|Year : 1991 | Volume
| Issue : 4 | Page : 148-150
Clinico-biochemical study of experimental complicated cataracts
R Sihota, Madan Mohan, SK Angra, RL Mathur
Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All Institute of Medical Sciences, New Delhi 110 029, India
Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All Institute of Medical Sciences, New Delhi 110 029
Source of Support: None, Conflict of Interest: None
Clinically observed complicated cataracts, generally do not have a definite causal factor. We studied the effects of E. coli toxin injected suprachoroidally, to simulate the effect of toxins released by extraocular organisms on the lens. 79.2% of eyes had a definable cataract at the end of the 6th week of observation. The biochemical changes portrayed an increased oxidative activity in the lens, evidenced by a fall in glutathione concentration, and the consequent tertiary reorientation of proteins to increase insoluble proteins, forming a cataract.
|How to cite this article:|
Sihota R, Mohan M, Angra S K, Mathur R L. Clinico-biochemical study of experimental complicated cataracts. Indian J Ophthalmol 1991;39:148-50
| Introduction|| |
In the majority of cases of complicated cataract, it is not possible to demonstrate a definitive pathology. Among cataractous patients at our hospital, stool examination was positive for intestinal pathogens in 48.57 of children with cataracts of unknown etiology. However, in the group of children with a known cause of the cataract, positive stool examinations were present in only 23.3%. We, therefore, felt that toxins liberated by organisms present in the body could act on the lens directly, or indirectly through structures such as the uvea, thereby causing formation of complicated cataracts.
Most of the patients studied by us showed no evidence of any ocular pathology in the other eye preoperatively or in the affected eye after surgery. Such cases, may be due to a subclinical ocular or systemic disease process liberating toxins.
Preliminary work had shown the occurrence of posterior subcapsular opacities in rabbit eyes, akin to complicated cataract, when injected suprachoroidally, not systemically with E. coli endotoxin. Therefore, further work was undertaken to delineate the progressive biochemical changes in the lens after intraocular injections of E. coli endotoxins and to correlate them with clinical observations.
| Material and methods|| |
Albino rabbits (New Zealand) weighing 1.5-2 kg. were examined by ophthalmoscopy and biomicroscopy to rule out any preexisting pathology. The animals were then anaesthetized with I.V. Nembutal 20-30 mg/Kg body weight. The conjunctiva was incised at the limbus and then undermined approximately 1 cm. to expose bare sclera. Using a tuberculin syringe and a 26G needle, 0.1 ml of toxin was injected into the suprachoroidal space in the right eye thrice at weekly intervals. The left eye was injected with a similar volume of normal saline, at the same instance.
Ophthalmoscopic and biomicroscopic observations were made on all the animals twice a week. The animals were killed by air embolism at 3, 4, 5 and 6 weeks after the first injection of E. coli endotoxin. Aqueous humour was drawn with a tuberculin syringe for analysis and the lenses were dissected out and placed in vials at 70 osub C, till used for analysis. The tissues, were analysed for glutathione by the method of Grunnert and Philips, 1950, ascorbic acid by that of Roe and Kuether, 1943 and proteins by the method of Lowry et al, 1951.
A known enteropathogenic strain of E. coli (ST 3) was subcultured on McConkey agar media and then suspended in normal saline. This suspension was kept in a water bath at 100 osub C for 30 min., cooled and centrifuged for 15 minutes at 10,000 `g'. The supernatant was dialysed and diluted with normal saline to a protein content of 5 ug/100 ul.
| Observations|| |
Clinically it was observed that the reaction at the site of injection of the toxin was, in all cases, associated with a mild local congestion and serous discharge. About half of the experimental eyes revealed fine keratic precipitates and aqueous flare, while all but two had a vitreous haze, the intensity of which correlated with the earlier onset of cataractous changes and greater final density of the cataract. No definite areas of active or healed choroiditis were seen, although later, histopathology revealed an inflammatory response in the choroid. In the lens, at first, there was a faint posterior subcapsular haze appearing commonly, i.e. in 66.6% of eyes, at the 4th week of observation, which developed into a definite granular opacity in a further seven to ten days [Figure 1]. On follow up, this irregular opacity spread both peripherally and axially [Figure 2]. Cataractous changes of varying densities were present in a total 79.2% of eyes injected with toxin, but even at the 6th week of observation only a small fraction of the posterior subcapsular area in the lens was involved.
We would like to emphasise that the cataractous changes were present in 66.6% of the eye at the 4th week of observation and the density of the changes as well as the number of eyes affected, increased gradually upto the 6th week, when 79.2% of eyes had a definable cataract.
The biochemical changes were evaluated from the 3rd week of observation, i.e. a week after the last injection of toxin [Table - 1]. The 3rd and 4th weeks were the period, at which clinically observable cataractous changes were noted.
Water soluble proteins, expressed on the wet weight basis of the lens, formed the largest fraction in both control and experimental eyes. The pool of water soluble proteins did not show a significant change in lenses of the experimental group when compared with control values, at any time.
The content of urea soluble proteins in the experimental eyes increased progressively at the 4th, 5th and 6th weeks respectively. Urea insoluble proteins in the lenses of the control group showed that this constitutes 1.4% of the total proteins. This fraction increased by 35.73% in experimental eyes at the 4th week of observation. The values for the 3rd, 5th and 6th weeks were also above normal, as seen in [Table - 1].
Glutathione levels in the lens decreased to 0.86 mg/g wet weight of the lens in the 3rd week, as compared to a control value of 1.33± 0.33 mg/g wet weight. The values were also decreased, at the 4th, 5th and 6th week, by 24.06, 21.05% and 21.05% respectively.
The ascorbic acid values in the aqueous and lens did not show a definite trend in the changes observed. In the Ienses of the experimental groups ascorbic acid ranged from 0.02±0.01 to 0.04± 0.01 mg/g wet weight, while the control value was 0.03±0.01 mg/g.
| Discussion|| |
In the majority of complicated cataracts, the presumed pathogenesis is a derangement of the metabolism of the lens by an interference with its permeability, or the diffusion into it, of toxins either from an inflammatory focus or from products of degeneration caused by disease. Owing to the thinness of the posterior capsule and its lack of a supporting epithelial barrier, the earliest clinical changes were typically seen in the region of the posterior pole.
In the present investigation we used the suprachoroidal route for injections of E. coli endotoxin to study the effect of the presence of toxins in the local circulation on lens transparency as the frequency of occurrence of cataractous changes is very low with systemically injected toxins. Moreover it is known that toxins in systemic circulation, enter the eye through the choroidal vessels and suprachoroidal injections are the nearest experimental approach to this effect. It is clear from the clinical observations that after three weekly injections of 0.1 ml of toxin, the initial opacities appear between 4-6 weeks of the 1st injection, a complicated cataract can be produced [Figure 1][Figure 2].
The biochemical observations suggest a moderate change in the pattern of various classes of proteins [Table - 1]. It is observed that water soluble proteins are not appreciably altered with cataractogenesis, while the urea soluble and insoluble proteins tend to increase. However, the changes at most points of observation are not statistically significant. This is due to the fact that only a small part of the lens, the posterior subcapsular region, undergoes opacification while the majority of lens fibers remain clear. Thus the biochemical alterations caused during opacification were masked by the larger contribution of the normal parts of the lens. The glutathione level is slightly decreased in the lenses of the toxin treated groups, while the ascorbic acid levels remain unchanged in the aqueous and lens of different groups.
Testa et a1 , studying experimental anaphylactic uveitis came to the conclusion that uveitis, seems to be an unmasking factor for the lens protein sulfhydryl groups. An increase in the reactivity of the protein sulfhydryl groups together with a significant drop in the glutathione level were the first changes that occur in the lenses of eyes with mild uveitis, which still maintained lens transparency. Oxidation of the gamma crystalline fraction and the insoluble protein was found to occur some weeks later when loss of transparency was observed. Further studies along similar lines by Auricchio et al , attributed the effect of uveitis to a breakdown of hydrogen bonds which usually mask the protein sulfhydryl groups. Barber  postulated that this exposure allowed oxidation of -SH groups and the rearrangement and realignment of alpha, beta and gamma crystalline to insoluble proteins. These studies were, however thought to be remote from clinical situations. Shimizu  successfully produced rapidly progressing lenticular opacities by the intravitreal injection of Shigella flexneri endotoxin in mice. The lenticular changes were initially confined to the posterior subcapsular regions.
We utilized the suprachoroidal approach as it was more physiological, the choroid being a common route of entry for intraocular pathogens and toxins. The biochemical changes observed, portray an increased oxidative activity in the lens, evidenced by a fall in glutathione concentration, and consequent reorientation of the protein structure to increase insoluble proteins, and form a cataract.
| References|| |
Angra. S.K., Kunnathur. S., Mathur, R.L., Mohan, M.: Effect of endotoxin on rabbit lens. Ind. J. Ophthal. 29:91-96, 1981.
Testa. M.. Fiore. C. Bocci, N., Calabro. S. : Effect of oxidation of sulfhydryl groups on lens proteins. Exp. Eye Res. 7: 276-290. 1965.
Testa, M., Fiore, C.. Bocci, N., Calabro. S., Auricchio, G. : Influence of experimental uveitis on the rabbit lens. Exp. Eye Res. 7: 473-480, 1968.
Auricchio, G., Bocci, N., Daniele, S., Fiore, S.: Early modifications of the soluble lens proteins in the course of experimental uveitis: Ophthalmologica, 15: 452-456, 1965.
Barber.G. W. : Human cataractogenesis: A review Exp. Eye Res. 16: 85-94, 1973.
Shimizn, M. : Folia ophthalmol Jap. 29: 562-564, 1978.
[Table - 1]