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Ginseng cas ayaa loo isticmaali jiray daawada dhaqanka Aasiya boqollaal sano.Daraasaddan, waxaan ku qiimeynay awoodda afar nooc oo ginseng cas ah (ginseng cas Shiinaha, ginseng cas ee Kuuriya, ginseng cas ee Kuuriya, iyo ginseng cas ee Kuuriya C) oo ku koray gobollo kala duwan si loo joojiyo sameynta iyo koritaanka sambabada kansarka. burooyinka.Tijaabada benzo (a) pyrene (B (a) P) ayaa lagu sameeyay jiirarka A/J, iyo ginseng cas ee Kuuriya ayaa la ogaaday inuu yahay kan ugu waxtarka badan ee dhimista culeyska buro ee afarta nooc ee ginseng cas.Intaa waxaa dheer, waxaan ku falanqeynay waxyaabaha ku jira ginsenosides kala duwan (Rg1, Re, Rc, Rb2, Rb3, Rb1, Rh1, Rd, Rg3, Rh2, F1, Rk1 iyo Rg5) ee afar ginseng cas oo casaan ah oo la ogaaday in ginseng cas ee Kuuriya heerarka ugu sarreeya ee ginsenoside Rg3 (G-Rg3), oo soo jeedinaya in G-Rg3 laga yaabo inay door muhiim ah ka qaadato waxtarkeeda daaweynta.Shaqadani waxay muujinaysaa in G-Rg3 uu leeyahay bioavailability yara hooseeya.Si kastaba ha noqotee, markii G-Rg3 lala xiriiriyay P-gp inhibitor verapamil, qulqulka G-Rg3 ee unugyada Caco-2 ayaa hoos u dhacay, heerka nuugista mindhicirka ee G-Rg3 ayaa lagu kordhiyay qaabka jiirka, iyo G-Rg3 ayaa la kordhiyey.Unugyada Caco-2, qulqulka Rg3 ayaa hoos u dhacaya, heerka fiirsashada Rg3 ayaa hoos u dhacaya.G-Rg3 waxaa lagu kordhiyaa mindhicirka iyo balaasmaha, iyo awooddeeda ka hortagga burooyinka ayaa sidoo kale lagu wanaajiyey qaabka jiirka ee B (a) tumorigenesis-ku-soo-baxa.Waxaan sidoo kale ogaanay in G-Rg3 ay hoos u dhigtay B (a) Cytotoxicity P-induced iyo DNA-da ee unugyada sambabada bini'aadamka, waxayna dib u soo celiyeen muujinta iyo waxqabadka wejiga II enzymes iyada oo loo marayo waddada Nrf2, taas oo laga yaabo inay la xiriirto habka suurtagalka ah ee ficilka. ee G ka-hortagga -Rg3..Ku saabsan dhacdooyinka burooyinka sambabada.Daraasaddeenu waxay muujinaysaa doorka suurtagalka ah ee G-Rg3 ee lagu beegsanayo burooyinka sambabada ee moodooyinka jiirka.Bioavailability-ka afka ee ginsenoside-kan ayaa lagu wanaajiyey iyadoo la beegsanayo P-glycoprotein, taasoo u oggolaanaysa molecule inuu sameeyo saameynaha kansarka.
Nooca ugu badan ee kansarka sanbabada waa kansarka sanbabada unugyada aan-yare ahayn (NSCLC), kaas oo ah mid ka mid ah sababaha ugu horreeya ee dhimashada kansarka ee Shiinaha iyo Waqooyiga Ameerika1,2.Qodobka ugu muhiimsan ee kordhiya khatarta ah inuu ku dhaco kansarka sanbabada unugyada aan yareyn waa sigaar cabista.Qiiqa sigaarka wuxuu ka kooban yahay in ka badan 60 kansar, oo ay ku jiraan benzo(a) pyrene (B(a)P), nitrosamines, iyo isotopes radioactive oo ka yimaada qudhunka radon.3 B(a)P polycyclic aromatic hydrocarbons qiiqa.Marka la soo bandhigo B (a) P, cytochrome P450 waxay u beddeshaa B (a) P-7,8-dihydrodiol-9,10-epoxide (BPDE), kaas oo la falgala DNA si uu u sameeyo BPDE-DNA 4. Intaa waxaa dheer, kuwan ducts induce tuberigenesis sambabada ee jiirarka leh heerka burada iyo histopathology la mid ah burooyinka sambabada bini'aadamka5.Habkani waxa uu ka dhigayaa nooca kansarka sanbabada ee B (a) P-ku-abuuray nidaam ku habboon in lagu qiimeeyo xeryahooda leh sifooyinka kansarka ee suurtogalka ah.
Mid ka mid ah xeeladaha suurtagalka ah ee looga hortagayo horumarinta kansarka sanbabada ee kooxaha khatarta sare leh, gaar ahaan kuwa sigaarka cabba, waa isticmaalka kiimikada ka hortagga ah si loo xakameeyo horumarinta nabarrada neoplastic ee intraepithelial oo sidaas darteed looga hortago horumarkooda xiga ee malignant.Daraasadaha xayawaanku waxay muujinayaan in kala duwan oo kiimiko ka hortag ah ay waxtar leeyihiin6.Warbixinteenii hore7 waxay iftiimisay saamaynta wanaagsan ee ka hortagga ginseng-ka cas ee kansarka sanbabada.Dhirtan ayaa qarniyo badan loo isticmaali jiray dawo dhaqameedka Aasiya si ay u dheeraato nolosha iyo caafimaadka, waxaana la caddeeyey inay leedahay saamaynta antitumor8.
Qodobka firfircoon ee ginseng waa ginsenoside, kaas oo loo isticmaalo calaamado isku dhafan si loo qiimeeyo tayada ginseng-ka.Falanqaynta tirada ee soosaarayaasha ginseng cayriin waxay caadi ahaan ku lug leedahay isticmaalka dhowr ginsenosides, oo ay ku jiraan RK1, Rg1, F1, Re, Rb1, Rb2, Rb3, Rd, Rh1, Rh2, Rg3, Rg5, iyo Rc9,10.Ginsenosides waxay leeyihiin isticmaal caafimaad oo yar sababtoo ah bayolojigooda afka oo aad u liita11.Inkasta oo habka loogu talagalay bioavailability-ga liita uusan caddayn, qulqulka ginsenosides ee uu keeno P-glycoprotein (P-gp) 12 ayaa laga yaabaa inay tahay sababta.P-gp waa mid ka mid ah gaadiidleyda qulqulka leh ee ugu muhimsan ee ATP-ku-xidha-qaadista cajaladaha superfamily, kaas oo isticmaala tamarta ATP hydrolysis si ay u sii daayaan walxaha gudaha gudaha deegaanka dibadda.Gaadiidleyda P-gp ayaa caadi ahaan si weyn loogu qaybiyaa mindhicirka, kelyaha, beerka iyo xannibaadda maskaxda-maskaxda13.P-gp waxay door muhiim ah ka ciyaartaa nuugista mindhicirka, iyo joojinta P-gp waxay kordhisaa nuugista afka iyo helitaanka qaar ka mid ah dawooyinka ka hortagga kansarka12,14.Tusaalooyinka horjoogayaasha horay loogu isticmaalay suugaanta waa verapamil iyo cyclosporine A15.Shaqadani waxay ku lug leedahay samaynta nidaamka jiirka ee lagu baranayo B (a) kansarka sanbabada ee P si loo qiimeeyo awoodda kala duwan ee ginseng cas ee Shiinaha iyo Kuuriya si ay u saameeyaan malignancies.Soosaarayaasha ayaa si gaar ah loo falanqeeyay si loo aqoonsado ginsenosides gaar ah oo saameyn kara kansarka.Verapamil ayaa markaa loo isticmaalay in lagu beegsado P-gp iyo hagaajinta bioavailability afka iyo waxtarka daaweynta kansarka-bartilmaameedka ginsenosides.
Habka ay ginseng saponins u sameeyaan saamaynta daaweynta kansarka kansarka ayaa weli ah mid aan caddayn.Cilmi baaris ayaa muujisay in ginsenosides kala duwan ay yareyn karaan waxyeelada DNA-da ee ay keento kansarka iyada oo la yareynayo cadaadiska oksaydhka iyo firfircoonida wejiga II detoxification enzymes, taas oo ka hortagaysa dhaawaca unugyada.Glutathione S-transferase (GST) waa wejiga II enzyme caadiga ah kaas oo loo baahan yahay si loo yareeyo waxyeelada DNA ee uu keeno kansarka.Nukliyeerka erythroid-ka 2-xiriirka 2 (Nrf2) waa arrin muhiim ah oo qoraal ah oo nidaamisa redox homeostasis waxayna kicisaa muujinta wejiga II enzymes iyo jawaabaha antioxidant cytoprotective18.Daraasaddeenu waxay sidoo kale baartay saameynta ginsenosides ee la aqoonsaday si loo yareeyo B (a) cytotoxicity P-induced iyo BPDE-DNA, iyo sidoo kale kicinta wajiga II enzymes iyadoo la beddelayo habka Nrf2 ee unugyada sambabada caadiga ah.
Samaynta qaabka jiirka ee B(a) Kansarka P-ku-dhacey wuxuu la socdaa shaqadii hore5.Jaantuska 1A wuxuu muujinayaa naqshadaynta tijaabada ah ee daaweynta 20-usbuuc ee qaabka kansarka jiirka ee ay keentay B (a) P, biyaha (xakamaynta), soosaarka ginseng cas ee Shiinaha (CRG), soosaarka ginseng cas ee Kuuriya A (KRGA), iyo casaan Kuuriya ginseng.Soosaar B (KRGB) iyo Kooriyaan Ginseng Extract C (KRGC).Kadib 20 toddobaad oo daawaynta ginseng cas ah, jiirarka ayaa lagu allabaryay CO2 neefta.Jaantuska 1B wuxuu muujinayaa burooyinka sambabada ee makroscopic ee xayawaanka lagu daaweeyay noocyo kala duwan oo ginseng cas ah, iyo Jaantuska 1C wuxuu muujinayaa micrograph iftiin wakiil ah oo ah muunadda burada.Culayska buro ee xayawaanka la daweeyay ee KRGB (1.5 ± 0.35) ayaa ka hooseeya kan xayawaanka kantaroolka (0.82 ± 0.2, P <0.05), sida ku cad sawirka 1D.Celcelis ahaan heerka xakamaynta culeyska buro waxay ahayd 45%.Soosaarayaasha kale ee ginseng-ka cas ee la tijaabiyay ma muujin isbeddellada la taaban karo ee culeyska buro (P> 0.05).Wax saameyn ah oo muuqda laguma arag qaabka jiirka inta lagu jiro toddobaadyada 20 ee daaweynta ginseng cas, oo ay ku jiraan wax isbeddel ah oo ku yimaada miisaanka jidhka (xogta aan la muujin) iyo beerka ama kelyaha sunta (Jaantus 1E, F).
Soosaarka ginseng-ka cas wuxuu daaweeyaa horumarinta burada sambabada ee jiirarka A/J.(A) Naqshadaynta tijaabada ah.(B) Burooyinka sambabada waaweyn ee qaabka jiirka.Burooyinka waxaa lagu tilmaamaa fallaadho.ah: Kooxda ginseng cas ee Shiinaha.b: kooxda A ee ginseng cas ee Kuuriya.c: Kooxda ginseng cas ee Kuuriya B. d: Kooxda ginseng cas ee Kuuriya C. d: Kooxda xakamaynta.(C) Micrograph khafiif ah oo muujinaya buro sambabada.Weynaynta: 100. b: 400. (D) Culayska buro ee kooxda soosaarta ginseng cas.(E) Heerarka Plasma ee enzyme beerka ALT.(F) Heerarka Plasma ee enzyme kelyaha Cr.Xogta waxaa lagu muujiyay celcelis ahaan ± weecasho caadi ah.P <0.05.
Soosaarayaasha ginseng-ka cas ee lagu aqoonsaday daraasaddan waxaa lagu falanqeeyay ultra-waxqabadka dareeraha chromatography tandem mass spectrometry (UPLC-MS / MS) si loo qiyaaso ginsenosides soo socda: Rg1, Re, Rc, Rb2, Rb3, Rb1, Rh1, Rd, Rg3, Rh2, F1, Rk1 iyo Rg5.Xaaladaha UPLC iyo MS ee loo adeegsaday cabbiraadda falanqaynta ayaa lagu sifeeyay warbixin hore19.UPLC-MS/MS chromatograms ee afar nooc oo ginseng cas ah ayaa lagu muujiyay sawirka 2A.Waxaa jiray kala duwanaansho weyn oo ku saabsan wadarta guud ee ginsenoside, oo leh wadarta guud ee ginsenoside ee CRG (590.27 ± 41.28 μmol / L) (Jaantus 2B).Marka la qiimeeyo ginsenosides shakhsi ahaaneed (Jaantus 2C), KRGB waxay muujisay heerka ugu sarreeya ee G-Rg3 marka la barbardhigo ginsenosides kale (58.33 ± 3.81 μmol / L ee G-Rg3s iyo 41.56 ± 2.88 μmol / L ee G -Rg3r) ).nooca ginseng cas (P <0.001).G-Rg3 waxay u dhacdaa sida lammaane stereoisomers G-Rg3r iyo G-Rg3s, kuwaas oo ku kala duwan booska kooxda hydroxyl ee kaarboon 20 (Jaantus. 2D).Natiijooyinku waxay muujinayaan in G-Rg3r ama G-Rg3 laga yaabo inay leeyihiin awood ka hortag ah oo muhiim ah oo ku jirta qaabka B(a) P-ku-abuuray qaabka jiirka kansarka.
Waxyaabaha ay ka kooban tahay ginsenosides ee noocyada kala duwan ee ginseng cas.(A) UPLC-MS/MS chromatograms oo afar nooc oo ginseng cas ah.(B) Qiyaasta wadarta ginsenoside-ka ku jira waxyaabaha la soo bandhigay.(C) Ogaanshaha ginsenosides-ka gaarka ah ee soosaarayaasha calaamadeysan.(D) Qaab-dhismeedka ginsenoside stereoisomers G-Rg3r iyo G-Rg3s.Xogta waxaa lagu muujiyey inay tahay celceliska ± weecanaanta caadiga ah ee go'aaminta saddex geesoodka ah.***P <0.001.
Daraasadda UPLC-MS/MS waxay u baahday qiyaasta ginsenosides ee mindhicirka iyo shaybaarka dhiigga ka dib 20 toddobaad oo daaweyn ah.Daawaynta KRGB waxay muujisay joogitaanka 0.0063 ± 0.0005 μg/ml Rg5 ee dhiiga.Ma jiro ginsenosides haray oo la ogaaday, taas oo muujinaysa bioavailability afka liidata oo sidaas darteed hoos u dhigista ginsenosides kuwan.
Khadka unugga xiidanka ee adenocarcinoma Caco-2 waa qaab-dhismeedka iyo biochemical ahaan la mid ah unugyada epithelial mindhicirka mindhicirka, taas oo muujinaysa waxtarkeeda qiimaynta gaadiidka enterocyte ee ka gudba xannibaadda epithelial xiidmaha.Falanqayntan waxay ku salaysan tahay daraasad hore 20.Jaantusyada 3A,B,C,D,E,F waxay muujinayaan sawirada wakiillada gaadiidka transcellular ee G-Rg3r iyo G-Rg3 iyadoo la isticmaalayo moodalka Caco-2 monolayer.Gaadiidka transcellular ee G-Rg3r ama G-Rg3 ee guud ahaan Caco-2 monolayers laga soo bilaabo basolateral ilaa dhinaca apical (Pb-a) ayaa aad uga sarreeyay dhinaca apical ilaa dhinaca hoose (Pa-b).G-Rg3r, celceliska qiimaha Pa-b wuxuu ahaa 0.38 ± 0.06, kaas oo kordhay 0.73 ± 0.06 ka dib daaweynta 50 μmol / L verapamil iyo 1.14 ± 0.09 ka dib daaweynta 100 μmol / L verapamil (p <0.01) siday u kala horreeyaan;3A).Kormeerka G-Rg3 wuxuu raacay qaab la mid ah (Sawir 3B), natiijadu waxay muujisay in daaweynta verapamil ay kor u qaadday gaadiidka G-Rg3r iyo G-Rg3.Daaweynta Verapamil waxay sidoo kale keentay hoos u dhac weyn oo ku yimid celceliska Pb-a iyo G-Rg3r iyo G-Rg3s saamiga dheecaanka (Jaantus 3C, D, E, F), taas oo muujinaysa in daaweynta verapamil ay yareyso maadada ginsenoside ee unugyada Caco-2 efflux..
Gaadiidka transcellular ee G-Rg3 ee Caco-2 monolayers iyo nuugista mindhicirka ee baaritaanka dheecaanka jiirka.(A) Qiimaha Pa-b ee kooxda G-Rg3r ee Caco-2 monolayer.(B) Qiimaha Pa-b ee kooxaha G-Rg3s ee Caco-2 monolayer.(C) Qiimaha Pb ee kooxda G-Rg3r ee Caco-2 monolayer.(D) Qiimaha Pb ee kooxaha G-Rg3s ee Caco-2 monolayer.(E) Saamiga wax-soo-saarka ee kooxaha G-Rg3r ee ku jira hal-abuurka Caco-2.(F) Saamiga wax-soo-saarka kooxaha G-Rg3 ee hal-abuurka Caco-2.(G) Boqolkiiba nuugista mindhicirka ee G-Rg3r ee baadhitaanka fayraska ee jiirka.(H) Boqolkiiba nuugista mindhicirka ee G-Rg3 ee qiimaynta fayraska ee jiirka.Permeability iyo nuugista ayaa la barbardhigay iyada oo aan lagu darin verapamil.Xogta waxaa lagu muujiyey inay tahay celceliska ± weecis heerka shan tijaabo oo madaxbannaan.P <0.05, **P <0.01, ***P <0.001.
Iyadoo la raacayo shaqada hore20, qulqulka mindhicirka mindhicirka ee orthotopic ee jiirka ayaa la sameeyay si loo go'aamiyo in nuugista G-Rg3 ee mindhicirka ay korodho ka dib daaweynta verapamil.Jaantusyada 3G,H waxay muujinayaan qiimaynta fayraska wakiilka si loo qiimeeyo boqolkiiba nuugista mindhicirka ee G-Rg3r iyo G-Rg3 ee jiirka moodada kansarka inta lagu jiro xilliyada sare.Boqolkiiba bilowga ah ee qaadashada daciifka ah ee G-Rg3r ee ku dhawaad 10% waxay korodhay in ka badan 20% ka dib daawaynta 50 μM verapamil iyo in ka badan 25% ka dib daawaynta 100 μM verapamil.Sidoo kale, G-Rg3, oo lahaa kor u kaca ugu horreeya ee 10%, ayaa sidoo kale muujisay in ka badan 20% ka dib daaweynta 50 μM verapamil iyo ku dhawaad 30% ka dib daaweynta 100 μM verapamil, oo soo jeedinaysa in joojinta P-gp ee verapamil ay kor u qaaddo. xiidmaha G-nuugista Rg3 ee qaabka jiirka ee kansarka sanbabada.
Marka loo eego habka kor ku xusan, B(a) Jiirarka nooca kansarka ee P-ku-dhacey ayaa si aan kala sooc lahayn loo qaybiyey lix kooxood, sida ku cad Jaantuska 4A.Ma jiro miisaan lumis weyn ama calaamadaha kiliinikada ee sunta lagu arkay kooxda daaweynta G-Rg3 marka la barbardhigo kooxda kantaroolka (xogta aan la muujin).Kadib 20 toddobaad oo daaweyn ah, sambabada jiir kasta ayaa la ururiyey.Jaantuska 4B wuxuu muujinayaa burooyinka sambabada makroscopic ee jiirarka ee kooxaha daawaynta ee sare, iyo Jaantuska 4C wuxuu muujinayaa micrograph iftiin wakiil ah buro matale ah.Marka la eego culeyska buro ee koox kasta (Sawir 4D), qiyamka jiirarka lagu daaweeyay G-Rg3r iyo G-Rg3s waxay ahaayeen 0.75 ± 0.29 mm3 iyo 0.81 ± 0.30 mm3, siday u kala horreeyaan, halka qiyamka G-jiirka lagu daweeyay. leh -Rg3s waxay ahaayeen 1.63 siday u kala horreeyaan ± 0.40 mm3.kantaroolka jiirarka (p <0.001), taasoo muujinaysa in daawaynta G-Rg3 ay hoos u dhigtay culayska buro ee jiirarka.Maamulka verapamil ayaa sii wanaajiyay hoos u dhacan: qiyamka jiirarka verapamil + G-Rg3r ayaa hoos uga dhacay 0.75 ± 0.29 mm3 ilaa 0.33 ± 0.25 mm3 (p <0.01), iyo qiyamka verapamil + laga bilaabo 0.81 ± 0.30 .2.2 mm3 ee jiirarka G. -Rg3s-la daaweeyay (p <0.05), taas oo muujinaysa in verapamil ay kor u qaadi karto saameynta xannibaadda ee G-Rg3 ee tumorigenesis.Culayska burooyinka ma muujin farqi weyn oo u dhexeeya kooxda kantaroolka iyo kooxda verapamil, kooxda G-Rg3r iyo kooxda G-Rg3s, iyo kooxda verapamil + G-Rg3r iyo kooxda verapamil + G-Rg3s.Waxaa intaa dheer, ma jirin wax sun ah oo beerka ama kelyaha ah oo la xidhiidha daawaynta la qiimeeyay (Jaantuska 4E,F).
Culayska buro ka dib daaweynta G-Rg3 iyo balasmaha ama mindhicirka G-Rg3r iyo G-Rg3 ee kooxaha la tilmaamay.(A) Naqshadaynta tijaabada ah.(B) Burooyinka makroscopic ee qaabka jiirka.Burooyinka waxaa lagu tilmaamaa fallaadho.ah: G-Rg3r.b: G-Rg3s.c: G-Rg3r oo lagu daray verapamil.d: G-Rg3 oo lagu daray verapamil.d: Verapamil.e: xakamayn.(C) Mikrogaraafka indhaha ee buro marka la weyneeyo.Jawaab: 100x.b: 400X.(D) Saamaynta G-Rg3 + daawaynta verapamil ee culayska buro ee jiirarka A/J.(E) Heerarka Plasma ee enzyme beerka ALT.(F) Heerarka Plasma ee enzyme kelyaha Cr.(G) Heerarka Plasma ee G-Rg3r ama G-Rg3 ee kooxaha la tilmaamay.(H) Heerarka G-Rg3r ama G-Rg3s ee xiidmaha kooxaha la tilmaamay.Xogta waxaa lagu muujiyey inay tahay celceliska ± weecanaanta caadiga ah ee go'aaminta saddex geesoodka ah.P <0.05, **P <0.01, ***P <0.001.
Heerarka G-Rg3 ee jiirarka nooca kansarka ee uu keeno B (a) P waxaa lagu qiimeeyay UPLC-MS/MS ka dib muddo daawaynta 20-usbuuc ah iyadoo loo eegayo habka lagu qeexay qaybta Hababka.Jaantusyada 4G iyo H waxay muujinayaan plasma iyo mindhicirka G-Rg3, siday u kala horreeyaan.Heerarka Plasma G-Rg3r waxay ahaayeen 0.44 ± 0.32 μmol/L waxayna kordheen 1.17 ± 0.47 μmol/L iyadoo la raacayo maamulka isku-dhafka ah ee verapamil (p <0.001), halka heerarka G-Rg3r mindhicirku ay ahaayeen 0.53 ± 0.08 .µl.Marka lagu daro verapamil, g wuxuu kordhay 1.35 ± 0.13 μg/g (p <0.001).G-Rg3, natiijadu waxay raacday qaab la mid ah, taas oo muujinaysa in daaweynta verapamil ay kordhisay bioavailability afka ee G-Rg3 ee jiirarka A / J.
Qiimaynta jiritaanka unugyada unugyada ayaa loo isticmaalay si loo qiimeeyo cytotoxicity B(a) P iyo G-Rg3 ee unugyada hEL.Sitotoxicity-ka ay keentay B(a) P ee unugyada hEL waxa lagu muujiyay Jaantuska 5A, halka sifooyinka aan sunta ahayn ee G-Rg3r iyo G-Rg3 lagu muujiyay jaantusyada 5A iyo 5B.5B, C. Si loo qiimeeyo saamaynta cytoprotective ee G-Rg3, B (a) P ayaa la isku daray iyada oo la raacayo uruurinta kala duwan ee G-Rg3r ama G-Rg3 ee unugyada hEL.Sida ku cad Jaantuska 5D, G-Rg3r ee xajmiyada 5 μM, 10 μM, iyo 20 μM ayaa dib loo soo celiyay ku noolaanshaha unugyada ilaa 58.3%, 79.3%, iyo 77.3%, siday u kala horreeyaan.Natiijooyin la mid ah ayaa sidoo kale lagu arki karaa kooxda G-Rg3s.Marka qiyaasta G-Rg3s ay ahaayeen 5 µM, 10 µM iyo 20 µM, awoodda unugga ayaa dib loogu soo celiyay 58.3%, 72.7% iyo 76.7%, siday u kala horreeyaan (Jaantuska 5E).).Joogitaanka xirmooyinka BPDE-DNA waxaa lagu cabbiray iyadoo la isticmaalayo xirmo ELISA ah.Natiijooyinkayagu waxay muujiyeen in heerarka dulsaarka BPDE-DNA lagu kordhiyay kooxda B (a) P-daaweeyay marka la barbar dhigo kooxda kantaroolka, laakiin marka la barbar dhigo wada-daaweynta G-Rg3, BPDE-DNA heerarka dulsaarka ee kooxda B (a) P. B ee kooxda la daaweeyay, heerarka dulsaarka DNA ayaa si weyn hoos loogu dhigay.Natiijooyinka daaweynta B (a) P oo keliya ayaa lagu muujiyay sawirka 5F (1.87 ± 0.33 vs. 3.77 ± 0.42 ee G-Rg3r, 1.93 ± 0.48 vs. 3.77 ± 0.42 ee G -Rg3s, p <0.001).
Jiritaanka unugyada iyo BPDE-DNA samaynta ku-meel-gaadhka ah ee unugyada hEL ee lagu daweeyay G-Rg3 iyo B(a) P.(A) Awood u yeelashada unugyada hEL ee lagu daweeyay B(a)P.(B) Awood u yeelashada unugyada hEL ee lagu daweeyay G-Rg3r.(C) Jiritaanka unugyada hEL ee lagu daweeyay G-Rg3.(D) Awood u yeelashada unugyada hEL ee lagu daaweeyay B(a)P iyo G-Rg3r.(E) Awood u yeelashada unugyada hEL ee lagu daweeyay B(a)P iyo G-Rg3.(F) Heerarka BPDE-DNA ee ku xidhan unugyada hEL ee lagu daweeyay B(a) P iyo G-Rg3.Xogta waxaa lagu muujiyey inay tahay celceliska ± weecanaanta caadiga ah ee go'aaminta saddex geesoodka ah.P <0.05, **P <0.01, ***P <0.001.
Muujinta ensaymka GST ayaa la ogaaday ka dib wada-daawayn 10 μM B(a) P iyo 10 μM G-Rg3r ama G-Rg3s.Natiijooyinkayagu waxay muujiyeen in B (a) P la xakameeyey muujinta GST (59.7 ± 8.2% ee kooxda G-Rg3r iyo 39 ± 4.5% ee kooxda G-Rg3s), iyo B (a) P waxay la xiriirtay midkood G-Rg3r , ama wata G-Rg3r, ama wata G-Rg3r.Wada-daawaynta G-Rg3s waxay soo celisay muujinta GST.GST muujinta (103.7 ± 15.5% ee kooxda G-Rg3r iyo 110 ± 11.1% ee kooxda G-Rg3s, p <0.05 iyo p <0.001, siday u kala horreeyaan, Fig. 6A, B, iyo C).Dhaqdhaqaaqa GST waxa lagu qiimeeyay iyada oo la isticmaalayo qalab qiimayn hawleed.Natiijooyinkayagu waxay muujiyeen in kooxda daawaynta isku-dhafka ah ay leeyihiin hawlo GST sare ah marka loo eego kooxda B (a) P kaliya (96.3 ± 6.6% vs. 35.7 ± 7.8% ee kooxda G-Rg3r vs. 92.3 ± 6.5 ee kooxda G-Rg3r ).% vs 35.7 ± 7.8% ee kooxda G-Rg3s, p <0.001, Jaantuska 6D).
Muujinta GST iyo Nrf2 ee unugyada hEL ee lagu daweeyay B(a) P iyo G-Rg3.(A) Ogaanshaha odhaahda GST ee ay reer galbeedku tirtirayaan.(B) Tibaaxaha tirada GST ee unugyada heel ee lagu daweeyay B(a) P iyo G-Rg3r.(C) Tirakoobka GST ee unugyada hEL ee lagu daweeyay B(a) P iyo G-Rg3s.(D) Hawlaha GST ee unugyada hEL ee lagu daweeyay B(a) P iyo G-Rg3.(E) Ogaanshaha Nrf2 muujinta reer galbeedka.(F) Tirakoobka Nrf2 ee unugyada hEL ee lagu daweeyay B(a) P iyo G-Rg3r.(G) Muujinta tirada Nrf2 ee unugyada hEL ee lagu daweeyay B(a) P iyo G-Rg3s.Xogta waxaa lagu muujiyey inay tahay celceliska ± weecanaanta caadiga ah ee go'aaminta saddex geesoodka ah.P <0.05, **P <0.01, ***P <0.001.
Si loo caddeeyo waddooyinka ku lug leh G-Rg3-dhexdhexaadinta xakamaynta B (a) P-induced tumorigenesis, muujinta Nrf2 waxaa lagu qiimeeyay xannibaadda Galbeedka.Sida lagu muujiyey jaantusyada 6E, F, G, marka la barbardhigo kooxda kantaroolka, kaliya heerka Nrf2 ee kooxda daaweynta B (a) P ayaa hoos loo dhigay;si kastaba ha ahaatee, marka la barbardhigo kooxda daaweynta B (a) P, B (a) Nrf2 heerarka kooxda PG-Rg3 ayaa la kordhiyay (106 ± 9.5% ee G-Rg3r vs. 51.3 ± 6.8%, 117 ± 6. 2% G-Rg3r vs. 41 ± 9.8% ee G-Rg3s, p <0.01).
Waxaan xaqiijinay doorka ka hortagga Nrf2 anagoo xakameynayna Nrf2 muujinta iyadoo la adeegsanayo RNA (siRNA) faragelin yar oo gaar ah.garaacista Nrf2 waxaa xaqiijiyay dhufashada Galbeedka (Jaantus. 7A,B).Sida lagu muujiyey jaantusyada 7C, D, wada-daawaynta unugyada hEL ee B (a) P iyo G-Rg3 waxay keentay hoos u dhac ku yimid tirada BPDE-DNA adcts (1.47 ± 0.21) marka la barbar dhigo daaweynta B (a) P keligiis kooxda siRNA ee xakamayntaSi kastaba ha noqotee, saameynta xannibaadda ee G-Rg3 ee samaynta BPDE-DNA ayaa la tirtiray garaacista Nrf2.Kooxda siNrf2, ma jirin farqi weyn oo u dhexeeya samaynta BPDE-DNA ee u dhexeeya B (a) P iyo G-Rg3 wada-daaweynta iyo B (a) daaweynta kaliya (3.0 ± 0.21 ee G-Rg3r vs. 3.56 ± 0.32 ).loogu talagalay G-Rg3r oo ka soo horjeeda 3.6 ee G-Rg3s oo ka soo horjeeda ± 0.45 oo ka soo horjeeda 4.0 ± 0.37, p> 0.05).
Saamaynta garaacista Nrf2 ee samaynta BPDE-DNA ee isku xidhka unugyada hEL.(A) garaacista Nrf2 waxaa xaqiijiyay burburka reer galbeedka.(B) Qiyaasta xoojinta band Nrf2.(C) Saamaynta Nrf2 garaacista heerarka BPDE-DNA ee unugyada hEL ee lagu daweeyay B (a) P iyo G-Rg3r.(D) Saamaynta Nrf2 garaacista heerarka BPDE-DNA ee unugyada hEL ee lagu daweeyay B (a) P iyo G-Rg3.Xogta waxaa lagu muujiyey inay tahay celceliska ± weecanaanta caadiga ah ee go'aaminta saddex geesoodka ah.P <0.05, **P <0.01, ***P <0.001.
Daraasadani waxay qiimeysay saamaynta kahortaga ee noocyada kala duwan ee ginseng-ka cas ee qaabka jiirka ee B (a) kansarka sanbabada ee P, iyo daaweynta KRGB waxay si weyn hoos ugu dhigtay culeyska buro.Iyadoo la tixgelinayo in G-Rg3 uu leeyahay nuxurka ugu sarreeya ee laga soosaaray ginseng-kan, doorka muhiimka ah ee ginsenoside this ee ka hortagga burooyinka ayaa la bartay.Labada G-Rg3r iyo G-Rg3 (laba epimers ee G-Rg3) waxay si weyn u yareeyeen culeyska buro ee qaabka jiirka ee B (a) kansarka uu keeno P.G-Rg3r iyo G-Rg3 waxay sameeyaan saameyn liddi ku ah kansarka iyaga oo keenaya apoptosis ee unugyada burooyinka21, xakameynaya koritaanka burada22, xiritaanka wareegga unugyada23 iyo saameynaya angiogenesis24.G-Rg3 ayaa sidoo kale lagu muujiyay in ay joojiso metastasis-ka gacanta25, iyo awoodda G-Rg3 si kor loogu qaado saameynta kemotherabi iyo daaweynta shucaaca ayaa la diiwaangeliyay26,27.Poon et al waxay muujiyeen in daaweynta G-Rg3 ay yarayn karto saameynta genotoxic ee B (a) P28.Daraasadani waxay muujinaysaa awoodda daaweynta ee G-Rg3 ee lagu beegsanayo molecules carcinogenic deegaanka iyo ka hortagga kansarka.
Inkasta oo ay awood u leeyihiin inay ka hortagaan, liidata bioavailability afka ee ginsenosides ayaa caqabad ku ah isticmaalka kiliinikada ee unugyadan.Falanqaynta farmacokinetic ee maamulka afka ee ginsenosides ee jiirka ayaa muujisay in bioavailability-geeda ay wali ka yar tahay 5%29.Tijaabooyinku waxay muujiyeen in ka dib muddada daaweynta 20-usbuuc, kaliya heerarka dhiigga ee Rg5 ayaa hoos u dhacay.Inkasta oo habka aasaasiga ah ee bioavailability liidata uu weli yahay in la caddeeyo, P-gp waxaa loo maleynayaa in ay ku lug leedahay qulqulka ginsenosides.Shaqadani waxay muujisay markii ugu horeysay in maamulka verapamil, xannibaadaha P-gp, uu kordhiyo bioavailability afka ee G-Rg3r iyo G-Rg3s.Sidaa darteed, natiijadani waxay soo jeedinaysaa in G-Rg3r iyo G-Rg3s ay u dhaqmaan sida substrates ee P-gp si loo xakameeyo qulqulka.
Shaqadani waxay muujinaysaa in daaweynta isku dhafan ee verapamil ay kordhiso bioavailability afka ee G-Rg3 ee qaabka jiirka ee kansarka sanbabada.Natiijooyinkan waxaa taageeraya kordhinta gaadiidka transcellular ee mindhicirka ee G-Rg3 ee xannibaadda P-gp, taas oo kordhinaysa nuugista.Qiimaynta unugyada Caco2 waxay muujisay in daaweynta verapamil ay hoos u dhigtay qulqulka G-Rg3r iyo G-Rg3s iyada oo hagaajinaysa qulqulka xuubka.Daraasad uu sameeyay Yang et al.Daraasaduhu waxay muujiyeen in daaweynta cyclosporine A (kale oo P-gp blocker) ay kordhiso bioavailability of ginsenoside Rh2 laga bilaabo qiimaha aasaasiga ah ee 1%20 ilaa in ka badan 30%.Xeryahooda Ginsenosides K iyo Rg1 ayaa sidoo kale muujiyay natiijooyin isku mid ah30,31.Marka verapamil iyo cyclosporin A la wada maamulo, qulqulka xarunta K ee unugyada Caco-2 ayaa si weyn hoos loogu dhigay min 26.6 ilaa in ka yar 3, halka heerarka intracellular ay kordheen 40-laab30.Joogitaanka verapamil, heerarka Rg1 ayaa kordhay unugyada epithelial sambabada jiirka, taas oo soo jeedinaysa doorka P-gp ee ginsenoside efflux, sida lagu muujiyay Meng et al.31.Si kastaba ha ahaatee, verapamil kuma yeelan saameyn isku mid ah qulqulka qaar ka mid ah ginsenosides (sida Rg1, F1, Rh1 iyo Re), taas oo muujinaysa in aysan saameyn ku yeelan substrates P-gp, sida lagu muujiyay Liang et al.32 .Indho-indhayntani waxa laga yaabaa inay la xidhiidho ka-qaybgalka gaadiidleyda kale iyo qaababka kale ee ginsenoside.
Habka ka hortagga G-Rg3 ee kansarka ma cadda.Daraasadihii hore waxay muujiyeen in G-Rg3 ay ka hortagto waxyeelada DNA-da iyo apoptosis iyada oo la yareynayo cadaadiska oksaydhka iyo caabuqa16,33, taas oo noqon karta habka asaasiga ah ee looga hortagayo B (a) P-induced tumorigenesis.Warbixinnada qaar ayaa tilmaamaya in genotoxicity ay keentay B(a) P lagu yarayn karo iyadoo la beddelayo wejiga II enzymes si loo sameeyo BPDE-DNA34.GST waa wejiga II ee enzyme-ka caadiga ah kaas oo xannibaya samaynta BPDE-DNA iyada oo kor u qaadaysa ku-xidhka GSH ilaa BPDE, taas oo yaraynaysa dhaawaca DNA ee B(a) P35.Natiijooyinkayagu waxay muujinayaan in daawaynta G-Rg3 ay yarayso B (a) Cytotoxicity P-induced iyo samaynta BPDE-DNA ee unugyada hEL waxayna dib u soo celisaa muujinta GST iyo dhaqdhaqaaqa gudaha vitro.Si kastaba ha noqotee, saameyntani waxay ka maqnayd maqnaanshaha Nrf2, oo soo jeedinaysa in G-Rg3 ay keento saamaynta cytoprotective iyada oo loo marayo waddada Nrf2.Nrf2 waa arrin weyn oo qoraal ah oo loogu talagalay wejiga II detoxification enzymes kaas oo kor u qaada nadiifinta xenobiotics36.Dhaqdhaqaaqa dariiqa Nrf2 waxay keentaa cytoprotection waxayna yaraynaysaa dhaawaca nudaha37.Waxaa intaa dheer, warbixinno dhowr ah ayaa taageeray doorka Nrf2 oo ah buro xakameynaya kansarka38.Daraasaddeenu waxay muujinaysaa in soo-saarka dariiqa Nrf2 ee G-Rg3 ay ka ciyaaraan door muhiim ah oo xakameynaya B (a) P-induced genotoxicity taasoo keenaysa B (a) P detoxification iyada oo la kicinayo wajiga II enzymes, taas oo joojinaysa habka tumorigenesis.
Shaqadeenu waxay muujinaysaa suurtagalnimada ginseng cas ee ka hortagga B (a) P-ku dhaca kansarka sanbabada ee jiirarka iyada oo loo marayo ka qaybgalka muhiimka ah ee ginsenoside G-Rg3.Biyo-helidda afka ee liidata ee molecule-kan ayaa caqabad ku ah codsigiisa caafimaad.Si kastaba ha ahaatee, daraasaddan ayaa muujinaysa markii ugu horeysay in G-Rg3 uu yahay substrate-ka P-gp, iyo maamulka P-gp inhibitor wuxuu kordhiyaa bioavailability ee G-Rg3 in vitro iyo in vivo.G-Rg3 waxay yaraynaysaa cytotoxicity B (a) P-ku-takoorka iyadoo nidaamisa dariiqa Nrf2, taas oo noqon karta hab suurtagal ah oo loogu talagalay hawsheeda ka hortagga.Daraasaddeenu waxay xaqiijinaysaa awoodda ginsenoside G-Rg3 ee ka hortagga iyo daaweynta kansarka sanbabada.
Jiirarka A/J oo lix toddobaad jir ah (20 ± 1 g) iyo 7-usbuuc jira jiirka Wistar ee lab ah (250 ± 20 g) ayaa laga helay Shaybaadhka Jackson (Bar Harbor, USA) iyo Machadka Wuhan ee Xayawaanka.Jaamacadda (Wuhan, Shiinaha).Xarunta Dhaqanka Nooca Shiinaha (Wuhan, Shiinaha) waxay na siisay unugyada Caco-2 iyo hEL.Sigma-Aldrich (St. Louis, USA) waa isha B(a) P iyo tricaprine.Ginsenosides la safeeyey G-Rg3r iyo G-Rg3s, dimethyl sulfoxide (DMSO), CellTiter-96 xirmada faafinta faafinta (MTS), verapamil, dhexdhexaad dhexdhexaad ah ugu yar (MEM), iyo serum bovine uurjiifka (FBS) ayaa laga soo iibsaday Chengdu Must Bio-Technology .Co., Ltd.(Chengdu, Shiinaha).Xirmada yar ee QIAamp DNA iyo BPDE-DNA xirmada ELISA waxaa laga soo iibsaday Qiagen (Stanford, CA, USA) iyo Cell Biolabs (San Diego, CA, USA).Qalabka qiimaynta dhaqdhaqaaqa GST iyo wadarta guud ee xidhmada qiimaynta borotiinka (habka caadiga ah ee BCA) ayaa laga soo iibsaday Solarbio (Beijing, Shiinaha).Dhammaan soosaarayaasha ginseng cas waxaa lagu kaydiyaa Mingyu Laboratory 7. Jaamacadda Baabtiisaha Hong Kong (Hong Kong, China) iyo Korea Cancer Center (Seoul, Korea) waa ilo ganacsi oo laga soosaaray CRG iyo noocyo kala duwan oo ginseng casaan ah oo asal ahaan Kuuriyaan ah (oo ay ku jiraan KRGA, KRGB) iyo KRGC).Ginseng cas waxaa laga sameeyaa xididdada ginseng cusub oo 6 jir ah.Soosaarka ginseng-ka cas waxaa lagu helaa iyadoo lagu dhaqo ginseng biyo saddex jeer, ka dibna xoogga la saaro soosaarka aqueous, iyo ugu dambeyntii qalajinta heerkulka hooseeya si loo helo budada ginseng.Unugyada difaaca (anti-Nrf2, anti-GST, iyo β-actin), horseradish peroxidase-conjugated anti-bakayle immunoglobulin G (IgG), reagent gudbinta, xakamaynta siRNA, iyo Nrf2 siRNA ayaa laga soo iibsaday Santa Cruz Biotechnology (Santa Cruz, CA) .), USA).
Unugyada Caco2 iyo hEL waxaa lagu beeray 100 mm2 suxuunta dhaqanka unugga oo leh MEM ka kooban 10% FBS at 37 °C jawi qoyan oo ah 5% CO2.Si loo go'aamiyo saameynta xaaladaha daawaynta, unugyada hEL waxaa lagu dhajiyay qiyaaso kala duwan oo B (a) P iyo G-Rg3 gudaha MEM ee 48 h.Unugyada waa la sii baadhi karaa ama la ururin karaa si loo diyaariyo wax-soosaar unug-la'aan ah.
Dhammaan tijaabooyinka waxaa ansixiyay Guddiga Anshaxa Xayawaanka Tijaabada ee Kulliyadda Caafimaadka ee Tongji, Jaamacadda Huazhong ee Sayniska iyo Teknolojiyada (Oggolaanshaha No. 2019; Diiwaangelinta No. 4587TH).Dhammaan tijaabooyinka waxaa loo sameeyay si waafaqsan tilmaamaha iyo xeerarka ku habboon, daraasadda waxaa lagu sameeyay si waafaqsan Cilmi-baarista Xayawaanka: Warbixinta Tijaabada Vivo (ARRIVE).Jiirarka A/J oo siddeed usbuuc jira ayaa markii ugu horreysay si xiniinyaha ah loogu duray B(a) P ee xalka tricaprine (100 mg/kg, 0.2 ml).Toddobaad ka dib, jiirarka ayaa si aan kala sooc lahayn loo qaybiyay kooxo kontorool ah iyo kooxo daaweyn oo kala duwan, 15 jiir koox kasta, waxaana la qaawiyay hal mar maalintii.Ka dib 20 toddobaad oo daaweyn ah, xoolaha ayaa lagu sadqeeyay CO2 neefta.Sambabada ayaa la ururiyay oo la hagaajiyay 24 saacadood.Tirada burooyinka dusha sare iyo cabbirada buro gaar ah ayaa lagu qiyaasay sambaba kasta iyadoo la hoos marinayo mikroskoob kala jeexaya.Qiyaasta mugga burada (V) ayaa la xisaabiyay iyadoo la adeegsanayo tibaaxaha soo socda: V (mm3) = 4/3πr3, halkaasoo r ay tahay dhexroorka burada.Wadarta saafiga ah ee dhammaan xajmiyada buro ee sambabada jiirarka ayaa ka dhigan wadarta mugga burada, iyo celceliska wadarta mugga buro ee koox kastaa waxay u taagan tahay culayska buro.Tijaabooyin dhiig oo dhan iyo mindhicirrada ayaa la ururiyey oo lagu kaydiyey -80°C si loo go'aamiyo UPLC-MS/MS.Serum waa la ururiyay waxaana la isticmaalay kimistari otomaatig ah falanqeeye si loo falanqeeyo alanine aminotransferase (ALT) iyo serum creatinine (Cr) heerarka si loo qiimeeyo shaqada beerka iyo kelyaha.
Muunado la ururiyey ayaa laga soo saaray kaydinta qaboojiyaha, waa la dhalaalay, la miisaamay, waxaana la geliyay tubooyinka sida kor lagu sharaxay.Tan waxaa lagu daray 0.5 μM phlorizin (heerka gudaha) oo ku jira 0.8 ml oo methanol ah.Unugyada ayaa markaa isku mid ah iyadoo la isticmaalayo Tissue-Tearor ka dibna isku-dhafka ayaa loo wareejiyay tuubo 1.5 ml oo microcentrifuge ah.Isku darka waxaa lagu meeleeyay 15500 rpm 15 daqiiqo.Ka dib markaad ka saarto 1.0 ml oo ah supernatant, ku qallaji nitrogen.Laba boqol oo mikroliter oo methanol ah ayaa loo isticmaalay soo kabashada.Dhiigga waxaa lagu ururiyaa oo lagu farsameeyaa hal xariiq waxaana loo isticmaalaa tixraac ahaan dhammaan cabbirada.
Taarikada 24-well Transwell ayaa lagu beeray 1.0 × 105 Caco-2 unug kasta si loo qiimeeyo kor u qaadida suurtagalka ah ee gaadiidka G-Rg3 iyadoo lagu daro verapamil.3 toddobaad oo dhaqan ka dib, unugyada waxaa lagu dhaqay HBSS waxaana lagu sii daayay 37°C.400 μL oo ah 10 μM G-Rg3 (G-Rg3r, G-Rg3s, ama isku dar ah 50 ama 100 μM verapamil) ayaa lagu duray dhinaca basolateral ama apical ee monolayer, iyo 600 μL ee xalka HBSS ayaa lagu daray kan kale dhinac.Ururi 100 µl oo dhexdhexaad ah dhaqameed wakhtiyada la cayimay (0, 15, 30, 45, 60, 90 iyo 120 minutes) kuna dar 100 µl HBSS si aad muggaan u samayso.Tusaalooyinka waxaa lagu kaydiyay -4 °C ilaa laga ogaado UPLC-MS/MS.Odhaahda Papp = dQ/(dT × A × C0) waxa loo istcmaalaa in lagu qiyaaso jiho aan toos ahayn oo muuqda iyo cagsigeeda (Pa-b iyo Pb-a, siday u kala horreeyaan);dQ / dT waa isbeddelka fiirsashada, A (0.6 cm2) waa aagga dusha sare ee monolayer, iyo C0 waa diiradda deeq-bixiyeyaasha bilowga ah.Saamiga qulqulka waxaa loo xisaabiyaa sida Pb-a/Pa-b, taasoo ka dhigan heerka daadad ee daawada daraasadda.
Jiirka labka ah ee Wistar ayaa la soomay 24 saacadood, waxa ay cabbeen biyo kaliya, waxaana lagu suuxiyay cirbad xididka ah oo ah 3.5% pentobarbital.Tuubada silikoon ee la geliyey waxay leedahay dhamaadka duodenumka sida albaabka laga soo galo iyo dhamaadka ileum sida bixida.Isticmaal bamka peristaltic si aad ugu shubto meesha laga soo galo 10 µM G-Rg3r ama G-Rg3s ee HBSS isotonic ah heerka socodka 0.1 ml/daqiiqo.Saamaynta verapamil waxaa lagu qiimeeyay iyadoo lagu daray 50 μM ama 100 μM ee xarunta ilaa 10 μM G-Rg3r ama G-Rg3s.UPLC-MS/MS waxaa lagu sameeyay faleebo laga soosaaray waqtiyada 60, 90, 120, iyo 150 daqiiqo kadib bilawga dhiig karka.Boqolkiiba nuugista waxaa lagu qiyaasaa qaacidada % nuugista = (1 - Cout/Cin) × 100%;fiirsashada G-Rg3 ee meesha laga soo galo iyo meesha laga soo galoba waxa muujiyay Cout iyo Cin, siday u kala horreeyaan.
Unugyada hEL waxaa lagu beeray taarikada 96-ceel cufnaanta 1 × 104 unugyadiiba waxaana lagu daaweeyay B(a) P (0, 1, 5, 10, 20, 30, 40 μM) ama G-Rg3 oo lagu milmay DMSO .Dawooyinka ayaa markaa lagu milmay dhexdhexaad dhaqameed ilaa xadyo kala duwan (0, 1, 2, 5, 10, 20 μM) 48 saacadood.Iyada oo la adeegsanayo xirmada baaritaanka MTS ee ganacsi ahaan loo heli karo, unugyadu waxaa la marsiiyay hab-maamuuska caadiga ah ka dibna lagu cabbiray iyadoo la isticmaalayo akhristaha microplate oo ah 490 nm.Heerka jiritaanka unugga ee kooxaha lala daweeyay B(a) P (10 μM) iyo G-Rg3 (0, 1, 5, 10, 20 μM) ayaa la qiimeeyay iyadoo la raacayo habka kor ku xusan waxaana la barbardhigay kooxda aan la daweyn.
Unugyada hEL waxaa lagu beeray taarikada 6-ceel cufnaanta 1 × 105 unug/si fiican waxaana lagu daaweeyay 10 μMB (a) P iyadoo ay jirto ama maqnaanshaha 10 μM G-Rg3.Kadib 48 saacadood oo daaweyn ah, DNA ayaa laga soo saaray unugyada hEL iyadoo la adeegsanayo QIAamp DNA Mini Kit sida uu qabo borotokoolka soo saaraha.Sameynta xirmooyinka BPDE-DNA waxaa lagu ogaadey iyadoo la adeegsanayo xirmada ELISA ee BPDE-DNA.Heerarka qaraabada ah ee BPDE-DNA ayaa lagu cabbiray iyadoo la isticmaalayo akhristaha microplate iyadoo lagu cabbirayo nuugista 450 nm.
Unugyada hEL waxaa lagu beeray taarikada 96-ceel cufnaanta 1 × 104 unugyadiiba waxaana lagu daweeyey 10 μMB(a) P maqnaanshaha ama joogitaanka 10 μM G-Rg3 ee 48 h.Dhaqdhaqaaqa GST waxa lagu cabiray iyada oo la isticmaalayo xidhmada qiimaynta dhaqdhaqaaqa GST ee ganacsiga sida uu qabo nidaamka soo saaraha.Dhaqdhaqaaqa GST-da ee qaraabada ah waxa lagu cabbiray nuugista 450 nm iyadoo la adeegsanayo akhristaha microplate.
Unugyada hEL waxaa lagu maydhay PBS qabow oo baraf ah ka dibna la nadiifiyey iyada oo la isticmaalayo qiyaasta qiyaasta radioimmunoprecipitation ee ka kooban protease inhibitors iyo fosfatase horjoogayaasha.Ka dib markii la qiyaaso borotiinka iyadoo la adeegsanayo wadarta xirmada baaritaanka borotiinka, 30 μg ee borotiinka muunad kasta waxaa loo kala saaray 12% SDS-PAGE waxaana loo wareejiyay xuubka PVDF iyadoo loo marayo electrophoresis.Xubnuhu waxa lagu xidhay 5% caano dufan waxaana lagu daray unugyada difaaca aasaasiga ah habeenkii 4°C.Ka dib markii lagu beeray malayga peroxidase-isku-dhafan unugyada difaaca jirka ee labaad, reagents chemiluminescence oo la xoojiyey ayaa lagu daray si loo arko calaamadda xidhitaanka.Xoogagga koox kasta oo borotiin ah ayaa lagu qiyaasay iyadoo la isticmaalayo barnaamijka ImageJ.
Software-ka GraphPad Prism 7.0 ayaa loo isticmaalay in lagu falanqeeyo dhammaan xogta, oo lagu muujiyay celcelis ahaan ± weecanka caadiga ah.Kala duwanaanshaha kooxaha daawaynta ayaa la qiimeeyay iyadoo la isticmaalayo imtixaanka ardayga t ama hal dhinac falanqaynta kala duwanaanshiyaha, oo leh P qiimaha <0.05 oo muujinaya muhiimada tirakoobka.
Dhammaan xogta la helay ama la falanqeeyay inta lagu guda jiro daraasaddan waxay ku jiraan maqaalkan la daabacay iyo faylasha macluumaadka dheeriga ah.
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Waqtiga boostada: Seb-17-2023