Experimental research of integrative Chinese medicine in multiple sclerosis

Ke-Ying Zhu,Zheng Kang,Yan Chen,Gen-Cheng Wu,Jun Wang*

1Department of Integrative Medicine and Neurobiology,Fudan University,Shanghai,China.

Introduction

Of all the neurodegenerative diseases being studied,Alzheimer’s disease(AD)and Amyotrophic lateral sclerosis(ALS)have been already highly valued,whereas Multiple sclerosis(MS)is far less known to the public.However,MSis also a severe neurodegenerative disease and the most common non-traumatic cause of neurological disability in young adults in Western countries,affecting over 2.5 million individuals worldwide. MS is an autoimmune neurological and demyelinating disease in the central nervous system(CNS),typically strikes young adults especially female,generally with onset at the age of 20-40[1].Individuals with MS could experience lifelong disability and poor quality of life for prolonged period,at the same time,the expense is pretty high for MS patients[2,3].Hence,it is of great necessity to further carry out studieson MS.

The signs and symptoms of MS vary a lot,depending on different type of peripheral nerve injuries,which include blurred vision,paresthesia or numbness,poor coordination,ataxia,fatigue,spasticity,sensory loss,sexual dysfunction,neuropathic pain,problems with memory and concentrations,and paralysis[6-8].

Although the etiology of MS is still unknown,numerous avenues of researches support the hypothesis of that autoimmune mechanism plays a pivotal role in the development of the diseases[9,10].

Due to its peculiar onset in young people,individuals with MSwould require treatments for decades or even for the rest of their life. With the successful introduction of ACTH corticosteroids in the 60’s,immunosuppressive drugs in the 70's and the introduction of interferon beta-1b[7],which is known as the first effective preventive remedy for MS,the treatment for MS has somehow progressed alot.During the past 20 years,researchers have made major breakthroughs in MS treatment.Disease-modifying treatments(DMTs)which are currently widely accepted worldwide,Treatment for MS have been applied to ameliorate symptoms,decrease the frequency of relapse,treat acute exacerbation,shorten the duration of MS attacks,and taken together,improve the quality of life of MS patients [11].The first-line injectable DMTs including glatiramer acetate and several interferons are shown to be effective and safe to a certain extent,though administration via injection are required[12].Meanwhile,the emergence of oral drugs such as azathioprine,dimethyl fumarate,fingolimod and some other second line DMTs(Natalizumab,Rituximab,etc.)opens up a new era for the treatment of MS.However,large,placebo-controlled trials as well as more researches about oral agentsarestill in need of conducting[13].

Traditional Chinese medicine[14]is an ancient healing system consisting of herbal compounds, acupuncture,moxibustion,bone setting,food therapy and physical therapy such as Taiji and Baduanjin Exercise[15].As one of the most important complementary and alternative medicine(CAM),Chinese medicine is popular in China and surrounding areas,and have long been applied to treat MS[16,17]and other nervous system diseases worldwide[18-20].

Despite these seemingly promising options,nearly all of the treatments are only partially contributing.Besides,there is still no approved option for patients with primary progressive MS hitherto,and the continued use of drugs in patients with secondary progressive MS is disputable.Additionally,the huge cost of DMTs is undoubtedly a heavy burden.

Accordingly,multiple effective approaches are needed to help repairing MS lesions and ameliorating progressive disabilities.In recent years,with the discovery of artemisin from Artemisia and sulfur trioxide from arsenic sublimate,much attention has been given to investigate Chinese medicine.Which it is a promising remedy for MS

The successful introduction and application of experimental autoimmune encephalomyelitis(EAE)[21-23],which is a intensively studied animal model of immune disease and widely used animal model that mimics the symptoms of MS in patient,provides a platform for integrative Chinese medicine to conduct systematic and evidence-based experimental studies on MS.Our review aims to summarize the findings of active ingredients of Chinese Herbal and acupuncture in the prevention and treatment on animal model of MS.

Database Search and Criteria

An online literature search for this review was adopted with scholar.google.com and PubMed.com by searching terms including “multiple sclerosis”,“MS”,experimental autoimmune encephalomyelitis,EAE,Chinese medicine,TCM,herbs,herbal medicine,acupuncture and various combinations of these terms.

Extracts and compounds of TCM herb

Extracts and compounds of Chinese herb,usually the effective ingredients,are novel application of ancient Chinese medicine.Some of the licensed extracts have already exerted significant efficacy on diseases like malaria(artemisinin)[24],cancer,etc.(extract of Hedyotis diffusa willd)[25-27].

Since the etiology and pathogenesis of multiple sclerosis involves a sophisticated network of multiple factors,compounds and extracts from traditional Chinese medicine,which exhibit multiple influences over autoimmune inflammation and neurodegenerative disease,are worth more attention.

Curcumin

Curcumin,a polyphenolic monomer,is the principal active ingredient extracted from Chinese crude drug rhizome Curcuma Longa.Curcuma Longa(also Turmeric,or Jiang Huang in TCM)is a tropical herbaceous rhizomatous plant as well as a medical herb widely used in China,India and other Asian areas for it functions in diseases like fractures,traumatic injuries,and different kinds of pain disorders[28].Curcumin has been found to possess multiple pharmacological activities and exhibit an extensive variety of anti-inflammatory, anti-oxidant,anti-carcinogenic and anti-toxic properties[29-34](Figure 1).Yet mounting evidence indicates that curcumin owns a significant effect on neural protection and inflammatory suppression of CNS[35-39].

Figure1 Chemical-protein interactions of curcumin.

Through its various anti-inflammatory function,Curcumin may exert a role in the treatment of Multiple Sclerosis.In vitro study of human astrocyte cell line(U373-MG)shows that curcumin down-regulates the release of IL-6 as well as the activity of Matrix metalloproteinase-9(MMP-9)induced by lipopolysaccharide(LPS)[40],indicative of its potential as a therapeutic agent in regulating CNS inflammatory diseases.Furthermore,in vivo study[41]of EAE mice model proved the role of curcumin in the suppression of IL-12 and IL-23 while an up-regulation of IL-4 and IL-10 expression,as well as increased CD4+,CD25+-,Foxp3+,and Treg cells,which taken together,indicating that curcumin may treat EAE mice by differentially regulating the response of CD4+T helper cell.Moreover,Feng et al.[42]explored the specific pathophysiological change concerning oligodendrocyte apoptosis and mitochondrial injury in MS and EAE model.The observations elucidated that curcumin can inhibit mitochondrial injury and suppress the apoptosis of oligodendrocyte occuring in the early phase of EAE[43].

Matrine

Matrine(MAT),which is a quinolizidine alkaloid extracted from a traditional Chinese medicine named Radix Sophorae Flave(Ku Shen in TCM),has been reported as a promising pharmaceutical for many diseases with side-effect rarely reported,such as liver cirrhosis[44,45],hepatitis B[46]and inflammation[47,48].Of note,MAT hasalso been approved as an adjuvant drug in Chinato prevent Cachexia.

Recently,MAT has been found to attenuate EAE and CNS inflammation.Li first proved the inhibiting role of MAT in suppressing immune activities of T cells,B cells and macrophages[49].Kan et al.[50]focused on the expression of adhesion molecules and chemokines and found that MAT effectively inhibited the production and expression of key adhesion molecules such as vascular cell adhesion molecule-1(VCAM-1)and intercellular adhesion molecular-1(ICAM-1)in the CNS.Key chemokines including CCL3 and CCL5 which attract inflammatory cells into CNS,were also suppressed by the treatment of MAT.Additionally,it is suggested that TLR4/MD2 pathway,which plays an important role in inducing Th1/Th17 cells in EAE,was also inhibited distinctively.In a recent study[51],Kan found that MAT attenuated EAE by downregulating glutamate and enhencing GABA levels as well as expression of two dependent glutamate transporters(GLAST and GLT-1).disruption based on the perspective that toxic molecules and proinflammatory cells may migrate into the brain via the damaged BBB in both EAE and MS,which will result in neural cell death,demyelination cerebral edema.And the results suggested that MAT reduced the leakage of BBB by strengthening basement membrane,inhibiting activities of matrix metalloproteinases(MMPs),including MMP-9 and MMP-2,and upregulating their inhibitors,which taken together,inhibited inflammatory infiltration into the CNS and protected the CNS tissues from proinflammatory cell/mediator-induced damage(Figure 2).

Figure2 Chemical-protein interactions of matrine.

The immunomodulatory mechanism of MAT on EAE was also observed[53,54].In this connection,it is suggested that MAT could suppress the production of IL-17/IL23 and increase the production of Th2 cytokines IL-4 and IL-5 in serum,and regulatory T cell(Treg)related cytokines IL-10,TGF-β1,as well as expression of Foxp3 in the spinal cord.Furthermore,MAT also shows a stimulating role in the CNS expression of Nrf2 and HO-1,which was related with oxidative stress and CNS inflammation.

Triptolide

There is a herb in traditional Chinese medicine named Tripterygium Wilfordii Hook F(TWHF,also Lei Gong Teng in TCM)that has long been used to treat systemic lupus erythematosus and rheumatoid arthritis in China[55].Triptolide(TPT),which is the major component that contributes mostly to the immunosuppressive effects of TWHF[56],also exerts immunoregulatory functions in EAE(Figure 3).It is indicated that TPT could inhibit the activation and inflammatory responses of T cells[57,58].Intriguingly,further observations[58]were obtained and showed that TPT could reduce pro-inflammatory cytokines and increase the expression of Foxp3,which is a marker for CD4+and CD25+regulatory T cells in the spleen.Hence,this finding indicated that TPT could induce antigen-specific immunomodulation by causing a regulatory T cell response rather than suppressing immune response through its effects on NF-kBmerely.

Another ground-breaking study manifested that TPT markedly increased the expression of heat shock protein 70(HSP70)mRNA and protein in the CNS.Besides,stabilization of the NF-κB/IκBαcomplex also accounted for the attenuation of inflammatory response in EAE as observed in this study[59].

Figue3 The chemical-protein interactions of triptolide.

Huperzine A

Figure4 The chemical-protein interactions of huperzine.A

Huperzine A(Hup A),a novel lycopodium alkaloid,was extracted from Chinese folk medicine Huperzia Serrata(named Qian Ceng Ta in TCM),which has been licensed as a treatment drug for Alzheimer’s Disease in China and a nutraceutical in the US[60](Figure 4).HupA is a reversible and selective acetylcholinesterase inhibitor which yields neuroprotective effects by suppressing the expression of proinflammatory mediators and the activation of glial cell[61,62].The mechanism of the neuroprotective effect of Hup A was mainly linked to glutamate toxicity.More recently,the potent anti-inflammatory property of Hup A has been explored.Several studies carried out in animal model including transient focal cerebral ischemia[63],chronic cerebral hypo-perfusion[64],and endotoxemia demonstrated that Hup A possess anti-inflammatory properties.Neuroinflammation plays a crucial role in disease initiation,which has been regarded as a common pathway of neurodegenerative diseases.Though it has already been licensed to treat AD in China,researches concerning HupA and its effect on MS are just starting on.

Our group[65]demonstrated Hup A could ameliorate EAE by suppressing autoimmune responses,inflammatory reactions,subsequent demyelination and axonal injury in the spinal cord.Administration of Hup A inhibited the expression of the pro-inflammatory cytokines(IFN-g and IL-17)and chemokines(MCP-1,TWEAK and RANTES)in the spinal cord of EAE mice while enhance the production of the anti-inflammatory cytokines (IL-4 and IL-10). Besides, proliferation of encephalomyelitic T-cells was inhibited as well as the inflammation of CNS,demyelination,and the axonal injury.Furthermore,Hup A have a strongly inhibitory activity on CCL2,IL-6,TNF-alpha,and IL-1beta production in vivo and in vitro[66].These results suggest Hup A may be a promising therapeutic candidate for treatment and prevention of neurodegenerative or autoimmune disorders,especially MS.

Eriocalyxin B

Figure5 The molecular formula of eriocalyxin B.

Isodon(Rabdosia),which is widely distributed in Yunnan Province of China,has been used as an anti-tumor and anti-inflammatory agent in folk medicine down the ages.Eriocalyxin B was initially extracted from Isodon eriocalyx var.laxiflora in 1982[67,68]With the potential of being developed into a broad-spectrum agent for the treatment of cancer.Eriocalyxin B has been studies on many kinds of cancer[69,70].Recently,its effect on EAE mice had gained concern(Figure5).

Lu et al.[71]have studied the effect of Eriocalyxin B on experimental autoimmune encephalomyelitis.It has been implicated that EriB treatment reduces CNS demyelination and inflammation,alleviates clinical symptoms and delays disease onset in EAE mice.Specifically,by down-regulating both Jak/STAT and NF-κB pathways,Eriocalyxin B remarkably suppressed the proliferation of both myelin oligodendrocyte glycoprotein(MOG)-reactive Th1 and Th17 cells.Moreover,the intervention of Eriocalyxin B given immediately after EAE induction resulted in a delayed disease onset,indicative of Eriocalyxin B`s effect on the early phase of immune responses.

Astragaloside

Figure 6 The molecular formula of astragaloside.

Astragaloside(AST),especially astragaloside IV(AS-TI),is an effective saponin component extracted from Astragali Radix(Huang Qi in TCM)(Figure 6).Reports had demonstrated an important role of AST in fighting against tumor[72],inflammation 73],heart failure[74],and hypertension[75].Concomitantly,inspired by the findings of AST in other fields,researchers then bridged the gap by investigating effects of AST on EAE.

He et al.[76]carried out studies of AST on EAE based on the theory that oxidative stress could induce neural apoptosis in the pathogenesis of EAE.From this research,it is disclosed that AST could modulate oxidative stress by reducing cellular ROS level,enhancing antioxidant defense system and increasing anti-apoptotic pathways.An anti-inflammatory function with the modulation of T-cell differentiation and infiltration into CNS was also found in this study.Besides,AST stopped the leakage of BBB in CNS,and inhibited the p53 and phosphorylation of tau by modulating the Bcl-2/Bax ratio,which all in all,demonstrated abeneficial function in EAE at multiple levels.

TCM compounds and decoctions

Herbal compounds and decoctions,which are important components of Chinese herbs,have long been used for centuries to cure different kinds of diseases.

Zuoguipill and Youguipill

Zuogui Pill and Yougui Pill,formulated by ancient doctor Jing-yue Zhang,are of great antiquity.The two pills are deemed to help strengthening body immunity,delaying senility and adjusting endocrine based on the theory of Chinese medicine.Wang et al.[77]have conducted are search on the mechanism of Yougui and Zuogui pill in promoting axonal regeneration in EAE rats.The study mainly focused on the change of expression of nerve growth factor(NGF),Nogo A,which is an axon growth inhibitor,and Nogo-66 receptor(Ng R),which is the receptor of Nogo A.And the study showed an alleviative effect of Zuogui pill and Yougui pill on axonal injury and a stimulative effect on axonal regeneration under the potential mechanism of reducing the expression of Nogo A and Ng R while increasing the expression of NGF.

There is another study concerning the effect of Zuogui Pill and Yougui Pill on experimental autoimmune encephalomyelitis[78].Dissimilarly, brain-derived neurotrophic factor(BDNF)and cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA)signaling of axonal regeneration were adopted as targets to observe the effect of Zuogui Pill and Yougui Pill.The results of this research indicated that the mRNA expression of BDNF in both brain and spinal cord were increased with the intervation of Zuogui pill and Yougui pill.As for cAMP/PKA signaling transduction pathway,the study suggest that Zuogui Pill and Yougui Pill may modulate cAMP/PKA pathway,and thereby,promote axonal regeneration.However,the correlation between the Chinese herbal medicine and cAMP/PKA pathway was not fully understood been in this study and the targets of molecular mechanism of Yougui Pill may differ from that of Zuogui Pill.

Bushen Yisuicapsule(Er Huang Formula)

Bushen Yisui Capsule,formerly called Erhuang Formula,is an effective Chinese patent drug for multiple sclerosis which consists of Radix Rehamanniae Preparata(Shudihuang),Radix Rehmanniae (Shengdihuang), Radix Polygoni Multifori(Heshouwu)and other Chinese medicine.A research was carried out to probe into the influence of Bushen Yisui Capsule on amyloid precursor protein expression(APP) and microtubule-associated protein 2(MAP2)regulation of EAE mice[79].And the research findings observed that the APP expression was down-regulated in brain and spinal cord of EAE while an increasing of MAP2 expression was detected with the administration of Bushen Yisui Capsule,which indicated this capsule may function in ameliorating axonal damage and promoting regeneration.

Li et al.[80]also looked into the mechanism of Erhuang Capsule for the treatment of MS.They found that Erhuang Capsule stimulate the expression of oligodendrocyte lineage gene 2 and enhance the differentiation of oligodendrocyte precursor cell in the brain and spinal cord of EAE mice,and thereby,improve clinical symptoms and inflammation of experimental autoimmune encephalomyelitis.

Acupuncture and Electro-acupuncture

Acupuncture is one of the most prevalent complementary and alternative treatment not only in China,but also in the United States,Europe,Southeastern Asia And so on.As one of the non-pharmaceutical therapies,acupuncture is regarded to be safe with side effects occurring rarely[81-83].Together with electro-acupuncture(EA),which is a modern and modified technique of acupuncture that integrate with electrical stimulation,acupuncture has been proved to play a role in many diseases clinically and experimentally,such as chronic pain[84],Bell`s palsy[85],stroke[86-88],and depression[89]and so forth.Moreover,the much progress of acupuncture on Parkinson`s disease[90,91]has reminded researchers to further investigate its role in other neurodegenerative diseases,including MS.Clinical studies have already shown a significant role of acupuncture or EA on improving symptoms of MS,such as reducing fatigue,spasticity and pain of MS [92,93],improving bladder dysfunction[94],and enhancing the quality of life[95].

Experimental researches have provided more convictive evidences about the effect of acupuncture or EA.In an ethidium bromide induced demyelinated spinal and injury rat model[96],which basically mimicked the pathological change of MS,amyotrophic lateral sclerosis(ALS),and other demyelinating diseases,two “Governor Vessel”acupoints Jizhong(GV6)and Zhiyang(GV9)are adopted to investigate the effect of EA.And it is suggested that EA functions in promoting neurotrophin-3(NT-3)expression,increasing the differentiation as well as the cell number of endogenous oligodendrocyte precursor cells(OPCs),and remyelinating.Besides,with the use of EAE mice model,Liu et al.[97,98]focused on the immune modulating functions of EA,which marked the first to elucidate the mechanism behind the effective properties of acupuncture on EAE.Zusanli acupoint(ST36)was adopted in their study.The former study[97]they did showed that EA reduced the severity of EAEby restoring the Th1/Th2/Th17/Treg Th cell balance and inhibiting myelin basic protein68-86(MBP68-86)-specific T cell proliferation, which resulted from stimulation of hypothalamus that increase ACTH secretion under the intervention of EAE.Later,the researchers conducted further study[98]and found that the beneficial anti-inflammatory effects of EA on EAE was connected with β-endorphin production which balanced the Th1/Th2 and Th17/Treg responses.Accordingly,the results promote the author to deem that β-endorphin could exert an important role in the development of EA-based therapies used for the treatment of MSor EAE.

Summary and Discussion

It is obvious that Chinese medicine is helpful in treating MSor EAE at least partially.Experimental studies above have also proved the efficacy of Traditional Chinese medicine in use.However,there are still many Chinese medicine decoctions widely used clinically in China except for the decoctions mentioned in this review lacking sufficient experimental study and the mechanism of many Chinese medicine decoctions treating MS still remains unclear.Besides,there are only few studies on the effect of Chinese herb extracts like Huperzine A or Eriocalyxin B in treating MSor EAE,though this extracts are found to have a therapeutic effect.More studies including clinical study are needed to thoroughly examine the efficacy and safety of this extracts to see if they are qualified to be used clinically in patients.Anyway,from this review,we can conclude that many herb extracts which originally being studied in cancer or other diseases,resulted in a remarkable effect in the treatment of EAE,which promoted us to guess whether other anti-cancer Chinese herb extracts could also exhibit functions in EAEor MS?And further studies should be conducted under this hypothesis.In addition,acupuncture including EA has an advantage in treating MS in many ways.Again,more studies about the effect of acupuncture or EA clinically and experimentally should be carried out in order to test and explain the mechanism of acupuncture. Treatment targeting remyelination rather than immune regulation in MS is now becoming more popular;however,no Chinese medicine-related article focus on the field of remyelination,whether there are any remedies to promote remyelination in Chinese medicine is still unknown.

Abbreviations:

AD:Alzheimer’sdisease

ALS:Amyotrophic lateral sclerosis

AMPA:2-amiino-3-propranoic acid

APP:Amyloid precursor protein

AST:Astragaloside

BBB:Blood-Brain barrier

BDNF:Brain-derived neurotrophic factor

CAM:Complementary and alternative medicine

cAMP:Cyclic adenosine monophosphate

CNS:Central nervous system

DMT:Disease modifying therapy

EA:Electro-acupuncture

EAE:Experimental autoimmune encephalomyelitis

GV:Governor vessel

HSP:Heat shock protein

HUP:Huperzine

ICAM:intercellular adhesion molecular

LPS:lipopolysaccharide

MAP:Microtubule-associated protein

MAT:Matrine

MBP:Myelin basic protein

MMP:Matrix metalloproteinase

MOG:Myelin oligodendrocyte glycoprotein

MS:Multiple sclerosis

NGF:Nerve growth factor

NMDA:N-methyl-D-aspartic acid

NT:Neurotrophin

OPC:Oligodendrocyte precursor cell

PKA:Protein kinase A

TCM:Traditional Chinese medicine

TPT:Triptolide

TWHF:Tripterygium Wilfordii Hook F

VCAM:vascular cell adhesion molecule

Competing interests:

The authors declare that they have no competing interests.

Author`s contributions:

Wu and Wang conceived and designed the study.Zhu and Chen collected and analyzed the references.Zhu wrote this manuscript under the guidance of Wu and Wang.All authors read and approved the final manuscript.

Acknowledgements:

This work was financially sponsored by the National Science Foundation of China(No.81202746)and the National Key Basic Research Program of China(2013CB531906).

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