Ta'sir of Sucrier Banan Peel Ekstraktlari On Inhibisyon Of Melanogenez orqali The ERK Signalizatsiya yo'li
Apr 25, 2023
Abstrakt
Hyperpigmentation is a type of pigmentary disorder induced by overexpression of melanin content activated severe esthetic problems such as melasma, freckle, ephelides, lentigo, and other forms on human skin. Several whitening agents have restricted use because of their side effects or stability such as kojic acid, ascorbic acid, and hydroquinone can act as cytotoxic substances associated with dermatitis and skin cancer. To find a safe substance, this study aimed to find the ability of several components in Sucrier banana peel (SBP) extracts to inhibit the melanogenesis process through the p38 signaling pathway in B16F10 mouse melanoma cells. Tyrosinase activity and the cellular melanin content were dose-dependent manners decreasing after SBP treatment. Furthermore, SBP decreased the expression of melanogenesis relate proteins as a microphthalmia-associated transcription factor (MITF) and tyrosinase protein after 24 hours of incubation with -melanocyte stimulating hormones (MSH) stimulating. The findings demonstrated that SBP contained an effective agent for hyperpigmentation inhibitors through p38 signaling pathways without any effect on the ERK pathway, and subsequently down-regulate MITF expression and tyrosinase enzyme family production.
cistanche is a keng tarqalgan o't ning nomi "the mo''jiza o't 22 umrni uzaytiradi. Uning asosiy komponenti hisoblanadisitastanozid% 2c turli taʼsir lar kabi taʼsirga egaantioksidant, yallig'lanishga qarshi% 2c va immunitet funksiya rag'batlantirish. The mexanizm orasidagi cistanche and skin whitening yotadi the antioksidant ta'siri of cistanche glikozidlar. Melanin in odam terisi tomonidan hosil bo'ladi tyrosin katalizator by tyrosinase, va the oksidlanish reaktsiya ning ishtiroki kislorod, shunday bo'ladi kislorodsiz radikallar ichida tana bo'ladi muhim omil ta'sir melanin ishlab chiqarish. Sitanche tarkibida sitstanosid% 2c qaysi an antioksidant va kamaytirish mumkin mumkin avlod erkin radikallar ichida tanasi, shunday inhibisyon melanin ishlab chiqarish.
Bosing on Qaerda Mumkin I Sotib olish Cistanche
david.deng@wecistanche.com WhatApp:86 13632399501
Kalit soʻzlar: sukri banan pech; fenolik birikmalar; tyrosinaza; mikrofaolmiya-bog'liq transkripsiya factor; melanogenez
Tanishuv
Melanin is produced within the melanosome which is an important organelle inside the melanocyte cells. Melanocyte is located in the basal layer of the skin epidermis. After the production process, melanin is the pigment responsible for skin, hair, and eye coloration which functions as a naturalized antioxidant, detoxification agent, and powerful-cation chelator and also can act as universal protection against UV damage [1]. Abnormal melanin production can cause pigmentary disorders such as hypopigmentation and hyperpigmentation. Hypopigmentation causes vitiligo, albinism, and abnormal hair problems while hyperpigmentation causes freckles, melasma, age spots, and post-inflammatory hyperpigmentation. Genetic predisposition, hormonal changes especially estrogen also others such as liver disease, tumor, cancer, malnutrition, or irregular function of the pituitary gland are the intrinsic factors including extrinsic factors like UV exposure, and toxic substances are the major cause of hyperpigmentation [2].
Tyrosinase is a rate-limiting step enzyme that targets the process of both stimulating and inhibiting the melanin production process. Melanin stimulation is used as a tanning agent or hair depigmentation treatment. Tyrosinase expression is controlled by microphthalmia-associated transcription factor (MITF) which is a melanocyte-specific transcription factor that controlled melanocyte differentiation, proliferation, and survival [3, 4]. The responsibility of MITF to melanogenesis is to increase tyrosinase expression by binding to DNA in the structure of homodimer or hetero dimer with MiT protein, these binding site involving E-Box and M-Box flanking thymidine nucleotide which stimulates the continuous steps to melanogenic enzyme production such as tyrosinase-related protein 1 (TRP-1), tyrosinase-related protein-2 (TRP-2) and dopachrome tautomerase. Regulation of MITF starts from signal transduction after -MSH binds to the MC1R receptor which stimulates mitogen-activated protein kinase (MARKs) which is serine/threonine kinase and also includes extracellular signaling-regulated kinase (ERK) and also p38 as well. Several studies found that melanin synthesis is controlled by several signaling pathways such as phosphotidylineasital-3-kinase (PI3K/AKT) can be suppressed by natural substances which contain polyphenol group through the ERK signaling pathway of B16F10 mouse melanoma cells [5,6].
Ushbu tadqiqot to lashga mo'ljallangan ta'sir c2�melanogenez inhibisyon dan Sucrier banan peel (SBP) ekstraktlar. S BBP may tarkibida bir nechta turdagi 2 polifenol birikmalar bo'lishi ki ishlaydi tyrosinaza inhibitorlar masalan katechin, procyanidin, ferulik kislota, va gallik kislota kislota qaysi kislota tomonidan aniqlanadi tomonidan erK signaling ta'sir qiladigan ta'sir qiladigan yo'llar ta'sir c2 melanogen fermentlar masalan TRP-1 va TRP% 7b{1}} [7-10].
Materiallar va usullari
Kimyoviy va materiallar
Yirtilgan (7 kun yig'im-terimdan keyin) Sucrier banana (Musa spp.) peels dan Kampangpetch Province, Tailand. Yuvilgan banan pechlari bilan suv then quyosh-pishirilgan qadar butunlay quritilgan%1 2c zamin bo'yicha blender, and qazib olingan by cho'ktirish bilan 60 foiz metanol uchun 24 hours (MW24))% 2c centrifuged bilan 95 foiz etanol uchun 10 minutes at at 4 degree (E4), qaynoq in DI suv at 60 degree for 20 minutes (W60) va qaynash ichida 95 foiz da metanol at 60 daraja for 20 daqiqa (M60). Keyingi qadam 2%% a0volume bilan a rotatary bugaporator at 40 daraja va c2�dry by freeze dryer Christ Alpha- 4 LD plus.
Katak madaniyat
B16F10 sichqoncha melanoma katak lari o'stirilgan in Dulbecco's modified Eagle's medium (DMEM)� �supplemented bilan 10 foiz homila sigir serum at 37 daraja da� an inkubatsiyalangan bilan 5 foiz CO2. � 24 soat dan so'ng inkubatsiya, then o'zgardi o'zgardi o'rta to serumsiz media bilan c2�different konsentratlari of Sucrier banan pech (SBP) extracts bilan -MSH stimulyatsiyasi bilan banan pech (SBP) extracts. Keyin hujayra hujayralari c2�harvested by trypsinization for the keyingi tajribalar.
Uyali hayotiylik tahlil
SBP ekstraktlarining sitotoksikligi MTT assay bilan o'lchangan. The hujayra (5 ×{{{}) urug'langan 96 quduq plates va o'stirilgan 0platlar olindi in DMEM for 24 hours at 37 degree in an inkubed with 5 percent CO2 then changed the 2 to serumsiz media with har xil konsentratlar c2�SBP ekstraktlari soat 48 soat davomida. davodan keyin, tashlab yuborilgan the orta va yuvilgan ikki marta sovuq PBS, 100 ul of MTT eritmasi (5mg/ml)) qo'shildi har bir quduq, inkubated da 37 daraja uchun 4 soat so'ngra o'lchandi c2�ELISA o'quvchi da 420 nm.
Melanin tarkibi assay
Yig'ib olingan hujayralar lised in sovuq lizis bufer (20 mM natriy fosfat pH 6,8, 1 foiz tri Xton{-100}, 1mM PMSF, va 1 mM EDTA) va keyin santrifüjlangan 2c%1 %1,200 rpm at 4 degree uchun 10 daqiqa alohida supernatant while melanin pellet lar eritildi eriydi in 200 ul 1N NaOH uchun 60 daqiqa at 80 daraja . ELISA o'quvchi uchun c2�used uchun o'lchash the the absorbance at 415 nm. and calculated compared the protein content from bicinchoninic acid (BCA) protein tarkibi assay (Pierce Biotechnology, Packford, IL, AQSh).
G'arbiy blot analiz
2 progression of hujayra lizisi SDS-PAGE va keyin o'tkazilgan ga Hybond kengaytirilgan cheluminescence nitrocellulose olindi membran (Amersham, Little Chalfont, UK) va probe bilan birlamchi antibody qarshi tirosinaza (Santa Cruz, Biotexnologiya, Santa Cruz, CA, U.S. A), MITF ( Merck Millipore Darmstadt, Germaniya), ERK (Merck Millipore Darmstadt, Germaniya), P38 ( Merk Darmstadt Millipore, Germany), va monoklonal antikorlar qarshi%c%c 2�B-Actin (AC{-15}, Sigma Aldrich). From then on, incubat membrana bilan Horseradish peroksidaza-konjugated ikkilamchi antikor kompleks vizualizatsiya� and kvantlangan the signals by ImageJ software.
Statistik tahlil All ma'lumot tahlil qilindi o'rtacha ± standard deviation (SD). The One-way ANOVA test is used to measure farqlar orasidagi farqlar har bir to'plam of informatsiya. A P-qiymati kam %{{0.05 c2�considered statistik ahamiyatga ega bo'ldi.
Natijalar
The objective of this experiment was to measure the efficacy of the solvents used for extraction and also a method that can extract phenolic compounds as much as possible. Most phenolic compounds dissolve very well when extract with high polar solutions such as water, methanol (MeOH), ethanol (EtOH), acetone, ethylacetate, propanol, acetic acid, or their mixture in various portions [11]. Figure 1A shows that W60 can extract phenolic compounds as much as 16.24 µg/ml followed by M60, E4, and MW24 with total phenol 13.89, 9.21, and 8.38 µg/ml respectively. Cytotoxicity of SBP was experimented with by MTT reduction assay which measured the reduction environment from the mitochondrial function of living cells by measuring the formazan formation to access cytotoxic effects. The result found that SBP at indicated concentrations (up to 500 µg/ml) with 48 hours incubation. Cytotoxicity shows no significant effect after SBP treatment (Fig. 1B).
The effects of SBP on enzyme activity and melanin content in B16F10 cells
To test melanogenesis suppression of SBP, the first step was to test the capability of mushroom tyrosinase enzyme inhibition compared to kojic acid which is a commercial whitening agent. SBP (MW24) has higher efficacy than Kojic acid and other extractions at the same concentration (100-500 µg/ml). MW24 has the highest percentage of tyrosinase inhibition (66.39-77.05 percent ) while kojic acid only has a percentage of tyrosinase inhibition at 44.68-59.93 percent as shown in Fig. 1C. From then on, MW24 with the highest percentage of tyrosinase inhibition was selected for anti-melanogenesis measurement by melanin production from B16F10 cells. The melanin content of B16F10 cells was reduced after SBP (MW24) treatment (Fig. 2A). The amount of melanin content of B16F10 cells decrease significantly after stimulated with -MSH and treated with SBP (MW24) for 24 hours in a dose-dependent manner as shown in Fig. 2B.
SBP (M24) ning oqsil ifodasi levels of tyrosinase va MITF
G'arbiy blot analiz ishlatilgan uchun tirosinaza and MITF faolligi by o'lchash oqsil ifoda levels dan so'ng davolangan bilan SBP bilan (MW24) for 24 soat. melanogen fermentlarning bunday TRP-1}}}%2%2%2%2%2%2%1%1%1%1%2%1 2c va TRP{{3{3}} natijada the transkriptional effect of MITF and tirosinaza dan kelib chiqdi oqsil darajasi. The results show that SBP (MW24) ning qobiliyati bo'lgan dozaga bog'liq down-tartibga solish ikkala tirosinaz va MITF oqsil expression as ko'rsatilgan in Fig 3.
SBP (M24) on melanogenez inhibition orqali mitogen-faollashtirilgan oqsil kinaza (MAPK) signalizatsiya yo'li
In melanogenez, the MAPKs kinase family especially ERK and p38 are related directly to this procedure %[12]. The ERK signalizatsiya will down-regulate w ammo p38 signalizatsiya will up-regulate during the occurrence of melanin production. However,� phosphorylation of ERK and p38 will have a reverse procedure of melanin production. Natijalar ko'rsatildi c2�clear effect after SBP (MW24) davolash bilan -MSH faollashtirilgan by immunoblotting to ning a MAPK signalizatsiya pathway. SBP (MW24) biroz kamaygan p38 oqsil expression va up-regulate fosph p38. natija ko'rsatdi yo'l to inhibit the melanogenez process through p3 yo'l yo'l siz hech ta'sir ta'sir � ERK lekin fosforilatsiya of ERK oqsil darajasi slightly kamaygan keyin -MSH va SBP (MW24) treatment as ko'rsatilgan in Fig. 4.
Munozara
Sucrier banana peels are agricultural waste that has an abundance of many active ingredient substances, especially in the group of phenolic compounds and carotenoids which are the secondary metabolite that plants create for self-defense mechanisms [7]. These substances can be found in leaves, bark, fruits, peels, and seeds of almost all plants. Phenolic compounds and carotenoids are phytochemical substances that have good properties for human health. The substances comprise anti-inflammatory, anti-viral, analgesic, antioxidant, anti-carcinogen, etc [13]. In general, phenolic compounds can be well dissolved in high-polar solvents. For the experiment, the extraction of MW24 should be able to extract the phenolic compounds the most because of the procedure of extraction from the Folin-Cicualteu assay. Low polar compounds were extracted first then follow by high polar substances depending on the final mixing ratio of solvent in the procedure. The results show that W60 can extract phenolic compounds the most, possibly due to 60 degrees Celsius temperature which could be the appropriate temperature to extract phenolic compounds [11]. It can also be assumed that SBP may contain high polar substances which dissolve well in water. When comparing the amount of total phenolic compounds as shown in Fig. 1A, W60, M60, E4, and MW24 demonstrated the possibility of substance solubility, significantly.
After the total amount of phenolic compound was known, we can proceed to pinpoint the type and amount of flavonoid and carotenoid substances. We found SBP (MW24) extracts contain ferulic acid as high as 906.62 mg/100g and also found a small amount of lutein and -carotene which were categorized in the carotenoid group at 1.23 and 2.37 mg/100g respectively. SBD (MW24) still has a high percentage of tyrosinase inhibition. Therefore, SBP (MW 24) was selected for further study.
Dan sitotoksiklik test, no ahamiyatli cytotoxic dan SBP dan topildi garchi c2�usage konsentratsiyasi dan 50 gacha bo'lgan 0 c2�g/ml. ehtimol because the small amount of karotenoid ichida extraction 2�extraction ning qobiliyati mavjud himoyalovchi hujayralar hujayralari. % c2�Substances in this group have antioxidant properties that can terminate poison through Cytochrome� %�P{{1{1}}} ayniqsa -karotin bor a qo'zg'atuvchi rol ichida c2�immune funktsiya 7d 7d 7d ayniqsa karotin bor c2�immune funktsiya larda qo'zg'atuvchi rol bilan o'zaro aloqa orqali reaktsiya Oxygen Species (ROS). Bu iroda qo'llab-quvvatlaydi the samaradorligi of a fenolik birikmalar mavjud bo'lgan ROS skavenging effect property to decrease yallig'lanish sitokin [14]. Shunday qilib, SBD (MW24) nafaqat toksik ta'sirga ega 2c u has roli hujayra himoyasi yaxshi.
Our study also found that SBP (MW24) has the capability of inhibiting the melanogenesis of B16F10 cells. The expression of tyrosinase and MITF were reduced after SBP (MW24) treatment in both assessed with mushroom tyrosinase and Western blot analysis in a dose-dependent manner. This test indicated that SBP (MW24) can suppress the creation of melanin by down-regulate p38 signaling pathway and up-regulate phosphorylation of p38 which activate MITF protein degradation and then effect to decrease the expression of melanogenic enzyme family such as tyrosinase. Suppression of p38 decreases cAMP Response Binding Protein (CREB) functions which is an important transcription factor that works with PAX3 and SOX10 that attaches to M-BOX of MITF gene to activate MITF protein expression that affects the melanin production process and cells survival activity [15]. In general, activating ERK leads to phosphorylation of MITF at serine 73 which causes ubiquitination and eventually degradation. Most natural compounds can inhibit melanogenesis through the stimulation process of the ERK signaling pathway, but SBP (MW24) was not significantly influenced the ERK signaling pathway [16].
Many research found that phenolic compounds and carotenoids can inhibit the production of melanin pigments because beta-carotene has the capability of absorbing UVB photons while also has the property of the lipid-soluble antioxidant and phenolic compound can activate Nrf2, blocking ROS receptors, decrease inflammatory cytokine production and induce y-GSC expression which support to our study but in the different way of melanogenesis inhibition [17]. Other than that ferulic acid found in SBP (MW24) was the main substance within the group of phenolic compounds confirmed by several studies that it has the property of anti-melanogenesis because it can modulate vascular endothelial growth factor (VEGF) expression, induces nitric oxide synthase, act as tumor suppressor gene and also relate to the procedure of melanin pigments production. In SBP (MW24) solution, ferulic acid can increase effectiveness when mixed with carotenoid or other phospholipids because the water solubility, lipid solubility, and bioavailability will be increased resulting to improve melanogenesis inhibition of B16F10 cells [18]. The implication of all the results concludes that SBP (MW24) could reduce melanogenesis effectively.
Qisqartmalar
MITF: mikrophthalmia-associated transkripsiya factor; -MSH: melanocytes stimullovchi gormon; MC1R: melanokortin{{4} } retseptor%; TRP1: tirosinaz bilan bog'liq oqsil 1; TRP2: tirazaza bilan bog'liq oqsil 2; MAPK: mitogen-faollashtirilgan oqsil kinaz.
E'tiroflar
Bu ish qo'llab-quvvatlandi grantlar dan the Royal Golden Jubayr Ph.D.: RGJPHD. Tailand. (PH.D./0017/2558).
Muallif hissalar
R.H., K.T., K.S.%, va U.P. homilador va mo'ljallangan the tajribalar; R.H. C. C.H. va M.C. bajarildi the experiments; 3b tajribalar R.H. va M.C. va C.H.L tahlil qildi c2�data; R.H. qo'shilgan reagentlar/material/analiz toollar; R.H. va C. H. L. qog'oz yozdi qog'oz.
Raqobatchi qiziqishlar
Mualliflar no raqobatchi interest mavjudligini e'lon qilishdi.
Maʼlumot
1.Regnault MR, Gadaud N, Boulinguez S, Tournier E, Lamant L, Gladieff L, et al. Kimyoterapiyaga oid retikulatsiya giperpigmentatsiya: a holat seriya va review of the literature. Dermatologiya 2015; 231: %312-8.
2. Ali A. Dermatologiya A Pictorial Review. McGraw Hill Education 2010. ISBN978-0-07-179323-0.
3. Garraway LA, Widlund R, Rubin MA, Getz G, Berger AJ, Ramaswamy S, Beroukhim R, et al. Integrative genomic analyses identify MITF as a lineage survival oncogene amplified in malignant melanoma. Nature 2005; 436: 117–22.
4. Busca R, Ballotti R. Cyclic AMP a key messenger in the regulation of skin pigmentation. Pigment Cell and Melanoma Research 2000; 13: 60-9.
5. Zhou J, Ren T, Li Y, Cheng A, Xie W, Xu L, et al. Oleoylethanolamide inhibitsiyalar -melanocyte stimulyatsiyalovchi gormon stimulyatsiyalangan melanogenez via ERK, Akt,%Akt, and CREB signalizatsiya yo'llari in B16 melanoma kataklar. Oncotarget 2017; 8: {56868-79}.
6. Onkoksoong T, Jeayeng S, Poungvarin N, Limsaengurai S, Thamsermsang O, Tripatara P, Akarasereenont P, Panich U. Tay o'simlik antipiretik 22 formula (APF22) inhibitlar UVA vositali melanogenez orqali � Nrf2-% 7dregulyatsiyalangan antioksidant himoya. Fitoterapiya tadqiqotlar 2018; 32: 1546-54}.
7. Singh B, Singx JP, Kaur A, Singh N. Bioaktiv birikmalar in banan va%c 2�their bog'liq sog'liq foyda foyda - A review. Oziq-ovqat kimyo 2015; 1: 1-11}.
8. Leerach N, Yakaew S, Phimnuan P, Soimee W, Nakyai 2c W, Luangbudnark W, Viyoch J. Effect of Thai banan (Musa AA group) in in kamaytirish the the to'planish of oksidlanish end mahsulotlar in UVB-nurlangan sichqon terisi. Journal of Fotokimyo %& Fotobiologiya, B: Biologiya 2017; 168: 50-58.
9. Hung BY, Kim SY, Jung JM, Won CH, Choi JH, Lee MW. The antimycotic agent clotrimazole inhibits melanogenez by tezlatgich ERK va PI3K-%/Akt-vositali tirosinaza degradatsiyasi. Eksperimental Dermatologiya 2015; 24: 381-400.
10. Shen T, Heo SI, Wang MH. Ishtirok of the p38 MAPK and ERK signalizatsiya pathway in of anti-melanogen effect of metil 3,{{4{4%. 7d} caffeoyl kvinat in B16F10 sichqoncha melanoma hujayra. Chemico-Biologik O'zaro ta'sir 2012; 199: 106-11.
11. Ruesgas-Ramón M, Figueroa-Espinoza MC, Durand E. Application of Deep Eutectic Solvents (DES) for Fenolic Compounds Extraction: Overview , Challenges, va Imkoniyatlar. Journal of Qishloq xoʻjaligi va oziq-ovqat kimyo 2017; 10: %3591-3601.
12. Tsao YT, Kuo CY, Kuan YD, Lin HC, Wu LH, Lee CH. The Extracts of Astragalus membranaceus Inhibit Melanogenez orqali the ERK Signaling Pathway. International Journal of Medical Sciences 2017; 3: 1049-53.
13. Chang TS. An yangilangan tekshirish tyrosinase inhibitörleri. International Journal of Molecular Sciences 2009; 26: 2440-75.
14. Juturu V, Bowman JP, Deshpande J. Umumiy teri tonusi va skin-lightening-yaxshilovchi ta'siri bilan oraliq qo'shimcha bilan lutein va zeaxanthin is 3a a ikki ko'r, placebo-boshqariladigan klinik sinov. Klinik, Cosmetic and Investigational Dermatologiya 2016; 7: 325-332}.
15. Bell RE, Levy C. The three m's: melanoma, mikro-oftalmiya-bog'liq transcription faktor%% 2c va microRNA. Pigment Cell Melanoma Research 2011; 24: 1088–106.
16. Zhou J, Ren T, Li Y, Cheng A, Xie W, Xu L, Peng L. Oleoylethanolamide inhibits -melanocyte stimulating gormon-stimulyatsiyalangan melanogenez via ERK, Akt, va CREB signalizatsiya yo'llari in B16 melanoma hujayralarida. Oncotarget 2017; 23: 56868-79}}.
17. Toews DPL, Hofmeister NR, Taylor SA. The Evolution and Genetics of Caro 3b 33: {{3{3}}}}}.; 3b 1 1 20 %3; 7d 7b} 7b%3; 1 3%3%1%1 1 1 1 3%2%1%1%1%1%1%1 3%33%1%1%1%
18. Li L, Liu Y, Xue Y, Zhu J, Wang X, Dong Y. Preparat of a ferulik acid-fosfolipid komplekt to improve solubility, dissotsiv, va B16F10 cellular melanogenez inhibisyon faolligi. Kimyo Markaziy Jurnal 2017; 22: 11-26}.
For more info: david.deng@wecistanche.com WhatApp:86 13632399501
