Lung inflammation is induced by renal ischemia and reperfusion injury as part of the systemic inflammatory syndrome

Ischemia and reperfusion injury (IRI) are mainly caused by leukocyte activation, endothelial dysfunction and production of reactive oxygen species. Moreover, IRI can lead to a systemic response affecting distant organs, such as the lungs. The objective was to study the pulmonary inflammatory systemic response after renal IRI. Male C57Bl/6 mice were subjected to 45 min of bilateral renal ischemia, followed by 4, 6, 12, 24 and 48 h of reperfusion. Blood was collected to measure serum creatinine and cytokine concentrations. Bronchoalveolar lavage fluid (BALF) was collected to determine the number of cells and PGE(2) concentration. Expressions of iNOS and COX-2 in lung were determined by Western blot. Gene analyses were quantified by real time PCR. Serum creatinine increased in the IRI group compared to sham mainly at 24 h after IRI (2.57 +/- A 0.16 vs. 0.43 +/- A 0.07, p < 0.01). The total number of cells in BAL fluid was higher in the IRI group in comparison with sham, 12 h (100 x 10(4) +/- A 15.63 vs. 18.1x10(4) +/- A 10.5, p < 0.05) 24 h (124 x 10(4) +/- A 8.94 vs. 23.2x10(4) +/- A 3.5, p < 0.05) and 48 h (79 x 10(4) +/- A 15.72 vs. 22.2 x 10(4) +/- A 4.2, p < 0.05), mainly by mononuclear cells and neutrophils. Pulmonary COX-2 and iNOS were up-regulated in the IRI group. TNF-alpha, IL-1 beta, MCP-1, KC and IL-6 mRNA expression were up-regulated in kidney and lungs 24 h after renal IRI. ICAM-1 mRNA was up-regulated in lungs 24 h after renal IRI. Serum TNF-alpha, IL-1 beta and MCP-1 and BALF PGE(2) concentrations were increased 24 h after renal IRI. Renal IRI induces an increase of cellular infiltration, up-regulation of COX-2, iNOS and ICAM-1, enhanced chemokine expression and a Th1 cytokine profile in lung demonstrating that the inflammatory response is indeed systemic, possibly leading to an amplification of renal injury.

THE ROLE OF INNATE IMMUNITY IN SEPTIC ACUTE KIDNEY INJURIES

Acute kidney injury (AKI) is an important clinical syndrome characterized by abnormalities in the hydroelectrolytic balance. Because of high rates of morbidity and mortality (from 15% to 60%) associated with AKI, the study of its pathophysiology is critical in searching for clinical targets and therapeutic strategies. Severe sepsis is the major cause of AKI. The host response to sepsis involves an inflammatory response, whereby the pathogen is initially sensed by innate immune receptors (pattern recognition receptors [PRRs]). When it persists, this immune response leads to secretion of proinflammatory products that induce organ dysfunction such as renal failure and consequently increased mortality. Moreover, the injured tissue releases molecules resulting from extracellular matrix degradation or dying cells that function as alarmines, which are recognized by PRR in the absence of pathogens in a second wave of injury. Toll-like receptors (TLRs) and NOD-like receptors (NLRs) are the best characterized PRRs. They are expressed in many cell types and throughout the nephron. Their activation leads to translocation of nuclear factors and synthesis of proinflammatory cytokines and chemokines. TLRs` signaling primes the cells for a robust inflammatory response dependent on NLRs; the interaction of TLRs and NLRs gives rise to the multiprotein complex known as the inflammasome, which in turn activates secretion of mature interleukin 1 beta and interleukin 18. Experimental data show that innate immune receptors, the inflammasome components, and proinflammatory cytokines play crucial roles not only in sepsis, but also in organ-induced dysfunction, especially in the kidneys. In this review, we discuss the significance of the innate immune receptors in the development of acute renal injury secondary to sepsis.

p38 alpha MAP Kinase Controls IL-17 Synthesis in Vogt-Koyanagi-Harada Syndrome and Experimental Autoimmune Uveitis

PURPOSE. Interleukin (IL)-17, which is responsible for the initial influx of leukocytes into the target tissue, was recently described as the main cytokine involved in autoimmune diseases. Vogt-Koyanagi-Harada (VKH) syndrome is a significant cause of noninfectious blindness in the world. Herein the authors aimed at unraveling the involvement of IL-17 in VKH and in experimental autoimmune uveitis, focusing on the signaling pathways involved in IL-17 synthesis. METHODS. Mice were immunized with 161-180 peptide and pertussis toxin. Draining lymph node cells, harvested 21 days after immunization, were cultured in the presence or absence of p38 alpha mitogen-activated protein kinase (MAPK) inhibitor (SB203580) and assayed for cytokine production and quantification of CD4(+)IL-17(+) cells. Mice received intraocular injections of SB203580, and disease severity was evaluated by histologic examination of the enucleated eyes at day 21. CD4(+) lymphocytes from MSK-1/2-deficient mice, human CD4(+) cells silenced with MSK1 siRNA, or peripheral blood mononuclear cells (PBMCs) from VKH patients were cultured in the presence or absence of p38 alpha MAPK inhibitor and then assayed for IL-17, IFN-gamma, and IL-4 production. RESULTS. The inhibition of p38 alpha MAPK fully blocked the synthesis of IL-17 by PBMCs from VKH patients and lymphocytes from EAU mice. The absence of the msk1/2 gene resulted in failure to produce IL-17 by murine and human lymphocytes. Interestingly, intraocular injections of SB203580 in EAU mice did not suppress development of the disease. CONCLUSIONS. These data show that p38 alpha MAPK-MSK1/2 is involved in the control of IL-17 synthesis by CD4(+) T cells and that inhibition of p38 alpha MAPK in vitro suppresses IL-17 synthesis but that inhibition of this kinase in vivo did not protect from EAU. (Invest Ophthalmol Vis Sci. 2010;51:3567-3574) DOI: 10.1167/iovs.09-4393

Early Phase of Allergic Airway Inflammation in Diabetic Rats: Role of Insulin on the Signaling Pathways and Mediators

Background: Allergic lung inflammation is impaired in diabetic rats and is restored by insulin treatment. In the present study we investigated the effect of insulin on the signaling pathways triggered by allergic inflammation in the lung and the release of selected mediators. Methods: Diabetic male Wistar rats (alloxan, 42 mg/kg, i.v., 10 days) and matching controls were sensitized by s.c. injections of ovalbumin (OA) in aluminium hydroxide, 14 days before OA (1 mg/0.4 ml) or saline intratracheal challenge. A group of diabetic rats were treated with neutral protamine Hagedorn insulin (NPH, 4 IU, s.c.), 2 h before the OA challenge. Six hours after the challenge, bronchoalveolar lavage (BAL) was performed for mediator release and lung tissue was homogenized for Western blotting analysis of signaling pathways. Results: Relative to non-diabetic rats, the diabetic rats exhibited a significant reduction in OA-induced phosphorylation of the extracellular signal-regulated kinase (ERK, 59%), p38 (53%), protein kinase B (Akt, 46%), protein kinase C (PKC)-alpha (63%) and PKC-delta (38%) in lung homogenates following the antigen challenge. Activation of the NF-kappa B p65 subunit and phosphorylation of I kappa B alpha were almost suppressed in diabetic rats. Reduced expression of inducible nitric oxide synthase (iNOS, 32%) and cyclooxygenase-2 (COX-2, 46%) in the lung homogenates was also observed. The BAL concentration of prostaglandin (PG)-E(2), nitric oxide (NO) and interleukin (IL)-6 was reduced in diabetic rats (74%, 44% and 65%, respectively), whereas the cytokine-induced neutrophil chemoattractant (CINC)-2 concentration was not different from the control animals. Treatment of diabetic rats with insulin completely or partially restored all of these parameters. This protocol of insulin treatment only partially reduced the blood glucose levels. Conclusion: The data presented show that insulin regulates MAPK, PI3K, PKC and NF-kappa B pathways, the expression of the inducible enzymes iNOS and COX-2, and the levels of NO, PGE(2) and IL-6 in the early phase of allergic lung inflammation in diabetic rats. It is suggested that insulin is required for optimal transduction of the intracellular signals that follow allergic stimulation. Copyright (C) 2010 S. Karger AG, Basel

Enhanced Neural Progenitor/Stem Cells Self-Renewal via the Interaction of Stress-Inducible Protein 1 with the Prion Protein

Prion protein (PrPC), when associated with the secreted form of the stress-inducible protein 1 (STI1), plays an important role in neural survival, neuritogenesis, and memory formation. However, the role of the PrP(C)-STI1 complex in the physiology of neural progenitor/stem cells is unknown. In this article, we observed that neurospheres cultured from fetal forebrain of wild-type (Prnp(+/+)) and PrP(C)-null (Prnp(0/0)) mice were maintained for several passages without the loss of self-renewal or multipotentiality, as assessed by their continued capacity to generate neurons, astrocytes, and oligodendrocytes. The homogeneous expression and colocalization of STI1 and PrP(C) suggest that they may associate and function as a complex in neurosphere-derived stem cells. The formation of neurospheres from Prnp(0/0) mice was reduced significantly when compared with their wild-type counterparts. In addition, blockade of secreted STI1, and its cell surface ligand, PrP(C), with specific antibodies, impaired Prnp(+/+) neurosphere formation without further impairing the formation of Prnp(0/0) neurospheres. Alternatively, neurosphere formation was enhanced by recombinant STI1 application in cells expressing PrP(C) but not in cells from Prnp(0/0) mice. The STI1-PrP(C) interaction was able to stimulate cell proliferation in the neurosphere-forming assay, while no effect on cell survival or the expression of neural markers was observed. These data suggest that the STI1-PrP(C) complex may play a critical role in neural progenitor/stem cells self-renewal via the modulation of cell proliferation, leading to the control of the stemness capacity of these cells during nervous system development. STEM CELLS 2011;29:1126-1136

INSULIN REGULATES CYTOKINES AND INTERCELLULAR ADHESION MOLECULE-1 GENE EXPRESSION THROUGH NUCLEAR FACTOR-kappa B ACTIVATION IN LPS-INDUCED ACUTE LUNG INJURY IN RATS

Diabetic patients have increased susceptibility to infection, which may be related to impaired inflammatory response observed in experimental models of diabetes, and restored by insulin treatment. The goal of this study was to investigate whether insulin regulates transcription of cytokines and intercellular adhesion molecule 1 (ICAM-1) via nuclear factor-kappa B (NF-kappa B) signaling pathway in Escherichia coli LIPS-induced lung inflammation. Diabetic male Wistar rats (alloxan, 42 mg/kg, iv., 10 days) and controls were instilled intratracheally with saline containing LPS (750 mu g/0.4 mL) or saline only. Some diabetic rats were given neutral protamine Hagedorn insulin (4 IU, s.c.) 2 h before LIPS. Analyses performed 6 h after LPS included: (a) lung and mesenteric lymph node IL-1 beta, TNF-alpha, IL-10, and ICAM-1 messenger RNA (mRNA) were quantified by real-time reverse transcriptase-polymerase chain reaction; (b) number of neutrophils in the bronchoalveolar lavage (BAL) fluid, and concentrations of IL-1 beta, TNF-alpha, and IL-10 in the BAL were determined by the enzyme-linked immunosorbent assay; and (c) activation of NF-kappa B p65 subunit and phosphorylation of I-kappa B alpha were quantified by Western blot analysis. Relative to controls, diabetic rats exhibited a reduction in lung and mesenteric lymph node IL-1 beta (40%), TNF-alpha (similar to 30%), and IL-10 (similar to 40%) mRNA levels and reduced concentrations of IL-1 beta (52%), TNF-alpha (62%), IL-10 (43%), and neutrophil counts (72%) in the BAL. Activation of NF-kappa B p65 subunit and phosphorylation of I-kappa B alpha were almost suppressed in diabetic rats. Treatment of diabetic rats with insulin completely restored mRNA and protein levels of these cytokines and potentiated lung ICAM-1 mRNA levels (30%) and number of neutrophils (72%) in the BAL. Activation of NF-kappa B p65 subunit and phosphorylation of I-kappa B alpha were partially restored by insulin treatment. In conclusion, data presented suggest that insulin regulates transcription of proinflammatory (IL-1 beta, TNF-alpha) and anti-inflammatory (IL-10) cytokines, and expression of ICAM-1 via the NF-kappa B signaling pathway.

Asthma management and asthma control in Sao Paulo, Brazil and Uppsala, Sweden: a questionnaire-based comparison

The Global Initiative Against Asthma (GINA) was developed to meet the global challenge of asthma. GINA has been adopted in most countries and comparison of asthma management in different parts of the world may be of help when assessing the global dissemination of the guideline. The overall goals in GINA include that asthma patients should be free of symptoms, acute asthma attacks and activity limitations. The aim of the present study was to compare asthma management and asthma control in Sao Paulo, Brazil and Uppsala, Sweden. Information was collected from asthmatics in Sao Paulo and Uppsala with a questionnaire. The questionnaire dealt with the following issues: symptoms, smoking, self-management, hospital visits, effect on school/work and medication. The Sao Paulo patients were more likely to have uncontrolled asthma (36% vs 13%, P < 0.001), having made emergency room visits (57% vs 29%, P < 0.001) and having lost days at school or work because of their asthma (46% vs 28%, P = 0.03) than the asthmatics from Uppsala. There were no difference in the use of inhaled corticosteroids, but the Brazilian patients were more likely to be using theophylline (18% vs 1%, P = 0.001) and less likely to be using long-acting beta-2 agonists (18% vs 37%, P < 0.001). We conclude that the level of asthma control was lower among the patients from Sao Paulo than Uppsala. Few of the patients in either city reached the goals set up by GINA. Improved asthma management may therefore lead to health-economic benefits in both locations. Please cite this paper as: Skorup P, Rizzo LV, Machado-Boman L and Janson C. Asthma management and asthma control in Sao Paulo, Brazil and Uppsala, Sweden: a questionnaire-based comparison. The Clinical Respiratory Journal 2009; 3: 22-28.

PAS-1, a protein from Ascaris suum, modulates allergic inflammation via IL-10 and IFN-gamma, but not IL-12

Helminths and their products have a profound immunomodulatory effect upon the inductive and effector phases of inflammatory responses, including allergy. We have demonstrated that PAS-1, a protein isolated from Ascaris strum worms, has an inhibitory effect on lung allergic inflammation due to its ability to down-regulate eosinophilic inflammation, Th2 cytokine release and IgE antibody production. Here, we investigated the role of IL-12, IFN-gamma and IL-10 in the PAS-1-induced inhibitory mechanism using a murine model of asthma. Wild type C57BL/6, IL-12(-/-), IFN-gamma(-/-) and IL-10(-/-) mice were immunized with PAS-1 and/or OVA and challenged with the same antigens intranasally. The suppressive effect of PAS-I was demonstrated on the cellular influx into airways, with reduction of eosinophil number and eosinophil peroxidase activity in OVA + PAS-1-immunized wild type mice. This effect well correlated with a significant reduction in the levels of IL-4, IL-5, IL-13 and eotaxin in BAL fluid. Levels of IgE and IgG1 antibodies were also impaired in serum from these mice. The inhibitory activity of PAS-I was also observed in IL-12(-/-) mice, but not in IFN-gamma(-/-) and IL-10(-/-) animals. These data show that IFN-gamma and IL-10, but not IL-12, play an important role in the PAS-1 modulatory effect. (C) 2008 Elsevier Ltd. All rights reserved.

A Role for galectin-3 in renal tissue damage triggered by ischemia and reperfusion injury

Ischemic-reperfusion injury (IRI) triggers an inflammatory response involving neutrophils/macrophages, lymphocytes and endothelial cells. Galectin-3 is a multi-functional lectin with a broad range of action such as promotion of neutrophil adhesion, induction of oxidative stress, mastocyte migration and degranulation, and production of pro-inflammatory cytokines. The aim of this study was evaluate the role of galectin-3 in the inflammation triggered by IRI. Galectin-3 knockout (KO) and wild type (wt) mice were subjected to 45 min of renal pedicle occlusion. Blood and kidney samples were collected at 6, 24, 48 and 120 h. Blood urea was analyzed enzymatically, while MCP-1, IL-6 and IL-1 beta were studied by real-time PCR. Reactive oxygen species (ROS) was investigated by flow cytometry. Morphometric analyses were performed at 6, 24, 48 and 120 h after reperfusion. Urea peaked at 24 h, being significantly lower in knockout animals (wt = 264.4 +/- 85.21 mg/dl vs. gal-3 KO = 123.74 +/- 29.64 mg/dl, P = 0.001). Galectin-3 knockout animals presented less acute tubular necrosis and a more prominent tubular regeneration when compared with controls concurrently with lower expression of MCP-1, IL-6, IL-1 beta, less macrophage infiltration and lower ROS production at early time points. Galectin-3 seems to play a role in renal IRI involving the secretion of macrophage-related chemokine, pro-inflammatory cytokines and ROS production.

Sustained activation of p53 in confluent nucleotide excision repair-deficient cells resistant to ultraviolet-induced apoptosis

p53 activation is one of the main signals after DNA damage, controlling cell cycle arrest, DNA repair and apoptosis. We have previously shown that confluent nucleotide excision repair (NER)-deficient cells are more resistant to apoptosis induced by ultraviolet irradiation (UV). Here, we further investigated the effect of cell confluence on UV-induced apoptosis in normal and NER-deficient (XP-A and XP-C) cells, as well as the effects of treatments with the ATWATR inhibitor caffeine, and the patterns of p53 activation. Strong p53 activation was observed in either proliferating or confluent cells. Caffeine increased apoptosis levels and inhibited p53 activation in proliferating cells, suggesting a protective role for p53. However, in confluent NER-deficient cells no effect of caffeine was observed. Transcription recovery measurements showed decreased recovery in proliferating XPA-deficient cells, but no recovery was observed in confluent cells. The levels of the cyclin/Cdk inhibitor, p21(Waf1/Cip1), correlated well with p53 activation in proliferating cells. Surprisingly, confluent cells also showed similar activation of p21(Waf1/Cip1). These results indicate that reduced apoptosis in confluent cells is associated with the deficiency in DNA damage removal, since this effect is not clearly observed in NER-proficient cells. Moreover, the strong activation of p53 in confluent cells, which barely respond to apoptosis, suggests that this protein, under these conditions, is not linked to UV-induced cell death signaling. (c) 2008 Elsevier B.V. All rights reserved.

Vibrios dominate as culturable nitrogen-fixing bacteria of the Brazilian coral Mussismilia hispida

Taxonomic characterization was performed on the putative N-2-fixing microbiota associated with the coral species Mussismilia hispida, and with its sympatric species Palythoa caribaeorum, P. variabilis, and Zoanthus solanderi, off the coast of Sao Sebastiao (Sao Paulo State, Brazil). The 95 isolates belonged to the Gammaproteobacteria according to the 16S rDNA gene sequences. In order to identify the isolates unambiguously, pyrH gene sequencing was carried out. The majority of the isolates (n = 76) fell within the Vibrio core group, with the highest gene sequence similarity being towards Vibrio harveyi and Vibrio alginolyticus. Nineteen representative isolates belonging to V. harveyi (n = 7), V. alginolyticus (n = 8), V. campbellii (n = 3), and V parahaemolyticus (n = 1) were capable of growing six successive times in nitrogen-free medium and some of them showed strong nitrogenase activity by means of the acetylene reduction assay (ARA). It was concluded that nitrogen fixation is a common phenotypic trait among Vibrio species of the core group. The fact that different Vibrio species can fix N, might explain why they are so abundant in the mucus of different coral species. (C) 2008 Published by Elsevier GmbH.

An interleukin-1 beta (IL-1 beta) single-nucleotide polymorphism at position 3954 and red complex periodontopathogens independently and additively modulate the levels of IL-1 beta in diseased periodontal tissues

Inflammatory cytokines such as interieukin-1 beta (IL-1 beta) are involved in the pathogenesis of periodontal diseases. A high individual variation in the levels of IL-10 mRNA has been verified, which is possibly determined by genetic polymorphisms and/or by the presence of periodontopathogens such as Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, and Aggregatibacter actinomycetemcomitans. In this study, we investigated the role of an IL-10 promoter single-nucleotide polymorphism at position 3954 [IL-1 beta(3954) SNP] and the presence of the periodontopathogens in the determination of the IL-1 beta levels in the periodontal tissues of nonsmoking chronic periodontitis (CP) patients (n = 117) and control (C) subjects in = 175) and the possible correlations with the clinical parameters of the disease. IL-1 beta(3954) SNP was investigated by restriction fragment length polymorphism, while the IL-1 beta levels and the presence of the periodontopathogens were determined by real-time PCR. Similar frequencies of IL-1 beta(3954) SNP were found in the C and CP groups, in spite of a trend toward a higher incidence of T alleles in the CP group. The IL-1 beta (3954) SNP CT and TT genotypes, as well as P. gingivalis, T. forsythia, and T. denticola, were associated with higher IL-1 beta levels and with higher values of the clinical parameters of disease severity. Concomitant analyses demonstrate that IL-1 beta(3954) and the red complex periodontopathogens were found to independently and additively modulate the levels of IL-1 beta in periodontal tissues. Similarly, the concurrent presence of both factors was associated with increased scores of disease severity. IL-1 beta(3954) genotypes and red complex periodontopathogens, individually and additively, modulate the levels of IL-1 beta in the diseased tissues of nonsmoking CP patients and, consequently, are potentially involved in the determination of the disease outcome.

Analysis of the genome of Spodoptera frugiperda nucleopolyhedrovirus (SfMNPV-19) and of the high genomic heterogeneity in group II nucleopolyhedroviruses

The genome of the most virulent among 22 Brazilian geographical isolates of Spodoptera frugiperda nucleopolyhedrovirus, isolate 19 (SfMNPV-1 9), was completely sequenced and shown to comprise 132 565 bp and 141 open reading frames (ORFs). A total of 11 ORFs with no homology to genes in the GenBank database were found. Of those, four had typical baculovirus; promoter motifs and polyadenylation sites. Computer-simulated restriction enzyme cleavage patterns of SfMNPV-1 9 were compared with published physical maps of other SfMNPV isolates. Differences were observed in terms of the restriction profiles and genome size. Comparison of SfMNPV-1 9 with the sequence of the SfMNPV isolate 3AP2 indicated that they differed due to a 1427 bp deletion, as well as by a series of smaller deletions and point mutations. The majority of genes of SfMNPV-1 9 were conserved in the closely related Spodoptera exigua NPV (SeMNPV) and Agrotis segetum NPV (AgseMNPV-A), but a few regions experienced major changes and rearrangements. Synthenic maps for the genomes of group 11 NPVs revealed that gene collinearity was observed only within certain clusters. Analysis of the dynamics of gene gain and loss along the phylogenetic tree of the NPVs showed that group 11 had only five defining genes and supported the hypothesis that these viruses form ten highly divergent ancient lineages. Crucially, more than 60% of the gene gain events followed a power-law relation to genetic distance among baculoviruses, indicative of temporal organization in the gene accretion process.

Molecular characterization and fumonisin production by Fusarium verticillioides isolated from corn grains of different geographic origins in Brazil

Gibberella moniliformis is most commonly associated with maize worldwide and produces high levels of fumonisins, some of the most agriculturally important mycotoxins. Studies demonstrate that molecular methods can be helpful for a rapid identification of Fusarium species and their levels of toxin production. The purpose of this research was to apply molecular methods (AFLP, TEF-1 alpha partial gene sequencing and PCR based on MAT alleles) for the identification of Fusarium species isolated from Brazilian corn and to verify if real time RT-PCR technique based on FUM1 and FUM19 genes is appropriated to estimate fumonisins B(1) and B(2) production levels. Among the isolated strains, 96 were identified as Fusarium verricillioides, and four as other Fusarium species. Concordant phylogenies were obtained by AFLP and TEF-1 alpha sequencing, permitting the classification of the different species into distinct clades. Concerning MAT alleles, 70% of the F. verricillioides isolates carried the MAT-1 and 30% MAT-2. A significant correlation was observed between the expression of the genes and toxin production r=0.95 and r=0.79 (correlation of FUM1 with FB(1) and FB(2), respectively, P < 0.0001): r=0.93 and r =0.78 (correlation of FUM19 with FB(1) and FB(2). respectively, P < 0.0001). Molecular methods used in this study were found to be useful for the rapid identification of Fusarium species. The high and significant correlation between FUM1 and FUM19 expression and fumonisins production suggests that real time RT-PCR is suitable for studies considering the influence of abiotic and biotic factors on expression of these genes. This is the first report concerning the expression of fumonisin biosynthetic genes in Fusarium strains isolated from Brazilian agricultural commodity. (c) 2010 Elsevier B.V. All rights reserved.

Hospital-based Surveillance to Evaluate the Impact of Rotavirus Vaccination in Sao Paulo, Brazil

Background: Brazil implemented routine immunization with the human rotavirus vaccine, Rotarix, in 2006 and vaccination coverage reached 81% in 2008 in Sao Paulo. Our aim was to assess the impact of immunization on the incidence of severe rotavirus acute gastroenteritis (AGE). Methods: We performed a 5-year (2004-2008) prospective surveillance at a sentinel hospital in Sao Paulo, with routine testing for rotavirus in all children less than 5 years of age hospitalized with AGE. Genotypes of positive samples were determined by reverse transcription polymerase chain reaction. Results: During the study, 655 children hospitalized with AGE were enrolled; of whom 169 (25.8%) were positive for rotavirus. In the post-vaccine period, a 59% reduction in the number of hospitalizations of rotavirus AGE and a 42.2% (95% confidence interval [CI], 18.6%-59.0%; P = 0.001) reduction in the proportion of rotavirus-positive results among children younger than 5 years were observed, with the greatest decline among infants (69.2%; 95% CI, 24.7%-87.4%; P = 0.004). Furthermore, the number of all-cause hospitalizations for AGE was reduced by 29% among children aged <5 years. The onset and peak incidences of rotavirus AGE occurred 3 months later in the 2007 and 2008 seasons compared with previous years. Genotype G2 accounted for 15%, 70%, and 100% of all cases identified, respectively, in 2006, 2007, and 2008. Conclusions: After vaccine implementation, a marked decline in rotavirus AGE hospitalizations was demonstrated among children younger than 5 years of age, with the greatest reduction in the age groups targeted for vaccination. The predominance of genotype G2P[4] highlights the need of continued postlicensure surveillance studies.