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Biomedical subjects

K R Hightower

Publications and source records attributed to K R Hightower.

At least 19 recordsLinked to original sources
PubMed1995

The role of the lens epithelium in development of UV cataract.

In view of renewed interest in the lens epithelium as the initiation site for cataract development, it seemed timely to review recent studies which appear to establish UV damage in the lens epithelium as the cause of UV cataract. While UV photons can and do interact with lens proteins in the cortex and nucleus, experimental results from cultured lenses and tissue cultured epithelial cells also demonstrate both mutagenic and cytotoxic effects in the epithelium. This minireview examines UV-induced changes in lens physiology that appear to follow epithelial cell damage, including inactivation of critical enzymes of transport and metabolic processes. Changes in membrane function include altered cation transport, increased permeability, and altered biosynthesis. One potential scenario for the propagation of damage from the epithelium to the underlying fiber cells includes calcium elevation, an early event in cataract development and critical to many physiological processes.

Animals↗
PubMed1994

Membrane damage in UV-irradiated lenses.

The purpose of this study was to investigate three possible causes of membrane damage following UV irradiation: photooxidation of membrane thiol (SH) groups, peroxidation of membrane lipids and inhibited synthesis of membrane proteins. Thiol loss was not observed. Thin-layer chromatography showed a four-fold increase in several primary lipid peroxidation products such as hydroperoxyl lipids in the epithelial membrane preparations isolated from irradiated lenses. The formation of new hydroxyl lipid bands not seen in control preparations was also observed in isolated membranes from irradiated lenses. Irradiation in the presence or absence of oxygen produced lipid peroxidation products. Aerobic irradiation produced small, but statistically significant increases in lipid hydroxyls and hydroperoxyls relative to controls. Repair of initial damage might be compromised by the observed 60% reduction in rate of protein synthesis measured in lens membranes following irradiation. Synthesis was affected by means other than depleted potassium or elevated calcium levels.

Animals↗
PubMed1991

Effect of thiol reagents on Ca-ATPase in rabbit lens epithelium.

The inhibitory effects of sulfhydryl reagents on Ca-ATPase activity in the rabbit lens epithelium were assessed. Test compounds used in this study were selected on their ability to cause a calcium increase in cultured lenses. Under conditions in which lenses were cultured in the presence of the test compound, epithelial Ca-ATPase was inhibited markedly by diamide, t-BHP, IAA and slightly by selenite. The findings demonstrated that hydrogen peroxide caused little inhibition of Ca-ATPase in the lens epithelium, both when the intact lens was cultured in the presence of the oxidant or when the epithelial homogenate contained peroxide during the assay of enzyme activity. The study suggests that if a thiol-modifying compound can reach Ca-ATPase or its critical SH groups, inhibition is likely.

Animals↗
PubMed1991

Effect of selenite on epithelium of cultured rabbit lens.

Selenite (Se) cataract in rabbit lenses was investigated in vitro to define target sites of Se that might be involved in calcium elevation and lens opacification. Experiments in which the anterior or the posterior surface of the lens was exposed to Se showed that anterior exposure led to ionic imbalances and opacification in the whole lens. Posterior exposure to Se (1 mM, 2 hr) had no effect. Se treatment (0.1 mM) of epithelial homogenates led to a 56% loss of thiol (SH) groups, and treatment of lenses cultured in Se led to a 22% loss. Experiments to assess the effects of Se on SH groups of Ca-ATPase showed that the transport enzyme was not affected by the poison. To determine whether this negative finding was due to the lack of accessibility of Se for SH sites in an ordered membrane, Ca-ATPase was also assayed in homogenate preparations treated with Se; still no inhibition of Ca-ATPase activity was observed. Therefore, an alternative explanation of calcium elevation was explored. The passive movement of labeled chloride (36Cl) was found to be twice as fast in Se-treated lenses as it was in control lenses. Measurement of the lens voltage indicated an 18-mV depolarization in Se-treated lenses, suggesting that Se increased membrane permeability. All cataractogenic changes that occurred after Se treatment were irreversible-despite intervention with external application of reduced glutathione or cysteine. This finding suggests that irreversible loss of SH groups in lens membranes is important in maintaining ion homeostasis.

Animals↗
PubMed1989

Regional distribution of calcium in alloxan diabetic rabbit lens.

A diabetic rabbit model was developed for investigation of cataractogenesis and other changes in the anterior segment. Rabbits were fasted, injected with 0.7 mg/kg alloxan, fed 1% glucose solution for 24 hrs and returned to a normal diet. Animals showing and maintaining blood glucose of greater than 300 mg% within two days were used in this study. Concomitant with increase in blood glucose was a rise in aqueous humor glucose and osmolality, together with a decrease in ascorbate concentration. Vacuoles and small discrete opacities developed, and in some cases, at longer time periods complete opacity of anterior or posterior aspects was found. Total calcium content of the whole lens increased up to 2-fold, especially after 60 days, and was correlated with a decrease in lens transmittance of a He/Ne laser beam and also with high osmolality of the aqueous humor. Free calcium was six-fold higher in opaque areas than clear areas, and was 100-fold higher in vacuoles. It is suggested that, in addition to the recognized role in sugar cataractogenesis of osmotic stress due to sorbitol accumulation in the lens, changes of intracellular calcium in localized areas of the lens and stresses imposed by changes in aqueous humor osmolality may also be important.

Alloxan↗
PubMed1989

Susceptibility of lens epithelial membrane SH groups to hydrogen peroxide.

Although membrane SH groups are thought to be targets of oxidative insults, no measurement of lens epithelial membrane SH groups following exposure to potentially damaging oxidants has been reported. Here we investigate the effect of hydrogen peroxide, an oxidant found in the aqueous humor, and of p-chloromercuriphenylsulfonic acid (p-chloromecuribenzene-sulfonic acid) (PCMBS), a relatively impermeant sulfhydryl probe, on membrane SH groups and ion homeostasis in cultured lens epithelial cells. Exposure to PCMBS caused a 10% loss of membrane SH groups, an increase in sodium and calcium levels, and a decrease in potassium, but did not affect the intracellular level of glutathione (GSH). After 5 min of exposure to an initial concentration of 1.0 mM hydrogen peroxide, GSH declined from 14.1 mM to 3 mM, there was a 20% loss of membrane SH groups and within 1 hr, potassium declined from 132 to 116 mM. Cells that were exposed to 0.1 or 0.5 mM peroxide did not exhibit significant loss of membrane SH groups and did not show a decrease in GSH comparable to that found in cells treated with 1 mM peroxide. The peroxide induced loss of membrane SH groups and subsequent change in ion homeostasis occurred only when there was a rapid and sustained loss of intracellular glutathione. Thus lens epithelial cell membrane SH groups are not only important in ion regulation but are targets of hydrogen peroxide when the intracellular level of GSH is significantly diminished.

4-Chloromercuribenzenesulfonate↗
PubMed1989

Human lens membrane cation permeability increases with age.

Parallel studies of the ionic balance and membrane permeability characteristics of normal human lenses were carried out in three countries (USA, England and Italy). Similar age-related changes were found in each laboratory. The lens membrane potential and resistance declined markedly with age while internal Na+ and free Ca2+ increased. There was a concomitant stimulation of Na+ and K+ transmembrane fluxes. These data indicate that in the ageing process there is an increasing contribution to membrane ion traffic from a channel, or channels, that permit Na+, K+ and Ca2+ to pass. The increase in permeability coincides exactly with the increase in optical density that occurs in the ageing human lens.

Adult↗
PubMed1989

Effects of selenium on ion homeostasis and transparency in cultured lenses.

Selenium toxicity was investigated in cultured rabbit lenses to provide further information about the role of Ca++ in Se cataract. At a dose of 0.1 mM for 20 hr, Se induces a 10% change in Na levels within 6 hr, a 30% increase after 20 hr, and a three-fold increase within 48 hr of subsequent culture after removal of Se. In contrast, Ca++ levels remained normal throughout the first 24 hr. Only a small, 25% decline in GSH was noted. Not until lenses begin to swell and become noticeably opaque and turbid were Ca++ levels found to be elevated. Thus, at 72 hr, 48 hr following the removal of selenium, Ca++ had increased to a concentration of 0.7 mM. Ca++ accumulation appears to be a consequence of osmotic stress rather than pump inhibition while Na accumulation is a direct consequence of Se-inhibited Na pump.

Animals↗
PubMed1988

p-chloro-mercuriphenyl sulphonate activates a quinine-sensitive potassium conductance in frog lens.

1. The effects of the sulphydryl-complexing reagent p-chloro-mercuriphenyl sulphonate (pCMPS) on membrane voltage and electrical conductance were studied on the isolated frog lens. 2. At low concentrations (0.1-50 microM) pCMPS induced a rapid and graded hyperpolarization of the lens membrane potential which saturated at -97 mV. 3. The lens conductance also showed a graded increase, but the initial changes were apparent only at concentrations above 1 microM. 4. Decreasing the external potassium concentration from 2.5 to 0.5 mM had little effect on the membrane potential in the absence of pCMPS, but increased the voltage from -97 to -110 mV when pCMPS was present. 5. Quinine (300 microM) had no effect when added in control solution, but depolarized the membrane potential and decreased the conductance when added to a pCMPS-treated preparation. 6. These data suggest that pCMPS activates voltage-sensitive potassium channels that are quiescent at the frog resting potential in control solution. 7. At pCMPS concentrations greater than or equal to 100 microM, the initial hyperpolarization is followed by a marked but slow depolarization of the membrane potential and a further increase in lens conductance. These data suggest that non-specific cation channels are activated in this case. 8. Cysteine (5 mM) added to a pCMPS-treated lens leads to a rapid recovery of membrane potential and conductance to near their resting values whether the lens had previously been exposed to low or high concentrations of pCMPS. 9. All of these changes in lens voltage and conductance occurred without apparent alteration in the lens internal sulphydryl content.

4-Chloromercuribenzenesulfonate↗
PubMed1987

Calcium-induced opacification is dependent upon lens pH.

The intracellular pH of a normal lens is 6.8 in the cortex and remains unchanged during culture in media buffered at pH 7.2. Incubation of rabbit lenses in calcium enriched media, either at 24 degrees C or 37 degrees C, results in lens opacification provided that the lens pH remains slightly acidic. Opacities are prevented in cultured lenses with an alkaline interior (pH 7.1-7.3) despite the accumulation of calcium (1.3 mM). The mechanism by which an intracellular pH shift from 6.8 to 7.1 prevents opacification in the presence of excess calcium is not known, but does not appear to depend upon the total level of bound calcium. This study provides the first data that opacification caused by calcium is associated with lens pH.

Animals↗
PubMed1987

Regional distribution of free calcium in selenite cataract: relation to calpain II.

The purpose of this experiment was to assess the roles of free, intracellular calcium and calcium-dependent neutral protease (calpain II, EC.34.22.17) in selenite nuclear cataract. Free calcium ion concentrations within lens nuclear fibers during selenite cataractogenesis increased to 3 microM on day 2 post-injection (clear lens) and to 108 microM at day 4 (nuclear cataract). Calpain II is known to be activated in vitro by calcium levels above 50 microM. Calpain II activity was present in the lens nucleus at time periods preceding formation of selenite cataract. These data suggested that after selenite injection, calpain II was activated by elevated free calcium in the nucleus, and that calpain II-induced proteolysis of nuclear proteins was an important mechanism in selenite cataract. Calpain II levels were also observed to decrease in the nucleus during selenite cataractogenesis, probably due to autolysis. This was supported by the finding that incubation of purified lens calpain II with 100 microM calcium caused partial inactivation of the protease.

Animals↗
PubMed1987

The influence of calcium on glucose metabolism in the rabbit lens.

Results were obtained which demonstrate that calcium accumulation in the rabbit lens may suppress glycolysis, not only by its inhibitory effect on cation transport but by its direct effect on glycolytic enzymes. In lenses cultured in calcium-enriched medium, lactate production declined in proportion to the increase in free and bound levels of calcium. In lens homogenates to which varying amounts of calcium were added, lactate production also decreased. To insure that excess calcium was not simply chelating ATP, homogenates were exposed to calcium and then dialyzed against a calcium-free buffer prior to addition of ATP. Under these conditions, lactate production diminished maximally by approximately 50% as bound calcium increased five-fold.

Animals↗
PubMed1986

Superficial membrane -SH groups inaccessible by intracellular GSH.

The importance of membrane -SH groups in the epithelium and posterior fiber cells of rabbit lens was demonstrated by employing a non-penetrating sulfhydryl reagent parachloromercuribenzoate sulfonic acid (PCMBS). Both fiber cell and epithelial membrane preparations contain substantial amounts of -SH, 31 nmoles/mg membrane protein. PCMBS-treatment of anterior and posterior surfaces of the lens leads to dramatic increases in the calcium influx across both anterior and posterior surfaces, indicating that the importance of membrane -SH groups is not limited to the epithelium. When the entire lens is bathed in PCMBS (0.1 mM) for short duration and transferred to normal medium, calcium continues to increase from 0.4 mM to nearly 1 mM over a 20 hr period. At this point in time, GSH levels are normal, indicating that intracellular GSH does not gain access to PCMBS-binding sites. In contrast, external GSH or cysteine, at lower levels (5 mM) quickly reverses PCMBS binding with membrane -SH groups and leads to near normal levels of lens calcium during subsequent culture. This in addition to the fact that PCMBS is not found in the cell interior where GSH levels are undiminished, suggests that the critical -SH groups involved in control of barrier properties are externally located where little protection from intracellular GSH is afforded. These data indicate that aqueous humor GSH may play a critical role in maintaining reduced -SH groups controlling membrane permeability located on the surface of membranes.

4-Chloromercuribenzenesulfonate↗
PubMed1985

Effects of intracellular calcium on lens membrane permeability.

The present investigation was designed to assess whether lens membrane permeability is affected by changes in levels of intracellular calcium. Lanthanum, an inhibitor of Ca-ATPase, affected an increase in the concentration of intracellular calcium (Cai) measured in cortical fiber cells. Preculture of lenses in lanthanum (1.0mM) caused an accumulation of 36Cl during subsequent culture at a rate three-fold higher than control lenses. Changes in calcium levels, however, were not responsible for the observed flux changes because a 40mV depolarization was observed to occur prior to a significant increase in calcium levels. The non-specific effects of lanthanum and other potential inhibitors of calcium transport were avoided by preculturing lenses in an ion-HEPES medium containing 20mM calcium chloride. In lenses with a six-fold increase in calcium levels there resulted only a 10% increase in 36Cl uptake over a 3 hr period. 86Rb efflux was also measured and the rate constant was unchanged compared to control lenses. Calcium accumulation did lead to a small (8mV) depolarization which may account for the small increase in chloride accumulation. By light microscopy, morphology of cortical lens fibers and the epithelium appeared unchanged in the calcium-loaded lens. The results provide little evidence that an increase in Cai leads to acute changes in lens membrane permeability.

Body Fluids↗
PubMed1985

Cytotoxic effects of internal calcium on lens physiology: a review.

While calcium is possibly involved in cataractogenesis, it is unquestionably involved in normal lens physiology. Numerous reports have documented the many cellular processes in other tissues affected by alterations in cellular levels of calcium. The homeostasis of the lens is no less dependent on the critical balance of intracellular calcium. With advances being made in calcium-sensitive microelectrodes and pioneering studies progressing in ion channel electrophysiology, interest in calcium metabolism in the lens has been intensified. This report is an attempt to review recent findings that deal solely with biochemical changes resulting from calcium imbalances in the lens interior.

Animals↗
PubMed1985

The importance of membrane sulfhydryl groups to calcium homeostasis in the lens.

This study focused on whether changes in lens levels of glutathione and calcium, early events associated with cataract formation, were related or that one might cause the other. The first part of the investigation was concerned with the extent to which an increase in levels of intracellular calcium might alter GSH levels in lens fiber and epithelial cells. The results demonstrate that calcium accumulation, either at 19 degrees C or 37 degrees C, did not diminish the concentration of GSH. More importantly, GSH levels did not decline in opaque regions of a calcium-loaded lens. The reciprocal part of the problem focused on whether a decline in lens thiol might lead to an increase in levels of calcium and subsequent opacification. In particular, it was shown that treatment of lenses with parachloromercuribenzene sulphonic acid (PCMBS), a nonpenetrating sulphydryl probe, resulted in a 10-30% loss of membrane SH groups in the epithelium. Diminished numbers of SH groups was accompanied by chloride fluxes and an increase in membrane permeability to sodium and calcium with an influx of sodium and calcium leading to opacities. It is important to note that these changes occurred in the absence of any change in cellular levels of soluble protein-SH or GSH. Additional experiments suggest that calcium transport was not impaired, as evidenced by lack of inhibition of Ca-ATPase activity in lenses treated with PCMBS. The results suggest that one explanation for opacification is that oxidative insults, which diminish GSH levels, leads to a loss of important membrane SH groups.(ABSTRACT TRUNCATED AT 250 WORDS)

4-Chloromercuribenzenesulfonate↗