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ARTICLE
Year : 1960  |  Volume : 8  |  Issue : 3  |  Page : 57-59

Relation between intraocular tension and hyaluronic acid content of aqueous


Medical Collage, Nagpur, India

Date of Web Publication5-May-2008

Correspondence Address:
K R Kesavachar
Medical Collage, Nagpur
India
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Source of Support: None, Conflict of Interest: None


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How to cite this article:
Kesavachar K R. Relation between intraocular tension and hyaluronic acid content of aqueous. Indian J Ophthalmol 1960;8:57-9

How to cite this URL:
Kesavachar K R. Relation between intraocular tension and hyaluronic acid content of aqueous. Indian J Ophthalmol [serial online] 1960 [cited 2019 Dec 7];8:57-9. Available from: http://www.ijo.in/text.asp?1960/8/3/57/40678

Table 1

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Table 1

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It was in 1934, that Meyer and Palm­er isolated a mucopoly-saccharide from the vitreous humor which they labelled Hyaluronic acid. Subsequently it was isolated from the umbilical cord by the same workers. Hyaluronic acid is a viscous inucopoly-saccharide com­posed of equal quantities of N-acetyl glucosamine and glucuronic acid. Hyaluronic acid is absent from the blood stream and because of its pre­sence in the aqueous it is only reason­able to assume that this substance is secreted into the eye by the ciliary epithelium ('Meyei, Smythe and Gallardo 1938).

In a paper published in Oct. 1938, Never Smvthe and Gallardo have given figures after estimation of Hyaluronic acid in the aqueous in normal and abnormal eyes. According to them, the concentration of Hyaluronic acid in the aqueous is too low for estimation as Hyaluronic acid. They developed a method for break­ing down Hyaluronic acid into Glucosamine and Glucnronic acid and estimating the Glucosamine content of the aqueous. This Glucosamine may come from two sources, namely Hyaluronic acid or Protein. As the Protein content of aqueous is negli­gible, it was felt that this method of estimating Hyaluronic acid by the Glucosamine content of the aqueous was sufficiently accurate for the pur­poses of this study. They tested, only 5 samples of normal human aqueous and obtained an average Glucosamine content of 16 micrograms per grain of aqueous. They similarly estimated the Glucosamine content of aqueous in senile cataracts and primary Glancomas of all types (9 samples)and found no appreciable difference from the normal aqueous.

Wyburn and Bacsich suggested that the constant production of Hyaluronic acid in the- eye and its removal, presumably by enzymatic hydrolysis, was in some way concerned with the maintenance of normal ocular tension and the condition of simple glaucoma may well be related to some upset in the Hyaluronic acid Enzyme system. Pierre showed (1951) that Hyaluronic acid is present in the ocular tissues which binds the water in the tissue spaces. According to them, it is being constantly destroyed by Hyaluroniase which is present in the Iris and Ciliary body. Barany (1955) has mentioned that the resistance to aqueous outflow is reduced to half by perfusing the rabbit's eye with small quantities of Hyaluronidase, suggest­ing thereby the presence of Hyaluronic acid in the outflow barrier. Similar experiments were conducted by Francis et al (1956) on rabbit and human eyes and they came to the conclusion that in rabbits the increase in the aqueous outflow through the angles during perfusion was 50% and in human eyes, 25%. It is interest­ing to note that these workers do not mention anything about the Hyahuronic acid content of the aqueous itself.


  Material Top


As only human aqueous was requir­ed and as tapping normal human eyes was out of question, aqueous was tap­ped from eyes which were normal apart from having cataract. Cases were carefully selected after slit lamp examination of each eye, to eliminate eyes with signs of past or present Iridocyclitis or any other inflammation.

Collection of aqueous :­

The eyes were anaesthetised with anethain. A small scratch with a Graefe Knife was made just inside the limbus and an aqueous needle fitted onto a syringe was pushed into the anterior chamber and the aqueous aspirated out. All care was taken to avoid contamination of aqueous with blood (blood contains large quantities of Glucosamine). Each eye would give about 1/4c.c. of aqueous, and as half c.c. of aqueous was required for each estimation, aqueous from different eyes was pooled together. A slightly modified method was used for collection of aqueous from eyes where the anterior chamber was very shallow. Samples of aque­ous from normal eyes and from eyes of all types of primary glaucoma, were analysed.


  Method of Estimation Top


The glucosaminc was estimated by utilising the Elson Morgan reaction after breaking down the Hyaluronic acid in the aqueous into its com­ponents namely glucosamine and glucuronic acid. Half c.c. of aqueous taken in a Pyrex glass test tube was hydrolysed with half c.c. 8 N hydrochloric acid for four hours in a boiling waterbath, after sealing the test tube.

After hydrolysis the excess of hydrochloric acid was neutralised with 4 N sodium hydroxide so as to leave the solution only slightly acidic. To this was added 0.5. c.c. of fresh acetyl acetone solution and the test tube, kept in a wire basket, was immersed to half depth in boiling water for fifteen minutes. The contents were then cool­ed under tap water and 2.5 c.c. Of aldehvde free alcohol added. 0.5 c.c. of Ehrlich's reagent (prepared by dis­solving 0.8 gm of p. dimethyl aminohenraldchyde in 3 c.c. of aldehvde free alcohol, adding 30 c.c. of Cone. Hcl and leaving in a refrigerator was then added and the test tube placed in a waterbath at 37°C for thirty minutes along with similarly treated blank solution and standard solutions. A purple colour would develop. Estimation was then carried out by using the Klet Sumer­,on Photo Electric Colorimeter.

It might be added here that there is no other substance in the aqueous which responds to the Elson 'Morgan reaction. The amount of Hyaluronic acid is expressed in terms of glucosamine in micrograms per gram of aqueous.


  Results Top


Out of the large number of samples of normal aqueous analysed, the glucosaminc content varied within fairly wide ranges from 15.0 micro­grams to 24.0 micrograms, giving an average reading of 18.54 micrograms per grain of aqueous (compare with the figure given by Meyer et al who obtained an average of 16.0 micro­grams per grail of aqueous). Thiry­four samples of aqueous from glaucomatous eyes were analysed. The results are tabulated as follows :-[Table - 1]

Barany (1955) found that cattle aqueous invitro in the presence of oxygen, gradually reduces the viscocity of Hyaluronic acid indicating depoly­imerization of the Hyaluronic acid. He is of the opinion that this invitro activity of the aqueous is due to its ascorbic acid content. With the idea of finding out whether with the in­crease in the content of Hyaluronic acid there would be a reduction in the ascorbic acid content of aqueous several samples of normal and abnor­mal aqueous were analysed. The normal ascorbic acid content of aque­ous varies within very large limits and as such to get an average is very diffi­cult. Amongst the 26 samples of nor­mal aqueous analysed, the minimum was o.66 miligrams and the maximum 7.5 miligrams per cent giving an average of 2.5 miligrams. Six samples of aqueous from chronic simple Glaucoma gave an average reading of 1.77 miligrams per cent. One sample from a chronic congestive Glaucoma showed an ascorbic acid content of 1.25 miligrams. On the face of it there does seem to be a reduction in the ascorbic acid content of aqueous in glaucomatous eyes, but it is felt that too much emphasis should not be plac­ed on this as there is a very large variation in the ascorbic acid content of aqueous even in normal eyes.


  Discussion Top


Some facts have got to be kept in mind while drawing any inference from the above results :­

1) Hyaluronic acid is present in the aqueous but not in the blood stream. This substance is probably secreted by the ciliary epithelium into the eye.

2) Perfusion studies with hy­aluronidase on the outflow of aqueous in enucleated human eyes show that there is a very definite increase in the outflow.

3) Hyaluronidase has not been demonstrated in the aqueous but has been demonstrated in the iris and ciliary epithelium and also in the angle of the anterior chamber.

Amongst the samples of abnormal aqueous analysed, there has been a very significant and very definite rise in the Hyaluronic acid content. The preceding table shows clearly that there is a definite rise in the Hyaluronic acid content of aqueous in eyes with chronic simple Glaucoma. It will also be noticed that the Hyaluronic acid content is more in congestive types of Glaucoma and maximum in the aque­ous from acute congestive Glaucomas.[7]


  Conclusion Top


One fact emerges from a study of the results. There is an undoubted and definite rise in the Hyaluronic acid content of aqueous in glaucomat­ous eyes. It is tempting to infer that this causes increased viscocity of aqueous thus rendering difficult the normal exit of aqueous, thereby in­creasing the Intraocular tension. What exactly is the cause of increase in Hyaluronic acid content is not known. It may be a phenomenon associated with secretomotor imbalance. It is towards this end that further study should be undertaken.

Before concluding I wish to acknow­ledge gratefully the grant given for this work by the Governing Body of the Indian Council of Medical Research.

 
  References Top

1.
Adler Physiology of the Eve Clinical Application 1950 Edition Page 86.  Back to cited text no. 1
    
2.
Atkinson (1949), Arch. of Ophthal.Vol. 42 , 588, 628.  Back to cited text no. 2
    
3.
Barany E, (1955), Arch. Ophthal. 53, 612.  Back to cited text no. 3
    
4.
Francis. J. (1956), Arch. Ophthal. 55, 193.  Back to cited text no. 4
    
5.
McClean D. (1943), Biochemical J. 37, 169.  Back to cited text no. 5
    
6.
Pierre. (1951), A.M.A. Arch. of ()phthal. 45, 493.  Back to cited text no. 6
    
7.
Von Sallman (1950), A.M.A. Arch. of ()phthal, 43, 638.  Back to cited text no. 7
    



 
 
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