Identification of Novel Immunoregulatory Molecules in Human Thymic Regulatory CD4+CD25+ T Cells by Phage Display

Thymic CD4+CD25+ cells play an important role in immune regulation and are continuously developed in the thymus as an independent lineage. How these cells are generated, what are their multiple pathways of suppressive activity and which are their specific markers are questions that remain unanswered. To identify molecules involved in the function and development of human CD4+CD25+ T regulatory cells we targeted thymic CD4+CD25+ cells by peptide phage display. A phage library containing random peptides was screened ex vivo for binding to human thymic CD4+CD25+ T cells. After four rounds of selection on CD4+CD25+ enriched populations of thymocytes, we sequenced several phage displayed peptides and selected one with identity to the Vitamin D Receptor (VDR). We confirmed the binding of the VDR phage to active Vitamin D in vitro, as well as the higher expression of VDR in CD4+CD25+ cells. We suggest that differential expression of VDR on natural Tregs may be related to the relevance of Vitamin D in function and ontogeny of these cells

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Abstract:  Current evidence on diet-cancer interaction remains incomplete, particularly in oral carcinoma (OC). This study aimed to evaluate the relationship among daily saturated, monounsaturated and polyunsaturated fatty acids (FAs) intake and eating patterns in people with / without OC, during a minimum period of five years, and its possible relationship with SNP TP53-R72P.  A retrospective case-control study (n = 79), matched by sex and age (21-85 years old), was carried out. Dietary information was collected using a quantitative food frequency survey validated for Cordoba population. TP53-R72P alleles were determined using conventional PCR. The Mann–Whitney U test and logistic regression were applied to assess the association between case/control status and FAs intake adjusted by alcohol/tobacco, respectively. Dietary patterns were identified by principal component factor analysis. The intake of saturated FAs such as myristic (p = 0.037), palmitic (p = 0.004), stearic (p = 0.009), arachidic (p = 0.012), monounsaturated such as palmitoleic (p = 0.013), oleic (p = 0.002) and polyunsaturated as linoleic (p = 0.0006) and alpha-linoleic (p = 0.003) was higher in cases. Omega6/omega3 ratio was significantly higher in cases (p = 0.003). The mutated C allele was more frequent in cases (p = 0.0061) than controls. All surveyed patients with OC were heterozygous for TP53-R72. In addition, they presented a high consumption of total fats in relation to controls. The studied population presented a dietary pattern described as “Western diet”;  related with a high daily intake of meat, eggs and alcohol; it was remarkable an association between red meat consumption and the presence of OC (p = 0.015). This diet has a high amount of omega-6 FAs, which leads to an increase in arachidonic acid and its derivatives, than have been related to the risk of OC. The presence of mutated TP53-R72P allele in OC patients is related to the loss of ability to induce apoptosis. The presence of both risk factors, diet and presence of a mutated polymorphic variant, could linked to an increaserisk of developing OC.Resumen:  La evidencia actual sobre la interacción dieta-cáncer sigue siendo incompleta, particularmente en el carcinoma oral (CO). El objetivo del estudio fue evaluar la relación entre la ingesta dietaria de ácidos grasos (AGs) saturados, monoinsaturados y poliinsaturados y patrones de alimentación en personas con/sin CO, durante un período mínimo previo de cinco años al momento de la encuesta; y su posible relación con la presencia del polimorfismo TP53-R72P. Se realizó un estudio retrospectivo de casos/controles (n=79), apareados por sexo y edad (21-85 años). Se recolectaron datos de un cuestionario cuantitativo de frecuencia alimentaria validado para la población cordobesa. TP53-R72P se genotipificó por PCR alelo específica. Se aplicaron la prueba Mann-Whitney y regresión logística para evaluar la asociación entre casos/controles y la ingesta de AGs ajustada por alcohol/tabaco, respectivamente. Los patrones alimentarios se identificaron por análisis factorial de componentes principales. La ingesta de AGs saturados como mirístico (p=0.037), palmítico (p=0.004), esteárico (p=0.009), araquídico (p=0.012), monoinsaturados como palmitoleico (p=0.013), oleico (p=0.002) y poliinsaturados como linoleico (p=0.0006) y alfa-linoleico (p=0.003) fue mayor en los casos. La relación omega-6/omega-3 fue significativamente mayor en los casos (p=0.003).  De los sujetos genotipificados, el alelo mutado C fue más frecuente en los casos (p=0.0061) que en los controles. De los sujetos encuestados y con genotipificación TP53-R72P se observó que todos los pacientes con CO eran heterocigotos y, además, presentaron un alto consumo de grasas en relación a los controles. La población total presentó un patrón alimentario reconocido como dieta occidental asociada a una ingesta diaria elevada en carnes, huevos y alcohol de la población estudiada; siendo notorio una asociación entre el consumo de carnes rojas y la presencia de CO (p=0.015). Esta dieta presenta una alta cantidad de AGs omega-6, lo que conduce a un aumento del ácido araquidónico y sus derivados, los cuales se han relacionado con el riesgo de CO. La presencia del alelo mutado TP53-R72P en pacientes con CO está relacionado con la pérdida de capacidad de conducir a la célula a apoptosis. La presencia de ambos factores de riesgo, dieta y presencia de variante polimórfica mutada, podría incrementar el riesgo de desarrollar CO.

Evidence of two viscous relaxation processes in the collective dynamics of liquid lithium

New inelastic X-ray scattering experiments have been performed on liquid lithium in a wide wavevector range. With respect to the previous measurements, the instrumental resolution, improved up to 1.5 meV, allows to accurately investigate the dynamical processes determining the observed shape of the the dynamic structure factor, $S(Q,\omega)$. A detailed analysis of the lineshapes shows the co-existence of relaxation processes with both a slow and a fast characteristic timescales, and therefore that pictures of the relaxation mechanisms based on a simple viscoelastic model must be abandoned.Comment: 5 pages, 4 .PS figure

Influence of photon-Induced photoacoustic streaming (PIPS) on root canal disinfection and post-Operative pain: a randomized clinical trial

The aim of this study was to evaluate the ability of a PIPS (photon-induced photoacoustic streaming) Er:YAG laser to reduce the root canal system bacterial count in vivo in comparison to the traditional irrigation technique. The post-operative patients’ quality of life (QoL) after endodontic therapy was evaluated through a questionnaire. Fifty-four patients affected by pulp necrosis with or without apical periodontitis biofilm disease were selected for endodontic treatment and randomly assigned to Group A (n = 27) with traditional irrigation and Group B (n = 27), with PIPS irrigation applied according to the protocol. Shaping was performed with ProGlider and ProTaper Next, and irrigation was performed with 5% NaOCl and 10% EDTA. Intracanal samples for culture tests were collected before and after irrigation. The microbiological analysis was evaluated by the Kolmogorov–Smirnov normality and Mann–Whitney tests (p < 0.05). A self-assessment questionnaire was used to evaluate the QoL during the 7 days after treatment; differences were analysed with Student’s t-test. Irrigation with the PIPS device was significantly effective in reducing bacterial counts, which were higher for facultative than obligate anaerobic strains, particularly for Gram-negative bacteria, without statistical significance (p > 0.05). There were no significant differences among the QoL indicators, except for the maximum pain (p = 0.02), eating difficulty (p = 0.03) and difficulty performing daily functions (p = 0.02) in the first few days post-treatment. PIPS may represent an aid to root canal disinfection not affecting the patients’ QoL, particularly for the first day after treatment

Evaluation of the bactericidal activity of a hyaluronic acid-vehicled clarithromycin antibiotic mixture by confocal laser scanning microscopy

Confocal laser scanning microscopy (CLSM) was used to evaluate the antibacterial effect and depth of action of a novel clarithromycin-containing triple antibiotic mixture, which was proposed for root canal disinfection in dental pulp regeneration. A previous study reported that this mixture had no tooth discoloration effects in vitro. After infection with Enterococcus faecalis for 3 weeks, the dentinal tubules in the cylindrical root specimens were exposed to different antibiotic mixtures: ciprofloxacin, metronidazole and minocycline (3-MIX); ciprofloxacin, metronidazole and clarithromycin (3-MIXC) and ciprofloxacin and metronidazole (2-MIX). Each antibiotic formulation was mixed with macrogol (MG) or hyaluronic acid (HA) vehicles. CLSM and viability staining were used to quantitatively analyze the mean depth of the antibacterial effect and the proportions of dead and live bacteria inside the dentinal tubules. The 3-MIX and 3-MIXC demonstrated a similar depth of action. The mean proportion of dead bacteria was similar in the 3-MIX and 3-MIXC groups, and both were statistically higher than that of 2-MIX (p = 0.014). Each antibiotic mixture showed a higher bactericidal efficacy if conveyed with HA, compared to MG (3-MIX, p = 0.019; 3-MIXC, p = 0.013 and 2-MIX, p = 0.0125). The depth of action and the antibacterial efficacy of 3-MIXC seemed comparable with 3-MIX