中华实验眼科杂志
中華實驗眼科雜誌
중화실험안과잡지
CHINESE JOURNAL OF EXPERIMENTAL OPHTHALMOLOGY
2014年
11期
1010-1013
,共4页
顾永昊%柯根杰%王林%何勤%周恩亮
顧永昊%柯根傑%王林%何勤%週恩亮
고영호%가근걸%왕림%하근%주은량
金属内肽酶类/拮抗剂和抑制剂%糖尿病/并发症%视网膜%血-视网膜屏障%渗透性%动物模型%SD大鼠
金屬內肽酶類/拮抗劑和抑製劑%糖尿病/併髮癥%視網膜%血-視網膜屏障%滲透性%動物模型%SD大鼠
금속내태매류/길항제화억제제%당뇨병/병발증%시망막%혈-시망막병장%삼투성%동물모형%SD대서
Metalloendopeptidase/antagonist and inhibitor%Diabetes/complications%Retina%Bloodretinal barrier%Permeability%Disease model,animal%Rat/Sprague-Dawley
背景 研究表明,多种基质金属蛋白酶(MMPs)家族成员在糖尿病视网膜病发(DR)的发病过程中发挥作用,但是否MMPs抑制剂能改善MMPs对血-视网膜屏障(BRB)的破坏作用尚不十分清楚.目的 探讨人工合成MMPs抑制剂GM6001对糖尿病大鼠BRB的影响.方法 24只成年清洁级SD大鼠按随机数字表法随机分为对照组、糖尿病组和糖尿病+GM6001组.采用链脲佐菌素诱导腹腔内注射法诱导大鼠糖尿病模型,对照组以相同的方法注射等体积枸橼酸盐缓冲液.造模成功后3d和14d,糖尿病+GM6001组大鼠玻璃体腔内注射10μl(100tμmol/L)GM6001各1次,对照组和糖尿病组大鼠以相同的方法注射等体积生理盐水.注射后1个月摘取大鼠一侧眼球,采用逆转录PCR(RT-PCR)法检测大鼠视网膜中MMP-2 mRNA和MMP-9 mRNA的相对表达量;用伊文思蓝(EB)灌注大鼠右侧颈静脉,120 min后以约120 mmHg(1 mmHg=0.133 kPa)的灌注压于大鼠左心室灌注枸橼酸钠缓冲液配制的质量分数1％多聚甲醛溶液,2 min后摘取大鼠另一侧眼球,观察大鼠视网膜EB的渗漏量.结果 RT-PCR检测显示MMP-2、MMP-9和GAPDH的反应条带分别位于436、536和484 bp.3个组大鼠视网膜中MMP-2 mRNA和MMP-9 mRNA相对表达量总体差异均有统计学意义(F=20.336,P=0.000;F=8.742,P=0.002);其中糖尿病组和糖尿病+GM6001组大鼠视网膜中MMP-2 mRNA和MMP-9 mRNA的相对表达量明显高于对照组大鼠,差异均有统计学意义(P＜0.01),而糖尿病+GM6001组大鼠视网膜中MMP-2 mRNA和MMP-9 mRNA的相对表达量明显低于糖尿病组大鼠,差异均有统计学意义(P=0.01、0.02).对照组、糖尿病组和糖尿病+GM6001组大鼠视网膜中标准化EB质量分数分别为(12.60±3.50)、(26.52±7.14)和(17.55±2.65) ng/mg,总体差异有统计学意义(F=17.032,P＜0.01),其中糖尿病组大鼠视网膜中标准化EB质量分数值明显高于对照组,而糖尿病+GM6001组较糖尿病组明显降低,差异均有统计学意义(P=0.003、0.020).结论 玻璃体腔内注射GM6001能下调大鼠视网膜中MMP-2和MMP-9的表达,从而对BRB发挥一定的保护作用.
揹景 研究錶明,多種基質金屬蛋白酶(MMPs)傢族成員在糖尿病視網膜病髮(DR)的髮病過程中髮揮作用,但是否MMPs抑製劑能改善MMPs對血-視網膜屏障(BRB)的破壞作用尚不十分清楚.目的 探討人工閤成MMPs抑製劑GM6001對糖尿病大鼠BRB的影響.方法 24隻成年清潔級SD大鼠按隨機數字錶法隨機分為對照組、糖尿病組和糖尿病+GM6001組.採用鏈脲佐菌素誘導腹腔內註射法誘導大鼠糖尿病模型,對照組以相同的方法註射等體積枸櫞痠鹽緩遲液.造模成功後3d和14d,糖尿病+GM6001組大鼠玻璃體腔內註射10μl(100tμmol/L)GM6001各1次,對照組和糖尿病組大鼠以相同的方法註射等體積生理鹽水.註射後1箇月摘取大鼠一側眼毬,採用逆轉錄PCR(RT-PCR)法檢測大鼠視網膜中MMP-2 mRNA和MMP-9 mRNA的相對錶達量;用伊文思藍(EB)灌註大鼠右側頸靜脈,120 min後以約120 mmHg(1 mmHg=0.133 kPa)的灌註壓于大鼠左心室灌註枸櫞痠鈉緩遲液配製的質量分數1％多聚甲醛溶液,2 min後摘取大鼠另一側眼毬,觀察大鼠視網膜EB的滲漏量.結果 RT-PCR檢測顯示MMP-2、MMP-9和GAPDH的反應條帶分彆位于436、536和484 bp.3箇組大鼠視網膜中MMP-2 mRNA和MMP-9 mRNA相對錶達量總體差異均有統計學意義(F=20.336,P=0.000;F=8.742,P=0.002);其中糖尿病組和糖尿病+GM6001組大鼠視網膜中MMP-2 mRNA和MMP-9 mRNA的相對錶達量明顯高于對照組大鼠,差異均有統計學意義(P＜0.01),而糖尿病+GM6001組大鼠視網膜中MMP-2 mRNA和MMP-9 mRNA的相對錶達量明顯低于糖尿病組大鼠,差異均有統計學意義(P=0.01、0.02).對照組、糖尿病組和糖尿病+GM6001組大鼠視網膜中標準化EB質量分數分彆為(12.60±3.50)、(26.52±7.14)和(17.55±2.65) ng/mg,總體差異有統計學意義(F=17.032,P＜0.01),其中糖尿病組大鼠視網膜中標準化EB質量分數值明顯高于對照組,而糖尿病+GM6001組較糖尿病組明顯降低,差異均有統計學意義(P=0.003、0.020).結論 玻璃體腔內註射GM6001能下調大鼠視網膜中MMP-2和MMP-9的錶達,從而對BRB髮揮一定的保護作用.
배경 연구표명,다충기질금속단백매(MMPs)가족성원재당뇨병시망막병발(DR)적발병과정중발휘작용,단시부MMPs억제제능개선MMPs대혈-시망막병장(BRB)적파배작용상불십분청초.목적 탐토인공합성MMPs억제제GM6001대당뇨병대서BRB적영향.방법 24지성년청길급SD대서안수궤수자표법수궤분위대조조、당뇨병조화당뇨병+GM6001조.채용련뇨좌균소유도복강내주사법유도대서당뇨병모형,대조조이상동적방법주사등체적구연산염완충액.조모성공후3d화14d,당뇨병+GM6001조대서파리체강내주사10μl(100tμmol/L)GM6001각1차,대조조화당뇨병조대서이상동적방법주사등체적생리염수.주사후1개월적취대서일측안구,채용역전록PCR(RT-PCR)법검측대서시망막중MMP-2 mRNA화MMP-9 mRNA적상대표체량;용이문사람(EB)관주대서우측경정맥,120 min후이약120 mmHg(1 mmHg=0.133 kPa)적관주압우대서좌심실관주구연산납완충액배제적질량분수1％다취갑철용액,2 min후적취대서령일측안구,관찰대서시망막EB적삼루량.결과 RT-PCR검측현시MMP-2、MMP-9화GAPDH적반응조대분별위우436、536화484 bp.3개조대서시망막중MMP-2 mRNA화MMP-9 mRNA상대표체량총체차이균유통계학의의(F=20.336,P=0.000;F=8.742,P=0.002);기중당뇨병조화당뇨병+GM6001조대서시망막중MMP-2 mRNA화MMP-9 mRNA적상대표체량명현고우대조조대서,차이균유통계학의의(P＜0.01),이당뇨병+GM6001조대서시망막중MMP-2 mRNA화MMP-9 mRNA적상대표체량명현저우당뇨병조대서,차이균유통계학의의(P=0.01、0.02).대조조、당뇨병조화당뇨병+GM6001조대서시망막중표준화EB질량분수분별위(12.60±3.50)、(26.52±7.14)화(17.55±2.65) ng/mg,총체차이유통계학의의(F=17.032,P＜0.01),기중당뇨병조대서시망막중표준화EB질량분수치명현고우대조조,이당뇨병+GM6001조교당뇨병조명현강저,차이균유통계학의의(P=0.003、0.020).결론 파리체강내주사GM6001능하조대서시망막중MMP-2화MMP-9적표체,종이대BRB발휘일정적보호작용.
Background Studies showed that some members of matrix metalloproteinases (MMPs) play an important role in the pathogenesis of diabetic retinopathy (DR).However,whether MMPs inhibitor can deter blood retinal barrier (BRB) from damage is below understood.Objective This study was to investigate the effects of GM6001,a MMPs inhibitor,on BRB permeability.Methods Twenty-four adult clean SD rats were randomized to the control group,diabetic group and diabetes+GM6001 group according to randomized number table.Diabetic models were induced by the intraperitoneal injection of streptozotocin in the rats of the diabetic group and the diabetes + GM6001 group,and equal volume of citrate buffer was used in the same way in the rats of the control group.GM6001 10 μ1 (100 μ mol/L) was intravitreously injected in the third and fourteenth day after modeling in the diabetes+ GM6001 group,and equal volume of normal saline solution was injected in the same way in the control group and the diabetic group.The rats were sacrificed and eyeballs were extracted 1 month after injection,and the relative expressions of MMP-2 mRNA and MMP-9 mRNA in rat retinas were detected by reverse transcription PCR (RT-PCR).Evens blue (EB) was infused via the right jugular vein,and paraformaldehyde solution 1％ was then infused via left ventricle at the perfusion pressure 120 mmHg.The eyeballs were extracted 2 minutes later,and the leakage of EB in rat retinas was examined.Results RT-PCR electrophoresis exhibited the response bands of MMP-2 mRNA,and MMP-9 mRNA and GAPDH,with the gene size of 436,536 and 484 bp,respectively.The difference of the MMP2 mRNA and MMP-9 mRNA was statisticaly significant (F =20.336,P =0.000 ; F =8.742,P =0.002) ; and the relative expressions of MMP-2 mRNA and MMP-9 mRNA were significantly higher in the diabetic group and diabetes +GM6001 group than those in the control group (all at P＜0.01),and the relative expressions of MMP-2 mRNA and MMP-9 mRNA in the diabetes+GM6001 group was significantly reduced in comparison with the diabetic group(both P=0.01,P=0.02).The standardized EB content in the retinas of the control group,diabetic group and diabetes+ GM6001 group was (12.60±3.50) ng/mg,(26.52±7.14) ng/mg and (17.55±2.65) ng/mg,showing a significant difference (F=17.032,P＜0.01),and EB content in rat retinas in the diabetic group was higher than that in the control group (P=0.003),and that in the diabetes+GM6001 group was lower in comparison with the diabetic group (P=0.020).Conclusions Intravitreal injection of GM6001 can down-regulate the expression of MMP-2 mRNA and MMP-9 mRNA in diabetic rats and therefore protect BRB.
