Puas yog cistanche tag nrho glycoside muaj teebmeem rau olfactory?

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Cov nyhuv ntawm cistanche tag nrho glycosides ntawm nas olfactory ensheathing hlwb thiab lawv tso tawm ntawm cov teeb meem neurotrophic

Li Chuangfeng, Guo Jinyu, Zhang Xu, Liu Yunnan

Spinal cord raug mob (SCI) yog ib qho kev raug mob loj ntawm lub hauv pauspaj hlwb, suav rau {{0}}.2 feem pua ​​— 0.5 feem pua ​​ntawm tag nrho cov kev raug mob. Hauv cov pob txha pob txha, 16 feem pua ​​​​-40 feem pua ​​​​yog qhov nyuaj los ntawm kev raug mob ntawm tus txha caj qaum, feem ntau ua rau muaj kev xiam oob khab, ua rau muaj kev nyuaj siab hauv lub neej niaj hnub cuam tshuam rau lub neej zoo ntawm cov neeg mob, thiab ua rau muaj lub nra hnyav rau tsev neeg thiab zej zog.Olfactoryensheathing cells (OECs) yog hom tshwj xeeb ntawm cov paj hlwb. Plasma hlwb tuaj yeem txav mus rau hauv lub hlwb nrogolfactorypaj hlwb axons thiab tuaj yeem zais ntau yam kev loj hlob xws li NGF, GDNF, BDNF, NT-3, neuropeptide-Y, thiab S-100. Vim cov yam ntxwv ntawmolfactoryensheathing hlwb, OECs raug suav hais tias yog ib qho ntawm cov noob cog qoob loo rau kev hloov pauv ntawm tes los kho SCI. Tam sim no, kev kho mob ntawm SCI hauv kev kho mob kho mob feem ntau yog ua raws li kev kho kom rov zoo. Yuav kom nrhiav tau ib txoj hauv kev zoo dua, kev tshawb fawb hauv qab no tau ua tiav ntawm cov nyhuv ntawmcistanchetag nrhoglycosidesntawm OECs:

Cistanche tag nrho glycoside rau Olfactory ensheathing cells

1 Cov khoom siv thiab cov txheej txheem

1.1 Cov ntaub ntawv sim

Wistar nas (muab los ntawm Animal Center ntawm Inner Mongolia University);cistanchetag nrhoglycosides, DMEM / F12 nruab nrab, fetal calf serum (FCS), poly-D-lysine, arabinoside (Ara-C), trypsin, D-Hanks tov, luav anti-p75NGFR (Santa Cruz), ELISA cov khoom siv (Promega), cell incubator (Thermo-HEPA Class 100), inverted phase-contrast microscope (Olympus CKX41, Nyiv), ultra-clean workbench (Suzhou Antai Air Technology Co., Ltd.) Lub tuam txhab, SW-CJ-2FD), enzyme Daim ntawv lo analyzer (Wellscan MKH), flow cytometer, thiab lwm yam.

1.2 Cov txheej txheem tshawb fawb

1.2.1 OECs kab lis kev cai, kev ua kom huv, thiab kev txheeb xyuas: cov nas tsuag Wistar tau muab txi los ntawm decapitation thiab soaked hauv cawv rau 5 feeb. Lub pob txha taub hau anterior tau qhib kom nthuav tawmolfactorynoob. Ob tugolfactoryqhov muag teev tau raug tshem tawm tag nrho nyob rau hauv ib lub tshuab nqus tsev dissecting. Ntxuav hauv D-Hanks tov; txiav covolfactorynoob nrog microsurgical txiab, ntxiv 2ml ntawm 0.25 feem pua ​​pancreatin thiab incubate ntawm 37 degree rau 20 feeb; ntxiv D MEM / F12 kab lis kev cai nruab nrab uas muaj 10 feem pua ​​​​calf cov ntshav mus rau 2ml centrifuge raj Kev zom zaub mov raug txiav tawm hauv nruab nrab; lub tsheb npav pipette aspirated cov ntaub so ntswg tsis tiav ntawm 800 rpm thiab centrifuged rau 5 min, pov tseg cov supernatant; Thaum kawg siv cov khoom nruab nrab (20 feem pua ​​FCS ntxiv rau 20|i, mol/LForskolin ntxiv rau 20|ig/mlBPE ntxiv rau DMEM /F12) Ua rau nws mus rau hauv lx 107ml ib leeg-cell ncua kev kawm ntawv, inoculate nws nyob rau hauv ib tug polylysine-coated cell kab lis kev cai hwj, qhov chaw nws nyob rau hauv ib tug 37 degree, 5 feem pua ​​CO2 incubator, thiab ces ntxiv Ara- C Purification yog nqa tawm ntawm 36h. Siv luav anti-p75E thiab FITC-labeled tshis los tiv thaiv luav IgG theem nrab antibody rau immunohistochemical cell txheeb xyuas thiab fluorescence microscope soj ntsuam.

1.2.2 Siv MTT thiab ntws cytometry los txiav txim qhov cuam tshuam ntawmcistanche tag nrho glycosidesntawmolfactoryensheathing hlwb: inoculate cov kab lis kev cai thiab purifiedolfactoryensheathing hlwb hauv 96-zoo phaj ntawm 1X106, thiab ntxivcistanche tag nrho glycosides(kev txiav txim siab yog Ijig / ml, 10mg / ml, 100mg / ml), ntsuas tus nqi OD thiab ntsuas cov xov tooj ntawm tes cytometry tom qab 24 teev ntawm kev ua. Teeb tsa Pawg Tswjhwm thiabCistanche tag nrho glycosidesCov pab pawg sib txawv: Pawg A (Control pab pawg): Cov kab lis kev cai ntawm cov ntshav qab zib yog siv los tswj qhov tsis zoo; Pab pawg B (cistanche tag nrho glycosidestsawg concentration pawg: llig / ml); Pawg C (cistanche tag nrho glycosides nruab nrab concentration Group: L10jxg/MK); Pawg D (cistanche tag nrho glycosidessiab concentration pawg: 100|xg/mlo): 15 nas nyob rau hauv txhua pab pawg.

1.2.3 Kev txiav txim siab ntawm cov ntsiab lus neurotrophic hauv cov supernatant los ntawm ELISA: inoculate cov kab lis kev cai thiab purifiedolfactoryensheathing hlwb hauv 6-zoo phaj ntawm 1x106, ntxivcistanche tag nrho glycosides(zoo ib yam li saum toj no) tom qab 24 teev thiab ua 24 teev, thiab sau cov supernatant nyias, Txuag ntawm -80 Ntev. Cov ntsiab lus ntawm cov paj hlwb loj hlob hauv cov kua dej tau txiav txim siab los ntawm txoj kev ELISA.

1.3 Kev txheeb cais

Cov ntaub ntawv hauv txoj kev tshawb fawb no raug kho raws li cov ntaub ntawv ntsuas, thiab cov ntaub ntawv tau txais yog qhia raws li qhov txhais tau tias ± tus qauv sib txawv, thiab SPSS 11.0 cov ntaub ntawv txheeb xyuas yog siv rau kev txheeb xyuas kev txheeb xyuas. Cov ntaub ntawv tau txais los ntawm kev txheeb xyuas qhov sib txawv, thiab P<0.05 indicates="" that="" there="" is="" a="" significant="">

2 Kev tshwm sim

2.1 Cov nyhuv ntawmcistanchetag nrho glycosides rauolfactoryensheathing hlwb

2.1.1 Methyl oxazolyl tetrazole method (MT) kev txiav txim siab: Pawg A: 0.617±0.149; Pab pawg B: 0.589±0.147; Pawg C: 0.761±{13}}.138; Pawg D: 0.732±{17}}.234. Kev sib piv ntawmCistanche tag nrho glycosides group (B) and Control group (A), P>0.05, qhov txawv tsis tseem ceeb;Cistanche tag nrho glycosides10mg / ml, 100mg / ml concentration pawg (C, D) piv nrog Tswj pawg (A) P<0.05, the="" difference="" is="">

2.1.2 Cov txiaj ntsig ntawm kev txiav txim siab ntws cytometry yog qhia hauv daim duab 1. Piv nrog rau Pawg Tswj Xyuas, covcistanche tag nrho glycosidespawg tau siab dua li Pawg Tswj Xyuas, thiab feem pua ​​​​ntawm cov hlwb apoptotic qis dua li Pawg Tswj Xyuas.

2.2 Enzyme-linked immunosorbent assay (ELISA) cov txiaj ntsig

Cov txiaj ntsig kev sim tau pom tias cov ntsiab lus ntawm glial-cell-line-derived neurotrophic factor (GDNF): ua ntej noj: 302.1 ± 16.6 pg / ml, pawg A: 340.5 ± 13.2pg / ml; pawg B: 333.2 ± 23.6pg / ml; Pab pawg C: 355.2 ± 17.6pg / ml; Pab pawg D; 379.2 ± 15.3 pg / ml. Piv nrog rau kev tswj hwm ua ntej, GDNF cov ntsiab lus tau nce ntxiv; piv nrog Pawg Tswj Xyuas (A), GDNF cov ntsiab lus ntawm covCistanche tag nrho glycosides100mg / ml concentration pawg (D) tau nce ntxiv.

3 Kev sib tham

Nyuaj hauv kev tsim dua tshiab ntawm axons yog ib qho ntawm cov laj thawj tseem ceeb vim li cas nws nyuaj rau rov zoo tom qab raug mob qaum qaum. OECs muaj lub luag haujlwm hauv kev tsim kho ntawm lub hauv paus neuron axons uas lwm hom hlwb nyuaj los hloov. Nyob rau hauv xyoo tas los no, kev tshawb fawb ntawmolfactoryensheathing hlwb tau dhau los ua qhov chaw kub.Olfactoryensheathing hlwb yog ib hom tshwj xeeb ntawm glial hlwb uas muaj cov yam ntxwv ntawm Schwann hlwb thiab astrocytes. Lawv pib los ntawm covolfactoryhauv qab daus membrane thiab yog dav faib rau saum npoo ntawm lubolfactorymucosa,olfactorynerv, thiabolfactorynoob. Nws muaj peev xwm los faib thiab rov tsim dua tshiab rau lub neej thiab tuaj yeem nrog rauolfactorypaj hlwb axons mus rau hauv lub hlwb. Cov axons ntawm covolfactorycov hlab ntsha tuaj yeem dhau los ntawm kev sib tshuam ntawm lub paj hlwb peripheral thiab lub paj hlwb hauv nruab nrab txhua 30-60 hnub, rov loj hlob mus rau hauv lub paj hlwb.olfactorysheath thiab tsim kom muaj kev sib txuas synaptic 4.Olfactoryensheathing hlwb tuaj yeem zais ntau yam kev loj hlob xws li NGF, BDNF, GDNF, NT-3, neuropeptide-Y, thiab S-100. Cov kev tshawb fawb tau pom tias cov paj hlwb loj hlob (NGF) tuaj yeem tiv thaiv tus txha caj qaum liab nucleus neurons los ntawm atrophy thiab ntxiv dag zog rau cov kab mob corticospinal Sprout thiab txhawb kev tsim kho ntawm cov liab nucleus qaum qaum paj hlwb txoj hauv kev; ob qho tib si lub hlwb-derived neurotrophic factor (BDNF) thiab neurotrophin-3 (NT-3) tuaj yeem tiv thaiv cov liab nucleus qaum qaum cell apoptosis nyob rau hauv cov laus nas nrog axotomized raug mob thiab cawm tuag neurons thiab txhawb axonal regeneration P. Ntxiv rau,olfactoryensheathing hlwb kuj qhia txog lwm yam ntawm lawv cov cell membranes, xws li paj hlwb adhesion factor (N-CAM), uas koom nrog hauv cell adhesion thiab axon kev loj hlob. Vim cov yam ntxwv ntawmolfactoryensheathing hlwb, OECs raug suav hais tias yog ib qho ntawm cov noob cog qoob loo rau kev hloov pauv ntawm tes los kho SCI.

Cov khoom siv tshuaj lom neeg ntawmCistancheFeem ntau suav nrog phenylethanoid glycosides, iridoids, volatile Cheebtsam, lignans, polysaccharides, thiab alkaloids. Txoj kev tshawb nrhiav pom tiasCistanchethiab nws cov tshuaj lom neeg muaj cov teebmeem neuroprotective. Deng et al. kawm txog cov nyhuv ntawm tubulosa B ntawm apoptosis ntawm SH-SY5Y neurons tshwm sim los ntawm qog necrosis factor (TNF). Cov txiaj ntsig tau pom tias tom qab 100ug / SH-SY5Y neurons tom qab 36 teev ntawm TNF kev kho mob los ntawm L tau pom cov yam ntxwv ntawm apoptosis, thaum kev kho tubulin B rau 2 teev tuaj yeem txo qhov apoptosis tshwm sim los ntawm TNF. Tsis tas li ntawd, cov kev tshawb fawb kuj tau pom tias phenylethanoid glycosides tuaj yeem cuam tshuam cov apoptosis ntawm nas cerebellar granule hlwb los ntawm inhibiting caspase-3 thiab caspase-8, thiab tawm tsam C57 nas dub tshwm sim los ntawm methyl phenyl tetrahydropyridine (MPTP). Kev puas tsuaj ntawm qhov zoo dopaminergic neurons muaj kev tiv thaiv zoo. Tsis tas li ntawd xwb, tab sisCistanchetuaj yeem txo ntau yam kev puas tsuaj xws li ischemia-reperfusion ntawm lub paj hlwb. Cov nas lub hlwb ischemia raug mob sim qhia tau hais tias qhov infarct ntau ntawm 24h thiab 48h tom qab ischemia hauv cerebral ischemia-reperfusion pawg yog 72.98 feem pua ​​thiab 60.45 feem pua ​​​​ntawm qhov nruab nrab. Nyob rau hauv sib piv, lubcistanchetag nrho cov glycosides loj, nruab nrab, thiab qis koob tshuaj pawg (250.0mg·kg/d; 125.0mg·kg/d; 62.5mg·kg/d) raws li tuaj yeem ua rau lub hlwb ntawm nas. Tom qab cerebral ischemia-reperfusion 24h Lub infarct loj tau txo mus rau 25.94 feem pua, 25.81 feem pua, thiab 31.13 feem pua, thiab cerebral infarction loj tau txo mus rau 22.14 feem pua, 20.06 feem pua, thiab 22.27 feem pua ​​tom qab 48 teev. Muaj qhov sib txawv tseem ceeb ntawm ob, uas qhia txog kev cuam tshuam tseem ceeb ntawm neuronal apoptosis.

Nyob rau hauv qhov kev tshawb fawb no, txawv concentrations ntawmcistanche tag nrho glycosidestau siv rau OECs. Txoj kev MTT thiab kev ntsuas cytometry tau lees paub tias 10mg / ml100mg / mlcistanche tag nrho glycosidestuaj yeem txhawb kev loj hlob ntawmolfactoryensheathing hlwb. Cov txiaj ntsig ntawm flow cytometry qhia tau tiascistanche tag nrho glycosidestuaj yeem txhawb kev loj hlob ntawmolfactoryensheathing hlwb Nws tsuas yog txhawb kev loj hlob ntawm G1 voj voog ntawmolfactorycov phev hlwb thiab txo cov hlwb apoptotic. Yog li ntawd,Cistanche tag nrho glycosidestuaj yeem tsis tsuas txhawb kev loj hlob ntawmolfactoryensheathing hlwb tab sis kuj nce cell kev ua si thiab inhibit lawv apoptosis. Enzyme-linked immunosorbent assay ua pov thawj tiasCistanche tag nrho glycosidestuaj yeem txhawb nqa qhov zais cia ntawm GDNF los ntawmolfactoryensheathing hlwb. Cov txiaj ntsig saum toj no muab kev sim hauv paus rau kev siv tshuaj kho mobCistanche tag nrho glycosides.

Cov ntaub ntawv

[1] Qiu Xu, Zhu Hongxiang, Zhang Baixiang, et al. Cov xwm txheej cuam tshuam rau lub neej zoo ntawm cov neeg mob uas raug mob txha caj qaum thaum rov kho dua [J]. Suav Journal of Rehabilitation Medicine, 2009, 4(24): 313.

[2] Richter MW, Fletcher PA, Liu J, et al. Lamina propria thiabolfactorybull ensheathing hlwb nthuav tawm qhov sib txawv ntawm kev sib koom ua ke thiab kev tsiv teb tsaws thiab txhawb nqa deferential axon sprouting nyob rau hauv lesioned qaum qaum [J]. J Neurosci, 2005.25(46): 10700 -10711.

[3] Fairless R, Bamet SC. olfactory ensheathing hlwb: lawv lub luag hauj lwm hauv nruab nrab paj hlwb kho [J.Int J Biochem Cell Biol. 2005, 37(4):{4}}.

[4] Chen X, Fang H. Schwob JE. Multipotency ntawm purified. transplanted qabzib basal hlwb hauvolfactoryepithelium[J]. J Comp Neurol, 2004, 46914): 457–474.

[5] La P, Jones LL. Tuszynski MH. BDNF-expressing marostromal cell; txhawb kev loj hlob ntawm axonal ntawm qhov chaw d txha caj qaum raug mob[J]. Exp Neurol, 2005,191(2):344–360.

[6] Deng M, Zhao JY, Ju XD, et al. Kev tiv thaiv ntawm tubulosa B ntawm TNF -induced apoptosis hauv cov hlwb hlwb [J]. Acta Pharmacol Sin, 2004, 25({4}}

[7] Tian Xue-Fei, PuXiao-Ping.Phenylethanoidglycosideslos ntawmCistanchessalsa inhibit apoptosis induced los ntawm 1-methyl-4-phenylpyridinium ion hauv neurons[J]. J Ethnopharmacol, 2005, 97 (1:59).

[8] Geng X, Song L.Pu X, et al. Neuroprotective nyhuv