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Inhibition of anion transport in human red blood cell : effect on the electrophoretic velocity
No full textThe electrophoretic mobility exhibited by the human red blood cells (RBC), under electric field, depends on: 1) the intensity of the applied electric field, 2) the shape and the net surface charge density of the cell, 3) the characteristics of the suspending medium (pH, ionic strength, viscosity and osmolarity).
Negative cell surface charge is due to both membrane glycolipids and glycoproteins.
We studied the influence of some inhibitors of anion exchange [phenylglyoxal (PG) and three PG derivatives: 4-hydroxy-3-nitrophenylglyoxal (HNPG), 3-methoxyphenylglyoxal (3MOPG) and 4-methoxyphenylgyoxal (4MOPG)] on electrophoretic mobility.
The experiments were performed in an horizontal cylindrical capillary by the microscope method at room temperature and physiological pH; the ionic strength was 0.262 mol/l.
After treatment with the inhibitors RBCs showed a reduced electrophoretic mobility: -16.7% by PG (5 mM), -14.7, -17.6% and -18.4% by HNPG (2 mM), 3MOPG(2mM) and 4MOPG (10 mM).
The surface electrical charge density was calculated by two models: assuming the RBC to be spherical or using a flat plate model. All inhibitors reduced the surface electrical charge density: -18.7 and -21.2 by PG, -14.6 and -16.9 by HNPG, -16.9 and -19.2 by 3MOPG, -19.4 and -21.8 by 4MOPG respectively.
The major role of inhibitors probably lies in their capability to react with arginine of Band 3 masking the glycoprotein charges and thus reducing the negative surface electrical charge.
In agreement with the characteristics of the inhibitory action on the anion exchange, we found a completely reversible HNPG reduction of electrophoretic mobility while it was not the case for PG, 3MOPG and 4MOPG reduction
EFFECTS OF HYPERTONIC SOLUTIONS AND OF SOME ANION TRANSPORT INHIBITORS ON THE ELECTROPHORETIC MOBILITY OF HUMAN ERYTHROCYTES
No full textUnder the conditions of physiological pH and ionic strength, the human erythrocytes exhibit a negative electrophoretic mobility (EM) due to the negative surface charge. In this research we studied the effects on the EM of: 1) the hypertonicity of the suspending medium, 2) the inhibition of anion exchange by means of some powerful molecules (phenylglyoxal, PG and two PG derivatives: HNPG and 3-MOPG kindly given by L. Zaki, Frankfurt). The experiments were performed in a horizontal cylindrical capillary by the microscope method at room temperature. In all the experiments the erythrocyte migration velocity resulted linearly related to the applied electric field (0-15 V/cm). In isotonic solution a mean velocity (± SE) of 1.47±0.04 (n=14) μm/s V/cm was obtained. The red blood cells suspended in hypertonic medium (400 mOsm) reduced their EM of about 16.2%. The incubation (30 min) of erythrocytes in solution containing PG (5mM), HNPG (2 mM) or 3-MOPG reduced EM respectively about 16.5%, 11.5% and 13.5%. HNPG reduction was reversible, those caused by PG and 3-MOPG were not. For the inhibitors the major role possibly lies in their ability to reduce the negative surface charge interacting with transport systems. For the hypertonic solution we must take into account the changes of size and shape which might mask partially the electrical charges for a new make up of the membrane surfac
La funzione gastrointestinale : capitolo 17
No full text17.1 La struttura del tubo digerente 17.2 Il controllo delle funzioni gastrointestinali 17.2.1 Il sistema nervoso 17.2.1.1 Il sistema nervoso enterico 17.2.2 Il sistema endocrino enterico 17.2.3 La peristalsi e la segmentazione 17.2.3.1 L’attività elettrica della muscolatura liscia gastrointestinale 17.3 La motilità dell’apparato gastrointestinale 17.3.1 Masticazione e deglutizione 17.3.2 La motilità gastrica 17.3.3 Movimenti dell’intestino tenue 17.3.4 Movimenti dell’intestino crasso 17.4 Le secrezioni del canale alimentare 17.4.1 Le ghiandole salivari 17.4.1.1 Modificazioni del secreto acinoso Scheda 17.1 Il muco 17.4.2 Le secrezioni gastriche 17.4.2.1 Il meccanismo di secrezione acida 17.4.2.2 La produzione di muco 17.4.2.3 Le proteasi 17.4.2.4 La chimosina e la coagulazione del latte Scheda 17.2 Ulcere gastriche e duodenali 17.4.2.5 Gli ormoni 17.4.3 La funzione esocrina del pancreas 17.4.3.1 Struttura del pancreas 17.4.3.2 Il succo pancreatico 17.4.3.3 Controllo della secrezione pancreatica 17.4.4 Il fegato e la produzione di bile 17.4.4.1 Il sistema vascolare epatico 17.4.4.2 Il sistema biliare e la bile 17.4.4.3 Regolazione della secrezione biliare 17.4.4.4 Le funzioni metaboliche del fegato 17.4.5 Le secrezioni intestinali 17.5 Digestione e assorbimento 17.5.1 L’enterocita e la superficie di assorbimento 17.5.2 Assorbimento di acqua ed elettroliti 17.5.2.1 Assorbimento di acqua 17.5.2.2 Assorbimento di Na+, K+, Cl– e HCO3– 17.5.2.3 Assorbimento del calcio 17.5.2.4 Assorbimento di ferro 17.5.3 Digestione e assorbimento dei carboidrati Scheda 17.3 Intolleranza al lattosio Scheda 17.4 Terapia orale di idratazione (ORT) 17.5.4 Digestione e assorbimento delle proteine 17.5.4.1 Assorbimento di proteine e peptidi 17.5.5 Digestione e assorbimento dei lipidi Scheda 17.5 Celiachia, una patologia da riconoscere 17.5.5.1 Riassorbimento degli acidi biliari 17.5.6 Assorbimento delle vitamine 17.5.6.1 Assorbimento della vitamina B12 17.6 Il sistema immunitario gastrointestinale 17.7 L’attività microbica nel canale alimentare Scheda 17.6 Il microbiota intestinale (Giovanna Traina
CALORIMETRIA DIFFERENZIALE A SCANSIONE DI MISCELE FOSFOLIPIDICHE DA MEMBRANE DI GLOBULI ROSSI
No full textNegli ultimi anni particolare attenzione è stata posta alle relazioni tra funzione delle membrane, le loro proprietà chimico-fisiche e la composizione in acidi grassi, prevalentemente della frazione fosfolipidica considerato il fatto che fosfolipidi, trovati nelle membrane biologiche, derivano le loro proprietà idrofobiche dagli acidi grassi costituenti.
Le correlazioni più frequentemente analizzate sono quelle tra componenti di membrana e i loro acidi grassi modificati, in vivo, da alterazioni della composizione lipidica della dieta o, in vitro, mediante opportune modifiche del mezzo di coltura.
Somministrando diete diverse a bovine da latte abbiamo creato una “banca” di campioni di membrane di globuli rossi a diversa composizione lipidica (1, 2).
Scopi di questo studio sono stati: 1) l’analisi della composizione delle miscele di fosfolipidi estratti dalle membrane, 2) la valutazione delle correlazioni tra composizione e proprietà termotropiche delle miscele.
I lipidi totali sono stati estratti dagli eritrociti bovini con miscele di cloroformio/metanolo in diversi rapporti e le miscele lipidiche sono state frazionate mediante cromatografia su colonna.
Le singole specie fosfolipidiche sono state separate in HPTLC e quantificate mediante densitometria a scansione.
La composizione in acidi grassi dei fosfolipidi di membrana è stata valutata mediante gas-liquido cromatografia e dispersioni acquose tamponate sono state esaminate mediante calorimetria differenziale a scansione.
Per ogni campione è stato calcolato il contenuto di acidi grassi saturi (S), insaturi (U) e l’indice di insaturazione (UI).
I principali acidi grassi presenti (mediamente ≥ 10% in peso) sono risultati: C16:0, C18:0, C18:1, C18:2 e C24:0.
Le miscele di fosfolipi mostrano una o due transizioni di fase a temperature comprese tra 17.7 e 40.0 °C. La prima temperatura di transizione è compresa tra 17.7 e 26.0 °C.
UI e U/S non risultano essere correlati al contenuto percentuale delle diverse specie molecolari di fosfolipidi (PC, PE, SM, PS+PI) e di PC+SM, costituenti principali del foglietto esterno del doppio strato lipidico.
Esiste, invece, una correlazione lineare tra la prima temperatura di transizione ed UI (P < 0.05) e U/S (P < 0.01). La prima temperatura di transizione diminuisce all’aumentare di UI o del rapporto U/S.
References
1. G. Monticelli, S. Rapelli, G. Montorfano, P. Magistretti and B. Berra- “Red blood cell membrane composition following diet manipulation in the cow” Riv. Ital. Sostanze Grasse 67: 507-515 (1990)
2. G. Monticelli, M. Masserini,G. Lercker, T. Beringhelli, P. Marciani, E. Calappi and B. Berra – “Red blood cell membrane composition following diet manipulation in the cow. II: phospholipid fatty acid distribution and physico-chemical characteristics of membrane and its constituents” Riv. Ital. Sostanze Grasse 69: 189-199 (1992
Meccanismi di trasporto e barriere biologiche : capitolo 3
No full text3.1 La membrana cellulare: composizione, struttura e funzione
3.1.1 I costituenti di membrana
3.1.2 Asimmetria della membrana cellulare
3.2 Meccanismi di trasporto passivi e attivi
3.2.1 Filtrazione
3.2.2 Osmosi
Scheda 3.1 Pressione osmotica, osmosi e osmolarità
Scheda 3.2 Diffusione e flussi unidirezionali
3.2.3 Filtrazione e osmosi
Scheda 3.3 Il rapporto superficie/volume
3.2.4 Osmosi e diffusione
3.2.5 Potenziale di flusso
3.2.6 Potenziale di diffusione
Scheda 3.4 Volume molare parziale
3.2.7 Presenza di una d.d.p.
Scheda 3.5 Il potenziale elettrochimico e la legge di Nernst
Scheda 3.6 Equilibrio di Gibbs-Donnan
3.2.8 Flussi e forze
3.3 Diffusione delle molecole lipofile attraverso la membrana
3.4 Trasporto facilitato
3.4.1 Cinetica del trasporto facilitato
3.5 Trasporto attivo. Il caso della pompa Na+-K+
3.6 Endocitosi ed esocitosi
3.7 Gli epiteli: barriere complesse
3.7.1 Struttura dell’epitelio
Scheda 3.7 Il profilo elettrico degli epiteli
Scheda 3.8 I canali per l’acqua, le acquaporin
bGH IN ANIMAL BREEDING (DAIRY COWS). POSSIBLE EFFECTS ON RBC MEMBRANE COMPOSITION AND PROPERTIES
No full textGrowth hormone (bGH) treatment, useful to improve milk production, can have nutritional and physiological implications. The improved milk production must be supported by an increased concentration of nutrients administered to treated cows. In fact bGH can induce fat mobilization with a resulting high level of blood NEFA.
The lipid structural organization is important for biological membrane function and it may change as a consequence of different factors. Red blood cell (RBC) membrane may exchange fats with plasma lipids and the membrane lipids greatly influence membrane properties.
The aim of this research has been to analyze the influence of bGH treatment on RBC membrane lipid content and compositio
Age related changes in function and physicochemical properties of rat jejunal brush border membrane after chronic ethanol administration
No full text1. We investigated the chronic effects of a 4 week treatment with ethanol on functions and physicochemical properties of BBM of young and adult rats (2 and 7 months old respectively). 2. In the ethanol treated groups the cholesterol/phospholipid and the protein/lipid ratios as well as the d-glucose uptake and lactase specific activity and Vmax were increased. In spite of a minor alcohol consumption the adult group was the more affected. 3. Membranes from the ethanol fed rats were less fluid and more tolerant to the in vitro addition of ethanol
ERYTHROCYTE MEMBRANE. EFFECTS OF AGE AND CHRONIC ETHANOL TREATMENT
No full textChronic ethanol consumption was demostrated to affect cell membrane lipid composition and fluidity.
Studies on red blood cell (RBC) membrane of alcoholics showed a decrease in sialic acid content and a disorganization of the outer membrane leaflet. In addition ageing seems to affect RBC survival in circulating blood and to modify the presence of sialic acid on the outer surface of RBC membrane.
In order to study the biochemical and biophysical properties of RBC membrane and the possible differences in the adaptive capability to ethanol chronic exposure during ageing we considered the membrane composition and the surface electric charge density.
Albino male rats (Crl:(WI)BR Charles River Italiana), aged 2 months (young) and 7 months (adult) at the time of the experiment, were divided in two groups (ethanol-treated and control) and fed for 24 days a liquid diet (Lieber - DeCarli formula), in which ethanol or carbohydrates represented 36 % of the caloric content. RBC membranes were prepared and analyzed as previously reported
(Monticelli et al. 1992). To estimate the membrane glycoprotein content, the defatted residue was assayed for neutral sugars and for sialic acid. Cell electrophoretic mobility measurements (MEF) were performed on RBC immediately after collected in a horizontal cylindrical capillary by the microscope method; electric surface charge density was calculated both assuming RBC to be spherical and using a flat plate model.
Our experiments showed that ethanol treatment does not affect the electrophoretic mobility and the electric surface charge density of RBC, which increase in adult animals aside from treatment.
Since electric surface charge is due to the presence of sialic acid on the outer surface of the membrane as expected after MEF experiments, gangliosidic sialic acid resulted not affected by ethanol treatment but it seems to be lower in adult treated animals than in the other groups. Nevertheless our data seem to indicate an increase of glycoprotein sialic acid in adult rats. According to the decreased galactosyltranspherase activity found in the enterocyte microsomes and in sinaptosomes of the same animals (Omodeo-Salè et al. 1994, Lindi et al. 1995), the glycoprotein neutral hexose content of RBC decreases while glycolipid neutral hexose content increases indicating that ethanol affects differently the various enzymatic systems.
In conclusion our experiments underlined the ethanol effect on membrane composition suggesting adult rat membranes react more than the young. It is not the ethanol but ageing exerting effects on RBC electrophoretic mobility.
Lindi et al. (1995) Alcohol: in press.
Monticelli et al. (1992) Riv. It. Sostanze Grasse 67: 507.
Omodeo-Salè et al. (1994) Alcohol 11: 301
IRON AND ZINC TRANSPORT : OXIDATIVE STRESS-MODULATION OF DCT1
No full textIn humans, iron plays a key role in a number of metabolic functions. Its deficiency as well as its overload leads to important pathologies. Excess of free iron can amplify the effect of oxidative stress, via Fenton's reaction, associated with a great number of diseases such as ischemia-reperfusion injury, inflammation, carcinogenesis and degenerative diseases of the SNC.
The functional properties and the localisation of the divalent cation transporter (DCT1/Nramp2/DMT1) in recycling endosomes and in plasma membranes indicate its key role in the direct cellular iron uptake and in the transferrin-receptor mediates pathway. Though DCT1 expression is regulated at a post-transcriptional level via the IRP/IRE system, in order to prevent iron overload in oxidative stress condition, a direct regulation of the protein in the plasma membrane could represent an important defensive mechanism of the cell.
To study the influence of redox reagents on DCT1 function we used the Xenopus laevis oocyte expression system and analysed the transport activity with radiotracer 55Fe2+ and 65Zn2+ -uptake assays and electrophysiology experiments.
Our radiotracer experiments confirmed the capability of DCT1 to mediate Zn uptake previously suggested on the basis of electrophysiology experiments. Since the characteristics of Zn2+ induced DCT1 mediated current resembled the characteristics of the iron current, we performed our experiments with Zn2+ which does not react with the used redox reagents.
Hydrogen peroxide treatment resulted in an inhibition of about 40 % of both Zn2+ uptake and Zn2+ induced DCT1 mediated current in oocytes. DCT1 sequence contains several amino acid residues that could be targets for oxidant reagents. Therefore Hg2+ administration exerted the same inhibition than H2O2 and, in both cases, DCT1 function was fully recovered after perfusion with dithiotreitol strongly indicating the involvement of cysteine residues. Mutagenesis experiments allowed us to identify a cysteine that is involved in the mechanism of DCT1 inhibition. C248 is located in the forth-outer loop of the predicted secondary structure of the transporter and could represent a sensor that allows the cell to activate defensive mechanism. In fact reactive oxygen species can be released from some kind of cells like phagocytes to affect target cells. The finding of a direct inhibition of iron transport appears to have relevant physiological interest to understand the regulation of iron metabolism and the defensive mechanisms to preserve cell from oxidative injury
Intestinal sugar transport during ageing
No full textAgeing effects on sugar intestinal transport were studied by using the everted sac and the brush-border membrane vesicle techniques. Four age groups of rats were used: very young, young, adult and old animals. Net transintestinal transport of d-glucose and intracellular sugar accumulation were greater in young than in very young, adult and old rats. Net Na+ transport was high in very young and young animals and then it declined with age. In brush-border membrane vesicle experiments d-glucose overshoot was smaller in the groups of animals where net sugar transport was less. In old rats, however, the overshoot did not occur. Short-circuiting of vesicles with valinomycin showed that the driving forces for sugar accumulation, i.e. the chemical potential gradient of Na+ and the electrical potential gradient, played different roles during ageing. In very young animals the chemical potential gradient seems to be responsible for d-glucose overshoot; in young rats both gradients are important while in adult animals the electrical potential gradient represents the main driving force
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