摘要
目的：
1. 研究四种不同波长（1064、532、810和755nm）激光对兔视网膜的损伤与波长的关系及损伤的相关因素。
2. 对兔视网膜损伤的自我修复功能进行观察和分析。
3. 对MLG-O1和MLG-O2型单兵用激光防护眼镜进行生物效应的实验研究。
4. 研究激光损伤视网膜时对细胞粘附分子-1（ICAM-1）、白细胞介素-1β（IL-1β）和髓过氧化酶（MPO）的影响，从而对激光视网膜损伤和修复机理进行初步探讨。
方法：
1. 设计激光兔眼底损伤光路，建立兔眼底损伤模型。
2. 用眼底镜、眼底照相、眼底组织切片观察分析四个波长激光损伤兔眼底的情况，并进行比较；还包括不同能量密度单脉冲激光对眼底的损伤；相同能量密度和脉宽，不同脉冲数对眼底的损伤。
3. 激光致伤眼底后，即刻、1、3、7、14天和1个月用眼底镜和眼底照相等观察分析眼底的自我修复功能。
4. 使兔戴上防护镜观察防护镜所能承受的四个波长激光的最大能量密度和功率密度。以及兔裸眼接受上述激光照射的眼底损伤程度，从而找出防护镜对激光的防护阈值。
5. 运用原位杂交方法观察ICAM-1表达变化，测定MPO的变化来衡量中性粒细胞（PMN）含量的变化，用放免法测定IL-1β含量的变化，对眼底损伤和修复机理进行初步分析。
结果：
1. 造成相同的视网膜损伤时，其能量密度532nm的最小，755nm居
中，1064nm最大。因810nm脉宽和上述三者相差太大，不具有可比性。
2. 同一波长相同脉宽的激光随能量密度的增加眼底损伤越重。
3. 同一波长相同脉宽的激光，每个脉冲能量密度相同时，随脉冲数增加眼底损伤程度越重。
4. 单脉冲1064nm激光，能量密度1924mJ/cm2，脉宽5ns，在切片中发现有些兔眼脉络膜出血，其它三个波长的激光未发现脉络膜出血。
5. 1064nm和532nm激光，能量密度一定时可造成眼屈光介质的机械损伤，视网膜损伤则减轻。
6. 在四个波长激光致使兔视网膜出血的实验中，出血情况不一，有点状出血、菊花状出血、不规则片状出血等不同出血。
7. 在没有任何药物的治疗干预下，直径为1/3PD的兔视网膜凝固斑1月后观察完全吸收。直径1PD的凝固斑1月后损伤部位视网膜形成灰白色小点。2-3PD凝固斑吸收缓慢， 1月后发现凝固斑直径变化不大。点状出血1周后完全吸收。菊花状或片状出血区半月后亦完全吸收。玻璃体出血吸收缓慢，1月后仍可见暗红色陈旧性出血。
8. 实验表明MLG-O1和 MLG-O2型激光防护镜有效地防护了致青紫兰灰黑兔兔眼玻璃体出血或大出血的激光剂量。
9. 激光损伤后第1天视网膜MPO含量稍有增加，损伤第3天，视网膜MPO含量达高峰（P<0.05），后逐渐下降；伤后1天IL-1β含量骤然增加（P<0.05），后逐渐下降；伤后1-3天，邻近伤口的视网膜神经节细胞层和内核层ICAM-1mRNA表达阳性，后表达随时间延长逐步降低。
结论：
1．激光对视网膜的损伤与激光的波长、能量密度、脉冲频率及眼底结构有关。
2．视网膜对损伤有自我修复能力，但自我修复能力较弱，轻度损伤可以完全修复，中度以上损伤则不能完全修复。
3．MLG-01和MLG-02型激光防护镜能有效防护532nm、755nm、810nm和1064nm激光对兔视网膜损伤。
4. ICAM-1、MPO和IL-1β介导视网膜激光损伤的炎性反应。
Purpose
1. To study the relationship between damages on rabbit retina after laser irradiation of four different wavelengths (532, 755,810and1064nm) and damage-related factors.
2. To observe and analyse the self-recovering function of the damages on rabbit retina.
3. to research on biological effects of MLG-01 and MLG-02 solo-soldier laser protective eyewear.
4. To starte a preliminary research on retina damage after laser irradiation and the recovering mechanism through the study of the influences on ICAM-1, IL-1β and MPO when retina being hurt by laser,.
Methods
1. Design light-paths of the damage of rabbit retina for laser irradiation and set up a model.
2. Observe, analyse and compare the different cases of the damages of the rabbit retina after irradiation by four different wavelength laser using funduscopy, fundusphotography and tissue section of fundus including damage on the retina by different energy-density solo-pulse laser; retina damage by same, energy-density and wavelength but different pulses; and the influences on the damage when laser functioning at the point in the fundus.
3. After the damage of the fundus of rabbit retina by laser, use funduscopy and fundusphotography to observe and analyse the self-recovering mechanism of the eye after 1, 3, 7, 14 days and a month.
4. Let the rabbit put on laser protective eyewear to observe the maximum energy-density and power-density which the spectacle can endure, compare the degree of damage on the rabbit retina when without spectacle thus find out the
protective point of the laser protective eyewear.
5. Use in situ hybridizationa to observe the changes of ICAM-1, measure the changes of MPO to weigh the changes in volume of PMN; use radio-immunity method to measure the volume changes of IL-1βand carry out a preliminary analysis of the retina damage and its recovering mechanism.
Results
1. When laser caused same retina damages 532nm energy-density was the weakest, 1064nm was the strongest, while 755nm was moderate.
2. Retina damages became heavier with the increase of energy-density of the laser of the same wavelength and pulse wideth.
3. Retina damages became heavier with the increase of pulse of the laser though with the same wavelength and same pulse energy-density.
4. In the damages caused by four different wavelength laser, bleeding of choroid in the tissue section of rabbit eye when irradiated by 1064nm wavelength, 5ns pulse-width, 1924mJ/cm2 density laser was found but no bleeding of choroid was found when irradiated by three other wavelength laser. retina to choroid.
5. In the experiments of rabbit retina bleeding by the four wavelength laser irradiation, there were different types of bleeding, some were spot-like, some were chrysanthemum-like and some were irregular block-like.
6. When the rabbit eyes was irradiated by 1064nm wavelength, 4998mJ/cm2 energy density, 5ns pulse-width, single pulse laser, 3×1mm2 wound paths was found in some rabbit lens. When the wound paths appeared, the damage of the retina decrease. When irradiated by 532nm wavelength, 756mJ/cm2 energy density, 4ns pulse-width single pulse laser, white linear mixture was found in rabbit lens.
7. Without any medical treatment, photocoagulation of rabbit retina (diameter 1/3PD) recovered in a month. Photocoagulation (diameter 1PD) beceome little white sport in a month. Photocoagulation (2-3PD) absorbed slowly, no obvious change in diameter in a month. Spot-like bleeding totally absorbed in a week. After laser irradiation, chrysanthemum-like or block-like bleeding area completely absorbed in half a month. vitreous hemorrhage began
to absorb slowly, dark red old bleeding could still be found in a month.
8. MLG-01 and MLG-02 solo-soldier laser protection eyewear was sufficient to protect hare against retinal hemorrhage and vitreous hemorrhage by laser of four wave-length.
9. The first day after the injuries, the volume of MPO of retina increased slightly and the volume of IL-1β increased greatly(p<0.05); the third day after
引文
1 高光煌,陈迹.激光辐射伤医学防护.军事医学科学出版社，北京,1998：74
2 Harlan JM. Leukocyte-endothelial interactions.Blood ,1985;65(3): 513-25
3 Leff JA, Bear JW, Kirkman JM, et al. Liposome-entrapped PGE1 posttreatment decreases IL-1 alpha-induced neutrophil accumulation and lung leak in rats. J Appl Physilo,1994;76(1):151-157
4 罗武生，郭兆贵.用髓过氧化酶法测定心肌组织中的中性粒细胞.中国药理学通报，1990,（4）：264-266
5 Krawixz, J. EP. Sharon, and W.F. Stenson. Quantitative assay for acute intestinal inflammation based on myeloperoxidase activity. Gastroenterlolgy, 1984;87:1344-1350
6 颜光涛等.特异性血清中白介素1β放射免疫分析的建立及初步应用.生物工程进展，1999，19（6）：68-70
7 Hazuda D et al. Purification and characterization of human recombinant precursor interleukin 1β. J Diol chem 1989;264:1689-1693
8 Platanias LC et al. Interleukin-1: biololgy, pathophysiology and clinica prospects. Am J Med,1990;89:621-629
9 Nagineni CN, Kutty-RK, Detrick-B et al. Inflammatory cytokines induce intercellular adhesion molecule-1 (ICAM-1) mRNA synthesis and protein secretion by human retinal pigment epithelial cell cultures. Cytokine, 1996;8(8):622-30
10 Larcher-C, Recheis-H, et al. Influence of viral in fection on expression of cell surface antigens in human retinal pigment epithelial cells. Graefes-Arch-Clin-Exp-Ophthalmol.1997 Nov; 235(11):709-16
11 Richardson-PR, et al. Immunocytochemical study of retinal diode laser photocoagulation in the rat. Br-J-Ophthalmol. 1996 Dec; 80(12):1092-8
高光煌,陈迹.激光辐射伤医学防护. 军事医学科学出版社，北京,1998：
12 75-76
13 唐建民主编.激光美容与皮肤病治疗学.军事医学科学出版社,北京,2000;4:66
14 唐建民主编.激光美容与皮肤病治疗学.军事医学科学出版社,北京,2000;4:72
15 Roider J, et al. Retinal photocoagulation with a pulsed: frequency-doubled Nd:YAG laser (532nm). Ophthalmologe,1994,91(6):777-782
16 唐建民等.防激光致盲镜动物实验报告.中国医学物理学杂志,1999,16(1):20-21
17 Gorgels TG, van Norren D. Ultravioler and green light cause different types of damage in rat retina. Invest Ophthmol vis Sci 1995;36:851
18 Roider J, Michaud NA, Flotte TJ, et al. Response of the retinal pigment epithelium to selective photocoagulation. Arch Ophthalmol 1992,110:1786-1792
19 Gorgels TG, van-Norren-D. Ultraviolet and green light cause different types of damage in rat retina. Invest Ophthalmol Vis Sci,1995;36(5): 851-63
20 Shahinfar S, Edward-DP, Tso-MO. A pathologic study of photoreceptor cell death in retinal photic injury. Curr Eye Res,1991;10(1): 47-59
21 Ryan SJ. Subretinal neovasculation: Nature history of an experimental model. Arch Ophthalmol,1982;100:1804
22 Ham WT, et al. Basic mechanisms underlying the production of photochemical lesions in the mammalian retina. Curr Eye Res.1984;3:165
23 Parvet LM, Auker CR, Fine BS, et al. Dexamethasone protection against photochemical retinal injure. Arch Ophthalmol,1984,102:772-777
24 Ishibashi T, Miki K, Sorgente N, et al. Effects of intravitreal adminstration of steroids on experimental subretinal neovascularization in the subhuman primate. Arch Ophthalmol,1985,103:708-711
25 曲治华，王朝清.激光对人的有害作用及防治措施.北京：人民卫生出版社，1985,247-249
26 Douki T, Lee S, Dorey K, et al. Stress-wave-induced injure to retinal pigment epithelium cells in vitro. Lasers Surg Med,1996,19:249-259
27 Pollack A, Korte GE. Restoration of the outer blood-retinal barrier after krypton laser photocoagulation. Ophthalmic Res,1993,25:201-209.
28 何守志等.视网膜色素上皮细胞间通讯功能激光损伤的防护.中华眼科杂志，1999，35（3）：216-218
29 惠延年.墨西哥第三届国际外伤研讨会.国外医学眼科学分册,1995;19(1):1-5
30 王迪深，金惠铭.病理生理学.北京：人民卫生出版社，1994，372-375
31 Karagouni-EE, Chryssikopoulos-A, Mantzavinos- T et al. Interleukin-1beta and interleukin-1alpha may affect the implantation rate of patients undergoing in vitro fertilization-embryo transfer. Fertil Steril, 1998; 70(3):553-59
32 De Los Santos MJ, Mercader A, Frances A, et al. Role of endometrial factors in regulating secretion of components of the immunoreactive human embryonic interleukin-1 system during embryonic development. Biol Reprod, 1996;54(2):563-74
33 Mendoza C, Cremades N. Ruiz-Requena E, et al. Relationship between fertilization results after intracytoplasmic sperm injection, and intrafollicular steroid, pituitary hormone and cytokine concentrations. Hum Reprod,1999;14(3):628-35
34 Rosenbaum,-J-T; Samples,-J-R; Hefeneider,-S-H et al. Ocular inflammatory effects of intravitreal interleukin 1. Arch-Ophthalmol. 1987 Aug; 105(8): 1117-20
Cominelli-F, Nast-C-C, Clark-B-D, et al. Interleukin 1 (IL-1) gene expression, synthesis, and effect of specific IL-1 receptor blockade in rabbit
35 immune complex colitis. J-Clin-Invest. 1990 Sep; 86(3): 972-80
36 Lavail MM, Unoki-K, Yasumura-D et al. Multiple growth factors, cytokines, and neurotrophins rescue photoreceptors from the damaging effects of constant light. Proc Natl Acad Sci USA,1992;89(23): 11249-53
37 Bevilacqua MP. Endothelial-leukocyte adhesion molecules. Ammu Rev Immunol,1993,11:767-804
38 Larson RS, Springer-TA. Structure and function of leukocyte integrins. Immunol Rev,1990;114:181-217
39 Harlan JM. Leukocyte-endothelial interactions.Blood ,1985;65(3): 513-25
40 Freedman-S-F, Hatchell-D-L. Enhanced superoxide radical production by stimulated polymorphonuclear leukocytes in a cat model of diabetes. Exp-Eye-Res. 1992 Nov; 55(5): 767-73
41 张晓红等.细胞粘附分子在眼部的作用.国外医学眼科学分册，1999，23，（6）：349-354
42 Martin SD, Springer TA. Purified ICAM-1 is a ligand of LFA-1. Cell, 1987,51:813-819
43 Staunton DE, Dustin ML, Springer TA. Function cloning of ICAM-1, a cell adhesion ligand for LFA-1 homologous ICAM-1. Nature,1987,57:813-819
44 Dustin ML, Rothleim R, Bhan AK, et al. Introduction by IL-1 and interferon-γ, tissue distribution, biochemistry, and function of a natural adhesion molecule(ICAM-1). J Immunol, 1986;137:245-54
45 Richardson-P-R, Boulton-M-E, Duvall-Young-J et al. Immunocytochemical study of retinal diode laser photocoagulation in the rat. Br-J-Ophthalmol. 1996 Dec; 80(12): 1092-8
46 卢信义主编.激光眼科学.北京：人民卫生出版社.1981：86
47 程凌云，范雪定。重度氩激光光凝后视网膜色素上皮屏障的修复。眼底病杂志。1992；3：14
Bamforth-S-D, Lightman-S-L, Greenwood-J. Ultrastructural analysis of
48 interleukin-1 beta-induced leukocyte recruitment to the rat retina. Invest-Ophthalmol-Vis-Sci. 1997 Jan; 38(1): 25-35
49 Pollack-A, Korte-G-E, Heriot-W-J, et al. Ultrastructure of Bruch's membrane after krypton laser photocoagulation. II. Repair of Bruch's membrane and the role of macrophages. Arch-Ophthalmol. 1986 Sep; 104(9): 1377-82
50 Boisjoly-H-M, Laplante-C, Bernatchez-S-F, et al. Effects of EGF, IL-1 and their combination on in vitro corneal epithelial wound closure and cell chemotaxis. Exp-Eye-Res. 1993 Sep; 57(3): 293-300
51 Rosenbaum JT, Samples-JR, Hefeneider-SH et al. Ocular inflammatory effects of intravitreal interleukin 1. Arch Ophthalmol,1987;105(8): 1117-20
52 高光煌, 陈迹.激光辐射伤医学防护.军事医学科学出版社，北京,1998：84