implications of type 2 diabetes in patients with myocardial infarction: the Polish Registry of Acute Coronary Syndromes (PL-ACS) annual 2018 report. Kardiol Pol. 2020;78:243–6. https://doi.org/10.33963/KP.15189.

    Article  PubMed  Google Scholar 

  5. Leander K, Blombäck M, Wallén H, He S. Impaired fibrinolytic capacity and increased fibrin formation associate with myocardial infarction. Thromb Haemost. 2012;106:1092–9. https://doi.org/10.1160/TH11-11-0760.

    Article  CAS  Google Scholar 

  6. Zalewski J, Bogaert J, Sadowski M, Woznicka O, Doulaptsis K, Ntoumpanaki M, et al. Plasma fibrin clot phenotype independently affects intracoronary thrombus ultrastructure in patients with acute myocardial infarction. Thromb Haemost. 2015;113:1258–69. https://doi.org/10.1160/TH14-09-0801.

    Article  PubMed  Google Scholar 

  7. Undas A, Wiek I, Stepien E, Zmudka K, Tracz W. Hyperglycemia is associated with enhanced thrombin formation, platelet activation, and fibrin clot resistance to lysis in patients with acute coronary syndrome. Diabetes Care. 2008;31:1590–5. https://doi.org/10.2337/dc08-0282.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Dunn EJ, Ariëns RAS, Grant PJ. The influence of type 2 diabetes on fibrin structure and function. Diabetologia. 2005;48:1198–206. https://doi.org/10.1007/s00125-005-1742-2.

    Article  CAS  PubMed  Google Scholar 

  9. Hess K, Alzahrani SH, Price JF, Strachan MW, Oxley N, King R, et al. Hypofibrinolysis in type 2 diabetes: the role of the inflammatory pathway and complement C3. Diabetologia. 2014;57:1737–41. https://doi.org/10.1007/s00125-014-3267-z.

    Article  CAS  PubMed  Google Scholar 

  10. Pieters M, Covic N, Loots DT, van der Westhuizen FH, van Zyl DG, Rheeder P, et al. The effect of glycaemic control on fibrin network structure of type 2 diabetic subjects. Thromb Haemost. 2006;96:623–9. https://doi.org/10.1160/TH06-07-0390.

    Article  CAS  PubMed  Google Scholar 

  11. Jörneskog G, Egberg N, Fagrell B, Fatah K, Hessel B, Johnsson H, et al. Altered properties of the fibrin gel structure in patients with IDDM. Diabetologia. 1996;39:1519–23.

    Article  Google Scholar 

  12. Konieczynska M, Fil K, Bazanek M, Undas A. Prolonged duration of type 2 diabetes is associated with increased thrombin generation, prothrombotic fibrin clot phenotype and impaired fibrinolysis. Thromb Haemost. 2014;111:685–93. https://doi.org/10.1160/TH13-07-0566.

    Article  CAS  PubMed  Google Scholar 

  13. Bochenek M, Zalewski J, Sadowski J, Undas A. Type 2 diabetes as a modifier of fibrin clot properties in patients with coronary artery disease. J Thromb Thrombolysis. 2013;35:264–70. https://doi.org/10.1007/s11239-012-0821-8.

    Article  CAS  PubMed  Google Scholar 

  14. Neergaard-Petersen S, Hvas A-M, Kristensen SD, Grove EL, Larsen SB, Phoenix F, et al. The influence of type 2 diabetes on fibrin clot properties in patients with coronary artery disease. Thromb Haemost. 2014;112:1142–50. https://doi.org/10.1160/TH14-05-0468.

    Article  CAS  PubMed  Google Scholar 

  15. Alessi MC, Juhan-Vague I. PAI-1 and the metabolic syndrome: links, causes, and consequences. Arterioscler Thromb Vasc Biol. 2006;26:2200–7. https://doi.org/10.1161/01.ATV.0000242905.41404.68.

    Article  CAS  PubMed  Google Scholar 

  16. Hori Y, Gabazza EC, Yano Y, Katsuki A, Suzuki K, Adachi Y, et al. Insulin resistance is associated with increased circulating level of thrombin-activatable fibrinolysis inhibitor in type 2 diabetic patients. J Clin Endocrinol Metab. 2002;87:660–5. https://doi.org/10.1210/jcem.87.2.8214.

    Article  CAS  PubMed  Google Scholar 

  17. Dunn EJ, Philippou H, Ariëns RAS, Grant PJ. Molecular mechanisms involved in the resistance of fibrin to clot lysis by plasmin in subjects with type 2 diabetes mellitus. Diabetologia. 2006;49:1071–80. https://doi.org/10.1007/s00125-006-0197-4.

    Article  CAS  PubMed  Google Scholar 

  18. Svensson J, Bergman A-C, Adamson U, Blombäck M, Wallén H, Jörneskog G. Acetylation and glycation of fibrinogen in vitro occur at specific lysine residues in a concentration dependent manner: a mass spectrometric and isotope labeling study. Biochem Biophys Res Commun. 2012;421:335–42. https://doi.org/10.1016/j.bbrc.2012.03.154.

    Article  CAS  PubMed  Google Scholar 

  19. Ajjan RA, Gamlen T, Standeven KF, Mughal S, Hess K, Smith KA, et al. Diabetes is associated with posttranslational modifications in plasminogen resulting in reduced plasmin generation and enzyme-specific activity. Blood. 2013;122:134–42. https://doi.org/10.1182/blood-2013-04-494641.

    Article  CAS  PubMed  Google Scholar 

  20. Wittbrodt E, Bhalla N, Andersson K, Qi S, Dong L, Cavender MA, et al. Assessment of the high risk and unmet need in patients with CAD and type 2 diabetes (ATHENA): US healthcare resource utilization, cost and burden of illness in the Diabetes Collaborative Registry. Endocrinol Diab Metab. 2020;3:e00133. https://doi.org/10.1002/edm2.133.

    Article  CAS  Google Scholar 

  21. Pignatelli P, Menichelli D, Pastori D, Violi F. Oxidative stress and cardiovascular disease: new insights. Kardiol Pol. 2018;76:713–22. https://doi.org/10.5603/KP.a2018.0071.

    Article  PubMed  Google Scholar 

  22. Ramanathan R, Gram JB, Sidelmann JJ, Dey D, Kusk MW, Nørgaard BL, et al. Sex difference in fibrin clot lysability: association with coronary plaque composition. Thromb Res. 2019;174:129–36. https://doi.org/10.1016/j.thromres.2018.12.020.

    Article  CAS  PubMed  Google Scholar 

  23. Zhang L, Xu C, Liu J, Bai X, Li R, Wang L, et al. Baseline plasma fibrinogen is associated with haemoglobin A1c and 2-year major adverse cardiovascular events following percutaneous coronary intervention in patients with acute coronary syndrome: a single-centre, prospective cohort study. Cardiovasc Diabetol. 2019;18:1–12. https://doi.org/10.1186/s12933-019-0858-5.

    Article  PubMed  PubMed Central  Google Scholar 

  24. Li M, Duan L, Cai YL, Li HY, Hao BC, Chen JQ, et al. Growth differentiation factor-15 is associated with cardiovascular outcomes in patients with coronary artery disease. Cardiovasc Diabetol. 2020;19:1–12. https://doi.org/10.1186/s12933-020-01092-7.

    Article  CAS  Google Scholar 

  25. Sumaya W, Wallentin L, James SK, Siegbahn A, Gabrysch K, Himmelmann A, et al. Impaired fibrinolysis predicts adverse outcome in acute coronary syndrome patients with diabetes: a PLATO sub-study. Thromb Haemost. 2020;120:412–22. https://doi.org/10.1055/s-0039-1701011.

    Article  PubMed  PubMed Central  Google Scholar 

  26. Neergaard-Petersen S, Larsen SB, Grove EL, Kristensen SD, Ajjan RA, Hvas AM. Imbalance between fibrin clot formation and fibrinolysis predicts cardiovascular events in patients with stable coronary artery disease. Thromb Haemost. 2020;120:75–82. https://doi.org/10.1055/s-0039-1700873.

    Article  PubMed  Google Scholar 

  27. Cosentino F, Grant PJ, Aboyans V, Bailey CJ, Ceriello A, Delgado V, et al. 2019 ESC guidelines on diabetes, pre-diabetes, and cardiovascular diseases developed in collaboration with the EASD. Eur Heart J. 2020;41:255–323. https://doi.org/10.1093/eurheartj/ehz486.

    Article  PubMed  Google Scholar 

  28. Stepień E, Plicner D, Branicka A, Stankiewicz E, Pazdan A, Sniezek-Maciejewska M, et al. Factors influencing thrombin generation measured as thrombin-antithrombin complexes levels and using calibrated automated thrombogram in patients with advanced coronary artery disease. Pol Arch Med Wewn. 2007;117:297–305. https://doi.org/10.20452/pamw.162.

    Article  PubMed  Google Scholar 

  29. Mills JD, Ariëns RAS, Mansfield MW, Grant PJ. Altered fibrin clot structure in the healthy relatives of patients with premature coronary artery disease. Circulation. 2002;106:1938–42.

    Article  CAS  Google Scholar 

  30. Undas A, Szułdrzynski K, Stepien E, Zalewski J, Godlewski J, Tracz W, et al. Reduced clot permeability and susceptibility to lysis in patients with acute coronary syndrome: effects of inflammation and oxidative stress. Atherosclerosis. 2008;196:551–7. https://doi.org/10.1016/j.atherosclerosis.2007.05.028.

    Article  CAS  PubMed  Google Scholar 

  31. Undas A, Podolec P, Zawilska K, Pieculewicz M, Jedliński I, Stgpień E, et al. Altered fibrin clot structure/function in patients with cryptogenic ischemic stroke. Stroke. 2009;40:1499–501. https://doi.org/10.1161/STROKEAHA.108.532812.

    Article  CAS  PubMed  Google Scholar 

  32. Undas A, Slowik A, Wolkow P, Szczudlik A, Tracz W. Fibrin clot properties in acute ischemic stroke: relation to neurological deficit. Thromb Res. 2010;125:357–61. https://doi.org/10.1016/j.thromres.2009.11.013.

    Article  CAS  PubMed  Google Scholar 

  33. Lisman T, Leebeek FW, Mosnier LO, Bouma BN, Meijers JC, Janssen HL, et al. Thrombin-activatable fibrinolysis inhibitor deficiency in cirrhosis is not associated with increased plasma fibrinolysis. Gastroenterology. 2001;121:131–9.

    Article  CAS  Google Scholar 

  34. Williams S, Fatah K, Ivert T, Blomback M. The effect of acetylsalicylic acid on fibrin gel lysis by tissue plasminogen activator. Blood Coagul Fibrinolysis. 1995;6:718–25. https://doi.org/10.1097/00001721-199512000-00004.

    Article  CAS  PubMed  Google Scholar 

  35. Collet JP, Mishal Z, Lesty C, Mirshahi M, Peynet J, Baumelou A, et al. Abnormal fibrin clot architecture in nephrotic patients is related to hypofibrinolysis: Influence of plasma biochemical modifications. A possible mechanism for the high thrombotic tendency? Thromb Haemost. 1999;82:1482–9. https://doi.org/10.1055/s-0037-1614859.

    Article  CAS  Google Scholar 

  36. Undas A, Celinska-Löwenhoff M, Löwenhoff T, Szczeklik A. Statins, fenofibrate, and quinapril increase clot permeability and enhance fibrinolysis in patients with coronary artery disease. J Thromb Haemost. 2006;4:1029–36. https://doi.org/10.1111/j.1538-7836.2006.01882.x.

    Article  CAS  PubMed  Google Scholar 

  37. Formanowicz D, Wanic-Kossowska M, Pawliczak E, Radom M, Formanowicz P. Usefulness of serum interleukin-18 in predicting cardiovascular mortality in patients with chronic kidney disease-systems and clinical approach. Sci Rep. 2015;5:1–13. https://doi.org/10.1038/srep18332.

    Article  CAS  Google Scholar 

  38. R Foundation for Statistical Computing. R: a language and environment for statistical computing; 2013.

  39. Carter AM, Cymbalista CM, Spector TD, Grant PJ. Heritability of clot formation, morphology, and lysis: the EuroCLOT study. Arterioscler Thromb Vasc Biol. 2007;27:2783–9. https://doi.org/10.1161/ATVBAHA.107.153221.

    Article  CAS  PubMed  Google Scholar 

  40. Chernysh IN, Nagaswami C, Kosolapova S, Peshkova AD, Cuker A, Cines DB, et al. The distinctive structure and composition of arterial and venous thrombi and pulmonary emboli. Sci Rep. 2020;10:5112. https://doi.org/10.1038/s41598-020-59526-x.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Badimon L, Vilahur G. Thrombosis formation on atherosclerotic lesions and plaque rupture. J Intern Med. 2014;276:618–32. https://doi.org/10.1111/joim.12296.

    Article  CAS  PubMed  Google Scholar 

  42. Okraska-Bylica A, Wilkosz T, Słowik L, Bazanek M, Konieczyńska M, Undas A. Altered fibrin clot properties in patients with premature peripheral artery disease. Pol Arch Med Wewn. 2012;122:608–15. https://doi.org/10.20452/pamw.1535.

    Article  CAS  PubMed  Google Scholar 

  43. Barco S, Konstantinides SV. Risk-adapted management of pulmonary embolism. Thromb Res. 2017;151:S92–6. https://doi.org/10.1016/S0049-3848(17)30076-2.

    Article  CAS  PubMed  Google Scholar 

  44. Kearney K, Tomlinson D, Smith K, Ajjan R. Hypofibrinolysis in diabetes: a therapeutic target for the reduction of cardiovascular risk. Cardiovasc Diabetol. 2017;16:34. https://doi.org/10.1186/s12933-017-0515-9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Standeven KF, Ariens RAS, Whitaker P, Ashcroft AE, Weisel JW, Grant PJ. The effect of dimethylbiguanide on thrombin activity, FXIII activation, fibrin polymerization, and fibrin clot formation. Diabetes. 2002;51:189–97. https://doi.org/10.2337/diabetes.51.1.189.

    Article  CAS  PubMed  Google Scholar 

  46. Ajjan RA, Grant PJ. Cardiovascular disease prevention in patients with type 2 diabetes: the role of oral anti-diabetic agents. Diabetes Vasc Dis Res. 2006;3:147–58. https://doi.org/10.3132/dvdr.2006.023.

    Article  Google Scholar 

  47. Weisel JW, Litvinov RI. Fibrin formation, structure and properties. Subcell Biochem. 2017;82:405–56. https://doi.org/10.1007/978-3-319-49674-0.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Lados-Krupa A, Konieczynska M, Chmiel A, Undas A. Increased oxidation as an additional mechanism underlying reduced clot permeability and impaired fibrinolysis in type 2 diabetes. J Diabetes Res. 2015. https://doi.org/10.1155/2015/456189.

    Article  PubMed  PubMed Central  Google Scholar 

  49. Bryk A, Konieczynska M, Rostoff P, Broniatowska E, Hohendorff J, Malecki M, et al. Plasma protein oxidation as a determinant of impaired fibrinolysis in type 2 diabetes. Thromb Haemost. 2019;119:213–22. https://doi.org/10.1055/s-0038-1676609.

    Article  PubMed  Google Scholar 

  50. Bhatt DL, Eikelboom JW, Conolly SJ, Steg PG, Anand SS, Verma S, et al. Role of combination antiplatelet and anticoagulation therapy in diabetes mellitus and cardiovascular disease Insights From the COMPASS Trial. Circulation. 2020;141:1841–54. https://doi.org/10.1161/CIRCULATIONAHA.120.046448.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Ajjan RA, Standeven KF, Khanbhai M, Phoenix F, Gersh KC, Weisel JW, et al. Effects of aspirin on clot structure and fibrinolysis using a novel in vitro cellular system. Arterioscler Thromb Vasc Biol. 2009;29:712–7. https://doi.org/10.1161/ATVBAHA.109.183707.

    Article  CAS  PubMed  Google Scholar 

  52. Carter RLR, Talbot K, Hur WS, Meixner SC, van der Gugten JG, Holmes D, et al. Rivaroxaban and apixaban induce clotting factor Xa fibrinolytic activity. J Thromb Haemost. 2018;16:2276–88. https://doi.org/10.1111/jth.14281.

    Article  CAS  PubMed  Google Scholar 

  53. Schneider JG, Isermann B, Kleber ME, Wang H, Boehm BO, Grammer TB, et al. Inverse association of the endogenous thrombin potential (ETP) with cardiovascular death: the Ludwigshafen Risk and Cardiovascular Health (LURIC) study. Int J Cardiol. 2014;176:139–44. https://doi.org/10.1016/j.ijcard.2014.07.026.

    Article  PubMed  Google Scholar 

  54. Van PPCS, Panova-noeva M, Van OR, Hermanns IM, Prochaska JH, Arnold N, et al. Thrombin generation in cardiovascular disease and mortality—results from the Gutenberg Health Study. Haematologica. 2020;105:2327–34. https://doi.org/10.3324/haematol.2019.221655.

    Article  CAS  Google Scholar 

  55. Tian R, Tian M, Wang L, Qian H, Zhang S, Pang H, et al. Cytokine C-reactive protein for predicting cardiovascular and all-cause mortality in type 2 diabetic patients: a meta-analysis. Cytokine. 2019;117:59–64. https://doi.org/10.1016/j.cyto.2019.02.005.

    Article  CAS  PubMed  Google Scholar 

  56. Castoldi E, Rosing J. Thrombin generation tests. Thromb Res. 2011;127:S21–5. https://doi.org/10.1016/S0049-3848(11)70007-X.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We would like to thank Dr. Joanna Natorska, PhD and Dr. Michal Zabczyk, PhD for technical support.

Funding

None.

Author information

Author notes

  1. Agata Hanna Bryk and Małgorzata Konieczyńska contributed equally to the authorship of this article

Authors and Affiliations

  1. Department of Endocrinology, Jagiellonian University Medical College, Krakow, Poland

    Agata Hanna Bryk

  2. John Paul II Hospital, Krakow, Poland

    Agata Hanna Bryk, Małgorzata Konieczyńska & Anetta Undas

  3. Department of Epidemiology and Population Studies, Institute of Public Health, Faculty of Health Sciences, Jagiellonian University Medical College, Krakow, Poland

    Maciej Polak

  4. Unit of Experimental Cardiology and Cardiac Surgery, Faculty of Medicine and Health Sciences, Andrzej Frycz Modrzewski Krakow University, Krakow, Poland

    Dariusz Plicner

  5. Clinic of Cardiac Transplantation and Mechanical Circulatory Support, Department of Heart Diseases, Wroclaw Medical University, Wrocław, Poland

    Maciej Bochenek

  6. Institute of Cardiology, Jagiellonian University Medical College, 80 Pradnicka St., 31-202, Krakow, Poland

    Anetta Undas

Authors
  1. Agata Hanna Bryk
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Małgorzata Konieczyńska
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Maciej Polak
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Dariusz Plicner
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Maciej Bochenek
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Anetta Undas
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

AHB contributed to the work by acquisition of data, analysis and interpretation of data, as well as writing of the manuscript. MK contributed to the work by acquisition of data, writing and revising the manuscript. MP provided statistical analysis of the data. DP and MB contributed to the work by acquisition of data and revised the manuscript. AU made substantial contributions to conception and design of the study, analysis and interpretation of data, revision of the manuscript and final approval of the version to be published. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Anetta Undas.

Ethics declarations

Ethics approval and consent to participate

The study complied with the principles of Good Clinical Practice rules and was approved by the Regional Medical Chamber Ethics Committee. Each study participant provided written informed consent.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Bryk, A.H., Konieczyńska, M., Polak, M. et al. Plasma fibrin clot properties and cardiovascular mortality in patients with type 2 diabetes: a long‐term follow‐up study. Cardiovasc Diabetol 20, 47 (2021). https://doi.org/10.1186/s12933-021-01230-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s12933-021-01230-9

Keywords

  • Type 2 diabetes
  • Fibrin clot
  • D-dimer
  • Cardiovascular mortality

Cardiovascular Diabetology

ISSN: 1475-2840