分子超光谱成像系统应用于糖网病药物疗效研究

将成像技术和光谱技术相结合,再配合显微镜技术,研制了基于AOTF(acousto-optic tunable filters)的分子超光谱成像系统.使用该系统采集了正常、糖尿病和EPO(erythropoietin,促红细胞生长素)药物治疗的大鼠视网膜组织切片的分子超光谱图像数据.通过对正常组、糖尿病组、药物治疗组共30例样本的分子超光谱图像数据进行处理,获得了3组样本的单波段图像和伪彩色合成图像,并提取了各组样本外核层的典型透射光谱曲线.从图像上分析各组ONL(outer nuclear layer,外核层)的厚度,由大到小依次为正常组、治疗组、糖尿病组,由于糖尿病会引起视网膜外核层细胞凋亡和厚度减少,实验结果表明经EPO治疗后可增加视网膜外核层的厚度.从光谱上分析各组的透射强度,糖尿病大鼠视网膜外核层组织在550～1000 nm光谱范围内的透射强度整体高于正常组,经EPO治疗后,透射强度介于正常组和糖尿病组之间;通过光谱相似性分析,治疗组与正常组之间的光谱相似性高于糖尿病与正常组.实验结果表明EPO能减少视网膜外核层细胞凋亡,对糖网病大鼠有一定的疗效.因而分子超光谱成像系统可以作为一种新的手段,辅助科研人员对糖网病的发病机理和致盲原因及药物疗效进行研究.     

角膜类脂环超微结构的初步观察

角膜类脂环是角膜缘内的一圈灰白色混浊环,一般认为其发生基础为脂滴沉积。我们通过对类脂环超微结构的研究,对其发生机理提出了新的补充。样品取自新鲜尸体角膜和摘除眼球的角膜,常规超薄切片技术制样。观察发现角膜上皮细胞有核畸形,呈锯齿状,核被膜波浪状扩张,核周间隙增宽、核孔扩大,核内染色质边集,胞浆形成微绒毛,细胞器少。固有层中     

氮激光照射兔眼后的实验性观察

作者用高低功率N2激光照射兔眼，发现角膜缘、角膜、玻璃体、视网膜和视乳头有慢性炎症。低功率组发现角膜缘球结膜下淋巴细胞浸漓为主兼有成纤维细胞和新生血管。高功率组发现视网膜视乳头的改变比低功率明显。因此，N2激光照射对兔眼可引起眼组织慢性炎症，并能损害视细胞。     

靶向bcl-2小分子化合物抑制角膜新生血管研究

目的探讨靶向bcl-2小分子化合物(Z24)对大鼠角膜新生血管的抑制作用。方法采用完全随机化方法将30只大鼠分为2组,每组15只鼠(15只眼),应用角膜基质缝线法诱导角膜新生血管,采用灌胃给药。治疗组:Z24混悬液(80mg/kg);对照组:等量不含Z24的空白液;自术后第1天起,1次/d,连续用药7d。每组预先标定8只大鼠进行新生血管的定量观察,其余大鼠进行组织学观察。结果缝线后第7天,治疗组新生血管的面积明显小于对照组(P<0.05)。结论全身应用Z24能够延缓角膜新生血管的生成。     

532-二极管激光光凝法诱导大鼠青光眼慢性高眼压模型的研究

目的：建立532-二极管激光光凝的大鼠慢性高眼压性青光眼模型,评价模型眼的相关组织病理学改变。方法：选用成年SD大鼠50只,采用532-二极管激光行角膜缘小梁网和角膜缘颞侧及颞上、颞下3条巩膜浅层静脉光凝。定期测量和记录双眼眼压,观察眼前节情况。于术后1、3,6个月时摘取双侧眼球,观察双眼视网膜组织形态学改变、视网膜厚度及视网膜神经节细胞数目的改变。结果：（1）大鼠眼内压于1个月时达到眼压峰值,高眼压状态持续2月余,然后眼压水平逐渐下降,6个月时眼压基本恢复到正常范围;（2）视网膜厚度明显变薄,视网膜神经节细胞数明显减少。结论：532-二极管激光光凝SD大鼠小梁网及巩膜浅层静脉血管可以建立长期眼压中度升高的慢性高眼压性青光眼模型。模型眼出现了青光眼眼底特征性的形态学改变。     

高强度聚焦超声辐照后的活体兔角膜组织学观察

目的:观察高强度聚焦超声(High Intensity Focused Ultrasound,HIFU)辐照活体兔角膜基质后角膜的组织形态学变化,检测CollagenⅢ及α平滑肌肌动蛋白(α-SMA)在角膜的表达情况,探讨HIFU作用于兔角膜的生物学改变.方法:HIFU以圆环状定位辐照兔角膜周边基质,辐照完后于不同时间段(术后1,3,10,30,60,90天)行兔眼裂隙灯观察及角膜组织切片观察,并用免疫组化方法检测CollenⅢ,α-SMA在角膜的表达.结果:HIFU辐照活体兔角膜后立即在基质层形成灰白色圈,辐照圈于术后10天开始变淡消失.术后10天至60天角膜组织切片HE染色后均可见辐照处基质纤维排列紊乱,基质层收缩变薄,上皮增生,术后90天组织形态逐渐恢复.辐照部位CollagenⅢ,α-SMA的表达均于术后3天逐渐增加,术后30天达高峰,然后逐渐减少趋于平稳.周围区域角膜组织无明显变化.结论:HIFU辐照活体兔角膜基质后辐照区组织形态学改变明显,成纤维细胞活化,新生胶原纤维产生,至术后90天辐照区组织形态学改变渐趋恢复.     

由安全极限确定激光散斑在角膜上的辐照度

激光束直接照射人眼时，对人眼角膜和视网膜会造成损伤，而焊接光的漫反射光和激光散斑在一定条件下照射到人眼上，也会引起人眼角膜和视网膜的损伤。本文根据角膜上辐照度的安全极限，导出了激光器光功率与强漫射板上光斑直径、角膜到漫射板之间距离的关系式。     

姜黄素对大鼠角膜碱烧伤的作用

目的:探讨不同浓度姜黄素在大鼠角膜碱烧伤中的作用。方法:建立SD大鼠角膜碱烧伤模型,大鼠分为4组(第1至4组),对照组(第1组)只给予羧甲基纤维素钠,第2,3,4组分别给予100mg/kg BW2、00mg/kg BW及400mg/kg BW姜黄素。在碱烧伤后第3,7,14d用硫代巴比土酸(TBA)法测丙二醛(MDA)含量,化学比色法检测角膜超氧化物歧化酶(SOD)活力。结果:第1,2组之间和第3,4组之间的SOD,MDA无统计学意义。第1,2组与第3,4组相比,后者SOD活力明显增加,MDA含量显著减少。结论:姜黄素对大鼠角膜碱烧伤具有保护作用且与姜黄素浓度有关。     

OCT在眼科的应用

通常,外部光是通过角膜、前房、晶状体、玻璃体到达视网膜。一旦光入射到视网膜最外层的圆锥细胞、杆状细胞,光便被转换成电信号。经过视神经传递到大脑,形成视力。眼球部分、直到视网膜均由透明的组织构成。因此,眼科医生在日常临床中可以利用检眼镜和狭缝灯显微镜观察中间透光体和眼底,相当于生物显微镜的水平。目前眼科采用激光治疗颇为盛行,这是由于激光被导入眼底对眼睛无任何伤害。对于糖尿病视网膜症为主的视网膜血管病变,最基本的治疗方法就是利用激光的视网膜凝固法。     

器官培养的长管状软骨超微结构观察

本文利用器官培养的方法,观察长管状软骨的分化,成熟和化骨过程。软骨的生长在第14天长径生长最迅速,此后渐趋停止。电镜观察,培养2—4天的软骨细胞核与胞浆的比值大,细胞内可见多数小泡体。培养6天时原始骨化中心开始出现,粗面内质网和线粒体丰富,细胞开始分泌基质,基质内有具单位膜的基质小泡形成。说明化骨准备工作已经完善。第14天时肥大层软骨细胞内粗面内质网和线粒体更加丰富,内质网扩张呈池状,其内含有合成的点状或丝状物。本文因此提出利用软骨器官培养方法,在培养8—10天时,施加实验因素最宜。     

小麦颖果发育过程中韧皮部细胞的结构变化观察

利用光学显微镜、透射电镜和扫描电镜技术．对小麦颖果发育过程中韧皮部细胞的结构变化进行了系统观察。结果表明，开花后7d颖果还没有形成完全分化的维管束。开花后12d韧皮部可见明显的筛分子。每个维管束有大约40个左右的筛分子，它们呈半圆形分布于维管束外侧。开花后14～20d，筛分子内有较丰富的线粒体、P^-型质体和许多无定型丝状物，有些筛分子有珠光壁。筛分子有典型的伴胞，并形成SE／CC复合物结构。伴胞有稠密的胞质和核质。韧皮薄壁细胞与中间细胞间有丰富的胞间连丝；韧皮薄壁细胞间的胞间连丝呈区域集中分布。开花后24-29d，筛分子内线粒体数目减少，伴胞和韧皮薄壁细胞电子染色变浅，说明它们的养分运输能力下降。中间细胞的细胞质内仍有丰富的线粒体和其它细胞器，它们沿胞壁周缘分布；此时，中间细胞形成类似筛分子的结构．因此中间细胞是一种适应于养分运输的特化伴胞，在颖果灌浆后期起重要作用。开花后39d，韧皮部细胞衰老变形，此时颖果发育成熟。     

Ultracytochemistry of enzymes in cell biology of the retina.

In matured chick retina, alkaline phosphatase activity is specifically localized in the outer plexiform layer and in the horizontal and Müller cells. In developing chick retina, the activity is recognized in growing neurites from horizontal cells, when synaptogenesis begins in the outer plexiform layer. Using levamisole, a potent inhibitor of alkaline phosphatase, on chick retina in vivo and in vitro, the enzyme was shown to play a significant role in retinal cell differentiation. 5'-Nucleotidase is used as a marker for the rod photoreceptors. It became apparent that the 'displaced' rod cells are localized in the inner nuclear layer of postnatal retina. High activity of glucose-6-phosphatase is confirmed in the cisternae of the smooth and rough endoplasmic reticulum, together with the space of the perinuclear envelope in the pigment epithelium of rat. The process of disc membrane recycling in the rod outer segment was investigated cytochemically to reveal sequential changes in lysosomal digestion both by conventional enzyme cytochemistry and by high voltage electron microscopy. With conventional enzyme histochemistry as well as with rapid freeze substitution enzyme cytochemistry, all enzyme for cGMP metabolism were observed to be on the cytoplasmic side of the disc membranes.     

黄蜂刺伤角膜新生血管与肥大细胞关系的电镜观察

观察黄蜂刺伤角膜后产生的肥大细胞与角膜新生血管（corneal neovascularization，CNV）的关系，为探索黄蜂刺伤角膜致盲的发生提供形态学依据。取黄蜂刺伤角膜组织，常规透射电镜样品制备及透射电镜观察。发现角膜固有层内出现肥大细胞及产生新生血管，肥大细胞可迁移至新生血管附近，肥大细胞多呈显著脱颗粒。黄蜂刺伤角膜经蜂毒作用后产生的肥大细胞促进了角膜新生血管的形成，是致盲的主要因素。     

软骨细胞光生物调节作用的体外实验

研究了低强度激光对兔软骨细胞增殖和变异的潜在性影响。选取3周龄新西兰白兔分离软骨细胞,在新生牛血清(NCS)体积分数分别为10%,5%,2.5%和无血清4种培养媒介中培养,采用功率6.5mW的He-Ne激光辐照(HNI)(5.74mW/cm2)软骨细胞。用四氮唑复合物法(XTT)检测细胞的活性。通过检测羟脯氨酸的(Hrp)含量了解细胞合成胶原的能力。在激光共聚焦显微镜下观察吖碇橙标记软骨细胞形态及DNA的表达。在营养缺乏培养状态中(5%,2.5%新生牛血清),辐照时间为16,30,45min的照射组细胞数量显著增加,具有明显的光生物调节作用(PBM);其中最佳辐照时间为30min,实际最佳辐照能量密度为9.42J/cm2;当光生物调节作用显著时,软骨细胞的DNA及胶原的合成也明显增强。He-Ne激光照射兔软骨细胞后在营养缺乏的媒介中增殖明显,所产生的光生物调节作用仅限在一定的剂量范围内产生,这一特性可为临床使用提供参考。     

Appearance of the reticular cells which trap antigens in the rat lymph node in postnatal development.

The cells binding and retaining immune complexes on their cell surface existed in rat lymph nodes with no germinal centers. This study attempted to clarify the relationship between the two types of cells, reticular cells and follicular dendritic cells (FDCs), in the rat lymph node at early stages of postnatal development by immuno-electron microscopy on anti-horseradish peroxidase (HRP) and HRP injected rat. On the 19th and 23rd day after birth, germinal centers were not yet constructed nor were typical FDCs visible. However, immune complex binding cells were observed on the 23rd day, and not on the 19th. HRP reactive materials (immune complexes) were localized between lymphocytes and large lucent cells, making meshworks. They were revealed by electron microscopy on the cell surface which invaginated into the cytoplasm. The HRP reactive cells extended their cytoplasmic processes and formed a connection by their processes. They were reticular cells which enclosed reticular fibers by their cytoplasmic processes or contacted with reticular fibers closely. The reticular cells may be precursors of the FDCs.     

准分子激光上皮下角膜磨镶术(LASEK)与准分子激光角膜切削术(PRK)术后兔角膜超微结构实验研究

目的:比较准分子激光上皮下角膜磨镶术(LASEK)与准分子激光角膜切削术(PRK)术后兔角膜创面愈合的情况,从组织学超微结构角度探讨角膜雾状混浊(Haze)及屈光度数回退的可能成因。方法:5 2只新西兰白兔分为8组,每只兔右眼行LASEK ,左眼行PRK。术后1天、7天及1、3、4、5、6月处死动物取角膜行透射电镜和扫描电镜观察,比较术后每一时相点角膜上皮及角膜基质愈合的动态变化过程。结果:LASEK术后角膜上皮细胞、基底膜、基质内胶原纤维和成纤维细胞的超微结构改变程度轻于PRK组。结论:术后LASEK组角膜创伤修复反应弱于PRK ,这可能是LASEK术后角膜恢复正常较早的原因。     

The structure of HeLa cell population after exposure to methylnitrosourea at different phases of culture growth

To study the methylnitrosourea (MNU) effect on the HeLa cells culture at different stages of its growth the method of subcultivation, i. e. dissemination of cells immediately after the MNU effect, has been used followed by the study of the growth pattern of the reinoculated culture. When dissemination was carried out on the 4th, 7th and 10th day of the culture growth after the action of MNU, the highest growth inhibition effect was observed at the late stationary growth stage of the culture (the 10th day). The study of the population structure of such a culture by the method of batch cytofluorimetry has shown that MNU exerts a cytotoxic effect on cells: there is a shift in cell distribution according to the DNA content towards 4c typical of the G2/M-period of the cell cycle.     

Development of cellular polarity and tight junctions in parotid acinar cells of postnatal rat

The morphogenesis of acino-tubular structures and cytodifferentiation of acinar cells in developing rat parotid glands from the day of birth to the 7th day after birth were studied by conventional ultrathin-section electron microscopy in conjunction with freeze replica and space tracer methods. An ultrathin-section study indicated that the acinar cells developed sequentially in the order of the following three stages: (1) the stage of undifferentiated cell immediately after birth, in which the presumptive acinar cells showed very scanty cell cytoplasm, poorly developed organellae, and no distinctive cellular polarity; (2) the stage about the 3rd day after birth, in which cells were arranged into a single layer, resulting in the establishment of three recognizable domains in the plasma membranes, and developing cellular organellae started to distribute with distinctive polarity; and (3) the stage of the 5th day after birth and thereafter, in which secretory granules were formed, indicating the beginning of exocrine functions. Freeze replica and a tracer study demonstrated that the formation of a sealing strand of tight junctional belt took place in correspondence to the establishment of cellular polarity. These results indicated that the development of cellular polarity, plasma membrane domains, tight junctions, and acino-tubular structure were closely interrelated to each other, and preceded the onset of secretory functions.     

Morphological changes of capillaries in the rat soleus muscle following experimental tenotomy

We examined the structural changes of capillaries in the rat soleus muscle 4, 7, 14, and 35 days after experimental limb tenotomy. In the soleus muscles after tenotomy, muscle fibres degenerated and some of them were destroyed; the muscle did not seem to recover until the 35th day. On the 14th day, some small muscle fibres, probably regenerating muscle fibres, started forming within the basal-lamina tube and remained after necrosis of a pre-existing muscle fibre. Most capillaries at each stage were of the continuous type. However, about 10% of the capillaries around degenerated muscle fibres at days 4, 7 and 14 consisted of endothelial cells with a small number of fenestrae bridged by a single-layered diaphragm. On the 14th day, capillaries around small regenerating muscle fibres also often had a small number of fenestrations. Even on the 35th day, capillaries occasionally had fenestrations. Additionally, some of the fenestrated capillaries formed small pores at the fenestrated portion of the endothelial cells. The untreated muscles contained only continuous capillaries. These findings suggest that fenestrations in the endothelial cells may occur in intramuscular capillaries not only around degenerated muscle fibres but also around regenerating muscle fibres after tenotomy.