scholarly journals Regulation of type-II collagen gene expression during human chondrocyte de-differentiation and recovery of chondrocyte-specific phenotype in culture involves Sry-type high-mobility-group box (SOX) transcription factors

2001 ◽  
Vol 360 (2) ◽  
pp. 461-470 ◽  
Author(s):  
David G. STOKES ◽  
Gang LIU ◽  
Rita DHARMAVARAM ◽  
David HAWKINS ◽  
Sonsoles PIERA-VELAZQUEZ ◽  
...  
Keyword(s):  
Transcription Factors ◽  
High Mobility ◽  
Type Ii Collagen ◽  
Culture Conditions ◽  
High Mobility Group ◽  
Collagen Gene ◽  
Type Ii ◽  
Reporter Construct ◽  
Monolayer Cultures

During ex vivo growth as monolayer cultures, chondrocytes proliferate and undergo a process of de-differentiation. This process involves a change in morphology and a change from expression of chondrocyte-specific genes to that of genes that are normally expressed in fibroblasts. Transfer of the monolayer chondrocyte culture to three-dimensional culture systems induces the cells to re-acquire a chondrocyte-specific phenotype and produce a cartilaginous-like tissue in vitro. We investigated mechanisms involved in the control of the de-differentiation and re-differentiation process in vitro. De-differentiated chondrocytes re-acquired their chondrocyte-specific phenotype when cultured on poly-(2-hydroxyethyl methacrylate) (polyHEMA) as assayed by morphology, reverse transcriptase PCR of chondrocyte-specific mRNA, Western-blot analysis and chondrocyte-specific promoter activity. Essentially, full recovery of the chondrocyte-specific phenotype was observed when cells that had been cultured for 4 weeks on plastic were transferred to culture on polyHEMA. However, after subsequent passages on plastic, the phenotype recovery was incomplete or did not occur. The activity of a gene reporter construct containing the promoter and enhancer from the human type-II collagen gene (COL2A1) was modulated by the culture conditions, so that its transcriptional activity was repressed in monolayer cultures and rescued to some extent when the cells were switched to polyHEMA cultures. The binding of Sry-type high-mobility-group box (SOX) transcription factors to the enhancer region was modulated by the culture conditions, as were the mRNA levels for SOX9. A transfected human type-II collagen reporter construct was activated in de-differentiated cells by ectopic expression of SOX transcription factors. These results underscore the overt change in phenotype that occurs when chondrocytes are cultured as monolayers on tissue-culture plastic substrata.

Induction of High Mobility Group I Architectural Transcription Factors in Proliferating Vascular Smooth Muscle in vivo and in vitro

1999 ◽  
Vol 31 (12) ◽  
pp. 2199-2205 ◽  
Author(s):  
Michael T Chin ◽  
Andrea Pellacani ◽  
Chung- Ming Hsieh ◽  
Sharon SJ Lin ◽  
Mukesh K Jain ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Transcription Factors ◽  
Vascular Smooth Muscle ◽  
High Mobility ◽  
High Mobility Group ◽  
Group I

scholarly journals Sp1 and Sp3 Transcription Factors Mediate Interleukin-1β Down-regulation of Human Type II Collagen Gene Expression in Articular Chondrocytes

2003 ◽  
Vol 278 (41) ◽  
pp. 39762-39772 ◽  
Author(s):  
Christos Chadjichristos ◽  
Chafik Ghayor ◽  
Magdalini Kypriotou ◽  
Grégoire Martin ◽  
Emmanuelle Renard ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Interleukin 1Β ◽  
Collagen Gene ◽  
Type Ii ◽  
Down Regulation ◽  
Human Type ◽  
Collagen Gene Expression

scholarly journals High-Mobility-Group Proteins NHP6A and NHP6B Participate in Activation of the RNA Polymerase IIISNR6 Gene

2001 ◽  
Vol 21 (9) ◽  
pp. 3096-3104 ◽  
Author(s):  
Sébastien Lopez ◽  
Magda Livingstone-Zatchej ◽  
Sabine Jourdain ◽  
Fritz Thoma ◽  
André Sentenac ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase ◽  
Rna Polymerase Iii ◽  
High Mobility ◽  
High Mobility Group ◽  
Wild Type ◽  
High Mobility Group Proteins ◽  
General Transcription Factors ◽  
Pol Iii

ABSTRACT Transcription of yeast class III genes involves the formation of a transcription initiation complex that comprises RNA polymerase III (Pol III) and the general transcription factors TFIIIB and TFIIIC. Using a genetic screen for positive regulators able to compensate for a deficiency in a promoter element of the SNR6 gene, we isolated the NHP6A and NHP6B genes. Here we show that the high-mobility-group proteins NHP6A and NHP6B are required for the efficient transcription of the SNR6 gene both in vivo and in vitro. The transcripts of wild-type and promoter-defectiveSNR6 genes decreased or became undetectable in annhp6AΔ nhp6BΔ double-mutant strain, and the protection over the TATA box of the wild-type SNR6 gene was lost innhp6AΔ nhp6BΔ cells at 37°C. In vitro, NHP6B specifically stimulated the transcription of SNR6 templates up to fivefold in transcription assays using either cell nuclear extracts from nhp6AΔ nhp6BΔ cells or reconstituted transcription systems. Finally, NHP6B activated SNR6transcription in a TFIIIC-independent assay. These results indicate that besides the general transcription factors TFIIIB and TFIIIC, additional auxilliary factors are required for the optimal transcription of at least some specific Pol III genes.

scholarly journals Collagen gene expression during limb cartilage differentiation.

1986 ◽  
Vol 102 (4) ◽  
pp. 1151-1156 ◽  
Author(s):  
R A Kosher ◽  
W M Kulyk ◽  
S W Gay
Keyword(s):  
Gene Expression ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Mesenchymal Cells ◽  
Type I ◽  
Collagen Gene ◽  
Type Ii ◽  
Collagen Mrna ◽  
Cytoplasmic Type

As limb mesenchymal cells differentiate into chondrocytes, they initiate the synthesis of type II collagen and cease synthesizing type I collagen. Changes in the cytoplasmic levels of type I and type II collagen mRNAs during the course of limb chondrogenesis in vivo and in vitro were examined using cloned cDNA probes. A striking increase in cytoplasmic type II collagen mRNA occurs coincident with the crucial condensation stage of chondrogenesis in vitro, in which prechondrogenic mesenchymal cells become closely juxtaposed before depositing a cartilage matrix. Thereafter, a continuous and progressive increase in the accumulation of cytoplasmic type II collagen mRNA occurs which parallels the progressive accumulation of cartilage matrix by cells. The onset of overt chondrogenesis, however, does not involve activation of the transcription of the type II collagen gene. Low levels of type II collagen mRNA are present in the cytoplasm of prechondrogenic mesenchymal cells at the earliest stages of limb development, well before the accumulation of detectable levels of type II collagen. Type I collagen gene expression during chondrogenesis is regulated, at least in part, at the translational level. Type I collagen mRNAs are present in the cytoplasm of differentiated chondrocytes, which have ceased synthesizing detectable amounts of type I collagen.

Human Umbilical Cord Mesenchymal Stem Cells Induce into Chondrocytes in 3D Cultured System

2013 ◽  
Vol 749 ◽  
pp. 198-205
Author(s):  
Li Yu ◽  
Jing Liu ◽  
Chao Xu ◽  
Er Mei Luo ◽  
Ming Qiao Tang
Keyword(s):  
Real Time ◽  
Alcian Blue ◽  
Culture System ◽  
Type Ii Collagen ◽  
Human Umbilical Cord ◽  
Type Ii ◽  
Pellet Culture ◽  
Hla Dr

Objective: To investigate a better method of inducing hUC-MSCs into chondrocytes in different culture system in vitro. Method: hUC-MSCs were isolated and cultured by tissue block culture, and the cells surface antigens were identified by flow cytometry, hUC-MSCs were cultured with chondrogenic media and stained with Alcian Blue. The production of matrix was estimated from the determination of hydroxyproline content and Alcian Blue method. Expressions of glycosaminoglycan (GAG), type II collagen and Sox-9 were assayed by real-time fluorescence quantitative PCR. Results: The cultured hUC-MSCs phenotype was CD105+/CD29+/CD44+/ CD31-/CD34-/ CD40-/CD45-/HLA-DR-. hUC-MSCs weakly expressed chondrocyte marker, which strongly expressed GAG and type II collagen after chondrogenic induction, and the cells were incubated in pellet culture with higher expression. Real-time PCR results demonstrated that chondrogenic induction cells were expressed GAG, type II collagen and Sox-9, and the cells were incubated in pellet culture with higher expression. Conclusion: hUC-MSCs incubated in pellet culture is more conducive to differentiate into chondrocytes than those cultured in monolayer culture system.

scholarly journals LPS Primes Brain Responsiveness to High Mobility Group Box-1 Protein

2021 ◽  
Vol 14 (6) ◽  
pp. 558
Author(s):  
Verena Peek ◽  
Lois M. Harden ◽  
Jelena Damm ◽  
Ferial Aslani ◽  
Stephan Leisengang ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Area Postrema ◽  
High Mobility ◽  
High Mobility Group ◽  
Direct Access ◽  
Significant Release ◽  
Factor Α ◽  
Culture Supernatants

High mobility group box (HMGB)1 action contributes to late phases of sepsis, but the effects of increased endogenous plasma HMGB1 levels on brain cells during inflammation are unclear. Here, we aimed to further investigate the role of HMGB1 in the brain during septic-like lipopolysaccharide-induced inflammation in rats (LPS, 10 mg/kg, i.p.). HMGB-1 mRNA expression and release were measured in the periphery/brain by RT-PCR, immunohistochemistry and ELISA. In vitro experiments with disulfide-HMGB1 in primary neuro-glial cell cultures of the area postrema (AP), a circumventricular organ with a leaky blood–brain barrier and direct access to circulating mediators like HMGB1 and LPS, were performed to determine the direct influence of HMGB1 on this pivotal brain structure for immune-to-brain communication. Indeed, HMGB1 plasma levels stayed elevated after LPS injection. Immunohistochemistry of brains and AP cultures confirmed LPS-stimulated cytoplasmatic translocation of HMGB1 indicative of local HMGB1 release. Moreover, disulfide-HMGB1 stimulation induced nuclear factor (NF)-κB activation and a significant release of interleukin-6, but not tumor necrosis factor α, into AP culture supernatants. However, only a few AP cells directly responded to HMGB1 with increased intracellular calcium concentration. Interestingly, priming with LPS induced a seven-fold higher percentage of responsive cells to HMGB1. We conclude that, as a humoral and local mediator, HMGB1 enhances brain inflammatory responses, after LPS priming, linked to sustained sepsis symptoms.

Down-regulation of chondrocyte aggrecan and type-II Collagen gene expression correlates with increases in static compression magnitude and duration

1999 ◽  
Vol 17 (6) ◽  
pp. 836-842 ◽  
Author(s):  
Paula M. Ragan ◽  
Alison M. Badger ◽  
Michael Cook ◽  
Vicki I. Chin ◽  
Maxine Gowen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Type Ii Collagen ◽  
Collagen Gene ◽  
Type Ii ◽  
Down Regulation ◽  
Static Compression ◽  
Collagen Gene Expression

scholarly journals Defective Calmodulin-Mediated Nuclear Transport of the Sex-Determining Region of the Y Chromosome (SRY) in XY Sex Reversal

2005 ◽  
Vol 19 (7) ◽  
pp. 1884-1892 ◽  
Author(s):  
Helena Sim ◽  
Kieran Rimmer ◽  
Sabine Kelly ◽  
Louisa M. Ludbrook ◽  
Andrew H. A. Clayton ◽  
...  
Keyword(s):  
Y Chromosome ◽  
Nuclear Import ◽  
High Mobility ◽  
Sex Reversal ◽  
High Mobility Group ◽  
Missense Mutations ◽  
Nuclear Localization Signals ◽  
Xy Sex Reversal

Abstract The sex-determining region of the Y chromosome (SRY) plays a key role in human sex determination, as mutations in SRY can cause XY sex reversal. Although some SRY missense mutations affect DNA binding and bending activities, it is unclear how others contribute to disease. The high mobility group domain of SRY has two nuclear localization signals (NLS). Sex-reversing mutations in the NLSs affect nuclear import in some patients, associated with defective importin-β binding to the C-terminal NLS (c-NLS), whereas in others, importin-β recognition is normal, suggesting the existence of an importin-β-independent nuclear import pathway. The SRY N-terminal NLS (n-NLS) binds calmodulin (CaM) in vitro, and here we show that this protein interaction is reduced in vivo by calmidazolium, a CaM antagonist. In calmidazolium-treated cells, the dramatic reduction in nuclear entry of SRY and an SRY-c-NLS mutant was not observed for two SRY-n-NLS mutants. Fluorescence spectroscopy studies reveal an unusual conformation of SRY.CaM complexes formed by the two n-NLS mutants. Thus, CaM may be involved directly in SRY nuclear import during gonadal development, and disruption of SRY.CaM recognition could underlie XY sex reversal. Given that the CaM-binding region of SRY is well-conserved among high mobility group box proteins, CaM-dependent nuclear import may underlie additional disease states.

scholarly journals Does high-mobility-group non-histone protein HMG 1 interact specifically with histone H1 subfractions?

1979 ◽  
Vol 183 (3) ◽  
pp. 657-662 ◽  
Author(s):  
P D Cary ◽  
K V Shooter ◽  
G H Goodwin ◽  
E W Johns ◽  
J Y Olayemi ◽  
...  
Keyword(s):  
Specific Interaction ◽  
High Mobility ◽  
Histone H1 ◽  
High Mobility Group ◽  
Chromosomal Protein ◽  
Histone Protein ◽  
Total Histone ◽  
Equilibrium Sedimentation ◽  
Histone H5

The interaction of the non-histone chromosomal protein HMG (high-mobility group) 1 with histone H1 subfractions was investigated by equilibrium sedimentation and n.m.r. sectroscopy. In contrast with a previous report [Smerdon & Isenberg (1976) Biochemistry 15, 4242–4247], it was found, by using equilibrium-sedimentation analysis, that protein HMG 1 binds to all three histone H1 subfractions CTL1, CTL2, and CTL3, arguing against there being a specific interaction between protein HMG 1 and only two of the subfractions, CTL1 and CTL2. Raising the ionic strength of the solutions prevents binding of protein HMG 1 to total histone H1 and the three subfractions, suggesting that the binding in vitro is simply a non-specific ionic interaction between acidic regions of the non-histone protein and the basic regions of the histone. Protein HMG 1 binds to histone H5 also, supporting this view. The above conclusions are supported by n.m.r. studies of protein HMG 1/histone H1 subfraction mixtures. When the two proteins were mixed, there was little perturbation of the n.m.r. spectra and there was no evidence for specific interaction of protein HMG 1 with any of the subfractions. It therefore remains an open question as to whether protein HMG 1 and histone H1 are complexed together in chromatin.

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