Phenylethanol Glycosides Los ntawm Herba Cistanche Txhim Kho Hypoxic Tumor Microenvironment Thiab Txhim Kho Cov Teebmeem Ntawm Oxaliplatin Ntawm HIF-1 Txoj Kev Qhia

Hu rau Tina kom paub ntxivtina.xiang@wecistanche.com

Abstract. Mob qog noj ntshavyog ib yam ntawm cov qog malignant ntau tshaj plaws thiab muaj cov yam ntxwv ntawm malignancy siab, kev loj hlob sai, kev mob siab, thiab kev tuag. Oxaliplatin (OXA) tau tshaj tawm tias muaj kev ua haujlwm zoo tiv thaiv qib siabmob qog noj ntshavnrog tolerable toxicity. Hauv cov qog nqaij hlav, lub hypoxic microenvironment txhawb kev hloov pauv ntawm epithelial-mesenchymal (EMT), uas tuaj yeem ua rau cov tshuaj tiv thaiv kab mob siab rau cov tshuaj platinum. Herba Cistanche (Cistanche tubulosa) tau nquag siv nyob rau hauv tsoos suav tshuaj thiab covphenyl ethanolglycosideslos ntawmHerba Cistanche(CPhGs) yog cov khoom tseem ceeb. Txoj kev tshawb fawb tam sim no tsom los tshawb xyuas qhov cuam tshuam ntawm CPhGs ntawm kev muaj peev xwm, apoptosis, tsiv teb tsaws, thiab cuam tshuam ntawm daim siab mob hlwb. Cov kab mob qog nqaij hlav hepG2 tau muab faib rau hauv kev tswj hwm, DMSO, CoCl2, OXA, OXA ntxiv rau CoCl2 thiab CPhGs ntxiv rau OXA ntxiv rau CoCl2 pawg. Tom qab ntawd, rov qab hloov pauv-quantitative PCR thiab western blot tsom xam tau ua los txiav txim siab qhia theem ntawm hypoxia-inducible factor 1 (HIF-1), lysyl oxidase-like 2 (LOXL2), thiab EMT-txog cov noob thiab cov proteins (ie, E. -cadherin thiab Twist), txhawm rau tshawb xyuas qhov cuam tshuam ntawm CPhGs ntawm daim siab mob cancer. Cov txiaj ntsig tau pom tias CPhGs tuaj yeem txhim kho qhov cuam tshuam ntawm OXA ntawm daim siab mob qog noj ntshav, thiab inhibit qhov kev tsiv teb tsaws, kev ntxeem tau, thiab apoptotic tus nqi ntawm daim siab mob qog noj ntshav. Tsis tas li ntawd, kev kho CPhGs tau ua rau muaj kev cuam tshuam ntawm HIF-1, LOXL2, thiab Twist, thiab kev tswj hwm ntawm E-cadherin. Cov kev tshawb pom tam sim no tau qhia tias CPhGs ua rau muaj kev cuam tshuam loj heev rau OXA thiab kev tawm tsam ntawm hypoxia-induced EMT hauv daim siab mob cancer los ntawm inhibiting HIF-1 signaling pathway. Yog li ntawd, CPhGs yuav raug suav hais tias yog cov tshuaj platinum sensitizer zoo, uas tuaj yeem txhim kho cov tshuaj khomob hauv cov neeg mob qog noj ntshav.

Taw qhia

Mob qog noj ntshavyog ib hom qog malignant uas feem ntau cuam tshuam nrog lub plab zom mov, thiab muaj cov hom thawj thiab theem nrab. Thawjmob qog noj ntshav is divided into hepatocellular carcinoma and intrahepatic cholangiocarcinoma (1,2). Notably, liver cancer is associated with a high incidence and mortality rate worldwide (3). According to the World Health Organization, it is predicted that there will be >1,000,000 daim siab mob qog noj ntshav txog kev tuag los ntawm 2030, thiab ntawm cov neeg mob tshiab tau lees paub, cov neeg nyob hauv Suav teb loj yuav suav txog 46.6 feem pua.

Oxaliplatin (OXA) tau tshaj tawm tias muaj kev cuam tshuam inhibitory ntawm kev loj hlob ntawm daim siab mob qog noj ntshav nrog rau kev mob toxicity hauv chaw kho mob. Txawm li cas los xij, tag nrho cov txiaj ntsig ntawm kev siv tshuaj kho mob platinum yog cuam tshuam los ntawm cov qog cell tsis kam (4). Nws tau lees paub tias mob qog noj ntshav siab ua rau muaj kev nkag siab zoo dua rau kev kho tshuaj khomob piv nrog rau lwm hom mob qog noj ntshav. Cov tshuaj tiv thaiv ntawm daim siab mob qog noj ntshav tau pab txhawb rau nws txoj kev tiv thaiv rau ntau tus neeg ua haujlwm kho mob (5). Hauv kev kawm dhau los, Xie thiab Zhong (6) tau tshaj tawm tiasHepG2 hlwbpom qhov tsis zoo rau adriamycin, 5-fluorouracil, thiab cisplatin nyob rau hauv cov xwm txheej hypoxic. Txawm hais tias muaj tseeb tias cov tshuaj siv tshuaj kho mob platinum yog cov kev kho mob tseem ceeb rau kev mob qog noj ntshav, cov tshuaj tiv thaiv no muaj nyob hauv cov neeg mob. Tsis tas li ntawd, qhov kev cia siab ntawm cov neeg mob qog nqaij hlav hauv siab tseem tsis zoo (7,8). Txog niaj hnub no, kev siv zog ntau ntxiv los tshawb xyuas cov tshuaj tiv thaiv thiab txhim kho cov tshuaj rhiab heev rau cov neeg mob qog noj ntshav.

Nyob rau hauv cov xwm txheej hypoxic, revascularization tshwm sim hauv cov qog nqaij hlav cancer, uas tuaj yeem ua rau kev hloov pauv ntawm epithelial-mesenchymal (EMT) thiab vascular mimicry (VM). EMT thiab VM tuaj yeem txhawb nqa kev ntxeem tau thiab nyob deb ntawm metastasis. Ntxiv mus, EMT tau raug kwv yees los ua lub luag haujlwm ua tus tsav tsheb rau kev mob qog noj ntshav (9). Tsis tas li ntawd, cov tshuaj hypoxia-inducible (HIFs) tau koom nrog hauv kev tsim neovessel, lub zog metabolism, cellular proliferation, ntxeem tau, thiab metastasis (10).

Lysyl oxidase (LOX) zoo li 2 (LOXL2) protein yog ib tug tswv cuab ntawm tsev neeg LOX thiab muaj feem cuam tshuam nrog covalent cross-linking ntawm collagen thiab elastin, uas tuaj yeem ua rau mob fibrosis thiab tseem ceeb heev rau kev ncaj ncees ntawm extracellular matrix ( 11). Hauv kev tshawb fawb yav dhau los, LOXL2 tau suav tias yog qhov cuam tshuam zoo rau cov kab mob qog noj ntshav (12). Tsis tas li ntawd, LOXL2 tau pom tias yuav hloov kho cov kab mob thiab kev loj hlob ntawm ntau hom mob qog noj ntshav los ntawm txoj hauv kev ntxiv- thiab sab hauv lub cev, uas yog qhov tseem ceeb rau kev ntsuas kev ntsuas tsis zoo (13,14).

Herba Cistancheyog cov tshuaj tonic uas feem ntau muab faib rau hauv thaj chaw suab puam, uas tau nquag siv hauv cov tshuaj suav tshuaj (15,16).Cistanche tubulosa(C. tubulosa) yog ib hom tshuaj ntsuab uas nquag cog rau hauv Xinjiang Autonomous Region. Cov phenyl ethanol glycosides los ntawm Herba Cistanche (CPhGs) yog ib qho ntawm cov khoom tseem ceeb ntawm Herba Cistanche. Yav dhau los, Hu et al (17) tau qhia tias CPhGs tuaj yeem ua rau lub siab raug mob hauv H22 cov qog nqaij hlav nas thiab inhibit qhov kev loj hlob ntawm cov qog nqaij hlav cancer. Nws tau pom tias lub hauv paus txheej txheem yuav cuam tshuam nrog kev txo qis hauv cov ntshav-fetoprotein thiab tuaj yeem txhim kho kev tiv thaiv kab mob hauv nas.

Hauv kev tshawb fawb tam sim no, tus qauv hypoxic ntawmHepG 2Cov kab mob qog nqaij hlav hauv siab tau raug ntxias siv CoCl2. Raws li lub hauv paus no, txoj kev tshawb fawb tam sim no tsom los tshawb xyuas qhov cuam tshuam ntawm OXA ntawm kev loj hlob, apoptosis, kev tsiv teb tsaws, thiab kev cuam tshuam ntawm cov qog nqaij hlav cancer nyob rau hauv lub xub ntiag ntawm CPhGs nyob rau hauv hypoxic mob. Tsis tas li ntawd, mRNA thiab protein qhia qib ntawm HIF-1, LOXL2, E-cadherin, thiab Twist tau kuaj pom. Tsis tas li ntawd, cov txheej txheem muaj tseeb hauv qab qhov cuam tshuam ntawm CPhGs ntawm pathogenesis ntawm daim siab mob cancer tau tshawb xyuas.

Cov ntaub ntawv thiab cov txheej txheem

Cell kab. Lub siab mob qog noj ntshav kab mob HepG2, raws li tau txheeb xyuas siv txoj kev STR, tau muab los ntawm Clinical Research Institution, First Affiliated Hospital of Xinjiang Medical University (Urumqi, China). Cov hlwb tau coj mus rau hauv high-glucose DMEM (HyClone; Cytiva) uas muaj 10 feem pua ​​​​fetal bovine serum (FBS; Hyclone; Cytiva) ntawm 37˚C hauv qhov chaw tsim khoom muaj 5 feem pua ​​CO2.

Kev npaj ntawm CPhGs. C. tubulosa extraction (CPhGs) was obtained from Hetian Dichen Biotech Co., Ltd. The content of CPhGs was >80 feem pua, cov ntsiab lus ntawm echinacoside thiab verbascose yog 44.5 thiab 16.1 feem pua, feem. Stems ntawm C. tubulosa (Schrenk) Wight tau sau thaum Lub Kaum Hli 2016 los ntawm Xinjiang, Suav. Cov nroj tsuag tau txheeb xyuas los ntawm Dr. Junping Hu. Tag nrho cov voucher specimens no (no. 201610) tau muab tso rau hauv Plant Herbarium, School of Pharmacy, Xinjiang Medical University, Xinjiang, China.

Kev sim tsim. HepG2 hlwb(5x104 / ml) tau kho nrog ntau qhov ntau ntawm CPhGs (5, 25, 50, 100, 200, thiab 500 ug / ml) rau 48 teev ntawm 37˚C (24 teev tom qab cog) mus kuaj CPhGs-L / M / H koob.HepG2 hlwb(5x104/ml) tau muab faib ua pawg hauv qab no: i) Pab pawg tswj hwm, kab lis kev cai hauv cov kua nplaum siab DMEM; ii) DMSO pawg, kab lis kev cai hauv 0.1 feem pua ​​DMSO (v/v); iii) CoCl2 pawg (hypoxia qauv pawg), kab lis kev cai hauv DMEM tsis pub muaj 100 μM CoCl2; iv) OXA pawg ( pab pawg tswj tau zoo), kab lis kev cai hauv 5 μM OXA (Beijing Solarbio Science & Technology Co., Ltd.); v) OXA plus CoCl2 pawg, kab lis kev cai hauv 5 μM OXA ua ke nrog 100 μM CoCl2; thiab vi) CPhGs pawg, kho nrog CPhGs‑L/M/H (25, 50 thiab 100 ug/ml, feem) ua ke nrog 5 μM OXA thiab 100 μM CoCl2.

Cell viability assay. Cell viability tau txiav txim siab siv Cell Counting Kit-8 (CCK-8) assay raws li cov chaw tsim khoom cov lus qhia (Beijing Solarbio Science & Technology Co., Ltd.) thiab raws li tau piav qhia yav dhau los (18). Cells (2.0x103 hlwb/zoo) tau noob nyob rau hauv 96-zoo daim hlau. Tom qab ntawd, ~ 24 teev tom qab cog, cov hlwb raug kho rau 48 teev raws li kev kho mob hauv txhua pab pawg. Qhov nruab nrab yog ces hloov nrog 100 ul high-glucose DMEM, ua raws li qhov sib ntxiv ntawm 10 ul CCK-8 reagent; cov hlwb tau incubated rau 1 h ntawm 37˚C. Optical density yog ntsuas los ntawm kev siv ntau qhov ntsuas microplate nyeem ntawv (Thermo Fisher Scientific, Inc.) ntawm 450 nm. Rau qhov kev hloov pauv tau npaj rau txhua qhov xwm txheej.

Kev txiav txim siab ntawm tus nqi apoptotic. Tus nqi Apoptotic tau txiav txim siab siv Annexin V/PI Apoptotic Detection Kit (Beijing Solarbio Science & Technology Co., Ltd.). HepG2 hlwb (5x105) tau inoculated hauv 6-zoo daim hlau ntawm 37˚C hauv 5 feem pua ​​CO2. Tom qab ntawd, ~ 24 teev tom qab kev kho mob, cov hlwb raug coj mus rau hauv cov ntshav tsis muaj DMEM rau 4 teev. Cov hlwb tom qab ntawd zom tau siv 0.25 feem pua ​​​​ntawm trypsinized (Gibco; Thermo Fisher Scientific, Inc.), tom qab ntawd tsawg kawg peb ntxuav nrog pre-txias PBS. Thaum centrifugation ntawm 167.7 xg rau 5 min ntawm 4˚C, cov hlwb tau rov ua dua nrog 1X khi tsis thiab cov concentration tau hloov mus rau 1 ~ 5x106 / ml, thiab tom qab ntawd stained nrog 5 ul Annexin V-FITC thiab 5 ul PI rau 15 min. ntawm chav tsev kub. Cov hlwb ces underwent flow cytometry siv BD LSRFortessa flow cytometer (BD Biosciences) thiab FlowJo 10.6.2 software (Tree Star, Inc.).

Kev ntsuam xyuas qhov txhab. Cov teebmeem inhibitory ntawm CPhGs ntawm kev tsiv teb tsaws ntawm tes raug tshuaj xyuas los ntawm qhov mob kho qhov mob (19). Cov hlwb tau muab cog rau hauv 6-zoo daim hlau kom txog rau thaum 100 feem pua ​​​​ntawm cov monolayer tau txais. Tom qab ntawd, cov hlwb raug mob siv 200-ul pipette taub hau, ntxuav nrog PBS, thiab incubated nrog kev kho mob nyob rau hauv ib tug nruab nrab tsis muaj serum. Tom qab kev kho tshuaj, qhov mob-kho ceev tau ntsuas ntawm 0, 12, 24, thiab 48 h, ntsig txog. Cov duab raug mob tau txais los ntawm kev siv lub tshuab ntsuas fluorescent (Nikon Ti‑S, Nyiv) hauv qab qhov loj ntawm x10. Kev kaw qhov txhab raug ntsuas los ntawm qhov chaw nyob deb ntawm txhua lub sijhawm thiab qhia raws li qhov feem pua ​​​​ntawm qhov chaw mob pib ntawm 0 h.

Transwell kev soj ntsuam. Kev cuam tshuam ntawm tes raug soj ntsuam los ntawm Transwell assay. Cells (1x105 cells/ml) raug tshem tawm hauv 200 ul high-glucose DMEM yam tsis muaj FBS. Tom qab ntawd cov noob tau muab cog rau hauv Matrigel-coated lub qhov dej sab saud uas npog nrog polyethylene terephthalate lim membrane (pore loj, 8.0 μm). Tag nrho ntawm 500 ul high-glucose DMEM muaj 10 feem pua ​​​​FBS tau muab tso rau hauv chav tsev qis. Cov paj rwb swabs tau siv los tshem tawm cov hlwb nyob rau saum npoo ntawm lub lim tom qab 48 teev ntawm 37˚C. Cov hlwb uas nkag los ntawm daim nyias nyias tau kho nrog 4 feem pua ​​paraformaldehyde rau 30 feeb. Tom qab ntawd, cov hlwb tau stained nrog 0.1 feem pua ​​​​ntawm crystal violet rau 15 feeb ntawm chav tsev kub. Invading hlwb raug pom nyob rau hauv lub fluorescent microscope (Nikon Ti‑S) ntawm ib tug magnification ntawm x100.

Reverse transcription-quantitative PCR (RT-qPCR). Tag nrho RNA tau muab rho tawm los ntawm HepG2 hlwb siv TRIzol® reagent (Invitrogen; Thermo Fisher Scientific, Inc.) thiab cDNA synthesis tau ua los ntawm PrimeScript RT reagent kit (Takara Bio, Inc.) raws li cov chaw tsim khoom raws tu qauv. qPCR tau ua tiav ntawm 7500 Real-Time PCR system (Applied Biosystems; Thermo Fisher Scientific, Inc.) siv TB ntsuab™ Premix Ex Taq™ (Takara Bio, Inc.) raws li cov chaw tsim khoom raws tu qauv. Cov primers siv rau qPCR yog teev nyob rau hauv Table I. PCR tej yam kev mob muaj denaturation ntawm 95˚C rau 30 sec, ua raws li 40 cycles ntawm denaturation ntawm 95˚C rau 5 sec thiab annealing ntawm 60˚C rau 30 sec. Thaum kawg, cov txiaj ntsig tau nthuav dav tau txheeb xyuas siv 2-ΔΔCq txoj kev (20).

Western blot tsom xam. Proteins raug muab rho tawm los ntawm cov hlwb kho rau 48 h los ntawm homogenization hauv RIPA lysis tsis (Thermo Fisher Scientific, Inc.) uas muaj cov tshuaj protease thiab phosphatase inhibitors. Cell protein cov ntsiab lus tau txiav txim siab siv txoj kev BCA. Cov proteins (40 ug) tom qab ntawd tau muab cais los ntawm SDS-PAGE ntawm 10 feem pua ​​​​gel thiab xa mus rau PVDF daim nyias nyias. Lub membrane raug thaiv hauv 5 feem pua ​​​​cov mis nyuj tsis muaj rog rau 1 teev ntawm 4˚C, thiab incubated nrog cov tshuaj tiv thaiv hauv qab no: -actin (1: 5, 000; cat. no. bs-0061R; BIOSS), HIF -1 (1:1,{13}}; cat. no. ab179483; Abcam), LOXL2 (1:500; cat. no. ab179810; Abcam), E-cadherin (1:1,{21}} ; cat. no. bs‑10009R; BIOSS) thiab Twist1 (1:500; cat. no. bs‑2441R; BIOSS) hmo ntuj ntawm 4˚C. Tom qab ntawd lub membrane tau incubated nrog tshis los tiv thaiv luav IgG H&L theem ob antibodies (1:2,000; cat. no. ab205718; Abcam) rau 4 h ntawm chav tsev kub. Tom qab ntxuav nrog TBS-0.05 feem pua ​​​​Tween-20, cov blots tau pom los ntawm kev siv Enhanced Chemiluminescence system (Amersham; Cytiva). Qhov txheeb ze ntawm cov bands yog ib nrab ntsuas los ntawm kev tsom xam densitometric siv ImageJ2x software (version 2.1.4.7; Rawak Software Inc.), thiab densitometric plots ntawm cov txiaj ntsig tau normalized rau qhov kev siv ntawm -actin.

Kev txheeb cais. SPSS 19.0 software (SPSS, Inc.) tau siv los txheeb xyuas cov ntaub ntawv. Cov ntaub ntawv raug nthuav tawm raws li qhov nruab nrab ± tus qauv sib txawv thiab tau txheeb xyuas los ntawm ib txoj kev ANOVA ua raws li Tukey's post hoc test. P<0.05 was="" considered="" to="" indicate="" a="" statistically="" significant="" difference.="" all="" experiments="" were="" performed="" at="" least="" in="">

Cov txiaj ntsig

Qhov cuam tshuam ntawm CPhGs ntawm cell viability. HepG2 hlwb raug kho nrog ntau qhov ntau ntawm CPhGs (5, 25, 50, 100, 200, thiab 500 ug / ml) rau 48 teev (24 teev tom qab cog). Raws li pom nyob rau hauv daim duab 1, muaj ib qho kev poob qis hauv kev muaj peev xwm ntawm cov hlwb kho nrog 200 thiab 500 ug / ml CPhGs piv nrog cov pab pawg tswj hwm (P<0.05). these="" findings="" indicated="" that="" cphgs="" could="" modulate="" cell="" viability="" in="" a="" dose‑dependent="">

CPhGs txhim kho cov teebmeem ntawm OXA ntawm daim siab mob cancer. Cov teebmeem ntawm CPhGs ntawm OXA-modulated HepG2 cell viability tau raug soj ntsuam tom qab (Fig. 2). Tom qab ~ 48 h, kev sib xyaw ua ke ntawm CPhGs thiab OXA txo qis kev muaj peev xwm ntawm HepG2 hlwb piv nrog rau hauv OXA ntxiv rau CoCl2 pawg. Tshwj xeeb, CPhGs-M ntxiv rau OXA ntxiv rau CoCl2 thiab CPhGs-H ntxiv rau OXA ntxiv rau CoCl2 cuam tshuam qhov muaj peev xwm ntawm HepG2 hlwb piv rau hauv pawg OXA ntxiv rau CoCl2 (P<0.05 and=""><0.01,>

CPhGs inhibits kev tsiv teb tsaws thiab cuam tshuam ntawm daim siab mob hlwb. Txhawm rau kawm ntxiv txog qhov muaj peev xwm cuam tshuam thiab muaj peev xwm tsiv teb tsaws ntawm daim siab mob qog noj ntshav tom qab kev kho mob nrog CPhGs thiab OXA, kev kho qhov txhab thiab Transwell tau ua. Qhov kev ntsuam xyuas qhov txhab kho tau qhia tias, piv nrog rau hauv pawg DMSO, kev tsiv teb tsaws ntawm HepG2 hlwb raug txo qis tom qab kev kho mob nrog CoCl2, OXA, thiab CPhGs (200 lossis 500 ug / ml) (P<0.05; fig.="" 3a="" and="" b).="" for="" the="" transwell="" assay,="" the="" invasive="" ability="" of="" cells="" was="" markedly="" inhibited="" in="" the="" co‑treatment="" groups="" (cphgs="" +="" oxa="" +="" cocl2)="" compared="" with="" that="" in="" the="" oxa="" +="" cocl2="" group=""><0.01; fig.="" 3c="" and="" d).="" conversely,="" in="" the="" co‑treatment="" groups="" (cphgs="" +="" oxa="" +="" cocl2),="" the="" invasive="" potential="" and="" migratory="" ability="" of="" cells="" were="" markedly="">

Cov teebmeem ntawm CPhGs thiab OXA ntawm apoptosis. Tom qab kev kho mob nrog kev sib xyaw ntawm CPhGs thiab OXA rau 48 h, HepG2 hlwb tau stained nrog Annexin V-FITC thiab PI, ua raws li kev ntws cytometry los txiav txim siab cellular apoptosis. Raws li pom nyob rau hauv daim duab 4A, cov pab pawg kho mob (CPhGs‑L/M/H ntxiv rau OXA ntxiv rau CoCl2) tau nthuav tawm qhov kev nce siab zuj zus hauv qhov feem pua ​​​​ntawm cov hlwb apoptotic nrog qhov nce hauv CPhGs concentration. Feem ntau ntawm cov hlwb kho nrog CPhGs thiab OXA tau nyob hauv cheeb tsam Q4, uas qhia tias kev sib xyaw ntawm CPhGs thiab OXA induced apoptosis thaum ntxov. Piv nrog rau hauv pab pawg OXA ntxiv rau CoCl2, qhov kev nce siab tseem ceeb hauv apoptotic tus nqi ntawm cov hlwb tau kuaj pom hauv cov pab pawg kho nrog OXA, CoCl2, thiab cov koob tshuaj nruab nrab lossis siab ntawm CPhGs (P<0.01; fig.="" 4b).="" these="" findings="" indicated="" that="" the="" combination="" of="" oxa="" and="" cphgs="" may="" contribute="" to="" the="" apoptosis="" of="" hepg2="">

mRNA qhia theem ntawm HIF-1, LOXL2, E-cadherin, thiab Twist tom qab CPhGs thiab OXA co-incubation. There were no statistical differences in the mRNA expression levels of HIF‑1α, LOXL2, E‑cadherin, and Twist between the control and DMSO groups (P>0.05; Fig. 5A-D). Hloov pauv, CoCl2 ua rau muaj kev nce ntxiv hauv mRNA qhia theem ntawm LOXL2, HIF-1, thiab Twist piv nrog cov hauv pawg DMSO (P<0.01; fig.="" 5a,="" b="" and="" d).="" compared="" with="" those="" in="" the="" oxa="" +="" cocl2="" group,="" the="" mrna="" expression="" levels="" of="" loxl2,="" hif‑1α="" and="" twist="" were="" significantly="" enhanced="" in="" the="" cphgs‑h="" +="" oxa="" +="" cocl2="" groups=""><0.01; fig.="" 5a,="" b="" and="" d).="" by="" contrast,="" cocl2="" induced="" a="" significant="" downregulation="" in="" the="" mrna="" expression="" levels="" of="" e‑cadherin="" compared="" with="" those="" in="" the="" dmso="" group=""><0.01; fig.="" 5c).="" all="" concentrations="" of="" cphgs="" combined="" with="" oxa="" and="" cocl2="" were="" able="" to="" upregulate="" the="" mrna="" expression="" levels="" of="" e‑cadherin="" compared="" with="" those="" in="" the="" oxa="" +="" cocl2="" group=""><0.01; fig.="" 5c).="" these="" results="" indicated="" that="" the="" combination="" of="" cphgs="" and="" oxa="" effectively="" inhibited="" the="" emt="" under="" hypoxic="">

Protein qhia theem ntawm HIF-1, LOXL2, E-cadherin, thiab Twist tom qab CPhGs thiab OXA co-incubation. Cov txiaj ntsig ntawm western blotting qhia tau hais tias cov protein qhia theem ntawm HIF-1 , LOXL2, thiab Twist tau raug kho raws li cov xwm txheej hypoxic piv nrog cov hauv pawg DMSO. Los ntawm qhov sib txawv, cov protein qhia theem ntawm E-cadherin raug txo qis hauv cov xwm txheej hypoxic (P<0.01; fig.="" 6a="" and="" b).="" notably,="" the="" protein="" expression="" levels="" of="" hif‑1α,="" loxl2,="" and="" twist="" were="" significantly="" decreased="" in="" the="" cphgs‑m="" +="" oxa="" +="" cocl2="" or="" cphgs‑h="" +="" oxa="" +="" cocl2="" groups="" compared="" with="" those="" in="" the="" oxa="" +="" cocl2="" group=""><0.01; fig.="" 6a="" and="" b).="" compared="" with="" the="" dmso="" group,="" the="" cocl2="" treatment="" significantly="" decreased="" the="" expression="" level="" of="" e‑cadherin.="" in="" the="" cphgs="" groups,="" the="" protein="" expression="" levels="" of="" e‑cadherin="" were="" significantly="" increased="" compared="" with="" those="" in="" the="" oxa="" +="" cocl2="" group=""><0.01; fig.="" 6b).="" these="" findings="" indicated="" that="" cphgs="" treatment="" could="" effectively="" inhibit="" the="" downregulation="" of="" e‑cadherin,="" and="" the="" upregulation="" of="" hif‑1α,="" loxl2,="" and="" twist="" induced="" by="">

Kev sib tham

Hypoxia yog ib qho tshwm sim hauv cov kab mob qog noj ntshav microenvironment; Qhov no feem ntau cuam tshuam nrog qhov tseeb tias kev loj hlob ntawm cov qog nqaij hlav cancer yog nrawm dua piv nrog cov vascular tsim ntawm aberrant neovessels. Tsis tas li ntawd, lwm cov txheej txheem lom neeg, suav nrog kev loj hlob, metastasis, thiab tshuaj rhiab heev raug cuam tshuam los ntawm hypoxia (21). Cov qog hypoxic microenvironment yog qhov tseem ceeb rau cov kab mob pathogenesis thiab kev mob qog noj ntshav, thiab nws kuj tseem ceeb hauv cov tshuaj tiv thaiv thiab vascularization ntawm daim siab mob cancer (22).

Hauv txoj kev tshawb no, HepG2 hlwb tau kho nrog ntau qhov ntau ntawm CPhGs, ntawm cov CPhGs (200 ug / ml) tuaj yeem ua rau txo qis hauv cell viability. Tshwj xeeb, CPhGs tuaj yeem hloov kho cellular viability nyob rau hauv koob tshuaj. Nyob rau hauv cov xwm txheej hypoxic, kev sib xyaw ntawm OXA thiab CPhGs (50 lossis 100 ug / ml) cuam tshuam qhov muaj peev xwm ntawm HepG2 hlwb piv nrog kev kho OXA ib leeg. Ib qho piv txwv ntawm kev noj tshuaj zoo sib xws tau pom nyob rau hauv kev tsiv teb tsaws thiab kev cuam tshuam ntawm cov kab mob qog nqaij hlav hauv siab. Tam sim no, cov kev tshawb fawb dav tau ua los tshawb xyuas lub luag haujlwm ntawm kev mob qog noj ntshav ntawm tes apoptosis hauv pathogenesis ntawm daim siab kab mob (23-26). Ntau lub tswv yim tau tsim los kho mob qog noj ntshav los ntawm kev txhawb nqa apoptosis (27-29); Yog li ntawd, kev cuam tshuam hauv HepG2 cell apoptosis tuaj yeem ua tus neeg sib tw zoo rau kev tiv thaiv thiab kho mob qog noj ntshav. Lub apoptosis ntawm HepG2 hlwb tau txhim kho tau zoo tom qab kev kho mob nrog kev sib xyaw ntawm CPhGs-M/-H thiab OXA piv nrog rau hauv cov hlwb kho nrog OXA ib leeg. Yog li ntawd, nws tau qhia tias CPhGs tuaj yeem txhim kho cov nyhuv antitumor ntawm OXA.

HIF-1 tuaj yeem txhim kho qib kev qhia ntawm E-cadherin, N-cadherin, thiab Vimentin, nrog rau qee qhov kev hloov pauv, xws li Snail1/2, Zeb1, thiab Twist1. Tom qab ntawd, qhov no yuav ua rau poob ntawm cellular polarity, loosening ntawm cell-cell junctions, alterations nyob rau hauv cytoskeletal protein, thiab tsiv teb tsaws thiab ntxeem tau ntawm cov qog nqaij hlav cancer, uas yuav ua rau translocation ntawm cov qog nqaij hlav cancer mus rau lub circulatory system los ntawm lub basilar membrane. thiab tom qab metastasis (30). Hauv txoj kev tshawb fawb tam sim no, CoCl2 tau siv los ntxias tus qauv ntawm hypoxia, uas ua rau muaj kev nce hauv kev muaj peev xwm ntawm HepG2 hlwb, nrog rau kev tsiv teb tsaws ntawm tes thiab ntxeem tau. Tsis tas li ntawd, mRNA thiab protein qhia qib ntawm HIF-1 tau nce ntxiv, qhia tias CoCl2 ua rau muaj kev tsim cov kab mob.

Twist thiab E-cadherin qhia; Yog li ntawd, cov kev tshawb fawb yav tom ntej tsom mus tsom rau ntau EMT cov cim ntsig txog, txhawm rau ntsuas qhov cuam tshuam ntawm CPhGs ntawm hypoxia-induced EMT hauv daim siab mob cancer.

HIF-1 tau tshaj tawm los txhawb kev nthuav qhia ntawm LOXL2, thiab txhim kho kev tsiv teb tsaws thiab kev cuam tshuam ntawm cov kab mob qog nqaij hlav hauv siab, uas yuav cuam tshuam zoo rau qhov tsis zoo ntawm kev mob qog noj ntshav siab (30). Hauv kev tshawb fawb yav dhau los, cov lus qhia theem ntawm LOXL2 nyob rau hauv cov ntaub so ntswg mob qog noj ntshav uas nyob ib sab tau nce ntau dua piv nrog cov qog nqaij hlav cancer (35). Tsis tas li ntawd, nws muaj feem cuam tshuam nrog kev cuam tshuam thiab metastasis ntawm daim siab mob qog noj ntshav. LOXL2 gene silencing los ntawm kev cuam tshuam me me RNA inhibited kev loj hlob ntawm HepG2 thiab SMCC-7721 hlwb, uas ua rau lub voj voog ntawm tes ntawm cov qog nqaij hlav cancer thiab nce apoptosis (36). Shao et al (35) tau tshawb xyuas qhov sib txheeb ntawm LOXL2 hauv cov qauv qog nqaij hlav hauv siab, thiab cov xwm txheej hauv tsev kho mob, VM, thiab kev kwv yees ntawm 201 tus neeg mob uas tau txais kev phais kho mob. Nws tau pom tias LOXL2 tau ua haujlwm tseem ceeb hauv kev tsim cov kab mob thiab kev loj hlob ntawm daim siab mob qog noj ntshav, uas tuaj yeem ua lub hom phiaj rau kev txhim kho tshuaj. Tsis tas li ntawd, Peng et al (37) tau qhia tias LOXL2 tuaj yeem ua rau Snail / E-cadherin thiab Src kinase / Focal adhesion kinase signaling txoj hauv kev, uas tuaj yeem ua rau muaj kab mob thiab kev loj hlob ntawm EMT ntawm cov qog nqaij hlav hauv plab. Hauv kev tshawb fawb tam sim no, nyob rau hauv cov xwm txheej hypoxic, mRNA thiab protein qhia qib ntawm LOXL2 tau nce ntxiv, thaum nws cov lus qhia raug txo qis tom qab kev kho mob nrog OXA. Tsis tas li ntawd, kev kho mob nrog kev sib xyaw ua ke ntawm CPhGs thiab OXA ua rau pom tseeb txo qis ntawm LOXL2 kev qhia, uas tuaj yeem pab tau zoo rau cov teebmeem ntawm OXA ntawm daim siab mob cancer.

Muaj qee qhov kev txwv rau txoj kev kawm tam sim no. Txoj kev tshawb fawb tam sim no yuav tsum tau siv ob txoj kab mob qog nqaij hlav hauv siab ntxiv, suav nrog SMCC-7721 cell kab, tab sis cov no tsis tuaj yeem siv vim nws tsis tuaj yeem yuav cov kab cell no vim lawv tau txheeb xyuas tsis raug thiab tau muab los ntawm HeLa hlwb. Tsis tas li ntawd, txoj kev tshawb fawb tam sim no tsis tau txheeb xyuas cov teebmeem antioxidant ntawm ntau qhov sib txawv ntawm CPhGs hauv cov hlwb.

Hauv kev xaus, CPhGs tuaj yeem hloov pauv qhov hypoxic qog microenvironment ntawm daim siab mob qog noj ntshav los ntawm kev hloov kho HIF-1 txoj kev taw qhia. Tsis tas li ntawd, qhov rhiab heev ntawm daim siab mob hlwb rau OXA tau nce siab hauv cov lus teb rau kev kho mob nrog kev sib xyaw ntawm CPhGs thiab OXA. Cov kev tshawb pom no tuaj yeem muab cov tswv yim kho tshiab los txhim kho qhov rhiab ntawm daim siab mob qog noj ntshav rau cov tshuaj kho mob.

LIMEI WEN1,2*, JUNPING HU1*, JIAWEI ZHANG1 thiab JIANHUA YANG1,2

1College of Pharmacy, Xinjiang Medical University, Urumqi, Xinjiang 830054; 2 Department of Pharmacy, Thawj Tsev Kho Mob, Xinjiang Medical University, Urumqi, Xinjiang 830011, PR Tuam Tshoj

Tau txais Lub Xya Hli 29, 2020; Tau txais Lub Peb Hlis 9, 2021

DOI: 10.3892/mmr.2021.12156

Kev lees paub

Tsis siv tau.

Nyiaj txiag

Txoj kev tshawb no tau txais kev txhawb nqa los ntawm Xinjiang Key Laboratory ntawm Natural Drug Active Components thiab Drug Release Technology (grant no. XJDX1713), Reserve Candidate Project for the Leader of Scientific and Technological Innovation in Xinjiang Uygur Autonomous Region (grant no. 2019XS14), National Natural Science Foundation of China (grant no. 81860735) thiab Bethune Charitable Foundation 'Bethune·Quest-construction of pharmaceutical scientific research capacity (grant no. B-19-H-20200622).

Muaj cov ntaub ntawv thiab cov ntaub ntawv

Cov ntaub ntawv siv thiab / lossis kev tshuaj xyuas thaum lub sijhawm kawm tam sim no muaj los ntawm tus kws sau ntawv raws li kev thov tsim nyog.

Cov neeg sau ntawv pab txhawb

LMW thiab JWZ tau ua qhov kev sim, tsim cov ntawv sau, thiab lees paub qhov tseeb ntawm tag nrho cov ntaub ntawv nyoos. JPH thiab JHY tsim txoj kev tshawb fawb tam sim no. Txhua tus kws sau ntawv tau nyeem thiab pom zoo cov ntawv sau kawg.

Kev pom zoo ntawm kev ncaj ncees thiab kev tso cai los koom

Tsis siv tau.

Tus neeg mob tso cai rau kev tshaj tawm

Tsis siv tau.

Kev nyiam sib tw

Cov kws sau ntawv tshaj tawm tias lawv tsis muaj kev sib tw nyiam.

Cov ntaub ntawv

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