|Year : 2018 | Volume
| Issue : 1 | Page : 69-75
Serum interleukin 23 and its associations with interstitial lung disease and clinical manifestations of scleroderma
Gamal A Hammad1, Refaat M Eltanawy1, Rasha M Fawzy1, Tahany M.A Gouda2, Mona A Eltohamy1
1 Department of Rheumatology, Rehabilitation and Physical Medicine, Faculty of Medicine, Benha University, Benha, Egypt
2 Chest Diseases Department, Faculty of Medicine, Benha University, Benha, Egypt
|Date of Submission||09-Apr-2017|
|Date of Acceptance||24-May-2017|
|Date of Web Publication||12-Jan-2018|
Tahany M.A Gouda
Chest Diseases Department, Faculty of Medicine, Benha University, Benha
Source of Support: None, Conflict of Interest: None
Introduction Systemic sclerosis (SSc) is a complex disease linked to immune system activation, vascular damage, associated with increased synthesis, and deposition of extracellular matrix, which contain excessive amounts of structurally normal collagen. Interleukin 23 (IL-23) might play a role in disease development and severity. This study aimed to assess the relationship between serum level of IL-23 and interstitial lung disease in SSc.
Patients and methods Thirty patients with SSc together with 30 age-matched and sex-matched healthy volunteers were recruited in this study. Serum IL-23 levels were measured by enzyme-linked immunosorbent assay. Functionally, lung involvement was assessed by pulmonary function tests and radiologically by chest radiography and high-resolution computed tomography of the lungs.
Results Mean serum IL-23 level was significantly highly elevated in SSc patients compared with healthy controls (P<0.005). Patients with elevated IL-23 levels exhibited shorter disease duration (P<0.05). Moreover, mean serum IL-23 level was elevated in diffuse SSc cases compared with limited SSc cases and in cases with pulmonary fibrosis (P<0.05), although they were not associated with other clinical features. Elevated mean serum IL-23 level was significantly higher in mild restrictive cases compared with moderate and severe restrictive cases. As regards high-resolution computed tomography, mean serum IL-23 level was statistically highly significantly elevated in cases with ground-glass appearance (P<0.001) compared with others.
Conclusion Alterations in serum concentrations of IL-23 support the hypothesis that IL-23 is associated with induction of SSc generally and SSc associated with interstitial lung disease specifically. Presumably, blockage of IL-23 could be used as a potential therapeutic target in early SSc.
Keywords: high-resolution computed tomography, interstitial lung disease, pulmonary function tests, systemic sclerosis
|How to cite this article:|
Hammad GA, Eltanawy RM, Fawzy RM, Gouda TM, Eltohamy MA. Serum interleukin 23 and its associations with interstitial lung disease and clinical manifestations of scleroderma. Egypt J Bronchol 2018;12:69-75
|How to cite this URL:|
Hammad GA, Eltanawy RM, Fawzy RM, Gouda TM, Eltohamy MA. Serum interleukin 23 and its associations with interstitial lung disease and clinical manifestations of scleroderma. Egypt J Bronchol [serial online] 2018 [cited 2018 Feb 20];12:69-75. Available from: http://www.ejbronchology.eg.net/text.asp?2018/12/1/69/223068
| Introduction|| |
Systemic sclerosis (SSc) is a heterogeneous chronic, autoimmune, multisystem connective tissue disorder characterized by vasculopathy, inflammation, and progressive fibrosis of the skin and internal organs . Interstitial lung disease (ILD) is a major common complication along with pulmonary arterial hypertension, which is the leading cause of morbidity and mortality in scleroderma patients . The diagnosis of ILD hinges on careful clinical evaluation together with pulmonary function tests (PFTs) and high-resolution computed tomography (HRCT) .
Interleukin 23 (IL-23) is a heterodynamic cytokine belonging to the IL-6/IL-12 family, which can have both proinflammatory and anti-inflammatory effects on the development of autoimmune pathology. This family mediates distinct roles and cellular functions, coordinating regulation of Th1 development and type 1 cell-mediated immunity .
The pathogenesis of the disease is still obscure, but strong evidence suggests an immune dysregulation in the development of SSc. CD8+ T cells can be found dominantly in lungs, and CD4+ T cells have cytokine profiles characterizing Th2, such as IL-4, IL-10, IL-13, IL-17, and IL-23 .
Another T-cell subtype is Th17, which is different from Th1 and Th2 that are involved in the development of many autoimmune diseases. IL-23 activates Th17 cells with subsequent secretion of IL-17 . The IL-23–IL-17 axis is a central player in the development of chronic inflammation and in the host defense against extracellular pathogens .
| Aim|| |
This study aimed to assess the relationship between serum level of IL-23 and ILD in SSc.
| Patients and methods|| |
In all, 30 SSc patients, diagnosed according to the American Colleague of Rheumatology (1980) criteria , together with thirty age-matched and sex-matched apparently healthy volunteers, were enrolled in this study as a control group. All patients gave their formal consent, The protocol was approved the Ethical committee of the faculty.
These patients were selected from those attending the inpatient and the outpatient clinics of the Rheumatology and Rehabilitation and Chest Departments of Benha University Hospitals between June 2014 and January 2016.
Patients who fulfilled the American Colleague of Rheumatology (1980) criteria were included in this study .
Exclusion criteria were as follows: patients who were smokers, known to be diabetics, those with a history of acute or chronic liver or kidney diseases, those who had a recent history of infection or other inflammatory or autoimmune diseases, those receiving cytotoxic or other immunosuppressive therapy, and patients with other causes of ILD.
All SSc patients were subjected to the following. Complete patient history was taken through clinical examination with complete respiratory system examination. Skin score was measured using the technique of the modified Rodnan total skin thickness score . Laboratory investigations included complete blood count, erythrocyte sedimentation rate, and C-reactive protein; antinuclear antibody (ANA) was determined by indirect immunofluorescence; antiscleroderma 70 (anti-Scl 70) antibody and anticentromere antibody were detected by enzyme-linked immunosorbent assay (ELISA); and serum IL-23 was detected by human IL-23 ELISA kits (R&D Systems, Minneapolis, Minnesota, USA) . About 2 ml of venous blood was collected by sterile venipuncture, allowed to clot, and sera were separated and kept frozen at − 20°C until they were used for the detection of serum IL-23 level. All samples were analyzed using the average of the optical density values to calculate concentrations. The minimum detection limit (cutoff) was calculated as the average value of the blanks plus 2 SD. The reaction is terminated by the addition of acid, and absorbance is measured at 450 nm.
Lung involvement was assessed functionally by PFT spirometry, forced expiratory volume in the 1 s (FEV1%), forced vital capacity% (FVC%), and FEV1/FVC. PFTs were done using sensor medics Spirolab III (Rome, Italy) according to American thoracic society, radiologically by chest radiograph posteroanterior view, and HRCT of both lungs as explained by Goh et al. . The computed tomography scans were performed with a scanner (Prospeed S Scaner, GE YMS Ltd, Tokyo, Japan) at 10-mm intervals from the apex of the lung to the diaphragm using 1–2 mm collimator with the HRCT technique. All patients underwent scanning in the supine position at full inspiration. No intravenous contrast agent was used. Images were obtained at a window width of 750 HU and a window level of −800 HU. Echocardiography was done to evaluate pulmonary artery pressure and to assess whether there was secondary pulmonary hypertension.
Data were tabulated and statistically analyzed using statistical package for the social science (SPSS, version 16; SPSS INC., Chicago, Illinois, USA) .
Qualitative data were represented as frequencies and relative percentages. χ2-test was used to calculate the difference between qualitative variables.
Quantitative data were expressed as mean±SD. Independent t test was used to calculate the difference between quantitative variables in two groups. Mann–Whitney test was used to calculate the difference between quantitative variables in two groups in non-normally distributed data.
Analysis of variance F-test test was used to calculate the difference between quantitative variables in more than two groups. Kruskal–Wallis K test was used to calculate the difference between quantitative variables in more than two groups in non-normally distributed data. Pearson’s correlation coefficient used to calculate the correlation between quantitative variables.
The significance level for all the above-mentioned statistical tests was calculated. The threshold of significance is fixed at 5% level (P-value): P value greater than0.05 indicates nonsignificant results, P value less than 0.05 indicates significant results, and P value less than 0.001 indicates highly significant results.
| Results|| |
This study included 30 SSc patients and 30 age-matched and sex-matched apparently healthy controls. Nine (30%) SSc patients had limited SSc (LSSc) and 21 (70%) had diffuse SSc (dSSc). Characteristics of the studied groups are shown in [Table 1]. There were no statistically significant differences between cases and controls as regards age and sex distribution (P>0.05). As regards mean serum IL-23 levels, there was a highly statistically significant difference − being higher in SSc patients. All differential leukocytic counts were within normal range, and no eosinophilia was detected.
[Table 2] showed the relation between mean serum IL-23 level and the studied variables in SSc patients.
|Table 2 Comparison between mean serum interleukin-23 levels and the studied variables in systemic sclerosis patients|
Click here to view
There was NO statistically significant difference as regards patients sex, statistically highly significant difference being higher in dSSc patients compared to LSSc patients (P=0.005). Mean serum IL-23 level was higher in cases with shorter disease duration (≤5 years) (P=0.02), with a statistically significant difference compared with patients with longer disease duration.
About 100% of SSc patients had cutaneous manifestations in the form of Raynaud’s disease, hypopigmentation and hyperpigmentations, digital ulcers, calcinosis, and telangiectasia; 23.3% had arthritis; 80% had gastroesophageal reflux; 80% had cough; 76.7% had dyspnea; 16.7% had pulmonary hypertension; and 6.7% had sicca manifestations, which classically combine dry eyes and dry mouth.
Mean serum IL-23 level was significantly higher in SSc patients with cardiopulmonary manifestation in the form of cough, dyspnea, and chest pain (P<0.001), with no statistically significant difference as regards PHT, skin manifestations, musculoskeletal manifestations, gastrointestinal manifestations, and sicca symptoms (dryness of the eyes and mouth) (P>0.05).
In this study, mean serum IL-23 levels were significantly higher in ANA and anti-Scl-70 positive patients as compared with others (P<0.05).
As regards radiological findings of SSc patients, there was a statistically significant difference (P=0.01) as regards mean serum IL-23 level in patients with pulmonary fibrosis by chest radiography, and there was a highly statistically significant difference in cases with reticular and ground-glass appearance (P<0.001).
[Table 3] shows PFTs of the two studied groups.
FVC% and FEV1% were significantly higher in the control group than in cases, with no statistically significant difference regarding FVC%/FEV1%.
[Figure 1] shows that SSc cases with interstitial fibrosis pattern in chest radiography have significantly higher mean serum levels of IL-23 than cases with normal chest radiography, and by HRCT; cases with ground-glass pattern and reticular pattern have significantly higher mean serum IL-23 levels than cases with normal, reticulonodular, and honeycomb patterns.
|Figure 1 Relation between mean serum interleukin 23 (IL-23) levels and radiological finding of the systemic sclerosis patients.|
Click here to view
[Figure 2] shows that mean Serum IL-23 levels were significantly higher in mild restrictive cases compared with moderate and severe cases, and also higher in moderate restrictive cases compared with severe restrictive cases.
|Figure 2 Relation between mean serum interleukin 23 (IL-23) levels and grading of pulmonary function tests of the systemic sclerosis patients.|
Click here to view
| Discussion|| |
SSc is a chronic, multisystem connective tissue disease of unknown etiology. IL-23 is a member of the fascinating family of cytokines, which can have both proinflammatory and anti-inflammatory effects on the development of autoimmune pathology .
This work showed that there was a highly statistically significant difference as regards mean serum IL-23 level in the studied groups, being higher in SSc patients. These results were in agreement with those of Murata et al. , who reported higher concentration of IL-23 in SSc than in healthy controls; as IL-23 production appears to be of great importance in the inflammatory reaction of SSc, it causes the activation of Th17 cell, producing IL-17, a proinflammatory cytokine that induces the production of other proinflammatory cytokines. In contrast, Mathian et al.  and Olewicz et al.  found decreased serum levels of IL-17 and IL-23 in SSc patients compared with healthy patients. These different results can result from differences in the treatment or differences in disease duration in the studied group or may be related to small sample size of SSc.
Moreover, Agarwal et al.  reported that the gene encoding for IL-23 receptor has been identified as a susceptibility gene for SSc development, and IL-23R polymorphisms are associated with antitopoisomerase-I positivity and lower frequency of pulmonary hypertension, which further support the increased mean serum IL-23 level in SSc patients.
In this study, mean serum IL-23 level was found to be statistically significantly elevated in dSSc patients compared with LSSc patients (P=0.005). In contrast to our results, Komura et al.  and Olewicz et al.  showed no statistically significant difference between mean serum IL-23 levels among dSSc and LSSc groups.
In this study, there was a statistically significant difference in cases with shorter disease duration (≤5 years) (P=0.02) than cases with longer disease duration. In agreement with our results, Komura et al.  and Olewicz et al.  found that SSc patients with disease duration less than 2 years had significantly elevated IL-23 level compared with those with duration of 2–5 years (P<0.05) and those with duration more than 5 years (P<0.01). This may be explained by decreased activity of Th17 cells along with disease progression and immunosuppressive drugs . Thus, the level of IL-17 was noted to be higher early in SSc patients, with subsequent progressive reduction of IL-17 levels along with disease duration. Moreover, IL-23 is known as a survival and proliferative factor for Th17 cells . Some reports recommend that not all Th17 cells are pathogenic, and exposure to antigen-presenting-cell-derived IL-23 is crucial for their ability to stimulate autoimmunity and inflammation .
In this study, mean serum IL-23 level was significantly elevated in SSc patients with cardiopulmonary manifestation in the form of cough, dyspnea, and chest pain) (P<0.001), with no statistically significant difference as regards PHT and other clinical manifestations (P>0.05). Similarly, Komura et al.  have described that elevated mean IL-23 level was related to the presence of pulmonary fibrosis, but not to other clinical features of SSc. This may be related to an increased number of Th17 cells with enhanced skin-homing and lung-homing properties in SSc individuals . In contrast, Radstake et al.  and Demir et al.  showed no statistically significant difference between mean serum IL-23 level and any clinical features of SSc.
In this study, as regards immunological profile, mean serum IL-23 level was significantly elevated in ANA-positive and anti-Scl-70-positive patients compared with others (P<0.05). Similarly, Gourh et al.  reported that IL-23 levels were more elevated in the anti-Scl-70 antibody and ANA-positive group. On the other hand, Demir et al.  found that there was no correlation between serum IL-23 levels and ANA, anti-Scl-70, and anti-centromere antibody positivity.
In this study, SSc cases with interstitial fibrosis pattern by chest radiography had significantly higher mean serum levels of IL-23 than cases with normal chest radiography. By HRCT, cases with mixed reticular and ground-glass patterns compared with reticulonodular and honeycomb patterns have significantly higher mean serum IL-23 levels than group of normal. By PFT, mean serum IL-23 levels were significantly higher in mild restrictive cases than in moderate and severe cases.
Komura et al.  and Olewicz et al.  found a significant correlation between serum IL-17and IL-23 concentration and the extent of lung damage in SSc patients as evaluated with HRCT. In contrast to these results, there was no difference between the frequency of Th17 bronchoalveolar lavage T cells in SSc patients with interstitial lung fibrosis, SSc patients without ILD, and healthy controls .
The study by Kurasawa et al.  revealed that IL-17 messenger RNA was expressed in unstimulated lymphocytes from the peripheral blood and lymphocytes from the skin and lungs of SSc patients, but not from patients with systemic lupus erythematosus or polymyositis/dermatomyositis or from healthy donors. IL-17 levels were also increased in the serum of SSc patients, but not in that of systemic lupus erythematosus patients or healthy donors. IL-17 overproduction was significantly related to the early stage of SSc, but not to other clinical features of SSc. Moreover, IL-17 enhanced the proliferation of fibroblasts and induced the expression of adhesion molecules and IL-1 production in endothelial cells in vitro and concluded that IL-17 is overproduced by T cells from the peripheral blood and fibrotic lesions of the skin and lungs in SSc patients. These results suggest that IL-17 overproduction has an important role in the pathogenesis of SSc, especially in the early stages of the disease, by inducing the proliferation of fibroblasts and the production of IL-1 and the expression of adhesion molecules on endothelial cells.
| Conclusion|| |
Alterations in serum concentrations of IL-23 support the hypothesis that IL-23 is associated with induction of SSc generally and SSc associated with ILD
Specifically and that blockage of IL-23 could be used as a potential therapeutic target in early SSc.
The authors thank all participants in this study, without whom the study would not have been completed.
Financial support and sponsorship
Conflicts of interest
There are no conflict of interest.
| References|| |
Awasthi A, Kuchroo VK. Th17 cells: from precursors to players in inflammation and infection. Int Immunol
Jain S, Anupama S, Chris TD. Interstitial lung disease in systemic sclerosis: pathophysiology, current and new advances in therapy. Inflamm Allergy Drug Targets
Sara RS, Flavia VC. Interstitial lung disease in scleroderma. Rheum Dis Clin North Am
Smith J, Colbert R. The interleukin-23/interleukin-17 axis in spondyloarthritis pathogenesis. Arthritis Rheumatol
Zhao XF, Pan HF, Yuan H, Zhang WH, Li XP, Wang GC et al.
Increased serum interleukin 17 in patients with systemic lupus erythematosus. Mol Biol Rep
Kastelein RA, Hunter CA, Cua DJ. Discovery and biology of IL-23 and IL-27: related but functionally distinct regulators of inflammation. Annu Rev Immunol
Dalila AS, Mohd Said MS, Shaharir SS, Asrul AW, Low SF, Shamsul AS et al.
Interleukin-23 and its correlation with disease activity, joint damage, and functional disability in rheumatoid arthritis. Kaohsiung J Med Sci
Masi AT, Rodnan GP, Medsger TA. Preliminary criteria for the classification of systemic sclerosis (scleroderma). Arthritis Rheum
Clements P, Lachenbruch P, Seibold J, White B, Weiner S, Martin R et al.
Inter- and intraobserver variability of total skin thickness score (modified Rodan TSS) in systemic sclerosis. J Rheumatol
Magyari L, Varszegi D, Kovesdi E, Sarlos P, Farago B, Javorhazy A et al.
Interleukins and interleukin receptors in rheumatoid arthritis: research, diagnosis and clinical implications. World J Orthop
; 2014; 5
Goh NS, Desai SR, Veeraraghavan S, Hansell DM, Copley SJ, Maher TM et al.
Interstitial lung disease in systemic sclerosis: a simple staging system. Am J Respir Crit Care Med
Greenberg RS, Daniels RS, Flanders D, Eley JW, Boring JR, editors. Diagnostic testings. In: Medical epidemiology
. 3rd ed. New York: McGraw-Hill; 1996. pp. 77–89.
Murata M, Fujimoto M, Matsushita T, Hamaguchi Y, Hasegawa M, Takehara K et al.
Clinical association of serum interleukin-17 levels in systemic sclerosis: is systemic sclerosis a Th17 disease? J Dermatol Sci
Mathian A, Parizot C, Dorgham K, Trad S, Arnaud L, Larsen M et al.
Activated and resting regulatory T cell exhaustion concurs with high levels of interleukin-22 expression in systemic sclerosis lesions. Ann Rheum Dis
Olewicz GA, Danczak PA, Kuznar KB, Pawel H. Interleukin-17 and interleukin-23: importance in the pathogenesis of lung impairment in patients with systemic sclerosis. Int J Rheum Dis
Agarwal SK, Gourh P, Shete S, Paz G, Divecha D, Reveille JD et al.
Association of interleukin 23 receptor polymorphisms with anti-topoisomerase-I positivity and pulmonary hypertension in systemic sclerosis. J Rheumatol
Komura K, Fujimoto M, Hasegawa M, Ogawa F, Hara T, Muroi E et al.
Increased serum interleukin 23 in patients with systemic sclerosis. J Rheumatol
Aggarwal S, Ghilardi N, Xie MH, de Sauvage FJ, Gurney AL. Interleukin-23 promotes a distinct CD4 T cell activation state characterized by the production of interleukin-17. J Biol Chem
Croxford AL, Mair F, Becher B. IL-23: one cytokine in control of autoimmunity. Eur J Immunol
Truchetet ME, Allanore Y, Montanari E, Chizzolini C, Brembilla NC. Prostaglandin I(2) analogues enhance already exuberant Th17 cell responses in systemic sclerosis. Ann Rheum Dis
Radstake TJ, Lenny van Bon, Jasper B, Anila H, Roger H, Dirk MW et al.
The pronounced Th17 profile in systemic sclerosis (SSc) together with intracellular expression of TGFb and IFNc distinguishes SSc phenotypes. PLoS One
Demir C, Sahin A, Kucuksahin O, Turgay M, Turkcapar N, Erten S, Kınıklı G. Sclerosis comparison of serum IL-23 and IL-17 levels in patients with systemic sclerosis and healthy subjects, sclerosis. J Clin Analytic Med
:83–87. This study was presented as a poster presentation on 5th Turkish-Greek Rheumatology Days, October 14–16, 2011.
Gourh P, Arnett F, Assassi S, Filemon KT, Mei H, Laura D et al.
Plasma cytokine profiles in systemic sclerosis: associations with autoantibody subsets and clinical manifestations. Arthritis Res Ther
Meloni F, Solari N, Cavagna L, Morosini M, Montecucco CM, Fietta AM. Frequency of Th1, Th2 and Th17 producing T lymphocytes in bronchoalveolar lavage of patients with systemic sclerosis. Clin Exp Rheumatol
Kurasawa K, Hirose K, Sano H, Endo H, Shinkai H, Nawata Y et al.
Increased interleukin-17 production in patients with systemic sclerosis. Arthritis Rheum
[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3]