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Year : 1992  |  Volume : 40  |  Issue : 2  |  Page : 53-55

Conjunctival and corneal peroxidases in vitamin A deficiency

Department of Pathology, Nizam's Institute of Medical Sciences, Punjagutta, Hyderabad-500 482, India

Correspondence Address:
K S Ratnakar
Department of Pathology, Nizam's Institute of Medical Sciences, Punjagutta, Hyderabad-500 482
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Source of Support: None, Conflict of Interest: None

PMID: 1452283

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Xerophthalmia is a commonly encountered nutritional disorder that affects the growing population of the world. Conjunctival and corneal epithelial cells contain peroxidase enzyme. In experimentally induced Vitamin A deficiency conjunctival and corneal peroxidases are markedly lowered indicating direct or indirect relation of Vitamin A to epithelial functional integrity.

How to cite this article:
Ratnakar K S. Conjunctival and corneal peroxidases in vitamin A deficiency. Indian J Ophthalmol 1992;40:53-5

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Ratnakar K S. Conjunctival and corneal peroxidases in vitamin A deficiency. Indian J Ophthalmol [serial online] 1992 [cited 2023 Dec 8];40:53-5. Available from: https://journals.lww.com/ijo/pages/default.aspx/text.asp?1992/40/2/53/24403

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  Introduction Top

Vitamin A deficiency, commonly known as xeroph­thalmia, is a frequently encountered ocular morbid condition of the developing world [1]. It primarily affects young growing children with deleterious effects on several organs, in addition to the eye. Clinical and experimental studies revealed that Vitamin A deficien­cy results in retardation of growth, impairment of vision, corneal and conjunctival dryness and depressed immune functions [2],[3],[4]. The influence of vitamin A on cellular kinetic and metabolic pathways, has been gaining importance in recent years. Inter­action of vitamin A and iron on erythropoiesis is current in the literature [5]. Clinical studies, recently, have shown dramatic reduction in size and number of Bitot's spots in retinol deficient children treated with combination of vitamin A and iron in appropriate dose [6]. These observations indicate the influence of iron and vitamin A in altering the developments of serious epithelial lesions. In this communication, iron containing haem enzyme, peroxidase of conjunctival, corneal is reported in experimental vitamin A deficien­cy.

  Material and methods Top

Weaning male wistar rats obtained from the central animal house were divided into two equal groups (I & II) and fed on retinol deficient and control diets. The composition and methods of feeding are ac­cording to Pirie et al [7]. At the end of four and eight weeks, identical sets - of animals were sacrificed for the histological and peroxidase studies.

The tissues for microscopic examination were fixed in 10% formaldehyde and processed by paraffin technique. The sections were subjected to H & E and PAS stains. For peroxidase study, the corneas and conjunctiva, of age and weight matched, animals were immediately transferred after sacrifice to ice cold Tris-Hcl buffer PH 8.5. The tissues were weighed and homogenised in 3m1 of the Tris-Hcl buffer. The mixture was allowed to stand for 3 hours at 4° C at the end of which it was centrifuged for 1 hour at 20,000 RPM 0° C. 1 ml of supernatant was transferred into a curette containing, Tris-Hcl buffer to which 0.05 ml of 0.02% H202 followed by 0.05 ml of 0.1 DAB were added. A control curette containing tissue supernatant without the last two reagents was also included in the study. Both the curettes were incubated for 10 minutes. The optical density was read at 360 mu for corneas and 338 mu for conjunctiva by spectrophotometer at intervals of 10 minutes for 30 minutes at 37° to observe the rate of reaction. In addition, DNA and RNA of the tissues were es­timated [8]. For the retinol estimations, blood samples were obtained from retro-orbital plexus of veins. Vitamin A was measured by Carr Price reaction [9].

  Observations Top

The retinol deficient rats grew slowly [Table - 1] com­pared to the controls. There was periocular porphyrin pigmentation [Figure - 1] observed in deficient animals at the end of four weeks. Cornea of vitamin A deprived rats showed superficial vascularisation by 4 weeks. The conjunctiva of the same animals exhibited loss of- luster and dryness at 4 and 8 weeks respectively.

  Histopathology Top

The conjunctiva of the deficient animals at 4 weeks showed atrophy of epithelium with paucity of goblet [Figure - 2]. The animals fed on deficient diet for 8 weeks showed keratinizing metaplasia with absence of mucus secreting cells. The subepithelial tissues contained scant lymphoid aggregates in the same set of animals.

The cornea of the retinol deficient animals showed attenuation of epithelial cells layer [Figure - 3]. The Bowman's layer was interrupted by invading vascular channels. There were few neutrophils and mononuclears accompanying the blood vessels in the same areas. The stroma and endothelium did not reveal any significant abnormality by routine histologi­cal examination at 4 and 8 weeks intervals in both deficient and control animals. The plasma retinol levels are given in [Table - 1].

  Peroxidase activity Top

The peroxidase activity was measured in pooled conjunctival and corneal tissues and expressed as optical density changes/min/gm of D N A. The max­imum activity in the conjunctiva was attained at 10 minutes and remained constant up to 30 minutes. The corneal activity, however showed a linear increase from 10 min to 30 minutes after which the activity declined. The readings were therefore taken between 10 and 30 minutes. There was significant reduction in conjunctival peroxidase activity at both 4 and 8 weeks. The corneal peroxidase was similarly low in retinol deficient rats. The results are given in [Table - 2].

  Discussion Top

The haem containing enzyme, peroxidase is widely distributed in several mammalian tissues which in­clude lactoperoxidase (salivary, lacrimal and mam­mary glands), myeloperoxidase (granulocytes), thyroid peroxidase, gluotathione peroxidase (erythrocytes). Recently peroxidase activity has also been demon­strated in conjunctival and corneal tissues [10]. Iwata et al (1976) [10] observed that goblet cells, in addition to mucin production, also show peroxidase activity. The corneal epithelial cells have also been shown to exhibit peroxidase activity. Marked reduction in the peroxidase activity of the corneal and conjunctival tissues in vitamin A deficient rats has been reported in the present investigation.

Vitamin A deficiency results in keratinizing metaplasia with loss of goblet cells [ 2 ] . The clinical xerosis is always accompanied by these morphological changes. The fall in peroxidase activity may therefore be due to loss of goblet cells in retinol deficiency. Despite lack of goblet cells, corneal epithelial cells have been shown to possess glycoprotein synthesis. Similar cells, like in conjunctiva may also be a source of peroxidase. There is always some degree of attenua­tion of cell layers in vitamin A deficiency. The reduction in cell number and functions in vitamin A deficiency may be responsible for decreased peroxidase activity.

Recently it has been demonstrated that iron transport is interfered in vitamin A deficient subjects, resulting in anaemia. It is possible that vitamin A deficiency may also produce reduction in iron containing en­zymes of the body secondarily. This area needs to be explored further.

  References Top

WHO. Chronicle.Nutrition a review of WHO programme 1972. 26, 160-164.  Back to cited text no. 1
Mc Laren D S : Malnutrition and Eye. Academic pies New York. 1965.  Back to cited text no. 2
Krishnan S. Bhuyan U N Talwar G P and Ramalingaswamy V The effect of Vitamin and Protein calorie undernutrition on the immune responses. Immunology 1974. 27:383-387.  Back to cited text no. 3
Maddy K H: Vitamin A in animal nutrition . Wayandotte Corporation, New York. 1978. P 12  Back to cited text no. 4
Mejia L A and Arroyave G Hypnvitaminosis A and its effect on haemotopoiesis and iron metabolism, Xerophthalmia club bulletin. 1979. 17  Back to cited text no. 5
Ratnakar K S. Dhingra K and Manthalkar Rao S: Influence of iron and vitamin A on the fate of Bilot's spots and xerosis . In Prep  Back to cited text no. 6
Pirie A. Werb Z and Burleish M C Collagenase and other proteases in the cornea of retinol deficient Brit. .J Nutr. 1975. 34:297.  Back to cited text no. 7
Webb J M and Levy 11 P, New developments in the chemical determination of nucleic acids In methods of Biochemical analysis Vol. VI Ed. David Glick 1958. P-1-30. Interscience publishers- Inc. New York.  Back to cited text no. 8
Varley H, Gowenlock A 1-1 and Boll M: Practical clinical chemistry Vol, William Heinemann Medical Books; Ltd. London 1976. p.210.  Back to cited text no. 9
Iwata T Cytochemical studies and endogenous peroxidase in con­junctival and corneal epitheliall cell;. Invest. Opth' 1976, 15:297-301.  Back to cited text no. 10


  [Figure - 1], [Figure - 2], [Figure - 3]

  [Table - 1], [Table - 2]


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