De-differentiation of primary human hepatocytes depends on the composition of specialized liver basement membrane

Despite the understanding of the importance of mitogen-activated protein (MAP) kinase activation in the stimulation of growth, little is known about the role of MAP kinase regulation during contact inhibited growth control. To investigate the role of the MAP kinase extracellular signal-regulated kinase (ERK) during the transition to a contact inhibited state, cultures of normal fibroblasts (BJ) were grown to different stages of confluency. The levels of MAP kinase phosphatase (MKP) expression and the amount of active ERK and MAP ERK kinase (MEK) in these cultures were assessed through western blot analysis and were compared to fibrosarcoma cell cultures (HT-1080), which lack contact inhibition. In normal fibroblasts, the amounts of active MEK and ERK decline at contact inhibition, concurrently with a rise in MKP-1, MKP-2, and MKP-3 protein levels. In contrast, fibrosarcoma cells appear to lack density-dependent regulation of the ERK pathway. Additionally, altering the redox environment of fibrosarcoma cells to a less reducing state, as seen during contact inhibition, results in increased MKP-1 expression. Taken together, these results suggest that the altered redox environment upon contact inhibition may contribute to the regulation of ERK inactivation by MKPs.     

In vitro antituberculosis activities of the constituents isolated from Haloxylon salicornicum

In vitro antituberculosis activities of fractions and pure compounds (1–20) including seven triterpenes, two alkaloids, two cycloheximide derivatives, two coumarins six sterol derivatives and a long chain alcohol, respectively, isolated from Haloxylon salicornicum were determined against Mycobecterium tuberculosis H37Rv. Actively growing cultures were tested by rapid colorimetric method while the stationary phase cultures were tested by drug exposure methods for bactericidal activity. The MIC values were found to be 50 μg/ml for compounds 15, 19 and 20 where as rest of the compounds invariably showed MIC value of 100 μg/ml against the logarithmic phase culture. These were compare to Isoniazid as a control drug. The compounds exhibited no activity against the stationary phase culture of M. tuberculosis H37Rv up to 200 μg/ml. Further studies are required to investigate the in vivo efficacies and activities of the compounds in combination with antimicrobials that are already being used for TB therapy.     

APUM23, a nucleolar Puf domain protein, is involved in pre-ribosomal RNA processing and normal growth patterning in Arabidopsis

Pumilio, an RNA-binding protein that contains tandemly repeated Puf domains, is known to repress translational activity in early embryogenesis and polarized cells of non-plant species. Although Pumilio proteins have been characterized in many eukaryotes, their role in plants is unknown. In the present study, we characterized an Arabidopsis Pumilio-encoding gene, APUM23. APUM23 is constitutively expressed, with higher levels in metabolically active tissues, and its expression is up-regulated in the presence of either glucose or sucrose. The T-DNA insertion mutants apum23-1 and apum23-2 showed slow growth, with serrated and scrunched leaves, an abnormal venation pattern, and distorted organization of the palisade parenchyma cells - a phenotype that is reminiscent of nucleolin and ribosomal protein gene mutants. Intracellular localization studies indicate that APUM23 predominantly localizes to the nucleolus. Based on this localization, rRNA processing was examined. In apum23, 35S pre-rRNA, and unprocessed 18S and 5.8S poly(A) rRNAs, accumulated without affecting the steady-state levels of mature rRNAs, indicating that APUM23 is involved in the processing and/or degradation of 35S pre-rRNA and rRNA maturation by-products. The apum23 mutant showed increased levels of 18S rRNA biogenesis-related U3 and U14 small nucleolar RNAs (snoRNAs) and accumulated RNAs within the nucleolus. Our data suggest that APUM23 plays an important role in plant development via rRNA processing.     

Insulin resistance,lipotoxicity and endothelial dysfunction

The number of people with the insulin-resistant conditions of type 2 diabetes mellitus (T2DM) and obesity has reached epidemic proportions worldwide. Eighty percent of people with T2DM will die from the complications of cardiovascular atherosclerosis. Insulin resistance is characterised by endothelial dysfunction, which is a pivotal step in the initiation/progression of atherosclerosis. A hallmark of endothelial dysfunction is an unfavourable imbalance between the bioavailability of the antiatherosclerotic signalling molecule nitric oxide (NO) and proatherosclerotic reactive oxygen species. In this review we discuss the mechanisms linking insulin resistance to endothelial dysfunction, with a particular emphasis on a potential role for a toxic effect of free fatty acids on endothelial cell homeostasis.     

Diesel Exhaust Particle Exposure In Vitro Alters Monocyte Differentiation and Function

Air pollution by diesel exhaust particles is associated with elevated mortality and increased hospital admissions in individuals with respiratory diseases such as asthma and chronic obstructive pulmonary disease. During active inflammation monocytes are recruited to the airways and can replace resident alveolar macrophages. We therefore investigated whether chronic fourteen day exposure to low concentrations of diesel exhaust particles can alter the phenotype and function of monocytes from healthy individuals and those with chronic obstructive pulmonary disease. Monocytes were purified from the blood of healthy individuals and people with a diagnosis of chronic obstructive pulmonary disease. Monocyte-derived macrophages were generated in the presence or absence of diesel exhaust particles and their phenotypes studied through investigation of their lifespan, cytokine generation in response to Toll like receptor agonists and heat killed bacteria, and expression of surface markers. Chronic fourteen day exposure of monocyte-derived macrophages to concentrations of diesel exhaust particles >10 µg/ml caused mitochondrial and lysosomal dysfunction, and a gradual loss of cells over time both in healthy and chronic obstructive pulmonary disease individuals. Chronic exposure to lower concentrations of diesel exhaust particles impaired CXCL8 cytokine responses to lipopolysaccharide and heat killed E. coli, and this phenotype was associated with a reduction in CD14 and CD11b expression. Chronic diesel exhaust particle exposure may therefore alter both numbers and function of lung macrophages differentiating from locally recruited monocytes in the lungs of healthy people and patients with chronic obstructive pulmonary disease.     

RNA deadenylation and decay in plants

In eukaryotic cells, RNA levels are tightly regulated in a spatio-temporal manner to maintain the protein levels necessary for cell growth, differentiation and division. To cope with developmental and rapid environmental changes, RNAs that are no longer required by the cell undergo degradation via the mRNA decay process. A number of players involved in RNA degradation pathways have been identified for the last two decades. A wealth of information about the process of mRNA deadenylation and decay in yeast and other model organisms is currently available; however, very limited information is available in plants, including Arabidopsis. Nevertheless, the efforts of various plant research groups are continuously extending our understanding of this complicated process. Here, we summarize our current knowledge of RNA decay in yeast and compare this information with the progress from Arabidopsis. This review will especially focus on the structure and function of the deadenylation complex, 5′ to 3′ exoribonuclease (XRN)-mediated decay pathways and the exosome-mediated 3′ to 5′ decay pathway in plants.     

Evaluation of the Antitumor Activity of Dacomitinib in Models of Human Bladder Cancer

Members of the human epidermal growth factor receptor (HER) family play a significant role in bladder cancer progression and may underlie the development of chemotherapy resistance. Dacomitinib is an irreversible tyrosine kinase inhibitor with structural specificity for the catalytic domains of epidermal growth factor receptor (EGFR), HER2 and HER4 that has exhibited vigorous efficacy against other solid tumors. We evaluated the antitumor activity of dacomitinib in human bladder cancer cell lines expressing varying levels of HER family receptors. These cell lines also were established as bladder cancer xenografts in nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice to assess dacomitinib activity in vivo. Significant cytotoxic and cytostatic effects were noted in cells expressing elevated levels of the dacomitinib target receptors with apoptosis and cell cycle arrest being the predominant mechanisms of antitumor activity. Cells expressing lower levels of HER receptors were much less sensitive to dacomitinib. Interestingly, dacomitinib was more active than either trastuzumab or cetuximab in vitro, and exhibited increased growth inhibition of bladder tumor xenografts compared with lapatinib. Pharmacodynamic effects of dacomitinib included decreased E-cadherin (E-cad) expression, reduction of EGFR and extracellular signal-regulated kinase (ERK) phosphorylation and reduced mitotic count. Dacomitinib also inhibited tumor growth in a chemotherapy-resistant xenograft and, when combined with chemotherapy in a sensitive xenograft, exhibited superior antitumor effects compared with individual treatments. Evaluation in xenograft-bearing mice revealed that this combination was broadly feasible and well tolerated. In conclusion, dacomitinib exhibited pronounced activity both as a single agent and when combined with chemotherapy in human bladder cancer models. Further investigation of dacomitinib in the preclinical and clinical trial settings is being pursued.