Acta Pharmacologica Sinica (2018) 39: 923–929 © 2018 CPS and SIMM All rights reserved 1671-4083/18 www.nature.com/aps Article Yangxin Tongmai Formula ameliorates impaired glucose tolerance in children with Graves’ disease through upregulation of the insulin receptor levels Yan-hong LUO1, Min ZHU1, Dong-gang WANG1, Yu-sheng YANG1, Tao TAN2, Hua ZHU2, Jian-feng HE1, * 1Children’s Hospital Chongqing Medical University, Chongqing 400000, China; 2Department of Surgery, Davis Heart and Lung Research Institute, the Ohio State University Wexner Medical Center, Columbus, OH 43210, USA Abstract Graves’ disease (GD) is the leading cause of hyperthyroidism, and the majority of GD patients eventually develop disorders of glucose handling, which further affects their quality of life. Yangxin Tongmai formula (YTF) is modified from a famous formula of traditional Chinese medicine for the treatment of cardiovascular diseases. In this study we investigated the potential effects of YTF in the treatment of pediatric GD patients with impaired glucose tolerance. Forty pediatric GD patients and 20 healthy children were recruited for this clinical study. Based on the glucose tolerance, the GD patients were divided into two groups: 20 patients displayed impaired glucose tolerance, while the other 20 patients displayed normal glucose tolerance. YTF was orally administered for 60 days. YTF administration significantly ameliorated the abnormal glucose tolerance and insulin sensitivity in the GD patients with impaired glucose tolerance. To determine the molecular mechanisms of this observation, the number of plasma insulin receptors was determined by ELISA. Before treatment, the fasting and postprandial levels of the insulin receptor were significantly lower in patients with impaired glucose tolerance compared with those in patients with normal glucose tolerance and healthy children. After YTF treatment, both the fasting and the postprandial circulating insulin receptor levels were upregulated, and close to those in healthy children. Therefore, YTF is a potential effective treatment to enhance glucose handling in GD children with impaired glucose tolerance. Keywords: Yangxin Tongmai formula; traditional Chinese medicine; Graves’ disease; impaired glucose tolerance; insulin receptor; insulin resistance; Fangjiomics Acta Pharmacologica Sinica (2018) 39: 923–929; doi: 10.1038/aps.2017.94; published online 23 Nov 2017 Introduction display insulin insensitivity and insulin antagonism[4-7], which Graves’ disease (GD) is a common endocrine disorder in pedi- significantly limit the options for treating GMD in Graves’ atric patients and is the leading cause of hyperthyroidism[1, 2]. disease patients. Thus, an effective treatment that ameliorates The incidence of GD is on the rise, and GD affects both the insulin sensitivity and impaired glucose tolerance in Graves’ physical and the mental health of children. Studies by our disease patients is highly desired. group and by others have reported that 44%-60% of patients It is well-known that the China have a long history of treat- with hyperthyroidism develop abnormal glucose tolerance ing diseases that have complex symptoms with Traditional and glucose metabolism disorders (GMD)[3], which further Chinese Medicine (TCM), for which Western medicine treat- compromise the quality of life of Graves’ disease patients, ments usually have limited beneficial effects. Although the especially pediatric patients. The current treatments for ultimate goal of both TCM and Western medicine is to cure abnormal glucose tolerance in Graves’s patients include exer- diseases, there are fundamental and conceptual differences cise, insulin administration and diet control. However, some between TCM and Western medicine. TCM treats the human of the major symptoms of Graves’ disease are fatigue, muscle body and disease as an integrated system, and the crosstalk weakness, and heat intolerance[1]. In addition, patients usually among different organs and the interplay of various symptoms are all taken into consideration when a TCM doctor diagnoses patients[8-11]. In TCM terminology, diabetes is called Xiao-ke, *To whom correspondence should be addressed. which is caused by an improper diet, emotional disturbances E-mail [email protected] and, most critically, Yin deficiency, in which patients exhibit Received 2017-05-08 Accepted 2017-06-17 fatigue, weakness and tiredness. www.nature.com/aps 924 Luo YH et al There has been two thousand years of history in TCM for who were recruited into this study were admitted into Chil- the development of an effective treatment for diabetes. As dren's Hospital Chongqing Medical University from June mentioned above, TCM focuses on the human body as an inte- 2008 to December 2012. The clinical information of patients grated system and treats diseases at their fundamental causes. and normal children is summarized in Table 2. The baseline In this clinical study, we used an established formula, the parameters of all patients and normal individuals are summa- Yangxin Tongmai Formula (YTF), to treat pediatric Graves’ rized in Supplemental Table S1. The diagnostic criteria for GD disease patients who had abnormal glucose tolerance. YTF was based on that of “Zhufutang Pediatric.” All patients had was modified from a famous TCM formula that is used to treat no history of diabetes. Patients who had other symptoms that cardiovascular diseases by Professor Zhaokai Yuan at Hunan induced metabolic disorders or diseases were excluded. Forty University of Traditional Chinese Medicine. The original for- GD patients were divided into 2 groups: a group of 20 patients mula of YTF was developed by a renowned TCM physician, who had impaired glucose tolerance (designated as GD1) Mr Bo-wei QIN, who used it to treat cardiovascular diseases and a group of 20 patients who had normal glucose tolerance based on the TCM theories of nourishing heart Qi (Tiao Yang (designated as GD2). There were initially 34 patients recruited Xin Qi) and reestablishing blood circulation (Huo Tong Xue into the GD1 group, 11 of whom (2 males and 9 females) were Mai). It uses Renshen (Radix Ginseng) combined with Guizhi lost during the study, while another 3 female patients did (Ramulus Cinnamomi) to invigorate the heart Qi and to warm not follow the instruction of taking YTF regularly and were the blood; Danshen (Radix Salviae Miltiorrhizae) to regulate excluded from the study. In addition, 20 healthy children the meridian and to promote blood circulation to remove were recruited into the normal group. blood coagulation; Zhishi (Fructus Aurantii Immaturus) to the break stagnation of qi and to remove food retention, to resolve Preparation and administration of Yangxin Tongmai Formula phlegm and to eliminate mass; and Zexie (Rhizoma Alisma- (YTF) tis) to eliminate phlegma-turbidity by promoting dieresis to The Yangxin Tongmai Formula (YTF) is composed of 5 crude resolve dampness from the lower energizer. All of these ingre- herbal medicines: 10 g of Renshen (Radix Ginseng), 15 g of dients aim to restore Yin , nourish the heart Qi and reestablish Danshen (Radix Salviae Miltiorrhizae), 6 g of Guizhi (Ramulus healthy blood circulation, which we believe is potentially Cinnamomi), 10 g of Zhishi (Fructus Aurantii Immaturus), suitable for the treatment of diabetes based on the theory of and 10 g of Zexie (Rhizoma Alismatis). To ensure the qual- TCM regarding the causes of Xiao-ke (diabetes). In the present ity, accuracy and consistency of the crude herbal components study, we used YTF to treat 20 pediatric GD patients who had in this study, the individual components were handpicked, impaired glucose tolerance for 60 days. Twenty normal chil- checked and weighed by a single board-certified pharmacist dren and twenty GD children with normal glucose tolerance and distributed to each patient. The YTF was prepared the were also recruited as controls. Glucose tolerance, insulin sen- night before its administration. Briefly, a mixture of cut crude sitivity index (ISI), and plasma insulin levels were measured. components (Table 1) was extracted in 50 mL of distilled water We found that plasma glucose, insulin and ISI were signifi- for 30 min at >90 °C. The extracted solution was filtered and cantly increased in GD children with impaired glucose toler- set aside. The extraction protocol was repeated two further ance compared with those in both normal children and GD times with the used crude components and combined with children with normal glucose tolerance. Interestingly, treat- the other extraction solutions to yield a total of 150 mL of ment with YTF significantly decreased the plasma glucose, the extraction solution. Warm YTF (about 50 °C) was orally insulin and ISI levels in GD children with impaired glucose administered three times per day at a dose of 50 mL each time. tolerance to the levels of control children. To test the potential YTF was administered for 60 days. There was an age cutoff mechanism underlying these observations, the plasma insulin criterion: if patients were younger than 7 years old, the dose of receptor levels were quantified by ELISA. We found that the YTF was reduced to 50% of the full dose, and if patients were decreased plasma insulin receptor level of GD children with older than 7 years old, the dose of YTF was the full dose (the impaired glucose tolerance was restored to the normal range same as described above). by the treatment with YTF. Thus, our study suggested that YTF could be an effective means to treat GD children with impaired