Echinacoside Txhim Kho Hematopoietic Function hauv 5-FU-induced Myelosuppression nas
Mar 19, 2022
Hu rau:joanna.jia@wecistanche.com/ WhatsApp: 008618081934791
Sai Sai, et al
Abstract
Hom phiaj:Peb tsom mus soj ntsuam cov teebmeem ntawmechinacoside cov tshuaj(ECH) yoghematopoieticmuaj nuj nqi hauv 5-FU-induced pob txha pob txha kev nyuaj siab nas.
Txoj kev tseem ceeb:Hauv vitro, tom qab stimulation nrog ECH, lub peev xwm loj hlob ntawm cov pob txha pob txha (BM) hlwb thiab pob txha pob txha stromal hlwb (BMSCs) muab los ntawmmyelosuppressionnas tau raug soj ntsuam los ntawm CCK8 kev soj ntsuam thiab morphology, feem. Hauv vivo, 5-FU-induced myelosuppression lossis tswj nas tau intragastrically tswj nrog ECH(echinacoside)ntawm 15 mg / kg los yog qhov sib npaug ntawm cov kua qaub ib hnub rau 12 hnub ua ntej BM hlwb raug cais tawm rau kev soj ntsuam colony-forming cell. Lub caij no, BMSCs tau coj mus kuaj rau 4 lub lis piam ua ntej cov hlwb raug soj ntsuam rau cov qauv kev loj hlob, cell culture supernatants tau sau rau GM-CSF secretion los ntawm ELISA, thiab RNA ntawm cov hlwb raug muab rho tawm rau EPO thiab GM-CSF RT-PCR. BM hlwb lossis BMSCs txhawb nqa nrog ECH rau 24 teev lossis 48 teev tau sau rau cov protein rho tawm thiab Western blotting.
Cov ntsiab lus tseem ceeb:ECH(echinacoside)txhawb kev loj hlob ntawm BM hlwb tab sis tsis yog BMSCs muab los ntawm 5-FU kho nas. Kev tswj hwm intragastric ntawm ECH(echinacoside)nyob rau hauv 5-FU kho nas tuaj yeem nce tus naj npawb ntawm tag nrhohematopoieticProgenitor cells thiab GM progenitor cells kom noj qab nyob zoo tswj cov nas, tab sis tsis yog BFU progenitor hlwb. BMSCs los ntawm ECH-khomyelosuppressionCov nas loj hlob zuj zus thiab nthuav tawm ntau dua GM-CSF, tab sis tsis yog EPO. ECH qhib txoj hauv kev PI3K teeb liab hauv 5-FU tua BM hlwb.
Qhov tseem ceeb:ECH(echinacoside)tuaj yeem txhim kho covhematopoietickev ua haujlwm ntawm cov pob txha hauv 5-FU-inducedmyelosuppressionnas. ECH(echinacoside)tuaj yeem suav hais tias yog lwm txoj kev kho mob zoo rau cov neeg mob thaum lub sijhawm siv tshuaj khomob lossis HSC hloov pauv.
Ntsiab lus:Echinacoside, Hematopoietic, Myelosuppression, GM-CSF, PI3K signaling pathway
Echinacoside hauvCistanche
Taw qhia
Fluorouracil los yog 5-FU yog ib qho ntawm cov kws khomob tshwjxeeb tshaj plaws rau kev kho mob qog noj ntshav thiab palliative [2,9,28]. Nws yog ib qho analog ntawm uracil nrog fluorine atom ntawm C-5 txoj hauj lwm nyob rau hauv qhov chaw ntawm hydrogen thiab ua hauj lwm los ntawm irreversible inhibiting cov kev ua ntawm thymidylate synthase thiab tom qab inhibiting DNA replication [20]. Leukopenia thiab atrophy ntawmhematopoieticCov kab mob hauv nruab nrog cev yog ob yam tshuaj lom neeg muaj feem xyuam rau 5-FU daim ntawv thov hauv tib neeg, nas, thiab npua [15,17,22].Myelosuppression, raws li qhov tshwm sim ntawm inhibitory thiab cytotoxic cuam tshuam rauhematopoieticprogenitor cells [10] thiab oxidative kev nyuaj siab ntawm cov pob txha pob txha [25], txwv tsis pub siv tshuaj kho mob ntawm 5-FU-based chemotherapies.
Cov pob txha pob txha raug tsim los ntawm 5-FU ua rau poob ntawm tag nrho cov qe ntshav. Cov tsos mob tshwm sim muaj xws li kub taub hau, anemia, thiab los ntshav, thiab cov kev kho mob feem ntau yog nyob ntawm kev tso ntshav thiab kev loj hlob ntxiv [4]. Kev hloov pauv tuaj yeem ua tau zoo hauv kev ntxiv cov qe ntshav liab thiab platelets, tab sis qhov kev pab no yog luv luv thiab xav tau kev kho mob dua. Kev loj hlob muaj xws li erythropoietin, granulocyte colony-stimulating factor (G-CSF), thiab granulocyte-macrophage colony-stimulating factor (GM-CSF). Cov yam ntxwv no yog cov tshuaj ntuj thiab tuaj yeem txhawb cov pob txha ua haujlwm [7,21], tab sis yog siv nyiaj thiab tsis muaj txiaj ntsig rau kev txhawb nqa cov ntshav dawb. Lwm txoj kev kho mob zoo yog xav tau. Cov tshuaj suav tshuaj yog ib qho kev xaiv uas tau txais txiaj ntsig [5]. Hauv ob peb xyoos dhau los, ntau cov kev tshawb fawb tau tsom mus rau cov txiaj ntsig ntawm cov tshuaj suav tshuaj ntsuab los tiv thaiv thiab kho cov kws khomob ntsig txog.myelosuppression[33,35].
Echinacoside(ECH) (Fig. 1) yog ib tug phenylethanoid compound muab rho tawm los ntawm tsoos suav tshuaj Herba Cistanches uas yog dav siv los ua tus neeg sawv cev rau cov ntshav ncig, tshuaj rau lub raum tsis ua hauj lwm raws li zoo raws li cov kab mob neurodegenerative [24]. ECH(echinacoside)muaj ntau yam tshuaj ua haujlwm ntawm cov pob txha rov tsim dua tshiab [18], tiv thaiv oxidation [8,14,27], tiv thaiv kab mob, tiv thaiv kab mob, thiab kev txhim kho tus cwj pwm [11,16]. Nws kuj tau tshwm sim los ua tus neeg sawv cev tiv thaiv kab mob siab hepatotoxicity [32] thiab mob ntsws mob [34]. Trk receptors thiab txoj hauv qab Trk-ERK txoj hauv kev tuaj yeem koom nrog hauv nws cov kev tiv thaiv neuronal tiv thaiv kev raug mob rotenone [36]. Lub luag haujlwm thiab cov txheej txheem tshiab ntawm ECH hauv kev txhawb nqamyelosuppressionkev loj hlob ntawm pob txha hlwb tsis to taub tag nrho. Hauv txoj kev tshawb no, peb tau soj ntsuam qhov cuam tshuam ntawm ECH(echinacoside)ntawmhematopoieticmuaj nuj nqi hauv 5-FU-induced pob txha hlwb kev nyuaj siab nas hauv vitro thiab hauv vivo thiab cov txheej txheem cuam tshuam.
Cov ntaub ntawv thiab cov txheej txheem
Tshuaj thiab biochemicals
Echinacosidetau yuav los ntawm Ronghe Medical Technology thiab Development Corporation (Shanghai, Tuam Tshoj) (purity Ntau dua lossis sib npaug li 98 feem pua, txiav txim los ntawm HPLC) thiab yaj hauv DMSO. 5-FU tau txais los ntawm HaoRan Biotechnology Corporation (Shanghai, Suav). Methylcellulose nruab nrab nrog recombinant cytokines (MethoCult nruab nrab) rau nas hlwb tau yuav los ntawm Stem Cell Technologies (Vancouver, Canada).
cistanche lub cev tsim
Tsiaj txhu thiab kev sim tsim
Cov kab mob tshwj xeeb-dawb, 5- rau 6-ib lub lis piam poj niam Balb/c nas, hnyav ntawm 15-18 g tau yuav los ntawm Shanghai Laboratory Tsiaj Tuam Txhab (SLAC, Shanghai, Suav). Cov nas tau khaws cia hauv qhov chaw tsiaj ntawm Zhejiang University, Suav. Cov tsiaj sim tau pom zoo los ntawm Pawg Saib Xyuas Tsiaj thiab Siv Tsiaj ntawm Zhejiang University thiab tau ua raws li Cov Lus Qhia rau kev saib xyuas thiab siv cov tsiaj kuaj [1,31].
Cov nas tau txhaj tshuaj intraperitoneally nrog ib koob ntawm 200 mg / kg 5-FU txhawm rau txhim kho pob txha pob txha kev nyuaj siab raws li tau piav qhia los ntawm Choi EH li al. [6]. Rau kev kawm hauv vitro, cov hlwb pob txha (BM) thiab cov pob txha pob txha stromal hlwb (BMSCs) raug cais tawm 2 hnub tom qab 5-FU txhaj. Hauv vivo, nas tau muab faib ua 4 pawg, 5–6 nas nyob rau hauv txhua pawg (n=5–6): 5-FU/ECH pawg, 5-FU/control pab pawg, ECH(echinacoside)pab pawg, thiab tswj pawg. Hauv 5-FU/ECH pawg, tom qab 5-FU txhaj, nas tau noj intragastrically noj txhua hnub nrog ECH ntawm 15 mg/kg ib hnub rau 12 hnub. Hauv 5-FU/control pab pawg, tsis yog ECH(echinacoside), qhov sib npaug ntawm cov kua qaub ib txwm tau muab rau 5-FU txhaj cov nas. Rau cov pab pawg ECH thiab pawg tswj hwm, ECH lossis cov tshuaj ntsev ib txwm raug tswj hwm los ntawm tib txoj kev rau 5-FU cov nas tsis kho. Hnub tim 12, cov nas tau euthanatized thiab cov pob txha pob txha tau muab rho tawm rau kev kawm ntxiv. Hauv kev nthuav dav, BM hlwb tau npaj rau kev soj ntsuam colony-forming cell (CFC), BMSCs yog rau kev soj ntsuam kev loj hlob, thiab cell culture supernatants ntawm BMSCs tau sau rau GM-CSF ELISA thiab RNA ntawm BMSCs tau muab rho tawm rau EPO thiab GM-CSF RT. -PCR. Kev sim tau rov ua dua 3 zaug.
Kev npaj ntawm BM hlwb thiab BMSCs
BM hlwb tau npaj los ntawm ib leeg-cell suspensions muab rho tawm los ntawm nas pob txha pob txha. Hauv luv luv, nas tau euthanatized thiab soaked hauv ethanol rau 5 feeb. Cells tau ntws tawm los ntawm cov kwj dej medullary ntawm tibia thiab cov pob txha femoral los ntawm kev rov ntxuav dua nrog 23- ntsuas koob thiab sieved los ntawm 100 μm cell strainer kom tau txais ib leeg-cell suspensions. Erythroid pob txha pob txha hlwb tau lysed los ntawm lub sij hawm luv luv incubation (30 s) nrog ammonium chloride tsis ua raws li rov qab ntxuav cov kauj ruam nrog cov kab lis kev cai nruab nrab. Viability thiab cell suav tau txiav txim siab los ntawm trypan xiav cais txoj kev siv lub hemocytometer.
Rau tiam ntawm BMSCs, BM hlwb raug tso cai kom loj hlob hauv qhov dej ntawm TC-phaj rau 24 teev, tom qab tshem tawm cov hlwb uas tsis yog-adherent thiab rov ntxuav nrog kab lis kev cai nruab nrab, cov hlwb adherent raug coj los ua BMSCs.
Cells raug coj los ntawm 37 degree nrog 5 feem pua CO2 nyob rau hauv ib qho chaw ntub dej hauv Dulbecco's Modified Eagle Medium (DMEM) ntxiv nrog 10 feem pua ntawm cov fetal bovine serum (FBS), penicillin / streptomycin (100 U / ml), thiab L-glutamine. (2 nM). Tag nrho cov cell culturing reagents tau yuav los ntawm Invitrogen (Shanghai, Suav).
Nyob rau hauv vitro BM hlwb proliferation los ntawm CCK8 assay
BM hlwb los ntawm 5-FU kho nas tau plated ntawm qhov ceev ntawm 1 × 103 hlwb/zoo nyob rau hauv ib tug 96-zoo phaj (Costar) thiab kab lis kev cai nyob rau hauv ib tug nruab nrab supplemented nrog txawv concentrations ntawm ECH(echinacoside)({{0}} μM, 0.1 μM, 1 μM, 10 μM, 25 μM, 50 μM, feem). Hnub 3 lossis hnub 6, cov hlwb tau "pulsed" nrog 20 ul ntawm CCK{10}} kua reagent (Beyotime, Jiangsu) thiab kab lis kev cai ntxiv rau 4 h. Cell proliferation tau soj ntsuam los ntawm kev ntsuas cov colorimetric CCK8 ntawm tus nyeem ntawv ultra microplate (Molecular Devices) ntawm 450 nm.
Echinacoside hauvCistanche
Hauv vitro BMSC proliferation thiab morphology
BMSCs tau npaj nyob rau hauv qhov dej ntawm 6-zoo phaj nrog thawj qhov ceev ntawm BM hlwb (tseem los ntawm 5-FU kho nas) yog 3.5 × 106 hlwb/zoo. Tom qab 24 teev, kab lis kev cai nruab nrab tau hloov los ntawm qhov nruab nrab uas muaj ECH (50 ug / ml). Txhua lub lim tiam, ib nrab ntawm cov supernatants raug tshem tawm thiab hloov los ntawm cov kab lis kev cai tshiab (nrog lossis tsis muaj ECH(echinacoside)raws li qhia). Tom qab 4 lub lis piam ntawm kab lis kev cai, morphology ntawm BMSCs tau pom nyob rau hauv ib qho inverted microscope.
Colony-forming cell (CFC) assay
Kev ntsuam xyuas CFC hauv MethoCult media tau ua raws li cov chaw tsim khoom cov lus qhia. Hauv luv luv, BM hlwb los ntawm ntau pawg nas tau npaj ntawm 2 × 105 ib ml. Tom qab ntawd, 0.4 ml ntawm cov hlwb tau ntxiv rau hauv 4 ml ntawm MethoCult nruab nrab los ua kom cov cell sib tov. Tom qab ua tib zoo sib xyaw nrog 16- ntsuas blunt-kawg koob, 1.1 ml ntawm cov xov tooj ntawm tes sib xyaw tau muab xa mus rau hauv 35 hli kab lis kev cai tais (Stem Cell Technologies), triplicated, thiab incubated ntawm 37 degree hauv cov av humidified ntawm 5 feem pua CO2. rau 7day. Tom qab 7 hnub ntawm kab lis kev cai, cov cheeb tsam uas muaj tsawg kawg yog 30 lub hlwb raug txheeb xyuas thiab suav.
Rov qab transcriptase-polymerase saw cov tshuaj tiv thaiv (RT-PCR) thiab GM-CSF ELISA
BMSCs muab tau los ntawm ntau pawg nas tau coj mus kuaj rau 4 lub lis piam raws li tau piav qhia saum toj no. Tom qab 4- lis piam kab lis kev cai, BMSCs tau sau tag nrho RNA rho tawm nrog RNAsimple Total RNA cov khoom siv (TianGen, Beijing). Thawj-strand cDNA tau tsim los siv 1 ug ntawm tag nrho RNA (DNase-treat-ed) siv iScript cDNA synthesis kit (Bio-Rad, Richmond, CA). Cov kev qhia ntawm granulocyte-macrophage colony-stimulator factor (GM-CSF) thiab Erythropoietin (EPO) tau txheeb xyuas los ntawm ib nrab-quantitative RT-PCR. PCR primer sequences yog raws li nram no: GM-CSF: 5′-GGCCTTGGAA GCATGTAGAGG-3′ (forward) thiab 5′-GGAGAACTCGTAGAGACGA CTT-3′ (reverse); EPO: 5′-CCTCATCTGCGACAGTCG AG-3′ (forward) and 5′-ACAACCCAT CGTGACATTTTCT-3′ (reverse); lub tsev tu gene Beta-actin: 5′-GTATCCCTGACCCT GAAGTACC-3′ (forward) and 5′-TGAA GGCTCCAAACATGATCT-3′ (reverse). PCR amplification tau ua tiav ntawm 2720 Thermal Cycler (Applied Biosystems) thiab cov khoom raug tshuaj xyuas siv agarose gel electrophoresis. MRNA qhia theem ntawm cov hom phiaj tau normalized rau ntawm beta-actin. Cov supernatants ntawm BMSCs tau sau rau GM-CSF ELISA kev soj ntsuam los ntawm kev siv cov khoom siv GM-CSF ELISA (JinSui, Shanghai) raws li cov chaw tsim khoom cov lus qhia.
Western blotting
BM hlwb thiab BMSCs los ntawm 5-FU inducedmyelosuppressionnas lossis nas tswj tau plated ntawm qhov pib ceev ntawm 5 × 106 rau ib qhov dej hauv 6-cov phiaj zoo. Tom qab 24 teev, ECH tau ntxiv rau cov kab lis kev cai nruab nrab ntawm 50ug / ml thiab cov hlwb raug tso cai kom loj hlob ntxiv rau 24 teev lossis 48 teev ua ntej cov hlwb raug lysed hauv 1 × RIPA tsis (CST, Danvers, MA) uas muaj 1 mM phenylmethylsulfonyl fluoride. (PMSF). Protein concentration tau txiav txim siab los ntawm kev ntsuam xyuas BCA protein. 30ug ntawm tag nrho cov proteins tau electrophoresed ntawm 10 feem pua sodium dodecyl sulfate-polyacrylamide gels thiab pauv mus rau PVDF daim nyias nyias (Millipore, MA, USA). Membranes raug thaiv nrog kev thaiv tsis pub ((Beyotime, Haimen, Tuam Tshoj) rau 1 h ntawm 37 degree, thiab tom qab ntawd incubated hmo ntuj ntawm 4 degree nrog cov tshuaj tiv thaiv -actin (#4967, CST), phospho-PI3 kinase p85 (Tyr458) / p55 (Tyr199) (#4228, CST), phospho-Akt (Ser473) (#4060, CST), Akt (yias) (C67E7) (#4691, CST), phosphor-Stat1 (Tyr701) (# 9167, CST), Phospho-Stat3 (Tyr705) (#9415, CST), Phospho-p38 MAPK (Thr180/Tyr182)(#9215, CST), Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204)(#4377, C. ), p38 MAPK(#9212, CST), p44/42 MAPK (Erk1/2)(#4695, CST), Phospho- SAPK/JNK (Thr183/Tyr185)(#4668, CST), and SAPK/JNK(# 9258, CST), ntsig txog. Tom qab ntxuav, cov daim nyias nyias tau soj ntsuam nrog horseradish-peroxidase-conjugated tshis los tiv thaiv tus luav theem nrab antibody (Lianke, Hangzhou, Suav teb) ua raws li kev kuaj pom cov teeb liab nrog FluorChem E System (Protein Yooj Yim, Santa Clara, CA, USA).
Kev txheeb cais
Cov ntaub ntawv tau nthuav tawm raws li qhov txhais tau tias ± SEM Qhov tseem ceeb ntawm qhov sib txawv ntawm txhua pab pawg tau txiav txim los ntawm Tub Kawm Ntawv t-test. A p-value b 0.05 raug suav hais tias yog qhov tseem ceeb.
cistanche lub cev tsim
Cov txiaj ntsig
Hauv vitro muab ECH(echinacoside)txhawb nqa 5-FU suppressed BM cell proliferation
BM hlwb los ntawm 5-FU inducedmyelosuppressionnas tau kho nrog ECH(echinacoside)ntawm ntau qhov sib txawv xws li 0.01 mus rau 50 μM rau 3 hnub lossis 6 hnub thiab kev loj hlob ntawm tes raug soj ntsuam los ntawm CCK8 kev soj ntsuam. Raws li pom nyob rau hauv daim duab 2, lub proliferation ntawm ECH-kho BM hlwb yog siab dua cov ECH-tsis kho hlwb ob qho tib si tom qab 3 los yog 6 hnub ntawm kab lis kev cai. Cov nyhuv stimulus pom ib yam tshuaj-nyob ntawm seb.
Hauv vitro muab ECH(echinacoside)tsis muaj kev cuam tshuam rau 5-FU suppressed BMSC proliferation
BMSCs los ntawm 5-FU inducedmyelosuppressionnas tau kho nrog ECH(echinacoside)ntawm qhov concentration ntawm 50 ug / ml rau 4 lub lis piam thiab cell proliferation raug soj ntsuam los ntawm kev soj ntsuam ntawm cell morphology nyob rau hauv ib lub teeb inverted microscope. Raws li pom nyob rau hauv daim duab 3, cov qauv kev loj hlob thiab morphology ntawm cov hlwb zoo sib xws nrog lossis tsis muaj ECH kho.
Intragastrically tswj ECH(echinacoside)txhawb kev ua haujlwm hematopoietic ntawm 5-FU induced myelosuppressive granulocyte-macrophage (CFU-GM), tab sis tsis yog erythroid (BFU-E) progenitor cells
Cov qauv kev loj hlob thiab kev sib txawv ntawm BM hlwb los ntawm 5-FU/ ECH, 5-FU/control, ECH(echinacoside), thiab pab pawg tswj tau raug soj ntsuam los ntawm CFC kev soj ntsuam. Tom qab 7 hnub ntawm kab lis kev cai ntawm BM hlwb hauv MethoCult nruab nrab, CFCs tau muab faib thiab suav nrog hauv qhov chaw nyob rau hauv lub teeb pom kev zoo. Raws li pom hauv daim duab 4, kev tshuaj ntsuam ntawm cov qib progenitor tau qhia qhov tseem ceeb (pb 0). piv nrog rau 5-FU/control pab pawg (169.3 ± 11.1 vs. 54.5 ± 3.1), nrog rau pawg tswj hwm (169.3 ± 11.1 vs. 129.3 ± 5.6) (pb 0.05, ntsig txog). Tus naj npawb ntawm BFU-E hauv txhua pab pawg tsis muaj qhov sib txawv (73.0 ± 11.1 vs. 74.7 ± 8.6 vs. 91.2 ± 5.5). Tus naj npawb ntawm CFU-GM thiab BFU-E hauv pawg ECH yog yuav luag tib yam li hauv pawg tswj hwm (cov ntaub ntawv tsis qhia).
Daim duab 4. Intragastrically tswj ECH(echinacoside)stimulates covhematopoietickev ua haujlwm ntawm 5-FU inducedmyelosuppressiongranulocyte-macrophage (CFU-GM), tab sis tsis yog erythroid (BFU-E) progenitor cells. (A) Ib daim duab sawv cev ntawm cov koom txoos ua ke. CFU-GM: colony-forming units-granulocyte macrophage. BFU-E: tawg tsim chav tsev-erythroid. (B) Cov duab sawv cev ntawm cov colony-forming units ntawm txhua pab pawg. (C) Cov naj npawb ntawm cov colony-forming units ntawm txhua pab pawg. BM hlwb raug cais tawm ntawm txhua pab pawg thiab noob rau CFU-GM thiab BFU-E progenitor cell colony-forming assay raws li tau piav qhia hauv Cov Khoom thiab cov txheej txheem. Cov txiaj ntsig tau txais los ntawm 3 qhov kev sim ywj pheej thiab qhia tias txhais tau tias ± SEM, n=5–6. * pb 0.05 piv nrog pawg tswj hwm, ## pb 0.0001 piv nrog 5-FU pawg.
Intragastrically tswj ECH(echinacoside)txhawb nqa 5-FU induced myelosuppression BMSC proliferation.
Tom qab 4 lub lis piam ntawm kab lis kev cai, kev loj hlob ntawm BMSCs tau soj ntsuam los ntawm kev soj ntsuam ntawm cov cell morphology nyob rau hauv lub teeb inverted microscope. Raws li pom hauv daim duab 5, BMSCs hauv 5-FU/ECH(echinacoside)pab pawg loj hlob ntau dua li pawg tswj hwm.
Intragastrically tswj ECH(echinacoside)txhawb nqa GM-CSF ntau lawm ntawm 5-FU induced myelosuppressive BMSCs
Tom qab 4 lub lis piam ntawm kab lis kev cai ntawm BMSCs, mRNA kab lus ntawm GM-CSF thiab EPO ntawm cov hlwb raug txiav txim los ntawm RT-PCR. Raws li pom nyob rau hauv daim duab 6A, GM-CSF qhia nyob rau hauv BMSCs los ntawm ECH-kho pob txha pob txha suppression nas tau nce ntxiv piv nrog rau 5-FU/control pab pawg. GM-CSF ntau lawm hauv cell culture supernatants tau txiav txim los ntawm ELISA. Raws li qhia hauv daim duab 6B, piv nrog rau 5-FU/ pawg tswj hwm, lub {{10}FU/ECH(echinacoside)pab pawg tau nthuav tawm ntau dua ntawm GM-CSF (pb 0.05) thiab yuav luag ntau npaum li pawg tswj hwm.
ECH(echinacoside)activates PI3K signaling pathway nyob rau hauv 5-FU suppressed BM hlwb
BM hlwb thiab BMSCs los ntawm 5-FU inducedmyelosuppressionlos yog tswj nas tau txhawb nqa nrog ECH(echinacoside)ntawm qhov concentration ntawm 50 ug / ml rau 24 teev lossis 48 teev ua ntej cov hlwb tau sau rau cov protein rho tawm thiab Western blotting. Raws li cov mechanisms ntawm ECH(echinacoside)tsis tau kawm ntau, peb tau tshuaj xyuas ib lub vaj huam sib luag ntawm txoj kev teeb liab, suav nrog JAK/STAT1, JAK/STAT3, SAPK/JNK, p-38 MAPK, p44/42 MAPK, thiab PI3K/Akt signaling pathways. Raws li pom hauv daim duab 7, ECH tuaj yeem ua rau muaj kev qhia siab ntawm p-PI3K p85/p55 hauv BM hlwb tau los ntawm 5-FU nas uas qhov induction tsis pom tseeb hauv BMSCs lossis hauv pawg tswj hwm. Rau tag nrho lwm txoj hauv kev, peb tsis pom qhov sib txawv loj (cov ntaub ntawv tsis qhia).
Kev sib tham
Zoo li muaj tseeb tias ECH(echinacoside)tsis muaj kev cuam tshuam rau cov nas uas muaj hematopoiesis li qub vim rau cov nas uas tsis muaj 5-FU kev kho mob, tus naj npawb ntawm cov hematopoietic progenitor hlwb hauv ECH(echinacoside)pab pawg neeg yuav luag zoo ib yam li cov pab pawg tswj hwm, yog li ua rau kev loj hlob thiab kev ua haujlwm ntawm BMSCs raws li qhia los ntawm morphology, GM-CSF thiab EPO secretions. Yog li, nyob rau hauv kev sib ntsib tshwm sim, cov ntaub ntawv los ntawm ECH(echinacoside)pab pawg tau ua ke nrog pawg tswj hwm thiab piav qhia tias yog kev tswj hwm.
5-FU ua cov tshuaj chemotherapy hauv kev kho mob qog noj ntshav muaj kev cuam tshuam rau cov pob txha ua haujlwm. Kev siv koob tshuaj ntau ntawm 5-FU tuaj yeem ua rau mob pob txha loj heev [19]. Hauv peb txoj kev tshawb fawb, tus qauv kev tiv thaiv kab mob hauv pob txha tau ua tiav hauv cov nas los ntawm 5-FU kev kho mob, raws li kev txhawb nqa los ntawm kev txo qis hauv pawg neeg ua haujlwm thiab morphology ntawm BMSCs.
Pob txha pob txha yog qhov lojhematopoieticLub cev muaj cov kab mob hemato-poietic qia hlwb (HSCs) thiab cov pob txha pob txha stromal hlwb (BMSCs) [13]. Cov hlwb qub tuaj yeem tsim cov qe ntshav nrog rau cov qe ntshav liab, platelets, monocytes, granulocytes, thiab lymphocytes. Thiab cov hlwb tom kawg tuaj yeem tsim cov rog, pob txha, thiab pob txha uas txhais tau tias yog mesenchymal lossis stromal hlwb [26]. Nws tau txais txiaj ntsig zoo tias BMSCs yog cov khoom tseem ceeb ntawm HSC niche qhov twg ob tus kab mob hlwb no tau npaj kom muaj kev sib raug zoo.hematopoieticthiab cov skeletal homeostasis [29] BMSCs tuaj yeem txhawb nqa thiab txhim kho kev loj hlob ntawm HSCs los ntawm kev tso cai tswj cov molecules thiab cytokines, xws li FL [23], GM-CSF [12], thiab SCF [12,3].
Cov ntaub ntawv hauv vitro tau pom tias ECH(echinacoside)tuaj yeem txhawb kev loj hlob ntawm BM hlwb los ntawmmyelosuppressionnas nyob rau hauv ib koob tshuaj-raws li, txawm hais tias, feem ntau, cov hlwb loj hlob ntau zog nyob rau hnub 3 tshaj li ntawm hnub 6, tej zaum yog vim nyob rau hauv vitro kab lis kev cai ib puag ncig. Ntawm qhov tsis sib xws, ECH(echinacoside)tsis muaj kev cuam tshuam rau kev loj hlob ntawm BMSCs los ntawm 5-FU kho cov nas. Txhawm rau tshem tawm qhov ua tau tias qhov kev cuam tshuam tsis zoo tsis yog vim qhov kev loj hlob ntawm tes, hauv qhov chaw sim, peb kuj suav nrog Rg1, cov tshuaj ntsuab uas tau raug pov thawj los txhawb BMSCs los ntawmmyelosuppressionnas loj hlob, raws li kev tswj zoo [33].
Hauv vivo cov ntaub ntawv pov thawj tias ECH(echinacoside)Nws tuaj yeem txhim kho hematopoiesis zoo, feem ntau yuav yog granulocytopoiesis thiab megakaryocytopoiesis tab sis tsis yog erythropoiesis,myelosuppressionnas. Kev tswj hwm intragastric ntawm ECH(echinacoside)rau 5-FU kho cov nas rau 12 hnub ua rau muaj tus lej ntawm ob qho tag nrhohematopoieticProgenitor cells thiab granulocyte-macrophage progenitor cells, tab sis tsis yog ntawm erythroid progenitor cells raws li tau hais los ntawm tus lej ntawm CFU-GM thiab BFU-E los ntawm CFC kev soj ntsuam. Qhov zoo siab, peb pom tias tus naj npawb ntawm CFU-GM hauv 5-FU/ECH(echinacoside)pab pawg tseem siab dua li hauv pawg tswj hwm. Peb kwv yees qhov kev nthuav dav sai ntawm CFU-GM uas tshwm sim raws li kev them nyiaj raumyelosuppressionxwm txheej coj los ntawm 5-FU. Peb cov ntaub ntawv kuj tau qhia tias ECH tuaj yeem txhawb nqa BMSC proliferation thiab GM-CSF tab sis tsis yog EPO pro-duction hauvmyelosuppressionnas, uas tau ua raws li CFC cov txiaj ntsig.
Cov txiaj ntsig tam sim no tau taw qhia txog txoj kev tshiab ECH(echinacoside)uas tej zaum yuav raug siv los txhim kho nwshematopoieticua haujlwm hauvmyelosuppressionnas (raws li qhia hauv daim duab 8): Nyob rau hauv ib qho chaw HSC niche, los ntawm kev sib xyaw BMs kab lis kev cai hauv vitro lossis hauv cov pob txha pob txha hauv qhov chaw, ECH txhawb nqa BMSC proliferation thiab zus tau tej cov GM-CSF, yog li stimulates thiab txhim khu kev loj hlob thiab sib txawv. ntawm HSCs. Yog tias tsis muaj HSCs, ECH tsis tuaj yeem txhawb nqa BMSC kev loj hlob.
GM-CSF receptor muaj (ligand binding) thiab (sib nqus) subunits. GM-CSF thawj khi rau lub subunit, ces induces downstream signaling cascades los ntawm kev sib cuam tshuam ntawm effector proteins nrog lub subunit [30]. subunit transactivated JAK2, uas pib pib qhov taw qhia qis. Cov teeb liab modules qhib muaj xws li Janus kinase / signal transducer thiab activator ntawm transcription (JAK/STAT), txoj kev mitogen-activated protein kinase (MAPK), thiab phosphatidylinositol 3-kinase (PI3K) txoj kev [30]. Qhov zoo siab, qhov tshwm sim los ntawm peb qhov kev tshuaj ntsuam tau pom tias txoj hauv kev PI3K tau qhib rau hauv BM hlwb tau los ntawm 5-FU-kho nas, tab sis tsis pom tseeb hauv 5-FU suppressed BMSCS thiab cov BM hlwb lossis BMSCs. Nws zoo li tias dhau ntawm kev ua kom GM-CSF / PI3K txoj hauv kev, ECH(echinacoside)selectively stimulates kev loj hlob ntawm BM hlwb.
Cov lus xaus
Hauv cov ntsiab lus, peb tau pom tias ECH(echinacoside)muaj kev cog lus txhawb nqa rauhematopoieticmuaj nuj nqi nyob rau hauv 5-FU-induced pob txha pob txha suppression nas, tshwj xeeb tshaj yog nyob rau hauv lub granulocytic thiab macrophagocytic hlwb, qhia tias ECH(echinacoside)tej zaum yuav muaj peev xwm rau kev siv tshuaj kho mob hauv pob txha pob txha inhibition thaum siv tshuaj khomob lossis HSC hloov pauv tom qab kho nrog tshuaj.
Kev tsis sib haum xeeb ntawm kev txaus siab nqe lus
Tsis muaj kev cuam tshuam ntawm kev txaus siab.
Cov ntaub ntawv
[1] ESD Alleva, Phau Ntawv Qhia rau kev saib xyuas thiab siv cov tsiaj kuaj, Ethology 103 (1997) 1072–1073.
[2] J. Bergh, T. Wiklund, B. Eriksen, E. Lidbrink, H. Lindman, P. Malmstrom, et al., Tailored fluorouracil, epirubicin, thiab cyclophosphamide piv nrog cov kab mob marrow-txhawb siab-dose chemotherapy raws li kev kho mob ntxiv. rau kev pheej hmoo mob qog noj ntshav mis: kev sim randomized. Scandinavian Breast Group 9401 kawm, Lancet 356 (2000) 1384–1391.
[3] A. Bernstein, L. Forrester, AD Reith, P. Dubreuil, R. Rottapel, The murine W/c-kit and steel loci and the control of hematopoiesis, Semin. Hematol. 28 (1991) 138–142.
[4] PJ Carey, Drug-inducedmyelosuppression: Diagnosis and Management, Drug Safe. 26 (2003) 691–706.
[5] F. Cheung, TCM: ua nyob rau hauv Tuam Tshoj, Xwm 480 (2011) S82–S83.
[6] EH Choi, HE Ok, Y. Yoon, BA Magnuson, MK Kim, HS Chun, Tiv thaiv cov nyhuv ntawm anthocyanin-nplua nuj extract los ntawm bilberry (Vaccinium myrtillus L.) tiv thaiv myelotoxicity induced by 5-uorouracil, BioFactors ( Oxf. England) 29 (2007) 55–65.
[7] J. Crawford, H. Ozer, R. Stoller, D. Johnson, G. Lyman, I. Tabbara, et al., Txo los ntawm granulocyte colony-stimulating factor ntawm kub taub hau thiab neutropenia induced los ntawm chemotherapy rau cov neeg mob me- cell lung cancer, N. Engl. J. Med. 325 (1991) 164–170.
[8] L. Dalby-Brown, H. Barrett, AK Landbo, AS Meyer, P. Molgaard, Synergistic antioxidative teebmeem ntawm alkamides, caffeic acid derivatives, thiab polysaccharide feem ntawm Echinacea purpurea nyob rau hauv vitro oxidation ntawm tib neeg tsawg-density lipoproteins. J. Agric. Khoom noj khoom haus Chem. 53 (2005) 9413–9423.
[9] WH de Vos tot Nederveen Cappel, HJ Meulenbeld, JH Kleibeuker, FM Nagengast,
FH Menko, G. Griffioen, et al., Ciaj sia nyob tom qab adjuvant 5-FU kev kho mob rau theem III mob qog noj ntshav nyob rau hauv hereditary nonpolyposis colorectal cancer, Int. J. Cancer 109 (2004) 468–471.
[10] LE Friberg, MO Karlsson, Mechanistic qauv raumyelosuppression, Tshawb nrhiav. New Drugs 21 (2003) 183–194.
[11] X. Geng, X. Tian, P. Tu, X. Pu, Neuroprotective effects ofechinacoside cov tshuajnyob rau hauv tus nas MPTP qauv ntawm Parkinson tus kab mob, Eur. J. Pharmacol. 564 (2007) 66–74.
[12] JY Han, RY Goh, SY Seo, TH Hwang, HC Kwon, SH Kim, et al., Cotransplantation of cord bloodhematopoieticstem cells thiab culture-expanded and GM-CSF-/SCF- transfected mesenchymal stem cells in SCID nas, J. Korean Med. Sci. 22 (2007) 242–247.
[13] EL Herzog, L. Chai, DS Krause, Plasticity of marrow-derived stem cells, Ntshav 102 (2003) 3483–3493.
[14] C. J., Li, H. W. Jin, J. W. Zhang, Y. Jiang, M. Zhao, et al., Metabolism of H.echinacoside cov tshuaj, zoo antioxidant, nyob rau hauv nas: cais thiab txheeb xyuas ntawm nws biliary metabolites, Tshuaj Metab. Dispos. 37 (2009) 431–438.
[15] .myelosuppressionhauv nas, Arch. Toxicol. 77 (2003) 285–290.
[16] R. Kuang, Y. Sun, W. Yuan, L. Li, X. Zheng, tiv thaiv los ntawmechinacoside cov tshuaj, ib qho ntawm phenylethanoid glycosides, ntawm H(2)O(2)-induced cytotoxicity hauv PC12 hlwb, Planta Med. 75 (2009) 1499–1504.
[17] JC Laurenz, M. Hadjisavas, GW Chovanec, FW Bazer,Myelosuppressionhauv npua (Sus scrofa): uteroferrin txo cov teebmeem myelosuppressive ntawm 5-uorouracil hauv cov npua hluas, Comp. Biochem. Physiol. Ib Physiol. 116 (1997) 369–377.
[18]
Li, F.Y. Yang, P. Zhu, W. Chen, D. Qi, X. Shi, et al.,Echinacosidetxhawb kev tsim cov pob txha los ntawm kev nce OPG/RANKL piv hauv MC3T3-E1 hlwb, Fitoterapia 83 (2012) 1443–1450.
[19] A. Lindner, D. Santilli, J. Hodgett, C. Nerlinger, Qhov cuam tshuam ntawm 5- fluorouracil ntawm lubhematopoieticsystem ntawm nas, Cancer Res. 20 (1960) 497–502.
[20] DB Longley, DP Harkin, PG Johnston, 5-uorouracil: mechanisms of action and clinical strategies, Nat. Rev. Cancer 3 (2003) 330–338.
[21] BR Luce, JW Singer, JM Weschler, CD Buckner, SH Sheingold, K. Shannon-Dorcy, thiab al., Recombinant human granulocyte-macrophage colony-stimulating factor after autologous bone marrow transplantation for lymphoid cancer: kev txheeb xyuas nyiaj txiag ntawm ib qho randomized, ob-dig muag, kev sim tshuaj placebo, PharmacoEconomics 6 (1994) 42–48.
[22] M. Malet-Martino, R. Martino, Kev tshawb fawb soj ntsuam ntawm peb qhov ncauj prodrugs ntawm 5-uorouracil (capecitabine, UFT, -1): kev tshuaj xyuas, Oncologist 7 (2002) 288–323 .
[23] S. Mishima, A. Nagai, S. Abdullah, C. Matsuda, T. Taketani, S. Kumakura, et al., Zoo ex vivo expansion ntawmhematopoieticqia hlwb siv osteoblast-differentiated mesenchymal qia hlwb yog CXCL12 nyob ntawm, Eur. J. Haematol. 84 (2010) 538–546.
[24] M. Mostafa, N. Nahar, M. Mosihuzzaman, T. Makhmoor, MI Choudhary, AU Rahman, Dawb radical scavenging phenylethanoid glycosides los ntawm Leucas indica Linn, Nat. Prod. Res. 21 (2007) 354–361.
[25] S. Numazawa, K. Sugihara, S. Miyake, H. Tomiyama, A. Hida, M. Hatsuno, et al., Muaj peev xwm koom nrog oxidative kev nyuaj siab hauv 5-uorouracil-mediatedmyelosuppressionhauv nas, Basic Clin. Pharmacol. Toxicol. 108 (2011) 40–45.
[26] DJ Prockop, Marrow stromal hlwb ua cov qia hlwb rau kev txuas ntxiv mus ntxiv ntawmnonhematopoieticcov ntaub so ntswg thiab raws li muaj peev xwm vectors rau kev kho noob caj noob ces, J. Cell. Biochem. (1998) 284–285.
[27] BD Sloley, LJ Urichuk, C. Tywin, RT Coutts, PK Pang, JJ Shan, Sib piv cov tshuaj tiv thaiv thiab antioxidants muaj peev xwm ntawm ntau hom Echinacea, J. Pharm. Pharmacol. 53 (2001) 849–857.
[28] C. Twelves, A. Wong, MP Nowacki, M. Abt, H. Burris III, A. Carrato, et al., Capecitabine as adjuvant treatment for stage III colon cancer, N. Engl. J. Med. 352 (2005) 2696–2704.
[29] M. Valtteri, A. Sorrentino, The mesenchymal stromal cell contribution to homeostasis, J. Cell. Physiol. 217 (2008) 296–300.
[30] L. van de Laar, PJ Coffer, AM Woltman, Regulation of dendritic cell development by GM-CSF: molecular control and implications for immune homeostasis and therapy, Blood 119 (2012) 3383–3393.
[31] JMBK Worley, M. Bloomsmith, K. Coleman, TL Koban, Phau ntawv thib yim ntawm Phau Ntawv Qhia Txog Kev Saib Xyuas thiab Kev Siv Cov Tsiaj Looj (2011); cuam tshuam rau kev tswj tus cwj pwm, Am. J. Primatol. 73 (2011) 98.
[32] Y. W. Li, T. Wen, YQ Li, tiv thaiv los ntawmechinacoside cov tshuajntawm carbon tetrachloride-induced hepatotoxicity hauv nas, Toxicology 232 (2007) 50–56.
[33] SF Xu, LM Yu, ZH Fan, Q. Wu, Y. Yuan, Y. Wei, et al., Txhim kho ginsenoside Rg1 ntawmhematopoieticua haujlwm hauv cyclophosphamide-inducedmyelosuppressionnas ,Eur. J. Pharmacol. 695 (2012) 7–12.
[34] Zhang, J. Xing, T. Ai, Wen, L. Guan, J. Zhao, tiv thaivechinacoside cov tshuajtiv thaiv mob ntsws mob los ntawm oleic acid nyob rau hauv nas, Free Radic. Res. 41 (2007) 798–805.
[35] AB Zhao, B. Yu, XL Wu, KJ Cao, EQ Li, QM Li, et al., Protective effects onmyelosuppressionnas kho los ntawm peb hom tshuaj classic suav tshuaj, Pharmacogn. Mag. 7 (2011) 133–140.
[36] M. Zhu, C. Lu, W. Li, Transient exposure toechinacoside cov tshuajNws txaus los qhib Trk signaling thiab tiv thaiv cov hlwb neuronal los ntawm rotenone, J. Neurochem. 124 (2013) 571–580.
