This is a prospective, assessor-blinded, randomized, controlled pilot trial with three parallel groups. A total of 90 post-ischemic stroke patients meeting the inclusion criteria will be recruited and then randomly allocated into 3 groups of 30 patients each. All patients will receive 4 weeks of standard care, and the patients assigned to 2 observational groups will receive tDCS and Acu-TENS treatments during a 4-week period (tDCS at 1 mA, Acu-TENS at 20 or 100 Hz) additionally. The stimulation waveform, sites, and duration will be kept the same throughout the study. This study will be performed in conformity to the principles of the Declaration of Helsinki. The “Standard Protocol Items: Recommendations for Interventional Trials” (SPIRIT) will be followed in reporting this protocol and the checklist is provided in Supplementary Material. The study protocol outline is presented as a flow chart in Figure 1, while the study timeline is illustrated in Table 1.

Post-ischemic stroke patients with spastic hemiplegia will be recruited from hospitals in Beijing, China. Posters and flyers will be used to recruit patients. Study information such as the purpose of the study, intervention methods, process and possible adverse reactions will be introduced to the potential participants. Patients will be screened according to inclusion and exclusion criteria by trained researchers. Informed consent will be signed prior to enrollment by patients who meet the requirements.

Eligible participants will be randomly assigned to 3 groups in a ratio of 1:1:1 by an independent statistician using a randomization sequence generated with SPSS 25.0 and will be concealed from the investigators. The random number of patients meeting criteria will be telephoned by the unaffiliated investigator. Due to the characteristics of interventions, participants in the standard care control group will be aware of their allocation ineluctably.

An independent researcher will pre-set the frequency of the Acu-TENS and then cover the frequency display site to ensure the operators remain blinded to the allocation (different frequencies of Acu-TENS) throughout the study. To reduce the risk of bias, the evaluators and statisticians will be blinded to group allocation during outcome assessment and data analysis.

Throughout the trial, all participants will receive standard care in modern medicine for ischemic stroke, corresponding symptomatic or supportive treatment, and regular rehabilitation treatment as well. Simultaneously, Acu-TENS combined with tDCS will be additionally applied to the participants in 2 observational groups. In this study, the acupuncture meridian points used for Acu-TENS and the stimulation locations of International Standard Scalp Acupuncture used for tDCS are referenced from the standards determined by the World Health Organization (38, 39). The locations of the acupoints used in this trial and the methods of electrodes connection are illustrated in Figure 2, while the detailed locations of acupoints are shown in Table 2. All acupoints-based treatment will be conducted independently by the same two acupuncturists, who have at least 5 years of work experience and to be trained specifically in a standard process prior to the study.

The investigators and operators will closely monitor the patients during the 4-week treatment period. If the patient feels any discomfort, the intervention will be stopped immediately, and the patient’s condition will be treated appropriately. Any adverse reaction caused by electrical stimulation, such as localized redness, swelling, pain, or other adverse events, will be recorded by operators.

This study is a small sample clinical trial, and as to be a preliminary for the further large sample clinical trial. Based on a previous study (46), PASS 15.0 software (NCSS Statistical Software, Kaysville, UT, USA) was used to determine the sample size, and by setting the MAS scores decreased in the three groups by 1, 0.75, and 0.45, respectively, with a standard deviation of 0.52, a two-sided α = 0.05 with 90% power, the minimum sample size was 24 cases per group. Considering a 20% dropout rate, 30 patients in each group and a total of 90 patients in three groups were needed.

The data required for the study of all patients will be recorded on the printed case report forms (CRFs), including observation time points, outcome measures, adverse reactions, and safety evaluations. The private information of each participant, including name, phone number and other identity will be protected from disclosure. An independent data manager will check the accuracy and timeliness of the data and keep all data safe. No artificial data modification will be allowed after the end of the trial.

To ensure the high quality of the trial, all investigators and operators will be trained uniformly and professionally before the start of the trial. The training includes understanding the trial protocol and process and clarifying the tasks of each recruitment, treatment and evaluation step. During the implementation of the trial, the outcome data and safety will be reviewed and supervised by experts from the Ethics Committee of the Beijing University of Chinese Medicine.

Statistical analyses will be performed by statisticians who are independent of the research team and have no knowledge of the study groupings or study settings. Statistical analyses will be performed using IBM SPSS Statistics 25.0 software and follow intention-to-treat (ITT) principles to ensure all subjects are included after randomization. Missing data will be handled by multiple imputations. Continuous variables that follow a normal distribution are presented as mean ± standard deviation, and those that do not follow a normal distribution are given as median and interquartile range. Comparisons within groups will be performed via paired t-tests, and difference among groups will be analyzed by the one-way ANOVA test. For measurements at different time points, differences were assessed using repeated-measures ANOVA followed by Bonferroni post hoc tests. The test level is 0.05, with p < 0.05 indicating a statistically significant difference.

Notably, spasticity rarely tends to resolve on its own but progressively worsens over time (50). Spasticity can exacerbate muscle weakness, leading to severe dyskinesia and further exacerbation of daily activities (51). Therefore, adequate and effective control of post-stroke spasticity is the key to break this vicious cycle, thereby fully facilitating motor recovery (48). Compared to medications and surgery, Acu-TENS therapy is a non-pharmaceutical and non-invasive modality that is more acceptable to patients.

The frequency used in an electrical stimulation therapy always plays a crucial role for its therapeutic effect. The frequency of Acu-TENS is worthy of exploring and researching, since it is used in variety, ranging from 2 to 100 Hz. Two Meta-analysis studies have shown that the frequency of TENS for post-ischemic stroke with spastic hemiplegia is mostly at 100 Hz (32, 33). TENS at 20 Hz has also been used in some studies, with positive effects of improving electromyography, relieving spasticity, and increasing walking speed (37, 52, 53). A very few studies compared the anti-spastic effects of TENS or TEAS at different frequencies in stoke patients. Only one study (36) investigated the effects of both 2 and 100 Hz of TEAS on muscle spasticity following brain injury, the results suggested that both low and high frequencies had anti-spastic effects while 100 Hz was better. Taken together, in this study, we will compare the effects of Acu-TENS at 20 and 100 Hz in treating post-ischemic stroke with spastic hemiplegia to explore the optimal Acu-TENS stimulation frequency. The results of this study could provide a reference for setting optimal rated parameters of noninvasive acupoint-based transcutaneous stimulator, which can be manipulated by patients themselves. This modulation may be an alternative option for some chronic diseases such as post-stroke sequelae, thereby promoting proactive rehabilitation of the elderly.

According to the classical theory of acupuncture, a formula of acupoints composed by the points with systemic or local regulation effects usually achieve better clinical results due to their synergistic effects (23, 54). In this study, we will choose Quchi (LI11), Waiguan (TE5), Yanglingquan (GB34), and Shangjuxu (ST37), which are located on the antagonistic muscles of upper and lower limbs respectively, as the stimulation points of Acu-TENS (21). Applying Acu-TENS on those four acupoints can better excite the antagonistic muscles, coordinate and balance the muscle tension of the active and antagonistic muscles, relieve spasticity, promote the transition from co-movement to separation movement, hence establish a normal movement pattern finally. Meanwhile, in terms of Traditional Chinese Medicine (TCM) theory, stimulating those acupoints has the effects of promoting the flow of the energy and balancing yin (medial side of the limbs) and yang (lateral side of the limbs) meridians, leading to functional recovery of the muscles and sinews. Recent study indicated that peripheral stimulation such as acupuncture, might produce both local spinal segmental reflexes and supra-spinal reflexes (55). Yanglingquan (GB34), also known as “confluential point of sinews,” has the effect of relaxing and strengthening sinews, thus it is considered as a classic acupoint for relieving spasticity. A fMRI study illustrated that acupuncture at GB34 may improve the motor function recovery in post-stroke hemiplegic patients via affecting the functional connectivity of the brain (56). It was found that the anti-spastic action of acupuncture at GB34 for post-stroke spasticity was related to its alleviation of spinal hyperreflexia via KCC2-mediated spinal GABA_A activation (57).

After a stroke, the body’s central nervous system immediately starts the stress repair function, which often leads to abnormal neuroplasticity and spasticity simultaneously (58). Although spastic hemiplegia is mainly manifested by contracture of limb muscles and joints, the root cause of spasticity lies in impaired brain function. Therefore, in terms of TCM, “co-morbidity of body and mind” can be considered as the basic clinical characteristic of spastic hemiplegia after stroke, while any modality based on the principle of “co-regulation of body and mind” should has certain clinical significance. Using TENS (34, 35) or tDCS (28, 30) alone has been proven effective for post-stroke spastic hemiplegia. A recent review (59) suggests that the combination of transcranial stimulation and peripheral transcutaneous stimulation techniques may have a synergic effect to optimize neuroplastic changes and improve motor recovery after stroke. Such an effect might be that tDCS depolarizes neuronal membrane potential and modulates GABA, Glutamate levels (60) and BDNF (brain-derived neurotrophic factor) expression levels in localized brain regions (61), whereas Acu-TENS activates KCC2-GABA_A pathway (57). Therefore, this synergic action of TENS combined with tDCS based on acupuncture points on post-stroke spastic hemiplegia needs to investigate further in clinical trials.

In this study, the anode of tDCS is placed on the upper 3/5 of the Anterior Oblique Line of Vertex-Temporal (MS6) of scalp acupuncture on the injured side. The location of MS6 is a projection of the motor cortex on the scalp, whose upper 1/5 corresponds to the lower limbs and trunk, and the middle 2/5 corresponds to the upper limbs. It has a good therapeutic effect on patients with post-stroke motor disorders and the locations of scalp acupuncture can also be applied to other neuromodulation modalities including tDCS (62). In addition, tDCS stimulation in stroke patients typically lasts for 20 min and uses frequencies of either 1 or 2 mA (63). However, research has shown that 2 mA stimulation poses a higher risk for adverse effects compared to 1 mA stimulation (64, 65). This study opted for the safer and more effective option of using a continuous 1 mA stimulation for a duration of 20 min. Thus, the results of this study are expected to provide new evidence of the effectiveness and safety of using both Acu-TENS and tDCS for post-stroke spastic hemiplegia.

To evaluate the efficacy of a treatment modality with more reliable evidence has been a challenge in many clinical trials. Even in some high quality studies with rigorous methodology, lack of objective outcomes is one of the most common limitations (66, 67). In this study, we attempt to observe the effects of combination of Acu-TENS and tDCS on spasticity and motor function in stroke patients from multiple perspectives. Various functional assessments for spasticity and motor function including MAS, Fugl–Meyer Motor Function Scale, MBI, and 10-meter walk test will be used. In addition to subjective measures, we will also detect the iEMG and CR value of SEMG of spastic and antagonistic muscles, as an objective assessment. The SEMG is a non-invasive and painless method for muscle function assessment, which is widely used in rehabilitation researches as an objective and reliable quantitative outcome measure (68, 69). Commonly used signal evaluation indexes of SEMG include frequency and time domain indexes, of which iEMG value is a time domain index. The iEMG value can respond to the total amount of discharges of the skeletal muscles involved in a unit of time, reflecting the contraction characteristics of the muscles in a unit of time, and its value is positively correlated with the muscle strength and muscle tension to a certain extent (70). Normal joint movement requires the coordinated contraction of active and antagonist muscles, and the CR value reflects the proportion of muscle activation of antagonist muscles during active muscle contraction. It has been shown that the co-contraction rate of antagonist muscles is significantly increased in stroke patients compared to healthy controls. SEMG is considered an objective and credible method to assess the muscle tone of spastic and antagonistic muscles. Therefore, we will combine the subjective and objective measures in this study, expecting for providing more reliable evidence of a new modality of Acu-TENS combined with tDCS for stroke rehabilitation.