一氧化氮和一氧化氮合酶基因多态性与糖尿病视网膜病变

糖尿病视网膜病变(diabetic retinopathy,DR)是重要致盲性眼病之一,其发生和进展与血糖水平、疾病病程、环境及遗传等多种因素有关。一氧化氮(NO)作为一种重要的血管活性物质,与DR中毛细血管闭塞、微循环障碍、内皮细胞增殖相关。一氧化氮合成酶(NOS)是NO合成过程中的关键酶,我们就NO和NOS基因多态性与DR的关系进行综述。     

糖尿病早期NOS亚型的蛋白表达及其在血流变化中作用的实验研究

目的探讨在糖尿病早期,3种亚型一氧化氮合酶(NOS)在大鼠视网膜不同组织细胞中蛋白表达的变化,及其与视网膜血流变化的关系。方法将Wistar大鼠随机分成正常对照组与链脲佐菌素(STZ)腹腔注射诱导的糖尿病组各10只,采用彩色多谱勒对大鼠视网膜中央动脉(CRA)进行血流参数的测量,运用免疫组织化学技术观察视网膜eNOS、iNOS、nNOS蛋白表达的变化。结果8wk糖尿病大鼠视网膜尚未出现病变,但血流参数已显著异常,表现为糖尿病组大鼠CRA的血流速度较对照组显著降低(P<0.05),而阻力指数则显著增高(P<0.05),搏动指数高于对照组,但尚未有显著意义。3种亚型NOS在糖尿病与正常对照组视网膜中表达部位一致;但与正常对照组比较,iNOS免疫反应的强度在糖尿病大鼠视网膜中的内核层显著增强,而eNOS与nNOS的免疫反应强度则未见显著变化。结论在糖尿病早期,大鼠视网膜已开始出现血流灌注不良,此时NOS亚型中iNOS在视网膜中的表达上调可能与之相关。     

脂联素对小鼠氧诱导视网膜病变的保护作用

目的：观察脂联素对C57BL/6J小鼠氧诱导视网膜病变的影响。

方法：将新生C57BL/6J小鼠随机分为3组,分别为常氧对照组、生理盐水注射氧诱导视网膜病变(oxygen-induced retinopathy,OIR)组和脂联素注射OIR组。将后两组小鼠于生后第7d(P7)至P12置于体积分数75%±2%高氧氧箱中以诱导OIR模型。脂联素注射OIR模型组在P7～P15每天接受腹腔注射重组鼠脂联素蛋白(3.0μg/g),生理盐水注射OIR组则于上述相同时间点注射同等剂量的生理盐水。三组小鼠均于P17时取右眼行视网膜铺片和Lectin染色,观察视网膜中央无血管区及病理性新生血管的情况; 取左眼行视网膜切片和HE染色,观察视网膜组织病理学变化; 应用Western-blot测定左眼视网膜组织中iNOS、nNOS、eNOS的表达; 取右眼视网膜组织定量检测ROS/RNS含量以及NO水平。

结果：脂联素注射OIR组小鼠视网膜中央无血管区面积较生理盐水注射OIR组明显减少(t=7.304,P<0.01),病理性新生血管数目明显减少(t=2.654,P<0.01); 脂联素注射OIR组小鼠视网膜组织ROS含量较生理盐水注射组明显降低(t=13.349,P<0.01); 脂联素注射OIR组小鼠视网膜组织NO含量明显高于生理盐水注射组(t=3.023,P<0.01),iNOS表达明显低于生理盐水注射组(t=5.112,P<0.01),eNOS表达明显高于生理盐水注射组(t=7.421,P<0.01),nNOS的表达无统计学差异(t=1.074,P>0.01)。

结论：脂联素可以通过激活内源性eNOS促进生理性NO生成,同时又能抑制ROS/RNS的产生,在OIR进程中发挥视网膜血管的保护作用。     

巨细胞病毒与2型糖尿病视网膜病变关系的临床研究

目的:探讨巨细胞病毒(cytomegalovirus,CMV)感染与2型糖尿病视网膜病变(diabetic retinopathy,DR)的关系。方法:采用聚合酶链反应(chain reaction technique,PCR)技术分别检测T2DM患者86例(单纯T2DM组40例、视网膜病变组46例)以及正常对照组30例外周血中的CMV-DNA,测定血浆内皮素(endothelin,ET)和一氧化氮(nitricox-ide,NO)。结果:T2DM视网膜病变组CMV-DNA检测阳性率(65.2%)明显高于单纯T2DM组(35.0%)和对照组(13.3%,P<0.01),T2DM视网膜病变组ET、NO含量与单纯T2DM组和对照组比较也有显著差异。结论:CMV感染与糖尿病视网膜病变的发生发展有关,CMV感染可能加重糖尿病视网膜病变。     

川芎嗪联合氨基胍对糖尿病大鼠视网膜保护作用的机制

目的　探讨川芎嗪联合氨基胍对糖尿病视网膜病变防治作用及其机制。方法　用链尿佐菌素 (STZ)制作糖尿病大鼠动物模型 ,分为正常对照组、糖尿病对照组、糖尿病氨基胍治疗组、糖尿病川芎嗪 +氨基胍治疗组 ,于第 12周测定大鼠视网膜组织NO的含量、NOS活性和AR的活性。结果　糖尿病川芎嗪 +氨基胍治疗组、糖尿病氨基胍治疗组大鼠视网膜组织NOS及AR活性显著低于糖尿病对照组 (P <0 0 1) ,NO的含量升高 (P <0 0 1) ;糖尿病氨基胍治疗组仍高于正常对照组 (P <0 0 1) ,NO的含量降低 (P <0 0 1) ;糖尿病川芎嗪 +氨基胍治疗组与正常组无显著差异。结论　川芎嗪联合氨基胍能够抑制糖尿病大鼠视网膜组织NOS、AR活性 ,从而对糖尿病视网膜病变起一定保护作用     

链脲佐菌素诱导的糖尿病大鼠早期视网膜NO的变化

目的利用链脲佐菌素(STZ)诱导的1型糖尿病大鼠模型,研究糖尿病早期视网膜一氧化氮(NO)浓度的变化。方法健康成年雄性SD大鼠腹腔注射STZ60mg/kg,7d后血糖>13.9mmol/L入选为糖尿病组,与正常对照组在无特定病原体(SPF)环境下饲养,每周监测体重与血糖。8周后氯胺酮全身麻醉下摘取眼球,分离视网膜,制成组织匀浆。应用硝酸还原酶法测定组织NO浓度。结果糖尿病组与对照组在8周末时的平均体重差异有极显著统计学意义(t=11,P<0.01)。两组在各观察时段的平均血糖水平差异均有显著统计学意义(P<0.05)。两组视网膜样品的蛋白浓度差异无显著统计学意义(t=0.765,P>0.05)。糖尿病组视网膜平均NO浓度为(0.352±0.256)μmol/gprot,对照组为(0.031±0.017)μmol/gprot,两组间差异有显著统计学意义(t=2.79,P<0.05)。结论STZ诱导的糖尿病大鼠早期视网膜NO浓度增加,是引起糖尿病早期视网膜血管功能性改变的重要因素。     

糖尿病大鼠视网膜组织中iNOS的表达及细胞凋亡的研究

目的:探讨诱导型一氧化氮合酶(iNOS)在糖尿病大鼠视网膜组织中的表达及其对细胞凋亡的影响。方法:建立糖尿病大鼠动物模型,分别在成模后第1,3,6mo时测定大鼠视网膜组织细胞凋亡指数(AI)和iNOS表达水平。结果:(1)与正常对照组(NC)相比,iNOS表达在糖尿病各组均升高,6mo时明显增强(P<0.05);(2)视网膜细胞凋亡情况:与正常对照组(NC)相比,糖尿病模型1mo时细胞凋亡指数差异无统计学意义,3,6mo时则明显升高(P<0.05)。结论:iNOS在糖尿病大鼠视网膜组织中的表达增强,推断iNOS在糖尿病视网膜病变(retinopathy of diabetes,DR)的发病机制中起一定作用。     

糖尿病视网膜病变患者高密度脂蛋白-3和总抗氧化能力以及NOx水平分析

目的:比较糖尿病视网膜病变组与对照组数据,以研究高密度脂蛋白-3(HDL3)、NOx和总抗氧化状态之间的关系.方法:前瞻性病例对照研究.106例受试者分为3组,84例2型糖尿病有或无视网膜病变患者,对照组为22例正常人.患者均行血清高密度脂蛋白-3的浓度检测和血清NOx水平测定.用铁还原法(FRAP)测量血浆总抗氧化能力.结果:糖尿病患者(DM)空腹血糖、糖化血红蛋白、甘油三酯显著高于对照组.正常人中HDL3水平为14.4(12.0)mg/dL,糖尿病视网膜病变患者为18.1(12.6)mg/dL,糖尿病无视网膜病变患者为14.0(12.5)mg/dL,组间差异无统计学意义(P=0.262).糖尿病患者FRAP水平低于对照组(P=0.003),但糖尿病视网膜病变组与非糖尿病视网膜病变组之间其差异无统计学意义.结论:研究发现:2型糖尿病视网膜病变患者、糖尿病无视网膜病变患者以及对照组间HDL和HDL3水平无明显不同.HDL3可能无法预测糖尿病视网膜病变患者的患病风险.糖尿病患者中血清NOx水平较高,FRAP水平较低.     

Relation Between Plasma Nitric Oxide Levels and Diabetic Retinopathy

Purpose Nitric oxide (NO) plays an important role in homeostatic vasodilation and the regulation of blood flow. On the other hand, excess release of NO causes various vascular complications. There are only a few reports on the relationship between plasma NO levels and microvascular complications, especially diabetic retinopathy (DR) in patients with type 2 diabetes. The purpose of this study was to determine the relationship between plasma NO levels and DR. Methods In a prospective study, blood samples were obtained from 36 patients with diabetes and no diabetic retinopathy (NDR), 43 patients with nonproliferative diabetic retinopathy (NPDR), 18 patients with proliferative diabetic retinopathy (PDR), and 40 subjects without diabetes mellitus, who served as controls. The levels of plasma NO_x (nitrite and nitrate), the stable metabolites of NO, were measured by high-performance liquid chromatography with the Griess method. Results The plasma NO_x levels were 92.8 ± 16.0, 70.2 ± 6.8, 90.3 ± 9.1, and 53.8 ± 6.1 μmol/l in patients with NDR, NPDR, or PDR, and in the controls, respectively. The plasma NO_x levels in the three diabetic groups were significantly higher than those in the control group (P < 0.05 in each case). Conclusion The increased plasma NO levels in patients with type 2 diabetes indicate that NO may be associated with the pathogenesis of DR. Jpn J Ophthalmol 2006;50:465–468 ? Japanese Ophthalmological Society 2006     

Effects of aminoguanidine on retinal apoptosis in mice with oxygen-induced retinopathy

AIM: To explore the protective effects of aminoguanidine (AG) on retinal apoptosis in mice with oxygen-induced retinopathy (OIR).METHODS:A total of 80 C57BL/6J mice, aged 7 days, were randomly divided into four groups:normal, high oxygen, high oxygen saline and high oxygen treated with AG. In the normal group, mice were housed in normoxic conditions from postnatal day P7 to P17. Mice in the other 3 groups were placed under hyperoxic conditions (75±2%O2) in an oxygen-regulated chamber for 5 days and subsequently placed in normoxic conditions for 5 days. Mice in the AG group were treated once daily, from P12 to P17, with AG hemisulfate (100mg/kg body weight, intraperitoneally) dissolved in physiological saline. An equivalent amount of 0.9% physiological saline was administered, as above, to mice in the high oxygen saline group. Ten mice were randomly selected from each group on P14 and on P17, euthanized and the retinas examined. Apoptotic cells in the retina were detected using the terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) method. The expression of nitric oxide synthase (iNOS) in the retina was detected by immunohistochemistry and changes in rod cells were observed using electron microscopy.RESULTS:TUNEL-positive cells and iNOS immunoreactive neurons were present in the inner nuclear and ganglion cell retinal layers of mice in the high oxygen group. The number of TUNEL-positive cells was significantly greater in the high oxygen group compared with the normal group (t=-20.81, P14d <0.05; t=-15.05, P17d<0.05). However, the number of TUNEL-positive cells in the AG treatment group was significantly lower (t=-13.21, P14d<0.05; t=-6.61,P17d <0.05) compared with the high oxygen group. The expression of iNOS was significantly higher in the high oxygen group compared with the normal group (t=-21.95, P14d<0.05; t=-17.30, P17d<0.05). However, the expression of iNOS in the AG treatment group was significantly lower (t=-12.17,P14d<0.05; t=-10.30,P17d<0.05) compared with the high oxygen group. The outer segments of the rods were disorganized and short in the high oxygen group. Rod morphology appeared to be slightly improved in the AG group.CONCLUSION:AG may protect retinal neurons in OIR by inhibiting apoptosis. The mechanism may be related to iNOS.