scholarly journals miR-34s inhibit osteoblast proliferation and differentiation in the mouse by targeting SATB2

2012 ◽  
Vol 197 (4) ◽  
pp. 509-521 ◽  
Author(s):  
Jianwen Wei ◽  
Yu Shi ◽  
Lihua Zheng ◽  
Bin Zhou ◽  
Hiroyuki Inose ◽  
...  
Keyword(s):  
Osteoblast Differentiation ◽  
Matrix Protein ◽  
Molecular Mechanisms ◽  
Loss Of Function ◽  
Osteoblast Proliferation ◽  
Nuclear Matrix Protein ◽  
Critical Determinant ◽  
Proliferation And Differentiation ◽  
Bone Mass Accrual

A screen of microRNAs preferentially expressed in osteoblasts identified members of the miR-34 family as regulators of osteoblast proliferation and/or differentiation. Osteoblast-specific gain- and loss-of-function experiments performed in vivo revealed that miR-34b and -c affected skeletogenesis during embryonic development, as well as bone mass accrual after birth, through two complementary cellular and molecular mechanisms. First, they inhibited osteoblast proliferation by suppressing Cyclin D1, CDK4, and CDK6 accumulation. Second, they inhibited terminal differentiation of osteoblasts, at least in part through the inhibition of SATB2, a nuclear matrix protein that is a critical determinant of osteoblast differentiation. Genetic evidence obtained in the mouse confirmed the importance of SATB2 regulation by miR-34b/c. These results are the first to identify a family of microRNAs involved in bone formation in vivo and to identify a specific genetic pathway by which these microRNAs regulate osteoblast differentiation.

scholarly journals Assessment of phagocytic activity in live macrophages-tumor cells co-cultures by Confocal and Nomarski Microscopy

2017 ◽  
Vol 2 (1) ◽  
Author(s):  
Dalia Martinez-Marin ◽  
Courtney Jarvis ◽  
Thomas Nelius ◽  
Stéphanie Filleur
Keyword(s):  
Tumor Microenvironment ◽  
Tumor Cells ◽  
Phagocytic Activity ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Functional Assay ◽  
Loss Of Function ◽  
Multiple Cancer

Abstract Macrophages have been recognized as the main inflammatory component of the tumor microenvironment. Although often considered as beneficial for tumor growth and disease progression, tumor-associated macrophages have also been shown to be detrimental to the tumor depending on the tumor microenvironment. Therefore, understanding the molecular interactions between macrophages and tumor cells in relation to macrophages functional activities such as phagocytosis is critical for a better comprehension of their tumor-modulating action. Still, the characterization of these molecular mechanisms in vivo remains complicated due to the extraordinary complexity of the tumor microenvironment and the broad range of tumor-associated macrophage functions. Thus, there is an increasing demand for in vitro methodologies to study the role of cell–cell interactions in the tumor microenvironment. In the present study, we have developed live co-cultures of macrophages and human prostate tumor cells to assess the phagocytic activity of macrophages using a combination of Confocal and Nomarski Microscopy. Using this model, we have emphasized that this is a sensitive, measurable, and highly reproducible functional assay. We have also highlighted that this assay can be applied to multiple cancer cell types and used as a selection tool for a variety of different types of phagocytosis agonists. Finally, combining with other studies such as gain/loss of function or signaling studies remains possible. A better understanding of the interactions between tumor cells and macrophages may lead to the identification of new therapeutic targets against cancer.

scholarly journals Direct observation of structure and dynamics during phase separation of an elastomeric protein

2017 ◽  
Vol 114 (22) ◽  
pp. E4408-E4415 ◽  
Author(s):  
Sean E. Reichheld ◽  
Lisa D. Muiznieks ◽  
Fred W. Keeley ◽  
Simon Sharpe
Keyword(s):  
Phase Separation ◽  
Matrix Protein ◽  
Molecular Mechanisms ◽  
Bulk Water ◽  
Extracellular Matrix Protein ◽  
Elastic Fibers ◽  
Cross Linking ◽  
Structure And Dynamics ◽  
Chemical Cross Linking

Despite its growing importance in biology and in biomaterials development, liquid–liquid phase separation of proteins remains poorly understood. In particular, the molecular mechanisms underlying simple coacervation of proteins, such as the extracellular matrix protein elastin, have not been reported. Coacervation of the elastin monomer, tropoelastin, in response to heat and salt is a critical step in the assembly of elastic fibers in vivo, preceding chemical cross-linking. Elastin-like polypeptides (ELPs) derived from the tropoelastin sequence have been shown to undergo a similar phase separation, allowing formation of biomaterials that closely mimic the material properties of native elastin. We have used NMR spectroscopy to obtain site-specific structure and dynamics of a self-assembling elastin-like polypeptide along its entire self-assembly pathway, from monomer through coacervation and into a cross-linked elastic material. Our data reveal that elastin-like hydrophobic domains are composed of transient β-turns in a highly dynamic and disordered chain, and that this disorder is retained both after phase separation and in elastic materials. Cross-linking domains are also highly disordered in monomeric and coacervated ELP3 and form stable helices only after chemical cross-linking. Detailed structural analysis combined with dynamic measurements from NMR relaxation and diffusion data provides direct evidence for an entropy-driven mechanism of simple coacervation of a protein in which transient and nonspecific intermolecular hydrophobic contacts are formed by disordered chains, whereas bulk water and salt are excluded.

Mesenchymal folliculin is required for alveolar development: implications for cystic lung disease in Birt-Hogg-Dubé syndrome

Thorax ◽  
2020 ◽  
Vol 75 (6) ◽  
pp. 486-493 ◽  
Author(s):  
Ling Chu ◽  
Yongfeng Luo ◽  
Hui Chen ◽  
Qing Miao ◽  
Larry Wang ◽  
...  
Keyword(s):  
Lung Disease ◽  
Molecular Mechanisms ◽  
Statistical Significance ◽  
Mesenchymal Progenitor Cells ◽  
Loss Of Function ◽  
Cystic Lung ◽  
Alveolar Development ◽  
Cystic Lung Disease ◽  
Pulmonary Cysts

BackgroundPulmonary cysts and spontaneous pneumothorax are presented in most patients with Birt-Hogg-Dubé (BHD) syndrome, which is caused by loss of function mutations in the folliculin (FLCN) gene. The pathogenic mechanisms underlying the cystic lung disease in BHD are poorly understood.MethodsMesenchymal Flcn was specifically deleted in mice or in cultured lung mesenchymal progenitor cells using a Cre/loxP approach. Dynamic changes in lung structure, cellular and molecular phenotypes and signalling were measured by histology, immunofluorescence staining and immunoblotting.ResultsDeletion of Flcn in mesoderm-derived mesenchymal cells results in significant reduction of postnatal alveolar growth and subsequent alveolar destruction, leading to cystic lesions. Cell proliferation and alveolar myofibroblast differentiation are inhibited in the Flcn knockout lungs, and expression of the extracellular matrix proteins Col3a1 and elastin are downregulated. Signalling pathways including mTORC1, AMP-activated protein kinase, ERK1/2 and Wnt-β-catenin are differentially affected at different developmental stages. All the above changes have statistical significance (p<0.05).ConclusionsMesenchymal Flcn is an essential regulator during alveolar development and maintenance, through multiple cellular and molecular mechanisms. The mesenchymal Flcn knockout mouse model provides the first in vivo disease model that may recapitulate the stages of cyst development in human BHD. These findings elucidate the developmental origins and mechanisms of lung disease in BHD.

scholarly journals IMMU-28. DEFINING MOLECULAR MECHANISMS OF RESISTANCE TO GLIOBLASTOMA (GBM) IMMUNITY USING A NOVEL CRISPR/CAS9 IN VIVO LOSS-OF-FUNCTION SCREENING PLATFORM

2017 ◽  
Vol 19 (suppl_6) ◽  
pp. vi118-vi118
Author(s):  
Martha R Neagu ◽  
Maria Carmela Speranza ◽  
Robert T Manguso ◽  
Sean E Lawler ◽  
Gordon J Freeman ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Loss Of Function ◽  
Mechanisms Of Resistance ◽  
Screening Platform

scholarly journals miR-26a/HOXC9 Dysregulation Promotes Metastasis and Stem Cell-Like Phenotype of Gastric Cancer

2018 ◽  
Vol 49 (4) ◽  
pp. 1659-1676 ◽  
Author(s):  
Xudong Peng ◽  
Qingjie Kang ◽  
Rui Wan ◽  
Ziwei Wang
Keyword(s):  
Gastric Cancer ◽  
Stem Cell ◽  
Molecular Mechanisms ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Promoting Effect ◽  
In Vivo Experiments ◽  
Direct Target ◽  
Cancer Tissues

Background/Aims: Previous studies demonstrated that HOXC9 acts as an oncogene in several tumors. The aim of this study was to explore whether HOXC9 promotes gastric cancer (GC) progression and elucidate the underlying molecular mechanisms. Methods: HOXC9 expression in GC tissues and adjacent non-cancer tissues was detected by quantitative RT-PCR (qRT-PCR) and immunohistochemistry. The functional effects of HOXC9 on proliferation, metastasis and stem cell-like phenotype were evaluated by relevant experiments in GC cells. The effect of miR-26a on HOXC9 was investigated by gain- and loss-of-function assays and luciferase reporter assay. Nude mouse models were established to test the effect of miR-26a and HOXC9 on tumorigenesis and metastasis of GC cells in vivo. Results: Herein, we showed that HOXC9 was upregulated in GC tissues and associated with a poor prognosis. HOXC9 knockdown inhibited the metastasis and stem cell-like phenotype of GC cells without significant effects on cell proliferation. In addition, we identifed HOXC9 as a direct target of miR-26a. Restoration of miR-26a in GC cells downregulated HOXC9 and reversed its promoting effect on metastasis and self-renewal, whereas miR-26a silencing upregulated HOXC9. In vivo experiments showed that HOXC9 knockdown suppressed tumorigenesis and lung metastasis of GC cells in nude mice, and these effects were mimicked by restoration of miR-26a. Conclusion: The present study demonstrates that HOXC9 promotes the metastasis and stem cell-like phenotype of GC cells, and this phenomenon can be reversed by restoration of miR-26a.

scholarly journals Effects of Titanium Micro-Nanopermeable Structures on Osteogenic Differentiation

2018 ◽  
Vol 2018 ◽  
pp. 1-11 ◽  
Author(s):  
Jiannan Liu ◽  
Mingjiang Jin ◽  
Zhen Zhang ◽  
Lihuang Wu ◽  
Xuejun Jin ◽  
...  
Keyword(s):  
Osteoblast Differentiation ◽  
Signal Pathway ◽  
Acid Etching ◽  
Porous Structures ◽  
Osteoblast Proliferation ◽  
Activity Assay ◽  
Alp Activity ◽  
Erk Phosphorylation ◽  
Proliferation And Differentiation ◽  
Mapk Signal Pathway

To evaluate the effects of different Ti surface micro-nanopermeable structures on osteoblast proliferation and differentiation and explore related mechanisms, hybrid technology of sandblast, acid etching, and hydrothermal (HT) was used to form the micro-nanopermeable surface of Ti. Scanning electron microscopy (SEM), surface profiler, and contact angle meter were utilized to assess the surface morphology, roughness, and hydrophilicity. MTT, SEM, alkaline phosphatase (ALP) activity assay, and real-time PCR were performed to investigate proliferation, adhesion and spreading, and differentiation of MC3T3-E1 cells grown on polished Ti (control), sandblast + acid etching- (SLA-) treated Ti, and SLA + HT-treated Ti. MAPK signal pathway activity was evaluated by Western blotting. The results showed that SLA + HT could result in not only formation of microscale groove containing submicroscale and nanoscale porous structures in Ti samples but also rough and hydrophilic surface. SLA + HT treatment has the best effects on cell adhesion and spreading. Significantly increased levels of ALP activity and osteogenic genes including Alp, Ocn, Opn, Runx2, and Bsp, as well as p38 but not ERK phosphorylation, were found in the SLA + HT group. In conclusion, sandblast, acid etching, and hydrothermal treatment on Ti regulates osteoblast differentiation, while activation of the MAPK p38 signaling pathway served as the mechanism.

The ubiquitin proteasome system in atrophying skeletal muscle: roles and regulation

2016 ◽  
Vol 311 (3) ◽  
pp. C392-C403 ◽  
Author(s):  
Philippe A. Bilodeau ◽  
Erin S. Coyne ◽  
Simon S. Wing
Keyword(s):  
Signaling Pathways ◽  
Muscle Atrophy ◽  
Molecular Mechanisms ◽  
Muscle Wasting ◽  
Clinical Problem ◽  
Ubiquitin Proteasome System ◽  
Mechanisms Of Action ◽  
Loss Of Function ◽  
Critical Determinant ◽  
Ubiquitin Proteasome

Muscle atrophy complicates many diseases as well as aging, and its presence predicts both decreased quality of life and survival. Much work has been conducted to define the molecular mechanisms involved in maintaining protein homeostasis in muscle. To date, the ubiquitin proteasome system (UPS) has been shown to play an important role in mediating muscle wasting. In this review, we have collated the enzymes in the UPS whose roles in muscle wasting have been confirmed through loss-of-function studies. We have integrated information on their mechanisms of action to create a model of how they work together to produce muscle atrophy. These enzymes are involved in promoting myofibrillar disassembly and degradation, activation of autophagy, inhibition of myogenesis as well as in modulating the signaling pathways that control these processes. Many anabolic and catabolic signaling pathways are involved in regulating these UPS genes, but none appear to coordinately regulate a large number of these genes. A number of catabolic signaling pathways appear to instead function by inhibition of the insulin/IGF-I/protein kinase B anabolic pathway. This pathway is a critical determinant of muscle mass, since it can suppress key ubiquitin ligases and autophagy, activate protein synthesis, and promote myogenesis through its downstream mediators such as forkhead box O, mammalian target of rapamycin, and GSK3β, respectively. Although much progress has been made, a more complete inventory of the UPS genes involved in mediating muscle atrophy, their mechanisms of action, and their regulation will be useful for identifying novel therapeutic approaches to this important clinical problem.

scholarly journals Sox2 induction by FGF and FGFR2 activating mutations inhibits Wnt signaling and osteoblast differentiation

2005 ◽  
Vol 168 (7) ◽  
pp. 1065-1076 ◽  
Author(s):  
Alka Mansukhani ◽  
Davide Ambrosetti ◽  
Greg Holmes ◽  
Lizbeth Cornivelli ◽  
Claudio Basilico
Keyword(s):  
Osteoblast Differentiation ◽  
Target Genes ◽  
Expression Profiles ◽  
Constitutive Expression ◽  
Growth Factor Receptor ◽  
Gene Expression Profiles ◽  
Down Regulation ◽  
Activating Mutations ◽  
Proliferation And Differentiation

Activating mutations in fibroblast growth factor receptor 2 (FGFR2) cause several craniosynostosis syndromes by affecting the proliferation and differentiation of osteoblasts, which form the calvarial bones. Osteoblasts respond to FGF with increased proliferation and inhibition of differentiation. We analyzed the gene expression profiles of osteoblasts expressing FGFR2 activating mutations (C342Y or S252W) and found a striking down-regulation of the expression of many Wnt target genes and a concomitant induction of the transcription factor Sox2. Most of these changes could be reproduced by treatment of osteoblasts with exogenous FGF. Wnt signals promote osteoblast function and regulate bone mass. Sox2 is expressed in calvarial osteoblasts in vivo and we show that constitutive expression of Sox2 inhibits osteoblast differentiation and causes down-regulation of the expression of numerous Wnt target genes. Sox2 associates with β-catenin in osteoblasts and can inhibit the activity of a Wnt responsive reporter plasmid through its COOH-terminal domain. Our results indicate that FGF signaling could control many aspects of osteoblast differentiation through induction of Sox2 and regulation of the Wnt–β-catenin pathway.

scholarly journals TCN1 Deficiency Inhibits The Malignancy of Colorectal Cancer Cells By Regulating The ITGB4 Pathway

2021 ◽  
Author(s):  
Xinqiang Zhu ◽  
Xuetong Jiang ◽  
Qinglin Zhang ◽  
Hailong Huang ◽  
Xiaohong Shi ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Molecular Mechanisms ◽  
Function Analysis ◽  
Hct116 Cell ◽  
Tumor Xenograft ◽  
Integrin Subunit ◽  
Filamin A ◽  
Loss Of Function ◽  
Colorectal Cancer Cells

Abstract Background: This study aimed to investigate the biological function and regulatory mechanism of TCN1 in colorectal cancer (CRC). Methods: We studied the biological functions of TCN1 using gain-of-function and loss-of-function analysis in HCT116 cell lines, and examined the effects of TCN1 on the proliferation, apoptosis, and invasion of CRC cells and determined its potential molecular mechanisms using CRC lines and mouse xenotransplantation models. Tumor xenograft and tumor metastasis studies were performed to detect the tumorigenicity and metastasis of cells in vivo. Results: TCN1-knockdown attenuated CRC cell proliferation, invasion and promoted cell apoptosis. Overexpression of TCN1 yielded the opposite effects. In addition, TCN1-knockdown HCT116 cells failed to form metastatic foci in the peritoneum after intravenous injection. Molecular mechanism studies showed that TCN1 interacts with integrin subunit β4 (ITGB4) to positively regulate the expression of ITGB4. TCN1-knockdown promoted the degradation of ITGB4 and increased the instability of ITGB4 and filamin A (FLNA). Downregulation of ITGB4 at the protein level resulted in the disassociation of the ITGB4/PLEC complex, leading to cytoskeletal damage. Conclusion: TCN1 might exert oncogenic role in CRC via regulating the ITGB4 signaling pathway.

Sign in / Sign up

Export Citation Format

Share Document