scholarly journals Alterations in Mitochondrial and Inflammasome Homeostasis by 2-Chloroethyl Ethyl Sulfide and their Mitigation by Curcumin: An In vitro Study

2021 ◽  
Author(s):  
Azam Kia ◽  
Mona Nadi ◽  
Vahideh Hajhasan ◽  
Jafar Salimian
Keyword(s):  
Sulfur Mustard ◽  
Fusion Gene ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
In Vitro Study ◽  
Inhibitory Effect ◽  
Epithelial Cell Line ◽  
Complete Suppression ◽  
Mitofusin 2

The mitochondrion has a substantial role in innate immunity and inflammasome signaling pathways. Sulfur mustard (SM) induces toxicity in cytoplasmic organelles. We aimed to evaluate the potential therapeutic effect of curcumin on the toxicity of SM analog through measuring gene expression levels of mitochondrial dynamics followed by induction of the inflammasome signaling pathway. After the treatment of pulmonary epithelial cell line (A549) by 2-chloroethyl ethyl sulfide (CEES) (2500 mM) for 48h, the transcriptional activity of mitochondrial fission and fusion genes such as dynamin-related protein 1 (Drp1), mitochondrial fission 1 protein (Fis1), mitofusin-1 (Mfn1), mitofusin-2 (Mfn2), and Dominant optic atrophy (Opa1) and inflammasome pathway genes including absent in melanoma 2 (AIM2), NLR family containing protein 3 (NLRP3), and Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) was measured. Furthermore, the inhibitory effect of curcumin (160 mM) concurrent with SM analog on the expression level of mitochondria and inflammasome genes was investigated. CEES was able to over-express the fission, fusion (Drp1 ~ 8, Fis1 4.5, Mfn2 15, and Opa1 16-fold) and inflammasome genes (AIM2, NLRP3, 8 and 6-fold, respectively), whereas Mfn1 was significantly decreased (0.5-fold) and a not statistically significant decrease was observed in the ASC gene. Curcumin could modulate the effect of CEES, mitigate the expression of fission, fusion, and inflammasome genes exceedingly. However, a major increase in the repairer fusion gene (Mfn1, 6-fold) and complete suppression of the ASC gene were the outcomes of using the curcumin. In conclusion, we suggest curcumin alleviates the disturbance of mitochondrial dynamics and downregulates the inflammasome genes exposed to the CEES.  

Abstract 433: The Role Of Claudin-5 on Mitochondrial Dynamics in Neonatal Rat Cardiomyocyte

2017 ◽  
Vol 121 (suppl_1) ◽  
Author(s):  
Tao Luo ◽  
Han Liu ◽  
Jin K Kim
Keyword(s):  
Cytochrome C ◽  
Western Blot ◽  
Protein Level ◽  
Mitochondrial Dynamics ◽  
Ischemia Reperfusion ◽  
Mitochondrial Fission ◽  
Neonatal Rat ◽  
Mitofusin 2

Background: Claudin-5, an integral transmembrane protein that controls the para-cellular permeability, is severely downregulated in end stage cardiomyopathy and during ischemia-reperfusion injury. Sustaining claudin-5 levels prevented myocardial damage and dysfunction. However, the underlying mechanism behind the cardioprotection associated with claudin-5 is unclear. The aim of this study is therefore to investigate the role of claudin-5 on mitochondria in vitro . Methods and Results: Western blot showed that claudin-5 was expressed in mouse heart tissue and neonatal rat cardiomyocyte (NRCM). Its protein level was severely decreased after myocardial ischemia/reperfusion (I/R; 30 min/24 hr) and after hypoxia/reoxygenation (H/R; 24 hr/4 hr), respectively. Further in-vitro confocal observation showed that claudin-5 was co-expressed on mitochondria in NRCM and its decrease was accompanied by mitochondrial fragmentation. We then detected the mitochondrial fusion and fission proteins in NRCM under the condition of H/R. Claudin-5 siRNA and claudin-5 overexpressing adenovirus were used to knockdown claudin-5 and overexpress claudin-5. The protein level of mitofusin 2 (responsible for mitochondrial outer membrane fusion) was dramatically decreased while the expression of Drp 1 (responsible for mitochondrial fission) was significantly increased after H/R; Claudin-5 siRNA pretreatment further decreased the level of mitofusin 2 and increased the level of Drp 1. However, overexpression of claudin-5 reversed these effects respectively. We further measured the apoptotic protein, cytochrome c, by western blot. The expression of cytochrome c in the cytoplasm of NRCM was significantly increased after H/R, which was reduced by overexpression of claudin-5 by adenovirus. Conclusion: In the present study, we first time introduce a new concept that claudin-5 may help maintaining mitochondrial dynamics by upregulating mitochondrial fusion and suppressing mitochondrial fission.

scholarly journals Oxyberberine Prevented Lipopolysaccharide-Induced Acute Lung Injury through Inhibition of Mitophagy

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Runmin Zhao ◽  
Bingxia Wang ◽  
Dasheng Wang ◽  
Benhe Wu ◽  
Peiyu Ji ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
A549 Cells ◽  
In Vitro Study ◽  
Epithelial Cell Line ◽  
Lung Edema ◽  
Neutrophil Accumulation ◽  
Mitofusin 2

Acute lung injury (ALI) is a serious respiratory syndrome characterized with uncontrolled inflammatory response. Oxyberberine has strong potential for clinical usage since it showed strong anti-inflammatory, antifungal, and antiarrhythmic effects in various diseases. In the present study, we evaluated whether oxyberberine can inhibit lipopolysaccharide- (LPS-) induced ALI in vivo and further evaluated the possible involvement of mitophagy in vitro by using A549 cells, a human lung epithelial cell line. Our in vivo study shows that oxyberberine significantly inhibited LPS-induced lung pathological injury and lung edema, as indicated by the changes in lung wet/dry ratio and total protein levels in the BALF in mice. Moreover, oxyberberine inhibited inflammation, as indicated by the changes of neutrophil accumulation and production of proinflammatory cytokines including tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), and IL-6 in both the lung and bronchoalveolar lavage fluid (BALF) in ALI mice. Our in vitro study shows that LPS significantly decreased the protein level of mitochondrial proteins, including cytochrome c oxidase subunit IV (COX IV), p62, and mitofusin-2 (Mfn2) in A549 cells. In addition, LPS induced significant Parkin1 translocation from cytoplasm to mitochondria. These changes were significantly inhibited by oxyberberine. Notably, the inhibitory effect of oxyberberine was almost totally lost in the presence of lysosome fusion inhibitor bafilomycin A1 (Baf), a mitophagy inhibitor. In conclusion, the present study demonstrated that oxyberberine alleviated LPS-induced inflammation in ALI via inhibition of Parkin-mediated mitophagy.

scholarly journals Mitochondrial fission and mitophagy are independent mechanisms regulating ischemia/reperfusion injury in primary neurons

2021 ◽  
Vol 12 (5) ◽  
Author(s):  
Anthony R. Anzell ◽  
Garrett M. Fogo ◽  
Zoya Gurm ◽  
Sarita Raghunayakula ◽  
Joseph M. Wider ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Ischemia Reperfusion Injury ◽  
Mitochondrial Dynamics ◽  
Ischemia Reperfusion ◽  
Mitochondrial Fission ◽  
Conditional Knockout ◽  
Mitochondrial Morphology ◽  
Primary Neurons ◽  
Mitochondrial Fragmentation

AbstractMitochondrial dynamics and mitophagy are constitutive and complex systems that ensure a healthy mitochondrial network through the segregation and subsequent degradation of damaged mitochondria. Disruption of these systems can lead to mitochondrial dysfunction and has been established as a central mechanism of ischemia/reperfusion (I/R) injury. Emerging evidence suggests that mitochondrial dynamics and mitophagy are integrated systems; however, the role of this relationship in the context of I/R injury remains unclear. To investigate this concept, we utilized primary cortical neurons isolated from the novel dual-reporter mitochondrial quality control knockin mice (C57BL/6-Gt(ROSA)26Sortm1(CAG-mCherry/GFP)Ganl/J) with conditional knockout (KO) of Drp1 to investigate changes in mitochondrial dynamics and mitophagic flux during in vitro I/R injury. Mitochondrial dynamics was quantitatively measured in an unbiased manner using a machine learning mitochondrial morphology classification system, which consisted of four different classifications: network, unbranched, swollen, and punctate. Evaluation of mitochondrial morphology and mitophagic flux in primary neurons exposed to oxygen-glucose deprivation (OGD) and reoxygenation (OGD/R) revealed extensive mitochondrial fragmentation and swelling, together with a significant upregulation in mitophagic flux. Furthermore, the primary morphology of mitochondria undergoing mitophagy was classified as punctate. Colocalization using immunofluorescence as well as western blot analysis revealed that the PINK1/Parkin pathway of mitophagy was activated following OGD/R. Conditional KO of Drp1 prevented mitochondrial fragmentation and swelling following OGD/R but did not alter mitophagic flux. These data provide novel evidence that Drp1 plays a causal role in the progression of I/R injury, but mitophagy does not require Drp1-mediated mitochondrial fission.

scholarly journals Non-alcoholic fatty liver disease in mice with hepatocyte-specific deletion of mitochondrial fission factor

Diabetologia ◽  
2021 ◽  
Author(s):  
Yukina Takeichi ◽  
Takashi Miyazawa ◽  
Shohei Sakamoto ◽  
Yuki Hanada ◽  
Lixiang Wang ◽  
...  
Keyword(s):  
Er Stress ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Primary Hepatocytes ◽  
Alcoholic Fatty Liver ◽  
Expression Of Genes ◽  
Specific Deletion

Abstract Aims/hypothesis Mitochondria are highly dynamic organelles continuously undergoing fission and fusion, referred to as mitochondrial dynamics, to adapt to nutritional demands. Evidence suggests that impaired mitochondrial dynamics leads to metabolic abnormalities such as non-alcoholic steatohepatitis (NASH) phenotypes. However, how mitochondrial dynamics are involved in the development of NASH is poorly understood. This study aimed to elucidate the role of mitochondrial fission factor (MFF) in the development of NASH. Methods We created mice with hepatocyte-specific deletion of MFF (MffLiKO). MffLiKO mice fed normal chow diet (NCD) or high-fat diet (HFD) were evaluated for metabolic variables and their livers were examined by histological analysis. To elucidate the mechanism of development of NASH, we examined the expression of genes related to endoplasmic reticulum (ER) stress and lipid metabolism, and the secretion of triacylglycerol (TG) using the liver and primary hepatocytes isolated from MffLiKO and control mice. Results MffLiKO mice showed aberrant mitochondrial morphologies with no obvious NASH phenotypes during NCD, while they developed full-blown NASH phenotypes in response to HFD. Expression of genes related to ER stress was markedly upregulated in the liver from MffLiKO mice. In addition, expression of genes related to hepatic TG secretion was downregulated, with reduced hepatic TG secretion in MffLiKO mice in vivo and in primary cultures of MFF-deficient hepatocytes in vitro. Furthermore, thapsigargin-induced ER stress suppressed TG secretion in primary hepatocytes isolated from control mice. Conclusions/interpretation We demonstrated that ablation of MFF in liver provoked ER stress and reduced hepatic TG secretion in vivo and in vitro. Moreover, MffLiKO mice were more susceptible to HFD-induced NASH phenotype than control mice, partly because of ER stress-induced apoptosis of hepatocytes and suppression of TG secretion from hepatocytes. This study provides evidence for the role of mitochondrial fission in the development of NASH. Graphical abstract

scholarly journals In vitro study on the effect of cornin on the activity of cytochrome P450 enzymes

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Qun Zhang ◽  
Zengqiang Qu ◽  
Yanqing Zhou ◽  
Jin Zhou ◽  
Junwei Yang ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Drug Interaction ◽  
Liver Microsomes ◽  
In Vitro Study ◽  
Inhibitory Effect ◽  
Dependent Manner ◽  
Cytochrome P450 Enzymes ◽  
Drug Drug Interaction ◽  
Dose Dependent

Abstract Background Cornin is a commonly used herb in cardiology for its cardioprotective effect. The effect of herbs on the activity of cytochrome P450 enzymes (CYP450s) can induce adverse drug-drug interaction even treatment failure. Therefore, it is necessary to investigate the effect of cornin on the activity of CYP450s, which can provide more guidance for the clinical application of cornin. Methods Cornin (100 μM) was incubated with eight isoforms of CYP450s, including CYP1A2, 2A6, 3A4, 2C8, 2C9, 2C19, 2D6, and 2E1, in pooled human liver microsomes. The inhibition model and corresponding parameters were also investigated. Results Cornin exerted significant inhibitory effect on the activity of CYP3A4, 2C9, and 2E1 in a dose-dependent manner with the IC50 values of 9.20, 22.91, and 14.28 μM, respectively (p < 0.05). Cornin inhibited the activity of CYP3A4 non-competitively with the Ki value of 4.69 μM, while the inhibition of CYP2C9 and 2E1 by cornin was competitive with the Ki value of 11.31 and 6.54 μM, respectively. Additionally, the inhibition of CYP3A4 by cornin was found to be time-dependent with the KI/Kinact value of 6.40/0.055 min− 1·μM− 1. Conclusions The inhibitory effect of cornin on the activity of CYP3A4, 2C9, and 2E1 indicated the potential drug-drug interaction between cornin and drugs metabolized by these CYP450s, which needs further investigation and validation.

Thrombin activatable fibrinolysis inhibitor (TAFI) affects fibrinolysis in a plasminogen activator concentration-dependent manner

2004 ◽  
Vol 91 (03) ◽  
pp. 473-479 ◽  
Author(s):  
Ana Guimarães ◽  
Dingeman Rijken
Keyword(s):  
Plasminogen Activator ◽  
In Vitro Study ◽  
Inhibitory Effect ◽  
Retardation Factor ◽  
Clot Lysis ◽  
Dependent Manner ◽  
Plasma Clot ◽  
Concentration Dependent Manner ◽  
Plasmin Inhibitor

SummaryTAFIa was shown to attenuate fibrinolysis. In our in vitro study, we investigated how the inhibitory effect of TAFIa depended on the type and concentration of the plasminogen activator (PA). We measured PA-mediated lysis times of plasma clots under conditions of maximal TAFI activation by thrombin-thrombomodulin in the absence and presence of potato carboxypeptidase inhibitor. Seven different PAs were compared comprising both tPA-related (tPA, TNK-tPA, DSPA), bacterial PA-related (staphylokinase and APSAC) and urokinase-related (tcu-PA and k2tu-PA) PAs. The lysis times and the retardation factor were plotted against the PA concentration. The retardation factor plots were bell-shaped. At low PA concentrations, the retardation factor was low, probably due to the limited stability of TAFIa. At intermediate PA concentrations the retardation factor was maximal (3-6 depending on the PA), with TNK-tPA, APSAC and DSPA exhibiting the strongest effect. At high PA concentrations, the retardation factor was again low, possibly due to inactivation of TAFIa by plasmin or to a complete conversion of glu-plasminogen into lys-plasminogen. Using individual plasmas with a reduced plasmin inhibitor activity (plasmin inhibitor Enschede) the bell-shaped curve of the retardation factor shifted towards lower tPA and DSPA concentrations, but the height did not decrease. In conclusion, TAFIa delays the lysis of plasma clots mediated by all the plasminogen activators tested. This delay is dependent on the type and concentration of the plasminogen activator, but not on the fibrin specificity of the plasminogen activator. Furthermore, plasmin inhibitor does not play a significant role in the inhibition of plasma clot lysis by TAFI.

scholarly journals Anti-Allergic Potential of Cinnamaldehyde via the Inhibitory Effect of Histidine Decarboxylase (HDC) Producing Klebsiella pneumonia

Molecules ◽  
2020 ◽  
Vol 25 (23) ◽  
pp. 5580
Author(s):  
Lorina I. Badger-Emeka ◽  
Promise Madu Emeka ◽  
Krishnaraj Thirugnanasambantham ◽  
Hairul Islam M. Ibrahim
Keyword(s):  
Mast Cell ◽  
Cell Activation ◽  
Histidine Decarboxylase ◽  
Cell Model ◽  
In Vitro Study ◽  
Inhibitory Effect ◽  
Mast Cell Activation ◽  
Klebsiella Pneumonia ◽  
Immunological Disorder

Allergy is an immunological disorder that develops in response to exposure to an allergen, and histamines mediate these effects via histidine decarboxylase (HDC) activity at the intracellular level. In the present study, we developed a 3D model of Klebsiella pneumoniae histidine decarboxylase (HDC) and analyzed the HDC inhibitory potential of cinnamaldehyde (CA) and subsequent anti-allergic potential using a bacterial and mammalian mast cell model. A computational and in vitro study using K. pneumonia revealed that CA binds to HDC nearby the pyridoxal-5′-phosphate (PLP) binding site and inhibited histamine synthesis in a bacterial model. Further study using a mammalian mast cell model also showed that CA decreased the levels of histamine in the stimulated RBL-2H3 cell line and attenuated the release of β-hexoseaminidase and cell degranulation. In addition, CA treatment also significantly suppressed the levels of pro-inflammatory cytokines TNF-α and IL-6 and the nitric oxide (NO) level in the stimulated mast cells. A gene expression and Western blotting study revealed that CA significantly downregulated the expressions of MAPKp38/ERK and its downstream pro-allergic mediators that are involved in the signaling pathway in mast cell cytokine synthesis. This study further confirms that CA has the potential to attenuate mast cell activation by inhibiting HDC and modifying the process of allergic disorders.

Demineralization Inhibitory Effects of Highly Concentrated Fluoride Dentifrice and Fluoride Gels/Solutions on Sound Dentin and Artificial Dentin Caries Lesions in vitro

2020 ◽  
pp. 1-14
Author(s):  
Daniel Erdwey ◽  
Hendrik Meyer-Lueckel ◽  
Marcella Esteves-Oliveira ◽  
Christian Apel ◽  
Richard Johannes Wierichs
Keyword(s):  
In Vitro Study ◽  
Inhibitory Effect ◽  
Negative Effects ◽  
Dentin Caries ◽  
Before And After ◽  
Amine Fluoride ◽  
Ph Cycling ◽  
Fluoride Dentifrice ◽  
Caries Lesions

<b><i>Objectives:</i></b> The aim of this in vitro study was to compare the demineralization inhibitory effect of gels/solutions used in combination with either standard or highly fluoridated dentifrices on sound dentin as well as on artificial dentin caries-like lesions. <b><i>Methods:</i></b> Bovine dentin specimens (<i>n</i> = 240) with two different surfaces each (sound [ST] and artificial caries lesion [DT]) were prepared and randomly allocated to twelve groups. Weekly interventions during pH-cycling (28 days, 6 × 120 min demineralization/day) were: the application of gels/solutions containing amine fluoride/sodium fluoride (12,500 ppm F [ppm]; pH = 4.4; AmF); NaF (12,500 ppm; pH = 6.6; NaF1); NaF (12,500 ppm; pH = 6.3; NaF2); silver diamine fluoride (14,200 ppm; pH = 8.7; SDF); acidulated phosphate fluoride (12,500 ppm; pH = 3.8; APF), and no intervention (standard control; S). Furthermore, half of the specimens in each group were brushed (10 s; twice per day) with dentifrice slurries containing either 1,450 ppm (e.g., AmF<sub>1450</sub>) or 5,000 ppm (e.g., AmF<sub>5000</sub>). Differences in integrated mineral loss (ΔΔZ) and lesion depth (ΔLD) were calculated between values before and after pH-cycling using transversal microradiography. <b><i>Results:</i></b> After pH-cycling Ss showed significantly increased ΔZ<sub>DT</sub> and LD<sub>DT</sub> values, indicating further demineralization. In contrast, except for one, all groups including fluoride gels/solutions showed significantly decreased ΔZ<sub>DT</sub> values. Additional use of most fluoride gels/solutions significantly enhanced mineral gain, mainly in the surface area; however, acidic gels/solutions seemed to have negative effects on lesion depths. <b><i>Significance:</i></b> Under the present pH-cycling conditions the highly fluoridated dentifrice significantly reduced caries progression and additional application of nearly all of the fluoride gels/solutions resulted in remineralization. However, there was no difference in the remineralizing capacity of fluoride gels/solutions when used in combination with either standard or highly fluoridated dentifrices.

scholarly journals Inhibitory effect of Salvadora persica extract (Miswak) on collagen degradation in demineralized dentin: In vitro study

2021 ◽  
Vol 16 (1) ◽  
pp. 208-213
Author(s):  
Sahar Khunkar ◽  
Ilnaz Hariri ◽  
Ehab Alsayed ◽  
Amal Linjawi ◽  
Sawsan Khunkar ◽  
...  
Keyword(s):  
In Vitro Study ◽  
Inhibitory Effect ◽  
Collagen Degradation ◽  
Vitro Study ◽  
Salvadora Persica ◽  
Demineralized Dentin

scholarly journals Tigecycline Interferes with Fibrinogen Polymerization Independent of Peripheral Interactions with the Coagulation System

Antibiotics ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 84 ◽  
Author(s):  
Anna Brandtner ◽  
Mirjam Bachler ◽  
Dietmar Fries ◽  
Martin Hermann ◽  
Jacqueline Ruehlicke ◽  
...  
Keyword(s):  
Qualitative Difference ◽  
In Vitro Study ◽  
Inhibitory Effect ◽  
Coagulation System ◽  
Mitochondrial Activity ◽  
Hepatic Cells ◽  
Spiked Plasma ◽  
Fibrin Network ◽  
Coagulation Tests

Tigecycline offers broad anti-bacterial coverage for critically ill patients with complicated infections. A described but less researched side effect is coagulopathy. The aim of this study was to test whether tigecycline interferes with fibrinogen polymerization by peripheral interactions. To study the effect of unmetabolized tigecycline, plasma of healthy volunteers were spiked with increasing concentrations of tigecycline. In a second experimental leg, immortalized human liver cells (HepG2) were treated with the same concentrations to test an inhibitory effect of hepatic tigecycline metabolites. Using standard coagulation tests, only the activated thromboplastin time in humane plasma was prolonged with increasing concentrations of tigecycline. Visualization of the fibrin network using confocal live microscopy demonstrated a qualitative difference in tigecycline treated experiments. Thrombelastometry and standard coagulation tests did not indicate an impairment of coagulation. Although the discrepancy between functional and immunologic fibrinogen levels increased in cell culture assays with tigecycline concentration, fibrinogen levels in spiked plasma samples did not show significant differences determined by functional versus immunologic methods. In our in vitro study, we excluded a direct effect of tigecycline in increasing concentrations on blood coagulation in healthy adults. Furthermore, we demonstrated a rapid loss of mitochondrial activity in hepatic cells with supra-therapeutic tigecycline dosages.

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