Inhibitory effect of Cyrtomium falcatum on melanogenesis in α-MSH-stimulated B16F10 murine melanoma cells

Objective:To explore the anti-melanogenic potential of Cyrtomium falcatum.Methods:The effects of Cyrtomium falcatum crude extract and its solvent fractions on tyrosinase activity,melanin content,and the expressions of melanogenesis-related genes and proteins were analyzed inα-melanocyte-stimulating hormone(α-MSH)-stimulated B16F10 cells.Results:α-MSH treatment significantly increased tyrosinase activity,and extracellular and intracellular melanin content,as well as the expression levels of tyrosinase,microphthalmia-associated transcription factor(MITF),tyrosinase-related protein(TRP)-1,and TRP-2 in B16F10 cells.Treatment with Cyrtomium falcatum crude extract and its solvent fractions reduced tyrosinase activity and extracellular and intracellular melanin content and downregulated the expression levels of tyrosinase,MITF,TRP-1,and TRP-2 in a dose-dependent manner.Conclusions:Cyrtomium falcatum has potential anti-melanogenesis effects and can be used as a potential source material in cosmeceutical industry for the research and development of novel lead molecules with whitening properties.

辣椒素对小鼠B16F10黑色素瘤抑制效应研究

目的观察辣椒素对荷B16F10细胞黑色素瘤实验小鼠的抗瘤效应,初步探讨其抑瘤效应机制。方法采用移植细胞瘤方法建立荷瘤小鼠模型,实验分为生理盐水组(0.9%氯化钠注射液,NS对照组)、阿拉伯胶组(5%阿拉伯胶水溶液)、辣椒素低剂量组〔5mg/(kg·d)〕、中剂量组〔10mg/(kg·d)〕和高剂量组〔20mg/(kg·d)〕5组。观察辣椒素对B16F10瘤细胞的体内抗瘤效应,通过检测细胞因子表达与NK效应细胞活性,分析小鼠免疫功能的变化。结果辣椒素治疗改善了小鼠一般状况,表现为体质量减轻变缓,NS组体质量减轻(2.32±0.61)g,辣椒素低剂量组、中剂量组和高剂量组分别减轻(1.49±0.65)、(0.80±0.33)和(1.58±0.39)g,辣椒素治疗组体质量减轻较小,与NS对照组相比较,差异均有统计学意义,P<0.05;其中中剂量组减轻最为明显,P<0.01。辣椒素显著抑制小鼠B16F10细胞瘤的增殖,表现为NS对照组终末瘤体质量为(4.02±0.56)g,辣椒素低剂量组、中剂量组和高剂量组分别为(3.37±0.25)、(2.21±0.22)和(3.10±0.20)g,辣椒素治疗组瘤体质量均减轻,与NS对照组比较,差异均有统计学意义,P<0.05;其中中剂量组瘤体质量减小最为明显,P<0.01。辣椒素增强了实验小鼠IFN-γ分泌,与NS对照组比较差异均有统计学意义,P<0.05;且中剂量治疗组达到(785.48±15.56)ng/L,变化尤为显著,P<0.01。而辣椒素各治疗组IL-4分泌水平则有所下降,但与NS对照组比较差异无统计学意义,P>0.05。辣椒素明显增强细胞移植瘤小鼠NK细胞活性,在效靶比为50∶1时,NK细胞免疫杀伤活性较为明显,与NS对照组比较,差异有统计学意义,P<0.05;且中剂量组NK细胞杀伤活性达47.36%,增强最为明显,P<0.01。结论辣椒素治疗显示出较强的抗B16F10黑色素细胞瘤效应,其中以辣椒素剂量10mg/(kg·d)的治疗效果最好,其机制可能与增强小鼠细胞免疫功能及NK细胞活性有关。

载芬维A铵脂质体体外对恶性黑素瘤细胞增殖、凋亡和迁移的影响

目的 探讨载芬维A铵脂质体(4-HPR-L)对A375及B16F10黑素瘤细胞增殖、凋亡和迁移的影响.方法 采用薄膜-超声分散法制备载4-HPR-L.体外分别培养A375及B16F10黑素瘤细胞,分为3组,空白对照组仅加入细胞和新鲜培养基,4-HPR组和4-HPR-L组分别加入同浓度4-HPR和4-HPR-L.细胞增殖抑制实验(CCK-8法)检测各组细胞增殖情况;Hoechst33258染色法和流式细胞仪检测细胞凋亡情况;细胞划痕实验检测细胞迁移能力;通过激光共聚焦显微镜观察4-HPR-L入胞情况.采用SPSS22.0软件进行统计分析,多组间数据比较采用单因素方差分析,两组间比较采用t检验.结果 4-HPR与4-HPR-L对A375和B16F10的增殖抑制作用均表现出浓度依赖,0.1、1、15、30、50、70 mg/L 4-HPR和4-HPR-L处理A375和B16F10细胞48 h后,4-HPR组A375细胞存活率分别为(94.3±1.4)%、(91.7±2.5)%、(84.4±2.5%)、(78.8±2.1)%、(59.0±1.1)%、(42.8±2.0)%,4-HPR-L组分另别为(86.0±0.2)%、(76.5±0.6)%、(60.9±1.5)%、(49.0±0.5)%、(32.9±0.2)%、(18.9±0.5)%,同浓度两组间比较,t值分别为8.019、8.298、11.455、19.978、33.672、16.314,均P<0.01;4-HPR组B16F10细胞存活率分别是(95.4±1.9)%、(90.5±2.6)%、(77.0±0.8%)、(64.4±3.5)%、(59.1±2.9)%、(49.9±1.9)%,4-HPR-L组分另别是(88.4±2.0)%、(80.9±3.4)%、(60.9±2.2)%、(51.5±2.9)%、(41.1±1.2)%、(33.5±2.4)%,同浓度两组间比较,均P<0.05.相同浓度下,4-HPR同浓度组A375和B16F10细胞存活率均高于4-HPR-L组.Hoechst33258染色显示,对照组、4-HPR组细胞无明显变化,而4-HPR-L组细胞体积变小,细胞质浓缩,细胞核裂解为碎块,产生凋亡小体.流式细胞仪检测显示,4-HPR-L组A375和B16F10细胞凋亡率均显著高于4-HPR组,均P<0.01.细胞划痕实验显示,4-HPR-L较4-HPR能更好地抑制细胞移行,显著降低划痕的愈合程度.激光共聚焦显微镜观察显示,C6脂质体入胞迅速.结论 4-HPR-L能更好地进入A375细胞、B16F10细胞,且能有效抑制A375、B16F10细胞的增殖和迁移,并诱导其凋亡.

Synthesis of poly(amidoamine)-poly(ethyleneglycol)-poly(amidoamine) and preparation of its hydrogel solution containing doxorubicin

Many pH-and temperature-responsive polymers have been designed for preparing hydrogel.In the present study,in order to decrease the pH sensitivity of reported poly(amidoamine)-poly(ethyleneglycol)-poly(amidoamine)(PAA1580-PEG4600-PAA1580),we designed and synthesized poly(amidoamine)-poly(ethyleneglycol)-poly(amidoamine)(PAA-PEG-PAA)with shorter length of PAA chain by decreasing reaction temperature for preparing PAA-PEG-PAA hydrogel solution containing doxorubicin(DOX).The PAA-PEG-PAA was synthesized via the Michael-addition polymerization.The characteristic of PAA-PEG-PAA was evaluated.The PAA-PEG-PAA hydrogel solution was prepared and investigated.DOX-loaded PAA-PEG-PAA hydrogel solution was prepared,and its in vitro DOX release and in vitro anti-tumor activity were evaluated.Our results indicated that the viscosity of PAA-PEG-PAA hydrogel solution was concentration-and temperature-dependent.The sol-gel transition temperature of PAA-PEG-PAA hydrogel solution(12%,w/w)ranged from 35 to 29℃,and its pH ranged from 6.0 to 7.4.The released DOX from DOX-loaded PAA-PEG-PAA hydrogel showed sustained release characteristics.The in vitro anti-tumor activity of DOX-loaded PAA-PEG-PAA hydrogel was confirmed in B16 F10 cell line.Considering the acidic tumor microenvironment,this DOX-loaded PAA-PEG-PAA hydrogel solution would be easy in situ administration for intra-tumor injection or para-tumor injection forming hydrogel at body temperature.We suggested that this DOX-loaded PAA-PEG-PAAhydrogel solution,if containing photothermal conversion agents,would have a potential further use for photothermal therapy.