球周注射壳聚糖甘油磷酸二钠对实验性白兔后巩膜的加固作用
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摘要
目的:探讨球周注射壳聚糖甘油磷酸二钠对实验性白兔后巩膜的加固效果。
方法:
1选用大耳白兔20只,雌雄各半,体重2-2.5kg,随机抽取5只为对照组。其余动物为处理组,均以左眼为手术眼。用注射器套自制的钝性针头,由切口处进针,沿筋膜下间隙紧贴着巩膜向后进针3mm-4mm,注入壳聚糖甘油磷酸二钠1.2ml-1.5ml。术毕用庆大地米注射液冲洗结膜囊,涂红霉素眼膏,包扎术眼。
2术后正常活动、定时饮食,测实验动物0h、24h、48h、72h餐前身体重量,计算每日重量改变量。
3术后2、4、8周测眼压、眼底荧光血管造影、眼眶CT、眼A超、眼B超。之后耳缘静脉注入空气处死白兔,冰冻兔头12小时后用自制小锯条沿兔眼眶做冠状切面和矢状切面,其余动物摘除眼球,冲洗干净,切取混合巩膜组织,先肉眼观察,再分别以10%福尔马林液固定,石蜡包埋切片,HE染色,光镜观察。
结果:
1 C/GP生物相容性:球周注射C/GP引起眼部轻度炎症反应。
2眼压测量结果:术后2周、4周、8周,2周组与对照组左眼眼压比较没有统计差异(P>0.05);2周组和4周组比较没有统计学差异(P>0.05);4周、8周组较对照组左眼眼压升高,有统计学差异(P<0.05);2周和8周组比较,8周组眼压升高,有统计学差异(P<0.05);4周和8周组比较,8周组眼压升高,有统计学差异(P<0.05);右眼各组间2、4、8周眼压比较没有统计学差异(P>0.05)。
3眼轴测量结果:动物眼部球周注射C/GP对眼轴影响较对照组无统计学差异(P>0.05);术后2、4、8周左眼眼轴比较无统计学差异(P>0.05),右眼眼轴比较无统计学差异(P>0.05),左眼和右眼眼轴比较无统计学差异(P>0.05)。
4眼眶CT检查结果:2周组左眼后极部偏颞上方巩膜较对照组左眼显著增厚,形成一规则环状“新巩膜”,密度均匀,与巩膜密度大致相同,低于眶骨密度,其与周围眼眶间隙清晰,视神经清晰规整。
5 B超检查结果:术后2周B超见左眼后部颞上方后巩膜有不规则条状低回声区,术后4周后此低回声区变为月牙状强回声区,术后8周此强回声区减弱,后部巩膜回声均匀一致,玻璃体未见异常回声,视网膜回声规整,视神经回声规整,眶内未见异常回声。
6冰冻眼球切面结果:术后8周冰冻眼球切面见,龙胆紫染色组织均匀地贴附于后部巩膜上,并与巩膜融为一体,与周围组织间隙清楚。
7巩膜组织病理学观察结果:术后2周见,巩膜表层胶原纤维排列粗细不均,炎性细胞增加,新生血管增生。术后4周见巩膜表层肉芽组织增生,新生血管增生。术后8周见后巩膜胶原纤维增粗,排列整齐,肉芽组织增生,后巩膜增厚。
8眼底荧光血管造影观察结果:对照组与实验组眼底荧光造影未发现明显变化。
结论:
1壳聚糖甘油磷酸二钠在眼部球周注射的炎性反应很小,生物相容性较好,可作为后巩膜加固术的一种新型材料。
2球周注射壳聚糖甘油磷酸二钠改变了巩膜的硬度及弹性,但对眼轴的影响不大。
3壳聚糖甘油磷酸二钠混合物球周注射后能够均匀的在球后巩膜分布,并与巩膜紧密的贴附,支撑着后巩膜。
4球周注射物对视网膜脉络膜及玻璃体无影响;壳聚糖甘油磷酸二钠与巩膜融合后发生炎症反应,新生血管增生,肉芽组织增生,胶原纤维增粗,排列整齐,巩膜重新塑形。
Objective:To discuss the effect on posterior scleral reinforcement of experimental rabbits with periorbitally injecting chitosan glycerophosphate disodium.
Method:
1 selected 20 rabbits, male and female in half, weight 2-2.5kg, were randomly selected five rabbits : the blank control group;the remaining rabbits were ( The left eye of treatment group rabbits were surgical eye.)Extraocular disinfection with 2% lidocaine retrobulbar anesthesia, 2% tetracaine surface anesthesia; injection needle were curved extraocular injection needle with consistent with the curvature of the eye. Open eyelid device opened eyelid . Conjunctival were dished with non-tooth forceps above the eye from the temporal limbus 3mm.We cut 1mm small incision with scalpel,peeled fascia exposed porcelain white sclera surface, blunt needle was entangled with syringe, needled along incision. Along fascia space , closing to the sclera, needled to the backward 3mm- 4mm,injected chitosan glycerophosphate disodium1.2mm-1.5mm, suture conjunctival、fascia and superficial sclera,then massaged eyeball,made drug evenly distributed periorbital.After the surgery, gentamicin injection and dexamethasone injection washed conjunctival sac, smeared erythromycin eye ointment, wrap the surgical eye.
2 After surgery,we observed normal activities, regular diet of rabbits in the first 4h, 24h, 48h, 72h .Observe all experimental animals, movement times, difficulty breathing and eye irritation. Measuring experimental animals 0h, 24h, 48h, 72h before eating, calculate weight change per day.
3 After surgery 2w,4w,6w,8w measure IOP, FFA, CT, A/B -ultrasonography. Rabbits were killed by injecting air embolism in ear vein, injected gentian violet 0.2ml cavity under the fascia of surgical eye, massaged eyeball,made gentian violet liquid and chitosan glycerophosphate disodium mixed fully.After frozen rabbit head 12h, along rabbits orbital made coronal section and sagittal Section with self-made small saw blade,the rest rabbits were removed eyeball,washed clean, Mixed scleral tissue was cut, the naked eye observed firstly,then fixed with 10%formalin respectively, used paraffin-embedded、slice,HE staining,light microscope observed.
Results:
1 C / GP biocompatibility: periorbital injection of C / GP eye caused mild inflammatory response. Weight changes of groups comparation were no significant difference (P> 0.05).
2 intraocular pressure measurement results: After 2 weeks, 4 weeks, 8 weeks, compared between 2 weeks group and the control group, there is no statistical difference in the left eye IOP (P> 0.05).With the control group,intraocular pressure of 4 weeks, 8 weeks increased,and the two groups were statistical difference (P <0.05).
Compared with 2 weeks, intraocular pressure of 8-week group increased, were statistically significant difference (P <0.05). Compared with 4 weeks, intraocular pressure of 8-week group increased, existed statistically significant difference (P <0.05).Comperaion of 2,4,8 weeks , the right eye of the three groups were not statistically significant compared intraocular pressure (P >0.05).
3 axial length measurements: Compared with the control group , axial length of operation groups were no significant difference (P> 0.05). Compared with axial length of the left eye of rabbits in 2,4,8 weeks were no significant difference (P> 0.05), compared with axial length of the right eye of rabbits in 2,4,8 weeks were no significant difference (P> 0.05), left and right eye of rabbits in 2,4,8 weeks were no significant difference between axial (P> 0.05).
4 orbital CT examination results: Compared with left eye of control group ,sclera of left eye over the posterior pole partial temporal of rabbits in 2 weeks were significant thickening, formed a annular“new sclera”, the new sclera was uniform density, was roughly the same density with the sclera, lowered than the orbital bone density.C/GP integrated with the sclera, and cleared space around the eyes, optic nerve cleared and structured.
5 B-ultrasonography test results: After surgical 2 weeks showed :back of the eye above the posterior temporal of left eye in surgical control group had irregular hypoechoic area , After surgical 4 weeks the hypoechoic area had changed into a crescent-shaped hyperechoic area. After surgical 8 weeks the hyperechoic area had weaked, posterior echo had becomed uniform, no abnormal echo in the vitreous body, echo s of retina tructured , optic nerve echo structured, orbital no abnormal echo.
6 Frozen section results of the eyeball: After surgical 8 weeks , frozen section of the eyeball shown: Crystal violet staining tissue uniformly attached on the posterior scleral,and integrated with the sclera, and had a clear boundaries with the surrounding tissue space.
7 Histopathological observation results: After surgical 2 weeks shown:scleral surface of the uneven thickness of collagen fibers arranged, inflammatory cells had increased , Neovascularization had regenerated. After surgical 4 weeks shown: Scleral surface of granulation tissue had regenerated, neovascularization had regenerated, scleral collagen fibers had thicken and ranged regularly.Granulation tissue had regenerated, Scleral had thicken.
8 Observation results of fundus fluorescein angiography:In the control group and test group had no significant change in fundus fluorescein angiography .
Conclusion:
1 periorbitally injecting of C / GP did not cause that the rabbits reflect systemic poisoning,however,could cause mild inflammatory response.The inflammatory response of chitosan glycerophosphate disodium in eyeball,it had a good biocompatibility,it could become a scleral reinforcement of a new material.
2 At different time points after surgical measured the results of intraocular pressure and axial length ,statistical analysis, with the observation period change, intraocular pressure of experimental group rabbits were increased more than control group, axial length of rabbits were no changed. The change of axial length, to some extent, could reflect the mechanical changes in the eye wall.periorbitally injecting chitosan glycerophosphate disodium had changed sclera of the hardness and elasticity,but it had little effects on the axial length .
3 Through the eyeballs of frozen after surgical coronal and Sagittal plane observation, B-ultrasonography and CT of orbital,we fund that the mixture of chitosan glycerophosphate disodium periorbitally injected,it could evenly distributed in the ball after the sclera, closely attached with the sclera, supported the posterior.
4 By fundus fluorescein angiography and histopathological observation of scleral,we had fund that periorbitally injecting chitosan glycerophosphate disodium had no effect on the retina, choroid, vitreous body; after chitosan glycerophosphate disodium had integated with the sclera, inflammatory response occurred, neovascularization had regenerated and granulation tissue regenerated, collagen fibers had thicken and ranged regularly, sclera reformed.
方法:
1选用大耳白兔20只,雌雄各半,体重2-2.5kg,随机抽取5只为对照组。其余动物为处理组,均以左眼为手术眼。用注射器套自制的钝性针头,由切口处进针,沿筋膜下间隙紧贴着巩膜向后进针3mm-4mm,注入壳聚糖甘油磷酸二钠1.2ml-1.5ml。术毕用庆大地米注射液冲洗结膜囊,涂红霉素眼膏,包扎术眼。
2术后正常活动、定时饮食,测实验动物0h、24h、48h、72h餐前身体重量,计算每日重量改变量。
3术后2、4、8周测眼压、眼底荧光血管造影、眼眶CT、眼A超、眼B超。之后耳缘静脉注入空气处死白兔,冰冻兔头12小时后用自制小锯条沿兔眼眶做冠状切面和矢状切面,其余动物摘除眼球,冲洗干净,切取混合巩膜组织,先肉眼观察,再分别以10%福尔马林液固定,石蜡包埋切片,HE染色,光镜观察。
结果:
1 C/GP生物相容性:球周注射C/GP引起眼部轻度炎症反应。
2眼压测量结果:术后2周、4周、8周,2周组与对照组左眼眼压比较没有统计差异(P>0.05);2周组和4周组比较没有统计学差异(P>0.05);4周、8周组较对照组左眼眼压升高,有统计学差异(P<0.05);2周和8周组比较,8周组眼压升高,有统计学差异(P<0.05);4周和8周组比较,8周组眼压升高,有统计学差异(P<0.05);右眼各组间2、4、8周眼压比较没有统计学差异(P>0.05)。
3眼轴测量结果:动物眼部球周注射C/GP对眼轴影响较对照组无统计学差异(P>0.05);术后2、4、8周左眼眼轴比较无统计学差异(P>0.05),右眼眼轴比较无统计学差异(P>0.05),左眼和右眼眼轴比较无统计学差异(P>0.05)。
4眼眶CT检查结果:2周组左眼后极部偏颞上方巩膜较对照组左眼显著增厚,形成一规则环状“新巩膜”,密度均匀,与巩膜密度大致相同,低于眶骨密度,其与周围眼眶间隙清晰,视神经清晰规整。
5 B超检查结果:术后2周B超见左眼后部颞上方后巩膜有不规则条状低回声区,术后4周后此低回声区变为月牙状强回声区,术后8周此强回声区减弱,后部巩膜回声均匀一致,玻璃体未见异常回声,视网膜回声规整,视神经回声规整,眶内未见异常回声。
6冰冻眼球切面结果:术后8周冰冻眼球切面见,龙胆紫染色组织均匀地贴附于后部巩膜上,并与巩膜融为一体,与周围组织间隙清楚。
7巩膜组织病理学观察结果:术后2周见,巩膜表层胶原纤维排列粗细不均,炎性细胞增加,新生血管增生。术后4周见巩膜表层肉芽组织增生,新生血管增生。术后8周见后巩膜胶原纤维增粗,排列整齐,肉芽组织增生,后巩膜增厚。
8眼底荧光血管造影观察结果:对照组与实验组眼底荧光造影未发现明显变化。
结论:
1壳聚糖甘油磷酸二钠在眼部球周注射的炎性反应很小,生物相容性较好,可作为后巩膜加固术的一种新型材料。
2球周注射壳聚糖甘油磷酸二钠改变了巩膜的硬度及弹性,但对眼轴的影响不大。
3壳聚糖甘油磷酸二钠混合物球周注射后能够均匀的在球后巩膜分布,并与巩膜紧密的贴附,支撑着后巩膜。
4球周注射物对视网膜脉络膜及玻璃体无影响;壳聚糖甘油磷酸二钠与巩膜融合后发生炎症反应,新生血管增生,肉芽组织增生,胶原纤维增粗,排列整齐,巩膜重新塑形。
Objective:To discuss the effect on posterior scleral reinforcement of experimental rabbits with periorbitally injecting chitosan glycerophosphate disodium.
Method:
1 selected 20 rabbits, male and female in half, weight 2-2.5kg, were randomly selected five rabbits : the blank control group;the remaining rabbits were ( The left eye of treatment group rabbits were surgical eye.)Extraocular disinfection with 2% lidocaine retrobulbar anesthesia, 2% tetracaine surface anesthesia; injection needle were curved extraocular injection needle with consistent with the curvature of the eye. Open eyelid device opened eyelid . Conjunctival were dished with non-tooth forceps above the eye from the temporal limbus 3mm.We cut 1mm small incision with scalpel,peeled fascia exposed porcelain white sclera surface, blunt needle was entangled with syringe, needled along incision. Along fascia space , closing to the sclera, needled to the backward 3mm- 4mm,injected chitosan glycerophosphate disodium1.2mm-1.5mm, suture conjunctival、fascia and superficial sclera,then massaged eyeball,made drug evenly distributed periorbital.After the surgery, gentamicin injection and dexamethasone injection washed conjunctival sac, smeared erythromycin eye ointment, wrap the surgical eye.
2 After surgery,we observed normal activities, regular diet of rabbits in the first 4h, 24h, 48h, 72h .Observe all experimental animals, movement times, difficulty breathing and eye irritation. Measuring experimental animals 0h, 24h, 48h, 72h before eating, calculate weight change per day.
3 After surgery 2w,4w,6w,8w measure IOP, FFA, CT, A/B -ultrasonography. Rabbits were killed by injecting air embolism in ear vein, injected gentian violet 0.2ml cavity under the fascia of surgical eye, massaged eyeball,made gentian violet liquid and chitosan glycerophosphate disodium mixed fully.After frozen rabbit head 12h, along rabbits orbital made coronal section and sagittal Section with self-made small saw blade,the rest rabbits were removed eyeball,washed clean, Mixed scleral tissue was cut, the naked eye observed firstly,then fixed with 10%formalin respectively, used paraffin-embedded、slice,HE staining,light microscope observed.
Results:
1 C / GP biocompatibility: periorbital injection of C / GP eye caused mild inflammatory response. Weight changes of groups comparation were no significant difference (P> 0.05).
2 intraocular pressure measurement results: After 2 weeks, 4 weeks, 8 weeks, compared between 2 weeks group and the control group, there is no statistical difference in the left eye IOP (P> 0.05).With the control group,intraocular pressure of 4 weeks, 8 weeks increased,and the two groups were statistical difference (P <0.05).
Compared with 2 weeks, intraocular pressure of 8-week group increased, were statistically significant difference (P <0.05). Compared with 4 weeks, intraocular pressure of 8-week group increased, existed statistically significant difference (P <0.05).Comperaion of 2,4,8 weeks , the right eye of the three groups were not statistically significant compared intraocular pressure (P >0.05).
3 axial length measurements: Compared with the control group , axial length of operation groups were no significant difference (P> 0.05). Compared with axial length of the left eye of rabbits in 2,4,8 weeks were no significant difference (P> 0.05), compared with axial length of the right eye of rabbits in 2,4,8 weeks were no significant difference (P> 0.05), left and right eye of rabbits in 2,4,8 weeks were no significant difference between axial (P> 0.05).
4 orbital CT examination results: Compared with left eye of control group ,sclera of left eye over the posterior pole partial temporal of rabbits in 2 weeks were significant thickening, formed a annular“new sclera”, the new sclera was uniform density, was roughly the same density with the sclera, lowered than the orbital bone density.C/GP integrated with the sclera, and cleared space around the eyes, optic nerve cleared and structured.
5 B-ultrasonography test results: After surgical 2 weeks showed :back of the eye above the posterior temporal of left eye in surgical control group had irregular hypoechoic area , After surgical 4 weeks the hypoechoic area had changed into a crescent-shaped hyperechoic area. After surgical 8 weeks the hyperechoic area had weaked, posterior echo had becomed uniform, no abnormal echo in the vitreous body, echo s of retina tructured , optic nerve echo structured, orbital no abnormal echo.
6 Frozen section results of the eyeball: After surgical 8 weeks , frozen section of the eyeball shown: Crystal violet staining tissue uniformly attached on the posterior scleral,and integrated with the sclera, and had a clear boundaries with the surrounding tissue space.
7 Histopathological observation results: After surgical 2 weeks shown:scleral surface of the uneven thickness of collagen fibers arranged, inflammatory cells had increased , Neovascularization had regenerated. After surgical 4 weeks shown: Scleral surface of granulation tissue had regenerated, neovascularization had regenerated, scleral collagen fibers had thicken and ranged regularly.Granulation tissue had regenerated, Scleral had thicken.
8 Observation results of fundus fluorescein angiography:In the control group and test group had no significant change in fundus fluorescein angiography .
Conclusion:
1 periorbitally injecting of C / GP did not cause that the rabbits reflect systemic poisoning,however,could cause mild inflammatory response.The inflammatory response of chitosan glycerophosphate disodium in eyeball,it had a good biocompatibility,it could become a scleral reinforcement of a new material.
2 At different time points after surgical measured the results of intraocular pressure and axial length ,statistical analysis, with the observation period change, intraocular pressure of experimental group rabbits were increased more than control group, axial length of rabbits were no changed. The change of axial length, to some extent, could reflect the mechanical changes in the eye wall.periorbitally injecting chitosan glycerophosphate disodium had changed sclera of the hardness and elasticity,but it had little effects on the axial length .
3 Through the eyeballs of frozen after surgical coronal and Sagittal plane observation, B-ultrasonography and CT of orbital,we fund that the mixture of chitosan glycerophosphate disodium periorbitally injected,it could evenly distributed in the ball after the sclera, closely attached with the sclera, supported the posterior.
4 By fundus fluorescein angiography and histopathological observation of scleral,we had fund that periorbitally injecting chitosan glycerophosphate disodium had no effect on the retina, choroid, vitreous body; after chitosan glycerophosphate disodium had integated with the sclera, inflammatory response occurred, neovascularization had regenerated and granulation tissue regenerated, collagen fibers had thicken and ranged regularly, sclera reformed.
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