Evaluación de los efectos de un dispositivo de entrenamiento de los músculos inspiratorios sobre la oxigenación muscular y la activación neuromuscular de los músculos respiratorios / [Evaluation of the effects of an inspiratory muscle training device on muscle oxygenation and neuromuscular activation of the respiratory muscles]

Objetivo: Comparar los cambios en la actividad electromiográfica y oxigenación muscular periférica de los músculos intercostales, en su condición de musculatura accesoria a la respiración, en pacientes con EPOC durante la realización de un test de marcha de 6 minutos (TM6M) con y sin el uso de un dispositivo FeelBreathe (FB).Material y métodos: Se seleccionaron a 20 sujetos diagnosticados de EPOC a los que se les realizaron dos TM6M separados al menos por 60 minutos. Aleatoriamente, cada uno de los pacientes realizó dos TM6M, uno usando el dispositivo FB y el otro sin FB (SFB) y se midieron durante la realización de ambos TM6M la actividad electromiográfica (EMG) obteniéndose la raíz de la media cuadrática (RMS), y por otro lado la oxigenación tisular de los músculos intercostales a través de la variable de oxihemoglobina (HbO2).Resultados: No hubo diferencias significativas en la distancia recorrida en ambos TM6M con FC a SFB. Tanto la RMS como la oxigenación tisular mostraron valores similares entre la condición FB vs. SFB al final de ambas pruebas (32,26 ± 101,94 μV vs 28,25 ± 87,02 μV; p = 0,16 y 70,63 ± 18,80 vs 70,74 ± 16,77; p = 0,975 respectivamente).Conclusiones: El estímulo de trabajo de la musculatura intercostal durante el TM6M con el dispositivo FB no compromete la aparición de la fatiga temprana por un exceso de activación o disminución de la oxigenación de dichos músculos al nivel de intensidad del TM6M. (AU)

Objective: To compare changes in electromyographic activity and peripheral muscle oxygenation of the intercostal muscles, in their condition as accessory muscles for respiration, in patients with COPD during a 6-minute walk test (6MWT) with and without the use of a FeelBreathe (FB) device.Material and methods: 20 subjects diagnosed with COPD who underwent two 6MTMs separated by at least 60 minutes were selected. Randomly, each of the patients underwent two 6MWT, one using the FB device and the other without FB (SFB) and electromyographic activity (EMG) was measured during the performance of both 6MWT, obtaining the root mean square (RMS), and on the other hand, tissue oxygenation of the intercostal muscles through the oxyhemoglobin (HbO2) variable.Results: There were no significant differences in the distance traveled in both 6MWT with HR to SFB. Both RMS and tissue oxygenation showed similar values between the FB condition vs. SFB at the end of both tests (32.26 ± 101.94 μV vs 28.25 ± 87.02 μV; p = 0.16 and 70.63 ± 18.80 vs 70.74 ± 16.77; p = 0.975 respectively).Conclusions: The work stimulus of the intercostal muscles during the 6MWT with the FB device does not compromise the appearance of early fatigue due to excessive activation or decreased oxygenation of these muscles at the intensity level of the 6MWT. (AU)

Differential IL 10 serum production between an arm-based and a leg-based maximal resistance test.

Bench press (i.e. arm-based) and half-squat (i.e. leg-based) are exercises commonly used to increase and evaluate muscular strength. In addition to differences in the location of the muscles that participate in each exercise, the total muscle mass required for the latter is larger than that involved in the former. The aim of this study is to analyze the effects of a maximal incremental strength test when performed by bench press and by half-squat on myocellular damage, oxidative damage and the inflammatory cytokine response. Ten male athletes were subjected to half-squat and bench press incremental strength tests. Blood samples were collected at rest, 15-minutes and 24 h post-test. Hydroperoxide and malondialdehyde concentrations were determined as lipid peroxidation markers. Lactate dehydrogenase (LDH) and creatine kinase isoenzyme MB (CK-MB) activities were determined as markers of muscle damage. α-Actin concentration was determined as a marker of sarcomeric damage. Serum interleukin (IL) 6, IL10, and tumor necrosis factor alpha (TNFα) were determined to assess the inflammatory response. LDH and CK-MB values were greater at 15 min and 24 h post bench press exercise (p < 0.05). No differences were found in lipid peroxidation or α-actin. Interestingly, IL10 values were greater in response to the press bench at 24 h post-test (p < 0.05). Our results suggest that, at equivalent workloads, an arm-based exercise induced higher anti-inflammatory effects and more severe muscle damage compared with a leg-based exercise.

Efficacy of environmental friendly disinfectants against the major postharvest pathogens of stone fruits on plastic and wood surfaces.

Disinfection of surface facilities during postharvest handling operation is an important practice to avoid secondary fruit infections at stone fruit packinghouses. The aim of this work was to evaluate the effect of six environmental friendly disinfectants against Monilinia fructicola, Penicillium expansum, Rhizopus spp., and Alternaria spp. on plastic and wood surfaces. Hydrogen peroxide, peracetic acid, sodium hypochlorite, Mico-E-pro®, Proallium FRD-N®, and DMC Clean-CNS® were used as the disinfectants. Untreated and surfaces treated with water were used as controls. Plastic and wood surfaces were sampled with Rodac plates at 2 and 24 h after treatments and the number of colonies were counted. In general, all disinfectants reduce the number of viable conidia from all studied surfaces. Hydrogen peroxide used in a concentration of 150 mg L-1 was the less effective disinfectant in all studied pathogens. The commercial product Mico-E-pro® composed of oregano, onion, and orange extract at a dose of 10 mg L-1 was the most effective disinfectant. Rhizopus spp. was the pathogen more resistant to the disinfectants followed by P. expansum, M. fructicola, and Alternaria spp. Water decreased the number of conidia adhered to the surface. In addition, the untreated control showed substantial conidia reduction after 24 h of artificial inoculation.

Influence of temperature on decay, mycelium development and sporodochia production caused by Monilinia fructicola and M. laxa on stone fruits.

Brown rot on peaches and nectarines caused by Monilinia spp. results in significant economic losses in Europe. Experiments were conducted to study the effects of temperature (0-33 °C) on the temporal dynamics of decay and mycelium development and the subsequent sporulation on peaches and nectarine fruit infected by M. laxa and M. fructicola. The rates of decay and mycelium development increased with temperature from 0 °C to 25 °C for both Monilinia species. At 0 °C, decay was faster for M. laxa (0.20 cm2 days-1) than for M. fructicola (0.07 cm2 days-1); indeed, M. laxa was able to develop mycelia and sporodochia, but M. fructicola was not. At 4 and 20 °C, there were no differences in decay and mycelia development between the two Monilinia species. When temperature increased from 25 to 33 °C, the rates of fungal decay and mycelium development decreased. At 30 and 33 °C, M. fructicola decayed faster (0.94 and 1.2 cm2 days-1, respectively) than M. laxa (0.78 and 0.74 cm2 days-1, respectively) and could develop mycelia and produce sporodochia, whereas M. laxa failed at 33 °C. These results indicated that M. fructicola is better adapted to high temperatures, whereas M. laxa is better adapted to low temperatures. These results can be used to predict the relative importance of the two species during the season at a given site and to improve management strategies for brown rot in areas where both species are present.

Ensayo clínico aleatorizado del asesoramiento nutricional en pacientes desnutridos hospitalizados / Randomized clinical trial of nutritional counseling for malnourished hospital patients

Introducción. La desnutrición se asocia a un mayor riesgo de mortalidad y morbilidad, a estancias hospitalarias más largas y a un deterioro general de la calidad de vida. Este estudio se propone evaluar en pacientes desnutridos hospitalizados el impacto del asesoramiento dietético. Pacientes y métodos. Estudio prospectivo, aleatorizado, abierto, en 106 pacientes hospitalizados con desnutrición (54 grupo control, 52 en el de «intervención»). El grupo «intervención» recibió asesoramiento dietético y el grupo control el tratamiento habitual. Se determinó el estado nutricional (índice de masa corporal, parámetros analíticos, Malnutrition Universal Screening Tool), el grado de dependencia (índice de Barthel), la calidad de vida (SF-12), el grado de satisfacción de los pacientes (CSQ-8), el número y días de reingresos hospitalarios y la mortalidad. Resultados. Los pacientes sometidos a «intervención» aumentaron de peso a los 6 meses, mientras que los controles perdieron peso (diferencia en el IMC de 2,14kg/m2 [p<0,001]). El grupo «intervención» mostró mejores resultados respecto al grupo control en las puntuaciones obtenidas en el Malnutrition Universal Screening Tool (diferencia –1,29; p<0,001), Barthel (diferencia 7,49; p=0,025), SF-12 (diferencia 13,72; p<0,001), y CSQ-8 (diferencia 4,34, p<0,001), y precisaron de un menor número de reingresos (diferencia –0,37; p=0,04) y de días de rehospitalización (diferencia –6,75; p=0,035). La mortalidad y los parámetros analíticos fueron similares en grupos. Conclusiones. El asesoramiento nutricional mejoró el estado nutricional, la calidad de vida y el grado de dependencia de los pacientes, además disminuyó el número de reingresos hospitalarios (AU)

Introduction. Malnutrition is associated with an increased risk of mortality and morbidity, longer hospital stays and general loss of quality of life. The aim of this study is to assess the impact of dietary counseling for malnourished hospital patients. Patients and methods. Prospective, randomized, open-label study of 106 hospital patients with malnutrition (54 in the control group and 52 in the intervention group). The intervention group received dietary counseling, and the control group underwent standard treatment. We determined the patients’ nutritional state (body mass index, laboratory parameters, malnutrition universal screening tool), degree of dependence (Barthel index), quality of life (SF-12), degree of satisfaction (CSQ-8), the number and length of readmissions and mortality. Results. The patients who underwent the “intervention” increased their weight at 6 months, while the controls lost weight (difference in body mass index, 2.14kg/m2; p<.001). The intervention group had better results when compared with the control group in the Malnutrition Universal Screening Tool scores (difference, –1.29; p<.001), Barthel index (difference, 7.49; p=.025), SF-12 (difference, 13.72; p<.001) and CSQ-8 (difference, 4.34, p<.001) and required fewer readmissions (difference, –0.37; p=.04) and shorter stays for readmissions (difference, –6.75; p=.035). Mortality and laboratory parameters were similar for the 2 groups. Conclusions. Nutritional counseling improved the patients’ nutritional state, quality of life and degree of dependence and decreased the number of hospital readmissions (AU)

Randomized clinical trial of nutritional counseling for malnourished hospital patients. / Ensayo clínico aleatorizado del asesoramiento nutricional en pacientes desnutridos hospitalizados.

INTRODUCTION: Malnutrition is associated with an increased risk of mortality and morbidity, longer hospital stays and general loss of quality of life. The aim of this study is to assess the impact of dietary counseling for malnourished hospital patients. PATIENTS AND METHODS: Prospective, randomized, open-label study of 106 hospital patients with malnutrition (54 in the control group and 52 in the intervention group). The intervention group received dietary counseling, and the control group underwent standard treatment. We determined the patients' nutritional state (body mass index, laboratory parameters, malnutrition universal screening tool), degree of dependence (Barthel index), quality of life (SF-12), degree of satisfaction (CSQ-8), the number and length of readmissions and mortality. RESULTS: The patients who underwent the "intervention" increased their weight at 6 months, while the controls lost weight (difference in body mass index, 2.14kg/m(2); p<.001). The intervention group had better results when compared with the control group in the Malnutrition Universal Screening Tool scores (difference, -1.29; p<.001), Barthel index (difference, 7.49; p=.025), SF-12 (difference, 13.72; p<.001) and CSQ-8 (difference, 4.34, p<.001) and required fewer readmissions (difference, -0.37; p=.04) and shorter stays for readmissions (difference, -6.75; p=.035). Mortality and laboratory parameters were similar for the 2 groups. CONCLUSIONS: Nutritional counseling improved the patients' nutritional state, quality of life and degree of dependence and decreased the number of hospital readmissions.

Prescripción de actividad física en pacientes con fibromialgia / Prescribing physical activity in patients with fibromyalgia

La realización de actividad física, de forma habitual, es esencial para mantener la salud física y psíquica, y constituye un pilar fundamental en el tratamiento de diversas enfermedades. En los pacientes con fibromialgia debe incluirse de forma sistemática la actividad física en el plan terapéutico, optimizando esta prescripción para obtener el máximo beneficio (AU)

The implementation of physical activity on a regular basis is essential to maintain physical and mental health, and is a mainstay in the treatment of several diseases. In patients with fibromyalgia physical activity should be routinely included in the treatment plan, and we must optimise the prescription to obtain the maximum benefit (AU)

Beneficial effects of synthetic KL4 surfactant in experimental lung transplantation.

The aim of this study was to investigate whether intratracheal administration of a new synthetic surfactant that includes the cationic, hydrophobic 21-residue peptide KLLLLKLLLLKLLLLKLLLLK (KL4), might be effective in reducing ischaemia-reperfusion injury after lung transplantation. Single left lung transplantation was performed in Landrace pigs 22 h post-harvest. KL4 surfactant at a dose of 25 mg total phospholipid·kg body weight⁻¹ (2.5 mL·kg body weight⁻¹) was instilled at 37°C to the donor left lung (n = 8) prior to explantation. Saline (2.5 mL·kg body weight⁻¹; 37°C) was instilled into the donor left lung of the untreated group (n = 6). Lung function in recipients was measured during 2 h of reperfusion. Recipient left lung bronchoalveolar lavage (BAL) provided native cytometric, inflammatory marker and surfactant data. KL(4) surfactant treatment recovered oxygen levels in the recipient blood (mean ± sd arterial oxygen tension/inspiratory oxygen fraction 424 ± 60 versus 263 ± 101 mmHg in untreated group; p=0.01) and normalised alveolar-arterial oxygen tension difference. Surfactant biophysical function was also recovered in KL4 surfactant-treated lungs. This was associated with decreased C-reactive protein levels in BAL, and recovery of surfactant protein A content, normalised protein/phospholipid ratios, and lower levels of both lipid peroxides and protein carbonyls in large surfactant aggregates. These findings suggest an important protective role for KL4 surfactant treatment in lung transplantation.

Effect of temperature and water activity on in vitro germination of Monilinia spp.

AIMS: This study evaluated the effect of temperature (0-38 degrees C) and water activity (a(w): 0.87-0.99) on the lag phase prior to germination and the percentage of germination over time for Monilinia laxa, Monilinia fructicola and Monilinia fructigena. METHODS AND RESULTS: More than 80% of viable conidia germinated at 25 degrees C and 0.99 a(w) within 2 h for M. fructicola and M. fructigena and 4 h for M. laxa. There was no germination at 38 degrees C, and all three Monilinia spp. germinated at 0 degrees C. At the lowest a(w) (0.87), none of the Monilinia spp. was able to germinate at any of the incubation temperatures studied. Whereas at 0.90 a(w), conidia were only able to germinate at 15, 25 and 30 degrees C for the three species studied, except for M. fructicola at 15 degrees C. In contrast, at 0.95, 0.97 and 0.99 a(w), germination occurred at all studied temperatures less 38 degrees C. Generally, the lag phase was longer at low levels of a(w) (0.90-095), and differences were more evident as temperatures were far from the optimum (0-5 degrees C). CONCLUSIONS: Germination and lag phase period were markedly influenced by temperature and a(w), and in general when conditions of temperature and a(w) were suboptimal, the lag phase was longer and the percentage of germination was lower. SIGNIFICANCE AND IMPACT OF THE STUDY: Knowledge of the germination requirements of this fungus is important in order to understand their behaviour in natural situations and to provide baseline data required for the construction of new prediction models. Our study might be used to develop a predictive model to understand and control the disease caused by Monilinia spp.

Improving low water activity and desiccation tolerance of the biocontrol agent Pantoea agglomerans CPA-2 by osmotic treatments.

AIMS: To study the improvement of tolerance to low water activity (aw) and desiccation during spray drying in Pantoea agglomerans cells subjected to mild osmotic stress during growth. METHODS AND RESULTS: The micro-organism was cultured in an unmodified liquid (control) or in aw-modified media, and viability of these cells was evaluated on unstressed (0.995) and 0.96 aw stressed solid media, in order to check total viability and aw stress tolerance respectively. Significant improvements in viability on unmodified medium were observed with cells grown for 24 h in NaCl 0.98 aw, glycerol 0.98 aw and 0.97 aw and for 48 h in NaCl 0.98 aw and 0.97 aw modified media. Both yield improvements and water stress tolerance were achieved with low aw media. Cells grown for 24 h in NaCl 0.98 aw or for 48 h in NaCl 0.98 aw, 0.97 aw and 0.96 aw, glucose 0.97 aw and glycerol 0.97 aw showed improved aw stress tolerance in comparison with control cells. The best results were obtained with NaCl treatments (0.98 aw and 0.97 aw) which also exhibited better survival rates than control cells during spray-drying process and maintained their efficacy against postharvest fungal pathogens in apples and oranges. CONCLUSIONS: NaCl treatments are very appropriate for improving P. agglomerans low aw tolerance obtaining high production levels and maintaining biocontrol efficacy. SIGNIFICANCE AND IMPACT OF THE STUDY: Improving stress tolerance of biocontrol agents could be an efficient way to obtain consistency and maintain efficacy of biological control under practical conditions.

Integron-mediated antibiotic multiresistance in Acinetobacter baumannii clinical isolates from Spain.

OBJECTIVE: To determine whether non-epidemiologically related, antibiotic-resistant isolates of Acinetobacter baumannii from different geographical origins possess common type 1 integrons. METHODS: The epidemiologic relationships between seven A. baumannii strains recovered from different Spanish hospitals were established by pulsed-field gel electrophoresis, the presence of integrons being determined by PCR and DNA sequencing. RESULTS: Integron analysis showed the presence of four different integrons, containing six different known genes (aacC1, aacA4, aadA1, aadB, oxa21 and oxa37) plus an ORF. It was found that the same integron was present in different unrelated strains and that related strains could have different integrons. CONCLUSION: These results show the potential risk of integron dissemination among different strains of A. baumannii.

Effect of hydroxylation and N187-linked glycosylation on molecular and functional properties of recombinant human surfactant protein A.

The objective of this study was to determine the effects of proline hydroxylation in the collagen-like domain and Asn(187)-linked glycosylation in the globular domain on the molecular and functional properties of human surfactant protein A1 (SP-A1). To address this issue, SP-A1 was in vitro expressed in insect and mammalian cells. Insect cells lack prolyl 4-hydroxylase activity. A glycosylation-deficient mutant SP-A1 was expressed in insect cells. In this report we present evidence that hydroxylation increased the T(m) of the collagen-like domain by 9 degrees C. Proline hydroxylation affected both the arrangement of disulfide bonding and the extent of oligomerization but did not affect conformational changes in the globular domain identified by intrinsic fluorescence. Both self-association and lipid-related functions of SP-A were clearly correlated with the thermal stability of the collagen domain and the degree of oligomerization. Structural properties and lipid-related characteristics of SP-A1 expressed in mammalian cells but not in insect cells were close to that of natural human SP-A. On the other hand, the lack of glycosylation did not affect either collagen domain stability or conformational changes induced by calcium in the globular domain. However, the lack of glycosylation favored nonspecific thermally induced aggregation of the protein.

Structural analysis and lipid-binding properties of recombinant human surfactant protein a derived from one or both genes.

Surfactant protein A (SP-A) constitutes an important part of the innate immune defense in the lung. In humans there are two functional genes (SP-A1 and SP-A2). The functional importance of having two distinct chain types in human SP-A is undefined. Amino acid substitutions in the primary structure of the protein may have effects on structural stability or on activity. To address this issue, SP-A1, SP-A2, and coexpressed SP-A1/SP-A2 variants were in vitro expressed in insect cells, purified, and used for study. We found the following: (1) Human SP-A variants expressed in insect cells, derived from one gene (SP-A1 or SP-A2) or both genes, differ in the relative extent and heterogeneity of oligomerization. SP-A1 and SP-A2 exist in small oligomeric forms, whereas coexpressed SP-A1/SP-A2 products favor the formation of larger oligomers. (2) Circular dichroic and fluorescence spectroscopic studies identified structural differences between SP-A variants in the collagen domain, with SP-A2 being more stable than SP-A1 but not in the calcium binding region. Recombinant human SP-A variants expressed in insect cells exhibit a lower melting temperature compared to native human SP-A. Oligomerization does not increase the thermal stability of the collagen domain of coexpressed SP-A1/SP-A2. (3) The ability of SP-A to undergo self-aggregation and induce phospholipid and bacterial lipopolysaccharide aggregation is greater for SP-A2 than for coexpressed SP-A1/SP-A2, which in turn is greater than that observed for SP-A1. The presence of SP-A1 polypeptide chains in coexpressed products modulates functional capabilities of SP-A, which depend on both the collagen and globular domains.

Superficial disposition of the N-terminal region of the surfactant protein SP-C and the absence of specific SP-B-SP-C interactions in phospholipid bilayers.

A dansylated form of porcine surfactant-associated protein C (Dns-SP-C), bearing a single dansyl group at its N-terminal end, has been used to characterize the lipid-protein and protein-protein interactions of SP-C reconstituted in phospholipid bilayers, using fluorescence spectroscopy. The fluorescence emission spectrum of Dns-SP-C in phospholipid bilayers is similar to the spectrum of dansyl-phosphatidylethanolamine, and indicates that the N-terminal end of the protein is located at the surface of the membranes and is exposed to the aqueous environment. In membranes containing phosphatidylglycerol (PG), the fluorescence of Dns-SP-C shows a 3-fold increase with respect to the fluorescence of phosphatidylcholine (PC), suggesting that electrostatic lipid-protein interactions induce important effects on the structure and disposition of the N-terminal segment of the protein in these membranes. This effect saturates above 20% PG molar content in the bilayers. The parameters for the interaction of Dns-SP-C with PC or PG have been estimated from the changes induced in the fluorescence emission spectrum of the protein. The protein had similar K(d) values for its interaction with the different phospholipids tested, of the order of a few micromolar. Cooling of Dns-SP-C-containing dipalmitoyl PC bilayers to temperatures below the phase transition of the phospholipid produced a progressive blue-shift of the fluorescence emission of the protein. This effect is interpreted as a consequence of the transfer of the N-terminal segment of the protein into less polar environments that originate during protein lateral segregation. This suggests that conformation and interactions of the N-terminal segment of SP-C could be important in regulating the lateral distribution of the protein in surfactant bilayers and monolayers. Potential SP-B-SP-C interactions have been explored by analysing fluorescence resonance energy transfer (RET) from the single tryptophan in porcine SP-B to dansyl in Dns-SP-C. RET has been detected in samples where native SP-B and Dns-SP-C were concurrently reconstituted in PC or PG bilayers. However, the analysis of the dependence of RET on the protein density excluded specific SP-B-Dns-SP-C associations.

Role of surfactant protein A (SP-A)/lipid interactions for SP-A functions in the lung.

Surfactant protein A (SP-A), an oligomeric glycoprotein, is a member of a group of proteins named collectins that contain collagen-like and Ca(2+)-dependent carbohydrate recognition domains. SP-A interacts with a broad range of amphipathic lipids (glycerophospholipids, sphingophospholipids, glycosphingolipids, lipid A, and lipoglycans) that are present in surfactant or microbial membranes. This review summarizes SP-A/lipid interaction studies regarding the lipid system used (i.e., phospholipid vesicles, phospholipid monolayers, and lipids immobilized on silica or adsorbed on a solid support). The effect of calcium, ionic strength, and pH on the binding of SP-A to lipids and the subsequent lipid aggregation process is discussed. Current evidence suggests that hydrophobic-binding forces are involved in the peripherical association of SP-A to membranes. It is also proposed that fluid and liquid-ordered phase coexistence in surfactant membranes might favor partition of SP-A into those membranes. The binding of SP-A to surfactant membranes containing hydrophobic surfactant peptides makes possible the formation of a membrane reservoir in the alveolar fluid that is protected by SP-A against inactivation and improves the rate of surfactant film formation. In addition, the interaction of SP-A with membranes might enhance the affinity of SP-A for terminal carbohydrates of glycolipids or glycoproteins on the surface of invading microorganisms.

Cholera outbreak in southern Tanzania: risk factors and patterns of transmission.

To identify risk factors and describe the pattern of spread of the 1997 cholera epidemic in a rural area (Ifakara) in southern Tanzania, we conducted a prospective hospital-based, matched case- control study, with analysis based on the first 180 cases and 360 matched controls. Bathing in the river, long distance to water source, and eating dried fish were significantly associated with risk for cholera. Toxigenic Vibrio cholerae O1, biotype El Tor, serotype Ogawa, was isolated in samples from Ifakara's main water source and patients' stools. DNA molecular analyses showed identical patterns for all isolates.

Selective labeling of pulmonary surfactant protein SP-C in organic solution.

Pulmonary surfactant protein SP-C has been isolated from porcine lungs and treated with dansyl isothiocyanate in chloroform:methanol 2:1 (v/v) solutions,under conditions optimized to introduce a single dansyl group covalently attached to the N-terminalamine group of the protein without loss of its native thioesther-linked palmitic chains. The resulting derivative Dans-SP-C conserves the secondary structure of native SP-C as well as the ability to promote interfacial adsorption of DPPC suspensions and to affect the thermotropic behavior of DPPC bilayers. This derivative can be used to characterize lipid-protein and protein-protein interactions of a native-like SP-C in lipid/protein complexes.

Intrinsic structural differences in the N-terminal segment of pulmonary surfactant protein SP-C from different species.

Predictive studies suggest that the known sequences of the N-terminal segment of surfactant protein SP-C from animal species have an intrinsic tendency to form beta-turns, but there are important differences on the probable location of these motifs in different SP-C species. Our hypothesis is that intrinsic structural determinants of the sequence of the N-terminal region of SP-C could define conformation, acylation and perhaps surface properties of the mature protein. To test this hypothesis we have synthesized peptides corresponding to the 13-residue N-terminal sequence of porcine and canine SP-C, and studied their structural behaviour in solution and in phospholipid bilayers and monolayers. In these peptides, leucine at position 1 of both sequences has been replaced by tryptophan in order to allow their study by fluorescence spectroscopy. Far-u.v. circular dichroism spectra of the peptides in aqueous and organic solutions and in phospholipid micelles or vesicles are consistent with predicted conformational differences between the porcine and the canine sequences. Both families of peptides showed changes in their fluorescence emission spectra in the presence of phospholipids that were consistent with spontaneous lipid/peptide interactions. Both canine and porcine peptides were able to form monolayers at air-liquid interfaces, the canine peptides occupying lower area/molecule and being compressible to higher pressures than the porcine sequences. The peptides also shifted the isotherms and perturbed the packing of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) monolayers, the effects being always higher in anionic than in zwitterionic lipids, and also substantially higher in films containing canine peptide in comparison to porcine peptide. Acylation of cysteines at the N-terminal end of SP-C may modulate these intrinsic conformational features and the changes induced could be important for the development of its surface activity.