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Secretory proteins are an essential component of interorgan communication networks that regulate animal physiology. Current approaches for identifying secretory proteins from specific cell and tissue types are largely limited to in vitro or ex vivo models which often fail to recapitulate in vivo biology. As such, there is mounting interest in developing in vivo analytical tools that can provide accurate information on the origin, identity, and spatiotemporal dynamics of secretory proteins. Here, we describe iSLET (in situ Secretory protein Labeling via ER-anchored TurboID) which selectively labels proteins that transit through the classical secretory pathway via catalytic actions of Sec61b-TurboID, a proximity labeling enzyme anchored in the ER lumen. To validate iSLET in a whole-body system, we express iSLET in the mouse liver and demonstrate efficient labeling of liver secretory proteins which could be tracked and identified within circulating blood plasma. Furthermore, proteomic analysis of the labeled liver secretome enriched from liver iSLET mouse plasma is highly consistent with previous reports of liver secretory protein profiles. Taken together, iSLET is a versatile and powerful tool for studying spatiotemporal dynamics of secretory proteins, a valuable class of biomarkers and therapeutic targets. The in vivo identification of proteins secreted from a specific cell type or tissue remains challenging. Here, the authors develop a proximity labeling-based method to selectively label secreted proteins and combine it with proteomics to identify liver secretory proteins in mouse plasma. [ABSTRACT FROM AUTHOR]

Bacteria can form biofilms that consist of multicellular communities embedded in an extracellular matrix (ECM). In Bacillus subtilis, the main protein component of the ECM is the functional amyloid TasA. Here, we study further the roles played by TasA in B. subtilis physiology and biofilm formation on plant leaves and in vitro. We show that ΔtasA cells exhibit a range of cytological symptoms indicative of excessive cellular stress leading to increased cell death. TasA associates to the detergent-resistant fraction of the cell membrane, and the distribution of the flotillin-like protein FloT is altered in ΔtasA cells. We propose that, in addition to a structural function during ECM assembly and interactions with plants, TasA contributes to the stabilization of membrane dynamics as cells enter stationary phase. The amyloid protein TasA is a main component of the extracellular matrix in Bacillus subtilis biofilms. Here the authors show that, in addition to a structural function during biofilm assembly and interactions with plants, TasA contributes to the stabilization of membrane dynamics during stationary phase. [ABSTRACT FROM AUTHOR]

The objective of this study was to evaluate the effects of photodynamic inactivation (PDI) on Candida albicans biofilms, evaluating its effects on gene expression of ALS3, HWP1, BCR1, TEC1, CPH1, and EFG1 by yeast. Three samples of C. albicans were used in this study: a clinical sample from a patient with HIV (39S), a clinical sample from a patient with denture stomatitis lesion (Ca30), and a standard strain ATCC 18804. The quantification of gene expression was related to the production of those genes in the samples referred above using quantitative polymerase chain reaction (qPCR) assay in real time. The photosensitizer methylene blue at 300 uM and erythrosine at 400 uM, sensitized with low-power laser (visible red, 660 nm) and green LED (532 nm), respectively, were used for PDI. Four groups of each sample and PDI protocol were evaluated: (a) P+L+: sensitization with the photosensitizer and irradiation with light, (b) P+L-: only treatment with the photosensitizer, (c) P-L+: only irradiation with light, and (d) P-L-: without sensitization with the dye and absence of light. The results were analyzed by t test, with a significance level of 5%. The photodynamic inactivation was able to reduce the expression of all genes for both treatments, laser and LED. The fold-decrease for the genes ALS3, HWP1, BCR1, TEC1, CPH1, and EFG1 were 0.73, 0.39, 0.77, 0.71, 0.67, and 0.60 for laser, respectively, and 0.66, 0.61, .050, 0.43, 0.54, and 0.66 for LED, respectively. It could be concluded that PDI showed a reduction in the expression of C. albicans genes, suggesting its virulence decrease. [ABSTRACT FROM AUTHOR]

Background: Curcumin, a pleiotropic substance used for centuries in traditional medicine, exhibits antioxidant, anti-inflammatory and antiproliferative efficacy against various tumours, but the role of curcumin in gastroprotection is little studied. We determined the effect of curcumin against gastric haemorrhagic lesions induced by 75% ethanol and alterations in gastric blood flow (GBF) in rats with cyclooxygenase-1 (COX-1) and COX-2 activity inhibited by indomethacin, SC-560 or rofecoxib, inhibited NO-synthase activity, capsaicin denervation and blockade of TRPV1 receptors by capsazepine.Methods: One hour after ethanol administration, the gastric mucosal lesions were assessed by planimetry, the GBF was examined by H2 gas clearance, plasma gastrin was determined by radioimmunoassay, and the gastric mucosal mRNA expression of Cdx-2, HIF-1α, HO-1 and SOD 2 was analysed by RT-PCR.Results: Curcumin, in a dose-dependent manner, reduced ethanol-induced gastric lesions and significantly increased GBF and plasma gastrin levels. Curcumin-induced protection was completely reversed by indomethacin and SC-560, and significantly attenuated by rofecoxib, L-NNA, capsaicin denervation and capsazepine. Curcumin downregulated Cdx-2 and Hif-1α mRNA expression and upregulated HO-1 and SOD 2, and these effects were reversed by L-NNA and further restored by co-treatment of L-NNA with L-arginine.Conclusions: Curcumin-induced protection against ethanol damage involves endogenous PG, NO, gastrin and CGRP released from sensory nerves due to activation of the vanilloid TRPV1 receptor. This protective effect can be attributed to the inhibition of HIF-1α and Cdx-2 expression and the activation of HO-1 and SOD 2 expression. [ABSTRACT FROM AUTHOR]

Low-level laser therapy (LLLT) has been used for more than 30 years to heal wounds. In recent years, LLLT or photostimulation has been indicated as an effective tool for regenerative and dental medicine by using monochromatic light. The aim of this study is to indicate the usability of plasma arc light source for bone regeneration. This is why we used polychromatic light source providing effective wavelengths in the range of 590-1500 nm for cellular response and investigated photostimulation effects on osteogenic differentiation of human mesenchymal stem cells (hMSCs) seeded on 3D silk scaffolds. Cellular responses were examined by using cell culture methods in terms of proliferation, differentiation, and morphological analyses. The results showed that photostimulation with a polychromatic light source (applied for 5 min from the 3rd day after seeding up to the 28th day in 2-day intervals with 92-mW/cm2 power from 10-cm distance to the cells) enhanced osteogenic differentiation of hMSCs according to higher alkaline phosphatase (ALP) activity, collagen and calcium content, osteogenic gene expressions, and matrix mineralization. In conclusion, we suggest that the plasma arc light source that was used here has a great potential for bone regeneration. [ABSTRACT FROM AUTHOR]

Conventional therapeutic approaches to post-transplant lymphoproliferative disorder (PTLD) occurring after solid-organ transplantation have shown only limited success in achieving durable response. Key factors driving the pathogenesis of PTLD include Epstein-Barr virus (EBV) reactivation and impaired immune surveillance due to prolonged immune suppression. Thus, EBV-specific cytotoxic T lymphocytes (EBV-CTLs) have emerged as an alternative therapeutic approach for the treatment of EBV-associated PTLD by enhancing EBV-specific immunity. We evaluated the safety and efficacy of EBV latent membrane proteins (LMP)-1- and 2-specific CTLs in two PTLD patients at high risk for relapse. Following diagnosis, patients were initially treated with a combination of chemotherapy and/or radiotherapy. Patients then received a total of eight doses of 2 × 107 EBV-CTLs/m2. Following initial therapy, both patients achieved complete remission confirmed by FDG-PET/CT imaging. Post-remission therapy using adoptive transfer of EBV-CTLs was safe without immediate or late toxicities. Infusion of EBV-CTLs led to an overall reduction in plasma EBV levels in the peripheral blood, which was associated with long-term remission of both patients during a follow-up of more than 65 months. Further prospective studies with larger number of patients will be needed to confirm the role of EBV-CTLs as post-remission therapy in high-risk PTLD. [ABSTRACT FROM AUTHOR]

Bone disease is the hallmark of multiple myeloma (MM), a hematological malignancy characterized by osteolytic lesions due to a severe uncoupled and unbalanced bone remodeling with pronounced osteoblast suppression. Bone metastasis is also a frequent complication of solid tumors including advanced breast or prostate cancer. In the past years, the ubiquitin-proteasome pathway has been proved critical in regulating the balance between bone formation and bone resorption. Proteasome inhibitors (PIs) are a new class of drugs, currently used in the treatment of MM, that affect both tumor cells and bone microenvironment. Particularly, PIs stimulate osteoblast differentiation by human mesenchymal stromal cells and increase bone regeneration in mice. Interestingly, in vitro data indicate that PIs block MM-induced osteoblast and osteocyte cell death by targeting both apoptosis and autophagy. The preclinical data are supported by the following effects observed in MM patients treated with PIs: increase of bone alkaline phosphatase levels, normalization of the markers of bone turnover, and reduction of the skeletal-related events. Moreover, the histomorphometric data indicate that the treatment with bortezomib stimulates osteoblast formation and maintains osteocyte viability in MM patients. This review updates the evidence on the effects of PIs on bone remodeling and on cancer-induced bone disease while focusing on MM bone disease. [ABSTRACT FROM AUTHOR]

Background: Glucocorticosteroid (GC) is one of the most effective drugs available for the treatment of primary nephrotic syndrome (PNS) in children. However, some patients show little or no response to GC. The purpose of our research was to observe and describe the different levels of histone deacetylase-2 (HDAC2) expression in peripheral blood lymphocytes in children with PNS compared with various responses to the GC treatment, with the primary aim to assess the correlation between HDAC2 and GC resistance in PNS children. Methods: Forty-eight patients with PNS suffering from their first attack prior to GC treatment were chosen as subjects. They were divided into two groups, those who had steroid-sensitive nephrotic syndrome (SSNS; n = 25) and those with steroid-resistant NS (SRNS; n = 23), according to their response to a 6-week course of oral prednisone. Twenty healthy children from the Physical Examination Center in the hospital served as the control group; Peripheral blood was collected at different time points prior to GC treatment and after regular therapy. RT-PCR, western blot, and enzyme-linked immunosorbent assay (ELISA) techniques were adopted to analyze HDAC2 mRNA, protein expression, and activity, respectively, in peripheral blood lymphocytes. The level of interleukin-8 (IL-8) in serum was measured by an ELISA. Results: Prior to GC treatment, HDAC2 expression level and activity were lower in the SRNS group than in the SSNS and control group. A statistically significant difference in HDAC2 expression and activity were observed after GC treatment between these groups, with HDAC2 expression and activity lower in the SRNS group than in the SSNS and control groups. In the SSNS group, the expression and activity of HDAC2 were higher following GC treatment than prior to GC treatment. There was a clear difference in HDAC2 expression and activity of SRNS at the different time points. No statistically significant difference was found between the two groups. The pre-treatment and post-treatment serum IL-8 levels in the SRNS group were significantly higher than those in the SSNS group. HDAC2 from children with PNS before GC treatment and after regular therapy for 6 weeks was negatively correlated with serum IL-8 level. Conclusion: The GC effect was influenced by the HDAC2 expression and activity, leading to decreased serum IL-8 levels in children with PNS. HDAC2 seems to be one of the markers of GC resistance in children with PNS. [ABSTRACT FROM AUTHOR]

Background: Tubulointerstitial nephritis (TIN) is an inflammatory disease of unknown pathogenesis. To evaluate a possible role of regulatory T cells (Tregs) in the pathophysiology of TIN with (TINU) and without uveitis, we investigated the presence and quantity of FOXP3 T regulatory lymphocytes in diagnostic kidney biopsies from pediatric patients. Methods: A total of 33 patients (14 TIN and 19 TINU) were enrolled. The quantity of CD4, FOXP3 and double-positive T cells in formalin-fixed kidney biopsies was determined using double label immunohistochemistry with anti-human CD4 and FOXP3 antibodies. Results: FOXP3 staining was successful in all 33 patients. In patients with chronic uveitis, the density of FOXP3 cells was significantly lower ( p = 0.046) than in TIN patients without uveitis or with uveitis lasting <3 months. CD4 staining was successful in 23 patients. The density of all lymphocytes (CD4, CD4FOXP3 and FOXP3 cells) was significantly lower ( p = 0.023) in patients with chronic uveitis than in other patients. Conclusions: FOXP3 T cells are present in kidney biopsy samples from TIN and TINU patients. In patients with chronic uveitis, the density of FOXP3 T cells is significantly lower than in other patients, suggesting a different pathomechanism for these clinical conditions. [ABSTRACT FROM AUTHOR]

The SOX4 transcription factor is involved in various cellular processes, such as embryonic development and differentiation. Deregulated expression of Sox4 in several human cancers has been reported to date, but its biological functions in the progression of osteosarcoma remain unclear. In this study, we found that the expression levels of SOX4 protein were significantly higher in high-grade osteosarcoma tissues and metastatic osteosarcoma tissues. Its overexpression was associated with poor prognosis in osteosarcoma. Knockdown of the SOX4 gene in the osteosarcoma cell lines resulted in decreased cell proliferation, migration, invasion, and induced apoptosis. After SOX4 gene silencing, the protein expression levels of Bax, Caspase-3, and P53 in osteosarcoma cells were significantly elevated, while the protein expression levels of Bcl-2, MMP2, and MMP9 were obviously decreased. In vivo analysis in nude mice further confirmed that knockdown of SOX4 suppressed tumor growth. In conclusion, SOX4 appears to be an important tumor oncogene in the regulation of osteosarcoma cell proliferation, apoptosis, and invasion, and it may be a potential target for effective osteosarcoma therapy. [ABSTRACT FROM AUTHOR]

Background: Meta-analysis shows that women with diabetes have a 20% increased risk of breast cancer and also an increased risk for distant metastasis and mortality. The molecular mechanisms for distant metastasis and mortality in breast cancer patients with diabetes are not very well understood.Methods: We compared the effect of physiological (5 mM) and diabetic (10 mM) levels of glucose on malignant breast epithelial cell invasion and stemness capabilities. We performed microRNA array to determine the dysregulated microRNAs in hyperglycaemic conditions and performed functional and molecular analysis of the gene targets.Results: Hyperglycaemia leads to hyperactivation of cancer stem cell pool and enhances invasive ability of breast cancer cells. MiR-424 seems to be a key regulator of cancer cell stemness and invasion. Knockdown of miR-424 in cancer cells under euglycaemic conditions leads to enhanced invasion and stem cell activity, whereas ectopic expression of miR-424 in cancer cells under hyperglycaemic conditions results in suppressed invasion and stem cell activity. Cdc42, a target of miR-424, influences cancer stem cell activity by positively regulating prdm14 through activation of pak1 (p-21-activated kinase 1) and stat5.Conclusions: Our findings establish miR-424→︀cdc42→︀prdm14 axis as a key molecular signalling cascade that might influence breast cancer progression in diabetic patients through hyperactivation of cancer stem cells. [ABSTRACT FROM AUTHOR]

Purpose: Regular physical activity induces oxidative stress but also causes adaptations in antioxidant defences including the nuclear factor κB (NF-κB) pathway, which activates target genes related to antioxidant defences such as uncoupling proteins (UCPs), and mitochondrial biogenesis mediated by peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). The aim of the study was to determine the effect of long-term training and acute exercise on oxidant/antioxidant status and the expression of mitochondrial biogenesis genes in peripheral blood mononuclear cells (PBMCs).Methods: Twelve professional football players performed an 8-week exercise programme comprising a daily 2-h football training session. Blood samples were taken before and after the training season.Results: The results reported a significant increase in antioxidant protein levels and in mitochondrial proteins in resting conditions after the 8-week training period. PGC1α, UCP-2 and mitofusin 2 protein levels also increased after acute exercise compared to pre-exercise levels. After the training, the expression of PGC1α, cytochrome c oxidase subunit IV and mitochondrial NADH dehydrogenase subunit 5 messenger RNA (mRNA) significantly augmented after the acute physical activity compared to pre-exercise levels; while no changes occurred in these mRNA in basal conditions. NF-κB activation and ROS production reported a significant increase after acute exercise.Conclusions: Training increases the levels of proteins related to mitochondrial biogenesis and improves the antioxidant capabilities of mitochondria in PBMCs among well-trained football players. Acute exercise may act as an inducer of mitochondrial biogenesis through NF-κB activation and PGC1α gene expression. [ABSTRACT FROM AUTHOR]

Clonorchiasis remains a nonnegligible public health problem in endemic areas. Cysteine protease of Clonorchis sinensis ( CsCP) plays indispensable roles in the parasitic physiology and pathology, and has been exploited as a promising drug and vaccine candidate. In recent years, development of spore-based vaccines against multiple pathogens has attracted many investigators' interest. In previous studies, the recombinant Escherichia coli (BL21) and Bacillus subtilis spores expressing CsCP have been successfully constructed, respectively. In this study, the immune effects of CsCP protein purified from recombinant BL21 (r CsCP) and B. subtilis spores presenting CsCP ( B.s- CsCP) in Balb/c mice model were conducted with comparative analysis. Levels of specific IgG, IgG1 and IgG2a were significantly increased in sera from both r CsCP and B.s- CsCP intraperitoneally immunized mice. Additionally, recombinant spores expressing abundant fusion CsCP (0.03125 pg/spore) could strongly enhance the immunogenicity of CsCP with significantly higher levels of IgG and isotypes. Compared with r CsCP alone, intraperitoneal administration of mice with spores expressing CsCP achieved a better effect of fighting against C. sinensis infection by slowing down the process of fibrosis. Our results demonstrated that a combination of Th1/Th2 immune responses could be elicited by r CsCP, while spores displaying CsCP prominently induced Th1-biased specific immune responses, and the complex cytokine network maybe mediates protective immune responses against C. sinensis. This work further confirmed that the usage of B. subtilis spores displaying CsCP is an effective way to against C. sinensis. [ABSTRACT FROM AUTHOR]

Reflux esophagitis damages the squamous epithelium that normally lines the esophagus, and promotes replacement of the damaged squamous lining by the intestinal metaplasia of Barrett's esophagus, the precursor of esophageal adenocarcinoma. Therefore, to prevent the development of Barrett's metaplasia and esophageal adenocarcinoma, the pathogenesis of reflux esophagitis must be understood. We have reported that reflux esophagitis, both in a rat model and in humans, develops as a cytokine-mediated inflammatory injury (i.e., cytokine sizzle), not as a caustic chemical injury (i.e., acid burn), as traditionally has been assumed. Moreover, reflux induces activation of hypoxia inducible factor (HIF)-2α, which enhances the transcriptional activity of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) causing increases in pro-inflammatory cytokines and in migration of T lymphocytes, an underlying molecular mechanism for this cytokine-mediated injury. In some individuals, reflux esophagitis heals with Barrett's metaplasia. A number of possibilities exist for the origin of the progenitor cells that give rise to this intestinal metaplasia including those of the esophagus, the proximal stomach, or the bone marrow. However, intestinal cells are not normally found in the esophagus, the stomach, or the bone marrow. Thus, the development of Barrett's intestinal metaplasia must involve some molecular reprogramming of key developmental transcription factors within the progenitor cell, a process termed transcommitment, which may be initiated by the noxious components of the gastric refluxate. This review will highlight recent studies on the pathogenesis of reflux esophagitis and on reflux-related molecular reprogramming of esophageal squamous epithelial cells in the pathogenesis of Barrett's metaplasia. [ABSTRACT FROM AUTHOR]

Purpose: We investigated the role of human platelet gel (PG) embedded within gelatin (Gel) scaffold on healing of critical-sized radial bone defects in rats.Methods: Twenty-five Sprague-Dawley rats were randomly divided into five equal groups. In each animal, critical-sized 5-mm bone defects were created in the radial bones of both forelimbs (n = 10/group). The defects were then either left untreated or filled with autograft, Gel, PG or Gel-PG. Before euthanasia, the healing defects were evaluated radiologically and clinically. The animals were euthanized after eight weeks and their radial bones evaluated by radiography, computed tomography (CT) scan, histology, biomechanical testing and ultrastructural evaluations.Results: PG implantation significantly increased cellular differentiation, osteoblastic proliferation and consequently new bone formation so that those defects treated with PG showed superior structural and biomechanical properties to the Gel and PG-Gel-treated defects. The PG-treated defects had radiological, morphological and mechanical properties closely comparable with those of the autograft group. In contrast, in the PG-Gel group, Gel significantly reduced the beneficial effects of PG on bone healing.Conclusions: Human PG had beneficial effects on bone regeneration, while combination of PG and Gel had no remarkable beneficial effect. Therefore, PG when used alone can be regarded as a promising osteoinductive and osteoconductive option in bone tissue engineering applications. [ABSTRACT FROM AUTHOR]

The transcription factor c-MYB is a well-known marker of undifferentiated cells such as haematopoietic cell precursors, but recently it has also been observed in differentiated cells that produce hard tissues. Our previous findings showed the presence of c-MYB in intramembranous bones and its involvement in the chondrogenic steps of endochondral ossification, where the up-regulation of early chondrogenic markers after c-myb overexpression was observed. Since we previously detected c-MYB in osteoblasts, we aimed to analyse the localisation of c-MYB during later stages of endochondral bone formation and address its function during bone matrix production. c-MYB-positive cells were found in the chondro-osseous junction zone in osteoblasts of trabecular bone as well as deeper in the zone of ossification in cells of spongy bone. To experimentally evaluate the osteogenic potential of c-MYB during endochondral bone formation, micromasses derived from embryonic mouse limb buds were established. Nuclear c-MYB protein expression was observed in long-term micromasses, especially in the areas around nodules. c-myb overexpression induced the expression of osteogenic-related genes such as Bmp2, Comp, Csf2 and Itgb1. Moreover, alizarin red staining and osteocalcin labelling promoted mineralised matrix production in c-myb-overexpressing cultures, whereas downregulation of c-myb by siRNA reduced mineralised matrix production. In conclusion, c-Myb plays a role in the osteogenesis of long bones by inducing osteogenic genes and causing the enhancement of mineral matrix production. This action of the transcription factor c-Myb might be of interest in the future for the establishment of novel approaches to tissue regeneration. [ABSTRACT FROM AUTHOR]

Purpose: The present work was aimed at the development of prospective positron emission tomography (PET) agents for infection imaging. Towards this aim, ubiquicidin (UBI) fragments conjugated with the macrocyclic NODAGA chelator were radiolabeled with Ga-68 and evaluated.Procedures: Conformations of custom synthesized NODAGA-UBI (29-41) and NODAGA-UBI (31-38) conjugates were compared with UBI (29-41) by circular dichroism (CD) spectroscopy. Optimization of labeling of NODAGA conjugates of UBI peptides with Ga-68 was performed and quality control analysis was carried out by chromatography techniques. In vitro uptake of [68Ga] NODAGA-UBI (29-41) and [68Ga]NODAGA-UBI (31-38) was studied in Staphylococcus aureus cells. In vivo distribution of [68Ga]GaCl3 and [68Ga]NODAGA-UBI complexes was performed in normal Swiss mice.Results: Conformations of NODAGA-UBI (29-41) and NODAGA-UBI (31-38) conjugates were found to be similar to UBI (29-41). NODAGA-UBI conjugates could be consistently labeled with Ga-68 in high radiochemical yields (>95 %) with high radiochemical purity (>95 %). [68Ga]NODAGA-UBI (29-41) and [68Ga]NODAGA-UBI (31-38) complexes showed retention time of 14 and 14.5 min, respectively, by HPLC radiochromatogram. Specific uptake of [68Ga]NODAGA-UBI fragments was observed in S.aureus cells. Greater than 64 % of the injected dose was cleared via the renal route at 1 h post injection, and no significant uptake in vital organs of mice was observed with both the agents.Conclusion: This is the first report on Ga-68 labeled NODAGA-UBI fragments for infection imaging and the agents hold tremendous prospect in PET imaging. [ABSTRACT FROM AUTHOR]

Summary: This study evaluated the levels of bone turnover markers (BTMs) and investigated relationships between them and bone mineral density (BMD) in postmenopausal women in China suburban district. The prevalence of osteoporosis was 25.03 % at lumbar spine and 6.23 % at femoral neck, and BTMs were negatively correlated with BMDs. Introduction: The aims of this study were to evaluate the levels of bone turnover markers (BTMs), including serum N-terminal procollagen of type I collagen (P1NP), beta C-terminal cross-linked of type I collagen (β-CTX), 25-hydroxyvitamin D [25(OH)D], and parathyroid hormone (PTH), and to investigate relationships between these markers and bone mineral density (BMD) as well the prevalence of osteoporosis in postmenopausal women of suburban district. Methods: A population of 4822 postmenopausal women aged 55-69 years old (62.22 ± 6.75) from the suburban district was recruited voluntarily. BMD was measured at the lumbar spine, femoral neck, and total hip using dual-energy X-ray absorptiometry; 2251 women in this group had the serum BTMs 25(OH)D and PTH tested. Results: The prevalence of osteoporosis was 25.03 % at lumbar spine and 6.23 % at femoral neck. The median (interquartile range) values of serum P1NP, β-CTX, 25(OH)D, and PTH were 59.3 ng/mL (44.7-75.52), 0.370 ng/mL (0.280-0.490), 23.0 ng/mL (17.1-30.5), and 31.4 pg/mL (24.9-39.7), respectively. Serum P1NP and β-CTX levels presented significantly negative correlations with BMDs at the all the sites (Beta = −0.098 to −0.208, respectively, P < 0.001), whereas PTH levels were negatively correlated with BMDs of the femoral neck and total hip (Beta = −0.062 and −0.054, P < 0.01, respectively). Serum 25(OH)D had positive associations with BMDs at total hip (Beta = 0.051, P < 0.01). Conclusions: The BMD of postmenopausal women in China suburban area is higher than that in downtown area, and over 60 % of the participants had their serum 25(OH)D level over 20 ng/mL. BTMs were negatively correlated with BMDs, suggesting that BTMs are reliable factors for early declines in BMD. [ABSTRACT FROM AUTHOR]

Conventional vaccine adjuvants enhance peptide-specific T-cell and B-cell responses by modifying peptide stability or uptake or by binding to pattern-recognition receptors on antigen-presenting cells (APCs). This article discusses the application of a distinct mechanism of adjuvant activity: the activation of type I, or invariant, natural killer T ( iNKT) cells to drive cellular and humoral immune responses. Using a semi-invariant T-cell receptor (TCR), iNKT cells recognize glycolipid antigens presented on cluster of differentiation (CD)-1d molecules. When their ligands are presented in concert with peptides, iNKT cells can provide T-cell help, 'licensing' APCs to augment peptide-specific T-cell and antibody responses. We discuss the potential benefits and limitations of exploiting iNKT cells as 'universal helpers' to enhance vaccine responses for the treatment and prevention of cancer and infectious diseases. [ABSTRACT FROM AUTHOR]

The acute toxicity of polychlorinated dibenzo- p-dioxins/dibenzofurans (PCDD/Fs) was investigated in the earthworm Eisenia andrei using filter paper toxicity test. Protein content, catalase (CAT) activity, and histology of intestinal wall (chloragogen cells and intestinal epithelium) were investigated in earthworms exposed for 48 h to 0 (control), 0.5, 1.0, and 1.5 ng/cm PCDD/Fs. The results showed an increase in the total protein content 1.56- ( p = 0.104), 1.66- ( p = 0.042), and 2.26-fold ( p < 0.001), respectively, compared to control. The average ± standard deviation of tissular CAT activity showed no significant differences; it was 36.01 ± 7.65, 36.17 ± 9.45, 36.08 ± 9.80, and 40.01 ± 6.98 U/g tissue, respectively. However, the average specific activity of CAT ± standard deviation was significantly decreased ( p < 0.001) at all doses compared to control; it was 2.93 ± 0.42, 1.93 ± 0.53, 1.80 ± 0.38, and 1.53 ± 0.44 U/mg protein, respectively. There was a progressive damage in both of the intestinal villi and the chloragogenous tissue associated with the incrementing doses. Since the toxic mixture altered the investigated biomarkers of E. andrei within 48 h, the cellular and molecular alterations resulted from the filter paper contact test could be utilized as a rapid toxicity assessment tool of environmental contamination with dioxins/furans and to assess consequent potential adverse effects on soil biota and other organisms in the ecosystem. [ABSTRACT FROM AUTHOR]

Although the response rates of chemotherapy in patients with acute T-lymphoblastic leukemia (T-ALL) have improved significantly, the outcome of these patients is still poor. Previous studies suggested that baicalein could inhibit the growth of several cancers, while its effect on T-ALL cells remains unclear. We used Jurkat cells as an in vitro model of T-ALL. Cell counting kit-8 assay and cytometric analysis with Annexin V-FITC/PI double staining were used to investigate the proliferation and apoptosis of Jurkat cells treated with increasing concentration of baicalein for indicated time. RT-PCR and western blotting was used to test the expression of Wnt/β-catenin associated genes and proteins. In cell viability assay, baicalein could inhibit the proliferation of Jurkat cells both in dose- and time-dependent manners. In cell apoptosis assay, baicalein could stimulate apoptosis of Jurkat cells both in dose- and time-dependent manners. Moreover, we demonstrated that baicalein could down-regulated the mRNA and protein levels of β-catenin and its widely accepted downstream targets (c-Myc, cyclin D1, and Axin2) in dose-dependent manners. These results proved that baicalein might be a potential choice for the treatment of T-ALL. [ABSTRACT FROM AUTHOR]

DNA methylation is the most common epigenetic modification in the mammalian genome. DNA methylation is governed by the DNA methyltransferases mainly DNMT1, DNMT3A, and DNMT3B. DNMT1 methylates hemimethylated DNA ensuring accurate DNA methylation maintenance. DNMT1 is involved in the proper differentiation of hematopoietic stem cells (HSCs) through the interaction with effector molecules. DNMT1 is deregulated in acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) as early as the leukemic stem cell stage. Through the interaction with fundamental transcription factors, non-coding RNAs, fusion oncogenes and by modulating core members of signaling pathways, it can affect leukemic cells biology. DNMT1 action might be also catalytic-independent highlighting a methylation-independent mode of action. In this review, we have gathered some current facts of DNMT1 role in AML and MDS and we also propose some perspectives for future studies. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the effect of Chinese medicine Haoqin Qingdan Decoction (蒿芩清胆汤, HQD) for febrile disease dampness-heat syndrome (FDDHS). Methods: Forty mice were divided into four groups, including normal control, FDDHS (induced by Radix et Rhizoma Rhei recipe and influenza virus A1 FM1 model), HQD, and the ribavirin groups (10 in each). The normal control and FDDHS groups were administered normal saline. HQD and the ribavirin groups were administered HQD and ribavirin intragastrically once daily at a dose of 64 g/(kg d) and 0.07 g/(kg d), respectively for 7 days. Lethargy, rough hair, diarrhea, tongue color and sole color were evaluated for pathological changes in morphology. The tongue and lung tissues were collected for histology. The CD14 and toll-like receptor 4 (TLR4) expression levels were measured using real-time quantitative polymerase chain reaction. Results: More than 80% of the FDDHS mice showed hypokinesia and lethargy, and pathological changes associated with rough hair, diarrhea, tongue color and sole color. With advanced treatment for 7 days, the thick greasy tongue fur of the HQD and ribavirin groups were thinner than that of the FDDHS group ( P<0.05), and it was the thinnest in the ribavirin group as compared with that in other groups ( P<0.05). The CD14 and TLR4 expression levels in the lung tissues of HQD and ribavirin groups significantly delined compared with the model group ( P<0.05 or P<0.01). CD14 was down-regulated more remarkably in the HQD group compared with the ribavirin group ( P<0.05), whereas the converse was true with TLR4 ( P<0.05). Conclusions: We established a FDDHS mouse model showing systemic clinical symptoms. Both HQD and ribavirin can inhibit the expression of CD14 and TLR4 in FDDHS mice, while the effect of ribavirin might be much more violent. The expression changes of CD14 and TLR4 consistently refers to lipopolysaccharide, the commonly and hotly inducing factor in FDDHS. [ABSTRACT FROM AUTHOR]

Macrophages are very important in the pathogenesis of rheumatoid arthritis (RA). The increase in the number of sublining macrophages in the synovium is an early hallmark of active rheumatic disease, and high numbers of macrophages are a prominent feature of inflammatory lesions. The degree of synovial macrophage infiltration correlates with the degree of joint erosion, and depletion of these macrophages from inflamed tissue has a profound therapeutic benefit. Research has now uncovered an unexpectedly high level of heterogeneity in macrophage origin and function, and has emphasized the role of environmental factors in their functional specialization. Although the heterogeneous populations of macrophages in RA have not been fully characterized, preliminary results in mouse models of arthritis have contributed to our understanding of the phenotype and ontogeny of synovial macrophages, and to deciphering the properties of monocyte-derived infiltrating and tissue-resident macrophages. Elucidating the molecular mechanisms that drive polarization of macrophages towards proinflammatory or anti-inflammatory phenotypes could lead to identification of signalling pathways that inform future therapeutic strategies. [ABSTRACT FROM AUTHOR]

Purpose: Our aim was to investigate lubricin, cartilage oligomeric matrix protein (COMP), and femoral cartilage deformation in response to different biomechanical loading of the knee joint (running vs cycling).Methods: Serum lubricin and COMP concentrations (enzyme-linked immunosorbent assay), and femoral cartilage thickness (suprapatellar transverse ultrasonography) were determined in 11 male runners (age: 40 ± 6 years; weight: 76 ± 8 kg) and 11 male cyclists (35 ± 12 years; 75 ± 5 kg) at baseline, immediately after, and 30 min after vigorous exercise (time trial: 10-km run or 25-km cycle).Results: At baseline, lubricin (runners: 104.0 ± 19.8 ng/ml; cyclists: 119.1 ± 23.9 ng/ml) and COMP (runners: 804.1 ± 87.5 ng/ml; cyclists: 693.0 ± 84.7 ng/ml) did not significantly differ; however, vigorous exercise was accompanied by an increase in lubricin (cyclists: 39.4 %; p < 0.05; runners: 56.9 %; p < 0.05) and COMP (cyclists: 32.1 %; p < 0.05; runners: 14.2 %; p = 0.14) that returned toward baseline following 30 min of rest (p < 0.05). No between-group differences were observed for baseline cartilage thickness at the intercondyle notch, medial condyle, and lateral condyle, and vigorous exercise did not result in significant change for either group.Conclusions: In the absence of ultrasonographic knee cartilage deformation, the response of serum lubricin and COMP following acute vigorous exercise indicates an increase in joint lubrication and cartilage metabolism, respectively, which appears largely independent of exercise modality. [ABSTRACT FROM AUTHOR]

Purpose: We assessed the role of monocarboxylate transporter 1 (MCT1) on lactate clearance during an active recovery after high-intensity exercise, by comparing genetic groups based on the T1470A (rs1049434) MCT1 polymorphism, whose influence on lactate transport has been proven.Methods: Sixteen young male elite field hockey players participated in this study. All of them completed two 400 m maximal run tests performed on different days, followed by 40 min of active or passive recovery. Lactate samples were measured immediately after the tests, and at min 10, 20, 30 and 40 of the recoveries. Blood lactate decreases were calculated for each 10-min period. Participants were distributed into three groups according to the T1470A polymorphism (TT, TA and AA).Results: TT group had a lower blood lactate decrease than AA group during the 10-20 min period of the active recovery (p = 0.018). This period had the highest blood lactate for the whole sample, significantly differing from the other periods (p ≤ 0.003). During the passive recovery, lactate declines were constant except for the 0-10-min period (p ≤ 0.003), suggesting that liver uptake is similar in all the genetic groups, and that the difference seen during the active recovery is mainly due to muscle lactate uptake.Conclusions: These differences according to the polymorphic variant T1470A suggest that MCT1 affects the plasma lactate decrease during a crucial period of active recovery, where the maximal lactate amount is cleared (i.e. 10-20 min period). [ABSTRACT FROM AUTHOR]

The novel FMS-like tyrosine kinase 3 (FLT3)-N676K point mutation within the FLT3 kinase domain-1 was recently identified in 6 % of de novo acute myeloid leukemia (AML) patients with inv(16). Because FLT3-N676K was encountered almost exclusively in inv(16) AML, we investigated the transforming potential of FLT3-N676K, the cooperation between FLT3-N676K and core binding factor ß-smooth muscle myosin heavy chain (CBFß-SMMHC) (encoded by the inv(16) chimeric gene CBFB-MYH11) in inducing acute leukemia, and tested the sensitivity of FLT3-N676K-positive leukemic cells to FLT3 inhibitors. Retroviral expression of FLT3-N676K in myeloid 32D cells induced AML in syngeneic C3H/HeJ mice (n = 11/13, median latency 58 days), with a transforming activity similar to FLT3-internal tandem duplication (ITD) (n = 8/8), FLT3-TKD D835Y (n = 8/9), and FLT3-ITD-N676K (n = 9/9) mutations. Three out of 14 (21.4 %) C57BL/6J mice transplanted with FLT3-N676K-transduced primary hematopoietic progenitor cells developed acute leukemia (latency of 68, 77, and 273 days), while no hematological malignancy was observed in the control groups including FLT3-ITD. Moreover, co-expression of FLT3-N676K/CBFß-SMMHC did not promote acute leukemia in three independent experiments (n = 16). In comparison with FLT3-ITD, FLT3-N676K induced much higher activation of FLT3 and tended to trigger stronger phosphorylation of MAPK and AKT. Importantly, leukemic cells carrying the FLT3-N676K mutant in the absence of an ITD mutation were highly sensitive to FLT3 inhibitors AC220 and crenolanib, and crenolanib even retained activity against the AC220-resistant FLT3-ITD-N676K mutant. Taken together, the FLT3-N676K mutant is potent to transform murine hematopoietic stem/progenitor cells in vivo. This is the first report of acute leukemia induced by an activating FLT3 mutation in C57BL/6J mice. Moreover, further experiments investigating molecular mechanisms for leukemogenesis induced by FLT3-N676K mutation and clinical evaluation of FLT3 inhibitors in FLT3-N676K-positive AML seem warranted. [ABSTRACT FROM AUTHOR]

Tissue culture under microgravity provides a venue which promotes cell-cell association while avoiding the detrimental effects of high shear stress. Hepatocytes cultured on carriers or entrapped within matrices under simulated microgravity conditions showed improved cell function and proliferation. In the present study, a new approach was adopted where a non-cell adherent scaffold was incorporated with hepatospheroids (HepG2) under microgravity. Gum arabic (GA) was cross-linked with gelatin (GA-Gel) and collagen (GA-Col) to prepare non-cell adherent scaffolds. Microgravity experiments with GA-Gel and GA-Col indicated that GA-Col is a better substrate compared to GA-Gel. Microgravity experiments of GA-Col scaffolds with HepG2 cells confirmed that the non-adherent surface with porous architecture can incorporate hepatocyte spheroids and maintain liver specific functions. Albumin and urea synthesis of hepatocytes was sustained up to 6 days under microgravity conditions in the presence of GA-Col scaffold. This new approach of using non-cell adherent matrix and microgravity environment for developing biological substitutes will be beneficial in tissue engineering, bioartificial liver devices and in vitro safety assessment of drugs. [ABSTRACT FROM AUTHOR]

Unconventional protein secretion (UPS) is a collective term for mechanisms by which cytosolic proteins that lack a signal peptide ('leaderless secretory proteins' (LSPs)) can gain access to the cell exterior. Numerous examples of UPS have been well documented in animal and yeast cells. In contrast, our understanding of the mechanism(s) and function of UPS in plants is very limited. This review evaluates the available literature on this subject. The apparent large numbers of LSPs in the plant secretome suggest that UPS also occurs in plants but is not a proof. Although the direct transport of LSPs across the plant plasma membrane (PM) has not yet been described, it is possible that as in other eukaryotes, exosomes may be released from plant cells through fusion of multivesicular bodies (MVBs) with the PM. In this way, LSPs, but also small RNAs (sRNAs), that are passively taken up from the cytosol into the intraluminal vesicles of MVBs, could reach the apoplast. Another possible mechanism is the recently discovered exocyst-positive organelle (EXPO), a double-membrane-bound compartment, distinct from autophagosomes, which appears to sequester LSPs. [ABSTRACT FROM AUTHOR]

Background: Regenerating islet-derived 3 (Reg3) is abnormally expressed in several human digestive system diseases, including chronic pancreatitis (CP) and pancreatic cancer (PC).Aim: The purpose of this study was to clarify the mechanism of the enhanced expression of Reg3 in inflammation-induced PC.Methods: C57BL/6 mice were treated with caerulein for 6 weeks to induce CP and then injected with pReg3g--a lentivirus system encoding for murine Reg3g--accompanied by dimethylbenzanthracene to induce PC. We detected pancreatic histopathological characteristics, tumor-related gene expression, inflammation-associated pathway activation, serum biochemical indicators, and immunological cell activities.Results: The mice that developed CP after caerulein treatment were marked by pronounced histologic lesions, elevated serum amylase levels, and activation of inflammation-related pathways. Mice given a high dose of pReg3g developed PC by 16 weeks, with recognizable tumors in the pancreas. While, both the low and high doses of pReg3g produced higher transcription of c-fos, k-ras, cytokeratin-19, and proliferating cell nuclear antigen, and a lower expression of caspase-3 compared to pNEG controls. Additionally, the higher dose of pReg3g increased the expressions of pSTAT3, NFκB (p65), and SOCS3 methylation during PC development. In addition, mice treated with pReg3g displayed higher levels of serum IL10 and TGFβ and suppressed T lymphocyte proliferation and DC function.Conclusion: The comprehensive analysis suggests enhanced Reg3g expression exacerbates PC in inflammation-associated cancer progression. Reg3g appears to promote CP-related PC in mice through multiple mechanisms, involving enhanced transcription of pancreatic tumor markers, repression of anti-tumor immunity, and activation of STAT3/p65 signal transduction pathways. [ABSTRACT FROM AUTHOR]

AC133 is a prominent surface marker of CD34+ and CD34- hematopoietic stem/progenitor cell (HSPC) subsets. AC133+ HSPCs contain high progenitor cell activity and are capable of hematopoietic reconstitution. Furthermore, AC133 is used for prospective isolation of tumor-initiating cells in several hematological malignancies. Nucleolin is a multifunctional factor of growing and cancer cells, which is aberrantly active in certain hematological neoplasms, and serves as a candidate molecular target for cancer therapy. Nucleolin is involved in gene transcription and RNA metabolism and is prevalently expressed in HSPCs, as opposed to differentiated hematopoietic tissue. The present study dissects nucleolin-mediated activation of surface AC133 and its cognate gene CD133, via specific interaction of nucleolin with the tissue-dependent CD133 promoter P1, as a mechanism that crucially contributes to AC133 expression in CD34+ HSPCs. In mobilized peripheral blood (MPB)-derived HSPCs, nucleolin elevates colony-forming unit (CFU) frequencies and enriches granulocyte-macrophage CFUs. Furthermore, nucleolin amplifies long-term culture-initiating cells and also promotes long-term, cytokine-dependent maintenance of hematopoietic progenitor cells. Active β-catenin, active Akt and Bcl-2 levels in MPB-derived HSPCs are nucleolin-dependent, and effects of nucleolin on these cells partially rely on β-catenin activity. The study provides new insights into molecular network relevant to stem/progenitor cells in normal and malignant hematopoiesis. [ABSTRACT FROM AUTHOR]

Diamond-Blackfan anemia (DBA) is a congenital erythroid hypoplasia caused by haploinsufficiency of genes encoding ribosomal proteins (RPs). Perturbed ribosome biogenesis in DBA has been shown to induce a p53-mediated ribosomal stress response. However, the mechanisms of p53 activation and its relevance for the erythroid defect remain elusive. Previous studies have indicated that activation of p53 is caused by the inhibition of mouse double minute 2 (Mdm2), the main negative regulator of p53, by the 5S ribonucleoprotein particle (RNP). Meanwhile, it is not clear whether this mechanism solely mediates the p53-dependent component found in DBA. To approach this question, we crossed our mouse model for RPS19-deficient DBA with Mdm2(C305F) knock-in mice that have a disrupted 5S RNP-Mdm2 interaction. Upon induction of the Rps19 deficiency, Mdm2(C305F) reversed the p53 response and improved expansion of hematopoietic progenitors in vitro, and ameliorated the anemia in vivo. Unexpectedly, disruption of the 5S RNP-Mdm2 interaction also led to selective defect in erythropoiesis. Our findings highlight the sensitivity of erythroid progenitor cells to aberrations in p53 homeostasis mediated by the 5S RNP-Mdm2 interaction. Finally, we provide evidence indicating that physiological activation of the 5S RNP-Mdm2-p53 pathway may contribute to functional decline of the hematopoietic system in a cell-autonomous manner over time. [ABSTRACT FROM AUTHOR]

Kv1.4 potassium channels are heavily glycosylated proteins involved in shaping action potentials and in neuronal excitability and plasticity. Kv1.4 N354Q, without an N-glycan, exhibited decreased protein stability and trafficking to the cell surface (Watanabe et al. in J Biol Chem 279:8879-8885, ). Here we investigated whether the composition of the N-glycan affected Kv1.4 cell surface expression. Kv1.4 proteins carrying N-glycans with different compositions were generated by adding glycosidase inhibitors or using N-glycosylation-deficient mutant cell lines. We found that oligomannose-type, hybrid-type, or incomplete complex-type N-glycans had a negative effect on surface protein expression of Kv1.4 compared with complex-type N-glycans. The decrease in surface protein level of Kv1.4 was mainly due to a reduction in total protein level, induced by altered N-glycan composition. Kv1.4 in CSTP-treated cells carried a unique oligomannose-type N-glycan that contains three glucose residues. This N-glycan had the most negative effect on cell surface expression of Kv1.4. It decreased Kv1.4 surface protein level by a combined mechanism of reducing total protein level and increasing ER-retention. Our data suggest that composition of the N-glycan plays an important role in protein stability and trafficking, and a sialylated complex-type N-glycan promoted high cell surface expression of Kv1.4. [ABSTRACT FROM AUTHOR]

Purpose: Exercise induces oxidative stress and causes adaptations in antioxidant defenses. The aim of the present study was to determine the effects of a 2-month diet supplementation with docosahexaenoic acid (DHA) on the pro-oxidant and antioxidant status of peripheral blood mononuclear cells (PBMCs) during football training and after acute exercise. Methods: Fifteen male football players, in a randomized double-blind trial, ingested a beverage enriched with DHA or a placebo for 8 weeks. Blood samples were collected in basal conditions before and after the training period and after an acute and intense exercise. Results: The training season increased the carbonyl and nitrotyrosine index but decreased the malondialdehyde (MDA) levels. Basal catalase activity decreased in both groups after 8 weeks of training, whereas glutathione peroxidase activity increased mainly in the placebo group. Protein levels of uncoupling proteins (UCP2 and UCP3) and inducible nitric oxide synthase significantly increased after the training period. Acute exercise induced redistribution in the number of circulating cells, increased the MDA levels and nitrotyrosine index, and decreased the levels of nitrate. Acute exercise also increased PBMCs reactive oxygen species (ROS) production after immune stimulation. Diet supplementation with DHA significantly increased the UCP3 levels after training and the superoxide dismutase protein levels after acute exercise, and reduced the production of ROS after acute exercise. Conclusion: Docosahexaenoic acid increased the antioxidant capabilities while reducing the mitochondrial ROS production in a regular football training period and reduced the oxidative damage markers in response to acute exercise. [ABSTRACT FROM AUTHOR]

Osteochondral injuries remain difficult to repair. We developed a novel photo-cross-linkable furfurylamine-conjugated gelatin (gelatin-FA). Gelatin-FA was rapidly cross-linked by visible light with Rose Bengal, a light sensitizer, and was kept gelled for 3 weeks submerged in saline at 37°C. When bone marrow-derived stromal cells (BMSCs) were suspended in gelatin-FA with 0.05% Rose Bengal, approximately 87% of the cells were viable in the hydrogel at 24 h after photo-cross-linking, and the chondrogenic differentiation of BMSCs was maintained for up to 3 weeks. BMP4 fusion protein with a collagen binding domain (CBD) was retained in the hydrogels at higher levels than unmodified BMP4. Gelatin-FA was subsequently employed as a scaffold for BMSCs and CBD-BMP4 in a rabbit osteochondral defect model. In both cases, the defect was repaired with articular cartilage-like tissue and regenerated subchondral bone. This novel, photo-cross-linkable gelatin appears to be a promising scaffold for the treatment of osteochondral injury. [ABSTRACT FROM AUTHOR]

MADS-domain proteins are important transcription factors involved in many biological processes of plants. In our study, a tomato MADS-box gene, SlFYFL, was isolated. SlFYFL is expressed in all tissues of tomato and significantly higher in mature leave, fruit of different stages, AZ (abscission zone) and sepal. Delayed leaf senescence and fruit ripening, increased storability and longer sepals were observed in 35S:FYFL tomato. The accumulation of carotenoid was reduced, and ethylene content, ethylene biosynthetic and responsive genes were down-regulated in 35S:FYFL fruits. Abscission zone (AZ) did not form normally and abscission zone development related genes were declined in AZs of 35S:FYFL plants. Yeast two-hybrid assay revealed that SlFYFL protein could interact with SlMADS-RIN, SlMADS1 and SlJOINTLESS, respectively. These results suggest that overexpression of SlFYFL regulate fruit ripening and development of AZ via interactions with the ripening and abscission zone-related MADS box proteins. [ABSTRACT FROM AUTHOR]

Epidermolysis bullosa (EB) represents a group of inherited blistering skin diseases, some forms of which are associated with considerable morbidity and increased mortality. Notably, in recessive dystrophic EB there can be extensive muco-cutaneous fragility and disease complications such as scars, contractures, anemia, malnutrition, and malignancy. Currently, there is no effective therapy or cure for EB. Over the last decade, however, a number of important advances have been made that are bringing new treatments closer to the clinic, including gene therapy, protein replacement therapy, cell therapies [allogeneic fibroblasts, mesenchymal stromal cells (MSCs), bone marrow stem cell transplantation, culturing/grafting revertant mosaic keratinocytes], gene editing/engineering, and clinical application of inducible pluripotent stem cells. Although a cure for EB still remains elusive, recent data on animal models and initial human clinical trials have raised the expectations of patients, clinicians, and researchers that disease modification and improved quality of life are feasible goals. Furthermore, the lessons learned in treating EB are likely to have significant implications for improving the management of other genetic diseases. [ABSTRACT FROM AUTHOR]

RNA-binding proteins are key regulators of gene expression, yet only a small fraction have been functionally characterized. Here we report a systematic analysis of the RNA motifs recognized by RNA-binding proteins, encompassing 205 distinct genes from 24 diverse eukaryotes. The sequence specificities of RNA-binding proteins display deep evolutionary conservation, and the recognition preferences for a large fraction of metazoan RNA-binding proteins can thus be inferred from their RNA-binding domain sequence. The motifs that we identify in vitro correlate well with in vivo RNA-binding data. Moreover, we can associate them with distinct functional roles in diverse types of post-transcriptional regulation, enabling new insights into the functions of RNA-binding proteins both in normal physiology and in human disease. These data provide an unprecedented overview of RNA-binding proteins and their targets, and constitute an invaluable resource for determining post-transcriptional regulatory mechanisms in eukaryotes. [ABSTRACT FROM AUTHOR]

CRAC (Calcium Release-Activated Calcium) channels represent the primary pathway for so-called "store-operated calcium entry" - the cellular entry of calcium induced by depletion of intracellular calcium stores. These channels play a key role in diverse cellular activities, most noticeably in the differentiation and activation of Tcells, and in the response of mast cells to inflammatory signals. CRAC channels are formed by members of the recently discovered Orai protein family, with previous studies indicating that the functional channel is formed by a tetramer of Orai subunits. However, a recent report has shown that crystals obtained from the purified Drosophila Orai protein display a hexameric channel structure. Here, by comparing the biophysical properties of concatenated hexameric and tetrameric human Orai1 channels expressed in HEK293 cells, we show that the tetrameric channel displays the highly calcium-selective conductance properties consistent with endogenous CRAC channels, whilst the hexameric construct forms an essentially non-selective cation channel. [ABSTRACT FROM AUTHOR]

Much of the focus on the transforming growth factor-β (TGFβ) superfamily in cancer has revolved around the TGFβ ligands themselves. However, it is now becoming apparent that deregulated signalling by many of the other superfamily members also has crucial roles in both the development of tumours and metastasis. Furthermore, these signalling pathways are emerging as plausible therapeutic targets. Their roles in tumorigenesis frequently reflect their function in embryonic development or in adult tissue homeostasis, and their influence extends beyond the tumours themselves, to the tumour microenvironment and more widely to complications of cancer such as cachexia and bone loss. [ABSTRACT FROM AUTHOR]

The heritability of specific phenotypical traits relevant for physical performance has been extensively investigated and discussed by experts from various research fields. By deciphering the complete human DNA sequence, the human genome project has provided impressive insights into the genomic landscape. The hope that this information would reveal the origin of phenotypical traits relevant for physical performance or disease risks has proven overly optimistic, and it is still premature to refer to a 'post-genomic' era of biological science. Linking genomic regions with functions, phenotypical traits and variation in disease risk is now a major experimental bottleneck. The recent deluge of genome-wide association studies (GWAS) generates extensive lists of sequence variants and genes potentially linked to phenotypical traits, but functional insight is at best sparse. The focus of this review is on the complex mechanisms that modulate gene expression. A large fraction of these mechanisms is integrated into the field of epigenetics, mainly DNA methylation and histone modifications, which lead to persistent effects on the availability of DNA for transcription. With the exceptions of genomic imprinting and very rare cases of epigenetic inheritance, epigenetic modifications are not inherited transgenerationally. Along with their susceptibility to external influences, epigenetic patterns are highly specific to the individual and may represent pivotal control centers predisposing towards higher or lower physical performance capacities. In that context, we specifically review how epigenetics combined with classical genetics could broaden our knowledge of genotype-phenotype interactions. We discuss some of the shortcomings of GWAS and explain how epigenetic influences can mask the outcome of quantitative genetic studies. We consider epigenetic influences, such as genomic imprinting and epigenetic inheritance, as well as the life-long variability of epigenetic modification patterns and their potential impact on phenotype with special emphasis on traits related to physical performance. We suggest that epigenetic effects may also play a considerable role in the determination of athletic potential and these effects will need to be studied using more sophisticated quantitative genetic models. In the future, epigenetic status and its potential influence on athletic performance will have to be considered, explored and validated using well controlled model systems before we can begin to extrapolate new findings to complex and heterogeneous human populations. A combination of the fields of genomics, epigenomics and transcriptomics along with improved bioinformatics tools and precise phenotyping, as well as a precise classification of the test populations is required for future research to better understand the inter-relations of exercise physiology, performance traits and also susceptibility towards diseases. Only this combined input can provide the overall outlook necessary to decode the molecular foundation of physical performance. [ABSTRACT FROM AUTHOR]

In 1817, James Parkinson described the symptoms of the shaking palsy, a disease that was subsequently defined in greater detail, and named after Parkinson, by Jean-Martin Charcot. Parkinson expected that the publication of his monograph would lead to a rapid elucidation of the anatomical substrate of the shaking palsy; in the event, this process took almost a century. In 1912, Fritz Heinrich Lewy identified the protein aggregates that define Parkinson disease (PD) in some brain regions outside the substantia nigra. In 1919, Konstantin Nikolaevich Tretiakoff found similar aggregates in the substantia nigra and named them after Lewy. In the 1990s, α-synuclein was identified as the main constituent of the Lewy pathology, and its aggregation was shown to be central to PD, dementia with Lewy bodies, and multiple system atrophy. In 2003, a staging scheme for idiopathic PD was introduced, according to which α-synuclein pathology originates in the dorsal motor nucleus of the vagal nerve and progresses from there to other brain regions, including the substantia nigra. In this article, we review the relevance of Lewy's discovery 100 years ago for the current understanding of PD and related disorders. [ABSTRACT FROM AUTHOR]

Field and laboratory studies of the iconic nectarivorous and 'pollenivorous' honey possum, Tarsipes rostratus, are reviewed with the aim of identifying aspects of its physiology that are as yet poorly understood and needed to implement management strategies for its long-term conservation. Dietary specialisations include the loss of teeth, a modified gut with a high rate of passage, exceptionally low minimum nitrogen requirements, an apparently high basal metabolic rate and a permanently polyuric kidney. In contrast, its reproductive physiology is plesiomorphic, combining aspects such as a post- partum oestrus, embryonic diapause, photoperiodicity and extended maternal care that are usually separate characteristics of other marsupial groups. In common with a number of other marsupials, the honey possum has the potential for trichromatic colour vision and has been the subject of several studies attempting to correlate visual quality with ecological realities. Field physiological studies have established its high rates of nectar and pollen intake needed to maintain energy balance and highlight the need for a constant intake from floral sources. Early allometric studies suggesting that the honey possum's relatively low reproductive rate may be linked to a diet limited in protein have not been supported and nitrogen intakes in the field exceed by a factor of 10 the animal's basic requirements for balance. Measurements of rates of protein turnover in field-caught lactating females suggest that they divert nitrogen from the protein pool to milk production by reducing rates of degradation, rather than by increasing rates of synthesis of protein. Although not yet an endangered species, the honey possum's habitat has been drastically reduced since European occupation of Australia and future-targeted research on the animal's unique physiology and habitat linkage is needed that can be translated into effective management practices. Only then will its long-term survival be assured. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) repress cellular protein levels to provide a sophisticated parameter of gene regulation that coordinates a broad spectrum of biological processes. Bone organogenesis is a complex process involving the differentiation and crosstalk of multiple cell types for formation and remodeling of the skeleton. Inhibition of mRNA translation by miRNAs has emerged as an important regulator of developmental osteogenic signaling pathways, osteoblast growth and differentiation, osteoclast-mediated bone resorption activity and bone homeostasis in the adult skeleton, miRNAs control multiple layers of gene regulation for bone development and postnatal functions, from the initial response of stem/progenitor cells to the structural and metabolic activity of the mature tissue. This Review brings into focus an emerging concept of bone-regulating miRNAs, the evidence for which has been gathered largely from in vivo mouse models and in vitro studies in human and mouse skeletal cell populations. Characterization of miRNAs that operate through tissue-specific transcription factors in osteoblast and osteoclast lineage cells, as well as intricate feedforward and reverse loops, has provided novel insights into the supervision of signaling pathways and regulatory networks controlling normal bone formation and turnover. The current knowledge of miRNAs characteristic of human pathologic disorders of the skeleton is presented with a future goal towards translational studies. [ABSTRACT FROM AUTHOR]

Telaprevir (Incivo®, Incivek®), an orally administered inhibitor of the hepatitis C virus non-structural protein NS3-4A serine protease, is used in combination with pegylated interferon (peginterferon)-alpha and ribavirin in the treatment of adults with genotype 1 chronic hepatitis C virus (HCV) infection (chronic hepatitis C). This article reviews data on the clinical efficacy and tolerability of telaprevir in adults with genotype 1 chronic hepatitis C and provides a summary of its pharmacological properties. In in vitro studies, telaprevir shows good activity against genotype 1 HCV, including viral isolates from patients with chronic hepatitis C. Numerous resistant variants of HCV have been identified in clinical isolates from patients receiving treatment with telaprevir-based therapy. However, the clinical relevance of viral variants emerging during treatment requires further study. Telaprevir administered for 12 weeks in combination with peginterferon-alpha-2a and ribavirin for up to 48 weeks was effective in the treatment of previously untreated or previously treated adults with genotype 1 chronic hepatitis C in three major randomized phase III trials. Sustained virological response rates (i.e. percentages of patients with undetectable HCV RNA levels 24 weeks after the last planned dose of study medication: the primary endpoint) achieved with the telaprevir-based regimens were significantly higher than those produced with peginterferon-alpha-2a and ribavirin alone (the 'current standard of care'). In the largest trial (the ADVANCE trial) in previously untreated patients, 24 weeks after the last planned dose of study drug, recipients of 12 weeks' treatment with telaprevir in combination with peginterferon-alpha-2a and ribavirin followed by treatment with peginterferon-alpha-2a and ribavirin for a further 12 or 36 weeks experienced significantly (p< 0.001) higher sustained virological response rates than patients who received peginterferon-alpha-2a and ribavirin dual therapy for 48 weeks (75% vs 44%). Adverse events were reported more frequently with telaprevir-based regimens than with peginterferon-alpha-2a and ribavirin dual therapy in the major trials. The most common adverse events included fatigue, rash, pruritus, anaemia and nausea. In conclusion, telaprevir in combination with peginterferon-alpha and ribavirin is an effective treatment for treatment-naive and previously treated adults with genotype 1 chronic hepatitis C - patient groups in whom peginterferon-alpha and ribavirin dual therapy may not be successful. Thus, telaprevir is a valuable new treatment option for use in combination with peginterferon-alpha and ribavirin in treatment-naive or previously treated adults with genotype 1 chronic hepatitis C. [ABSTRACT FROM AUTHOR]

An important issue concerning the use of soft contact lenses is comfort, which, among other factors, has been related to the level of friction between the anterior side of the lens and the inner eyelid. Although several studies have been carried out to investigate the frictional properties of contact lenses, these have not taken the physiological environment of the eye into account. In use, lenses are in contact with proteins present in tears, with corneal cells and with the palpebral conjunctiva (clear membrane on inner eyelid). The focus of this study was to establish a biologically relevant measurement protocol for the investigation of friction of contact lenses that would mimic the eye's physiological environment. By optimizing parameters such as the composition of the friction counter surface, the lubricant solution, the normal load and the velocity, an ideal protocol and setup for microtribological testing could be established and used to perform a comparative study of various commercially available soft contact lenses. [ABSTRACT FROM AUTHOR]

Background and Objectives: Existing methods for the prediction of human clearance of therapeutic proteins involve the use of allometry approaches. In general, these approaches have concentrated on the role of body weight, with only occasional attention given to more specific physiological parameters. The objective of this study was to develop a mechanism-based model of hepatic clearance (CLH), which combines a single-species scaling approach with liver physiology, for predicting CLH of selected glycoprotein derivate therapeutics, and to compare the outcome of this novel method with those of two empirical methods obtained from the literature- namely, the single-exponent theory and multiple-species allometry. Thus, this study was designed as an explanatory study to verify if the addition of physiological information is of benefit for extrapolating clearance of selected therapeutic proteins from one species to another. Methods: Five glycoprotein derivate therapeutics that are known to be principally eliminated by asia-loglycoprotein receptors (ASGPRs) under in vivo conditions were selected. It was assumed that the interspecies differences in CLH reported for these compounds are reflected by the interspecies differences in the abundance of these receptors. Therefore, key scaling factors related to these differences were integrated into one model. Fourteen extrapolation (prediction) scenarios across species were used in this study while comparing the single-species model, based on physiology, with the single-exponent theory. In addition, the physiological model was compared with multiple-species allometry for three proteins. Results: In general, the novel physiological model is superior to the derived allometric methods. Overall, the physiological model produced a predicted CLH value with levels of accuracy of 100% within 3-fold, 100% within 2-fold and about 82% within 1.5-fold, compared with the observed values, whereas the levels of accuracy decreased to 93%, 77% and 53%, respectively, for allometry. The proposed physiological model is also superior to allometry on the basis of the root mean square error and absolute average fold error values. Conclusions: It has been demonstrated that interspecies differences in the abundance of ASGPRs principally govern interspecies variations in CLH of compounds that are principally eliminated by ASGPRs. Overall, the proposed physiological model is an additional tool, which should facilitate investigation and prediction of human CLH of specific glycoproteins solely on the basis of clearance data determined in a single preclinical species. [ABSTRACT FROM AUTHOR]

The tectorial membrane (TM) connects to the stereociliary bundles of outer hair cells (OHCs). Humans with an autosomal dominant C1509G mutation in alpha-tectorin, a protein constituent of the TM, are born with a partial hearing loss that worsens over time. The Tecta(C1509/+) transgenic mouse with the same point mutation has partial hearing loss secondary to a shortened TM that only contacts the first row of OHCs. As well, Tecta(C1509G/+) mice have increased expression of the OHC electromotility protein, prestin. We sought to determine whether these changes impact OHC survival. Distortion product otoacoustic emission thresholds in a quiet environment did not change to 6 months of age. However, noise exposure produced acute threshold shifts that fully recovered in Tecta (+/+) mice but only partially recovered in Tecta(C1509G/+) mice. While Tecta(+/+) mice lost OHCs primarily at the base and within all three rows, Tecta(C1509G/+) mice lost most of their OHCs in a more apical region of the cochlea and nearly completely within the first row. In order to estimate the impact of a shorter TM on the forces faced by the stereocilia within the first OHC row, both the wild type and the heterozygous conditions were simulated in a computational model. These analyses predicted that the shear force on the stereocilia is ~50% higher in the heterozygous condition. We then measured electrically induced movements of the reticular lamina in situ and found that while they decreased to the noise floor in prestin null mice, they were increased by 4.58 dB in Tecta(C1509G/+) mice compared to Tecta(+/+) mice. The increased movements were associated with a fourfold increase in OHC death as measured by vital dye staining. Together, these findings indicate that uncoupling the TM from some OHCs leads to partial hearing loss and places the remaining coupled OHCs at higher risk. Both the mechanics of the malformed TM and the increased prestin-related movements of the organ of Corti contribute to this higher risk profile. [ABSTRACT FROM AUTHOR]

Background: Naturally oncolytic reovirus preferentially kills cancer cells, making it a promising cancer therapeutic. Mutations in tumour suppressor p53 are prevalent in cancers, yet the role of p53 in reovirus oncolysis is relatively unexplored.Methods: Human cancer cell lines were exposed to Nutlin-3a, reovirus or a combination of the two and cells were processed for reovirus titration, western blot, real-time PCR and apoptosis assay using Annexin V and 7-AAD staining. Confocal microscopy was used to determine translocation of the NF-κB p65 subunit.Results: We show that despite similar reovirus replication in p53(+/+) and p53(-/-) cells, stabilisation of p53 by Nutlin-3a significantly enhanced reovirus-induced apoptosis and hence virus release and dissemination while having no direct effect on virus replication. Enhanced apoptosis by Nutlin-3a was not observed in p53(-/-) or p53 knockdown cells; however, increased expression of Bax and p21 are required. Moreover, elevated NF-κB activation in reovirus-infected cells following Nutlin-3a treatment was necessary for enhanced reovirus-induced apoptosis, as synergistic cytotoxicity was overcome by specific NF-κB inhibitors.Conclusion: Nutlin-3a treatment enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-κB activation, and combination of reovirus and Nutlin-3a might represent an improved therapy against cancers harbouring wild-type p53. [ABSTRACT FROM AUTHOR]