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1.
Physiol Res ; 71(5): 643-656, 2022 11 28.
Article in English | MEDLINE | ID: mdl-36047722

ABSTRACT

Natural compounds may bear promising therapeutic benefits against metabolic diseases such as type 2 diabetes mellitus (T2DM), which are characterized by a state of insulin resistance and mitochondrial dysfunction. Here, we examined the cellular mechanisms by which aspalathin, a dihydrochalcone C-glucoside unique to rooibos, may ameliorate palmitate-induced insulin resistance and mitochondrial dysfunction in cultured C2C12 myotubules. This current study demonstrated that aspalathin remains effective in improving glucose uptake in insulin-resistant skeletal muscle cells, supported by the upregulation of insulin-dependent signaling that involves the activation of insulin receptor (IR) and direct phosphorylation of protein kinase B (AKT). Interestingly, aspalathin also improved mitochondrial respiration and function, which was evident by an increased expression of carnitine palmitoyltransferase 1 (Cpt1), fatty acid transport protein 1 (Fatp1), sirtuin 1 (Sirt1), nuclear respiratory factor 1 (Nrf1), and transcription factor A, mitochondrial (Tfam). Importantly, our results showed that aspalathin treatment was effective in ameliorating the devastating outcomes of insulin resistance and mitochondrial dysfunction that are linked with an undesired pro-inflammatory response, by reducing the levels of well-known pro-inflammatory markers such as interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and protein kinase C-theta (PKC-theta). Thus, beyond improving glucose uptake and insulin signaling, the current study brings a new perspective in the therapeutic benefits of aspalathin in improving mitochondrial respiration and blocking inflammation to attenuate the detrimental effect of palmitate in skeletal muscle cells.


Subject(s)
Diabetes Mellitus, Type 2 , Insulin Resistance , Humans , Insulin Resistance/physiology , Diabetes Mellitus, Type 2/metabolism , Insulin/pharmacology , Muscle, Skeletal/metabolism , Muscle Fibers, Skeletal/metabolism , Palmitates , Glucose/metabolism , Mitochondria/metabolism
2.
Oxid Med Cell Longev ; 2018: 8936251, 2018.
Article in English | MEDLINE | ID: mdl-30473743

ABSTRACT

Aging skeletal muscles are characterized by a progressive decline in muscle mass and muscular strength. Such muscular dysfunctions are usually associated with structural and functional alterations of skeletal muscle mitochondria. The senescence-accelerated mouse-prone 8 (SAMP8) model, characterized by premature aging and high degree of oxidative stress, was used to investigate whether a combined intervention with mild physical exercise and ubiquinol supplementation was able to improve mitochondrial function and preserve skeletal muscle health during aging. 5-month-old SAMP8 mice, in a presarcopenia phase, have been randomly divided into 4 groups (n = 10): untreated controls and mice treated for two months with either physical exercise (0.5 km/h, on a 5% inclination, for 30 min, 5/7 days per week), ubiquinol 10 (500 mg/kg/day), or a combination of exercise and ubiquinol. Two months of physical exercise significantly increased mitochondrial damage in the muscles of exercised mice when compared to controls. On the contrary, ubiquinol and physical exercise combination significantly improved the overall status of the skeletal muscle, preserving mitochondrial ultrastructure and limiting mitochondrial depolarization induced by physical exercise alone. Accordingly, combination treatment while promoting mitochondrial biogenesis lowered autophagy and caspase 3-dependent apoptosis. In conclusion, the present study shows that ubiquinol supplementation counteracts the deleterious effects of physical exercise-derived ROS improving mitochondrial functionality in an oxidative stress model, such as SAMP8 in the presarcopenia phase.


Subject(s)
Mitochondrial Diseases/drug therapy , Mitochondrial Diseases/therapy , Ubiquinone/analogs & derivatives , Animals , Autophagy/drug effects , Blotting, Western , Cell Survival/drug effects , Disease Models, Animal , Flow Cytometry , Mice , Mitochondria, Muscle/drug effects , Mitochondria, Muscle/metabolism , Mitochondrial Diseases/metabolism , Oxidative Stress/drug effects , Physical Conditioning, Animal , Ubiquinone/pharmacology , Ubiquinone/therapeutic use
3.
J Tissue Eng Regen Med ; 9(6): 691-701, 2015 Jun.
Article in English | MEDLINE | ID: mdl-23109511

ABSTRACT

The osteoblast is the bone-forming cell and is derived from mesenchymal stem cells (MSCs). Osteo-inductive substances could represent a useful therapeutic approach during the fracture repair process. The aim of this work was to evaluate the effects of vitamin MK-7, alone or in association with vitamin D3, in differentiating human MSCs (hMSCs) in vitro along the osteoblastic lineage. In particular, primary endpoints of the study include gene and protein markers of osteoblast differentiation. Considering genes involved in bone formation and mineralization, our data show that vitamin MK-7 enhances vitamin D3 gene induction of osteocalcin (OC). Among genes related to cell growth and differentiation, a specific effect of vitamin MK-7 was observed for growth differentiation factor-10 (GDF10) and insulin-like growth factor 1 (IGF1), the latter being also involved in the induction of vascular endothelial growth factors (VEGFA). Accordingly, vitamin co-supplementation greatly affected VEGFA and its receptor fms-related tyrosine kinase 1 (FLT1), a key factor in both angiogenic and osteogenic processes. These results stress the relevance of MK-7 and D3 co-supplementation in the bone-healing process as able to modulate the expression of genes involved in both mineralization and angiogenesis. Moreover, at the protein level co-association of vitamins might provide an optimal balance between induction and carboxylation of osteocalcin, essential for its functionality in the extracellular matrix (ECM). Our results may provide hints for therapeutic application of hMSCs in bone disease, clarifying mechanisms involved in stem cell-mediated bone development, and they also highlight the relevance of co-supplementation strategies, since single supplementations might result in a suboptimal effect.


Subject(s)
Calcification, Physiologic/drug effects , Cholecalciferol/pharmacology , Mesenchymal Stem Cells/metabolism , Neovascularization, Physiologic/drug effects , Osteogenesis/drug effects , Vitamin K 2/analogs & derivatives , Adult , Biomarkers/metabolism , Cell Survival/drug effects , Enzyme-Linked Immunosorbent Assay , Flow Cytometry , Gene Expression Regulation/drug effects , Humans , Male , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/drug effects , Middle Aged , Osteocalcin/metabolism , Osteogenesis/genetics , Vitamin K 2/pharmacology
4.
Andrologia ; 46(7): 805-7, 2014 Sep.
Article in English | MEDLINE | ID: mdl-23964789

ABSTRACT

Many conditions associated with male infertility are inducers of oxidative stress, including varicocele. Antioxidants, such as coenzyme Q10, may be useful in this case. To evaluate the antioxidant capacity of seminal plasma of infertile men with varicocele before and after an oral supplementation with coenzyme Q10 , 38 patients were recruited from a pilot clinical trial. A standard semen analysis was also performed at baseline and 3 months after an oral supplementation with exogenous coenzyme Q10 100 mg per die. Seminal plasma antioxidant capacity was measured using a spectroscopic method. Coenzyme Q10 therapy improved semen parameters and antioxidant status. This study highlights the importance of oxidative stress in the pathogenesis of male infertility, namely in varicocele, and strengthens the possibility of the usefulness of the antioxidant therapy.


Subject(s)
Infertility, Male/drug therapy , Ubiquinone/analogs & derivatives , Varicocele/complications , Dietary Supplements , Humans , Infertility, Male/etiology , Male , Pilot Projects , Ubiquinone/administration & dosage
5.
J Biotechnol ; 156(3): 153-62, 2011 Dec 10.
Article in English | MEDLINE | ID: mdl-21963587

ABSTRACT

Data normalization of gene expression on human dermal fibroblasts (HDF) exposed to UVA has commonly been done using either GAPDH or ß-actin as reference genes without any validation of their expression stability. Since this aspect, important for accurate normalization, has been overlooked, we aimed to establish a suitable set of reference genes for studies on UVA-treated HDF cultured under both standard atmospheric oxygen tension (normoxia, 21%) and under a physiological, low oxygen tension for these cells (hypoxia, 5%). The stability of six commonly used reference genes was assessed using the geNorm and NormFinder softwares subsequent to reverse-transcription quantitative real-time PCR (RT-qPCR). GAPDH/SDHA were found to be the most stable genes under normoxia, while SDHA/TBP or HPRT1/ß2M were the most stable ones under hypoxia in HDF exposed to 18 J/cm(2) UVA. ß-Actin was always the most unstable reference gene. To emphasize the importance of selecting the most stably expressed reference genes for obtaining reliable results, mRNA expression levels of MMP-1 and COL1A1 were analyzed vs the best reference genes and the worst one. These reference genes are hence recommended for future qPCR analyses in studies concerning photo-damage on UVA-treated HDF.


Subject(s)
Actins/genetics , Cell Hypoxia , Fibroblasts , Gene Expression Profiling/methods , Cell Hypoxia/genetics , Cell Hypoxia/radiation effects , Cells, Cultured , Collagen Type I/genetics , Collagen Type I, alpha 1 Chain , Fibroblasts/metabolism , Fibroblasts/radiation effects , Humans , Matrix Metalloproteinase 1/genetics , Real-Time Polymerase Chain Reaction , Reference Standards , Reverse Transcriptase Polymerase Chain Reaction , Skin/cytology , Ultraviolet Rays
6.
Neurobiol Aging ; 32(11): 2103-5, 2011 Nov.
Article in English | MEDLINE | ID: mdl-20022408

ABSTRACT

Down syndrome (DS) is a chromosomal abnormality (trisomy 21) associated with a complex phenotype. Oxidative stress is known to play a major role in this pathology both due to genetic and epigenetic factors, suggesting that oxidative imbalance contributes to the clinical manifestation of DS. In particular, the implications of oxidative DNA damage in Down syndrome has been linked with neurodegeneration. Here we report the results of a double blind controlled trial aimed at investigating the protective effect of Coenzyme Q(10) on DNA oxidation in this clinical setting using the single cell gel electrophoresis technique.


Subject(s)
DNA Damage/drug effects , Down Syndrome/drug therapy , Oxidative Stress/drug effects , Ubiquinone/analogs & derivatives , Adult , Double-Blind Method , Down Syndrome/metabolism , Humans , Treatment Outcome , Ubiquinone/pharmacology , Ubiquinone/therapeutic use
7.
J Endocrinol Invest ; 32(7): 626-32, 2009 Jul.
Article in English | MEDLINE | ID: mdl-19509475

ABSTRACT

We had previously demonstrated that Coenzyme Q10 [(CoQ10) also commonly called ubiquinone] is present in well-measurable levels in human seminal fluid, where it probably exerts important metabolic and antioxidant functions; seminal CoQ10 concentrations show a direct correlation with seminal parameters (count and motility). Alterations of CoQ10 content were also shown in conditions associated with male infertility, such as asthenozoospermia and varicocele (VAR). The physiological role of this molecule was further clarified by inquiring into its variations in concentrations induced by different medical or surgical procedures used in male infertility treatment. We therefore evaluated CoQ10 concentration and distribution between seminal plasma and spermatozoa in VAR, before and after surgical treatment, and in infertile patients after recombinant human FSH therapy. The effect of CoQ10 on sperm motility and function had been addressed only through some in vitro experiments. In two distinct studies conducted by our group, 22 and 60 patients affected by idiopathic asthenozoospermia were enrolled, respectively. CoQ10 and its reduced form, ubiquinol, increased significantly both in seminal plasma and sperm cells after treatment, as well as spermatozoa motility. A weak linear dependence among the relative variations, at baseline and after treatment, of seminal plasma or intracellular CoQ10, ubiquinol levels and kinetic parameters was found in the treated group. Patients with lower baseline value of motility and CoQ10 levels had a statistically significant higher probability to be responders to the treatment. In conclusion, the exogenous administration of CoQ10 increases both ubiquinone and ubiquinol levels in semen and can be effective in improving sperm kinetic features in patients affected by idiopathic asthenozoospermia.


Subject(s)
Infertility, Male , Ubiquinone/analogs & derivatives , Vitamins , Animals , Asthenozoospermia/enzymology , Double-Blind Method , Humans , Infertility, Male/drug therapy , Infertility, Male/enzymology , Male , Oxidation-Reduction , Randomized Controlled Trials as Topic , Semen/enzymology , Sperm Motility/physiology , Spermatozoa/enzymology , Ubiquinone/metabolism , Ubiquinone/therapeutic use , Vitamins/metabolism , Vitamins/therapeutic use
8.
Biofactors ; 32(1-4): 161-7, 2008.
Article in English | MEDLINE | ID: mdl-19096112

ABSTRACT

Down syndrome (DS) is a chromosomal abnormality (trisomy 21) associated with mental retardation and Alzheimer-like dementia, characteristic change of the individual's phenotype and premature ageing. Oxidative stress is known to play a major role in this pathology since a gene dose effect leads to elevated ratio of superoxide dismutase to catalase/glutathione peroxidase compared to controls in all age categories suggesting that oxidative imbalance contributes to the clinical manifestation of DS. Hyperuricemia is another feature of DS that has an interesting relationship with oxidative stress since uric acid represents an important free radical scavenger. However its formation is connected to the conversion of Xanthine dehydrogenase (XDH) to Xanthine oxidase (XO) which leads to concomitant production of free radicals. Here we report that plasma samples from DS patients in pediatric age, despite an increased total antioxidant capacity, largely due to elevated Uric acid content (UA), present significantly elevated markers of oxidative damage such as increased allantoin levels. Moreover DS plasma samples do not differ from healthy control ones in terms of Coenzyme Q10 and susceptibility to peroxidative stimuli. On the contrary, lymphocyte and platelet CoQ10 content was significantly lower in DS patients, a fact that might underlie oxidative imbalance at a cellular level.


Subject(s)
Down Syndrome/metabolism , Oxidative Stress/drug effects , Ubiquinone/analogs & derivatives , Allantoin/blood , Child , Child, Preschool , Humans , Ubiquinone/metabolism , Uric Acid/blood
9.
Free Radic Biol Med ; 45(3): 345-54, 2008 Aug 01.
Article in English | MEDLINE | ID: mdl-18485919

ABSTRACT

This paper reports on the synthesis and properties of a new UV-absorber (OC-NO) based on the most popular UV filter worldwide, ethylhexyl methoxycinnamate (OMC) in which the methoxy group has been replaced with a pyrrolidine nitroxide bearing antioxidant activity. This sunscreen active has therefore both UV-absorbing and antioxidant properties which could ideally address both the UV-B and UV-A skin photo-damage. For broad-spectrum coverage, the combinations of OC-NO with two commonly used UV-A absorbers (BMDBM and DHHB) were also studied. The results obtained reveal that OC-NO: (a) is as photostable as OMC after UV-A exposure; (b) acts as free radical scavenger as demonstrated by EPR and chemical studies; (c) reduces UV-A and UV-A+BMDBM induced lipid peroxidation in liposomes and cells, measured as reduced TBARS levels and increased C11-BODIPY red fluorescence, respectively; (d) has comparable antioxidant activity to that of vitamin E and BHT commonly used in skin care formulations; (e) is non-cytotoxic to human skin fibroblasts as assessed with the MTT assay when exposed to increasing doses of UV-A; and (f) OC-NO+DHHB is a promising, photostable broad spectrum UV-filter combination that concomitantly reduces UV-induced free radical damage. These results suggest that nitroxide/antioxidant-based UV-absorbers may pave the way for the utilization of 'multi-active' ingredients in sunscreens thereby reducing the number of ingredients in these formulations.


Subject(s)
Cinnamates/chemistry , Cyclic N-Oxides/chemistry , Radiation Protection/methods , Sunscreening Agents/chemical synthesis , Antioxidants/chemical synthesis , Antioxidants/chemistry , Antioxidants/pharmacology , Cinnamates/pharmacology , Fibroblasts/drug effects , Free Radical Scavengers/chemical synthesis , Free Radical Scavengers/pharmacology , Humans , Lipid Peroxidation/drug effects , Sunburn/prevention & control , Sunscreening Agents/chemistry , Sunscreening Agents/pharmacology
10.
Biofactors ; 25(1-4): 241-54, 2005.
Article in English | MEDLINE | ID: mdl-16873953

ABSTRACT

Numerous changes occur post-mortem in fish, affecting its chemical composition and nutritional quality. In the present paper we describe the effect of storage on ice or at -30 degrees C or -80 degrees C on 10 species of Mediterranean fish. Water and lipid soluble antioxidants, lipid pattern and products of oxidative attack on lipids, proteins and DNA were quantified for 7 consecutive days on homogenates of fish light muscle. The earliest events were oxidation of ubiquinol and vitamin C, which disappeared almost completely within 48 hours. Ubiquinol oxidation gave rise to an initial increase of ubiquinone, which peaked at the second day: thereafter ubiquinone itslef decreased, more rapidly and to a greater extent than vitamin E. The decrease in antioxidants was accompanied by significant oxidative damage to lipids, proteins and DNA. TBARS significantly increased beginning from the third day of storage in all species and were linked to a significant reduction in the n-3 PUFA of triglycerides (TG) and phospholipid fractions (PL). A remarkable elevation of protein carbonyls and 8OHdG occurred approximately 24 hours later than PUFA oxidation. For SOD, GPX and GSH significant depletions occurred for all species only at 6th or 7th day, but the final values were always higher than 50% compared to the initial ones. Deep-freezing of the same species at -30 degrees C and -80 degrees C for up to 12 months did not significantly affect the levels of enzymatic antioxidants, the redox couple GSH/GS-SG, n-3 and n-6 PUFA of TG and PL fractions of the light muscle. The only antioxidants, which at -30 degrees C and -80 degrees C appeared to be degraded after 6 and 12 months were ubiquinol and vitamin C. As expected their degradation was higher at -30 degrees C than at -80 degrees C. In fact the average decrease for ubiquinol at -80 degrees C was 42% at 6 and 12 months respectively, whereas at -30 degrees C the decrease was 61% and 87% For vitamin C the average decrease at -80 degrees C was 36% and 67% at 6 and 12 months respectively, and at -30 degrees C it was 61% and 82%. Vitamin E was considerably more stable than ubiquinol and vitamin C. The relative stability of the antioxidants, with the exceptions of ubiquionols, vitamin C and, to a certain extent, vitamin E, was accompanied by a very limited increase in oxidation products. In addition no significant hydrolysis of TG and PL fractions were observed throughout the storage time. The dynamics of lipid, protein and DNA oxidation is discussed in the light of depletion of the various antioxidant systems.


Subject(s)
Antioxidants/metabolism , Fishes/metabolism , Food Preservation/methods , Postmortem Changes , Animals , Ascorbic Acid/metabolism , Frozen Foods/analysis , Lipid Metabolism , Muscles/metabolism , Oxidation-Reduction , Ubiquinone/analogs & derivatives , Ubiquinone/metabolism , Vitamin E/metabolism
11.
Biofactors ; 18(1-4): 51-63, 2003.
Article in English | MEDLINE | ID: mdl-14695920

ABSTRACT

UV-C radiation is able to impair cellular functions by directly damaging DNA, and by inducing an increased formation of reactive oxygen species that leads to a condition of oxidative stress. In this study we evaluated different responses to UV insult of two leukemia cell lines, HL-60 and Raji, and the relationship with their CoQ10 content. DNA damage was monitored by means of the alkaline single cell gel electrophoresis (Comet assay); intracellular levels of ROS, mitochondrial depolarization and cell viability was measured by flow cytometry. Raji cells appeared more resistant to the UV insult; moreover, they did not show any increase in ROS content and the extent of mitochondrial depolarisation was much lower than in HL 60 cell line. Raji cells also contained significantly higher levels of CoQ10 and their ability to incorporate and to reduce exogenous CoQ10 added to the culture medium was remarkably elevated compared with HL 60.


Subject(s)
Leukemia/metabolism , Oxidative Stress , Ubiquinone/analogs & derivatives , Ubiquinone/analysis , Ultraviolet Rays , Cell Survival , Coenzymes , DNA Damage/radiation effects , Electrophoresis, Agar Gel , HL-60 Cells , Humans , Membrane Potentials , Mitochondria/ultrastructure , Oxidation-Reduction , Propidium , Reactive Oxygen Species/metabolism , Tumor Cells, Cultured , Ubiquinone/metabolism
12.
Comp Biochem Physiol B Biochem Mol Biol ; 130(4): 559-64, 2001 Dec.
Article in English | MEDLINE | ID: mdl-11691632

ABSTRACT

The peroxidative activity of trout hemoglobins, HbI and HbIV, which differ in their conformation, was compared with that of HbA. Artificial substrates (guaiacol and dopamine) and more physiological substrates such as model lipid membranes containing unsaturated fatty acids were used. The results indicate that all the hemoglobin molecules assayed show different levels of peroxidative activity. The capability to act as peroxidases is greater in HbIV than in HbI and HbA. In contrast, native globins did not show peroxidase activity. The different peroxidative activity of the Hbs is discussed in relation to stability both vs. protein oxidation and protein dissociation. The results confirm the view that hemoglobin may be of importance in establishing the life span of the erythrocyte itself.


Subject(s)
Erythrocytes/metabolism , Hemoglobins/chemistry , Hemoglobins/physiology , Lipid Peroxidation , Animals , Cell Membrane/metabolism , Dose-Response Relationship, Drug , Hemin/chemistry , Lipid Metabolism , Models, Chemical , Protein Conformation , Time Factors , Trout
13.
Biochim Biophys Acta ; 1505(2-3): 226-37, 2001 Jun 01.
Article in English | MEDLINE | ID: mdl-11334787

ABSTRACT

Previous literature reports have demonstrated that nucleated trout erythrocytes in condition of oxidative stress are subjected to DNA and membrane damage, and inactivation of glutathione peroxidase. The present study was undertaken to investigate if mitochondrial membrane potential in stressed conditions was also influenced. Density-separated trout erythrocyte fractions, obtained using a discontinuous Percoll gradient, were submitted to stress conditions and the mitochondrial membrane potential was determined by means of cytofluorimetric analysis after incubation of each subfraction with JC-1, a mitochondrial specific fluorescent probe. The results clearly show that the mitochondrial membrane potential decreased significantly in all erythrocyte fractions, also if the oxidative effect on mitochondria is more severe with increased density (age) of the cell. Ebselen was very effective in preventing mitochondrial depolarization in young as well as in old erythrocytes.


Subject(s)
Erythrocytes/chemistry , Intracellular Membranes/chemistry , Mitochondria/chemistry , Animals , Antioxidants/pharmacology , Azoles/pharmacology , Benzimidazoles , Carbocyanines , Cellular Senescence , Chromans/pharmacology , Cyclic N-Oxides/pharmacology , Flow Cytometry , Fluorescent Dyes , Free Radicals/analysis , In Vitro Techniques , Isoindoles , Membrane Potentials , Microscopy, Confocal , Molecular Structure , Organoselenium Compounds/pharmacology , Oxidative Stress/drug effects , Reactive Oxygen Species , Trout
14.
Prostaglandins Other Lipid Mediat ; 62(4): 351-66, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11060899

ABSTRACT

Alterations in the functional activities of platelets (PLT) in type I diabetes have been widely observed. These changes play a key role in the development of cardiovascular complications in diabetes. Various functional activities of PLT are the result of the interaction of numerous stimuli with PLT plasma membrane. This study was designed to evaluate the oxidative response and membrane modifications of diabetic PLT stimulated by platelet activating factor (PAF). The oxidative response was assessed by employing luminol- and lucigenin-amplified chemiluminescence. Luminol-amplified chemiluminescence is sensitive to the release of hydrogen peroxide whereas lucigenin-amplified chemiluminescence is sensitive to the production of superoxide anion. Membrane fluidity and polarity were studied using fluorescence spectroscopy. Membrane fluidity was investigated by measuring steady-state fluorescence anisotropy of 1-[4-trimethylammonium-phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH) and membrane polarity was studied by measuring the steady-state fluorescence emission and excitation spectra of 2-dimethylamino[6-lauroyl]-naphthalene (Laurdan). The diabetic group consisted of 20 type I diabetic children with good metabolic control. Our results show a significant decrease in the luminol- and lucigenin-amplified chemiluminescence of PAF stimulated PLT in the diabetic group with respect to controls. These data indicate a decrement in the release of reactive oxygen species by diabetic PLT. We observed a significant increase in steady-state fluorescence anisotropy of diabetic PLT membrane that reflects a decrease in membrane fluidity. Laurdan showed a blue shift of the fluorescence emission and excitation spectra in diabetic PLT with respect to the control group, indicating a decrease in membrane polarity. The addition of PAF to PLT induced a red shift of Laurdan spectra in both groups, indicating an increase in membrane polarity. Our study [table: see text] demonstrates an altered oxidative response to PAF stimulation of diabetic PLT, probably due to altered generation or handling of reactive oxygen species, and alterations in the physico-chemical properties of the plasma membrane which could influence various functional activities of PLT.


Subject(s)
Blood Platelets/physiology , Cell Membrane/physiology , Diphenylhexatriene/analogs & derivatives , Platelet Activating Factor , Reactive Oxygen Species/metabolism , Acridines , Adolescent , Blood Platelets/ultrastructure , Child , Female , Fluorescence Polarization , Humans , Hydrogen Peroxide/blood , Kinetics , Luminescent Measurements , Luminol , Male , Membrane Fluidity , Spectrometry, Fluorescence , Superoxide Dismutase/blood , Superoxides/blood
15.
Biochim Biophys Acta ; 1457(3): 118-28, 2000 Apr 21.
Article in English | MEDLINE | ID: mdl-10773157

ABSTRACT

Density separated trout erythrocytes, using a discontinuous Percoll gradient, yielded three distinct subfractions (top, middle and bottom) since older cells are characterized by increasing density. Cells from each subfraction were incubated with mitochondria-specific fluorescent probe Mitotracker and JC-1 in order to assess mitochondrial mass and membrane potential by means of cytofluorimetric analysis, confocal microscopy and subsequent computer-aided image analysis allowing a detailed investigation at single cell level. Both cytofluorimetric data and image analysis revealed changes in size and redistribution of mitochondria starting from the light fraction to the bottom. In particular in young erythrocytes small mitochondria were detected localized exclusively around the nucleus in a crown-like shape, the middle fraction revealed enlarged mitochondria partially scattered throughout the cytosol, whereas the last fraction represented again mitochondria with reduced size being distinctly dispersed throughout the cytosol in the cells. Concerning membrane potential considerations, our study revealed a dramatic decrease of DeltaPsi(m) in the bottom layer cell mitochondria compared to the top and unusual membrane potential increase of a subpopulation of enlarged mitochondria. DeltapH was also investigated in the three fractions by pretreating the cells with nigericin, allowing to confirm a mitochondrial energetic impairment in older cells.


Subject(s)
Erythrocytes/cytology , Mitochondria/physiology , Oncorhynchus mykiss/blood , Animals , Apoptosis , Benzimidazoles , Carbocyanines , Erythrocyte Aging , Erythrocytes/classification , Flow Cytometry , Fluorescent Dyes , Membrane Potentials , Microscopy, Confocal , Mitochondria/ultrastructure , Nigericin , Xanthenes
16.
Mutat Res ; 464(2): 269-77, 2000 Jan 24.
Article in English | MEDLINE | ID: mdl-10648914

ABSTRACT

Previous literature reports have demonstrated that nucleated trout erythrocytes in conditions of oxidative stress are subjected to DNA and membrane damage, and inactivation of glutathione peroxidase. The present study was undertaken to evaluate the ability of three diaryl tellurides and the organoselenium compound ebselen to protect trout (Salmo irideus) erythrocytes against oxidative stress, induced thermally and by a variation of pH. The antioxidant ability of these molecules was evaluated through chemiluminescence. Impairment of DNA was assessed using the comet assay, a rapid and sensitive single cell gel electrophoresis technique, used to detect primary DNA damage in individual cells. At low concentrations (<10 microM), all the compounds used presented a protective effect on DNA damage without altering the hemolysis rate. In higher concentrations, they accelerated the hemolysis rate and two of the diaryl tellurides were strongly genotoxic.


Subject(s)
Antioxidants/toxicity , Azoles/toxicity , Erythrocytes/drug effects , Erythrocytes/metabolism , Organometallic Compounds/toxicity , Organoselenium Compounds/toxicity , Oxidative Stress/drug effects , Animals , Comet Assay , DNA Damage/drug effects , Dose-Response Relationship, Drug , Erythrocytes/cytology , Hemolysis/drug effects , Hot Temperature/adverse effects , Hydrogen-Ion Concentration , Isoindoles , Luminescent Measurements , Mutagenicity Tests , Oncorhynchus mykiss , Organometallic Compounds/chemistry , Tellurium/chemistry
17.
Eur J Histochem ; 43(2): 113-20, 1999.
Article in English | MEDLINE | ID: mdl-10439214

ABSTRACT

The chromatin structure dynamics of the Colpoda inflata macronucleus have been investigated in relation to its functional condition, concerning chromatin body extrusion regulating activity. Samples of 2- and 25-day-old resting cysts derived from a standard culture, and of 1-year-old resting cysts derived from a senescent culture, were examined by means of histogram analysis performed on acquired optical microscopy images. Three groups of histograms were detected in each sample. Histogram classification, clustering and matching were assessed in order to obtain the mean histogram of each group. Comparative analysis of the mean histogram showed a similarity in the grey level range of 25-day- and 1-year-old cysts, unlike the wider grey level range found in 2-day-old cysts. Moreover, the respective mean histograms of the three cyst samples appeared rather similar in shape. All this implies that macronuclear chromatin structural features of 1-year-old cysts are common to both cyst standard cultures. The evaluation of the acquired images and their respective histograms evidenced a dynamic state of the macronuclear chromatin, appearing differently condensed in relation to the chromatin body extrusion regulating activity of the macronucleus. The coexistence of a chromatin-decondensed macronucleus with a pycnotic extrusion body suggests that chromatin unable to decondense, thus inactive, is extruded. This finding, along with the presence of chromatin structural features common to standard and senescent cyst populations, supports the occurrence of 'rejuvenated' cell lines from 1-year-old encysted senescent cells, a phenomenon which could be a result of accomplished macronuclear renewal.


Subject(s)
Chromatin/metabolism , Ciliophora/metabolism , Animals , Ciliophora/genetics , Ciliophora/growth & development
18.
Am J Law Med ; 6(1): 125-49, 1980.
Article in English | MEDLINE | ID: mdl-7435506

ABSTRACT

In Parham v. J.R., 442 U.S. 584 (1979), the U.S. Supreme Court held that a parent or a guardian can commit a minor to a mental institution if a staff physician certifies that the minor should be committd, even if the minor strenuously opposes their decision. The Court specifically rejected claims that commitment of a minor by a parent or guardian without an adversary hearing is a deprivation of the minor's liberty without due process of law. This Note reviews the Parham opinion, with special attention to its impact on "mature minors" and wards of the state and to its definition of a neutral factfinder. The Note argues first that the Court's failure to establish special safeguards for "mature minors" and wards of the state is inconsistent with constitutional standards of due process, and second, that the Court's acceptance of staff physicians as neutral fact-finders may be unwarranted. The Note recommends the creation of more stringent procedural safeguards for the commitment of minors by parents and guardians, including the use of independent mental health professionals as "neutral factfinders."


Subject(s)
Child Advocacy/legislation & jurisprudence , Commitment of Mentally Ill/legislation & jurisprudence , Forensic Psychiatry/legislation & jurisprudence , Minors , Age Factors , Child , Georgia , Hospitals, Psychiatric/legislation & jurisprudence , Humans , Legal Guardians , Male , Mentally Ill Persons , Parental Consent , Supreme Court Decisions , United States
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