Papers by Fábio Fernandes
Biochimica Et Biophysica Acta-biomembranes, 2008
We present a combined theoretical (molecular dynamics, MD) and experimental (differential scannin... more We present a combined theoretical (molecular dynamics, MD) and experimental (differential scanning calorimetry, DSC) study of the effect of 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) acyl chain-labeled fluorescent phospholipid analogs (C6-NBD-PC and C12-NBD-PC) on 1,2dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayers. DSC measurements reveal that b 1 mol% of NBD-PC causes elimination of the pretransition and a large loss of cooperativity of the main transition of DPPC. Labeling with C6-NBD-PC or C12-NBD-PC shifts the main transition temperature to lower or higher values, respectively. Following our recent report on the location and dynamics of these probes (BBA 1768(BBA (2007 467-478) in fluid phase DPPC, we present a detailed analysis of 100-ns MD simulations of systems containing either C6-NBD-PC or C12-NBD-PC, focused on their influence on several properties of the host bilayer. Whereas most monitored parameters are not severely affected for 1.6 mol% of probe, for the higher concentration studied (6.2 mol%) important differences are evident. In agreement with published reports, we observed that the average area per phospholipid molecule increases, whereas DPPC acyl chain order parameters decrease. Moreover, we predict that incorporation of NBD-PC should increase the electrostatic potential across the bilayer and, especially for C12-NBD-PC, slow lateral diffusion of DPPC molecules and rotational mobility of DPPC acyl chains.
Biophysical Journal, 2003
M13 major coat protein was derivatized with BODIPY (n-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-dia... more M13 major coat protein was derivatized with BODIPY (n-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-sindacene-3-yl)methyl iodoacetamide), and its aggregation was studied in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and DOPC/1,2-dioleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DOPG) or 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE)/DOPG (model systems of membranes with hydrophobic thickness matching that of the protein) using photophysical methodologies (time-resolved and steady-state self-quenching, absorption, and emission spectra). It was concluded that the protein is essentially monomeric, even in the absence of anionic phospholipids. The protein was also incorporated in pure bilayers of lipids with a strong mismatch with the protein transmembrane length, 1,2-dierucoyl-sn-glycero-3-phosphocholine (DEuPC, longer lipid) and 1,2-dimyristoleoyl-sn-glycero-3-phosphocholine (DMoPC, shorter lipid), and in lipidic mixtures containing DOPC and one of these lipids. The protein was aggregated in the pure vesicles of mismatching lipid but remained essentially monomeric in the mixtures as detected from BODIPY fluorescence emission self-quenching. From fluorescence resonance energy transfer (FRET) measurements (donor-n-(iodoacetyl)aminoethyl-1-sulfonaphthylamine (IAEDANS)-labeled protein; acceptor-BODIPY labeled protein), it was concluded that in the DEuPC/DOPC and DMoPC/DOPC lipid mixtures, domains enriched in the protein and the matching lipid (DOPC) are formed.
Journal of Lipid Research, 2006
Phosphatidylinositol-(4,5)-bisphosphate [PI(4,5) P 2 ] plays a key role in the modulation of acti... more Phosphatidylinositol-(4,5)-bisphosphate [PI(4,5) P 2 ] plays a key role in the modulation of actin polymerization and vesicle trafficking. These processes seem to depend on the enrichment of PI(4,5)P 2 in plasma membrane domains. Here, we show that PI(4,5)P 2 does not form domains when in a fluid phosphatidylcholine matrix in the pH range of 4.8-8.4. This finding is at variance with the spontaneous segregation of PI(4,5)P 2 to domains as a mechanism for the compartmentalization of PI(4,5)P 2 in the plasma membrane. Water/bilayer partition of PI(4,5)P 2 is also shown to be dependent on the protonation state of the lipid.-Fernandes, F., L. M. S. Loura, A. Fedorov, and M. Prieto. Absence of clustering of phosphatidylinositol-(4,5)bisphosphate in fluid phosphatidylcholine. J. Lipid Res. 2006. 47: 1521-1525.
Biophysical Journal, 2004
Quantification of lipid selectivity by membrane proteins has been previously addressed mainly fro... more Quantification of lipid selectivity by membrane proteins has been previously addressed mainly from electron spin resonance studies. We present here a new methodology for quantification of protein-lipid selectivity based on fluorescence resonance energy transfer. A mutant of M13 major coat protein was labeled with 7-diethylamino-3((4#iodoacetyl)amino)phenyl-4-methylcoumarin to be used as the donor in energy transfer studies. Phospholipids labeled with N-(7-nitro-2-1,3benzoxadiazol-4-yl) were selected as the acceptors. The dependence of protein-lipid selectivity on both hydrophobic mismatch and headgroup family was determined. M13 major coat protein exhibited larger selectivity toward phospholipids which allow for a better hydrophobic matching. Increased selectivity was also observed for anionic phospholipids and the relative association constants agreed with the ones already presented in the literature and obtained through electron spin resonance studies. This result led us to conclude that fluorescence resonance energy transfer is a promising methodology in protein-lipid selectivity studies.
Biochimica Et Biophysica Acta-biomembranes, 2006
The macrolide antibiotic bafilomycin and the related synthetic compound SB 242784 are potent inhi... more The macrolide antibiotic bafilomycin and the related synthetic compound SB 242784 are potent inhibitors of the vacuolar H + -ATPases (V-ATPase). It is currently believed that the site of action of these inhibitors is located on the membrane bound c-subunits of V-ATPases. To address the identification of the critical inhibitors binding domain, their specific binding to a synthetic peptide corresponding to the putative 4th transmembrane segment of the c-subunit was investigated using fluorescence resonance energy transfer (FRET), and for this purpose a specific formalism was derived. Another peptide of the corresponding domain of the c′ isoform, was checked for binding of bafilomycin, since it is not clear if V-ATPase inhibition can also be achieved by interaction of the inhibitor with the c′-subunit. It was concluded that bafilomycin binds to the selected peptides, whereas SB 242784 was unable to interact, and in addition for bafilomycin, its interaction with the peptides either corresponding to the c-or the c′-subunit isoforms is identical. Since the observed interactions are however much weaker as compared to the very efficient binding of both bafilomycin and SB 242784 to the whole protein, it can be concluded that assembly of all V-ATPase transmembrane segments is required for an efficient interaction.
Biophysical Journal, 2008
A group of proteins with cell membrane remodeling properties is also able to change dramatically ... more A group of proteins with cell membrane remodeling properties is also able to change dramatically the morphology of liposomes in vitro, frequently inducing tubulation. For a number of these proteins, the mechanism by which this effect is exerted has been proposed to be the embedding of amphipathic helices into the lipid bilayer. For proteins presenting BAR domains, removal of an N-terminal amphipathic a-helix (H0-NBAR) results in much lower membrane tubulation efficiency, pointing to a fundamental role of this protein segment. Here, we studied the interaction of a peptide corresponding to H0-NBAR with model lipid membranes. H0-NBAR bound avidly to anionic liposomes but partitioned weakly to zwitterionic bilayers, suggesting an essentially electrostatic interaction with the lipid bilayer. Interestingly, it is shown that after membrane incorporation, the peptide oligomerizes as an antiparallel dimer, suggesting a potential role of H0-NBAR in the mediation of BAR domain oligomerization. Through monitoring the effect of H0-NBAR on liposome shape by cryoelectron microscopy, it is clear that membrane morphology is not radically changed. We conclude that H0-NBAR alone is not able to induce vesicle curvature, and its function must be related to the promotion of the scaffold effect provided by the concave surface of the BAR domain.
Biochimica Et Biophysica Acta-biomembranes, 2007
The outer membrane protein F (OmpF) is known to play an important role in the uptake of fluoroqui... more The outer membrane protein F (OmpF) is known to play an important role in the uptake of fluoroquinolone antibiotics by bacteria. In this study, the degree of binding of the fluoroquinolone antibiotic ciprofloxacin to OmpF in a lipid membrane environment is quantified using a methodology based on Förster resonance energy transfer (FRET). Analysis of the fluorescence quenching of OmpF is complex as each OmpF monomer presents two tryptophans at different positions, thus sensing two different distributions of acceptors in the bilayer plane. Specific FRET formalisms were derived accounting for the different energy transfer contributions to quenching of each type of tryptophan of OmpF, allowing the recovery of upper and lower boundaries for the ciprofloxacin-OmpF binding constant (K B ). log (K B ) was found to lie in the range 3.15-3.62 or 3.58-4.00 depending on the location for the ciprofloxacin binding site assumed in the FRET modelling, closer to the centre or to the periphery of the OmpF trimer, respectively. This methodology is suitable for the analysis of FRET data obtained with similar protein systems and can be readily adapted to different geometries.
Arquivos Brasileiros De Cardiologia, 2001
To analyze clinical and histologic findings of 50 patients with primary neoplasms of the heart in... more To analyze clinical and histologic findings of 50 patients with primary neoplasms of the heart in a tertiary referral center. From 1980 to 1998, we retrospectively analyzed 50 patients, 32 of whom were females, whose ages ranged from 9 to 73 years (mean age = 44.16+/-18 years). Most tumors were located in the left side of the heart (72%), myxoma being the most common (84%) histologic type. The other histologic types found were as follows: fibroma (4%), lipoma (2%), rhabdomyosarcoma (2%), hemangioma (2%), sarcoma (2%), angiosarcoma (2%), and lymphoma (2%). Diagnosis was established by echocardiography in 94% of the cases. Clinical findings were as follows: dyspnea (36%), weight loss (20%), palpitations (18%), chest pain (16%), fever (8%), and arthralgia (6%). All patients with thromboembolic phenomena (10%) had left atrial myxoma. Approximately 20% of the patients were asymptomatic at the initial clinical assessment. Primary cardiac tumors are a rare entity with diverse clinical and histologic findings, requiring, therefore, a high level of clinical suspicion.
Echocardiography-a Journal of Cardiovascular Ultrasound and Allied Techniques, 2006
Left ventricular outflow tract (LVOT) obstruction is predictive of a worse outcome in hypertrophi... more Left ventricular outflow tract (LVOT) obstruction is predictive of a worse outcome in hypertrophic cardiomyopathy (HCM). In a detailed Doppler echocardiographic study of 178 selected HCM patients, the group of patients (n = 73) with the obstructive form (resting peak gradient ≥ 30 mmHg) presented more hypertrophy and poorer systolic and diastolic left ventricular (LV) functions than the HCM group (n = 105) without obstruction. LVOT peak gradient was positively correlated with hypertrophy (P < 0.0001) and negatively to tissue Doppler mitral annulus systolic (P = 0.0001) and early diastolic (P < 0.0001) velocities. The gradient significantly correlated with E/Ea ratio (r = 0.67; P < 0.0001). By multiple regression, LVOT gradient was related to E/Ea, LV maximal thickness and left atrial size. In comparison with patients without obstruction, patients with obstruction presented greater hypertrophy (P < 0.0001), lower systolic and early diastolic mitral annulus velocities (both P < 0.0001), higher E/Ea ratio (P < 0.0001) and higher global function index (P < 0.0001). In HCM, beyond the effects on hypertrophy, LVOT obstruction is an independent determinant of LV functional abnormalities.
Journal of Cardiac Failure, 2003
Background: Systemic sclerosis (SSc) is a disease characterized by the fibrosis of the skin and i... more Background: Systemic sclerosis (SSc) is a disease characterized by the fibrosis of the skin and internal organs. The nature and functional significance of myocardial damage is controversial. Systematic endomyocardial biopsy in this disease has not yet been performed. Methods: The hypothesis that increased myocardial fibrous tissue deposition occurs in patients with systemic sclerosis with no signs or symptoms of heart failure and normal left systolic ventricle function was tested in 16 SSc patients and 10 controls. Endomyocardial biopsy specimens were obtained from the right ventricular septum in SSc patients. Myocardial specimens were obtained from the same area in controls. Masson's trichrome staining was used for collagen fiber identification. Interstitial (ICVF) and perivascular collagen volume fraction (PCVF) was quantified by videomorphometry. There was a significant increase in the ICVF in patients with SSc compared with the controls, in spite of normal systolic left ventricular function. However, it was not observed in the PCVF. Conclusions: It is possible to identify cardiac remodeling, characterized by myocardial fibrosis deposits, particularly within the interstitium in SSc patients before the any signs or symptoms of heart failure appear.
Journal of Cardiac Failure, 2003
Background: Endothelins are associated with cardiac remodeling. These peptides are the most power... more Background: Endothelins are associated with cardiac remodeling. These peptides are the most powerful vasoconstrictor described-whether this remodeling is a direct effect of this hormone or indirect response to a relative ischemia promoted by vasoconstrictor effect. We evaluated the role of endothelin upon myocardial fibrosis despite of its hemodynamic effects and the benefits of its antagonism. Methods and Results: We used 40 Wistar rats: control, sham operated, rats had undergone myocardial infarction (MI) and MI rats treated with SB209670 which is an ET A /ET B endothelin antagonist. We evaluated tail systolic blood pressure (BP) and left ventricular end diastolic pressure (LVED) before surgery, just after, and at the end of the study. Remodeling was studied based on interstitial collagen and MI size by an image system analysis. BP decreased in MI groups after surgery, but did not differ between treated and untreated animals. LVED had increased levels in MI groups after surgery and did not differ between them. However, ICVF had an increase in MI group but significantly less in MIϩSB209670. MI size was similar in both groups. Conclusions: Endothelin may have a pivotal role in the myocardial fibrosis by direct stimulation of collagen accumulation despite of its hemodynamic effects.
Echocardiography-a Journal of Cardiovascular Ultrasound and Allied Techniques, 2006
Background: A global function index (GFI) derived from tissue Doppler imaging (TDI) has been prop... more Background: A global function index (GFI) derived from tissue Doppler imaging (TDI) has been proposed to improve the diagnosis of hypertrophic cardiomyopathy (HCM). We aimed to evaluate the usefulness of this index in a large selected HCM population. Methods: GFI =[E/Ea]/Sa, was calculated at mitral annulus lateral and septal borders in 164 HCM patients and in 40 healthy volunteers. Group comparisons and correlations between GFI and other variables were performed. Results: Of the 164 patients, 69 (42%) had a peak gradient >30 mmHg in the left ventricle outflow tract (LVOT). GFI (lateral or septal) was not normally distributed. There were differences among controls, obstructive HCM, and nonobstructive HCM (P < 0.0001), but significant overlap of GFI values were observed between groups. GFI was correlated to septal thickness (r = 0.44; P < 0.0001), left atrial diameter (r = 0.52; P < 0.0001), and LVOT gradient (r = 0.58; P < 0.0001). Conclusion: In a selected HCM population, GFI was limited by its asymmetrical distribution and significant overlap of values between groups. Further studies are necessary to verify the reliability of GFI in the clinical practice and its position among other tissue Doppler indices.
Arquivos Brasileiros De Cardiologia, 2002
This study evaluated the effects of a new method of mitral valve replacement on left ventricular ... more This study evaluated the effects of a new method of mitral valve replacement on left ventricular (LV) remodeling and heart failure functional class. Eight patients (6 men) with severe mitral regurgitation from end-stage dilated cardiomyopathy underwent surgery. Five patients were in functional class (FC) IV, 2 were in FC III and 1 was in FC III/IV. Age ranged from 33 to 63 years. Both the anterior and posterior leaflets of the mitral valve were divided into hemileaflets. The resultant 4 pedicles were displaced under traction toward the left atrium and anchored between the mitral annulus and an implanted valvular prosthesis. The beating heart facilitated ideal chordae tendineae positioning. All patients survived and were discharged from the hospital. After a mean follow-up period of 6.5 months (1-12 m), 5 patients were in FC I; 2 in FC I/II; and 1 in FC II. The preoperative ejection fraction ranged from 19% to 30% (mean: 25.7 +/- 3.4 %), and the postoperative ejection fraction ranged from 21% to 40% (mean: 31.1 +/- 5.8%). Doppler echocardiography showed evidence of LV remodeling in 4 patients, including lateral wall changes and a tendency of the LV cavity to return to its elliptical shape. This technique of mitral valve replacement, involving new positioning of the chordae tendineae, allowed LV remodeling and improvement in FC during this brief follow-up period.
Arquivos Brasileiros De Cardiologia, 2003
Benign cardiac tumors are rare, and lipomas are among those less frequently found. We report the ... more Benign cardiac tumors are rare, and lipomas are among those less frequently found. We report the case of a 48-year-old male complaining of high blood pressure and epistaxis in the last 2 months, with a diagnosis of right atrial lipoma established on echocardiography, magnetic resonance imaging, and anatomicopathological examination. The tumor was successfully removed, and up to 42 months after surgical excision, no evidence of tumor relapse was observed.
American Heart Journal, 2005
Selected patients with hypertrophic cardiomyopathy (HCM) have 3% to 4% annual mortality as compar... more Selected patients with hypertrophic cardiomyopathy (HCM) have 3% to 4% annual mortality as compared to only 0.5% to 1.5% in nonselected patients. Our aim was to evaluate survival and prognostic factors in HCM in patients in a tertiary care center.From 1980 to 1997, 214 patients were prospectively studied, with a mean follow-up of 7 years (range 1-25 years); there were 102 male and 112 female patients, aged 37 ± 16 years (range 3-76 years). All patients had 12-lead electrocardiogram, 24-hour Holter monitor, and surface echocardiography. Univariate analysis was performed for known adverse factors such as young age, family history, syncope, functional class, atrial fibrillation, ventricular hypertrophy, left ventricular outflow tract obstruction, and nonsustained ventricular tachycardia.There were 22 deaths (10%), 15 directly related to HCM (sudden in 11). The cumulative survival rates were 94.5% at 5 years, 91% at 10 years, and 87.9% at 15 years. The annual mortality rate was 1%. Only New York Heart Association functional class III/IV and maximal ventricular wall thickness >30 mm were associated with HCM-related cardiac death.We concluded that even a referred population of HCM patients may have a relatively benign outcome. Prognosis is related to advanced functional class and degree of left ventricular hypertrophy.
Thoracic and Cardiovascular Surgeon, 2006
The aim of the study was to correlate cardiac tumors with implantation site, embolic potential an... more The aim of the study was to correlate cardiac tumors with implantation site, embolic potential and long-term evolution in patients undergoing surgery. From 1981 to 2004, 78 patients underwent surgery for cardiac tumors. 64 % were female, with a mean age of 44 years. 14 % had embolic episodes (91 % CVA). The sizes of the tumors ranged from 0.4 cm to 10 cm. The most frequent tumors were myxomas (70 %), papillary fibroelastomas and fibromas. The preferential location was the left atrium (72 %). Data were analyzed using chi (2) and Student&amp;amp;amp;amp;amp;amp;#39;s T-tests, and the Kaplan-Meyer method and curves were compared with the log-rank test. Hospital mortality was 6 % and 8.2 % in the follow-up period. Myxoma was predominantly found in the left atrium ( P &amp;amp;amp;amp;amp;amp;lt; 0.0001; OR = 12.8 and 4 - 41 95 % CI). Left atrial tumors showed less mortality ( P = 0.1; OR = 3 and 0.7 - 11.6 95 % CI), with a lower mortality for myxomas ( P &amp;amp;amp;amp;amp;amp;lt; 0.001), fewer recurrences ( P = 0.03; OR = 8.7 and 0.8 - 89 95 % CI), and a greater embolic potential than in other locations ( P = 0.03 % OR = 0.7 and 0.6 - 0.8 95 % CI). Left atrial tumors showed greater embolic potential, fewer recurrences and less mortality.
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Papers by Fábio Fernandes