Microprecision Delivery of Oligonucleotides in a 3D Tissue Model and Its Characterization Using Optical Imaging
Microprecision Delivery of Oligonucleotides in a 3D Tissue Model and Its Characterization Using Optical Imaging
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DOI：
10.1021/mp300717f
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发表时间：
2013-08-01
影响因子：
4.9
通讯作者：
Nitin, N.
中科院分区：
文献类型：
--
作者：
Luo, Zhen;Ye, Ting;Nitin, N.
尽管寡核苷酸(ONS)在治疗和诊断应用方面具有巨大的潜力，但ONS在皮肤、头颈部和眼睛等组织中的细胞内快速和广泛的递送尚未实现。本研究旨在评价微针(MN)阵列和生化方法在3D组织模型活体细胞内定位递送寡核苷酸方面的协同作用。这种协同组合是基于MNS能够准确地将ONS输送到组织中以实现广泛分布的能力，以及生化试剂(链球菌素O(SLO)和胆固醇与ONS的结合)增强细胞内输送的能力。本研究的结果表明，ON探针被均匀地涂覆在微针阵列上，并在插入组织模体后有效地从微针表面释放。将包裹了ONS和SLO的微针共同插入3D组织模型，导致ONS同时进入细胞质和细胞核。在很短的孵育时间内(35min)，在距离微针插入点500微米的3D组织的外侧和沿深度观察到ONS，其分布类似于输送ON-胆固醇结合物时广泛分布的ONS。随着植入后孵育时间的延长(24小时)，ON在组织中的传递均匀度提高。使用胆固醇结合的ONS，ON探针的传递仅限于组织内细胞的细胞质。最后，转导胆固醇结合的抗GFP抗体导致HeLa细胞中GFP表达降低。综上所述，本研究结果为ONS在组织中的高效细胞内传递提供了一种新的途径。
Despite significant potential of oligonucleotides (ONs) for therapeutic and diagnostic applications, rapid and widespread intracellular delivery of ONs in cells situated in tissues such as skin, head and neck cavity, and eye has not been achieved. This study was aimed at evaluating the synergistic combination of microneedle (MN) arrays and biochemical approaches for localized intratissue delivery of oligonucleotides in living cells in 3D tissue models. This synergistic combination was based on the ability of MNs to precisely deliver ONs into tissues to achieve widespread distribution, and the ability of biochemical agents (streptolysin O (SLO) and cholesterol conjugation to ONs) to enhance intracellular ON delivery. The results of this study demonstrate that ON probes were uniformly coated on microneedle arrays and were efficiently released from the microneedle surface upon insertion in tissue phantoms. Co-insertion of microneedles coated with ONs and SLO into 3D tissue models resulted in delivery of ONs into both the cytoplasm and nucleus of cells. Within a short incubation time (35 min), ONs were observed both laterally and along the depth of a 3D tissue up to a distance of 500 mu m from the microneedle insertion point Similar widespread intratissue distribution of ONs was achieved upon delivery of ON-cholesterol conjugates. Uniformity of ON delivery in tissues improved with longer incubation times (24 h) postinsertion. Using cholesterol-conjugated ONs, delivery of ON probes was limited to the cytoplasm of cells within a tissue. Finally, delivery of cholesterol-conjugated anti-GFP ON resulted in reduction of GFP expression in HeLa cells. In summary, the results of this study provide a novel approach for efficient intracellular delivery of ONs in tissues.