Structural characterization of Campylobacter jejuni lipooligosaccharide outer cores associated with Guillain-Barre and Miller Fisher syndromes

Molecular mimicry between lipooligosaccharides (LOS) of Campylobacter jejuni and gangliosides in peripheral nerves plays a crucial role in the pathogenesis of C. jejuni-related Guillain-Barré syndrome (GBS). We have analyzed the LOS outer core structures of 26 C. jejuni strains associated with GBS and its variant, Miller Fisher syndrome (MFS), by capillary electrophoresis coupled with electrospray ionization mass spectrometry. Sixteen out of 22 (73%) GBS-associated and all 4 (100%) MFS-associated strains expressed LOS with ganglioside mimics. GM1a was the most prevalent ganglioside mimic in GBS-associated strains (10/22, 45%), and in eight of these strains, GM1a was found in combination with GD1a mimics. All seven strains isolated from patients with ophthalmoplegia (GBS or MFS) expressed disialylated (GD3 or GD1c) mimics. Three out of 22 GBS-associated strains (14%) did not express sialylated ganglioside mimics because their LOS locus lacked the genes necessary for sialylation. Three other strains (14%) did not express ganglioside mimics because of frameshift mutations in either the cstII sialyltransferase gene or the cgtB galactosyltransferase gene. It is not possible to determine if these mutations were already present during C. jejuni infection. This is the first report in which mass spectrometry combined with DNA sequence data were used to infer the LOS outer core structures of a large number of neuropathy-associated C. jejuni strains. We conclude that molecular mimicry between gangliosides and C. jejuni LOS is the presumable pathogenic mechanism in most cases of C. jejuni-related GBS. However, our findings suggest that in some cases, other mechanisms may play a role. Further examination of the disease etiology in these patients is mandatory.     

Prolonged hypoxia concomitant with serum deprivation induces massive human mesenchymal stem cell death

Mesenchymal stem cells (MSCs) have been proposed for the repair of damaged tissue including bone, cartilage, and heart tissue. Upon in vivo transplantation, the MSCs encounter an ischemic microenvironment characterized by reduced oxygen (O2) tension and nutrient deprivation that may jeopardize viability of the tissue construct. The aim of this study was to assess the effects of serum deprivation and hypoxia on the MSC survival rates in vitro. As expanded MSCs are transferred from plastic to a scaffold in most tissue engineering approaches, possibly inducing loss of survival signals from matrix attachments, the effects of a scaffold shift on the MSC survival rates were also assessed. Human MSCs were exposed for 48 hours to (i) a scaffold substrate shift, (ii) serum deprivation, and (iii) O2 deprivation. MSCs were also exposed to prolonged (up to 120 hours) hypoxia associated with serum deprivation. Cell death was assessed by Live/Dead staining and image analysis. The MSC death rates were not affected by the shift to scaffold or 48-hour hypoxia, but increased with fetal bovine serum (FBS) starvation, suggesting that between the two components of ischemia, nutrient deprivation is the stronger factor. Long-term hypoxia combined with serum deprivation resulted in the complete death of MSCs (99 +/- 1%), but this rate was reduced by half when MSCs were exposed to hypoxia in the presence of 10% FBS (51 +/- 31%). These results show that MSCs are sensitive to the concurrent serum and O2 deprivation to which they are exposed when transplanted in vivo, and call for the development of new transplantation methods.     

Sol gel derived SiO(2)-CaO-MgO-P(2)O(5) bioglass system--preparation and in vitro characterization

The synthesis of SiO(2)-CaO-MgO-P(2)O(5) bioactive glass was carried out by the sol-gel method. Sol-gel derived bioglass material was crushed into powder to produce pellet disks by uniaxial pressing, followed by sintering at 900 degrees C. The biocompatibility evaluation of the formed glass was assessed through in vitro cell culture experiments and immersion studies in simulated body fluid (SBF) for different time intervals while monitoring the pH changes and the concentration of calcium and magnesium in the SBF medium. Scanning electron microscopy, X-ray energy dispersive spectroscopy, X-ray diffraction analysis, and FT-IR spectroscopy studies were conducted before and after contact of the material with SBF. At first, an amorphous calcium phosphate was formed; after 21 days this surface consisted of deposited crystalline spheres of apatite. The present investigation also revealed that the sol-gel derived quaternary bioglass system has the ability to support the growth of osteoblast-like cells in vitro and to promote osteoblast differentiation by stimulating the expression of major phenotypic markers.     

Caveolin-1(-/-)- and caveolin-2(-/-)-deficient mice both display numerous skeletal muscle abnormalities, with tubular aggregate formation

Here, we examine the role of "non-muscle" caveolins (Cav-1 and Cav-2) in skeletal muscle biology. Our results indicate that skeletal muscle fibers from male Cav-1(-/-) and Cav-2(-/-) mice show striking abnormalities, such as tubular aggregates, mitochondrial proliferation/aggregation, and increased numbers of M-cadherin-positive satellite cells. Notably, these skeletal muscle defects were more pronounced with increasing age. Because Cav-2-deficient mice displayed normal expression levels of Cav-1, whereas Cav-1-null mice exhibited an almost complete deficiency in Cav-2, these skeletal muscle abnormalities seem to be due to loss of Cav-2. Thus, Cav-2(-/-) mice represent a novel animal model-and the first genetically well-defined mouse model-that can be used to study the pathogenesis of tubular aggregate formation, which remains a poorly understood age-related skeletal muscle abnormality. Finally, because Cav-1 and Cav-2 were not expressed within mature skeletal myofibers, our results indicate that development of these abnormalities probably originates in stem/precursor cells, such as satellite cells or myoblasts. Consistent with this hypothesis, skeletal muscle isolated from male Cav-3(-/-) mice did not show any of these abnormalities. As such, this is the first study linking stem cells with the genesis of these intriguing muscle defects.     

Sleep-induced hypotension precipitates severe myocardial ischemia

STUDY OBJECTIVES: Epidemiologic studies have shown a high frequency of major cardiac events at night in patients with coronary artery disease. This has been attributed to the sympathetic surges accompanying rapid eye movement (REM) sleep; the role of non-REM sleep, which comprises 80% of total sleep duration, has been largely neglected. Accordingly, we evaluated the effect of non-REM sleep on contractile function in a region of the left ventricular wall supplied by a flow-limiting coronary stenosis. DESIGN: Eight domestic pigs were chronically instrumented to measure regional left ventricular contractile function (wall thickening), coronary blood flow, and systemic hemodynamic variables. Measurements were obtained: (1) during wakefulness, i.e., conscious condition, prior to imposition of coronary stenosis; (2) during wakefulness following imposition of coronary stenosis (30% reduction of baseline coronary blood flow from 40 +/- 4 to 27 +/- 3 mL/min); and (3) during non-REM sleep with coronary stenosis maintained. RESULTS: During wakefulness, coronary stenosis reduced wall thickening (from 23.3 +/- 3.4% to 15.7 +/- 2.0%), whereas mean arterial pressure and heart rate were unchanged. With coronary stenosis maintained, the onset of non-REM sleep caused 20% decreases in mean arterial pressure and coronary blood flow, accompanied by a cessation of regional wall thickening, i.e., akinesis (wall thickening = 0.2 +/- 2.8%), indicating severe myocardial ischemia. CONCLUSIONS: The arterial hypotension, and associated reduction in coronary blood flow, during non-REM sleep precipitated severe myocardial ischemia in a region of the left ventricular wall supplied by flow-limiting coronary stenosis. Such episodes would occur repeatedly during the sleep cycle and could potentially set the stage for a major cardiac event during the sympathetic activation accompanying REM sleep or morning activities.     

Anticonvulsant effect of phytol in a pilocarpine model in mice

The present study investigated the effects of phytol in pilocarpine-induced seizures. The latency for development of convulsions and mortality rate was recorded in this model using mice. The results revealed that phytol (25, 50 and 75 mg/kg, i.p.) increased latency to first seizure and decreased percentage of these seizures. Moreover, phytol also protected the animals against status epilepticus induced by pilocarpine, and decreased the mortality rate. Mice treated with pilocarpine (n = 24) presented 100% of mortality during the first hour of observation. In turn, phytol-pretreated animals within 30 min before the administration of pilocarpine (400 mg/kg) remained alive during the first hour of observation. On the other hand, 6–8 h after administration of pilocarpine it was observed that 10 (41.66%), 8 (33.33%) and 4 (16.66%) animals died (respectively). Thus, the pretreatment with phytol was able to block mortality rate during the first hour in acute phase of seizures, and significantly reduced this rate in a dose-dependent manner (p < 0.05), suggesting an anticonvulsant effect. In addition, none of the phytol effects was blocked by pre-treatment with flumazenil, an antagonist of benzodiazepine receptors. In conclusion, phytol exhibits anticonvulsant activity by modulating of neurotransmitter systems, but further investigations are in progress to confirm this pharmacological property.     

Developments in the field of allergy in 2011 through the eyes of Clinical and Experimental Allergy

As in previous years, we felt it would be of value to our readership to summarize the new information provided by the authors who have published in Clinical and Experimental Allergy in 2011 and set this in the context of recent advances in our understanding of the pathogenesis and management of allergic disease in all its many manifestations. In 2011, about 210 articles were published in Clinical and Experimental Allergy including editorials, reviews, opinion articles, guidelines, letters, book reviews and of course at the heart of the journal, papers containing original data. As before, this review is divided into sections based on the way the journal is structured, although this year we have grouped together all the papers dealing with mechanisms of allergic disease, whether they involve patients (clinical mechanisms), pure in vitro studies (basic mechanisms) or animal models (experimental models), as we felt this was a more coherent way to deal with the subject. In the field of asthma and rhinitis, the relationship between airway inflammation and airway dysfunction was of perennial interest to investigators, as were phenotypes and biomarkers. Aspirin hypersensitivity appeared in studies in several papers and there was new interest in asthma in the elderly. The mechanisms involved in allergic disease describe advances in our understanding of T cell responses, the relationship between inflammation and disease, mast cell and basophil activation, steroid resistance and novel therapies. In the section dealing with epidemiology, studies seeking to identify risk factors for allergic disease including vitamin D are prominent, as once again are studies investigating gene–environment interactions. The clinical allergy section focuses on drug allergy, food allergy and immunotherapy. The area of oral immunotherapy for food allergy is well covered and we were grateful to Stephen Durham for guest editing an outstanding special issue on immunotherapy in the centenary year of Leonard Noon's pioneering work. Lastly, in the field of allergens, the interest in component‐resolved diagnosis continues to grow and there are also articles describing important novel cultivars and the effect of food processing on the allergenic properties of foods. Another terrific year, full of important and high‐quality work,which the journal has been proud to bring to the allergy community.     

Assessing the interest to participate in a dengue vaccine efficacy trial among residents of Puerto Rico

Dengue, endemic in Puerto Rico, is a major public health problem. Vaccines are thought the best means to prevent dengue because vector control alone has been largely ineffective. We implemented qualitative studies in 2006 and 2010 to determine the acceptability of conducting placebo-controlled dengue vaccine efficacy trials in Puerto Rican children. Key informant interviews and focus groups with parents and children were conducted in municipalities with high dengue incidence. We used structured open-ended questions to determine motivators and attitudes regarding vaccine trial participation. Knowledge about dengue risk and prevention, and knowledge, attitudes, and beliefs regarding vaccines and vaccine trials were assessed. Using grounded theory, we conducted content analysis and established categories and sub-categories of participant responses. All participants were knowledgeable about dengue prevention and perceived children as most affected age groups. Participants were aware of vaccines benefits and they thought a vaccine could prevent dengue. However, most would not allow their children to participate in a placebo-controlled vaccine trial. Barriers included lack of trust in new vaccines and vaccine trial procedures; fear of developing dengue or side effects from the vaccine and lack of information about candidate dengue vaccines. Participants thought information, including results of previous trials might overcome barriers to participation. Motivators for participation were altruism, protection from dengue, free medical attention, and compensation for transportation and participation. Parents would consider children participation if accurate vaccine trial information is provided.     

Hepatoprotective treatment attenuates oxidative damages induced by carbon tetrachloride in rats

The present study evaluated the hepatoprotective effect of an N-acetyl dl-methionine + choline chloride + caffeine + thiamine hydrochloride + nicotinamide + pyridoxine hydrochloride compound at doses of 0.2, 0.6 and 1.0 mL/kg of b.w., and the assessment was done by the investigation of serum-enzymatic activity, metabolic functions of the liver and histophatological changes in female Wistar rats, which were subjected to experimental intoxication with CCl₄. One hundred and nineteen rats were randomly distributed into 17 groups, performing five different treatments, being evaluated seven animals per treatment in four periods: 2, 4, 6 and 8 days after CCl₄-induced intoxication. Treated rats with the hepatoprotective medicine (HM) presented a significant reduction in infiltration of inflammatory cells, steatosis, necrosis and liver congestion when compared to non-treated rats (control). Beside these results, the treatment showed a positive effect on circulatory alterations in the intoxicated animals, with reduction of spleen and renal congestion, as well as, promotion of a significant improvement in ALT, AST, LDH, ALP, GGT enzymatic serum activity reduction and in recovering liver function regarding the metabolism of urea, triglycerides and glucose. These findings indicate therapeutic usefulness of the compound when administered at dose 0.6 and 1.0 mL/kg of b.w. in female Wistar rats.     

Characterization of CD4(+) Cytotoxic Lymphocytes and Apoptosis Markers Induced by Trypanossoma cruzi Infection

Although the pathophysiology of Chagas disease is not completely understood, it is widely accepted that involvement of the immune response is critical in determining the outcome of the disease. In this context, CD4⁺ T cells may play an important role in generating different mechanisms of protection. In addition to effector and regulatory functions, CD4⁺ T cells may be also involved with lytic activities against the parasite and may have a relevant role on control of the infection. In this study, we have evaluated CD4⁺ T cells expressing cytotoxic and apoptosis markers in response to Trypanossoma cruzi infection in indeterminate (IND) and cardiac (CARD) patients with Chagas disease and non‐infected individuals (NI). Our data demonstrated that: (1) CD4⁺ T cells presented higher ex vivo granzyme B expression in patients with Chagas disease compared with healthy individuals and that antigen induced a greater granzyme B expression in IND patients; (2) CD95L expression in CD4⁺ CD95⁺ T cells from IND patients is higher than in CARD and NI; (3) IND and CARD patients had an increased frequency of caspase‐3 after in vitro stimulation and also expressed a high frequency of annexinV⁺ 7ADD⁺ within CD4⁺ T cells; (4) Lastly, a positive correlation was seen between cytotoxic molecules and CD45RO memory marker in CD4⁺ T cells and between caspase‐3 and CD95L within CD4⁺ CD95⁺ T cells. These results suggest new insights into the functional competence of CD4⁺ T cells among the different clinical forms of Chagas disease, which will lead to a better understanding of their influence during immune responses against T. cruzi.