Chidule cha Ma Protein Factors Ophatikizidwa mu Hematopoietic Stem Cell Culture

Gwero: T&L Biotechnology nthawi yotulutsidwa: 2023-07-13

Mawu Oyamba
Mzaka zaposachedwa, Stem CellZakhala zikugwiritsidwa ntchito mochulukira muzachipatala. Komabe, kuchuluka ndi kuchuluka kwa ma stemcell m'thupi la munthu ndizochepa kwambiri, zomwe sizingakwaniritse zosowa zachipatala. Chifukwa chake, kukula kwa in vitro ndikukulitsa ma cell stem kwakhala kofunika kwambiri. Chifukwa chazifukwa zamakhalidwe ndi zamakono, chithandizo cha maselo oyambira amakumanabe ndi mavuto ambiri. Zolemba zapamwamba za maselo a hematopoietic stem / progenitor ndi mibadwo yosiyanasiyana ya maselo a magazi ndi omveka bwino, ndipo zizindikiro za phenotypic za maselo zimatha kusankhidwa mochulukira, kupatukana, ndipo ntchito zawo zingakhale zaulere, popanda kufunikira kwa njira zovuta zapansi monga "biological scaffolds" kwa mitsempha, mitsempha, ndi kupatsirana opaleshoni. Chifukwa chake, ndiye chitsanzo chabwino kwambiri pakukulitsa ndi kusiyanitsa kwa stem cell, komanso ndi yabwino kugwiritsa ntchito mwachindunji kuchipatala.
Ma cell a hematopoietic stem ali ndi kuchuluka kwakukulu kwa kudzikonzanso komanso kusiyanitsa kangapo. Amatha kupanga maselo onse okhwima a magazi, monga maselo ofiira a magazi, maselo oyera a magazi, mapulateleti, ndi ma lymphocyte, ndipo amatha kumanganso dongosolo lonse la hematopoietic. Kukula kwa in vitro kwa maselo amtundu wa hematopoietic kumafuna kukhalabe ndi luso la kudzikonzanso ndikulepheretsanso kusiyanitsa, ndikupangitsa kuti ikhale ukadaulo wovuta kwambiri. M'zaka zaposachedwa, zoyeserera zambiri zawonetsa kuti kusiyanitsa kwa maselo amtundu wa hematopoietic kumadalira ma cytokines. Tifotokoza mwachidule zinthu ndi zotsatira zake zomwe zimagwiritsidwa ntchito mu chikhalidwe cha ma cell a hematopoietic.

Stem cell factor (SCF)
Stem cell factor (SCF) ndi chinthu chomwe chimagwira ntchito ndikumangirira ndikuwonetsa tyrosine receptor c-Kit pamwamba pa ma HSC onse. Kusalongosoka kwa c-Kit kumabweretsa kuchepa kwa kuchuluka kwa HSC. Pakadali pano, pafupifupi mitundu yonse ya ma cytokine omwe amagwiritsidwa ntchito poyesa zachikhalidwe cha HSC ali ndi SCF. Kuphatikiza apo, onse a SCF ndi FL3 ali m'gulu la tyrosine kinase receptor TKR, lomwe limagwira ntchito pakukulitsa ma cell a hematopoietic. Pomangiriza ku TKR yeniyeni, SCF imatumiza zizindikiro ku maselo, kuyambitsa kugawanika koyambirira ndi kufalikira kwa maselo a progenitor, kulola kuti maselo ayambe kukula ndi kuletsa apoptosis akamaliza gawo la G0.

ThrombopoietinTpo）
TPO poyamba ankakhulupirira kuti ndi chinthu chokulirapo cha megakaryocytes, cha m'gulu la ma cytokines omwe amatha kupitiriza kukula, kusiyanitsa, kusasitsa, kugawa, ndi kupanga mapulateleti ogwira ntchito mu megakaryocytes. Ndilo chinthu chomwe chimakondedwa pakukulitsa megakaryocytes. M'zaka zaposachedwapa, zoyesera zatsimikizira kuti TPO imagwira ntchito yofunika kwambiri polimbikitsa maphunziro a HSC mu vitro, ndipo ikaphatikizidwa ndi ma cytokines ena, imatha kuonjezera chiwerengero cha mayunitsi opangira koloni ndi kukula kwa maselo a CD34 +. Makamaka muzophatikiza za FL3, zimatha kukhalabe ndikukula kwanthawi yayitali komanso kufalikira kwa zingwe zamagazi a CD34 +.

Interleukin-3 (IL-3)
Interleukin-3 (IL-3), yomwe imadziwikanso kuti masT cell kukula factor, ndi pleiotropic cytokine makamaka opangidwa ndi activated T lymphocytes, amene angathe kulimbikitsa kuchulukira ndi kusiyanitsa Pluripotent HSCs ndi osiyana mizera oriented maselo obadwa. Pambuyo pa IL-3 kuyambitsa heterodimerization ya stem cell cell receptors, imatha kumangirira ku mapuloteni ambiri otulutsa ma sign, monga njira ya Janus kinase signing transducer ndi transcriptal activator (JAK/STAT), potero kumalimbikitsa kutsika kwa ma siginecha ndikutenga nawo gawo pakuwongolera kukula kwa cell cell. IL-3 imathanso kuyambitsa njira ya extracellular regulated kinase (ERK) ndi c-jun aminotransferase (JNK) njira, kupangitsa kukula, kufalikira, ndi kupulumuka kwa maselo a tsinde.

Interleukin-6 (IL-6)
IL-6 ndi multidirectional cytokine yomwe imagwira ntchito yofunika kwambiri pachitetezo cha alendo poyendetsa mayankho a chitetezo chamthupi ndi kutupa. IL-6 imapangidwa ndi T cell, monocytes, fibroblasts, endothelial cell, ndi keratinocytes, ndipo imakhala ndi ntchito zambiri zamoyo. Itha kulimbikitsa kusiyana kwa ma cell a B ndi kupanga ma antibody, synergistically IL-3 imathandizira pakukula kwa megakaryocytes ndi kupanga mapulateleti, kulimbikitsa kuwonekera kwa mapuloteni owopsa m'chiwindi, ndikuwongolera kagayidwe ka mafupa. IL-6 imatumiza zizindikiro kudzera mu IL-6 receptor system, yomwe imakhala ndi maunyolo awiri: IL-6Rα ndi gp130. STAT3 ndiye molekyulu yotsimikizika pakusunga mawonekedwe osagwirizana a ma embryonic stem cell, pomwe IL-6 ndiye woyambitsa woyamba wa JAK/STAT3 njira yolumikizira.

LFLT3 ligand (FL)
FLT3 ligand ndi chinthu chakukula chomwe chimayang'anira kukula kwa ma cell a hematopoietic. FLT3 ligand imamangiriza ku maselo akuwonetsa tyrosine kinase receptor FLT3. FLT3 ligand palokha sichilimbikitsa kukula kwa maselo oyambirira a hematopoietic, koma m'malo mwake imapangitsa kukula ndi kusiyana ndi ma CSF ena ndi interleukins. Mosiyana ndi SCF, FLT3 ligand ilibe mphamvu pama cell a mast. Ma subtypes angapo a FLT3 ligand adziwika. Mawonekedwe akuluakulu a bioactive amakhazikika pama cell ngati gawo la extracellular la protein transmembrane (209a.). Isomeromu yomangidwa ndi nembanemba imatha kung'ambika ndi mapuloteni kuti apange ma isomers osungunuka a biologically.

Mowa wa FMS-Chinese 3
FMS-liketyrosine kinease 3 (FL3) yowonetsedwa kwambiri mu CD34 + CD38dim maselo amatumiza zizindikiro ku selo pomangirira ku tyrosine kinase active receptors (TKRs). FL3 imagwira ntchito pa HSC/HPC ndipo imagwiritsa ntchito hematopoietic regulation pomanga TKR pa cell. FL3 ndiyonso yofunika kwambiri yoyambitsa ma cell cell stimulating factor, yomwe ili ndi kulimbikitsa kwambiri pakukulitsidwa kwa in vitro kwa HSC/HPC. Itha kuletsa ma CD34+ stem cell kuti asasiyanitse pang'onopang'ono ndikuchepetsa HPC panthawi yakukulitsa kwa in vitro.

Kusintha kukula kwa chinthu- β
Kusintha kukula kwa chinthu- β, Kusintha kukula kwa chinthu- β 1 β 2 ndi β 3 subtypes za zinyama zimatulutsa zizindikiro kudzera mu cholandirira chomwecho, kuchititsa kuyankhidwa kofanana kwachilengedwe. Ndi ma cytokines omwe amagwira ntchito zambiri omwe amawongolera kukula kwa maselo, kukula, kusiyanitsa, ndi kuyenda, komanso kaphatikizidwe ndi kuyika kwa matrix owonjezera. Kusintha kukula kwa chinthu- β (TGF- β) kumapangidwa ndi maselo a mafupa a stromal.Amalepheretsa HSC / HPC oyambirira kulowa mu gawo la S, zomwe zimapangitsa kuti HSC / HPC yambiri ikhale mu gawo la G0.

Macrophage yotupa mapuloteni-1 α
Macrophage yotupa mapuloteni-1β (MIP-1β) ndi mdani wachilengedwe wa MIP-1α, omwe amatsagana nawo. Ikhoza kuthetsa zotsatira zolepheretsa za MIP-1α pa HSC/HPC yoyambirira ndikuletsa HSC kuti ibwerere kudera labata.

p38
P38, ngati molekyulu yozindikiritsa ya banja la mitogen activated protein kinase (MAPK), imalepheretsa kukula kwa ma HSC mu vitro pansi pazikhalidwe za normoxic. Kuyesera kwawonetsa kuti ma HSC akawonjezedwa ku media media yopanda seramu yokhala ndi TPO, SCF, ndi FL3, kupsinjika kwa okosijeni kumayambitsa p38 ndi p16, zomwe zimapangitsa kuchepa kwakukulu kwa ma HSC a mbewa.

Granulomacrophage colony-stimulating factor (GM-CSF)
Granulocyte macrophage colony-stimulating factor ndi mankhwala omwe amagwiritsidwa ntchito pochiza matenda osiyanasiyana a leukopenia kapena granulocytopenia. Maselo amakono olimbikitsa ma cell ndi granulocyte macrophage colony-stimulating factor (GM-CSF), zomwe sizimangowonjezera kuchuluka kwa maselo amtundu wa hematopoietic m'magazi ozungulira, komanso zimathandiza pa ntchito ya mtima ndi ntchito zina.

Granulocyte colony stimulating factor (G-CSF)
Zotsatira za granulocyte colony-stimulating factor nthawi zambiri zimaphatikizapo kuwonetseredwa kwa antigen, kupititsa patsogolo ntchito ya macrophage, ndi kulimbikitsa kukula kwa maselo amtundu wa hematopoietic. Granulocyte colony-stimulating factor ndi mphamvu yolimbikitsira maselo a m'mafupa, omwe amatha kulimbikitsa kukula kwa maselo amtundu wa autologous ndikuwalimbikitsa kuchoka ku mafupa kupita kumagazi ozungulira.

Erythropoietin (EPO)
Erythropoietin (EPO) ndiye chinthu chachikulu chothandizira kusiyanitsa kwa hematopoietic, chomwe chingalimbikitse kusiyanitsa kwa maselo ofiira a m'magazi kukhala maselo ofiira a magazi, kufulumizitsa kugawanika ndi kufalikira kwa maselo ofiira aang'ono, kulimbikitsa kaphatikizidwe ka hemoglobini, komanso kutenga gawo lofunikira pakuphunzira kusiyanasiyana kwa maselo ofiira a magazi.
Ma cell a hematopoietic stem ali ndi kuthekera kodzikonzanso komanso kusiyanitsa njira zambiri. Kuphatikiza kwa zinthu zosiyanasiyana kumakhala ndi zotsatira zosiyanasiyana pakukula kwa maselo amtundu wa hematopoietic. Kugwiritsiridwa ntchito kwa maselo a tsinde a hematopoietic kungapangitse kupanga maselo angapo, omwe mosakayikira amapereka malingaliro atsopano pakukula kwa maselo a NK. Pakalipano, ma cell a NK amatha kupangidwa kuchokera ku maselo a embryonic stem ndi iPSCs, koma maselo onse a embryonic stem cell ndi induced pluripotent stem cell ayenera kusinthidwa kukhala maselo a hematopoietic asanayambe kusiyanitsa mu maselo a NK. Chifukwa chake, ma cell a tsinde a hematopoietic amasewera mlatho wofunikira kwambiri pakuchita izi. Ubwino wa maselo a NK omwe amachokera ku maselo a tsinde ndikuti amatha kugwiritsidwa ntchito pofunidwa, kukhala ndi homogeneity yamphamvu, kumasulidwa kwa cytokine kochepa, ndi ntchito yakupha yamphamvu. Chifukwa chake, padakali chidwi chachikulu chofufuza ma cell a NK omwe amachokera ku maselo oyambira pamsika. Pali mabuku okhudza kulowetsedwa kwa tsinde mu maselo a NK. Timayang'ana makamaka pazinthu zomwe zatchulidwa m'mabuku.

1. Maselo a NK ochokera ku maselo amtundu wa embryonic

HESCs adasamutsidwa ku coculture ndi murine bone marrow stromal cell line M210-B4 mkati mwa sing'anga yomwe ili ndi RPMI 1640, 15% defined fetal bovine serum, 2 mM L-glutamine, 1% osafunikira amino acid, 1% penicillin / streptomyocin, ndi 0.1 sing'anga pamasiku 2 mM - zomwe zafotokozedwa m'mbuyomu 2 mM - kusintha kwa masiku atatu. Pambuyo pa masiku 17 mpaka 20, kuyimitsidwa kwa selo limodzi kunakonzedwa ndipo ma CD34 + CD45 + anali olekanitsidwa, monga momwe tafotokozera kale. Maselo odzipatula adasamutsidwira ku coculture yachiwiri ndi murine fetal chiwindi-derived stromal cell line AFT024 mkati mwa sing'anga yomwe ili ndi 1: 2 osakaniza a Dulbecco modified Eagle medium/Ham F12, 20% kutentha-inactivated human seramu AB, 2 mM L-glutamine, 1% penicillin / streptomL 5mL0mcinyomyo, sodium 5mL, sodium 5mL, streptom, streptomyom ethanolamine, 25μM -mercaptoethanol, 20 mg/mL ascorbic acid, interleukin-3, stem cell factor, IL-15, Fms-monga tyrosine kinase 3 ligand, ndi IL-7. Maselo amadyetsedwa ndi sing'anga yatsopano ndikusintha theka lapakati pamasiku 5 mpaka 6 aliwonse. Pambuyo pa masiku 30 mpaka 35 pachikhalidwe, ma cell adakololedwa, kusefedwa kudzera mu fyuluta ya 70-μm, ndikugwiritsidwa ntchito kuwunikanso.

2. Maselo a NK ochokera ku maselo amtundu wa pluripotent

Timakolola ma cell kwa masiku 18-21 a CD34+ CD45+ progenitor cell enrichment Zikhalidwe zama cell a NK tidatsitsimula ndi 0,5 ml ya sing'anga yokhala ndi cytokine masiku 4-5 aliwonse. Maselo okhwima a NK adayesedwa pa 28-35 masiku a chikhalidwe pa EL08-1D2.

T&L stem cell zokhudzana ndi ma cytokines & factor factor

Za T&L
T&L Biotechnology Ltd., yomwe idakhazikitsidwa mchaka cha 2011, imayang'ana kwambiri kafukufuku ndi chitukuko cha zida za GMP zopangira ma cell and gene therapy (CGT). Timadzipereka kupereka zinthu zodalirika ndi ntchito za sayansi ya moyo