Effects of Compressive Loading on Human Synovium-derived Cells

2007 ◽  
Vol 86 (8) ◽  
pp. 786-791 ◽  
Author(s):  
Y. Muroi ◽  
K. Kakudo ◽  
K. Nakata
Keyword(s):  
Protein Expression ◽  
Temporomandibular Joint ◽  
Inflammatory Cytokines ◽  
Apoptotic Cell ◽  
Compressive Loading ◽  
Cell Number ◽  
Joint Disease ◽  
Mrna Levels ◽  
Dna Amount

Compressive stress may be involved in temporomandibular joint (TMJ) synovitis, but its mechanism has not been fully elucidated. We hypothesized that mechanical stress to the synovial cells of the TMJ potentially causes degenerative changes in temporomandibular joint disease. We examined the effect of cyclic compressive loading on three-dimensionally engineered constructs using human TMJ synovium-derived cells in vitro. Human TMJ synovium-derived cells were cultured onto collagen scaffolds, resulting in three-dimensional constructs. Cyclic compression loading was applied to the constructs by means of a custom-designed apparatus. DNA amount, apoptotic cells, and mRNA levels for inflammatory cytokines were analyzed. The protein expression and activity of MMPs were examined. DNA amount or apoptotic cell number was unchanged by loading. MMP-2, -3, and IL-8 mRNA expression was up-regulated by the compression, and both MMP-1 and -3 protein expression and MMP-2 activity were detected. Thus, compression of human TMJ synovium-derived cells appears to modulate inflammatory cytokines.

scholarly journals Polyphenols Derived from Lychee Seed Suppress Aβ (1-42)-Induced Neuroinflammation

2018 ◽  
Vol 19 (7) ◽  
pp. 2109 ◽  
Author(s):  
Yong Tang ◽  
Rui Xiong ◽  
An-Guo Wu ◽  
Chong-Lin Yu ◽  
Ya Zhao ◽  
...  
Keyword(s):  
Liquid Chromatography ◽  
Protein Expression ◽  
Inflammatory Cytokines ◽  
Amyloid Β ◽  
Neuronal Cell ◽  
Future Research ◽  
Mrna Levels ◽  
Bax Protein

Amyloid-β (Aβ) is commonly recognized as the most important factor that results in neuronal cell death and accelerates the progression of Alzheimer’s disease (AD). Increasing evidence suggests that microglia activated by Aβ release an amount of neurotoxic inflammatory cytokines that contribute to neuron death and aggravate AD pathology. In our previous studies, we found that lychee seed fraction (LSF), an active fraction derived from the lychee seed, could significantly improve the cognitive function of AD rats and inhibit Aβ-induced neuroinflammation in vitro, and decrease neuronal injuries in vivo and in vitro. In the current study, we aimed to isolate and identify the specific components in LSF that were responsible for the anti-neuroinflammation effect using preparative high performance liquid chromatography (pre-HPLC), liquid chromatography-mass spectrometry (LC-MS), and nuclear magnetic resonance (NMR) methods. To this end, we confirmed two polyphenols including catechin and procyanidin A2 that could improve the morphological status of BV-2 cells and suppress the release, mRNA levels, and protein expression of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) through downregulating the nuclear factor-κB (NF-κB) signaling pathway using ELISA, RT-PCR, and Western blotting methods. Furthermore, catechin and procyanidin A2 could inhibit Aβ-induced apoptosis in BV-2 cells by upregulating Bcl-2 and downregulating Bax protein expression. Therefore, the current study illustrated the active substances in lychee seed, and first reported that catechin and procyanidin A2 could suppress neuroinflammation in Aβ-induced BV-2 cells, which provides detailed insights into the molecular mechanism of catechin and procyanidin A2 in the neuroprotective effect, and their further validations of anti-neuroinflammation in vivo is also essential in future research.

scholarly journals Sestrin 2 Attenuates Rat Hepatic Stellate Cell (HSC) Activation and Liver Fibrosis via an mTOR/AMPK-Dependent Mechanism

2018 ◽  
Vol 51 (5) ◽  
pp. 2111-2122 ◽  
Author(s):  
Yi-Bing Hu ◽  
Xiao-Ting Ye ◽  
Qing-Qing Zhou ◽  
Rong-Quan Fu
Keyword(s):  
Liver Fibrosis ◽  
Protein Expression ◽  
Hepatic Fibrosis ◽  
Transforming Growth Factor ◽  
Histopathological Examination ◽  
Stellate Cell ◽  
Smooth Muscle Actin ◽  
Mrna Levels ◽  
Alpha Smooth Muscle Actin

Background/Aims: Sestrin 2 is associated with the pathophysiology of several diseases. The aim of this study was to investigate the effects and potential mechanisms of Sestrin 2 in rat hepatic stellate cells (HSCs) during liver fibrogenesis. Methods: In this study, Sestrin 2 protein expression was detected in rat HSC-T6 cells challenged with transforming growth factor-β (TGF-β) and in mice treated with carbon tetrachloride (CCl4), a well-known model of hepatic fibrosis. Next, HSC-T6 cells and fibrotic mice were transfected with lentivirus. The mRNA expression levels of markers of liver fibrosis [alpha-smooth muscle actin (α-SMA) and collagen 1A1 (Col1A1)] were analyzed by quantitative reverse transcription–polymerase chain reaction (RT-PCR). Cell death and proliferation were evaluated by the MTT assay, and biochemical markers of liver damage in serum [alanine transaminase (ALT) and aspartate transaminase (AST)] were also measured using a biochemical analyzer. Histopathological examination was used to evaluate the degree of liver fibrosis, and protein expression [phospho-adenosine monophosphate-activated protein kinase (p-AMPK), AMPK, phospho-mammalian target of rapamycin (p-mTOR), and mTOR] was determined by western blotting. Results: We found that Sestrin 2 was elevated in both the HSC-T6 cell and hepatic fibrosis models. In vitro, overexpression of Sestrin 2 attenuated the mRNA levels of α-SMA and Col1A1, suppressed α-SMA protein expression, and modulated HSC-T6 cell proliferation. In vivo, overexpression of Sestrin 2 reduced the ALT and AST levels as well as the α-SMA and Col1A1 protein expression in the CCl4 model of liver fibrosis. Moreover, the degree of liver fibrosis was ameliorated. Interestingly, overexpression of Sestrin 2 increased p-AMPK but decreased p-mTOR protein expression. Conclusion: Our findings indicate that Sestrin 2 may attenuate the activation of HSCs and ameliorate liver fibrosis, most likely via upregulation of AMPK phosphorylation and suppression of the mTOR signaling pathway.

scholarly journals Bioinformatic and experimental findings to indicate anti-cancer targets and mechanisms of calycosin against nasopharyngeal carcinoma

2020 ◽  
Author(s):  
Fangxian Liu ◽  
Qijin Pan ◽  
Liangliang Wang ◽  
Shijiang Yi ◽  
Peng Liu ◽  
...  
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Molecular Mechanisms ◽  
Apoptotic Cell ◽  
Cell Number ◽  
Network Pharmacology ◽  
Tunel Staining ◽  
Pharmacological Targets ◽  
Experimental Findings

Abstract Background: Calycosin is a naturally-occurring phytoestrogen that reportedly exerts anti- nasopharyngeal carcinoma (NPC) effects. Nevertheless, the molecular mechanisms for anti-NPC using calycosin remain unrevealed. Methods: Thus, a network pharmacology was used to uncover anti-NPC pharmacological targets and mechanisms of calycosin. Additionally, validated experiments were conducted to validate the bioinformatic findings of calycosin for treating NPC. Results: As results, bioinformatic assays showed that the predictive pharmacological targets of calycosin against NPC were TP53, MAPK14, CASP8, MAPK3, CASP3, RIPK1, JUN, ESR1, respectively. And the top 20 biological processes and pharmacological mechanisms of calycosin against NPC were identified accordingly. In clinical data, NPC samples showed positive expression of MAPK14, reduced TP53, CASP8 expressions. In studies in vitro and in vivo, calycosin-dosed NPC cells resulted in reduced cell proliferation, promoted cell apoptosis. In TUNEL staining, calycosin exhibited elevated apoptotic cell number. And immunostaining assays resulted in increased TP53, CASP8 positive cells, and reduced MAPK14 expressions in calycosin-dosed NPC cells and tumor-bearing nude mice. Conclusion: Altogether, these bioinformatic findings reveal optimal pharmacological targets and mechanisms of calycosin against NPC, following with representative identification of human and preclinical experiments. Notably, some of original biotargets may be potentially used to treat NPC.

scholarly journals Hexarelin exerts neuroprotective and antioxidant effects against hydrogen peroxide-induced toxicity through the modulation of MAPK and PI3K/Akt patways in Neuro-2A cells

2020 ◽  
Author(s):  
Ramona Meanti ◽  
Laura Rizzi ◽  
Elena Bresciani ◽  
Laura Molteni ◽  
Vittorio Locatelli ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Apoptotic Cell ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Inhibitory Influence ◽  
No Production ◽  
Mrna Levels

AbstractHexarelin, a synthetic hexapeptide, protects cardiac and skeletal muscles by inhibiting apoptosis, both in vitro and in vivo. Moreover, evidence suggests that hexarelin could have important neuroprotective bioactivity.Oxidative stress and the generation of free radicals has been implicated in the etiologies of several neurodegenerative diseases, including amyotrophic lateral sclerosis, Parkinson’s disease, Alzheimer’s disease, Huntington’s disease and multiple sclerosis. In addition to direct oxidative stress, exogenous hydrogen peroxide (H2O2) can penetrate biological membranes and enhance the formation of other reactive oxygen species.The aim of this study was to examine the inhibitory influence of hexarelin on H2O2-induced apoptosis in Neuro-2A cells, a mouse neuroblastoma cell line. Our results indicate that H2O2 reduced the viability of Neuro-2A cells in a dose-related fashion. Furthermore, H2O2 induced significant changes in the morphology of Neuro-2A cells, reflected in the formation of apoptotic cell bodies, and an increase of nitric oxide (NO) production. Hexarelin effectively antagonized H2O2 oxidative damage to Neuro-2A cells as indicated by improved cell viability, normal morphology and reduced nitrite (NO2−) release. Hexarelin treatment of Neuro-2A cells also reduced mRNA levels of caspases−3 and −7 and those of the pro-apoptotic molecule Bax; by contrast, hexarelin treatment increased anti-apoptotic Bcl-2 mRNA levels. Hexarelin also reduced MAPKs phosphorylation induced by H2O2 and concurrently increased p-Akt protein expression.In conclusion, our results identify several neuroprotective and anti-apoptotic effects of hexarelin. These properties suggest that further investigation of hexarelin as a neuroprotective agent in an investigational and therapeutic context are merited.

scholarly journals Cytokine-Mediated Downregulation of Coxsackievirus-Adenovirus Receptor in Endothelial Cells

2004 ◽  
Vol 78 (15) ◽  
pp. 8047-8058 ◽  
Author(s):  
Theresa Vincent ◽  
Ralf F. Pettersson ◽  
Ronald G. Crystal ◽  
Philip L. Leopold
Keyword(s):  
Endothelial Cells ◽  
Epithelial Cells ◽  
Protein Expression ◽  
Western Blot ◽  
Inflammatory Cytokines ◽  
Cell Physiology ◽  
Mrna Levels ◽  
Cytokine Treatment ◽  
Adenovirus Receptor ◽  
The Impact

ABSTRACT Endothelial cells have the ability to change their complement of cell surface proteins in response to inflammatory cytokines. We hypothesized that the expression of the coxsackievirus-adenovirus receptor (CAR), a viral receptor and putative cell-cell adhesion molecule, may be altered during the response of endothelial cells to inflammation. To test this hypothesis, we evaluated CAR protein and mRNA levels in human umbilical vein endothelial cells after they were exposed to tumor necrosis factor alpha, gamma interferon, or a combination of the two cytokines. Flow cytometric and Western blot analyses indicated that cytokine treatment led to a synergistic decrease in CAR protein expression. A Western blot analysis showed that CAR levels decreased to 16% ± 4% or 1% ± 4% of the CAR protein levels in untreated cells with either 24 or 48 h of cytokine treatment, respectively. Quantitative reverse transcription-PCR demonstrated that the combination treatment caused CAR mRNA levels to decrease to 21% ± 12% or 5% ± 3% of the levels in untreated cells after a 24- or 48-h cytokine treatment, respectively. Reduced CAR expression led to a decrease in adenovirus (Ad) binding of 80% ± 3% (compared with untreated endothelial cells), with a subsequent decrease in Ad-mediated gene transfer that was dependent on the dose and duration of cytokine treatment but not on the dose of Ad. A similar decrease in CAR protein level and susceptibility to Ad infection was observed in human microvascular endothelial cells, while CAR expression on normal human bronchial epithelial cells or A549 lung epithelial cells was less affected by cytokine treatments. Taken together, the data demonstrate that inflammatory cytokines decrease CAR mRNA and protein expression with a concomitant decrease in Ad binding, reflecting the impact of cell physiology on the function of CAR and the potential effect of inflammation on the ability of Ad to transfer genes to endothelial cells.

scholarly journals Aldosterone up-regulates voltage-gated potassium currents and NKCC1 protein membrane fractions

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Parveen Bazard ◽  
Bo Ding ◽  
Harish K. Chittam ◽  
Xiaoxia Zhu ◽  
Thomas A. Parks ◽  
...  
Keyword(s):  
Protein Expression ◽  
Mineralocorticoid Receptor ◽  
Therapeutic Potential ◽  
Potassium Currents ◽  
Chloride Ions ◽  
Mrna Levels ◽  
Mineralocorticoid Receptor Antagonist ◽  
Voltage Gated ◽  
Age Related

Abstract Na+–K+–2Cl− Cotransporter (NKCC1) is a protein that aids in the active transport of sodium, potassium, and chloride ions across cell membranes. It has been shown that long-term systemic treatment with aldosterone (ALD) can enhance NKCC1 protein expression and activity in the aging cochlea resulting in improved hearing. In the present work, we used a cell line with confirmed NKCC1 expression to demonstrate that in vitro application of ALD increased outward voltage-gated potassium currents significantly, and simultaneously upregulated whole lysate and membrane portion NKCC1 protein expression. These ALD-induced changes were blocked by applying the mineralocorticoid receptor antagonist eplerenone. However, application of the NKCC1 inhibitor bumetanide or the potassium channel antagonist Tetraethyl ammonium had no effect. In addition, NKKC1 mRNA levels remained stable, indicating that ALD modulates NKCC1 protein expression via the activation of mineralocorticoid receptors and post-transcriptional modifications. Further, in vitro electrophysiology experiments, with ALD in the presence of NKCC1, K+ channel and mineralocorticoid receptor inhibitors, revealed interactions between NKCC1 and outward K+ channels, mediated by a mineralocorticoid receptor-ALD complex. These results provide evidence of the therapeutic potential of ALD for the prevention/treatment of inner ear disorders such as age-related hearing loss.

335 EFFECT OF VITAMIN E ON THE DEVELOPMENT OF BOVINE OOCYTES AFTER CHEMICAL ACTIVATION

2007 ◽  
Vol 19 (1) ◽  
pp. 283
Author(s):  
J. I. Park ◽  
Y. Jang ◽  
E. S. Lee
Keyword(s):  
Vitamin E ◽  
Apoptotic Cell ◽  
Chemical Activation ◽  
Calf Serum ◽  
Cell Number ◽  
Total Cell ◽  
Chi Square ◽  
Cell Numbers ◽  
Humidified Air

Oxidative stress is known to induce apoptotic cell death by reactive oxygen species (ROS) generated from in vitro culture systems. This study was conducted to evaluate the effect of Vitamin E (VitE), as antioxidant, on development of bovine embryos activated in vitro. Bovine ovaries were collected from slaughtered cows at a local abattoir. Oocytes were aspirated from follicles 3-8 mm in diameter and transferred to maturation medium: tissue culture medium (TCM)-199 supplemented with 10% (v/v) fetal calf serum, 100 mg/mL-1 l-cysteine, 20 mg/mL-1 sodium pyruvate, gonadotropins (250 IU each of eCG and hCG/mL), 10 mg/mL-1 epidermal growth factor, and 100 �M VitE. Oocytes were cultured at 38.9�C in 5% CO2 in humidified air. After 22 hours of culture, oocytes with polar bodies were selected and subjected to activation treatments. Oocytes were exposed to calcium ionomycin (5 �M for 5 min), followed by incubation with 6-DMAP (2 mM) for 3.5 hours in medium supplemented with or without VitE (100 �M). After activation, oocytes were cultured in mSOF medium containing 0.8% BSA at 38.9�C in 5% CO2, 5% O2 in humidified air for 7–8 days. Cell numbers were counted by the number of nuclei of blastocysts stained with Hoechst 33342, and apoptosis was detected by TUNEL assay using a MK500 kit (Takara Bio, Inc., Otsu, Shiga, Japan). Total cell and apoptotic cell number were determined under a fluorescence microscope. Data were analyzed using Student's t-test and chi-square test. The cleavage and blastocyst rates were significantly higher (P < 0.05) after activation with VitE (78.1% and 16.3%, n = 80) than without VitE (66.7% and 11.0%, n = 60). Total cell numbers were also significantly higher (P < 0.05) in blastocysts after activation with VitE (143.0 ± 34.02, n = 21) than in those without VitE (127.63 ± 40.25, n = 20). However, the percentage of TUNEL-positive (apoptotic) cells was similar between blastocysts activated with VitE (5.38 ± 2.22) and those without VitE (6.76 ± 1.98). The results of the present study demonstrate that vitamin E added to activation medium promoted further development of activated embryos, although its role in the alleviation of apoptosis remains unclear.

scholarly journals Long-Term Exposure to Urban Particulate Matter on the Ocular Surface and the Incidence of Deleterious Changes in the Cornea, Conjunctiva and Retina in Rats

2020 ◽  
Vol 21 (14) ◽  
pp. 4976
Author(s):  
Wan Seok Kang ◽  
Hakjoon Choi ◽  
Goeun Jang ◽  
Ki Hoon Lee ◽  
Eun Kim ◽  
...  
Keyword(s):  
Particulate Matter ◽  
Apoptotic Cell ◽  
Tear Film ◽  
Cell Number ◽  
Corneal Epithelial Cell ◽  
Urban Particulate Matter ◽  
Epithelium Thickness ◽  
The Right

We investigated the time-dependent deleterious ocular changes induced by urban particulate matter (UPM) in vitro and in vivo. UPM treatment decreased human corneal epithelial cell migration and survival. Fluorescein scores were consistently increased by UPM application for 16 weeks. One week of rest at 2 or 4 weeks led to a recovery trend, whereas two weeks of rest at 8 weeks induced no change. UPM treatment decreased the tear film break-up time at 2 weeks, which was thereafter maintained until 16 weeks. No changes were found after periods of rest. UPM-treated eyes exhibited greater corneal epithelium thickness than normal eyes at 2 weeks, which recovered to normal at 4 and 8 weeks and was significantly decreased at 16 weeks. Apoptotic cell number in the epithelium was increased at 2 weeks, which remained constant except at 8 weeks. IL-6 expression in the cornea of the right eye continually increased for 16 weeks, and significant recovery was only observed at 8 weeks after 2 weeks of rest. Ocular pressure was significantly increased in the right eye at 12 and 16 weeks. Topical UPM application to the eye induced deleterious changes to various closely related parts of the eye.

scholarly journals High-Frequency Near-Infrared Diode Laser Irradiation Attenuates IL-1β-Induced Expression of Inflammatory Cytokines and Matrix Metalloproteinases in Human Primary Chondrocytes

2020 ◽  
Vol 9 (3) ◽  
pp. 881 ◽  
Author(s):  
Shuzo Sakata ◽  
Ryo Kunimatsu ◽  
Yuji Tsuka ◽  
Ayaka Nakatani ◽  
Tomoka Hiraki ◽  
...  
Keyword(s):  
Matrix Metalloproteinases ◽  
Protein Expression ◽  
Laser Irradiation ◽  
Diode Laser ◽  
High Frequency ◽  
Near Infrared ◽  
Mrna Levels ◽  
Induced Expression ◽  
Tnf Α

High-frequency near-infrared diode laser provides a high-peak output, low-heat accumulation, and efficient biostimulation. Although these characteristics are considered suitable for osteoarthritis (OA) treatment, the effect of high-frequency near-infrared diode laser irradiation in in vitro or in vivo OA models has not yet been reported. Therefore, we aimed to assess the biological effects of high-frequency near-infrared diode laser irradiation on IL-1β-induced chondrocyte inflammation in an in vitro OA model. Normal Human Articular Chondrocyte-Knee (NHAC-Kn) cells were stimulated with human recombinant IL-1β and irradiated with a high-frequency near-infrared diode laser (910 nm, 4 or 8 J/cm2). The mRNA and protein expression of relevant inflammation- and cartilage destruction-related proteins was analyzed. Interleukin (IL) -1β treatment significantly increased the mRNA levels of IL-1β, IL-6, tumor necrosis factor (TNF) -α, matrix metalloproteinases (MMP) -1, MMP-3, and MMP-13. High-frequency near-infrared diode laser irradiation significantly reduced the IL-1β-induced expression of IL-1β, IL-6, TNF-α, MMP-1, and MMP-3. Similarly, high-frequency near-infrared diode laser irradiation decreased the IL-1β-induced increase in protein expression and secreted levels of MMP-1 and MMP-3. These results highlight the therapeutic potential of high-frequency near-infrared diode laser irradiation in OA.

scholarly journals A Pilot Study on the Beneficial Effects of the Additional Selenium Supplementation to Methimazole for Treating Patients with Graves? disease

2019 ◽  
Author(s):  
BIN XU ◽  
DI WU ◽  
HONG YING ◽  
YING ZHANG
Keyword(s):  
Protein Expression ◽  
Combination Group ◽  
Thyroid Peroxidase ◽  
Graves Disease ◽  
Mrna Levels ◽  
Thyroid Cells ◽  
Beneficial Effects ◽  
Thyroid Peroxidase Antibody ◽  
Thyroglobulin Antibody

Backgroud/aim: The aim of this study was to assess the effect of a combination use of methimazole (MMI) and selenium (Se) in the treatment of Graves’ disease (GD). Materials and methods: A total of 103 newly-diagnosed hyperthyroidism patients were randomized to MMI and MMI+Se combination group. After treatment for six months, the levels of triiodothyronine (FT3), free thyroxine (FT4), thyrotrophin receptor antibody (TRAb), thyroid peroxidase antibody (TPOAb), and thyroglobulin antibody (TGAb) were observed. Besides, an in vitro culture model of thyroid cells was established and the protein expression and mRNA levels of TRAb, TPOAb and TGAb were determined by western blot and RT-PCR. Results: A significant decrease in the levels of FT3, FT4, TRAb, TPOAb and TGAb were observed in both groups along with a marked increase in TSH levels. Furthermore, the in vitro experiments showed that the protein expression and mRNA levels of TRAb, TPOAb and TGAb decreased significantly. Also, compared to the MMI group, there was a greater improvement of these indices in the MMI+Se group. Conclusion: We suggest that the combined use of MMI and Se could improve the thyroid activity in patients which may provide effective therapy for the treatment of GD in clinical settings. Key words: Graves’ disease; methimazole; selenium; TRAb; TPOAb; TGAb

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