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

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（80 μg·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0.980 μg、银3.510μg；Epikutis^®角质层中金0.270 μg、银0.750 μg，细胞层中金0.540 μg、银0.570 μg）；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3.650 μg、银5.590 μg；Epikutis^®角质层中金0.780 μg、银1.770 μg，细胞层中金1.650 μg、银3.490 μg）；下层培养基中也均检测到少量金、银元素（HaCaT培养基中金0.034 μg、银0.053μg；Epikutis^®培养基中金0.004 μg、银0.046 μg）；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 dosedependent 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.

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