Photodynamic therapy on keloid fibroblasts in tissue-engineered keratinocyte-fibroblast co-culture
Photodynamic therapy on keloid fibroblasts in tissue-engineered keratinocyte-fibroblast co-culture
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DOI：
10.1002/lsm.20213
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发表时间：
2005-09-01
影响因子：
2.4
通讯作者：
Wong, BJF
中科院分区：
文献类型：
--
作者：
Chiu, LL;Sun, CH;Wong, BJF
背景和目的：疤痕疙瘩是一种毁容性增生性疤痕，是皮肤损伤的病理反应。使用器官型组织培养系统（Raft 模型 [1-10]）来研究使用光动力疗法（PDT）作为手术切除后疤痕疙瘩的辅助疗法的可行性。 Raft 共培养系统通过在嵌入胶原基质的成纤维细胞顶部分层角质形成细胞来模拟皮肤。 PDT 使用在光照射下原位产生单线态氧的药物，并可能对活组织产生从生长调节到细胞凋亡等多种影响。 PDT 已用于治疗皮肤、视网膜和食道等器官的多种良性和恶性疾病。 研究设计/材料和方法：从接受选择性手术的患者皮肤中分离出正常成人、新生儿和瘢痕疙瘩成纤维细胞和角质形成细胞，并用于构建 Raft。将成熟的筏（4天后）与光敏剂5-氨基乙酰丙酸（5-AlA）一起孵育3小时，并进行激光照射（635 nm）以获得5 J/cm（2）、10 J/cm（2）或20 J/cm（2）的总能量传递。 24小时和14天后对筏进行检查。使用共焦成像结合活死荧光染料测定细胞活力。多光子显微镜 (MPM) 对胶原结构和密度进行成像。随着筏随时间的推移而收缩，每天使用光学测微法测量表面积。结果：在 10 和 20 J/cm(2) 下，所有构建体中都观察到细胞几乎全部死亡，而在 5 J/cm(2) 下，细胞活力与对照组相当。瘢痕疙瘩和新生儿筏中的细胞活力高于正常成人筏中观察到的细胞活力。与对照相比，经过处理的筏在 14 天期间收缩较少。疤痕疙瘩和新生儿筏的收缩和胶原密度最大。结论：建立了 PDT 剂量范围，可减少组织收缩和胶原密度，同时最大限度地减少对成纤维细胞的损伤。
Background and Objectives: Keloids are disfiguring, proliferative scars that are a pathologic response to cutaneous injury. An organotypic tissue culture system (the Raft model [1-10]) was used to investigate the feasibility of using photodynamic therapy (PDT) as an adjunctive therapy to treat keloids following surgical excision. The Raft co-culture system mimics skin by layering keratinocytes on top of fibroblasts embedded in a collagen matrix. PDT uses drugs that produce singlet oxygen in situ when irradiated by light, and may lead to a number of effects in living tissues varying from the modulation of growth to apoptosis. PDT is already used to treat several benign and malignant diseases in organs such as the skin, retina, and esophagus.Study Design/Materials and Methods: Normal adult, neonatal, and keloid fibroblasts and keratinocytes were isolated from skin obtained from patients undergoing elective procedures and used to construct the Rafts. Mature Rafts (after 4 days) were incubated with 5-amino levulinic acid (5-AlA), a photosensitizer, for 3 hours and were laser-irradiated (635 nm) for total energy delivery of 5 J/cm(2), 10 J/cm(2), or 20 J/cm(2). Rafts were examined 24 hours and 14 days later. Cell viability was determined using confocal imaging combined with live-dead fluorescent dyes. Multiphoton microscope (MPM) imaged collagen structure and density. As Rafts contract over time, surface area was measured using optical micrometry daily.Results: At 10 and 20 J/cm(2), near-total cell death was observed in all constructs, while at 5 J/cm(2) cell viability was comparable to controls. Cell viability in keloid and neonatal Rafts was greater than that observed in normal adult Rafts. Treated Rafts contracted less over the 14-day period compared to controls. Contraction and collagen density were greatest in keloid and neonatal Rafts.Conclusions: A PDT dosimetry range was established, which reduces tissue contraction and collagen density while minimizing injury to fibroblasts.