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慢病毒介导的果蝇脱氧核苷激酶联合(E)-5-(2-溴乙烯基)-2'-脱氧尿苷或 1-β-D-阿拉伯呋喃糖胸腺嘧啶治疗人瘢痕成纤维细胞的疗效。

Efficacy of lentivirus‑mediated Drosophila melanogaster deoxyribonucleoside kinase combined with (E)‑5‑(2‑bromovinyl)‑2'‑deoxyuridine or 1‑β‑D‑arabinofuranosylthymine therapy in human keloid fibroblasts.

机构信息

Department of Plastic Surgery, Aoyang Hospital, Zhangjiagang, Jiangsu 215600, P.R. China.

Department of Breast Surgery, First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China.

出版信息

Mol Med Rep. 2018 Aug;18(2):1660-1665. doi: 10.3892/mmr.2018.9101. Epub 2018 May 30.

DOI:10.3892/mmr.2018.9101
PMID:29901093
Abstract

Keloid scarring is a type of fibroproliferative disease with a high recurrence rate. However, no effective treatment is currently available. Combined therapy with recombinant lentivirus‑mediated Drosophila melanogaster deoxyribonucleoside kinase (Dm‑dNK) and prodrug has been widely studied and used for cancer treatment. Due to the similarities between keloid scars and tumors, the aim of the present study was to investigate the efficacy of a Dm‑dNK/nucleoside analog system for the treatment of keloid scars. Recombinant lentivirus expression of the Dm‑dNK suicide gene was assessed. Western blotting was used to examine the protein expression of lentivirus mediated Dm‑dNK in keloid fibroblasts. Enzyme activity assays were conducted using [3H]‑labeled substrates. Furthermore, cytotoxicity and bystander effects were evaluated using MTT assays. The expression of green fluorescent protein was observed using fluorescence microscope and results indicated that there was no notable difference in lentivirus infectivity between the multiplicity of infection (MOI) of 1 and 10 in cells. Notably, western blotting revealed that Dm‑dNK was stably expressed in keloid fibroblasts and the enzymatic activity assays revealed that the enzyme was activated following introduction into the keloid fibroblasts via the lentivirus. The cytotoxicity and bystander effects of Dm‑dNK combined with cytotoxic nucleoside analogs were both observed in Dm‑dNK+ keloid fibroblasts. These results demonstrated that the lentivirus‑mediated Dm‑dNK therapy may be effective in treating keloid fibroblasts, which provides some evidence for the use of Dm‑dNK/prodrug therapy for keloid treatment in vivo in the future.

摘要

瘢痕疙瘩是一种纤维增生性疾病,具有很高的复发率。然而,目前尚无有效的治疗方法。重组慢病毒介导的果蝇脱氧核苷激酶(Dm-dNK)与前药联合治疗已广泛研究和用于癌症治疗。由于瘢痕疙瘩和肿瘤之间存在相似性,本研究旨在探讨 Dm-dNK/核苷类似物系统治疗瘢痕疙瘩的疗效。评估了重组慢病毒表达的 Dm-dNK 自杀基因。使用 Western blot 检测慢病毒介导的 Dm-dNK 在瘢痕疙瘩成纤维细胞中的蛋白表达。使用 [3H]标记的底物进行酶活性测定。此外,通过 MTT 测定评估细胞毒性和旁观者效应。使用荧光显微镜观察绿色荧光蛋白的表达,结果表明,在细胞中,慢病毒感染率的感染复数(MOI)为 1 和 10 之间,慢病毒的感染率没有明显差异。值得注意的是,Western blot 显示 Dm-dNK 在瘢痕疙瘩成纤维细胞中稳定表达,酶活性测定显示,通过慢病毒导入瘢痕疙瘩成纤维细胞后,酶被激活。在 Dm-dNK+瘢痕疙瘩成纤维细胞中观察到 Dm-dNK 与细胞毒性核苷类似物联合的细胞毒性和旁观者效应。这些结果表明,慢病毒介导的 Dm-dNK 治疗可能对治疗瘢痕疙瘩成纤维细胞有效,为将来在体内使用 Dm-dNK/前药治疗瘢痕疙瘩提供了一定的证据。

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