Biomedical subjects

Marta C Yappert

Publications and source records attributed to Marta C Yappert.

7 recordsLinked to original sources

PubMed2006

Oxidation-induced changes in human lens epithelial cells. 1. Phospholipids.

Lipid compositional changes in lens epithelial cells (HLE B-3) grown in a hyperoxic atmosphere were studied to determine if oxidation could cause changes in the amount and type of phospholipid similar to those found in vivo with age and cataract. The phosphatidylcholines in HLE B-3 cells were 8 times more unsaturated than the sphingomyelins. Cell viability was the same for cells grown for up to 48 h in a normoxic or hyperoxic atmosphere. Lipid oxidation was about three times higher after growth in a hyperoxic atmosphere compared with cells grown in a normoxic atmosphere. The lack of change in the relative amount of sphingomyelin and the decrease in phosphatidylcholine coupled with the increase in lysophosphatidylcholine support the idea that similar mechanisms may be responsible for the lipid compositional changes in both lens epithelial and fiber cells. It is postulated that lipases eliminate oxidized unsaturated glycerolipids, leaving a membrane increasingly composed of more ordered and more saturated sphingolipids. Oxidative stress leads to changes in membrane composition that are consistent with those seen with age in human epithelial cells. Oxidation-induced epithelial phospholipid change is an area of research that has gone virtually unexplored in the human lens and could be relevant to all cell types and may be important to lens clarity.

Aging↗

PubMed2006

Oxidation-induced changes in human lens epithelial cells 2. Mitochondria and the generation of reactive oxygen species.

The relationships among reactive oxygen species (ROS) generation, lipid compositional changes, antioxidant power, and mitochondrial membrane potential were determined in a human lens epithelial cell line, HLE-B3. Cells grown in a hyperoxic atmosphere grew linearly for about 3 days, and then progressively died. Total antioxidant power and ROS generation increased by 50 and 43%, respectively, in cells grown in a hyperoxic atmosphere compared to those cultured in a normoxic atmosphere. By specifically uncoupling the mitochondrial proton gradient, we determined that the mitochondria are most likely the major source of ROS generation. ROS generation correlated inversely with mitochondrial membrane potential and the amount of cardiolipin, factors likely to contribute to loss of cell viability. Our results support the idea that hyperoxic damage to HLE-B3 cells derives from enhanced generation of ROS from the mitochondrial electron transport chain resulting in the oxidation of cardiolipin. With extended hyperoxic insult, the oxidants overwhelm the antioxidant defense system and eventually cell death ensues.

Carbonyl Cyanide m-Chlorophenyl Hydrazone↗

PubMed2005

alpha-Crystallin binding in vitro to lipids from clear human lenses.

The association of alpha-crystallin to lens membranes increases with age and cataract. Lipid compositional changes also occur with age, cataract, and diabetes. In this study we determined the influence of lipid compositional differences on the binding capacity of alpha-crystallin to lipid vesicles in vitro. Lipids were extracted from pools of human lenses from younger (22+/-4 y, n=30) and older (69+/-3 y, n=26) nondiabetic donors as well as from diabetics taking insulin (60+/-9 y, n=26) and diabetics not taking insulin (58+/-9 y, n=20). Diabetics were insulin dependent for an average of 6 years. Extracted lipids were extruded into large unilamellar vesicles. alpha-Crystallin was mixed with the lipid at 36 degrees C, allowed to bind for about 12 h, and centrifuged at 14,000 g. This centrifugal force was low enough to not pellet free alpha-crystallin but high enough to pellet the lipid and bound alpha-crystallin. alpha-Crystallin-lipid binding was characterized by comparing the amount alpha-crystallin in the pellets of samples with and without lipid. Protein was measured using an assay that minimized interference from lipids. Lipid composition was determined by 31P-NMR spectroscopy. The binding capacity of alpha-crystallin to lipids was 12, 19, 8.9, 17 microg bound/mg lipid for lens lipids extracted from younger, older, insulin-treated and nontreated diabetic donors, respectively. The amount of alpha-crystallin in the pellet (bound alpha-crystallin) was significantly lower for the lipids from the younger group of lenses, p=0.033 and insulin-treated group, p=0.006, compared with the older group of lenses. Higher binding capacity was associated with a higher relative amount of sphingolipid and lower relative amounts of phosphatidylethanolamine-related lipid and phosphatidylcholine. The binding capacity of alpha-crystallin to lens lipids, measured in vitro, increases with age and decreases in diabetic donors that were treated with insulin. Our data support the idea that with age and perhaps certain types of diabetes, more alpha-crystallin is bound to the membrane and serves as a condensation point to which other crystallins bind and then become oxidized.

Adult↗

PubMed2005

Human lens phospholipid changes with age and cataract.

PURPOSE: To determine the phospholipid changes responsible for the increase in membrane lipid hydrocarbon chain order, or stiffness, with age and cataract in the human lens. METHODS: Clear human lenses were pooled into four groups, with donors ranging in age from 15 to 29, 30 to 49, 50 to 64, and 65 to 74 years. Whole human cataractous lenses were obtained from donors after extracapsular cataract extraction. Cataractous lenses were grouped into four classifications: mature, mixed cortical and nuclear, immature nuclear sclerotic, mature posterior subcapsular, and mature nuclear. Lipids were extracted and quantified gravimetrically. The relative phospholipid composition was determined by (31)P-nuclear magnetic resonance spectroscopy. RESULTS: The relative and absolute amount of sphingolipids, including dihydrosphingomyelin and sphingomyelin, increased with age, whereas glycerolipids, including phosphatidylcholine and two phosphatidylethanolamine-related phospholipids, decreased. These changes were exacerbated by the presence of cataract and were substantial, greater than the changes in lipid levels reported in any organ in association with any disease. CONCLUSIONS: The changes in the amount of lipids with age and cataract support the idea that glycerolipids are selectively oxidized over lipids with fewer double bonds, such as sphingolipids. As a result of the elevation of sphingolipid levels with species, age, and cataract, lipid hydrocarbon chain order, or stiffness, increases. Increased membrane stiffness may increase light-scattering, reduce calcium pump activity, alter protein-lipid interactions, and perhaps slow fiber cell elongation.

Adolescent↗

PubMed2004

Lens lipids and maximum lifespan.

Unlike in most organs, the lipid composition of lenses varies dramatically among species and with age. The focus of this study is to assess how these changes relate to lifespan. Studies on cataract suggest that the lens may serve as a window into the processes leading to accelerated mortality. As a first step toward elucidating cellular processes in the lens that may serve as markers for accelerated mortality, we examined the correlation between species-dependent and age-related lens lipid compositional differences and maximum life span. We included data from camels, which, even in old age, rarely develop cataracts although they live under adverse conditions. Camel lens lipids were mainly composed of sphingolipids (77%) and phosphatidylcholines (23%). Bovine lens lipid composition was comparable to a previous study, and both bovine lens sphingolipids, phosphatidylcholines and camel lens phosphatidylcholines content fit well (within the 95% confidence limits) in the curve obtained by plotting maximum life spans of other species with sphingolipids and phosphatidylcholines. Lifespan was directly related to lens sphingolipid content and indirectly related to lens phosphatidylcholine content. The camel lens sphingolipid value was significantly above the curve for other species. Except for the camel lens nucleus, lipid order and sphingolipid content were linearly related, p < 0.005 with a slope of 0.85+/-0.07, and intercept of 6.9+/-3.8. Lipid phase transition temperature and sphingolipid content were also linearly related, p = 0.01 with a slope of 0.20+/-0.07, and intercept of 21.7+/-5.3. Our data support the hypothesis that humans have adapted so that their lens membranes have a high sphingolipid content that confers resistance to oxidation, allowing these membranes to stay clear for a relatively longer time than is the case in many other species. Age-related changes in human lens lipid composition may serve as a marker for oxidative stress and may reflect systemic oxidative insult, providing a window into the health of an individual.

Animals↗

PubMed2003

Light scattering of human lens vesicles in vitro.

In passing through the lens, light crosses thousands of cell membranes. To explore the possible contribution of lipids to the scattering properties of the lens, we have carried out in vitro studies with lipids extracted from human lenses 1-90 years of age. Sphingomyelin and human lens lipids were extruded into large unilamellar vesicles (LUVs). The intensity of light scattered by human lens LUVs increased with age and lipid hydrocarbon chain order. Hydrocarbon chain order also correlated with light scattering intensity by sphingomyelin LUVs. Light scattered by LUVs composed of sphingomyelin (1-30 mg ml(-1)) was 20 to 100 times more intense than that scattered by the same concentration of alpha-crystallin in aqueous media. Increased lipid hydrocarbon chain order as well as variations in the headgroup and interfacial region of bilayers resulting from lipid compositional changes can influence membrane light scattering properties. In vitro measurements suggest that the contribution to light scattering by lipids may be significant and should not be disregarded in the investigation of factors and components that lead to the increase in light scattering by human lenses with age and cataract.

Adolescent↗

PubMed2003

Influence of age, diabetes, and cataract on calcium, lipid-calcium, and protein-calcium relationships in human lenses.

PURPOSE: Calcium is elevated in most cataractous human lenses and may contribute to cataractogenesis. In this study, age-related changes were examined in the total calcium content of clear human lenses and the binding of calcium to lens lipids and proteins. METHODS: Total lens calcium was determined by atomic absorption spectroscopy. Calcium binding was measured by light scattering and measurement of calcium by atomic absorption spectroscopy in bound and unbound fractions. RESULTS: The calcium content of clear human lenses decreased between 18 and 55 years of age and increased between 55 and 75 years, as well as in the presence of cataract. Total calcium levels in clear lenses from subjects with insulin-dependent diabetes did not differ from that in lenses of age-matched control subjects. In vitro binding studies have shown that lens lipids can bind nearly all the calcium present in the human lens. Age and cataract diminished the capacity of lens lipids to bind calcium. Calcium-induced light-scattering, measured in vitro for lens proteins, correlated with increasing age and cataract. CONCLUSIONS: The data support the hypothesis that increased intracellular calcium concentrations and a diminished capacity of lens lipids to bind to calcium initiate a cascade of events that culminates in increased light-scattering from lipids and especially proteins. Calcium binding to lipid membranes cannot directly contribute to light-scattering in cataractous lenses. It has been suggested that most of the diffusible calcium in the lens is in the intercellular spaces and that lens lipids in the outer leaflet of the bilayer bind to that calcium. If so, this could account for the 150-fold difference between free and bound calcium levels in the lens.

Adolescent↗