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Summary: Activated calcium‐/calmodulin‐dependent protein kinaseII (CaMKII) is important to promote chondrocytes from proliferative to pre‐hypertrophic state, which probably plays a role in osteoarthritis (OA), a widespread degeneration disease with enhanced aberrant chondrocyte differentiation. Our aim was to detect the role of CaMKII, and its relationship with the feedback loop of Indian hedgehog (Ihh) and Parathyroid‐related peptide (PTHrP) in the temporomandibular joints (TMJs) OA. KN93, the competitive inhibitor of CaMKII, was added to the culture medium in vitro and was locally injected to rats TMJs (n = 54, female) every other day for 4 weeks from the beginning of the 5th and 9th week after installing of unilateral anterior crossbite (UAC), termed as 4 wk+4 wk and 8 wk+4 wk, accordingly. The RNA expression of CaMKII α (1.49 ± 0.09), CaMKII β (3.36 ± 0.20), Ihh (1.88 ± 0.06) and PTHrP (1.87 ± 0.12) was all enhanced, especially at 24 dyn/cm2 in vitro (all P < .05), accompanied with downregulated expression of cartilage matrix, but upregulated markers of chondrocytes differentiation (all P < 0.05). Similarity was observed in the 4 wk+4 wk group in vivo. In the 8 wk+4 wk group, UAC upregulated the RNA expression of CaMKII α (1.81 ± 0.24), CaMKII β (1.36 ± 0.07) and Ihh (1.70 ± 0.21), however, down‐regulated PTHrP (0.53 ± 0.04) (all P < .05), in consonance with the protein expression. All these changes were attenuated by KN93 (all P < .05). In conclusion, CaMKII took a role, via Ihh and PTHrP pathways, in promoting biomechanically induced TMJ chondrocytes differentiation, the initiation issue of UAC stimulated osteoarthritic changes in rodent TMJs. Inhibiting CaMKII is helpful to rescue the biomechanically stimulated cartilage degradation and prospective to be a target treatment of OA. [ABSTRACT FROM AUTHOR]

While regulatory T cells (Tregs) constitutively express programmed cell death-1 (PD-1), the role of PD-1 expression in Tregs of patients with sepsis remains unclear. Thus, we determined PD-1 expression in Tregs from the peripheral blood of patients with severe sepsis and septic shock. Seventy-eight patients with severe sepsis and 40 with septic shock, as well as 21 healthy subjects, were enrolled in this study. The percentage of peripheral blood PD-1+ Tregs, as well as absolute Treg counts, were compared between these three groups. PD-1 expression in Tregs and absolute Treg counts were also compared between survivors and non-survivors in patients with sepsis. PD-1 expression in Tregs increased in patients with sepsis compared to healthy controls. Conversely, absolute Treg counts were significantly decreased in patients with sepsis compared to healthy controls; patients with septic shock had the lowest absolute Treg counts. Among patients with sepsis, survivors had lower levels of PD-1 expression in Tregs, as well as higher absolute Treg counts, than non-survivors. Additionally, the percentage of PD-1+ Tregs correlated positively with lactate levels as well as the Acute Physiology and Chronic Health Evaluation II and Sequential Organ Failure Assessment scores in patients with sepsis. PD-1 was upregulated in Tregs of patients with sepsis and may indicate a state of immune dysfunction. Overexpression of PD-1 in Tregs was associated with more severe sepsis as well as poor outcomes. [ABSTRACT FROM AUTHOR]

Abstract: To provide a tool for research on regulating adipocyte differentiation, tetracycline inducible (Tet on) lentiviral expression vectors under the control of an adipose-specific promoter were constructed. The lowest basal expression in the absence of doxycycline and most efficient dose-dependent, doxycycline-induced transient overexpression was observed using vectors constructed with a combination of Tetracycline Responsive Element (TRE) and reverse tetracycline-controlled TransActivator advanced (rtTAadv), transfected in white (3T3-L1) and brown (HIB-1B) preadipocytes cell lines. The results demonstrate that doxycycline adipogenic inducible expression can be achieved using a pLenti TRE / rtTA adv under the control of the truncated aP2 promoter in HIB-1B preadipocytes. [Copyright &y& Elsevier]

Expression of the brown adipocyte-specific gene, uncoupling protein 1 (UCP1), is increased by both PPARγ stimulation and cAMP activation through their ability to stimulate the expression of the PPAR coactivator PGC1α. In HIB1B brown preadipocytes, combination of the PPARγ agonist, rosiglitazone, and the cAMP stimulator forskolin synergistically increased UCP1 mRNA expression, but PGC1α expression was only increased additively by the two drugs. The PPARγ antagonist, GW9662, and the PKA inhibitor, H89, both inhibited UCP1 expression stimulated by rosiglitazone and forskolin but PGC1α expression was not altered to the same extent. Reporter studies demonstrated that combined rosiglitazone and forskolin synergistically activated transcription froma full length 3.1 kbp UCP1 luciferase promoter construct, but the response was only additive andmuch reducedwhen a minimal 260 bp proximal UCP1 promoter was examined. Rosiglitazone and forskolin in combination were able to synergistically stimulate promoters comprising of tandem repeats of either PPREs or CREs. We conclude that rosiglitazone and forskolin act together to synergistically activate the UCP1 promoter directly rather than by increasing PGC1α expression and by a mechanism involving cross-talk between the signalling systems regulating the CRE and PPRE on the promoters. [ABSTRACT FROM AUTHOR]

Abstract: A critical period in respiratory network development occurs in the rat around postnatal days (P) 12–13, when abrupt neurochemical, metabolic, and physiological changes were evident. As serotonin and its receptors are involved in respiratory modulation, and serotonergic abnormality is implicated in sudden infant death syndrome, we hypothesized that 5-HT receptors are significantly downregulated during the critical period. This was documented recently for 5-HT2AR in several respiratory nuclei. The present study represents a comprehensive analysis of postnatal development of 5-HT1AR and 5-HT1BR in 10 brain stem nuclei and 5-HT2AR in six nuclei not previously examined. Optical densitometric analysis of immunohistochemically-reacted neurons from P2 to P21 indicated four developmental patterns of expression: (1) Pattern I: a high level of expression at P2-P11, an abrupt and significant reduction at P12, followed by a plateau until P21 (5-HT1AR and 5-HT1BR in raphé magnus [RM], raphé obscurus [ROb], raphé pallidus [RP], pre-Bötzinger complex [PBC], nucleus ambiguus [Amb], and hypoglossal nucleus [XII; 5-HT1AR only]). (2) Pattern II: a high level at P2-P9, a gradual decline from P9 to P12, followed by a plateau until P21 (5-HT1AR and 5-HT1BR in the retrotrapezoid nucleus (RTN)/parafacial respiratory group (pFRG)). (3) Pattern III: a high level at P2-P11, followed by a gradual decline until P21 (5-HT1AR in the ventrolateral subnucleus of solitary tract nucleus [NTSVL] and the non-respiratory cuneate nucleus [CN]). (4) Pattern IV: a relatively constant level maintained from P2 to P21 (5-HT1AR in the commissural subnucleus of solitary tract nucleus (NTSCOM); 5-HT1BR in XII, NTSVL, NTSCOM, and CN; and 5-HT2AR in RM, ROb, RP, RTN/pFRG, NTSVL, and NTSCOM). Thus, a significant reduction in the expression of 5-HT1AR, 5-HT1BR, and 5-HT2AR in multiple respiratory-related nuclei at P12 is consistent with reduced serotonergic transmission during the critical period, thereby rendering the animals less able to respond adequately to ventilatory distress. [Copyright &y& Elsevier]

A comprehensive analysis of sequence patterns around the stop codons was performed, by using more than 26,000 rice full-length cDNA sequences. Here it is shown that the bias was most outstanding at the position immediately before the stop codons (-1 codon), where the AAC codon was strongly preferred among ANC codons. Compared with other positions, the codon immediately after the stop codons (+1 codon) also displayed an apparent difference, and had a strong consensus for base A at the first, C at the second, and A at the third letters, respectively. Notably, the base biases at the positions directly downstream of the stop codons, such as the +4, +5 and +6 positions, were much stronger than other positions in the 3'-UTR region, suggesting that those base positions might act as an extended stop signal in the process of protein synthesis. Examination of the relationship between sequence pattern and gene expression level, assessed by CAI values and EST counting, revealed a tendency towards bigger base biases for highly expressed genes. It could be inferred that the translation stop signal is possibly involved in many sequence recognition elements other than the stop codons; highly expressed genes should hold strong sequence consensus around the stop codons for efficient translation termination. [ABSTRACT FROM AUTHOR]

High amylose content (AC) in rice endosperm is correlated with poor grain quality, particularly in indica hybrid rice. We have generated several homozygous transgenic parent lines of indica hybrid rice carrying an antisense Waxy ( Wx) gene and demonstrated that the AC in seeds of these lines decreased dramatically. Two transgenic maintainer lines (L25B and L18B), derived from one of the key maintainer parents of an indica hybrid rice in China, Long-te-fu B (LTF-B), were selected and the antisense Wx gene was subsequently introgressed into the male-sterile counterpart, LTF-A, with the aim to generate improved indica hybrids. The indica hybrids derived from the selected transgenic male-sterile lines and restorer lines were tested for quality and agronomic performance under normal field conditions. Our results demonstrated that the reduction of AC in the homozygous transgenic maintainer lines stably passed down in five successive generations and the improved quality was also found in their relevant transgenic hybrids produced. The other two key characters of rice cooking and eating quality, the gel consistence (GC) and gelatinization temperature (GT), were also improved in the grains of both the transgenic maintainer lines and their relevant hybrids. In addition, no change was observed for most of the agronomic characters of the transgenic maintainer lines and the relevant transgenic hybrids. Although the grain weight of the transgenic line was reduced, the grain yield of the homozygous transgenic parent lines and the transgenic hybrids was similar when compared with that of the wild-type controls. These results suggest that transgenic approaches are an effective way to obtain rice lines with both improved qualities and high yield, especially for indica hybrid rice. [ABSTRACT FROM AUTHOR]

Aims: Multidrug resistance (MDR) is a major obstacle in the treatment of lymphoma. The expression of MDR-1 mRNA and P-glycoprotein (MDR-1/P-gp) has been linked to MDR. We aimed to investigate the expression of MDR-1/P-gp in B-cell lymphoma. Methods and results: Samples at diagnosis and relapse from 10 patients with B-cell lymphoma were obtained. We also obtained 14 unselected control cases of B-cell lymphoma at diagnosis. The expression of mRNA and protein were determined semiquantitatively by RT-PCR and immunohistochemistry. High MDR-1 and P-gp expressions were found in seven and seven of 10 samples obtained at diagnosis, eight and eight of 10 samples obtained at relapse, and three and four of 14 control cases at diagnosis, respectively. The results of RT-PCR paralleled those of immunohistochemistry. Concordance of high MDR-1/P-gp expression was noted in 27 of 34 samples (r = 0.73, P = 0.001). There were no significant changes in MDR-1/P-gp expression in all cases at relapse and during the clinical course following chemotherapy. In the 14 control cases, the average survival time was 12.7 months in MDR-1/P-gp positive cases and 29.0 months in the MDR-1/P-gp negative cases (P = 0.20). Conclusions: Our results showed that at least some B-cell lymphomas express MDR-1/P-gp, which could be detected by different methods, and suggested that high MDR-1/P-gp expression in tumour cells may be associated with a high probability of relapse and poor prognosis. [ABSTRACT FROM AUTHOR]

Background: Though the CXCR2 chemokine receptor is known to play a key role in cancer growth and response to therapy, a direct link between expression of CXCR2 in tumor progenitor cells during induction of tumorigenesis has not been established. Methods: To characterize the role of CXCR2 during melanoma tumorigenesis, we generated tamoxifen-inducible tyrosinase-promoter driven BrafV600E/Pten−/−/Cxcr2−/− and NRasQ61R/INK4a−/−/Cxcr2−/− melanoma models. In addition, the effects of a CXCR1/CXCR2 antagonist, SX-682, on melanoma tumorigenesis were evaluated in BrafV600E/Pten−/− and NRasQ61R/INK4a−/− mice and in melanoma cell lines. Potential mechanisms by which Cxcr2 affects melanoma tumorigenesis in these murine models were explored using RNAseq, mMCP-counter, ChIPseq, and qRT-PCR; flow cytometry, and reverse phosphoprotein analysis (RPPA). Results: Genetic loss of Cxcr2 or pharmacological inhibition of CXCR1/CXCR2 during melanoma tumor induction resulted in key changes in gene expression that reduced tumor incidence/growth and increased anti-tumor immunity. Interestingly, after Cxcr2 ablation, Tfcp2l1, a key tumor suppressive transcription factor, was the only gene significantly induced with a log2 fold-change greater than 2 in these three different melanoma models. Conclusions: Here, we provide novel mechanistic insight revealing how loss of Cxcr2 expression/activity in melanoma tumor progenitor cells results in reduced tumor burden and creation of an anti-tumor immune microenvironment. This mechanism entails an increase in expression of the tumor suppressive transcription factor, Tfcp2l1, along with alteration in the expression of genes involved in growth regulation, tumor suppression, stemness, differentiation, and immune modulation. These gene expression changes are coincident with reduction in the activation of key growth regulatory pathways, including AKT and mTOR. [ABSTRACT FROM AUTHOR]

Dementia is most frequently caused by Alzheimer’s disease. The objective of this study was to investigate the potential targets of Alzheimer’s disease and its associated biological processes. We obtained gene expression profiles of the GSE5281, GSE48350 and GSE11882 datasets from the gene expression Omnibus database. Among the total datasets, 263 were of Alzheimer’s disease tissues and 204 were of healthy tissues. We selected differentially expressed genes from Alzheimer’s disease tissues and healthy tissues using the gene expression Omnibus 2R tool and Venn diagram software. Next, we used database for annotation, visualization and integrated discovery database, gene ontology database and Kyoto encyclopedia of genes and genomes pathways for analysis. Then, by using Cytoscape’s search tool for the retrieval of interacting genes, it is possible to see how these differentially expressed genes and protein-protein interactions interact with one another. In total, there were 270 consistently expressed genes, containing 193 down-regulated genes and 77 up-regulated genes. Additionally, 270 consistently expressed genes were found by chip analysis as the test set and 242 acquisitions of causative differentially expressed genes were sought via gene list automatically derived for you as the training set. ToppGene was then utilized to optimize these genes. Of the protein-protein interaction network, analysed by the molecular complex detection plugin, 14 regulated genes were selected. In conclusion, using integrated bioinformatics techniques, we have attempted to discover the substantially altered expression genes associated with a poorer prognosis in Alzheimer’s disease and the investigations may benefit their predictive and prognostic value in Alzheimer’s disease. [ABSTRACT FROM AUTHOR]

Oxygen deficiency is a critical limiting factor for nosiheptide production in Streptomyces actuosus during fermentation. To alleviate oxygen limitation and enhance the yield of nosiheptide, haemoprotein from Sinorhizobium meliloti (SmHb) was overexpressed in S. actuosus with overexpression of haemoglobin from Vitreoscilla ( VHb) as a positive control. The expression of SmHb and VHb in S. actuosus was confirmed by SDS- PAGE and CO-difference spectra analysis. The results showed that S. actuosus recombinant strain with SmHb expression had higher nosiheptide production (increased by 151%) than the wild strain ( WT) under the low aeration conditions, which was similar with S. actuosus mutant strain with VHb expression. Furthermore, two copies of SmfHb gene were integrated in S. actuosus, which further increased the nosiheptide production by approx. 1·9-fold compared with original strain, and final nosiheptide yield was up to 2352 μg ml−1. These results suggested that engineering of SmHb expression could be used as an efficient method for constructing a high nosiheptide-accumulating strain. Significance and Impact of the Study The significant improvement of nosiheptide production was found in recombinant strain with overexpressed sm gene. And further improvement was obtained in the two copies of sm overexpressing strain. These results suggested that engineering of SmHb expression could be used as an efficient method for constructing a high nosiheptide-accumulating strain. [ABSTRACT FROM AUTHOR]

Abstract: Background: Livers exposed to warm ischemia (WI) are increasingly used for transplantation. The molecular mechanisms activated by WI alone (prior to procurement and transplantation) are not understood. To elucidate the pathways involved, we used microarrays to investigate the gene expression in porcine livers exposed to WI. Methods: Porcine livers (n = 6 group) were randomly subjected to WI periods of 15, 30 or 45 minutes. mRNA was extracted and gene expression determined by microarray analysis. Using bioinformatics software, we identified differentially expressed genes and related molecular pathways. We used the corresponding human annotation of the porcine microarray for the functional analysis. Results: Between 0 and 15 minutes of WI, 3530 genes were altered with a 2-log-fold change of <−0.58 or >+0.58 and P < .05. Between 0 and 30 minutes of WI, 4141 genes were differentially expressed; and between 0 and 45 minutes of WI, 2814 genes. At each time point, ∼50% of genes were up-regulated, whereas 50% were down-regulated. After pathway mapping, we found that the same pathways were induced for observed clustering of in the three WI periods: cell death, proliferation, inflammation, and metabolism pathways. Among the top genes that were up-regulated after 15 minutes of WI, the majority started to return to but did not reach baseline expression with increasing WI. A similar pattern was observed for the top suppressed genes. Conclusions: WI causes rapid changes in gene expression that affect several molecular pathways. This phenomenon seems to plateau at 15 to 30 minutes of WI. These new insights in the timing and the nature of molecular pathways induced by WI alone may help to design specific interventions to alter these changes and improve the outcome of livers from cardiac death donors. [Copyright &y& Elsevier]

Abstract: Proteolytic activity is important in the lifecycles of parasites and their interactions with hosts. Cysteine proteases have been best studied in Giardia, but other protease classes have been implicated in growth and/or differentiation. In this study, we employed bioinformatics to reveal the complete set of putative proteases in the Giardia genome. We identified 73 peptidase homologs distributed over 5 catalytic classes in the genome. Serial analysis of gene expression of the G. lamblia lifecycle found thirteen protease genes with significant transcriptional variation over the lifecycle, with only one serine protease transcript upregulated late in encystation. The translated gene sequence of this encystation-specific transcript was most similar to eukaryotic subtilisin-like proprotein convertases (SPC), although the typical catalytic triad was not identified. Epitope-tagged gSPC protein expressed in Giardia under its own promoter was upregulated during encystation with highest expression in cysts and it localized to encystation-specific secretory vesicles (ESV). Total gSPC from encysting cells produced proteolysis in gelatin gels that co-migrated with the epitope-tagged protease in immunoblots. Immuno-purified gSPC also had gelatinase activity. To test whether endogenous gSPC activity is involved in differentiation, trophozoites and cysts were exposed to the specific serine proteinase inhibitor 4-(2-aminoethyl)-benzenesulfonyl fluoride hydrochloride (AEBSF). After 21h encystation, a significant decrease in ESV was observed with 1mM AEBSF and by 42h the number of cysts was significantly reduced, but trophozoite growth was not inhibited. Concurrently, levels of cyst wall proteins 1 and 2, and AU1-tagged gSPC protein itself were decreased. Excystation of G. muris cysts was also significantly reduced in the presence of AEBSF. These results support the idea that serine protease activity is essential for Giardia encystation and excystation. [Copyright &y& Elsevier]

Abstract: B-cell activating factor belonging to the tumor necrosis factor superfamily (BAFF) is a critical regulator of B-cell maturation and survival. We investigated the expression of BAFF in peripheral blood sample from kidney transplant recipients. Results of flow cytometry showed that at 5 years or more posttransplantation, cell-surface BAFF was significantly expressed on peripheral CD3+ T lymphocytes kidney transplant recipients in compared with other groups (P < .05). BAFF expression was noted on CD4+ and CD8+ T cells. The BAFF messenger RNA level in peripheral mononuclear cells was consistent with the protein level. However, the serum soluble BAFF level varied among individuals in each group. Stratified by renal function, the cell-surface BAFF level was significantly higher in recipients with abnormal renal function compared with those with normal renal function (P < .05). Enzyme-linked immunosorbent assay results showed that expression level of cell-surface BAFF significantly correlated with panel reactive antibody. These results indicate that BAFF may be involved in the development of graft loss. [Copyright &y& Elsevier]

Abstract: Cathepsin K (CatK) is a cysteine protease expressed predominantly in osteoclasts, that plays a prominent role in degrading Type I collagen. Growing CatK null mice have osteopetrosis associated with a reduced ability to degrade bone matrix. Bone strength and histomorphometric endpoints in young adult CatK null mice aged more than 10 weeks have not been studied. The purpose of this paper is to describe bone mass, strength, resorption, and formation in young adult CatK null mice. In male and female wild-type (WT), heterozygous, and homozygous CatK null mice (total N =50) aged 19 weeks, in-life double fluorochrome labeling was performed. Right femurs and lumbar vertebral bodies 1–3 (LV) were evaluated by dual-energy X-ray absorptiometry (DXA) for bone mineral content (BMC) and bone mineral density (BMD). The trabecular region of the femur and the cortical region of the tibia were evaluated by histomorphometry. The left femur and sixth lumbar vertebral body were tested biomechanically. CatK (−/−) mice show higher BMD at the central and distal femur. Central femur ultimate load was positively influenced by genotype, and was positively correlated with both cortical area and BMC. Lumbar vertebral body ultimate load was also positively correlated to BMC. Genotype did not influence the relationship of ultimate load to BMC in either the central femur or vertebral body. CatK (−/−) mice had less lamellar cortical bone than WT mice. Higher bone volume, trabecular thickness, and trabecular number were observed at the distal femur in CatK (−/−) mice. Smaller marrow cavities were also present at the central femur of CatK (−/−) mice. CatK (−/−) mice exhibited greater trabecular mineralizing surface, associated with normal volume-based formation of trabecular bone. Adult CatK (−/−) mice have higher bone mass in both cortical and cancellous regions than WT mice. Though no direct measures of bone resorption rate were made, the higher cortical bone quantity is associated with a smaller marrow cavity and increased retention of non-lamellar bone, signs of decreased endocortical resorption. The relationship of bone strength to BMC does not differ with genotype, indicating the presence of bone tissue of normal quality in the absence of CatK. [Copyright &y& Elsevier]

Contents The consistent failure to isolate bona fide pluripotent cell lines from livestock indicates that the underlying mechanisms of early lineage specification are poorly defined. Unlike other species, the contrivances of segregation have been comprehensively studied in the mouse. In mouse, FGF/ MAPK signalling pathway dictates the segregation of hypoblast (primitive endoderm). However, it is not evident whether this mechanism is also conserved in livestock. Here, in this study, we examined the roles of FGF/ MAP kinase signalling pathways in porcine parthenogenetic embryos during the early development. Porcine parthenogenetic embryos were cultured in the medium addition with FGFR inhibitor BGJ398 (10 μ m) or DEMOS. Pluripotency- and lineage-related gene expressions in the early porcine embryos were determined. Compared to control, total cell numbers on day 7 were significantly higher (55 ± 5.96 vs 47 ± 1.97, p < 0.05) in embryos cultured in the presence of BGJ398, but had no significant effect on the rate of blastocyst development (47% vs 44%, p > 0.05). Nonetheless, BGJ398 treatment significantly augmented the expression of pluripotency and trophoblast marker genes ( SOX2, OCT4, KLF4 and CDX2), but did not significantly change the expression of NANOG and hypoblast marker gene ( GATA4). Furthermore, the addition of FGF signalling agonist ( FGF2) during the embryo development significantly decreased the expression of pluripotency and trophoblast marker genes ( SOX2, NANOG, KLF4 and CDX2), but no significant effect on the expression of OCT4 and GATA4 was observed. Here, we exhibit that inhibition of FGF signalling could improve the quality of the porcine embryo and escalate the chance to capture pluripotency. Besides, it also promotes the trophoblast development of porcine parthenogenetic embryo. In addition, the data suggested that FGF signalling pathway is dispensable for the segregation of hypoblast and epiblast lineages in porcine embryo during the early development. [ABSTRACT FROM AUTHOR]

Papillary thyroid carcinoma (PTC) is the most common type of thyroid carcinoma, and our understanding of its pathogenesis is incomplete. To elucidate the mechanisms underlying such progression and identify novel diagnostic markers, we aimed to discover the underlying gene associated with PTC. Integrated analysis of microarray datasets was performed to identify differentially expressed genes (DEGs) between PTCs and normal tissues. GO enrichment analysis and KEGG pathway enrichment analysis were then performed to uncover the functions of DEGs. Furthermore, the protein-protein interaction (PPI) network of DEGs was constructed. Five GEO datasets were obtained. Totally, 154 DEGs across the studies were identified, including 26 upregulated and 128 downregulated DEGs. In the PPI network, MLLT1, DLG2, and EFEMP1 were the hub proteins, in which DLG2 and EFEMP1 were involved in tumor progression. Among the top 10 up- and down-regulated genes, the dysregulation genes of TPO, CDH16, and MPPED2 may be closely related to the tumorigenesis of PTC. By integrated analysis of multiple gene expression profiles, we propose that the dysregulation genes of TPO and MPPED2 will be the promising diagnostic markers for PTCs. Supporting Information for this article is available online at http://www.thieme-connect.de/products [ABSTRACT FROM AUTHOR]

Background:Aberrant Smad7 expression contributes to the invasion and metastasis of pancreatic cancer cells. However, the potential mechanism underlying aberrant Smad7 expression in human pancreatic ductal adenocarcinoma (PDAC) remains largely unknown.Methods:Bioinformatic prediction programmes and luciferase reporter assay were used to identify microRNAs regulating Smad7. The association between miR-367 expression and the overall survival of PDAC patients was evaluated by Kaplan-Meier analysis. The effects of miR-367 and Smad7 on the invasion and metastasis of pancreatic cancer cells were investigated both in vitro and in vivo.Results:We found that miR-367 downregulated Smad7 expression by directly targeting its 3′-UTR in human pancreatic cancer cells. High level of miR-367 expression correlated with poor prognosis of PDAC patients. Functional studies showed that miR-367 promoted pancreatic cancer invasion in vitro and metastasis in vivo through downregulating Smad7. In addition, we showed that miR-367 promoted epithelial-to-mesenchymal transition by increasing transforming growth factor-β (TGF-β)-induced transcriptional activity.Conclusions:The present study identified and characterised a signalling pathway, the miR-367/Smad7-TGF-β pathway, which is involved in the invasion and metastasis of pancreatic cancer cells. Our results suggest that miR-367 may be a promising therapeutic target for the treatment of human pancreatic cancer. [ABSTRACT FROM AUTHOR]

In this study, the gene and promoter sequences of turbot Scophthalmus maximus (Sm) toll-like receptor 3 (Tlr3) were cloned and its mRNA tissue distribution and gene expression in response to polyinosinic:polycytidylic acid (poly I:C) and turbot reddish body iridovirus ( TRBIV) challenges were studied in vivo. The smtlr3 gene spans over 4·4 kb with a structure of five exons-four introns and encodes a peptide of 916 amino acids. The putative protein shares the highest sequence identity of 52·8-78·5% with fish Tlr3 and contains a signal peptide sequence, 13 leucine-rich repeat ( LRR) motifs, a transmembrane region and a toll/interleukin-1 receptor ( TIR) domain. Phylogenetic analysis grouped it with other teleost Tlr3s. A number of transcription factor binding sites were identified in the 1538 bp 5′ flanking region of smtlr3, including interferon-stimulated response element ( ISRE) and those for interferon regulatory factors ( IRF) and signal transducer and activator of transcriptions ( STATs) smtlr3 transcripts were expressed ubiquitously with higher levels in the head kidney, heart and digestion organs. They were up-regulated by both poly I:C and TRBIV in immune and non-immune organs, but most strongly in the head kidney. Finally, the smtlr3 exhibited a two-wave induced expression during a five day time course when exposure of S. maximus to poly I:C. These findings provide insights into the role of SmTlr3 in antiviral response. [ABSTRACT FROM AUTHOR]

TGF-β has paradoxical effects on cancer cell proliferation, as it suppresses proliferation of normal epithelial and low-invasive cancer cells, but enhances that of high-invasive cancer cells. However, how cancer cells acquire the ability to evade the tumor-suppressing effects of TGF-β, yet still take advantage of its tumor-promoting effects, remains elusive. Here, we identified miR-106b as a molecular switch to determine TGF-β effects on cell proliferation. TGF-β1 enhances the transcription of miR-106b via a promoter independent of its host gene MCM7 by activating c-jun. In high-invasive breast cancer cells, miR-106b is upregulated by TGF-β1 at a much higher level than that in normal or low-invasive cancer cells. Accumulation of miR-106b counterbalances TGF-β growth-inhibiting effects by eliminating activated retinoblastoma (RB) and results in enhanced proliferation. Furthermore, miR-106b mediates TGF-β effects on tumor growth and metastasis in breast cancer xenografts. In addition, miR-106b expression is elevated in higher stage tumors and correlated with tumor progression in breast cancer patients. These findings suggest that high level of miR-106b induced by TGF-β determines the tumor-promoting effects of TGF-β in breast cancer. [ABSTRACT FROM AUTHOR]

The evpP gene in fish pathogen Edwardsiella tarda, coding the T6 SS secretory protein Evp P and carrying an evpA-evpO independent promoter region, was crucial for host cell invasion. The transcription of evpP was positively regulated by either the two-component system Esr A- Esr B or iron concentration, and its overexpression was known to enhance the invasion ability in our previous study. This work demonstrated that the H- NS protein, a pleiotropic regulator of gene expression, was a new transcriptional modulator of evpP gene. The results showed that in vivo the transcriptional level of evpP was downregulated by H- NS and in vitro this global regulator interacted directly with evpP promoter region. Moreover, DNase I footprinting experiments mapping the interaction regions of H- NS and evpP revealed that this global regulator bound to evpP promoter and neighbouring areas at multiple sites. We provided a new insight into evpP regulation network and demonstrated the repression of H- NS to the transcription of evpP gene. Significance and Impact of the Study Recently, the devastating fish disease edwardsiellosis caused by Edwardsiella tarda has been widely concerned. The xenogeneic silencing of the classic regulator H-NS to the T6SS secretory protein EvpP, which played an important role in the virulence of Edw. tarda, was firstly reported in this study. It raised a better understanding of the virulence regulation of EvpP and provided more information about the complex infection mechanism of this pathogen. Our findings would contribute to the development of live attenuated vaccines against edwardsiellosis thus reducing the economic losses caused by this bacterium. [ABSTRACT FROM AUTHOR]

Hepatitis B virus X protein (HBx) plays critical roles in the pathogenesis of hepatocellular carcinoma (HCC). Here, we were interested in knowing whether the oncogene Lin28A and its homolog Lin28B are involved in the hepatocarcinogenesis mediated by HBx. We showed that the expression levels of Lin28A and Lin28B were increased in clinical HCC tissues, HepG2.2.15 cell line and liver tissues of p21-HBx transgenic mice. Interestingly, the expression levels of HBx were positively associated with those of Lin28A/Lin28B in clinical HCC tissues. Moreover, the overexpression of HBx resulted in the upregulation of Lin28A/Lin28B in hepatoma HepG2/H7402 cell lines by transient transfection, suggesting that HBx was able to upregulate Lin28A and Lin28B. Then, we examined the mechanism by which HBx upregulated Lin28A and Lin28B. We identified that the promoter region of Lin28A regulated by HBx was located at nt −235/−66 that contained Sp-1 binding element. Co-immunoprecipitation showed that HBx was able to interact with Sp-1 in HepG2-X cells. Moreover, chromatin immunoprecipitation (ChIP) demonstrated that HBx could bind to the promoter of Lin28A, which failed to work when Sp-1 was silenced. Electrophoretic mobility shift assay (EMSA) further identified that HBx was able to interact with Sp-1 element in Lin28A promoter via transcription factor Sp-1. In addition, we found that c-Myc was involved in the activation of Lin28B mediated by HBx. In function, Lin28A/Lin28B played important roles in HBx-enhanced proliferation of hepatoma cells in vitro and in vivo. In conclusion, HBx activates Lin28A/Lin28B through Sp-1/c-Myc in hepatoma cells. Lin28A/Lin28B serves as key driver genes in HBx-induced hepatocarcinogenesis. [ABSTRACT FROM AUTHOR]

Rac1b, an alternative splice form of Rac1, has been previously shown to be upregulated in colon and breast cancer cells, suggesting an oncogenic role for Rac1b in these cancers. Our analysis of NSCLC tumor and matched normal tissue samples indicates Rac1b is upregulated in a significant fraction of lung tumors in correlation with mutational status of K-ras. To directly assess the oncogenic potential of Rac1b in vivo, we employed a mouse model of lung adenocarcinoma, in which the expression of Rac1b can be conditionally activated specifically in the lung. Although expression of Rac1b alone is insufficient to drive tumor initiation, the expression of Rac1b synergizes with an oncogenic allele of K-ras resulting in increased cellular proliferation and accelerated tumor growth. Finally, we show that in contrast to our previous findings demonstrating a requirement for Rac1 in K-ras-driven cell proliferation, Rac1b is not required in this context. Given the partially overlapping spectrum of downstream effectors regulated by Rac1 and Rac1b, our findings further delineate the signaling pathways downstream of Rac1 that are required for K-ras driven tumorigenesis. [ABSTRACT FROM AUTHOR]

SUMMARY The goal of this study was to explore the molecular mechanism of hypoxia inducible factor-1 alpha (HIF-1 alpha) action on migration and invasion of esophageal carcinoma cells. We used cobalt chloride (CoCl2) to mimic tumor hypoxic microenvironment and analyzed the expressions of E-cadherin, matrix metalloproteinase-2 (MMP-2), and HIF-1 alpha in esophageal carcinoma cells under hypoxia by reverse transcription polymerase chain reaction and Western blotting. To analyze the function of HIF-1 alpha in Eca109 and TE1 cells, we established stable HIF-1 alpha knockdown cells using small interfering RNA. Blocking effect was detected by Western blotting. The concentrations of MMP-2 protein in the conditioned medium were also determined by enzyme-linked immunosorbent assay. Wound-healing and cell invasion assay were used to evaluate the migration and invasion of esophageal carcinoma cells. After exposure to hypoxia, expressions of HIF-1 alpha protein in Eca109 and TE1 cells were upregulated, both mRNA and protein levels of E-cadherin were downregulated, and MMP-2 were upregulated ( P < 0.05), whereas HIF-1 alpha mRNA had no significant change ( P > 0.05). Small interfering RNA could block HIF-1 alpha effectively under hypoxia, then enhanced E-cadherin expression and inhibited MMP-2 expression, respectively. Furthermore, expression of HIF-1 alpha protein was stable even though MMP-2 repressed by BB2516. Compared with that in normoxia, Snail expression was enhanced when Eca109 or TE1 cells exposed to hypoxia. Once HIF-1 alpha blocked, Snail expressions were inhibited accordingly. Wound recovery and the number of invading cells decreased ( P < 0.05) after HIF-1 alpha blocked. The hypoxia suppresses E-cadherin expression and enhances MMP-2 expression favoring esophageal carcinoma migration and invasion via HIF-1 alpha activation. Our observations suggest that HIF-1 alpha inhibition might be an effective strategy to weaken the migration and invasion of esophageal carcinoma cells. [ABSTRACT FROM AUTHOR]

The alterations induced in dendritic cells (DCs) in the cancer microenvironment have not been extensively explored. We found that the tumor-associated factor TGF-β may selectively upregulate the expression of miR-27a via the SP1 transcription factor. Importantly, miR-27a altered the activity of NF-κB and MAPKs (mitogen-activated protein kinases) p38, JNK (c-Jun N-terminal kinases) and ERK (extracellular signal-regulated kinase 1/2). It influences the production of proinflammatory cytokines by targeting TAB3, p38 MAPK, MAP2K4 and MAP2K7. As a consequence, miR-27a hampered the DC-mediated differentiation of Th1 and Th17 cells in vitro and in vivo, but it promoted the DC-mediated accumulation of Tr1 (CD4+IL-10+) and Treg (CD4+CD25+Foxp3+) cells in vivo. The repeated infusion of miR-27a-engineered DCs into tumor tissues accelerated tumor growth, indicating that miR-27a is a potential target for tumor immunotherapy. [ABSTRACT FROM AUTHOR]

Two recent genome-wide association studies of East Asian populations revealed three genetic variants in WDFY4/LRRC18 associated with systemic lupus erythematosus (SLE). To identify the gene contributing to this disease susceptibility, we examined the mRNA expression of WDFY4 and LRRC18 in patients with SLE and healthy controls. WDFY4 was significantly downregulated in SLE patients as compared with controls. We used allelic expression and dual-luciferase assays to identify the functional variant. Transcriptional activity was lower for the rs877819A than -G allele. Electrophoretic mobility shift and supershift assays revealed that the transcription factor Yinyang1 (YY1) binds to rs877819, with lower affinity to the A allele, which explained the reduced transcriptional activity. This effect was further confirmed by YY1 small interfering RNA knockdown, overexpression and chromatin immunoprecipitation experiments. rs877819 in WDFY4 might be the functional site associated with SLE by reduced binding of YY1 and downregulating WDFY4 expression. [ABSTRACT FROM AUTHOR]

Objectives: To explore the expression of manganese superoxide dismutase (MnSOD) in esophageal squamous cell carcinoma (ESCC) and its relationship with clinicopathological characteristics and biological behavior. Methods: On 45 patients with ESCC, immunohistochemistry (SP method), reverse transcription-polymerase chain reaction (RT-PCR), and Western blot were used to detect MnSOD protein and mRNA expression in ESCC and in its adjacent normal tissue 5 cm apart from the edge of cancer lesion and without documented microscopic invasive cancer. Meanwhile, the relationship between the pathological features of esophageal cancer and its biological behavior was analyzed. Results: In ESCC and normal esophageal tissue, MnSOD protein expression was identified 31.1% (14/45) and 86.7% (31/45) ( P= 0.000), respectively, with the relative expression levels of MnSOD mRNA 0.310 ± 0.036 and 0.482 ± 0.053 ( P= 0.000), relatively. Western blot study showed that the relative expressions of MnSOD protein in cancer lesion and in adjacent normal tissue were 0.384 ± 0.038 and 0.766 ± 0.041, respectively ( P= 0.000). With longer lesion, deeper invasion, and poorer differentiation, the expression of MnSOD would get lower, indicating that the levels of MnSOD protein and mRNA expression were closely related to the length of lesion, depth of invasion, and degree of differentiation in ESCC ( P < 0.05). Nevertheless, the result showed no association with the presence of lymph node metastasis, cancer location, and macroscopic classification ( P > 0.05). Conclusions: MnSOD protein and mRNA expression both decreased in ESCC, which may be related to carcinogenesis and the development of esophageal cancer. Therefore, detecting the expression of MnSOD in esophageal carcinoma would be of clinical significance in understanding its biological behavior and in guiding therapeutic strategy of esophageal cancer. [ABSTRACT FROM AUTHOR]

Hepatitis C is a devastating disease worldwide. Proteins encoded by the etiologic agent, hepatitis C virus (HCV), are believed to play important roles in HCV-associated pathogenesis. However, the biological functions of the non-structural protein-2 (NS2) encoded by HCV are not well characterized. Here, we show that HCV NS2 protein activates CXCL-8 (interleukin-8, IL-8) transcription in HepG2 cells as measured by reverse transcription-polymerase chain reaction and IL-8 promoter-luciferase reporter assays. Furthermore, when the κB site on the IL-8 promoter was eliminated by mutagenesis or when intracellular NF-κB activity was suppressed by an inhibitor, NS2 did not activate the IL-8 promoter, suggesting a role of NF-κB in this process. These results prompted us to hypothesize that HCV NS2 might be able to activate NF-κB. This hypothesis was tested by determination of NF-κB-driven reporter gene expression and NF-κB p65 subunit subcellular localization after HCV NS2 expression. Indeed, NS2 could up-regulate NF-κB-driven luciferase activity and was associated with p65 nuclear localization. These results demonstrate that HCV NS2 up-regulates IL-8 transcription through NF-κB. This newly identified function increases our understanding of the role of HCV NS2 protein in virus-host interactions. [ABSTRACT FROM AUTHOR]

Background and Purpose: Liver X receptors (LXRs) activate genes that regulate lipid and cholesterol metabolism. LXR agonists were shown recently to also increase murine renin gene expression in vivo. To further examine a link between lipid metabolism, the renin-angiotensin-aldosterone-system and blood pressure regulation, we investigated the effect of a LXR agonist (GW3965) on angiotensin II (Ang II)-mediated vasoreactivity and vascular angiotensin II receptor (ATR) gene expression. Experimental Approach: Arterial blood pressure (BP) was measured during Ang II infusions (1.5 min duration; 0.001-3 microg kg(-1)) in pentobarbital-anesthetized male Sprague-Dawley rats (n = 6-9) after oral administration of GW3965 (10 mg kg(-1), q.d.) or vehicle for 7 - 15 days. Mesenteric arteries and plasma were collected to analyze ATR gene expression and to measure plasma renin activity (PRA) and lipid profile, respectively. Key Results: Basal mean arterial pressure (MAP) was similar between groups. GW3965 dosing blunted the vasopressor effect of Ang II, which was significantly different with the 0.3 and 3 microg kg(-1) doses. No difference in heart rate, PRA or lipid profile was observed between groups. A time-course indicated that ATR type 1 and 2 gene expression of GW3965-treated vs. vehicle-treated rats decreased by 50%, reaching significance for ATR type 2, but not for ATR type 1, at time-points coinciding with BP measurements. Conclusions and Implications: GW3965 decreased Ang II-mediated vasopressor responses coincident with a trend toward reduced ATR gene expression, suggesting that LXR agonists could affect vascular reactivity. [ABSTRACT FROM AUTHOR]

• Both sodium selenite (SS) and coated SS (CSS) addition increased dry matter intake. • Only CSS supplement increased average daily gain and reduced feed conversion ratio. • Ruminal fermentation and nutrient digestibility were promoted by SS and CSS supplements. • The CSS supplement increased serum total protein, albumin, triglyceride and total antioxigenic capacity. • The CSS addition was more efficient in stimulating hepatic gene expression related to lipid metabolism than SS addition. This study evaluated the effects of sodium selenite (SS) and coated SS (CSS) addition on performance, ruminal fermentation, nutrient digestibility and hepatic gene expression related to lipid metabolism in dairy bulls. Thirty-six Holstein bulls were randomly divided into three groups: without Se addition, addition of 0.3 mg/kg dry matter (DM) of Se from SS and addition of 0.3 mg/kg DM of Se from CSS, respectively. Dietary Se addition increased DM intake. Higher average daily gain and lower feed conversion ratio was observed for CSS addition. Ruminal pH and acetate to propionate ratio decreased, total volatile fatty acids concentration increased but ammonia-N concentration was unchanged with dietary Se addition. Apparent total-tract digestibility of DM, organic matter and crude protein was highest for CSS addition, followed by SS addition and then control. Bulls receiving Se addition had higher digestibility of ether extract, neutral detergent fibre and acid detergent fibre. Serum glucose and glutathione peroxidase and hepatic expression of peroxisome proliferator-activated receptor-α, sterol regulatory element-binding protein-1, carnitine palmitoyl transferase-1 and acetyl-CoA carboxylase-α was highest for CSS addition, followed by SS addition and then control. Bulls receiving CSS addition had higher serum total protein, albumin, triglyceride and total antioxigenic capacity. Addition of Se increased serum cholesterol, insulin and Se. Hepatic expression of stearoyl-CoA desaturase and 3‑hydroxy‑3-methylglutaryl-CoA reductase and Se content increased but content of total lipid and triglyceride was unchanged with Se addition. The data indicated that addition of Se increased performance and hepatic expression of gene related to lipid metabolism in bulls. [ABSTRACT FROM AUTHOR]