To evaluate the effect of blocking the angiotensin II AT-1 receptor by the systemic administration of candesartan on the expression of intercellular adhesion molecule-1 in the sclera and choroid of hypercholesterolemic rabbits.
New Zealand rabbits were divided into 3 groups, as follows: GI, which was fed a rabbit standard diet; GII, which was fed a hypercholesterolemic diet; and GIII, which received hypercholesterolemic diet plus candesartan. Samples of the rabbits' sclera and choroid were then studied by hematoxylin-eosin staining and histomorphometric and immunohistochemical analyses for intercellular adhesion molecule-1 expression.
Histological analysis of hematoxylin- and eosin-stained sclera and choroid revealed that macrophages were rarely present in GI, and GII had significantly increased macrophage numbers compared to GIII. Moreover, in GII, the sclera and choroid morphometry showed a significant increase in thickness in comparison to GI and GIII. GIII presented a significant increase in thickness in relation to GI. Sclera and choroid immunohistochemical analysis for intercellular adhesion molecule-1 expression revealed a significant increase in immunoreactivity in GII in relation to GI and GIII. GIII showed a significant increase in immunoreactivity in relation to GI.
Candesartan reduced the expression of intercellular adhesion molecule-1 and consequently macrophage accumulation in the sclera and choroid of hypercholesterolemic rabbits.
Renin-Angiotensin System; Cholesterol; Cell Adhesion Molecules; Macrophages; Choroid; Sclera; Macular Degeneration
Oxidative stress and mitochondrial dysfunction have been implicated in the pathology of HD, however the precise mechanisms by which mutant huntingtin modulates levels of oxidative damage in turn resulting in mitochondrial dysfunction are not known. We hypothesize that mutant huntingtin increases oxidative mtDNA damage leading to mitochondrial dysfunction. We measured nuclear and mitochondrial DNA lesions and mitochondrial bioenergetics in the STHdhQ7 and STHdhQ111 in vitro striatal model of HD. Striatal cells expressing mutant huntingtin show higher basal levels of mitochondrial-generated ROS and mtDNA lesions and a lower spare respiratory capacity. Silencing of APE1, the major mammalian apurinic/apyrimidinic (AP) endonuclease that participates in the base excision repair (BER) pathway, caused further reductions of spare respiratory capacity in the mutant huntingtin-expressing cells. Localization experiments show that APE1 increases in the mitochondria of wild type Q7 cells but not in the mutant huntingtin Q111 cells after treatment with hydrogen peroxide. Moreover, these results are recapitulated in human HD striata and HD skin fibroblasts that show significant mtDNA damage (increased lesion frequency and mtDNA depletion) and significant decreases in spare respiratory capacity, respectively. These data suggest that mtDNA is a major target of mutant huntingtin-associated oxidative stress and may contribute to subsequent mitochondrial dysfunction and that APE1 (and, by extension, BER) is an important target in the maintenance of mitochondrial function in HD.
mitochondrial DNA; Huntington’s disease; mitochondrial respiration base excision repair; AP endonuclease; mitochondrial dysfunction
We have previously proposed triosephosphate isomerase of Giardia lamblia (GlTIM) as a target for rational drug design against giardiasis, one of the most common parasitic infections in humans. Since the enzyme exists in the parasite and the host, selective inhibition is a major challenge because essential regions that could be considered molecular targets are highly conserved. Previous biochemical evidence showed that chemical modification of the non-conserved non-catalytic cysteine 222 (C222) inactivates specifically GlTIM. The inactivation correlates with the physicochemical properties of the modifying agent: addition of a non-polar, small chemical group at C222 reduces the enzyme activity by one half, whereas negatively charged, large chemical groups cause full inactivation.
In this work we used mutagenesis to extend our understanding of the functional and structural effects triggered by modification of C222. To this end, six GlTIM C222 mutants with side chains having diverse physicochemical characteristics were characterized. We found that the polarity, charge and volume of the side chain in the mutant amino acid differentially alter the activity, the affinity, the stability and the structure of the enzyme. The data show that mutagenesis of C222 mimics the effects of chemical modification. The crystallographic structure of C222D GlTIM shows the disruptive effects of introducing a negative charge at position 222: the mutation perturbs loop 7, a region of the enzyme whose interactions with the catalytic loop 6 are essential for TIM stability, ligand binding and catalysis. The amino acid sequence of TIM in phylogenetic diverse groups indicates that C222 and its surrounding residues are poorly conserved, supporting the proposal that this region is a good target for specific drug design.
The results demonstrate that it is possible to inhibit species-specifically a ubiquitous, structurally highly conserved enzyme by modification of a non-conserved, non-catalytic residue through long-range perturbation of essential regions.
The prevalence of spontaneous mutations increases with age in the male germline; consequently, older men have an increased risk of siring children with genetic disease due to de novo mutations. The lacI transgenic mouse can be used to study paternal age effects, and in this system, the prevalence of de novo mutations increases in the male germline at old ages. Mutagenesis is linked with DNA repair capacity, and base excision repair, which can ameliorate spontaneous DNA damage, decreases in nuclear extracts of spermatogenic cells from old mice. Mice heterozygous for a null allele of the Apex1 gene, which encodes apurinic/apyrimidinic endonuclease I (APEN), an essential base excision repair enzyme, display an accelerated increase in spontaneous germline mutagenesis early in life. Here, the consequences of lifelong reduction of APEN on genetic instability in the male germline were examined, for the first time, at middle and old ages. Mutation frequency increased earlier in spermatogenic cells from Apex1+/− mice (by 6 months of age). Nuclear DNA damage increased with age in the spermatogenic lineage for both wild-type and Apex1+/− mice. By old age, mutation frequencies were similar for wild-type and APEN-deficient mice. Mitochondrial genome repair also depends on APEN, and novel analysis of mitochondrial DNA damage revealed an increase in the Apex1+/− spermatogenic cells by middle age. Thus, Apex1 heterozygosity results in accelerated damage to mitochondrial DNA and spontaneous mutagenesis, consistent with an essential role for APEN in maintaining nuclear and mitochondrial DNA integrity in spermatogenic cells throughout life.
paternal age; mutagenesis; base excision repair; AP endonuclease; lacI; transgenic mice
Many forms of neurodegeneration are associated with oxidative stress and mitochondrial dysfunction. Mitochondria are prominent targets of oxidative damage, however, it is not clear whether mitochondrial DNA (mtDNA) damage and/or its lack of repair are primary events in the delayed onset observed in Huntington’s disease (HD). We hypothesize that an age-dependent increase in mtDNA damage contributes to mitochondrial dysfunction in HD. Two HD mouse models were studied, the 3-nitropropionic acid (3-NPA) chemically induced model and the HD transgenic mice of the R6/2 strain containing 115–150 CAG repeats in the huntingtin gene. The mitochondrial toxin 3-NPA inhibits complex II of the electron transport system and causes neurodegeneration that resembles HD in the striatum of human and experimental animals. We measured nuclear and mtDNA damage by quantitative PCR (QPCR) in striatum of 5- and 24-month-old untreated and 3-NPA treated C57BL/6 mice. Aging caused an increase in damage in both nuclear and mitochondrial genomes. 3-NPA induced 4–6 more damage in mtDNA than nuclear DNA in 5-month-old mice, and this damage was repaired by 48 h in the mtDNA. In 24-month-old mice 3NPA caused equal amounts of nuclear and mitochondrial damage and this damage persistent in both genomes for 48 h. QPCR analysis showed a progressive increase in the levels of mtDNA damage in the striatum and cerebral cortex of 7–12-week-old R6/2 mice. Striatum exhibited eight-fold more damage to the mtDNA compared with a nuclear gene. These data suggest that mtDNA damage is an early biomarker for HD-associated neurodegeneration and supports the hypothesis that mtDNA lesions may contribute to the pathogenesis observed in HD.
Mitochondrial DNA repair; Huntington’s disease; R6/2; 3-Nitropropionic acid
The Saccharomyces cerevisiae APN1 gene that participates in base excision repair has been localized both in the nucleus and the mitochondria. APN1 deficient cells (apn1Δ) show increased mutation frequencies in mitochondrial DNA (mtDNA) suggesting that APN1 is also important for mtDNA stability. To understand APN1-dependent mtDNA repair processes we studied the formation and repair of mtDNA lesions in cells exposed to methyl methanesulfonate (MMS). We show that MMS induces mtDNA damage in a dose-dependent fashion and that deletion of the APN1 gene enhances the susceptibility of mtDNA to MMS. Repair kinetic experiments demonstrate that in wild-type cells (WT) it takes 4 hr to repair the damage induced by 0.1% MMS, whereas in the apn1Δ strain there is a lag in mtDNA repair that results in significant differences in the repair capacity between the two yeast strains. Analysis of lesions in nuclear DNA (nDNA) after treatment with 0.1% MMS shows a significant difference in the amount of nDNA lesions between WT and apn1Δ cells. Interestingly, comparisons between nDNA and mtDNA damage show that nDNA is more sensitive to the effects of MMS treatment. However, both strains are able to repair the nDNA lesions, contrary to mtDNA repair, which is compromised in the apn1Δ mutant strain. Therefore, although nDNA is more sensitive than mtDNA to the effects of MMS, deletion of APN1 has a stronger phenotype in mtDNA repair than in nDNA. These results highlight the prominent role of APN1 in the repair of environmentally induced mtDNA damage.
base excision repair; mitochondrial DNA; alkylating agent
We review the potential impact of DDT on public health in Mexico. DDT production and consumption patterns in Mexico during the last 20 years are described and compared with those in the United States. In spite of the restrictions on DDT use in antimalaria campaigns in Mexico, use of DDT is still higher than in other Latin American countries. We analyzed information from published studies to determine accumulated levels of this insecticide in blood, adipose tissue, and breast milk samples from Mexican women. Current lipid-adjusted DDE levels from women living in Mexico City are 6.66 ppb in mammary adipose tissue and 0.594 ppm in total breast milk. Finally, the methodological limitations of existing epidemiological studies on DDT exposure and breast cancer are discussed. We conclude that DDT use in Mexico is a public health problem, and suggest two solutions: identification of alternatives for the control of malaria and educational intervention to reduce DDT exposure. We also recommend strengthening epidemiological studies to evaluate the association between accumulated DDT levels in adipose tissue and breast cancer incidence among Mexican women.
Several clinical trials have substantiated the efficacy of the co-administration of statins like atorvastatin (ATO) and fibrates. Without information currently available about the interaction between the two drugs, a pharmacokinetic study was conducted to investigate the effect when both drugs were co-administered. The purpose of this study was to investigate the pharmacokinetic profile of tablets containing ATO 20 mg, or the combination of ATO 20 mg with fenofibrate (FNO) 160 mg administered to healthy Mexican volunteers. This was a randomized, two-period, two-sequence, crossover study; 36 eligible subjects aged between 20–50 years were included. Blood samples were collected up to 96 h after dosing, and pharmacokinetic parameters were obtained by non-compartmental analysis. Adverse events were evaluated based on subject interviews and physical examinations. Area under the concentration-time curve (AUC) and maximum plasma drug concentration (Cmax) were measured for ATO as the reference and ATO and FNO as the test product for bioequivalence design. The estimation computed (90% confidence intervals) for ATO and FNO combination versus ATO for Cmax, AUC0-t and AUC0-∞, were 102,09, 125,95, and 120,97%, respectively. These results suggest that ATO and FNO have no relevant clinical-pharmacokinetic drug interaction.
atorvastatin; fenofibrate; pharmacokinetic interaction; combination; LC-MS-MS
Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH) and the aldehyde dehydrogenase (ALDH). We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa) of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2–C6) and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde.
bifunctional enzyme-active alcohol dehydrogenase (ADHa); ethanol-acetaldehyde-oxidation; Gluconacetobacter diazotrophicus; acetic acid bacteria; alcohol aldehyde dehydrogenase
The recognition of the limits between normal and pathological aging is essential to start preventive actions. The aim of this paper is to compare the Cambridge Neuropsychological Test Automated Battery (CANTAB) and language tests to distinguish subtle differences in cognitive performances in two different age groups, namely young adults and elderly cognitively normal subjects.
We selected 29 young adults (29.9±1.06 years) and 31 older adults (74.1±1.15 years) matched by educational level (years of schooling). All subjects underwent a general assessment and a battery of neuropsychological tests, including the Mini Mental State Examination, visuospatial learning, and memory tasks from CANTAB and language tests. Cluster and discriminant analysis were applied to all neuropsychological test results to distinguish possible subgroups inside each age group.
Significant differences in the performance of aged and young adults were detected in both language and visuospatial memory tests. Intragroup cluster and discriminant analysis revealed that CANTAB, as compared to language tests, was able to detect subtle but significant differences between the subjects.
Based on these findings, we concluded that, as compared to language tests, large-scale application of automated visuospatial tests to assess learning and memory might increase our ability to discern the limits between normal and pathological aging.
aging; neuropsychological tests; cluster analysis; discriminant analysis
Multiple Sclerosis (MS) is an immune-mediated disease of the Central Nervous System with two major underlying etiopathogenic processes: inflammation and neurodegeneration. The latter determines the prognosis of this disease. MS is the main cause of non-traumatic disability in middle-aged populations.
The MS-VisualPath Cohort was set up to study the neurodegenerative component of MS using advanced imaging techniques by focusing on analysis of the visual pathway in a middle-aged MS population in Barcelona, Spain. We started the recruitment of patients in the early phase of MS in 2010 and it remains permanently open. All patients undergo a complete neurological and ophthalmological examination including measurements of physical and disability (Expanded Disability Status Scale; Multiple Sclerosis Functional Composite and neuropsychological tests), disease activity (relapses) and visual function testing (visual acuity, color vision and visual field). The MS-VisualPath protocol also assesses the presence of anxiety and depressive symptoms (Hospital Anxiety and Depression Scale), general quality of life (SF-36) and visual quality of life (25-Item National Eye Institute Visual Function Questionnaire with the 10-Item Neuro-Ophthalmic Supplement). In addition, the imaging protocol includes both retinal (Optical Coherence Tomography and Wide-Field Fundus Imaging) and brain imaging (Magnetic Resonance Imaging). Finally, multifocal Visual Evoked Potentials are used to perform neurophysiological assessment of the visual pathway.
The analysis of the visual pathway with advance imaging and electrophysilogical tools in parallel with clinical information will provide significant and new knowledge regarding neurodegeneration in MS and provide new clinical and imaging biomarkers to help monitor disease progression in these patients.
Multiple Sclerosis; Visual pathway; Neurodegeneration; Cohort studies
Giardiasis is highly prevalent in the developing world, and treatment failures with the standard drugs are common. This work deals with the proposal of omeprazole as a novel antigiardial drug, focusing on a giardial glycolytic enzyme used to follow the cytotoxic effect at the molecular level. We used recombinant technology and enzyme inactivation to demonstrate the capacity of omeprazole to inactivate giardial triosephosphate isomerase, with no adverse effects on its human counterpart. To establish the specific target in the enzyme, we used single mutants of every cysteine residue in triosephosphate isomerase. The effect on cellular triosephosphate isomerase was evaluated by following the remnant enzyme activity on trophozoites treated with omeprazole. The interaction of omeprazole with giardial proteins was analyzed by fluorescence spectroscopy. The susceptibility to omeprazole of drug-susceptible and drug-resistant strains of Giardia lamblia was evaluated to demonstrate its potential as a novel antigiardial drug. Our results demonstrate that omeprazole inhibits giardial triosephosphate isomerase in a species-specific manner through interaction with cysteine at position 222. Omeprazole enters the cytoplasmic compartment of the trophozoites and inhibits cellular triosephosphate isomerase activity in a dose-dependent manner. Such inhibition takes place concomitantly with the cytotoxic effect caused by omeprazole on trophozoites. G. lamblia triosephosphate isomerase (GlTIM) is a cytoplasmic protein which can help analyses of how omeprazole works against the proteins of this parasite and in the effort to understand its mechanism of cytotoxicity. Our results demonstrate the mechanism of giardial triosephosphate isomerase inhibition by omeprazole and show that this drug is effective in vitro against drug-resistant and drug-susceptible strains of G. lamblia.
AIM: To determine the frequency of occult hepatitis B infection (OHBI) in a group of human immunodeficiency virus (HIV)-1+/ hepatitis B surface antigen negative (HBsAg)- patients from Mexico.
METHODS: We investigated the presence of OHBI in 49 HIV-1+/HBsAg- patients. Hepatitis B virus (HBV) DNA was analyzed using nested PCR to amplify the Core (C) region and by real-time PCR to amplify a region of the S and X genes. The possible associations between the variables and OHBI were investigated using Pearson’s χ2 and/or Fisher’s exact test.
RESULTS: We found that the frequency of OHBI was 49% among the group of 49 HIV-1+/HBsAg- patients studied. The presence of OHBI was significantly associated with the HIV-1 RNA viral load [odds ratio (OR) = 8.75; P = 0.001; 95%CI: 2.26-33.79] and with HIV-antiretroviral treatment with drugs that interfere with HBV replication (lamivudine, tenofovir or emtricitabine) (OR = 0.25; P = 0.05; 95%CI: 0.08-1.05).
CONCLUSION: The OHBI frequency is high among 49 Mexican HIV-1+/HBsAg- patients and it was more frequent in patients with detectable HIV RNA, and less frequent in patients who are undergoing HIV-ARV treatment with drugs active against HBV.
Hepatitis B virus; Occult hepatitis B virus infection; Human immunodeficiency virus; Hepatitis B surface antigen negative; Risk factors; Molecular diagnostics
Reactive and pharmaceutical-grade heparins were used as biologically compatible reducing and stabilizing agents to photochemically synthesize colloidal gold nanoparticles. Aggregates and anisotropic shapes were obtained photochemically under UV black-light lamp irradiation (λ = 366 nm). Heparin-functionalized gold nanoparticles were characterized by Scanning Electron Microscopy and UV-Vis spectroscopy. The negatively charged colloids were used for the Surface Enhanced Raman Spectroscopy (SERS) analysis of differently charged analytes (dyes). Measurements of pH were taken to inspect how the acidity of the medium affects the colloid-analyte interaction. SERS spectra were taken by mixing the dyes and the colloidal solutions without further functionalization or addition of any aggregating agent.
heparin; gold nanoparticles; photochemistry; SERS; organic dyes
Nephrotic syndrome is associated with a hypercoagulable state and an increased risk
of thromboembolic complications. Cerebral venous sinus thrombosis is a rare
complication of nephrotic syndrome, with few cases described in the literature,
although the disease may be under-diagnosis. The true incidence of cerebral venous
sinus thrombosis may be underestimated because many events are asymptomatic or are
not diagnosed in time. Here, we describe the case of a male child, 2 years and 10
months old, with nephrotic syndrome presenting with headache, epileptic seizures and
sensory inhibition who was diagnosed with superior sagittal and transverse sinuses
thrombosis. An international literature review was performed with a defined search
strategy in the PubMed, SciELO and Lilacs databases using the terms ‘nephrotic
syndrome’ and ‘cerebral sinovenous thrombosis’. The diagnosis of venous thrombosis
should be considered in any patient with nephrotic syndrome who presents with
neurological signs and symptoms, as early clinical diagnosis promotes favorable
Sinus thrombosis, intracranial/etiology; Venous thrombosis; Nephrotic syndrome/complications; Child; Case reports
The amount of epicardial adipose tissue (EAT) around the heart has been identified as an independent predictor of coronary artery disease (CAD), potentially through local release of inflammatory cytokines. Ethnic differences have been observed, but no studies have investigated this relationship in the Mexican population. The objective of the present study was to evaluate whether a relationship exist between EAT thickness assessed via echocardiography with CAD and adiponectin levels in a Mexican population.
We studied 153 consecutive patients who underwent coronary angiography and transthoracic echocardiography (TTE). EAT thickness on the free wall of the right ventricle was measured at the end of systole from parasternal long and short axis views of three consecutive cardiac cycles. Coronary angiograms were analyzed for the presence, extent and severity of CAD. Serum adiponectin, lipids, glucose, C-reactive protein and fibrinogen were determined.
EAT thickness was greater in patients with CAD than in those without CAD from both parasternal long (5.39 ± 1.75 mm vs 4.00 ± 1.67 mm p < 0.0001) and short-axis views (5.23 ± 1.67 vs 4.12 ± 1.77, p = 0.001). EAT thickness measured from parasternal long and short-axis showed a statistically significant positive correlation with age (r = 0.354, p < 0.001; r = 0.286, p < 0.001 respectively), and waist circumference (r = 0.189, p = 0.019; r = 0.217, p = 0.007 respectively). A significant negative correlation between EAT thickness from the parasternal long axis with cholesterol-HDL was observed (r = -0.163, p = 0.045). No significant correlation was found between epicardial fat thickness and serum adiponectin or with the severity of CAD.
EAT thickness was greater in patients with CAD. However, no correlation was observed with the severity of the disease or with serum adiponectin levels. EAT thickness measured by echocardiography might provide additional information for risk assessment and prediction of CAD.
Epicardial adipose tissue; Coronary artery disease; Echocardiography; Adiponectin
During the 2013–14 influenza season, we assessed characteristics of 102 adults with suspected influenza pneumonia in a hospital in Mexico; most were unvaccinated. More comorbidities and severity of illness were found than for patients admitted during the 2009–10 influenza pandemic. Vaccination policies should focus on risk factors.
Influenza; A(H1N1)pdm09; viruses; vaccine; pandemic; pneumonia; viral pneumonia; diabetes; intensive care; obesity; overweight; body mass index; BMI; ventilator; zoonoses
The beneficial effects of the Mediterranean diet have been amply proven in adults with cardiovascular risk factors. The effects of this diet have not been extensively assessed in pediatric populations with obesity, insulin resistance (IR) and metabolic syndrome (MetS). The aim of this study was to assess the efficacy of the Mediterranean style diet (MSD) to decrease cardiovascular risk factors in children and adolescents with obesity.
Participants were randomly assigned to a MSD rich in polyunsaturated fatty acids, fiber, flavonoids and antioxidants (60% of energy from carbohydrate, 25% from fat, and 15% from protein, (n = 24); or a standard diet (55% of carbohydrate, 30% from fat and 15% from protein, (n = 25), the caloric ingest was individualized. At baseline and 16-week of intervention, the glucose, triglycerides (TG), total cholesterol (TC), HDL-C, LDL-C were measured as well as the body composition and anthropometric data. The diet compliance was determined by the 24-hour recalls.
Paired Student’s t and Macnemar’s test were used to compare effects in biochemical, body composition, anthropometric, and dietary variables.
The MSD group had a significantly decrease in BMI, lean mass, fat mass, glucose, TC, TG, HDL-C and LDL-C. (p < 0.05); the diet compliance increased consumption of omega 9 fatty acids, zinc, vitamin E, selenium, and decreased consumption of saturated fatty acids (p < 0.05). The standard diet group decrease in glucose levels and frequency of glucose >100 mg/dL (p < 0.05).
The MSD improves the BMI, glucose and lipid profile in children and adolescents with obesity and any MetS component.
Obesity; Metabolic syndrome; Mediterranean diet; Children; Adolescents
We have previously demonstrated that a DNA vaccine encoding HIV-p55gag in association with the lysosomal associated membrane protein-1 (LAMP-1) elicited a greater Gag-specific immune response, in comparison to a DNA encoding the native gag. In vitro studies have also demonstrated that LAMP/Gag was highly expressed and was present in MHCII containing compartments in transfected cells. In this study, the mechanisms involved in these processes and the relative contributions of the increased expression and altered traffic for the enhanced immune response were addressed. Cells transfected with plasmid DNA constructs containing p55gag attached to truncated sequences of LAMP-1 showed that the increased expression of gag mRNA required p55gag in frame with at least 741 bp of the LAMP-1 luminal domain. LAMP luminal domain also showed to be essential for Gag traffic through lysosomes and, in this case, the whole sequence was required. Further analysis of the trafficking pathway of the intact LAMP/Gag chimera demonstrated that it was secreted, at least in part, associated with exosome-like vesicles. Immunization of mice with LAMP/gag chimeric plasmids demonstrated that high expression level alone can induce a substantial transient antibody response, but targeting of the antigen to the endolysosomal/secretory pathways was required for establishment of cellular and memory response. The intact LAMP/gag construct induced polyfunctional CD4+ T cell response, which presence at the time of immunization was required for CD8+ T cell priming. LAMP-mediated targeting to endolysosomal/secretory pathway is an important new mechanistic element in LAMP-mediated enhanced immunity with applications to the development of novel anti-HIV vaccines and to general vaccinology field.
In 2010, and due to a quality problem identified in the vaccine manufacture, the rotavirus (RV) vaccination was withheld in Spain during 5 months. Our study aimed to evaluate the impact that this sudden cease had on rotavirus acute gastroenteritis (RAGE) hospitalizations. An increase in RAGE hospitalization was observed in parallel to the drop in vaccine coverage. Here, we report the first reverse evidence of rotavirus vaccine impact.
gastroenteritis/epidemiology; hospitalization; rotavirus infections/prevention & control; rotavirus vaccines; vaccine effectiveness
Deletion of Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) envelope (E) gene attenuates the virus. E gene encodes a small multifunctional protein that possesses ion channel (IC) activity, an important function in virus-host interaction. To test the contribution of E protein IC activity in virus pathogenesis, two recombinant mouse-adapted SARS-CoVs, each containing one single amino acid mutation that suppressed ion conductivity, were engineered. After serial infections, mutant viruses, in general, incorporated compensatory mutations within E gene that rendered active ion channels. Furthermore, IC activity conferred better fitness in competition assays, suggesting that ion conductivity represents an advantage for the virus. Interestingly, mice infected with viruses displaying E protein IC activity, either with the wild-type E protein sequence or with the revertants that restored ion transport, rapidly lost weight and died. In contrast, mice infected with mutants lacking IC activity, which did not incorporate mutations within E gene during the experiment, recovered from disease and most survived. Knocking down E protein IC activity did not significantly affect virus growth in infected mice but decreased edema accumulation, the major determinant of acute respiratory distress syndrome (ARDS) leading to death. Reduced edema correlated with lung epithelia integrity and proper localization of Na+/K+ ATPase, which participates in edema resolution. Levels of inflammasome-activated IL-1β were reduced in the lung airways of the animals infected with viruses lacking E protein IC activity, indicating that E protein IC function is required for inflammasome activation. Reduction of IL-1β was accompanied by diminished amounts of TNF and IL-6 in the absence of E protein ion conductivity. All these key cytokines promote the progression of lung damage and ARDS pathology. In conclusion, E protein IC activity represents a new determinant for SARS-CoV virulence.
Several highly pathogenic viruses encode small transmembrane proteins with ion-conduction properties named viroporins. Viroporins are generally involved in virus production and maturation processes, which many times are achieved by altering the ion homeostasis of cell organelles. Cells have evolved mechanisms to sense these imbalances in ion concentrations as a danger signal, and consequently trigger the innate immune system. Recently, it has been demonstrated that viroporins are inducers of cytosolic macromolecular complexes named inflammasomes that trigger the activation of key inflammatory cytokines such as IL-1β. The repercussions of this system in viral pathogenesis or disease outcome are currently being explored. SARS-CoV infection induces an uncontrolled inflammatory response leading to pulmonary damage, edema accumulation, severe hypoxemia and eventually death. In this study, we report that SARS-CoV E protein ion channel activity is a determinant of virulence, as the elimination of this function attenuated the virus, reducing the harmful inflammatory cytokine burst produced after infection, in which inflammasome activation plays a critical role. This led to less pulmonary damage and to disease resolution. These novel findings may be of relevance for other viral infections and can possibly be translated in order to find therapies for their associated diseases.
To determine the effects of acute and chronic administration of
methylprednisolone on oxidative stress, as quantified by measuring lipid
peroxidation (LPO) and total reactive antioxidant potential (TRAP), in rat lungs.
Forty Wistar rats were divided into four groups: acute treatment, comprising rats
receiving a single injection of methylprednisolone (50 mg/kg i.p.); acute control,
comprising rats i.p. injected with saline; chronic treatment, comprising rats
receiving methylprednisolone in drinking water (6 mg/kg per day for 30 days); and
chronic control, comprising rats receiving normal drinking water.
The levels of TRAP were significantly higher in the acute treatment group rats
than in the acute control rats, suggesting an improvement in the pulmonary
defenses of the former. The levels of lung LPO were significantly higher in the
chronic treatment group rats than in the chronic control rats, indicating
oxidative damage in the lung tissue of the former.
Our results suggest that the acute use of corticosteroids is beneficial to lung
tissue, whereas their chronic use is not. The chronic use of methylprednisolone
appears to increase lung LPO levels.
Lung; Methylprednisolone; Glucocorticoids; Lipid peroxidation; Antioxidant response elements
Carbon dioxide pneumoperitoneum used during laparoscopic surgeries alters the integrity of the peritoneum and results in denudation of the basal lamina that might cause altered immune response, inhibited fibrinolysis, hypoxia, and acidosis. The changes in the structure of pneumoperitoneum were described as bulging of mesothelial cells, irregular cell junction’s cell membrane degradation, and mesodermal edema. As denaturation of peritoneal proteins reflects overall condition of its structure and interactions with the surrounding molecules, the physical status of collagen was assessed on the basis of parameters of thermal denaturation measured by DSC method.
Twenty-four female patients operated on due to cholelithiasis were enrolled in this study. Laparoscopic cholecystectomy was performed using standard four-trocar technique, and standard values of insufflated carbon dioxide pneumoperitoneum were used. After trocar placement, the first collection of peritoneal sample (sample A) was performed. The second peritoneal sample (sample B) was collected after the removal of gall bladder. Differential scanning calorimetry (Q200 calorimeter, TA Instruments) was performed on samples defrosted at room temperature.
In all samples of peritoneum, a nonreversible endothermal process recognized as denaturation was observed. Sample B obtained at the end of surgery did not differ from sample A obtained at the beginning in terms of all parameters under study. Temperature of denaturation in A and B was correlated only marginally, but enthalpy and specific heat were significantly correlated. The analysis of data from DSC measurements did not reveal differences in physical stability of collagen in peritoneal samples obtained at the beginning and at the end of surgery. Significant negative correlations between duration of CO2 pneumoperitoneum and enthalpy of denaturation in sample B were found.
Differences in enthalpy of denaturation may reflect a quantitative relation between amount of native collagen molecules in the sample and other, non-collagenous components or impaired collagen.
Laparoscopy; Peritoneum; Calorimetry; Surgery
The correlations of genotypic and phenotypic tests with treatment, clinical history and the significance of mutations in viruses of HIV-infected patients are used to establish resistance mutations to protease inhibitors (PIs). Emerging mutations in human immunodeficiency virus type 1 (HIV-1) protease confer resistance to PIs by inducing structural changes at the ligand interaction site. The aim of this study was to establish an in silico structural relationship between natural HIV-1 polymorphisms and unusual HIV-1 mutations that confer resistance to PIs.
Protease sequences isolated from 151 Mexican HIV-1 patients that were naïve to, or subjected to antiretroviral therapy, were examined. We identified 41 unrelated resistance mutations with a prevalence greater than 1%. Among these mutations, nine exhibited positive selection, three were natural polymorphisms (L63S/V/H) in a codon associated with drug resistance, and six were unusual mutations (L5F, D29V, L63R/G, P79L and T91V). The D29V mutation, with a prevalence of 1.32% in the studied population, was only found in patients treated with antiretroviral drugs. Using in silico modelling, we observed that D29V formed unstable protease complexes when were docked with lopinavir, saquinavir, darunavir, tipranavir, indinavir and atazanavir.
The structural correlation of natural polymorphisms and unusual mutations with drug resistance is useful for the identification of HIV-1 variants with potential resistance to PIs. The D29V mutation likely confers a selection advantage in viruses; however, in silico, presence of this mutation results in unstable enzyme/PI complexes, that possibly induce resistance to PIs.
Antiretroviral agents; Bioinformatics; Molecular docking simulation; Drug resistance; HIV protease; In silico; Polymorphism; Mutations
Tumor expression of estrogen receptor (ER) is an important marker of prognosis, and is predictive of response to endocrine therapy in breast cancer. Several studies have observed that epigenetic events, such methylation of cytosines and deacetylation of histones, are involved in the complex mechanisms that regulate promoter transcription. However, the exact interplay of these factors in transcription activity is not well understood. In this study, we explored the relationship between ER expression status in tumor tissue samples and the methylation of the 5′ CpG promoter region of the estrogen receptor gene (ESR1) isolated from free circulating DNA (fcDNA) in plasma samples from breast cancer patients.
Patients (n = 110) with non-metastatic breast cancer had analyses performed of ER expression (luminal phenotype in tumor tissue, by immunohistochemistry method), and the ESR1-DNA methylation status (fcDNA in plasma, by quantitative methylation specific PCR technique).
Our results showed a significant association between presence of methylated ESR1 in patients with breast cancer and ER negative status in the tumor tissue (p = 0.0179). There was a trend towards a higher probability of ESR1-methylation in those phenotypes with poor prognosis i.e. 80% of triple negative patients, 60% of HER2 patients, compared to 28% and 5.9% of patients with better prognosis such as luminal A and luminal B, respectively.
Silencing, by methylation, of the promoter region of the ESR1 affects the expression of the estrogen receptor protein in tumors of breast cancer patients; high methylation of ESR1-DNA is associated with estrogen receptor negative status which, in turn, may be implicated in the patient’s resistance to hormonal treatment in breast cancer. As such, epigenetic markers in plasma may be of interest as new targets for anticancer therapy, especially with respect to endocrine treatment.
Breast cancer; Methylation; Luminal phenotypes