微脉冲半导体激光对兔眼视网膜损伤的形态学观察

目的:研究微脉冲半导体激光对兔眼视网膜损伤的生物学效应.方法:有色兔8只16眼,每眼上、下方视网膜随机分别行810nm半导体激光连续波阈值光凝及微脉冲阈下光凝,于光凝后即刻观察光斑反应和眼底荧光造影后荧光素渗漏情况,并在光镜和电镜下观察其对视网膜和脉络膜造成的组织学改变.结果:微脉冲激光阈下光凝后光斑不可见,亦无荧光素渗漏,视网膜损伤不明显;阈值光凝后可见光斑反应并有荧光素渗漏,视网膜损伤明显,外核层细胞数量减少,内、外核层均出现少量核固缩和胞质空泡化,视网膜色素上皮细胞增生,Bruch膜完整无损.结论:微脉冲激光光凝视网膜不会损伤视网膜,但确定阈能量时要从低能量起,且选择周边视网膜.     

微脉冲半导体激光与氩激光对兔视网膜损伤的形态学比较

目的:研究微脉冲半导体激光和氩激光对兔眼视网膜损伤的形态学改变。方法:有色兔8只16眼,每只兔的两眼随机分别行810nm微脉冲阈下光凝和514nm氩绿激光光凝,在黄斑水肿治疗参数下,光凝后即刻观察光斑反应并在光镜和电镜下观察其对视网膜和脉络膜造成的组织学改变。结果:微脉冲激光阈下光凝后光斑不可见,氩激光光凝后可见光斑反应;激光后即刻两者光镜下视网膜感光细胞损伤不明显但脉络膜毛细血管、小静脉轻度充血,电镜下氩激光光凝处外节排列稀疏,色素上皮细胞和节细胞肿胀,外颗粒层细胞异染色质增多。微脉冲光凝处外节排列较整齐,外颗粒层变化不明显。结论:微脉冲激光光凝视网膜较氩激光损伤更轻微。     

微脉冲半导体激光与氩激光对兔视网膜损伤及修复的组织学研究

目的观察微脉冲半导体激光与氩激光对兔视网膜损伤和修复的组织学反应。方法在黄斑水肿治疗参数下分别对兔眼行微脉冲阈下光凝和氩激光光凝,观察视网膜组织学改变。结果微脉冲阈下光斑不可见而氩激光可见;激光后两者光镜下视网膜感光细胞损伤不明显但脉络膜小静脉轻度充血,4周后消退;电镜下氩激光光斑处外节排列稀疏,色素上皮细胞（RPE）和节细胞肿胀,外核层细胞异染色质增多,4周后RPE和胶原增生明显,脉络膜内色素颗粒增生;而微脉冲光斑变化不明显。结论微脉冲激光光凝视网膜较氩激光损伤更轻微。     

577nm阈下微脉冲激光与577nm激光光凝对兔视网膜组织影响的比较

背景 近年来研究表明,577 nm阈下微脉冲激光治疗视网膜疾病可达到传统577 nm激光光凝的治疗作用且对视网膜组织损伤小,但其具体作用机制和敏感的靶细胞尚未完全阐明. 目的 探讨和比较577 nm阈下微脉冲激光与577 nm激光光凝视网膜后成年中华黑兔视网膜组织形态学变化,为577 nm阈下微脉冲激光光凝在临床上的应用提供依据.方法 采用抽签法按照视网膜光凝条件不同将26只中华黑兔分为正常对照组(2只)、577 nm激光组(6只)和阈下微脉冲激光组(18只),其中阈下微脉冲激光组按照激光工作负载率的不同亚分为9％、12％和15％阈下微脉冲激光组,每组各6只,正常对照组不做任何处理.光凝后行彩色眼底照相及OCT检查,摘取兔眼球壁行苏木精-伊红染色,光学显微镜下观察兔脉络膜和视网膜组织结构的变化.结果 彩色眼底照相和OCT显示正常对照组兔眼视网膜组织结构清晰.9％阈下微脉冲激光组OCT扫描可见视网膜神经上皮层稍模糊;12％阈下微脉冲激光组光凝斑处视网膜神经上皮层轻度水肿,视网膜色素上皮(RPE)层稍模糊;15％阈下微脉冲激光组可见光凝斑处视网膜神经上皮层明显水肿,RPE层局限性隆起;各阈下微脉冲激光组彩色眼底照相均未见光凝斑.577 nm激光组兔眼彩色眼底照相可见灰白色光凝斑,OCT扫描层面可见视网膜呈多灶性隆起,视网膜各层组织结构模糊,伴浆液性神经上皮层脱离.视网膜组织病理学检查可见,与正常对照组兔眼相比,9％和12％阈下微脉冲激光组兔眼脉络膜血管变形或出血,但视细胞形态结构、双极细胞层和视网膜神经节细胞(RGC)层未见明显改变;15％阈下微脉冲激光组视细胞扁平状膜盘肿胀,双极细胞层和RGC层未见明显改变;577 nm激光组兔眼视细胞层、双极细胞层和RGC层结构紊乱,RPE层变薄.结论 577 nm阈下微脉冲激光对脉络膜层和RPE层具有高度选择性,视网膜光凝后对视网膜神经上皮层的损伤程度轻微,既可发挥治疗作用,又不损伤视网膜神经上皮;577 nm激光视网膜光凝可对视网膜全层造成损伤.     

兔视网膜在不同阈值激光作用后MMP-9表达的变化

目的:观察兔视网膜在不同阈值577nm激光作用后MMP-9的表达变化.方法:将色素兔26只采用抽签法随机分为正常对照组2只、常规光凝组6只和阈下微脉冲光凝组18只.正常对照组不做任何处理,常规光凝组行577nm激光光凝,阈下微脉冲光凝组又亚分为三个亚组,分别行9%、12%、15%工作负载率的577nm阈下微脉冲激光光凝.采用免疫组织化学法检测各组兔眼视网膜上MMP-9的表达情况.结果:常规光凝组:RPE层及视细胞层MMP-9呈强阳性表达,较阈下微脉冲组明显增多,差异有统计学意义(P<0.05);阈下微脉冲组:9%工作负载率见RPE层及视细胞层MMP-9少许阳性表达,12%工作负载率见RPE层及视细胞层阳性表达增多、胞核也出现少许阳性表达,15%工作负载率RPE层及视细胞层中度阳性表达,三个亚组间差异无统计学意义(P>0.05).结论:577nm阈下微脉冲光凝在9%、12%、15%工作负载率下对视网膜色素上皮层具有高度选择性,对视网膜神经纤维层无明显损伤,较常规577nm激光光凝更加安全.     

兔眼经瞳孔温热疗法阈值能量治疗的组织病理反应和细胞凋亡

目的:观察兔眼视网膜经瞳孔温热疗法(transpupillary thermotherapy,TTT)阈值能量照射后组织病理反应和细胞凋亡情况.方法:健康青紫兰兔20只,采用阈值能量对实验眼采用1.2mm光斑的810nm激光照射60s.采用眼底镜和眼底照相对光斑进行形态学研究,采用光镜和电镜的方法研究光斑的组织病理和超微结构的改变,采用TUNEL法、荧光素标记Annexin Ⅴ-FITC/PI双染色流式细胞测定法观察视网膜细胞的凋亡.结果:TTT后1d可见视网膜轻度灰白色水肿,后视网膜逐渐出现色素沉着.组织病理学切片显示神经节细胞无显著性破坏.TUNEL染色可见视网膜全层均有细胞凋亡的发生,以内颗粒层为主,流式细胞双染测定显示以凋亡为主.结论:阈能级TTT照射未引起神经节细胞严重损伤,较安全,其作用机制以细胞凋亡为主.     

577nm阈下微脉冲激光治疗黄斑水肿时光凝斑的影像学观察

目的 用多波长照相技术(F-10)观察577 nm阈下微脉冲激光光凝治疗黄斑水肿时光凝斑的形态学改变.方法 前瞻性系列病例研究.黄斑水肿患者17例(19眼)纳入研究.所有患者均经眼底荧光素血管造影(FFA)及光学相干断层扫描(OCT)检查确诊,其中,糖尿病黄斑水肿患者8例(10眼),视网膜中央静脉阻塞引起的黄斑水肿患者5例(5眼),视网膜分支静脉阻塞引起的黄斑水肿患者4例(4眼).首先在传统连续波激光下对患眼进行阈能量P测定,然后在F-10的retro mode模式检测下对这19眼进行阈下微脉冲光凝测试及治疗,微脉冲激光所用能量的调整参照阈能量P.留取患眼光凝前后眼底彩照及自发荧光成像,并随访观察术后1个月患眼中心凹厚度(CMT)及最佳矫正视力(BCVA)变化.分别采用配对t检验、配对秩和检验对数据进行分析.结果 术后即刻retro mode成像中,6眼在122.4％P能量下出现光凝斑;8眼在153.1％P能量下出现光凝斑;5眼在183.7％P能量下出现光凝斑.术后即刻在眼底彩照及自发荧光成像下光凝区未发生改变.术后1个月患眼CMT较术前明显降低,差异有统计学意义(t=4.238,P＜0.01),BCVA与术前相比无明显变化.结论 Retro mode 成像可检测到阈下微脉冲激光光凝产生的不可见光凝斑,在治疗时可用于指导光凝能量大小的选择.     

微脉冲半导体激光照射对正常大鼠视网膜损伤的观察

目的观察810 nm微脉冲半导体激光照射对正常棕色挪威大鼠（BN 大鼠）视网膜的损伤。方法使用不同能量及负载系数（duty cycle, D C）的810 nm微脉冲半导体激光对130只BN大鼠眼进行照射。分别于激光照射后第1、3、7、1 4、28 d进行彩色眼底照相、荧光素眼底血管造影及组织病理学观察，并检测热休克蛋白(HSP-70)在视网膜的表达情况，用TdT介导dUTP缺口末端标记法(TUNEL)检查细胞凋亡 。结果阈值及阈上能量条件下，低DC时激光照射部位无光学显微镜下的组织病理学改变，高DC时出现可累及视网膜内核层组织的严重损伤；微脉冲半导体激光照 射后1 d大鼠视网膜内核层细胞HSP-70阳性表达细胞即较正常视网膜明显增加，3 d时达到高峰，以后逐渐下降，14 d时恢复近正常水平。HSP-70阳性细胞数量 随激光能量提高而增加。TUNEL染色可见激光照射部位凋亡细胞主要存在于视网膜色素上皮（RPE）层、外核层、内核层，甚至脉络膜层，其数量随激光能量增高而增多。在激光照射后第3 d，凋亡细胞数量最多。结论810 nm微脉冲半导体激光照射后，视网膜损伤程度与激光能量及DC呈正相关。低能量高负载系数（50 mW，50％）或高能量低负载系数（100 mW，5％～15％）时，损伤限于RPE层，避免了神经上皮层的损伤。激光照射后HSP-70高表达及细胞凋亡可能在组织损伤修复过程发挥重要作用。 （中华眼底病杂志，2008，24：122-126）     

微脉冲半导体激光的临床应用现状

微脉冲半导体激光是一种阈下光凝技术 ,它能选择性地作用于视网膜色素上皮细胞 ,避免损伤视网膜神经层 ,在产生治疗作用的同时最大限度地保持视功能 ,对治疗黄斑水肿、中心性浆液性脉络膜视网膜病变 (中浆 )和玻璃膜疣具有较好的疗效。本研究阐述了微脉冲半导体激光的原理、临床应用及激光光凝斑检测等方面的进展。     

微脉冲半导体激光治疗CSC的初步观察

目的:探讨微脉冲半导体激光治疗中心性浆液性脉络膜视网膜病变(central serous chorioretinopathy,CSC)的疗效。方法:自身对照病例研究。将经荧光素眼底血管造影(fundus fluorescein angiography,FFA)和光学相干断层扫描(optic coherence tomography,OCT)检查确诊的典型性CSC患者12例12眼行微脉冲半导体激光治疗前及治疗后随访观察1,2wk;1,3mo的临床资料进行分析,主要以治疗前后的视力、自觉症状、眼底情况、FFA及OCT检查的改变为观察指标,以评价微脉冲半导体激光对CSC的治疗效果。结果:所有患者自觉症状减轻或消退,视力及视觉质量均有不同程度改善。激光光凝2wk和1mo后,分别有9眼(75.0%)和11眼(91.7%)的黄斑区水肿完全消退,3mo后全部患者黄斑区水肿消退。光凝2wk后,FFA检查发现有9眼荧光素渗漏完全消失,3眼渗漏减轻;OCT检查有9眼黄斑区视网膜下积液消失,中心凹视网膜厚度恢复正常,3眼黄斑区视网膜下积液大部分吸收。光凝1mo后,FFA检查有11眼荧光素渗漏完全消失;OCT检查有11眼黄斑区视网膜下积液消失,中心凹视网膜厚度恢复正常。光凝3mo后,FFA显示所有患眼的荧光素渗漏完全消失,未见明显"窗样缺损"样荧光;OCT检查发现所有患眼黄斑区视网膜下积液完全吸收。结论:微脉冲半导体激光治疗CSC是一种安全、有效的治疗方法,能有效缩短病程,提高患者视力,避免并发症的发生。     

氩绿激光致兔视网膜感光细胞凋亡的实验研究

目的:观察氩绿激光对兔视网膜损伤及修复的组织学改变及对兔视网膜感光细胞凋亡的影响。方法:将8只有色兔(16眼)中每只眼的上、下方视网膜随机分配为激光光凝区及空白对照区,光凝后24h、4wk通过光镜和电镜及TUNEL技术观察视网膜改变及感光细胞凋亡。结果:(1)光镜观察:光凝处视网膜损伤不明显但脉络膜小静脉充血,4wk后消退。(2)电镜观察:光凝处外节膜盘排列稀疏,神经节细胞轻微肿胀,外核层细胞异染色质增多,4wk后色素增殖,胶原增生。(3)TUNEL染色观察:光凝后24h,光凝处外颗粒层可见较多阳性细胞,内颗粒层偶见,4wk后接近正常。(4)感光细胞凋亡率:光凝后24h,感光细胞凋亡率较正常对照组增加,差异有统计学意义(P<0.01)。光凝后4wk较24h凋亡率显著降低,差异有统计学意义(P<0.01)。结论:轻度氩绿激光光凝视网膜损伤轻微。     

Micropulse and continuous wave diode retinal photocoagulation: visible and subvisible lesion parameters

BACKGROUND/AIM: Subvisible micropulse diode laser photocoagulation localises retinal laser damage because brief micropulses allow little time for heat conduction to spread temperature rise from the retinal pigment epithelium to the neural retina. Treatment power is often chosen as a multiple of that needed for visible continuous wave lesions. The authors measured clinical laser powers needed for visible end point micropulse and continuous wave diode laser retinal photocoagulation. METHODS: Six parallel rows of 10 diode laser (810 nm) burns were made in the superior peripheral retina of six consecutive patients undergoing their initial frequency doubled Nd:YAG (532 nm) panretinal photocoagulation for proliferative or severe non-proliferative diabetic retinopathy. All photocoagulation exposures were 125 microm in retinal diameter and 0.2 seconds in duration. Micropulse exposures were performed with 500 Hz, 0.3 ms micropulses. The minimal power needed (1) for visible continuous wave diode photocoagulation was determined from two adjacent rows of laser lesions and (2) for visible micropulse diode photocoagulation from four additional adjacent rows of laser lesions. Fluorescein angiograms and red-free fundus photographs were obtained immediately and 6 days after laser photocoagulation in each patient. Calculations were performed to determine the extent to which clinical parameters exceeded ANSI Z136.1-2000 maximal permissible exposure (MPE) levels for laser exposure. RESULTS: Continuous wave and micropulse lesions typically required 300 mW (60 mJ) and 1800 mW (54 mJ), respectively. Visible continuous wave and micropulse lesions exceeded MPE levels by multiples of 36 x and 133 x, respectively. Laser energies were similar for visible continuous wave and micropulse lesions. CONCLUSION: Visible micropulse lesions require 6 x more power but roughly the same energy as visible continuous wave lesions. No significant difference was demonstrable in the minimal power needed for photographically and angiographically apparent diode micropulse lesions. MPE levels are designed to provide a 10 x safety margin. This safety margin was 3.7 x greater for micropulse than continuous wave diode laser photocoagulation.     

Subthreshold micropulse diode laser treatment in diabetic macular oedema

BACKGROUND/AIM: Enlargement of laser scars after retinal argon laser photocoagulation can give rise to deterioration in visual acuity. Subthreshold micropulse diode laser may decrease this risk. The aim of this study was to compare the effectiveness of subthreshold micropulse diode laser (810 nm) and conventional argon laser (514 nm) photocoagulation for the treatment of clinically significant macular oedema in diabetic patients. METHODS: 23 eyes of 16 patients were randomised to either treatment. Follow up was conducted for a minimum of 5 months. Changes in visual acuity and macular oedema measured by optical coherence tomography were examined. RESULTS: Visual acuity remained stable in all treatment groups throughout the observation period. Changes in retinal thickness were small both foveally and perifoveally. In patients with focal macular oedema a significant reduction in retinal thickness (9% approximately -26 microm, p = 0.02) was seen foveally 3 months after diode laser photocoagulation. CONCLUSION: Subthreshold micropulse diode laser and conventional argon laser treatment showed an equally good effect on visual acuity. Subthreshold micropulse diode laser showed a stabilising or even improving effect on macular oedema. The combination of primary diode laser and supplementary argon laser might be particularly favourable in reducing diabetic macular oedema.     

Predictors of drusen reduction after subthreshold infrared (810 nm) diode laser macular grid photocoagulation for nonexudative age-related macular degeneration

PURPOSE: To determine the predictors of drusen reduction in eyes with nonexudative age-related macular degeneration (ARMD) treated with subthreshold infrared (810 nm) diode laser macular grid photocoagulation. Additionally, to determine the relationship of laser-induced drusen reduction and best-corrected visual acuity (BCVA) 18 months after laser treatment.DESIGN: Randomized controlled clinical trial.METHODS: Fifty patients (100 eyes) with bilateral nonexudative ARMD were enrolled at two centers. One eye of each patient was randomized to the observation; the other eye was treated with 48 subthreshold (invisible end point) applications of infrared (810 nm) diode laser in a macular grid pattern. The eyes that received subthreshold laser treatment were compared with the eyes that received no treatment. The baseline fundus characteristics (number, size, and distribution of drusen, as well as focal hyperpigmentation) from two macula areas (central 1500 micro diameter, pericentral 1500 micro ring area) on stereo color photographs, the number of laser-induced lesions, and the area of laser induced retinal pigment epithelial (RPE) lesions on fluorescein angiography 3 months after treatment were studied as predictors of major drusen reduction (> or = 50% drusen reduction from baseline) 18 months after laser treatment. BCVA at baseline and 18 months later was compared in observation eyes and in laser-treated eyes.RESULTS: Eighteen months after randomization, 24 (48%) of 50 eyes treated with subthreshold laser had major drusen reduction compared with three (6%) of 50 observation eyes (P =.00001). At 3 months post-treatment in laser-treated eyes with major drusen reduction, the mean number of laser-induced lesions on fluorescein angiography was 30.7 and the mean area of RPE change was 0.81 mm(2) compared with 14.8 laser-induced lesions and 0.35 mm(2) area of RPE change in eyes without major drusen reduction (P =.0001 and P =.0003, respectively). At baseline, fundus characteristics were not significantly different between observation eyes and laser-treated eyes or between the major drusen reduction group and the nonmajor drusen reduction group. At 18 months after treatment, BCVA was not significantly different in laser-treated eyes and in observation eyes.CONCLUSIONS: Subthreshold infrared (810 nm) diode laser macular grid photocoagulation in eyes with nonexudative ARMD significantly reduced drusen 18 months after laser treatment. Both the number of subthreshold laser lesions and the area of RPE changes visible on fluorescein angiography 3 months after treatment appeared to be predictors for major drusen reduction 18 months after treatment. However, it remains to be determined whether laser-induced drusen reduction is beneficial for visual acuity or reduces the incidence of choroidal neovascularization (CNV) in eyes with nonexudative ARMD.     

Subthreshold transpupillary thermotherapy reduces experimental choroidal neovascularization in the mouse without collateral damage to the neural retina

PURPOSE: Transpupillary thermotherapy (TTT) is currently being evaluated for treatment of choroidal neovascularization (CNV) in age-related macular degeneration. To optimize TTT for CNV, the effect was analyzed of invisible (subthreshold) or visible (threshold) doses of TTT on the normal mouse retina and on experimental CNV. METHODS: TTT was delivered to the normal retina of 42 mice with a diode laser at increasing power settings (50, 60, 70, or 80 mW), to obtain thermal lesions ranging from invisible (subthreshold) to visible (threshold) burns. CNV was induced in 53 mice by krypton laser photocoagulation of the fundus, after which the CNV lesions were treated with TTT (50, 60, or 80 mW). Eyes were enucleated 7 days after TTT and prepared for histology, and the CNV complex was evaluated on hematoxylin-eosin stained serial sections by measuring the maximum height of the CNV lesions. Ultrastructural changes were examined by transmission electron microscopy. RESULTS: Increasing the TTT laser power yielded gradually more visible effects. At 50 mW, which induced subthreshold burns, no damage was seen in the neural retina, retinal pigment epithelium (RPE), or choroid at any time point. By contrast, eyes treated with higher power exhibited progressively more damage to the neural retina, including a complete disruption of the outer nuclear layer. When TTT was applied to the laser-induced CNV lesions, the height of lesions was significantly reduced (P < 0.001) in response to all three power settings at 7 days after treatment. The mean relative thickness of the CNV lesion was 3.29 +/- 0.89 in untreated mice, whereas in TTT-treated mice it was 1.69 +/- 0.35, 1.69 +/- 0.41 and 1.70 +/- 0.17 at power settings of 50, 60, and 80 mW, respectively. The overlying neural retina showed no apparent damage with the 50- or 60-mW settings, whereas outer nuclear layer disruption occurred with a power of 80 mW. Electron microscopy confirmed the presence of vascular occlusion at 1 day and a fibrotic scar at 7 days after TTT. CONCLUSIONS: Subthreshold TTT can effectively occlude newly formed vessels and cause regression of the experimental CNV complex without damaging the neural retina. The results demonstrate the importance of using subthreshold laser power in experimental and clinical evaluation of TTT.