Dexamethasone preconditioning improves the response of collagen-induced arthritis to treatment with short-term lipopolysaccharide-stimulated collagen-loaded dendritic cells.

Background. Pharmacologically modulated dendritic cells (DCs) have been shown to restore tolerance in type II collagen-(CII-) induced arthritis (CIA). We examined the effect of dexamethasone (DXM) administration as a preconditioning agent, followed by an injection of lipopolysaccharide-(LPS-) stimulated and CII-loaded DCs on the CIA course. Methods. After CIA induction, mice pretreated with DXM were injected with 4-hour LPS-stimulated DCs loaded with CII (DXM/4hLPS/CII/DCs). Results. Mice injected with DXM/4hLPS/CII/DCs displayed significantly less severe clinical disease compared to animals receiving 4hLPS/CII/DCs alone or those in which only DXM was administered. Cytokine profile evaluation showed that CD4+ T cells from DXM/4hLPS/CII/DCs and 4hLPS/CII/DCs groups release higher IL-10 levels than those from mice receiving DXM alone or CIA mice. CD4+ T cells from all DC-treated groups showed less IL-17 release when compared to the CIA group. On the contrary, CD4+ T cells from DXM/4hLPS/CII/DCs and 4hLPS/CII/DCs groups released higher IFN- γ levels than those from CIA group. Conclusion. A combined treatment, including DXM preconditioning followed by an inoculation of short-term LPS-stimulated CII-loaded DCs, provides an improved strategy for attenuating CIA severity. Our results suggest that this benefit is driven by a modulation in the cytokine profile secreted by CD4+ T cells.

Rrn7 protein, an RNA polymerase I transcription factor, is required for RNA polymerase II-dependent transcription directed by core promoters with a HomolD box sequence.

The region in promoters that specifies the transcription machinery is called the core promoter, displaying core promoter elements (CPE) necessary for establishment of a preinitiation complex and the initiation of transcription. A classical CPE is the TATA box. In fission yeast, Schizosaccharomyces pombe, a new CPE, called HomolD box, was discovered. Collectively, 141 ribosomal protein genes encoding the full set of 79 different ribosomal proteins and more than 60 other housekeeping genes display a HomolD box in the core promoter. Here, we show that transcription directed by the HomolD box requires the RNA polymerase II machinery, including the general transcription factors. Most intriguingly, however, we identify, by DNA affinity purification, Rrn7 as the protein binding to the HomolD box. Rrn7 is an evolutionary conserved member of the RNA polymerase I machinery involved in transcription initiation of core ribosomal DNA promoters. ChIP shows that Rrn7 cross-links to a ribosomal protein gene promoter containing the HomolD box but not to a promoter containing a TATA box. Taken together, our results suggest that Rrn7 is an excellent candidate to be involved in the coordination of ribosomal DNA and ribosomal gene transcription during ribosome synthesis and, therefore, offer a new perspective to study conservation and evolvability of regulatory networks in eukaryotes.

Schizosaccharomyces pombe positive cofactor 4 stimulates basal transcription from TATA-containing and TATA-less promoters through Mediator and transcription factor IIA.

The positive cofactor 4 (PC4) protein has an important role in transcriptional activation, which has been proposed to be mediated by transcription factor IIA (TFIIA) and TATA-binding protein-associated factors. To test this hypothesis, we cloned the Schizosaccharomyces pombe PC4 gene and analysed the role of the PC4 protein in the stimulation of basal transcription driven by TATA-containing and TATA-less promoters. Sc. pombe PC4 was able to stimulate basal transcription from several TATA-containing promoters and from the Initiator sequences of the highly transcribed Sc. pombe nmt1 gene. Moreover, it was demonstrated that Sc. pombe PC4 stimulates formation of the transcription preinitiation complex. Activation of transcription by PC4 was dependent on the Mediator complex and TFIIA, but was independent of TATA-binding protein-associated factor. PC4 binds to double-stranded and single-stranded DNA and interacts with TATA-binding protein, TFIIB, TFIIA, Mediator, TFIIH and the transcriptional activator protein VP16.

Short-term lipopolysaccharide stimulation induces differentiation of murine bone marrow-derived dendritic cells into a tolerogenic phenotype.

Dendritic cells (DCs) are professional, antigen-presenting cells, which induce and regulate T cell reactivity. DCs are crucial in innate and adaptive immune responses, and are also involved in central and peripheral tolerance induction. Tolerance can be mediated by immature and semi-mature DCs expressing low levels of co-stimulator and major histocompatibility complex (MHC) molecules. The aim of this study was to investigate the ability of short-term lipopolysaccharide (LPS) stimulation to modulate the stage of differentiation of bone marrow-derived DCs. For this purpose, DCs obtained from DBA1/lacJ mice were stimulated for four (4hLPS/DCs) or 24 (24hLPS/DCs) hours with LPS, using DCs without stimulation (0hLPS/DCs) as a control. Flow cytometry analysis of 4hLPS/DCs showed intermediate CD40 and MHC class II expression, lower than that of 24hLPS/DCs (fully mature), and greater than that of 0hLPS/DCs (immature). A functional assay showed that 4hLPS/DCs displayed increased endocytotic ability compared to 24hLPS/DCs, indicating a semi-mature state. 4hLPS/DCs were greater producers of IL-10 protein and TGFbeta1 mRNA than 24hLPS/DCs and immature DCs, displaying a cytokine production pattern that is characteristic of tolerogenic DCs. An assay for antigen-presenting capacity demonstrated that 4hLPS/DCs induced secretion of IL-2 from an OTH4 T cell hybridoma, indicating a functional presenting activity. Finally, the tolerogenic phenotype of 4hLPS/DCs was demonstrated by their ability to interfere with the progression of bovine type II collagen (bII)-induced arthritis (CIA) when they were loaded with bCII antigen and injected into mice with established CIA. We conclude that the stimulation of murine bone marrow-derived DCs with LPS for four hours generates semi-mature DCs with tolerogenic capability.

The -308 polymorphism in the promoter region of the tumor necrosis factor-alpha (TNF-alpha) gene and ex vivo lipopolysaccharide-induced TNF-alpha expression in patients with aggressive periodontitis and/or type 1 diabetes mellitus.

Several single-nucleotide polymorphisms (SNPs) have been identified in the TNF-alpha gene promoter. The transition G-->A at position -308 generates the TNF-alpha1 (G/G) and TNF-alpha2 (G/A or A/A) alleles, where the polymorphic TNF-alpha2 allele is associated with a high, in vitro TNF-alpha expression and an increased susceptibility to diverse illnesses. Here we study the association of the -308 TNF-alpha SNP with the susceptibility for developing aggressive periodontitis (AP), AP combined with type 1 diabetes mellitus (DM) and DM. We also explore the TNF-alpha capability expression and the presence of the -308 polymorphism. For this purpose we recruited 27 individuals with AP (AP+ group), 27 individuals with AP combined with DM (AP+/DM+ group), and 27 individuals with DM without signs of periodontitis upon clinical examination (DM+ group). The control group was comprised of 30 subjects. Genotyping for TNF-alpha promoter was performed by PCR-RFLP analysis. For TNF-alpha expression we used a blood culture system.

Transcription directed by human core promoters with a HomolD box sequence requires DDB1, RECQL and RNA polymerase II machinery.

TATA box is the most studied core promoter element and has a well-described transcription mechanism. However, most metazoan promoters lack TATA box and contain other core promoter elements. One of such elements is HomolD box, which was first described in promoters of ribosomal protein genes in Schizosaccharomyces pombe, and studies performed in this model showed that transcription directed by HomolD box is dependent on RNAPII machinery, and the HomolD-binding protein was Rrn7, a component of RNAPI core factor. Nevertheless, the mechanisms that underlie HomolD-dependent transcription are still unknown. The purpose of this study is to determine the mechanism of transcription directed by human HomolD box. By stepwise purification through different ion exchange columns and affinity chromatography, we purified two proteins: DDB1 and RECQL (DNA damage-binding protein 1 and ATP-dependent DNA helicase Q1 respectively). These proteins showed specific HomolD-binding activity and were required for in vitro HomolD-directed transcription. Recombinant RECQL, but not DDB1, presented HomolD-binding activity in vitro. Both proteins bound to HomolD box in vivo, which could be explained because these proteins co-immunoprecipitated. Additionally, RNAPII machinery was also required to transcription. Collectively, these data suggest that HomolD-containing promoters require the RNAPII machinery and the proteins DDB1 and RECQL for an accurate transcription.

Anemia hemolítica secundaria a plastía mitral: caso clínico local y revisión de la literatura / Hemolytic anemia after mitral valve repair: case report

Una mujer de 54 años sometida a una reparación mitral se presenta 2 meses después con anemia hemolítica. El mecanismo de ésta correspondió a dehiscencia de la cuadrantectomía y a falta de endotelización de un sector del anillo, que se evidenció mediante ecografía transesofágica y se confirmó por hallazgos operatorios. Se resecó el anillo mitral y se implantó una prótesis de Saint Jude, y no hubo recurrencia de la hemólisis. Se incluye una discusión con revisión de la literatura.

A 54 year old woman was subjected to a mitral valve repair of a mixomatous degenerative valve with severe mitral insufficiency. Quadrantectomy, mitral chords transfer from P2 to a A2 and implantation of a rigid Edwards 28 ring were performed. Two months later the patient was diagnosed with severe hemolytic anemia. Trans esophageal echocardiography revealed severe mitral insufficiency and at surgery a dehiscence at the base of the qudrantectomy in addition to a non endothelysed sector of the mitral ring were found. A Saint-Jude mitral prosthesis was inserted. At follow up, no recurrence of hemolysis occurred.

[Emergent therapies for rheumatoid arthritis]. / Terapias emergentes en artritis reumatoide.

The use of biological agents such as etanercept, infliximab, adalimumab and anakinra has been recently approved for the treatment of rheumatoid arthritis. All are effective controlling signs and symptoms and inhibiting disease progression. To overcome the problems generated by their high costs and possible participation in reactivating latent infections, other therapeutic tools are being developed. Gene therapy using expression vectors carrying genes coding for specific proteins, may interfere in key points involved in the pathogenesis of the disease. Intra-articular administration of cDNA coding for soluble TNF receptors, IL-1, or IL-1Ra decreases signs of the disease in animal models. Vectors, expressing inhibitors of signal transduction pathways involving to NF-kB and JAK-STAT-3, are effective in modulating joint inflammation in mice. The use of antigen-pulsed antigen presenting cells or dendritic cells (DC) bound to apoptosis-inducing molecules, specifically eliminates autoreactive T cells. Other novel approach attempts the development of T regulatory-inducing tolerogenic DC-based vaccines that inhibit autoreactive T cells, through the secretion of suppressing cytokines or by other mechanisms to be elucidated. Oral tolerance induction to auto-antigens is also a successful experimental strategy under study. Current research aims to control peripheral tolerance in rheumatoid arthritis patients.

Abortos espontáneos en Hospital de Llay-Llay y su relación con labores agrícolas de la madre / Spontaneous abortions in the Llay -Llay hospital an their relation to mother's agricultural labors

Los pesticidas son sustancias químicas usadas con frecuencia en las zonas agrícolas de nuestro país. Es un hecho bien establecido que estas sustancias pueden ser dañinas para la salud no sólo del trabajador agrícola sino también para su descendencia. Su objetivo es estudiar la asociación entre abortos espontáneos y la ocupación agrícola de la madre. Se efectuó un estudio descriptivo, recopilando las fichas clínicas de aquellas pacientes que presentaron aborto espontáneo registrados en el libro de procedimientos de la maternidad del Hospital San Francisco de Llay-Llay entre enero del 2002 y diciembre del 2003. Se calcula la tasa de incidencia de aborto espontáneo comparándola con las últimas cifras nacionales disponibles, y el porcentaje de ocupación agrícola de la madre en relación con la población femenina ocupada de 15 años o más de la misma área geográfica según Censo del 2002. Se realizó la prueba Z para comparación de proporciones, e intervalos de confianza para la razón entre las densidades de incidencia. La tasa de incidencia de aborto espontáneo fue 81,02 casos/1.000 RNV versus 9,5 casos/1.000 RNVen Chile 1996, otorgando una razón de incidencia de 8,5 veces (IC = 6,72-10,65). El porcentaje de ocupación agrícola es mayor que la población femenina ocupada de 15 años o más de la misma área geográfica. (p <0,0001). El riesgo de presentar aborto espontáneo en Llay-Llay y Catemu es mayor que el resto del país. La ocupación agrícola está relacionada con un mayor riesgo de aborto espontáneo, probablemente debida a la exposición a pesticidas.

Pesticides are chemical substances frequently used in agricultural areas of our country. It is a well established fact that these substances can be harmful to the health, not only of agricultural workers, but also to that of their offspring. The objective of this paper is that of studying the relation between spontaneous abortions and the agricultural occupation of the mother. A descriptive study was carried out assembling the clinical records of those patients presenting sponrtaneous abortion registered in the maternity book of procedures of the San Francisco de Llay-Llay Hospital between January 2002 and December 2003. The incidence of spontaneous abortion is calculated comparing it to the last available national figures, and the percentage of agricultural occupation of the mother in relation to the 15 year old and over female population of the same geographic area, according to the 2002 Census. The Z test was performed for comparison of proportions and confidence intervals for the ratio between incidence densities. The incidence rate for spontaneous abortion was 81.02 cases/1,000 live-newborn registrations (RNV) versus 9.5 cases/1,000 live-born registrations in Chile 1996, granting an incidence ratio of 8.5 (CI=6.72-10.65). The percentage of agricultural occupation is higher than the female 15 year old and over population of the same geographic area (p<0.0001). The risk of spontaneous abortion occurring in Llay-Llay or Catemu is higher than in the rest of the country. The agricultural occupation is related to a higher risk of spontaneous abortion, probably due to the exposure to pesticides.

Terapias emergentes en artritis reumatoide / Emergent therapies for rheumatoid arthritis

The use of biological agents such as etanercept, infliximab, adalimumab and anakinra has been recently approved for the treatment of rheumatoid arthritis. All are effective controlling signs and symptoms and inhibiting disease progression. To overcome the problems generated by their high costs and possible participation in reactivating latent infections, other therapeutic tools are being developed. Gene therapy using expression vectors carrying genes coding for specific proteins, may interfere in key points involved in the pathogenesis of the disease. Intra-articular administration of cDNA coding for soluble TNF receptors, IL-1, or IL-1Ra decreases signs of the disease in animal models. Vectors, expressing inhibitors of signal transduction pathways involving to NF-kB and JAK-STAT-3, are effective in modulating joint inflammation in mice. The use of antigen-pulsed antigen presenting cells or dendritic cells (DC) bound to apoptosis-inducing molecules, specifically eliminates autoreactive T cells. Other novel approach attempts the development of T regulatory-inducing tolerogenic DC-based vaccines that inhibit autoreactive T cells, through the secretion of suppressing cytokines or by other mechanisms to be elucidated. Oral tolerance induction to auto-antigens is also a successful experimental strategy under study. Current research aims to control peripheral tolerance in rheumatoid arthritis patients.

Strongyloidosis no autóctona en Chile: descripción de un brote familiar / Non autochthonous strongyloidosis in Chile: report of a familiar outbrake

Se presenta un brote familiar de stron-gyloidosis que afectó a inmigrantes peruanos que habían llegado a Chile en noviembre del año 2001. En marzo del 2002, se confirmó en el Laboratorio de Parasitología Básico-Clínico de la Facultad de Medicina de la Universidad de Chile que el caso índice presentaba infección por Strongyloides stercoralis. Correspondía a una menor de 11 años que estaba siendo estudiada en el Hospital Roberto del Río por artritis reumatoide juvenil. Además de esta patología la niña refería leves molestias abdominales, siendo derivada a nuestra unidad, donde se le efectuó un examen parasitológico seriado de deposiciones (EPSD), observándose abundantes larvas rabditoides de S. stercoralis, por lo cual se procedió a estudiar al grupo familiar integrado por los padres y una hermana. A todos ellos se les efectuó EPSD e inmunodiagnóstico mediante ELISA para detectar anticuerpos anti S.stercoralis. Resultaron positivos para ambos exámenes la madre y las dos hijas. El padre fue negativo. Se decidió tratar con ivermectina y controlar los resultados del tratamiento anti-parasitario al grupo familiar al cabo de un año. Después de ese periodo permanecía positiva serológicamente, solamente la hermana del caso índice debido a que no ingirió los medicamentos de la manera indicada.