金核/银壳纳米棒在三维表皮模型中的透皮行为和对组织活力的影响
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摘要
目的:应用三维(3D)组织工程表皮模型(HaCaT,无角质层和Epikutis~?,有角质层)评价金核/银壳纳米棒(gold core/silver shell nanorods,Au@Ag NRs)的透皮行为及对模型组织活力的影响。方法:将Au@AgNRs(80μg·mL~(-1))暴露于表皮模型24 h和72 h。通过电感耦合等离子体-质谱法(ICP-MS)测定进入和穿透表皮模型的金、银元素的量,评价Au@AgNRs在HaCaT和Epikutis~?中的透皮行为。通过将不同浓度Au@AgNRs暴露于表皮模型24 h,用甲基噻唑四唑(methylthiazole tetrazolium,MTT)试验和乳酸脱氢酶(lactate dehydrogenase,LDH)释放试验测定Au@AgNRs对表皮模型组织活力的影响。结果:Au@AgNRs暴露24 h后,HaCaT和Epikutis~?模型中均检测到金、银元素(HaCaT中金0.980μg、银3.510μg;Epikutis~?角质层中金0.270μg、银0.750μg,细胞层中金0.540μg、银0.570μg);仅HaCaT模型的下层培养基中检测到极少量银元素;Au@AgNRs暴露72 h后,HaCaT和Epikutis~?模型中金、银元素含量较24 h明显增多(HaCaT中金3.650μg、银5.590μg;Epikutis~?角质层中金0.780μg、银1.770μg,细胞层中金1.650μg、银3.490μg);下层培养基中也均检测到少量金、银元素(HaCaT培养基中金0.034μg、银0.053μg;Epikutis~?培养基中金0.004μg、银0.046μg);MTT实验显示,Au@AgNRs可引起HaCaT及Epikutis~?模型组织活力下降(分别为86.34%～20.02%和95.69%～85.43%),并呈剂量-效应关系;在LDH实验中,随着Au@AgNRs浓度的增加,HaCaT模型的LDH释放量从11.31%升高至116.90%,呈剂量-效应关系;Epikutis~?模型LDH释放量无明显变化。结论:无论角质屏障是否健全,Au@AgNRs均可进入甚至透过表皮模型,其透皮行为呈现时间-效应关系;并引起表皮模型剂量依存性组织活力下降。角质层屏障可减轻和减缓Au@AgNRs在表皮模型中的透过率及对皮肤的损伤。
Objective:To evaluate the transdermal behavior of gold core/silver shell nanorods(Au@AgNRs) and its effect on tissue activity by using tissue engineered three dimensional epidermal model(HaCaT,without cuticle and Epikutis~? with cuticle).Methods:The epidermal models were exposed with 80 μg·mL-1 of Au@AgNRs for 24 h and 72 h,and the gold and silver elements entered and penetrated into the epidermal model were determined by inductively coupled plasma-mass spectrometry(ICP-MS),to valuat of the transdermal behavior of Au@AgNRs in HaCaT and Epikutis~?.The epidermal models were exposed with different concentrations of Au@AgNRs(10,20,40,80,160 μg·mL~(-1)),and MTT(methylthiazole tetrazolium) test and lactate dehydrogenase(LDH) release test were used to assess the effect of Au@AgNRs on the tissue viability at 24 h exposure.Results:After exposure of Au@AgNRs~? for 24 h,gold and silver were detected both in HaCaT and Epikutis~?(in HaCaT,gold 0.980 μg,silver 3.510 μg;in Epikutis~? cuticle,gold 0.270 μg,silver 0.750 μg and in cell layer,gold 0.540 μg,silver 0.570 μg);Onlya few silver were detected in the medium of the HaCaT model.;After 72 h exposure of Au@AgNRs,the gold and silver contents were increase both in HaCaT and Epikutis~? compared to 24 h exposure(in HaCaT,gold 3.650 μg,silver 5.590 μg;in Epikutis~? cuticle,gold 0.780 μg,silver 1.770 μg and in cell layer,gold 1.650 μg,silver 3.490 μg).Afew silverwere detected in the medium of both HaCaT and Epikutis~?(in HaCaT medium,gold 0.034 μg,silver 0.053 μg;in Epikutis~? medium,gold 0.004 μg,silver 0.046 μg).The MTT results suggest that Au@AgNRs can cause a decrease in tissue viability of HaCaT and Epikutis~?(86.34%-20.02% and 95.69%-85.43 %,respectively) with a dose-dependent manner.In the LDH test,with the increase of Au@AgNRs concentration,the LDH release in HaCaT model increased from 11.31% to 116.90% with a dose-dependent manner;but no significant change in Epikutis~?.Conclusion:Regardless whether the keratin barrier is exist or intact(with or without cuticle),the Au@AgNRs can enter and even penetrate through the epidermal models,and the transdermal behavior presents a time-dependent manner.And Au@AgNRs can cause decrease in the tissue viability of the epidermal models with dose-dependent manner.The cuticle barrier can reduce and slow down the transmissivity and skin damage of Au@AgNRs on the epidermis models.
Objective:To evaluate the transdermal behavior of gold core/silver shell nanorods(Au@AgNRs) and its effect on tissue activity by using tissue engineered three dimensional epidermal model(HaCaT,without cuticle and Epikutis~? with cuticle).Methods:The epidermal models were exposed with 80 μg·mL-1 of Au@AgNRs for 24 h and 72 h,and the gold and silver elements entered and penetrated into the epidermal model were determined by inductively coupled plasma-mass spectrometry(ICP-MS),to valuat of the transdermal behavior of Au@AgNRs in HaCaT and Epikutis~?.The epidermal models were exposed with different concentrations of Au@AgNRs(10,20,40,80,160 μg·mL~(-1)),and MTT(methylthiazole tetrazolium) test and lactate dehydrogenase(LDH) release test were used to assess the effect of Au@AgNRs on the tissue viability at 24 h exposure.Results:After exposure of Au@AgNRs~? for 24 h,gold and silver were detected both in HaCaT and Epikutis~?(in HaCaT,gold 0.980 μg,silver 3.510 μg;in Epikutis~? cuticle,gold 0.270 μg,silver 0.750 μg and in cell layer,gold 0.540 μg,silver 0.570 μg);Onlya few silver were detected in the medium of the HaCaT model.;After 72 h exposure of Au@AgNRs,the gold and silver contents were increase both in HaCaT and Epikutis~? compared to 24 h exposure(in HaCaT,gold 3.650 μg,silver 5.590 μg;in Epikutis~? cuticle,gold 0.780 μg,silver 1.770 μg and in cell layer,gold 1.650 μg,silver 3.490 μg).Afew silverwere detected in the medium of both HaCaT and Epikutis~?(in HaCaT medium,gold 0.034 μg,silver 0.053 μg;in Epikutis~? medium,gold 0.004 μg,silver 0.046 μg).The MTT results suggest that Au@AgNRs can cause a decrease in tissue viability of HaCaT and Epikutis~?(86.34%-20.02% and 95.69%-85.43 %,respectively) with a dose-dependent manner.In the LDH test,with the increase of Au@AgNRs concentration,the LDH release in HaCaT model increased from 11.31% to 116.90% with a dose-dependent manner;but no significant change in Epikutis~?.Conclusion:Regardless whether the keratin barrier is exist or intact(with or without cuticle),the Au@AgNRs can enter and even penetrate through the epidermal models,and the transdermal behavior presents a time-dependent manner.And Au@AgNRs can cause decrease in the tissue viability of the epidermal models with dose-dependent manner.The cuticle barrier can reduce and slow down the transmissivity and skin damage of Au@AgNRs on the epidermis models.
引文
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[14]卢永波,张荣芳,许珊珊,等.一种用于体外测试的中国汉族人表皮替代模型[A]//中国毒理学会第六届全国毒理学大会论文摘要,2013:1LU YB,ZHANG RF,XU SS,et al.An epidermal replacement model of Chinese Han people used for in vitro testing[A]//The Sixth National Toxicology Conference of the Chinese Society of Toxicology,2013:1
[2]THOMAS R,JANARDHANAN A,VARGHENSE RT,et al.Antibacterial properties of silver nanoparticles synthesized by marine Ochrobactrum sp.[J].Braz J Microbiol,2014,45(4):1221
[3]CHERNOUSIVA S,EPPLE M.Silver as antibacterial agent:ion,nanoparticle,and metal[J].Angew Chem Int Ed,2013,52(6):1636
[4]ZANETTE C,PELIN M,CROSERA M,et al.Silver nanoparticles exert a long-lastingantiproliferative effect on human keratinocyte HaCaT cell line[J].Toxicol Vitro,2011,25(5):1053
[5]SZMYD R,GORALVZYK AG,SKALNIAK L,et al.Effect of silver nanoparticles on human primary keratinocytes[J].Biol Chem,2013,394(1):113
[6]BOONKAEW B,KEMPF M,KIMBLE R,et al.Cytotoxicity testing of silver-containing burn treatments using primary and immortal skin cells[J].Burns,2014,40(8):1562
[7]KORANI M,REZAYAT SM,ARBABI BS.Sub-chronic dermal toxicity of silver Nanoparticles in Guinea pig:special emphasis to heart,bone and kidney toxicities[J].Iran J Pharm Res,2013,12(3):511
[8]程祥,赵玉云,邵安良,等.含银敷料的表征和银的体外释放实验方法研究及其应用[J].药物分析杂志,2015,35(3):491CHENG X,ZHAO YY,SHAO AL,et al.Study on the characterization of silver-containing dressings and experimental methods of in vitro release of silver and its application[J].Chin JPharm Anal,2015,35(3):491
[9]BIANCO C,KEZIC S,CROSERA M,et al.In vitro percutaneous penetration andcharacterization of silver from silver-containing textiles[J].Int J Nanomedicine,2015,10(10):1899
[10]TAURAN Y,BRIOUDE A,COLEMAN AW,et al.Molecular recognition by gold,silver and copper nanoparticles[J].World JBiol Chem,2013,4(3):35
[11]MENG J,LI YL,LIU J,et al.Using gold nanorods core/silver shell nanostructures as model material to probe biodistribution and toxic effects of silvernanoparticles inmice[J].Nanotoxicology,2014,8(6):686
[12]CHEN XL,WU XC,XU HY,et al.Revealing silver cytotoxicity using Au nanorods/Ag shellnanostructures:disrupting cell membrane and causing apoptosis through oxidative damage[J].RSC Adv,2013,3(3):2296
[13]WU MY,CHEN L,LI RU,et al.Bio-distribution and bioavailability of silver and gold in rat tissues with silver/gold nanorodadministration[J].RSC Adv,2018,8:12260
[14]卢永波,张荣芳,许珊珊,等.一种用于体外测试的中国汉族人表皮替代模型[A]//中国毒理学会第六届全国毒理学大会论文摘要,2013:1LU YB,ZHANG RF,XU SS,et al.An epidermal replacement model of Chinese Han people used for in vitro testing[A]//The Sixth National Toxicology Conference of the Chinese Society of Toxicology,2013:1