Ishophloroglucin Ib Leeg Los Ntawm Ishige Okamurae Tshem Tawm Melanogenesis Induced Los Ntawm -MSH: Hauv Vitro Thiab Hauv Vivo Part 1

Abstract:Diphlorethohydroxycarmalol (DPHC) cais tawm los ntawm Ishige Okamura (IO) pom tias muaj peev xwm ua kom tawv nqaij tiv thaiv UV-B hluav taws xob. Txawm li cas los xij, cov khoom ntawm IO thiab lawv cov txheej txheem molecular tiv thaiv -melanocyte-stimulating hormone (-MSH) tseem tsis tau tshawb xyuas. Yog li, txoj kev tshawb no tsom mus tshawb xyuas cov teebmeem inhibitory ntawm Ishophloroglucin A (IPA), phlorotannin cais los ntawm cov xim av algae IO, thiab nws cov extract (IOE), nyob rau hauv melanogenesis hauv vivo nyob rau hauv ib tug-MSH-induced zebrafish qauv thiab hauv B16F10 melanoma hlwb. hauv vitro. Kev tshawb fawb molecular docking ntawm phlorotannins tau ua tiav los txiav txim siab lawv cov teebmeem inhibitory thiab nthuav tawm lawv cov kev cuam tshuam nrog tyrosinase, glycoprotein ntsig txog melanogenesis. Tsis tas li ntawd, morphological kev hloov pauv thiab cov ntsiab lus melanin poob qis hauv -MSH-induced zebrafish qauv tom qab kev kho IPA thiab IOE. Tsis tas li ntawd, Western blotting tau tshaj tawm tias IPA tau tswj hwm cov kab mob ntsig txog cov protein nyob hauv -MSH-stimulated B16F10 hlwb. Yog li, cov txiaj ntsig no qhia tias IPA cais tawm ntawm IOE muaj peev xwm siv tau hauv kev lag luam tshuaj thiab tshuaj pleev ib ce.

Raws li kev tshawb fawb cuam tshuam,cistancheyog ib hom tshuaj ntsuab hu ua "cov tshuaj ntsuab miracle uas ua rau lub neej ntev". Nws lub ntsiab tivthaiv yogcistanoside, uas muaj ntau yam teebmeem xws liantioxidant, anti-inflammatory, thiab tiv thaiv kev ua haujlwm nce qib. Cov txheej txheem ntawm cistanche thiab tawv nqaij dawb yog nyob 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 ntawm tyrosinase. 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 qhov 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.

1. Taw qhia

Marine algae tau txais txiaj ntsig zoo los ntawm cov khoom noj khoom haus, tshuaj pleev ib ce, thiab kev lag luam tshuaj thaum lawv siv cov khoom siv bioactive xws li phlorotannins, fucoidans, polysaccharides, thiab proteins [1-7]. Raws li kev tshawb fawb yav dhau los, Ishige Okamurae (IO) tau tshuaj xyuas rau nws cov teebmeem inhibitory ntawm tyrosinase kev ua haujlwm [8]. Txawm hais tias nws cov phlorotannin, diphlorethohydroxycarmalol (DPHC), tau raug soj ntsuam rau kev ua haujlwm los tiv thaiv melanogenesis tshwm sim los ntawm UV-B hluav taws xob hauv ib qho qauv hauv vitro [8], tsis muaj kev tshawb fawb ntxiv los piav qhia txog kev cuam tshuam ntawm DPHC thiab cov protein uas cuam tshuam txog melanogenesis. Nyob rau hauv txoj kev tshawb no, ib tug tshiab polyphenol compound, Ishophloroglucin A (IPA) [9], tau soj ntsuam rau nws cov kev sib cuam tshuam nrog tyrosinase nyob rau hauv ib tug molecular docking kawm, piv nrog cov DPHC.

Nyem rau ntawm Organic Cistanche Tubulosa rau Whitening

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Molecular docking yog ib txoj hauv kev ua kom muaj txiaj ntsig zoo los kwv yees cov peev txheej sib txuas ntawm cov molecules me lossis ligands nyob rau hauv qhov chaw ua haujlwm ntawm cov protein lossis cov receptor ntawm cov qauv peb-dimensional rau kev kawm lawv cov qauv kev sib cuam tshuam lossis rau kev tsim tshuaj [10–12]. Cov kev tshawb fawb yav dhau los tau tshaj tawm tyrosinase enzyme qhov chaw sib cuam tshuam thiab lub zog muaj nqis raws li kev tshawb fawb los ntawm cov kev tshawb fawb molecular docking [13,14]. Functionally, tyrosinase yog tus nqi-txheej enzyme uas catalyzes qeeb cov tshuaj tiv thaiv tus nqi hauv txoj kev taw qhia thiab ua lub luag haujlwm tseem ceeb hauv melanin biosynthesis hauv cov organelles tshwj xeeb hu ua melanosomes, tshwj xeeb los ntawm cov hlwb melanocytic [15]. Yog li, kev ua haujlwm enzymatic ntawm tyrosinase raws li qauv los ntawm hauv silico molecular docking simulations yog lub hom phiaj rau kev tshawb nrhiav cov tshuaj tiv thaiv kom tsis txhob muaj cov kab mob ntawm daim tawv nqaij hyperpigmented. Raws li qhov no, hauv kev sim thiab suav teb, ntau cov phenolic tebchaw los yog inhibitors muaj peev xwm hlau chelating tau dav kawm raws li tyrosinase inhibitors [16].

Melanin feem ntau hloov kho cov xim ntawm daim tawv nqaij thiab ua raws li cov xim tiv thaiv thaum daim tawv nqaij raug UV hluav taws xob [17]. Kev raug UV hluav taws xob ua rau kev tso tawm ntawm -melanocyte-stimulating hormone (-MSH) los ntawm cutaneous keratinocytes thiab melanocytes kom induce melanin synthesis thiab tom qab ntawd tiv thaiv daim tawv nqaij los ntawm kev puas tsuaj los ntawm hnub ci hluav taws xob [18]. Txawm li cas los xij, ntev mus rau UV hluav taws xob ua rau muaj qhov tseem ceeb melanin synthesis los ntawm -MSH, ua rau cov tawv nqaij puas xws li freckles, sunspots, thiab cov pob dub ntawm cov epidermis, ua rau DNA photodamage thiab mob qog noj ntshav [19]. Raws li qhov no, kev tswj hwm ntawm -MSH rau hauv melanocytes tau kawm dav dav kom ua rau muaj kev hloov pauv ntau hauv cov haujlwm tyrosinase rau melanogenesis hauv melanoma [20].

Zebrafish pom kev hloov pauv ntawm cov tawv nqaij xim zoo li tib neeg. Lawv yog cov qauv tsiaj siv los nkag siab txog keeb kwm caj ces ntawm tib neeg cov tawv nqaij xim, sib txuas cov qauv kab mob biology nrog tib neeg caj ces [21]. Tsis tas li ntawd, cov pigments ntawm lub ventral thiab lateral cheeb tsam ntawm zebrafish gastrula thiab lawv cov pob tshab thaum lub sij hawm larval theem tso cai yooj yim kev soj ntsuam ntawm cov txheej txheem pigmentation, muab lub sij hawm los tsim cov qauv siv tau rau kev nkag siab ntawm daim tawv nqaij mob uas tshwm sim los ntawm qhov tsis xws luag nyob rau hauv melanocyte txoj kev loj hlob [22- 24] ib.

Melanin synthesis tshwm sim los ntawm kev ua haujlwm ntawm ntau cov proteins uas cuam tshuam nrog melanogenesis, xws li tyrosinase, microphthalmia-associated transcription factor (MITF), tyrosinase-related protein-1 (Trp-2), thiab tyrosinase-related protein{ {6}} (Trp-1) [15]. Nyob rau hauv txoj kev tshawb no, peb tsom mus soj ntsuam cov nyhuv inhibitory ntawm IPA thiab IO crude extract (IOE) nyob rau hauv tyrosinase kev ua thiab melanogenesis ntawm -MSH-induced zebrafish nyob rau hauv vivo thiab B16F10 melanoma hlwb hauv vitro los soj ntsuam lawv cov peev xwm siv los kho cov tawv nqaij pigmentation thiab melanoma. .

2. Cov txiaj ntsig

2.1. Molecular Docking Study

Ib txoj hauv kev docking tau ua los simulate kev khi ntawm kev lag luam tyrosinase enzyme nrog IPA, DPHC, thiab arbutin [25]. Cov hom kev khi ntawm IPA, DPHC, lossis arbutin nrog nceb tyrosinase tau nthuav tawm hauv daim duab 1A-C. Raws li pom nyob rau hauv daim duab 3D ntawm tyrosinase-IPA complex (Daim duab 1A), tyrosinase-DPHC (Daim duab 1B), thiab tyrosinase-arbutin (Daim duab 1C), txawm hais tias IPA, DPHC, thiab arbutin docked rau qhov chaw nquag, tyrosinase- IPA complex pom qhov loj tshaj plaws kev sib khi nto thaj tsam. Tsis tas li ntawd, raws li pom nyob rau hauv daim duab 2D, tyrosinase-IPA complex pom muaj kev cuam tshuam ntau dua nrog ntau cov amino acids ntawm tyrosinase, piv rau tyrosinase-DPHC lossis tyrosinase-arbutin. Xya lub nplhaib benzene (1st, 2nd, 3rd, 8th, 9th, 12th, thiab 16th) thiab lawv cov pa atoms muaj π-π cuam tshuam nrog Histidine60, Methionine61, Lysine157, Glutamate158, Proline1160, Proline220, . Tshwj xeeb, 16th benzene nplhaib tau ua ke nrog Histidine60 thiab Histidine204, cov amino acids tseem ceeb ntawm qhov chaw ua haujlwm. Tsis tas li ntawd, ntau cov pa atoms hauv IPA cuam tshuam los ntawm hydrogen bonds nrog Glutamate158, Proline160, Aspartate167, Methionine184, Phenylalanine197, Asparagine199, Glutamine202, Asparagine205, thiab Arginine209. Tsis tas li ntawd, raws li cov txiaj ntsig kev soj ntsuam docking, nws tau suav tias tag nrho cov kev sib txuas zog thiab CDOCKER kev sib cuam tshuam lub zog siv CDOCKER kev sib cuam tshuam lub zog ntawm DS 3.0 (Daim duab 1D) tau raws li hauv qab no: kev sib cuam tshuam lub zog nrog tyrosinase: Ishophloroglucin A (IPA): - 152.154 kcal/mol, diphlorethohydroxycarmalol (DPHC): −65.5221 kcal/mol thiab arbutin: −33.6835 kcal/mol thiab lub zog khi nrog tyrosinase: Ishophloroglucin A (IPA): −546.504 kcal / mol 7 DP, DPHC mol thiab arbutin: -79.0913 kcal / mol.

2.2. Cov teebmeem ntawm Melanogenic Inhibitors ntawm Melanin Synthesis hauv Zebrafish Larvae

Lub hom phiaj ntawm txoj kev tshawb no yog los txiav txim seb IPA thiab IOE puas tuaj yeem cuam tshuam melanogenesis hauv zebrafish larvae hauv vivo. Hauv peb txoj kev tshawb fawb yav dhau los, IPA thiab IOE pom tias tsis muaj tshuaj lom nyob rau hauv zebrafish larvae [26]. Peb kho zebrafish larvae nrog zebrafish (0.05, 0.15, thiab 0.5 nM) thiab IOE (3, 10, thiab 30 µg/mL ) los txiav txim lawv cov ntsiab lus melanin. Txhawm rau kwv yees cov haujlwm inhibitory, peb ntsuas tag nrho cov ntsiab lus melanin ntawm zebrafish extracts. Cov nyhuv phenotypic ntawm zebrafish melanin tau pom los ntawm kev tshuaj xyuas lawv lub cev pigmentation nyob rau hauv lub tshuab tsom iav ntawm chav tsev kub. Nto melanin tau txo qis los ntawm ntau qhov ntau ntawm cov hom phiaj inhibitors (Daim duab 2). Kev kho mob nrog 3 nM -MSH ua rau muaj zog melanin cov ntsiab lus ntawm zebrafish piv nrog cov pab pawg tsis kho. Qhov siab tshaj plaws ntawm IPA (0.5 nM) thiab IOE (30 µg / mL) txo tag nrho cov ntsiab lus melanin los ntawm yuav luag 28.56 feem pua ​​​​thiab 30.36 feem pua ​​​​, feem, uas nthuav tawm cov teebmeem zoo ib yam li arbutin (200 µM)-kho pab pawg (Daim duab 2A, B).

2.3. Cov teebmeem ntawm IPA thiab IOE ntawm Melanin Synthesis hauv B16F10 Cells

Cov teebmeem cytotoxic ntawm IPA thiab IOE ntawm B16F10 hlwb raug ntsuas los ntawm MTT (3-(4,5-dimethylthiazol-2-yl)-2,{{ 7}}diphenyltetrazolium bromide) kev soj ntsuam. Peb pib tshuaj xyuas cov teebmeem cytotoxic ntawm IPA thiab IOE ntawm B16F10 hlwb kho nrog ntau qhov sib txawv yam tsis muaj -MSH stimulation. IPA (0.15, 0.5, 1.5, thiab 5 nM) tsis qhia qhov tseem ceeb cytotoxicity ntawm B16F10 hlwb, thaum IOE (1, 3, 10, thiab 30 µg / mL) tsis pom muaj cytotoxicity (Daim duab 3A, B). Cov concentrations ntawm IPA thiab IOE uas tsis muaj tshuaj lom tau siv rau kev kawm ntxiv.

Peb tau tshuaj xyuas seb qhov cuam tshuam ntawm melanin inhibition los ntawm IPA lossis IOE kev kho mob hauv vivo puas rov qab los yam tsis muaj -MSH stimulation hauv B16F10 hlwb. Cov ntsiab lus melanin tau txo qis thaum kho nrog IPA (5 nM) lossis IOE (30 µg / mL) piv rau pawg tswj hwm (Daim duab 3C, D). Cov txiaj ntsig no tau qhia tias IPA thiab IOE inhibited melanin synthesis yam tsis muaj -MSH stimulation hauv B16F10 hlwb, yog li ua rau nws cov nyhuv anti-melanogenesis.

2.4. Cov teebmeem ntawm IPA thiab IOE ntawm Tyrosinase Kev Ua Haujlwm thiab Melanin Synthesis hauv -MSH-Stimulated B16F10 Cells

Yog li ntawd, koob tshuaj IPA (0.5, 1.5, thiab 5 nM) thiab IOE (3, 10, thiab 30 µg/mL) tau tsim nrog B16F10 melanoma hlwb los txiav txim siab lawv cov bioactivity hauv cellular tyrosinase kev ua haujlwm thiab -MSH - kho melanogenesis.

Qhov siab ntawm -MSH thiab arbutin tau txiav txim siab raws li cov txiaj ntsig ntawm lawv cov cytotoxicity thiab melanin ntau lawm hauv B16F10 hlwb (Daim duab S2A-D). Los ntawm kev txheeb xyuas cov ntaub ntawv ntawm kev ciaj sia, 1 nM ntawm -MSH thiab 100 µM ntawm arbutin tau siv rau kev sim ntxiv.

Raws li kev ua haujlwm ntawm cellular tyrosinase, txawm hais tias cov teebmeem inhibitory ntawm 1.5 nM ntawm IPA kuj qis dua li cov uas tau piav qhia hauv kev tswj tau zoo, qhov sib txawv ze li ntawm 10 npaug, nrog rau qhov siab dua ntawm arbutin ntawm 100 µM (Daim duab 4A). IOE txo qis kev ua haujlwm tyrosinase nrog kev hloov pauv me ntsis ntawm qhov cuam tshuam ntawm ntau qhov sib txawv piv rau -MSH-kho pab pawg (Daim duab 4B). Raws li rau -MSH-mediated melanogenesis, qhov tseem ceeb txo cov ntsiab lus melanin tau pom nyob rau hauv 1.5 thiab 5 nM IPA-kho pab pawg (Daim duab 4C). Tsis tas li ntawd, 30 µg / mL IOE inhibited pigmentation mus rau qhov chaw uas zoo li cov pab pawg dawb paug (Daim duab 4D). Cov txiaj ntsig no qhia tias IPA thiab IOE txo qis kev ua haujlwm ntawm tyrosinase thiab tias cov nyhuv inhibitory no tuaj yeem ua rau txo qis cellular melanin synthesis hauv B16F10 hlwb.

2.5. Cov teebmeem ntawm IPA thiab IOE ntawm Molecular Mechanism hauv -MSH-Stimulated B16F10 Cells

Txhawm rau elucidate lub luag haujlwm rau lawv cov nyhuv melanogenesis inhibitory, peb txiav txim siab qhov cuam tshuam ntawm IPA thiab IOE ntawm kev qhia theem ntawm kev taw qhia cov molecules koom nrog hauv melanin synthesis (Daim duab 5). ERK (extracellular signal-regulated kinase), JNK (Jun N-terminal kinase), thiab p38 belongs rau mitogen-activated protein kinase (MAPK) intracellular signal transduction cascade [27,28]. Raws li tau hais hauv daim duab 5A, B, ERK phosphorylation tau txhim kho nrog kev kho IPA, qhov phosphorylation ntawm JNK tau qis me ntsis tom qab kev kho nrog IPA (1.5 thiab 5 nM) lossis IOE (30 µg / mL). Tsis tas li ntawd, raws li tau hais hauv daim duab 5C, p-p38 qib tau nce siab hauv pawg -MSH-stimulated, nyob ib ncig ntawm 80 feem pua ​​​​piv rau kev tswj hwm. Tsis tas li ntawd, tom qab kev kho mob nrog IPA, IOE, lossis arbutin, phosphorylation ntawm p38 tau raug tshem tawm me ntsis. Cov kev tshawb pom no qhia tias cov teebmeem melanogenic inhibitory ntawm IPA thiab IOE tuaj yeem cuam tshuam nrog JNK thiab p38 MAPK txoj kev taw qhia.

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