通过5-ALA(δ-氨基-γ-酮戊酸)诱导的原卟啉Ⅸ对C6细胞进行的双光子光敏疗法
J Photochem Photobiol B. 2007 Apr 4; [Epub ahead of print]Two-photon photodynamic therapy of C6 cells by means of 5-aminolevulinic acid induced protoporphyrin IX.
通过5-ALA(δ-氨基-γ-酮戊酸)诱导的原卟啉Ⅸ对C6细胞进行的双光子光敏疗法
Beck TJ, Burkanas M, Bagdonas S, Krivickiene Z, Beyer W, Sroka R, Baumgartner R, Rotomskis R.
Laser Research Laboratory, Ludwig-Maximilians-University, Munich, Germany.
Photodynamic therapy (PDT) has received increased attention as a treatment modality for malignant tumors as well as non-oncologic diseases such as age-related macular degeneration (AMD). An alternative to excite the photosensitizer by the common one-photon absorption is the method of two-photon excitation (TPE). This two-photon photodynamic therapy has the potential of improving the therapeutic outcome due to a highly localized photodynamic effect. The present study investigated the two-photon excited PDT performing in vitro experiments where C6 rat glioma cells were irradiated with a pulsed and focused fs Ti:sapphire laser emitting light at 800nm. The irradiance distribution of the laser beam was carefully analyzed before the experiment and the applied irradiance was known for each position within the irradiated cell layer. Cells were divided into four groups and one group was incubated with 5-ALA and irradiated 4-5h later. The survival of this group was tested after irradiation by means of ethidium bromide and acridine orange staining and compared to a control group, which was irradiated under the same conditions, but not incubated with 5-ALA before. Both groups showed necrotic areas depending on the applied irradiance, the value of which at the margin of the necrotic area could be deduced from its size. 5-ALA incubated cells became necrotic after irradiation with a mean irradiance above 6.1x10(10)W/cm(2), while non-incubated cells remained viable. Cells of both groups became necrotic when treated with an irradiance above 10.9x10(10)W/cm(2). The observed affected area of the cell layers was between 0.13mm(2) and 1.10mm(2). Since the irradiation of non-incubated cells below the mean power density of 10.9x10(10)W/cm(2) induced no necrosis, apparently no thermal damage was induced in the cells and necrosis of the 5-ALA incubated cells can be ascribed to the photodynamic effect induced by two-photon excitation. The successful photodynamic treatment of a large area of a monolayer cell culture induced by two-photon excitation offers new perspectives for photodynamic treatment modalities.
光敏疗法不仅在与年龄相关的黄斑退行性变等非肿瘤疾病中，在恶性肿瘤治疗中也作为一种治疗方法得到越来越多的关注。对光敏剂的激发也已经从普通的单光子激发发展到双光子激发。这种双光子光敏疗法由于可以产生高度集中而局限的光动力效应，因此被认为在改善疗效上有很大的潜力。本实验用双光子疗法进行C6大鼠胶质瘤的体外实验，用800NM波长发射光照射胶质瘤细胞。辐射度的分配在实验前严格进行计算分配，不同部位细胞的辐射度也在实验前确定。细胞被分为四组，其中一组和5-ALA一起进行培养，4-5小时后进行照射。照射后对本组的存活细胞行溴乙锭和吖啶橙染色并与照射条件相同，但未加5-ALA进行培养的对照组比较观察。两组在照射部位都可以见到坏死区，在坏死区边缘的辐射值可以从坏死区的大小推出。与5-ALA共培养的细胞在平均功率密度6.1×10(10)W/CM2以上就出现坏死，而对照组细胞仍然存活。当功率密度达到10.9×10(10)W/CM2以上，两组细胞均出现坏死。被观察到的细胞层受累区在0.13mm2 和 1.10mm2之间。由于对照组在10.9×10(10)W/CM(2)的功率密度以下，不产生坏死，因此对正常细胞不会产生热损伤；而5-ALA共培养的细胞的坏死可以被归于双光子激发产生的光动力效应。对单层细胞培养区域通过双光子激发进行光疗，为光疗这种治疗方法展现出了新的前景。
PMID: 17513121 [PubMed - as supplied by publisher]
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