FGIN -1-27 yog ib qho hluavtaws mitochondrial diazepam binding inhibitor receptor (MDR)agonist uas tau ua pov thawj pro-apoptotic, tiv thaiv kev ntxhov siab, thiab kev ua haujlwm steroidogenicnyob rau hauv ntau yam kev tshawb fawb. Ntawm no peb qhia, rau lubua ntejlub sij hawm, lub anti-melanogenic efficiencyntawmFGIN-1-27hauv vitrothiabhauv vivo. FGIN-1-27 tseem ceeb heevinhibited basal thiab - melanocyte-stimulating hormone ( -MSH)-, 1-Oleoyl-2-acetyl-sn-glycerol (OG)-thiab Endothelin-1 (ET-1)-induced melanogenesis yam tsis muaj cellular toxicity.Mushroom tyrosinase kev ua ub no qhia tau tias FGIN -1-27 tsis ncaj qha inhibittyrosinase kev ua, qhia tias FGIN -1-27 tsis yog ib qho ncaj qha tyrosinase inhibitor. Txawm hais tias nws tsis tuaj yeem hloov kho cov haujlwm catalytic ntawmnceb tyrosinasehauv vitro, FGIN-1-27 downregulated cov theem qhia ntawmcov proteins tseem ceeb uas ua haujlwm hauv melanogenesis. FGIN{0}} ua si cov haujlwm nolos ntawm suppressing PKA/CREB, PKC- , thiab MAPK txoj kev. Thaum inactivated, nwstxo qis kev qhia ntawm MITF, tyrosinase, TRP-1, thiab TRP-2, thiab inhibitedkev ua haujlwm ntawm tyrosinase,thaum kawginhibiting melanogenesis. Thaumhauv vivokev sim,FGIN{0}} inhibited lub cev pigmentation ntawm zebrafishthiab txo UVB-inducedhyperpigmentation nyob rau hauv daim tawv nqaij guinea npua, tab sis tsis txo qis ntawm ib tug xov tooj ntawm melanocytes.Pebkev tshawb pomqhia tias FGIN -1-27 tsis muaj cytotoxicity thiab inhibitedmelanogenesis nyob rau hauv ob qho tib sihauv vitrothiabhauv vivoqauv. Nws yuav ua pov thawj zoo heev li ibkev nyab xeeb ntawm daim tawv nqaij-dawb huv.

Raws li cov kev tshawb fawb cuam tshuam,cistancheyog ib hom tshuaj ntsuab uas hu ua "cov tshuaj ntsuab miracle uas ua rau lub neej ntev". Nws lub ntsiab tivthaiv yog cistanoside, uas muaj ntau yam teebmeem xws liantioxidant, anti-inflammatory, thiab kev tiv thaiv kev ua haujlwm nce qib. Lub mechanism ntawm cistanche thiabtawv nqaij dawbnyob rau hauv cov nyhuv antioxidant ntawm cistanche glycosides.MelaninNyob rau hauv tib neeg daim tawv nqaij yog tsim los ntawm oxidation ntawm tyrosine catalyzed los ntawmtyrosinase, thiab cov tshuaj tiv thaiv oxidation yuav tsum muaj kev koom tes ntawm oxygen, yog li cov pa oxygen dawb radicals hauv lub cev dhau los ua ib qho tseem ceeb cuam tshuam rau kev tsim melanin. Cistanche muaj cistanoside, uas yog ib qho antioxidant thiab tuaj yeem txo qhov tsim ntawm cov dawb radicals hauv lub cev, yog liinhibiting melanin ntau lawm.

Nyem rau ntawm Cistanche Tubulosa

Yog xav paub ntxiv: david.deng@wecistanche.com WhatApp:86 13632399501

Taw qhia

Melanogenesis yog lub luag haujlwm tseem ceeb tshaj plaws ntawm melanocytes hauv daim tawv nqaij. Ntau yam tseem ceeb uas cuam tshuam nrog melanogenesis tau qhia meej (Raposo thiab Marks, 2007; Noguchi li al., 2014). Lub enzyme tseem ceeb uas tswj cov tshuaj melanogenesis yog tyrosinase thiab nws ua nrog tyrosinase-related protein 1 (TRP1) thiab tyrosinase-related protein 2 (TRP2) los txhawb melanin synthesis (Zhu li al., 2015). Lwm qhov tseem ceeb ntawm cov pigmentation yog microphthalmia-associated transcription factor (MITF), uas tsis tsuas yog tswj kev loj hlob thiab ciaj sia ntawm melanocytes tab sis kuj tswj tyrosinase, TRP1, thiab TRP2 qhia (Kawasaki li al., 2008; Levy thiab Fisher, 2011. ). Environmental stimuli, xws li ultraviolet (UV) irradiation, ua lub luag haujlwm tseem ceeb hauv melanogenesis (Abdel-Naser li al., 2003). Thaum raug UV hluav taws xob, activated keratinocytes thiab melanocytes tsim ib qho -melanocyte-stimulating hormone (-MSH), diacylglycerol (DAG), thiab endothelin-1 (ET-1) (D'Mello li al., 2016; Bae Harboe and Park, 2012). Thaum -melanocyte-stimulating hormone (-MSH) khi rau melanocortin -1 receptor (MC1R) hauv melanocytes, nws tuaj yeem ua rau cov qib hauv cov cell ntawm cyclic adenosine monophosphate (cAMP) thiab qhib cov protein kinase A (PKA) / (CREB) ) txoj kev, thaum kawg txhawb melanogenesis (Corre li al., 2004; Rzepka li al., 2016). Diacylglycerol (DAG) yog qhov tseem ceeb rau kev ua kom cov protein kinase C- (PKC- ) hauv melanocytes, uas ua rau muaj kev ua haujlwm ntawm tyrosinase thiab induces pigmentation (Kim li al., 2006; Kawaguchi li al., 2012; Yuan and Jin, 2018. ). 1-Oleoyl-2- acetyl-sn-glycerol (OOG) yog ib qho hluavtaws, membrane-permeable DAG analog uas tau siv dav los ua tshuaj siv tshuaj ntawm PKC- (Rainbow et al., 2011). Endothelin-1 (ET- 1) induces melanogenesis hauv melanocytes los ntawm kev ua kom cov mitogen-activated protein kinase (MAPK) txoj kev (Park et al., 2015; Regazzetti et al., 2015).

Melanin plays lub luag haujlwm tseem ceeb hauv kev tiv thaiv ntawm daim tawv nqaij los ntawm kev phom sij ntawm ultraviolet hluav taws xob, xws li mob qog noj ntshav thiab kev laus (Slominski et al., 2004). Txawm li cas los xij, kev tsim khoom ntau dhau thiab qhov sib txawv ntawm melanin ua rau muaj teeb meem ntawm daim tawv nqaij, xws li melasma, freckles, thiab qhov chaw tsaus nti (Jadotte and Schwartz, 2010). Ob peb lub active whitening tebchaw, nrog rau kojic acid thiab arbutin, twb tau pom zoo raws li cosmetic additives (Kim li al., 2008). Tab sis nws tau raug tshaj tawm tias arbutin muaj cov teebmeem cytotoxic thiab kojic acid tuaj yeem ua rau mob qog noj ntshav (Singh li al., 2016). Yog li, qhov kev tshawb pom ntawm cov khoom siv tau zoo dua thiab muaj kev nyab xeeb ntawm daim tawv nqaij dawb yog qhov tseem ceeb.

Mitochondrial diazepam binding inhibitor receptor (MDR) plays lub luag haujlwm tseem ceeb hauv kev tswj hwm ntawm lipid thiab steroid synthesis thiab nthuav dav hauv cov ntaub so ntswg nrog rau ntawm daim tawv nqaij (Alho li al., 1993). Hauv peb cov kev tshawb fawb yav dhau los, diazepam, ib tus MDR agonist, tswj cov pigmentation ntawm PKA/CREB txoj hauv kev (Lv li al., 2019). Tsis tas li ntawd, kev tshawb pom tsis ntev los no tau qhia tias kev ua kom MDR me ntsis nce tus naj npawb ntawm melanocytes tshiab hauv zebrafish (Allen li al., 2020). Hauv kev sib piv nrog diazepam, FGIN-1-27 (N, N-Dihexyl-2-(4-fluorophenyl)indole-3- acetamide) nthuav tawm qhov kev sib raug zoo dua thiab qhov tshwj xeeb rau MDR (Freeman thiab Young, 2000). FGIN-1–27 tau qhia txog kev ua haujlwm pro-apoptotic thiab steroidogenic (Lacapère thiab Papadopoulos, 2003). Tsis ntev los no, nws tau tshaj tawm tias FGIN{13}} ua rau muaj kev tiv thaiv kev ntxhov siab thiab tiv thaiv kev ntshai hauv vivo (Lima-Maximino li al., 2018). Txawm li cas los xij, cov nyhuv ntawm FGIN{18}} ntawm pigmentation hauv vitro thiab hauv vivo tsis tau tshaj tawm.

KHOOM PLIG THIAB MUAB

Khoom siv

FGIN-1-27(N, N-Dihexyl{2}}(4-fluorophenyl)indole-3-acetamide), OAG (1-Oleoyl-2-acetyl-sn -glycerol), thiab cov tshuaj tiv thaiv PKC- (1:200), p-PKA miv (phospho T197)(1:200), PKA cat (1:500), p-CREB (1:200), CREB(1:200), p-ERK(1:200), ERK (1:200), p-p38 (1:200), p38 (1:200), p-JNK (1:200),thiab JNK (1:200) tau txais los ntawm Santa Cruz Biotechnology(CA, USA). Tyrosinase los ntawm nceb, -MSH, ES-1 (Endothelin-1), thiab PTU (N-Phenylthiourea) tau txaislos ntawm Aladdin (Shanghai, Suav). Cov tshuaj tiv thaivtyrosinase (1:1,{2}}), TRP-1(1:1,000), TRP-2 (1:1,{10}}), MITF(1:2000), thiab -actin (1: 3, 000) tau txais los ntawm Abcam(Cambridge, UK). Masson-Fontana melanin staining tovtau yuav los ntawm SenBeiJia Biological Technology (Nanjing,Tuam Tshoj). RT-qPCR cov khoom siv tau yuav los ntawm Takara BiomedicalTechnology (Beijing, Tuam Tshoj). Cell lysis buffer thiab BCA proteinCov khoom siv tshuaj ntsuam xyuas tau txais los ntawm Beyotime Biotechnology (Shanghai,Tuam Tshoj).

Cells Culture

Cell Viability Assay

Qhov kev muaj peev xwm ntawm SK-MEL-2 hlwb thiab HEM raug ntsuas los ntawm MTT kev xeem (Zhou et al., 2014). SK-MEL-2 hlwb thiab HEM tau raug plated nyob rau hauv 96-zoo daim hlau ntawm qhov ntom ntawm 2 × 104 cell ntawm ib qhov dej. FGIN-1-27 (0, 1, 2, 4, 8, 16 μM) tau siv rau SK-MEL-2 hlwb thiab HEM rau 48 teev. MTT tov (20 μL) tau ntxiv rau txhua qhov dej ntawm 96-zoo phaj rau lwm 4 h. Tom qab tshem tawm cov tshuaj, DMSO (200 μL) tau ntxiv rau txhua qhov dej thiab qhov nqus tau raug tshuaj xyuas ntawm 570 nm siv lub microplate spectrophotometer (BioTek Instruments).

Melanin Assay

SK-MEL-2 hlwb thiab HEM tau raug plated nyob rau hauv 6-zoo daim hlau ntawm ib tug concentration ntawm 5 × 105 hlwb rau ib qhov dej. Tom qab 24 teev incubation ntawm 37 degree, FGIN-1-27 (0, 1, 2, 4 μM) tau siv rau SK-MEL-2 hlwb thiab HEM. Cov hlwb raug incubated rau lwm 48 h, ntxuav nrog PBS, ua raws li lysis nrog cell lysis tsis rau 20 min ntawm 4 degree C, thiab cov lysates tau centrifuged ntawm 13, 000 rpm rau 10 min ntawm 4 degree C. Tag nrho cov melanin nyob rau hauv lub cell pellet yog yaj nyob rau hauv 100 μL NaOH tov (1 mol / L, muaj 10 feem pua ​​DMSO) ntawm 80 degree C rau 1 h (Lv li al., 2015). Optical absorbance tau ntsuas ntawm 405 nm siv lub microplate spectrophotometer (BioTek Instruments).

Tyrosinase Activity Assay

Cov kev ua ntawm cellular tyrosinase tau ua raws li tau piav qhia dhau los (Lv li al., 2020). SK-MEL-2 cov hlwb tau tsim nrog ntau qhov sib txawv ntawm FGIN-1-27 (0, 1, 2, 4 μM) lossis OAG (200 μM) rau 12 h lossis 48 h thiab tom qab ntawd lysed nrog cell lysis buffer. Cell lysates tau centrifuged ntawm 13, 000 rpm rau 10 min ntawm 4 degree C kom tau txais supernatant rau tyrosinase kev ua ub no. Protein concentrations tau txiav txim siab los ntawm cov khoom siv BCA nrog bovine serum albumin (BSA) raws li tus qauv.100 μL PBS (0.1 M, pH 6.5) muaj 30 ug cov protein sib xyaw nrog 100 μL L-DOPA (0.1 feem pua) thiab tom qab ntawd incubated ntawm 37 degree C rau 1 h. Optical absorbance tau ntsuas ntawm 475 nm siv lub microplate spectrophotometer (BioTek Instruments).

Qhov cuam tshuam ncaj qha ntawm FGIN{{0}} ntawm kev ua tyrosinase tau ua raws li txoj hauv kev dhau los (Tomita li al., 2010; Lv et al., 2020). 100 μL PBS (0.1 M, pH 6.5) muaj qhov sib txawv ntawm FGIN-1-27 sib xyaw nrog nceb tyrosinase (10 units) thiab 50 μL L-tyrosine (0.05 feem pua) thiab tom qab ntawd incubated ntawm 37 degree rau 10 min. Optical absorbance tau ntsuas ntawm 475 nm siv lub microplate spectrophotometer (BioTek Instruments).

Masson-Fontana Ammoniacal Silver Staining

Rau kev kuaj pom cov xim melanin, SK-MEL-2 hlwb raug plated rau ntawm 6-zoo phaj uas muaj 13- hli iav npog hauv 2.0 ml ntawm kab lis kev cai nruab nrab (10 feem pua ​​​​fetal bovine serum) hauv txhua qhov dej. Tom qab 48 teev tom qab kev tswj hwm, cov hlwb raug kho nrog Formalin (10 feem pua ​​​​ntxhiab buffered) rau 20 feeb.

Cells thiab daim tawv nqaij swb tau stained raws li Masson-Fontana ammoniacal silver staining method (Lee et al., 2013). Cells thiab daim tawv nqaij swb tau incubated nrog Masson-Fontana melanin staining tov rau 16 h ntawm chav tsev kub. Tom qab ntawd cov hlwb thiab cov tawv nqaij swb tau muab yaug hauv dej distilled 3 zaug thiab tau incubated nrog hypo daws rau 7 min. Tom qab ntxuav tag huv si hauv dej distilled, hlwb, thiab daim tawv nqaij slides raug counterstained nrog ib tug nruab nrab liab stain rau 7 min. Cells thiab daim tawv nqaij swb tau soj ntsuam siv Nikon Ti2-U microscope.

Immunohistochemistry rau S-100

S-100 immunohistochemistry tau ua raws li tau piav qhia yav dhau los (Lee et al., 2013). Cov swb raug thaiv nrog 5 feem pua ​​​​BSA rau 1 teev ntawm chav tsev kub thiab tom qab ntawd incubated hauv anti-S-100 thawj cov tshuaj tiv thaiv (Dako, Carpentaria, CA) diluted hauv thaiv kev daws thaum hmo ntuj ntawm 4 degree C. Hnub tom qab, cov Slides tau ntxuav nrog TBST 4 zaug, thiab incubated nrog ib tug thib ob antibody (Dako, Carpentaria, CA). Tom qab ntawd, cov slides tau kho nrog aminoethyl carbazole los tsim cov ntu. Cov tawv nqaij swb tau soj ntsuam siv Nikon Ti2-U microscope.

Rov qab Transcription-Quantitative PCR (RT-qPCR)

RT-qPCR tau ua raws li tau piav qhia dhau los (Lv li al., 2020). Luv luv, SK-MEL-2 cell tag nrho RNA raug muab rho tawm siv TRIzol reagent thiab quantified spectrophotometrically. Ib leeg-stranded cDNA tau tsim los siv SuperScript II Reverse transcriptase raws li cov chaw tsim khoom cov lus qhia. SYBR-Green quantitative PCR tsom xam tau ua nrog ABI PRISM Sequence Detection System (Applied Biosystems) thiab pre-validated primer sets. Tag nrho cov qauv tau khiav hauv triplicate. Lub voj voog pib tau muab tso rau hauv lub logarithmic feem ntawm qhov kev nthuav dav nkhaus, thiab cov txiaj ntsig tau zoo li qub rau GAPDH. Qhov sib txawv ntawm ob qho piv txwv tau txiav txim siab los ntawm txoj kev delta-delta Cq. Cov primers sequences ntawm Mitf gene yog 5′-AGAGCAGGCAGAGAGTGAGTG-3′, 5′-AACTTGATT CCAGGCTGATGATGTC-3′.

Western Blot Analysis

Qhov kev tshem tawm cov protein tau txais raws li txoj kev tau hais los saum toj no. Cov proteins uas tau muab rho tawm hauv lub cev (30 ug) raug cais los ntawm SDS-PAGE (10 feem pua) thiab tom qab ntawd xa mus rau polyvinylidene L fluoride (PVDF) daim nyias nyias, ua raws li cov txheej txheem ntub dej hloov pauv. Cov membrane raug thaiv siv 2.5 feem pua ​​​​BSA ntawm chav tsev kub rau 1 h, raug rau cov thawj cov tshuaj tiv thaiv uas tsim nyog ntawm 4 degree C thaum hmo ntuj, thiab ntxuav nrog TBST 4 zaug. Cov daim nyias nyias tau raug rau HRP-labeled Tshis Anti-Lavbit IgG (1: 1, 000) lossis Tshis Anti-Mouse IgG (1: 1,000) ntawm chav tsev kub rau 1 h thiab ntxuav nrog TBST 4 zaug (Liao et al., 2017). Cov daim nyias nyias tau pom los ntawm kev siv lub tshuab ua haujlwm ntau yam chemiluminescence (Tanon 4600). Internal tswj los ntawm tib blot raug soj ntsuam nrog anti-actin.

Phenotype-raws li kev ntsuam xyuas thiab kev txiav txim siab ntawm Melanin Cov ntsiab lus hauv Zebrafish

Cov neeg laus zebrafifishes tau khaws cia rau hauv lub tank nrog cov xwm txheej hauv qab no: 28.5 degree, nrog rau 14/10 h lub teeb / tsaus voj voog. Embryos tau txais los ntawm ntuj spawning uas yog induced thaum sawv ntxov los ntawm tig rau lub teeb. Kev sau ntawm embryos tau ua tiav hauv 30 min. Kev ntsuam xyuas raws li phenotype tau ua raws li tau piav qhia yav dhau los (Choi et al., 2007). Luv luv, synchronized embryos tau sau thiab arrayed los ntawm pipette (3-4 embryos ib qhov dej nyob rau hauv 96-zoo daim hlau uas muaj 200 ul ntawm embryo nruab nrab). FGIN{10}} thiab PTU tau yaj hauv 0.1 feem pua ​​DMSO, tom qab ntawd ntxiv rau hauv nruab nrab embryo los ntawm 35 txog 60 h (25 h raug). Cov teebmeem ntawm melanogenesis ntawm zebrafish tau pom nyob rau hauv stereomicroscope.

UVB-Induced Hyperpigmentation Guinea Pig Model

Yim xim av guinea npua (6 lub lis piam, kwv yees li 300 g) tau txais los ntawm Lub Tsev Haujlwm Saib Xyuas Kev Tshawb Fawb Tsiaj (Beijing, Suav). Cov npua guinea tau nyob ib leeg hauv chav tswj qhov kub thiab txias raws li cov txheej txheem kuaj, nrog rau 12 teev lub teeb / 12 teev tsaus ntuj (lub teeb ntawm 9:00 AM txog 9:00 PM) nrog qhov kub thiab txias (23 ± 3 degree) thiab av noo (50 ± 10%). Cov cheeb tsam sib cais (1 cm diametrical voj voog) ntawm sab nraub qaum ntawm txhua tus tsiaj tau raug 500 mJ / cm2 UVB (Sigma SH-4, Shanghai, Tuam Tshoj) ib hnub ib zaug rau 1 lub lis piam (Fujimoto li al., 1988; Lee et al., 2013). Tom qab ntawd lub tsheb (PEG400 / EtOH 7: 3) thiab FGIN-1-27 (1 feem pua) tau muab rau cov cheeb tsam hyperpigmented (20 ul tov ib lub voj voog) ob zaug hauv ib hnub rau 3 lub lis piam. Lub degree ntawm pigmentation yog ntsuas los ntawm Spectrophotometer (YS3010, 3nh, Shenzhen, Tuam Tshoj). Qhov ΔL-tus nqi tau txiav txim siab raws li L-tus nqi ntsuas los ntawm spectrophotometer, raws li hauv qab no: ΔL L (ntawm txhua hnub ntsuas)-L (thaum hnub 0). Tag nrho cov txheej txheem tsiaj rau txoj kev tshawb fawb no tau ua raws li "NIH Phau Ntawv Qhia rau Kev Saib Xyuas thiab Kev Siv Cov Tsiaj Ua Haujlwm" thiab pom zoo los ntawm pawg saib xyuas tsiaj thiab siv ntawm Changzhou University.

Kev txheeb cais

TSEEM CEEB

FGIN-1-27 Inhibited -MSH, OAG lossis ET-1-Induced Melanin Synthesis hauv SK-MEL-2 Cells

Ua ntej tshawb xyuas qhov cuam tshuam ntawm FGIN-1-27 (Daim duab 1A) ntawm pigmentation, ib qho kev ntsuam xyuas ntawm tes tau ua los tshuaj xyuas cov teebmeem ntawm FGIN-1-27 ntawm SK-MEL-2 kev loj hlob ntawm tes. Raws li pom nyob rau hauv daim duab 1B, FGIN-1-27 tsis muaj kev cuam tshuam rau kev loj hlob ntawm SK-MEL- 2 hlwb ntawm qhov ntau npaum li cas ntawm 1-16 μM tom qab 48 teev. Hauv kev ntsuam xyuas cov ntsiab lus melanin, FGIN -1-27 inhibited basal melanogenesis (Daim duab 1C). -MSH yog lub elevator ntawm intracellular cAMP, uas ua rau muaj kev cuam tshuam melanogenesis (Rzepka li al., 2016; Corre et al., 2004; Lee et al., 2013). FGIN-1-27 kuj inhibited -MSH-induced melanin synthesis (Daim duab 1C). Raws li txoj cai, Masson-Fontana ammoniacal silver staining qhia tias FGIN -1-27 txo qis melanin concentration hauv SK-MEL-2 hlwb nrog lossis tsis muaj -MSH (Daim duab 1D). Tsis tas li ntawd, OAG yog lub elevator ntawm PKC thiab ET-1 yog tus activator ntawm MAPK txoj hauv kev, ob qho tib si tuaj yeem txhawb nqa melanin synthesis (Kim li al., 2006; Park et al., 2015; Regazzetti et al., 2015). Raws li pom nyob rau hauv daim duab 1E thiab 1F, FGIN-1-27 kuj tseem inhibited OAG los yog ET-1-induced melanogenesis.

FGIN-1-27 Tshem Tawm Tyrosinase, TRP-1, TRP-2 Kev nthuav qhia thiab txo qis Cellular Tyrosinase Kev Ua Haujlwm

Melanogenesis yog tswj los ntawm peb qhov tseem ceeb enzymes: tyrosinase, TRP-1, thiab TRP-2 (Zhu li al., 2015). -MSH thiab ET-1 txhawb nqa melanin synthesis los ntawm kev nce peb qhov tseem ceeb melanogenic enzymes qhia (Regazzetti li al., 2015; Lee et al., 2013). Txhawm rau tshawb xyuas seb qhov ua kom tawv nqaij dawb ntawm FGIN-1-27 puas cuam tshuam nrog cov proteins cov lus qhia, peb tau kawm txog cov txiaj ntsig ntawm FGIN-1-27 ntawm kev qhia ntawm tyrosinase, TRP-1, thiab TRP{{10} }} ntawm western-blot tsom xam. Raws li pom hauv daim duab 2A, FGIN-1-27 suppressed tyrosinase, TRP-1, thiab TRP{15}} qhia nrog lossis tsis muaj -MSH. FGIN-1-27 kuj tseem hloov pauv ET-1- induced tyrosinase, TRP-1, thiab TRP-2 kev nthuav qhia nce (Daim duab 2B). Tsis zoo li -MSH thiab ET-1, OAG txhawb nqa melanogenesis los ntawm kev ua kom cov cellular tyrosinase ncaj qha (D'Mello li al., 2016). Raws li pom hauv daim duab 2C thiab 2D, FGIN-1-27 kev kho mob 12 h inhibited OAG-induced cellular tyrosinase kev ua haujlwm nce thiab tsis cuam tshuam rau kev qhia ntawm tyrosinase. Tsis tas li ntawd, ib qho kev ntsuam xyuas nceb tyrosinase ua haujlwm los tshuaj xyuas qhov cuam tshuam ncaj qha ntawm FGIN-1-27 ntawm kev ua tyrosinase. Raws li pom hauv daim duab 2E, FGIN-1-27 tsis cuam tshuam rau cov haujlwm enzymatic ntawm nceb tyrosinase. Cov txiaj ntsig no tau qhia tias FGIN -1-27 txwv tsis pub cov lus qhia ntawm tyrosinase, TRP-1, thiab TRP-2, thiab txo qis kev ua haujlwm ntawm cellular tyrosinase, tab sis tsis muaj kev cuam tshuam ncaj qha rau cov kev ua ub no ntawm tyrosinase.

FGIN-1-27 Txo MITF kev nthuav qhia

MITF yog tus tswv transcriptional yam uas induces kev qhia ntawm tyrosinase, TRP-1, thiab TRP-2 (Kawasaki li al., 2008; Levy thiab Fisher, 2011). Raws li pom hauv daim duab 3A, FGIN-1-27 inhibited MITF transcription. Peb kuj tau ntsuas qhov qhia ntawm MITF tom qab SK-MEL-2 hlwb raug kho nrog -MSH nyob rau hauv lub xub ntiag los yog tsis muaj FGIN-1-27. Raws li pom hauv daim duab 3B, -MSH tau txhawb nqa qhov kev qhia ntawm MITF. Qhov zoo siab, qhov kev qhia ntawm MITF tau txo qis thaum muaj FGIN-1-27. FGIN-1-27 kuj tseem txwv tsis pub ET-1-induced MITF nthuav qhia (Daim duab 3C). Cov txiaj ntsig no tau qhia tias FGIN-1-27 inhibited qhov kev qhia ntawm MITF.

FGIN-1-27 Txo kev nthuav qhia ntawm PKC- , p-PKA Cat, p-CREB, p-p38, thiab p-ERK

PKA, PKC, thiab MAPK yog txoj hauv kev hloov pauv tseem ceeb hauv melanogenesis (D'Mello li al., 2016). Tus tub txib thib ob cAMP tuaj yeem qhib PKA, uas phosphorylates CREB, thaum kawg txhawb kev qhia ntawm MITF (Corre li al., 2004; Rzepka li al., 2016). PKC- -Txoj kev nyob ntawm kev tswj hwm melanogenesis los ntawm kev ua kom cov tyrosinase (Kim li al., 2006; Kawaguchi li al., 2012; Yuan thiab Jin, 2018). MAPK, suav nrog p38, extracellular signal-regulated protein kinase (ERK), thiab c-jun N-terminal kinase (JNK), tau tshaj tawm los tswj cov pigmentation (Zhou li al., 2014). Txhawm rau nkag siab ntxiv txog cov txheej txheem molecular ntawm kev tswj hwm melanogenesis los ntawm FGIN-1-27, peb tau ntsuas txoj hauv kev hloov pauv uas cuam tshuam nrog pigmentation. Raws li pom hauv daim duab 4, FGIN-1-27 tau txo qis kev qhia ntawm PKC-, p-PKA miv, p-CREB, p-p38, thiab p-ERK.

FGIN-1-27 Inhibited Melanogenesis hauv tib neeg Melanocytes

Qhov inhibition ntawm FGIN -1-27 ntawm melanogenesis raug soj ntsuam hauv tib neeg epidermis melanocytes (HEM). Raws li qhia hauv Daim Duab Ntxiv S1, FGIN-1-27 tsis muaj kev cuam tshuam rau kev loj hlob ntawm HEM ntawm kev noj ntau ntawm 1-16 μM tom qab 48 teev. Raws li pom hauv daim duab 5A, FGIN-1-27 cuam tshuam melanogenesis hauv HEM. Western blotting tsom xam pom tias FGIN-1-27 txo qis kev qhia theem ntawm tyrosinase, TRP-1, thiab TRP-2 hauv HEM (Daim duab 5B). Raws li qhia hauv Daim Duab Ntxiv S2, FGIN- 1-27 kev kho mob 12 h inhibited OAG-induced cellular tyrosinase kev ua haujlwm nce hauv HEM. Tsis tas li ntawd, peb ntsuas qhov kev qhia ntawm MITF tom qab HEM kho nrog -MSH nyob rau hauv lub xub ntiag los yog tsis muaj FGIN-1-27. Raws li tau pom hauv daim duab 5C, qhov kev qhia ntawm MITF poob qis heev thaum muaj FGIN-1-27. Tsis tas li ntawd, ib yam li hauv SK-MEL-2 hlwb, kev koom tes ntawm PKC- , PKA, thiab MAPK hauv FGIN-1-27-kev kho cov tshuaj tiv thaiv melanogenic kuj tau lees paub hauv HEM (Daim duab 5D).

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