Somatic mutations in exocrine pancreatic tumors: association with patient survival.

KRAS mutations are major factors involved in initiation and maintenance of pancreatic tumors. The impact of different mutations on patient survival has not been clearly defined. We screened tumors from 171 pancreatic cancer patients for mutations in KRAS and CDKN2A genes. Mutations in KRAS were detected in 134 tumors, with 131 in codon 12 and only 3 in codon 61. The GGT>GAT (G12D) was the most frequent mutation and was present in 60% (80/134). Deletions and mutations in CDKN2A were detected in 43 tumors. Analysis showed that KRAS mutations were associated with reduced patient survival in both malignant exocrine and ductal adenocarcinomas (PDAC). Patients with PDACs that had KRAS mutations showed a median survival of 17 months compared to 30 months for those without mutations (log-rank P = 0.07) with a multivariate hazard ratio (HR) of 2.19 (95%CI 1.09-4.42). The patients with G12D mutation showed a median survival of 16 months (log-rank-test P = 0.03) and an associated multivariate HR 2.42 (95%CI 1.14-2.67). Although, the association of survival in PDAC patients with CDKN2A aberrations in tumors was not statistically significant, the sub-group of patients with concomitant KRAS mutations and CDKN2A alterations in tumors were associated with a median survival of 13.5 months compared to 22 months without mutation (log-rank-test P = 0.02) and a corresponding HR of 3.07 (95%CI 1.33-7.10). Our results are indicative of an association between mutational status and survival in PDAC patients, which if confirmed in subsequent studies can have potential clinical application

Evidence for Regulated Interleukin-4 Expression in Chondrocyte-Scaffolds under In Vitro Inflammatory Conditions

OBJECTIVE: To elucidate the anti-inflammatory and anabolic effects of regulated expression of IL-4 in chondrocyte-scaffolds under in vitro inflammatory conditions. METHODS: Mature articular chondrocytes from dogs (n = 3) were conditioned through transient transfection using pcDNA3.1.cIL-4 (constitutive) or pCOX-2.cIL-4 (cytokine-responsive) plasmids. Conditioned cells were seeded in alginate microspheres and rat-tail collagen type I matrix (CaReS®) to generate two types of tissue-engineered 3-dimensional scaffolds. Inflammatory arthritis was simulated in the packed chondrocytes through exogenous addition of recombinant canine (rc) IL-1β (100 ng/ml) plus rcTNFα (50 ng/ml) in culture media for 96 hours. Harvested cells and culture media were analyzed by various assays to monitor the anti-inflammatory and regenerative (anabolic) properties of cIL-4. RESULTS: cIL-4 was expressed from COX-2 promoter exclusively on the addition of rcIL-1β and rcTNFα while its expression from CMV promoter was constitutive. The expressed cIL-4 downregulated the mRNA expression of IL-1β, TNFα, IL-6, iNOS and COX-2 in the cells and inhibited the production of NO and PGE(2) in culture media. At the same time, it up-regulated the expression of IGF-1, IL-1ra, COL2a1 and aggrecan in conditioned chondrocytes in both scaffolds along with a diminished release of total collagen and sGAG into the culture media. An increased amount of cIL-4 protein was detected both in chondrocyte cell lysate and in concentrated culture media. Neutralizing anti-cIL-4 antibody assay confirmed that the anti-inflammatory and regenerative effects seen are exclusively driven by cIL-4. There was a restricted expression of IL-4 under COX-2 promoter possibly due to negative feedback loop while it was over-expressed under CMV promoter (undesirable). Furthermore, the anti-inflammatory /anabolic outcomes from both scaffolds were reproducible and the therapeutic effects of cIL-4 were both scaffold- and promoter-independent. CONCLUSIONS: Regulated expression of therapeutic candidate gene(s) coupled with suitable scaffold(s) could potentially serve as a useful tissue-engineering tool to devise future treatment strategies for osteoarthritis

Low frequency of TERT promoter mutations in gastrointestinal stromal tumors (GISTs).

Somatic mutations in the promoter region of telomerase reverse transcriptase (TERT) gene, mainly at positions c. − 124 and c. − 146 bp, are frequent in several human cancers; yet its presence in gastrointestinal stromal tumor (GIST) has not been reported to date. Herein, we searched for the presence and clinicopathological association of TERT promoter mutations in genomic DNA from 130 bona fide GISTs. We found TERT promoter mutations in 3.8% (5/130) of GISTs. The c. − 124C4T mutation was the most common event, present in 2.3% (3/130), and the c. − 146C4T mutation in 1.5% (2/130) of GISTs. No significant association was observed between TERT promoter mutation and patient’s clinicopathological features. The present study establishes the low frequency (4%) of TERT promoter mutations in GISTs. Further studies are required to confirm our findings and to elucidate the hypothetical biological and clinical impact of TERT promoter mutation in GIST pathogenesis.This project was partially supported by Barretos Cancer Hospital internal research funds (PAIP) and CNPq Universal Grant (476192/2013-7) to RMR. NCC is a recipient of an FAPESP Doctoral Fellowship (2013/25787-3). Further funding from the project ‘Microenvironment, metabolism and cancer’ that was partially supported by Programa Operacional Regional do Norte (ON.2—O Novo Norte) under the Quadro de Referência Estratégico Nacional (QREN) and the Fundo Europeu de Desenvolvimento Regional (FEDER). IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology and Higher Education that is partially supported by the FCT

Acid Stability of the Hemagglutinin Protein Regulates H5N1 Influenza Virus Pathogenicity

Highly pathogenic avian influenza viruses of the H5N1 subtype continue to threaten agriculture and human health. Here, we use biochemistry and x-ray crystallography to reveal how amino-acid variations in the hemagglutinin (HA) protein contribute to the pathogenicity of H5N1 influenza virus in chickens. HA proteins from highly pathogenic (HP) A/chicken/Hong Kong/YU562/2001 and moderately pathogenic (MP) A/goose/Hong Kong/437-10/1999 isolates of H5N1 were found to be expressed and cleaved in similar amounts, and both proteins had similar receptor-binding properties. However, amino-acid variations at positions 104 and 115 in the vestigial esterase sub-domain of the HA1 receptor-binding domain (RBD) were found to modulate the pH of HA activation such that the HP and MP HA proteins are activated for membrane fusion at pH 5.7 and 5.3, respectively. In general, an increase in H5N1 pathogenicity in chickens was found to correlate with an increase in the pH of HA activation for mutant and chimeric HA proteins in the observed range of pH 5.2 to 6.0. We determined a crystal structure of the MP HA protein at 2.50 Å resolution and two structures of HP HA at 2.95 and 3.10 Å resolution. Residues 104 and 115 that modulate the acid stability of the HA protein are situated at the N- and C-termini of the 110-helix in the vestigial esterase sub-domain, which interacts with the B loop of the HA2 stalk domain. Interactions between the 110-helix and the stalk domain appear to be important in regulating HA protein acid stability, which in turn modulates influenza virus replication and pathogenesis. Overall, an optimal activation pH of the HA protein is found to be necessary for high pathogenicity by H5N1 influenza virus in avian species

Telomerase promoter mutations in cancer: an emerging molecular biomarker?

João Vinagre, Vasco Pinto and Ricardo Celestino contributed equally to the manuscript.Cell immortalization has been considered for a long time as a classic hallmark of cancer cells. Besides telomerase reactivation, such immortalization could be due to telomere maintenance through the “alternative mechanism of telomere lengthening” (ALT) but the mechanisms underlying both forms of reactivation remained elusive. Mutations in the coding region of telomerase gene are very rare in the cancer setting, despite being associated with some degenerative diseases. Recently, mutations in telomerase (TERT) gene promoter were found in sporadic and familial melanoma and subsequently in several cancer models, notably in gliomas, thyroid cancer and bladder cancer. The importance of these findings has been reinforced by the association of TERT mutations in some cancer types with tumour aggressiveness and patient survival. In the first part of this review, we summarize the data on the biology of telomeres and telomerase, available methodological approaches and non-neoplastic diseases associated with telomere dysfunction. In the second part, we review the information on telomerase expression and genetic alterations in the most relevant types of cancer (skin, thyroid, bladder and central nervous system) on record, and discuss the value of telomerase as a new biomarker with impact on the prognosis and survival of the patients and as a putative therapeutic target

New developments in osteoarthritis. Posttraumatic osteoarthritis: pathogenesis and pharmacological treatment options

Joint trauma can lead to a spectrum of acute lesions, including osteochondral fractures, ligament or meniscus tears and damage to the articular cartilage. This is often associated with intraarticular bleeding and causes posttraumatic joint inflammation. Although the acute symptoms resolve and some of the lesions can be surgically repaired, joint injury triggers a chronic remodeling process in cartilage and other joint tissues that ultimately manifests as osteoarthritis in a majority of cases. The objective of the present review is to summarize information on pathogenetic mechanisms involved in the acute and chronic consequences of joint trauma and discuss potential pharmacological interventions. The focus of the review is on the early events that follow joint trauma since therapies for posttraumatic joint inflammation are not available and this represents a unique window of opportunity to limit chronic consequences

Mechanisms of human telomerase reverse transcriptase (hTERT) regulation: clinical impacts in cancer

Background Limitless self-renewal is one of the hallmarks of cancer and is attained by telomere maintenance, essentially through telomerase (hTERT) activation. Transcriptional regulation of hTERT is believed to play a major role in telomerase activation in human cancers. Main body The dominant interest in telomerase results from its role in cancer. The role of telomeres and telomere maintenance mechanisms is well established as a major driving force in generating chromosomal and genomic instability. Cancer cells have acquired the ability to overcome their fate of senescence via telomere length maintenance mechanisms, mainly by telomerase activation. hTERT expression is up-regulated in tumors via multiple genetic and epigenetic mechanisms including hTERT amplifications, hTERT structural variants, hTERT promoter mutations and epigenetic modifications through hTERT promoter methylation. Genetic (hTERT promoter mutations) and epigenetic (hTERT promoter methylation and miRNAs) events were shown to have clinical implications in cancers that depend on hTERT activation. Knowing that telomeres are crucial for cellular self-renewal, the mechanisms responsible for telomere maintenance have a crucial role in cancer diseases and might be important oncological biomarkers. Thus, rather than quantifying TERT expression and its correlation with telomerase activation, the discovery and the assessment of the mechanisms responsible for TERT upregulation offers important information that may be used for diagnosis, prognosis, and treatment monitoring in oncology. Furthermore, a better understanding of these mechanisms may promote their translation into effective targeted cancer therapies. Conclusion Herein, we reviewed the underlying mechanisms of hTERT regulation, their role in oncogenesis, and the potential clinical applications in telomerase-dependent cancers.info:eu-repo/semantics/publishedVersio

The evaluation of acoustic characteristic performance on natural sound absorbing materials from cogon grass waste

In the past few decades, synthetic fibers are been used widely in the field of sound absorption due to their superior characteristics such as durable and chemical resistant. However, there are several disadvantages of synthetic fibers such as non-biodegradability and hazards to the health of human. In this research, the natural sound absorber from cogon grass was investigated. The objective of the research was to evaluate the performance of cogon grass physical characteristics on its acoustical behavior, to evaluate the effect of sodium hydroxide (NaOH) treatment times on physical and acoustical characteristics of cogon grass, to investigate the decay effects after it was left over for twelve months and lastly to compare and verify the acoustical results with theoretical models based on (Delany-Bazley and Miki Model). The measurement of acoustical characteristics which are sound absorption coefficient (SAC) and noise reduction coefficient (NRC) were done by using impedance tube method (ITM). The samples of cogon grass were tested in a way of the untreated and treated with NaOH in varied soaked hours which are one, two, three, four and five hours. Scanning electron microscope (SEM) and density kit were used to investigate physical characteristics. The research confirmed that physical characteristics of tortuosity and airflow resistivity values tend to increase with the increment of treatment times, but the density and porosity tend to decrease. Untreated samples were tested with varied thicknesses of 10, 20, 30, 40 and 50mm. The results show SAC value increases when the thickness of the sample was increased. Treated samples results show the least treated sample (1 hour) reached the maximum SAC value and indicated the highest value of NRC which is 0.50. The results also show a reduction in sound absorption value after the samples were left for twelve months. Verification parts demonstrated that Delany-Bazley and Miki Model can predict approximately pattern compared with ITM results because of the theoretical models are developed by a simple empirical model approach. Overall, cogon grass samples have the good characteristics to be an acoustic material component