Cistanche Tubulosa Tiv Thaiv Dopaminergic Neurons Los Ntawm Kev Cai Ntawm Apoptosis Thiab Glial Cell-Drived Neurotrophic Factor: Hauv Vivo Thiab hauv Vitro-Ⅰ

Taw qhia

Parkinson's disease (PD) yog ib hom kab mob neurodegenerative tshwm sim hauv cov neeg laus uas muaj cov kab mob tshwm sim ntawm kev poob ntawm dopaminergic neurons hauv substantia nigra (SN) vim degeneration. Qhov hnyav ntawm tus kab mob yog cuam tshuam nrog dopamine (DA) neuronal cell poob hauv SN, uas yog raws li kev pom tias cov neurodegenerative.Cov txheej txheem tau nce ntau xyoo ua ntej cov tsos mob tshwm sim (Sawle thiab Myers, 1993). Qhov xwm txheej ntawm tus kab mob no qhia txog qhov muaj txiaj ntsig zoo rau kev kho mob los ntawm kev thaiv cov txheej txheem neurodegenerative hauv qab. Kev tshawb nrhiav kev kho mob-vim muaj zog thiab kev ua haujlwm tshwj xeeb ntawm cov teeb meem neurotrophic ntawm DA neuron ciaj sia yog qhov txaus siab heev.

NATURAL CISTANCHE TUBULOSA FORTECT DOPAMINERGIC NEURONS PHGS75% ECH 30% ACT 12%

Neurotrophic yam tseem ceeb yog cov proteins uas tseem ceeb, suav nrog cov paj hlwb loj hlob (NGF), lub hlwb-derived neurotrophic factor (BDNF), thiab glial cell-derived neurotrophic factor (GDNF), uas txhawb kev loj hlob ntawm paj hlwb, kev loj hlob ntawm neurological, kev taw qhia axonal, thiab kev ua haujlwm ntawm cov paj hlwb. Ntawm tag nrho cov neurotrophic yam uas tiv thaiv thiab txhawb kev kho cov dopaminergic neurons, GDNF muaj qhov cuam tshuam zoo tshaj plaws (Hong li al., 2008; Rangasamy li al., 2010; Allen et al., 2013). GDNF tau pom tias muaj cov teebmeem neurotrophic muaj zog ntawm DA neurons hauv vitro (Lin li al., 1993) thiab kom siv cov teebmeem neuroprotective hauv vivo. GDNF tau pom tias yuav cawm tau nigral DA neurons los ntawm cov kab mob tuag tes tuag taw tom qab phais- lossis cov tshuaj lom-induced axotomy hauv nas (Beck li al., 1995; Kearns thiab Gash, 1995; Sauer et al., 1995) thiab qee qhov tom qab. systemic tswj ntawmN-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)hauv nas (Tomac et al., 1995). Qhov tshwm sim ntau ntxiv ntawm neuronal apoptosis thiab txo kev tiv thaiv los ntawm cov teeb meem neurotrophic, uas tshwm sim los ntawm ntau yam kab mob pathological, underlie degeneration ntawm dopaminergic neurons (Holden li al., 2006).

C. tubulosa yog ib hom tshuaj ntsuab uas muaj los ntawm ntau cov nroj tsuag ntawm genus Cistanche. Nws yog ib qho kev xaiv kho mob loj rau lub raum tsis muaj zog uas cuam tshuam nrog cov tshuaj androgen hauv cov tshuaj suav tshuaj (TCM). Txog rau tam sim no, ntau qhov chaw kho mob thiab kev tshawb fawb ntawm C. tubulosa tau qhia txog cov haujlwm ntawm cov kab mob neurodegenerative. Kev txheeb xyuas ntawm TCM raum-tonifying cov tshuaj hauv kev kho mob PD tuaj yeem yog li muab lwm txoj kev kho mob rau PD.Echinacoside (ECH)yog ib qho tseem ceeb bioactive tivthaiv nyob rau hauv cov tshuaj ntsuab C. tubulosa. Cov kev tshawb fawb tau pom cov kev kho mob ntawm glycosides ntawm Cistanche thiab ECH,lus verbascoside(VER), thiab icariin (ICA) ntawm Alzheimer's disease (AD), PD, thiab lwm cov neeg mob vascular dementia (Urano thiab Tohda, 2010; Wang et al., 2013; Wu et al., 2014). Wu et al. (2014) tau qhia tias C. tubulosa extracts uas muaj ECH txaus thiab acteoside ameliorated qhov kev paub tsis meej tshwm sim los ntawm A -42 los ntawm kev thaiv cov amyloid deposition thiab thim rov qab cholinergic thiab hippocampal dopaminergic neuronal muaj nuj nqi. Tau et al. (2015) pom tiasphenylethanoid glycosideslos ntawm C. tubulosa (Ph Gs-Ct) tiv thaiv high-altitude cerebral edema los ntawm kev txo cov protein thiab mRNA qhia ntawm AQP4 nyob rau hauv lub hlwb cov ntaub so ntswg ntawm nas qauv.

NATURAL CISTANCHE TUBULOSA EXTRACT SNACKS txhim kho nco PHGS75% ECH 30% ACT 12%

Cov kev tshawb fawb yav dhau los tau pom tias suav tshuaj ntsuab suav nrog, suav nrog peb cov khoom xyaw ntawm C. tubulosa, epimedium, thiab rhizoma polygonati, alleviated kev puas tsuaj rau dopaminergic neurons thiab nce qib ntawm dopamine los ntawm kev tswj cov kev qhia ntawm neurotrophic yam (Wu li al., 2013). Yog li, nws tseem tsis tau paub tias C. tubulosa-induced neuroprotective cov teebmeem tau ntev npaum li cas thiab qhov kev cawmdim ntawm nigral DA neurons los ntawm kev tswj hwm ntawm GDNF tuaj yeem them taus qhov tseem ceeb ntawm kev txuag ntawm kev coj tus cwj pwm ntawm qhov cuam tshuam rau cov tsos mob ntawm PD cov tsiaj. Txoj kev tshawb no tau siv TCM raum-tonifying daim ntawv qhia, C. tubulosa nanopowder, uas tau txais lub teb chaws patent (patent naj npawb: 2011103028541) nyob rau hauv Suav teb thiab yav dhau los tau pom muaj qee yam kev kho mob hauv PD. Txoj kev tshawb no, yog li ntawd, tau tsim los tshuaj xyuas cov neuroprotective thiab regenerative cuam tshuam ntawm C. tubulosa kev kho mob thiab tshawb xyuas cov apoptosis ntawm MES23.5 hlwb thiab tus cwj pwm zoo nas, thiab cov kev cai ntawm GDNF, raws li ntsuas los ntawm lub roj teeb ntawm lub hom phiaj kev xeem. .

KHOOM PLIG THIAB MUAB

Khoom siv, Reagents, thiab khoom siv

C. tubulosa tau yuav los ntawm Beijing Tong Ren Tang Group, Co., Ltd., Beijing, Suav. ECH, VER, thiab ICA tuaj ntawm National Institutes for Food and Drug Control, Tuam Tshoj. MES23.5 dopaminergic neuronal cell kab yog khoom plig los ntawm xibfwb Biao Chen, Laboratory of Neurobiology, Capital Medical University, Beijing, Suav. Tsib caug C57BL / 6 txiv neej nas (qhov hnyav 20–25 g txhua) tau yuav los ntawm Shanghai SLAC Laboratory Tsiaj Co., Ltd. (daim ntawv tso cai: SCXK2012-0002), Shanghai, Suav.

MPP+, MTT, thiab glutamine tau yuav los ntawm Sigma-Aldrich (Carlsbad, CA, USA); DMEM/F12 nruab nrab thiab fetal bovine serum tau yuav los ntawm Gibco Co. (Life Technologies, Carlsbad, CA, USA); thiab MPTP, DA tus qauv thiab homovanillic acid (HVA) tus qauv tau yuav los ntawm Sigma-Aldrich (Carlsbad, CA, USA). -actin, Bax, Bcl2, GDNF, GDNF tsev neeg receptor alpha (GFR 1) thiab Ret antibodies tau muas los ntawm Cell Signaling Technology, Inc. (Beverly, MA, USA); 3,3'-diaminobenzidine (DAB) staining reagent cov khoom tau yuav los ntawm Fuzhou Maixin Biotech., Ltd. (Fujian, Tuam Tshoj); thiab SDS-PAGE gel cov qauv npaj cov khoom siv, cov khoom siv kuaj xyuas cov khoom siv ultrasensitive enhanced chemiluminescence (ECL) thiab bicinchoninic acid (BCA) assay tau yuav los ntawm Beyotime Institute of Biotechnology (Beijing, Suav).

Txoj kev tshawb no siv cov cuab yeej hauv qab no: ELX800 microplate nyeem ntawv (Bio Tek Winooski, VT, USA); CO2 incubator (Heraeus, Hanau, Lub teb chaws Yelemees); Gel DOC 2000 gel imaging analysis system, electrophoresis cell thiab electrophoresis tank (Bio-Rad, Hercules, CA, USA); DU-650 protein analyzer (Beckman Coulter, Inc., Fullerton, CA, USA); 5417R high-speed refrigerated centrifuge (Eppendorf, Hamburg, Lub teb chaws Yelemees); SXQM dual planetary pob zeb (Changsha, Tencan Powder Technology Co., Ltd, Hunan, Suav); MM400 tov khoom zeb (Retsch GmbH, Haan, Lub teb chaws Yelemees); Agilent 1200 high-performance kua chromatography (HPLC; Agilent Technologies, Santa Clara, CA, USA); PowerPac Basic electrophoresis, PowerPac Basic transmembrane transfer system, Universal Hood II chemiluminescence imager, S1000 Thermal Cycler RNA thim rov qab transcription system (Bio-Rad); Motic Med 6.0 cov ntaub so ntswg thiab cov duab tsom xam ntawm tes (Motic China Group, Co., Ltd, Xiamen, Suav).

Kev npaj ntawmC. tubulosaNanopowder

C. tubulosa tau hnyav, ces purified thiab lub cev qhuav dej. Tom qab cov pa pulverization, C. tubulosa hmoov zoo tau dhau los ntawm 200-mesh sieve thiab khov-qhuav. Lub tshuab nqus tsev ntsuas kub thiab lub zog siab pob zeb tau siv los npaj C. tubulosa nanopowder. Ua ntej, raw C. tubulosa nanopowder tau muab tso rau hauv lub tshuab nqus tsev pob-milling tank uas tau ntim nrog carbide sib tsoo pob. Qhov sib piv ntawm cov khoom sib tsoo thiab C. tubulosa nanopowder ranged ntawm 15: 1 txog 5: 1. Txhawm rau kom tau txais cov hmoov zoo, qhov ceev thiab lub sijhawm ntawm lub tshuab hluav taws xob siab tau teem rau 300 rpm thiab 20 min, raws li. Cov hmoov zoo tau hnyav rau kev ua cov ntaub ntawv nanoscale thiab ua tiav hauv lub tshuab tov khoom nrog ntau zaus ntawm 25 / s thiab oscillation ntawm 20s rau peb rov ua dua. PBS tau siv los yaj thiab npaj cov tshuaj 25 mg / mL, ua raws li 30 min ntawm ultrasonication, autoclaving, thiab thaum kawg cia ntawm −20◦C.

Kev Tswjhwm Zoo ntawm Cov Txheej Txheem Ua Haujlwm ntawmC. tubulosalos ntawm HPLC

Lub ECH thiab VER muaj gradient elution nrog octadecylsilane-bonded silica li muab tub lim, methanol li mobile theem A, thiab 0.1% formic acid tov raws li mobile theem B. Qhov kev tshawb nrhiav wavelength yog 330 nm. ICA muaj gradient elution nrog octadecylsilane-bonded silica li muab tub lim thiab acetonitrile-dej (30:70) raws li theem mobile. Kev kuaj pom wavelength yog 270 nm. Cov qauv, kev tswj hwm, thiab kev tswj tsis zoo tau ntsuas li 10 µL txhua rau qhov kev xeem.

Cell Culture thiab MTT Assay los ntsuas kev muaj peev xwm ntawm MPP+- Kho Cells

MES23.5 hlwb raug inoculated nrog 5% fetal calf serum, 1% glutamine, 2% 50 × Sato cov tshuaj, thiab DMEM / F12 nruab nrab nrog 2% penicillin / streptomycin. Lawv tau muab tso rau ntawm 37 ◦C hauv 5% CO2 incubator nrog cov av noo txaus. Cov hlwb raug cais tawm thiab dhau mus nrog 0.25% trypsin thiab kev ncua ntawm tes tau sau rau hauv theem kev loj hlob ntawm logarithmic. Cov hlwb cais nrog qhov ceev ntawm 1 × 105 tau muab noob rau hauv polylysine-coated 96- cov phiaj zoo, ua raws li qhov sib ntxiv ntawm qhov sib txawv zaum kawg (6.25, 12.5, 25, 50, 100, 200, 400 thiab 800 µmol / L. ) ntawm MPP+ media. MES23.5 hlwb uas tau tsim nrog kab lis kev cai nruab nrab rau 24 h thiab 48 h tau siv los ua kev tswj tsis zoo hauv kev sim hauv vitro. Cov hlwb los ntawm ntau pab pawg kho mob tau incubated nrog MTT reagent rau 4 h. Cov tshuaj nyob rau hauv lub qhov dej tom qab ntawd muab pov tseg thiab 150 µL ntawm DMSO tau ntxiv thiab oscillated rau 10 min. Kev nqus ntawm txhua tus qauv ntawm lub wavelength ntawm 570 nm tau ntsuas los ntawm kev nyeem ntawv tsis siv neeg microplate. Qhov feem pua ​​​​ntawm cell viability (%)=txhais tau tias absorbance ntawm pawg sim / txhais tau tias absorbance ntawm pawg tswj tsis zoo × 100%.

Qhov tseem ceeb ntawm MPP + nruab nrab tau ntxiv rau 24-teev kev kho mob hauv MES23.5 hlwb siv tib txoj hauv kev raws li kev coj noj coj ua hauv vitro. Tom qab kev kho mob, cov tshuaj hauv lub qhov dej tau muab pov tseg. Media uas muaj cov concentrations sib txawv (10, 50, 100, 200, 250, 500, thiab 1000 µg/mL) ntawm C. tubulosa nanopowder tau ntxiv rau MES23.5 hlwb hauv cov qhov dej sib txawv thiab sab laug rau incubate rau 24 h thiab 48 h. MES23.5 hlwb uas tau tsim nrog cov kab lis kev cai ib txwm muaj rau 24 h thiab 48 h tau siv los ua kev tswj tsis zoo. MES23.5 hlwb uas tau tsim nrog MPP + nruab nrab tau siv los ua lub tsheb. Kev ntsuas tau ua hauv triplicate rau txhua tus qauv. Lub absorbance ntawm cov kev kho mob thiab cov pab pawg tswj tau ntsuas los xam cov cell viability.

NATURAL CISTANCHE TUBULOSA EXTRACT ANTI FATIGUE PHGS75% ECH 30% ACT 12%

TH Expression Ntsuas los ntawm Immunocytochemistry

Thaum C. tubulosa nanopowder nyob rau hauv 100, 200, thiab 250 µg/ml, cov cell ciaj sia taus nce ntau (Daim duab 2G). Yog li, hauv kev sim tom ntej, peb tau sim peb qhov kev xav tau: cov tshuaj qis qis, nruab nrab koob, thiab cov tshuaj siab ntau. Peb qhov kev hloov pauv tau raug sim rau txhua pawg C. tubulosa. Sterilized, polylysine-coated npog npog tau muab tso rau hauv 6- cov phiaj zoo. Tom ntej no, 5 × 104 hlwb tau muab noob rau hauv txhua qhov dej thiab incubated rau 24 teev. Cov nruab nrab tshiab tau hloov pauv hauv pawg tswj hwm ib txwm muaj thiab qhov kawg ntawm 100 µmol / L MPP + nruab nrab tau hloov pauv hauv cov pab pawg kho mob ntxiv rau incubate rau 24 teev. Cov ntawv xov xwm tshiab tau hloov pauv hauv cov kev tswj hwm ib txwm muaj thiab pab pawg tsheb, thiab qhov kawg ntawm 100, 200, thiab 250 µg / mL C. tubulosa nanopowder tau tsim nrog cov hlwb rau 24 teev hauv qhov qis-, nruab nrab- thiab qib siab. C. tubulosa cov pab pawg kho mob, feem. MES23.5 hlwb nyob rau hauv cov pab pawg sib txawv tau ntxuav peb zaug hauv PBS kom tshem tawm cov supernatant thiab ntxiv kho rau hauv 4% paraformaldehyde rau 15 min. Tom qab ntxuav nrog PBS, cov hlwb raug incubated nrog ib tug peroxidase blocker ntawm 37◦C rau 30 min thiab ces ntxuav dua nrog PBS. Cov tshuaj 0.2% Triton X-100 tau siv rau cell permeabilization rau 10 min, tom qab ntawd ntxuav nrog PBS. Ib txwm tshis ntshav tau ntxiv rau txhua tus qauv thiab incubated ntawm chav tsev kub rau 30 min. Tom qab ntawd cov tshis cov ntshav ib txwm raug muab tshem tawm thiab thawj cov tshuaj tiv thaiv diluted 1:400 hauv PBS tau ntxiv rau txhua tus qauv thiab incubated ntawm 4◦C thaum hmo ntuj. Cov hlwb hauv pawg tswj tsis zoo tau raug tsim nrog PBS ntawm 4◦C thaum hmo ntuj. Tom qab ntxuav nrog PBS, cov hlwb raug incubated nrog biotin-labeled thib ob cov tshuaj tiv thaiv nyob rau hauv cov dej noo chamber ntawm 37◦C rau 20 min. Txhua tus qauv raug ntxuav hauv PBS thiab sau nrog horseradish peroxidase-streptavidin conjugates (ua haujlwm daws C) ntawm 37◦C rau 20 min. Tom qab ntxuav nrog PBS, cov hlwb tau stained nrog DAB reagent nyob rau hauv qhov tsaus ntuj rau kwv yees li 1-10 min thiab kev loj hlob ntawm cov xim xim av tau soj ntsuam nyob rau hauv lub teeb microscopy. Tom qab ntawd txhua tus qauv raug ntxuav ob zaug hauv dej distilled rau 1-2 feeb thiab cov nuclei raug counterstained nrog hematoxylin tov rau 0.5-1 min. Tom qab ntxuav txhua tus qauv hauv dej, cov hlwb tau muab tso rau hauv 1% hydrochloric acid cawv rau kev sib txawv thiab 1% aqueous ammonia, tom qab ntawd ntxuav cov hlwb hauv dej. Cov hlwb los ntawm txhua tus qauv yog tom qab lub cev qhuav dej hauv 70% ethanol rau 2 min, 80% ethanol rau 2 min, 90% ethanol rau 2 min ob zaug, 95% ethanol rau 2 min ob zaug, thiab 100% ethanol rau 2 min ob zaug. Tom qab ntawd cov hlwb tau muab tso rau hauv cov tshuaj xylene rau 2 min ob zaug thiab muab tso rau ntawm ib lub iav swb nrog nruab nrab resins. Nyob rau hauv lub teeb microscopy, txhua tus qauv underwed duab capture thiab random xaiv ntawm 5-10 zoo pom teb los txiav txim qhov kev qhia ntawm xaiv proteins nyob rau hauv dopaminergic neurons qhia los ntawm kev siv ntawm cov xim av hais thiab mus rau ib nrab kom muaj nuj nqis ntawm cov protein cov ntsiab lus los ntawm nws qhov nruab nrab grey nqi. .

Apoptosis tus nqi ntawm MES23.5 cElls ntsuas los ntawm Flow Cytometry

Adherent cells raug ntxuav ib zaug nrog PBS. Rau kev sib cais ntawm tes, qhov tsim nyog ntawm EDTA-free trypsin tov tau ntxiv rau ntawm chav tsev kub thiab cov tshuaj tau maj mam muab tso rau hauv cov hlwb kom tshem tawm. Tom qab ntawd, ib qho cell kab lis kev cai nruab nrab tau ntxiv los nres qhov trypsinization. Cov sib tov tau raug xa mus rau lub raj centrifuge tshiab thiab tom qab ntawd centrifuged rau 5 min ntawm 1500 rpm kom sau cov hlwb cais. Tom qab muab pov tseg cov supernatant, lub cell pellet tau maj mam rov qab siv PBS, thiab cov hlwb raug suav. Kwv yees li ntawm 1 × 105 rau 5 × 105 resuspended hlwb tau centrifuged rau 5 min ntawm 1500 rpm thiab cov supernatant raug muab pov tseg. Tsib microliters ntawm Annexin V-FITC binding tov tau ntxiv rau lub cell pellet kom maj mam resuspend lub hlwb. Lwm 5 µL ntawm Annexin V-FITC tau ntxiv thiab sib xyaw kom huv si. Tsib microliters ntawm propidium iodide tov tau siv rau cov xov tooj ntawm tes los ntawm incubating nyob rau hauv chav tsev kub rau 10 min sai ua ntej lub cytometry ntws.

Western Blot Analysis rauhauv vitro

Cov hlwb tau muab faib ua ib pab pawg, MPP + pab pawg kho mob, koob tshuaj tsawgC. tubulosapab pawg kho mob, pab pawg kho mob C. tubulosa, thiab pab pawg kho mob siab C. tubulosa. Kev qhia ntawm Bcl2 thiab Bax tau ntsuas hauv txhua qhov. Tag nrho ntawm 1 × 105 hlwb rau ib qhov dej hauv 6-cov phaj zoo tau siv los ua qauv thiab kev kho mob hauv txhua pab pawg ua ntej cov cell sau. Ib qho sib xyaw lysate, muaj RIPA tsis txwv, protease inhibitor, thiab phosphatase inhibitor, tau ntxiv rau lyse cov hlwb rau 30 min ntawm dej khov. Tom qab centrifugation, lub supernatant tau siv rau kev tsom xam cov protein. Tag nrho cov protein tau suav nrog kev kuaj BCA thiab sib cais los ntawm 10% SDS-PAGE. Cov proteins sib cais tau hloov mus rau ib daim nyias nyias thiab incubated nrog 5% skim mis-thaiv tsis pub nyob rau hauv chav tsev kub rau 2 teev. Tom qab ntawd lub membrane tau incubated nyob rau hauv thawj antibody (Bcl-2 0.34 mg/ml, Bax 0.11 mg/ml, 1:200 dilution) ntawm 4◦C thaum hmo ntuj. Tom qab ib kauj ruam ntxuav, daim nyias nyias tau muab tso rau hauv cov tshuaj tiv thaiv kab mob thib ob (0.5 mg / ml, 1: 5000 dilution) ntawm 4◦C rau 1 h thiab tom qab ntawd hauv ECL tus tsim tawm rau 2 min rau kev loj hlob zoo. Quantity Ib software tau siv rau kev tsom xam ib nrab ntawm cov protein qhia.

NATURAL CISTANCHE TUBULOSA FOR ANTI FATIGUE PROTECT LIVER PHGS75% ECH 30% ACT 12%

Kev sim tsiaj ua qauv thiab Kev Tswj Xyuas Tshuaj

Tsib caug tshwj xeeb-tsis muaj kab mob 8- cov nas uas muaj hnub nyoog ib lub lim tiam tau muab faib ua tsib pawg: ib pab pawg ib txwm muaj, MPTP pab pawg kho mob (Tsheb), pawg tshuaj qis C. tubulosa kho mob, koob tshuaj C. tubulosa kho pab pawg, thiab koob tshuaj C. tubulosa kho mob pawg. Cov tsiaj tau nyob ntawm 20-22 ◦C nrog kev nkag mus rau zaub mov thiab dej dawb. Cov nas nyob rau hauv ib txwm pab pawg tau txhaj tshuaj intraperitoneally nrog qhov sib npaug ntawm cov kua qaub ib txwm rau xya hnub sib law liag. Cov nas hauv lwm pab pawg kho mob tau txhaj tshuaj MPTP (30 mg / kg / d) rau xya hnub sib law liag los tsim cov tsheb.

Thaum lub sij hawm PD qauv, cov nas nyob rau hauv cov koob tshuaj tsawg, koob tshuaj nruab nrab, thiab koob tshuaj C. tubulosa cov pab pawg kho mob tau muab tshuaj intragastrically sib npaug ntawm qhov chaw kho mob ntawm 4 g / kg / d, 8 g / kg / d, thiab 16 g /. kg/dC. tubulosa nanopowder, feem, rau 14 hnub sib law liag. Cov nas nyob hauv pawg tswj hwm thiab cov tsheb hauv pawg tau tswj hwm qhov sib npaug ntawm cov kua qaub li qub rau 14 hnub sib law liag. Tag nrho cov txheej txheem sim tau pom zoo los ntawm Pawg Neeg Saib Xyuas Kev Ncaj Ncees ntawm Fujian University of TCM thiab tau ua raws li kev lees paub thoob ntiaj teb rau kev siv thiab saib xyuas tsiaj hauv chav kuaj. Txhua qhov kev siv zog tau ua kom txo qis kev txom nyem tsiaj hauv txoj kev tshawb no.