Abstract
Differentiated thyroid carcinoma (DTC) is the most common endocrine malignancy. Surgical removal with radioactive iodine therapy is recommended for recurrent thyroid carcinoma, and the postsurgical thyroid removal is critical. This study evaluated the clinical values of radiofrequency ablation (RFA) in the postsurgical thyroid removal for DTC. 35 DTC patients who had been treated by subtotal thyroidectomy received RFA for postsurgical thyroid removal. Before and two weeks after RFA, the thyroid was examined by ultrasonography and 99mTcO4 - thyroid imaging, and the serum levels of free triiodothyronine (FT3), free thyroxin (FT4), thyroid stimulating hormone (TSH) and thyroglobulin (Tg) were detected. The efficacy and complications of RFA were evaluated. Results showed that, the postsurgical thyroid removal by RFA was successfully performed in 35 patients, with no significant complication. After RFA, the average largest diameter and volume were significantly decreased in 35 patients (P > 0.05), and no obvious contrast media was observed in ablation area in the majority of patients. After RFA, the serum FT3, FT4 and Tg levels were markedly decreased (P < 0.05), and TSH level was significantly increased (P < 0.05). After RFA, radioiodine concentration in the ablation area was significantly reduced in the majority of patients. The reduction rate of thyroid update was 0.69±0.20%. DTC staging and interval between surgery and RFA had negative correlation (Pearson coefficient = -0.543; P = 0.001), with no obvious correlation among others influential factors. RFA is an effective and safe method for postsurgical thyroid removal of DTC.
Keywords: Radiofrequency ablation, ultrasonography, differentiated thyroid carcinoma, removal
Introduction
Differentiated thyroid carcinoma (DTC) is the most common endocrine malignancy, and has good prognosis after surgery [1,2]. However, the frequent metastasis in local cervical lymph nodes and remote organs are usually found. The follow-up period of 11.3 years shows that, the overall recurrence and mortality rates for DTC are 20.5% and 8.5% respectively [3]. It is necessary to find the effective way to treat recurrent thyroid carcinoma, for improving the survival rate and life quality of patients.
Surgical removal with radioactive iodine therapy is recommended for recurrent thyroid carcinoma [3]. Postsurgical thyroid removal is critical for the radioactive iodine therapy. For the repeated surgery, the neck dissection is difficult for surgeons due to distortion in the normal tissue plane, postsurgical fibrosis and severe adhesion, and is associated with the complications such as nerve injury, hypoparathyroidism and cosmetic concerns rated to re-incision [4]. For radioactive iodine therapy, the efficacy depends on many influential factors such as time of iodine ablation, surgical method, residual thyroid tissue, thyroid stimulating hormone (TSH) level, metastatic status, and so on. The remission rate of postsurgical thyroid removal by radioactive iodine therapy is not satisfactory, and there are many complications such as excessive iodine intake in thyroid tissue and high risk of radioiodine concentration in the local area [5]. As the first radioactive iodine therapy for patients with total thyroidectomy, the remission rate is just 64.5% [6]. For post-surgical DTC patients without far metastasis, the remission rate can reach 84.6%, but if the lymphatic metastasis and far metastasis occurs, the rate changes to 45.5% and 27.3%, respectively [7]. Radiofrequency ablation (RFA) is usually used in patients at high surgery risk and in those who refuse to undergo repeated surgery [8]. Recently, several studies report that ethanol injection with RFA can be the effective and valuable treatment option for original thyroid carcinoma and recurrent disease [9-11]. However, application of RFA to thyroid removal of after surgery in DTC patients has seldom been reported. In this study, the efficacy and safety of RFA in postsurgical thyroid removal of DTC was evaluated. The objective was to more formally define the role of RFA in treatment of DTC.
Patients and methods
Patient recruitment
Thirty-five DTC patients (15 males and 20 females; aged 27-78 years, with mean age of 43.49±13.01 years) who received ultrasonography-guided RFA for postsurgical thyroid removal from January 2014 to May 2015 were enrolled in this study. All patients had been treated by subtotal thyroidectomy, and the interval between surgery and RFA was 1-3 months. The histologic type confirmed by postsurgical pathology was papillary thyroid carcinoma. The size of thyroid carcinomas was 7-46 mm (average 28.71±10.32 mm).
According to the results from US and 99mTcO4 - thyroid nuclear medical imaging, there was no obvious nodule in thyroid after surgery. 25 patients needed unilateral removal of residual thyroid tissue (left, 11 cases; right, 14 cases), and 10 patients needed bilateral removal. In 35 patients, the maximum unilateral diameter and volume of residual thyroid tissue were 15-39 mm (average 25.14±5.87 mm), and 0.9-4.4 mL (average 2.38±0.86 mL), respectively. The TSH level of patients was < 30 mU/L. This study was approved by the Institutional Review Board in XXXXXX Hospital. Informed consent was obtained from each patient.
Radiofrequency ablation
Under guidance with Logiq E9 ultrasound machine with 6-16 MHz linear probe (GE Healthcare Inc., WS, USA), the RFA was performed in an outpatient operating room by 2 ultrasound intervention doctors with 5 years of RFA experience. Radiofrequency generator (VRS01, STARmed Inc., Gyeonggi-do, Korea) and thyroid-dedicated, internally cooled radiofrequency electrode (STARmed Inc., Gyeonggi-do, Korea) were used for ablation. A 7-cm long, 18-gauge electrode with a 0.5- or 1.0-cm active tip was chosen according to postsurgical thyroid size. The radiofrequency power was 20-45 w, and the radiofrequency work time was 1.0-4.5 min. The patient blood pressure, blood O2 saturation, heart rate and electrocardiography were monitored during the RFA procedure.
The patient was in supine position, with pillow under shoulder and head back to expose the neck. After iodophor disinfection and sterile towel, lidocaine (2%, up to 10 mL; Hebei Tiancheng Pharmaceutical Co., Ltd., Shijiazhuang, China) we carefully injected to the skin, needle track and anterior thyroid capsule for local anesthesia. Several important methods were performed to prevent the unnecessary compliances during RFA. Firstly, US-guidance was used to trace the electrode tip. Secondly, totally 20-30 mL of physiological saline (North China Pharmaceutical Co., Ltd., Shijiazhuang, China) was injected to front edge of residual thyroid tissue, inner carotid sheath and back of thyroid gland to create a protective barrier to radiofrequency energy (i.e., the hydrodissection technique) [10,12].
The electrode tip was initially positioned in the deepest, most remote portion of the lesion, and then the thyroid tissue removal was executed gradually and anteriorly with the moving short technique [8]. Ablation was started with low power (5 w for 0.5-cm active tip or 20 w for 1.0-cm active tip, respectively), and the radiofrequency power was gradually and carefully increased (up to 10 w in 0.5-cm active tip and 45 w in 1.0-cm active tip, respectively) [3]. During the ablation, the patient voice and pain were checked frequently. If the patient could not tolerate pain, the radiofrequency power was reduced or turned off immediately. Ablation was terminated when the residual thyroid tissue changed to a transient hyperechoic zone or the hoarseness developed [8,12]. Finally, following the electrode trace, the ablation was completed, and the pinprick on the skin was packed with woundplaster. After RFA, all patients were observed for 1-2 h in the resting room, and the discomfort or complications associated with RFA were observed. All patients stopped taking thyroid hormone tablets and continued to take low-iodine diet for two weeks before and after RFA.
Ultrasonography
Logiq E9 ultrasound machine with 6-16 MHz linear probe was used for pre- and post-RFA routine ultrasound and contrast-enhanced ultrasound (CEUS) examination, and guidance during the ablation. The size of residual thyroid tissue, internal echo and color flow was observed and recorded by routine ultrasound. During CEUS, the contrast media SonoVue (Bracco Corp., Milan, Italy) was used, which consisted of inert gas (SF6) with lecithin. The microbubble suspension of SonoVue with physiological saline was prepared. The contrast mode on the machine was activated, and the mechanical index was set as 0.06. After shaking for 20 s, 2.4 mL of SonoVue suspension was quickly injected through the elbow vein, followed by continued quick injection of 5 mL physiological saline. 3-min contrast dynamic pictures were observed and stored for analysis.
Follow-up and assessment of RFA efficacy
After RFA, any discomfort or complication associated with treatment was observed. Patients were followed up two weeks after RFA. The size, internal echo and color flow of residual thyroid tissue were observed by routine ultrasonography. The contrast media performance of residual thyroid tissue was examined by CEUS. No obvious contrast media in the area of postsurgical thyroid was regarded as the diagnosis standard of successful postsurgical thyroid removal by RFA [13]. If there was still residual thyroid tissue, the additional RFA was performed, until till no obvious contrast media was found. The 99mTcO4 thyroid imaging was performed, and radioiodine concentration in the residual thyroid tissue was significantly reduced that was regarded as the diagnosis standard of successful RFA removal. The reduction rate of thyroid uptake was assessed, and the final conclusion for further radioactive iodine treatment was determined by two radiologists with 5 years of diagnosis experience. In addition, the serum levels of free triiodothyronine (FT3), free thyroxin (FT4) and TSH were detected using radioimmunoassay, and thyroglobulin (Tg) was detected using double-antibody radioimmunoassay. All procedures were performed according to the instructions of kit manufactures.
Statistical analysis
All statistical analysis was carried out using SPSS 17.0 software (SPSS Inc., Chicago, IL, USA). The measurement data were presented as mean ± SD. Wilcoxon signed-rank test was used for comparison of maximum diameter and volume of residual thyroid tissues, serum FT3, FT4, TSH and Tg level before RFA and two weeks after RFA. P < 0.05 was considered as statistically significant. According to previous study on iodine therapy [6], some significant influential factors such as DTC staging, pre-RFA thyroid volume, interval between surgery and RFA, post-RFA TSH level and post-RFA Tg level were selected for Pearson’s correlation coefficient analysis. Scatter plots were obtained to indicate the correlations among these variables.
Results
General information of patients
Total 35 DTC patients received RFA for postsurgical thyroid removal. The patient and thyroid carcinoma characteristics were summarized in Table 1. The DTC staging included T2, T3 and T4 or M1 (distant metastasis) and N1 (lateral neck lymphatic metastasis, without central lymphatic metastasis).
Table 1.
Patients and thyroid carcinoma characteristics according to DTC stage
| Stage | T2 | T3 | T4 | M1 | N1 |
|---|---|---|---|---|---|
| n (%) | 5 (14.3%) | 7 (20.0%) | 7 (20.0%) | 8 (22.9%) | 8 (22.9%) |
| Age, year | 55.8±18.16 | 44.71±10.94 | 47.71±14.8 | 41.0±7.43 | 33.5±6.3 |
| Male | 3 | 1 | 4 | 6 | 1 |
| DTC size, mm | 32.6±3.51 | 43.4±2.3 | 27.1±7.76 | 24.1±5.57 | 19.4±8.35 |
DTC, differentiated thyroid carcinoma.
Overall treatment outcome
All 35 patients successfully received the post-surgical thyroid removal by RFA, including 25 cases of unilateral postsurgical thyroid removal (left, 11 cases; right 14 cases) and 10 cases of bilateral postsurgical thyroid removal for in patients. The mean time of hospitalization for all patients was 1.2 days.
Complications
There was no significant complication for the RFA procedure. Two patients experienced voice change immediately after RFA and recovered soon. No any voice change and nerve complication was observed in other patients. A burning sensation, pain, or both came from 10 patients, but the symptoms were relieved by reducing RF power or stopping RFA procedure for several seconds.
Ultrasonography examination results
Routine ultrasonography showed that, before RFA, there were low-level echo and relatively rich blood flow signal in thyroid tissue (Figure 1A, 1B). After RFA, the high-level internal echo with light spots in the ablation area was observed, without obvious color flow inside (Figure 1C, 1D).
Figure 1.
Routine ultrasonography on thyroid before and after RFA. Before RFA, the size of residual thyroid tissue was 32.2 mm*9.2 mm*10.7 mm (A), and rich blood flow was observed in thyroid tissue (B); After RFA, the size of residual thyroid tissue was 30.0 mm*10.0 mm*10.5 mm (C), and there was no obvious blood flow in thyroid tissue (D).
CEUS showed that, before RFA, there were irregular enhancement in the whole residual thyroid tissue (Figure 2A); After RFA, no obvious contrast media was observed in the ablation area in 34 patients (Figure 2B), with contrast media filling in 1 patient. The second RFA was executed for this patient, and the active ablation time was 45 s. The CEUS was performed again, until no obvious contrast media was observed.
Figure 2.
CEUS on thyroid before and after RFA. Before RFA, there was irregular enhancement in the residual thyroid tissue (A); After RFA, no obvious contrast media was observed in the ablation area (B).
Changes of maximum diameter/volume of residual thyroid tissue and serum biochemical indexes
The unilateral maximum diameter and volume of residual thyroid tissue and FT3, FT4, TSH and Tg levels were detected before RFA and two weeks after RFA. Results were shown in Table 2. Compared with before RFA, the maximum diameter and volume of residual thyroid tissue after RFA decreased (from 25.14±5.87 mm to 24.73±5.61 mm and from 2.38±0.86 mm3 to 2.33±0.71 mm3, respectively), but the difference was not significant (P > 0.05). Compared with before RFA, the serum FT3, FT4 and Tg level after RFA significantly decreased (from 3.19±0.32 mm to 2.82±0.63, from 1.03±0.2 mm to 0.93±0.26 mm, from 44.45±14.30 mm to 27.79±23.65 mm, respectively) (P < 0.05), and the TSH level significantly increased (from 18.78±6.28 mm to 33.37±14.66 mm) (P < 0.05).
Table 2.
Maximum thyroid diameter/volume and serum biochemical indexes before and after RFA
| Diameter mm | Volume ml | FT3 pg/mL | FT4 ng/dL | TSH U/L | Tg ng/mL | |
|---|---|---|---|---|---|---|
| Before | 25.14±5.87 | 2.38±0.86 | 3.19±0.32 | 1.03±0.20 | 18.78±6.28 | 44.45±14.30 |
| After | 24.73±5.61 | 2.33±0.71 | 2.82±0.63 | 0.93±0.26 | 33.37±14.66 | 27.79±23.65 |
| P | 0.064 | 0.080 | 0.003 | 0.011 | 0.000 | 0.001 |
RFA, radiofrequency ablation; FT3, free triiodothyronine; FT4, free thyroxin; TSH, thyroid stimulating hormone; Tg, thyroglobulin.
Changes of radioiodine concentration in thyroid
99mTcO4 - thyroid nuclear medical imaging showed that, before RFA, there was obvious radioiodine concentration in thyroid, which indicated the residual thyroid tissue. After RFA, radioiodine concentration in the ablation area was significantly reduced, the reduction rate of thyroid update was 0.69±0.20% (Figure 3). Only for 2 patients, the reduction rate of thyroid uptake was 0.4%, others were more than 0.5%. In addition, the Tg levels of these 2 patients were 2.31 and 1.76 ng/mL, respectively. Therefore, two radiologists gave the final conclusion that all 35 patients could receive the radioactive iodine treatment.
Figure 3.
99mTcO4 - thyroid nuclear medical imaging on thyroid before and after RFA. Before RFA, there was obvious radioiodine concentration in thyroid (A); Two weeks after RFA, only a little radioiodine concentration was observed (B).
Correlations among DTC staging, pre-RFA thyroid volume, interval between surgery and serum biochemical indexes
Results of Pearson’s correlation coefficient analysis showed that, among the influential factors including DTC staging, pre-RFA thyroid volume, interval between surgery and RFA, post-RFA TSH level and post-RFA Tg level, the DTC staging and interval between surgery and RFA had negative correlation (Pearson coefficient = -0.543; P = 0.001), and no obvious correlation among others factors. Scatter plots were used to indicate the correlations between these variables, and the results were shown in Figures 4, 5.
Figure 4.
Scatter plots for DTC staging, thyroid volume, interval between surgery and RFA, post-RFA TSH and post-RFA Tg. RFA, radiofrequency ablation; Tg, thyroglobulin; TSH, thyroid stimulating hormone; DTC, differentiated thyroid carcinoma.
Figure 5.
Scatter plots for DTC staging and interval between surgery and RFA. RFA, radiofrequency ablation; DTC, differentiated thyroid carcinoma.
Discussion
Thyroid carcinoma is the most common endocrine malignant tumor. The prevalence of thyroid nodules is 18.6%, and 5-15% thyroid nodules are malignant, of which 90% is DTC. The incidence of DTC continues to increase in these 30 years [5,14]. The surgical resection of tumor is often used for DTC, and there are two ways for removal of residual thyroid tissue after surgery. One is secondary surgical removal, and the other is radioactive iodine treatment [15,16]. There are many significant complications for repeated surgical therapy, including nerve injury, hypoparathyroidism and cosmetic concerns rated to re-incision [4]. Although the radioactive iodine thyroid removal is the most common treatment method in clinic, but the efficacy depends on many influential factors. This treatment is also associated with many complications [5]. Therefore, it is necessary to seek better methods for postsurgical thyroid removal of DTC. In this study, RFA is used for postsurgical thyroid removal of DTC. All 35 patients have successfully received RFA, and obtained satisfactory efficacy. After RFA, the maximum diameter and volume of residual thyroid tissue are decreased slightly. The serum FT3, FT4 and Tg level significantly drop down in the majority of patients, and the TSH level markedly increases. These are the intuitionistic results of the efficacy of RFA.
In this study, RFA was performed in the patients who require radioactive iodine treatment after thyroid surgery. The inclusion standard for radioactive iodine treatment is recommended as TSH ≥ 30 mU/L, and the lower TSH level will decrease the treatment efficacy [17]. In this study the TSH level in all patients is under 30 mU/L, so these patients cannot acquire satisfied efficacy by directly radioactive iodine treatment. After RFA, the TSH level increases to above 30 mU/L, so the subsequent radioactive iodine treatment can be conducted. During all the procedures, one patient receives additional RFA, because of the border residual thyroid tissue around the ablation area. The additional RFA depends on the size of residual thyroid tissue. For this patient, there is a spot of residual thyroid tissue, with maximum diameter of 39 mm, so second RFA is performed. According to our experience, RFA is just a treatment before radioactive iodine treatment. Some thyroid tissue adjacent to the trachea and laryngeal nerve should be retained, for ensuring the safety of RFA.
Results of Pearson’s correlation coefficient analysis show that, DTC staging and interval between surgery and RFA have negative correlation. The reason may be that, for the patients with T4 staging or far lymph node metastasis, the interval time between surgery and RFA should be shortened, and the removal of residual thyroid should be executed as soon as possible, based on the condition of the patients. Just as we mention that the aim of RFA is to inactive the residual normal thyroid tissue, there is no obvious correlation among pre-RFA thyroid volume, post-RFA TSH level and post-RFA Tg level.
From previous report [18], except no obvious radioiodine concentration by nuclear medical imaging in the whole body, Tg < 2.0 ng/mL is another diagnosis standard of successful removal of thyroid tissue under radioactive iodine treatment. In this study, Tg < 2.0 ng/ml is not considered as the referred standard indicator. After RFA, the Tg level in 35 patients is 27.79±23.65 ng/mL. There are two reasons. Firstly, the treatment is for inactivated normal residual thyroid tissue, but not the metastasis of DTC in the whole body, which has the function of Tg secretion. This is also the key point of subsequent radioactive iodine therapy. Secondly, some thyroid tissues adjacent to the trachea and laryngeal nerve are retained, which affects the Tg level. Also for the 2 patients that the reduction rate of thyroid uptake was 0.4%, two radiologists considered the Tg levels that were 2.31 and 1.76 ng/mL the final conclusion was determined that all patients could receive the radioactive iodine treatment. RFA is a new method for postsurgical thyroid removal, the diagnostic standard for Tg level and nuclear medicine imaging also need be explored in the further study in the future.
In this study, there are 2 patients (5.7%, 2/35) with voice change. The complication rate (5.7%, 2/35) is somewhat superior to the results reported in previous studies [8,11], in which are RFA is applied to treating thyroid cancer or local recurrence of DTC. A multicenter study of Korean Society of Thyroid Radiology (KSThR) reports that the voice change occurs in 1% patients after RFA [15]. The relatively higher complication rate in this study is probably due to the superabundant thyroid tissue ablation. In addition, the maximum diameter and volume of residual thyroid tissue are related to voice change, and the conservation treatment or RF power may also be the reason of voice change. In this study, the two patients just have immediate voice change, and both of them recover soon. The physiological salin is used to create a protective barrier as prevention procedure, which may be the good way to avoid voice change and decrease complication.
Until now we have not inquired any related literature for RFA treated post-surgical thyroid removal. As Lee et al. [3] report that, FRA can be effective in treating loco-regional, recurrent, and DTC in patients with high surgical risk. After 30 months of follow-up, the maximum diameter and volume of recurrent DTC were decreased by 93.2% and 96.4%, respectively. According to more previous RFA studies (Table 3), after FRA the volume reduction rate of thyroid is from 50.9% to 93%, with complete disappearance rate from 25% to 50%. All of them are the evidence to ensure that RFA is effective and safe for thyroid carcinoma. According to previous study [8] for RFA to treat local recurrence of DTC, a mean volume reduction is 96.4%, which is extremely higher than this study. The postsurgical thyroid removal is the treatment of normal thyroid tissue, and the residual thyroid tissue is just inactivated, which is different from RFA treatment for thyroid carcinoma or local tumor recurrence.
Table 3.
Clinical findings and outcomes of RFA for recurrent DTC (literatures)
| Authors | Number of Tumor | Follow-up range/month | VRR/% | CDR/% | Tg | |
|---|---|---|---|---|---|---|
|
| ||||||
| Patient number | Reduction Rate/% | |||||
| Dupuy et al. [19] | 11 (8 patients) | 6-26 (10.3) | NA | 25 | 6 | 92.4 |
| Monchik et al. [9] | 24 (16 patients) | 10-68 (40.7) | NA | 50 | 11 | 95 |
| Baek et al. [10] | 12 (10 patients) | 16-31 (23.6) | 93 | 50 | 7 | 73 |
| Park et al. [20] | 16 (11 patients) | 1-14 (6) | 50.9 | NA | 9 | 58.4 |
RFA, radiofrequency ablation; DTC, differentiated thyroid carcinoma; VRR, volume reduction rate; CDR, complete disappearance rate; Tg, thyroglobulin.
The literatures on the efficacy of radioactive iodine ablation are reviewed, and the detailed information is shown in Table 4. The low- and high-dose radioactive iodine groups are separated for meta analysis [26]. The success rates of treatment in low- and high-dose groups are 63%-93% and 74%-95%, respectively. RFA thyroid removal is a local ablation, and is based on the diagnosis criteria from the results of CEUS and 99mTcO4 - thyroid imaging. In this study, the removal efficacy was satisfied for all 35 patients and all of them could receive the further radioactive iodine treatment. However, for radioactive iodine thyroid removal, the success rate usually is based on the whole body scanning result and Tg level. Although RFA cannot be simply compared with radioactive iodine ablation, it can be concluded that RFA is effective for postsurgical thyroid removal.
Table 4.
Meta analysis results of radioactive iodine thyroid removal
| Authors | Time | Low-dose group | SR | High-dose group | SR | Weight factor % | ||
|---|---|---|---|---|---|---|---|---|
|
|
|
|||||||
| Successful cases | Patients number | Successful cases | Patients number | |||||
| Johansen et al. [21] | 1991 | 21 | 36 | 58.3% | 14 | 27 | 51.9% | 4.7 |
| Bal et al. [22] | 1996 | 17 | 27 | 63.0% | 28 | 38 | 73.7% | 6.9 |
| Maenpaa et al. [23] | 2008 | 49 | 77 | 63.6% | 55 | 72 | 76.4% | 16.8 |
| Mallick et al. [24] | 2012 | 198 | 214 | 92.5% | 197 | 207 | 95.2% | 59.3 |
| Caglar et al. [25] | 2012 | 36 | 47 | 76.6% | 42 | 48 | 87.5% | 12.3 |
Low-dose group, 1110 MBq; High -dose group, 3700 MBq; SR, success rate.
This study also has some limitations. Although 35 patients are included in the study, the sample size is relatively small. In addition, it is the primary study for RFA to treat postsurgical thyroid removal, and the research methods are not very mature. There are some points which should be improved for the study. Firstly, a relatively large sample size is needed for patient selection, adaptation disease indication and complication. Secondly, considering the characteristics of thyroid nuclear medical imaging and Tg level, longer follow-up period is also needed. Thirdly, the efficacy of subsequent radioactive iodine treatment should be observed for comprehensive analysis. Finally, the standardized procedures for RFA thyroid removal should be established. Therefore, further large-size, long-term, prospective studies are required to provide advanced evidence for the efficacy of RFA for postsurgical thyroid removal.
In conclusion, RFA is the effective and safe method for postsurgical thyroid removal of DTC. This study has provided a basis for further application of RFA to treatment of thyroid carcinoma.
Acknowledgements
This study was supported by Zhejiang Provincial Natural Science Foundation of China (No. LY15H180001).
Disclosure of conflict of interest
None.
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