Immature Platelet Fraction and Clinical Outcomes in Patients Undergoing Transcatheter Aortic Valve Implantation

Abstract

Introduction

Immature platelets or reticulated platelets are newly released thrombocytes. They can be identified by their large size and high RNA cytoplasm concentration. Immature platelet fraction (IPF) represents the percentage of immature circulative platelets relative to the total number of platelets. The role of IPF in patients undergoing transcatheter aortic valve implantation (TAVI) is unknown. The aim of the current trial was to assess the levels of IPF in patients undergoing TAVI and correlation with clinical outcomes.

Material and Methods

Immature platelet fraction levels were measured 3 times in all patients (preprocedure, 1-2 days post-procedure and 1-month post-procedure). Immature platelet fraction measurement was carried out using an autoanalyzer (Sysmex XE-2100). Patients were followed for 12 months. Primary outcomes were defined as complications during hospitalizations, rehospitalization, and mortality.

Results

Fifty-one patients were included in the study. Mean age was 79.8 (±9.6), and 28 (55%) were women. Twenty-one patients (41%) had complications: Of them, 6 of 21 (29%) occurred during hospitalizations (2—vascular complications; 2—sepsis, 2—implantation of a pacemaker), 9 of 21 (43%) patients were rehospitalized after the index admission, and 6 patients died during the follow-up period. Multivariate Cox regression analysis found that IPF < 7% in at least one of the 3 tests was associated with worse outcomes (hazard ratio 3.42; 95% CI 1.11-10.5, P = .032).

Conclusion

Immature platelet fraction >7% in patients undergoing TAVI is associated with worse outcomes. Further studies are needed to better understand this phenomenon.

Introduction

Platelets play an essential role in atherothrombotic events, and platelet hyperactivity is associated with adverse outcomes in patients with acute coronary syndrome (ACS). ¹ Platelet populations are heterogeneous in terms of size, reactivity, and responsiveness to antiplatelet therapies. ² Reticulated platelets (RPs) or immature platelets are “young” platelets with content of dense granules, a residual amount of megakaryocyte-derived messenger ribonucleic acid and increased mean volume when compared to older circulating platelets. ³ Reticulated platelets are believed to more active and thus more prothrombotic than mature platelets. ⁴ Detection of these platelets in the bloodstream reflects increased platelet production from megakaryocytes in the bone marrow, suggesting a high rate of platelet turnover. ⁵ Immature platelet fraction (IPF) represents the percentage of RP's compared with the total number of platelets.

Immature platelet fraction is increased in patients with diseases characterized by increased platelet turnover, such as bleeding and thrombocytopenic purpura. High IPF has also been documented in other conditions such as in diabetes and smoking.^6,7 Increased IPF was reported in patients with arterial thrombotic events including ACS.^8,9 Recently, IPFs were found to be independent predictors of poor cardiovascular prognosis and cardiovascular death. ¹⁰ In patients with atrial fibrillation, ablation therapy has been shown to decrease IPF. ¹¹

Increased IPF has been correlated with impaired response to various antiplatelet drugs in patients with coronary disease. ⁸ Although there are only limited data regarding high residual platelet reactivity,^12,13 knowledge regarding the effect of transcatheter aortic valve implantation (TAVI) on IPF is lacking. Severe aortic stenosis patients are usually elderly patients with comorbidities, who are predisposed to inflammation and platelet dysfunction likely due to disturbed flow and shear exposition and activation. ¹⁴

The aim of the current study was to examine IPF levels and its stability over time in patients undergoing TAVI, and to evaluate the prognostic implications of IPF levels on 30-day clinical outcomes.

Methods

The prospective study was conducted from July 2021 to October 2021 and comprised 51 patients undergoing TAVI and admitted to a tertiary care medical center intensive cardiac care unit. All patients were treated with either aspirin and/or clopidogrel for antiplatelet therapy or with novel oral anticoagulants if indicated, at the discretion of the treating physician. Exclusion criteria: (1) age < 18 years; (2) active bleeding or bleeding diathesis; (3) history of gastrointestinal or genitourinary bleeding within 2 months; (4) major surgery within 6 weeks prior to enrollment; (5) history of intracranial bleeding or intracranial structural abnormalities; (6) known severe liver disease or renal failure (glomerular filtration rate < 30 mL/min); (7) pregnancy; (8) human immunodeficiency virus treatment; (9) a psychiatric disorder or inability to sign a consent form, and (10) bleeding during TAVI requiring blood transfusion or any blood transfusion within one month prior to enrollment.

Data Collection

Data were prospectively and anonymously documented in an electronic case report form. Data were checked for accuracy and out-of-range values by the study coordinator. Demographic data, presenting symptoms, comorbid conditions, and physical examination were systematically recorded. Laboratory, imaging, hemodynamic, and clinical course data were collected as well.

The study was approved by the SZMC Institutional Review Board (approval number 0094-19-SZMC). All patients provided written informed consent.

Immature Platelet Fraction Testing

Venous blood samples were drawn from each patient at 3 time points: (1) prior to the TAVI procedure, (2) 1 to 2 days after the procedure (prior to discharge), and (3) 1-month after the TAVI procedure. These time points were chosen in order to capture preprocedural baseline IPF levels, and because the first 30 days following cardiac events or procedures has been associated with increased platelet reactivity, reduced response to antiplatelet drugs, and increased risk for recurrent ischemic events.^12,15

Immature Platelet Fraction Measurement

The level of IPF was determined as a part of complete blood cell count by an autoanalyzer (Sysmex XN-3000, Sysmex America Inc), which uses fluorescent dyes containing oxazine and ethylene glycol. ¹⁵ This system discriminates between mature and immature platelets and reports the IPF as a percentage. Reticulated platelets are displayed as a percentage of the total optical platelet count. Immature platelet count (IPC) is calculated by multiplying IPF and total platelet count, representing the absolute count of immature platelets (×10³/μL). The normal reference range for IPF% and IPC used in our laboratory (as defined by Sysmex) is 1.2% to 8.6% and 3.6-20.0/103 ×µL, respectively. The analytical error in measuring IPF is 3.6%.

Follow-Up

Thirty-day clinical follow-up information was obtained at an outpatient follow-up visit and by chart review. The composite outcome of cardiovascular events was defined as complications during hospitalizations (vascular complications, infection, arrhythmia, requirement of permanent pacemaker), rehospitalization, or death during the study period.

Statistical Analysis

Patient characteristics were presented as numbers (%) for categorical variables, and as means (standard deviation) or medians (interquartile range) for normal and non-normal distributed continuous variables, respectively. Comparison of categorical variables was done by Chi-square test and Fisher exact tests. Student t test and Mann-Whitney test were performed for comparison of normally and non-normally distributed continuous variables, respectively. The Friedman test (nonparametric alternative to the 1-way ANOVA with repeated measures) was used for analyzing the measures of IPF. Post hoc analysis with Wilcoxon signed-rank tests was conducted with a Bonferroni correction applied. The probability of composite outcome according to the study groups was calculated according to the method of Kaplan-Meier and was displayed using the adjusted cox survival function (ggadjustedcurves function from the Survminer package in R). Univariate and multivariate Cox proportional hazards regression modeling were used to compare patients who had at least one IPF test >7% to patients who did not, with adjustments made for parameters who were found to be significant in the univariate model or are recognized to impact the survival of patients after TAVI. The cutoff value of 7% was based on the literature review^16-18 and by the 75th quantile of IPF values, and for a grouping of the patients in a ratio of 1:2. The adjusted cox model incorporated the following variables: age, sex, body mass index, smoking, prior coronary artery events, and diabetes mellitus (DM). All analyses were performed R software version 3.4.4 (R Foundation for Statistical Computing). An association was considered statistically significant with a 2-sided P value less than .05.

Results

Patient Enrollment and Characteristics

Fifty-one patients fulfilled the study inclusion criteria and comprised the study population. Baseline characteristics were similar in both groups and are presented in Table 1. Mean age was 77 (±7.7) and 23 (45%) patients were men. A total of 19 (37.2%) patients had DM, 13 (25.4%) patients had atrial fibrillation, and 21 (41.1%) patients had prior coronary artery disease (CAD).

Table 1.

Patient Baseline Characteristics.

	N = 51
Age(years)	77.86 (±7.75)
Male gender	23 (45.1%)
Body mass index	28.50(±7.16)
Current smoker	7 (13.7%)
Prior coronary disease	23 (45.1%)
Atrial fibrillation	13 (25.5%)
Diabetes mellitus	19(37.3%)
Malignancy	8 (15.7%)
IPF 1	5.239 (±2.95)
IPF 2	5.12 (±3.18)
IPF 3	4.698 (±3.196)

Abbreviation: IPF, immature platelet fraction.

In-Hospital Complications

Five patients (9.8%) had in-hospital complications: 2 had vascular complications (minor dissection, pseudoaneurysm), 2 had infections, and 1 required insertion of a permanent pacemaker.

Adverse Outcomes During Follow-Up

During 1 year of follow up, 16 (31.3%) patients had adverse outcomes: 10 (20%) patients required rehospitalization and 6 (12%) patients died.

Immature Platelet Fraction Values and Outcomes

The crude number of positive outcomes among patients with IPF >7 was 8 of 17 (47%) compared to 13 of 34 (38%) among patients with IPF lower than or equal to 7. Kaplan-Meier survival analysis revealed that the cumulative probability of the composite outcome at 1 year of follow-up was 47.1% ± 12.1%, and 42% ± 9.2%, respectively. Differentiation by IPF values alone failed to demonstrate statistical significance between groups, as seen in Table 2. However, multivariate Cox analysis demonstrated that compared with patients that all there 3 IPF test were lower than or equal to 7, those with at least one test above 7 were 3 times more likely to show adverse outcomes, including death during follow-up (hazard ratio = 3.42, 95% CI 1.11-10.5, P = .032). Figure 1 displays the survival rates based on the adjusted Cox model. In addition, the multivariate model identified several other independent predictors of adverse outcomes, including male sex and atrial fibrillation (Figure 2).

Table 2.

Primary Outcomes.

	Adverse out comes (N = 21)	No adverse outcomes (N = 30)	P value
Sex (male)mean(SD)	13 (61.9%)	10 (33.3%)	.083
BMI	27.5 (7.72)	29.2 (6.52)	.183
Smoking	4 (19.0%)	3 (10.0%)	.61
Prior CAD	11 (52.4%)	12 (40.0%)	.556
Atrial fibrillation	10 (47.6%)	3 (10.0%)	.007
Malignancy	3 (14.3%)	5 (16.7%)	1
Diabetes mellitus	11 (52.4%)	8(26.7%)	.115
Age	78.4 (8.92)	77.5 (6.96)	.443
IPF > 7	8 (38.1%)	9 (30.0%)	.763
IPF 1	4.81 (2.84)	5.54 (3.05)	.353
IPF 2	4.84 (2.59)	5.32 (3.57)	.962
IPF 3	4.65 (2.98)	4.73 (3.39)	.954

Abbreviations: BMI, body mass index; CAD, coronary artery disease; IPF, immature platelet fraction.

Figure 1.

Adjusted COX survival.

Figure 2.

Multivariate COX model is demonstrating hazard ratio (HR) and the 95% CI of HR.

Discussion

To our knowledge, this is the first study to examine the dynamic changes in platelet turnover indices expressed as IPF in patients undergoing TAVI to evaluate long-term prognostic implications. Our study revealed that IPF levels in patients who experienced adverse outcomes were not significantly different from those seen in patients with better outcomes at 1-year follow-up. However, our results demonstrated that an IPF < 7% in at least one test was associated with worse prognosis in this population.

Our findings are supported by other studies and laboratory reports which found the cutoff of IPF > 7% to be associated with worse outcomes.^16-18

The study enrolled relatively older patients with multiple underlying diseases, who may have decreased bone marrow production of platelets. As a result, the IPF levels in these patients may be lower compared to younger patients with the same condition.

As IPF is a relatively a recent biomarker requiring additional research, there is limited evidence for its role in various cardiovascular conditions. Existing research predominantly focuses on its utility as a prognostic indicator and its association with adverse events in individuals with CAD, cardiac surgery and cyanotic heat defects.^{4,7-10,15,18,19}

Previous studies investigating morbidity and mortality predictors in patients with ACS have identified IPF as an independent predictor of cardiovascular mortality, with IPF values above 3.3% predicting cardiovascular death. ¹⁰ However, our study investigated patients with chronic valvular malfunction who underwent elective procedures, and as a result may exhibit lower inflammation and potentially different IPF levels when compared to ACS patients. Our findings indicated that a single IPF measurement <7% increases the risk for morbidity and mortality in patients undergoing TAVI, and that this could be a useful predictor of outcome in this patient population.

Several platelet parameters necessitate further investigation in future studies. Such as platelet count, which may indicate thrombocytopenia following the procedure. Research has established a correlation between thrombocytopenia and adverse outcomes in TAVI patients. Where IPF may offer insights into the mechanism of thrombocytopenia. ¹⁸

Another potentially significant parameter is platelet size, as evidenced by Pizzulli et al ²⁰ study revealing alterations in platelet size and count in patients with ACS. ²¹ However, there is currently a gap of literature addressing this specific aspect in the context of TAVI.

The present study has some limitations: First, it was a single-center study that involved a relatively small sample size. Nevertheless, we included all-comers in a real-world setting. Notably, due to the small study size and the use of multivariate analysis the establishment of our conclusions require further research. Second, the relationship between IPF and mean platelet volume, which has been shown to exhibit a positive correlation, was not evaluated. ²² Additionally, IPF levels were only measured one month after the procedure, and therefore, any temporal changes in IPF levels beyond this time frame remain unknown.

Conclusions

Immature platelet fraction >7% in patients undergoing TAVI is associated with worse outcomes. Further studies are warranted to evaluate IPF and other hematological indices which correlate with adverse prognoses in this patient population.