Cartilage oligomeric matrix protein in idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a progressive and life threatening disease with median survival of 2.5-3 years. The IPF lung is characterized by abnormal lung remodeling, epithelial cell hyperplasia, myofibroblast foci formation, and extracellular matrix deposition. Analysis of gene expression microarray data revealed that cartilage oligomeric matrix protein (COMP), a non-collagenous extracellular matrix protein is among the most significantly up-regulated genes (Fold change 13, p-value <0.05) in IPF lungs. This finding was confirmed at the mRNA level by nCounter® expression analysis in additional 115 IPF lungs and 154 control lungs as well as at the protein level by western blot analysis. Immunohistochemical analysis revealed that COMP was expressed in dense fibrotic regions of IPF lungs and co-localized with vimentin and around pSMAD3 expressing cells. Stimulation of normal human lung fibroblasts with TGF-β1 induced an increase in COMP mRNA and protein expression. Silencing COMP in normal human lung fibroblasts significantly inhibited cell proliferation and negatively impacted the effects of TGF-β1 on COL1A1 and PAI1. COMP protein concentration measured by ELISA assay was significantly increased in serum of IPF patients compared to controls. Analysis of serum COMP concentrations in 23 patients who had prospective blood draws revealed that COMP levels increased in a time dependent fashion and correlated with declines in force vital capacity (FVC). Taken together, our results should encourage more research into the potential use of COMP as a biomarker for disease activity and TGF-β1 activity in patients with IPF. Hence, studies that explore modalities that affect COMP expression, alleviate extracellular matrix rigidity and lung restriction in IPF and interfere with the amplification of TGF-β1 signaling should be persuaded. © 2013 Vuga et al

Optimization of BY2 cell suspension as a stable transformable system

Tobacco (Nicotiana tabacum) cv. ‘Bright Yellow 2’ (BY2) cell suspension is a useful system to study the structure and function of plant cell. However, low efficiency of Agrobacterium-mediated transformation, and transgene silencing during subculture limit its application. Here we present optimization of the traditional protocols of Agrobacterium-mediated transformation and genomic DNA extraction. The transforming efficiency and recovery ratio of genomic DNA extraction were substantially increased by these improvements. Southern assay demonstrated that copy number of transgene could be determined unambiguously. Meanwhile by monitoring the GFP fluorescence we found that the GFP expression can keep stable in suspension culture cells for at least 20 days in liquid medium. Finally, applicability of constitutive promoters of Arabidopsis thaliana UBIQUITIN10 (AtUBQ10) and ARABIDOPSISSKP1 HOMOLOGUE1 (AtASK1) also can drive stable GFP expression in vivo of BY2 cells like CaMV 35S promoter in this plant system./span

Transcriptional Regulation of Frizzled-1 in Human Osteoblasts by Sp1.

The wingless pathway has a powerful influence on bone metabolism and is a therapeutic target in skeletal disorders. Wingless signaling is mediated in part through the Frizzled (FZD) receptor family. FZD transcriptional regulation is poorly understood. Herein we tested the hypothesis that Sp1 plays an important role in the transcriptional regulation of FZD1 expression in osteoblasts and osteoblast mineralization. To test this hypothesis, we conducted FZD1 promoter assays in Saos2 cells with and without Sp1 overexpression. We found that Sp1 significantly up-regulates FZD1 promoter activity in Saos2 cells. Chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift (EMSA) assays identified a novel and functional Sp1 binding site at -44 to -40 from the translation start site in the FZD1 promoter. The Sp1-dependent activation of the FZD1 promoter was abolished by mithramycin A (MMA), an antibiotic affecting both Sp1 binding and Sp1 protein levels in Saos2 cells. Similarly, down-regulation of Sp1 in hFOB cells resulted in less FZD1 expression and lower alkaline phosphatase activity. Moreover, over-expression of Sp1 increased FZD1 expression and Saos2 cell mineralization while MMA decreased Sp1 and FZD1 expression and Saos2 cell mineralization. Knockdown of FZD1 prior to Sp1 overexpression partially abolished Sp1 stimulation of osteoblast differentiation markers. Taken together, our results suggest that Sp1 plays a role in human osteoblast differentiation and mineralization, which is at least partially mediated by Sp1-dependent transactivation of FZD1

A new species of Argentina (Rosaceae, Potentilleae) from Southeast Tibet, with reference to the taxonomic status of the genus

Argentina songzhuensis, a new species of tribe Potentilleae (Rosaceae) from Southeast Tibet, is described and illustrated. Placement of this species within Argentina was assessed based on morphological comparisons with related species and on phylogenetic analyses of nuclear ITS and plastid trnL-F sequences. The new species is similar to A. glabriuscula, but can be easily distinguished by its densely hairy leaflets, number of stamens (ca.10-12) and decurrent bases of the uppermost pair of leaflets. Our results support the generic status of Argentina and suggest that ventral stipular auricles may be a good synapomorphy for the genus. The recent transfer to Argentina of several Sibbaldia species as well as the genera Piletophyllum and Tylosperma is also confirmed by our molecular phylogenetic analyses. A taxonomic key to A. songzhuensis and other Argentina species in Tibet is provided

Chloroplast phylogeography of the East Asian Arcto-Tertiary relict Tetracentron sinense (Trochodendraceae)

Aim: A phylogeographical study of the widespread but phylogenetically isolated East Asian endemic tree species Tetracentron sinense (Trochodendraceae) was performed to evaluate whether and how Pleistocene and pre-Pleistocene climate changes helped to influence current phylogeographical patterns, and to describe the current patterns of genetic diversity and their implications for conservation. Location: Southwestern and central subtropical China. Methods: Sequences of four chloroplast spacer regions were obtained from 157 individuals of T. sinense. A haplotype network was constructed using tcs. Genetic diversity and differentiation, spatial analysis of molecular variance (SAMOVA) and analysis of molecular variance (AMOVA) were used to test for genetic structure. beast was used to estimate the divergence times between haplotypes. Historical demographic expansion was tested using pairwise mismatch distribution analysis. Results: Of the 21 recovered haplotypes, three were widely distributed, but most were restricted to particular regions. Populations with high haplotype diversity were located in western Hubei, southern Sichuan and southern Chongqing. The two earliest-diverging haplotypes were found in southwestern China. The haplotype distribution of T. sinense demonstrated significant phylogeographical structure (NST \u3e GST; P \u3c 0.05). The best partitioning of genetic diversity by SAMOVA (K = 5) produced groups that matched the main tcs-derived clades. Two independent range expansions within SAMOVA-derived groups 2 and 3 were dated to approximately 399 and 311 ka, respectively. The time to the most recent common ancestor of all haplotypes was 9.6 (95% highest posterior density: 27.0–2.2) Ma, but most of the haplotype diversity appeared during the Quaternary. Main conclusions: The extant distribution of T. sinense is likely to have been shaped by both pre-Quaternary and Pleistocene climate changes. Southwestern China may have served as an important refugium for T. sinense throughout the Neogene, while the species also occupied multiple refugia during the late Pleistocene glacial periods. Populations of T. sinense were resolved into five allopatric groups, between which there is apparently no seed movement

Efficient generation of male germ-like cells derived during co-culturing of adipose-derived mesenchymal stem cells with Sertoli cells under retinoic acid and testosterone induction

Abstract Background Adipose-derived mesenchymal stem cells (ADMSCs) are considered an efficient and important candidate for male infertility treatment because they contain pluripotent stem cells, which can differentiate into all cells from three germ layers. However, the efficient generation of male germ-like cell (MGLCs) is one of the key issues, and little is known about the mechanisms underlying generation of MGLCs. Herein, we attempt to improve the efficient generation of MGLCs derived during co-culturing of rat ADMSCs with SCs under retinoic acid (RA) and testosterone (T) treatment. Methods ADMSCs isolated from male SD rat were induced into generation of MGLCs by using respective methods in vitro. Transwell insert system was used for co-culturing. Busulfan-induced non-obstructive azoospermia rat mode was used to evaluate spermatogenic recovery ability of treated ADMSCs. Besides, the relative gene expression level was detected by reverse transcription PCR, quantitative RT-PCR. The relative protein expression level was detected by western blot (WB) and immunostaining analysis. Results The results showed that ADMSCs co-cultured with TM4 cells under RA and T induction enhanced the formation of bigger and tightly packed MGLCs feature colonies in vitro. Moreover, the expression of male germ cell-related markers (Oct4, Stella, Ddx4, Dazl, PGP9.5, Stra8, and ITGα6) is significantly upregulated in TM4 cell-co-cultured ADMSCs in vitro and in busulfan-treated rat testis after injecting TM4 cell-treated ADMSCs for 2 months. Comparatively, the ADMSCs treated by TM4 cell with RA and T exhibited the highest expression of male germ cell-related markers. RA- and T-treated TM4 cell showed fewer dead cells and higher cytokine secretion than untreated groups. The protein expression level of TGFβ-SMAD2/3, JAK2-STAT3, and AKT pathways in ADMSCs co-cultured with TM4 cells under RA and T was higher than others. Whereas, downregulation of male germ cell-related marker expression subsequently inhibited the phosphorylation of SMAD2/3, JAK2, STAT3, and AKT. Conclusion These results suggested that TM4 cells could efficiently stimulate in vitro generation of MGLCs during co-culturing of ADMSCs under RA and T treatment. Conclusively, the ADMSCs co-cultured with TM4 cell under RA and T induction stimulate the efficient generation of MGLCs in vitro through activating TGFβ-SMAD2/3, JAK2-STAT3, and AKT pathways. Among them, JAK2-STAT3 and AKT pathways are being first reported to show involvement of in vitro generation of MGLCs during ADMSC co-culturing with SCs

Low-carbon and energy-efficient strategy to convert CO2 into carbons with tunable graphitization degree as lithium storage materials toward ultra-long cycle life

Converting greenhouse gas CO2 into high-performance energy storage materials is of great significance due to its capability of simultaneously addressing environmental issues and energy crises. However, great challenges still remain for the low-carbon and energy-efficient conversion of CO2. Herein, we report an energy-efficient and time-saving strategy for converting CO2 into carbon materials by the reaction of CO2 with Mg(AlH4)2 at about 126–136 ℃ in seconds. The chemical reaction of CO2 with Mg(AlH4)2 is reported for the first time. Importantly, the graphitization degree and pore structure of as-synthesized carbon materials are found to be regulated by CO2 pressure. As a lithium storage application, the graphitization degree-dependent electrochemical performance is revealed for the carbon anodes. The highly graphitized carbon prepared at high CO2 pressure exhibits high capacity and ultra-long cycle life, delivering a high reversible capacity of 487 mAh g−1 at 1.0 A g−1 after 3500 cycles. This study develops a green and low-temperature strategy to synthesize CO2-derived carbon materials with tunable graphitization degree for energy storage

Sp1 knockdown by siRNA in hFOB and Saos2 cells reduces FZD1 expression and osteoblast differentiation.

<p>(A) Western blot analysis of Sp1 and FZD1 expression in hFOB cells transfected with 50 nM Sp1 siRNA or scramble siRNA as a control. (B) Real-time quantitative PCR for Sp1, FZD1 and osteoblast differentiation associated genes in hFOB cells transfected with 50 nM Sp1 siRNA or scramble siRNA as a control. (C) ALP activity staining with BCIP/NBT substrate solution (Sigma, USA) in Saos2 and hFOB cells transfected with 50 nM Sp1 siRNA or scramble siRNA. (D) Quantitative results of the ALP staining for Fig 4C by densitometry. All siRNA knockdown experiments were conducted in triplicates and repeated for 2 independent times. * indicates significant difference between the control siRNA and Sp1 specific siRNA treated cells (<i>P</i> < 0.05).</p