|Year : 2018 | Volume
| Issue : 1 | Page : 1-4
Cytokines and immunoglobulin derangement in egyptian children with primary immune thrombocytopenic purpura
Ebeid S. E Fatma1, Khalifa S Ahmed1, El-Kinawy S Nihal2, Helmy Y Salwa1
1 Department of Paediatrics, Ain Shams University, Cairo, Egypt
2 Department of Clinical Pathology, Ain Shams University, Cairo, Egypt
|Date of Submission||05-Sep-2017|
|Date of Acceptance||12-Oct-2017|
|Date of Web Publication||3-Aug-2018|
Ebeid S. E Fatma
Department of Paediatrics, Ain Shams University, Cairo, 11566
Source of Support: None, Conflict of Interest: None
Background Immune thrombocytopenic purpura (ITP) is a heterogeneous immune-mediated disorder in which the immune system reacts with cytokine-mediated T lymphocytes and a platelet autoantigen(s).
Aim The aim of this study was to detect helper T-1 and T-2 response through assessment of tumor necrosis factor (TNF)-α and interleukin (IL)-6, respectively, and immunoglobulin (Ig)M, IgG, and IgA in patients with acute and chronic ITP and to detect their possible roles as predictors of the course of the disease.
Patients and methods This cross-sectional case–control study, conducted at Ain Shams University, recruited 20 patients with ITP (10 acute and 10 chronic) and 10 age-matched and sex-matched healthy controls. All were subjected to detailed clinical assessment, assessment of serum IL-6 and TNF-α using enzyme-linked immunosorbent assay, and serum IgA, IgM, and IgG using nephlometric method.
Results Mean serum values of IgM and IgG were statistically lower in patients with acute ITP, and statistically higher serum values of TNF-α were detected in patients with acute and chronic ITP compared with controls. IL-6 levels were statistically higher in acute ITP when compared with chronic ITP and with controls and a higher significant value in chronic ITP than controls. There were significant positive correlations between IgM, IgG, and platelet count and a significant negative correlation between both TNF-α and IL-6 and platelet count. On follow-up of patients with acute ITP, one of them developed a chronic progressive course, and this patient was found to have low serum levels of IgA and IgG at presentation.
Conclusion Children with primary ITP had dysfunction at cytokine and serum Ig level.
Keywords: cytokines, Egypt, immune thrombocytopenic purpura, immunoglobulins
|How to cite this article:|
Fatma ES, Ahmed KS, Nihal EKS, Salwa HY. Cytokines and immunoglobulin derangement in egyptian children with primary immune thrombocytopenic purpura. Egypt J Haematol 2018;43:1-4
|How to cite this URL:|
Fatma ES, Ahmed KS, Nihal EKS, Salwa HY. Cytokines and immunoglobulin derangement in egyptian children with primary immune thrombocytopenic purpura. Egypt J Haematol [serial online] 2018 [cited 2020 Jul 14];43:1-4. Available from: http://www.ehj.eg.net/text.asp?2018/43/1/1/238541
| Introduction|| |
In immune thrombocytopenic purpura (ITP), platelet membrane proteins become antigenic and stimulate the immune system to produce autoantibodies and cytotoxic T cells. Once produced, autoantibody may bind either to platelets, causing their destruction, or to megakaryocytes, resulting in decreased thrombopoiesis .
Dysfunction of T cells in ITP may be contributed to loss of tolerance, and impairment of the delicate balance of specific cytokine and serum cytokines may play a role in the pathogenesis of ITP . T lymphocytes polarize into helper T-1 response characterized primarily by the presence of cytokines interleukin (IL)-2, interferon-γ, and tumor necrosis factor (TNF)-α and helper T-2 response produces IL-4, IL-5, IL-6, IL-10, and IL-13 ,. Understanding the role of T-cell subsets will permit a better control of autoimmunity through manipulation of their cytokine network . Dysfunction at cellular immunities is evaluated by the levels of cytokines, e.g. TNF-α and IL-6. Measurement of levels of cytokines may help in prediction of the course of ITP .
This study was designed to assess helper T-1 and helper T-2 response through assessment of serum level of TNF-α and IL-6, respectively, and to assess the serum level of immunoglobulin (Ig)G, IgM, and IgA in patients with acute and chronic ITP. Moreover, it was conducted to detect their correlations to each other and assess their possible role as a predictor of the course of the disease.
| Patients and methods|| |
This is a case–control study conducted at Pediatric Hematology/Oncology Department, Ain Shams University, Egypt. Children younger than 18 years with primary ITP who were regular attendee were recruited during the period from June 2015 to September 2016. Their diagnosis was based on presence of bruising and/or petechiae, presence of isolated true thrombocytopenia (platelet count <100×109/l) without any other underlying disease, absence of splenomegaly and lymphadenopathy, and normal or increased megakaryocytes in the bone marrow. Patients with chronic infections, those with known autoimmune or immune deficiency disorders, and those with past history of undiagnosed recurrent bleeding were excluded. Patients were classified according to disease duration into acute ITP (<3 months) and chronic ITP (>12 months) . Ten age-matched and sex-matched healthy children selected from the outpatient clinic were enrolled in this study.
All investigations were performed in accordance with the Ain Shams University, Health and Human Ethical Clearance Committee guidelines for Clinical Research. The local regulatory authority approved the study protocol. Verbal consents were obtained from participants and/or their caregivers before the study.
All patients and controls were subjected to thorough clinical assessment with stress on disease duration, presence of preceding infection, history of vaccination, drug intake, presence of organomegaly or lymph node enlargement, bleeding manifestations, and assessment of bleeding score . Bone marrow examination and other tests, e.g. viral markers such as cytomegalovirus IgM and IgG, were recorded from patient’s files.
Blood samples were collected under complete aseptic procedure for performing complete blood count using Sysmex XT-1800i (Syemex Corporation, Norderstedt, Germany), and 2 ml was allowed to clot naturally and the serum was separated and then frozen at −20°C until used with exclusion of grossly hemolyzed or lipemic specimens. Serum IL-6 and TNF-α were assessed using IL-6 AviBion Human IL-6 ELISA kit(Orgenium Laboratories, Vantaa, Finland) and AssayMax™ Human TNF-alpha ELISA Kit (Assaypro LLC, St. Charles, Missouri, United States), respectively. Serum IgA, IgM, and IgG were detected according to nephlometric method using MININEPH Human kit (IgA code: ZK010.R, IgM code: ZK012.R, IgG code: ZK004.R; The Binding Site Group Ltd, Brimingham, UK), and then normal ranges were stratified according to age .
Statistical analysis was performed using statistical program for social science version 20.0 (SPSS; SPSS Inc., Chicago, Illinois, USA). Quantitative data were expressed as mean±SD. Student’s t-test was used to compare two means. Qualitative data were expressed as frequency and percentage. χ2-test with Fisher’s exact test, if needed, was used to compare proportions between two qualitative parameters. Pearson’s correlation coefficient (r) test was used to detect correlations between parameters. The confidence interval was set to 95%, so the P value was considered significant at the level of less than 0.05.
| Results|| |
There were 13 male and seven female patients, with a mean age of 6.83±4.41 years. Patients with chronic ITP were statistically older, with mean age of 7.7±4.19, than those with acute ITP, with mean age of 3.95±2.21 (χ2=3.83, P=0.001). Other demographic data and clinical manifestations showed no significant difference between the two patient groups. None of our patients received vaccination in the period before the study, whereas six patients with acute ITP and two with chronic ITP had viral infections in that period. All patients with either acute or chronic ITP were presented by cutaneous bleeding, whereas six patients with acute ITP and seven with chronic ITP presented by mucous bleeding. Only one patient with chronic ITP had internal bleeding, and none had intracranial hemorrhage. In the current study, patients with acute ITP were treated according to the standard guidelines: 30% received intravenous Ig (IG), 60% received steroids, and one was kept under observation with no treatment.
According to bleeding score for ITP, patients with acute ITP had scores from 3 to 8 (one patient had a score of 3, two had 4, three of them had 5, one had a score of 6, other one had 7, and two had 8), whereas patients with chronic ITP had scores from 6 to 11 (two of them had score of 6, one had 7, six of them had 8, and one had 11).
Regarding serum Igs as illustrated in [Table 1], in spite that serum IgM levels were found to be within normal ranges in both patient and control groups, mean serum values of IgM were statistically lower in patients with acute ITP than controls. A statistically significant higher frequency of low level of serum IgG was present in acute ITP than controls. Serum IgG mean values were statistically lower in patients with acute ITP compared with patients with chronic ITP and controls (P<0.01 and 0.001, respectively).
|Table 1 Comparison between patients groups and controls regarding laboratory parameters|
Click here to view
Regarding cytokines, statistically higher serum values of TNF-α were detected in patients with acute and chronic ITP compared with controls (P=0.02 and 0.03, respectively). IL-6 levels were statistically higher in patients with acute ITP when compared with those with chronic ITP and with controls (P<0.002 and 0.001, respectively) and a higher significant value in chronic ITP than controls (P<0.001).
Correlation studies were performed to detect statistical correlation between serum levels of Igs, TNF-α, and IL-6 among patients groups. These studies showed no significant correlations between studied parameters except for the negative statistical correlation detected between IL-6 and IgG (r=−0.48; P=0.03).
On correlating the studied parameters with the blood picture of the patients, there was significant positive correlations between IgM and IgG and platelet count (r=0.40 and 0.42 and P=0.03 and 0.02, respectively) and a significant negative correlation between both TNF-α and IL-6 and platelet count (r=−0.47 and −0.7 and P=0.009 and <0.001, respectively).
On follow-up of patients with acute ITP over the study period, 70% of the patients had a complete remission, two were lost to follow-up at hematology clinic, and only one patient had a chronic progressive course. This patient was found to have decreased serum levels of IgA and IgG at presentation.
| Discussion|| |
ITP is a complex heterogeneous disorder involving autoantibodies, cytokines, antigen-presenting cells, costimulatory molecules T-lymphocytes , and auto-reactive B-lymphocytes that produce antiplatelet autoantibodies . We aimed to detect helper T-1 and T-2 response through assessment of TNF-α and IL-6, respectively, and Igs in patients with acute and chronic ITP and to detect their possible roles as predictors of the course of the disease.
In this study, patients with chronic ITP were significantly older than those with acute ITP, and this is most probably because of the chronic nature of the disease. All studied patients presented with cutaneous bleeding, and only one of the patients with chronic ITP developed intracranial hemorrhage owing to head trauma. The bleeding score of the patients with chronic ITP tend to be higher than patients with acute ITP, indicating severity of bleeding as previously described .
IL-6 plays a role in megakaryocytopoiesis and has a potent thrombopoietic action, and also has a role in regulation of the inflammatory response . In the present study, increased serum IL-6 level was significantly associated with ITP suggesting that both autoimmune system and megakaryocyte maturation are stimulated at the same time in the pathogenesis of ITP. Previous studies speculated that elevated IL-6 may be related to compensatory megakaryopoiesis and thrombopoiesis , and became decreased with progression of treatment . In the present study, TNF-α was increased in all patients with acute ITP and in 70% of those with chronic ITP showing a highly statistical significant association compared with controls, and this may highlight its role in the pathogenesis of ITP as previously reported ,.
In the current study, we revealed significant association between decreased serum Igs levels and ITP. The decrease in Igs levels may be explained by rapid clearance of some types of antibody–platelet complexes which may decrease serum antiplatelet antibody titers to below the threshold of detection. Again, antibodies directed against minor or cryptic antigens on platelets or antigens that present primarily on megakaryocytes are difficult to be detected; moreover, there may be a subset of patients with ITP who do not have antiplatelet antibodies . In contrast, a previous study showed high Ig levels with acute ITP and suggested that high levels at disease onset were considered a good prognostic factor ,.
In the present study, correlation analysis showed positive correlation between both serum IgM and serum IgG levels compared with platelet count. This in line with other researcher who  detected lower platelet counts in patients with low IgM without increase in bleeding manifestations and noted that patients with abnormal IgG levels have significantly lower platelet counts than patients with normal IgG levels. We also found a statistically significant negative correlations between both TNF-α and IL-6 when compared with platelet count. This supports the theory that cytokine imbalance plays a role in the pathogenesis of ITP . Furthermore, we found a negative correlation detected between IL-6 and IgG, indicating the involvement of activation of immune system in the pathogenesis of ITP by producing autoantibodies together with polarization of helper T cell with cytokine production .
On follow-up of patients with acute ITP over the study period, only one patient had a chronic progressive course, and this patient was found to have decreased serum levels of IgA and IgG at presentation. This was in line with previous Egyptian study that reported that levels of Igs differ according to type of ITP and that mean IgG levels tend to be low in acute ITP and return to normal with recovery and are persistently low in chronic ITP .
Our study limitation is the small number of patients and the lack of repeated analysis of Igs and cytokines during the course of the disease, which may reveal their influence on the disease course, response to treatment, and the severity of bleeding.
| Conclusion|| |
Children with primary ITP had dysfunctions at cytokine and serum Ig levels.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
McMillan R. The pathogenesis of chronic immune thrombocytopenic purpura. Semin Hematol
Zhong H, Bao W, Li X, Miller A, Seery C, Haq N et al.
CD16+ monocytes control T-cell subset development in immune thrombocytopenia. Blood
Saitoh T, Kasamatsu T, Inoue M, Mitsui T, Koiso H, Yokohama A et al.
Interleukin-10 gene polymorphism reflects the severity of chronic immune thrombocytopenia in Japanese patients. Int J Lab Hematol
Chang WW, Su H, He L, Zhao KF, Wu JL, Xu ZW. Association between transforming growth factor-β1 T869C polymorphism and rheumatoid arthritis: a meta-analysis. Rheumatology
Nugent D, McMillan R, Nichol JL, Slichter SJ. Pathogenesis of chronic immune thrombocytopenia: increased platelet destruction and/or decreased platelet production. Br J Haematol
Guo XH, Zhao F, Shi W, Ma XM, Xu Q, Patiguli AB, Halida YS. Detection and clinical significance of Th1/Th2 cytokines in patients with idiopathic thrombocytopenic purpura. Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi
Rodeghiero F, Stasi R, Gemsheimer T, Michel M, Provan D, Arnold DM et al.
Standardization of terminology, definitions and outcome criteria in immune thrombocytopenic purpura of adults and children: report from an international group. Blood
Rodeghiero F, Michel M, Gernsheimer T, Ruggeri M, Blanchette V, Bussel JB et al.
Standardization of bleeding assessment in immune thrombocytopenia: report from the International Working Group. Blood
Lentner C. Geigy scientific tables. Physical chemistry, composition of blood, hematology, somatometric data
. 8th ed. Publ. Basle, Switzerland: Ciba-Geigy International Medical and Pharmaceutical Information, 1984. 3
Choi PY, Roncolato F, Badoux X, Ramanathan S, Ho S-J, Chong BH. A novel triple therapy for ITP using high dose dexamethasone, low dose rituximab and cyclosporine (TT4). Blood
Blanchette V, Bolton-Maggs P. Childhood immune thrombocytopenic purpura: diagnosis and management. Hematol Oncol Clin North Am
Hunter CA, Jones SA. IL-6 as a keystone cytokine in health and disease. Nat Immunol
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). British Society for Immunology. Clin Exp Immunol
Olcay L, Yenicesu I, Yetgin S. Soluble P-selectin, interleukin 6, and thrombopoietin levels in children with acute and chronic idiopathic thrombocytopenic purpura and their relationship with mega-dose methylprednisolone therapy: a pilot study. J Pediatr Hematol Oncol
Wada H, Minamikawa K, Ohiwa M, Kaneko T, Mori Y, Tamaki S et al.
Elevated plasma interleukin-6 in patient with idiopathic thrombocytopenic purpura. Rinsho Ketsueki
Pesmatzoglou M, Lourou M, Goulielmos GN, Stiakaki E. DNA methyltransferase 3B gene promoter and interleukin-1 receptor antagonist polymorphisms in childhood immune thrombocytopenia. Clin Dev Immunol
Neunert C, Lim W, Crowther M, Cohen A, Solberg L Jr, Crowther MA; American Society of Hematology. The American Society of Hematology 2011 evidence-based practice guideline for immune thrombocytopenia. Blood
Kubota M, Adachi S, Usami I, Okada M, Kitoh T, Shiota M et al.
Characterization of chronic idiopathic thrombocytopenic purpura in Japanese children: a retrospective multi-center study. Int J Hematol
Kubota M, Usami I, Kobayashi K, Tsutsui T, Matsubara K. Serum immunoglobulin levels at onset: association with the prognosis of childhood idiopathic thrombocytopenic purpura. Int J Hematol
Arnason JE, Campigotto F, Neuberg D, Bussel JB. Abnormalities in IgA and IgM are associated with treatment-resistant ITP. Blood
Ogawara H, Handa H, Morita K, Hayakawa M, Kojima J, Amagai H et al.
High Th1/Th2 ratio in patients with chronic idiopathic thrombocytopenic purpura. Eur J Haematol
Khalifa AS, Lusher JM, Cejka J, Zuelzer WW. Immunoglobulins in idiopathic thrombocytopenic purpura in childhood. Acta Haematol
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