Analysis of correlation between cerebrospinal fluid and plasma HIV-1 RNA levels in patients with neurological opportunistic diseases

The question of whether HIV-1 RNA in cerebrospinal fluid (CSF) is derived from viral replication in the central nervous system or simply reflects the transit of infected lymphocytes from the blood compartment has long been a matter of debate. Some studies found no correlation between CSF and plasma viral load, whereas others did. The lack of a correlation between the two compartments suggests that the presence of HIV-1 RNA is not simply due to the passive passage of the virus from blood to CSF but rather due to intrathecal replication. To evaluate the correlation between plasma and CSF HIV-1 RNA levels and to identify situations in which there is no correlation between the two compartments, seventy patients were prospectively studied. The association between CSF and plasma viral load was evaluated in the total population and in subgroups of patients with similar characteristics. A correlation between the CSF and plasma compartments was observed for patients undergoing highly active antiretroviral therapy (HAART), those with a CD4 T lymphocyte count lower than 200 cells/mm³, and those with increased CSF protein content. On the other hand, no correlation was observed for patients without adequate virological control, who had a CD4 count higher than 200 cells/mm³ and who did not use HAART. The correlation between the two compartments observed in some patients suggests that CSF HIV-1 RNA levels may reflect plasma levels in these subjects. In contrast, the lack of a correlation between the two compartments in patients who were not on HAART and who had normal CSF proteins and a poor virological control possibly indicates compartmentalization of the virus in CSF and, consequently, plasma-independent intrathecal viral replication.

Lack of Correlation between Seminal and Plasma HIV-1 Viral Loads is Associated with CD4T Cell Depletion in Therapy-naïve HIV-1+ Patients

The present study was conducted to investigate a possible correlation between plasma (PVL) and seminal viral load (SVL) on treatment-naïve HIV-1-infected patients in Vitória, ES, Brazil. We also evaluated whether the progressive immunosuppression associated with HIV disease (as evidenced by declining CD4 T cell counts) has any impact on the correlation between PVL and SVL HIV-1. Viral load on paired blood and semen samples from 56 consecutive treatment-naïve patients were evaluated and compared to CD4 cell counts. Viral load and T cell counts (cells/µl) were determined by NASBA and by flow cytometry, respectively. Overall, a strong positive correlation between PVL and SVL (rho = 0.438, p = 0.001) was observed. However, when patients were grouped according to their CD4 counts, this correlation was only significant among patients with CD4 counts > 200 cells/µl. Results presented here demonstrate the existence of a strong correlation between PVL and SVL on patients with CD4 cell counts > 200 cells/µl, suggesting that this association may correlate with disease progression.

The role of platelet and plasma markers of antioxidant status and oxidative stress in thrombocytopenia among patients with vivax malaria

Malaria remains an important health problem in tropical countries like Brazil. Thrombocytopenia is the most common hematological disturbance seen in malarial infection. Oxidative stress (OS) has been implicated as a possible mediator of thrombocytopenia in patients with malaria. This study aimed to investigate the role of OS in the thrombocytopenia of Plasmodium vivax malaria through the measurement of oxidant and antioxidant biochemical markers in plasma and in isolated platelets. Eighty-six patients with P. vivax malaria were enrolled. Blood samples were analyzed for total antioxidant and oxidant status, albumin, total protein, uric acid, zinc, magnesium, bilirubin, total thiols, glutathione peroxidase (GPx), malondialdehyde (MDA), antibodies against mildly oxidized low-density lipoproteins (LDL-/nLDL ratio) and nitrite/nitrate levels in blood plasma and GPx and MDA in isolated platelets. Plasma MDA levels were higher in thrombocytopenic (TCP) (median 3.47; range 1.55-12.90 µmol/L) compared with the non-thrombocytopenic (NTCP) patients (median 2.57; range 1.95-8.60 µmol/L). Moreover, the LDL-/nLDL autoantibody ratio was lower in TCP (median 3.0; range 1.5-14.8) than in NTCP patients (median 4.0; range 1.9-35.5). Finally, GPx and MDA were higher in the platelets of TPC patients. These results suggest that oxidative damage of platelets might be important in the pathogenesis of thrombocytopenia found in P. vivax malaria as indicated by alterations of GPx and MDA.

CCR5 genotype and plasma ß-chemokine concentration of Brazilian HIV-infected individuals

The 32-bp deletion in the HIV-1 co-receptor CCR5 confers a high degree of resistance to HIV-1 infection in homozygous individuals for the deleted allele and partial protection against HIV-1 during disease progression in heterozygotes. Natural ligands for CCR5, MIP-1alpha, MIP-1ß and RANTES, have been shown to inhibit HIV replication in CD4+ T cells. In the present study, we examined the CCR5 genotype by PCR and the plasma levels of RANTES and MIP-1alpha by ELISA among blood donors (N = 26) and among HIV-1-infected individuals (N = 129). The control group consisted of healthy adult volunteers and HIV-1-infected subjects were an asymptomatic and heterogeneous group of individuals with regard to immunologic and virologic markers of HIV-1 disease. The frequency of the CCR5 mutant allele (delta32ccr5) in this population was 0.032; however, no delta32ccr5 homozygote was detected. These results could be related to the intense ethnic admixture of the Brazilian population. There was no correlation between circulating ß-chemokines (MIP-1alpha, RANTES) and viral load in HIV-infected individuals. RANTES concentrations in plasma samples from HIV+ patients carrying the homozygous CCR5 allele (CCR5/CCR5) (28.23 ng/ml) were higher than in the control samples (16.07 ng/ml; P<0.05); however, this HIV+ patient group (mean 26.23 pg/ml) had significantly lower concentrations of MIP-1alpha than those observed in control samples (mean 31.20 pg/ml; P<0.05). Both HIV-1-infected and uninfected individuals heterozygous for the delta32ccr5 allele had significantly lower concentrations of circulating RANTES (mean 16.07 and 6.11 ng/ml, respectively) than CCR5/CCR5 individuals (mean 28.23 and 16.07 ng/ml, respectively; P<0.05). These findings suggest that the CCR5 allele and ß-chemokine production may affect the immunopathogenesis of HIV-1.

Changes in red blood cell osmotic fragility induced by total plasma and plasma fractions obtained from rats bearing progressive and regressive variants of the Walker 256 tumor

Two variants (A and B) of the widely employed Walker 256 rat tumor cells are known. When inoculated sc, the A variant produces solid, invasive, highly metastasizing tumors that cause severe systemic effects and death. We have obtained a regressive variant (AR) whose sc growth is slower, resulting in 70-80% regression followed by development of immunity against A and AR variants. Simultaneously with the beginning of tumor regression, a temporary anemia developed (~8 days duration), accompanied by marked splenomegaly (~300%) and changes in red blood cell osmotic fragility, with mean corpuscular fragility increasing from 4.1 to 6.5 g/l NaCl. The possibility was raised that plasma factors associated with the immune response induced these changes. In the present study, we identify and compare the osmotic fragility increasing activity of plasma fractions obtained from A and AR tumor bearers at different stages of tumor development. The results showed that by day 4 compounds precipitating in 60% (NH4)2SO4 and able to increase red blood cell osmotic fragility appeared in the plasma of A and AR tumor bearers. Later, these compounds disappeared from the plasma of A tumor bearers but slightly increased in the plasma of AR tumor bearers. Furthermore, by day 10, compounds precipitating between 60 and 80% (NH4)2SO4 and with similar effects appeared only in plasma of AR tumor bearers. The salt solubility, production kinetics and hemolytic activity of these compounds resemble those of the immunoglobulins. This, together with their preferential increase in rats bearing the AR variant, suggest their association with an immune response against this tumor.

Follicle profile and plasma gonadotropin concentration in pubertal female ponies

Twelve female ponies were examined daily for 30 days and classified as ovulating (OV; N = 6; 197 ± 6 kg) or prepubertal (PP; N = 6; 196 ± 9 kg). Follicles were detected by ultrasound and gonadotropins quantified by radioimmunoassay. The mean diameter of the largest follicles was significantly larger in OV (38 ± 1 mm) than in PP (26 ± 2 mm) but there was no difference between groups in the size of the second largest follicle. There were more small follicles (<24 mm) in the PP than in the OV group, but PP fillies had a smaller number of follicles >29 mm than the OV fillies. Follicle-stimulating hormone (FSH) levels did not differ between groups but PP fillies had lower luteinizing hormone (LH) peak (8 ± 1 ng/ml) and basal (4 ± 0.5 ng/ml) levels, lower peak magnitude (2 ± 0.2 ng/ml) and period average (5 ± 0.6 ng/ml) than OV fillies (32 ± 4.5, 8 ± 1.2, 17.1 ± 6, and 15 ± 2.3 ng/ml, respectively). The PP group, in contrast to the OV group, showed no relationship between FSH surge and follicle wave emergence. We conclude that an LH concentration higher than 8 ng/ml is needed for follicle growth to a preovulatory size. Wave emergence and FSH secretion seem to be independent events, probably due to an inhibitory neural system in these PP animals. PP fillies may provide a physiological model for the study of follicle wave emergence which apparently does not depend on gonadotropin levels.

Concentration of CCL11, CXCL8 and TNF-alpha in sputum and plasma of patients undergoing asthma or chronic obstructive pulmonary disease exacerbation

Asthma and chronic obstructive pulmonary disease (COPD) are common respiratory illnesses characterized by chronic inflammation of the airways. The characterization of induced or spontaneously produced sputum is a useful technique to assess airway inflammation. In the present study, we compared the concentrations of CCL2, CCL11, CXCL8, and tumor necrosis factor-alpha (TNF-alpha) in plasma and induced sputum of patients with severe asthma or COPD and correlated the levels of these mediators with inflammatory cells in sputum. Asthmatic patients had elevated levels of eosinophils (40.1 ± 6.24%) in sputum whereas neutrophils (63.3 ± 4.66%) predominated in COPD patients. The levels of the chemokine CCL11 were markedly increased in sputum (708.7 ± 330.7 pg/ml) and plasma (716.6 ± 162.2 pg/ml) of asthmatic patients and correlated with the percentage of eosinophils in induced sputum. The concentrations of CXCL8 (817.0 ± 105.2 pg/ml) and TNF-alpha (308.8 ± 96.1 pg/ml) were higher in sputum of COPD patients and correlated with the percentage of neutrophils in induced sputum. There was also an increase in the concentrations of CXCL8 (43.2 ± 6.8 pg/ml) in sputum of asthmatic patients. These results validate that sputum is a suitable method to assess chemokines and cytokines associated with asthma and COPD. Moreover, the mechanisms involved in the synthesis of CCL11 and CXCL8/TNF-alpha would be helpful to better understand the inflammatory profile associated with asthma and COPD, respectively.

Inflammatory cytokines and plasma redox status responses in hypertensive subjects after heat exposure

Hypertension is characterized by a pro-inflammatory status, including redox imbalance and increased levels of pro-inflammatory cytokines, which may be exacerbated after heat exposure. However, the effects of heat exposure, specifically in individuals with inflammatory chronic diseases such as hypertension, are complex and not well understood. This study compared the effects of heat exposure on plasma cytokine levels and redox status parameters in 8 hypertensive (H) and 8 normotensive (N) subjects (age: 46.5±1.3 and 45.6±1.4 years old, body mass index: 25.8±0.8 and 25.6±0.6 kg/m2, mean arterial pressure: 98.0±2.8 and 86.0±2.3 mmHg, respectively). They remained at rest in a sitting position for 10 min in a thermoneutral environment (22°C) followed by 30 min in a heated environmental chamber (38°C and 60% relative humidity). Blood samples were collected before and after heat exposure. Plasma cytokine levels were measured using sandwich ELISA kits. Plasma redox status was determined by thiobarbituric acid reactive substances (TBARS) levels and ferric reducing ability of plasma (FRAP). Hypertensive subjects showed higher plasma levels of IL-10 at baseline (P<0.05), although levels of this cytokine were similar between groups after heat exposure. Moreover, after heat exposure, hypertensive individuals showed higher plasma levels of soluble TNF receptor (sTNFR1) and lower TBARS (P<0.01) and FRAP (P<0.05) levels. Controlled hypertensive subjects, who use angiotensin-converting-enzyme inhibitor (ACE inhibitors), present an anti-inflammatory status and balanced redox status. Nevertheless, exposure to a heat stress condition seems to cause an imbalance in the redox status and an unregulated inflammatory response.

Relationship between acute diarrhoea and low plasma levels of vitamin A and retinol binding protein

The objective of this study was to assess vitamin A status and association between acute diarrhoea and plasma levels of vitamin A through cross-sectional comparison in children. Plasma vitamin A was measured by colorimetric method of Neeld & Pearson and RBP by radial immunodiffusion technique. Seventy eight children (aged 18-119 months), 26 with current history of diarrhoea and 52 children as controls (outpatient from the Santa Casa de Misericórdia Hospital in metropolitan area of São Paulo City, Brazil) were studied. Children with history of diarrhoea showed significant low levels (mean ± s.e.) as compared to controls, vitamin A (15.87 ± 1.4 µg/dl vs. 21.14 ± 1.15 µg/dl, p < 0.007) and RBP (1.70 ± 0.2 mg/dl vs. 2.52 ±0.11 mg/dl). Multivariate logistic regression adjusted by sex, age, nutritional status and mother education revealed association between diarrhoea and inadequate levels of vitamin A and RBP.

Characterization of mannose-binding lectin plasma levels and genetic polymorphisms in HIV-1-infected individuals

INTRODUCTION: The present study investigated the association between mannose-binding lectin (MBL) gene polymorphism and serum levels with infection by HIV-1. METHODS: Blood samples (5mL) were collected from 97 HIV-1-infected individuals resident in Belém, State of Pará, Brazil, who attended the Special Outpatient Unit for Infections and Parasitic Diseases (URE-DIPE). CD4+ T-lymphocyte count and plasma viral load were quantified. A 349bp fragment of exon 1 of the MBL was amplified via PCR, using genomic DNA extracted from controls and HIV-1-infected individuals, following established protocols. MBL plasma levels of the patients were quantified using an enzyme immunoassay kit. RESULTS: Two alleles were observed: MBL*O, with a frequency of 26.3% in HIV-1-infected individuals; and the wild allele MBL*A (73.7%). Similar frequencies were observed in the control group (p > 0.05). Genotype frequencies were distributed according to the Hardy-Weinberg equilibrium in both groups. Mean MBL plasma levels varied by genotype, with statistically significant differences between the AA and AO (p < 0.0001), and AA and OO (p < 0.001) genotypes, but not AO and OO (p = 0.17). Additionally, CD4+ T-lymphocytes and plasma viral load levels did not differ significantly by genotype (p > 0.05). CONCLUSIONS: The results of this study do not support the hypothesis that MBL gene polymorphism or low plasma MBL concentrations might have a direct influence on HIV-1 infection, although a broader study involving a large number of patients is needed.

Correlations between ANP concentrations in atria, plasma and cerebral structures and sodium chloride preference in Wistar rats

We determined whether ANP (atrial natriuretic peptide) concentrations, measured by radioimmunoassay, in the ANPergic cerebral regions involved in regulation of sodium intake and excretion and pituitary gland correlated with differences in sodium preference among 40 Wistar male rats (180-220 g). Sodium preference was measured as mean spontaneous ingestion of 1.5% NaCl solution during a test period of 12 days. The relevant tissues included the olfactory bulb (OB), the posterior and anterior lobes of the pituitary gland (PP and AP, respectively), the median eminence (ME), the medial basal hypothalamus (MBH), and the region anteroventral to the third ventricle (AV3V). We also measured ANP content in the right (RA) and left atrium (LA) and plasma. The concentrations of ANP in the OB and the AP were correlated with sodium ingestion during the preceding 24 h, since an increase of ANP in these structures was associated with a reduced ingestion and vice-versa (OB: r = -0.3649, P<0.05; AP: r = -0.3291, P<0.05). Moreover, the AP exhibited a correlation between ANP concentration and mean NaCl intake (r = -0.4165, P<0.05), but this was not the case for the OB (r = 0.2422). This suggests that differences in sodium preference among individual male rats can be related to variations of AP ANP level. Earlier studies indicated that the OB is involved in the control of NaCl ingestion. Our data suggest that the OB ANP level may play a role mainly in day-to-day variations of sodium ingestion in the individual rat

Effect of an aqueous extract of Scoparia dulcis on blood glucose, plasma insulin and some polyol pathway enzymes in experimental rat diabetes

The effects of an aqueous extract of the plant Scoparia dulcis (200 mg/kg) on the polyol pathway and lipid peroxidation were examined in the liver of streptozotocin adult diabetic male albino Wistar rats. The diabetic control rats (N = 6) presented a significant increase in blood glucose, sorbitol dehydrogenase, glycosylated hemoglobin and lipid peroxidation markers such as thiobarbituric acid reactive substances (TBARS) and hydroperoxides, and a significant decrease in plasma insulin and antioxidant enzymes such as glutathione peroxidase (GPx), glutathione-S-transferase (GST) and reduced glutathione (GSH) compared to normal rats (N = 6). Scoparia dulcis plant extract (SPEt, 200 mg kg-1 day-1) and glibenclamide (600 µg kg-1 day-1), a reference drug, were administered by gavage for 6 weeks to diabetic rats (N = 6 for each group) and significantly reduced blood glucose, sorbitol dehydrogenase, glycosylated hemoglobin, TBARS, and hydroperoxides, and significantly increased plasma insulin, GPx, GST and GSH activities in liver. The effect of the SPEt was compared with that of glibenclamide. The effect of the extract may have been due to the decreased influx of glucose into the polyol pathway leading to increased activities of antioxidant enzymes and plasma insulin and decreased activity of sorbitol dehydrogenase. These results indicate that the SPEt was effective in attenuating hyperglycemia in rats and their susceptibility to oxygen free radicals.

Toluene permeabilization differentially affects F- and P-type ATPase activities present in the plasma membrane of Streptococcus mutans

Streptococcus mutans membrane-bound P- and F-type ATPases are responsible for H+ extrusion from the cytoplasm thus keeping intracellular pH appropriate for cell metabolism. Toluene-permeabilized bacterial cells have long been used to study total membrane-bound ATPase activity, and to compare the properties of ATPase in situ with those in membrane-rich fractions. The aim of the present research was to determine if toluene permeabilization can significantly modify the activity of membrane-bound ATPase of both F-type and P-type. ATPase activity was assayed discontinuously by measuring phosphate release from ATP as substrate. Treatment of S. mutans membrane fractions with toluene reduced total ATPase activity by approximately 80% and did not allow differentiation between F- and P-type ATPase activities by use of the standard inhibitors vanadate (3 µM) and oligomycin (4 µg/mL). Transmission electron microscopy shows that, after S. mutans cells permeabilization with toluene, bacterial cell wall and plasma membrane are severely injured, causing cytoplasmic leakage. As a consequence, loss of cell viability and disruption of H+ extrusion were observed. These data suggest that treatment of S. mutans with toluene is an efficient method for cell disruption, but care should be taken in the interpretation of ATPase activity when toluene-permeabilized cells are used, because results may not reflect the real P- and F-type ATPase activities present in intact cell membranes. The mild conditions used for the preparation of membrane fractions may be more suitable to study specific ATPase activity in the presence of biological agents, since this method preserves ATPase selectivity for standard inhibitors.

Effects of 174 G/C polymorphism in the promoter region of the interleukin-6 gene on plasma IL-6 levels and muscle strength in elderly women

We investigated the effect of -174 G/C single-nucleotide polymorphism in the promoter region of the IL6 gene on plasma IL-6 levels and muscle strength, and the relationship between IL-6 levels and muscle strength in elderly women. The sample consisted of 199 elderly residents (73.0 ± 7.8 years old) from rest homes and the community in Belo Horizonte, MG, Brazil. -174 G/C polymorphism was determined by direct sequencing of the product by PCR, and plasma IL-6 concentrations were measured by ELISA. Muscle strength in the knee joint was evaluated using a Biodex System 3 Pro® isokinetic dynamometer. ANCOVA was used to determine the effect of polymorphism on IL-6 levels and muscle strength, and the Pearson correlation coefficient to assess the relationship between IL-6 levels and muscle strength. -174 G/C polymorphism was associated with the plasma IL-6 levels of elderly women (P < 0.01) since homozygotes for the G allele showed high IL-6 levels (GG 3.85 pg/mL, GC + CC 2.13 pg/mL). There was no association of polymorphism on muscle strength (P > 0.05). No association was found between IL-6 levels and knee extensor muscle (r = 0.087, P = 0.306) or flexor (r = -0.011, P = 0.894) strength. An interaction between -174 G/C polymorphism and housing conditions of the sample of elderly women was identified, with the effect of genotype on IL-6 levels being higher in the institutionalized elderly. These results support the evidence that -174 G/C polymorphism of the IL6 gene associates with individual variability of plasma IL-6 levels in elderly women.

Plasma concentrations and placental immunostaining of interleukin-10 and tumornecrosis factor-α as predictors of alterations in the embryo-fetal organism and the placental development of diabetic rats

Interleukin-10 (IL-10) appears to be the key cytokine for the maintenance of pregnancy and inhibits the secretion of inflammatory cytokines such as tumor necrosis factor-α (TNF-α). However, there are no studies evaluating the profile of these cytokines in diabetic rat models. Thus, our aim was to analyze IL-10 and TNF-α immunostaining in placental tissue and their respective concentrations in maternal plasma during pregnancy in diabetic rats in order to determine whether these cytokines can be used as predictors of alterations in the embryo-fetal organism and in placental development. These parameters were evaluated in non-diabetic (control; N = 15) and Wistar rats with streptozotocin (STZ)-induced diabetes (N = 15). At term, the dams (100 days of life) were killed under anesthesia and plasma and placental samples were collected for IL-10 and TNF-α determinations by ELISA and immunohistochemistry, respectively. The reproductive performance was analyzed. Plasma IL-10 concentrations were reduced in STZ rats compared to controls (7.6 ± 4.5 vs 20.9 ± 8.1 pg/mL). The placental scores of immunostaining intensity did not differ between groups (P > 0.05). Prevalence analysis showed that the IL-10 expression followed TNF-α expression, showing a balance between them. STZ rats also presented impaired reproductive performance and reduced plasma IL-10 levels related to damage during early embryonic development. However, the increased placental IL-10 as a compensatory mechanism for the deficit of maternal regulation permitted embryo development. Therefore, the data suggest that IL-10 can be used as a predictor of changes in the embryo-fetal organism and in placental development in pregnant diabetic rats.