T cell apoptosis and induction of foxp3+ regulatory T cells underlie the therapeutic efficacy of CD4 blockade in experimental autoimmune encephalomyelitis

The pathogenesis of multiple sclerosis requires the participation of effector neuroantigen-specific T cells. Thus, T cell targeting has been proposed as a promising therapeutic strategy. However, the mechanism underlying effective disease prevention following T cell targeting remains incompletely known. We found, using several TCR-transgenic strains, that CD4 blockade is effective in preventing experimental autoimmune encephalopathy and in treating mice after the disease onset. The mechanism does not rely on direct T cell depletion, but the anti-CD4 mAb prevents the proliferation of naive neuroantigen-specific T cells, as well as acquisition of effector Th1 and Th17 phenotypes. Simultaneously, the mAb favors peripheral conversion of Foxp3(+) regulatory T cells. Pre-existing effector cells, or neuroantigen-specific cells that undergo cell division despite the presence of anti-CD4, are committed to apoptosis. Therefore, protection from experimental autoimmune encephalopathy relies on a combination of dominant mechanisms grounded on regulatory T cell induction and recessive mechanisms based on apoptosis of neuropathogenic cells. We anticipate that the same mechanisms may be implicated in other T cell-mediated autoimmune diseases that can be treated or prevented with Abs targeting T cell molecules, such as CD4 or CD3.     

Cutting edge: adaptive versus innate receptor signals selectively control the pool sizes of murine IFN-γ- or IL-17-producing γδ T cells upon infection

γδ T lymphocytes are commonly viewed as embracing properties of both adaptive and innate immunity. Contributing to this is their responsiveness to pathogen products, either with or without the involvement of the TCR and its coreceptors. This study clarifies this paradoxical behavior by showing that these two modes of responsiveness are the properties of two discrete sets of murine lymphoid γδ T cells. Thus, MyD88 deficiency severely impaired the response to malaria infection of CD27((-)), IL-17A-producing γδ T cells, but not of IFN-γ-producing γδ cells. Instead, the latter compartment was severely contracted by ablating CD27, which synergizes with TCRγδ in the induction of antiapoptotic mediators and cell cycle-promoting genes in CD27((+)), IFN-γ-secreting γδ T cells. Hence, innate versus adaptive receptors differentially control the peripheral pool sizes of discrete proinflammatory γδ T cell subsets during immune responses to infection.     

Profiles of Steroid Hormones in Canine X-Linked Muscular Dystrophy via Stable Isotope Dilution LC-MS/MS

Golden retriever muscular dystrophy (GRMD) provides the best animal model for characterizing the disease progress of the human disorder, Duchenne muscular dystrophy (DMD). The purpose of this study was to determine steroid hormone concentration profiles in healthy golden retriever dogs (control group - CtGR) versus GRMD-gene carrier (CaGR) and affected female dogs (AfCR). Therefore, a sensitive and specific analytical method was developed and validated to determine the estradiol, progesterone, cortisol, and testosterone levels in the canine serum by isotope dilution liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). To more accurately understand the dynamic nature of the serum steroid profile, the fluctuating levels of these four steroid hormones over the estrous cycle were compared across the three experimental groups using a multivariate statistical analysis. The concentration profiles of estradiol, cortisol, progesterone, and testosterone revealed a characteristic pattern for each studied group at each specific estrous phase. Additionally, several important changes in the serum concentrations of cortisol and estradiol in the CaGR and AfCR groups seem to be correlated with the status and progression of the muscular dystrophy. A comprehensive and quantitative monitoring of steroid profiles throughout the estrous cycle of normal and GRMD dogs were achieved. Significant differences in these profiles were observed between GRMD and healthy animals, most notably for estradiol. These findings contribute to a better understanding of both dog reproduction and the muscular dystrophy pathology. Our data open new venues for hormonal behavior studies in dystrophinopathies and that may affect the quality of life of DMD patients.     

Establishment of Murine Gammaherpesvirus Latency in B Cells Is Not a Stochastic Event

Murid γ-herpesvirus-4 (MuHV-4) promotes polyclonal B cell activation and establishes latency in memory B cells via unclear mechanisms. We aimed at exploring whether B cell receptor specificity plays a role in B cell susceptibility to viral latency and how this is related to B cell activation. We first observed that MuHV-4-specific B cells represent a minority of the latent population, and to better understand the influence of the virus on non-MuHV-4 specific B cells we used the SW_HEL mouse model, which produce hen egg lysozyme (HEL)-specific B cells. By tracking HEL⁺ and HEL⁻ B cells, we showed that in vivo latency was restricted to HEL⁻ B cells while the two populations were equally sensitive to the virus in vitro. Moreover, MuHV-4 induced two waves of B cell activation. While the first wave was characterized by a general B cell activation, as shown by HEL⁺ and HEL⁻ B cells expansion and upregulation of CD69 expression, the second wave was restricted to the HEL⁻ population, which acquired germinal center (GC) and plasma cell phenotypes. Antigenic stimulation of HEL⁺ B cells led to the development of HEL⁺ GC B cells where latent infection remained undetectable, indicating that MuHV-4 does not benefit from acute B cell responses to establish latency in non-virus specific B cells but relies on other mechanisms of the humoral response. These data support a model in which the establishment of latency in B cells by γ-herpesviruses is not stochastic in terms of BCR specificity and is tightly linked to the formation of GCs.     

Overexpression of Escherichia coli nucleotide excision repair genes after cisplatin-induced damage

Cisplatin is currently used in tumor chemotherapy to induce the death of malignant cells through blockage of DNA replication. It is a commonly used chemotherapeutic agent binding mono- or bifunctionally to guanines in DNA. Escherichia coli K12 mutant strains deficient in nucleotide excision repair (NER) were submitted to increasing concentrations of cisplatin, and the results revealed that uvrA and uvrB mutants are sensitive to this agent, while uvrC and cho mutants remain as the wild type strain. The time required for both gene expression turn-off and return to normal weight DNA in wild-type E. coli was not accomplished even after 4 h post-treatment with cisplatin, while the same process takes place within 1.5 h after ultraviolet radiation (UV). Besides, a heavily damaging action of cisplatin can be seen not only by persistent nicks on genomic DNA, but also by NER gene expression exceeding manifold that seen after equivalent lethal doses of UV. Moreover, cisplatin caused an increase in uvrB gene expression from its putative upstream promoter P3 in an SOS-independent manner.     

Effect of commercial herbal toothpastes and mouth rinses on the prevention of enamel demineralization using a microcosm biofilm model

Abstract

This work evaluated the effects of commercial toothpastes and mouth rinses containing natural/herbal agents on biofilm viability, extracellular polysaccharide (EPS) production and on enamel demineralization in vitro. Microcosm biofilm was produced on bovine enamel for 5?days and treated daily with: Orgânico natural^® (toothpaste/mouth rinse), Boni Natural Menta &; Malaleuca^® (toothpaste/mouth rinse), Propolis &; Myrrh^® (toothpaste), Colgate Total 12 Clean Mint^® (toothpaste, positive control), Malvatricin^® Plus (mouth rinse), PerioGard^® (mouth rinse, positive control) or PBS (negative control). Tom’s Propolis &; Myrrh^® and Colgate Total 12^® toothpastes and Malvatricin^® Plus and PerioGard^® mouth rinses significantly reduced biofilm viability (p?® had significant effects on biofilm thickness and EPS. Despite the indication that Tom’s Propolis &; Myrrh^® significantly reduced lesion depth, only Colgate Total 12^® significantly reduced mineral loss. Malvatricin^® Plus significantly reduced mineral loss and lesion depth, as did PerioGard^®. Some herbal products, Malvatricin^® Plus and Tom’s Propolis &; Myrrh^®, showed anticaries effects.     

Synthesis of potentially bioactive PABA-related <Emphasis Type="Italic">N</Emphasis>-(aminoalkyl)lactamic amino acids and esters via selective S<Subscript>N</Subscript>Ar reactions

Abstract  

Potentially bioactive N-(aminoalkyl)lactamic amino acids and esters were synthesized in satisfactory to good yields by S_NAr reactions of aromatic acids with N-(3-aminopropyl)lactams followed by esterification with tertiary amino alcohols. The addition–elimination S_NAr mechanism was confirmed by NMR and MS measurements.     

Structure of a heteroxylan of gum exudate of the palm Scheelea phalerata (uricuri)

The polysaccharide isolated from the gum exudate of palm Scheelea phalerata (SPN) was water-insoluble and composed of Fuc, Ara, Xyl, and uronic acid moieties in a 5:34:54:7 molar ratio: 12% of phenolics were also present. A soluble polysaccharide (SPNa) was obtained after alkaline treatment, which contained Fuc, Ara, Xyl and uronic acid in a 7:44:42:7 molar ratio, with only 2% phenolics. SPNa had an M(W) approximately 1.04 x 10(5) g mol(-1) and was almost monodisperse (M(W)/M(N) : 1.25 +/-0.22). It had a branched structure with side chains of 2-O-substituted Xylp (approximately 8%) and 3-O-substituted Araf (12%) units, and a large proportion of nonreducing end-units of Araf (15%), Fucp (10%), Xylp (4%), and Arap (6%). The (1 --> 4)-linked beta-Xylp main-chain units were 3-O- (9%), 2-O- (13%), and 2,3-di-O- (13%) substituted. Its (13)C NMR spectrum contained at least 9 C-1 signals, those at delta 108.6 and 107.7 arising from alpha-Araf units. Others were present at delta 175.4 from C-6 of alpha-GlcpA and delta 15.6 from C-6 of Fucp units. The main chain of SPNa was confirmed by analysis of a Smith-degraded polysaccharide (SPDS): methylation analysis provided a 2,3-Me(2)-Xyl (65%) derivative and its (13)C NMR spectrum showed five main signals typical of a (1 --> 4)-linked beta-Xylp units. Methylation analysis of a carboxy-reduced polysaccharide (SPN-CR) revealed a 2,3,4,6-Me(4)-Glc derivative (4%) arising from nonreducing end-units of GlcpA. Alpha-GlcpA-(1 --> 2)-alphabeta-Xy1p and alpha-GlcpA-(1 --> 2)-beta-Xylp-(1 --> 4)-alphabeta-Xylp were obtained via partial acid hydrolysis of SPN, showing the structure of side-chain substituents on O-2 of the main-chain units.