EFFECT OF THE TEMPERATURE AND pH ON METHANOL RELEASE IN COFFEE BREW SWEETENED WITH ASPARTAME

Methanol is an alcohol which is metabolized to folmaldehyde in humans. It is a very toxic substance, responsible for blindness in cases of methanol intoxication. This study shows the production of methanol when aspartame is used to sweeten coffee brew. The temperature versus pH binomium was also tested. When assayed at 90 degrees C coffee brew+aspartame and aspartame solution showed an increase in methanol release of 17.8 and 19.3%, respectively, as compared with coffee brew. At 180 degrees C, the increase was 32.5 and 26.3%, respectively. Our data revealed a protective effect of the pH of coffee on the degradation of aspartame and formation of methanol; an important fact, mainly for specific populations that use aspartame, like diabetics.

Hypoxia modulates phenotype, inflammatory response, and leishmanial infection of human dendritic cells

Development of hypoxic areas occurs during infectious and inflammatory processes and dendritic cells (DCs) are involved in both innate and adaptive immunity in diseased tissues. Our group previously reported that macrophages exposed to hypoxia were infected with the intracellular parasite Leishmania amazonensis, but showed reduced susceptibility to the parasite. This study shows that although hypoxia did not alter human DC viability, it significantly altered phenotypic and functional characteristics. The expression of CD1a, CD80, and CD86 was significantly reduced in DCs exposed to hypoxia, whereas CD11c, CD14, CD123, CD49 and HLA-DR expression remained unaltered in DCs cultured in hypoxia or normoxia. DC secretion of IL-12p70, the bioactive interleukin-12 (IL-12), a cytokine produced in response to inflammatory mediators, was enhanced under hypoxia. In addition, phagocytic activity (Leishmania uptake) was not impaired under hypoxia, although this microenviroment induced infected DCs to reduce parasite survival, consequently controlling the infection rate. All these data support the notion that a hypoxic microenvironment promotes selective pressure on DCs to assume a phenotype characterized by pro-inflammatory and microbial activities in injured or inflamed tissues and contribute to the innate immune response.

Targeting the Acute Myeloid Leukemia Stem Cells

The idea that within the bulk of leukemic cells there are immature progenitors which are intrinsically resistant to chemotherapy and able to repopulate the tumor after treatment is not recent. Nevertheless, the term leukemia stem cells (LSCs) has been adopted recently to describe these immature progenitors based on the fact that they share the most relevant features of the normal hematopoetic stem cells (HSCs), i.e. the self-renewal potential and quiescent status. LSCs differ from their normal counterparts and from the more differentiated leukemic cells regarding the default status of pathways regulating apoptosis, cell cycle, telomere maintenance and transport pumps activity. In addition, unique features regarding the interaction of these cells with the microenvironment have been characterized. Therapeutic strategies targeting these unique features are at different stages of development but the reported results are promising. The aim of this review is, by taking acute myeloid leukemia (AML) as a bona fide example, to discuss some of the mechanisms used by the LSCs to survive and the strategies which could be used to eradicate these cells.

Membrane transporter proteins are involved in Trichophyton rubrum pathogenesis

Trichophyton rubrum is a dermatophyte responsible for the majority of human superficial mycoses. The functional expression of proteins important for the initial step and the maintenance of the infection process were identified previously in T. rubrum by subtraction suppression hybridization after growth in the presence of keratin. In this study, sequences similar to genes encoding the multidrug-resistance ATP-binding cassette (ABC) transporter, copper ATPase, the major facilitator superfamily and a permease were isolated, and used in Northern blots to monitor the expression of the genes, which were upregulated in the presence of keratin. A sequence identical to the TruMDR2 gene, encoding an ABC transporter in T rubrum, was isolated in these experiments, and examination of a T rubrum Delta TruMDR2 mutant showed a reduction in infecting activity, characterized by low growth on human nails compared with the wild-type strain. The high expression levels of transporter genes by T. rubrum in mimetic infection and the reduction in virulence of the Delta TruMDR2 mutant in a disease model in vitro suggest that transporters are involved in T. rubrum pathogenicity.

Developing human minor salivary glands: morphological parallel relation between the expression of TGF-beta isoforms and cytoskeletal markers of glandular maturation

Morphogenesis of salivary glands involves complex coordinated events. Synchronisation between cell proliferation, polarisation and differentiation, which are dependent on epithelial-mesenchymal interactions and on the microenvironment, is a requirement. Growth factors mediate many of these orchestrated biological processes and transforming growth factor-beta (TGF-beta) appear to be relevant. Using immunohistochemistry and immunofluorescence, we have mapped the distribution of TGF-beta 1, 2 and 3 and compared it with the expression of maturation markers in human salivary glands obtained from foetuses ranging from weeks 4 to 24 of gestation. TGF-beta 1 first appeared during canalisation stage in the surrounding mesenchyme and, in the more differentiated stages, was expressed in the cytoplasm of acinar cells throughout the adult gland. TGF-beta 2 was detected since the bud stage of the salivary gland. Its expression was observed in ductal cells and increased along gland differentiation, TGF-beta 3 was detected from the canalisation stage of the salivary gland, being weakly expressed on ductal cells, and it was the only factor detected on myoepithelial cells. The data suggest that TGF-beta have a role to play in salivary gland development and differentiation.

A role for COX2-derived PGE2 and PGE2-receptor subtypes in head and neck squamous carcinoma cell proliferation

The overexpression of cyclooxygenase (COX)-2 is a frequent event in squamous cell carcinomas of the head and neck (HNSCC), and non-steroidal anti-inflammatory drugs, which are potent inhibitors of COX-1 and COX-2, exert chemopreventive effects on HNSCC cancer development. COX-2 promotes the release of the pro-inflammatory mediator prostaglandin E2 (PGE2), which acts on its cell surface G protein-coupled receptors EP1, EP2, EP3, and EP4. Here, we investigated the role of PGE2 and its receptors in cellular proliferation in HNSCC. The expression of COX-2 and EP1-4 was examined in immortalized oral epithelial cells and in a representative panel of HNSCC cell lines, and based on these data EP1-EP3 and COX-2 expression were evaluated by immunohistochemistry in a large clinical sample collection using HNSCC tissue microarrays. The ability of selective COX-2 inhibition to block PGE2 secretion was measured by ELISA specific assays. The effects of PGE2 on cell proliferation were evaluated using PGE2, its stable analog, and EP2 and EP3-specific synthetic agonists. The results presented here show that HNSCC tumoral lesions and their derived cell lines constitutively express COX-2 and the EP1, EP2 and EP3 receptors for PGE2. HNSCC cells secrete PGE2, which can be suppressed by low concentrations of COX-2 selective inhibitors, without inhibiting cell proliferation. Exogenously added stable PGE2 and EP3-specific agonists induce DNA synthesis in all HNSCC cell lines tested. Overall, our study supports the emerging notion that PGE2 produced in the tumor microenvironment by the overexpression of COX-2 in tumoral and inflammatory cells may promote the growth of HNSCC cells in an autocrine and paracrine fashion by acting on PGE2 receptors that are widely expressed in most HNSCC cancer cells. In particular, our findings suggest that EP3 receptor may play a more prominent role in HNSCC cell growth promotion, thus providing a rationale for the future evaluation of this PGE2 receptor as a target for HNSCC prevention strategies. Published by Elsevier Ltd.

A possible mechanism of low molecular weight protein tyrosine phosphatase (LMW-PTP) activity modulation by glutathione action during human osteoblast differentiation

Objective: Low molecular weight protein tyrosine phosphatases (LMW-PTPs) are a family of enzymes strongly involved in the regulation of cell growth and differentiation. Since there is no information concerning the relationship between osteoblastic differentiation and LMW-PTP expression/activity, we investigated its involvement during human osteoblast-like cells (hFOB 1.19) differentiation. It is known that LMW-PTP is regulated by an elegant redox mechanism, so we also observed how the osteoblastic differentiation affected the reduced glutathione levels. Design: hFOB 1.19 cells were cultured in DMEM/F12 up to 35 days. The osteoblast phenotype acquisition was monitored by measuring alkaline phosphatase activity and mineralized nodule formation by Von Kossa staining. LMW-PTP activity and expression were measured using the p-nitrophenylphosphate as substrate and Western blotting respectively. Crystal violet assay determined the cell number in each experimental point. Glutathione level was determined by both HPLC and DNTB assays. Results: LMW-PTP modulation was coincident with the osteoblastic differentiation biomarkers, such as alkaline phosphatase activity and presence of nodules of mineralization in Vitro. Likewise LMW-PTP, the reduced glutathione-dependent microenvironment was modulated during osteoblastic differentiation. During this process, LMW-PTP expression/activity, as well as alkaline phosphatase and glutathione increased progressively up to the 21st day (p < 0.001) of culturing, decreasing thereafter. Conclusions: Our results clearly suggest that LMW-PTP expression/activity was rigorously modulated during osteoblastic differentiation, possibly in response to the redox status of the cells, since it seems to depend on suitable levels of reduced glutathione. in this way, we pointed out LMW-PTP as an important signaling molecule in osteoblast biology and bone formation. (C) 2009 Elsevier Ltd. All rights reserved.

Enhanced programmed death 1 (PD-1) and PD-1 ligand (PD-L1) expression in patients with actinic cheilitis and oral squamous cell carcinoma

PD-1 and PD-L1 can be involved in tumor escape, and little is known about the role of these molecules in oral tumors or pre-malignant lesions. In the present study, we investigated the expression of PD-1 and PD-L1 in the blood and lesion samples of patients with actinic cheilitis (AC) and oral squamous cell carcinoma (OSCC). Our results showed that lymphocytes from peripheral blood and tissue samples exhibited high expression of PD-1 in both groups analyzed. Patients with AC presented higher percentage as well as the absolute numbers of CD4(+)PD-1(+) and CD8(+)PD-1(+) lymphocytes in peripheral blood mononuclear cells (PBMC) than healthy individuals, while patients with OSCC presented an increased frequency of CD8(+)PD1(+) in PBMC when compared with controls. On the other hand, increased frequency of CD4(+) and CD8(+) T cells expressing PD-1(+) accumulate in samples from OSCC, and the expression of PD-L1 was intense in OSCC and moderate in AC lesion sites. Lower levels of IFN-gamma and higher levels of TGF-beta were detected in OSCC samples. Our data demonstrate that PD-1 and PD-L1 molecules are present in blood and samples of AC and OSCC patients. Further studies are required to understand the significance of PD-1 and PD-L1 in oral tumors microenvironment.

Tartrate-resistant acid phosphatase activity and glutathione levels are modulated during hFOB 1.19 osteoblastic differentiation

Tartrate-resistant acid phosphatase (TRAP) is a well-known marker of osteoclasts and bone resorption. Here we have investigated whether osteoblast-like cells (hFOB 1.19) present TRAP activity and how would be its pattern of expression during osteoblastic differentiation. We also observed how the osteoblastic differentiation affected the reduced glutathione levels. TRAP activity was measured using the p-nitrophenylphosphate substrate. The osteogenic potential of hFOB 1.19 cells was studied by measuring alkaline phosphatase activity and mineralized nodule formation. Oxidative stress was determined by HPLC and DNTB assays. TRAP activity and the reduced glutathione-dependent microenvironment were modulated during osteoblastic differentiation. During this phase, TRAP activity, as well as alkaline phosphatase and glutathione increased progressively up to the 21st day, decreasing thereafter. We demonstrate that TRAP activity is modulated during osteoblastic differentiation, possibly in response to the redox state of the cell, since it seemed to depend on suitable levels of reduced glutathione.