【作者】 黄平；

【导师】 刘承宜；

【作者基本信息】 华南师范大学 ， 生理学， 2004， 硕士

【摘要】 随着激光研究理论和生产技术的不断发展，不但高强度激光手术在外科、眼科、牙科和皮肤科等领域获得了越来越为广泛的应用，而且低强度激光的光生物调节作用研究也已经获得相当的进展，但很少有人注意到两个领域的交叉。高强度激光在生物组织中迅速衰减，激光束边缘强度低到无法损伤细胞，这部分激光对已损伤组织边缘存活细胞的光生物调节作用（例如，对细胞增殖的促进或抑制效应，对免疫细胞功能的调节）是决定高强度激光手术后生物修复状况的主要因素之一。为了尽量减少激光束对存活细胞的热效应，本文用单层成纤维细胞作为模型，首先用高强度激光对单层成细胞造成局部破坏，然后研究高强度激光束边缘对周围细胞的光生物调节作用对单层细胞生物修复的影响。 在本文第一章中，首先讲述了机械创伤愈合以及高强度激光手术后创口愈合的生理过程。指出高强度激光在切割组织的同时，还对创口附近组织产生了两种一般外科手术所没有的效应：一是热损伤效应，二是光生物调节效应。这两种效应都会影响术后创口愈合过程；然后，介绍成纤维细胞体外培养在创伤愈合研究中的应用；最后，简述人皮肤成纤维细胞的培养方法。 在第二章中，本文提出建立细胞模型，用MTT比色法研究高强度Nd：YAG和KTP倍频激光照射对单层人成纤维细胞的损伤及其生物修复过程。同时通过多项式数学模型r_d=A+B₁×E_pulse+B₂×E_pulse²【r_d是损伤半径（mm），E_pulse是脉冲能量（mJ）】，对实验结果进行数值拟合，探究激光照射造成的损伤和照射参数的关系。实验结果表明，无论是1064nm照射还是532nm照射，即时存活率都随着脉冲能量的增加或者脉冲频率的升高而降低；损伤半径随着脉冲能量的增加或者脉冲频率的升高而增大。而在相同参数的条件下，1064nm照射组的即时存活率比532nm照射组的小；前者的损伤半径比后者的较大。数值拟合结果表明，无论是红外光，还是绿光，脉冲频率越高，参数A越大，表示脉冲能量较低时，高频率的照射比低频且相同脉冲能量的照射所造成的细胞损伤范围更广；脉冲频率越高，参数B₁越小，表示高频照射时，不同的脉冲能量引起损伤半径的差异比低频照射时较小。 在第三章中，首先综述了低强度激光对离体细胞的光生物调节作用，指出低高强度Nd涅人G及其KrP倍频激光对单层成纤维细胞损伤作用和光生物调节作用的初步研究强度激光对生物细胞的刺激作用的普遍性。在“可见光对细胞的光生物调节作用”这一节中，重点讲述了激光对淋巴细胞、巨噬细胞和成纤维细胞的刺激作用。旨在突出低强度激光对伤口愈合的影响。然后运用刘承宜等提出的低强度激光的信息生物模型（BIMP），从协同光信号转导的角度解释了低强度激光的信息功能。 本文的第四章探讨了高强度Nd:YAG和KTP倍频激光束边缘对单层人成纤维细胞的光生物调节作用。实验结果表明 1064刊m激光照射后24小时，1仪腼J/脉冲组和15OtnJ/脉冲组的细胞显著增殖印<0 .05);而532钊叮激光照射后24小时，1侃haJ/脉冲组的细胞显著抑制印<0.05)。其他各组和对照组相比有差异但不明显。光照48小时后，各组结果与24小时的结果相同，但1以讲刊m照射组与对照组的差异增大，而532钊m照射组与对照组的差异减少。激光束损伤半径以外强度分布的分析和BIMP的应用研究表明，在高强度激光束照射细胞单层时，激光束损伤半径以外的边缘部分对存活的细胞可能具有光生物调节作用。 本文的研究表明，在用高强度激光束照射细胞单层时，激光束除了对细胞单层有局部损伤作用外，不同参数的激光束对存活细胞具有不同的光生物调节作用。因此，在高强度激光的临床应用中，对于激光的种类和强度的选择应给与足够的重视和考虑。更多还原

【Abstract】 With the development of the theoretical and technical study of laser, the usage of laser in the surgery and medicine is more and more extensive. The lasers used in surgeries, such as ocular surgery, dental surgery, dermatic surgery and medical surgery, are because their functional thermal ablation and melting ablation. Meanwhile, The biomodulation of low intensity laser irradiation was found in many years ago. However there is still little of study focusing on the intersection of the two fields. In fact, the intensity of high power laser beem decreases rapidly when laser is transmissing in bio-tissue and the intensity of the edge of laser beem is so low that photobiomodulation , which might effect the healing process after laser surgery, but not thermal effect might work. In this thesis, monolayer human skin fibroblasts wasused as models to study the damage effect of high intensity laser irradiation and photobiomodulation of marginal irradiation of high intensity laser beam.In the first part of this thesis, the healing processes after mechanical wounded and laser surgery were discussed first. In this section, the thesis showed that the effects, which were thermal effect and photobiomodulation, induced by laser ablation responded to the healing delay. Then the thesis summoned the effects of fibroblasts on wound healing. How to cultivate human skin fibroblasts in vitro was also summaried in this thesis.In the second part, with the help of MTT assay, monolayer human skin fibroblasts were used as models to study the damage effects of Nd:YAG and KTP laser irradiation. In order to understand the relationship between damage effect and,irradiation parameter, polynomial model rd = A + B1 X Epulse + B2 x E2pulse (the rd is damage radius (mm) and the Epulse is pulse energy (mJ)) was used to fit out the damageradius-pulse energy curves. The results showed that the survival rate (damage radius) decreased (increased) with the increase (decrease) of pulse energy or pulse rate immediately after 532nm or 1064 nm laser irradiation, and the survival rate (damage radius) immediately after 532nm laser irradiation was slightly larger (smaller) than the one immediately after 1064 nm laser irradiation at the same pulse energy and pulse rate. The fitness results indicated that coefficient A increased but coefficient Bl decreased with the increase of frequency, regardless the wavelength.Recent advance of effects of low intensity laser irradiation on cells were summarized in this thesis. The effects of low intensity laser irradiation onlymphocyte , macrophage, fibroblasts and the biostimulation effects of low intensitylaser irradiation were included in the third part. Then the biological information model of photobiomodulation (BIMP) was introduced to explain photobiomodulation and the information function of low intensity laser irradiation.Monolayer human skin fibroblasts were used again to study the photobiomodulation of marginal irradiation of high intensity laser beam in the fourth part. The results showed that HSFs proliferation was significantly promoted 24hours/48 hours after 1064 run laser treatment at 100mJ and 150mJ pulse energy, but significantly inhibited 24 hours/48 hours after 532 run laser treatment at lOOmJ pulse energy. These results indicated that when cells were irradiating, the energy of laser not only transformed into heat energy, which responded to the thermal effects, but also modulated the cell function as the photo signal.In conclusion, when high power laser irradiation is used on monolayer cells, thermal effect might works in the central area and the photobiomodulation, which depends on the parameters of the laser output, might works in the rest. The category of laser and the parameters of output are important to laser surgery.更多还原