Apigenin attenuates tetrabromobisphenol A-induced cytotoxicity in neuronal SK-N-MC cells.

Tetrabromobisphenol A (TBBPA) is a reactive brominated flame retardant widely used in various industrial and household products. This compound is persistent in the environment and accumulates in living organisms through the food chain, and is toxic to animals and human beings. Studies have shown that TBBPA is toxic to various human cell lines, including neuronal cells. Apigenin is a dietary flavonoid that exhibits various beneficial health effects on biological activities, including antioxidant, anti-inflammatory, and neuroprotective effects. This study investigated the cytoprotective effects of apigenin against TBBPA-mediated cytotoxicity in SK-N-MC cells. Our results demonstrated that treatment of SK-N-MC cells with apigenin increased the cell viability, which was decreased by TBBPA, and reduced apoptosis and autophagy induced by TBBPA. Although we did not observe any change in the levels of IL-1ß and nitrite in cultured cells after TBBPA treatment, apigenin was found to decrease the production of these pro-inflammatory mediators. Apigenin decreased the intracellular Ca2+ concentration, NOX4 level, oxidative stress, and mitochondrial membrane potential loss and increased the mitochondrial biogenesis and nuclear Nrf2 levels that were reduced by TBBPA. Finally, apigenin treatment decreased Akt and ERK induction in cells exposed to TBBPA. Based on these results, apigenin could be a promising candidate for designing natural drugs to treat or prevent TBBPA-related neurological disorders.

Protective effects of sciadopitysin against methylglyoxal-induced degeneration in neuronal SK-N-MC cells.

The accumulation of advanced glycation end products (AGEs) causes metabolic dysfunction and neuronal cell damage. Methylglyoxal (MG) is a major glycating agent that reacts with basic residues present in proteins and promotes the formation of AGEs. Sciadopitysin, a type of biflavonoid, exerts protective effects against neuronal cell damage; however, the underlying mechanisms have not been studied. This study aimed to investigate the mechanisms underlying the protective effects of sciadopitysin against MG-mediated cytotoxicity in SK-N-MC neuroblastoma cells. Our results demonstrated that pretreatment of SK-N-MC cells with sciadopitysin improved the cell viability that was inhibited by MG and inhibited the apoptosis induced by MG. Sciadopitysin attenuated intracellular Ca2+ , NOX4 levels, oxidative stress, and MG-protein adduct levels, and increased nuclear Nrf2 and glyoxalase 1 levels in the presence of MG. These results suggest that sciadopitysin exerts neuroprotective effects against MG-induced death of human SK-N-MC cells via its antioxidative action. This study highlights sciadopitysin as a promising candidate for antioxidant therapy and designing natural drugs against AGE-induced neurodegenerative disorders.

Spironolactone Attenuates Methylglyoxal-induced Cellular Dysfunction in MC3T3-E1 Osteoblastic Cells.

BACKGROUND: Methylglyoxal (MG) is associated with the pathogenesis of age- and diabetes-related complications. Spironolactone is a competitive antagonist of aldosterone that is widely employed in the treatment of hypertension and heart failure. This study examined the effects of spironolactone on MG-induced cellular dysfunction in MC3T3-E1 osteoblastic cells. METHODS: MC3T3-E1 cells were treated with spironolactone in the presence of MG. The mitochondrial function, bone formation activity, oxidative damage, inflammatory cytokines, glyoxalase I activity, and glutathione (GSH) were measured. RESULTS: Pretreatment of MC3T3-E1 osteoblastic cells with spironolactone prevented MG-induced cell death, and improved bone formation activity. Spironolactone reduced MG-induced endoplasmic reticulum stress, production of intracellular reactive oxygen species, mitochondrial superoxides, cardiolipin peroxidation, and inflammatory cytokines. Pretreatment with spironolactone also increased the level of reduced GSH and the activity of glyoxalase I. MG induced mitochondrial dysfunction, but markers of mitochondrial biogenesis such as mitochondrial membrane potential, adenosine triphosphate, proliferator-activated receptor gamma coactivator 1α, and nitric oxide were significantly improved by treatment of spironolactone. CONCLUSION: Spironolactone could prevent MG-induced cytotoxicity in MC3T3-E1 osteoblastic cells by reduction of oxidative stress. The oxidative stress reduction was explained by spironolactone's inhibition of advanced glycation end-product formation, restoring mitochondrial dysfunction, and anti-inflammatory effect.

Oleuropein attenuates the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-perturbing effects on pancreatic ß-cells.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an endocrine disrupting compound and persistent organic pollutant that has been associated with diabetes in several epidemiological studies. Oleuropein, a major phenolic compound in olive fruit, is a superior antioxidant and radical scavenger. This study aimed to examine the effects of oleuropein against TCDD-induced stress response in a pancreatic beta cell line, INS-1 cells. Cells were pre-incubated with various concentrations of oleuropein and then stimulated with TCDD (10 nM) for 48 hrs. When treated with TCDD, INS-1 cells produced robust amounts of prostaglandin E2 (PGE2) compared to the untreated control, and this increase was inhibited by oleuropein treatment. TCDD increased Ca2+-independent phospholipase A2 (iPLA2ß) level, but had no effect on Group 10 secretory phospholipase A2 (PLA2G10) level, while oleuropein deceased the levels of iPLA2ß and PLA2G10 in the presence of TCDD. Cyclooxygenase-1 (COX-1) was significantly increased by TCDD treatment and attenuated with oleuropein pretreatment. Oleuropein decreased TCDD-mediated production of JNK, TNF-α, and ROS. In addition, oleuropein increased Akt and GLUT2 levels suppressed by TCDD in INS-1 cells. Thus, the results suggest that oleuropein prevents pancreatic beta cell impairment by TCDD.

Catalpol protects against 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced cytotoxicity in osteoblastic MC3T3-E1 cells.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a well-known environmental contaminant that produces a wide variety of adverse effects in humans. Catalpol, a major bioactive compound enriched in the dried root of Rehmannia glutinosa, is a major iridoid glycoside that alleviates bone loss. However, the detailed mechanisms underlying the effects of catalpol remain unclear. The present study evaluated the effects of catalpol on TCDD-induced cytotoxicity in osteoblastic MC3T3-E1 cells. Catalpol inhibited TCDD-induced reduction in cell viability and increases in apoptosis and autophagic activity in osteoblastic MC3T3-E1 cells. Additionally, pretreatment with catalpol significantly decreased the nitric oxide and nitrite levels compared with a control in TCDD-treated cells and significantly inhibited TCDD-induced increases in the levels of cytochrome P450 1A1 and extracellular signal-regulated kinase. Pretreatment with catalpol also effectively restored the expression of superoxide dismutase and extracellular signal-regulated kinase 1 and significantly enhanced the expression of glutathione peroxidase 4 and osteoblast differentiation markers, including alkaline phosphatase and osterix. Taken together, these findings demonstrate that catalpol has preventive effects against TCDD-induced damage in MC3T3-E1 osteoblastic cells.

Tetrabromobisphenol A Promotes the Osteoclastogenesis of RAW264.7 Cells Induced by Receptor Activator of NF-kappa B Ligand In Vitro.

BACKGROUND: Tetrabromobisphenol A (TBBPA), one of the most widely used brominated flame-retardants, is a representative persistent organic pollutants group. Studies on TBBPA toxicity have been conducted using various target cells; however, few studies have investigated TBBPA toxicity in bone cells. Therefore, this study investigated the in vitro effects of TBBPA on osteoclasts, a cell type involved in bone metabolism. METHODS: RAW264.7 cells were cultured in medium containing 50 ng/mL receptor activator of nuclear factor kappa B ligand (RANKL) and varying concentrations of TBBPA. To evaluate the effects of TBBPA on the differentiation and function of osteoclasts, osteoclast-specific gene expression, tartrate-resistant acid phosphatase (TRAP) activity, bone resorbing activity, mitochondrial membrane potential (MMP) and mitochondrial superoxide were measured. RESULTS: The presence of 20 µ TBBPA significantly increased TRAP activity in RANKL-stimulated RAW264.7 cells, the bone resorbing activity of osteoclasts, and the gene expression of Akt2, nuclear factor of activated T-cells cytoplasmic 1, and chloride channel voltage-sensitive 7. However, TBBPA treatment caused no change in the expression of carbonic anhydrase II, cathepsin K, osteopetrosis-associated transmembrane protein 1, Src, extracellular signal-related kinase, GAB2, c-Fos, or matrix metalloproteinase 9. Furthermore, 20 µ TBBPA caused a significant decrease in MMP and a significant increase in mitochondrial superoxide production. CONCLUSION: This study suggests that TBBPA promotes osteoclast differentiation and activity. The mechanism of TBBPA-stimulated osteoclastogenesis might include increased expression of several genes involved in osteoclast differentiation and reactive oxygen species production.

Biochanin A prevents 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced adipocyte dysfunction in cultured 3T3-L1 cells.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental pollutant. TCDD accumulates in the food chain, mainly in the fatty tissues of the human body where it causes various toxic effects. Biochanin A is a natural organic compound in the class of phytochemicals known as flavonoids. We investigated whether biochanin A suppresses TCDD-induced loss of adipogenic action using 3T3-L1 adipocytes as a cell culture model of wasting syndrome. In the present study, biochanin A suppressed TCDD-induced loss of lipid accumulation. Pretreating the cells with biochanin A increased the levels of the adipogenesis-associated factors peroxisome proliferator-activated receptor γ and adiponectin, which were inhibited by TCDD. TCDD decreased insulin-stimulated glucose uptake, which was effectively restored by pretreatment with biochanin A. Biochanin A also inhibited the TCDD-driven decrease in production of insulin receptor substrate-1 and glucose transporter 4. These results suggest a preventive effect of biochanin A against TCDD in the development of insulin resistance and diabetes. TCDD increased production of intracellular calcium ([Ca2+]i), prostaglandin E2, cytosolic phospholipase A2, and cyclooxygenase-1, while reducing the level of peroxisome proliferator-activated receptor gamma coactivator 1-alpha. However, biochanin A inhibited these TCDD-induced effects. We conclude that biochanin A is an attractive compound for preventing TCDD-induced wasting syndrome.

Cytoprotective effects of xanthohumol against methylglyoxal-induced cytotoxicity in MC3T3-E1 osteoblastic cells.

Methylglyoxal (MG) has been suggested to be a major source of intracellular reactive carbonyl compounds, and has been implicated in increasing the levels of advanced glycation end products in age-related diseases. Xanthohumol is a prenylated flavonoid found in hops (Humulus lupulus) and beer. In the present study, we investigated the effects of xanthohumol on MG-induced cytotoxicity in osteoblastic MC3T3-E1 cells. Xanthohumol attenuated MG-induced cytotoxicity, as evidenced by improved cell viability, and prevented MG-induced MG-protein adducts, inflammatory cytokines, reactive oxygen species and mitochondrial superoxide production. In addition, xanthohumol increased glyoxalase I activity, glutathione, heme oxygenase-1 and nuclear factor erythroid 2-related factor 2 levels in the presence of MG. Pretreatment with xanthohumol before MG exposure reduced MG-induced mitochondrial dysfunction. Furthermore, xanthohumol treatment resulted in a significant reduction in the levels of endoplasmic reticulum stress and autophagy induced by MG. Notably, the autophagy-reducing effect of xanthohumol was abolished after the addition of Ex527, a selective inhibitor of sirtuin 1, suggesting that xanthohumol is an effective sirtuin 1 activator for reducing autophagy. Taken together, our findings suggest xanthohumol as a promising new strategy for preventing diabetic osteopathy.

The protective effects of sciadopitysin against methylglyoxal-induced cytotoxicity in cultured pancreatic ß-cells.

Increased glycation of macromolecules via the reactive dicarbonyl and α-oxoaldehyde methylglyoxal (MG) has shown an association with diabetes and its complications. In the present study, the protective effects of sciadopitysin against MG-induced oxidative cell damage were investigated in the insulin-producing pancreatic ß-cell line, RIN-m5F cells. When exposed to MG for 48 hours, RIN-m5F cells experienced significant loss of viability and impaired insulin secretion; however, treatment with sciadopitysin protected RIN-m5F cells against MG-induced cell death and decreased insulin secretion. Treatment of RIN-m5F cells with sciadopitysin prevented MG-induced production of interleukin-1ß, intracellular reactive oxygen species and cardiolipin peroxidation. Furthermore, sciadopitysin increased adenosine monophosphate-activated protein kinase phosphorylation of RIN-m5F cells. Treatment of cells with sciadopitysin increased the activity of glyoxalase I and decreased the levels of MG-protein adducts, indicating that sciadopitysin protects against MG-induced protein glycation by increasing MG detoxification. Taken together, the results indicated the potential utility of sciadopitysin as an intervention against MG-induced cell damage in pancreatic ß-cells.

Glabridin attenuates antiadipogenic activity induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in murine 3T3-L1 adipocytes.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) has various toxicological effects in adipose tissue. Evidence is accumulating that glabridin, a flavonoid extracted from licorice, has beneficial effects on the regulation of glucose homeostasis. In this study, we investigated whether glabridin suppresses TCDD-induced loss of adipogenic action using 3T3-L1 adipocytes as a cell culture model of wasting syndrome. Glabridin effectively suppressed TCDD-induced loss of lipid accumulation in this model. Pretreating cells with glabridin increased the gene expression of not only the adipogenesis-associated key transcription factors peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer binding protein alpha, but also lipoprotein lipase in the presence of TCDD. TCDD decreased insulin-stimulated glucose uptake, which was effectively restored by pretreatment with glabridin. Glabridin also inhibited the TCDD-driven decreased production of insulin receptor substrate 1 and glucose transporter 4. TCDD increased the production of mitochondrial superoxides, prostaglandin E2 , phospholipase A2 , cyclooxygenase-1 and intracellular calcium concentrations, while reducing the production of PPARγ coactivator 1 alpha and glycolysis. However, glabridin treatment reduced these TCDD-induced effects. We conclude that glabridin suppresses the TCDD-induced loss of lipid accumulation in 3T3-L1 adipocytes by regulating the levels of PPARγ, CCAAT/enhancer binding protein alpha, lipoprotein lipase, glucose uptake, prostaglandin E2 and energy metabolism. These results also provide in vitro evidence of the effects of glabridin on adipocyte metabolism, which suggests a protective effect against dioxin exposure in the development of insulin resistance and diabetes.

Xanthohumol ameliorates 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced cellular toxicity in cultured MC3T3-E1 osteoblastic cells.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is an environmental contaminant. Xanthohumol is a prenylated flavonoid found in hops (Humulus lupulus) and beer. The aim of the current study was to explore the role of xanthohumol in modulating the toxicity of TCDD in MC3T3-E1 osteoblastic cells. In cells treated with TCDD alone, intracellular Ca2+ concentrations, mitochondrial membrane potential disruption, reactive oxygen species production, cardiolipin peroxidation, nitric oxide release and cytochrome P450 1A1 expression were significantly increased. TCDD treatment increased the mRNA levels of extracellular signal-regulated kinase 1 and nuclear factor kappa B, and significantly decreased the level of protein kinase B (AKT) in MC3T3-E1 osteoblastic cells. However, the presence of xanthohumol alleviated the pathological effects of TCDD. In addition, xanthohumol treatment significantly increased the expression of genes associated with osteoblast differentiation (alkaline phosphatase, osteocalcin, osteoprotegerin and osterix). We conclude that xanthohumol has a beneficial influence and may antagonize TCDD toxicity in osteoblastic cells.

27-Deoxyactein prevents 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced cellular damage in MC3T3-E1 osteoblastic cells.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a well-known environmental contaminant that exerts its toxicity through a variety of signaling mechanisms. The present study evaluated the effects of 27-deoxyactein, one of the major constituents isolated from Cimicifuga racemosa, on TCDD-induced toxicity in osteoblastic MC3T3-E1 cells. TCDD reduced cell survival, markedly increased apoptosis, and enhanced autophagy activity. However, pre-treatment with 27-deoxyactein attenuated all TCDD-induced effects and significantly decreased intracellular calcium (Ca2+) concentrations, the collapse of the mitochondrial membrane potential (MMP), the level of reactive oxygen species (ROS), and cardiolipin peroxidation compared to the TCDD-treated controls. Additionally, TCDD-induced increases in the levels of aryl hydrocarbon receptor (AhR), cytochrome P450 1A1 (CYP1A1), and extracellular signal-regulated kinase (ERK) were significantly inhibited by 27-deoxyactein. The mRNA levels of superoxide dismutase (SOD), ERK1, and nuclear factor kappa B (NF-κB) were also effectively restored by pre-treatment with 27-deoxyactein. Furthermore, 27-deoxyactein significantly increased the expressions of genes associated with osteoblast differentiation, including alkaline phosphatase (ALP), osteocalcin, bone sialoprotein (BSP), and osterix. Taken together, the present findings demonstrate the preventive effects of 27-deoxyactein on TCDD-induced damage in osteoblasts.

Actein alleviates 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated cellular dysfunction in osteoblastic MC3T3-E1 cells.

The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is known to affect bone metabolism. We evaluated the protective effects of the triterpene glycoside actein from the herb black cohosh against TCDD-induced toxicity in MC3T3-E1 osteoblastic cells. We found that TCDD significantly reduced cell viability and increased apoptosis and autophagy in MC3T3-E1 osteoblastic cells (P < .05). In addition, TCDD treatment resulted in a significant increase in intracellular calcium concentration, mitochondrial membrane potential collapse, reactive oxygen species (ROS) production, and cardiolipin peroxidation, whereas pretreatment with actein significantly mitigated these effects (P < .05). The effects of TCDD on extracellular signal-related kinase (ERK), aryl hydrocarbon receptor, aryl hydrocarbon receptor repressor, and cytochrome P450 1A1 levels in MC3T3-E1 cells were significantly inhibited by actein. The levels of superoxide dismutase, ERK1, and nuclear factor kappa B mRNA were also effectively restored by pretreatment with actein. Furthermore, actein treatment resulted in a significant increase in alkaline phosphatase (ALP) activity and collagen content, as well as in the expression of genes associated with osteoblastic differentiation (ALP, type I collagen, osteoprotegerin, bone sialoprotein, and osterix). This study demonstrates the underlying molecular mechanisms of cytoprotection exerted by actein against TCDD-induced oxidative stress and osteoblast damage.

Actein protects against methylglyoxal-induced oxidative damage in osteoblastic MC3T3-E1 cells.

BACKGROUND: Methylglyoxal (MG) is an endogenous product of glucose metabolism known to be toxic to cells and to be present in elevated concentrations under certain pathophysiological conditions. In the present study the effect of actein isolated from black cohosh on MG-induced cytotoxicity was investigated in MC3T3-E1 osteoblastic cells. RESULTS: Treatment of MC3T3-E1 osteoblastic cells with actein prevented MG-induced cell death and the production of intracellular reactive oxygen species (ROS), mitochondrial superoxide, inflammatory cytokines and soluble receptor for advanced glycation end-products (sRAGE). In addition, actein increased the activity of glyoxalase I and levels of reduced glutathione (GSH) and the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2). These findings suggest that actein protects against MG-induced cell damage by reducing oxidative stress and increasing MG detoxification. Treatment with actein prior to MG exposure reduced MG-induced mitochondrial dysfunction by preventing mitochondrial membrane potential dissipation and adenosine triphosphate (ATP) loss. Additionally, peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1) and nitric oxide (NO) levels were significantly increased by actein, suggesting that actein may induce mitochondrial biogenesis. CONCLUSION: This study demonstrates that actein reduces MG-induced damage in osteoblastic MC3T3-E1 cells by enhancing antioxidant defenses, the glyoxalase system and mitochondrial biogenesis. © 2016 Society of Chemical Industry.

Perfluorooctanoic acid induces mitochondrial dysfunction in MC3T3-E1 osteoblast cells.

Perfluorooctanoic acid (PFOA), a stable organic perfluorinated compound, is an emerging persistent organic pollutant, found widely in human and wildlife populations. Recent evidence suggests that exposure to environmental toxicants can be associated with higher risks of osteoporosis and fractures. We studied the cellular toxicology of PFOA in MC3T3-E1osteoblast cells. To examine the effect of PFOA, we measured cell viability, reactive oxygen species (ROS), mitochondrial superoxide, and mitochondrial parameters including adenosine triphosphate (ATP) level, mitochondrial membrane potential (MMP), cardiolipin content, and cytochrome c release in MC3T3-E1 cells. Incubating MC3T3-E1 cells in different concentrations of PFOA for 48 h resulted in a concentration-dependent decrease in cell viability and significant inductions of ROS and mitochondrial superoxide. Moreover, PFOA induced MMP collapse, cardiolipin peroxidation, cytochrome c release, and decreased ATP levels, which in turn induced apoptosis or necrosis. When osteoblast differentiation markers were assessed, PFOA treatment caused a significant reduction in alkaline phosphatase activity, collagen synthesis, and mineralization in the cells. In summary, we found an ROS- and mitochondria-mediated pathway for the induction of cell damage by PFOA in MC3T3-E1 cells. Together, our results indicate that mitochondrial toxicity could be a plausible mechanism for the toxic effects of PFOA on osteoblast function.

Tetrabromobisphenol A induces cellular damages in pancreatic ß-cells in vitro.

Tetrabromobisphenol A (TBBPA) is a well-known organobrominated flame retardant. TBBPA has been detected in the environment. The roles played by environmental pollutants in increasing the prevalence of metabolic syndrome are attracting increasing concern. In the present work, we investigated the effects of TBBPA on rat pancreatic ß-cells (the RIN-m5F cell line). RIN-m5F cells were incubated with different concentrations of TBBPA for 48 h, and cell viability and the extent of apoptosis were determined. We also measured the levels of inflammatory cytokines, reactive oxygen species (ROS), mitochondrial adenosine triphosphate (ATP), and cardiolipin, as well as the extent of cytochrome c release from mitochondria. TBBPA reduced the ATP level, induced cardiolipin peroxidation and cytochrome c release, and triggered apoptotic cell death. Moreover, TBBPA increased the levels of inflammatory cytokines (TNF-α and IL-1ß), nitric oxide, intracellular ROS, and mitochondrial superoxide. Together, our results indicate that TBBPA damages pancreatic ß-cells by triggering mitochondrial dysfunction and inducing apoptosis.

Exposure to tetrabromobisphenol A induces cellular dysfunction in osteoblastic MC3T3-E1 cells.

This study was undertaken to investigate the possible involvement of oxidative stress in tetrabromobisphenol A (TBBPA)-induced toxicity in osteoblastic MC3T3-E1 cells. To examine the potential effect of TBBPA on cultured osteoblastic cells, we measured cell viability, apoptosis, reactive oxygen species (ROS), mitochondrial superoxide, and mitochondrial parameters including adenosine triphosphate (ATP) level, cardiolipin content, cytochrome c release, cyclophilin levels, and differentiation markers in osteoblastic MC3T3-E1 cells. TBBPA exposure for 48 h caused the apoptosis and cytotoxicity of MC3T3-E1 cells. TBBPA also induced ROS and mitochondrial superoxide production in a concentration-dependent manner. These results suggest that TBBPA induces osteoblast apoptosis and ROS production, resulting in bone diseases. Moreover, TBBPA induced cardiolipin peroxidation, cytochrome c release, and decreased ATP levels which induced apoptosis or necrosis. TBBPA decreased the differentiation markers, collagen synthesis, alkaline phosphatase activity, and calcium deposition in cells. Additionally, TBBPA decreased cyclophilin A and B releases. Taken together, these data support the notion that TBBPA inhibits osteoblast function and has detrimental effects on osteoblasts through a mechanism involving oxidative stress and mitochondrial dysfunction.

Inhibitory effect of apocynin on methylglyoxal-mediated glycation in osteoblastic MC3T3-E1 cells.

Methylglyoxal (MG), a highly reactive metabolite of hyperglycemia, can enhance protein glycation, oxidative stress or inflammation. The present study investigated the effects of apocynin on the mechanisms associated with MG toxicity in osteoblastic MC3T3-E1 cells. Pretreatment of MC3T3-E1 cells with apocynin prevented the MG-induced protein glycation and formation of intracellular reactive oxygen species and mitochondrial superoxide in MC3T3-E1 cells. In addition, apocynin increased glutathione levels and restored the activity of glyoxalase I inhibited by MG. These findings suggest that apocynin provide a protective action against MG-induced cell damage by reducing oxidative stress and by increasing the MG detoxification system. Apocynin treatment decreased the levels of proinflammatory cytokines such as tumor necrosis factor-α and interleukin-6 induced by MG. Additionally, the nitric oxide level reduced by MG was significantly increased by apocynin. These findings indicate that apocynin might exert its therapeutic effects via upregulation of glyoxalase system and antioxidant activity. Taken together, apocynin may prove to be an effective treatment for diabetic osteopathy.

Liquiritigenin restores osteoblast damage through regulating oxidative stress and mitochondrial dysfunction.

We investigated the protective effect of liquiritigenin, one of the flavonoids present in Glycyrrhizae radix, against antimycin A-induced mitochondrial dysfunction in MC3T3-E1 osteoblast cells. Osteoblastic MC3T3-E1 cells were pre-incubated with liquiritigenin before treatment with antimycin A, and markers of mitochondrial function and oxidative damage were examined. In addition, the effects of liquiritigenin on the activation of phosphoinositide 3-kinase (PI3K) were examined in MC3T3-E1 cells. Liquiritigenin protected MC3T3-E1 cells from antimycin A-induced cell death. However, the PI3K inhibitor, LY294002, significantly attenuated liquiritigenin-mediated cell survival, indicating the involvement of PI3K in the cytoprotective effect of liquiritigenin. Pretreatment with liquiritigenin prior to antimycin A exposure significantly reduced antimycin A-induced PI3K inactivation, mitochondrial membrane potential dissipation, complex IV inactivation, and ATP loss. Liquiritigenin also reduced mitochondrial superoxide generation, nitrotyrosine production, and cardiolipin peroxidation during mitochondrial complex inhibition with antimycin A. Taken together, the results of this study show that modulation of PI3K, antioxidant effects, and the attenuation of mitochondrial dysfunction by liquiritigenin represent an important mechanism for its protection of osteoblasts against cytotoxicity resulting from mitochondrial oxidative stress.

Pinacidil stimulates osteoblast function in osteoblastic MC3T3-E1 cells.

This study examined the effect of pinacidil, a nonselective adenosine triphosphate-sensitive potassium channel opener, on the function of osteoblastic MC3T3-E1 cells. Pinacidil caused a significant elevation of collagen synthesis, alkaline phosphatase activity, osteocalcin synthesis and mineralization in the cells (p < 0.05). Pinacidil significantly decreased the production of osteoclast differentiation inducing factors such as TNF-α, IL-6 and receptor activator of nuclear factor-κB ligand in the presence of antimycin A, which inhibits mitochondrial electron transport. Moreover, pinacidil prevented antimycin A-induced reactive oxygen species and nitrotyrosine production. These results demonstrate that pinacidil may have positive effects on skeletal structure.