Mucin 5B, carbonic anhydrase 9 and claudin 18 are potential theranostic markers of gallbladder carcinoma.

AIMS: Gallbladder cancer (GBC) is an aggressive tumour that is usually diagnosed at advanced stages and is characterised by a poor prognosis. Using public data of normal human tissues, we found that mRNA and protein levels of mucin 5B (MUC5B) and carbonic anhydrase 9 (CA9) were highly increased in gallbladder tissues. In addition, previous evidence has shown that claudin 18 (CLDN18) protein expression is higher in GBC. The aim of this study was to perform an analysis of these cell surface proteins during the histological progression of GBC in order to identify their theranostic potential. METHODS AND RESULTS: MUC5B expression, CA9 expression and CLDN18 expression were examined by immunohistochemistry in a series of 179 chronic cholecystitis (including 16 metaplastic tissues), 15 dysplasia and 217 GBC samples by the use of tissue microarray analysis. A composite staining score was calculated from staining intensity and percentage of positive cells. Immunohistochemical analysis showed high expression of MUC5B and CA9 among normal epithelium, metaplastic tissues, and dysplastic tissues. However, expression of both proteins was observed in roughly 50% of GBC samples. In contrast, CLDN18 was absent in normal epithelium, but its expression was higher in metaplastic cells. Among GBC cases, approximately half showed high CLDN18 expression. No associations were found between MUC5B, CA9 and CLDN18 expression and any clinicopathological features. CONCLUSIONS: CLDN18 is a new metaplasia marker in gallbladder tissues, and is conserved in approximately half of GBC cases. MUC5B and CA9 are highly conserved during GBC histological progression. The three markers are potential theranostic markers, in particular CA9 and CLDN18, for which there are already targeted therapies available.

[Recurrent respiratory papillomatosis in a pediatric patient: case report]. / Papilomatosis respiratoria recurrente en paciente pediátrico: reporte de un caso.

Genetic variability related to the host immune system has been proposed as one of the most influential factors in the development of diseases caused by HPV. CLINICAL CASE: We report the case of a 5-year-old child in whom chronic laryngeal papillomatosis, probably acquired vertically during labor, was detected. The diagnosis of laryngeal papillomatosis was confirmed with a biopsy after a first surgery to remove the papillomas. The Derkay classification system was used to assess the severity of papillomatosis. Biopsy genotyping was performed by demonstrating HPV-6. Later, HLA-DQA1 * 0505, -DQB1 * 0301, -DRB1 * 1101 alleles were homozygous for HLA allele typing. CONCLUSIONS: Further studies are needed to identify the most prevalent HLA alleles in the Latino population and their potential association with genetic susceptibility in Recurrent Respiratory Papillomatosis.

The low-abundance transcriptome reveals novel biomarkers, specific intracellular pathways and targetable genes associated with advanced gastric cancer.

Studies on the low-abundance transcriptome are of paramount importance for identifying the intimate mechanisms of tumor progression that can lead to novel therapies. The aim of the present study was to identify novel markers and targetable genes and pathways in advanced human gastric cancer through analyses of the low-abundance transcriptome. The procedure involved an initial subtractive hybridization step, followed by global gene expression analysis using microarrays. We observed profound differences, both at the single gene and gene ontology levels, between the low-abundance transcriptome and the whole transcriptome. Analysis of the low-abundance transcriptome led to the identification and validation by tissue microarrays of novel biomarkers, such as LAMA3 and TTN; moreover, we identified cancer type-specific intracellular pathways and targetable genes, such as IRS2, IL17, IFNγ, VEGF-C, WISP1, FZD5 and CTBP1 that were not detectable by whole transcriptome analyses. We also demonstrated that knocking down the expression of CTBP1 sensitized gastric cancer cells to mainstay chemotherapeutic drugs. We conclude that the analysis of the low-abundance transcriptome provides useful insights into the molecular basis and treatment of cancer.

High-resolution melting of the cytochrome B gene in fecal DNA: A powerful approach for fox species identification of the Lycalopex genus in Chile.

Easy, economic, precise species authentication is currently necessary in many areas of research and diagnosis in molecular biology applied to conservation studies of endangered species. Here, we present a new method for the identification of three fox species of the Lycalopex genus in Chile. We developed an assay based on high-resolution melt analysis of the mitochondrial cytochrome B gene, allowing a simple, low cost, fast, and accurate species determination. To validate the assay applicability for noninvasive samples, we collected fecal samples in the Atacama Desert, finding unexpectedly one species outside of its known distribution range. We conclude that the assay has a potential to become a valuable tool for a standardized genetic monitoring of the Lycalopex species in Chile.

Contribution of Fcγ Receptor-Mediated Immunity to the Pathogenesis Caused by the Human Respiratory Syncytial Virus.

The human Respiratory Syncytial Virus (hRSV) is the leading cause of severe acute lower respiratory tract infections (ALRTIs) in humans at all ages and is the main cause of hospitalization due to pneumonia, asthma, and bronchiolitis in infants. hRSV symptoms mainly develop due to an excessive host immune and inflammatory response in the respiratory tissue. hRSV infection during life is frequent and likely because of non-optimal immunological memory is developed against this virus. Vaccine development against this pathogen has been delayed after the detrimental effects produced in children by vaccination with a formalin-inactivated hRSV preparation (FI-hRSV), which caused enhanced disease upon natural viral infection. Since then, several studies have focused on understanding the mechanisms underlying such disease exacerbation. Along these lines, several studies have suggested that antibodies elicited by immunization with FI-hRSV show low neutralizing capacity and promote the formation of immune complexes containing hRSV (hRSV-ICs), which contribute to hRSV pathogenesis through the engagement of Fc gamma receptors (FcγRs) expressed on the surface of immune cells. Furthermore, a role for FcγRs is supported by studies evaluating the contribution of these molecules to hRSV-induced disease. These studies have shown that FcγRs can modulate viral clearance by the host and the inflammatory response triggered by hRSV infection. In addition, ICs can facilitate viral entry into host cells expressing FcγRs, thus extending hRSV infectivity. In this article, we discuss current knowledge relative to the contribution of hRSV-ICs and FcγRs to the pathogenesis caused by hRSV and their putative role in the exacerbation of the disease caused by this virus after FI-hRSV vaccination. A better understanding FcγRs involvement in the immune response against hRSV will contribute to the development of new prophylactic or therapeutic tools to promote virus clearance with limited inflammatory damage to the airways.

Contribution of IDO to human respiratory syncytial virus infection.

IDO is an enzyme that participates in the degradation of tryptophan (Trp), which is an essential amino acid necessary for vital cellular processes. The degradation of Trp and the metabolites generated by the enzymatic activity of IDO can have immunomodulating effects, notably over T cells, which are particularly sensitive to the absence of Trp and leads to the inhibition of T cell activation, cell death, and the suppression of T cell effector functions. Noteworthy, T cells participate in the cellular immune response against the human respiratory syncytial virus (hRSV) and are essential for viral clearance, as well as the total recovery of the host. Furthermore, inadequate or non-optimal polarization of T cells is often seen during the acute phase of the disease caused by this pathogen. Here, we discuss the capacity of hRSV to exploit the immunosuppressive features of IDO to reduce T cell function, thus acquiring relevant aspects during the biology of the virus. Additionally, we review studies on the influence of IDO over T cell activation and its relationship with hRSV infection.

Description of Echinococcus granulosus genotypes in human hydatidosis in a region of southern Chile.

INTRODUCTION: Echinococcus granulosus species has a wide variety in both geography and hosts; indeed, 10 genotypes have been reported in studies on material of animal origin. The aim of this study was to genotype E. granulosus obtained from human hydatid cysts. MATERIALS AND METHODS: The hydatid fluid and sand was collected from patients who underwent surgery for hepatic and pulmonary hydatidosis at Hospital Regional in Temuco, Chile, between 2004 and 2005. Two PCR systems were used: PCR Eg 9 and PCR Eg 16. The RsaI enzyme was used for RFLP. The genotype was confirmed using the sequence of one fragment of 366 bp from a mitochondrial gene (cox1). RESULTS: The DNA of protoscolices from 24 samples was analyzed, 4 of them from pulmonary cysts and 20 from hepatic cysts. The 366 bp fragment was amplified in 20 out of 24 samples (83.3%). Enzymatic digestion revealed the presence of 3 possible genotypes: in 20 out of 21 samples (95,2%), a restriction was observed corresponding to the G1 or G7 genotypes; in the remaining sample genotype G4 or G7 was observed. Sequencing confirmed the presence of G1 genotype for 19 samples and G6 genotype for the remaining sample (G4 or G7 according to PCR-RFLP). CONCLUSION: The PCR-RFLP technique enabled three possible genotypes present (G1 or G7, G4 or G7) to be established. Sequencing allowed us to decisively identify the G1 and G6 genotypes in our study group. Previous studies agree with the identification of the G1 genotype in our country. We consider it significant that the G6 genotype is present in Chile for its epidemiological implications.

Human Metapneumovirus: Mechanisms and Molecular Targets Used by the Virus to Avoid the Immune System.

Human metapneumovirus (hMPV) is a respiratory virus, first reported the year 2001. Since then, it has been described as one of the main etiological agents that causes acute lower respiratory tract infections (ALRTIs), which is characterized by symptoms such as bronchiolitis, wheezing and coughing. Susceptible population to hMPV-infection includes newborn, children, elderly and immunocompromised individuals. This viral agent is a negative-sense, single-stranded RNA enveloped virus, that belongs to the Pneumoviridae family and Metapneumovirus genus. Early reports-previous to 2001-state several cases of respiratory illness without clear identification of the responsible pathogen, which could be related to hMPV. Despite the similarities of hMPV with several other viruses, such as the human respiratory syncytial virus or influenza virus, mechanisms used by hMPV to avoid the host immune system are still unclear. In fact, evidence indicates that hMPV induces a poor innate immune response, thereby affecting the adaptive immunity. Among these mechanisms, is the promotion of an anergic state in T cells, instead of an effective polarization or activation, which could be induced by low levels of cytokine secretion. Further, the evidences support the notion that hMPV interferes with several pattern recognition receptors (PRRs) and cell signaling pathways triggered by interferon-associated genes. However, these mechanisms reported in hMPV are not like the ones reported for hRSV, as the latter has two non-structural proteins that are able to inhibit these pathways. Several reports suggest that viral glycoproteins, such as G and SH, could play immune-modulator roles during infection. In this work, we discuss the state of the art regarding the mechanisms that underlie the poor immunity elicited by hMPV. Importantly, these mechanisms will be compared with those elicited by other common respiratory viruses.

Effects of clopidogrel on inflammatory cytokines and adhesion molecules in human endothelial cells: Role of nitric oxide mediating pleiotropic effects.

INTRODUCTION: Clopidogrel is commonly used in prevention and treatment of atherothrombosis. Some previous studies have suggested a pleiotropic effect of clopidogrel; however, when this drug causes platelet-independent effects on endothelial function remains unclear. AIMS: To evaluate the influence of clopidogrel on inflammatory biomarkers and adhesion molecules in human endothelial cells and the role of nitric oxide (NO) in this process. METHODS: TNF-α-induced human umbilical vein endothelial cells (HUVEC) were exposed to clopidogrel. Gene expression and protein expression of ICAM-1, P-selectin, IL-8, IL-6, and MCP-1 were evaluated by qPCR, flux cytometry, or milliplex technology. Expression of endothelial nitric oxide synthase (NOS3) and NO release were also evaluated. Influence of clopidogrel was further evaluated in NOS3 downregulated HUVEC by RNAi. RESULTS: Clopidogrel at 20 µmol/L induced NO release in HUVEC after 24-hours treatment. Gene expressions of inflammatory markers IL-8 and MCP1 were reduced after clopidogrel treatment (P<.05); however, only MCP-1 remained reduced at protein level. IL-6 was not modified by clopidogrel treatment. Gene expression and protein expression of ICAM-1 were diminished by 24-hours clopidogrel exposure, whereas P-selectin was not modified. NOS3 downregulated HUVEC model revealed that ICAM-1 modification by clopidogrel is dependent of this via, whereas MCP-1 is modulated in an NO-independent form. CONCLUSIONS: Our results support new evidence for pleiotropic effects of clopidogrel on inflammation and endothelial function. Reduction in ICAM-1 and MCP-1 in human endothelium is an important extent of the use of this drug for treatment of cardiovascular diseases, and NO has an important role in this process.

Papilomatosis respiratoria recurrente en paciente pediátrico: reporte de un caso / Recurrent respiratory papillomatosis in a pediatric patient: case report

La variabilidad genética relacionada al sistema inmune del huésped ha sido propuesta como uno de los factores más influyentes en el desarrollo de enfermedades causadas por HPV. Caso clínico: Reportamos el caso de un niño de 5 años en cuyo estudio por disfonía crónica se encuentra papilomatosis laríngea probablemente adquirida por vía vertical durante el parto. El diagnóstico de papilomatosis laríngea se confirmó con una biopsia tras una primera cirugía orientada a remover los papilomas. Se utilizó el sistema de clasificación Derkay para evaluar la severidad de la papilomatosis. Se realizó genotipificación en biopsia demostrándose HPV-6. Posteriormente mediante tipificación de alelos HLA se demostró homocigosis para los alelos HLA-DQA1*0505, -DQB1*0301, -DRB1*1101. Conclusiones: Se necesitan estudios adicionales que permitan identificar los alelos HLA más prevalentes en población latina y su potencial asociación con la susceptibilidad genética en Papilomatosis Respiratoria Recurrente.

Genetic variability related to the host immune system has been proposed as one of the most influential factors in the development of diseases caused by HPV. Clinical case: We report the case of a 5-year-old child in whom chronic laryngeal papillomatosis, probably acquired vertically during labor, was detected. The diagnosis of laryngeal papillomatosis was confirmed with a biopsy after a first surgery to remove the papillomas. The Derkay classification system was used to assess the severity of papillomatosis. Biopsy genotyping was performed by demonstrating HPV-6. Later, HLA-DQA1 * 0505, -DQB1 * 0301, -DRB1 * 1101 alleles were homozygous for HLA allele typing. Conclusions: Further studies are needed to identify the most prevalent HLA alleles in the Latino population and their potential association with genetic susceptibility in Recurrent Respiratory Papillomatosis.

Effects of clopidogrel on inflammatory cytokines and adhesion molecules in human endothelial cells: Role of nitric oxide mediating pleiotropic effects

NTRODUCTION: Clopidogrel is commonly used in prevention and treatment of atherothrombosis. Some previous studies have suggested a pleiotropic effect of clopidogrel; however, when this drug causes platelet-independent effects on endothelial function remains unclear. AIMS: To evaluate the influence of clopidogrel on inflammatory biomarkers and adhesion molecules in human endothelial cells and the role of nitric oxide (NO) in this process. METHODS: TNF-α-induced human umbilical vein endothelial cells (HUVEC) were exposed to clopidogrel. Gene expression and protein expression of ICAM-1, P-selectin, IL-8, IL-6, and MCP-1 were evaluated by qPCR, flux cytometry, or milliplex technology. Expression of endothelial nitric oxide synthase (NOS3) and NO release were also evaluated. Influence of clopidogrel was further evaluated in NOS3 downregulated HUVEC by RNAi. RESULTS: Clopidogrel at 20 μmol/L induced NO release in HUVEC after 24-hours treatment. Gene expressions of inflammatory markers IL-8 and MCP1 were reduced after clopidogrel treatment (P<.05); however, only MCP-1 remained reduced at protein level. IL-6 was not modified by clopidogrel treatment. Gene expression and protein expression of ICAM-1 were diminished by 24-hours clopidogrel exposure, whereas P-selectin was not modified. NOS3 downregulated HUVEC model revealed that ICAM-1 modification by clopidogrel is dependent of this via, whereas MCP-1 is modulated in an NO-independent form...