Consensus statement of the European guidelines on clinical management of HIV-1 tropism testing

Introduction: Testing for HIV tropism is recommended before prescribing a chemokine receptor blocker. To date, in most European countries HIV tropism is determined using a phenotypic test. Recently, new data have emerged supporting the use of a genotypic HIV V3-loop sequence analysis as the basis for tropism determination. The European guidelines group on clinical management of HIV-1 tropism testing was established to make recommendations to clinicians and virologists. Methods: We searched online databases for articles from Jan 2006 until March 2010 with the terms: tropism or CCR5-antagonist or CCR5 antagonist or maraviroc or vicriviroc. Additional articles and/or conference abstracts were identified by hand searching. This strategy identified 712 potential articles and 1240 abstracts. All were reviewed and finally 57 papers and 42 abstracts were included and used by the panel to reach a consensus statement. Results: The panel recommends HIV-tropism testing for the following indications: i) drug-naïve patients in whom toxicity or limited therapeutic options are foreseen; ii) patients experiencing therapy failure whenever a treatment change is considered. Both the phenotypic Enhanced Trofile assay (ESTA) and genotypic population sequencing of the V3-loop are recommended for use in clinical practice. Although the panel does not recommend one methodology over another it is anticipated that genotypic testing will be used more frequently because of its greater accessibility, lower cost and shorter turnaround time. The panel also provides guidance on technical aspects and interpretation issues. If using genotypic methods, triplicate PCR amplification and sequencing testing is advised using the G2P interpretation tool (clonal model) with an FPR of 10%. If the viral load is below the level of reliable amplification, proviral DNA can be used, and the panel recommends performing triplicate testing and use of an FPR of 10%. If genotypic DNA testing is not performed in triplicate the FPR should be increased to 20%. Conclusions: The European guidelines on clinical management of HIV-1 tropism testing provide an overview of current literature, evidence-based recommendations for the clinical use of tropism testing and expert guidance on unresolved issues and current developments. Current data support both the use of genotypic population sequencing and ESTA for co-receptor tropism determination. For practical reasons genotypic population sequencing is the preferred method in Europe.

A Change in the Epidemiology of Infections Due to Extended-Spectrum b-Lactamase–Producing Organisms

Extended-spectrum β-lactamases (ESBLs) form a heterogeneous group that share the property of hydrolytic activity against the oxyimino-β-lactams while remaining susceptible to inhibition by β-lactamase inhibitors, such as clavulanic acid. From a clinical point of view, they are important because they confer resistance to penicillins, aztreonam, and cephalosporins, and ESBL-producing organisms are typically also resistant to aminoglycosides, trimethoprim-sulfamethoxazole, and quinolones [1]. Until recently, the main problem posed by ESBLs was related to nosocomial outbreaks caused by ESBL-producing Klebsiella species. These outbreaks are usually clonal, the strains are mainly spread through cross-transmission, and the risk factors are similar to those found for other multidrug-resistant nosocomial pathogens [2]. In Europe and the United States, most ESBL-producing Klebsiella isolates harbored enzymes belonging to the TEM and SHV families [3]. Detection of colonized patients by performing surveillance cultures within affected units, isolation precautions for colonized patients, and restriction of oxyimino-β-lactam use are frequently useful for the control of these outbreaks [1]. There is no evidence that hospital-acquired ESBL-producing klebsiellae are decreasing in importance—in fact, data from the Centers for Disease Control and Prevention show that 20.6% of Klebsiella pneumoniae isolates from United States intensive care units in 2003 were probable producers of ESBL [4]. This represented a 47% increase, compared with the preceding 5 years. However, during the last few years, an impressive increase in the number of ESBL-producing Escherichia coli (and, less frequently, other Enterobacteriaceae) is being described in several parts of the world [5–8]. This emergent phenomenon shows some differences from the problem posed by Klebsiella species; many of these ESBL-producing E. coli are isolated …

High ACSL5 transcript levels associate with systemic lupus erythematosus and apoptosis in Jurkat T lymphocytes and peripheral blood cells

BACKGROUND: Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease in which increased apoptosis and decreased apoptotic cells removal has been described as most relevant in the pathogenesis. Long-chain acyl-coenzyme A synthetases (ACSLs) have been involved in the immunological dysfunction of mouse models of lupus-like autoimmunity and apoptosis in different in vitro cell systems. The aim of this work was to assess among the ACSL isoforms the involvement of ACSL2, ACSL4 and ACSL5 in SLE pathogenesis. FINDINGS: With this end, we determined the ACSL2, ACSL4 and ACSL5 transcript levels in peripheral blood mononuclear cells (PBMCs) of 45 SLE patients and 49 healthy controls by quantitative real time-PCR (q-PCR). We found that patients with SLE had higher ACSL5 transcript levels than healthy controls [median (range), healthy controls =16.5 (12.3-18.0) vs. SLE = 26.5 (17.8-41.7), P = 3.9x10 E-5] but no differences were found for ACSL2 and ACSL4. In in vitro experiments, ACSL5 mRNA expression was greatly increased when inducing apoptosis in Jurkat T cells and PBMCs by Phorbol-Myristate-Acetate plus Ionomycin (PMA+Io). On the other hand, short interference RNA (siRNA)-mediated silencing of ACSL5 decreased induced apoptosis in Jurkat T cells up to the control levels as well as decreased mRNA expression of FAS, FASLG and TNF. CONCLUSIONS: These findings indicate that ACSL5 may play a role in the apoptosis that takes place in SLE. Our results point to ACSL5 as a potential novel functional marker of pathogenesis and a possible therapeutic target in SLE

Giant proliferating trichilemmal malignant tumor

Proliferating trichilemmal cyst is a benign tumor that originates in the outer root sheath of hair follicle. Usually, it is located on the scalp in older women, but also have been reported in other sites such as back, chest, axilla, groin, gluteal region, thigh, vulva, and face. Malignant transformation of proliferating trichilemmal cyst is confirmed only on histological findings. This tumor has a variable biologic behavior with local recurrence and lymph node metastasis.

Granada Virus: a Natural Phlebovirus Reassortant of the Sandfly Fever Naples Serocomplex with Low Seroprevalence in Humans

A new member of the phlebovirus genus, tentatively named Granada virus, was detected in sandflies collected in Spain. By showing the presence of specific neutralizing antibodies in human serum collected in Granada, we show that Granada virus infects humans. The analysis of the complete genome of Granada virus revealed that this agent is likely to be a natural reassortant of the recently described Massilia virus (donor of the long and short segments) with ayet unidentified phlebovirus (donor of the medium segment)

Diarrea crónica refractaria y malabsorción secundaria a hipogammaglobulinemia común variable, infestación crónica por giardia lambila y gastrectomía total por adenocarcinoma gástrico: un manejo nutricional complejo

Gastric cancer is a frequent cause of cancer-related mortality in the world. Surgery is the only potentially curative therapy, although the adverse effects of surgery are common and considerable. Common variable immunodeficiency is in many cases cause of gastrointestinal system problems such as chronic diarrhea caused by infestation with giardia lamblia, nodular lymphoid hiperplasia ad loss of villi leading frequently to malapsortion and malnutrition. Nutritional deficiencies due to malapsorption (postgastrectomy and secondary to loss of villi, giardiasis and common variable inmunodeficiency) are common. We present the case of a patient with gastric cancer who underwent a gastrectomy with common variable hipogammaglobulinemia and chronic infestation by giardia lamblia, with serious diarrhea resistant to treatment and malabsorption.

The immunomodulator PSK induces in vitro cytotoxic activity in tumour cell linesvia arrest of cell cycle and induction of apoptosis

BACKGROUND Protein-bound polysaccharide (PSK) is derived from the CM-101 strain of the fungus Coriolus versicolor and has shown anticancer activity in vitro and in in vivo experimental models and human cancers. Several randomized clinical trials have demonstrated that PSK has great potential in adjuvant cancer therapy, with positive results in the adjuvant treatment of gastric, esophageal, colorectal, breast and lung cancers. These studies have suggested the efficacy of PSK as an immunomodulator of biological responses. The precise molecular mechanisms responsible for its biological activity have yet to be fully elucidated. METHODS The in vitro cytotoxic anti-tumour activity of PSK has been evaluated in various tumour cell lines derived from leukaemias, melanomas, fibrosarcomas and cervix, lung, pancreas and gastric cancers. Tumour cell proliferation in vitro was measured by BrdU incorporation and viable cell count. Effect of PSK on human peripheral blood lymphocyte (PBL) proliferation in vitro was also analyzed. Studies of cell cycle and apoptosis were performed in PSK-treated cells. RESULTS PSK showed in vitro inhibition of tumour cell proliferation as measured by BrdU incorporation and viable cell count. The inhibition ranged from 22 to 84%. Inhibition mechanisms were identified as cell cycle arrest, with cell accumulation in G0/G1 phase and increase in apoptosis and caspase-3 expression. These results indicate that PSK has a direct cytotoxic activity in vitro, inhibiting tumour cell proliferation. In contrast, PSK shows a synergistic effect with IL-2 that increases PBL proliferation. CONCLUSION These results indicate that PSK has cytotoxic activity in vitro on tumour cell lines. This new cytotoxic activity of PSK on tumour cells is independent of its previously described immunomodulatory activity on NK cells.

CD209 in inflammatory bowel disease: a case-control study in the Spanish population

BACKGROUND The etiology of Ulcerative Colitis (UC) and Crohn's Disease (CD), considered together as Inflammatory Bowel Diseases (IBD), involves environmental and genetic factors. Although some genes are already known, the genetics underlying these diseases is complex and new candidates are continuously emerging. The CD209 gene is located in a region linked previously to IBD and a CD209 functional polymorphism (rs4804803) has been associated to other inflammatory conditions. Our aim was to study the potential involvement of this CD209 variant in IBD susceptibility. METHODS We performed a case-control study with 515 CD patients, 497 UC patients and 731 healthy controls, all of them white Spaniards. Samples were typed for the CD209 single nucleotide polymorphism (SNP) rs4804803 by TaqMan technology. Frequency comparisons were performed using chi2 tests. RESULTS No association between CD209 and UC or CD was observed initially. However, stratification of UC patients by HLA-DR3 status, a strong protective allele, showed that carriage of the CD209_G allele could increase susceptibility in the subgroup of HLA-DR3-positive individuals (p = 0.03 OR = 1.77 95% CI 1.04-3.02, vs. controls). CONCLUSION A functional variant in the CD209 gene, rs4804803, does not seem to be influencing Crohn's disease susceptibility. However, it could be involved in the etiology or pathology of Ulcerative Colitis in HLA-DR3-positive individuals but further studies are necessary.

Histological and histochemical evaluation of human oral mucosa constructs developed by tissue engineering

Reconstruction of large oral mucosa defects is often challenging, since the shortage of healthy oral mucosa to replace the excised tissues is very common. In this context, tissue engineering techniques may provide a source of autologous tissues available for transplant in these patients. In this work, we developed a new model of artificial oral mucosa generated by tissue engineering using a fibrin-agarose scaffold. For that purpose, we generated primary cultures of human oral mucosa fibroblasts and keratinocytes from small biopsies of normal oral mucosa using enzymatic treatments. Then we determined the viability of the cultured cells by electron probe quantitative X-ray microanalysis, and we demonstrated that most of the cells in the primary cultures were alive and had high K/Na ratios. Once cell viability was determined, we used the cultured fibroblasts and keratinocytes to develop an artificial oral mucosa construct by using a fibrin-agarose extracellular matrix and a sequential culture technique using porous culture inserts. Histological analysis of the artificial tissues showed high similarities with normal oral mucosa controls. The epithelium of the oral substitutes had several layers, with desmosomes and apical microvilli and microplicae. Both the controls and the oral mucosa substitutes showed high suprabasal expression of cytokeratin 13 and low expression of cytokeratin 10. All these results suggest that our model of oral mucosa using fibrin-agarose scaffolds show several similarities with native human oral mucosa.

Analysis of TNFAIP3, a feedback inhibitor of nuclear factor-κB and the neighbor intergenic 6q23 region in rheumatoid arthritis susceptibility

INTRODUCTION Genome-wide association studies of rheumatoid arthritis (RA) have identified an association of the disease with a 6q23 region devoid of genes. TNFAIP3, an RA candidate gene, flanks this region, and polymorphisms in both the TNFAIP3 gene and the intergenic region are associated with systemic lupus erythematosus. We hypothesized that there is a similar association with RA, including polymorphisms in TNFAIP3 and the intergenic region. METHODS To test this hypothesis, we selected tag-single nucleotide polymorphisms (SNPs) in both loci. They were analyzed in 1,651 patients with RA and 1,619 control individuals of Spanish ancestry. RESULTS Weak evidence of association was found both in the 6q23 intergenic region and in the TNFAIP3 locus. The rs582757 SNP and a common haplotype in the TNFAIP3 locus exhibited association with RA. In the intergenic region, two SNPs were associated, namely rs609438 and rs13207033. The latter was only associated in patients with anti-citrullinated peptide antibodies. Overall, statistical association was best explained by the interdependent contribution of SNPs from the two loci TNFAIP3 and the 6q23 intergenic region. CONCLUSIONS Our data are consistent with the hypothesis that several RA genetic factors exist in the 6q23 region, including polymorphisms in the TNFAIP3 gene, like that previously described for systemic lupus erythematosus.

New aspects in celiac disease

Celiac disease (CD) is a common autoimmune disorder characterized by an immune response to ingested gluten and has a strong HLA association with HLA-DQ2 and HLA-DQ8 molecules, but human HLA-DQ risk factors do not explain the entire genetic susceptibility to gluten intolerance. CD is caused by the lack of immune tolerance (oral tolerance) to wheat gluten. In this sense, the expression of soluble HLA-G in CD is of special interest because the molecule plays an important role in the induction of immune tolerance. The enhanced expression of soluble HLA-G found in CD may be part of a mechanism to restore the gluten intolerance. In this editorial, we review recent progress in understanding CD in relation to its prevalence, diagnosis and possible mechanisms of pathogenesis.

Alveolar fluid clearance in healthy pigs and influence of positive end-expiratory pressure

INTRODUCTION The objectives were to characterize alveolar fluid clearance (AFC) in pigs with normal lungs and to analyze the effect of immediate application of positive end-expiratory pressure (PEEP). METHODS Animals (n = 25) were mechanically ventilated and divided into four groups: small edema (SE) group, producing pulmonary edema (PE) by intratracheal instillation of 4 ml/kg of saline solution; small edema with PEEP (SE + PEEP) group, same as previous but applying PEEP of 10 cmH2O; large edema (LE) group, producing PE by instillation of 10 ml/kg of saline solution; and large edema with PEEP (LE + PEEP) group, same as LE group but applying PEEP of 10 cmH2O. AFC was estimated from differences in extravascular lung water values obtained by transpulmonary thermodilution method. RESULTS At one hour, AFC was 19.4% in SE group and 18.0% in LE group. In the SE + PEEP group, the AFC rate was higher at one hour than at subsequent time points and higher than in the SE group (45.4% vs. 19.4% at one hour, P < 0.05). The AFC rate was also significantly higher in the LE + PEEP than in the LE group at three hours and four hours. CONCLUSIONS In this pig model, the AFC rate is around 20% at one hour and around 50% at four hours, regardless of the amount of edema, and is increased by the application of PEEP.

Role of wireless capsule endoscopy in inflammatory bowel

Capsule endoscopy (CE) offers state-of-the-art imaging of the small bowel. In Crohn's disease its clinical role is still uncertain. This report analyses the usefulness of CE in patients with suspected Cronh's disease, in patients with established Crohn's disease (when assessing severity, occult gastrointestinal bleeding and/or as a guide to therapy), in patients with inflammatory bowel disease unclassified (IBDU), and in individuals with ulcerative colitis. The first item in this group is the most important although there is no strong evidence to establish the position of CE in the diagnostic workup. In patients with established Crohn's disease, recently developed activity scores are promising tools for an accurate assessment of severity. As a guide to therapy, CE should be focused on patients with unexplained symptoms when other investigations are inconclusive. In postoperative Crohn's Disease, international consensus recommended considering CE only if ileocolonoscopy is contraindicated or unsuccessful. In the case of IBDU, studies have shown a significant proportion of patients reclassified with Crohn's disease. In this setting, CE could have a role determining small bowel involvement. The role of CE in ulcerative colitis is limited. Some authors advocate CE before colectomy for refractory cases in order to exclude Crohn's disease. In summary, CE offers a new horizon in inflammatory bowel disease, and a better knowledge of mucosal abnormalities that could offer a paradigm shift: changing from symptom-based disease activity estimation to direct mucosal healing monitoring. Nevertheless, randomized controlled studies are still needed to provide stronger evidence in this setting.

The Presence of Precursors of Benign Pre-B Lymphoblasts (Hematogones) in the Bone Marrow of a Paediatric Patient with Cytomegalovirus Infection

Hematogones are normal B-lymphoid precursors that multiply in the bone marrow of small children and of adults with ferropenic anaemia, neuroblastoma or idiopathic thrombocytopenic purpura. They are not normally found in peripheral blood, and the immunophenotype is virtually indistinguishable from that of B lymphoblasts. We discuss the case of a 3-month infant with an active cytomegalovirus infection, with hepatitis and pancytopenia associated with 13% hematogones in the bone marrow

Mutation analysis of genes that control the G1/S cell cycle in melanoma: TP53, CDKN1A, CDKN2A, and CDKN2B.

BACKGROUND The role of genes involved in the control of progression from the G1 to the S phase of the cell cycle in melanoma tumors in not fully known. The aim of our study was to analyse mutations in TP53, CDKN1A, CDKN2A, and CDKN2B genes in melanoma tumors and melanoma cell lines METHODS We analysed 39 primary and metastatic melanomas and 9 melanoma cell lines by single-stranded conformational polymorphism (SSCP). RESULTS The single-stranded technique showed heterozygous defects in the TP53 gene in 8 of 39 (20.5%) melanoma tumors: three new single point mutations in intronic sequences (introns 1 and 2) and exon 10, and three new single nucleotide polymorphisms located in introns 1 and 2 (C to T transition at position 11701 in intron 1; C insertion at position 11818 in intron 2; and C insertion at position 11875 in intron 2). One melanoma tumor exhibited two heterozygous alterations in the CDKN2A exon 1 one of which was novel (stop codon, and missense mutation). No defects were found in the remaining genes. CONCLUSION These results suggest that these genes are involved in melanoma tumorigenesis, although they may be not the major targets. Other suppressor genes that may be informative of the mechanism of tumorigenesis in skin melanomas should be studied.