兔眼脉络膜黑色素瘤的第二代光敏剂光动力治疗

目的 观察第二代光敏剂CASPc（chloroaluminum su lfonated phthalocyanine,CASPc）对B16F10黑色素瘤动物模型的光动力治疗（PDT）作用 。 方法 38只新英格兰大白兔经免疫抑制后种植B16F10黑色素瘤碎片于脉络膜下腔，B型超声、间接检眼镜随访肿瘤生长至2.0 ~3.8 mm厚时30只兔眼给予PDT治疗。耳静脉注射CASPc 5 mg/kg, 24 h后给予氩-染料激光照射，激光波长675 nm, 输出功率600 nm/cm2，激光照射剂量20~70 J/cm2；8只对照组兔眼只给予激光或光敏物质。治疗后观察6~8周。 结果 38只兔眼建模成功。30只PDT治疗兔眼中21只眼肿瘤得到控制，9只眼未能控制；对照组兔眼肿瘤治疗后2周充满整个玻璃体。 结论 第二代光敏剂CASPc对实验性眼B16F10黑色素瘤具有一定的PDT治疗作用。 （中华眼底病杂志，2004，20：67-132）     

兔脉络膜黑色素瘤光动力学治疗的研究

目的 评价第二代光敏物质（benzoporphyrin derivative,BPD)在兔脉络膜色素性黑色素瘤光动力学治疗中的疗效。方法 对44只新英格兰大白兔眼,建立脉络膜色素性黑色素瘤模型,每日注射环孢素A行免疫抑制治疗。当黑色素瘤生长厚度达2.0-4.6mm时,将44只兔分成3组,治疗I组（14只兔）及治疗Ⅱ（20只兔）,静脉给予BPI1mg/kg,通过氩离子－染料激光进行光动力学治疗,长为692nm,照射剂量为60－150J /cm^2,对照组（10只兔）中的6只兔仅给予激光照射,4只兔为空白对照。治疗后4－6周外死兔,取其眼球观察脉络膜黑色素瘤光动力学治疗效果。结果 34只脉络膜黑色素瘤兔经光动力学治疗后,治疗I组肿瘤厚度为＜3mm,激光照射剂量60－80J/cm^2,与对照组比较,差异有非常显著意义（P＜0．001）;治疗Ⅱ组肿瘤厚度为3.0-4.6mm,激光照射剂量为＞80J/cm^2,与对照组比较,差异有非常显著意义（P＜0．001）。对照组兔眼肿瘤在2－3周时充满整个玻璃体腔。结论 光动力学治疗可能成为一种选择性治疗脉络膜色素性黑色素瘤的方法。     

血啉甲醚在兔脉络膜黑色素瘤组织中的分布

目的应用血管造影和组织冰冻切片荧光显微镜观察的方法,研究国产新型光敏剂血啉甲醚(hematoporphyrin monomethyl ether,HMME)在兔脉络膜黑色素瘤眼部组织的分布。方法建立改良的兔脉络膜黑色素瘤动物模型。肿瘤生长3~4mm后进行药物分布实验。28只兔眼(肿瘤20眼,正常眼底4眼,虹膜4眼)静脉注射HMME(10mg·kg-1)后96h内血管造影;于兔静脉注射HMME(10mg·kg-1)后的5min、15min、30min、60min、180min、480min和24h、48h、72h各取兔眼球2眼制作组织冰冻切片,荧光显微镜下观察药物在肿瘤及眼部各组织的分布,并进行半定量分析。结果于静脉注射HMME后3~4h在肿瘤组织选择性聚积达到最高。结论HMME可以在兔脉络膜黑色素瘤中选择性聚集、滞留,静脉注射后3~4h是光动力治疗的适宜时间。     

应用血啉单醚光动力疗法治疗角膜新生血管的研究

目的观察应用血啉单醚行光动力疗法(PDT)治疗角膜新生血管的效果。设计前瞻性随机对照实验。研究对象成年有色兔18只。方法制作碱烧伤角膜新生血管模型。血啉单醚5mg/kg静脉注射,不同能量密度(7.6￣152.8J/cm2)的氩绿激光照射角膜新生血管根部,不注射血啉单醚只单纯行同等能量密度的激光照射组作为对照组。主要指标PDT的能量密度,行角膜新生血管荧光素造影观察新生血管封闭情况。结果PDT后3天,角膜新生血管荧光素血管造影显示:应用61.2J/cm2及以上的能量,有67%以上的眼角膜新生血管被完全封闭,33%的眼部分有效。PDT后1周,61.2J/cm2及以上组仍有66.7%(16/24眼)的眼角膜新生血管完全封闭。激光后1个月5/24眼无新生血管出现,其余眼再次出现新生血管。结论采用激光能量密度在(61.2￣152.8)J/cm2照射的PDT治疗能完全或部分封闭兔碱烧伤角膜新生血管模型中的角膜新生血管,但有新生血管再生。(眼科,2006,15:180-183)     

应用血啉甲醚对体外培养鼠皮肤黑色素瘤细胞PDT治疗的实验研究

目的探讨应用国产光敏剂血啉甲醚（hematoporphyrin monomethyl ether，HMME）对鼠皮肤黑色素瘤细胞株（B16FIO细胞株）行光动力疗法（photodynamic therapy，PDT）的效应和机制。设计实验性研究。研究对象B16FIO细胞株。方法采用HMME作光敏剂，对B16FlO细胞用630nm波长的氦氖激光机作光源（功率密度5mW／cm^2），以2．5、5．0、10,0、20、0、40．0、80．0uml的HMME经2.4、3．6、4、8、6．0、7．2J/cm^2的激光能量照射后，MTT0法测定PDT对B16F10的杀伤率，Annexin—V／PI双染流式细胞仪测定B16F10的凋亡率。主要指标吸光度（OD值）、凋亡率。结果随光敏剂浓度升高和照光剂量增加，光动力作用对细胞的相对杀伤率逐渐增大，在低光动力剂量下明显上升，随后上升渐趋缓慢，杀伤曲线呈“S”形。在对细胞杀伤率约70％的PDT剂量时，细胞死亡机制以凋亡为主，在对细胞杀伤率约99％的过量PDT剂量时，细胞可能损伤过重发生坏死。结论应用HMME行PDT治疗对体外培养鼠皮肤黑色素瘤细胞具有明确的杀伤效应，其对细胞的杀伤率具有显著的剂量．效应关系；在一定PDT剂量范围内细胞死亡机制主要以凋亡为主。     

光动力学疗法治疗角膜新生血管后角膜的光电镜改变

目的观察应用国产光敏剂行光动力学疗法(PDT)治疗角膜新生血管后角膜的组织学改变。方法碱烧伤有色兔角膜制作角膜新生血管模型。血啉单醚5mg/kg自耳缘静脉注射,不同的能量密度61.2-52.8J/cm2的氲绿激光照射角膜新生血管根部,不注射血啉单醚并单纯行同等能量密度的激光照射组作为对照组,PDT治疗后3h、1周、1个月行角膜光电镜检查。结果PDT后3h可见角膜急性炎症反应,有大量中性粒细胞浸润和少量嗜酸性白细胞浸润,虹膜组织无损伤。PDT后1周角膜炎症反应大部分消失,可见新生血管腔内有无定形物质填塞和许多影子血管。透射电镜显示:PDT组角膜新生血管内皮细胞内线粒体显示出空泡样变,细胞形态不完整。结论血啉单醚作为光敏剂,应用氩绿激光对角膜新生血管行PDT治疗,导致新生血管内皮细胞损伤,能有效封闭角膜新生血管,对周围组织无明显损伤。     

兔葡萄膜黑色素瘤模型建立方法的实验研究

目的建立兔眼脉络膜黑色素瘤动物模型,观察肿瘤的生长特性。设计实验研究。研究对象新西兰大白兔20只（20跟）。方法将培养的B16F10鼠皮肤黑色素瘤细胞悬液种植到C57BL／6黑鼠大腿内侧皮下,7～12天孵育成直径10mm团块后制成肿瘤碎片,种植到新西兰大白兔右眼脉络膜上腔,自种植之日起予环孢霉素A20mg／kg肌肉注射以抑制淋巴细胞免疫功能。每周行间接检眼镜、眼底照相、B超、彩色超声多普勒观察并记录兔眼底肿瘤大小、形状、眼部其他组织的改变以及肿瘤内部血液供应情况。观察6周后眼球摘除,行HE染色组织病理学检查。主要指标肿瘤最大基底径、高度、细胞类型、肿瘤内血管分布情况以及眼部其他组织肿瘤浸润情况。结果除1只（5％）兔因呕吐物窒息死亡外,余19只兔19眼（95％）肿瘤存活并迅速生长。其中1眼（5％）自种植4周开始肿瘤自行萎缩,第6周12眼（60％）肿瘤充满玻璃体腔。2周、3周、4周B超测量肿瘤平均最大基底径分别为3．1mm、6．1mm、12．2mm,平均高度分别为2．2mm、2．7mm、8．5mm。HE染色示肿瘤以梭形细胞为主,排列规则紧密,部分坏死,血管充盈。18眼（90％）均可见眼部其他组织受肿瘤细胞浸润,3跟（15％）肿瘤突破巩膜向球外蔓延。结论B16FIO鼠皮肤黑色素瘤细胞株兔眼脉络膜上腔种植后肿瘤迅速生长,模型建造成功率高。f跟科,2010,19：130—134）     

光动力治疗兔角膜新生血管的研究

目的：观察光动力学治疗(photodynamic therapy,PDT)缝线法诱导的兔角膜新生血管(cornaeal neovascularization,CNV)的效果和安全性。方法：取新西兰白兔5只10眼,用5．0号的丝线在角膜上缝线诱导角膜新生血管的生长,注射光敏剂Photofrin 48h后行DIOMED630PDT激光照射。裂隙灯显微镜观察角膜NV生长情况,并且计算其面积、生长长度,照射后28d处死实验兔,取其眼球做组织病理学检查。结果：角膜缝线后3d开始有新生血管长入,3wk后长到缝线的尖端。激光照射后2d开始角膜新生血管变得模糊不清,2wk后角膜新生血管近完全消失。组织病理学检查见角膜,视网膜以及虹膜、睫状体、脉络膜等组织正常。结论：光动力治疗角膜新生血管有效、安全。     

应用血啉甲醚的光动力学疗法治疗实验性虹膜新生血管

目的观察以国产血啉甲醚(HMME)为光敏剂,应用光动力学疗法(PDT)封闭实验性虹膜新生血管(INV)的疗效和并发症。方法应用氩离子激光封闭猴眼全部视网膜分枝静脉的方法建立INV模型。静脉注射HMME(5 mg/kg),用514 nm的氩绿激光照射,设定能量密度分别为107.46、143.28、275.22 J/cm23组,行虹膜彩像、虹膜荧光造影(IFA)、虹膜吲哚青绿造影(IICGA)及光镜和电镜观察疗效。结果PDT治疗后48 h,治疗区INV基本闭塞;ICGA显示基质层血管有不同程度的闭塞。PDT后3周,治疗区域新生血管仍闭塞,虹膜基质血管重新开放;非治疗区新生血管退行。光镜显示中能量组虹膜基质轻度萎缩,高能量组虹膜可见大量色素增殖。治疗组均未发生眼压升高。结论以HMME为光敏剂的PDT在一定时期内可有效封闭实验性虹膜新生血管,减少新生血管性青光眼的发生,但对虹膜基质有一定损伤。     

HMME-PDT药物剂量和光剂量对正常视网膜和脉络膜结构的影响

目的观察血卟啉单甲醚-光动力疗法（HMME-PDT）过程中不同药物剂量和激光剂量对照光区域视网膜和脉络膜组织结构的影响。方法以新西兰兔为实验动物，药物剂量分别为2．5、5和10mg／kg体重耳缘静脉注射，在注射药物结束后5min内分别使用光斑能量密度5、20、40和60J／cm^2进行照射，并在第1d、7d行眼底观察、荧光素眼底血管造影（FAA）和组织病理学方法检测照射部位的结构变化。结果在3种药物剂量、4种能量密度情况下，在PDT后第1d都有脉络膜毛细血管闭塞现象发生。随着药物剂量和激光剂量的增加，发生脉络膜毛细血管闭塞的光斑增多，维持到第7d。结论提高药物剂量或能量密度能够促进并保持脉络膜毛细血管闭塞，但同时使非选择性损伤加重。药物剂量和激光剂量的良好匹配能获得良好治疗效果，而同时对视网膜损伤最小。     

Evaluation of the new photosensitizer Stakel (WST-11) for photodynamic choroidal vessel occlusion in rabbit and rat eyes

PURPOSE: To evaluate the photodynamic potential of a new hydrosoluble photosensitizer (WST-11, Stakel; Steba Biotech, Toussus-Le-Noble, France), for use in occlusion of normal choroidal vessels in the rabbit eye and CNV (choroidal neovascularization) in the rat eye. METHODS: Occlusive and nonocclusive parameters of Stakel and verteporfin photodynamic therapy (PDT) were investigated in pigmented rabbits. Eyes were followed by fluorescein angiography (FA) and histology at various intervals after PDT. RESULTS: When occlusive parameters (fluence of 50 J/cm(2), 5 mg/kg drug dose and DLI [distance to light illumination] of 1 minute) were used, Stakel PDT was efficient immediately after treatment without associated structural damage of the RPE and retina overlying the treated choroid in the rabbit eye. Two days later, total occlusion of the choriocapillaries was seen in 100% of the treated eyes, along with accompanying histologic structural changes in the overlying retina. When the occlusive parameters (fluence, 100 J/cm2; drug dose, 12 mg/m2; and DLI, 5 minutes) of verteporfin PDT were used, occlusion of the choriocapillaries was observed in 89% of the treated eyes. Histology performed immediately after treatment demonstrated structural damage of the overlying retina and RPE layer. Weaker, nonocclusive Stakel PDT parameters (25 J/cm2, 5 mg/kg, and DLI of 10 minutes) did not induce choriocapillary occlusion or retinal lesions on FA or histology. Weaker, nonocclusive verteporfin PDT parameters (10 J/cm2, 0.2 mg/kg, and DLI of 5 minutes) did not induce choriocapillary occlusion. However, histology of these eyes showed the presence of damage in the retinal and choroidal tissues. Moreover, preliminary results indicate that selective CNV occlusion can be achieved with Stakel PDT in the rat eye. CONCLUSIONS: Unlike verteporfin PDT, Stakel PDT does not cause direct damage to the RPE cell layer or retina. These observations indicate that Stakel PDT may have a high potential for beneficial therapeutic outcomes in treatment of AMD.     

Experimental photodynamic effects of hypocrellin A on the choriocapillaris

OBJECTIVE: To investigate the use of the photosensitizer hypocrellin A as a photodynamic agent to occlude the choriocapillaris in the eyes of pigmented rabbits. METHODS: Hypocrellin A (500 microg/kg) incorporated into liposomes was injected intravenously in pigmented rabbits. Ten to 15 minutes later, the retinas were irradiated with a dye laser (spot size, 2 mm; 10-60 mW; 10-60 seconds; fluences 3.2 to 95.5 J/cm2) at two wavelengths: yellow (568 nm) and red (647 nm). Fluorescein angiography was performed 2 and 24 hours later. Parallel controls were irradiated without a photosensitizer. Histology was performed immediately after the 24-hour angiogram. observed 24 hours after PDT in eyes subjected to yellow laser irradiation at 20 mW for 60 seconds (fluence, 38.2 J/cm2) and at higher powers for lesser durations: 30 mW for 40 seconds (fluence, 38.2 J/cm2), 40 mW for 20 seconds (fluence, 25.4 J/cm2), and 50 and 60 mW for 10 seconds (fluences, 15.9 and 19.1 J/cm2, respectively). Light microscopy showed occlusion of the choriocapillaris in all eyes with angiographic responses. Control eyes and eyes irradiated with the red laser revealed no angiographic or histologic changes. CONCLUSION: Photodynamic therapy using liposomal hypocrellin A and the yellow laser of 568 nm was effective in occluding the choriocapillaris of pigmented rabbits.     

Threshold power levels for NPe6 photodynamic therapy

OBJECTIVE: To determine the threshold power levels for producing retinal and choroidal vascular occlusion using mono-L-aspartyl chlorin e6 (NPe6) photodynamic therapy; to evaluate its efficacy with longer intervals between photosensitizer injection and laser application; to determine the elapsed time between light application and appearance of angiographic changes. METHODS: Pigmented and nonpigmented rabbits were injected intravenously with 2 mg/kg of NPe6 before laser irradiation of the retina-choroid. Group 1 was treated at increasing power levels; fluorescein angiograms were obtained at each fluence. Group 2 animals were exposed to laser irradiation at 5 minutes, and 1 and 3 hours postinjection to determine (by fluorescein angiography 24 hours post-treatment) if increasing the interval affected outcome. Group 3 animals underwent fluorescein angiography at 30 minutes, 1 hour, 2 hours, and 24 hours posttreatment to document the time between laser application and subsequent vessel closure. RESULTS: Choroidal vessel occlusion was angiographically evident in all lesions at fluences of > or = 2.65 J/cm2 in pigmented rabbits and at > or = 0.88 J/cm2 in nonpigmented rabbits. Lesion diameter decreased as the time between injection and treatment increased. Vessel occlusion was documented at least 2 hours after treatment. CONCLUSION: Choroidal vessel occlusion can occur at very low fluence.     

Choriocapillaris photodynamic therapy using indocyanine green

PURPOSE: To evaluate the potential of photodynamic therapy using indocyanine green for occlusion of choroidal neovascularization, the authors studied efficiency and collateral damage of photodynamic therapy-induced photothrombosis in the rabbit choriocapillary layer. METHODS: Fundus photography, fluorescein angiography, and light and transmission electron microscopy were used to study the efficiency of photodynamic therapy-induced photothrombosis using indocyanine green as the photosensitizer, and to assess the resultant collateral damage. The delivery system consisted of a modified infrared diode laser tuned to 810 nm, near the maximum absorption peak of indocyanine green. RESULTS: Choriocapillary occlusion was achieved at indocyanine green doses of 10 and 20 mg/kg and a radiant as low as 6.3 J/cm(2). When photodynamic therapy was performed with indocyanine green doses of 10 mg/kg, damage to the neural retina was minimal. Only inner photoreceptor segments showed degeneration, probably secondary to choroidal ischemia. Bruch membrane remained intact. Retinal pigment epithelium was invariably damaged, as seen with other photosensitizers. Temporary occlusion of large choroidal vessels occurred at both dye doses. CONCLUSIONS: In this experimental study, photodynamic therapy using indocyanine green and 810-nm light irradiation produced endothelium-bound intraluminal photothrombosis, with preservation of the retinal architecture and minimal loss of visual cells. Membrane targetability, hydrophilic and fluorescent properties, and activation at 805 nm suggest indocyanine green as a potential photosensitizer for choroidal neovascularization. These combined considerations point toward further study of photodynamic therapy using indocyanine green for the treatment of choroidal vascular disease.     

Experimental assessment of the capacities of use of photosense. Communication 2. Photodynamic therapy for epibulbar and choroid tumors

The study was undertaken to evaluate the efficiency of photodynamic therapy of pigmented choroid and epibulbar melanoma, by using the agent Photosense. Pigmented choroid and epibulbar melanoma were identified in 50 rabbit eyes. After intravenous injection of Photosense, 0.7 mg/kg, the tumors were irradiated at 675 nm with an argon-pumped dye laser at 150 J/cm2. Photodynamic therapy with Photosense may be effective in treating pigmented choroid and epibulbar melanomas.     

超声造影对兔眼脉络膜黑色素瘤血流循环的定量评估

目的 探索改良兔眼脉络膜黑色素瘤原位肿瘤模型的建立方法及成瘤影响因素;观察超声造影在定性和定量评估脉络膜黑色素瘤血液循环模式中的应用.方法 将脉络膜黑色素瘤的瘤组织碎片植入10只兔的右眼,左眼作为对照.4周后采集兔眼普通超声、多普勒彩色超声、超声造影图像并自动记录,使用SonoLiver分析眼内肿块血流充盈曲线.结果 兔眼内脉络膜黑色素瘤成瘤率为80.0％(2只动物死亡).肿块体积最大切面两条垂直线测量值为(0.92±0.28)cm、(0.66±0.30) cm,肿瘤的峰值强度为(263.56±143.33)％,上升时间为(5.01±1.42)s,达峰时间为(6.17±2.18)s,渡越时间为(18.01±7.65)s,曲线下面积为(4505.13±1491.75)％,上升支斜率为59.48±75.18,下降支斜率为-7.21±5.76.肿瘤组织与正常眼球壁组织比较,峰值强度、曲线下面积、下降支斜率均较大(均为P＜0.05),上升时间、上升支斜率、达峰时间、平均渡越时间差异均无统计学意义(均为P＞0.05).结论 脉络膜下腔植入肿瘤组织碎片是一种稳定兔眼脉络膜黑色素瘤模型建立方法,超声造影在定性和定量判断眼脉络膜黑色素瘤内血流微循环模式中具有重要的临床价值.