scholarly journals Tert-Butylhydroquinone Prevents Oxidative Stress-Mediated Apoptosis and Extracellular Matrix Degradation in Rat Chondrocytes

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Bo Yang ◽  
Haisheng Huang ◽  
Qisong He ◽  
Wei Lu ◽  
Lu Zheng ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Extracellular Matrix ◽  
Matrix Metalloproteinase ◽  
Protein Expression ◽  
Cell Viability ◽  
Cell Counting ◽  
Expression Levels ◽  
Extracellular Matrix Components ◽  
Mrna And Protein Expression

Oxidative stress-induced chondrocyte apoptosis and degradation of the extracellular matrix (ECM) play an important role in the progression of osteoarthritis (OA). In addition, tert-butylhydroquinone (TBHQ) is an activator of the nuclear factor erythroid derived-2-related factor 2 (Nrf2). The present study aimed to determine the effectiveness of TBHQ in preventing the apoptosis of chondrocytes and degradation of the extracellular matrix, induced by oxidative stress, in vitro. Therefore, rat chondrocytes were exposed to 20 μM tert-butyl hydroperoxide (TBHP) for 24 h to establish an oxidative damage model, in vitro. Thereafter, cell viability was evaluated using the Cell Counting Kit-8 assay. Moreover, the level of ROS was determined through 2′,7′-dichlorofluorescein diacetate staining. The mitochondrial membrane potential of chondrocytes was also measured using JC-1. Furthermore, cell apoptosis was assessed through Annexin V-fluorescein isothiocyanate/propidium iodide staining. The study also performed Western blotting and qPCR to evaluate the expression of extracellular matrix components, matrix catabolic enzymes, and changes in signalling pathways. The results showed that 2.5 and 5 μM of TBHQ reduced the TBHP-induced generation of excessive ROS and improved cell viability. Additionally, 2.5 and 5 μM of TBHQ prevented mitochondrial damage and apoptosis in rat chondrocytes. Treatment with TBHQ also increased the mRNA and protein expression levels of aggrecan and collagen II. However, TBHQ reduced the mRNA and protein expression levels of matrix metalloproteinase 3 (MMP3) and matrix metalloproteinase 13 (MMP13) in rat chondrocytes. In addition, treatment with TBHQ enhanced the protein expression levels of Nrf2, NADPH quinone oxidoreductase 1 (NQO-1), and hemeoxygenase-1 (HO-1) in rat chondrocytes. The current study showed that TBHQ was not only effective in protecting against TBHP-induced oxidative stress but also inhibited the apoptosis of rat chondrocytes and degradation of the ECM by activating the Nrf2 pathway. The results therefore suggest that TBHQ holds potential for use in the treatment of OA.

scholarly journals Extracellular SPARC cooperates with TGF-β signalling to induce pro-fibrotic activation of systemic sclerosis patient dermal fibroblasts

Rheumatology ◽  
2019 ◽  
Vol 59 (9) ◽  
pp. 2258-2263 ◽  
Author(s):  
Tiago Carvalheiro ◽  
Beatriz Malvar Fernández ◽  
Andrea Ottria ◽  
Barbara Giovannone ◽  
Wioleta Marut ◽  
...  
Keyword(s):  
Protein Expression ◽  
Therapeutic Target ◽  
Dermal Fibroblasts ◽  
Dependent Manner ◽  
Healthy Controls ◽  
Multiple Organs ◽  
Extracellular Matrix Components ◽  
Mrna And Protein Expression

Abstract Objectives SSc is an autoimmune disease characterized by inflammation, vascular injury and excessive fibrosis in multiple organs. Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein that regulates processes involved in SSc pathology, such as inflammation and fibrosis. In vivo and in vitro studies have implicated SPARC in SSc, but it is unclear if the pro-fibrotic effects of SPARC on fibroblasts are a result of intracellular signalling or fibroblast interactions with extracellular SPARC hampering further development of SPARC as a potential therapeutic target. This study aimed to analyse the potential role of exogenous SPARC as a regulator of fibrosis in SSc. Methods Dermal fibroblasts from both healthy controls and SSc patients were stimulated with SPARC alone or in combination with TGF-β1, in the absence or presence of a TGF receptor 1 inhibitor. mRNA and protein expression of extracellular matrix components and other fibrosis-related mediators were measured by quantitative PCR and western blot. Results Exogenous SPARC induced mRNA and protein expression of collagen I, collagen IV, fibronectin 1, TGF-β and SPARC by dermal fibroblasts from SSc patients, but not from healthy controls. Importantly, exogenous SPARC induced the activation of the tyrosine kinase SMAD2 and pro-fibrotic gene expression induced by SPARC in SSc fibroblasts was abrogated by inhibition of TGF-β signalling. Conclusion These results indicate that exogenous SPARC is an important pro-fibrotic mediator contributing to the pathology driving SSc but in a TGF-β dependent manner. Therefore, SPARC could be a promising therapeutic target for reducing fibrosis in SSc patients, even in late states of the disease.

scholarly journals MicroRNA-204-5p regulates apoptosis by targeting Bcl2 in rat ovarian granulosa cells exposed to cadmium†

2020 ◽  
Vol 103 (3) ◽  
pp. 608-619
Author(s):  
Ping Zhong ◽  
Jin Liu ◽  
Hong Li ◽  
Senbin Lin ◽  
Lingfeng Zeng ◽  
...  
Keyword(s):  
Protein Expression ◽  
Granulosa Cells ◽  
B Cell Lymphoma ◽  
Experimental Conditions ◽  
Expression Levels ◽  
Ovarian Granulosa Cells ◽  
Mrna And Protein Expression ◽  
Induced Apoptosis ◽  
Apoptosis Regulation

Abstract This study aimed to investigate whether cadmium (Cd) cytotoxicity in rat ovarian granulosa cells (OGCs) is mediated through apoptosis or autophagy and to determine the role of microRNAs (miRNAs) in Cd cytotoxicity. To test this hypothesis, rat OGCs were exposed to 0, 10, and 20 μM CdCl2 in vitro. As the Cd concentration increased, OGC apoptosis increased. In addition, Cd promoted apoptosis by decreasing the mRNA and protein expression levels of inhibition of B-cell lymphoma 2 (Bcl2). However, under our experimental conditions, no autophagic changes in rat OGCs were observed, and the mRNA and protein expression levels of the autophagic markers microtubule-associated protein 1 light chain 3 alpha (Map1lc3b) and Beclin1 (Becn1) were not changed. Microarray chip analysis, miRNA screening, and bioinformatics approaches were used to further explore the roles of apoptosis regulation-related miRNAs. In total, 19 miRNAs putatively related to Cd-induced apoptosis in rat OGCs were identified. Notably, miR-204-5p, which may target Bcl2, was identified. Then, rat OGCs were cultured in vitro and used to construct the miR-204-5p-knockdown cell line LV2-short hairpin RNA (shRNA). LV2-shRNA cells were exposed to 20 μM Cd for 12 h, and the mRNA and protein expression levels of Bcl2 were increased. Our findings suggest that Cd is cytotoxic to rat OGCs, and mitochondrial apoptosis rather than autophagy mediates Cd-induced damage to OGCs. Cd also affects apoptosis-related miRNAs, and the underlying apoptotic mechanism may involve the Bcl2 gene.

scholarly journals Cytochrome c oxidase III as a mechanism for apoptosis in heart failure following myocardial infarction

2009 ◽  
Vol 297 (4) ◽  
pp. C928-C934 ◽  
Author(s):  
Changgong Wu ◽  
Lin Yan ◽  
Christophe Depre ◽  
Sunil K. Dhar ◽  
You-Tang Shen ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Myocardial Infarction ◽  
Heart Failure ◽  
Cytochrome C ◽  
Protein Expression ◽  
Cell Viability ◽  
Cytochrome C Oxidase ◽  
Mitochondrial Gene

Cytochrome c oxidase (COX) is composed of 13 subunits, of which COX I, II, and III are encoded by a mitochondrial gene. COX I and II function as the main catalytic components, but the function of COX III is unclear. Because myocardial ischemia affects mitochondrial oxidative metabolism, we hypothesized that COX activity and expression would be affected during postischemic cardiomyopathy. This hypothesis was tested in a monkey model following myocardial infarction (MI) and subsequent pacing-induced heart failure (HF). In this model, COX I protein expression was decreased threefold after MI and fourfold after HF ( P < 0.05 vs. sham), whereas COX II expression remained unchanged. COX III protein expression increased 5-fold after MI and further increased 10-fold after HF compared with sham ( P < 0.05 vs. sham). The physiological impact of COX III regulation was examined in vitro. Overexpression of COX III in mitochondria of HL-1 cells resulted in an 80% decrease in COX I, 60% decrease in global COX activity, 60% decrease in cell viability, and threefold increase in apoptosis ( P < 0.05). Oxidative stress induced by H2O2 significantly ( P < 0.05) increased COX III expression. H2O2 decreased cell viability by 47 ± 3% upon overexpression of COX III, but only by 12 ± 5% in control conditions ( P < 0.05). We conclude that ischemic stress in vivo and oxidative stress in vitro lead to upregulation of COX III, followed by downregulation of COX I expression, impaired COX oxidative activity, and increased apoptosis. Therefore, upregulation of COX III may contribute to the increased susceptibility to apoptosis following MI and subsequent HF.

scholarly journals In vitro and in vivo tenocyte-protective effectiveness of dehydroepiandrosterone against high glucose-induced oxidative stress

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Shintaro Mukohara ◽  
Yutaka Mifune ◽  
Atsuyuki Inui ◽  
Hanako Nishimoto ◽  
Takashi Kurosawa ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Matrix Metalloproteinase ◽  
Protein Expression ◽  
Achilles Tendon ◽  
Mrna Expression ◽  
High Glucose ◽  
Control Group ◽  
Type I

Abstract Background Dehydroepiandrosterone (DHEA), an adrenal steroid, has a protective role against diabetes. This study aimed to investigate the in vitro and in vivo protective effects of DHEA against high glucose-induced oxidative stress in tenocytes and tendons. Methods Tenocytes from normal Sprague-Dawley rats were cultured in low-glucose (LG) or high-glucose (HG) medium with or without DHEA. The experimental groups were: control group (LG without DHEA), LG with DHEA, HG without DHEA, and HG with DHEA. Reactive oxygen species (ROS) production, apoptosis, and messenger RNA (mRNA) expression of NADPH oxidase (NOX) 1 and 4, and interleukin-6 (IL-6) were determined. Further, diabetic rats were divided into a control group and a DHEA-injected group (DHEA group). NOX1 and NOX4 protein expression and mRNA expression of NOX1, NOX4, IL-6, matrix metalloproteinase (MMP)-2, tissue inhibitors of matrix metalloproteinase (TIMP)-2, and type I and III collagens in the Achilles tendon were determined. Results In rat tenocytes, DHEA decreased the expression of NOX1 and IL-6, ROS accumulation, and apoptotic cells. In the diabetic rat Achilles tendon, NOX1 protein expression and mRNA expression of NOX1, IL-6, MMP-2, TIMP-2, and type III collagen were significantly lower while type I collagen expression was significantly higher in the DHEA group than in the control group. Conclusions DHEA showed antioxidant and anti-inflammatory effects both in vitro and in vivo. Moreover, DHEA improved tendon matrix synthesis and turnover, which are affected by hyperglycemic conditions. DHEA is a potential preventive drug for diabetic tendinopathy.

scholarly journals Trehalose can effectively protect sheep epididymis epithelial cells from oxidative stress

2021 ◽  
Vol 64 (2) ◽  
pp. 335-343
Author(s):  
Zhaojin Luan ◽  
Xiaomei Fan ◽  
Yongchao Zhao ◽  
Huizi Song ◽  
Wei Du ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Glutathione Peroxidase ◽  
Marker Gene ◽  
Membrane Integrity ◽  
Cell Counting ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cell Membrane Integrity ◽  
Mrna And Protein Expression

Abstract. Trehalose, a naturally nontoxic disaccharide that does not exist in mammals, stabilizes cell membrane integrity under oxidative stress conditions, the mechanism of which is still unclear. Here, we analyzed the effects of trehalose on sheep epididymis epithelial cell (EEC) proliferation and its possible mechanisms. To study the effect of trehalose on EECs, EECs were isolated from testes of 12-month-old sheep; cell counting kit-8 (CCK-8) was used to measure the growth of the cells. Cell proliferation was evaluated by assaying cell cycle and apoptosis, and RT-PCR was utilized to identify the epididymal molecular markers glutathione peroxidase 5 (GPX5) and androgen receptor (AR). Next, reactive oxygen species (ROS) content was evaluated by a dichloro-dihydro-fluorescein diacetate (DCFH-DA) probe. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) activities were evaluated by enzyme chemistry methods, and GPX5 expression was evaluated by qRT-PCR and enzyme-linked immunosorbent assay (ELISA). The results showed that 100 mM trehalose significantly improved the proliferation potential of EECs, in which the cells could be serially passaged 14 times with continued normal GPX5 and AR marker gene expression in vitro. The trehalose can increase significantly a proportion of EECs in S phase (P<0.01) and decrease significantly the apoptotic rate of EECs (P<0.01) compared to the control. Moreover, the trehalose decreased ROS significantly (P<0.01) and increased CAT (P<0.01) and GSH-Px (P<0.05) activities significantly in EECs. GPX5 mRNA and protein expression were also significantly upregulated in trehalose-treated EECs (P<0.05 and P<0.01 respectively). Our study suggested that exogenous trehalose exhibited antioxidant activity through increasing the activities of CAT, GSH-Px, and the expression level of GPX5 and could be employed to maintain vitality of sheep EECs during long-term in vitro culture.

scholarly journals In Vitro and in Vivo Tenocyte-protective Effectiveness of Dehydroepiandrosterone Against High Glucose-induced Oxidative Stress

2021 ◽  
Author(s):  
Shintaro Mukohara ◽  
Yutaka Mifune ◽  
Atsuyuki Inui ◽  
Hanako Nishimoto ◽  
Takashi Kurosawa ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Matrix Metalloproteinase ◽  
Protein Expression ◽  
Achilles Tendon ◽  
Mrna Expression ◽  
High Glucose ◽  
Control Group ◽  
Type I

Abstract BackgroundDehydroepiandrosterone (DHEA), an adrenal steroid, has a protective role against diabetes. The aim of this study was to investigate the in vitro and in vivo protective effects of DHEA against high glucose-induced oxidative stress in tenocytes and tendons. Methods In an in vitro study, tenocytes from normal Sprague-Dawley rats were cultured in low-glucose (LG) or high-glucose (HG) medium with or without DHEA. The experimental groups were: control group (LG without DHEA), LG with DHEA, HG without DHEA, and HG with DHEA. Reactive oxygen species (ROS) production, apoptosis, and messenger RNA (mRNA) expression of NADPH oxidase (NOX) 1 and 4, and interleukin-6 (IL-6) were determined. In the in vivo study, diabetic rats were divided into a control group and a DHEA-injected group (DHEA group). NOX1 and NOX4 protein expression and mRNA expression of NOX1, NOX4, IL-6, matrix metalloproteinase (MMP)-2, tissue inhibitors of matrix metalloproteinase (TIMP)-2, and type I and III collagens in the Achilles tendon were determined. Results In rat tenocytes, DHEA decreased the expression of NOX1 and IL-6, ROS accumulation, and apoptotic cells. In the diabetic rat Achilles tendon, NOX1 protein expression and mRNA expression of NOX1, IL-6, MMP-2, TIMP-2, and type III collagen were significantly lower, while type I collagen expression was significantly lower in the DHEA group.Conclusions DHEA showed antioxidant and anti-inflammatory effects both in vitro and in vivo. Moreover, DHEA improved tendon matrix synthesis and turnover which are affected by hyperglycemic conditions. DHEA could be a preventive drug for the diabetic tendinopathy.

scholarly journals MiR-103a-3p antagonism attenuates pneumonia via inactivating the PI3K/AKT/NF-κB signaling pathway by targeting PTEN

2020 ◽  
Author(s):  
Lin Xu ◽  
Qingying Song ◽  
Zhanghong Ouyang ◽  
Xiangyan Zhang ◽  
Cheng Zhang
Keyword(s):  
Cell Viability ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Expression Levels ◽  
Tensin Homolog ◽  
Factor Α

Abstract Pneumonia accounts for approximately 15% mortalities in adolescents worldwide. MicroRNAs (miRNAs) regulate numerous diseases including pneumonia. miRNA and mRNA expression levels were detected by real time polymerase chain reaction (RT-qPCR). Protein expression levels were determined by enzyme-linked immunosorbent assay (ELISA) and western blot. The interaction between phosphatase and tensin homolog on chromosome ten (PTEN) and miR-103a-3p was explored by dual luciferase reporter assay. Cell viability and cell apoptosis were detected by cell Counting Kit-8 (CCK-8) and flow cytometry. Herein, we discovered that PTEN was decreased and miR-103a-3p was overexpressed in Ana-1 cells of in vitro pneumonia model. miR-103a-3p downregulated the expression levels of PTEN. AntagomiR-103a-3p reversed the increased cell apoptosis and decreased cell viability and inflammatory cytokine expression levels (tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6) induced by LPS in Ana-1 cells by PTEN. AntagomiR-103a-3p inhibited the activation of PTEN/PI3K/AKT/NF-κB signaling pathway induced by LPS in Ana-1 cells. Taken together, our findings exhibited that miR-103a-3p attenuated LPS induced pneumonia by blocking the activation of PTEN/PI3K/AKT/NF-κB signaling pathway and the following cell apoptosis as well as release of proinflammatory cytokines, suggesting that miR-103a-3p might serve as a novel therapeutic target for the treatment of pneumonia.

scholarly journals Copper Induces Oxidative Stress and Apoptosis in the Mouse Liver

2020 ◽  
Vol 2020 ◽  
pp. 1-20 ◽  
Author(s):  
Huan Liu ◽  
Hongrui Guo ◽  
Zhijie Jian ◽  
Hengmin Cui ◽  
Jing Fang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Protein Expression ◽  
Signaling Pathway ◽  
Mouse Liver ◽  
B Cell Lymphoma ◽  
Death Domain ◽  
Expression Levels ◽  
Mrna And Protein Expression ◽  
High Doses ◽  
Induced Apoptosis

Copper (Cu) is an essential trace element involved in the normal physiological processes of animals. However, excessive exposure to Cu can produce numerous detrimental impacts. The aim of this study was to investigate the effects of Cu on oxidative stress and apoptosis as well as their relationship in the mouse liver. Four-week-old ICR mice (n=240) were randomly assigned to different Cu (Cu2+-CuSO4) treatment groups (0, 4, 8, and 16 mg/kg) for periods of 21 and 42 days. The high doses of Cu exposure could induce oxidative stress, by increasing the levels of reactive oxygen species (ROS) and protein carbonyls (PC) and decreasing the activities of antisuperoxide anion (ASA) and antihydroxyl radical (AHR) and content of glutathione (GSH), as well as activities and mRNA expression levels of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px). Moreover, high doses of Cu exposure induced hepatic apoptosis via the mitochondrial apoptotic pathway, as characterized by the depolarization of mitochondrial membrane potential (MMP); significantly increased mRNA and protein expression levels of cytosolic cytochrome (Cyt c), apoptosis-inducing factor (AIF), endonuclease G (Endo G), apoptosis protease-activating factor-1 (Apaf-1), cleaved caspase-9, cleaved caspase-3, cleaved PARP, Bcl-2 antagonist killer (Bak), Bcl-2-associated X protein (Bax), and Bcl-2-interacting mediator of cell death (Bim); and decreased mRNA and protein expression levels of B-cell lymphoma-2 (Bcl-2) and Bcl-extra-large (Bcl-xL). Furthermore, the activation of the tumor necrosis factor receptor-1 (TNF-R1) signaling pathway was involved in Cu-induced apoptosis, as characterized by the significantly increased mRNA and protein expression levels of TNF-R1, Fas-associated death domain (FADD), TNFR-associated death domain (TRADD), and cleaved caspase-8. These results indicated that exposure to excess Cu could cause oxidative stress triggered by ROS overproduction and diminished antioxidant function, which in turn promoted hepatic apoptosis via mitochondrial apoptosis and that the TNF-R1 signaling pathway was also involved in the Cu-induced apoptosis.

scholarly journals Regulation of A Disintegrin And Metalloproteinase with ThromboSpondin Repeats-1 Expression in Human Endometrial Stromal Cells by Gonadal Steroids Involves Progestins, Androgens, and Estrogens

2006 ◽  
Vol 91 (12) ◽  
pp. 4825-4835 ◽  
Author(s):  
Jiadi Wen ◽  
Hua Zhu ◽  
Shuko Murakami ◽  
Peter C. K. Leung ◽  
Colin D. MacCalman
Keyword(s):  
Protein Expression ◽  
Stromal Cells ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Endometrial Stromal Cells ◽  
Expression Levels ◽  
Mrna And Protein Expression ◽  
Regulatory Effects ◽  
A Disintegrin And Metalloproteinase

Abstract Context: Gonadal steroids are key regulators of the extracellular matrix remodeling events that occur in the human endometrium during each menstrual cycle. The spatiotemporal expression of A Disintegrin And Metalloproteinase with ThromboSpondin repeats (ADAMTS)-1 in human endometrial stroma in vivo suggests that this novel metalloproteinase may contribute to this tightly regulated developmental process. Objective: The objective of the study was to determine whether progesterone (P4), 17β-estradiol (E2), or the nonaromatizable androgen dihydrotestosterone (DHT), alone or in combination, is capable of regulating ADAMTS-1 mRNA and protein levels in human endometrial stromal cells in a concentration- and time-dependent manner. Design: A real-time quantitative PCR strategy and Western blotting were used to examine ADAMTS-1 mRNA and protein expression levels in primary cultures of human endometrial stromal cells. Results: P4 and DHT but not E2 increased the levels of the ADAMTS-1 mRNA transcript and protein species (110 kDa) present in endometrial stromal cells in vitro in a concentration- and time-dependent manner. A combination of P4 and DHT resulted in an additional increase in stromal ADAMTS-1 expression, whereas E2 attenuated the regulatory effects of P4 and DHT in a concentration-dependent manner. The antisteroidal compounds, mifepristone (RU486) and hydroxyflutamide, were also found to inhibit specifically the P4- and DHT-mediated increase in ADAMTS-1 mRNA and protein expression levels in these primary cell cultures in a concentration-dependent manner, respectively. Conclusions: These studies demonstrate that progestins, androgens, and estrogens, alone and in combination, have distinct regulatory effects on ADAMTS-1 mRNA and protein expression levels in human endometrial stromal cells in vitro.

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