The effect of steroid treatment and thymectomy on bone age and height development in juvenile myasthenia gravis

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1 DOI /s ORIGINAL ARTICLE The effect of steroid treatment and thymectomy on bone age and height development in juvenile myasthenia gravis Haiyan Wang Zhe Su Chuanming Luo Yan Li Huiyu Feng Wei Fang Chunyan Du Juan Deng Fei Yu Weibin Liu Received: 25 December 2012 / Accepted: 11 April 2013 Ó Springer-Verlag Italia 2013 Abstract Juvenile myasthenia gravis (JMG) is usually associated with growth retardation; however, no data have been published to date in this regard. The goal of this study was to analyze the developmental status of bone age (BA) and height in JMG patients and to subsequently identify the clinical factors, particularly thymectomy, associated with it to guide clinical practice. We conducted a cross-sectional study of 76 JMG patients to examine whether they had abnormalities of height and bone development according to the distribution of the height standard deviation score (Ht SDS) and BA. Correlation analysis and linear regression analysis were conducted to explore the related factors that may affect bone Haiyan Wang and Zhe Su contributed equally to this work. H. Wang Y. Li H. Feng J. Deng F. Yu W. Liu (&) Department of Neurology, The First Affiliated Hospital of Sun Yat-sen University, 58 Zhongshan 2nd Road, Guangzhou , China neurologymg@163.com Z. Su Department of Pediatrics, The First Affiliated Hospital of Sun Yat-sen University, 58 Zhongshan 2nd Road, Guangzhou , China C. Luo Department of Neurology, The Second Clinical College of Guangdong Medical College, Songshan Lake, Dongguan, China W. Fang Department of Biology, Long Island University, 720 Northern Blvd, Brookville, NY 11548, USA C. Du Department of Preventive Medicine, School of Public Health, Sun Yat-sen University, 58 Zhongshan 2nd Road, Guangzhou , China and height development. The mean BA was delayed 0.13 ± 1.42 years compared with the patients chronological age (CA). The Ht SDS was also lower than the healthy peers. In multivariate analysis, the age at onset was negatively associated with delayed BA (R 2 = 0.08, p = 0.019), whereas the cumulative intake of prednisone was negatively associated with Ht CASDS (Ht SDS based on CA) (R 2 = 0.168, p = 0.003). There was no significant association between thymectomy and delayed BA and Ht CA SDS. Delayed BA and growth retardation existed in JMG patients. The age at onset of JMG was a correlating factor for delayed BA and the cumulative prednisone use may be a determinant of height retardation. Thymectomy had no impact on the growth of bone and height. Monitoring BA and height should become routine care in JMG patients to take appropriate therapeutic interventions. Keywords Myasthenia gravis Thymectomy Glucocorticoids Growth Bone age Juvenile Abbreviations MG Myasthenia gravis JMG Juvenile myasthenia gravis OMG Ocular myasthenia gravis GMG Generalized myasthenia gravis BA Bone age CA Chronological age Ht CA SDS Height standard deviations score based on chronological age Ht BA SDS Height standard deviations score based on bone age NF-Ht SDS Near finial height standard deviations score based on adult BMI SDS Body mass index standard deviations score based on chronological age AChR-Ab Acetylcholine receptor antibody

2 Introduction Myasthenia gravis (MG) is an autoimmune disorder that is mostly caused by the binding of the acetylcholine receptor antibody (AChR-Ab) at the neuromuscular junction and is a disease prone to volatility and relapse [1]. A majority of the MG patients in China have juvenile myasthenia gravis (JMG), and more than half of these cases are of the ocular myasthenia gravis (OMG) [2]. The first-line therapy is immunoregulation, and prednisone is also commonly used[3]. Previous studies have suggested that the thymus is the target organ that produces the pathogenic antibody AChR-Ab, and abnormal histopathology of the thymus is a frequent finding in MG patients [4]. Therefore, thymectomy is an important treatment for disease management, as it removes the source of pathogenic antibodies and alters the sensitivity of the cholinesterase inhibitor [5]. Thymectomy is a controversial treatment in children with MG, especially in the youngest patients and the OMG patients [6]. It is believed that the thymus is not only an immune organ but also an endocrine organ that can secrete thymosin, which has a variety of biological functions. Previous animal experiments demonstrated that the damaged secretion of the growth hormoneinathymicmiceledtogrowthstunting[7]. However, whether thymectomy affects the growth and development of JMG patients is not yet known. Given the distribution characteristics of JMG in China, the selection of thymectomy as a treatment for JMG is of great significance. Children with autoimmune disease being administered glucocorticoids frequently suffer from stature retardation. One of the hypotheses with regard to stature retardation is that exogenous glucocorticoids may delay the growth rate of growth plate chondrocytes [8]. JMG patients with height retardation are common, and some of them have delayed bone age (BA), although not all patients present with this phenomenon. Therefore, we conducted a cross-sectional study to explore this clinical issue. To the best of our knowledge, there have been no studies targeting growth and development in JMG patients. So far, the assessment of the BA of the left wrist is regarded as an objective indicator of skeletal maturity and growth status [9, 10]. Moreover, BA is helpful in predicting residual growth potential. Therefore, our study aims to (1) determine the distribution of growth scores in JMG patients; (2) identify clinical features, especially thymectomy, associated with them; and (3) explain the causes that may contribute to the correlation to help to guide clinical practice. Patients and methods Patients We conducted a cross-sectional study of 76 consecutive JMG patients receiving treatment in the neurological outpatient clinic of the First Affiliated Hospital of Sun Yatsen University, China, from July 2011 to August The patients and/or their parents/guardians were provided informed consent to take part in the study. Ethical approval for this study protocol was obtained from the Ethics Committee and Institutional Review Board of the First Affiliated Hospital of Sun Yat-sen University. Among those patients, 52 had received thymectomy, and 71 had received treatment with prednisone. The diagnostic criteria of JMG were as follows: (1) below 18 years of age, (2) typical clinical features of muscle weakness that worsen after repeated activity and improve with rest; and (3) one or more of the following: (a) a definite positive response to anticholinesterase, (b) decrements of 10 % on repetitive nerve stimulation and (c) abnormal jitter or blocking on single fiber electromyography. Excluded criteria were as follows: (1) thymoma; (2) bulbar type MG that affects eating; (3) other autoimmune diseases, such as hyperthyroidism and Crohn s disease; (4) congenital MG and neonatal temporary MG and (5) other diseases that may affect growth and metabolism, e.g., diabetes and congenital heart disease. Methods General clinical data (e.g., gender, age, onset age, duration, age at thymectomy) were collected as comprehensively as possible. The duration of prednisone therapy and the daily dose per unit of weight were recorded. The cumulative intake of prednisone per unit of weight was also recorded. Anti-acetylcholine receptor antibodies (AChR-Ab) were tested in all 76 patients (the upper limit of the normal range in serum is 0.45 nmol/l). The weight and height of the patients were measured by one investigator in the morning. Standing height was measured using an electronic wall-mounted stadiometer (Seca 242, Hamburg, Germany) to the nearest 0.1 cm. Weight was measured on a standard physician s beam scale to the nearest 0.1 kg. Body mass index (BMI) was calculated as weight (kilograms) divided by height (meters) squared. BA was determined by two experienced pediatric endocrinologists applying the TW3 method (Tanner- Whitehouse method edition 3) through the assessment of left wrist radiography, taking the average value [11]. Both endocrinologists were aware of the gender but were blinded to the chronological age of the patients. BA delay was represented by the difference between the chronological age (CA) and BA. Anthropometric measures of each of the JMG patients were expressed as the score of the number of standard deviation score (SDS) compared with the mean for a large

3 normal population of the same age [e.g., height, weight and BMI SDS (Ht SDS, Wt SDS and BMI SDS)]. The Ht SDS was the standardized score in pediatric endocrinology research that reflected the distribution of the individual height among the population. The determination of single patient SDS was as follows: (actual measured value-mean)/standard deviation. The mean and standard deviation of the height, weight and BMI were based on the National Physical Fitness and Health Survey Report [12, 13]. The Ht BA SDS was the Ht SDS, of which the mean and standard deviation were based on BA but not CA. The JMG patients who had caught their near finial height (NF-Ht) need to satisfy: (1) the height velocity was \2 cm/year, calculated over a minimum of 6 months and (2) BA [ 16 years in boys and BA [ 14 years in girls [14]. NF-Ht SDS = (actual height-mean of adult height)/ standard deviation of adult height. Table 1 Clinical characteristics and anthropometric data in JMG patients Variable Total Boys (mean ± SD) Girls (mean ± SD) P Age ± ± ± BA (years) 9.04 ± ± ± Duration (years) 5.26 ± ± ± Age at onset (years) 4.78 ± ± ± Age at thymectomy 8.28 ± ± ± Prednisone duration (month) ± ± ± Cumulative prednisone (mg/kg) ± ± ± BA delay (CA BA) 0.13 ± ± ± Ht CA SDS ± ± ± Ht BA SDS ± ± ± NF-Ht SDS ± ± ± BMI SDS 0.23 ± ± ± BA bone age, CA chronological age, Ht CA SDS height standard deviations score based on chronological age, Ht BA SDS height standard deviations score based on bone age, NF-Ht SDS near finial height standard deviations score based on adult, BWI SDS body mass index Fig. 1 Frequency distribution of JMG patients according to anthropometric indicators. The percentage of patients according to a Ht CA SDS, b Ht BA SDS, c NF-Ht SDS and d BMI SDS is shown in histogram. The Ht CASDS was lower than the Ht BASDS. The patients NF-Ht SDS were lower than 0, but none were \-2 SDS. The distribution of BMI SDS range from -1 to 1 accounted for 72 %. Ht CA SDS height standard deviations score based on chronological age; Ht BA SDS height standard deviations score based on bone age; NF-Ht SDS near finial height standard deviations score based on adult; BWI SDS body mass index standard deviations score based on chronological age

4 The determination of single patient genetic height (genetic-ht) was as follows: the average of the parents heights plus 6.5 cm for boys and minus 6.5 cm for girls [15]; genetic-ht SDS = (genetic-ht-mean of adult height)/standard deviation of adult height. In our study the Ht SDS (e.g., Ht CA SDS, Ht BA SDS and NF-Ht SDS) was all adjusted by genetic-ht SDS (Ht SDS -genetic-ht SDS). Statistical analysis Data were expressed as the mean ± SD. Differences in measurement data were compared with t tests and variance analyses, and the comparison of counted data was performed with Chi square and Rank sum tests. The analysis was conducted using SPSS 17.0 (SPSS, Chicago, IL). Correlation analysis and multiple factors linear regression models were performed to examine the relationship between clinical features and anthropometric parameters with BA and Ht CA SDS. P \ 0.05 was considered statistically significant in this study. Results General clinical information at enrollment Seventy-six JMG patients participated in the study, including 45 girls and 31 boys. The median age was years, and the mean age at onset was 4.78 years. According to the Osserman classification, 62 patients were OMG, and 14 were generalized MG (GMG). The duration of symptoms before enrollment ranged from 0.17 to 13 years, and the mean duration was 5.26 years. The comparison of the age at enrollment, duration, the age at onset, the duration of prednisone, and the cumulative dose of prednisone did not exhibit a statistically significant differences between boys and girls. The mean Ht CA SDS, Ht BA SDS and NF-Ht SDS were all below 0, and the mean BA delay was 0.13 ± 1.42 years. Compared with a large sample of healthy peers, the mean BMI SDS was 0.23 ± 1.07 (Table 1). The BA delay accounted for 50 % patients, and 22.1 % were delayed for more than 1 year. The distribution of Ht CASDS, Ht BA SDS, NF-Ht SDS and BMI SDS was shown in Fig. 1. The clinical characteristics in thymectomy group To explore the effect of thymectomy on the skeletal maturation and height development in JMG patients, we must first understand the distribution of the general clinical data between the two groups. The results showed that the thymectomy group had a longer duration of disease than the non-thymectomy group: the statistical difference was significant (p \ 0.001), and the patients with a duration of \3 years were prone to accept non-thymectomy therapy. There were no significant differences in gender, onset age and clinical classification between the thymectomy group and the non-thymectomy group (Table 2). Fifty-two patients had undergone thymectomy in this study, the distribution of preoperative duration (from onset to accept thymectomy) and postoperative duration (from being taken thymectomy to the date that being enrolled in our study) is shown in Fig. 2. The effect of thymectomy and other clinical features on BA and height In order to understand the effect of prednisone therapy and the duration and age at onset, as well as the age at Table 2 The distribution of clinical features in thymectomy and non-thymectomy groups v 2 v 2 test, OMG ocular myasthenia gravis, GMG generalized myasthenia gravis * Significance was set at P \ 0.05 Variable Thymectomy Non-thymectomy N v 2 P Sex Female 18/31 13/31 31 Male 34/45 11/45 45 Age at onset /29 12/ /22 7/22 22 [5 20/25 5/25 25 Clinical classification OMG 40/62 22/62 62 GMG 12/14 2/14 14 Duration (years) \0.001* \3 10/29 19/ /20 4/20 20 [7 26/27 1/27 27

5 Fig. 2 Frequency distribution of preoperative and postoperative duration of JMG patients in thymectomy group. Postoperative duration was from onset to accept thymectomy, and postoperative duration was from being taken thymectomy to the date that being enrolled in our study. a The distribution of preoperative duration in thymectomy group, and the majority percentage accounted for 50 % in 2 5group. b The distribution of postoperative duration in thymectomy group, and the duration longer than 1 year accounted for 71 % Table 3 Correlation analysis of BA delay, Ht CA SDS and Ht BA SDS with clinical features Item BA delay Ht CA SDS Ht BA SDS R p R p R p Age at onset * Duration Age at thymectomy * Accumulative prednisone * * BA bone age, Ht CA SDS height standard deviations score based on chronological age, Ht BA SDS height standard deviations score based on bone age, R correlation coefficient * Significance was set at P \ 0.05 thymectomy, on the patients bone and height development, we conducted a bivariate correlation analysis. It showed that the delayed BA was correlated with the cumulative intake of prednisone per unit of weight and the age at onset. The Ht CA SDS had a correlation with the cumulative intake of prednisone, as well as the age at thymectomy. However, the Ht BA SDS had no correlation with the above clinical indicators (Table 3). Next, we set BA delay as the dependent variable and the age at onset, thymectomy (yes or no) and the cumulative intake of prednisone as the independent variables; we similarly set Ht CA SDS as a dependent variable and the age at thymectomy, thymectomy (yes or no) and the cumulative intake of prednisone as independent variables to conduct the multiple regression analysis. The age at onset and the cumulative prednisone were each entered into the regression model (Table 4). Discussion Our data showed that delayed BA and height retardation existed in JMG patients, which were consistent with clinical findings. The majority of HT CA SDS and Ht BA SDS were below 0, and Ht CA SDS was even lower. The distribution of general clinical features exhibited no significant difference between genders. Our study suggests that delayed BA and height retardation are related to past cumulative prednisone intake and that onset age might be a factor in BA delay. However, neither the duration nor thymectomy was risk factors for growth stunting. Glucocorticoid is a cornerstone of MG therapy, exerting a broad immunosuppressive effect [16], and it is also widely used in pediatric patients. The association between glucocorticoid exposure and BA delay, as well as height retardation found in our study, has been found in many immune diseases, such as Crohn s disease, asthma and Cushing s syndrome [17 19]. Glucocorticoid may decrease the proliferation rate of the growth plate chondrocytes and thereby delay the fusion of the epiphyses [20]. In addition, the reduction in growth hormone secretion and the barriers to skeletal maturity caused by exogenous glucocorticoid may affect stature [8]. Moreover, the somatomedin inhibitors that are induced by steroids could cause a drop in somatomedin. Additionally, the mitosis of chondrocytes and collagen synthesis both contribute to impaired growth [21, 22]. Correlation analysis shows that delayed BA has a positive correlation with the onset age, which means the

6 Table 4 Multi-factor linear regression models with BA delay and Ht CA SDS as dependent variable Dependent variable Independent variables b t p 95 % CI R 2 BA delay Age at onset * * Cumulative prednisone Thymectomy (yes/no) * Ht CA SDS Age at thymectomy Cumulative prednisone * * Thymectomy (yes/no) * BA bone age, CI confidence interval, Ht CA SDS height standard deviations score based on chronological age * Significance was set at P \ 0.05 younger the age at onset, the greater the BA delay. Younger patients with an earlier onset of disease were more sensitive to the side-effects of prednisone, as they usually had a longer disease course and longer exposure to the cumulative effects of prednisone, so skeletal maturation was more easily affected. Moreover, the long-term presence of inflammatory mediators may affect skeletal development in autoimmune diseases. The patients who were younger at disease onset suffered from muscle weakness that may have caused a lack of exercise, leading to delayed skeletal growth. Though our study did not focus on nutritional status, the majority of patients exhibited a moderate BMI SDS compared to healthy peers, suggesting that their nutritional state was normal. One study concerning asthma showed that short stature was associated with an early onset (before the age of 3 years) of the disease, which was consistent with our data [23]. A recent study confirmed that inhaled glucocorticoid in lower doses appeared to lead to impaired final adult height. Although the height decrease was not progressive or cumulative, the deficit may persist into adulthood [18]. It is still unclear how the long-term effect of glucocorticoid therapy affects the attainment of adult height of JMG patients. We analyzed the near final height of 11 older patients: this analysis suggested that the near final height was impaired, and all of the NF- Ht SDS were below 0. However, the mean value of NF- Ht SDS (-0.95 ± 0.46) was higher than Ht CA SDS (-1.27 ± 1.04), as catch-up growth presumably played a role. None of the NF- Ht SDS was \-2SDS (\-2SDS could satisfy the diagnosis of short stature syndrome), this suggested that adult height might not be significantly limited. Future studies should conduct an in-depth analysis. Even in the state of chronic inflammation, parental height was a powerful determinant of linear growth [17]. We therefore adjusted the Ht SDS by one s genetic height to minimize the effect of parental height. Ht BA SDS is the most accurate indicator for height growth because it excludes the impact of delayed BA on height. Our data showed the mean Ht BA SDS was also below 0, slightly larger than Ht CA SDS, but it had no correlation with clinical indicators. Our results demonstrate that thymectomy has no significant impact on BA and height development. Although previous animal experiments demonstrated that the thymus participates in growth and development [24], the deficiency in serum thymulin during early life might compromise body growth [25]. However, because the mechanism for growth retardation related to serum thymulin has not yet been confirmed in human studies, it may be that thymulin is not a key factor in human growth and development, or it may play a role earlier in life. In the present study, the patients were all beyond 3 years old at thymectomy, so their bone and height development may not have been affected. The timing of thymectomy is debated in many circumstances in JMG therapy. It has been thought that thymectomy may affect children s growth and development, as well as immune function [26]. Delayed surgery may require large doses of prednisone to control symptoms, so children are at risk for steroid side effects, including growth failure, susceptibility to severe infection, and delayed vaccinations [27]. However, more recent studies have demonstrated that thymectomy has no effect on mature T cell function and immune function [28, 29]. A large number of clinical studies showed that thymectomy easily induced remission [30 32]. Opting for thymectomy at early onset (with in 1 year) may be more effective than delayed surgery, as early thymectomy induced remission much more easily [5, 33]. Therefore, we believe that thymectomy has no significant effect on growth and development. Instead, the appropriate timing of surgery should be decisively chosen to promote recovery. Glucocorticoids should be used at the

7 lowest effective dose, and a steroid-sparing drug (e.g., azathioprine) should be started simultaneously with glucocorticoids, with the goal of reducing the duration of steroid treatment. In summary, growth retardation and delayed BA occur frequently in JMG patients and are related to exposure to glucocorticoids. Onset at an early age may be a factor in delayed BA. Thymectomy did not affect either the development of bone or height. Therefore, steroid-sparing drugs should be started as soon as possible to reduce glucocorticoid-related side effects. The timing of appropriate thymectomy should be chosen decisively to promote recovery. Determination of BA and height should become routine care in JMG patients to help optimize therapeutic interventions to both reduce delay in skeletal maturation and contribute to optimal catch-up growth. Acknowledgments The study was supported by grants from the National Natural Science Foundation of China ( , ), the Clinical study of 5010 plan of Sun Yat-sen University ( ), and the Postdoctoral Initial Funding of Guangdong province (s ). The work is attributed to the Department of Neurology, the First Affiliated Hospital, Sun Yat-sen University, Guangdong Key Laboratory for Diagnosis and Treatment of Major Neurological Diseases, National Key Clinical Department, National Key Discipline, Guangzhou , China. References 1. Meriggioli MN, Sanders DB (2009) Autoimmune myasthenia gravis: emerging clinical and biological heterogeneity. Lancet Neurol 8: Huang X, Liu WB, Men LN, Feng HY, Li Y, Luo CM, Qiu L (2012) Clinical features of myasthenia gravis in southern China: a retrospective review of 2,154 cases over 22 years. Neurol Sci 3. 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