Expansion of the CD8 memory T cells : implications for the self-renewal gene Hoxb4

Les cellules CD8 T mémoire (Tm) offrent une protection tout au long de la vie contre les infections récurrentes. Elles sont maintenues grâce à des mécanismes d'auto-renouvellement. Les cellules souches hématopoïétiques (CSH) peuvent aussi s'auto-renouveller lentement, en assurant leur maintenance à long terme. Les deux types de cellules utilisent la moelle osseuse comme principale niche de prolifération. CD8 Tm et CSH partagent partiellement un profil transcriptionnel, y compris certains gènes connus pour contrôler l'auto-renouvellement. Les gènes Hox, dont Hoxb4 qui est un activateur puissant de l’expansion des CSH in vitro et in vivo, sont exprimés par les CSH. Basé sur les similitudes entre les CSH et les cellules Tm, nous émettons l'hypothèse que les gènes impliqués dans l’auto-renouvellement des CSH, comme Hoxb4, favoriseront l'expansion des cellules CD8 Tm. Pour tester cette hypothèse, nous avons déterminé l'effet de la surexpression de Hoxb4 dans les cellules T à partir de souris transgéniques jeunes et âgées, et sur la prise de greffe et le maintien des cellules CD8 Tm après leur transplantation dans des souris immunocompétentes ou immunosupprimées. À l'état basal, la composition et le phénotype des cellules T naïves et mémoire n’ont pas été affectés par la surexpression de Hoxb4. Pour étudier le maintien des cellules CD8 Tm, les souris transgéniques Hoxb4 et OT-1 ont été croisées. Les cellules T des souris OT-1, entre autre, expriment un récepteur de lymphocyte T qui reconnaît le peptide d'ovalbumine (OVA), présenté par le complexe majeur d’histocompatibilité de classe I. En utilisant ce modèle, nous avons généré in vitro un grand nombre de cellules CD8 Tm pour les expériences du transfert adoptif. Ces cellules sont monoclonales et spécifiques pour la reconnaissance d’OVA. Le nombre et le phénotype des cellules CD8 Tm générées en culture n’ont pas été affectés par la surexpression de Hoxb4. Ces cellules ont été transplantées dans des souris de type sauvage afin d'évaluer leur prise de greffe et leur maintien à long terme. Après la transplantation chez des souris soit immunocompétentes ou immunosupprimées, la prise de greffe et le maintien des cellules CD8 Tm surexprimant Hoxb4 n'a pas été améliorée lorsque comparée aux cellules CD8 Tm de type sauvage. Ces résultats suggèrent que Hoxb4 ne favorise pas l’auto-renouvellement des cellules CD8 Tm. D'autres études vont essayer d'identifier les facteurs contrôlant l’auto-renouvellement des cellules CD8 Tm.CD8 memory T (Tm) cells provide life long protection against recurrent infections and are maintained through self-renewal mechanisms. Hematopoietic stem cells (HSC) also possess the capacity to slowly self-renew assuring their long-term maintenance. Both cell types use the bone marrow as their preferred proliferation niche. CD8 Tm and HSC partially share a transcriptional profile including some genes known to control self-renewal. Hox genes are expressed in HSCs, among them Hoxb4 is a potent enhancer of HSC expansion in vitro and in vivo. Based on the similarities between HSC and Tm cells, we hypothesize that genes involved in HSC self-renewal, like Hoxb4, will promote CD8 Tm cell expansion. To test this hypothesis, we have determined the effect of Hoxb4 overexpression in T cells from young and old transgenic mice, and on the engraftment and maintenance of CD8 Tm cells after transplantation into immunocompetent or immunodeficient mice. At the steady-state, the composition and phenotype of the naïve and memory T cells were not affected by Hoxb4 overexpression. To study CD8 Tm cell maintenance, the Hoxb4 and the OT-1 transgenic mice were crossed; the latter express a T cell receptor that recognizes the ovalbumin (OVA) peptide, presented by major histocompatibility complex class I. Using this model, we generated in vitro large numbers of monoclonal and OVA specific CD8 Tm cells for adoptive transfer experiments. The number and the phenotype of Tm cells generated in culture were not affected by Hoxb4 overexpression. These cells were transplanted into wild type mice to evaluate their engraftment and long-term maintenance. After transplantation in either immunocompetent or immunodeficient mice, Hoxb4 overexpressing Tm cell engraftment or maintenance was not enhanced when compare to wild-type Tm cells. These results demonstrate that Hoxb4 does not promote CD8 Tm cell self-renewal. Further studies will try to identify the factors controlling CD8 Tm cell self-renewal

vacA genotypes in oral cavity and Helicobacter pylori seropositivity among adults without dyspepsia

Objective: The aims of this research were to determine the prevalence of Helicobacter pylori and its vacA genotypes in oral cavity in persons without dyspepsia and to establish the association between the presence of H. pylori in oral cavity and oral hygiene. The seroprevalence of anti-H. pylori antibodies and its associated factors were analyzed too. Study design: For the study, 200 adults without dyspepsia symptoms were selected. Dental plaque and saliva samples from each subject were obtained. H. pylori detection in oral samples was carried out by polymerase chain reaction (PCR) and for vacA genotyping a semi-nested and nested PCR was used. The enzyme-linked immunosorbent assay (ELISA) was used to detect anti-H. pylori IgG and IgM. The data were analyzed with Chi square and Fisher exact test and the statistical significance was set to 0.05. Results: Of 200 subjects tested, 124 (62%) were seropositive. H. pylori was detected in the oral cavity of 34 subjects (17%) and vacA allelotypes were typified in 12 of those samples. The s1 allele was detected in 8 (66.7%) samples and in one of them m1 and m2 alleles were found. In four subjects vacA m1 subtypes were found and in two of those both m1 and m2 alleles were detected. The prevalence of H. pylori in oral cavity was higher (l8.5%) among seropositive subjects compared with seronegative persons. No association was found between the presence of H. pylori and oral hygiene habits. Conclusions: The presence of H. pylori in oral cavity is more frequent in seropositive subjects without dyspepsia symptoms and could represent the source of gastric infection and bacterial transmission. The data suggest that more than one H. pylori strain may exist in the mouth of asymptomatic persons

Experimental arthritis induced by a clinical Mycoplasma fermentans isolate

BACKGROUND: Mycoplasma fermentans has been associated with rheumatoid arthritis. Recently, it was detected in the joints and blood of patients with rheumatoid arthritis, but it is not clear yet how the bacteria enter the body and reach the joints. The purpose of this study was to determine the ability of M. fermentans to induce experimental arthritis in rabbits following inoculation of the bacteria in the trachea and knee joints. METHODS: P-140 and PG-18 strains were each injected in the knee joints of 14 rabbits in order to evaluate and compare their arthritogenicity. P-140 was also injected in the trachea of 14 rabbits in order to test the ability of the bacteria to reach the joints and induce arthritis. RESULTS: M. fermentans produced an acute arthritis in rabbits. Joint swelling appeared first in rabbits injected with P-140, which caused a more severe arthritis than PG-18. Both strains were able to migrate to the uninoculated knee joints and they were detected viable in the joints all along the duration of the experiment. Changes in the synovial tissue were more severe by the end of the experiment and characterized by the infiltration of neutrophils and substitution of adipose tissue by connective tissue. Rabbits intracheally injected with P-140 showed induced arthritis and the bacteria could be isolated from lungs, blood, heart, kidney, spleen, brain and joints. CONCLUSION: M. fermentans induced arthritis regardless of the inoculation route. These findings may help explain why mycoplasmas are commonly isolated from the joints of rheumatic patients

Receptores noradrenérgicos en las células estelares hepáticas

Las células estelares hepáticas (HSC) juegan un papel muy importante en el desarrollo del daño y la regeneración del hígado. El entendimiento de su activación por causa de un daño hepático resulta de gran importancia para poder tener nuevas estrategias terapéuticas. Para ello un modelo comúnmente utilizado son las células estelares hepáticas de rata. Se tienen líneas celulares obtenidas de hígados cirróticos de rata que presentan características bien definidas, y con frecuencia se utiliza el cultivo primario, ya que las células durante el cultivo presentan las características de células activadas de manera espontánea. Actualmente existen algunos trabajos que plantean que el sistema nervioso está jugando un papel importante en los procesos de daño y regeneración hepática. Por ello consideramos que es relevante saber si en el modelo de células estelares de rata, éstas son capaces de responder a los estímulos del sistema nervioso. Siendo la norepinefrina (NE) uno de los principales neurotransmisores, además de que se ha encontrado elevada en pacientes con cirrosis hepática, decidimos estudiar si las células pueden responder a este neurotransmisor. El objetivo del trabajo fue determinar si las HSC de cultivo primario de rata y de la línea celular CFSC2G eran capaces de reconocer el estímulo de la NE. Las HSC fueron cultivadas y tratadas con Fura-2AM para poder observar el calcio intracelular y con isoproterenol 10µM por 5 min para realizar la medición de AMPc. En experimentos realizados para medir la liberación de calcio intracelular ante el estímulo de la NE encontramos que las células estelares activadas en cultivo primario y de la línea celular CFSC2G no presentan una respuesta a NE, mientras que en la medición de la producción de AMPc sí encontramos una respuesta estadísticamente significativa. Por ello concluimos que la respuesta a NE en las HSC de cultivo primario de rata y de la línea celular CFSC2G es mediada por los receptores b adrenérgicos, con una producción de AMPc como segundo mensajero

Hoxb4 overexpression in CD4 memory phenotype T cells increases the central memory population upon homeostatic proliferation.

Memory T cell populations allow a rapid immune response to pathogens that have been previously encountered and thus form the basis of success in vaccinations. However, the molecular pathways underlying the development and maintenance of these cells are only starting to be unveiled. Memory T cells have the capacity to self renew as do hematopoietic stem cells, and overlapping gene expression profiles suggested that these cells might use the same self-renewal pathways. The transcription factor Hoxb4 has been shown to promote self-renewal divisions of hematopoietic stem cells resulting in an expansion of these cells. In this study we investigated whether overexpression of Hoxb4 could provide an advantage to CD4 memory phenotype T cells in engrafting the niche of T cell deficient mice following adoptive transfer. Competitive transplantation experiments demonstrated that CD4 memory phenotype T cells derived from mice transgenic for Hoxb4 contributed overall less to the repopulation of the lymphoid organs than wild type CD4 memory phenotype T cells after two months. These proportions were relatively maintained following serial transplantation in secondary and tertiary mice. Interestingly, a significantly higher percentage of the Hoxb4 CD4 memory phenotype T cell population expressed the CD62L and Ly6C surface markers, characteristic for central memory T cells, after homeostatic proliferation. Thus Hoxb4 favours the maintenance and increase of the CD4 central memory phenotype T cell population. These cells are more stem cell like and might eventually lead to an advantage of Hoxb4 T cells after subjecting the cells to additional rounds of proliferation

Change of naïve and MP T cell populations in <i>Hoxb4</i> transgenic and wt mice with age.

<p>Scatter plots showing the percentage of (<b>A</b>) CD4 and (<b>B</b>) CD8 T cells that are naïve (CD44^lo/CD62L^hi), MP (CD44^hi) or are a subpopulation of MP T cells (CD44^hi/Ly6C^hi) for LN, Spl and BM derived from individual <i>Hoxb4</i> transgenic and wt (n = 6–8) age matched mice. Young mice are between 3–4 months and old mice are all older than 28 months. Naïve and MP populations change significantly with age, but not between <i>Hoxb4</i> and wt mice. *P<0.05, 2-tailed Student ttest. MP = memory phenotype, Wt = wild type, LN = lymph node, Spl = spleen and BM = bone marrow.</p

Percentage of T cell populations in hematopoietic organs of young adult mice.

<p>Note that no significant differences were observed between T cell populations of <i>Hoxb4</i> and wild type mice. 1-tailed Student ttest, comparing <i>Hoxb4</i> vs. wild type mice. LN = Lymph node; BM = bone marrow.</p

Total donor cells contribution to hematopoietic organs and the proportion of <i>Hoxb4</i> versus wild type cells.

<p>LN = Lymph node; BM = bone marrow, wt = wild type.</p