ACTA 0 P HT HA LM0 LOG ICA
70 (1992) 839-841
S H 0 R T C 0 M M U N I CAT I 0N
Role of trace elements in senile cataract V. K. Srivastava, N. Varshney and D. C. Pandey Department of Chemistry, University of Gorakhpur, Gorakhpur, India
Abstract. The contents of copper, zinc and protein were determined in cortex and nucleus sections of cortical and nuclear cataracts. The results show that copper and zinc content increased in both types of cataracts, although more in the cortex than nucleus sections of the cataractous lenses. Key words: senile cataract - copper - zinc - cortex- nucleus.
The human lens contains a number of elements which constitute only 0.2% of inorganic material in the tissue (Underwood 1977). The implication of these elements in senile cataract formation is currently an area of intensive research. Some of them may have vital roles in the processes of chemical reactions (Shlopak 1962) which may be responsible for life phenomenon of the lens. In recent times, due to increased industrialization,studies of environmental pollution by trace elements and their effect on human health have shown a possibility that trace elements may initiate cataractogenic processes (Murata & Taura 1975). It is thus speculated that a possible way of obtaining information about the significance of trace elements in relation to cataractogenesis, is to gain knowledge of their exact loci and association within the lens. Amongst a number of trace elements which are present in the lens, copper and zinc have recently attracted attention, as besides being a redox catalyst, they are important constituentsof various metalloenzymes (Hock & Vallee 1958; Li & Vallee 1958; Nielsen 1984)which have their intricate relationship with senile cataract formation. The purpose of the present investigation was thus to determine the concentrations of copper and zinc in the lens of cortical and nuclear types of
cataracts. It is expected that a knowledge of their concentration levels may show a relationship with the extent of coloration in human cataractous lenses.
Materials and Methods Normal lenses Normal lenses were obtained from Eye Bank PostGraduate Institute of Medical Education and Research Chandigarh. Cataractous lenses Lenses were extracted intracapsularly from patients admitted in Eye Hospital, Gorakhpur. The age of the patients was recorded from their personal history. Clinical examinations were done in detail to determine the type of cataract. Patients having any obvious metabolic or nutritional disturbances as evidenced by routine biochemical tests were not included in this study. Classification of lenses The lenses were classified on the basis of slit-lamp examination and visual acuity into one of the following groups: 1) predominantly cortical; and 2 ) predominantly nuclear. Dissection of lenses. Frozen lenses were divided into cortex and nucleus sections using a trephine with bore of 7 n m diameter. Estimation of protein Lenses were individually and separately homogenized in 0.1 M Tris buffer (pH 7.4). The homoge-
839
S1. No.
Normal lens
Parameters
nate was processed for the determination of soluble and insoluble proteins by Lowry method (Lowry et al. 1951). Estimation of copper and zinc
Approximately 500-600 mg lyophilized samples were digested separately with 3.0 ml digestion mixture containing concentrated nitric acid and perchloric acid in the ratio of (5:l) for half an hour at room temperature. The digested samples were then placed on sand bath and evaporated to dryness. The residue so obtained was dissolved in 3.0 mll0 mM nitric acid, mixed well and left overnight at room temperature. A blank solution 3.0 ml acid mixture was also processed in the same manner. The metal content was then determined by Atomic Absorption Spectrophotometer (StanojevicPaovic et al. 1987). Sigma Standard was used for calibration purpose. All measurementswere made by using GBC-902 Atomic Absorption Spectrophotometer with wavelengths of 324.7 (Cu), 213.9 (%) and slit width of 0.5 nm.
Results The results showed that the concentration of copper and zinc increased in both cortical and nuclear
Discussion There has been a good amount of controversy regarding the status of copper and zinc in senile cataractogenesis as both increased and decreased levels have earlier been reported (Swanson & Truesdale 1971; Racz & Ordogh 1977; Racz & Erdohelyi 1988). The higher level of copper in the lens nucleus shows that this is not associated with the more metabolically active area of the lens. It has,
Cortical cataract
Nuclear cataract
Parameters Cortex
840
Nuclear cataract
types. The increase was more significant with copper and was more pronounced with nuclear cataract. Such an increase in concentration was signified by greater insolubilization of soluble protein (Table 1). Although there was no significant difference in the level of copper and zinc in cortex and nucleus sections of the normal lens, their levels were found to be of greater magnitude in cortex and nucleus sections of cataractous lenses. Interestingly enough, the level of zinc in cortical and nuclear cataract was 45% and 49% higher in the cortex section compared to the nucleus section; the level of copper in cortical and nuclear cataract was 48% and 43% higher in nucleus section than cortex section (Table 2). The other parameters followed the same pattern as observed in whole lenses.
Normal lens S1. No.
Cortical cataract
Nucleus
Cortex
Nucleus
Cortex
Nucleus
however, been reported that the nucleus section of the lens has a lower level of copper (Shlopak 1962). On the other hand, the presence of a higher concentration of zinc in the cortex section of the lens signifies its differential behaviour as compared to copper (Racz & Ordogh 1977). It is suggested that the presence of elevated levels of copper and zinc may damage the lens through different pathways. Their elevated levels might lead to oxidation of sulphydryl groups (Srivastava & A n s a r i 1980)which will damage the permeability of the membrane eventually leading to the formationof cataract, and there also appears to be a link with the presence of HMW proteins which are considered to be precursors of insoluble protein aggregate. However, excess level of zinc in the lens may induce formation of metallothionein, the low molecular weight protein (Reinhold 1984). The increased level of copper and zinc in the cataractous lenses may be due to an outcome of their inherent biological variability. It is not understood whether their high levels in cataractous lenses are a result of the presence of metalloproteins (Srivastava et al. 1988) or from the association of metals with small molecules. It is worth mentioning that zinc from the aqueous environment is readily taken up into ocular tissues. This is exemplified by the observation that the cortex region, being more active metabolically, accumulates more zinc than the nucleus section of the lens. Such an observation has also been found in calf lenses where zinc was found to be higher in the cortex than nucleus section of the lens (Murata& Taura 1975).Unfortunately, till now, there are no systemicinvestigations regarding the level of copper and zinc and their implication in different types of cataract. Besides, the role of zinc in the regulation of lens transparency is also not known. It is possible that the variation in the content of copper and zinc in the cortex and nucleus sections of cataractous lenses possibly reflects a build up of metal associated molecules that accumulate with age and increased polution.
Acknowledgments The authors are thankful to the Indian Council of Medical Research, New Delhi for providing financial assistance.
References Hock F L & Vallee B L (1958):The biochemical and nutritional role of trace elements. In: Lamb C A, Bentley 0 G & Beattie J M (eds). ‘Trace elements’. Proc Cont Wooster Ohio, Academic Press, New York. p 337. Li T K & Vallee B L (1958): Section B the biochemical and nutritional role of trace elements. In: Lamb C A, Bentley 0 G & Beattie J M (eds). ‘Trace elements’. Proc Cont Wooster Ohio, Academic Press, New York. p 372. Lowry D H, Rosebrough N J, Farr A L & Randall R L (1951):Protein measurement with the folin phenol reagent. J Biol Chem 193: 265. Murata & Taura Y (1975): Study of trace metallic elements in the lens. Ophthalmic Res ’I: 8. Nielsen F H (1984): Ultratrace elements in nutrition. Annu Rev Nutr 4: 21. Racz P & Erdohelyi A (1988): Cadmium, Lead and Copper concentrations in normal and senile Cataractous human lenses. Ophthalmic Res 20: 10. Racz P & Ordogh M (1977): Investigations in trace elements in normal and senile cataractous lenses. Graefes Arch Klin Exp Ophthalmol204: 67. Reinhold J G (1984): The phylate hypothesis. Proc Int Congr Nutr. p 98. Shlopak T V (1964): Normal and pathological metabolism of trace elements in the lens. Chem Abs 60: 9755b. Srivastava S K, Ansari N H et al. (1980): Role of sulphydrylds and disulphides in lens and red cell metabolism. Elsevier North Holland. Srivastava V K, Chaturvedi N, Garg M, Afaq Z & Pandey D C (1988): Copper and zinc in human senile cataract. Curr Sci 57: 1288. Stanojevic Paovic A, Hristic V, Caperlovic M, Javonovic S & Krsmanovic (1987):Macro and Microelements in the Cataractous eye lens. Ophthalmic Res 19: 230. SwansonA & Truesdale A W (1971):Elemental analysisin normal and Cataractous human lens tissue. Biochem Biophys Res Commun 45: 1488. Underwood EJ (1977):Trace elements in human and animal nutrition 3rd Ed Academic Press, New York.
Received on December loth, 1991. Author’s address:
Dr. V. K. Srivastava, Cataract Research Laboratory, Department of Chemistry, University of Gorakhpur, Gorakhpur-273009, India.
841
70 (1992) 839-841
S H 0 R T C 0 M M U N I CAT I 0N
Role of trace elements in senile cataract V. K. Srivastava, N. Varshney and D. C. Pandey Department of Chemistry, University of Gorakhpur, Gorakhpur, India
Abstract. The contents of copper, zinc and protein were determined in cortex and nucleus sections of cortical and nuclear cataracts. The results show that copper and zinc content increased in both types of cataracts, although more in the cortex than nucleus sections of the cataractous lenses. Key words: senile cataract - copper - zinc - cortex- nucleus.
The human lens contains a number of elements which constitute only 0.2% of inorganic material in the tissue (Underwood 1977). The implication of these elements in senile cataract formation is currently an area of intensive research. Some of them may have vital roles in the processes of chemical reactions (Shlopak 1962) which may be responsible for life phenomenon of the lens. In recent times, due to increased industrialization,studies of environmental pollution by trace elements and their effect on human health have shown a possibility that trace elements may initiate cataractogenic processes (Murata & Taura 1975). It is thus speculated that a possible way of obtaining information about the significance of trace elements in relation to cataractogenesis, is to gain knowledge of their exact loci and association within the lens. Amongst a number of trace elements which are present in the lens, copper and zinc have recently attracted attention, as besides being a redox catalyst, they are important constituentsof various metalloenzymes (Hock & Vallee 1958; Li & Vallee 1958; Nielsen 1984)which have their intricate relationship with senile cataract formation. The purpose of the present investigation was thus to determine the concentrations of copper and zinc in the lens of cortical and nuclear types of
cataracts. It is expected that a knowledge of their concentration levels may show a relationship with the extent of coloration in human cataractous lenses.
Materials and Methods Normal lenses Normal lenses were obtained from Eye Bank PostGraduate Institute of Medical Education and Research Chandigarh. Cataractous lenses Lenses were extracted intracapsularly from patients admitted in Eye Hospital, Gorakhpur. The age of the patients was recorded from their personal history. Clinical examinations were done in detail to determine the type of cataract. Patients having any obvious metabolic or nutritional disturbances as evidenced by routine biochemical tests were not included in this study. Classification of lenses The lenses were classified on the basis of slit-lamp examination and visual acuity into one of the following groups: 1) predominantly cortical; and 2 ) predominantly nuclear. Dissection of lenses. Frozen lenses were divided into cortex and nucleus sections using a trephine with bore of 7 n m diameter. Estimation of protein Lenses were individually and separately homogenized in 0.1 M Tris buffer (pH 7.4). The homoge-
839
S1. No.
Normal lens
Parameters
nate was processed for the determination of soluble and insoluble proteins by Lowry method (Lowry et al. 1951). Estimation of copper and zinc
Approximately 500-600 mg lyophilized samples were digested separately with 3.0 ml digestion mixture containing concentrated nitric acid and perchloric acid in the ratio of (5:l) for half an hour at room temperature. The digested samples were then placed on sand bath and evaporated to dryness. The residue so obtained was dissolved in 3.0 mll0 mM nitric acid, mixed well and left overnight at room temperature. A blank solution 3.0 ml acid mixture was also processed in the same manner. The metal content was then determined by Atomic Absorption Spectrophotometer (StanojevicPaovic et al. 1987). Sigma Standard was used for calibration purpose. All measurementswere made by using GBC-902 Atomic Absorption Spectrophotometer with wavelengths of 324.7 (Cu), 213.9 (%) and slit width of 0.5 nm.
Results The results showed that the concentration of copper and zinc increased in both cortical and nuclear
Discussion There has been a good amount of controversy regarding the status of copper and zinc in senile cataractogenesis as both increased and decreased levels have earlier been reported (Swanson & Truesdale 1971; Racz & Ordogh 1977; Racz & Erdohelyi 1988). The higher level of copper in the lens nucleus shows that this is not associated with the more metabolically active area of the lens. It has,
Cortical cataract
Nuclear cataract
Parameters Cortex
840
Nuclear cataract
types. The increase was more significant with copper and was more pronounced with nuclear cataract. Such an increase in concentration was signified by greater insolubilization of soluble protein (Table 1). Although there was no significant difference in the level of copper and zinc in cortex and nucleus sections of the normal lens, their levels were found to be of greater magnitude in cortex and nucleus sections of cataractous lenses. Interestingly enough, the level of zinc in cortical and nuclear cataract was 45% and 49% higher in the cortex section compared to the nucleus section; the level of copper in cortical and nuclear cataract was 48% and 43% higher in nucleus section than cortex section (Table 2). The other parameters followed the same pattern as observed in whole lenses.
Normal lens S1. No.
Cortical cataract
Nucleus
Cortex
Nucleus
Cortex
Nucleus
however, been reported that the nucleus section of the lens has a lower level of copper (Shlopak 1962). On the other hand, the presence of a higher concentration of zinc in the cortex section of the lens signifies its differential behaviour as compared to copper (Racz & Ordogh 1977). It is suggested that the presence of elevated levels of copper and zinc may damage the lens through different pathways. Their elevated levels might lead to oxidation of sulphydryl groups (Srivastava & A n s a r i 1980)which will damage the permeability of the membrane eventually leading to the formationof cataract, and there also appears to be a link with the presence of HMW proteins which are considered to be precursors of insoluble protein aggregate. However, excess level of zinc in the lens may induce formation of metallothionein, the low molecular weight protein (Reinhold 1984). The increased level of copper and zinc in the cataractous lenses may be due to an outcome of their inherent biological variability. It is not understood whether their high levels in cataractous lenses are a result of the presence of metalloproteins (Srivastava et al. 1988) or from the association of metals with small molecules. It is worth mentioning that zinc from the aqueous environment is readily taken up into ocular tissues. This is exemplified by the observation that the cortex region, being more active metabolically, accumulates more zinc than the nucleus section of the lens. Such an observation has also been found in calf lenses where zinc was found to be higher in the cortex than nucleus section of the lens (Murata& Taura 1975).Unfortunately, till now, there are no systemicinvestigations regarding the level of copper and zinc and their implication in different types of cataract. Besides, the role of zinc in the regulation of lens transparency is also not known. It is possible that the variation in the content of copper and zinc in the cortex and nucleus sections of cataractous lenses possibly reflects a build up of metal associated molecules that accumulate with age and increased polution.
Acknowledgments The authors are thankful to the Indian Council of Medical Research, New Delhi for providing financial assistance.
References Hock F L & Vallee B L (1958):The biochemical and nutritional role of trace elements. In: Lamb C A, Bentley 0 G & Beattie J M (eds). ‘Trace elements’. Proc Cont Wooster Ohio, Academic Press, New York. p 337. Li T K & Vallee B L (1958): Section B the biochemical and nutritional role of trace elements. In: Lamb C A, Bentley 0 G & Beattie J M (eds). ‘Trace elements’. Proc Cont Wooster Ohio, Academic Press, New York. p 372. Lowry D H, Rosebrough N J, Farr A L & Randall R L (1951):Protein measurement with the folin phenol reagent. J Biol Chem 193: 265. Murata & Taura Y (1975): Study of trace metallic elements in the lens. Ophthalmic Res ’I: 8. Nielsen F H (1984): Ultratrace elements in nutrition. Annu Rev Nutr 4: 21. Racz P & Erdohelyi A (1988): Cadmium, Lead and Copper concentrations in normal and senile Cataractous human lenses. Ophthalmic Res 20: 10. Racz P & Ordogh M (1977): Investigations in trace elements in normal and senile cataractous lenses. Graefes Arch Klin Exp Ophthalmol204: 67. Reinhold J G (1984): The phylate hypothesis. Proc Int Congr Nutr. p 98. Shlopak T V (1964): Normal and pathological metabolism of trace elements in the lens. Chem Abs 60: 9755b. Srivastava S K, Ansari N H et al. (1980): Role of sulphydrylds and disulphides in lens and red cell metabolism. Elsevier North Holland. Srivastava V K, Chaturvedi N, Garg M, Afaq Z & Pandey D C (1988): Copper and zinc in human senile cataract. Curr Sci 57: 1288. Stanojevic Paovic A, Hristic V, Caperlovic M, Javonovic S & Krsmanovic (1987):Macro and Microelements in the Cataractous eye lens. Ophthalmic Res 19: 230. SwansonA & Truesdale A W (1971):Elemental analysisin normal and Cataractous human lens tissue. Biochem Biophys Res Commun 45: 1488. Underwood EJ (1977):Trace elements in human and animal nutrition 3rd Ed Academic Press, New York.
Received on December loth, 1991. Author’s address:
Dr. V. K. Srivastava, Cataract Research Laboratory, Department of Chemistry, University of Gorakhpur, Gorakhpur-273009, India.
841
