Role of regulatory T cells in long-term immune dysfunction associated with severe sepsis

Objective: To investigate the role of regulatory T cells in the modulation of long-term immune dysfunction during experimental sepsis. It is well established that sepsis predisposes to development of a pronounced immunosuppression. Nevertheless, the mechanisms underlying the immune dysfunction after sepsis are still not well understood. Design: Prospective experimental study. Setting: University research laboratory. Interventions: Wild-type mice underwent cecal ligation and puncture and were treated with antibiotic during 3 days after surgery. On days 1, 7, or 15 after cecal ligation and puncture, the frequency of regulatory T cells, proliferation of CD4(+) T cells and bacterial counts were evaluated. Fifteen days after cecal ligation and puncture, surviving mice underwent secondary pulmonary infection by intranasal inoculation of nonlethal dose of Legionella pneumophila. Some mice received agonistic glucocorticoid-induced tumor necrosis factor receptor antibody (DTA-1) before induction of secondary infection. Measurements and Main Results: Mice surviving cecal ligation and puncture showed a markedly increased frequency of regulatory T cells in thymus and spleen, which was associated with reduced proliferation of CD4(+) T cells. Fifteen days after cecal ligation and puncture, all sepsis-surviving mice succumbed to nonlethal injection of L. pneumophila. Treatment of mice with DTA-1 antibody reduced frequency of regulatory T cells, restored CD4(+) T cell proliferation, reduced the levels of bacteria in spleen, and markedly improved survival of L. pneumophila infection. Conclusion: These findings suggest that regulatory T cells play an important role in the progression and establishment of immune dysfunction observed in experimental sepsis. (Crit Care Med 2010; 38: 1718-1725)

Essential role of platelet-activating factor receptor in the pathogenesis of Dengue virus infection

Severe dengue infection in humans causes a disease characterized by thrombocytopenia, increased levels of cytokines, increased vascular permeability, hemorrhage, and shock. Treatment is supportive. Activation of platelet-activating factor (PAF) receptor (PAFR) on endothelial cells and leukocytes induces increase in vascular permeability, hypotension, and production of cytokines. We hypothesized that activation of PAFR could account for the major systemic manifestations of dengue infection. Inoculation of adult mice with an adapted strain of Dengue virus caused a systemic disease, with several features of the infection in humans. In PAFR(-/-) mice, there was decreased thrombocytopenia, hemoconcentration, decreased systemic levels of cytokines, and delay of lethality, when compared with WT infected mice. Treatment with UK-74,505, an orally active PAFR antagonist, prevented the above-mentioned manifestations, as well as hypotension and increased vascular permeability, and decreased lethality, even when started 5 days after virus inoculation. Similar results were obtained with a distinct PAFR antagonist, PCA-4246. Despite decreased disease manifestation, viral loads were similar (PAFR(-/-)) or lower (PAFR antagonist) than in WT mice. Thus, activation of PAFR plays a major role in the pathogenesis of experimental dengue infection, and its blockade prevents more severe disease manifestation after infection with no increase in systemic viral titers, suggesting that there is no interference in the ability of the murine host to deal with the infection. PAFR antagonists are disease-modifying agents in experimental dengue infection.

Cigarette smoke exposure impairs respiratory epithelial ciliogenesis

Background: Cigarette smoke exposure is considered an important negative prognostic factor for chronic rhinosinusitis (CRS) patients. However, there is no clear mechanistic evidence implicating cigarette smoke exposure in the poor clinical evolution of the disease or in the maintenance of the inflammatory state characterizing CRS. This study aimed to evaluate the effects of cigarette smoke exposure on respiratory cilia differentiation. Methods: Monse nasal septal epithelium cultures grown at an air-liquid interface were used as a model of respiratory epithelium. After 5 days of cell growth, cultures were exposed to air on the apical surface. Additionally, cigarette smoke condensate (CSC; the particulate phase of tobacco smoke) or cigarette smoke extract (CSE; the volatile phase) Were diluted in the basolateral compartment in different concentrations. After 15 days of continuous exposure, scanning electron microscopy and immunofluorescence for type IV tubulin were used to determine presence and maturation of cilia. Transepithelial resistance was also recorded to evaluate confluence and physiological barrier integrity. Results: CSC and CSE impair ciliogenesis in a dose-dependent manner with notable effects in concentrations higher than 30 mu g/mL, yielding >70% nonciliation and shorter cilia compared With control. No statistical difference on transepithelial resistance was evident. Conclusion: CSC and CSE exposure negatively impacts ciliogenesis of respiratory cells at concentrations not effecting transepithelial resistance. The impairment on ciliogenesis reduce the mucociliary clearance apparatuts after injury and/or infection and may explain the poor response to therapy for CRS patients exposed to tobacco smoke.

Evidence of Renal Infection in Fatal Cases of 2009 Pandemic Influenza A (HI NI)

The 2009 pandemic influenza A (H1N1) caused significant morbidity and mortality. Acute lung injury is the hallmark of the disease, but multiple organ system dysfunction can develop and lead to death. Therefore, we sought to investigate whether there was postmortem evidence of H1N1 presence and virus-induced organ injury in autopsy specimens. Five cases in which patients died of influenza A (H1N1) virus infection were studied. The lungs of all patients showed macroscopic and microscopic findings already described for H1N1 (consolidation, edema, hemorrhage, alveolar damage, hyaline membrane, and inflammation), and H1N1 viruses were present in alveolar cells in immunochemical studies. Acute tubular necrosis was present in all cases, but there was no evidence of direct virus-induced kidney injury. Nevertheless, H1N1 viruses were found in the cytoplasm of glomerular macrophages in the kidneys of 4 patients. Therefore, our data provide strong evidence that H1N1 presence is not restricted to the lungs.

Gastroesophageal Reflux in Severe Cases of Robin Sequence Treated With Nasopharyngeal Intubation

Objective: To study the prevalence of abnormal gastroesophageal reflux in infants with Robin sequence who had severe respiratory obstruction treated with nasopharyngeal intubation and to evaluate the efficacy of nonsurgical treatment. Design: Longitudinal prospective study. Setting: Hospital de Reabilitacao de Anomalias Craniofaciais, University of Sao Paulo, Brazil. Patients: Twenty infants with severe isolated Robin sequence treated with nasopharyngeal intubation. Interventions: We performed 24-hour esophageal pH monitoring on each child at 2, 4, and 6 months of age. Respiratory and feeding status were evaluated. We considered abnormal gastroesophageal reflux as reflux index values above the 95th percentile of the Vandenplas reference for normal children. Results: The prevalence of reflux index above the 95th percentile at the first exam was 6/20, a value significantly higher than the reference (5/103, p < .01). At the second and third exams, reflux index values were decreased. Ninety percent of the infants showed improvement of respiratory difficulty and developed oral feeding capacity. Conclusions: The prevalence of abnormal gastroesophageal reflux is higher in infants with severe cases of Robin sequence than in normal infants. Nonsurgical procedures improved respiratory and feeding difficulties of most of these infants.

Cutting Edge: Nucleotide-Binding Oligomerization Domain 1-Dependent Responses Account for Murine Resistance against Trypanosoma cruzi Infection

An effective innate immune recognition of the intracellular protozoan parasite Trypanosoma cruzi is critical for host resistance against Chagas disease, a severe and chronic illness that affects millions of people in Latin America. In this study, we evaluated the participation of nucleotide-binding oligomerization domain (Nod)like receptor proteins in host response to T cruzi infection and found that Nod1-dependent, but not Nod2-dependent, responses are required for host resistance against infection. Bone marrow-derived macrophages from Nod1(-/-) mice showed an impaired induction of NF-kappa B-dependent products in response to infection and failed to restrict T cruzi infection in presence of IFN-gamma. Despite normal cytokine production in the sera, Nod1(-/-) mice were highly susceptible to T cruzi infection, in a similar manner to MyD88(-/-) and NO synthase 2(-/-) mice. These studies indicate that Nod1-dependent responses account for host resistance against T cruzi infection by mechanisms independent of cytokine production. The Journal of Immunology, 2010, 184: 1148-1152.

Effects of progressive exercise during phase I cardiac rehabilitation on the heart rate variability of patients with acute myocardial infarction

Purpose. aEuro integral Heart rate variability (HRV) decreases after an acute myocardial infarction (AMI) due to changes in cardiac autonomic balance. The purpose of the present study, therefore, was to evaluate the effects of a progressive exercise protocol used in phase I cardiac rehabilitation on the HRV of patients with post-AMI. Material and methods. aEuro integral Thirty-seven patients who had been admitted to hospital with their first non-complicated AMI were studied. The treated group (TG, n == 21, age == 52 +/-+/- 12 years) performed a 5-day programme of progressive exercise during phase I cardiac rehabilitation, while the control group (CG, n == 16, age == 54 +/-+/- 11 years) performed only respiratory exercises. Instantaneous heart rate (HR) and RR interval were acquired by a HR monitor (Polar (R) A (R) S810i). HRV was analysed by frequency domain methods. Power spectral density was expressed as normalised units (nu) at low (LF) and high (HF) frequencies, and as LF/HF. Results. aEuro integral After 5 days of progressive exercise, the TG showed an increase in HFnu (35.9 +/-+/- 19.5 to 65.19 +/-+/- 25.4) and a decrease in LFnu and LF/HF (58.9 +/-+/- 21.4 to 32.5 +/-+/- 24.1; 3.12 +/-+/- 4.0 to 1.0 +/-+/- 1.5, respectively) in the resting position (p < 0.05). No changes were observed in the CG. Conclusions. aEuro integral A progressive physiotherapeutic exercise programme carried out during phase I cardiac rehabilitation, as supplement to clinical treatment increased vagal and decreased sympathetic cardiac modulation in patients with post-AMI.

Impact of implementing an exclusively dedicated respiratory isolation room in a Brazilian tertiary emergency department

Background Occupational risk due to airborne disease challenges healthcare institutions. Environmental measures are effective but their cost-effectiveness is still debatable and most of the capacity planning is based on occupational rates. Better indices to plan and evaluate capacity are needed. Goal To evaluate the impact of installing an exclusively dedicated respiratory isolation room (EDRIR) in a tertiary emergency department (ED) determined by a time-to-reach-facility method. Methods A group of patients in need of respiratory isolation were first identified-group I (2004; 29 patients; 44.1 +/- 3.4 years) and the occupational rate and time intervals (arrival to diagnosis, diagnosis to respiratory isolation indication and indication to effective isolation) were determined and it was estimated that adding an EDRIR would have a significant impact over the time to isolation. After implementing the EDRIR, a second group of patients was gathered in the same period of the year-group II (2007; 50 patients; 43.4 +/- 1.8 years) and demographic and functional parameters were recorded to evaluate time to isolation. Cox proportional hazard models adjusted for age, gender and inhospital respiratory isolation room availability were obtained. Results Implementing an EDRIR decreased the time from arrival to indication of respiratory isolation (27.5 +/- 9.3 X 3.7 +/- 2.0; p = 0.0180) and from indication to effective respiratory isolation (13.3 +/- 3.0 X 2.94 +/- 1.06; p = 0.003) but not the respiratory isolation duration and total hospital stay. The impact on crude isolation rates was very significant (8.9 X 75.4/100.000 patients; p < 0.001). The HR for effective respiratory isolation was 26.8 (95% CI 7.42 to 96.9) p < 0.001 greater for 2007. Conclusion Implementing an EDRIR in a tertiary ED significantly reduced the time to respiratory isolation.

Recognition by toll-like receptor 2 induces antigen-presenting cell activation and Th1 programming during infection by Neospora caninum

Neospora caninum is an apicomplexan parasite responsible for major economic losses due to abortions in cattle. Toll-like receptors (TLRs) sense specific microbial products and direct downstream signaling pathways in immune cells, linking innate, and adaptive immunity. Here, we analyze the role of TLR2 on innate and adaptive immune responses during N. caninum infection. Inflammatory peritoneal macrophages and bone marrow-derived dendritic cells exposed to N. caninum-soluble antigens presented an upregulated expression of TLR2. Increased receptor expression was correlated to TLR2/MyD88-dependent antigen-presenting cell maturation and pro-inflammatory cytokine production after stimulation by antigens. Impaired innate responses observed after infection of mice genetically deficient for TLR2((-/-)) was followed by downregulation of adaptive T helper 1 (Th1) immunity, represented by diminished parasite-specific CD4(+) and CD8(+) T-cell proliferation, IFN-gamma:interleukin (IL)-10 ratio, and IgG subclass synthesis. In parallel, TLR2(-/-) mice presented higher parasite burden than wild-type (WT) mice at acute and chronic stages of infection. These results show that initial recognition of N. caninum by TLR2 participates in the generation of effector immune responses against N. caninum and imply that the receptor may be a target for future prophylactic strategies against neosporosis. Immunology and Cell Biology (2010) 88, 825-833; doi:10.1038/icb.2010.52; published online 20 April 2010

Hemodynamic effects of PEEP in a porcine model of HCl-induced mild acute lung injury

Positive end-expiratory pressure (PEEP) and sustained inspiratory insufflations (SI) during acute lung injury (ALI) are suggested to improve oxygenation and respiratory mechanics. We aimed to investigate the hemodynamic effects of PEEP with and without alveolar recruiting maneuver in a mild ALI model induced by inhalation of hydrochloric acid. Thirty-two pigs were randomly allocated into four groups (Control-PEEP, Control-SI, ALI-PEEP and ALI-SI). ALI was induced by intratracheal instillation of hydrochloric acid. PEEP values were progressively increased and decreased from 5, 10, 15 and 20 cmH(2)O in all groups. Three SIs maneuvers of 30 cmH(2)O for 20 s were applied to the assignable groups between each PEEP level. Transesophageal echocardiography (TEE), global hemodynamics, oxygenation indexes and gastric tonometry were measured 5 min after the maneuvers had been concluded and at each established value of PEEP (5, 10, 15 and 20 cmH(2)O). The cardiac index, ejection fraction and end-diastolic volume of right ventricle were significantly (P < 0.001) decreased with PEEP in both Control and ALI groups. Left ventricle echocardiography showed a significant decrease in end-diastolic volume at 20 cmH(2)O of PEEP (P < 0.001). SIs did not exert any significant hemodynamic effects either early (after 5 min) or late (after 3 h). In a mild ALI model induced by inhalation of hydrochloric acid, significant hemodynamic impairment characterized by cardiac function deterioration occurred during PEEP increment, but SI, probably due to low applied values (30 cmH(2)O), did not exert further negative hemodynamic effects. PEEP should be used cautiously in ALI caused by acid gastric content inhalation.

Detection and Molecular Characterization of Gene 3 and 5 of Turkey Coronavirus from Turkeys with Severe Enteritis in Brazil

Turkey coronavirus (TCoV) is a causative agent associated with poult enteritis and mortality syndrome (PEMS) in turkeys worldwide. The disease is an acute, highly contagious enteric disease that is characterized by depression, anorexia, diarrhea, and high mortality in commercial turkey flocks. The presence of TCoV in 12 intestinal-content samples, from turkey flocks aged between 10 and 104 days and exhibiting severe enteritis, was monitored during the period of 2004 to 2006. TCoV detection was accomplished by a reverse transcriptase-polymerase chain reaction (RT-PCR) through amplification of the 3` UTR region, followed by amplification of genes 3 and 5. Molecular characterization of the viruses was done through amplification of genes 3 and 5 and showed evidence of genetic similarity between them, although they differed from sequences of other TCoVs described in the literature. In relation to gene 3, samples showed a greater relationship with chicken infectious bronchitis virus (IBV), while gene 5 showed greater identity with pheasant coronavirus (PhCoV). Our results suggest that the strategy of amplification of the 3` UTR region, followed by sequencing of genes 3 and 5, has proven to be an effective means of detecting TCoV in intestinal contents.

Hematopoietic response of rats exposed to the impact of an acute psychophysiological stressor on responsiveness to an in vivo challenge with Listeria monocytogenes: Modulation by Chlorella vulgaris prophylactic treatment

In this study, we investigated the hematopoietic response of rats pretreated with CV and exposed to the impact of acute escapable, inescapable or psychogenical stress on responsiveness to an in vivo challenge with Listeria monocytogenes. No consistent changes were observed after exposure to escapable footshock. Conversely, the impact of uncontrollable stress (inescapable and psychogenical) was manifested by an early onset and increased severity and duration of myrelossuppression produced by the infection. Small size CFU-CM colonies and increased numbers of clusters were observed, concurrently to a greater expansion in the more mature population of bone marrow granulocytes. No differences were observed between the responses of both uncontrollable stress regimens. CV prevented the myelossuppression caused by stress/infection due to increased numbers of CFU-GM in the bone marrow. Colonies of cells tightly packed, with a very condensed nucleus; in association with a greater expansion in the more immature population of bone marrow granulocytes were observed. Investigation of the production of colony-stimulating factors revealed increased colony-stimulating activity (CSA) in the serum of normal and infected/stressed rats treated with the algae. CV treatment restored/enhanced the changes produced by stress/infection in total and differential bone marrow and peripheral cells counts. Further studies demonstrated that INF-gamma is significantly reduced, whereas IL-10 is significantly increased after exposure to Uncontrollable stress. Treatment with CV significantly increased INF-gamma levels and diminished the levels of IL-10. Uncontrollable stress reduced the protection afforded by CV to a lethal dose of L. monocytogenes, with survival rates being reduced from (50%) in infected rats to 20% in infected/stressed rats. All together, our results suggest Chlorella treatment as an effective tool for the prophylaxis of post-stress myelossupression, including the detrimental effect of stress on the course and outcome of infections. (C) 2008 Elsevier Inc. All rights reserved.

The essential role of toll like receptor-4 in the control of Aggregatibacter actinomycetemcomitans infection in mice

Objective: Aggregatibacter actinomycetemcomitans is an oral Gram-negative bacterium that contributes to periodontitis progression. Isolated antigens from A. actinomycetemcomitans could be activating innate immune cells through Toll-like receptors (TLRs). In this study, we evaluated the role of TLR4 in the control of A. actinomycetemcomitans infection. Material and Methods: We examined the mechanisms that modulate the outcome of A. actinomycetemcomitans-induced periodontal disease in TLR4(-/-) mice. The production of cytokines was evaluated by ELISA. The bacterial load was determined by counting the number of colony-forming units per gram of tissue. Results: The results showed that TLR4-deficient mice developed less severe periodontitis after A. actinomycetemcomitans infection, characterized by significantly lower bone loss and inflammatory cell migration to periodontal tissues. However, the absence of TLR4 facilitated the A. actinomycetemcomitans dissemination. Myeloperoxidase activity was diminished in the periodontal tissue of TLR4(-/-) mice. We observed a significant reduction in the production of tumour necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1 beta in the periodontal tissue of TLR4(-/-) mice. Conclusion: The results of this study highlighted the role of TLR4 in controlling A. actinomycetemcomitans infection.

CCR5-dependent regulatory T cell migration mediates fungal survival and severe immunosuppression

Paracoccidioidomycosis, a debilitating pulmonary mycosis, is caused by the dimorphic fungus Paracoccidioides brasiliensis. The infection results in the formation of granulomas containing viable yeast cells that are the fungal sources for disease reactivation. Because CD4(+)CD25(+) regulatory T cells (Tregs) are in the lesions of patients with paracoccidioidomycosis, the migration of Treg cells is dependent on the axis chemokine-chemokine receptors, and CCR5 ligands are produced in P. brasiliensis-induced lesions, we investigated the role of CCR5 in the control of the infection. The results showed that CCR5(-/-) mice are more efficient in controlling fungal growth and dissemination and exhibited smaller granulomas than wild-type (WT) mice. In the absence of CCR5, the percentage of CD4(+)CD25(+) T cells expressing Foxp3, glucocorticoid-induced TNFR (GITR), CD103, CD45(low), and CTLA-4 in the granulomas was significantly decreased. Interestingly, P. brasiliensis infection resulted in an absence of T cell proliferation in response to Con A in WT but not CCR5(-/-) mice that was abrogated by anti-CTLA-4 mAb and anti-GITR mAb. Moreover, the adoptive transfer of CD4(+)CD25(+) but not CD4(+)CD25(-) T cells from infected WT to infected CCR5(-/-) mice resulted in a significant increase in fungal load. Overall, CCR5 is a key receptor for the migration of Treg cells to the site of P. brasiliensis infections leading to down-modulation of effector immune response and the long-term presence of the fungus in the granulomas. Thus, a tight control of Treg cell migration to the granulomatous lesions could be an important mechanism for avoiding exacerbation and reactivation of the disease.