Long-term memory in Staphylococcus aureus α-hemolysin ion channel kinetics.

The kinetics of an ion channel are classically understood as a random process. However, studies have shown that in complex ion channels, formed by multiple subunits, this process can be deterministic, presenting long-term memory. Staphylococcus aureus α-hemolysin (α-HL) is a toxin that acts as the major factor in Staphylococcus aureus virulence. α-HL is a water-soluble protein capable of forming ion channels into lipid bilayers, by insertion of an amphipathic  ß-barrel. Here, the α-HL was used as an experimental model to study memory in ion channel kinetics. We applied the approximate entropy (ApEn) approach to analyze randomness and the Detrended Fluctuation Analysis (DFA) to investigate the existence of long memory in α-HL channel kinetics. Single-channel currents were measured through experiments with α-HL channels incorporated in planar lipid bilayers. All experiments were carried out under the following conditions: 1 M NaCl solution, pH 4.5; transmembrane potential of + 40 mV and temperature 25 ± 1 °C. Single-channel currents were recorded in real-time in the memory of a microcomputer coupled to an A/D converter and a patch-clamp amplifier. The conductance value of the α-HL channels was 0.82 ± 0.0025 nS (n = 128). The DFA analysis showed that the kinetics of α-HL channels presents long-term memory ([Formula: see text] = 0.63 ± 0.04). The ApEn outcomes showed low complexity to dwell times when open (ApEno = 0.5514 ± 0.28) and closed (ApEnc = 0.1145 ± 0.08), corroborating the results of the DFA method.

A new SYBR Green real-time PCR to detect SARS-CoV-2.

Phylogenetic analysis has demonstrated that the etiologic agent of the 2020 pandemic outbreak is a betacoronavirus named SARS-CoV-2. For public health interventions, a diagnostic test with high sensitivity and specificity is required. The gold standard protocol for diagnosis by the Word Health Organization (WHO) is RT-PCR. To detect low viral loads and perform large-scale screening, a low-cost diagnostic test is necessary. Here, we developed a cost-effective test capable of detecting SARS-CoV-2. We validated an auxiliary protocol for molecular diagnosis with the SYBR Green RT-PCR methodology to successfully screen negative cases of SARS-CoV-2. Our results revealed a set of primers with high specificity and no homology with other viruses from the Coronovideae family or human respiratory tract pathogenic viruses, presenting with complementarity only for rhinoviruses/enteroviruses and Legionella spp. Optimization of the annealing temperature and polymerization time led to a high specificity in the PCR products. We have developed a more affordable and swift methodology for negative SARS-CoV-2 screening. This methodology can be applied on a large scale to soften panic and economic burden through guidance for isolation strategies.

Saccharomyces boulardii as therapeutic alternative in experimental giardiasis.

AIMS: The objective of this study was to evaluate the effect of treatment with the probiotic Saccharomyces boulardii with or without metronidazole in experimental giardiasis. METHODS AND RESULTS: The effect of treatment with S. boulardii with or without metronidazole on the intestinal mucosa, the antioxidant defence system and the parasitic load was determined in experimental giardiasis. Eight groups of animals with infection and/or treatment with the probiotic and/or drugs for 1 week after infection with Giardia lamblia were used. A reduction of approximately 90% in the parasitic load was observed in all the treated groups. Saccharomyces boulardii attenuated the damage caused by infection in the intestinal mucosa preserving its architecture and inhibiting the oxidative stress induced by parasite and metronidazole. CONCLUSIONS: Saccharomyces boulardii was effective alone or in combination with metronidazole in resolving already established G. lamblia infection. SIGNIFICANCE AND IMPACT OF THE STUDY: These results suggest the use of S. boulardii as an alternative treatment for giardiasis mainly in cases of resistance or intolerance to conventional treatment.

Invasive Lactococcus lactis producing mycobacterial Hsp65 ameliorates intestinal inflammation in acute TNBS-induced colitis in mice by increasing the levels of the cytokine IL-10 and secretory IgA.

AIMS: To investigate the anti-inflammatory activity of an invasive and Hp65-producing strain Lactococcus lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) in acute 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis in mice as an innovative therapeutic strategy against Crohn's disease (CD). METHODS AND RESULTS: The pXYCYT:Hsp65 plasmid was transformed into the L. lactis NCDO2118 FnBPA+ strain, resulting in the L. lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) strain. Then, the functionality of the strain was evaluated in vitro for Hsp65 production by Western blotting and for invasion into Caco-2 cells. The results demonstrated that the strain was able to produce Hsp65 and efficiently invade eukaryotic cells. Subsequently, in vivo, the anti-inflammatory capacity of the recombinant strain was evaluated in colitis induced with TNBS in BALB/c mice. Oral administration of the recombinant strain was able to attenuated the severity of colitis by mainly reducing IL-12 and IL-17 levels and increasing IL-10 and secretory immunoglobulin A levels. CONCLUSIONS: The L. lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) strain contributed to a reduction in inflammatory damage in experimental CD. SIGNIFICANCE AND IMPACT OF THE STUDY: This study, which used L. lactis for the production and delivery of Hsp65, has scientific relevance because it shows the efficacy of this new strategy based on therapeutic protein delivery into mammalian enterocytes.

Notch signaling in human iPS-derived neuronal progenitor lines from Focal Cortical Dysplasia patients.

Focal Cortical Dysplasia (FCD), a common type of Malformations of cortical development, may result from an early disturbance in the migration and final arrangement of the cortical architecture of immature neurons. FCD type II is now known to be due to a post-zygotic somatic mutation that involves the mTOR and AKT pathways. The aim of the present study was to investigate the possible differences in neurogenesis and neurodifferentiation of iPSCs (induced pluripotent stem cells) from fibroblasts of individuals affected by FCD type II (2) and normal individuals (2). iPSCs were generated from skin fibroblasts of FCD individuals and healthy individuals. The reprogramming was done through the fibroblasts exposure to viral vectors containing the OCT4, KLF4, SOX2, and c-MYC genes and the clones were characterized by immunohistochemistry. iPSCs were neurodifferentiated and analyzed at the 14th, 22nd and 35th days. We also analyzed the cerebral cortex tissue, fibroblasts and iPSCs cells from the individuals. Through qRT-PCR, the expression of 4 genes involved in Notch signaling process were quantified. In general, individuals with dysplasia presented increase and decrease in the relative quantification in the most genes analyzed compared to control individuals in all processes and study groups. We suggest that, during embryonic neurogenesis, the neural precursor cells of FCD type II individuals present increase and decrease in gene expression in the Notch signaling pathway causing cortical formation disorders and can be seen as a candidate for the developmental changes observed in the cerebral cortex of individuals with FCD type II. This altered gene expression may be related to brain formation with dysplasia.

Sensitivity for palpating lumbopelvic soft- tissues and bony landmarks and its associated factors: A single-blinded diagnostic accuracy study.

BACKGROUND: Evidence on the diagnostic performance of palpatory methods and possible confounding factors is scarce. OBJECTIVES: To examine the sensitivity of palpatory methods for location of lumbopelvic landmarks and to assess its association with personal characteristics. METHODS: Eighty-three participants (41 men, 55.6 (16.5) years, 25.9 (4.8) kg/m2 [mean (SD)]) were enrolled in this single-blinded study. Fourteen body and softy-tissue landmarks were sequentially palpated from the spinous process of L4 to the ischial tuberosity. CT-scan images were used to assess what landmark was located. RESULTS: Sensitivity for location was in range 22-86% for soft-tissues and 26-69% for bony landmarks. Reduction in sensitivity was observed from the quadratus lumborum to the inferior and lateral angle of the sacrum (86-26% and 75-33%, left and right sides, respectively). Palpations of L4 and L5 spinous processes were systematically more cephalic than other landmarks. Gender was weakly correlated to almost all landmarks (rp⁢b= 0.333 or weaker). Body mass index was weakly correlated to the accurate location of ILAS and quadratus lumborum, great trochanter, PSIS, and piriformis (rp⁢b=-0.307 or weaker). CONCLUSIONS: Systematic and propagation errors were present using sequential palpatory methods. Palpation in men was more sensitive and higher BMI was associated with lower sensitivity for lumbopelvic landmarks.

Correlations between forced oscillation technique parameters and pulmonary densitovolumetry values in patients with acromegaly.

The aims of this study were to evaluate the forced oscillation technique (FOT) and pulmonary densitovolumetry in acromegalic patients and to examine the correlations between these findings. In this cross-sectional study, 29 non-smoking acromegalic patients and 17 paired controls were subjected to the FOT and quantification of lung volume using multidetector computed tomography (Q-MDCT). Compared with the controls, the acromegalic patients had a higher value for resonance frequency [15.3 (10.9-19.7) vs 11.4 (9.05-17.6) Hz, P=0.023] and a lower value for mean reactance [0.32 (0.21-0.64) vs 0.49 (0.34-0.96) cm H2O/L/s2, P=0.005]. In inspiratory Q-MDCT, the acromegalic patients had higher percentages of total lung volume (TLV) for nonaerated and poorly aerated areas [0.42% (0.30-0.51%) vs 0.25% (0.20-0.32%), P=0.039 and 3.25% (2.48-3.46%) vs 1.70% (1.45-2.15%), P=0.001, respectively]. Furthermore, the acromegalic patients had higher values for total lung mass in both inspiratory and expiratory Q-MDCT [821 (635-923) vs 696 (599-769) g, P=0.021 and 844 (650-945) vs 637 (536-736) g, P=0.009, respectively]. In inspiratory Q-MDCT, TLV showed significant correlations with all FOT parameters. The TLV of hyperaerated areas showed significant correlations with intercept resistance (rs=-0.602, P<0.001) and mean resistance (rs=-0.580, P<0.001). These data showed that acromegalic patients have increased amounts of lung tissue as well as nonaerated and poorly aerated areas. Functionally, there was a loss of homogeneity of the respiratory system. Moreover, there were correlations between the structural and functional findings of the respiratory system, consistent with the pathophysiology of the disease.

Correlations between forced oscillation technique parameters and pulmonary densitovolumetry values in patients with acromegaly

The aims of this study were to evaluate the forced oscillation technique (FOT) and pulmonary densitovolumetry in acromegalic patients and to examine the correlations between these findings. In this cross-sectional study, 29 non-smoking acromegalic patients and 17 paired controls were subjected to the FOT and quantification of lung volume using multidetector computed tomography (Q-MDCT). Compared with the controls, the acromegalic patients had a higher value for resonance frequency [15.3 (10.9-19.7) vs 11.4 (9.05-17.6) Hz, P=0.023] and a lower value for mean reactance [0.32 (0.21-0.64) vs 0.49 (0.34-0.96) cm H2O/L/s2, P=0.005]. In inspiratory Q-MDCT, the acromegalic patients had higher percentages of total lung volume (TLV) for nonaerated and poorly aerated areas [0.42% (0.30-0.51%) vs 0.25% (0.20-0.32%), P=0.039 and 3.25% (2.48-3.46%) vs 1.70% (1.45-2.15%), P=0.001, respectively]. Furthermore, the acromegalic patients had higher values for total lung mass in both inspiratory and expiratory Q-MDCT [821 (635-923) vs 696 (599-769) g, P=0.021 and 844 (650-945) vs 637 (536-736) g, P=0.009, respectively]. In inspiratory Q-MDCT, TLV showed significant correlations with all FOT parameters. The TLV of hyperaerated areas showed significant correlations with intercept resistance (rs=−0.602, P<0.001) and mean resistance (rs=−0.580, P<0.001). These data showed that acromegalic patients have increased amounts of lung tissue as well as nonaerated and poorly aerated areas. Functionally, there was a loss of homogeneity of the respiratory system. Moreover, there were correlations between the structural and functional findings of the respiratory system, consistent with the pathophysiology of the disease.

Effect of different helminth extracts on the development of asthma in mice: The influence of early-life exposure and the role of IL-10 response.

It is not currently clear whether different parasites have distinct effects on the airway inflammatory response in asthma and whether exposure in early life to helminths have a stronger impact in a potential inhibitory effect on asthma. The aim of this study is to evaluate the effect of exposure to different helminth extracts on the development of allergic pulmonary response in mice, including early-life exposure. Different helminth extracts (Angiostrongylus costaricensis, Angiostrongylus cantonensis and Ascaris lumbricoides) were studied in female adult BALB/c and C57BL/6 IL-10-deficient mice in a protocol of murine asthma, injected intraperitoneally in different periods of exposure (early, pre-sensitization and post-sensitization). Cell counts in bronchoalveolar lavage (BAL), eosinophil peroxidase (EPO) from lung tissue, cytokine levels from BAL/spleen cell cultures, and lung histology were analyzed. Airway cellular influx induced by OVA was significantly inhibited by extracts of A. cantonensis and A. lumbricoides. Extracts of A. lumbricoides and A. costaricensis led to a significant reduction of IL-5 in BAL (p < 0.001). Only the exposure to A. lumbricoides led to an increased production of IL-10 in the lungs (p < 0.001). In IL-10-deficient mice exposed to A. costaricensis pre-sensitization, eosinophil counts and IL-5 levels in BAL and EPO in lung tissue were significantly reduced. In the early exposure to A. cantonensis, lung inflammation was clearly inhibited. In conclusion, different helminth extracts inhibit allergic lung inflammation in mice. IL-10 may not play a central role in some helminth-host interactions. Early exposure to helminth extracts could be a potential strategy to explore primary prevention in asthma.

Relationships between the pulmonary densitometry values obtained by CT and the forced oscillation technique parameters in patients with silicosis.

OBJECTIVE: To evaluate the correlations between pulmonary densitometry values and forced oscillation technique (FOT) parameters in patients with silicosis. METHODS: This cross-sectional study comprised 36 non-smoker patients with silicosis and 20 matched control subjects who were submitted to FOT and multidetector CT (MDCT). RESULTS: Compared with the control subjects, the MDCT evaluation demonstrated that patients with silicosis exhibited greater total lung mass. These patients also had larger non-aerated and poorly aerated compartments, which included nodules and scarring. Compared with the control subjects, FOT evaluation demonstrated that patients with silicosis exhibited changes in both reactive and resistive properties of the respiratory system. In these patients, there was a greater heterogeneity of the respiratory system and increased work of breathing. Significant correlations between non-aerated compartment size and FOT parameters that reflect the non-homogeneity of the respiratory system were observed. The dynamic compliance of the respiratory system was negatively correlated with non-aerated compartment size, while the impedance at 4 Hz was positively correlated with non-aerated compartment size. CONCLUSION: Patients with silicosis have heavier lungs. In these patients, a larger non-aerated compartment is associated with a worsening of lung function. A more significant pulmonary involvement is associated with a loss of homogeneity and increased mechanical load of the respiratory system. Advances in knowledge The findings provided by both pulmonary densitometry and FOT may add valuable information to the subjective analysis of silicosis; however, more studies are necessary to evaluate the potential use of these methods for assessing disease progression.

The expression of pluripotency genes and neuronal markers after neurodifferentiation in fibroblasts co-cultured with human umbilical cord blood mononuclear cells.

Human umbilical cord blood is an attractive source of stem cells; however, it has a heterogeneous cell population with few mesenchymal stem cells. Cell reprogramming induced by different methodologies can confer pluripotency to differentiated adult cells. The objective of this study was to evaluate the reprogramming of fibroblasts and their subsequent neural differentiation after co-culture with umbilical cord blood mononuclear cells. Cells were obtained from four human umbilical cords. The mononuclear cells were cultured for 7 d and subsequently co-cultured with mouse fibroblast NIH-3T3 cells for 6 d. The pluripotency of the cells was evaluated by RT-PCR using primers specific for pluripotency marker genes. The pluripotency was also confirmed by adipogenic and osteogenic differentiation. Neural differentiation of the reprogrammed cells was evaluated by immunofluorescence. All co-cultured cells showed adipogenic and osteogenic differentiation capacity. After co-cultivation, cells expressed the pluripotency gene KLF4. Statistically significant differences in cell area, diameter, optical density, and fractal dimension were observed by confocal microscopy in the neurally differentiated cells. Contact in the form of co-cultivation of fibroblasts with umbilical cord blood mononuclear fraction for 6 d promoted the reprogramming of these cells, allowing the later induction of neural differentiation.

Assessment of extended-spectrum ß-lactamase, KPC carbapenemase and porin resistance mechanisms in clinical samples of Klebsiella pneumoniae and Enterobacter spp.

The emergence and spread of resistance mechanisms in Gram-negative bacilli has complicated the treatment of serious nosocomial infections. Current automated systems for detection of Klebsiella pneumoniae carbapenemase (KPC)-producing isolates are unreliable. One possible straightforward alternative method is evaluation of ertapenem resistance. However, the accuracy of this method is affected by other resistance mechanisms such as AmpC gene expression or extended-spectrum ß-lactamase production associated with porin loss. This study included 128 samples of K. pneumoniae and Enterobacter spp. that were non-susceptible to ertapenem. The disk diffusion and Etest method were applied to determine susceptibility to imipenem, meropenem and ertapenem. Isolates exhibiting intermediate or complete resistance to ertapenem were evaluated for resistance mechanisms. bla(TEM), bla(SHV), bla(CTX-M), bla(CTX-M-2) and bla(KPC) genes were tested for by PCR, and the presence of outer membrane protein was investigated by dot-blot assay. bla(TEM) was detected in 52.9% and 10.3%, bla(SHV) in 29.4% and 0.94%, bla(CTX-M) in 41.4% and 1.9% and bla(CTX-M-2) in 23.5% and 1.9% of K. pneumoniae and Enterobacter cloacae isolates, respectively. The bla(KPC) gene was present in 12.6% of Enterobacter spp. isolates. OmpC and OmpF were present in 6.6% of E. cloacae isolates. These results indicate that several resistance mechanisms contribute to potential therapeutic failure of carbapenem therapy and point to the need for better detection methods and surveillance strategies.

The protective effects of guaraná extract (Paullinia cupana) on fibroblast NIH-3T3 cells exposed to sodium nitroprusside.

The antioxidant effects of the hydro-alcoholic guaraná extract (Paullinia cupana var. sorbilis Mart.) on nitric oxide (NO) and other compounds generated from the degradation of sodium nitroprusside (SNP) in an embryonic fibroblast culture (NIH-3T3 cells) were evaluated. The guaraná bioactive compounds were initially determined by high-performance liquid chromatography: caffeine=12.240 mg/g, theobromine=6.733 mg/g and total catechins=4.336 mg/g. Cells were exposed to 10 µM SNP during a 6 h period because the cells exhibited >90% mortality at this concentration. Guaraná was added to the cultures in five concentrations (0.5, 1, 5, 10 and 20 mg/mL). The guaraná antioxidant effect was evaluated by viability assays, biochemical oxidation [lipid peroxidation, catalase and superoxide dismutase (SOD) activity] and genotoxicity (DNA Comet assay) analysis. Additionally, oxidative stress was evaluated by a 2,7-dihydrodichlorofluorescein diacetate fluorescence assay. Guaraná reverted the SNP toxicity mainly at lower concentrations (<5 mg), which decreased cell mortality, lipid peroxidation, DNA damage and cell oxidative stress as well as increased the SOD levels. These results demonstrate that guaraná has an antioxidant effect on NO metabolism in situations with higher cellular NO levels.

The dose-response effect of acute intravenous transplantation of human umbilical cord blood cells on brain damage and spatial memory deficits in neonatal hypoxia-ischemia.

Despite the beneficial effects of cell-based therapies on brain repair shown in most studies, there has not been a consensus regarding the optimal dose of human umbilical cord blood cells (HUCBC) for neonatal hypoxia-ischemia (HI). In this study, we compared the long-term effects of intravenous administration of HUCBC at three different doses on spatial memory and brain morphological changes after HI in newborn Wistar rats. In addition, we tested whether the transplanted HUCBC migrate to the injured brain after transplantation. Seven-day-old animals underwent right carotid artery occlusion and were exposed to 8% O(2) inhalation for 2 h. After 24 h, randomly selected animals were assigned to four different experimental groups: HI rats administered with vehicle (HI+vehicle), HI rats treated with 1×10(6) (HI+low-dose), 1×10(7) (HI+medium-dose), and 1×10(8) (HI+high-dose) HUCBC into the jugular vein. A control group (sham-operated) was also included in this study. After 8 weeks of transplantation, spatial memory performance was assessed using the Morris water maze (MWM), and subsequently, the animals were euthanized for brain morphological analysis using stereological methods. In addition, we performed immunofluorescence and polymerase chain reaction (PCR) analyses to identify HUCBC in the rat brain 7 days after transplantation. The MWM test showed a significant spatial memory recovery at the highest HUCBC dose compared with HI+vehicle rats (P<0.05). Furthermore, the brain atrophy was also significantly lower in the HI+medium- and high-dose groups compared with the HI+vehicle animals (P<0.01; 0.001, respectively). In addition, HUCBC were demonstrated to be localized in host brains by immunohistochemistry and PCR analyses 7 days after intravenous administration. These results revealed that HUCBC transplantation has the dose-dependent potential to promote robust tissue repair and stable cognitive improvement after HI brain injury.

Doença granulomatosa sistêmica em bovinos no estado de Santa Catarina associada ao pastoreio de ervilhaca peluda (Vicia villosa) / Systemic granulomatous disease in cattle in Santa Catarina, Brazil, associated with grazing of vetch (Vicia villosa)

Spontaneous intoxication in three dairy cows grazing pasture contaminated with Vicia villosa in two different farms was reported. Hyperthermia, skin alopecia and pruritus were the main clinical signs. Macroscopically, gray to white up to 5cm nodules were detected, especially in kidney and lymph nodes, which correspond to mild to severe multifocal granulomatous infiltrate. This is the first report of systemic granulomatous disease due to consumption of hairy vetch in the State of Santa Catarina, Brazil.

Can local supply of bone marrow mononuclear cells improve the outcome from late tubular repair of human median and ulnar nerves?

The purpose of this non-randomised retrospective study was to compare nerve regeneration after reconnection with silicone tubes with two different strategies. A total of 44 patients with injured median or ulnar nerves in the forearm were surgically treated. In one group of patients, a silicone tube alone was placed in the nerve gap. In a second group, the silicone tube was filled with autologous bone marrow mononuclear cells obtained by aspiration from the iliac crest. Motor function, sensation and the effect of pain on function were assessed 1 year after surgery. The tubes filled with bone marrow cells showed better recovery than the empty tubes. The use of bone marrow mononuclear cells in addition to tube re-connection may promote better nerve regeneration than conventional tubular repair.

TNF-alpha and IL-10 levels in tracheobronchial lavage of ventilated preterm infants and subsequent lung function.

The role of airway inflammation in ventilated preterm newborns and the risk factors associated with the development of chronic lung disease are not well understood. Our objective was to analyze the association of the airway inflammatory response in ventilated preterm infants by serial measurements of TNF-alpha and IL-10 in tracheobronchial lavage (TBL) with perinatal factors and lung function measured early in life. A series of TBL samples were collected from ventilated preterm infants (less than 32 weeks of gestational age) and concentrations of TNF-alpha and IL-10 were measured by ELISA. Pulmonary function tests were performed after discharge by the raised volume rapid compression technique. Twenty-five subjects were recruited and 70 TBL samples were obtained. There was a significant positive association between TNF-alpha and IL-10 levels and length of time between the rupture of the amniotic membranes and delivery (r = 0.65, P = 0.002, and r = 0.57, P < 0.001, respectively). Lung function was measured between 1 and 22 weeks of corrected age in 10 patients. Multivariable analysis with adjustment for differences in lung volume showed a significant negative association between TNF-alpha levels and forced expiratory flow (FEF(50); r = -0.6; P = 0.04), FEF(75) (r = -0.76; P = 0.02), FEF(85) (r = -0.75; P = 0.03), FEF(25-75) (-0.71; P = 0.02), and FEV(0.5) (r = -0.39; P = 0.03). These data suggest that TNF-alpha levels in the airways during the first days of life were associated with subsequent lung function abnormalities measured weeks or months later.

TNF-a and IL-10 levels in tracheobronchial lavage of ventilated preterm infants and subsequent lung function

The role of airway inflammation in ventilated preterm newborns and the risk factors associated with the development of chronic lung disease are not well understood. Our objective was to analyze the association of the airway inflammatory response in ventilated preterm infants by serial measurements of TNF-a and IL-10 in tracheobronchial lavage (TBL) with perinatal factors and lung function measured early in life. A series of TBL samples were collected from ventilated preterm infants (less than 32 weeks of gestational age) and concentrations of TNF-a and IL-10 were measured by ELISA. Pulmonary function tests were performed after discharge by the raised volume rapid compression technique. Twenty-five subjects were recruited and 70 TBL samples were obtained. There was a significant positive association between TNF-a and IL-10 levels and length of time between the rupture of the amniotic membranes and delivery (r = 0.65, P = 0.002, and r = 0.57, P < 0.001, respectively). Lung function was measured between 1 and 22 weeks of corrected age in 10 patients. Multivariable analysis with adjustment for differences in lung volume showed a significant negative association between TNF-a levels and forced expiratory flow (FEF50; r = -0.6; P = 0.04), FEF75 (r = -0.76; P = 0.02), FEF85 (r = -0.75; P = 0.03), FEF25-75 (-0.71; P = 0.02), and FEV0.5 (r = -0.39; P = 0.03). These data suggest that TNF-a levels in the airways during the first days of life were associated with subsequent lung function abnormalities measured weeks or months later.

Evaluation of the humoral immune response in BALB/c mice immunized with a naked DNA vaccine anti-methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is the major pathogen involved in nosocomial infections, leading to high rates of morbidity and mortality in hospitals worldwide. The methicillin resistance occurs due to the presence of an additional penicillin-binding protein, PBP2a, which has low affinity for beta-lactam antibiotics. In the past few years, vancomycin has been the only antibiotic option for treatment of infections caused by multiresistant MRSA; however, reports of vancomycin-resistant strains have generated great concerns regarding the treatment to overcome these infections. In the present study, we report preliminary results regarding the humoral immune response generated in BALB/c mice by two different doses of naked DNA vaccine containing an internal region, comprising the serine-protease domain, of the PBP2a of MRSA. The immunization procedure consisted of four immunizations given intramuscularly within 15-day intervals. Blood was collect weekly and anti-PBP2a-specific antibodies were screened by ELISA. BALB/c mice immunized with DNA vaccine anti-PBP2a have shown higher antibody titers mainly after the fourth immunization, and intriguingly, no correlation between the humoral immune response and DNA dose was observed. Our results suggest that the DNA vaccine anti-PBP2a induced an immune response by production of specific antibodies anti-MRSA in a non-dose-dependent manner, and it could represent a new and valuable approach to produce specific antibodies for passive immunization to overcome MRSA infections.

Evaluation of the humoral immune response in BALB/c mice immunized with a naked DNA vaccine anti-methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is the major pathogen involved in nosocomial infections, leading to high rates of morbidity and mortality in hospitals worldwide. The methicillin resistance occurs due to the presence of an additional penicillin-binding protein, PBP2a, which has low affinity for b-lactam antibiotics. In the past few years, vancomycin has been the only antibiotic option for treatment of infections caused by multiresistant MRSA; however, reports of vancomycin-resistant strains have generated great concerns regarding the treatment to overcome these infections. In the present study, we report preliminary results regarding the humoral immune response generated in BALB/c mice by two different doses of naked DNA vaccine containing an internal region, comprising the serine-protease domain, of the PBP2a of MRSA. The immunization procedure consisted of four immunizations given intramuscularly within 15-day intervals. Blood was collect weekly and anti-PBP2a-specific antibodies were screened by ELISA. BALB/c mice immunized with DNA vaccine anti-PBP2a have shown higher antibody titers mainly after the fourth immunization, and intriguingly, no correlation between the humoral immune response and DNA dose was observed. Our results suggest that the DNA vaccine anti-PBP2a induced an immune response by production of specific antibodies anti-MRSA in a non-dose-dependent manner, and it could represent a new and valuable approach to produce specific antibodies for passive immunization to overcome MRSA infections.