Effect of antibodies to intercellular adhesion molecule type 1 on the protection of distant organs during reperfusion syndrome in rats

We investigated kidney and lung alterations caused by intercellular adhesion molecule type 1 (ICAM-1) blockade after ischemia and reperfusion of hind limb skeletal muscles. Rats were submitted to ligature of the infrarenal aorta for 6 h. The animals were randomized into three groups of 6 rats each: group I, sacrificed after ischemia; group II, reperfusion for 24 h, and group III, reperfusion for 24 h after receiving monoclonal anti-ICAM-1 antibodies. At the end of the experiment, blood samples were collected for creatinine, lactate dehydrogenase, creatine phosphokinase, potassium, pH and leukocyte counts. Samples were taken from the muscles of the hind limbs and from the kidneys and lungs for histological analysis and measurement of the neutrophil infiltrate by myeloperoxidase staining. The groups did not differ significantly with regard to the laboratory tests. There were no major histological alterations in the kidneys. An intense neutrophil infiltrate in the lungs, similar in all groups, was detected. Myeloperoxidase determination showed that after reperfusion there was significantly less retention of polymorphonuclear neutrophils in the muscles (352 ± 70 vs 1451 ± 235 × 10² neutrophils/mg; P<0.01) and in the kidneys (526 ± 89 vs 852 ± 73 × 10² neutrophils/mg; P<0.01) of the animals that received anti-ICAM-1 before perfusion compared to the group that did not. The use of anti-ICAM-1 antibodies in this experimental model minimized neutrophil influx, thus reducing the inflammatory process, in the muscles and kidneys after ischemia and reperfusion of the hind limbs.

Clinical features of panic patients sensitive to hyperventilation or breath-holding methods for inducing panic attacks

Our aim was to compare the clinical features of panic disorder (PD) patients sensitive to hyperventilation or breath-holding methods of inducing panic attacks. Eighty-five PD patients were submitted to both a hyperventilation challenge test and a breath-holding test. They were asked to hyperventilate (30 breaths/min) for 4 min and a week later to hold their breath for as long as possible, four times with a 2-min interval. Anxiety scales were applied before and after the tests. We selected the patients who responded with a panic attack to just one of the tests, i.e., those who had a panic attack after hyperventilating (HPA, N = 24, 16 females, 8 males, mean age ± SD = 38.5 ± 12.7 years) and those who had a panic attack after breath holding (BHPA, N = 20, 11 females, 9 males, mean age ± SD = 42.1 ± 10.6 years). Both groups had similar (chi² = 1.28, d.f. = 1, P = 0.672) respiratory symptoms (fear of dying, chest/pain disconfort, shortness of breath, paresthesias, and feelings of choking) during a panic attack. The criteria of Briggs et al. [British Journal of Psychiatry, 1993; 163: 201-209] for respiratory PD subtype were fulfilled by 18 (75.0%) HPA patients and by 14 (70.0%) BHPA patients. The HPA group had a later onset of the disease compared to BHPA patients (37.9 ± 11.0 vs 21.3 ± 12.9 years old, Mann-Whitney, P < 0.001), and had a higher family prevalence of PD (70.8 vs 25.0%, chi² = 19.65, d.f. = 1, P = 0.041). Our data suggest that these two groups - HPA and BHPA patients - may be specific subtypes of PD.

Th-1 and Th-2 cytokine production in infants with virus-associated wheezing

Wheezing associated with respiratory viral infections in infancy is very common and results in high morbidity worldwide. The Th1/Th2 pattern of immune response in these patients remains unclear and previous studies have shown controversial results. The aim of the present study was to compare the type of Th1/Th2 cytokine response between infants with acute bronchiolitis, recurrent wheezing and upper respiratory infections from a developing country. Infants younger than 2 years of age admitted to Hospital São Lucas, Porto Alegre, RS, Brazil, between May and November 2001, with an acute episode of wheezing associated with viral respiratory infection were selected. Subjects with upper respiratory infections from the emergency department were selected for the control group. Interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) levels from nasal aspirates were determined by ELISA from peripheral mononuclear cell cultures. Twenty-nine subjects with acute bronchiolitis, 18 with recurrent wheezing and 15 with upper respiratory infections were enrolled. There were no differences in family history of atopy or parental smoking between groups. Oxygen requirement was similar for the acute bronchiolitis and recurrent wheezing groups. The percentage of positive tests for the cytokines studied and the IFN-gamma/IL-4 ratio was similar for all groups. Comparison of the polarized Th1/Th2 cytokine results for the various groups showed no specific pattern of cytokine production. Infants with wheezing from a developing country do not show any specific predominant pattern of Th1/Th2 cytokine production, suggesting that multiple factors may be involved in the pathogenesis of this illness.

Identification of anaerobic threshold using heart rate response during dynamic exercise

The objective of the present study was to characterize the heart rate (HR) patterns of healthy males using the autoregressive integrated moving average (ARIMA) model over a power range assumed to correspond to the anaerobic threshold (AT) during discontinuous dynamic exercise tests (DDET). Nine young (22.3 ± 1.57 years) and 9 middle-aged (MA) volunteers (43.2 ± 3.53 years) performed three DDET on a cycle ergometer. Protocol I: DDET in steps with progressive power increases of 10 W; protocol II: DDET using the same power values as protocol 1, but applied randomly; protocol III: continuous dynamic exercise protocol with ventilatory and metabolic measurements (10 W/min ramp power), for the measurement of ventilatory AT. HR was recorded and stored beat-to-beat during DDET, and analyzed using the ARIMA (protocols I and II). The DDET experiments showed that the median physical exercise workloads at which AT occurred were similar for protocols I and II, i.e., AT occurred between 75 W (116 bpm) and 85 W (116 bpm) for the young group and between 60 W (96 bpm) and 75 W (107 bpm) for group MA in protocols I and II, respectively; in two MA volunteers the ventilatory AT occurred at 90 W (108 bpm) and 95 W (111 bpm). This corresponded to the same power values of the positive trend in HR responses. The change in HR response using ARIMA models at submaximal dynamic exercise powers proved to be a promising approach for detecting AT in normal volunteers.

The inactivation of the basolateral nucleus of the rat amygdala has an anxiolytic effect in the elevated T-maze and light/dark transition tests

Pharmacological evidence indicates that the basolateral nucleus of the amygdala (BLA) is involved in the mediation of inhibitory avoidance but not of escape behavior in the elevated T-maze test. These defensive responses have been associated with generalized anxiety disorder (GAD) and panic disorder, respectively. In the present study, we determined whether the BLA plays a differential role in the control of inhibitory avoidance and escape responses in the elevated T-maze. Male Wistar rats (250-280 g, N = 9-10 in each treatment group) were pre-exposed to one of the open arms of the maze for 30 min and 24 h later tested in the model after inactivation of the BLA by a local injection of the GABA A receptor agonist muscimol (8 nmol in 0.2 µL). It has been shown that a prior forced exposure to one of the open arms of the maze, by shortening latencies to withdrawal from the open arm during the test, improves the escape task as a behavioral index of panic. The effects of muscimol in the elevated T-maze were compared to those caused by this GABA agonist in the avoidance reaction generated in the light/dark transition test. This defensive behavior has also been associated with GAD. In the elevated T-maze, intra-BLA injection of muscimol impaired inhibitory avoidance (control: 187.70 ± 14.90 s, muscimol: 37.10 ± 2.63 s), indicating an anxiolytic effect, without interfering with escape performance. The drug also showed an anxiolytic effect in the light/dark transition test as indicated by the increase in the time spent in the lighted compartment (control: 23.50 ± 2.45 s, muscimol: 47.30 ± 4.48 s). The present findings point to involvement of the BLA in the modulation of defensive responses that have been associated with GAD.

Gender-related pathways for behavior problems in the offspring of alcoholic fathers

The objective of the present study was to examine gender differences in the influence of paternal alcoholism on children's social-emotional development and to determine whether paternal alcoholism is associated with a greater number of externalizing symptoms in the male offspring. From the Mannheim Study of Risk Children, an ongoing longitudinal study of a high-risk population, the developmental data of 219 children [193 (95 boys and 98 girls) of non-alcoholic fathers, non-COAs, and 26 (14 boys, 12 girls) of alcoholic fathers, COAs] were analyzed from birth to the age of 11 years. Paternal alcoholism was defined according to the ICD-10 categories of alcohol dependence and harmful use. Socio-demographic data, cognitive development, number and severity of behavior problems, and gender-related differences in the rates of externalizing and internalizing symptoms were assessed using standardized instruments (IQ tests, Child Behavior Checklist questionnaire and diagnostic interviews). The general linear model analysis revealed a significant overall effect of paternal alcoholism on the number of child psychiatric problems (F = 21.872, d.f. = 1.217, P < 0.001). Beginning at age 2, significantly higher numbers of externalizing symptoms were observed among COAs. In female COAs, a pattern similar to that of the male COAs emerged, with the predominance of delinquent and aggressive behavior. Unlike male COAs, females showed an increase of internalizing symptoms up to age 11 years. Of these, somatic complaints revealed the strongest discriminating effect in 11-year-old females. Children of alcoholic fathers are at high risk for psychopathology. Gender-related differences seem to exist and may contribute to different phenotypes during development from early childhood to adolescence.

Health in early adulthood: the contribution of the 1978/79 Ribeirão Preto birth cohort

The increase in non-communicable chronic diseases of adults is due to demographic changes and changes in the risk factors related to physical activity, smoking habits and nutrition. We describe the methodology for the evaluation of persons at 23/25 years of age of a cohort of individuals born in Ribeirão Preto in 1978/79. We present their socioeconomic characteristics and the profile of some risk factors for chronic diseases. A total of 2063 participants were evaluated by means of blood collection, standardized questionnaires, anthropometric and blood pressure measurements, and methacholine bronchoprovocation tests. The sexes were compared by the chi-square test, with alpha = 0.05. Obesity was similar among men and women (12.8 and 11.1%); overweight was almost double in men (30.3 vs 17.7%). Weight deficit was higher among women than among men (8.6 and 2.6%). Women were more sedentary and consumed less alcohol and tobacco. Dietary fat consumption was similar between sexes, with 63% consuming large amounts (30 to 39.9 g/day). Metabolic syndrome was twice more frequent among men than women (10.7 vs 4.8%), hypertension was six times more frequent (40.9 vs 6.4%); altered triglyceride (16.1 vs 9.8%) and LDL proportions (5.4 vs 2.7%) were also higher in men, while women had a higher percentage of low HDL (44.7 vs 39.5%). Asthma and bronchial hyper-responsiveness were 1.7 and 1.5 times more frequent, respectively, among women. The high prevalence of some risk factors for chronic diseases among young adults supports the need for investments in their prevention.

Responses of IgE, IgG1, and IgG4 to concanavalin A-binding Blomia tropicalis antigens in allergic patients

Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the prevalent house dust mites in tropical countries and are associated with allergic diseases. Glycosylated antigens are highly immunogenic and involved in different pathologies. We evaluated the presence of IgE, IgG1, and IgG4 to concanavalin A-binding antigens (Bt-Con-A) isolated from Bt-total extract in sera of allergic and non-allergic subjects. Bt-total and Bt-Con-A extracts were evaluated by SDS-PAGE and ELISA for reacting with IgE, IgG1, and IgG4 in sera of 121 patients with allergic rhinitis and 36 non-allergic individuals. All subjects were skin prick tested with Bt-total extract and inhibition tests were performed for IgE, IgG1, and IgG4 using both extracts (Bt-total and Bt-Con-A). Skin prick test showed that 58% of the patients were sensitized to Bt (Bt+), with 52% reactive to both mites (Bt and Dp) and 6% to Bt only. A broad spectrum of proteins (14-152 kDa) was visualized in Bt-total and components >27 kDa for the Bt-Con-A extract. ELISA showed a similar profile of IgE, IgG1 and IgG4 levels in response to Bt-total and Bt-Con-A extracts in different groups, although Bt+ patients showed a lower IgG4 reactivity to Bt-Con-A extract. Specific IgG1 levels were higher in Bt+ patients than in control subjects, and IgG4 levels showed no significant difference among groups. ELISA inhibition showed a partial IgE and total IgG1 and IgG4 cross-reactivity with Dp extract for Bt-total and Bt-Con-A extracts. We conclude that Con-A-binding components isolated from Bt constitute major allergens and are involved in both allergen sensitization (IgE response) and homeostasis maintenance (IgG1 and IgG4 responses).

Anti-tumor properties of blackseed (Nigella sativa L.) extracts

The objective of the present study was to evaluate the in vitro and in vivo anti-cancer effect of Nigella sativa L. seed extracts. The essential oil (IC50 = 0.6%, v/v) and ethyl acetate (IC50 = 0.75%) extracts were more cytotoxic against the P815 cell line than the butanol extract (IC50 = 2%). Similar results were obtained with the Vero cell line. Although all extracts had a comparable cytotoxic effect against the ICO1 cell line, with IC50 values ranging from 0.2 to 0.26% (v/v), tests on the BSR cell line revealed a high cytotoxic effect of the ethyl acetate extract (IC50 = 0.2%) compared to the essential oil (IC50 = 1.2%). These data show that the cytotoxicity of each extract depends on the tumor cell type. In vivo, using the DBA2/P815 (H2d) mouse model, our results clearly showed that the injection of the essential oil into the tumor site significantly inhibited solid tumor development. Indeed, on the 30th day of treatment, the tumor volume of the control animals was 2.5 ± 0.6 cm³, whereas the tumor volumes of the essential oil-treated animals were 0.22 ± 0.1 and 0.16 ± 0.1 cm³ when the animals were injected with 30 µL (28.5 mg)/mouse and 50 µL (47.5 mg)/mouse per 48 h (six times), respectively. Interestingly, the administration of the essential oil into the tumor site inhibited the incidence of liver metastasis development and improved mouse survival.

Breathing pattern, thoracoabdominal motion and muscular activity during three breathing exercises

The objective of the present study was to evaluate breathing pattern, thoracoabdominal motion and muscular activity during three breathing exercises: diaphragmatic breathing (DB), flow-oriented (Triflo II) incentive spirometry and volume-oriented (Voldyne) incentive spirometry. Seventeen healthy subjects (12 females, 5 males) aged 23 ± 5 years (mean ± SD) were studied. Calibrated respiratory inductive plethysmography was used to measure the following variables during rest (baseline) and breathing exercises: tidal volume (Vt), respiratory frequency (f), rib cage contribution to Vt (RC/Vt), inspiratory duty cycle (Ti/Ttot), and phase angle (PhAng). Sternocleidomastoid muscle activity was assessed by surface electromyography. Statistical analysis was performed by ANOVA and Tukey or Friedman and Wilcoxon tests, with the level of significance set at P < 0.05. Comparisons between baseline and breathing exercise periods showed a significant increase of Vt and PhAng during all exercises, a significant decrease of f during DB and Voldyne, a significant increase of Ti/Ttot during Voldyne, and no significant difference in RC/Vt. Comparisons among exercises revealed higher f and sternocleidomastoid activity during Triflo II (P < 0.05) with respect to DB and Voldyne, without a significant difference in Vt, Ti/Ttot, PhAng, or RC/Vt. Exercises changed the breathing pattern and increased PhAng, a variable of thoracoabdominal asynchrony, compared to baseline. The only difference between DB and Voldyne was a significant increase of Ti/Ttot compared to baseline. Triflo II was associated with higher f values and electromyographic activity of the sternocleidomastoid. In conclusion, DB and Voldyne showed similar results while Triflo II showed disadvantages compared to the other breathing exercises.

Mercury toxicity in the Amazon: contrast sensitivity and color discrimination of subjects exposed to mercury

We measured visual performance in achromatic and chromatic spatial tasks of mercury-exposed subjects and compared the results with norms obtained from healthy individuals of similar age. Data were obtained for a group of 28 mercury-exposed subjects, comprising 20 Amazonian gold miners, 2 inhabitants of Amazonian riverside communities, and 6 laboratory technicians, who asked for medical care. Statistical norms were generated by testing healthy control subjects divided into three age groups. The performance of a substantial proportion of the mercury-exposed subjects was below the norms in all of these tasks. Eleven of 20 subjects (55%) performed below the norms in the achromatic contrast sensitivity task. The mercury-exposed subjects also had lower red-green contrast sensitivity deficits at all tested spatial frequencies (9/11 subjects; 81%). Three gold miners and 1 riverine (4/19 subjects, 21%) performed worse than normal subjects making more mistakes in the color arrangement test. Five of 10 subjects tested (50%), comprising 2 gold miners, 2 technicians, and 1 riverine, performed worse than normal in the color discrimination test, having areas of one or more MacAdam ellipse larger than normal subjects and high color discrimination thresholds at least in one color locus. These data indicate that psychophysical assessment can be used to quantify the degree of visual impairment of mercury-exposed subjects. They also suggest that some spatial tests such as the measurement of red-green chromatic contrast are sufficiently sensitive to detect visual dysfunction caused by mercury toxicity.

Cost-effective analysis of different algorithms for the diagnosis of hepatitis C virus infection

We compared the cost-benefit of two algorithms, recently proposed by the Centers for Disease Control and Prevention, USA, with the conventional one, the most appropriate for the diagnosis of hepatitis C virus (HCV) infection in the Brazilian population. Serum samples were obtained from 517 ELISA-positive or -inconclusive blood donors who had returned to Fundação Pró-Sangue/Hemocentro de São Paulo to confirm previous results. Algorithm A was based on signal-to-cut-off (s/co) ratio of ELISA anti-HCV samples that show s/co ratio ³95% concordance with immunoblot (IB) positivity. For algorithm B, reflex nucleic acid amplification testing by PCR was required for ELISA-positive or -inconclusive samples and IB for PCR-negative samples. For algorithm C, all positive or inconclusive ELISA samples were submitted to IB. We observed a similar rate of positive results with the three algorithms: 287, 287, and 285 for A, B, and C, respectively, and 283 were concordant with one another. Indeterminate results from algorithms A and C were elucidated by PCR (expanded algorithm) which detected two more positive samples. The estimated cost of algorithms A and B was US$21,299.39 and US$32,397.40, respectively, which were 43.5 and 14.0% more economic than C (US$37,673.79). The cost can vary according to the technique used. We conclude that both algorithms A and B are suitable for diagnosing HCV infection in the Brazilian population. Furthermore, algorithm A is the more practical and economical one since it requires supplemental tests for only 54% of the samples. Algorithm B provides early information about the presence of viremia.

Moderate-term reproducibility of heart rate variability during rest and light to moderate exercise in children

Previous studies have demonstrated the high reproducibility of heart rate variability (HRV) measures in adults while little information exists concerning HRV reproducibility in children. Subsequently, the aim of the current study was to examine the moderate-term reproducibility of heart rate and frequency domain measures of HRV during rest and light to moderate exercise in children. Ten healthy children (6 males, 4 females) aged between 7 and 12 years of age volunteered for this study with HRV recordings obtained during supine rest and three treadmill walking exercise work rates (≤60% maximum heart rate), initially and then 8 weeks later. Differences (P < 0.05) between variables were examined using paired t-tests or Wilcoxon signed rank tests while reliability and reproducibility were examined by intraclass correlation coefficients (ICC), coefficients of variation (CV), and mean bias ratio and ratio limits of agreement (LOA). Heart rate and all measures of HRV at rest and exercise were unchanged after 8 weeks. Significant ICC were documented primarily during rest (0.72-0.85) while weaker relationships (-0.02-0.87) were evident during exercise. A large range of CV was identified during rest (6-33%) and exercise (3-128%) while the ratio LOA were variable and substantial (1.04-2.73). Despite similar HRV over an 8-week period, variable ICC and sizable CV and ratio LOA indicate moderate to poor reproducibility of HRV in children, particularly during light to moderate exercise. Studies examining HRV in children should include age- or maturation stage-matched control participants to address the age-related change in HRV and inadequate HRV reliability.

Dose-response effects of systemic anandamide administration in mice sequentially submitted to the open field and elevated plus-maze tests

The endocannabinoid system is involved in the control of many physiological functions, including the control of emotional states. In rodents, previous exposure to an open field increases the anxiety-like behavior in the elevated plus-maze. Anxiolytic-like effects of pharmacological compounds that increase endocannabinoid levels have been well documented. However, these effects are more evident in animals with high anxiety levels. Several studies have described characteristic inverted U-shaped dose-response effects of drugs that modulate the endocannabinoid levels. However, there are no studies showing the effects of different doses of exogenous anandamide, an endocannabinoid, in animal models of anxiety. Thus, in the present study, we determined the dose-response effects of exogenous anandamide at doses of 0.01, 0.1, and 1.0 mg/kg in C57BL/6 mice (N = 10/group) sequentially submitted to the open field and elevated plus-maze. Anandamide was diluted in 0.9% saline, ethyl alcohol, Emulphor® (18:1:1) and administered ip (0.1 mL/10 g body weight); control animals received the same volume of anandamide vehicle. Anandamide at the dose of 0.1 mg/kg (but not of 0.01 or 1 mg/kg) increased (P < 0.05) the time spent and the distance covered in the central zone of the open field, as well as the exploration of the open arms of the elevated plus-maze. Thus, exogenous anandamide, like pharmacological compounds that increase endocannabinoid levels, promoted a characteristic inverted U-shaped dose-response effect in animal models of anxiety. Furthermore, anandamide (0.1 mg/kg) induced an anxiolytic-like effect in the elevated plus-maze (P < 0.05) after exposing the animals to the open field test.

The performance of semi-quantitative differential PCR is similar to that of real-time PCR for the detection of the MYCN gene in neuroblastomas

Amplification of the MYCN gene in neuroblastomas is a potent biological marker of highly aggressive tumors, which are invariably fatal unless sound clinical management is applied. To determine the usefulness of semi-quantitative differential PCR (SQ-PCR) for accurate quantification of MYCN gene copy number, we evaluated the analytical performance of this method by comparing the results obtained with it for 101 tumor samples of neuroblastoma to that obtained by absolute and relative real-time PCR. Similar results were obtained for 100 (99%) samples, no significant difference was detected between the median log10 MYCN copy number (1.53 by SQ-PCR versus 1.55 by absolute real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). In the comparison of SQ-PCR and relative real-time PCR, SQ-PCR versus relative real-time PCR concordant results were found in 100 (99%) samples, no significant difference was found in median log10 MYCN copy number (1.53 by SQ-PCR versus 1.27 by relative real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). These findings indicate that the performance of SQ-PCR was comparable to that of real-time PCR for the amplification and quantification of MYCN copy number. Thus, SQ-PCR can be reliably used as an alternative assay in laboratories without facilities for real-time PCR.