Methotrexate Promotes Platelet Apoptosis via JNK-Mediated Mitochondrial Damage: Alleviation by N-Acetylcysteine and N-Acetylcysteine Amide

Paul, Manoj; Hemshekhar, Mahadevappa; Thushara, R. M.; Sundaram, Mahalingam S.; NaveenKumar, Somanathapura K.; Naveen, S.; Sannaningaiah, Devaraja; Somyajit, Kumar; West, Robert; Basappa, Basappa; Nayaka, S. Chandra; Zakai, Uzma I.; Nagaraju, Ganesh; Rangappa, Kanchugarakoppal S.; Kemparaju, K.; Girish, K. S.

doi:10.1371/journal.pone.0127558

Cited by 62 publications

(43 citation statements)

References 53 publications

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“…Moreover, isoliensinine-induced apoptosis in human breast cancer cells was mediated by p38 MAPK and JNK pathways [40]. The level of p-JNK increased in methotrexate (MTX)-treated platelet apoptosis, and it activated Bad and Bax and inhibited Bcl-2, thus, contributing toward mitochondrial dysfunction [41]. Serum-free media could increase the H 2 O 2 production and activate JNK in MLO-Y4 osteocyte-like cells, resulting in cell apoptosis [42].…”

Section: Discussionmentioning

confidence: 99%

Oxidative Stress Induces Mouse Follicular Granulosa Cells Apoptosis via JNK/FoxO1 Pathway

Weng

Liu

et al. 2016

PLoS ONE

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The c-Jun N-terminal protein kinase (JNK) plays an important role in the regulation of cell apoptosis. Forkhead box O (FoxO) transcription factors are involved in diverse biological processes, including cellular metabolism, cell apoptosis, and cell cycle. However, the JNK/FoxO1 pathway involved in the process of apoptosis induced by oxidative stress remains to be elucidated. Here, we demonstrated that the JNK activity significantly increased in response to oxidative stress in mouse follicular granulosa cells (MGCs). SP600125, a selective JNK inhibitor, attenuated the oxidative stress-induced MGCs apoptosis. Oxidative stress enhanced the FoxO1 nuclear translocation by activating the JNK activity. Moreover, JNK mediated the dissociation of FoxO1 from 14-3-3 proteins in MGCs after the treatment with H2O2. Finally, oxidative stress up-regulated the expression of FoxO1 via JNK mediation of FoxO1 self-regulation in MGCs. Taken together, our findings suggest that JNK/FoxO1 is involved in the regulation of oxidative stress-induced cell apoptosis in MGCs.

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Section: Discussionmentioning

confidence: 99%

Oxidative Stress Induces Mouse Follicular Granulosa Cells Apoptosis via JNK/FoxO1 Pathway

Weng

Liu

et al. 2016

PLoS ONE

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“…The suspended platelets in each group were pre-treated with a 1-mM dose of the ROS scavenger N-acetylcysteine (NAC) (Sigma-Aldrich, USA) [28], a 0.2-μM dose of the mTOR inhibitor Rapamycin (Sigma-Aldrich, USA) [16] or a 1-mM dose of the autophagy inhibitor 3-MA (Sigma-Aldrich, USA) [16] for 30 min, and then 50 μg/ml ox-LDL was added to platelets and incubated for 10 min.…”

Section: Grouping and Treatment Of Washed Plateletsmentioning

confidence: 99%

ROS Promote Ox-LDL-Induced Platelet Activation by Up-Regulating Autophagy Through the Inhibition of the PI3K/AKT/mTOR Pathway

Wang

Liu

et al. 2018

Cell Physiol Biochem

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Background/Aims: Oxidized low-density lipoprotein (oxLDL) promotes unregulated platelet activation in patients with dyslipidemic disorders. Although oxLDL stimulates activating signaling, researchers have not clearly determined how these events drive accelerated thrombosis. Here, we describe the mechanism by which ROS regulate autophagy during ox-LDL-induced platelet activation by modulating the PI3K/AKT/mTOR signaling pathway. Methods: For in vitro experiments, ox-LDL, the ROS scavenger N-acetylcysteine (NAC), the mTOR inhibitor rapamycin and the autophagy inhibitor 3-MA were used alone or in combination with other compounds to treat platelets. Then, platelet aggregation was evaluated on an aggregometer and platelet adhesion was measured under shear stress. The levels of a platelet activation marker (CD62p) were measured by flow cytometry, reactive oxygen species (ROS) levels were then quantified by measuring DCFH-DA fluorescence intensity via flow cytometry. Nitric oxide (NO) and superoxide (O₂^·-) levels were determined by the nitric acid deoxidize enzyme method and lucigenin-enhanced chemiluminescence (CL), respectively. Transmission electron microscopy was used to observe the autophagosome formation, immunofluorescence staining was employed to detect LC3 expression and western blotting was used to measure the levels of PI3K/AKT/mTOR pathway- and autophagy-related proteins. Results: Ox-LDL-induced platelets showed a significant increase in platelet aggregation and adhesion, CD62p expression, ROS level and O₂^·- content, with an elevated LC3II/LC3I ratio and Beclin1 expression, but a dramatic reduction in the levels of p62 and pathway-related proteins (all P < 0.05). However, platelet activation and autophagy were aggravated by the Rapamycin treatment, and decreased following treatment with NAC, 3-MA, or NAC and 3-MA, together with increased activity of the PI3K/AKT/mTOR pathway. Additionally, decreased platelet activation and autophagy were observed in platelets treated with NAC and Rapamycin or Rapamycin and 3-MA compared with platelets treated with Rapamycin alone, suggesting that both NAC and 3-MA reversed the effects of Rapamycin. Conclusion: Inhibition of ROS production may reduce autophagy to suppress ox-LDL-induced platelet activation by activating PI3K/AKT/mTOR pathway.

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“…Recently emerging evidence confirmed NACA as a protective agent against oxidative stress in vitro and in vivo [13–16]. Our previous study also indicated NACA could protect renal tubular epithelial cell against contrast-induced apoptosis in vitro [6].…”

Section: Introductionmentioning

confidence: 78%

Nephroprotective Effects of N‐Acetylcysteine Amide against Contrast‐Induced Nephropathy through Upregulating Thioredoxin‐1, Inhibiting ASK1/p38MAPK Pathway, and Suppressing Oxidative Stress and Apoptosis in Rats

Gong

Duan

Zheng

et al. 2016

Oxidative Medicine and Cellular Longevity

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Contrast-induced nephropathy (CIN) is a leading cause of hospital-acquired acute kidney injury (AKI) due to apoptosis induced in renal tubular cells. Our previous study demonstrated the novel N-acetylcysteine amide (NACA); the amide form of N-acetyl cysteine (NAC) prevented renal tubular cells from contrast-induced apoptosis through inhibiting p38 MAPK pathway in vitro. In the present study, we aimed to compare the efficacies of NACA and NAC in preventing CIN in a well-established rat model and investigate whether thioredoxin-1 (Trx1) and apoptosis signal-regulating kinase 1 (ASK1) act as the potential activator for p38 MAPK. NACA significantly attenuated elevations of serum creatinine, blood urea nitrogen, and biomarkers of AKI. At equimolar concentration, NACA was more effective than NAC in reducing histological changes of renal tubular injuries. NACA attenuated activation of p38 MAPK signal, reduced oxidative stress, and diminished apoptosis. Furthermore, we demonstrated that contrast exposure resulted in Trx1 downregulation and increased ASK1/p38 MAPK phosphorylation, which could be reversed by NACA and NAC. To our knowledge, this is the first report that Trx1 and ASK1 are involved in CIN. Our study highlights a renal protective role of NACA against CIN through modulating Trx1 and ASK1/p38 MAPK pathway to result in the inhibition of apoptosis among renal cells.

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Methotrexate Promotes Platelet Apoptosis via JNK-Mediated Mitochondrial Damage: Alleviation by N-Acetylcysteine and N-Acetylcysteine Amide

Cited by 62 publications

References 53 publications

Oxidative Stress Induces Mouse Follicular Granulosa Cells Apoptosis via JNK/FoxO1 Pathway

Oxidative Stress Induces Mouse Follicular Granulosa Cells Apoptosis via JNK/FoxO1 Pathway

ROS Promote Ox-LDL-Induced Platelet Activation by Up-Regulating Autophagy Through the Inhibition of the PI3K/AKT/mTOR Pathway

Nephroprotective Effects of N‐Acetylcysteine Amide against Contrast‐Induced Nephropathy through Upregulating Thioredoxin‐1, Inhibiting ASK1/p38MAPK Pathway, and Suppressing Oxidative Stress and Apoptosis in Rats

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