难治性青光眼的Ahmed青光眼阀植入治疗：并发症与生物相容性

背景：近年来房水引流物的出现在治疗难治性青光眼上已经取得了较好的效果,但植入后会出现一些与材料有关的并发症,如炎症反应导致房水生成减少等原因引起低眼压及浅前房等。 目的：观察Ahmed青光眼阀植入治疗难治性青光眼的效果及其生物相容性。 方法：对32例(32眼)难治性青光眼实施Ahmed阀植入治疗,其中包括新生血管性青光眼12例,无晶状体眼青光眼6例,葡萄膜炎性青光眼4例,外伤性青光眼4例,有滤过手术失败史的青光眼6例,术后随访6~24个月。 结果与结论：术前眼压4.655~9.044 kPa,平均(5.61±1.29) kPa,末次随访时平均眼压(2.85±1.16) kPa,总有效率84.4%,其中新生血管性青光眼有效率83.3%,其他类型有效率85%。术后10眼视力较前有不同程度提高,最好视力达0.3。出现短暂性低眼压6例,浅前房6例,前房积血4例,内流管堵塞3例,经对症治疗后好转。结果表明Ahmed青光眼阀植入后降低眼压效果较好,但出现并发症比率较大,可能为引流装置设计和材料上的不足引起。 关键词：青光眼;Ahmed青光眼阀;植入术;眼内压;视力 doi:10.3969/j.issn.1673-8225.2010.29.020     

组织工程角膜内皮载体材料的研究进展★

学术背景：前房注射法移植角膜内皮细胞遭到失败，而采用载体法移植角膜内皮获得成功，提示了载体在组织工程角膜内皮体外构建与移植中的重要性。以各种材料为载体进行角膜内皮细胞体外培养与移植的实验研究广泛开展，获得了初步成功。但材料的免疫相容性、降解/溶解特性、亲细胞性、术后排斥反应、移植疾病传播等难题限制了角膜内皮体外构建与移植的发展，至今无法应用于临床实践。所以，寻找一种理想的载体材料一直是组织工程角膜内皮研究领域的重要课题和研究热点。 目的：分析组织工程角膜内皮各种载体材料的优缺点，探索该领域还存在的难题和进一步发展方向。 检索策略：应用计算机检索MEDLINE数据库和中国期刊全文数据库1973-01/2007-12的有关文献，检索词为“角膜内皮，组织工程，载体，材料，角膜移植，角膜构建”，限定文章语言分别为英语和中文。共检索到986篇相关文献，其中中文文献645篇，英文文献341篇。根据本文文题和研究目的对文献进行筛选。纳入标准：①选取针对性强，相关度高的文献；②对同一领域的文献选择近期发表或权威杂志的文献；排除重复研究和Meta分析类文章。最后35篇论文被选用。 文献评价：选用35篇文献，其中3篇为综述，其余均为临床与实验研究。 资料综合：早期开始使用的材料主要有：异体角膜片、明胶薄膜、胶原凝胶、聚羟基乙酸和聚乳酸羟基乙酸、水凝胶膜等，而角膜基质/后弹力膜、羊膜基底膜、角膜基质片、复合生物膜是目前的研究热点。组织工程角膜内皮材料投入临床实践目前还存在许多难题。 结论：组织工程角膜内皮载体材料各有优缺点，改善材料的亲细胞性、免疫原性和降解/溶解特性是今后的发展方向，并且应该加强对材料的人体效应和远期影响的研究。     

碱性成纤维细胞生长因子促进猫角膜内皮细胞的增殖作用☆

背景：有实验报道,碱性成纤维细胞生长因子能改变角膜内皮细胞形状、促进其分裂和趋化,刺激活体和体外角膜内皮细胞的损伤修复。 目的：观察碱性成纤维细胞生长因子对猫角膜内皮细胞增殖的影响。 设计、时间及地点：细胞形态学观察实验,2005-01/2006-12在青岛大学医学院附属医院中心实验室完成。 材料：家猫购买自青岛大学医学院附属医院动物实验中心。 方法：猫角膜内皮细胞原代培养,行NSE免疫组织化学染色鉴定细胞的类型、纯度及生理特性。传1代后,分别在培养液中加1,10,100 μL的碱性成纤维细胞生长因子,继续卵化1~5 d。 主要观察指标：加药后第1,3,5天分别利用MTT法测定在490 nm处的吸光度值来判断角膜内皮细胞增殖情况;同时应用倒置相差显微镜观察细胞形态及透射电镜检测细胞超微结构。 结果：碱性成纤维细胞生长因子作用组加药后1,3,5 d,MTT法测定猫角膜内皮细胞在490 nm处的吸光度值明显高于对照组,其中10 μg/L作用组差别最显著。 结论：碱性成纤维细胞生长因子作用能促进猫角膜内皮细胞的增殖,相对有效浓度为10 μg/L。     

组织工程角膜内皮载体材料的研究现状

组织工程角膜内皮载体材料的研究历程，总体上经历了从天然材料到人造材料，然后再回到天然材料这样一个循环，取得了宝贵的正反两方面经验。早期开始使用的材料主要有：异体角膜片、明胶薄膜、胶原凝胶、聚羟基乙酸和聚乳酸羟基乙酸、水凝胶膜等，而角膜基质/后弹力膜、羊膜基底膜、角膜基质片、复合生物膜是目前的研究热点。改善材料的亲细胞性、免疫原性和降解/溶解特性是今后的发展方向，并且应该加强对材料的人体效应和远期影响的研究。     

角膜内不同部位植入壳聚糖-胶原复合膜的生物相容性

背景：角膜基质层间植入是评价材料角膜内生物相容性的理想模型,但将材料植入角膜基质层间的位置不同,可能会影响材料的降解以及角膜的代谢,最终影响到材料在角膜内生物相容性评价结果。 目的：观察壳聚糖胶原复合膜(胶原含量为30%)植入兔角膜不同部位基质层的组织相容性。 方法：制备厚度为20 μm的壳聚糖胶原复合膜。将新西兰大白兔以抽签法随机分为两组：一组角膜周边基质层间植入壳聚糖胶原复合膜材料;另一组角膜中央基质层间植入壳聚糖胶原复合膜材料。裂隙灯显微镜观察材料植入后眼表及前房的反应、材料植入部位角膜和材料的相互反应及植入部位角膜透明性改变。6周后取角膜做组织切片,观察植入部位角膜组织和材料的组织学改变。 结果与结论：角膜周边植入组的复合膜逐渐降解,最终完全降解,并对角膜结构无明显影响,显示了良好的角膜组织相容性;中央植入组复合膜植入早期眼表稳定,也显示了良好的生物相容性,但是2周过后伴随着材料的缓慢降解,材料周边变得不透明,伴随着炎症细胞浸润,最终在第34,42天分别有一只发生角膜溶解。结果说明薄的壳聚糖胶原复合膜材料透明性好、力学性能匹配,植入角膜周边基质层间后在短时间内能完全降解,适宜作为角膜修复材料,特别是作为角膜缘干细胞的载体可以用来修复角膜缘干细胞损伤引起的角膜疾病。     

软性角膜接触镜矫正近视低阶像差后高阶像差的变化

背景：低阶像差矫正之后,高阶像差成为影响视觉质量的主要方面。作为一种屈光不正的矫治方法,角膜接触镜矫正了低阶像差,而高阶像差随之的变化值得观察探讨。目的: 分析佩戴软性角膜接触镜矫正青年单纯近视眼低阶离焦像差对高阶像差的影响。方法: 使用依据Hartmann-Shack原理设计的COAS波前像差仪对47只单纯性近视眼分别测量角膜接触镜矫正低阶离焦像差前后的高阶像差,提取瞳孔直径为6 mm时高阶像差的均方根值进行分析。比较矫正低阶离焦像差前后3阶至6阶各项高阶像差以及屈光度的变化值、二阶离焦像差的变化值与球差C40Zenike系数变化值的关系。结果与结论: 矫正后较矫正前,垂直慧差C3-1正向变化趋势(t =3.843,P=0.000);球差C40负向变化趋势(t=–8.319,P=0.000);3阶到6阶总高阶像差均方根值增加(t =2.125,P=0.039),球差C40的变化量与屈光度变化值呈正相关(r=0.536,P=0.000)。结果提示矫正低阶像差影响高阶像差改变。佩戴软性角膜接触镜后眼总高阶像差增加,矫正低阶近视离焦像差后球差C40减小。关键词：近视;高阶像差;软性角膜接触镜;屈光;矫正     

准分子激光角膜上皮瓣下磨镶术中角膜上皮瓣的活性☆

背景：保证角膜上皮瓣活性的最佳乙醇浸润时间是准分子激光角膜上皮瓣下磨镶术研究的热点。目的：观察准分子激光角膜上皮瓣下磨镶术后影响角膜上皮瓣活性的相关因素。方法：应用前瞻性研究方法选取50例(100眼)行准分子激光角膜上皮瓣下磨镶术患者,术眼按度数分为轻度近视(<-3.00 m-1) 40眼和中度近视(-3.00~-6.00 m-1)60眼。术中乙醇作用时间分别为15,25 s。结果与结论：术后1周、1,6,12个月时,相同近视度数下,与乙醇作用25 s比较,术中乙醇作用15 s眼的裸眼视力明显提高,角膜上皮愈合时间加快(P < 0.01),角膜荧光素染色评分降低,雾状角膜混浊程度降低(P < 0.05或P < 0.01)。相关性分析显示,角膜荧光素染色评分与雾状角膜混浊程度呈正相关(P < 0.01)。说明准分子激光角膜上皮瓣下磨镶术中切削深度相当情况下,乙醇浸润时间越短,角膜上皮瓣制作质量越高,术后角膜上皮瓣活性越好,愈合较快、视力易恢复。     

帕金森病患者角膜神经的临床分析

目的 应用角膜共聚焦显微镜(CCM)探讨帕金森病(PD)患者的角膜神经纤维变化特点及角膜神经与运动症状侧别、运动亚型的关系。方法 选取2022年1月至2022年12月就诊于徐州医科大学附属医院的PD患者50例作为PD组,同期来院体检的健康老年人56例作为对照组。两组受检者均行CCM检查,比较PD组与对照组间的角膜神经纤维参数差异,比较角膜神经在不同运动亚型和躯体运动症状严重侧别的差异。结果 PD组的角膜神经纤维长度(CNFL)、角膜神经纤维密度(CNFD)及角膜神经分形维数(CNFrD)较对照组均减少(P<0.05)。两组间角膜神经分支密度(CNBD)比较差异无统计学意义(P>0.05)。PD患者的运动症状较重侧眼的CNFD、CNFL、CNBD及CNFrD较症状轻侧眼比较差异无统计学意义(P>0.05)。不同运动亚型间CNFD、CNFL、CNBD及CNFrD比较差异无统计学意义(P>0.05)。结论 PD患者的角膜神经纤维存在损伤,双眼对称性发生,并且这种损伤与运动亚型无关。     

碱性成纤维细胞生长因子联合透明质酸钠对角膜保存液中内皮细胞的作用*

背景：作者前期将透明质酸钠和碱性成纤维细胞生长因子分别加入基础培养基,制成改良角膜保存液并证实其对角膜内皮细胞活性的保存作用。目的：探讨碱性成纤维细胞生长因子及透明质酸钠在角膜中期保存液中的作用及其有效浓度,评价该两种物质联合对角膜内皮细胞的保护作用。方法：在以基础培养基 MEM、M-199为主要成分,添加 6%葡聚糖、HEPES缓冲系、抗生素等制成角膜保存液的基础上,加入不同浓度的透明质酸钠和碱性成纤维细胞生长因子,并设立未加透明质酸钠及碱性成纤维细胞生长因子的对照组。各组于4 ℃下保存新西兰大白兔角膜片,于3,5,7,10,14 d,行裂隙灯检查、锥虫蓝-茜素红染色、角膜组织病理切片及扫描电镜观察检测角膜内皮细胞活性,评价保存质量。 结果与结论：加入碱性成纤维细胞生长因子及透明质酸钠的中期角膜保存液保存的角膜片其透明度、内皮细胞活性、显微结构、超微结构均明显优于未加组,各实验组同期比较,浓度较高组效果优于浓度较低组。碱性成纤维细胞生长因子及透明质酸钠同时加入基础保存液中能够对角膜内皮细胞进行有效的保护,其效果与浓度有关。     

大鼠外周血单个核细胞体外向内皮细胞诱导分化时的表型变化

背景：内皮祖细胞在贴壁分化过程中其干细胞标记以及内皮细胞表型的变化过程将有助于了解内皮祖细胞的分化特性。但目前还未找到区别于成熟内皮细胞特异性的细胞标记。 目的：观察大鼠外周血单个核细胞外分化为内皮细胞的过程中干细胞标志物以及内皮细胞表型的变化。 设计、时间及地点：细胞观察实验,于2004-06/2008-12在青岛市市立医院心脏外科实验室完成。 材料：抽取雄性SD大鼠外周血,应用Ficoll密度梯度离心的方法获得单个核细胞。 方法：单个核细胞体外培养于纤连蛋白处理的培养皿中,同时应用血管内皮生长因子和碱性成纤维细胞生长因子诱导,使之向内皮细胞转化。 主要观察指标：以免疫组织化学方法检测细胞CD31、CD34、Flk-1、vWF在第1~7天的表达。 结果：贴壁细胞的CD31、CD34、Flk-1、vWF在不同时段呈不同程度的表达。干细胞标志物CD31、CD34在培养的第2天开始表达,第4天最强,以后逐渐减弱,第6天消失。而Flk-1在第3天开始表达,以后逐渐增强,第7天时最强。vWF表达是逐步出现的,第7天时表达为100%。 结论：大鼠外周血干细胞向内皮祖细胞分化的过程中,干细胞标志物逐渐消失,内皮细胞的表型逐步出现。     

Effect of glycemic control on corneal nerves and peripheral neuropathy in streptozotocin‐induced diabetic C57Bl/6J mice

We sought to determine the impact that duration of hyperglycemia and control has on corneal nerve fiber density in relation to standard diabetic neuropathy endpoints. Control and streptozotocin‐diabetic C57Bl/6J mice were analyzed after 4, 8, 12, and 20 weeks. For the 20‐week time point, five groups of mice were compared: control, untreated diabetic, and diabetic treated with insulin designated as having either poor glycemic control, good glycemic control, or poor glycemic control switched to good glycemic control. Hyperglycemia was regulated by use of insulin‐releasing pellets. Loss of corneal nerves in the sub‐epithelial nerve plexus or corneal epithelium progressed slowly in diabetic mice requiring 20 weeks to reach statistical significance. In comparison, slowing of motor and sensory nerve conduction velocity developed rapidly with significant difference compared with control mice observed after 4 and 8 weeks of hyperglycemia, respectively. In diabetic mice with good glycemic control, average blood glucose levels over the 20‐week experimental period were lowered from 589 ± 2 to 251 ± 9 mg/dl. All diabetic neuropathy endpoints examined were improved in diabetic mice with good glycemic control compared with untreated diabetic mice. However, good control of blood glucose was not totally sufficient in preventing diabetic neuropathy.     

Reduced association between dendritic cells and corneal sub‐basal nerve fibers in patients with fibromyalgia syndrome

In our study, we aimed at investigating corneal langerhans cells (LC) in patients with fibromyalgia syndrome (FMS) and small fiber neuropathy (SFN) as potential contributors to corneal small fiber pathology. We enrolled women with FMS (n = 134) and SFN (n = 41) who underwent neurological examination, neurophysiology, prostaglandin analysis in tear fluid, and corneal confocal microscopy (CCM). Data were compared with those of 60 age‐matched female controls. After screening for dry eye disease, corneal LC were counted and sub‐classified as dendritic (dLC) and non‐dendritic (ndLC) cells with or without nerve fiber association. We further analyzed corneal nerve fiber density (CNFD), length (CNFL), and branch density (CNBD). Neurological examination indicated deficits of small fiber function in patients with SFN. Nerve conduction studies were normal in all participants. Dry eye disease was more prevalent in FMS (17%) and SFN (28%) patients than in controls (5%). Tear fluid prostaglandin levels did not differ between FMS patients and controls. While corneal LC density in FMS and SFN patients was not different from controls, there were fewer dLC in association with nerve fibers in FMS and SFN patients than in controls (P < .01 each). Compared to controls, CNFL was lower in FMS and SFN patients (P < .05 each), CNFD was lower only in FMS patients (P < .05), and CNBD was lower only in SFN patients (P < .001). There was no difference in any CCM parameter between patients with and without dry eyes. Our data indicate changes in corneal innervation and LC distribution in FMS and SFN, potentially based on altered LC signaling.     

Functional deterioration of endothelial nitric oxide synthase after focal cerebral ischemia

Endothelial nitric oxide synthase (eNOS) dysfunction is related to secondary injury and lesion expansion after cerebral ischemia. To date, there are few reports about postischemic alterations in the eNOS regulatory system. The purpose of the present study was to clarify eNOS expression, Ser1177 phosphorylation, and monomer formation after cerebral ischemia. Male Wistar rats were subjected to transient focal cerebral ischemia. Endothelial nitric oxide synthase messenger RNA (mRNA) and protein expression increased ∼8-fold in the ischemic lesion. In the middle cerebral artery core, eNOS-Ser1177 phosphorylation increased 6 hours after ischemia; however, there was an approximately 90% decrease in eNOS-Ser1177 phosphorylation observed 24 hours after ischemia that continued until at least 7 days after ischemia. Endothelial nitric oxide synthase monomer formation also increased 24 and 48 hours after ischemia (P<0.05), and protein nitration progressed in parallel with monomerization. To assess the effect of a neuroprotective agent on eNOS dysfunction, we evaluated the effect of fasudil, a Rho-kinase inhibitor, on eNOS phosphorylation and dimerization. Postischemic treatment with fasudil suppressed lesion expansion and dephosphorylation and monomer formation of eNOS. In conclusion, functional deterioration of eNOS progressed after cerebral ischemia. Rho-kinase inhibitors can reduce ischemic lesion expansion as well as eNOS dysfunction in the ischemic brain.     

螺内酯对碱烧伤诱导大鼠角膜新生血管形成的影响及其机制研究

背景：螺内酯为醛固酮受体拮抗剂,近来有实验证实螺内酯在体内外能够有效地抑制新生血管的形成。 目的：验证螺内酯对碱烧伤诱导大鼠角膜新生血管的抑制作用。 方法：取SD大鼠36只,采用碱烧伤方法制备大鼠角膜新生血管模型,造模后以数字表法随机分成实验组和对照组。实验组术后灌胃给予螺内酯100 mg/kg,1次/d,对照组灌胃给予等量生理盐水。另取大鼠6只不作任何处理作为正常对照组。于造模后4,7,14 d运用裂隙灯观察各组大鼠角膜新生血管并计算面积,并每组随机处死6只大鼠,运用免疫组化染色方法和计算机图像分析系统检测观察血管内皮细胞生长因子及基质金属蛋白酶2的表达。 结果与结论：正常角膜组织未见炎症细胞与新生血管,角膜上皮及基质层仅见微弱血管内皮细胞生长因子表达,未见基质金属蛋白酶2表达。与对照组比较,实验组各个时期新生血管面积减小,血管内皮细胞生长因子和基质金属蛋白酶2表达降低(P < 0.05)。提示螺内酯可能通过参与下调血管内皮生长因子和基质金属蛋白酶2表达,从而有效地抑制角膜新生血管的形成。     

Effect of diet‐induced obesity or type 1 or type 2 diabetes on corneal nerves and peripheral neuropathy in C57Bl/6J mice

We determined the impact diet‐induced obesity (DIO) and types 1 and 2 diabetes have on peripheral neuropathy with emphasis on corneal nerve structural changes in C57Bl/6J mice. Endpoints examined included nerve conduction velocity, response to thermal and mechanical stimuli and innervation of the skin and cornea. DIO mice and to a greater extent type 2 diabetic mice were insulin resistant. DIO and both types 1 and 2 diabetic mice developed motor and sensory nerve conduction deficits. In the cornea of DIO and type 2 diabetic mice there was a decrease in sub‐epithelial corneal nerves, innervation of the corneal epithelium, and corneal sensitivity. Type 1 diabetic mice did not present with any significant changes in corneal nerve structure until after 20 weeks of hyperglycemia. DIO and type 2 diabetic mice developed corneal structural damage more rapidly than type 1 diabetic mice although hemoglobin A₁C values were significantly higher in type 1 diabetic mice. This suggests that DIO with or without hyperglycemia contributes to development and progression of peripheral neuropathy and nerve structural damage in the cornea.     

Chloroquine and primary amines inhibit the internalization of antithrombin III. Trypsin complex in cultured cells

Bovine corneal endothelial (BCE) cells have been shown to specifically bind, internalize and degrade antithrombin III (AT III) protease complexes as well as thrombin. Previous studies have indicated that chloroquine has no effect on the internalization of thrombin or other cell surface-bound ligands, but it inhibits their subsequent degradation. In contrast, the present study demonstrates the unique inhibitory effect of chloroquine on the internalization of ¹²⁵I-AT III. trypsin complex by BCE cultures. Similarly, the primary amines, monodansylcadaverine and methylamine, inhibit the internalization of ¹²⁵I-AT III . trypsin complex, but not the internalization of ¹²⁵I-thrombin. The various amines used in this study revealed:

(1) differences in the process of cellular binding and internalization between AT III . protease complex and thrombin, although the degradation of both internalized ligands proceed in an analogous manner; and (2) the unique sensitivity to chloroquine of ¹²⁵I-AT III . trypsin complex internalization by cultured cells. These results might indicate that AT III . protease complexes are internalized via a distinct receptor and/or a different mechanism from thrombin.     

The culture of vascular endothelial cells to confluence on microporous membranes

We have designed a two compartment system in which upper and lower compartments are separated by a layer of vascular endothelial cells grown on a porous membrane. The culture of pig aortic endothelial cells and their growth to confluence on porous PTFE membranes is described and the principal characteristics of membrane-cultured endothelium are compared with those of similar cells cultured on solid surfaces. While growth of endothelium to visual confluence was slower on the PTFE membranes than on solid surfaces, gross morphology and ratio of 6 keto prostaglandin F₁ to prostaglandin E₂ production by the cells was similar. A decrease in fluid flow across the membrane-cultured cells was associated with their growth to visual confluence: from low levels, fluid flow could be increased markedly by altering the environmental temperature of the cells. The possible uses and limitations of such a system in the interpretation of the role of endothelial cells in selective transport and compartmentation of fluid and cellular components of the blood are discussed.     

Pro-angiogenic effects of resveratrol in brain endothelial cells: nitric oxide-mediated regulation of vascular endothelial growth factor and metalloproteinases

Resveratrol may be a powerful way of protecting the brain against a wide variety of stress and injury. Recently, it has been proposed that resveratrol not only reduces brain injury but also promotes recovery after stroke. But the underlying mechanisms are unclear. Here, we tested the hypothesis that resveratrol promotes angiogenesis in cerebral endothelial cells and dissected the signaling pathways involved. Treatment of cerebral endothelial cells with resveratrol promoted proliferation, migration, and tube formation in Matrigel assays. Consistent with these pro-angiogenic responses, resveratrol altered endothelial morphology resulting in cytoskeletal rearrangements of β-catenin and VE-cadherin. These effects of resveratrol were accompanied by activation of phosphoinositide 3 kinase (PI3-K)/Akt and Mitogen-Activated Protein Kinase (MAPK)/ERK signaling pathways that led to endothelial nitric oxide synthase upregulation and increased nitric oxide (NO) levels. Subsequently, elevated NO signaling increased vascular endothelial growth factor and matrix metalloproteinase levels. Sequential blockade of these signaling steps prevented resveratrol-induced angiogenesis in cerebral endothelial cells. These findings provide a mechanistic basis for the potential use of resveratrol as a candidate therapy to promote angiogenesis and neurovascular recovery after stroke.