[Show abstract][Hide abstract] ABSTRACT: The fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH) and DPH derivatives have been used to characterize structural and physicochemical properties of specific membrane domains. Steady-state and fluorescence decay measurements of three probes, DPH (1,6-diphenyl-1,3,5-hexatriene), TMA-DPH [1-(4-trimethyl-ammonium-phenyl)-6-phenyl-1,3,5-hexatriene], and a phosphatidylcholine derivative of DPH, DPH-pPC [2-(3-(diphenylhexatriene)propanoyl)-3-pamitoyl-L--phosphatidyl choline], have been performed in erythrocyte membranes and in lymphocyte plasma membranes. The steady-state fluorescence polarization of the three probes showed a similar trend in both membranes. In fact either in erythrocyte or in lymphocyte plasma membranes the fluorescence polarization values of DPH-pPC and TMA-DPH were similar, but significantly higher with respect to DPH. A better characterization of erythrocyte and lymphocyte plasma membranes was possible by using fluorescence decay measurements. The data suggest the possible use of different DPH derivatives to characterize specific domains in biological membranes.
No preview · Article · Dec 1994 · Journal of Fluorescence
[Show abstract][Hide abstract] ABSTRACT: The interaction between lipoproteins and the platelet membrane has been proved to cause a modification in cellular functions. We studied 12 men with insulin-dependent diabetes mellitus (IDDM), 14 men with noninsulin-dependent diabetes mellitus (NIDDM), and 26 age-matched healthy men on the same diet. We determined fluidity by measuring the fluorescence polarization (P) of the probe 1,6-diphenyl-1,3,5-hexatriene (DPH) both in platelet membranes and in lipoproteins isolated by ultracentrifugation in NaBr density gradient. The lipid composition of lipoproteins and of platelet membranes was determined by enzymatic methods. The fluidity of platelet membranes was significantly increased both in patients affected by NIDDM and in subjects with IDDM compared with normal subjects. Low-density lipoproteins (LDL) showed an increased fluidity only in NIDDM patients. A percent increase in the triglyceride content was observed in all lipoprotein fractions in diabetic subjects. Increased phospholipid content was found in the platelet membranes from IDDM and NIDDM patients. The change in LDL fluidity observed in NIDDM patients might determine altered interactions between the lipoprotein and cellular receptors. The role of lipoproteins in the modulation of the platelet membrane properties in diabetes mellitus deserves further studies.
No preview · Article · Nov 1994 · Clinical Biochemistry
[Show abstract][Hide abstract] ABSTRACT: Hepatic ethanol (EtOH) metabolism has been assumed to involve hepatocytes differently, according to their location in the hepatic acinus. This study's aim was to gain information on plasma membrane (PM) order parameter in periportal (PP) and perivenular (PV) hepatocyte-enriched fractions isolated by a digitonin-collagenase perfusion technique from rats pair-fed for 6-8 wk liquid diets containing either EtOH or isocaloric carbohydrates. Fluorescence polarization (P) studies have been performed to measure PM order parameter of PP and PV hepatocytes cultured for 2-6 h on glass cover slips and labeled with 1-[4-(trimethylamino)phenyl]-6-phenylhexa-1,3,5-triene (TMA-DPH), a specific probe for PM of living cells. Fluorescence polarization and microscopy indicated that TMA-DPH is a suitable probe to study PM order parameter in subconfluent rat hepatocyte monolayers where it labeled, after a rapid incorporation, PM of cells. In pair-fed control rats, PM order parameter was lower in PP hepatocytes than in PV cells (P = 0.366 +/- 0.013 vs. 0.381 +/- 0.021, respectively, P < 0.02; n = 7). In EtOH-treated rats, these zonal differences tended to disappear (P = 0.419 +/- 0.012 in PP cells vs. 0.417 +/- 0.007 in PV cells; n = 7). In addition, the order parameter was significantly higher either in PP or PV hepatocytes compared with pair-fed control animals (P < 0.002 and 0.003, respectively). A 30-min culture of cells in the presence of 40-200 mM EtOH significantly decreased the PM order parameter of hepatocytes isolated from pair-fed control rats with respect to EtOH-treated animals both in PP and PV cells (P < 0.01 and 0.02, respectively; n = 4).(ABSTRACT TRUNCATED AT 250 WORDS)
No preview · Article · Feb 1994 · The American journal of physiology
[Show abstract][Hide abstract] ABSTRACT: Using static and dynamic fluorescence we studied the fluorescence properties of a phosphatidylcholine analog of 1,6-diphenyl-1,3,5-hexatriene (DPH-PC) incorporated in lymphocyte plasma membranes with respect to DPH and its cationic derivative 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), in order to study if phospholipid derivatives of DPH may be used to investigate structural and physicochemical properties of specific membrane lipid domains. DPH-PC and TMA-DPH showed similar fluorescence polarization values that were significantly higher with respect to DPH, suggesting a localization of the fluorescent portion of DPH-PC in a more ordered region of the membrane which was probably due to the elecrostatic interactions between phospholipid head-groups. The localization of the fluorescent moiety of DPH-PC near the membrane surface was also supported by the study of the fluorescence decay of the three probes using frequency-domain fluorometry. The main lifetime component of DPH-PC was rather similar to that of TMA-DPH (6.74 versus 6.24, ns) but considerably lower with respect to DPH (10.52 ns), in agreement with data obtained from exponential analysis. In lymphocyte membranes obtained from concanavalin A treated cells, a significant decrease of fluorescence polarization has been shown with DPH and its phosphatidylcholine derivative, but not with TMA-DPH. In liposomes obtained from total lipids extracted from lymphocyte membranes, a decrease of fluorescence polarization has been observed only with DPH. Our results suggest that DPH-PC localizes the fluorescent portion of its molecule in membrane microenvironments of different properties with respect to those probed by DPH and TMA-DPH. The use of DPH-phospholipid derivatives and other DPH-probes may represent an useful tool to study plasma membrane heterogeneity in biological membranes.
No preview · Article · Jan 1993 · Membrane biochemistry
[Show abstract][Hide abstract] ABSTRACT: In this study we have demonstrated a transfer of a phosphatidylcholine derivative of 1,6-diphenyl-1,3,5-hexatriene (DPH-PC) from self-quenched lipid vesicles to intact lymphocytes. Membrane labeling was followed measuring the time dependent reexpression of fluorescence. The results of fluorescence quenching by 2,4,6-trinitrobenzene sulfonate and the decrease of fluorescence polarization values during incubation at 37 degrees C, suggest that the probe could remain localized at level of the plasma membrane until 20-30 minutes. DPH-PC is identical to the natural phospholipid with respect to head group structure and polarity therefore we suggest that under appropriate experimental conditions, it could represent an useful tool to study the physico-chemical properties of specific phospholipid domains in the plasma membrane of intact cells.
No preview · Article · Mar 1992 · Biochemistry international
[Show abstract][Hide abstract] ABSTRACT: Many drugs and chemicals have been shown to induce modifications of the physicochemical properties of cellular membranes. In this study we investigated the changes in fluidity of erythrocyte membrane from epileptic patients under different pharmacological treatments, with respect to healthy controls, by using trimethylammonium-1,6-diphenyl-1,3,5-hexatriene (TMA-DPH) fluorescence polarization. The increase in TMA-DPH fluorescence polarization values observed in epileptic patients indicated a decrease in membrane fluidity. Since the analysis of erythrocyte membrane composition did not reveal significant differences between the two groups studied, a correlation with membrane lipoperoxide content was tried, as different drugs and chemicals elicit in vivo alterations resulting in peroxidation of membrane lipids. Therefore the presence of peroxidation products in the blood and the possible correlation with membrane lipoperoxide were studied. Although a direct causal linkage cannot be proved we can hypothesize that exogenous compounds such as antiepileptic drugs could modify membrane fluidity by increasing membrane lipid peroxidation. Moreover the increase of peroxidative products in the blood could indicate that the peroxidative damage might propagate through the formation of new free radical species. The possibility of using erythrocyte membrane as a model system to analyze antiepileptic drug side effects is advanced.
No preview · Article · Nov 1991 · Experimental and Molecular Pathology
[Show abstract][Hide abstract] ABSTRACT: Mobilization and aggregation of intramembrane particles (IMPs) are physiological events observed in various cells. In erythrocyte membranes, aggregation of IMPs can be induced by the exposure of partially desprectrinized erythrocyte membranes to acidic pH. We investigated the association between IMPs aggregation, protein mobility, and membrane fluidity in erythrocyte membranes of healthy controls and Duchenne muscular dystrophy (DMD) patients by using electron spin resonance and specific spin labels for membrane proteins and lipids. In erythrocyte membranes of control subjects, the partial spectrin removal induced a decreased segmental motion of protein spin label indicating an increase of protein-protein interactions. Stearic acid spin labels 5- and 16-(N-oxyl-4,4'-dimethyloxazolidine) showed that the treatment induces an increase of membrane fluidity. In DMD patients, both treated and untreated erythrocyte membranes showed changes of membrane fluidity when compared to those of the controls. Our results suggest that defects in the interactions between skeletal proteins and/or between membrane and skeleton components may contribute to the alterations of erythrocyte membranes in DMD.
No preview · Article · Dec 1990 · Experimental Cell Research
[Show abstract][Hide abstract] ABSTRACT: The percentage of echinocytes induced after red cell treatment with L-α-lysophosphatidylcholine in the blood of 16 patients affected by Duchenne muscular dystropy (DMD) has been evaluated. Moreover, 15 mothers, 10 sisters, and 15 fathers were also included in the study. We found an increased level of echinocytes in dystrophic patients and in known and possible carriers. Correlations were also evaluated between echinocytes and serum enzymes used in DMD diagnosis, showing an increase of echinocytes also in DMD carriers with normal levels of serum creatine kinase, lactate dehydrogenase, and aldolase.
Our results suggest that the sensitivity of erythrocytes to L-α-lysophosphatidylcholine in DMD could be used as a diagnostic test for carrier detection.
No preview · Article · Apr 1989 · American Journal of Medical Genetics
[Show abstract][Hide abstract] ABSTRACT: Using 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5,hexatriene (TMA-DPH) as fluorescent probe, we have studied platelet membrane fluidity and thrombin induced exocytosis in ten obese children and fifteen controls. Our results indicate a decrease of membrane fluidity, as shown by an increase of fluorescence anisotropy, in platelets from obese patients in comparison with the controls. Associated with the changes in membrane fluidity, platelets of obese subjects showed a decreased sensitivity to thrombin stimulation.
No preview · Article · Dec 1988 · Biochemistry international