|Year : 2017 | Volume
| Issue : 4 | Page : 148-154
Role of measurement of interleukin 10 in idiopathic (immune) thrombocytopenic purpura
Hanan Hamed1, Mohamed Moussa1, Hanaa Fathey1, Heba Tolba2
1 Department of Hematology, Faculty of Medicine, Ain Shams University, Cairo, Egypt
2 Hematology & BMT Unit, El-Shiekh Zayed Specialized Hospital, Giza, Egypt
|Date of Submission||19-May-2017|
|Date of Acceptance||29-May-2017|
|Date of Web Publication||9-Feb-2018|
Hematology & BMT Unit, El-Shiekh Zayed Specialized Hospital, Giza, 12555
Source of Support: None, Conflict of Interest: None
Aim Biological markers useful for defining newly diagnosed patient with immune thrombocytopenic purpura (ITP) who are likely to develop the chronic form of the disease are partially lacking. The purpose of this study was to assess the clinical role of a cytokine in patients with ITP and correlate its levels with different stages of the disease.
Patients and methods A total of 20 patients with ITP at the onset stage, 40 patients with chronic ITP, and 30 healthy matched controls were enrolled in this study. Serum levels of interleukin 10 (IL-10) were measured in all patients using quantitative immunoenzymatic assays.
Result Serum IL-10 levels were significantly higher in patients with an acute evolution of ITP than in either healthy controls or patients with chronic ITP.
Conclusion IL-10 seems to predict the clinical course of ITP, as it is significantly higher at the onset of the disease in patients who obtain disease remission in less than 1 year.
Keywords: adult, cytokines, interleukin 10, immune thrombocytopenic purpura
|How to cite this article:|
Hamed H, Moussa M, Fathey H, Tolba H. Role of measurement of interleukin 10 in idiopathic (immune) thrombocytopenic purpura. Egypt J Haematol 2017;42:148-54
|How to cite this URL:|
Hamed H, Moussa M, Fathey H, Tolba H. Role of measurement of interleukin 10 in idiopathic (immune) thrombocytopenic purpura. Egypt J Haematol [serial online] 2017 [cited 2018 Apr 20];42:148-54. Available from: http://www.ehj.eg.net/text.asp?2017/42/4/148/225086
| Introduction|| |
Autoimmune diseases are characterized by various factors that contribute to a breakdown in self-tolerance, that is, the ability of the immune system to effectively distinguish self from non-self and to refrain from attacking self. Autoimmune diseases include a broad spectrum of disorders, such as idiopathic thrombocytopenic purpura, systemic lupus erythematosus, rheumatoid arthritis, systemic sclerosis, and inflammatory bowel disease. Although significant progress has been achieved in the development of approaches to the treatment of autoimmune diseases, the etiologies, and pathogenesis of autoimmune diseases remain obscure .
Primary immune thrombocytopenic purpura (ITP) is an autoimmune-mediated bleeding disorder characterized by thrombocytopenia which results owing to increased platelet destruction and platelet production decrease by antiplatelet autoantibodies .
The pathophysiology of ITP is heterogeneous and complex. Research advances highlight that a complex dysregulation of the immune system is involved in the pathogenesis of this condition . Many studies in recent years have shown that the abnormities of T lymphocyte, dendritic cell, natural killer cell, cytokines, programmed cell death, oxidative stress, infection, pregnancy, and drugs play an important role in the pathogenesis of ITP .
Among these abnormalities include the increased number of the T helper 1 (Th1) cells , the decreased number or defective suppressive function of regulatory T cells (Tregs) (Yu et al., 2008) , and the platelet destruction by cytotoxicity T lymphocytes . Moreover, dysregulated T cells in patients with ITP may enable the development of platelet autoantibodies, have a direct cytotoxic effect on platelets, and impair platelet production by megakaryocytes .
Cytokines are molecules responsible for controlling intracellular communication and directing the immunological reaction .
Interleukin 10 (IL-10) is an anti-inflammatory cytokine and suppresses immune responses . IL-10 is secreted by macrophages, Th2 cells, and mast cells. Cytotoxic T cells also release IL-10 to inhibit viral infection stimulated natural killer cell activity . IL-10 inhibits the synthesis of a number of cytokines involved in the inflammatory process including IL-2, IL-3, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor α (TNF-α), and interferon γ (IFN-γ) . It can promote the activity of mast cells, B cells, and certain T cells .
| Patients and methods|| |
A total of 20 patients were referred to our department for primary ITP. They were enrolled in this study at the onset stage of their disease before starting any treatment along with 40 patients with chronic ITP (lasting at least 2 years) and 30 age-matched and sex-matched healthy controls. The diagnosis of ITP was made following standardized guidelines ,,,. The site and extent of bleeding symptoms at the onset of the disease were recorded, and three different clinical phenotypes were identified according to national consensus 11: (i) type I (asymptomatic–paucisymptomatic ITP), in the presence of clinical symptoms ranging from no bleeding to few petechiae and some bruises without mucosal hemorrhages; (ii) type II (intermediate ITP), in the presence of petechiae, bruising, and mucosal hemorrhages; and (iii) type III (severe ITP), characterized by severe bleeding with organ impairment or life-threatening conditions (retinal or intracranial hemorrhage, other severe internal hemorrhages, and shock).
All patients gave their consent to participate in the study, which was approved by the local ethics committee.
A blood specimen was obtained from all patients at the time of first visit. Serum levels of IL-10 were measured using a quantitative enzyme-linked immunosorbent assay (ELISA; R&D Systems, Minneapolis, Minnesota, USA) following the manufacturer’s instructions for sample collection, storage, and assay procedure. The detection limits for the IL-10 were 3.9 pg/ml.
| Results|| |
The demographic and clinical characteristics of all enrolled patients, which included 20 patients with ITP who were enrolled at the onset of the disease, 40 patients who had chronic ITP, and the 30 healthy control individuals, are summarized in [Table 1],[Table 2],[Table 3],[Table 4],[Table 5] and [Figure 1].
|Table 1 Comparison between groups according to demographic data and presentation|
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|Table 4 Correlation between interleukin 10 and other parameters using Pearson’s correlation coefficient in the acute group|
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|Table 5 Correlation between interleukin 10 and other parameters using Pearson correlation coefficient in the chronic group|
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| Discussion|| |
In this study, we recruited a series of newly diagnosed ITP cases at the onset of the disease, before starting any treatment, and a group of patients already having chronic ITP who received treatment (intravenous immunoglobulin or steroids) for their condition. In all these patients, we sought for significant variations in cytokines level during the clinical progression (newly diagnosed or chronic) of their disease to determine the potential diagnostic and prognostic role of the serum biomarker in adult ITP.
The demographic characteristics of our population were similar to those reported by other authors. In line with the other studies, our patients with chronic ITP showed a higher prevalence of asymptomatic–paucisymptomatic form of ITP compared with patients with newly diagnosed ITP.
We found that among patients with ITP, 15.6% were males and 84.4% were females, showing a female predominance, with age range of 18–50 years with mean of 31.8 years, which was in agreement with the results of Zhan et al. ; Hua et al. ; Del Vecchio et al. ; Giordano et al. ; Li et al. ; Talaat et al. ; and Elsalakawy et al. . All these studies were performed on patients with ITP and showed that ITP among adults affects females more than males, with no specific age predominance. On comparing our three groups, we found no statistically significant difference between them.
On the contrary, Arandi et al.  showed that the incidence of ITP is predominant in males in childhood.
Regarding the presentation in our patients, we observed that 100% of patients with acute ITP were presented by petechial rash, 90% had epistaxis, and 80% had combined skin and mucous membrane bleeding, whereas 12.5% of patients with chronic ITP were presented by petechial rash, 5% had epistaxis, and 6% had combined skin and mucous membrane bleeding. In contrast, the control group did not show any picture of bleeding. There was highly statistically significant difference between the three groups (P<0.001).
These data are comparable to the classical ITP presentation reported by Talaat et al.  who found that clinical features were present in all patients in the form of petechial rash in 100% of patients and epistaxis in 91.4% (32 patients of 35 patients) of patients. In agreement with our study was the study by Vianelli et al. ; the study was performed on 310 patients with ITP over a period of 10 years, and only one hemorrhagic death had occurred. Others reported that petechiae and ecchymosis were present in 99% of patients with childhood ITP, and epistaxis was the most frequent hemorrhagic manifestation.
Regarding disease course, chronic ITP was reported in 40 (71.1%) patients, whereas acute ITP was reported in 20 (28.9%) patients. Likewise, the study of Del Vecchio et al.  included 28 (79.4%) patients with acute ITP and 11 (20.6%) patients with chronic ITP.
In the present study, there is no statistically significant difference between our groups with respect to any other comorbidities (diabetes mellitus, hypertension, etc.).
Our study showed that one (5%) of the acute cases resorted to splenectomy and 12 (30%) patient with chronic ITP underwent splenectomy as a line of treatment in comparison with the control group, in which the spleen was present in all individuals, showing statistically significant difference between the three groups (P<0.003). This finding is comparable with the study by Del Vecchio et al.  that reported that 33% of cases underwent splenectomy compared with the healthy group.
Assessment of laboratory data in the current study revealed a highly statistically significant difference between the groups regarding platelets count, with lower counts in patients with acute ITP than those with chronic ITP (P<0.001) in comparison with healthy controls, which is in agreement with the former study by Elsalakawy et al. .
Moreover, the study revealed statistically significant difference between erythrocyte sedimentation rate levels among the groups, which is increased in patients with acute ITP than those with chronic ITP in comparison with healthy controls.
The study revealed that there is no statistically significant difference between the three groups concerning coagulation profile and liver and kidney functions, which is in agreement with the former study by Elsalakawy et al.  and Del Vecchio et al. .
Regarding the levels of serum IL-10, our results revealed the following.
In the acute ITP group, IL-10 levels ranged from 70.1 to 221.4 ng/ml. The mean was 103.3 ng/ml, and the SD was 43.2 ng/ml.
In the chronic ITP group, IL-10 levels ranged from 39.4 to 128.6 ng/ml. The mean was 68.3 ng/ml, and the SD was 16.4 ng/ml.
The comparison between the cases and control groups with respect to IL-10 levels showed that there was a statistically significant difference between the three studied groups, where the IL-10 levels in the cases were significantly higher (P<0.001).
These results were in accordance with Del Vecchio et al.  who investigated 28 acute ITP cases and 11 chronic ITP cases for their levels of serum IL-10 and found the levels to be statistically significantly higher in the patients than the control groups (P<0.001).
The study by Lee and Yoon Kim (2016) was conducted to assess the cytokine imbalances such as serum levels of IL-2, IL-10, and transforming growth factor β1 (TGF-β1) in childhood chronic ITP during the thrombocytopenic and spontaneous remission phases.
The findings of this study offer new insights into the pathogenesis of childhood chronic ITP and the prognostic factors involved in spontaneous remission. Of the 10 children with chronic ITP, six thrombocytopenic, and four recovered cases were enrolled. Moreover, five healthy children and eight healthy adults were included in this study as controls. The serum levels of IL-2, IL-10, and TGF-β1 were measured by ELISA technique during this study; four patients achieved spontaneous remission.
The serum level of IL-10 was higher in the patients who recovered spontaneously compared with the patients with persistent chronic thrombocytopenia. The serum levels of IL-2 were not detectable, as the levels were too low for analysis. TGF-β1 showed no significant differences among the groups in this study.
These data suggest that increased serum level of IL-10 could have prognostic significance in the natural course and long-term outcome of childhood with chronic ITP.
Talaat et al.  also studied 35 children with ITP (15 acute and 20 chronic) and 10 healthy controls. Plasma levels of Th1 cytokines (IFN-γ and TNF-α) and Th2 (IL-4, IL-6, and IL-10) cytokines were measured using ELISA. All evaluated cytokines (IFN-γ, TNF-α, IL-4, IL-6, and IL-10) were elevated significantly in patients with ITP (P<0.001, <0.05, <0.05, <0.05, and <0.001, respectively) compared with controls.
They concluded that the immune cells and their related cytokines have fundamental role in patients with ITP and are predictors of ITP progression.
Tesse et al.  found derangement of genetic and immunological factors seems to have a pivotal role in the pathophysiology of ITP. They investigated IL-10 genetically determined expression in children with an acute progression of ITP (n=41) compared with young patients with chronic ITP (n=44) and healthy controls (n=60), and attempted to correlate IL-10 production with the course of the disease. They genotyped their study population for three single nucleotide polymorphisms at positions −1082 (A/G), −819 (C/T), and −592 (C/A) in the promoter region of the IL-10 gene. IL-10 levels were measured by enzyme-linked immunoassay.
The IL-10 production in this study population was significantly higher in patients carrying the GCC haplotype than those bearing ACC and ATA haplotypes.
The serum concentration of IL-10 was significantly higher in patients with an acute course of their disease, who mainly carried the GCC haplotype (92%), compared with patients with chronic form of the disease, bearing the non-GCC haplotypes, and controls [17 pg/ml (1.7–18) vs. 3.5 pg/ml (0.6–11) vs. 3 pg/ml (1–7), P<0.01)].
Their findings show that patients carrying the GCC-high producer IL-10 haplotype have an acute development of ITP and that IL-10 levels might represent a useful predictive biomarker of the disease course.
Liet et al. (2014) studied 22 adult patients with newly diagnosed ITP and 16 sex-matched and age-matched healthy human patients enrolled as normal controls. Their study showed that the percentages of IL-10 Tregs in Tregs’ population were found elevated dramatically in patients with ITP but decreased in the remitted patients.
Moreover, Giordano et al.  studied 28 children with ITP at the onset of their disease, who were followed up for a whole year, and they were divided according to whether their disease resolved within the 12 months or became chronic. The study included 11 patients with chronic ITP off therapy for at least 1 month at the time of enrollment, and 30 healthy matched controls. Serum levels of Th1 and Th2 and Tregs-associated cytokines, such as IFN-γ, TNF-α, IL-2, IL-6, IL-10, and thrombopoietin, were measured in all children using quantitative immunoenzymatic assays, whereas reticulated platelets were evaluated by flow cytometric analysis.
They concluded that serum IL-10 levels were significantly higher in patients with an acute evolution of ITP than in either healthy controls (P<0.001) or patients with chronic progression of ITP (P<0.05).
Therefore, IL-10 seems to predict the clinical course of ITP, as it is significantly higher at the onset of disease in patients who obtain disease remission in less than 1 year.
Regarding the levels of serum IL-10 levels, our results revealed the following:
In the acute patient group, IL-10 levels ranged from 70.1 to 221.4 ng/ml. The mean was 103.3 ng/ml, and the SD was 43.2 ng/ml.
In the chronic group, IL-10 levels ranged from 39.4 to 128.6 ng/ml. The mean was 68.3 ng/ml, and the SD was 16.4 ng/ml.
In the control group, IL-35 levels ranged from 29.0 to 67.9 ng/ml. The mean was 48.1 ng/ml.
The comparison between cases and control groups with respect to IL-10 levels showed that there was a statistically significant difference between the two studied groups, where the levels in cases were significantly higher (P<0.001).
Our results were different from those of El Ghannam et al.  who investigated the relationship of IL-10 promoter (IL-10 −1082, −819, and −592) polymorphisms with the susceptibility, severity, and outcome of adult chronic ITP in a cohort of Egyptian population.
Typing of IL-10 promoter polymorphisms was done using restriction fragment length polymorphism for 62 adult patients with chronic ITP and 73 age-matched and sex-matched healthy controls.
No significant differences were found between patients with ITP and controls regarding the frequency of IL-10 promoter genotypes, alleles, or haplotypes. IL-10 −592 AA genotype and ATA (IL-10 −1082, −819, and −592) haplotype were associated with severe ITP (P=0.003 and 0.043, respectively).
They concluded that the IL-10 promoter polymorphisms are unlikely to affect the development or treatment outcome of chronic adult ITP in Egyptian population, but IL-10-592 AA genotype and IL-10 (−1082, −819, and −592) ATA haplotype may be associated with disease severity, This is because ITP is a complex disease.
Moreover, our results were different from Makhlouf and Abd Elhamid .
The study showed that IL-10 A alleles were detected more frequently among patients with ITP compared with controls.
A statistically significant difference was observed in IL-10 and IL-4 gene polymorphism distribution between patients with acute ITP and those with chronic ITP, with higher A allele and RP2 allele among patients with chronic ITP versus patients with acute ITP.
According to Mouzaki et al. , increased levels of IL-10 were found in children with chronic ITP, further suggesting Th1 involvement in the pathology of ITP illustrated by an increase in the Th1 cytokines; these findings may be related to ongoing immune activation related to autoimmunity.
| Conclusion|| |
In conclusion, it is intriguing that high serum levels of IL-10 were found at the onset of ITP in patient with an acute disease course (and thus resolution), in agreement with the role of this cytokine in immunotolerance. The serum level of IL-10 in patient with ITP at the onset stage may provide a promising indicator of the clinical progression of the disease.
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Conflicts of interest
There are no conflicts of interest.
| References|| |
Tao JH, Cheng M, Tang JP et al.
Foxp3, regulatory T cell, and autoimmune diseases. Inflammation
Zhan Y, Hua F, Ji L et al.
Polymorphisms of the IL-23R gene are associated with primary immune thrombocytopenia but not with the clinical outcome of pulsed high-dose dexamethasone therapy. Ann Hematol
Consolini R, Legitimo A, Caparello MC. The centenary of immune thrombocytopenia − part 1: revising nomenclature and pathogenesis. Front Pediatr
Yang ZZ, Novak AJ, Ziesmer SC et al.
Attenuation of CD8+ T-cell function by CD4+CD25+ regulatory T cells in B-cell non-Hodgkin’s lymphoma. Cancer Res
Panitsas FP, Theodoropoulou M, Kouraklis A et al.
Adult chronic idiopathic thrombocytopenic purpura (ITP) is the manifestation of a type-1 polarized immune response. Blood
Yu J, Heck S, Patel V et al.
Defective circulating CD25 regulatory T cells in patients with chronic immune thrombocytopenic purpura. Blood
Zhao C, Li X, Zhang F et al.
Increased cytotoxic T-lymphocyte-mediated cytotoxicity predominant in patients with idiopathic thrombocytopenic purpura without platelet autoantibodies. Haematologica
Poniatowski LA, Wojdasiewicz P, Gasik R et al.
Transforming growth factor beta family: insight into the role of growth factors in regulation of fracture healing biology and potential clinical applications. Mediators Inflamm
Wei AH, Schoenwaelder SM, Andrews RK, Jackson Sp. New insights into the haemostatic function of platelets. Br J Haematol
Ouyang W, Rutz S, Crellin NK et al.
Regulation and functions of the IL-10 family of cytokines in inflammation and disease. Annu Rev Immunol
Kuhn R, Lohler J, Rennick D et al.
Interleukin-10-deficient mice develop chronic enterocolitis. Cell
Iyer and Cheng, et al.
.. Role of interleukin 10 transcriptional regulation in inflammation and autoimmune disease. Crit Rev Immunol
Zhan Y, Zou S, Hua F et al.
High dose dexamethasone modulates serum cytokine profile in patients with primary immune thrombocytopenia. Immunol let
Hua F, Ji L, Zhan Y et al.
Pulsed high-dose dexamethasone improves interleukin 10 secretion by CD5+ B cells in patients with primary immune thrombocytopenia. J Clin Immunol
Del Vecchio GC, Giordano P, Tesse R et al.
Clinical significance of serum cytokine levels and thrombopoietic markersin childhood idiopathic thrombocytopenic purpura Blood Transfus
Giordano P et al.
Association of interleukin-(IL)10 haplotypes and serum IL-10 levels in the progression of childhood immune thrombocytopenic purpura. Gene
Li X, Zhong H, Bao W et al.
Defective regulatory B-cell compartment in patients with immune thrombocytopenia. Blood
Talaat RM, Elmaghraby AM, Barakat SS et al.
Alterations in immune cell subsets and their cytokine secretion profile in ITP. Clinical and Experimental Immunology
Elsalakawy WA, Ali MA, Hegazy MG et al.
Value of vanin-1 assessment in adult patients with primary immune thrombocytopenia. Platelets
Arandi N, Mirshafiey A, Jeddi-Tehrani M et al.
Alteration in frequency and function of CD4 +CD25 +FOXP3+regulatory T cells in patients with immune thrombocytopenic purpura. Iran J Allergy Asthma Immunol
Vianelli N, Valdre L, Fiacchini M et al.
Long-term follow-up of idiopathic thrombocytopenic purpura in 310 patients. Haematologica
Talaat RM, Elmaghraby AM, Barakat SS, El-Shahat M. Alterations in immune cell subsets and their cytokine secretion profile in childhood idiopathic thrombocytopenic purpura (ITP). Clin Exp Immunol
Tesse R, Del Vecchio GC, De Mattia D et al.
Association of interleukin-(IL)10 haplotypes and serum IL-10 levels in the progression of childhood ITP. Gene
El Ghannam D et al.
Relation of interleukin-10 promoter polymorphisms to adult chronic immune thrombocytopenic purpura in a cohort of Egyptian population. Immunol Invest
Makhlouf M, Abd Elhamid S. Expression of IL4 (VNTR intron 3) and IL10 (−627) genes polymorphisms in childhood immune thrombocytopenic purpura. Lab Med
Mouzaki A, Theodoropoulou M, Gianakopoulos I, Vlaha V, Kyrtsonis MC et al.
Expression patterns of Th1 and Th2 cytokine genes in childhood idiopathic thrombocytopenic purpura (ITP) at presentation and their modulation by intravenous immunoglobulin G (IVIg) treatment: their role in prognosis. Blood
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]