Month: June 2020

In the present study, we did not find an increase of TBARS levels in whole saliva in response to an acute psychosocial stressor that might indicate resilience to lipid peroxidation in saliva of young people at stress, possibly because of the enhanced activity of catalase. No difference in TBARS levels was observed between men and women at rest as well as at stress. The side chains of all amino acid residues of proteins are susceptible to oxidation by ROS. Excessive oxidative stress results in carbonylation of proteins that is an irreversible oxidative modification often leading to decrease or loss of protein function. The concentration of carbonyl GDC-0879 groups is a good measure of ROS-mediated protein oxidation. Unexpectedly, in the present study we found that levels of oxidatively modified proteins responded to stress with a substantial decrease that was larger in women than in men. The decrease might be explained by augmented catalase activity and is consistent with the observation that stress caused a greater rise in catalase in women compared to men. In the present study we investigated how psychosocial stress effects levels of free sialic acids in whole saliva. Sialic acids are believed to be important in antioxidant defense. They occupy the terminal position of many glycoproteins, e.g. mucins and have important roles in their functioning. Mucins have been found to be hydroxyl radical scavengers, with sialic acid moieties essential for the role. However, excessive quantities of hydroxyl radicals cause depolimerization of native mucin, while Eguchi et al. showed that the glycosidic linkage of sialic acid is a potential target for superoxide and other related ROS that therefore cause liberation of free sialic acids. Iijima et al. showed that free Nacetylneuraminic acid scavenges hydrogen peroxide under physiological conditions and is a potent defense molecule against oxidative stress. We have found a significant stress-induced increase of free sialic acids in whole saliva of young people at stress. Interestingly, baseline levels of free sialic acids were significantly lower in women than in men. An exposure to a psychosocial stressor caused a sharper increase of sialic acids in saliva of women and resulted in similar concentration of sialic acids in both sexes. The mechanism and the role of the observed stress-induced rise of free sialic acids in saliva is unclear. The rise might be attributed to the intensified damage of mucins at stress. Alternatively, the phenomenon might indicate an additional protective mechanism against oxidative lesions. We tested effects of psychosocial stress on salivary flow rate and total protein concentration to eliminate the possibility that they are confounding factors in the research. Up to now research of effects of stress on salivary flow rate or on total protein concentration in saliva has produced contradictory results. A decreased flow rate was found in an academic stress paradigm.

The successful introduction of anti-angiogenic therapies into clinical trials requires the development of reliable non-invasive methods for assessing angiogenesis and its modulation or inhibition in-vivo. Thus, in the last few years, a broad range of MRI techniques have been developed to provide feedback and surrogate markers for therapeutic response including tumor blood volume, perfusion, vessel permeability, oxygenation and vessel size. These methods, aimed at the early detection of vascular changes in response to therapy, may guide patient management based on the individual response pattern. Contrast enhanced – MRI is widely established and currently is the preferred method for brain tumor assessment. However, CE-MRI does not adequately assess LEE011 disease status especially during Bevacizumab therapy for recurrent glioblastoma since recurrence is commonly associated with nonenhancement on CE-MRI. Blood oxygenation level-dependent MRI uses the paramagnetic nature of deoxygenated hemoglobin versus the diamagnetic nature oxygenated hemoglobin. Using this method, hemoglobin can serve as an endogenous contrast agent which indirectly represents changes in blood flow, volume and oxygenation. BOLD MRI is the basis for the well-established functional MRI method, in which hemodynamic changes due to neuronal activation are monitored. Changes in BOLD signal can also occur due to respiratory challenges of hyperoxia or hypercapnia. Pure oxygen inhalation causes increased blood oxygenation and reduced blood flow, while inhalation of a mixture of oxygen and CO2 has been shown to increased blood oxygenation and flow. Typically, when respiratory challenges are viewed, a T2* sequence is used. Previously, we utilized Hemodynamic Response Imaging, an fMRI method combined with hypercapnic and hyperoxic challenges for functional analysis of vessel maturation, and vessel density and functionality. Using changes in the BOLD signal induced by respiratory challenges is not limited to brain imaging and can be used as a tool for assessing the hemodynamic response in different organs and pathologies. Several animal studies were performed to estimate treatment response in liver metastases ; classify liver fibrosis ; evaluate renal perfusion and hemodynamics during acute kidney injury ; and study tumor vasculature. Also, several studies utilized rodent brain models in order to distinguish neural from non-neural contributions to fMRI signals and evaluated BOLD signal changes due to hyperoxia and hypercapnia. Recently, our group reported a clinical study demonstrating the use of HRI for the assessment of angiogenesis in brain tumors both in untreated tumors and in tumors treated with chemotherapy and radiation therapy. The aim of the current study focused on the development of equivalent mouse model, which will enable us to perform more fundamental research in order to achieve better understanding of the involved cellular processes and to elucidate the imaging characteristics. In this context an orthotopic mouse model for brain glioblastoma tumors was established. In this model, the effect of an anti-angiogenic treatment with B20-4.1.1 was evaluated.

Many genetic alterations have been reported at this locus including HhAntag691 structure deletions, duplications, nonsense, and missense mutations. Genotype/phenotype correlations have been described, with some variation. In general, mutations that abolish enzyme function are associated with a relatively uniform clinical and biochemical presentation. Animal models of OTCD that have been characterized and are readily available include the spf mouse and the spf-ash mouse. These models are currently maintained on a mixed background, B6EiC3Sn, which may limit certain issues of experimental design. Herein, we describe a new spontaneous hypomorphic missense mutation in the C57BL/6J-Otcspf-J/J that produces a new model of mild OTCD. Spf-J mice display normal plasma ammonia and plasma orotic acid at baseline and milder amino acid perturbations when compared to their predecessors. Despite this mild plasma biochemical phenotype, cerebral amino acid concentrations were elevated at baseline and, unlike WT, were depleted during a systemic immune response suggesting altered cerebral amino acid metabolism or transport. Overall, this mouse model is not only useful for further characterization of functional domains of the OTC enzyme, but is also useful for investigating the pathophysiology of cerebral amino acid metabolism in UCD. Due to the inheritance of OTCD, the phenotypic expression of the disease severity in patients is dependent upon the nature of the mutation, genetic background and in females, Xinactivation in the liver. In males, this disorder classically manifests with symptomatic hyperammonemia in infancy, although milder alleles have been described. In addition, phenotypic variability can be seen in males within the same family with the same mutation suggesting modifier alleles may play a role. Late-onset symptoms of OTCD, due to milder enzyme defects, may manifest in men as late as the fifth or sixth decade given the right precipitant, such as the Atkins’ diet. Galloway et al. described a previously healthy male who experienced his first documented hyperammonemic episode at 13 years of age. The precipitant of this episode was unknown. On admission to hospital, his ammonia was 750 μmol/L and hemodialysis was initiated. Urine orotate was 813 μmol/mmol creatinine, suggesting a diagnosis of OTCD. A subsequent liver biopsy demonstrated OTC enzyme activity to be 11% of normal. Molecular investigation revealed a lysine substitution for asparagine in the OTC protein. This amino acid substitution in the OTC enzyme is identical to the mutation described here in the spf-J mouse. Similar to the reported patient, the K80N mutation results in a reduction in enzyme activity to *11–12% in spf-J. The mutation lies outside the substrate binding and catalytic regions of the enzyme and may be related to homo-oligomerization or stability of the enzyme. In our studies, we demonstrated normal levels of OTC mRNA and OTC protein that was below the limit of detection by immunoblot. These findings lead us to suggest that the K80N mutation may destabilize the enzyme, leading to its early degradation.

In CF patients, S. aureus predominates in the lung of children and teenagers, while P. aeruginosa prevails in adults. Exopolysaccharide-enriched biofilms produced by P. aeruginosa increase the mucus viscosity, resistance to antibiotics and host immune effectors. Chronic bacterial infections are common in CF patients and facilitates lung inflammation, mucous obstruction and tissue remodeling, resulting in fatal loss of function. CF lungs display excessive inflammatory response, especially with increased neutrophil recruitment, the mechanism of this phenomenon is not adequately explained. However, intervention in this process likely will benefit CF patients. The role of the pro-inflammatory signaling cytokine Interleukin 1b in CF lung disease has been reported before. P. aeruginosa induces IL-1b or IL-18 production through NLRC4 inflammasome activation. P. aeruginosa flagellin and highly acylated LPS is recognized by TLR5 and TLR4 respectively. Human polymorphisms observed in the IL1B gene were associated with CF disease. CFTR deficient mice were found to be more susceptible to acute and chronic P. aeruginosa infection and display an exacerbated inflammatory response to LPS and P. aeruginosa activated alveolar macrophages from F508del mutant mice have enhanced expression of IL-1b. Huaux et al recently showed a deregulated inflammatory and fibrotic response in F508del mutant mice to bleomycin, which is IL-1R1 signaling dependent. Here we revisited the role of IL-1b in the resolution of P. aeruginosa infection, in a murine model based on mice carrying the most common CF mutation F508del CFTR. In this study, we show that excessive activation of IL-1b correlates with increased bacterial load, Epoxomicin inflammation and lung damage in F508del CFTR mice. Further, we show that IL-1b antibody neutralization attenuates the inflammatory response to P. aeruginosa infection. CF patients display increased susceptibility to chronic infections with opportunistic bacteria, excessive lung inflammation and fibrosis leading to fatal loss of function eventually. P. aeruginosa has been shown to induce IL-1b through NLRC4 inflammasome activation. CFTR deficient mice were found to be more susceptible to P. aeruginosa infection and have an exacerbated inflammatory response to LPS and P. aeruginosa. Furthermore, F508del CFTR mutant alveolar macrophages display an enhanced IL-1b production in response to LPS stimulation. Thus, we were interested in clarifying the role of IL-1b in the resolution of P. aeruginosa infection, in a murine model with the most common CF mutation F508del CFTR. Here, we show that, after P. aeruginosa infection, excessive activation of IL-1b in F508del CFTR mice compared to WT was accompanied by increased CFU in BALF, inflammation and lung damage. Further we show that a therapeutic antibody administration attenuates inflammatory response in an acute model of infection using WT mice. In CF patients bacterial infections persist and their chronicity facilitates lung inflammation, and subsequent tissue remodeling with severe loss of function.

Though, no evidence was reported regarding their role in oral cancer. Perlecan is a large proteoglycan harboring five distinct structural domains, to which long chains of heparan sulfate and/or WZ4002 chondroitin sulfate are attached. This molecule is present in all vascularized tissues with a distribution that is primarily confined to basement membranes. Also, other studies have also identified perlecan in the stromal spaces of various pathophysiological conditions. Agrin shares a rather intriguing multimodular organization with perlecan, but more complexity to agrin can be added due to at least four sites of alternative splicing. The amino acid sequence of agrin encodes a protein with a molecular size of 220 kDa, but the observed molecular weight is around 400 kDa due to the long heparan sulfate and chondroitin sulfate glycosaminoglycans attached to the core protein. Although originally discovered in the neuromuscular junctions, agrin has been observed in numerous other tissues, and it is described as highly expressed in hepatocellular carcinomas and cholangiocellular carcinomas. Nevertheless, little is known about its role at locations other than the neuromuscular junctions, and even less information is known about its role in tumor tissues. In the present study, we focused on understanding the role of the proteoglycans agrin and perlecan in oral cancer. First, we sought to validated the overexpression of agrin and perlecan in oral cancer tissues compared to normal tissues and in cell lines with different site of origin: oral squamous carcinoma originated from human tongue, oral squamous carcinoma SCC-9 isolated from lymph nodes and a skin-derived squamous carcinoma. Next, we showed that oral squamous carcinoma cell line had a reduced ability to adhere to extracellular matrix proteins and increased sensibility to cisplatin when treated with chondroitinase. By specific target agrin and perlecan protein levels with siRNA, we showed that OSCC cells have decreased cell adhesion and migration and increased sensibility to cisplatin treatment. Overall, our findings opened new avenues to better understand the role of agrin and perlecan, as well as their involvement in carcinogenesis, which may offer a novel approach to cancer therapy by targeting the tumor microenvironment. Proteoglycans, essential macromolecules of the tumor microenvironment, have their expression altered during malignant transformation and tumor progression. Agrin and perlecan are two of the major HSPG identified in the basement membrane, and their functional roles in modulation of cancer growth have been reviewed elsewhere. In this study, we showed for the first time that agrin and perlecan are highly expressed in OSCCs, and the function of these proteins in oral cancer associated processes was investigated. Not only the expression of agrin and perlecan was shown to be higher in OSCC tissues compared to control tissues, but also their expression might be associated with different sites of origin, where higher expression of agrin was found in cell line originated from primary site.