Clinical utility of immature platelet fraction in newly diagnosed immune thrombocytopenia: a prospective observational study of diagnostic performance, bleeding risk, and corticosteroid response

¹Department of Hematology, Cho Ray Hospital, Ho Chi Minh City, Vietnam, ²Australian Centre for Blood Diseases, School of Translational Medicine, Monash University, Melbourne, Australia

^&Corresponding author
Ngoc-Sang Thi Nguyen, Department of Hematology, Cho Ray Hospital, Ho Chi Minh City, Vietnam

Introduction    

Immune thrombocytopenia (ITP) is a common hematologic disorder characterised by a transient or persistent reduction in platelet count, resulting in an increased risk of bleeding. Clinical manifestations range from asymptomatic disease or mild bruising and petechiae to severe mucosal or internal bleeding that may be life-threatening [1-3]. The condition is primarily associated with immune-mediated platelet destruction; however, the precise pathophysiological mechanisms underlying ITP have not been fully elucidated [2]. In particular, platelet-specific autoantibodies promote peripheral platelet destruction and inhibit platelet release from megakaryocytes, leading to thrombocytopenia [4]. Current first-line treatment strategies, including corticosteroids, intravenous immunoglobulins in emergency settings, splenectomy, and other immunosuppressive therapies, aim to reduce immune-mediated platelet destruction and improve platelet survival [3].

ITP is characterised by abnormally low platelet counts resulting from a combination of increased peripheral platelet destruction and impaired platelet production in the bone marrow [5]. Nevertheless, the relative contribution of these two mechanisms remains incompletely understood [6]. This distinction is clinically important, as it may influence disease progression and responsiveness to first-line corticosteroid therapy [7,8]. Therefore, the availability of a reliable and quantitative marker to assess platelet production in the bone marrow is essential for improving the understanding of ITP pathophysiology and optimising individualised treatment strategies. In response to increased platelet destruction in ITP, immature platelets are released prematurely from the bone marrow into the peripheral circulation. These platelets are larger in size and exhibit greater reactivity compared with mature platelets [9]. Because the proportion of immature platelets reflects the rate of thrombopoiesis, it serves as a surrogate marker of platelet production.

The immature platelet fraction (IPF) is a peripheral blood parameter designed to quantify the proportion of immature platelets within the total platelet population. IPF is useful in differentiating the underlying causes of thrombocytopenia [10]. In addition, IPF has demonstrated clinical value in predicting platelet recovery in various conditions, including post-transplantation, chemotherapy-induced thrombocytopenia, and infectious diseases [11-16]. Reported IPF values in patients with ITP vary considerably across studies, ranging from 5.7% to 22.3% [17]. Moreover, Takami et al. [18] demonstrated that pretreatment IPF may serve as a predictor of corticosteroid responsiveness in newly diagnosed ITP patients [18]. At present, corticosteroids remain the standard first-line therapy for newly diagnosed ITP. Treatment response at predefined time points is commonly used to evaluate therapeutic efficacy and guide corticosteroid tapering while balancing clinical benefits against potential adverse effects [19,20]. In this context, early treatment response (1 week), initial treatment response (1 month), and progression to persistent ITP according to international consensus criteria (disease duration between 3 and 12 months) represent clinically relevant outcome measures.

In Vietnam, no studies to date have evaluated the clinical utility of IPF in newly diagnosed ITP. Therefore, this prospective observational study aimed to evaluate the diagnostic and prognostic value of immature platelet fraction (IPF) in newly diagnosed immune thrombocytopenia (ITP), including its ability to differentiate ITP from thrombocytopenia caused by bone marrow pathology, predict clinically significant bleeding, and assess corticosteroid treatment response and disease progression. Specifically, the study addressed three research questions: (1) Can IPF differentiate newly diagnosed ITP from thrombocytopenia caused by bone marrow pathology? (2) Can baseline IPF predict corticosteroid treatment response and progression to persistent ITP? and (3) Is baseline IPF associated with clinically significant bleeding in newly diagnosed ITP?

Methods     

Study design: this prospective observational analytical study evaluated immature platelet fraction (IPF) dynamics and clinical characteristics in patients with newly diagnosed primary immune thrombocytopenia (ITP) compared with patients with non-ITP thrombocytopenia.

Setting: the study was conducted at a tertiary hospital hematology service. All clinical and laboratory assessments were performed as part of routine diagnostic and treatment care. IPF and hematologic measurements were analysed in the hospital clinical laboratory using standardised procedures. The study was conducted between May 2022 and September 2023 at the Department of Hematology, Cho Ray Hospital, Ho Chi Minh City, Vietnam.

Participants: patients aged ≥16 years with newly diagnosed primary immune thrombocytopenia were consecutively enrolled. Primary ITP was diagnosed according to the 2019 American Society of Hematology (ASH) guidelines [2]. Secondary causes of thrombocytopenia were excluded through comprehensive clinical and laboratory evaluation, including autoimmune serologic testing and bone marrow aspiration when clinically indicated. Disease duration terminology was defined according to the International Working Group consensus classification as newly diagnosed ITP (<3 months), persistent ITP (3-12 months), and chronic ITP (>12 months) [1-3]. Patients with refractory response to corticosteroids or progression to persistent or chronic ITP underwent additional investigations, including repeat bone marrow examination and extended autoimmune testing, to exclude alternative hematologic or systemic etiologies. A comparison group consisted of patients with platelet counts <50 G/L due to other hematologic causes of thrombocytopenia, including acute leukaemia, chronic leukaemia, myelodysplastic syndrome, and bone marrow pathology. Exclusion criteria were evidence of chronic inflammation, liver fibrosis, septicemia, current antiplatelet therapy, or platelet transfusion within 48 hours before blood sampling. Chronic inflammation was defined by clinical history of chronic inflammatory or autoimmune disease (e.g., rheumatoid arthritis, systemic lupus erythematosus, chronic infection) or persistently elevated inflammatory markers at enrollment (C-reactive protein >10 mg/L or erythrocyte sedimentation rate >30 mm/h).

Variables: the primary laboratory variable was immature platelet fraction (IPF). Baseline IPF measured at admission before corticosteroid therapy was defined as IPF₀. IPF measured on day 7 after corticosteroid initiation was defined as IPF₁. The change in IPF (IPF₀-IPF₁) represented an early thrombopoietic response. Additional hematologic variables included platelet count and mean platelet volume (MPV). Clinical variables included age, sex, hemorrhagic status at admission, and clinically significant bleeding. Bleeding severity and treatment response were classified according to International Working Group (IWG) ITP criteria [1].

Data sources and measurement: peripheral venous blood samples were collected at hospital admission using standard aseptic technique into ethylenediaminetetraacetic acid (EDTA) anticoagulated tubes. Samples were analysed within 2 hours of collection to minimise pre-analytical variation. Platelet count, MPV, and IPF were measured using a Sysmex XN-3000 automated hematology analyser (Sysmex Corporation, Kobe, Japan) according to the manufacturer's instructions and laboratory quality control standards. IPF testing was performed before initiation of corticosteroid therapy. After confirmation of primary ITP, all patients received first-line corticosteroid treatment with methylprednisolone 1-2 mg/kg/day in accordance with ASH 2019 recommendations.

Bias: selection bias was minimised by enrolling consecutive eligible patients and applying standardised diagnostic criteria for ITP. Information bias was reduced by using automated hematology analysis and predefined laboratory timing (baseline and day 7). Exclusion of conditions affecting platelet kinetics (inflammation, transfusion, antiplatelet therapy) further limited confounding. Residual bias related to observational design and clinical heterogeneity may remain.

Study size: the study sample comprised all eligible ITP and non-ITP thrombocytopenia patients meeting inclusion criteria during the study period. A total of 106 patients were included in the final analysis.

Quantitative variables: continuous variables included platelet count, MPV, IPF₀, IPF₁, and IPF change. These were summarised as mean ± standard deviation for normally distributed data or median with interquartile range for non-normal data. A platelet count <50 G/L defined severe thrombocytopenia for control group eligibility. IPF change was calculated as the difference between day 7 and baseline values.

Statistical methods: categorical variables were summarised as frequencies and percentages. Continuous variables were presented as mean ± standard deviation or median and interquartile range, depending on data distribution. Comparisons between two independent groups were performed using the Mann-Whitney U test for non-normally distributed continuous variables. Comparisons among more than two independent groups were performed using one-way analysis of variance for normally distributed variables and the Kruskal-Wallis test for non-normally distributed variables. A two-sided p-value <0.05 was considered statistically significant. To explore the differentiation between immune thrombocytopenia (ITP) and bone marrow pathology (BMP)-related thrombocytopenia, baseline immature platelet fraction (IPF) values were compared between groups using the Mann-Whitney U test. To evaluate factors associated with clinically significant bleeding, univariate and multivariable binary logistic regression analyses were performed. To assess corticosteroid response and subsequent disease course, baseline IPF and day-7 IPF values were compared between responders and non-responders, followed by logistic regression analyses to identify predictors of early and initial treatment non-response. Variables with p <0.10 in univariate analysis, together with clinically relevant variables based on prior literature, were entered into multivariable binary logistic regression models using the enter method. Odds ratios (ORs) with 95% confidence intervals (CIs) were reported. Receiver operating characteristic (ROC) curve analysis was performed to determine optimal IPF cutoff values for predicting clinically significant bleeding and corticosteroid treatment response. Optimal thresholds of 10.4% and 14.25% were selected based on the maximum Youden index. Statistical analyses were performed using IBM SPSS Statistics version 25.0.

Ethical consideration statement: the study protocol was reviewed and approved by the Ethics Committee of Cho Ray Hospital, Ho Chi Minh City, Vietnam (Approval No. 602/HDDD-BVCR, issued on 15^th April 2022). Written informed consent was obtained from all participants before enrollment. The study was conducted in accordance with the Declaration of Helsinki and national regulations governing biomedical research involving human participants. The study was observational and did not alter routine diagnostic or therapeutic management. IPF measurements were obtained from the same blood samples collected for routine complete blood count testing, and no additional blood sampling was required.

Results     

Baseline characteristics of the study population: patients with ITP had a younger age distribution, with a median age of 38.5 years, and showed a marked female predominance (72.6%) (Table 1). Bleeding manifestations were observed in 81.3% of ITP patients, of whom 25.8% experienced clinically significant bleeding. The median platelet count in the ITP group was 6 × 10⁹/L (IQR: 2.8-11.0), indicating severe thrombocytopenia at presentation. Mean platelet volume (MPV) was 10.2 fL (IQR: 8.5-11.4) and did not differ significantly between the ITP and BMP groups. In contrast, IPF was significantly higher in ITP patients than in those with BMP-related thrombocytopenia (16.8% vs. 4.0%, p <0.001), suggesting higher platelet production activity in ITP than in thrombocytopenia associated with bone marrow pathology (Table 1).

Baseline IPF and corticosteroid response: patients who did not achieve early or initial treatment response had significantly lower baseline IPF values than responders, whereas IPF measured on day 7 did not differ significantly between groups (Table 2). For early treatment non-response, both age ≥65 years and baseline IPF <14.25% were independent predictors in multivariable analysis. Similarly, baseline IPF <14.25% remained independently associated with initial treatment non-response, whereas age and platelet count did not retain statistical significance (Table 3). Among 87 patients followed for 3 months, baseline IPF differed significantly according to disease course. Patients who maintained treatment response had higher median baseline IPF values than those who progressed to persistent ITP or lost response (p = 0.01), supporting the prognostic relevance of baseline IPF in predicting early disease evolution.

Baseline IPF and clinically significant bleeding: baseline IPF was significantly associated with bleeding risk. In univariate analysis, IPF <10.4% was strongly associated with clinically significant bleeding (OR: 4.7; 95% CI: 1.7-13.2; p = 0.003), and this association remained significant after multivariable adjustment (OR: 4.5; 95% CI: 1.3-16.2; p = 0.02). Although older age and low platelet count were associated with bleeding in univariate analysis, they were not independent predictors in the multivariable model (Table 3).

Discussion     

The present study addressed three clinically relevant research questions regarding the utility of IPF in newly diagnosed ITP: diagnostic discrimination from BMP thrombocytopenia, association with bleeding severity, and prediction of corticosteroid response. Our findings suggest that baseline IPF may provide complementary diagnostic and prognostic information in routine clinical practice.

In this study, 106 patients with newly diagnosed immune thrombocytopenia (ITP) underwent baseline blood testing, including assessment of the immature platelet fraction (IPF), at hospital admission. The median baseline IPF (IPF₀) was 16.8%, supporting previous observations that elevated IPF at diagnosis is associated with preserved thrombopoietic activity in ITP. Our findings are consistent with prior studies demonstrating that higher IPF values are associated with a more favourable response to corticosteroid therapy in patients with primary ITP. Takami et al. [18] reported an IPF cutoff value of 12% with high sensitivity and specificity for predicting treatment response, and our results further support the utility of IPF as a predictive biomarker in this clinical setting.

Beyond baseline assessment, dynamic changes in IPF during treatment may provide additional clinical insight. An increase in IPF typically precedes platelet count recovery by several days, allowing early anticipation of treatment response, whereas a secondary rise in IPF accompanied by declining platelet counts may indicate relapse or renewed marrow stress [21]. In line with previous reports, our study also confirms the diagnostic value of IPF in differentiating ITP from other causes of thrombocytopenia [1,22-24]. Moreover, the observation that IPF is markedly elevated and stable in hereditary macrothrombocytopenia (HM) suggests that IPF may assist not only in distinguishing hyper- from hypoproliferative thrombocytopenias, but also in differentiating ITP from HM, thereby reinforcing its role in diagnostic algorithms [24,25].

Consistent with these findings, we observed significantly higher IPF values in ITP patients compared with those with bone marrow pathology, supporting the role of IPF in distinguishing hyperproliferative thrombocytopenia from conditions characterised by impaired platelet production. Previous studies have shown that IPF levels in HM may even exceed those observed in ITP, despite differing pathophysiological mechanisms-immune-mediated peripheral platelet destruction in ITP versus defective proplatelet formation related to megakaryocyte cytoskeletal abnormalities in HM [24,25]. Structural alterations, such as abnormal alpha-tubulin distribution in megakaryocytes, further explain these differences [25]. Collectively, these findings underscore the utility of IPF as a non-invasive marker in the initial evaluation of thrombocytopenia.

Our results also demonstrate a significant association between lower IPF values and increased bleeding risk in ITP patients. McDonnell et al. reported that an IPF <10.4% independently predicted severe and life-threatening bleeding [26], while Bride et al. observed significantly lower IPF values in pediatric patients with major bleeding [27]. Similarly, in our cohort, patients with clinically significant bleeding had a lower median IPF₀ compared with those without significant bleeding, with a statistically significant difference. Minor discrepancies in cutoff values between studies may reflect differences in patient populations and bleeding definitions, as previous studies primarily focused on life-threatening hemorrhage, whereas our definition included both moderate and severe bleeding [26]. These findings reinforce the role of IPF not only as a diagnostic indicator but also as a potential tool for bleeding risk stratification in ITP, particularly in patients with profound thrombocytopenia.

Multivariate analysis further confirmed that IPF₀ <10.4% was independently associated with clinically significant bleeding, with an odds ratio of 4.5. This finding is concordant with previous studies identifying low IPF as a predictor of severe bleeding [26]. In addition, age ≥65 years was associated with a higher bleeding risk in our cohort. This observation is supported by data from a large French study demonstrating a linear relationship between age and the risk of gastrointestinal and intracranial hemorrhage in ITP, although no specific age threshold was defined [4]. Taken together, these results suggest that both low IPF and advanced age are relevant factors in bleeding risk assessment, while current evidence does not support a direct association between age and IPF levels in ITP populations [18,26,28].

Assessment of treatment response at early time points, particularly at 1 and 4 weeks, is a critical component of corticosteroid-based ITP management. In the Takami et al. study of 46 newly diagnosed ITP patients, median IPF values were significantly higher in responders than in non-responders, with an optimal cutoff of 12% [18]. In our study, both median IPF values and optimal cutoff thresholds were higher than those reported previously. Although our cohort had a younger median age and a larger sample size, these differences alone are unlikely to fully explain the observed variability. Given the limited number of studies evaluating IPF as a predictor of treatment response, further research is required to establish standardised cutoff values across different populations [17,18].

Beyond the initial response, disease persistence and loss of treatment response remain major clinical challenges in adult ITP. Among the 87 patients followed for more than 3 months, nearly half progressed to the persistent phase, and a substantial proportion experienced loss of response during follow-up. Patients with persistent disease had significantly lower baseline IPF₀ values than those who maintained treatment response. Adult ITP is known to carry a higher risk of chronicity and treatment refractoriness, particularly during corticosteroid tapering or discontinuation [2,29]. Reported rates of chronic or refractory progression in adults range from 20% to 50%, consistent with our findings [29].

This study has several limitations. First, the study was conducted at a single tertiary referral center, which may limit the generalizability of the findings to other clinical settings and populations. Second, the sample size was relatively modest, particularly for subgroup analyses evaluating treatment response and persistent disease progression. Third, the follow-up duration was limited, preventing assessment of long-term progression to chronic ITP and long-term corticosteroid outcomes. Fourth, external validation of the proposed IPF cutoff values was not performed. Finally, although multivariable analyses were conducted, residual confounding inherent to observational study designs cannot be completely excluded.

Overall, monitoring treatment response and disease progression in ITP remains challenging, particularly in patients who fail to respond to first-line therapy or experience relapse. Although no single biomarker has been established as a gold standard for prognostic stratification, IPF measurement at diagnosis offers several advantages: it is non-invasive, low-cost, and readily available as part of automated complete blood count testing. Our findings, together with existing evidence, support the integration of IPF into the diagnostic and prognostic evaluation of ITP to improve risk stratification and guide individualised management strategies [2,17,24].

Conclusion     

Baseline immature platelet fraction was associated with diagnostic differentiation between ITP and BMP-related thrombocytopenia, clinically significant bleeding, corticosteroid treatment response, and short-term disease course in patients with ITP. Lower IPF at diagnosis was related to higher bleeding risk, poorer response to first-line corticosteroid therapy, and greater likelihood of persistent disease or loss of response. Because IPF can be obtained from routine automated blood count analysis without additional blood sampling, it may provide a practical adjunctive marker for risk stratification in patients with thrombocytopenia. Larger prospective studies are needed to validate these findings and define standardised IPF thresholds for clinical use.

Competing interests     

The authors declare no competing interests.

Authors' contributions     

Tung Thanh Tran: Study design, patient recruitment, data collection, and manuscript drafting. Ngoc-Sang Thi Nguyen: Conceptualization, supervision of study implementation, interpretation of results, and critical revision of the manuscript. Tuan Huu Nguyen: Statistical analysis, data interpretation, and manuscript revision. Thao Thi Nguyen: Clinical data acquisition and patient assessment. Van Thi Kim Pham: Laboratory coordination and hematologic data validation. Thoa Thi Nguyen: Data management and quality control. All authors read and approved the final version of the manuscript.

Acknowledgments     

The authors thank the medical and laboratory staff of the Department of Hematology, Cho Ray Hospital, for their assistance in patient management and data collection.

Tables     

Table 1: demographic, clinical, and laboratory characteristics of patients with newly diagnosed immune thrombocytopenia (ITP) and thrombocytopenia due to bone marrow pathology (BMP), Department of Hematology, Cho Ray Hospital, Ho Chi Minh City, Vietnam, May 2022 to September 2023 (N = 106)

Table 2: comparison of baseline and day-7 immature platelet fraction (IPF) values according to corticosteroid treatment response among patients with newly diagnosed immune thrombocytopenia, Department of Hematology, Cho Ray Hospital, Ho Chi Minh City, Vietnam, May 2022 to September 2023

Table 3: univariate and multivariable logistic regression analyses of factors associated with clinically significant bleeding and corticosteroid treatment non-response in patients with newly diagnosed immune thrombocytopenia, Department of Hematology, Cho Ray Hospital, Ho Chi Minh City, Vietnam, May 2022 to September 2023

References