严重眼烧伤早期角膜移植的实验研究
http://www.100md.com
37c医学网
作者：S.A.Yakimenko R.I.Chalanova N.F.Leus S.G.Kolomiychuk
单位：乌克兰费拉托夫眼科研究所，270061
关键词：眼烧伤；早期角膜移植；溶解酶
眼科研究000201
摘要 目的 研究角膜碱性烧伤后，早期用蛋白溶解酶清除坏死组织，然后即行板层角膜移植，动态观察其疗效。 方法 实验兔36只，用10%?NaOH造成兔角膜烧伤模型，早期应用水解蛋白酶冲洗除去表层坏死组织后，即行板层角膜移植。 结果 有70.5%实验兔角膜透明或半透明愈合。 结论 在严重眼烧伤时，早期应用水解蛋白酶创造一个板层角膜移植的良好环境可以获得较好的治疗效果。
分类号 R772.2
, 百拇医药
Urgent keratoplasty in the treatment of severe eye burns
S.A.Yakimenko，R.I.Chalanova，N.F.Leus，S.G.Kolomiychuk
(Institute of Eye Diseases and Tissue Therapy by V.P.Filatov，Odessa，270061 Ukraine)
Abstract ObjectiveTo examine the efficiency of urgent lamellar keratoplastic surgery with preliminary ferment necrectomy of corneal isolated alkaline burns and its influence on proteolytical indices in serum and camera moisture.MethodsExperiments were conducted on both eyes of rabbits(n＝36)using an alkaline burn model of 10% NaOH with 10-s exposure time.The doses of proteases(lekozim，alkaline proteases)were established.ResultsThe research showed that with the aid of proteolitical ferments it is possible to remove necrotic corneal layers and prepare the cornea for lamellar keratoplasty.This allows improved wound healing after lamellar keratoplasty in eye burns.More favorable course of eye burn disease reflected positively also on the proteolitical activity(PA) indices of the serum and camera moisture.Urgent keratoplasty with preliminary necrectomy resulted in transparent and half-transparent healing of the transplant in 70.5% of cases.ConclusionThe application of proteolitical ferments in the early terms allows to create better conditions for the lamellar keratoplasty with the severe eye burns.Urgent keratoplasty with preliminary ferment necrectomy appears to be more effective for the treatment of chemical burns of rabbit corneas.
, 百拇医药
Key words eye burn urgent keratoplasty proteolytical ferments
INTRODUCTION
Clinical displays of eye burn disease are stipulated by those violations which cause damages in biological eye structures.Violations of innervation，blood supply，suppression of all types of metabolism，the phenomena of autosensibilization and autointoxication of the organism with products of tissue necrosis lead to deepening of structural violations，burdening of course of eye burn disease and to severe invalidizing results.
, 百拇医药
Taking all this into consideration N.A.Puchkovskaya and co-authors[1]suggested during first 24 h after severe eye burn to carry out urgent lamellar keratoplasty with preliminary necrectomy of damaged corneal layers.Authors’ experimental investigations showed that the efficacy of urgent keratoplasty depends on the burn’s severeness.The more severe the burn，the more quickly lamellar keratoplasty should be performed.However，despite positive results of these investigations，the operation of urgent keratoplasty did not find application.The reason of it is the lack of diagnostic methods of severeness and depth of corneal affection.
, http://www.100md.com
We studied necrolitical properties of a number of proteases on a model of severe alkaline burns to the cornea and ascertained that with the help of ferments，it is possible to achieve full removal of necrotic layers of the cornea and prepare the cornea for keratoplasty.We also ascertained concentrations and exposure time of proteases necessary to achieve removal of changed corneal tissues[2].
Study of the efficiency of urgent lamellar keratoplasty with preliminary ferment necrectomy on isolated alkaline burns of the cornea and its influence upon indices of proteolytical activity in blood serum and camera moisture in the dynamics of burn disease.
, 百拇医药
MATERIALS AND METHODS
Experiments were carried out on both eyes of 36 rabbits.Using a special applicator，an isolated corneal burn with 10% NaOH was reproduced with exposure time of 10 s(Fig.1).19 rabbits formed I control group(CG)，where the treatment was carried out with thrice-repeated instillations of desinfecting means daily.In the second experimental group(EG)，of 17 rabbits in 3 h after burn a ferment processing of cornea was carried out with following urgent lamellar keratoplasty.Applications of 27 units of alkaline protease in 2.0 ml of polyvinyl spirit or lekozym 17.5 Fip in 2.0 ml 3.5% of urea were carried out with the help of eye-bath-eyelid-dilator for 5 min.In 25～30 min full tearing away of necrotic layers of cornea was seen (Fig.2).A layer-by-layer transplant 10 mm in diameter was cut out and was fixed with main stitches to the edge of the dislike bed(Fig.3).After the operation the treatment was carried out the same way as in the control group with thrice-repeated instillations of desinfecting means daily.The observation over animals was carried out for 1.5 months with the help of biomicroscopy and fluorescein sample.At the same time the following symptoms of burn disease were evaluated：the duration of the inflammatory reaction，the terms of the beginning and finishing of the epithelization of the defect of the cornea and the transplant，the development of such complications as ulcerization or fusion of the cornea or the transplant，outcomes of the burn disease according to the intensity of cornea dimness.
, 百拇医药
Fig.1 The alkaline model burn of the cornea.The cornea was dimness accordance the “porcelain plate” Fig.2 The cornea after ferments necrectomy.Full tearing away of necrotic layers Fig.3 Lamellar transplant was fixed to the edge of the bed Fig.4 The result of keratoplasty is the half-transparent grafting of the transplant
In both groups on d 1，3，7，14 and 21，the blood and camera moisture were taken for biochemical analysis.The study of proteolytical activity of these biological liquids were carried out according to methods based on splitting of the substratum bensoyl-arginin-r-nitroanilyn with proteolitical ferments.
, 百拇医药
RESULTS
As a result of cornea burn in the optical zone a dense primary dimness is formed according to the “porcelain plate”type 10 mm in diameter.In the damaged section the cornea loses an epithelial cover.In the control group，taking into account an isolated character of the damage with the presence of a rim of undamaged cornea the process of epithelization of the defect proceeded in a wavelike way.In connection with presence of the regeneration source from the undamaged paralimbal section of the cornea for 3?d an almost full cover of deepithelized section occured.However，weakly solded with underlying burnt stroma，the neoplasm epithelium was easily seized making a vast defect by 6-d observation.During the 2nd～3rd week of the observation an exacerbation peak of erosion process was marked，connected with progressing trophic disorders in the damaged eye tissues.Almost in a half of the cases (18?eyes：47.3%)those were deep，up to Descemet’s membrane，defects of the cornea stroma.From d 13 to d 15 of observation the epithelization activized and was over by observation for 35～38 d.
, 百拇医药
The inflammatory reaction of the front section of the eye kept up to 38～40 d.The outcome of the burn process of this group of animals in 100% cases was the formation of a dense wall-eye.The course of the burn disease in the group of the operated animal was different.The inflammatory reaction increased on d?1，reaching maximum on d 7～14 and gradually reduced with almost complete liqudation of observation by d?30.The erosion of the transplant was kept from d 6～7 to d 21～23 of observation. The data are presented in Tab.1.
, 百拇医药
Tab.1 Eye burn disease symptoms cessation terms
in observed groups of animals(
±s) Rabbits
n
Inflammation cessation
Erosionepithelialization
of cornea or transplant
Beginning
Finishing
, 百拇医药
Beginning
Finishing
groupControl
38
20.68±1.04
38.63±1.20
13.91±0.79
35.21±0.89
Experimental
34
15.22±1.18
30.91±1.66
, 百拇医药
8.51±0.482
2.32±1.39
P
<0.001
<0.001
<0.001
<0.001
In the operated group of animals complications from the transplant in the form of deep erosions were observed on 5 eyes (14.7%).In the outcome of the burn process in the experimental group on 24 eyes (70%) transparent and half transparent dimnesses were formed.Only on 10 eyes (29.5%) dense wall-eyes were formed.
, http://www.100md.com
The study of the proteolytical activity(PA) level in the blood serum showed the following(Tab.2).During d?1 of observation on the background of the developing phenomena of inflammation in the front section of the eye PA increases in both groups：twice as much in the experimental group and three times as much in the control group.Tab.2 Blood serum proteolitical activity dynamics in the
control and experimental animals group(n/kat/l) Observation
, 百拇医药
terms (d)
Controlgroup
Experimental group
P
n±s
n±s
Before burn
5
1.15±0.22
5
, 百拇医药
0.98±0.22
>0.05
1
19
3.74±0.93
10
2.47±0.82
>0.05
3
8
4.76±0.92
9
3.16±0.9
, http://www.100md.com
>0.05
7
9
8.03±0.96
7
2.62±1.17
<0.05
14
10
6.21±0.72
8
83.81±1.02
>0.05
, http://www.100md.com
21
6
6.50±1.22
7
0.9±0.18
<0.001
30
6
2.46±0.77
6
0.8±0.081
>0.05
F21urther increase of PA reached maximum in the control group on d 7 and till d 14～d 21 this index remained high corresponding to mostly expressed inflammatory reaction.By observation for 30 d，the PA index reduced without reaching the initial level.
, 百拇医药
In the experimental group the PA index was higher than the initial level from d 3 to d 14.However，the degree of the PA increase in the experimental group was lower than in the control one.From d 21 to d 30 of observation， the PA index in the main group returned to the initial level，while in the control group it remained high.
The PA indices in the camera moisture were 4 times lower than in the blood serum (Tab.3).On d 1 these indices increase twice as much and then gradually grow reaching maximum on d 14.The PA indices in the control group were higher than in the experimental one with the reliable difference on d 7 and d 14.Further in both groups the PA level decrease was observed，reaching initial values by observation for 30 d.Tab.3 Camera moisture proteolytical activity dynamics in the
, 百拇医药
control and experimental groups of animals(n/kat/l) Observation
terms(d)
Control group
Experimental group
P
n±s
n±s
Before burn
, http://www.100md.com 4
0.31±0.40
4
0.29±0.15
>0.05
1
5
0.62±0.07
5
0.52±0.06
>0.05
3
4
, 百拇医药
0.47±0.045
4
0.34±0.04
>0.05
7
5
0.61±0.022
5
0.44±0.058
<0.05
14
5
1.33±0.18
, 百拇医药
5
0.82±0.082
<0.05
21
4
0.9±0.07
5
0.7±0.067
>0.05
30
4
0.32±0.02
4
, 百拇医药
0.24±0.08
>0.05
DISCUSSION
The results of these experiments showed that with the help of proteolytical ferments，it is possible to obtain tearing away of necrotically changed cornea layers and thus prepare the cornea for the following lamellar keratoplasty.The carrying out of such operation allows to improve the course of the eye burn disease.
In the experimental group the cessation of the inflammatory reaction began earlier and finished earlier.The epithelization of the transplant erosion was also achieved earlier in comparison with the cornea erosion in the control group of animals.More favorable course of the eye burn disease reflected positively also on the PA indices of the blood serum and camera moisture.As a whole，the dynamics of these indices corresponds to the dynamics of the inflammatory reaction in both groups.The increased level of the PA of the blood serum in the control group remained increased during the entire term of observation with the highest indices from d 7 till d 21.In the experimental group，the PA indices of blood serum were increased only to 14-d observation.In the camera moisture the maximum increase of the PA in both groups was marked from d 7 to d 21，but in the main group this index on d 7 and d 14 was reliably lower than in the control group.More favorable course of the burn disease in the main group led to its better results，in 70.5% it was allowed to obtain the transparent and half-transparent grafting of the transplant(Fig.4)，while in the control group in 100% of the cases the cornea wall-eyes were formed.
, 百拇医药
The application of proteolytical ferments in the early terms allows to create better conditions for the lamellar keratoplasty with the severe eye burns.
The urgent keratoplasty with the preliminary ferment necrectomy promotes more favorable course of the burn disease and achieves better outcome in the treatment.
REFERENCES
1，Puchkovskaya NA，Sulgina NS，Nepomyachaja VM.The pathogenesis and treatment of eye burns and their seguelae.Kiev，1973，197
2，Chalanova RI.Izychenie necroliticheskix svojstv nekotorix proteoliticheskix fermentov pri schelochnix ojogax glaz.Ophthalmology，1998，4∶277
(收稿：1999-12-10), 百拇医药
单位：乌克兰费拉托夫眼科研究所，270061
关键词：眼烧伤；早期角膜移植；溶解酶
眼科研究000201
摘要 目的 研究角膜碱性烧伤后，早期用蛋白溶解酶清除坏死组织，然后即行板层角膜移植，动态观察其疗效。 方法 实验兔36只，用10%?NaOH造成兔角膜烧伤模型，早期应用水解蛋白酶冲洗除去表层坏死组织后，即行板层角膜移植。 结果 有70.5%实验兔角膜透明或半透明愈合。 结论 在严重眼烧伤时，早期应用水解蛋白酶创造一个板层角膜移植的良好环境可以获得较好的治疗效果。
分类号 R772.2
, 百拇医药
Urgent keratoplasty in the treatment of severe eye burns
S.A.Yakimenko，R.I.Chalanova，N.F.Leus，S.G.Kolomiychuk
(Institute of Eye Diseases and Tissue Therapy by V.P.Filatov，Odessa，270061 Ukraine)
Abstract ObjectiveTo examine the efficiency of urgent lamellar keratoplastic surgery with preliminary ferment necrectomy of corneal isolated alkaline burns and its influence on proteolytical indices in serum and camera moisture.MethodsExperiments were conducted on both eyes of rabbits(n＝36)using an alkaline burn model of 10% NaOH with 10-s exposure time.The doses of proteases(lekozim，alkaline proteases)were established.ResultsThe research showed that with the aid of proteolitical ferments it is possible to remove necrotic corneal layers and prepare the cornea for lamellar keratoplasty.This allows improved wound healing after lamellar keratoplasty in eye burns.More favorable course of eye burn disease reflected positively also on the proteolitical activity(PA) indices of the serum and camera moisture.Urgent keratoplasty with preliminary necrectomy resulted in transparent and half-transparent healing of the transplant in 70.5% of cases.ConclusionThe application of proteolitical ferments in the early terms allows to create better conditions for the lamellar keratoplasty with the severe eye burns.Urgent keratoplasty with preliminary ferment necrectomy appears to be more effective for the treatment of chemical burns of rabbit corneas.
, 百拇医药
Key words eye burn urgent keratoplasty proteolytical ferments
INTRODUCTION
Clinical displays of eye burn disease are stipulated by those violations which cause damages in biological eye structures.Violations of innervation，blood supply，suppression of all types of metabolism，the phenomena of autosensibilization and autointoxication of the organism with products of tissue necrosis lead to deepening of structural violations，burdening of course of eye burn disease and to severe invalidizing results.
, 百拇医药
Taking all this into consideration N.A.Puchkovskaya and co-authors[1]suggested during first 24 h after severe eye burn to carry out urgent lamellar keratoplasty with preliminary necrectomy of damaged corneal layers.Authors’ experimental investigations showed that the efficacy of urgent keratoplasty depends on the burn’s severeness.The more severe the burn，the more quickly lamellar keratoplasty should be performed.However，despite positive results of these investigations，the operation of urgent keratoplasty did not find application.The reason of it is the lack of diagnostic methods of severeness and depth of corneal affection.
, http://www.100md.com
We studied necrolitical properties of a number of proteases on a model of severe alkaline burns to the cornea and ascertained that with the help of ferments，it is possible to achieve full removal of necrotic layers of the cornea and prepare the cornea for keratoplasty.We also ascertained concentrations and exposure time of proteases necessary to achieve removal of changed corneal tissues[2].
Study of the efficiency of urgent lamellar keratoplasty with preliminary ferment necrectomy on isolated alkaline burns of the cornea and its influence upon indices of proteolytical activity in blood serum and camera moisture in the dynamics of burn disease.
, 百拇医药
MATERIALS AND METHODS
Experiments were carried out on both eyes of 36 rabbits.Using a special applicator，an isolated corneal burn with 10% NaOH was reproduced with exposure time of 10 s(Fig.1).19 rabbits formed I control group(CG)，where the treatment was carried out with thrice-repeated instillations of desinfecting means daily.In the second experimental group(EG)，of 17 rabbits in 3 h after burn a ferment processing of cornea was carried out with following urgent lamellar keratoplasty.Applications of 27 units of alkaline protease in 2.0 ml of polyvinyl spirit or lekozym 17.5 Fip in 2.0 ml 3.5% of urea were carried out with the help of eye-bath-eyelid-dilator for 5 min.In 25～30 min full tearing away of necrotic layers of cornea was seen (Fig.2).A layer-by-layer transplant 10 mm in diameter was cut out and was fixed with main stitches to the edge of the dislike bed(Fig.3).After the operation the treatment was carried out the same way as in the control group with thrice-repeated instillations of desinfecting means daily.The observation over animals was carried out for 1.5 months with the help of biomicroscopy and fluorescein sample.At the same time the following symptoms of burn disease were evaluated：the duration of the inflammatory reaction，the terms of the beginning and finishing of the epithelization of the defect of the cornea and the transplant，the development of such complications as ulcerization or fusion of the cornea or the transplant，outcomes of the burn disease according to the intensity of cornea dimness.
, 百拇医药
Fig.1 The alkaline model burn of the cornea.The cornea was dimness accordance the “porcelain plate” Fig.2 The cornea after ferments necrectomy.Full tearing away of necrotic layers Fig.3 Lamellar transplant was fixed to the edge of the bed Fig.4 The result of keratoplasty is the half-transparent grafting of the transplant
In both groups on d 1，3，7，14 and 21，the blood and camera moisture were taken for biochemical analysis.The study of proteolytical activity of these biological liquids were carried out according to methods based on splitting of the substratum bensoyl-arginin-r-nitroanilyn with proteolitical ferments.
, 百拇医药
RESULTS
As a result of cornea burn in the optical zone a dense primary dimness is formed according to the “porcelain plate”type 10 mm in diameter.In the damaged section the cornea loses an epithelial cover.In the control group，taking into account an isolated character of the damage with the presence of a rim of undamaged cornea the process of epithelization of the defect proceeded in a wavelike way.In connection with presence of the regeneration source from the undamaged paralimbal section of the cornea for 3?d an almost full cover of deepithelized section occured.However，weakly solded with underlying burnt stroma，the neoplasm epithelium was easily seized making a vast defect by 6-d observation.During the 2nd～3rd week of the observation an exacerbation peak of erosion process was marked，connected with progressing trophic disorders in the damaged eye tissues.Almost in a half of the cases (18?eyes：47.3%)those were deep，up to Descemet’s membrane，defects of the cornea stroma.From d 13 to d 15 of observation the epithelization activized and was over by observation for 35～38 d.
, 百拇医药
The inflammatory reaction of the front section of the eye kept up to 38～40 d.The outcome of the burn process of this group of animals in 100% cases was the formation of a dense wall-eye.The course of the burn disease in the group of the operated animal was different.The inflammatory reaction increased on d?1，reaching maximum on d 7～14 and gradually reduced with almost complete liqudation of observation by d?30.The erosion of the transplant was kept from d 6～7 to d 21～23 of observation. The data are presented in Tab.1.
, 百拇医药
Tab.1 Eye burn disease symptoms cessation terms
in observed groups of animals(
±s) Rabbitsn
Inflammation cessation
Erosionepithelialization
of cornea or transplant
Beginning
Finishing
, 百拇医药
Beginning
Finishing
groupControl
38
20.68±1.04
38.63±1.20
13.91±0.79
35.21±0.89
Experimental
34
15.22±1.18
30.91±1.66
, 百拇医药
8.51±0.482
2.32±1.39
P
<0.001
<0.001
<0.001
<0.001
In the operated group of animals complications from the transplant in the form of deep erosions were observed on 5 eyes (14.7%).In the outcome of the burn process in the experimental group on 24 eyes (70%) transparent and half transparent dimnesses were formed.Only on 10 eyes (29.5%) dense wall-eyes were formed.
, http://www.100md.com
The study of the proteolytical activity(PA) level in the blood serum showed the following(Tab.2).During d?1 of observation on the background of the developing phenomena of inflammation in the front section of the eye PA increases in both groups：twice as much in the experimental group and three times as much in the control group.Tab.2 Blood serum proteolitical activity dynamics in the
control and experimental animals group(n/kat/l) Observation
, 百拇医药
terms (d)
Controlgroup
Experimental group
P
n
±sn
±sBefore burn
5
1.15±0.22
5
, 百拇医药
0.98±0.22
>0.05
1
19
3.74±0.93
10
2.47±0.82
>0.05
3
8
4.76±0.92
9
3.16±0.9
, http://www.100md.com
>0.05
7
9
8.03±0.96
7
2.62±1.17
<0.05
14
10
6.21±0.72
8
83.81±1.02
>0.05
, http://www.100md.com
21
6
6.50±1.22
7
0.9±0.18
<0.001
30
6
2.46±0.77
6
0.8±0.081
>0.05
F21urther increase of PA reached maximum in the control group on d 7 and till d 14～d 21 this index remained high corresponding to mostly expressed inflammatory reaction.By observation for 30 d，the PA index reduced without reaching the initial level.
, 百拇医药
In the experimental group the PA index was higher than the initial level from d 3 to d 14.However，the degree of the PA increase in the experimental group was lower than in the control one.From d 21 to d 30 of observation， the PA index in the main group returned to the initial level，while in the control group it remained high.
The PA indices in the camera moisture were 4 times lower than in the blood serum (Tab.3).On d 1 these indices increase twice as much and then gradually grow reaching maximum on d 14.The PA indices in the control group were higher than in the experimental one with the reliable difference on d 7 and d 14.Further in both groups the PA level decrease was observed，reaching initial values by observation for 30 d.Tab.3 Camera moisture proteolytical activity dynamics in the
, 百拇医药
control and experimental groups of animals(n/kat/l) Observation
terms(d)
Control group
Experimental group
P
n
±sn
±sBefore burn
, http://www.100md.com 4
0.31±0.40
4
0.29±0.15
>0.05
1
5
0.62±0.07
5
0.52±0.06
>0.05
3
4
, 百拇医药
0.47±0.045
4
0.34±0.04
>0.05
7
5
0.61±0.022
5
0.44±0.058
<0.05
14
5
1.33±0.18
, 百拇医药
5
0.82±0.082
<0.05
21
4
0.9±0.07
5
0.7±0.067
>0.05
30
4
0.32±0.02
4
, 百拇医药
0.24±0.08
>0.05
DISCUSSION
The results of these experiments showed that with the help of proteolytical ferments，it is possible to obtain tearing away of necrotically changed cornea layers and thus prepare the cornea for the following lamellar keratoplasty.The carrying out of such operation allows to improve the course of the eye burn disease.
In the experimental group the cessation of the inflammatory reaction began earlier and finished earlier.The epithelization of the transplant erosion was also achieved earlier in comparison with the cornea erosion in the control group of animals.More favorable course of the eye burn disease reflected positively also on the PA indices of the blood serum and camera moisture.As a whole，the dynamics of these indices corresponds to the dynamics of the inflammatory reaction in both groups.The increased level of the PA of the blood serum in the control group remained increased during the entire term of observation with the highest indices from d 7 till d 21.In the experimental group，the PA indices of blood serum were increased only to 14-d observation.In the camera moisture the maximum increase of the PA in both groups was marked from d 7 to d 21，but in the main group this index on d 7 and d 14 was reliably lower than in the control group.More favorable course of the burn disease in the main group led to its better results，in 70.5% it was allowed to obtain the transparent and half-transparent grafting of the transplant(Fig.4)，while in the control group in 100% of the cases the cornea wall-eyes were formed.
, 百拇医药
The application of proteolytical ferments in the early terms allows to create better conditions for the lamellar keratoplasty with the severe eye burns.
The urgent keratoplasty with the preliminary ferment necrectomy promotes more favorable course of the burn disease and achieves better outcome in the treatment.
REFERENCES
1，Puchkovskaya NA，Sulgina NS，Nepomyachaja VM.The pathogenesis and treatment of eye burns and their seguelae.Kiev，1973，197
2，Chalanova RI.Izychenie necroliticheskix svojstv nekotorix proteoliticheskix fermentov pri schelochnix ojogax glaz.Ophthalmology，1998，4∶277
(收稿：1999-12-10), 百拇医药