Lipofuscin Content Research Articles

Autofluorescent Granules of the Human Retinal Pigment Epithelium: Phenotypes, Intracellular Distribution, and Age-Related Topography.

PurposeThe human retinal pigment epithelium (RPE) accumulates granules significant for autofluorescence imaging. Knowledge of intracellular accumulation and distribution is limited. Using high-resolution microscopy techniques, we determined the total number of granules per cell, intracellular distribution, and changes related to retinal topography and age.MethodsRPE cells from the fovea, perifovea, and near-periphery of 15 human RPE flat mounts were imaged using structured illumination microscopy (SIM) and confocal fluorescence microscopy in young (≤51 years, n = 8) and older (>80 years, n = 7) donors. Using custom FIJI plugins, granules were marked with computer assistance, classified based on morphological and autofluorescence properties, and analyzed with regard to intracellular distribution, total number per cell, and granule density.ResultsA total of 193,096 granules in 450 RPE cell bodies were analyzed. Based on autofluorescence properties, size, and composition, the RPE granules exhibited nine different phenotypes (lipofuscin, two; melanolipofuscin, five; melanosomes, two), distinguishable by SIM. Overall, lipofuscin (low at the fovea but increases with eccentricity and age) and melanolipofuscin (equally distributed at all three locations with no age-related changes) were the major granule types. Melanosomes were under-represented due to suboptimal visualization of apical processes in flat mounts.ConclusionsLow lipofuscin and high melanolipofuscin content within foveal RPE cell bodies and abundant lipofuscin at the perifovea suggest a different genesis, plausibly related to the population of overlying photoreceptors (fovea, cones only; perifovea, highest rod density). This systematic analysis provides further insight into RPE cell and granule physiology and links granule load to cell autofluorescence, providing a subcellular basis for the interpretation of clinical fundus autofluorescence.

Quantifying lipofuscin in retinal pigment epithelium in vivo by visible-light optical coherence tomography-based multimodal imaging

Lipofuscin in the retinal pigment epithelium (RPE) is the major source of fundus autofluorescence (FAF). A technical challenge to accurately quantify the FAF intensities, thus the lipofuscin concentration, is to compensate the light attenuation of RPE melanin. We developed the VIS-OCT-FAF technology to accomplish optical coherence tomography (OCT) and FAF simultaneously with a single broadband visible light source. We demonstrated that light attenuation by RPE melanin can be assessed and corrected using the depth-resolved OCT signals. FAF images from albino and pigmented rats showed that without compensation, FAF signals from pigmented rats are lower than that from albinos. After compensation, however, FAF signals from pigmented rats are higher. This finding is supported by measurements of lipofuscin fluorophore A2E in the RPE using liquid chromatography/mass spectrometry (LC/MS) showing that compensated FAF intensities correlate linearly with A2E contents. The present work represents an important step toward accurately assessing RPE lipofuscin concentrations by FAF.

OXIDATIVE AND ANTIOXIDANT PROCESSES IN THE FOCUS OF THE SKIN EXPOSED TO LOCAL ULTRAVIOLET IRRADIATION

The purpose of this study was to investigate the effect of local ultraviolet radiation on the content of lipid peroxidation products, lipofuscin, and the functioning of antioxidant enzymes in the skin of guinea pigs. Materials and methods. The studies were performed on 24 albino guinea pigs exposed to a single local ultraviolet radiation. The control group was made up of intact animals. The content of primary (diene conjugates) and secondary (TAC-active products) lipid peroxidation products, the activity of antioxidant enzymes (superoxide dismutase and catalase), and the content of lipofuscin were assessed in the skin in 2 hours, 4 hours, and on the 3rd and 8th day following the UV irradiation. Results. Under the influence of local ultraviolet radiation, all guinea pigs develop erythema (its peak is observed in 4 hours, and on the 3rd day), which disappears on the 8th day. There is a sharp enhancement of lipid peroxidation processes due to the accumulation of lipid peroxidation primary and secondary products in the skin of guinea pigs in all periods of the experiment. In parallel with the increase in lipid peroxidation products, there is a decrease in the activity of antioxidant enzymes (catalase and superoxide dismutase) in the focus of radiation. The content of lipofuscin in the skin progressively increases throughout the study period. Conclusions. The early erythema period following the local ultraviolet irradiation in the skin of guinea pigs, is characterized by increased content of diene conjugates and TBA-active products (in 2, 4 hours, on the 3rd day); decreased catalase activity and increased lipofuscin concentration (on the 3rd day after irradiation). The early post-erythema period (8th day) following the local ultraviolet irradiation is characterized by the accumulation of diene conjugates, TBA-active products and lipofuscin against the background of a decrease in the activity of catalase and superoxide dismutase in the skin of guinea pigs.

Verification of ageing methods for the burrowing shrimp, Neotrypaea californiensis, using extractable lipofuscin and gastric mill cuticular features

AbstractThe burrowing shrimp Neotrypaea californiensis is an important ecosystem engineer that inhabits estuaries along the US Pacific Northwest coast. This species plays an important role in the estuarine ecosystem but negatively impacts oyster aquaculture through its burrowing activities. Development of population models for burrowing shrimp management requires more detailed life history information and accurate estimates of age. Ageing studies have been limited for crustaceans because it is generally believed that they do not retain structures with annual deposits commonly used to age other marine organisms, when they moult their exoskeletons. A mesocosm growth experiment and field surveys were combined to compare the performance of two ageing techniques, quantification of autofluorescent lipofuscin and gastric mill ossicular lamellae, for estimating age in N. californiensis. Animals of known age were grown in outdoor mesocosms and sampled regularly to correlate age metrics with body size and true age. Lipofuscin concentration increased with time across multiple cohorts at the rate of 1.430 ± 0.060 ng µg−1 year−1. Lamellae counts also increased with time (4.922 ± 0.337 lamellae year−1). While age estimates based on lipofuscin concentration and lamellae counts generally agreed, carapace size did not correlate to either age metric. Lamellae counts from field collections suggest they are added sequentially with age but the relationship can vary by location. When used together, the application of both techniques may provide robust estimates of crustacean age especially when size-based measurements are imprecise.

CONSISTENT PATTERNS OF POSTMORTAL STRUCTURAL AND BIOCHEMICAL CHANGES IN DIAPHRAGM MUSCULAR TISSUE: IMPORTANCE FOR FORENSIC MEDICAL ESTIMATION OF TIME OF DEATH COMING

The issue on estimating the time of death coming still remains a challenge that requires a lot of questions to be answered, therefore, the development of new scientific methods aimed at overcoming these difficulties is a top priority in solving the problem of estimating the time of death coming.
 The purpose of this study was to investigate the structural and biochemical changes in the muscle tissue of the diaphragm in the early post-mortem period.
 The muscular tissue samples of the diaphragm just below the lungs taken from 30 human corpses in the early post-mortem period (in time intervals of 3-13 hours from the onset of death) were investigated. We studied structural and biochemical markers in the homogenates of the muscles of the diaphragm: the content of glycogen, acid phosphatase, lactate, lactate dehydrogenase, lipofuscin, and cholinesterase. It was found out that in the early post-mortem period structural and biochemical changes in the muscle tissue of the diaphragm are characterized by regular morphological biochemical and biophysical changes. In particular, the post-mortem dynamic changes in glycogen, acid phosphatase and lipofuscin levels is most significant; i.e. within the time interval of 3–9 hours from the time of death coming, the assessment of the “cholinesterase / glycogen or acidic phosphatase / glycogen” ratio is the most diagnostically valuable, then after 9 hours the only “cholinesterase / glycogen” ratio is meaningful.
 The content of glycogen in the homogenates of the diaphragm muscle tissue depending on the time of death coming ranged from (4.765 ± 0.058) mg / g in 3 hours, to (1.883 ± 0.027) mg / g in 13 hours after the death coming, reliably ( p <0.001) differing in time intervals. The inverse pattern was characterized by the acid phosphatase content: (2.748 ± 0.019) u / g in 3 hours and (4.49 ± 0.032) u / g in 13 hours. These markers are the most significant to demonstrate the patterns and can be used as critically important signs in forensic diagnosis.

Combination of chick embryo and nutrient mixture prevent D-galactose-induced cognitive deficits, immune impairment and oxidative stress in aging rat model

Aging is spontaneous and inevitable processes that lead to changes in biological systems. The present paper was designed to investigate the anti-aging roles of chick embryo (CE) and nutrient mixture (NM) in aging rats. Aging was induced by administration of D-galactose (D-gal, 500 mg/kg/day for 90 days). CE and NM were administered to aging rats through different dose gavage once a day. Cognitive function assessment was performed using the Morris water maze test. At the end of experiment, serum and tissues were collected for immunity and antioxidation function. The organs and tissues were excised for histological study. The results demonstrated that CE plus NM was superior treatment to improve the histopathologic changes and reverse learning and memory impairment of the aging rats. CE plus NM also increased the spleen and thymus index as well as splenocyte proliferation, and reversed inflammatory cytokine levels. In addition, the biochemical index showed that CE plus NM could improve the antioxidant enzyme activity of the aging rats, decrease lipofuscin (LF) and glutamate content. CE plus NM also inhibited the activation of TLR4/NF-κB pathway stimulated by LPS in splenic B lymphocytes. Overall, these results seem to be implying that CE plus NM was used as potentially natural supplement or functional food for preventing aging.

Kidney anatomy, histology and histometric traits associated to renosomatic index in Gymnotus inaequilabiatus (Gymnotiformes: Gymnotidae)

ABSTRACT The Gymnotus inaequilabiatus is a Neotropical fish widely distributed in marginal areas of bays. The aim of this study was to describe the main histological and histopathological traits in the head and exocrine kidney. Here, histometric and structural density techniques were associated with renosomatic index (RSI). The kidney was processed for light microscopy. Lipofuscin and hemosiderin content were visually estimated in the melonomacrophages centers (MMCs). All the biometric body variables were correlated with RSI, especially the kidney weight and gross lesions count. The general architecture of head and exocrine kidney was similar to that described for other teleost species. MMCs were prevalent in both portions and correlated with RSI in the head and exocrine kidney. Granulomatous structures were often observed in both portions; however, they were associated only in the exocrine kidney with RSI. Of all the structures hystometrically estimated, only proximal tubular diameter and thickness, and distal tubular thickness were correlated to renosomatic index. The RSI is an useful biometric variable that represent some physiological and morphological characteristics of kidney in G. inaequilabiatus. These findings may be used in future studies to evaluate the effects of environmental stressors on the renal adaptative physiological process.

Mitochondrial Lipofuscin and Thermomitochondrial Lipofuscin in Homogenates of Rat Organs

Abstract—The amounts of lipofuscin (age pigment) were compared in four organs of rats: the liver, the kidney, the heart muscle, and the brain, as well as in a suspension of ghosts of hepatic mitochondria. It was shown that liver lipofuscin, which absorbs UV light at 360 nm and fluoresces at 460 nm, is predominantly formed due to mitochondrial processes. The lowest lipofuscin content was observed in the heart muscle; in the other organs studied, the levels of lipofuscin calculated per mg of protein were approximately the same. The spectral data and the lifetime of the excited state suggest that a considerable contribution to the fluorescence of lipofuscin in various organs belongs to mitochondrial lipofuscin. Along with lipofuscin, a fluorescent pigment, all organs were shown to contain non-fluorescent protein aggregates: their level was the highest in the heart muscle and the lowest in the liver. Among them, a significant portion were covalently cross-linked aggregates that were not destroyed by detergent treatment. Moderate heating of the organ homogenates caused the formation of thermolipofuscin. The level of thermolipofuscin production was the highest in the brain and the lowest in the heart. Apparently, a significant portion of the thermolipofuscin that formed in organ homogenates was thermomitochondrial lipofuscin from the membranes of damaged mitochondria. The formation of thermolipofuscin strongly involved aromatic residues: tyrosine and tryptophan.

Organotropism and biomarker response in oyster Crassostrea gigas exposed to platinum in seawater.

Platinum (Pt) is a technology critical element (TCE) for which biogeochemical cycles are still poorly understood. This lack of knowledge includes Pt effects on marine organisms, which proved to be able to bioconcentrate this trace element. Oysters Crassostrea gigas were exposed to stable Pt isotope spiked daily in seawater for 35days. Seawater was renewed daily and spiked (with Pt(IV)) to three nominal Pt concentrations (50, 100, and 10,000ngL-1) for two replicate series. Organotropism study revealed that gills, and to a lesser extent mantle, are the key organs regarding Pt accumulation, although a time- and concentration-dependent linear increase in Pt levels occurred in all the organs investigated (i.e., digestive gland, gonads, gills, mantle, and muscle). In oysters exposed to Pt concentrations of 10,000ngL-1, significant biomarker impairments occurred, especially at cellular levels. They reflect altered lipofuscin and neutral lipid contents, as well as intralysosomal metal accumulation. These observations were attributed to activation of excretion/detoxification mechanisms, including Pt elimination through feces and clearly support the importance of the digestive gland in the response to direct Pt exposure. Despite relatively constant condition index, the integrative biological response (IBR) index suggests a generally decreasing health status of oysters.

Toxic effects of prolonged exposure to [C14mim]Br on Caenorhabditis elegans

Ionic liquids (ILs) are gradually concerned due to their potential environmental and health risks. In this work, the chronic effects of imidazolium-based ILs, using [C14mim]Br as a representative, were evaluated using model animal Caenorhabditis elegans. Our results show that prolonged exposure (72 h) of ILs to the nematodes at concentrations of 5 and 10 mg/L induced adverse effects on the growth, locomotive behaviors and development. To explore the toxicity mechanism, lipofuscin content, ROS level and the expressions of five superoxide dismutase (SOD) genes were determined after the prolonged exposure. The lipofuscin content, ROS level and expressions of SOD genes did not show significant changes except that the expression of sod-5 was reduced by 2.7-fold following the treatment of 10 mg/L of [C14mim]Br. These results suggest that oxidative stress may not be responsible for the adverse physiological effects induced by relatively low concentrations of imidazolium-based ILs. We further determined the gene expressions of phase I detoxification enzyme cytochrome P450 (CYP), phase II detoxification enzyme UDP-glucuronosyltransferase (UGT) and ATP-binding cassette (ABC) transporter P-glycoprotein (PGP). The results demonstrate that CYP, UGT and PGP may be involved in the detoxification of ILs. Our findings will aid in understanding the mechanisms of both toxicity and detoxification of imidazolium-based ILs in animals.

Hepatotoxic responses of 4-nonylphenol on African catfish (Clarias gariepinus): antixoidant and histochemical biomarkers.

4-Nonylphenol (NP) toxicity in fish attracts much attention due to its ability in targeting several organs; however, the researches regarding its potential hepatotoxicity are conflicting and still require further investigation. Therefore, the objective of this study is to focus on this issue from the histophysiological point of view using NP intoxicated African catfish (Clarias gariepinus) as a model of hepatotoxicity. Twelve adult fish (6 per group) were divided into two groups; the first was considered as a control and the second was exposed to NP dissolved in water at a dose of 0.1mg/kg BW for 3weeks. A significant reduction in the hepatic alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase levels was observed in NP-exposed fish. Concerning the oxidant/antioxidant balance, a significant depletion in superoxide dismutase, catalase, and glutathione peroxidase was found along with a significant elevation in total peroxide and malondialdhyde. The histopathological examination of the hepatic tissues revealed that NP had marked hepatotoxic effects including hepatitis, centrilobular and focal hydropic and fatty degeneration, fatty change (steatosis), hepatic coagulative necrosis, and nuclear alterations in addition to apoptosis of hepatocytes and necrosis of endothelial cells. Depletion of the glycogen and increased in pigments (lipofuscin and hemosiderin) content in the hepatocytes were also recorded. Hemosiderosis and proliferation of the connective tissue around the blood vessels and branches of bile ducts and in the portal areas were also observed. In light of these findings, it was concluded that NP has a well-defined hepatotoxic impact paving the road towards other studies to investigate other detrimental cyto-physiological influences of this aquatic pollutant.

Chlorogenic acid inhibits non-enzymatic glycation and oxidation of low density lipoprotein

: To investigate the effect of chlorogenic acid (CGA) on non-enzymatic glycation and oxidation of low density lipoprotein (LDL). : The non-enzymatic glycation incubation system of LDL-glucose was established. The contents of early glycation products (Amodori product) and intermediate products (dicarbonyl compound) were determined by ultraviolet-visible spectrophotometry, and the content of advanced glycation end products (AGEs) was determined by fluorescence spectrophotometry. The LDL oxidation incubation system was established. The contents of thiobarbituric acid reactive substances(TBARS) and conjugated diene were determined by ultraviolet-visible spectrophotometry. The tryptophan fluorescence quenching, and the content of lipofuscin, total fluorescence products, active aldehydes and malondialdehyde were determined by fluorescence spectrophotometry, and further verified by three-dimensional fluorescence spectroscopy. : In the LDL glycation experiment, 150 μg/mL and 300 μg/mL CGA inhibited the formation of Amadori product, dicarbonyl compounds and AGEs. In the LDL oxidation experiment, 15 μg/mL and 25 μg/mL CGA inhibited the formation of TBARS effectively; 5 μg/mL and 10 μg/mL CGA inhibited tryptophan fluorescence quenching, and the formation of active aldehydes, malondialdehyde, total fluorescence products, lipofuscin and conjugated diolefine. And the three-dimensional fluorescence spectroscopy showed the same results. : CGA can inhibit non-enzymatic glycation and oxidation of LDL.

Clinical application of ultra-wide field fundus autofluorescence imaging

Ultra-wide field fundus autofluorescence (FAF) imaging is a new noninvasive technique with an imaging range of about 200 °. It can detect peripheral retinal lesions that cannot be found in previous FAFs and more objectively reflect intracellular content and distribution of lipofuscin in the retinal pigment epithelium (RPE) and RPE cell metabolic status. The ultra-wide field FAF can find the abnormal autofluorescence (AF) in the peripheral retina of the eyes of age-related macular degeneration (AMD), and different AF manifestations may have an impact on the diagnosis and treatment of the different AMD subtypes. It is helpful to evaluate subretinal fluid in the eyes of central serous choroidal retinopathy and can accurately detect the changes in the outer retina of the eyes without subretinal fluid. It can help to determine the type of uveitis and fully display the evolution of the disease. It can also assess the peripheral photoreceptor cell layer and RPE in patients with retinal dystrophy and retinitis pigmentosa, and comprehensively evaluate their retinal function and monitor the progress of disease. It can also assist in the evaluation of the short-term efficacy and RPE cell function after the scleral buckling surgery for patients with rhegmatogenous retinal detachment. In the future, ultra-wide field FAF may change the knowledge and intervention strategy of ocular fundus diseases and promote the clinical and scientific research in this field. Key words: Retinal diseases/diagnosis; Choroid diseases/diagnosis; Macular degeneration/diagnosis; Diagnostic techniques, ophthalmological; Fluorescence; Review

Vulnerability of marsh frog Pelophylax ridibundus to the typical wastewater effluents ibuprofen, triclosan and estrone, detected by multi-biomarker approach

Pharmaceutical and personal care products (PPCPs) are the environmental pollutants of growing concern. The aim of this study was to indicate the effects of typical PPCPs on the marsh frog Pelophylax ridibundus. We treated male frogs with waterborne ibuprofen (IBU, 250ng·L−1), triclosan (TCS, 500ng·L−1), or estrone (E1, 100ng·L−1) for 14days. Common vulnerability of the frogs was detected from dramatic decrease of Zn, total and metalated metallothionein (MT) concentrations, Zn/Cu ratio, the elevation of activity of glutathione-S-transferase, cathepsin D and DNA instability in the liver, the depletion of cholinesterase in the brain and cortisol in the blood plasma in all exposures. Nevertheless, lipofuscin concentration in the liver was always decreased. The groups were best distinguished by cytochrome P450 (CYP450) activity determined by ELISA. The exposure to IBU caused lesser damage, but elevated the levels of oxyradicals and glutathione (GSH and GSSG) and lysosomal membrane instability. Exposures to TCS and E1 provoked the endocrine disturbance (increased levels of vitellogenin and thyrotropin in blood plasma), decreased lactate dehydrogenase activity and increased level of pyruvate in the liver. TCS caused the increase of GSSG by 7.3 times and lactate levels. Only E1 lead to decrease of deiodinase activity in the liver, activation of CYP450 and caspase-3 and efflux of cathepsin D from lysosomes. Spectrophotometric and ELISA assays of MTs and CYP450 gave distinct results in E1-group. Broad disruption of the hormonal pathways caused by E1 could be of concern for the health status of frogs in their habitats.

Assessing the impact of Benzo[a]pyrene on Marine Mussels: Application of a novel targeted low density microarray complementing classical biomarker responses.

Despite the increasing use of mussels in environmental monitoring and ecotoxicological studies, their genomes and gene functions have not been thoroughly explored. Several cDNA microarrays were recently proposed for Mytilus spp., but putatively identified partial transcripts have rendered the generation of robust transcriptional responses difficult in terms of pathway identification. We developed a new low density oligonucleotide microarray with 465 probes covering the same number of genes. Target genes were selected to cover most of the well-known biological processes in the stress response documented over the last decade in bivalve species at the cellular and tissue levels. Our new ‘STressREsponse Microarray’ (STREM) platform consists of eight sub-arrays with three replicates for each target in each sub-array. To assess the potential use of the new array, we tested the effect of the ubiquitous environmental pollutant benzo[a]pyrene (B[a]P) at 5, 50, and 100 μg/L on two target tissues, the gills and digestive gland, of Mytilus galloprovincialis exposed invivo for three days. Bioaccumulation of B[a]P was also determined demonstrating exposure in both tissues. In addition to the well-known effects of B[a]P on DNA metabolism and oxidative stress, the new array data provided clues about the implication of other biological processes, such as cytoskeleton, immune response, adhesion to substrate, and mitochondrial activities. Transcriptional data were confirmed using qRT-PCR. We further investigated cellular functions and possible alterations related to biological processes highlighted by the microarray data using oxidative stress biomarkers (Lipofuscin content) and the assessment of genotoxicity. DNA damage, as measured by the alkaline comet assay, increased as a function of dose.DNA adducts measurements using 32P-postlabeling method also showed the presence of bulky DNA adducts (i.e. dG-N2-BPDE). Lipofiscin content increased significantly in B[a]P exposed mussels. Immunohistochemical analysis of tubulin and actin showed changes in cytoskeleton organisation. Our results adopting an integrated approach confirmed that the combination of newly developed transcriptomic approcah, classical biomarkers along with chemical analysis of water and tissue samples should be considered for environmental bioimonitoring and ecotoxicological studies to obtain holistic information to assess the impact of contaminants on the biota.

P 162 - Impact of the age-related protein aggregate lipofuscin on β-cell functionality

The formation of cross-linked protein aggregates such as lipofuscin is a typical hallmark of cellular aging and especially affects long-living and postmitotic cells such as neurons and skeletal muscle cells. Lipofuscin can neither be degraded nor exocytosed, it is a prominent source of oxidants and able to inhibit proteolytic systems such as the 20 S proteasome and the autophagy-lysosomal pathway. Adult pancreatic β-cells are also considered to be long-living cells which may last for years or even a life-time. It was shown, that these cells are also characterized by an age-dependent increase in lipofuscin content, but little is known about the process of accumulating protein aggregates and their impact on β-cell functionality. We used the pancreatic β-cell line MIN6 with a well-established artificial lipofuscin to investigate different parameters of β-cell functionality and the impact of lipofuscin on the insulin-degrading enzyme (IDE). Interestingly, we observed both, a significantly higher insulin amount and secretion in the aggregate-fed MIN6 cells. While the expression of insulin does not seem to be affected by lipofuscin treatment, the presence of lipofuscin is able to decrease the activity of IDE by an unknown mechanism.

Assessment of health status of oysters (Crassostreagigas) exposed to environmentally relevant concentrations of Ag and Cu in brackish waters

Human activities have altered estuarine environments leading to increased presence of different pollutants including metals. Although the implementation of new environmental policies has caused a considerable decrease in trace metal concentrations in estuaries around the Bay of Biscay, some elements such as copper (Cu) and silver (Ag) are still present in relatively high concentrations. Oysters have been widely used in environmental biomonitoring programs as sentinel organisms. Oysters Crassostrea gigas from an uncontaminated estuary were exposed to sublethal, environmentally relevant concentrations of Cu (2000ng Cu/L) and Ag (500ng Ag/L) during 14days in brackish water (S=18). A battery of cell and tissue level (exposure) biomarkers at different levels of biological complexity was applied and integrated into the Integrative Biological Response (IBR) index including: metallothionein contents, intralysosomal metal accumulation, digestive gland atrophy and digestive gland tissue integrity. Condition Index (CI) was incorporated into the IBR index as a complementary parameter that reflects the general physiological condition of oysters (organism level). Results indicated an increase in intralysosomal metal accumulation after 7 and 14days of exposure to Ag together with an increase in the digestive epithelium atrophy and lipofuscin content after 7days of exposure to Ag. The responses detected with the aid of biomarkers integrated in the IBR index showed higher toxicity in oysters exposed to Ag, inducing the clear onset of detoxification processes which also occurred, to a lower extent, in Cu-exposed oysters.

Lipofuscin accumulation in tissues of Arctica islandica indicates faster ageing in populations from brackish environments

Environmental factors can affect the rate of ageing and shape the lifespan in marine ectotherms. The mechanisms and the degree of - environmental influence on aging can best be studied in species with wide ranging biogeographic distribution. One of the biomarkers of physiological ageing is the fluorescent age pigment lipofuscin, which accumulates over lifetime in tissues of bivalves. We compared lipofuscin accumulation rate in muscles and respiratory tissues of the extremely long lived bivalve Arctica islandica from five geographically distinct populations (Northern Norway, White Sea, Kiel Bay, German Bight and Iceland). Maximum investigated chronological age across different populations in the present study differed from 40 years in Kiel Bay to 192 years at Iceland. An inverse association between lipofuscin deposition rate and recorded maximum age was observed through inter-population comparisons. In most cases lipofuscin accumulated exponentially over age in a tissue specific manner. The age specific lipofuscin content was significantly higher in respiratory than muscles tissues in all populations. Cellular lipofuscin granule area can be used as indicator of aging across A. islandica populations with the variance in granule accumulation depending on the annual variations of salinity in different marine regions, but not on the habitat specific thermal envelope.

168 - Impact of Artificial Lipofuscin on β-Cell Functionality

Aging is a multi-causal irreversible process characterized by a progressive degeneration affecting all higher organisms. The formation of insoluble protein aggregates such as lipofuscin is one typical hallmark of aging. The accumulation of lipofuscin has already been shown in many different tissues, but especially long-living and postmitotic cells such as neurons and skeletal muscle cells are affected by the intralysosomal accumulation of this protein aggregate. Lipofuscin can neither be degraded by lysosomal enzymes nor exocytosed, furthermore postmitotic cells are not able to protect themselves from lipofuscin accumulation by cell division. Lipofuscin is a source of reactive oxygen species, able to inhibit proteolytic systems leading to a decline in proteasomal activity. Therefore, lipofuscin is an important contributor limiting the lifespan of especially postmitotic cells. Pancreatic β-cells are also considered to be long-living cells in adults and it was shown, that these cells are characterized by an age-dependent increase in lipofuscin content. However, even though β-cell replication is a rare event in aged humans and mice, little is known about the accumulation of protein aggregates and the intracellular effects of lipofuscin in β-cells. Therefore the pancreatic β-cell line MIN6 was treated with artificial lipofuscin to investigate the impact of this aggregate on the insulin-degrading enzyme (IDE) and different parameters of β-cell functionality. Interestingly, we observed a significant rise in insulin secretion and insulin content in the aggregate-fed MIN6 cells. While the expression of insulin does not seem to be affected by lipofuscin treatment, the prescence of lipofuscin is able to decrease the activity of IDE by an unknown mechanism.

Changes in the Interaction of the Pineal Gland and Immune System Organs in Rats in Response to Administration of Melatonin in Conditions of an Altered Light Regime

The aim of the present work was to use correlation analysis to identify the complex reaction of the pineal gland (PG) and immunocompetent organs in response to administration of melatonin (MT) in conditions of an abnormal light regime. Animals were kept for 14 days with natural or round-the-clock illumination (RTCI). Animals then received 0.9% sodium chloride solution or MT for seven days, after which they were decapitated and measurements were made of body, PG, thymus, and spleen weight. Thymus and spleen lymphocyte subpopulations were assessed by flow cytometry. PG lipofuscin content was determined in terms of autofluorescence intensity on frozen sections at wavelengths over the range 560–600 nm. The number of intraorgan relationships within the immune system was found to increase in response to MT regardless of light regime. Administration of MT to animals subjected to RTCI decreased the number of interorgan relationships, with the appearance of negative correlations between PG lipofuscin content and the cellular composition of the spleen. The most marked synchronizing and adaptogenic effects of MT were seen in RTCI conditions.