Molecular mechanisms of the response to mechanical stimulation during chondrocyte differentiation

2010 ◽  
pp. 53-59
Author(s):  
Ichiro Takahashi ◽  
Taisuke Masuda ◽  
Kumiko Kohsaka ◽  
Fumie Terao ◽  
Takahisa Anada ◽  
...  
Keyword(s):  
Mechanical Stimulation ◽  
Molecular Mechanisms ◽  
Chondrocyte Differentiation

scholarly journals Effect of mechanical loading on insulin-like growth factor-I gene expression in rat tibia

2007 ◽  
Vol 192 (1) ◽  
pp. 131-140 ◽  
Author(s):  
Christianne M A Reijnders ◽  
Nathalie Bravenboer ◽  
Annechien M Tromp ◽  
Marinus A Blankenstein ◽  
Paul Lips
Keyword(s):  
Bone Formation ◽  
Mrna Expression ◽  
Mechanical Loading ◽  
Mechanical Stimulation ◽  
Molecular Mechanisms ◽  
Bone Marrow Cells ◽  
Mechanical Stimuli ◽  
Igf I ◽  
Female Wistar Rats ◽  
Tibia Shaft

Mechanical loading plays an essential role in maintaining skeletal integrity. Mechanical stimulation leads to increased bone formation. However, the cellular and molecular mechanisms that are involved in the translation of mechanical stimuli into bone formation, are not completely understood. Growth factors and osteocytes, which act as mechanosensors, play a key role during the bone formation after mechanical stimulation. The aim of this study was to characterize the role of IGF-I in the translation of mechanical stimuli into bone formation locally in rat tibiae. Fifteen female Wistar rats were randomly assigned to three groups (n = 5): load, sham-loaded, and control. The four-point bending model of Forwood and Turner was used to induce a single period of mechanical loading on the tibia shaft. The effects of mechanical loading on IGF-I mRNA expression were determined with non-radioactive in situ hybridization on decalcified tibiae sections, 6 h after the loading session. Endogenous IGF-I mRNA was expressed in trabecular and cortical osteoblasts, some trabecular and sub-endocortical osteocytes, intracortical endothelial cells of blood vessels, and periosteum. Megakaryocytes, macrophages, and myeloid cells also expressed IGF-I mRNA. In the growth plate, IGF-I mRNA was located in proliferative and hypertrophic chondrocytes. Mechanical loading did not affect the IGF-I mRNA expression in osteoblasts, bone marrow cells, and chondrocytes, but the osteocytes at the endosteal side of the shaft showed a twofold increase of IGF-I mRNA expression. The proportion of IGF-I mRNA positive osteocytes in loaded tibiae was 29.3 ± 12.9% (mean ± s.d.; n = 5), whereas sham-loaded and contra-lateral control tibiae exhibited 16.7 ± 4.4% (n = 5) and 14.7 ± 4.2% (n = 10) respectively (P < 0.05). Lamellar bone formation after a single mechanical loading session was observed at the endosteal side of the shaft. In conclusion, a single loading session results in a twofold up-regulation of IGF-I mRNA synthesis in osteocytes which are present in multiple layers extending into the cortical bone of mechanically stimulated tibia shaft 6 h after loading. This supports the hypothesis that IGF-I, which is located in osteocytes, is involved in the translation of mechanical stimuli into bone formation.

scholarly journals Dexamethasone enhances SOX9 expression in chondrocytes

2001 ◽  
Vol 169 (3) ◽  
pp. 573-579 ◽  
Author(s):  
I Sekiya ◽  
P Koopman ◽  
K Tsuji ◽  
S Mertin ◽  
V Harley ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Molecular Mechanisms ◽  
Consensus Sequence ◽  
Chondrocyte Differentiation ◽  
Sox9 Expression ◽  
Newborn Mice ◽  
Col2a1 Gene ◽  
Dose Dependent ◽  
Sox9 Protein

SOX9 is a transcription factor that activates type II procollagen (Col2a1) gene expression during chondrocyte differentiation. Glucocorticoids are also known to promote chondrocyte differentiation via unknown molecular mechanisms. We therefore investigated the effects of a synthetic glucocorticoid, dexamethasone (DEX), on Sox9 gene expression in chondrocytes prepared from rib cartilage of newborn mice. Sox9 mRNA was expressed at high levels in these chondrocytes. Treatment with DEX enhanced Sox9 mRNA expression within 24 h and this effect was observed at least up to 48 h. The effect of DEX was dose dependent, starting at 0.1 nM and maximal at 10 nM. The half life of Sox9 mRNA was approximately 45 min in the presence or absence of DEX. Western blot analysis revealed that DEX also enhanced the levels of SOX9 protein expression. Treatment with DEX enhanced Col2a1 mRNA expression in these chondrocytes and furthermore, DEX enhanced the activity of Col2-CAT (chloramphenicol acetyltransferase) construct containing a 1.6 kb intron fragment where chondrocyte-specific Sry/Sox- consensus sequence is located. The enhancing effect of DEX was specific to SOX9, as DEX did not alter the levels of Sox6 mRNA expression. These data suggest that DEX promotes chondrocyte differentiation through enhancement of SOX9.

scholarly journals The Effect of Curcumin on the Differentiation of Mesenchymal Stem Cells into Mesodermal Lineage

Molecules ◽  
2019 ◽  
Vol 24 (22) ◽  
pp. 4029
Author(s):  
Armita Mahdavi Gorabi ◽  
Nasim Kiaie ◽  
Saeideh Hajighasemi ◽  
Tannaz Jamialahmadi ◽  
Muhammed Majeed ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Molecular Mechanisms ◽  
Stem Cell Differentiation ◽  
Signal Transduction Pathways ◽  
Chondrocyte Differentiation ◽  
Pharmacological Effects ◽  
Structural Modifications ◽  
Mechanism Of Inhibition ◽  
Mesodermal Lineage

Curcumin has been placed at the forefront of the researcher’s attention due to its pleiotropic pharmacological effects and health benefits. A considerable volume of articles has pointed out curcumin’s effects on the fate of stem cell differentiation. In this review, a descriptive mechanism of how curcumin affects the outcome of the differentiation of mesenchymal stem cells (MSCs) into the mesodermal lineage—i.e., adipocyte, osteocyte, and chondrocyte differentiation—is compiled from the literature. The sections include the mechanism of inhibition or induction of MSCs differentiation to each lineage, their governing molecular mechanisms, and their signal transduction pathways. The effect of different curcumin doses and its structural modifications on the MSCs differentiation is also discussed.

scholarly journals Role of CaMKK2 in Mechanically Induced Bone Formation

2019 ◽  
Vol 2 (1) ◽  
Author(s):  
Adam J. Warrick ◽  
Uma Sankar
Keyword(s):  
Protein Kinase ◽  
Bone Density ◽  
Bone Formation ◽  
Mechanical Stimulation ◽  
Internal Control ◽  
Molecular Mechanisms ◽  
Bone Growth ◽  
Lower Limbs ◽  
Alizarin Red ◽  
Bone Formation Rate

Background and Hypothesis: Mechanical stimulation of bone results in the translation of external forces into a cascade of structural and biochemical changes which work to increase bone density and decrease fracture healing time. The specific mechanisms contributing to these processes are areas of active investigation. Ca2+/calmodulin-dependent protein kinase kinase 2 (CaMKK2) is a serine-threonine protein kinase with key roles in both the anabolic and catabolic pathways of bone remodeling. We hypothesize that the absence of CaMKK2 potentiates an increase in bone density as a response to mechanical stimulation. Experimental Design or Project Methods: The right ulna of anesthetized C57BL/6 mice were loaded for 220 cycles at 2 Hz and with peak forces specific to both sex and genotype. Loading was completed using an electro actuator (Bose ElectroForce 3200; EnduraTEC, Minnetonka, MN, USA) and was repeated on days 3, 5, 8 and 10 after the initial procedure. The non-loaded left ulna served as an internal control. Calcein and alizarin red were administered intraperitoneally on days 9 and 16 respectively. Mice were sacrificed on day 19 after the initial load; blood and long bones of the lower limbs were collected for analysis. Results: Bone volumetric analyses will be measured using microcomputed tomography, bone formation rate will be assessed using dynamic histomorphometry measurements of double fluorochrome labeling, and cellular and molecular mechanisms will be assessed using histology, immunohistochemistry and real-time reverse transcription-polymerase chain reaction. These data are currently forthcoming. Conclusion and Potential Impact: Clinical outcomes of conditions ranging from stress fractures to osteoporosis may be improved by an increased understanding of the mechanisms through which bone growth is augmented. Expanded knowledge of these pathways may provide opportunities for the development of novel therapies which decrease healing times in the event of injury and increase bone density to combat degenerative disease states.

scholarly journals Arid5a cooperates with Sox9 to stimulate chondrocyte-specific transcription

2011 ◽  
Vol 22 (8) ◽  
pp. 1300-1311 ◽  
Author(s):  
Katsuhiko Amano ◽  
Kenji Hata ◽  
Shuji Muramatsu ◽  
Makoto Wakabayashi ◽  
Yoko Takigawa ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Gene Promoter ◽  
Transcriptional Regulators ◽  
Chondrocyte Differentiation ◽  
Col2a1 Gene ◽  
Organ Culture System ◽  
Histone 3 ◽  
Col2a1 Expression ◽  
Essential Transcription Factor

SRY-box–containing gene 9 (Sox9) is an essential transcription factor in chondrocyte lineage determination and differentiation. Recent studies demonstrated that Sox9 controls the transcription of chondrocyte-specific genes in association with several other transcriptional regulators. To further understand the molecular mechanisms by which Sox9 influences transcriptional events during chondrocyte differentiation, we attempted to identify transcriptional partners of Sox9 and to examine their roles in chondrocyte differentiation. We isolated AT-rich interactive domain–containing protein 5a (Arid5a; also known as Mrf1) as an activator of the Col2a1 gene promoter from an ATDC5 cDNA library. Arid5a was highly expressed in cartilage and induced during chondrocyte differentiation. Furthermore, Arid5a physically interacted with Sox9 in nuclei and up-regulated the chondrocyte-specific action of Sox9. Overexpression of Arid5a stimulated chondrocyte differentiation in vitro and in an organ culture system. In contrast, Arid5a knockdown inhibited Col2a1 expression in chondrocytes. In addition, Arid5a binds directly to the promoter region of the Col2a1 gene and stimulates acetylation of histone 3 in the region. Our results suggest that Arid5a may directly interact with Sox9 and thereby enhance its chondrocyte-specific action.

scholarly journals Investigating the mechanistic basis of biomechanical input controlling skeletal development: exploring the interplay with Wnt signalling at the joint

2018 ◽  
Vol 373 (1759) ◽  
pp. 20170329 ◽  
Author(s):  
Rebecca A. Rolfe ◽  
Claire A. Shea ◽  
Pratik Narendra Pratap Singh ◽  
Amitabha Bandyopadhyay ◽  
Paula Murphy
Keyword(s):  
Mechanical Stimulation ◽  
Wnt Pathway ◽  
Molecular Mechanisms ◽  
Skeletal Development ◽  
Wnt Signalling ◽  
Mechanical Stimuli ◽  
Mechanistic Basis ◽  
Bmp Signalling ◽  
Canonical Wnt ◽  
Mechanical Influence

Embryo movement is essential to the formation of a functional skeleton. Using mouse and chick models, we previously showed that mechanical forces influence gene regulation and tissue patterning, particularly at developing limb joints. However, the molecular mechanisms that underpin the influence of mechanical signals are poorly understood. Wnt signalling is required during skeletal development and is altered under reduced mechanical stimulation. Here, to explore Wnt signalling as a mediator of mechanical input, the expression of Wnt ligand and Fzd receptor genes in the developing skeletal rudiments was profiled. Canonical Wnt activity restricted to the developing joint was shown to be reduced under immobilization, while overexpression of activated β-catenin following electroporation of chick embryo limbs led to joint expansion, supporting the proposed role for Wnt signalling in mechanoresponsive joint patterning. Two key findings advance our understanding of the interplay between Wnt signalling and mechanical stimuli: first, loss of canonical Wnt activity at the joint shows reciprocal, coordinated misregulation of BMP signalling under altered mechanical influence. Second, this occurs simultaneously with increased expression of several Wnt pathway component genes in a territory peripheral to the joint, indicating the importance of mechanical stimulation for a population of potential joint progenitor cells. This article is part of the Theo Murphy meeting issue ‘Mechanics of Development’.

scholarly journals Live cell imaging of mechanotransduction

2010 ◽  
Vol 7 (suppl_3) ◽  
Author(s):  
Bo Liu ◽  
Tae-Jin Kim ◽  
Yingxiao Wang
Keyword(s):  
Mechanical Stimulation ◽  
Molecular Mechanisms ◽  
Fluorescent Proteins ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Stem Cell Differentiation ◽  
Cellular Functions ◽  
Signalling Molecules ◽  
Signalling Transduction ◽  
Systematic Understanding

Mechanical forces play important roles in the regulation of cellular functions, including polarization, migration and stem cell differentiation. Tremendous advancement in our understanding of mechanotransduction has been achieved with the recent development of imaging technologies and molecular biosensors. In particular, genetically encoded biosensors based on fluorescence resonance energy transfer (FRET) technology have been widely developed and applied in the field of mechanobiology. In this article, we will provide an overview of the recent progress of FRET application in mechanobiology, specifically mechanotransduction. We first introduce fluorescent proteins and FRET technology. We then discuss the mechanotranduction processes in different cells including stem cells, with a special emphasis on the important signalling molecules involved in mechanotransduction. Finally, we discuss methods that can allow the integration of simultaneous FRET imaging and mechanical stimulation to trigger signalling transduction. In summary, FRET technology has provided a powerful tool for the study of mechanotransduction to advance our systematic understanding of the molecular mechanisms by which cells respond to mechanical stimulation.

scholarly journals p38 MAP kinase signalling is required for hypertrophic chondrocyte differentiation

2004 ◽  
Vol 378 (1) ◽  
pp. 53-62 ◽  
Author(s):  
Lee-Anne STANTON ◽  
Shalev SABARI ◽  
Arthur V. SAMPAIO ◽  
T. Michael UNDERHILL ◽  
Frank BEIER
Keyword(s):  
Molecular Mechanisms ◽  
P38 Map Kinase ◽  
Limb Bud ◽  
Marker Genes ◽  
Chondrocyte Differentiation ◽  
P38 Kinase ◽  
Matrix Mineralization ◽  
Myocyte Enhancer Factor ◽  
Mitogen Activated Protein ◽  
Enhancer Factor

Longitudinal growth of endochondral bones is accomplished through the co-ordinated proliferation and hypertrophic differentiation of growth plate chondrocytes. The molecular mechanisms and signalling cascades controlling these processes are not well understood. To analyse the expression and roles of p38 mitogen-activated protein kinases in this process, we have established a micromass system for the reproducible hypertrophic differentiation of mouse mesenchymal limb bud cells. Our results show that all four mammalian p38 kinase genes are expressed during the chondrogenic programme, as well as their upstream regulators MKK3 (mitogen-activated protein kinase kinase 3) and MKK6. Treatment of micromass cultures with pharmacological inhibitors of p38 results in a marked delay in hypertrophic differentiation in micromass cultures, indicating a requirement for p38 signalling in chondrocyte differentiation. Inhibition of p38 kinase activity leads to reduced and delayed induction of alkaline phosphatase activity and matrix mineralization. In addition, p38 inhibition causes reduced expression of hypertrophic marker genes such as collagen X, matrix metalloproteinase 13 and bone sialoprotein. The function of p38 in hypertrophic differentiation appears to be mediated, at least in part, by the transcription factor myocyte enhancer factor 2C. In summary, we have demonstrated a novel requirement for p38 signalling in hypertrophic differentiation of chondrocytes and identified myocyte enhancer factor 2C as an important regulator of chondrocyte gene expression.

scholarly journals Pannexin 1-Mediated ATP Signaling in the Trigeminal Spinal Subnucleus Caudalis Is Involved in Tongue Cancer Pain

2021 ◽  
Vol 22 (21) ◽  
pp. 11404
Author(s):  
Ryo Koyama ◽  
Koichi Iwata ◽  
Yoshinori Hayashi ◽  
Suzuro Hitomi ◽  
Ikuko Shibuta ◽  
...  
Keyword(s):  
Cancer Pain ◽  
Mechanical Stimulation ◽  
Calcium Binding ◽  
Molecular Mechanisms ◽  
Tongue Cancer ◽  
Interleukin 1 ◽  
Inhibitory Peptide ◽  
Nociceptive Neurons ◽  
Pannexin 1 ◽  
Withdrawal Threshold

Pain is one of the most severe concerns in tongue cancer patients. However, the underlying mechanisms of tongue cancer pain are not fully understood. We investigated the molecular mechanisms of tongue cancer-induced mechanical allodynia in the tongue by squamous cell carcinoma (SCC) inoculation in rats. The head-withdrawal threshold of mechanical stimulation (MHWT) to the tongue was reduced following SCC inoculation, which was inhibited by intracisternal administration of 10Panx, an inhibitory peptide for pannexin 1 (PANX1) channels. Immunohistochemical analyses revealed that the expression of PANX1 was upregulated in the trigeminal spinal subnucleus caudalis (Vc) following SCC inoculation. The majority of PANX1 immunofluorescence was merged with ionized calcium-binding adapter molecule 1 (Iba1) fluorescence and a part of it was merged with glial fibrillary acidic protein (GFAP) fluorescence. Spike frequencies of Vc nociceptive neurons to noxious mechanical stimulation were significantly enhanced in SCC-inoculated rats, which was suppressed by intracisternal 10Panx administration. Phosphorylated extracellular signal-regulated kinase (pERK)-immunoreactive (IR) neurons increased significantly in the Vc after SCC inoculation, which was inhibited by intracisternal 10Panx administration. SCC inoculation-induced MHWT reduction and increased pERK-IR Vc neuron numbers were inhibited by P2X7 purinoceptor (P2X7R) antagonism. Conversely, these effects were observed in the presence of P2X7R agonist in SCC-inoculated rats with PANX1 inhibition. SCC inoculation-induced MHWT reduction was significantly recovered by intracisternal interleukin-1 receptor antagonist administration. These observations suggest that SCC inoculation causes PANX1 upregulation in Vc microglia and adenosine triphosphate released through PANX1 sensitizes nociceptive neurons in the Vc, resulting in tongue cancer pain.

Transcription factor/DNA interactions visualized by electron spectroscopic imaging

1992 ◽  
Vol 50 (1) ◽  
pp. 450-451
Author(s):  
David P. Bazett-Jones ◽  
Mark L. Brown
Keyword(s):  
Transcription Factor ◽  
Dna Sequences ◽  
Transcription Initiation ◽  
Molecular Mechanisms ◽  
Phosphorus Content ◽  
Spectroscopic Imaging ◽  
Electron Spectroscopic Imaging ◽  
Dna Backbone ◽  
Two Parameters ◽  
High Level

A multisubunit RNA polymerase enzyme is ultimately responsible for transcription initiation and elongation of RNA, but recognition of the proper start site by the enzyme is regulated by general, temporal and gene-specific trans-factors interacting at promoter and enhancer DNA sequences. To understand the molecular mechanisms which precisely regulate the transcription initiation event, it is crucial to elucidate the structure of the transcription factor/DNA complexes involved. Electron spectroscopic imaging (ESI) provides the opportunity to visualize individual DNA molecules. Enhancement of DNA contrast with ESI is accomplished by imaging with electrons that have interacted with inner shell electrons of phosphorus in the DNA backbone. Phosphorus detection at this intermediately high level of resolution (≈lnm) permits selective imaging of the DNA, to determine whether the protein factors compact, bend or wrap the DNA. Simultaneously, mass analysis and phosphorus content can be measured quantitatively, using adjacent DNA or tobacco mosaic virus (TMV) as mass and phosphorus standards. These two parameters provide stoichiometric information relating the ratios of protein:DNA content.

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