中华眼科杂志
中華眼科雜誌
중화안과잡지
Chinese Journal of Ophthalmology
2011年
3期
252-259
,共8页
蔡蓉蓉%王艳%徐建江%张朝然
蔡蓉蓉%王豔%徐建江%張朝然
채용용%왕염%서건강%장조연
渗透压%眼%上皮%黏蛋白5AC%泪器
滲透壓%眼%上皮%黏蛋白5AC%淚器
삼투압%안%상피%점단백5AC%루기
Osmotic pressure%Eye%Epithelium%Mucin 5AC%Lacrimal apparatus
目的 探讨高渗透压对兔眼表上皮和黏蛋白(MUC)5AC表达的影响.方法 实验研究.将18只健康新西兰大白兔采用计算机随机数字法分为3组,分别为高渗盐溶液(500 mmol/L)、生理盐水(308 mmol/L)滴眼组及空白对照组,每组6只兔.实验第0、7、14天测定泪液分泌量与泪膜破裂时间(BUT),结膜印迹细胞学测定杯状细胞密度.实验第7、14天处死动物,行角结膜光镜和电镜观察,结膜组织末端脱氧核苷酸转移酶介导的脱脲苷三磷酸缺口末端标记法(TUNEL)检测,第14天的结膜组织另行MUC5AC表达的免疫组织化学染色和免疫印迹法检测.同一时间点上泪液分泌量值、BUT值和杯状细胞密度的组间比较采用单因素方差分析法,进一步两两比较采用Bonferroni法;若方差不齐则采用Kruskal-Wallis H检验.组内不同时间点比较采用配对t检验.结果 实验第7和第14天,高渗液组BUT分别为(7.6±2.5)和(7.0±2.3)s,较第0天的(10.3±2.5)s显著缩短(t=5.800,4.950;均P＜0.01),与空白对照和生理盐水组相比亦明显缩短(F=8.030,P＜0.01).高渗液组结膜杯状细胞密度明显减少,第7和第14天分别为(19.5±16.6)和(32.3±18.2)个/mm2,与第0天(75.7±43.4)个/mm2相比,差异有统计学意义(t=5.319,2.970;均P＜0.05).高渗盐溶液滴眼第14天,光镜下可见结膜上皮下炎性细胞浸润,角膜上皮部分脱落;扫描电镜下可见角膜上皮细胞肿胀、脱屑、微绒毛减少;透射电镜下可见角膜上皮细胞问连接松弛,基底层上皮细胞内自噬空泡增多,结膜上皮微绒毛减少,杯状细胞内分泌颗粒减少;TUNEL检测结膜上皮可见明显的阳性染色;结膜组织免疫组织化学染色和免疫印迹检测则发现MUC5AC表达明显低于其他两组.生理盐水组与空白对照组的结膜杯状细胞密度及角结膜上皮细胞结构在实验中则均无明显变化.结论 眼表高渗压力可破坏角结膜上皮细胞结构,造成杯状细胞密度降低和MUC5AC表达水平下降,并导致泪膜稳定性下降.
目的 探討高滲透壓對兔眼錶上皮和黏蛋白(MUC)5AC錶達的影響.方法 實驗研究.將18隻健康新西蘭大白兔採用計算機隨機數字法分為3組,分彆為高滲鹽溶液(500 mmol/L)、生理鹽水(308 mmol/L)滴眼組及空白對照組,每組6隻兔.實驗第0、7、14天測定淚液分泌量與淚膜破裂時間(BUT),結膜印跡細胞學測定杯狀細胞密度.實驗第7、14天處死動物,行角結膜光鏡和電鏡觀察,結膜組織末耑脫氧覈苷痠轉移酶介導的脫脲苷三燐痠缺口末耑標記法(TUNEL)檢測,第14天的結膜組織另行MUC5AC錶達的免疫組織化學染色和免疫印跡法檢測.同一時間點上淚液分泌量值、BUT值和杯狀細胞密度的組間比較採用單因素方差分析法,進一步兩兩比較採用Bonferroni法;若方差不齊則採用Kruskal-Wallis H檢驗.組內不同時間點比較採用配對t檢驗.結果 實驗第7和第14天,高滲液組BUT分彆為(7.6±2.5)和(7.0±2.3)s,較第0天的(10.3±2.5)s顯著縮短(t=5.800,4.950;均P＜0.01),與空白對照和生理鹽水組相比亦明顯縮短(F=8.030,P＜0.01).高滲液組結膜杯狀細胞密度明顯減少,第7和第14天分彆為(19.5±16.6)和(32.3±18.2)箇/mm2,與第0天(75.7±43.4)箇/mm2相比,差異有統計學意義(t=5.319,2.970;均P＜0.05).高滲鹽溶液滴眼第14天,光鏡下可見結膜上皮下炎性細胞浸潤,角膜上皮部分脫落;掃描電鏡下可見角膜上皮細胞腫脹、脫屑、微絨毛減少;透射電鏡下可見角膜上皮細胞問連接鬆弛,基底層上皮細胞內自噬空泡增多,結膜上皮微絨毛減少,杯狀細胞內分泌顆粒減少;TUNEL檢測結膜上皮可見明顯的暘性染色;結膜組織免疫組織化學染色和免疫印跡檢測則髮現MUC5AC錶達明顯低于其他兩組.生理鹽水組與空白對照組的結膜杯狀細胞密度及角結膜上皮細胞結構在實驗中則均無明顯變化.結論 眼錶高滲壓力可破壞角結膜上皮細胞結構,造成杯狀細胞密度降低和MUC5AC錶達水平下降,併導緻淚膜穩定性下降.
목적 탐토고삼투압대토안표상피화점단백(MUC)5AC표체적영향.방법 실험연구.장18지건강신서란대백토채용계산궤수궤수자법분위3조,분별위고삼염용액(500 mmol/L)、생리염수(308 mmol/L)적안조급공백대조조,매조6지토.실험제0、7、14천측정루액분비량여루막파렬시간(BUT),결막인적세포학측정배상세포밀도.실험제7、14천처사동물,행각결막광경화전경관찰,결막조직말단탈양핵감산전이매개도적탈뇨감삼린산결구말단표기법(TUNEL)검측,제14천적결막조직령행MUC5AC표체적면역조직화학염색화면역인적법검측.동일시간점상루액분비량치、BUT치화배상세포밀도적조간비교채용단인소방차분석법,진일보량량비교채용Bonferroni법;약방차불제칙채용Kruskal-Wallis H검험.조내불동시간점비교채용배대t검험.결과 실험제7화제14천,고삼액조BUT분별위(7.6±2.5)화(7.0±2.3)s,교제0천적(10.3±2.5)s현저축단(t=5.800,4.950;균P＜0.01),여공백대조화생리염수조상비역명현축단(F=8.030,P＜0.01).고삼액조결막배상세포밀도명현감소,제7화제14천분별위(19.5±16.6)화(32.3±18.2)개/mm2,여제0천(75.7±43.4)개/mm2상비,차이유통계학의의(t=5.319,2.970;균P＜0.05).고삼염용액적안제14천,광경하가견결막상피하염성세포침윤,각막상피부분탈락;소묘전경하가견각막상피세포종창、탈설、미융모감소;투사전경하가견각막상피세포문련접송이,기저층상피세포내자서공포증다,결막상피미융모감소,배상세포내분비과립감소;TUNEL검측결막상피가견명현적양성염색;결막조직면역조직화학염색화면역인적검측칙발현MUC5AC표체명현저우기타량조.생리염수조여공백대조조적결막배상세포밀도급각결막상피세포결구재실험중칙균무명현변화.결론 안표고삼압력가파배각결막상피세포결구,조성배상세포밀도강저화MUC5AC표체수평하강,병도치루막은정성하강.
Objective To investigate the effects of hyperosmotic stress on rabbit ocular surface and mucin 5AC (MUC5AC) expression. Methods Experimental study. Eighteen New Zealand white rabbits were randomly divided into three groups with equal number as hyperosmolar saline solution (HOSS,500 mmol/L) group, normal saline (NS, 308 mmol/L) group and blank control group respectively. In HOSS and NS groups, the HOSS and NS eye drops were instilled on bilateral eyes six times every day for 14 days. On day 0, 7 and 14, Schirmer Ⅰ test and tear break-up time (BUT) were measured and conjunctival impression cytology specimens were collected. On day 7 and 14, cornea and conjunctiva were harvested for Hematoxylin and Eosin (HE) staining, scanning and transmission electron microscopy observation and conjunctival TUNEL examination. On day 14, the conjunctiva were also harvested for immunology histological staining and western blot to evaluate the expression of MUC5AC. Results In HOSS group, the BUT on day 7 and 14 was (7.6±2. 5) and (7.0±2. 3) s respectively which was significantly shorter than the (10.3±2. 5) s on day 0(t=5. 800,4. 950; P ＜ 0.01), and also significantly shorter than the BUT in NS and control groups (F=8.030, P ＜ 0.01). But the Schirmer Ⅰ test value did not change obviously in and between all those three groups. The mean conjunctival goblet cell (GC) density in HOSS group on day 7 and 14 was (19.5±16.6) and (32.3±18.2) cells/mm2 respectively which was also significantly lower than the (75.7±43.4) cells/mm2 on day 0 (t=5.319,2. 970; P ＜ 0.05). However the GC density did not change obviously in other two groups with time. After instillation of HOSS for 14 days,subepithelial inflammatory cell infiltration was showed on conjunctival tissue specimens and decreased epithelial layers and evident desquamation were found in the cornea specimens by the HE staining. Under the electron microscope, decreased microvilli and loosened intercellular junction in the superficial epithelium and increased autophagic vesicles in basal epithelium were observed in the cornea in HOSS group; and decreased microvilli and mucous granules were found in the conjunctiva in HOSS group. Obvious TUNEL positive staining was showed in the conjunctiva in the HOSS group. Also the MUC5AC immunology histological staining and western blot indicated decreased MUC5AC protein expression in HOSS group. Conclusion Hyperosmotic stress destroyed the structure of ocular surface epithelium, induced the decrease of conjunctival goblet cell density and MUC5AC expression, and led to the decreased tear film stability.
