背景 研究证实,紫外光-核黄素光动力疗法可通过改变角膜的理化性质控制圆锥角膜的进展,羊膜移植已广泛用于眼表疾病,羊膜胶原成分与角膜相似,目前关于紫外光-核黄素诱导的胶原交联用于羊膜组织的研究尚少见.目的 探讨紫外光-核黄素对冻存羊膜的作用. 方法 在知情同意条件下,获取并常规处理人羊膜组织,-80℃保存.制备成2 mm×15 mm大小,用简单随机法中的抽签法将其分为4个组,每组6片羊膜,前3组均给予紫外光-核黄素(质量分数0.1％)处理30 min(波长为370 nm,紫外光功率分别为1、2、3 mW/cm2,光源距羊膜10 mm),第4组不做任何处理,为对照组.利用微力材料试验机对处理后的各组羊膜分别进行生物力学测量,记录羊膜长度改变5％、10％、15％时各组样本所需的拉力值(mN).用环钻钻取直径7 mm的羊膜,用抽签法将其分为4个组,每组5片羊膜,各组处理情况同上,将处理后的各组羊膜浸于质量分数0.1％胶原蛋白酶Ⅰ溶液中进行消化,对羊膜透明度进行评分,并记录羊膜完全溶解时间.关于羊膜的组织学观察,用抽签法将羊膜分为紫外光-核黄素组、核黄素组及对照组,每组3片羊膜,各组分别给予紫外光-核黄素(0.1％)处理30 min(波长为370 nm,紫外光功率为3 mW/cm2,光源距羊膜10 mm)、0.1％核黄素浸泡30 min和生理盐水浸泡30 min,透射电子显微镜下进行观察. 结果 羊膜长度改变5％、10％、15％时,对照组和1、2、3 mW/cm2紫外光组4个组所需拉力值差异均有统计学意义(F=3.411,P=0.037；F=9.927,P=0.001;F=11.118,P=0.000)；交联后抗拉力性能明显增强,且紫外光功率越大,羊膜的抗拉性增加越明显,差异均有统计学意义(P＜0.05).对照组羊膜完全酶解时间为(8.6±1.8)h,1、2、3 mW/cm2紫外光组分别为(39.6±2.3)、(71.4±0.9)、(78.8±1.8)h,交联后抗酶解能力增强,且紫外光功率越大,羊膜的抗酶解能力增强越明显,差异有统计学意义(P＜0.01).透射电子显微镜超微结构观察,交联后羊膜基质层胶原纤维密度增加、胶原间连接增多、胶原与上皮间的连接加强.结论 紫外光-核黄素可使冻存羊膜发生胶原交联,从而使其组织结构发生变化,生物力学性质改变,抗酶解能力加强.
揹景 研究證實,紫外光-覈黃素光動力療法可通過改變角膜的理化性質控製圓錐角膜的進展,羊膜移植已廣汎用于眼錶疾病,羊膜膠原成分與角膜相似,目前關于紫外光-覈黃素誘導的膠原交聯用于羊膜組織的研究尚少見.目的 探討紫外光-覈黃素對凍存羊膜的作用. 方法 在知情同意條件下,穫取併常規處理人羊膜組織,-80℃保存.製備成2 mm×15 mm大小,用簡單隨機法中的抽籤法將其分為4箇組,每組6片羊膜,前3組均給予紫外光-覈黃素(質量分數0.1％)處理30 min(波長為370 nm,紫外光功率分彆為1、2、3 mW/cm2,光源距羊膜10 mm),第4組不做任何處理,為對照組.利用微力材料試驗機對處理後的各組羊膜分彆進行生物力學測量,記錄羊膜長度改變5％、10％、15％時各組樣本所需的拉力值(mN).用環鑽鑽取直徑7 mm的羊膜,用抽籤法將其分為4箇組,每組5片羊膜,各組處理情況同上,將處理後的各組羊膜浸于質量分數0.1％膠原蛋白酶Ⅰ溶液中進行消化,對羊膜透明度進行評分,併記錄羊膜完全溶解時間.關于羊膜的組織學觀察,用抽籤法將羊膜分為紫外光-覈黃素組、覈黃素組及對照組,每組3片羊膜,各組分彆給予紫外光-覈黃素(0.1％)處理30 min(波長為370 nm,紫外光功率為3 mW/cm2,光源距羊膜10 mm)、0.1％覈黃素浸泡30 min和生理鹽水浸泡30 min,透射電子顯微鏡下進行觀察. 結果 羊膜長度改變5％、10％、15％時,對照組和1、2、3 mW/cm2紫外光組4箇組所需拉力值差異均有統計學意義(F=3.411,P=0.037；F=9.927,P=0.001;F=11.118,P=0.000)；交聯後抗拉力性能明顯增彊,且紫外光功率越大,羊膜的抗拉性增加越明顯,差異均有統計學意義(P＜0.05).對照組羊膜完全酶解時間為(8.6±1.8)h,1、2、3 mW/cm2紫外光組分彆為(39.6±2.3)、(71.4±0.9)、(78.8±1.8)h,交聯後抗酶解能力增彊,且紫外光功率越大,羊膜的抗酶解能力增彊越明顯,差異有統計學意義(P＜0.01).透射電子顯微鏡超微結構觀察,交聯後羊膜基質層膠原纖維密度增加、膠原間連接增多、膠原與上皮間的連接加彊.結論 紫外光-覈黃素可使凍存羊膜髮生膠原交聯,從而使其組織結構髮生變化,生物力學性質改變,抗酶解能力加彊.
배경 연구증실,자외광-핵황소광동력요법가통과개변각막적이화성질공제원추각막적진전,양막이식이엄범용우안표질병,양막효원성분여각막상사,목전관우자외광-핵황소유도적효원교련용우양막조직적연구상소견.목적 탐토자외광-핵황소대동존양막적작용. 방법 재지정동의조건하,획취병상규처리인양막조직,-80℃보존.제비성2 mm×15 mm대소,용간단수궤법중적추첨법장기분위4개조,매조6편양막,전3조균급여자외광-핵황소(질량분수0.1％)처리30 min(파장위370 nm,자외광공솔분별위1、2、3 mW/cm2,광원거양막10 mm),제4조불주임하처리,위대조조.이용미력재료시험궤대처리후적각조양막분별진행생물역학측량,기록양막장도개변5％、10％、15％시각조양본소수적랍력치(mN).용배찬찬취직경7 mm적양막,용추첨법장기분위4개조,매조5편양막,각조처리정황동상,장처리후적각조양막침우질량분수0.1％효원단백매Ⅰ용액중진행소화,대양막투명도진행평분,병기록양막완전용해시간.관우양막적조직학관찰,용추첨법장양막분위자외광-핵황소조、핵황소조급대조조,매조3편양막,각조분별급여자외광-핵황소(0.1％)처리30 min(파장위370 nm,자외광공솔위3 mW/cm2,광원거양막10 mm)、0.1％핵황소침포30 min화생리염수침포30 min,투사전자현미경하진행관찰. 결과 양막장도개변5％、10％、15％시,대조조화1、2、3 mW/cm2자외광조4개조소수랍력치차이균유통계학의의(F=3.411,P=0.037；F=9.927,P=0.001;F=11.118,P=0.000)；교련후항랍력성능명현증강,차자외광공솔월대,양막적항랍성증가월명현,차이균유통계학의의(P＜0.05).대조조양막완전매해시간위(8.6±1.8)h,1、2、3 mW/cm2자외광조분별위(39.6±2.3)、(71.4±0.9)、(78.8±1.8)h,교련후항매해능력증강,차자외광공솔월대,양막적항매해능력증강월명현,차이유통계학의의(P＜0.01).투사전자현미경초미결구관찰,교련후양막기질층효원섬유밀도증가、효원간련접증다、효원여상피간적련접가강.결론 자외광-핵황소가사동존양막발생효원교련,종이사기조직결구발생변화,생물역학성질개변,항매해능력가강.
Background Studies confirmed that ultraviolet A (UVA)- riboflavin photodynamic therapy can control keratoconus progresses by altering the physicochemical property of cornea.The collagen components of amniotic membrane transplantation is similar to that of cornea and amniotic membrane transplantation has been widely used to ocular surface reconstruction.However,the study on UVA riboflavin-induced-collagen crosslinking for amniotic tissue is less now. Objective This study was to investigate the role of UVA-riboflavin on frozen-preserved human amniotic membrane. Methods Human amnions were obtained in informed consent and prepared into 2 mm×15 mm pieces and were then divided into 4 groups using lottery method and 6 pieces for each group.The first 3 groups were treated with the photosensitizer riboflavin and UVA-irradiation ( wavelength:370 nm ; irradiation energy:1,2 or 3 mW/cm2,distance:10 mm) for 30 minutes,and the untreated fourth group was as control group.Biomechanical stress-strain test was performed using a microcomputer-controlled biomaterial tester and the stress(mN) was recorded when the strains were set to 5％,10％ and 15％.The 7 mm diameter of human amniotic membrane pieces were trephined and divided into 4 groups(5 pieces for each group) with the treated method as mentioned above,and then the buttons were exposed to 0.1％ collagenase Ⅰ solution.The transparency was scored and the complete dissolving time was record.In histological evaluation,three groups (3 pieces for each group) of human amniotic membranes were treated using UVAriboflavin(3 mW/cm2),0.1％ riboflavin,normal saline for 30 minutes respectively and examined under the transmission electron microscopy.This study was performed under the permission of the Ethic Commission of Beijing Tongren Hospital. Results When the strain was 5％,10％,15％,the stress of control group and 1,2,3 mW/cm2UVA group were statistically signifcantly different ( F =3.411,P =0.037; F =9.927,P =0.001;F=11.118,P=0.000).The tensile strength of human amniotic membrane cross-linked with UVA-riboflavin was statistically significantly increased in comparison to the control group (P＜0.05 ),and the tensile strength of human amniotic membrane became stronger as UVA power increased.The complete dissolve time was (8.6± 1.8 ) hours for the control group,(39.6± 2.3 ) hours for 1 mW/cm2 UVA group,(71.4±0.9 ) hours for 2 mW/cm2 UVA group,(78.8± 1.8 ) hours for 3 mW/cm2 UVA group,showing the enhanced anti-enzyme ability of human amniotic membrane after cross-linking(P＜0.01 ).The collagen density in the UVA-riboflavin treated group was increased,the connection among the collagen fibers as well as between the stroma and the epithelium became tighter than those of control group.Conclusions Collagen cross-linking with UVA-riboflavin make the biomechanical strength and enzymatic resistance of human amniotic membrane enhance and ultrastructure change of human amniotic membrane.
