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Role of calcium ions in the aggregation process of hIAPP. Possible implications in T2DM.
No full textType 2 diabetes (T2DM), Alzheimer’s disease and other age-related neurodegenerative and systemic disorders are classified as amyloid disorders. hIAPP appears to be involved in the mechanisms underlying the pathogenesis of T2DM due to its tendency to aggregate and self-assemble. The aggregation of hIAPP is a dynamic process that leads to the formation of transient and heterogeneous oligomers which evolve to form insoluble fibrils. Oligomers appear to be more toxic to β cells than monomers and fibrils. The main target of hIAPP aggregates seems to be the plasma membrane which can undergo different types of damage, destruction of its integrity or alteration of the physical and biological properties that lead to oxidative stress. Numerous factors, both intrinsic and extrinsic to the peptide molecule, appear to influence the hIAPP aggregation process. Different metal ions are able to interact with the hIAPP molecule, modulating its secondary structure and subsequently the peptide’s capacity to aggregate. In this study, the effect of different Ca2+ concentrations on the hIAPP aggregation process was evaluated. The formation and morphology of hIAPP aggregate were respectively evaluated with Thioflavin T assays and electron microscopy, while the ability of the peptide to incorporate into POPC PLMs and form ion channels was monitored by single-channel current measurements. Ca2+ seems to favor the formation of soluble oligomers, suggesting a possible mechanism of hIAPP toxicity
Educazione alimentare a cinque sensi. Il progetto Edueat
No full textL’articolo presenta i contenuti e alcuni risultati di un progetto d’educazione alimentare realizzato in
una scuola primaria di Canosa di Puglia e in una di Foggia nell’anno scolastico 2016-17, incentrato
sul percorso Edueat, un progetto educativo nato dalla collaborazione tra l’Università di Macerata
e il Laboratorio delle Idee di Fabriano. L’esperienza affronta i temi dell’educazione alimentare
basata sui cinque sensi e della cooperazione tra scuole e soggetti esterni nella promozione di un
approccio positivo e consapevole all’alimentazione. Lo studio mette in luce alcuni miglioramenti
a livello conoscitivo rispetto al consumo di cibi salutari nel gruppo dei bambini coinvolti nelle
attività, rispetto al gruppo di controllo
Kinetic parameters of porin incorporation into black lipid membranes of oxidized cholesterol evaluated by means of alternating current
No full textIn this work we report the results of systematical investigation of the time course of porin incorporation into bilayer membranes made of oxidized cholesterol using an ac method (E. Gallucci et al., Biophys. J. 71: 824-831, 1996). This method has the advantages of continuously monitoring resistance and capacitance as a function of time during pore formation until a steady state of the phenomenon is reached. In our case the bilayer membranes are represented, as usual, by an equivalent electrical circuit consisting of one resistance and one capacitance connected in parallel. The total current passing through the membranes is easily monitored, during the insertion process and is interpreted as a probe of the process involved during channel incorporation and assembly. By means of a simple mathematical model, we provide evidence that two concurrent processes are present during the incorporation time course which can be interpreted either as insertion/deinsertion or as positive/negative cooperativity. We observe a phase transition (or similae phenomenon) which seems to take place during the insrtion process, an aspect which has been neglected in most previous investigation of insertion into bilayer membranes. Moreover, the continuous monitoring of capacitance may prove useful in tracking membrane parameters, such as thickness and dielectrical constant, that depend on the lipid and incorporated proteins
Cholesterol modulates amyloid beta peptide 1-42 channel formation in planar lipid membranes
No full textCholesterol, an integral component of eukaryotic cell membranes, is assuming a pivotal role as a target in some neurodegenerative diseases such as Alzheimer, although its role remains elusive Recent studies indicate that Ch can favour the conformational transition of many peptides in model membranes and vesicles. ABP1-42 is one of the pathological features of Alzheimer's disease and, together with ABP1-40, is among the major components of senile plaques. Notably, the ABP1-42 peptide is more neurotoxic than ABP1-40, owing to its much stronger aggregation propensity. In this work, we studied the role of cholesterol in the incorporation and channel formation of ABP1-42 in planar lipid membranes of different composition
Evidence for a different assembly of Gramicidin A in oxidized cholesterol black lipid membrane
No full textGramicidin A (GA) is a hydrophobic linear pentadocapeptide that forms cation- selective channels in natural and black lipid membranes (BLM). The channels are believed to consist of single-stranded helices with the NH2 terminals linked together by hydrogen bonds. Linear gramicidins require a lipid bilayer to fold properly. If the phospholipid acyl chains are less than eight carbons long, channel folding is not observed; if the acyl chains are too long, the membrane- spanning channels are destabilized. Besides, embedded proteins might influence the molecular packing lipids (R. E. Koeppe II et al. Biol. Skr. Dan. Selsk, 49-93-98, 1998). In our experiments, we chose oxidized cholesterol black lipid membranes because their thickness (40±10 A) (Ti Tien and A. L. Diana, In Chemistry and Physics of Lipids, 2, 1968) is suited to both gramicidin length (25-30 A) and folding. The time course of gramicidin incorporation into BLM, studied by means of alternating current (E. Gallucci et al., Biophys. J. 71:824- 831, 1986), was sigmoidal independently of the KCI concentration used, indicating a cooperative process of incorporation into BLM. By means of a simple mathematical model (S. Micelli et al, submitted) we estimated the tm value, i.e. the parameter that describes a phase transition process (or similar phenomenon). This phenomenon has been observed by AA as a function of temperature and GA concentration (J. A. Killian and B. De Kruijff, Biophys. 1. 53: 111-117, 1988; Mingtao Ge and J. H. Freed, Biophys. J., 76: 266-280, 1999). On the other hand, the average values of the steady-state conductance against gramicidin concentration, for different KCI concentrations, shows a sigmoidal behavior too. These curves were fitted by means of a four-parameter equation which allowed us to obtain the Hill coefficient, D, or slope factor. The Hill coefficient gives the number of single-stranded helices which form the channel. Our results suggest that the number of channel sub-units depends on the KCI concentration. In our case, at a KCI concentration of 1M, the channel is characterized by two sub-units, in accordance with AA's finding (D. U. Urry, Proc. Natl. Acad. Sci. USA, 86:672, 1971, W. R. Veatch and R. Blond, Biochemisty, 13:5249, 1974), while for KCI concentrations of 0.1 and 0.5 M. the channel is formed by more than two sub-units
Water permeability of rat liver mitochondria: A biophysical study
No full textThe movement of water accompanying solutes between the cytoplasm and the mitochondrial spaces is central for mitochondrial volume homeostasis, an important function for mitochondrial activities and for preventing the deleterious effects of excess matrix swelling or contraction. While the discovery of aquaporin water channels in the inner mitochondrial membrane provided valuable insights into the basis of mitochondrial plasticity, questions regarding the identity of mitochondrial water permeability and its regulatory mechanism remain open. Here, we use a stopped flow light scattering approach to define the water permeability and Arrhenius activation energy of the rat liver whole intact mitochondrion and its membrane subcompartments. The water permeabilities of whole brain and testis mitochondria as well as liposome models of the lipid bilayer composing the liver inner mitochondrial membrane are also characterized. Besides finding remarkably high water permeabilities for both mitochondria and their membrane subcompartments, the existence of additional pathways of water movement other than aquaporins are suggested. (c) 2006 Elsevier B.V. All rights reserved
Porin incorporation in BLM controlled by surface potencial
No full textPorin incorporation and channel formation in bilayer membranes have been investigated under different incorporation conditions (Gallucci et al Biophys. J. 71 824-831, 1996; Micelli et al., Biophys J.,1998 submitted) We have identified a process of positive/negative cooperativity which can be described by simple mathematical model depending. among others, on two growth rate parameters: K1 describing the insertion or positive cooperativity and K2 describing the negative cooperativity. This model takes into account for the kinetics growth and steady state phase of the insertion, using appropriate data acquisition (dependence of steady state conductance on porin concentration) and analytical techniques, we have provided indirect evidence that the porin channel shows well-defined properties of assambly into the bilayers. In this note we pursue our study in order to answer the following question: are the above-mentioned characteristics dependent on the surface charge ie membrane potential) of the bilayers?
For this reason, using alternating current, we investigate the kinetics of porin incorporation into black lipid membranes (BLM) as a function of ionic strength, the BLM were made up of phosphatidylinositol (PI), oxidized cholesterol (Ox Ch).
Our results indicate that bilayers with higher negative charge density are associated with a slower insertion rate K, and an almost constant negative cooperativity rate K1. These results are corroborated, especially at lower ionic strength, by experiments in which Ba++ is added to the solution as a screening agent. Indeed, the K, parameter for Ox Ch, a neutral bilayer, is higher than that of the PI charged bilayer, and adding Ba++ to Ox Ch does not seem to affect either K1 or K2. Finally, the assembly properties of porin, as evaluated by fitting the data regarding the dependence of steady state conductance Gmss on porin concentration for three different ionic strength (0.1, 0.5 and 1 M), seem to be dependent on the ionic strength and membrane substrat
The role of external applied voltage on kinetic parameters of porin incorporation into black lipid membranes of oxidized cholesterol
No full textThe time course of porin incorporation into black lipid membranes of oxidized cholesterol, as a function of on external applied voltage, was studied by means of an ac method previously reported (E. Gallucci et al., Biophys. J. 71: 824-831, 1996). The kinetic parameters was derived by means of a simple mathematical model before described (D. Meleleo et al., SIF, 1998, Р 53).
At the different voltages Vs examined, the kinetics remained unmodified in its cooperative behavior. The K1 or growth rate parameter describing porin insertion or positive cooperativity seems to be independent of the applied voltage. The K2 or growth rate parameter related to porin deinsertion or negative cooperativity is strongly dependent on applied voltage. In particular K2 decreases with increasing applied voltages (Vs). The v(tm) or value of the maximum depolarization rate, and the Vll(tm) or value of the output voltage when the derivative v(tm) reaches the maximum, both increase with applied voltage (Vs). The kinetic values found indicated that mitochondrial porin is not driven by potential in the membrane, but its insertion depends (for fixed porin concentration, temperature and pH) on the lipid-protein affinity. This result supports previous authors' findings of porin insertion into mitochondrial membranes and asymmetric lipid bilayers (H. Freitag et al., Eur.J. Biochem. 74:483-492, 1993; S.M. Gasser et al.,J. Biol. Chem. 25: 3427- 3430,1983, A. Viese et al., Biophys. J., 70:321-329,1996). On the other hand, as K2 was found to be related with the conductance at the end of porin incorporation, its correlation with Vs reflects the voltage- dependence characteristics of the porin channel
The Effect of Calcium Ions on hIAPP Channel Activity: Possible Implications in T2DM
Full text linkThe calcium ion (Ca2+) has been linked to type 2 diabetes mellitus (T2DM), although the role of Ca2+ in this disorder is the subject of intense investigation. Serum Ca2+ dyshomeostasis is associated with the development of insulin resistance, reduced insulin sensitivity, and impaired glucose tolerance. However, the molecular mechanisms involving Ca2+ ions in pancreatic β-cell loss and subsequently in T2DM remain poorly understood. Implicated in the decline in β-cell functions are aggregates of human islet amyloid polypeptide (hIAPP), a small peptide secreted by β-cells that shows a strong tendency to self-aggregate into β-sheet-rich aggregates that evolve toward the formation of amyloid deposits and mature fibrils. The soluble oligomers of hIAPP can permeabilize the cell membrane by interacting with bilayer lipids. Our study aimed to evaluate the effect of Ca2+ on the ability of the peptide to incorporate and form ion channels in zwitterionic planar lipid membranes (PLMs) composed of palmitoyl-oleoyl-phosphatidylcholine (POPC) and on the aggregation process of hIAPP molecules in solution. Our results may help to clarify the link between Ca2+ ions, hIAPP peptide, and consequently the pathophysiology of T2DM
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