The alpha and gamma crystallin content in aqueous humor of eyes with clear lenses and with cataracts

doi:10.1016/0014-4835(79)90047-2

Experimental Eye Research

Volume 28, Issue 5, May 1979, Pages 601-610

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Abstract

Specific radioimmunoassays were used to measure the concentration of α- and γ-crystallins in human aqueous humor. It was demonstrated that these crystallins are normally present in aqueous humor from healthy eyes. The crystallin concentration did not seem to increase with age. The normal upper limit for the α-crystallin concentration was found to be 10 ng/ml and for the γ-crystallin concentration 60 ng/ml. In the aqueous humor of eyes with cortical cataract the concentration of both crystallins was increased. With nuclear cataracts the α-crystallin concentration was increased while the γ-crystallin concentration was decreased. Experiments in rabbits showed that the crystallins in the aqueous humor left the anterior chamber at the same rate as the aqueous bulk flow. The demonstration of lens crystallins in the aqueous humor is compatible with the hypothesis that they leak from the lens.

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Cited by (44)

The Proteome of Cataract Markers: Focus on Crystallins
2018, Advances in Clinical Chemistry
Citation Excerpt :
Several publications have also reported on the proteome of cataract markers in the aqueous humor. Sandberg et al. reported the α- and γ-crystallin content in the aqueous humor of eyes with clear lenses and with cataract as early as 1979 [76]. In a study by Fujiwara, the possibility of soluble protein leakage through the capsule into the aqueous humor was studied.
Cataract is a major cause of blindness worldwide. It is characterized by lens opacification and is accompanied by extensive posttranslational modifications (PTMs) in various proteins. PTMs play an essential role in lens opacification. Several PTMs have been described in proteins isolated from relatively old human lenses, including phosphorylation, deamidation, racemization, truncation, acetylation, and methylation.
An overwhelming majority of previous cataract proteomic studies have exclusively focused on crystallin proteins, which are the most abundant proteome components of the lens. To investigate the proteome of cataract markers, this chapter focuses on the proteomic research on the functional relevance of the major PTMs in crystallins of human cataractous lenses. Elucidating the role of these modifications in cataract formation has been a challenging task because they are among the most difficult PTMs to study analytically. The proteomic status of some amides presents similar properties in normal aged and cataractous lenses, whereas some may undergo greater PTMs in cataract. Therefore, it is of great importance to review the current proteomic research on crystallins, the major protein markers in different types of cataract, to elucidate the pathogenesis of this major human-blinding condition.
Secreted leukocyte protease inhibitor is present in aqueous humours from cataracts and other eye pathologies
2006, Experimental Eye Research
Previous studies identified serine, cysteine and metalloproteases in normal aqueous humours (AH) and suggested that a balance between proteases and their inhibitors may play a role in the modulation of the AH outflow. We aimed to determine whether secretory leukocyte protease inhibitor (SLPI), a serine protease inhibitor, is present in AH of patients with cataract and other eye pathologies. AH was collected from 117 cataract patients of which 55 were diagnosed with more when one eye disease: cataract only (n=62), pseudoexfoliation (PEX) (n=26), glaucoma (n=6), diabetes retinopathy (n=4), iritis–uveitis (n=4) and macular degeneration (n=28). The total protein in AH was determined by a Bradford assay and SLPI was analyzed by Western blot and ELISA methods. The average concentration of total protein and SLPI in AH samples was 160±15 μg/ml (n=117, ±SEM) and 500±94 pg/ml (n=105), respectively. The cataract patients with additional eye disease(s) showed higher protein levels (201±35 μg/ml) than cataract (controls) (128±31 μg/ml), P<0.01. It is noteworthy that no correlation was found between SLPI and the total protein concentrations in AH, but SLPI was positively correlated with age (r=0.2, P<0.05). No statistical difference in SLPI levels was found between controls (cataract) and other pathologies, while patients with iritis/uveitis had higher SLPI levels compared to those with diabetes (P<0.05). We show here for the first time that SLPI is present in AH and may play a role as well as serve as a marker in pathological states.
Micropurification of β- and γ-crystallins from rabbit aqueous humor
1999, International Journal of Biological Macromolecules
Soluble crystallins are normally present in the aqueous humor, originating from the lens, and their concentration may increase in certain conditions such as cataract, possibly contributing to aqueous outflow pathway obstruction, leading to glaucoma. Whether the stability and the tendency of aqueous crystallins to aggregate are different in patients with certain forms of open-angle glaucoma has not so far been established, mainly due to the lack of a suitable purification procedure from this fluid in which crystallins are present at very low concentration together with dozens of other proteins. About 4 μg each of β- and γ-crystallins were obtained from 20 ml of rabbit aqueous humor by C₈ reversed-phase high-performance liquid chromatography (HPLC) and high-performance electrophoresis chromatography (HPEC). The identity of the proteins was confirmed by amino acid analysis following sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and electrophoretic blotting onto polyvinylidene fluoride membranes, with or without previous digestion with Staphylococcus aureus protease V₈.
Failure to withstand oxidative stress induced by phospholipid hydroperoxides as a possible cause of the lens opacities in systemic diseases and ageing
1996, Biochimica et Biophysica Acta - Molecular Basis of Disease
Lipid peroxidation (LPO) is a causative factor of cataract. The increased concentrations of primary molecular LPO products (diene conjugates, lipid hydroperoxides) and end fluorescent LPO products, were detected in the lipid moieties of the aqueous humor samples obtained from patients with senile and complicated cataracts as compared to normal donors. The degrees of lens clouding were assessed quantitatively by measuring the optical density indices and areas of equidensities using digital image analysis. Human cataractous lenses showed decreased activity of glutathione peroxidase (GPX, catalyzing reduction of organic hydroperoxides including hydroperoxides of lipids). The apparent K_m for tert-butylhydroperoxide was 0.434 mM for human normal and cataractous lens GPX. When lenses were exposed for 1 h at 37°C to linoleic acid hydroperoxide (LOOH, 0.5 mM) or egg phosphatidyl-choline hydroperoxide (PLOOH, 1 μmol per 112 μmol of phospholipid) in liposomes suspended in the incubation medium, normal, immature and mature human cataractous lenses showed a significant loss in the residual content of liberated LOOH to 62%, 38% or 17%, correspondingly, but little or no reduction was observed with PLOOH in liposomal membranes. Human, rabbit or mice transparent or immature cataractous lenses induced significantly more absorbance changes in conjugated diene, iodometric and TBA-reactive substance measurements when incubated with liposomal membranes which were decreased in the presence of free radical scavengers and antioxidant enzymes (EDTA, SOD, l-carnosine, chelated iron, catalase). Injection into the vitreous body of the rabbit eye of a suspension of liposomes prepared from phospholipids containing LPO products induced the development of posterior subcapsular cataract. Saturated liposomes did not cause clouding of the lens. This modelling of cataract was accompanied by accumulation of fluorescing LPO products in the vitreous body, aqueous humor and the lens and also by a fall in the concentration of GSH in the lens. The peroxidative damage to the lens cell membranes and biomolecules induced in the lack of reductive detoxification of phospholipid hydroperoxides is proposed as the triggering mechanism of cataractogenesis.
Distribution and type of morphological damage in human nuclear age-related cataracts
1996, Experimental Eye Research
The distribution and type of fiber cell damage was evaluated in human age-related nuclear cataracts and in aged normal (non-cataractous) lenses. Ten age-related nuclear cataracts (53 to 89 years old) and four normal lenses (59 to 67 years old) were examined by electron microscopy of fixed Vibratome sections. Images from the adult, juvenile, fetal and embryonic nuclear regions were compared. Each cataractous lens contained a central region of increased light scattering which involved the embryonic and fetal regions with progressively less involvement in the juvenile and adult nuclear regions. Some damaged fiber cells were observed in all specimens, although damage was minor and infrequent in the normal lenses. Degeneration of single or groups of fiber cells was noted in all the adult nuclei of the cataractous lenses, becoming less frequent in the juvenile nuclei. The types of damage included localized voids, multilamellar membrane aggregates, globular bodies, enlarged cells and regions of highly convoluted membranes. The fetal and embryonic nuclei of the cataractous lenses exhibited rare and minor morphological defects, and were virtually identical to the equivalent regions of the normal aged lenses. Examination of cell interfaces in opaque regions of cataractous lenses revealed that the oldest fiber cells sustained apparent membrane loss. Extracellular spaces in the embryonic, fetal and juvenile regions of the cataractous lenses often contained dense deposits, presumably cytoplasmic material lost from adjacent fibers. The results indicate that the region of greatest nuclear opacity, located in the lens center, does not contain any significant cellular damage. This suggests that older fiber cells respond differently to pathological and senescent changes than younger cells made after fetal development. The observed loss of membranes and cytoplasmic material from the oldest fiber cells may be a contributory mechanism in the formation of age-related human nuclear cataracts.
Cataractogenesis in transgenic mice containing the HIV-1 protease linked to the lens αA-crystallin promoter
1996, Journal of Biological Chemistry
Several lines of transgenic mice were generated with either active or inactive forms of the human immunodeficiency virus type 1 (HIV-1) protease gene under the control of the mouse lens αA-crystallin promoter. Mice bearing the inactive protease coding sequence displayed no gross abnormalities in the lens, while mice with the active protease developed time-dependent bilateral cataracts. One line, TG₆₁, developed cataracts in utero while the second line, TG₇₂, developed cataracts postnatally. TG₆₁ mice, homozygous for the transgene, developed severe microphthalmia and were significantly smaller than the control mice at postnatal day 30. two-dimensional-polyacrylamide gel electrophoresis analysis of the protein profiles of TG₇₂ and TG₆₁ lenses revealed extensive modifications in the lens crystallins. Proteolysis in the homozygous TG₇₂ mouse lenses began at postnatal day 20 with the disappearance or partial loss of βB1-, βB3-, and βA3-crystallins and the appearance of crystallin fragments. Protein leakage and the gradual breakdown of cytoskeletal elements also occurred. In contrast, the opacification of the homozygous TG₆₁ lenses appeared to have been influenced by differentiation and developmental processes. It apppears that HIV-1 protease expression activates other proteases, and these enzymes, in concert with the HIV-1 protease, are responsible for the protein modifications that eventually result in the opacification of the lens.

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Experimental Eye Research

Abstract

References (15)

An investigation into the loss of proteins of low molecular size from the lens in senile cataract

Exp. Eye Res.

The α-crystallin content of aqueous humor in cortical, nuclear and complicated cataracts

Exp. Eye Res.

Radioimmunoassay of sheep alpha and gamma crystallins. I. ¹²⁵I labelling of sheep gamma crystallin. Characterization of the assays in detecting homologous and heterologous lens crystallins

Exp. Eye Res.

Radioimmunoassay of sheep alpha and gamma crystallins. II. Organ specificity of lens crystallins

Exp. Eye Res.

Characterization of human lens antigens by crossed immunoelectrophoresis

Exp. Eye Res.

The human lens. II. Some observations on cataracts removed in Oxford, England

Exp. Eye Res.

Influence of cultivation conditions on the production of extracellular proteins by Staphylococcus aureus

Acta Pathol. Microbiol. Scand. (B)

Cited by (44)

The Proteome of Cataract Markers: Focus on Crystallins

Secreted leukocyte protease inhibitor is present in aqueous humours from cataracts and other eye pathologies

Micropurification of β- and γ-crystallins from rabbit aqueous humor

Failure to withstand oxidative stress induced by phospholipid hydroperoxides as a possible cause of the lens opacities in systemic diseases and ageing

Distribution and type of morphological damage in human nuclear age-related cataracts

Cataractogenesis in transgenic mice containing the HIV-1 protease linked to the lens αA-crystallin promoter