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Year : 1962  |  Volume : 10  |  Issue : 3  |  Page : 55-60

Amniotic membrane grafts in corneal ulcer

Ophthalmic Department, G. R. Medical College, Gwalior, India

Date of Web Publication18-Mar-2008

Correspondence Address:
I M Shukla
Ophthalmic Department, G. R. Medical College, Gwalior
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Source of Support: None, Conflict of Interest: None

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How to cite this article:
Shukla I M. Amniotic membrane grafts in corneal ulcer. Indian J Ophthalmol 1962;10:55-60

How to cite this URL:
Shukla I M. Amniotic membrane grafts in corneal ulcer. Indian J Ophthalmol [serial online] 1962 [cited 2020 Aug 14];10:55-60. Available from: http://www.ijo.in/text.asp?1962/10/3/55/39559

Table 2

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Table 2

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Table 1

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Table 1

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This study was undertaken with a view to assess the role of amnio­tic membrane grafts in the healing of corneal ulcers and to evaluate the degree of symptomatic relief obtained by covering the raw sur­face of the ulcer.

  Review of Literature Top

From time to time various sub­stances both from heterogenous and autogenous sources have been used as a very useful adjunct in treat­ing a few types of corneal ulcers. Bruckner tried conjunctival flap hooding for serpiginous ulcers. Miklos (1950) ,found total conjunc­tival hooding most efficacious for healing of serpiginous ulcers. Siegal (1944) used buccal mucosa to replace injured conjunctiva in cases of ocular burns. He was of opinion that by inserting a graft, the nutri­tion of cornea and its metabolism was re-established. Egg membrane for chemical injuries of eye was used by Croll and Croll (1952). These authors felt that insertion of egg membrane between the con­juctiva and cornea served as a pro­tective cover for cornea, and delay­ed corneal healing was avoided. Besides avoiding formation of ad­hesions between burnt conjunctiva and cornea, it enhanced rapid epi­thelialization and sequalae such as corneal scarring etc., were avoided.

Amniotic membrane was first proposed by Chao, Humphreys and Penfield (1940) as a means of filling defect in meninges. But subsequent clinical and experimental studies have proved disappointing to Neuro-Surgeons. Later, amniotic membrane was used in the eye for the first time, by Sorsby and Sym­mons (1946) in the treatment of caustic burns. Another series of work was done by Sorsby, Hay­thorne and Read (1947) in Caustic burns. Sorsby while discussing the action of this membrane in cases of chemical burns suggests that such a graft over a raw surface will not prevent the growth of the proliferating edge of a wound and moreover it will guide such prolife­rations towards the wounded sur­face.

  Healing of Corneal Wounds Top

The healing of wounds is a com­plex phenomenon which probably requires the co-ordinated activity of many agents and which is influen­ced by numerous factors both ex­trinsic and intrinsic-Arey (1936). Some of them have only general effects whereas others specifically determine the completeness and smooth march of the pathological processes. The methods involved in repair of epithelial defects of cornea have recently been the subject of detailed study as also have been the influences exerted by various physical, chemical and therapeutic agents. Of the various factors, pro­tection by pad and bandage has long been advocated. A wound completely protected from mecha­nical, chemical and bacterial irrita­tion does not heal for many days. The healing period is materially shortened in the presence of a mild irritant or slight infection. It thus appears that the stimulus for heal­ing under ordinary conditions is not an internal factor. The reported beneficial action of pressure dres­sings is probably based on the mechanism. However, increased mechanical damage causes inhibition of mitosis and consequently regeneration of epithelial cells for several hours, - Duke-Elder and Goldsmith (1951), which delays the normal process of wound healing. According to Duke-Elder (0000) :im­mobility of lids is of great value not only in allowing rapid epitheli­alization but also most essential from the point of view of relieving pain. With a view to achieve rapid epithelialization and immediate symptomatic relief by covering the exposed nerve endings in a raw surface, directly by this membrane, we undertook the present study.

  Material and Methods Top

This study was divided into two parts (1) Clinical, (2) Experimental. In the clinical part seventy-two patients of corneal ulcers have been studied. They comprised both superficial and deep ulcers. Ten cases of superficial and twelve of deep served as control and were put deep on routine line of treatment (Drops and sub-conjunctival inject­ion of penicillin, atropine drops, fomentations, pad and bandage) while in fifty cases amniotic mem­brane grafts were applied in addi­tion. The above classification of superficial and deep ulcers is arbi­trary irrespective of the fact whether the ulcer was sloughing or non-sloughing, clear, or infected. The classification was made after staining the lession with fluroscein and is purely based on a subjective examination with oblique illumina­tion by a corneal loupe. Lesions which seemed to involve only the anterior half lavers of substantia propria were classified as superfi­cial while those that involved the layers deeper than half-thickness were termed as deep.

  Preparation of Membrane Top

Amniotic membrane used in these cases was taken from maternity room of Agra Medical College out of fresh placenta. It was then pre­pared for use by the method of Sorsby with slight modification. A healthy placenta immediately after delivery was washed and placed in normal saline and within 24 hours amnion was separated from chorion carefully, commencing from the fringe to the cord. After washing thoroughly in running water it was placed in saturated solution at common salt for 12 to 24 hours, charging the solution, once or twice in between. Any surface fat was removed with a piece of gauze by spreading the membrane on a black-backed plate of glass. A thorough washing was again done and the membrane was kept in dis­tilled water for half an hour fre­quently agitating the solution. Salt free amnion was then put into 1 in 150 solution of potassium hydroxide for one to two hours frequently agitating the solution, the time fact­or depending upon the change in colour of the membrane, which gradually took a white shine. A thorough washing was repeated by spreading the membrane till it was free of any soapy material. It was again immersed in ' distilled water for 12 hours. The membrane thus prepared was cut into pieces of size 1" x 1" by folding the bigger pieces and these pieces were stored in a sterilised bottle containing Penicillin solution which was charged, every 24 hours. Once prepared, the membrane could be stored for a week in a refrigerator.

  Method of Grafting Top

At the time of grafting, one piece was taken out by sterilised forceps and was allowed to dry lying in a petridish. The dried membrane, folded twice or thrice was ready for grafting.

The patient's eye was prepared by irrigating it with normal saline and cleansing the conjunctival sac by putting penicillin drops 2 hour­ly for 24 to 48 hours or intermittent instillation of sodium sulphaceta­mide 30% along with atropine 1%. In most of the cases the eye was fit for grafting after 24 hours of this preliminary treatment as there was no discharge after that.

The eye was anaesthetised by 1% Anethaine 2 or 3 drops and the facial nerve was blocked with 2% novocain. The grafts were applied by one of the two methods:­

1. Suturing Method:-A dried piece of membrane about the required size to cover the ulcerated part of the cornea and surrounding conjunctiva was removed from the petridish and spread on a sterilised dry towel. A No. 6 ophthalmic needle threaded with No. 0 black silk was passed through each corner of the membrane. A self retaining speculum was applied to the anaesthetised eye. The towel with graft and threaded needles was brought upto the eye and the needles were then in turn passed through the bulbar conjunctiva, thus suturing the graft in place. Liquid paraffin was put and both eyes were bandaged. This method was used only in 10 cases as this consumed more time and sutures were difficult to apply in the sore conjunctiva and also because sutures caused irritation.

2. Spreading Method was ap­plied in rest of the cases. After taking the required size of the membrane as judged by the con­junctival sac, we made a slit shaped or circular opening into the membrane in such a place that after applying the graft the opening corresponded to the healthy portion of the cornea, while the ulcer was completely covered by the mem­brane. The above precaution we took to avoid the temporary hazi­ness of the cornea which occurs otherwise, We spread the merz­brane from the upper fornix while the patient was looking down, to the lower fornix, while the patient was asked to look up. Later we carefully reposited the membrane over the required part of the cornea and conjunctival sac with an iris repo­sitar. Liquid paraffin was put and eye was bandaged.

The bandage was opened after 48 hours. It was noted that graft was no longer visible as it had got absorbed. Staining of the- ulcer was done and its size and depth were recorded. In cases where sutures were applied they were removed on the third day, when bandage was removed. After 48 hours, dressing with atropine and penicillin eye ointment was done. In cases where it was found necessary a second or a third graft was applied. The day­to-day observations on the relief of pain, photophobia, congestion and extent of scar tissue formation were recorded in tabular form.

  Experimental Study Top

For experimental study, four albino rabbits were chosen and an attempt was made to study the effect of amniotic membrane grafts by producing identical corneal breach of substance and surface in each eye. In two animals a superficial wound and in the other two, a deep wound were made by a 4.5 mm. trephine and keratome. The depth of wound could be adjusted by giving two rotations in an anaesthetised eye in the case of superficial wound and four rotations of trephine in the case of deep wound, removing the epithe­lium and substantia propria by a keratome. A uniform stain taken by 2% Fluorescein could give an idea of the uniformity of depth of wound.

The opposite eye of the rabbit served as control whereas on the experimental eye, routine treatment and amniotic grafts were applied by the spreading :method. In this expe­rimental study the amniotic mem­brane behaved as a foreign body. It became swollen and hard parch­ment like and had to be removed on the seventh day. The ulcer heal­ed on ninth day resulting in a dense opacity.

  Observations Top

The observations were recorded in tabular form separately in the superficial and deep ulcer groups.

[Table - 1] shows the beneficial effects of using amniotic grafts in both the superficial and deep ulcer groups. Denoting nebular scar as 1, macular scar as 2 and leucoma as 3, the average scar-density has been worked out in numericals. In the treated group of deep ulcers, four cases showed formation of facets and so have been excluded for as­certaining the density.

  Discussion Top

With a view to achieve rapid epithelialization and immediate symptomatic relief by covering the raw surface with exposed nerve end­ings directly by amniotic mem­brane, we ventured to take up the present study. Being very much impressed by the results of Sorsby in cases of chemical burns of the eye treated by amniotic membrane, we were led to the idea of trying this membrane in cases of corneal ulcers, more so because any materi­al which does not behave as a foreign body over the eye should provide the most suitable coverage material. It has an added advant­age that it can be easily prepared and stored in any hospital or laboratory.

Prior to inserting such a graft it is of utmost importance to combat the infection because this immediate covering material worsens the con­dition of an ulcer, if it is grossly infected or sloughing.

From the above data it was very remarkable to find such quick relief of pain. No doubt in the first two hours this relief of pain was due to the effect of anaesthesia but later on when the effect wore off 70 per cent of cases of superficial ulcers and 40 per cent of the deep ulcer group did not complain of pain at all. This cause of immediate relief of pain in these cases must be due to the protective covering of the exposed nerve endings of the ulcerated corneal surface. This immediate covering material pro­bably acted by not allowing any irritant, mechanical, chemical or toxic to reach the nerve endings. The cases where pain was continu­ed for a longer period may be due to an inflammation of the iris.

In cases where amniotic mem­brane grafting was done there was marked reduction in the healing time. In superficial ulcer group the average healing time came down to 6 days as against 11.8 days in control cases and similarly in deep ulcer cases the average time came down from 19.5 days to 9.6 days where grafts were applied. Sorsby while discussing the action of this membrane in cases of chemical burns suggested that such a graft over a raw surface while not preventing growth of proliferat­ing edge of the wound, would guide such proliferations towards the wounded surface. Such phenomenon can explain the favourable effect of this membrane on healing. Be­sides, as the ulcer is protected, the mitosis of corneal cells and produc­tion of fibroblasts to fill up the ulcer proceeded unimpaired. More­over the mechanical trauma on the diseased cornea by blinking of the lids is also eliminated, thus allow­ing the process of healing to proceed unimpaired or even to hasten. An interesting temporary phenomenon noted in cases treated with amniotic membrane grafts was sustained corneal vascularization. Congestion persisted almost upto the time the ulcer healed or even in some cases it was continued after the healing. It is therefore quire pos­sible and reasonable to presume that there may be increased local supply of protective substances and other metabolites, helping in corneal respiration and metabolism with consequent regeneration of damag­ed corneal tissue.

Thus increased metabolic activity, and avoidance of irritants may be factors in enhancing the healing process. But then how much sub­stantive mechanical effect can be achieved by amniotic membrane is open to discussion because of the rabidity with which the membrane gets absorbed. In this series of clinical cases it was obvious that the healing process was definitely quickened by the grafting.

In our observations on experiment­al study it was shown that amniotic membrane when applied on rabbit cornea ' behaves as an irritating foreign body and does not get absorbed and rather gets swollen up and hard. On the other hand in our clinical cases this human amniotic membrane was invariably absorbed in about 48 hours. This above observation definitely proves that this membrane is species specific as has also been pointed out by Sorsby. Sorsby has further suggested a specific biological action between the damaged tissue, of conjunctival sac in cases of chemical burns and the amniotic membrane. This species specificity and rapid absorption of amniotic membrane in conjunctival sac de­finitely show that there is some biological action between the raw surface of cornea and amniotic membrane. Thus besides other things, this specific biological action in its turn may also be responsible for stimulating such a rapid heal­ing of ulcer.

Application of amniotic membrane in ulcer cases leads to the formation of thinner opacities than otherwise. It may be explained that due to the rapid growth of epithelium over the surface there is lesser formation of fibroblasts, there­by making the wound less dense. That may explain the formation of facets in certain cases of deer ulcers.

  Summary Top

A study of 50 cases of superficial and deep corneal ulcers with amniotic membrane grafts has been presented. The method of prepara­tion of amniotic membrane and grafting it on corneal ulcers has been described. It has been possible to conclude that amniotic membrane grafts give immediate relief from pain in most cases. An explanation on biochemical grounds is suggest­ed for the reduction in healing time and density of scar formation with amniotic grafting on clean ulcers.

Amniotic membrane has got its limitations of not being effective in the presence of slough and infection.

The grafts were also applied in experimental animals. It was found that amniotic membrane is species specific.[10]

  Acknowledgement Top

I am grateful to Dr. H. N. Bhatt, Principal Medical College, Agra for allowing me to work in that institu­tion. I am specially thankful to Professor K. N. Mathur whose const­ant guidance and direction helped me to go through this work.

  References Top

Arev, L. B., (iv3h). Phisol Rev.: 16, 327.  Back to cited text no. 1
Bruckner, A. Ophthalmologica, Basel, 117, 236.  Back to cited text no. 2
Croll, M., and Croll, L. J.; (1952), Am. J. Ophthal. 35: 1585-96.  Back to cited text no. 3
Duke-Elder, S. W. and Goldsmith, A. J. B., (1951), Recent Advances in Oph­thalmology, 283-88.  Back to cited text no. 4
Duke-Elder, S. W. Text Book of Ophthalmology, Vol. II. Kimpton London.  Back to cited text no. 5
Miklos, (1950), Brit. J. Ophth. 34: 335.  Back to cited text no. 6
Siegal, R., (rg44), Arch. Ophth. 32 104.  Back to cited text no. 7
Sorsby Modern Trends in Ophthal­mology, Vol. No. II, 504­  Back to cited text no. 8
Sorsby, A. and Symmons, H. M., (1946), Brit. J. Ophthal. 3, 337.  Back to cited text no. 9
Sorsby, A. Haythorne, J. and Read, H., (1947), Brit. Jourl. Ophthal., 31, 409.  Back to cited text no. 10


  [Table - 1], [Table - 2]


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  In this article
Review of Literature
Healing of Corne...
Material and Methods
Preparation of M...
Method of Grafting
Experimental Study
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