scholarly journals Dysfunction of Collagen Synthesis and Secretion in Chondrocytes Induced byWisp3Mutation

2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
Min Wang ◽  
Xiao-Fei Man ◽  
Ya-Qing Liu ◽  
Er-Yuan Liao ◽  
Zhi-Feng Shen ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Collagen Synthesis ◽  
Underlying Mechanism ◽  
Expression Vectors ◽  
Protein Distribution ◽  
Collagen Secretion ◽  
Proline Incorporation ◽  
Spondyloepiphyseal Dysplasia Tarda ◽  
Peak Phase ◽  
Intracellular Collagen

Wisp3gene mutation was shown to cause spondyloepiphyseal dysplasia tarda with progressive arthropathy (SRDT-PA), but the underlying mechanism is not clear. To clarify this mechanism, we constructed the wild and mutatedWisp3expression vectors and transfected into human chondrocytes lines C-20/A4;Wisp3proteins subcellular localization, cell proliferation, cell apoptosis, andWisp3-mediated gene expression were determined, and dynamic secretion of collagen in transfected chondrocytes was analyzed by14C-proline incorporation experiment. MutatedWisp3protein increased proliferation activity, decreased apoptosis of C-20/A4 cells, and aggregated abnormally in cytoplasm. Expression of collagen II was also downregulated in C-20/A4 cells transfected with mutatedWisp3. Wild typeWisp3transfection increased intracellular collagen content and extracellular collagen secretion, but the mutatedWisp3lost this function, and the peak phase of collagen secretion was delayed in mutatedWisp3transfected cells. Thus abnormal protein distribution, cell proliferation, collagen synthesis, and secretion inWisp3mutated chondrocytes might contribute to the pathogenesis of SEDT-PA.

scholarly journals Glomerular Mesangial Cell pH Homeostasis Mediates Mineralocorticoid Receptor-Induced Cell Proliferation

Biomedicines ◽  
2021 ◽  
Vol 9 (9) ◽  
pp. 1117
Author(s):  
Michael Gekle ◽  
Sigrid Mildenberger
Keyword(s):  
Cell Proliferation ◽  
Mineralocorticoid Receptor ◽  
Mesangial Cells ◽  
Cytosolic Calcium ◽  
Cell Counting ◽  
Glomerular Mesangial Cells ◽  
Ph Homeostasis ◽  
Glomerular Mesangial Cell ◽  
Collagen Secretion ◽  
Proline Incorporation

Mineralocorticoids (e.g., aldosterone) support chronic inflammatory tissue damage, including glomerular mesangial injury leading to glomerulosclerosis. Furthermore, aldosterone leads to activation of the extracellular signal-regulated kinases (ERK1/2) in rat glomerular mesangial cells (GMC). Because ERK1/2 can affect cellular pH homeostasis via activation of Na+/H+-exchange (NHE) and the resulting cellular alkalinization may support proliferation, we tested the hypothesis that aldosterone affects pH homeostasis and thereby cell proliferation as well as collagen secretion also in primary rat GMC. Cytoplasmic pH and calcium were assessed by single-cell fluorescence ratio imaging, using the dyes BCECF or FURA2, respectively. Proliferation was determined by cell counting, thymidine incorporation and collagen secretion by collagenase-sensitive proline incorporation and ERK1/2-phosphorylation by Western blot. Nanomolar aldosterone induces a rapid cytosolic alkalinization which is prevented by NHE inhibition (10 µmol/L EIPA) and by blockade of the mineralocorticoid receptor (100 nmol/L spironolactone). pH changes were not affected by inhibition of HCO3− transporters and were not dependent on HCO3−. Aldosterone enhanced ERK1/2 phosphorylation and inhibition of ERK1/2-phosphorylation (10 µmol/L U0126) prevented aldosterone-induced alkalinization. Furthermore, aldosterone induced proliferation of GMC and collagen secretion, both of which were prevented by U0126 and EIPA. Cytosolic calcium was not involved in this aldosterone action. In conclusion, our data show that aldosterone can induce GMC proliferation via a MR and ERK1/2-mediated activation of NHE with subsequent cytosolic alkalinization. GMC proliferation leads to glomerular hypercellularity and dysfunction. This effect presents a possible mechanism contributing to mineralocorticoid receptor-induced pathogenesis of glomerular mesangial injury during chronic kidney disease.

cAMP-elevating agents and adenylyl cyclase overexpression promote an antifibrotic phenotype in pulmonary fibroblasts

2004 ◽  
Vol 286 (5) ◽  
pp. C1089-C1099 ◽  
Author(s):  
Xiaoqiu Liu ◽  
Rennolds S. Ostrom ◽  
Paul A. Insel
Keyword(s):  
Cell Proliferation ◽  
Pulmonary Fibrosis ◽  
Adenylyl Cyclase ◽  
Collagen Synthesis ◽  
Lung Fibroblasts ◽  
Prostaglandin E ◽  
Pulmonary Fibroblasts ◽  
Adenosine Receptor Agonist ◽  
Proline Incorporation ◽  
Camp Levels

Pulmonary fibroblasts are recruited to sites of lung injury, where they are activated to produce extracellular matrix proteins and to facilitate repair. However, these cells become dysregulated in pulmonary fibrosis, producing excess collagen at sites of injury and forming fibrotic loci that impair lung function. In this study, we used WI-38 human lung fibroblasts and evaluated the ability of G protein-coupled receptor agonists to increase cAMP production and regulate cell proliferation and collagen synthesis. WI-38 cells increase cAMP in response to the β-adrenergic agonist isoproterenol (Iso), prostaglandin E2 (PGE2), certain prostanoid receptor-selective agonists (beraprost, butaprost), an adenosine receptor agonist, and the direct adenylyl cyclase (AC) activator forskolin (Fsk). Responses to Iso, PGE2, and Fsk were studied in more detail. Each induced a dose-dependent inhibition of serum-stimulated cell proliferation (as measured by [3H]thymidine incorporation) and collagen synthesis (as measured by [3H]proline incorporation, collagenase-sensitive [3H]proline incorporation, or levels of procollagen type 1 C-peptide). Quantitative RT-PCR analyses indicated that elevation in cellular cAMP levels decreases expression of collagen types 1α(II) and 5α(I) and increases expression and activity of matrix metalloproteinase 2 (MMP-2). Overexpression of AC type 6 or inhibition of cyclic nucleotide phosphodiesterases also increased cellular cAMP levels and decreased cell proliferation and collagen synthesis. Thus multiple approaches that increase cAMP signaling reduce proliferation and differentiated function in human pulmonary fibroblasts. These results suggest that therapies that raise cAMP levels may prove useful in the treatment of pulmonary fibrosis.

Characterization of YY1 OPB Peptide for its Anticancer Activity

2019 ◽  
Vol 19 (6) ◽  
pp. 504-511 ◽  
Author(s):  
Yige Qi ◽  
Ting Yan ◽  
Lu Chen ◽  
Qiang Zhang ◽  
Weishu Wang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Amino Acids ◽  
Cell Proliferation ◽  
Cancer Cell ◽  
Breast Cancer Cell ◽  
Expression Vectors ◽  
Cancer Cell Proliferation ◽  
Breast Cancer Cell Proliferation ◽  
Oncogene Products ◽  
And Migration

Background:The oncoprotein binding (OPB) domain of Yin Yang 1 (YY1) consists of 26 amino acids between G201 and S226, and is involved in YY1 interaction with multiple oncogene products, including MDM2, AKT, EZH2 and E1A. Through the OPB domain, YY1 promotes the oncogenic or proliferative regulation of these oncoproteins in cancer cells. We previously demonstrated that a peptide with the OPB sequence blocked YY1-AKT interaction and inhibited breast cancer cell proliferation.Objective:In the current study, we characterized the OPB domain and determined a minimal region for peptide design to suppress cancer cellMethods:Using alanine-scan method, we identified that the amino acids at OPB C-terminal are essential to YY1 binding to AKT. Further studies suggested that serine and threonine residues, but not lysines, in OPB play a key role in YY1-AKT interaction. We generated GFP fusion expression vectors to express OPB peptides with serially deleted N-terminal and found that OPB1 (i.e. G201-S226) is cytoplasmic, but OPB2 (i.e. E206-S226), OPB3 (i.e. E206-S226) and control peptide were both nuclear and cytoplasmic.Results:Both OPB1 and 2 inhibited breast cancer cell proliferation and migration, but OPB3 exhibited similar effects to control. OPB1 and 2 caused cell cycle arrest at G1 phase, increased p53 and p21 expression, and reduced AKT(S473) phosphorylation in MCF-7 cells, but not in MDA-MB-231 cells.Conclusion:: Overall, the serines and threonines of OPB are essential to YY1 binding to oncoproteins, and OPB peptide can be minimized to E206-S226 that maintain inhibitory activity to YY1- promoted cell proliferation.

scholarly journals Knockdown of long noncoding RNA HOTAIR inhibits osteoarthritis chondrocyte injury by miR-107/CXCL12 axis

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Jipeng Lu ◽  
Zhongxiong Wu ◽  
Ying Xiong
Keyword(s):  
Cell Proliferation ◽  
Western Blot ◽  
Cell Apoptosis ◽  
Noncoding Rna ◽  
Underlying Mechanism ◽  
Joint Disease ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Cxcl12 Expression ◽  
Ecm Degradation

Abstract Background Osteoarthritis (OA) is a joint disease characterized via destruction of cartilage. Chondrocyte damage is associated with cartilage destruction during OA. Long noncoding RNAs (lncRNAs) are implicated in the regulation of chondrocyte damage in OA progression. This study aims to investigate the role and underlying mechanism of lncRNA homeobox antisense intergenic RNA (HOTAIR) in OA chondrocyte injury. Methods Twenty-three OA patients and healthy controls without OA were recruited. Chondrocytes were isolated from OA cartilage tissues. HOTAIR, microRNA-107 (miR-107) and C-X-C motif chemokine ligand 12 (CXCL12) levels were measured by quantitative real-time polymerase chain reaction and western blot. Cell proliferation, apoptosis and extracellular matrix (ECM) degradation were measured using cell counting kit-8, flow cytometry and western blot. The target interaction was explored by bioinformatics, luciferase reporter and RNA immunoprecipitation assays. Results HOTAIR expression was enhanced, and miR-107 level was reduced in OA cartilage samples. HOTAIR overexpression inhibited cell proliferation, but induced cell apoptosis and ECM degradation in chondrocytes. HOTAIR knockdown caused an opposite effect. MiR-107 was sponged and inhibited via HOTAIR, and knockdown of miR-107 mitigated the effect of HOTAIR silence on chondrocyte injury. CXCL12 was targeted by miR-107. CXCL12 overexpression attenuated the roles of miR-107 overexpression or HOTAIR knockdown in the proliferation, apoptosis and ECM degradation. CXCL12 expression was decreased by HOTAIR silence, and restored by knockdown of miR-107. Conclusion HOTAIR knockdown promoted chondrocyte proliferation, but inhibited cell apoptosis and ECM degradation in OA chondrocytes by regulating the miR-107/CXCL12 axis.

scholarly journals ANXA4 promotes trophoblast invasion via the PI3K/Akt/eNOS pathway in preeclampsia

2019 ◽  
Vol 316 (4) ◽  
pp. C481-C491 ◽  
Author(s):  
Yalan Xu ◽  
Lili Sui ◽  
Bintao Qiu ◽  
Xiuju Yin ◽  
Juntao Liu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Western Blotting ◽  
Underlying Mechanism ◽  
Trophoblast Invasion ◽  
Pathological Examination ◽  
Trophoblast Cells ◽  
Human Trophoblast ◽  
Annexin A4 ◽  
Extravillous Cytotrophoblasts ◽  
Phosphoinositide 3 Kinase

The inadequate trophoblast invasion is associated with the development of preeclampsia (PE). Considering that annexin A4 (ANXA4) enhances tumor invasion, we aimed to explore the functional role of ANXA4 in trophoblast cells and to examine the underlying mechanism. ANXA4 expression in PE placentas was analyzed using immunohistochemistry and Western blotting. Cell proliferation, invasion, and apoptosis were determined using a MTT assay, Transwell assay, and flow cytometry, respectively. The expression levels of matrix metalloproteinase (MMP)-2, MMP-9, phosphoinositide 3-kinase (PI3K), Akt, phosphorylated (p)-Akt, and phosphorylated endothelial nitric oxide synthase (p-eNOS) were detected by Western blotting. Placentas were prepared for pathological examination using hematoxylin and eosin staining and apoptosis determination using the TUNEL method. Expression of ANXA4, PI3K, p-Akt and p-eNOS was downregulated in human PE placentas and PE placenta-derived extravillous cytotrophoblasts (EVCTs). Furthermore, ANXA4 overexpression promoted cell proliferation and invasion, inhibited cell apoptosis, and upregulated protein expression of PI3K, p-Akt, and p-eNOS in human trophoblast cells HTR-8/SVneo and JEG-3. By contrast, ANXA4 knockdown exerted the opposite effects. Furthermore, inhibition of the PI3K/Akt pathway by LY294002 abrogated the ANXA4 overexpression-mediated effects on trophoblast behavior. Furthermore, eNOS knockdown abrogated the ANXA4 overexpression-induced promotion of cell invasion and MMP2/9 expression. Additionally, in N-nitro-l-arginine methyl ester (l-NAME)-induced PE rats, ANXA4 overexpression alleviated PE progression, accompanied by an increase in expression of PI3K, p-Akt, and p-eNOS in rat placentas. Our findings demonstrate that ANXA4 expression is downregulated in PE. ANXA4 may promote trophoblast invasion via the PI3K/Akt/eNOS pathway.

scholarly journals [3H]proline incorporation and hydroxyproline concentration in articular cartilage during the development of osteoarthritis caused by immobilization. A study in vivo with rabbits

1981 ◽  
Vol 200 (2) ◽  
pp. 435-440 ◽  
Author(s):  
T Videman ◽  
I Eronen ◽  
T Candolin
Keyword(s):  
Total Protein ◽  
Collagen Synthesis ◽  
Weight Bearing ◽  
Specific Radioactivity ◽  
Collagen Content ◽  
Synthetic Activity ◽  
Synthesis Rate ◽  
Hydroxyproline Concentration ◽  
Proline Incorporation

Proline metabolism in vivo was studied during the development of immobilization osteoarthritis in rabbits. Collagen content was measured as the hydroxyproline concentration of the tissue in question. The incorporation of [3H]proline was used as the indicator for total protein synthesis; collagen synthesis rate was estimated from measurements of the specific radioactivity of hydroxyproline. Cartilage samples from knee and hip joints were analysed after 3, 7, 11, 18, 35 and 56 days of immobilization. The total protein and collagen synthesis rates of the immobilized legs increased and reached a maximum after 11-35 days. Although they decreased thereafter, these rates remained elevated to the end of the experiment. A slight increase in the synthetic activity of the non-immobilized contralateral legs was also detected after 7--18 days of immobilization. The isotope incorporation was markedly higher in tibial marginal tissue than in weight-bearing cartilage. In spite of the increased synthesis, no clear changes were found in the collagen content of the tissues studied during the experiment.

scholarly journals Asiaticoside induces cell proliferation and collagen synthesis in human dermal fibroblasts

2015 ◽  
Vol 34 (2) ◽  
pp. 96
Author(s):  
Linda Yuliati ◽  
Etik Mardliyati ◽  
Kusmarinah Bramono ◽  
Hans Joachim Freisleben
Keyword(s):  
Cell Proliferation ◽  
Wound Healing ◽  
Retinoic Acid ◽  
Collagen Synthesis ◽  
Active Agent ◽  
Dermal Fibroblasts ◽  
Type Iii Collagen ◽  
Type I ◽  
Human Dermal Fibroblasts ◽  
Type Iii

Background<br />Asiatiocoside, a saponin component isolated from Centella asiatica can improve wound healing by promoting the proliferation of human dermal fibroblasts (HDF) and synthesis of collagen. The skin-renewing cells and type I and III collagen synthesis decrease with aging, resulting in the reduction of skin elasticity and delayed wound healing. Usage of natural active compounds from plants in wound healing should be evaluated and compared to retinoic acid as an active agent that regulates wound healing. The aim of this study was to compare and evaluate the effect of asiaticoside and retinoic acid to induce greater cell proliferation and type I and III collagen synthesis in human dermal fibroblast.<br /><br />Methods<br />Laboratory experiments were conducted using human dermal fibroblasts (HDF) isolated from human foreskin explants. Seven passages of HDF were treated with asiaticoside and retinoic acid at several doses and incubated for 24 and 48 hours. Cell viability in all groups was tested with the MTT assay to assess HDF proliferation. Type I and III collagen synthesis was examined using the respective ELISA kits. Analysis of variance was performed to compare the treatment groups. <br /><br />Results<br />Asiaticoside had significantly stronger effects on HDF proliferation than retinoic acid (p&lt;0.05). The type III collagen production was significantly greater induction with asiaticoside compared to retinoic acid (p&lt;0.05). <br /><br />Conclusion<br />Asiaticoside induces HDF proliferation and type I and III collagen synthesis in a time- and dose-dependent pattern. Asiaticoside has a similar effect as retinoic acid on type I and type III collagen synthesis.

A Rapid Degradation of Calponin 2 Is Required for Cytokinesis

2021 ◽  
Author(s):  
Airong Qian ◽  
Tzu-Bou Hsieh ◽  
M. Moazzem Hossain ◽  
Jim J.-C. Lin ◽  
J.-P. Jin
Keyword(s):  
Cell Proliferation ◽  
Cell Division ◽  
Underlying Mechanism ◽  
Fluorescent Imaging ◽  
Hek293 Cells ◽  
Computer Assisted ◽  
Over Expression ◽  
Dividing Cells ◽  
Ubiquitin Proteasome ◽  
Dynamic Image Analysis

Calponin 2 is an actin cytoskeleton-associated protein and plays a role in regulating cell motility-related functions such as phagocytosis, migration and division. We previously reported that the expression of calponin 2 inhibits the rate of cell proliferation. To investigate the underlying mechanism, our present study found that the levels of endogenous calponin 2 in NIH3T3 and HEK293 cells rapidly decreased prior to cell division characterized by an absence at the actin contractile ring. In cells lacking endogenous calponin 2, transfective expression of GFP-fusion calponin 2 inhibited cell proliferation similar to that of non-fusion calponin 2. Fluorescent imaging studies of mitotic cells indicated that a proper level of calponin 2 expression and effective degradation during cytokinesis are necessary for normal cell division. Computer-assisted dynamic image analysis of dividing cells revealed that over-expression of calponin 2 significantly affects motile and shape behaviors of cells only on the interval from the start of anaphase to the start of cytokinesis, i.e., the pre-cytokinesis phase, but not on the interval from the start of cytokinesis to 50% completion of cytokinesis. The pre-cytokinesis degradation of calponin 2 was attenuated by MG132 inhibition of the ubiquitin proteasome and inhibitor of protein kinase C (PKC), suggesting that PKC phosphorylation-triggered degradation of calponin 2 could determine the rate of cytokinesis. The novel role of calponin 2 in regulating the rate of cytokinesis may be targeted for therapeutic applications such as in an inhibition of malignant tumor growth.

LncRNA PCAT18/miR-301a/TP53INP1 axis is involved in gastric cancer cell viability, migration and invasion

2020 ◽  
Vol 168 (5) ◽  
pp. 547-555
Author(s):  
Jin Dou ◽  
Daoyuan Tu ◽  
Haijian Zhao ◽  
Xiaoyu Zhang
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Cell Viability ◽  
Cell Lines ◽  
Underlying Mechanism ◽  
Migration And Invasion ◽  
Invasion And Migration ◽  
Proliferation And Metastasis ◽  
And Migration ◽  
Cell Proliferation And Metastasis

Abstract MiR-301a is as an oncogene involved in the regulation of gastric cancer (GC) progression, but the underlying mechanism is unclear. This study was to explore the lncRNA PCAT18/miR-301a/TP53INP1 axis in regulating the GC cell proliferation and metastasis. In the present study, GC tissues and cell lines were collected for the detection of PCAT18 expression. Herein, we found that PCAT18 is significantly decreases in human GC tissues and five GC cell lines. Overexpression of PCAT18 inhibits cell viability, invasion and migration of GC cells and tumour growth of GC xenograft tumours. PCAT18 negatively regulates the expression level of miR-301a. The interaction between PCAT18 and miR-301a is confirmed by RIP and RNA pull down. MiR-301a mimic increases cell viability and promotes cell migration and invasion and reverses the inhibitory action of PCAT18. TP53INP1 expression is negatively regulated by miR-301a and TP53INP1/miR-301a is involved in GC viability, migration and invasion. The promoting of PCAT18 on TP53INP1 expression is abolished by miR-301a overexpression. In conclusion, lncRNA PCAT18 acts as a tumour suppressor for GC and lncRNA PCAT18, miR-301a and TP53INP1 comprise a signal axis in regulating GC cell proliferation, migration and invasion.

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