Structural magnetic resonance imaging (MRI) studies found some abnormalities of the CSTC circuit with corresponding changes in RRBs. The orbitofrontal cortex (OFC) gray matter volume was positively associated with the severity of RRBs (Hegarty et al., 2020). However, right caudal anterior cingulate U-fiber volume was negatively associated with RRBs (Hau et al., 2019). Interestingly, sex differences in brain structure were associated with RRBs symptoms in autism. The female twin with more severe RRBs had increased thickness of the right intraparietal sulcus and decreased volume of the right orbital gyrus. However, increased volume of the bilateral pallidum was related to more severe RRBs in males (Van’t Westeinde et al., 2020). In addition, structural covariance describes the anatomical association in brain regions, which partially recapitulate networks of synchronized brain activity and is connected with the coordinated rates of developmental change in co-varying regions (Alexander-Bloch et al., 2013). Aberrant structural covariance in subcortical regions, such as thalami and basal ganglia, occurred in children with ASD and predicted the severity of RRBs, which suggested that abnormities of coordinating development of subcortical regions played an essential role in RRBs (Duan et al., 2020). Similarly, Mei et al. (2020) also reported that the structural covariation in brain areas associated with the CSTC circuit was significantly correlated with RRBs in individuals with ASD. A latest preclinical neuroimaging study on RRBs showed that reduced volume in key cortical and basal ganglia regions, including the motor cortex, striatum, globus pallidus, and subthalamic nucleus, was associated with repetitive behaviors in C58/J mice (Wilkes et al., 2020).

There were some new reports of functional connectivity changes of the CSTC circuit in recent years. For example, the over-connectivity pattern primarily in networks involving the fronto-temporal nodes related to RRBs occurred in individuals with ASD (Conti et al., 2017). Ma et al. (2021) showed the increased cortico-striatal intrinsic functional connectivities (iFC) with age in ASD and significant correlations between ADOS-RRB scores and iFC of the dorsal attention network-posterior cingulate cortex/precuneus. Furthermore, Akkermans et al. (2019) showed that increased functional connectivity between the left nucleus accumbens (NAcc) and a cluster in the right premotor cortex/middle frontal gyrus was correlated to more severe RRBs in children with ASD.

Langen et al. suggested the cortico-striatal circuit could be functionally divided into three “macro-circuits.” Each circuit was comprised of discrete, essentially non-overlapping subcortical structures, such as the striatum, globus pallidus, and thalamus, and received multiple inputs from functionally related and interconnected cortexes (Langen et al., 2011b). The CSTC circuit mainly included the sensorimotor circuit (comprising the motor and oculomotor loops), the associative circuit (dorsolateral prefrontal loop), and the limbic circuit (lateral orbitofrontal and anterior cingulate loops) (Groenewegen et al., 2003; Langen et al., 2011a). The abnormities of any circuit could give rise to different RRBs types. Some studies focused on these discrete loops. For example, Abbott et al. reported that the individuals with ASD and high RRBs showed depressed frontoparietal/limbic and motor/limbic circuit ratios. In other words, RRBs seemed to be linked to the imbalance of cortico-striatal connectivity, which showed increased connectivity of limbic circuits, but reduced connectivity of frontoparietal and motor circuits (Abbott et al., 2018). Moreover, complex motor stereotypies (CMS) were rhythmic, repetitive, fixed, and purposeless movements (Oakley et al., 2015). Augustine et al. (2021) found reduced functional connectivity between the prefrontal cortex and striatal regions in children with CMS. However, functional connectivity between motor/premotor cortex and striatal regions was no different from the control group. In a word, these findings offered evidence of the role of discrete loops in RRBs.

How this circuit regulates repetitive behaviors, here we describe the underlying neurobiological mechanisms of RRBs. Based on the neurobiological studies, the CSTC circuit is composed mainly of the direct pathway (cerebral cortex-striatum-internal segment of the globus pallidus/substantia nigra-thalamus-cerebral cortex) and indirect pathway (cerebral cortex-striatum-external segment of the globus pallidus-subthalamic nucleus-internal segment of the globus pallidus/substantia nigra-thalamus-cerebral cortex) (Figure 1) (Kim et al., 2016). There are two major classes of medium spiny neurons (MSNs) in the striatum [i.e., D1R-expressing direct pathway MSNs (dMSNs) and D2R-expressing indirect-pathway MSNs (iMSNs)], and MSNs respectively project to different brain areas (Figure 1). The behavioral result of activation of the direct pathway is motor activation/movements. However, activating the indirect pathway will reduce motor activity and movement (Calabresi et al., 2014). There was abundant evidence of dysfunction of direct and indirect pathways in ASD. Either of dysfunctions gave rise to the imbalance of both pathways (Kim et al., 2016; Gandhi and Lee, 2020), which might underlie RRBs (Lewis and Kim, 2009; Burguière et al., 2015; Monteiro and Feng, 2016).

Recently, some studies reported abnormal activation of the direct pathway in RRBs. In conditional knockout mice, the overactivation of dMSNs caused excessive self-grooming (a pathological repetitive behavior in mice), which suggested the role of direct-pathway deficiency in RRBs (Shonesy et al., 2018). Similarly, optogenetic activation of dMSNs also resulted in sustained and chronic repetitive behaviors (Bouchekioua et al., 2018). Engeln et al. (2021) found chemogenetic inhibition of dMSN can reduce repetitive rotations. In addition, the increased RRBs related to aberrant dMSNs have been found in Neuroligin 1 and Neuroligin-3 mutant mice (Rothwell et al., 2014; Espinosa et al., 2015).

Other studies revealed the indirect pathway role in RRBs. Shank3 deletion preferentially caused synaptic defects in iMSNs in Shank3B-KO mice, which provided direct evidence that a primary dysfunction of indirect pathway brought about the RRBs in ASD mice (Wang et al., 2017). In addition, Brandenburg et al. (2020) reported the increased dopamine type 2 gene expression in the dorsal striatum in postmortem brain tissue from an individual with ASD, which implied the alteration of indirect pathway in ASD. When it comes to the indirect pathway, STN, a vital part of the indirect pathway, has to be mentioned (Tanimura et al., 2011; Wilkes et al., 2020). In C58/J mice, reduced volume in STNs was associated with repetitive behaviors (Wilkes et al., 2020). The pharmacological studies on the indirect pathway also implied the indirect-pathway role in RRBs. For example, sub-chronic drug treatment targeting the indirect pathway reduced repetitive behavior in C58 mice and improved the STN dysfunction (Muehlmann et al., 2020). Similarly, adenosine A2A receptor agonist treatment attenuated increased grooming behaviors in BTBR mice (Amodeo et al., 2018). Selectively enhancing the indirect striatal pathway activation also corrected the RRBs in Shank3B-KO mice (Wang et al., 2017). Moreover, exposure to environmental enrichment retarded the development of stereotypy and recovered the decreased STN activation related to RRBs in high-stereotypy mice (Tanimura et al., 2010). Subsequently, the same team reported that this effect was based on the increased neuronal activation and dendritic spine densities in STN (Bechard et al., 2016). Besides, high-frequency stimulation at STN significantly alleviated RRBs in rodents (Aliane et al., 2012; Chang et al., 2016) and primates (Baup et al., 2008). However, there was a lack of neuroimaging studies that revealed the relation between RRBs and STN in humans. To sum up, the activations of direct and indirect pathways determine behavioral responses, and the imbalance of activations will give rise to RRBs.

The hyperdirect pathway, an integral component of the CSTC circuit, may be involved in the occurrence of RRBs (Figure 1). The STN is considered to receive fast monosynaptic projections from motor areas of the cortex via the hyperdirect pathway (Nambu et al., 2002). This pathway has been verified in humans (Brunenberg et al., 2012; Kelley et al., 2018) and animal models (Haynes and Haber, 2013; Averbeck et al., 2014). Activation of hyperdirect pathway will inhibit ongoing motor movements (Bahuguna et al., 2015). Increasing studies on the role of the hyperdirect pathway in inhibitory control have been reported in humans and animals (Eagle et al., 2008; Rae et al., 2015; Pasquereau and Turner, 2017; Jahfari et al., 2019). Cai et al. (2019) showed that the hyperdirect pathway predicted the inhibitory control in children. As mentioned earlier, inhibitory control may be another manifestation of RRBs. Thus, we suggested the hyperdirect pathway is related to RRBs, although there is a lack of direct evidence for the correlation between hyperdirect pathway and RRBs in ASD.

Moreover, the CSTC circuit is modulated by endogenous neuropeptides, including cannabinoids, opioids, and several other neurotransmitters. Shonesy et al. (2018) found that regulating endocannabinoid signaling in the direct pathway influenced the level of RRBs. Endogenous opioids in the frontal cortex (i.e., the starting site of projections in direct and indirect pathways) were negatively correlated to RRBs (Augustine et al., 2020). In addition, some studies have found abnormalities in the CSTC circuit in genetically mutated mice with particular signal transduction deficits, such as Dlg2 deletion mice (Yoo et al., 2020) and xCT −/− mice (Bentea et al., 2020). These studies gave support to the importance of the CSTC circuit in RRBs.

Risperidone and aripiprazole, the atypical antipsychotic drugs acting on the D2 dopamine receptor, have been approved to reduce irritability, agitation, aggression, and self-harm in ASD by the Food and Drug Administration (FDA) (McCracken et al., 2002; Owen et al., 2009). Although risperidone has been reported to reduce RRBs in salt-induced kinase 1 (SIK1)-mutant mice via attenuating neural excitability and excitatory synaptic transmission (Badawi et al., 2021), a meta-analysis published in 2020 showed that antipsychotics were not beneficial to RRBs in clinical trials (Yu et al., 2020). Subsequently, another meta-analysis including more RCTs pointed out a slight improvement in RRBs after administration of antipsychotics (Zhou et al., 2021). This finding has no practical value because clinicians must weigh these moderate benefits of antipsychotics against the considerable side effects (Correll et al., 2006). Compared with other clinical pharmacological trials of ASD, the studies of antipsychotics have a smaller sample size and more significant heterogeneity in estimated treatment effect.

A great number of studies reported the critical role of oxytocin in human social interaction (Insel et al., 1999; Yatawara et al., 2016). Intranasal administration of the neuropeptide oxytocin (IN-OT) has been regarded as a potential therapy for the core symptoms of ASD. However, its effect in RRB has received less attention. An early study revealed oxytocin infusion reduced RRBs in adults with autism and Asperger syndrome (Hollander et al., 2003). This invasive method is not ideal and replaced by IN-OT for researchers and clinicians (Guastella et al., 2013). After 4 weeks of daily oxytocin administration (24 IU/day), RRBs were significantly reduced in 40 adult men with high-functioning autism (Bernaerts et al., 2020). Interestingly, a preliminary trial showed that 6 weeks of IN-OT had a significant effect on social communication rather than RRBs in 18 men with ASD (Watanabe et al., 2015). However, the same research team found that oxytocin reduced ADOS-RRB score in a larger sample (n = 106), which applied the same study design (Yamasue et al., 2020). This difference may be caused by sampling error and less placebo effect on RRBs related to less expectation for effects. In the same year, another randomized controlled trial (RCT) study also verified the benefits of oxytocin to RRBs (Alaerts et al., 2020). Regarding the underlying neural mechanism of the IN-OT effect, Alaerts et al. (2020) suggested that IN-OT might cause long-term alterations in the internal functional connectivity of the amygdala to the OFC, which was related to RRBs improvement. In addition, Watanabe et al. (2015) reported that the improvement of the core social symptoms in ASD was accompanied by oxytocin-induced enhancement of task-independent resting-state functional connectivity between the anterior cingulate cortex (ACC) and dorsomedial prefrontal cortex. As an increasing number of trials have evaluated the clinical response of multiple doses of IN-OT in ASD, Peled-Avron et al. (2020) conducted a meta-analysis that showed that IN-OT was well tolerated and supported that oxytocin could improve RRBs in ASD, although the effect size was small. However, some studies with small sample sizes have not found the benefits of oxytocin to RRBs (Anagnostou et al., 2012; Dadds et al., 2014; Guastella et al., 2015; Kosaka et al., 2016), it is necessary to conduct multi-center RCT studies with a larger sample and focus on the improvement of RRBs.

Bumetanide is an effective diuretic. As mentioned above, GABAergic signals play a vital role in regulating RRBs (Cellot and Cherubini, 2014). Convincing evidence has shown that defects in inhibitory GABAergic signals led to ASD, and the level of GABAergic inhibition depended on the concentration of intracellular chloride [(Cl-)i] (Schulte et al., 2018). NKCC1 (Na-K-Cl cotransporter 1) is the primary transporter responsible for regulating (Cl-)i, and its activity controls the level of chloride in neurons, which further affects the post-synaptic effect of GABAergic transmission (Schulte et al., 2018). Bumetanide, a selective NKCC1antagonist (Ben-Ari, 2017; Kharod et al., 2019), could restore GABAergic inhibition and weaken behavioral and electrophysiological characteristics in various diseases (e.g., ASD and Fragile X syndrome) by regulating the concentration of neuronal chloride (Payne et al., 2003; He et al., 2014; Kaila et al., 2014; Tyzio et al., 2014; Juarez-Martinez et al., 2021). However, there is a lack of reports on the effects of bumetanide on RRBs in animal models. Increasing studies exhibited a positive effect of bumetanide in children with ASD or Fragile X syndrome (Lemonnier et al., 2013; Du et al., 2015; Zhang et al., 2020; Dai et al., 2021). After a pilot study reported the benefits of bumetanide to RRBs (Lemonnier and Ben-Ari, 2010), Lemonnier et al. conducted two RCT studies to test bumetanide in 60 and 88 patients. Both trials showed a significant reduction in scores of RRBs (Lemonnier et al., 2012; Lemonnier et al., 2017). Similarly, another Phase-2 Superiority Trial also revealed significant effects on RRBs in children aged 7–15, despite no superior effects on the primary outcome of social communication and social interaction (Sprengers et al., 2021). Crutel et al. (2021) described a design of two Phase III studies to evaluate the efficacy/safety of bumetanide oral liquid in ASD, which will provide strong evidence to support the benefits of bumetanide to RRBs. In addition, bumetanide could improve emotional face perception and increase the time spent in spontaneous eye gaze in ASD, with alterations of the activation level in corresponding brain regions (Hadjikhani et al., 2015; Hadjikhani et al., 2018). Current studies were mainly conducted in children and adolescents under 18, and this potential effect in adults should be further verified.

Based on the possible neurobiological mechanism of RRBs, some emerging treatment methods, such as pioglitazone (Capano et al., 2018), pioglitazone (Chugani et al., 2016), intranasal administration of vasopressin (Parker et al., 2019), and IGF-1 injection (Kolevzon et al., 2014), could significantly improve RRBs in individuals with ASD. Some anti-inflammatory drugs targeting abnormal immune functions in ASD showed the benefits to RRBs, such as memantine plus risperidone (Ghaleiha et al., 2013), org 2766 (a synthetic analog of the adrenocorticotrophic hormone) (Buitelaar et al., 1990; Buitelaar et al., 1992), and celecoxib plus risperidone (Asadabadi et al., 2013). In 2020, a meta-analysis of pharmacological interventions for RRBs in ASD included 64 different trials conducted before November 2019. Except for the drugs mentioned above, divalproex sodium, leucovorin, and guanfacine as monotherapies have a more significant positive effect on RRBs of ASD (Zhou et al., 2021). Most findings of the above drugs were based on studies with a small sample and required to clarify their effects further. No pharmacological drug has shown significant clinical benefits and a solid evidence base of effectiveness.

More drug studies are in preclinical stage, and a variety of ASD animal models have become the essential tools for preclinical studies (Lewis et al., 2007), which provides the theoretical basis for following clinical trials. Increasing drug trials in animal models are based on the hypotheses of potential mechanisms in ASD, especially abnormal neurotransmitter/neuromodulator systems. In SHANK3 mutant mice, acute administration of tandospirone, a 5-HT_1A receptor agonist, reduced self-grooming behavior (Dunn et al., 2020). In addition, N-methyl-D-aspartate (NMDA) receptor plays an important role in the balance of E/I and postnatal low-dose MK-801, an NMDA receptor blocker, improved ASD-related behaviors in valproic acid (VPA)-treated rats (Kim et al., 2017; Mohammadi et al., 2020). Another potential function mechanism of the NMDA receptor antagonist was to ameliorate immune dysfunction. For example, dextromethorphan rescued the impaired behavioral patterns in VPA-induced autistic rats and decreased the levels of various oxidative stress and inflammatory markers (Singla et al., 2021).

Another area that receives much attention is the drugs targeting abnormal immune function. 5-aminoisoquinolinone (5-AIQ) has the effects of neuroprotection and down-regulated inflammatory responses (Alhosaini et al., 2021). In BTBR mice, the 5-AIQ treatment significantly prevented self-grooming and marble burying behaviors and ameliorated neuroimmune dysfunctions (Ahmad et al., 2020). Similarly, the benefits to RRBs were reported in the study of the administration of catechin hydrate and pioglitazone in VPA-induced rats (Mirza and Sharma, 2019a; Mehta et al., 2021) and sulforaphane in BTBR mice (Nadeem et al., 2019b). They corrected immune dysfunction and oxidant-antioxidant imbalance in periphery and brain in mice. Beyond that, Zhang et al. (2019) showed that folic acid reduced RRBs in BTBR mice via mitigation of oxidative stress, inflammation, and ferroptosis. Another nutritional supplement of gestational B-vitamin alleviated mitochondrial damage in the hippocampus and PM2.5-induced autism-like behaviors in mice offspring (Wang et al., 2019).

Since a large proportion of people with neurodevelopmental disorders such as ASD are disturbed in their daily sleep/wake cycles (Robinson-Shelton and Malow, 2016). In response to this phenomenon, researchers tried melatonin treatment in CNTNAP2 KO mice and found that it improved excessive grooming in mice (Wang et al., 2020). Furthermore, MTHFR polymorphism was associated with an increased risk of ASD, and the offspring of Mthfr + / − mice (Pu et al., 2013), whether wild-type or heterozygous, exhibited autism-like behaviors. It is surprising that after 14 days of choline supplementation, the characteristics of RRBs were offset (Agam et al., 2020). In addition, other supplementary treatments showed promising effects in ASD animals. For example, abnormally high levels of homocysteine (Hcy) were considered to have a relation with ASD (Kałużna-Czaplińska et al., 2013). Administration of betaine, a methyl group donor in Hcy metabolism, significantly ameliorates RRBs in VPA-induced autistic mice (Huang et al., 2019). Furthermore, exposure to VPA might alter zinc metabolism resulting in a transient deficiency of zinc. Cezar et al. (2018) showed that zinc supplements reduced the transient zinc deficiency and prevented VPA-induced RRBs in rats. Autistic children with similar genetic or metabolic alterations would benefit from similar supplementary treatment if these results are replicated. Other drug studies showed initial outcomes of RRBs in animal models (Bhandari and Kuhad, 2015; Luhach et al., 2021). For example, treatment with medical cannabis alleviated RRBs by over 70% in Shank3 mice (Poleg et al., 2021). Administration of beta-carotene (Avraham et al., 2019; Avraham et al., 2021) and curcumin (Zhong et al., 2020) reduced RRBs in BTBR mice. However, the limitations are that these drugs have multi-target effects and their specific mechanisms are unclear, hindering their use in clinical trials. Moreover, some drugs approved to treat other diseases revealed a new therapeutic effect on RRBs (Román et al., 2021; Ryu et al., 2021; Wu et al., 2021). Chinese herbal medicine also positively influenced RRBs in BTBR mice (Park et al., 2021).

Studies have reported decreased vitamin D levels in the blood of patients with ASD (Wang T. et al., 2016). Vitamin D3 seemed to have therapeutic potential in ASD (Jia et al., 2015). After 3-month vitamin D3 supplementation, RRBs improved in 37 children with ASD, particularly in younger children (Feng et al., 2017). Another study involving 83 children with ASD also found an improvement in RRBs after 3-month treatment (Saad et al., 2016). In contrast, Kerley et al. (2017) conducted an RCT study including 38 children with ASD, which found that vitamin D3 did not affect RRBs after 5-month supplementation.

The abnormal metabolism of folic acid is related to ASD (Castro et al., 2016), and folic acid deficiency have been found in the brains of individuals with ASD (Karin et al., 2017). Supplementing high doses of folic acid in maternal mice can significantly reduce RRBs of offspring (Di et al., 2021). A recent RCT study evaluated the effect of high-dose leucovorin supplementation in ASD. Forty-eight children with ASD and language barriers were randomly given leucovorin or placebo, and RRBs were significantly improved in the leucovorin group (Frye et al., 2018).

Omega-3 fatty acid is associated with mood disorders. Some preliminary studies suggested that Omega-3 fatty acid could effectively treat various mental disorders, such as ASD (Yehuda et al., 2005). Yui conducted a 12-week, 240 mg/day DHA + 240 mg/day arachidonic acid (ARA) intervention on 13 individuals with ASD, and significant improvements in RRBs were observed (Yui et al., 2012). However, the latest meta-analysis, including six studies, suggested no significant effect of omega-3 fatty acids (Zhou et al., 2021).

Gastrointestinal problems and unique gut flora in individuals with ASD are related to the development and severity of ASD symptoms (Hughes et al., 2018). Changing gut flora is considered a promising treatment for related behavioral disorders. Seventeen children with ASD (3–16 years old) were supplemented with Lactobacillus Plantarum WCSF1. After a 12-week intervention, the behavioral score was significantly improved (Parracho et al., 2010). Additionally, some dietary patterns have shown advantages of RRBs in animal models (Castro et al., 2017; Lee et al., 2018a). Gluten-free diets (Ghalichi et al., 2016), casein-free diets (Lucarelli et al., 1995), and ketogenic diets (El-Rashidy et al., 2017) have been verified to have positive effects on ASD symptoms. However, another study reported no significant differences in RRBs (Harris and Card, 2012; Navarro et al., 2015). So, there are still uncertainties about the effects of dietary approaches. Further investigations are needed to confirm these dietary interventions’ specific efficacy and safety for RRBs with a larger sample.

The studies of epigenetics, neuroimmunology, and neurobiology in ASD indicated that cell therapy was an effective approach for treating the core symptoms of ASD (Vaccarino et al., 2011; Siniscalco et al., 2012b; Liu et al., 2019; Zhang et al., 2021). Two outstanding features of stem cells are the intense immunosuppressive activity that allows them to be used in autologous or heterologous transplantation (Siniscalco et al., 2012a) and paracrine actions (Baraniak and McDevitt, 2010; Siniscalco, 2012). Stem cells usually synthesize and release a variety of cytokines, chemokines, and growth factors (Beyth et al., 2005; Siniscalco et al., 2012b), which can reduce the proinflammatory state observed in children with ASD (Gupta et al., 2010) and activate endogenous repair mechanism to recover the damaged function of related cells and tissues (Siniscalco et al., 2012b).

Stem cell therapy has shown benefits to RRBs in various ASD models. Intraventricular administration of mesenchymal stem cells (MSCs) significantly improved core ASD-like symptoms in BTBR mice, including social interaction and RRBs (Segal-Gavish et al., 2016). In addition, intranasal administration of human exosomes derived from mesenchymal stem cells (MSC-exos) was effective on all core ASD behaviors in two different mice (BTBR and SHANK3 KO) (Perets et al., 2018, 2020). Transplantation of mesenchymal stem cells has been proven safe in many clinical trials (Gupta et al., 2010), and whether mesenchymal stem cells apply to individuals with ASD and clinically improve the ASD-like symptoms deserves to be further explored. Regarding clinical trials on cell therapy, Lv et al. conducted a single-center phase I/II trial to assess the safety and efficacy of combined transplantation of human cord blood mononuclear cells (CBMNCs) and umbilical cord-derived mesenchymal stem cells (UCMSCs) in 37 children with ASD (3–12 years of age). Individual transplantation of CBMNCs was demonstrated to remarkably decrease repetitive behaviors compared to the control group. In addition, combined transplantation of CBMNCs and UCMSCs showed better therapeutic effects (Lv et al., 2013). Similarly, Nguyen Thanh et al. (2021) found that transplantation of mononuclear cells of bone marrow combined with educational intervention showed that RRBs and hyperactivity were significantly reduced in children with ASD. These stem cell trials were proved excellent safety, with no safety issues noted during injection and the whole follow-up period (Lv et al., 2013; Nguyen Thanh et al., 2021). Significantly, the benefits of cell therapy to RRBs are necessary to be clarified because most clinical studies in ASD focused on social deficits rather than RRBs or regarded the alterations of RRBs as a secondary result (Bradstreet et al., 2014; Chez et al., 2018; Villarreal-Martínez et al., 2021).

There is still a long way before cell therapy becomes an approved treatment for RRBs in ASD. More in-depth and detailed studies on stem cell biology are necessary to understand the mechanism in RRBs. In addition, the exact dose, time, and site of stem cell infusion, as well as the fatal side effects and long-term safety, need to be further determined. For example, some researchers are concerned about intravenous administration because animal models have shown that it was difficult for transplanted cells to pass through organs (e.g., spleen and kidney) via intravenous administration (Steiner et al., 2012). Moreover, there is a correlation between the dose of transplanted stem cells and the subsequent clinical improvement (Rocha et al., 2002), which emphasized the importance of choosing the exact dose. What should not be ignored is ethical issues in stem cell studies. Such as the acquisition of stem cells, the safety of stem cell collection and administration, the tumorigenicity of stem cells, and other ethical risks similar to other clinical studies (Siniscalco, 2012). The life expectancy of children with ASD is close to normal, and potential risks of children’s medication are difficult to define. Therefore, interventional stem cell therapy is morally untenable unless more studies prove that the apparent benefits outweigh the risks (Yeo-Teh and Tang, 2021).

Transcranial Magnetic Stimulation is a non-invasive brain stimulation used to treat depression and other mental illnesses via changing the excitability of neural circuits and reorganizing the functions of cortex. Previous pilot studies reported positive effects of repeated TMS (rTMS) in individuals with ASD (Sokhadze et al., 2009; Casanova et al., 2012; Wang Y. et al., 2016). Irritability, hyperactivity, and RRBs were decreased in 27 participants with ASD after 18-rounds rTMS on the dorsolateral prefrontal cortex, and the latest study also drew a similar conclusion (Sokhadze et al., 2014; Abujadi et al., 2018). In addition, adults with autism and major depressive disorder reported improvements in repetitive behaviors after 25-session rTMS (Gwynette et al., 2020). A consensus statement of rTMS for ASD showed that rTMS was a potential treatment for ASD and suggested that existing studies have significant limitations, and more definitive studies needed to be conducted to clarify the safety and efficacy of rTMS in ASD (Cole et al., 2019).