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Interferon regulatory factor-1 together with reactive oxygen species promotes the acceleration of cell cycle progression by up-regulating the cyclin E and CDK2 genes during high glucose-induced proliferation of vascular smooth muscle cells.

Zhang X, Liu L, Chen C, Chi YL, Yang XQ, Xu Y, Li XT, Guo SL, Xiong SH, Shen MR, Sun Y, Zhang CS, Hu KM - Cardiovasc Diabetol (2013)

Bottom Line: The levels of cyclin/CDK expression in two cell models involving Irf-1 knockdown and overexpression were quantified to explore the relationship between Irf-1 and its downstream effectors under normal or high glucose conditions.We found that Irf-1 overexpression led to down-regulation of cyclin D1/CDK4 and inhibited cell cycle progression in VSMCs under normal glucose conditions.These results demonstrate that the downstream effectors of Irf-1 are cyclin E/CDK2 during the high glucose-induced proliferation of VSMCs, whereas they are cyclin D1/CDK4 in normal glucose conditions.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Biomedical Engineering, Second Military Medical University, Shanghai, China. zhangxicyl1126@163.com.

ABSTRACT

Background: The high glucose-induced proliferation of vascular smooth muscle cells (VSMCs) plays an important role in the development of diabetic vascular diseases. In a previous study, we confirmed that Interferon regulatory factor-1 (Irf-1) is a positive regulator of the high glucose-induced proliferation of VSMCs. However, the mechanisms remain to be determined.

Methods: The levels of cyclin/CDK expression in two cell models involving Irf-1 knockdown and overexpression were quantified to explore the relationship between Irf-1 and its downstream effectors under normal or high glucose conditions. Subsequently, cells were treated with high glucose/NAC, normal glucose/H₂O₂, high glucose/U0126 or normal glucose/H₂O₂/U0126 during an incubation period. Then proliferation, cyclin/CDK expression and cell cycle distribution assays were performed to determine whether ROS/Erk1/2 signaling pathway was involved in the Irf-1-induced regulation of VSMC growth under high glucose conditions.

Results: We found that Irf-1 overexpression led to down-regulation of cyclin D1/CDK4 and inhibited cell cycle progression in VSMCs under normal glucose conditions. In high glucose conditions, Irf-1 overexpression led to an up-regulation of cyclin E/CDK2 and an acceleration of cell cycle progression, whereas silencing of Irf-1 suppressed the expression of both proteins and inhibited the cell cycle during the high glucose-induced proliferation of VSMCs. Treatment of VSMCs with antioxidants prevented the Irf-1 overexpression-induced proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression in high glucose conditions. In contrast, under normal glucose conditions, H₂O₂ stimulation and Irf-1 overexpression induced cell proliferation, up-regulated cyclin E/CDK2 expression and promoted cell cycle acceleration. In addition, overexpression of Irf-1 promoted the activation of Erk1/2 and when VSMCs overexpressing Irf-1 were treated with U0126, the specific Erk1/2 inhibitor abolished the proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression under high glucose or normal glucose/H₂O₂ conditions.

Conclusions: These results demonstrate that the downstream effectors of Irf-1 are cyclin E/CDK2 during the high glucose-induced proliferation of VSMCs, whereas they are cyclin D1/CDK4 in normal glucose conditions. The Irf-1 overexpression-induced proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression are associated with ROS/Erk1/2 signaling pathway under high glucose conditions.

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The intracellular ROS production was measured by flow cytometry analysis under high glucose or normal glucose conditions. Values are showed as relative fold of fluorescence intensity measurement (High glucose/2 days =100). *P <0.05; **P <0.01 versus corresponding values in VSMCs under normal glucose conditions. 2 days, 4 days and 6 days = VSMCs incubated with high glucose or normal glucose for 2 days, 4 days and 6 days.
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Figure 3: The intracellular ROS production was measured by flow cytometry analysis under high glucose or normal glucose conditions. Values are showed as relative fold of fluorescence intensity measurement (High glucose/2 days =100). *P <0.05; **P <0.01 versus corresponding values in VSMCs under normal glucose conditions. 2 days, 4 days and 6 days = VSMCs incubated with high glucose or normal glucose for 2 days, 4 days and 6 days.

Mentions: ROS production in VSMCs was measured by flow cytometric analysis. The results showed that the level of intracellular ROS under high glucose conditions was higher than that under normal glucose conditions (P < 0.01, n = 12; Figure 3 and Table 2), which indicates that high glucose treatment increased significantly the level of intracellular ROS in VSMCs.


Interferon regulatory factor-1 together with reactive oxygen species promotes the acceleration of cell cycle progression by up-regulating the cyclin E and CDK2 genes during high glucose-induced proliferation of vascular smooth muscle cells.

Zhang X, Liu L, Chen C, Chi YL, Yang XQ, Xu Y, Li XT, Guo SL, Xiong SH, Shen MR, Sun Y, Zhang CS, Hu KM - Cardiovasc Diabetol (2013)

The intracellular ROS production was measured by flow cytometry analysis under high glucose or normal glucose conditions. Values are showed as relative fold of fluorescence intensity measurement (High glucose/2 days =100). *P <0.05; **P <0.01 versus corresponding values in VSMCs under normal glucose conditions. 2 days, 4 days and 6 days = VSMCs incubated with high glucose or normal glucose for 2 days, 4 days and 6 days.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3852693&req=5

Figure 3: The intracellular ROS production was measured by flow cytometry analysis under high glucose or normal glucose conditions. Values are showed as relative fold of fluorescence intensity measurement (High glucose/2 days =100). *P <0.05; **P <0.01 versus corresponding values in VSMCs under normal glucose conditions. 2 days, 4 days and 6 days = VSMCs incubated with high glucose or normal glucose for 2 days, 4 days and 6 days.
Mentions: ROS production in VSMCs was measured by flow cytometric analysis. The results showed that the level of intracellular ROS under high glucose conditions was higher than that under normal glucose conditions (P < 0.01, n = 12; Figure 3 and Table 2), which indicates that high glucose treatment increased significantly the level of intracellular ROS in VSMCs.

Bottom Line: The levels of cyclin/CDK expression in two cell models involving Irf-1 knockdown and overexpression were quantified to explore the relationship between Irf-1 and its downstream effectors under normal or high glucose conditions.We found that Irf-1 overexpression led to down-regulation of cyclin D1/CDK4 and inhibited cell cycle progression in VSMCs under normal glucose conditions.These results demonstrate that the downstream effectors of Irf-1 are cyclin E/CDK2 during the high glucose-induced proliferation of VSMCs, whereas they are cyclin D1/CDK4 in normal glucose conditions.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Biomedical Engineering, Second Military Medical University, Shanghai, China. zhangxicyl1126@163.com.

ABSTRACT

Background: The high glucose-induced proliferation of vascular smooth muscle cells (VSMCs) plays an important role in the development of diabetic vascular diseases. In a previous study, we confirmed that Interferon regulatory factor-1 (Irf-1) is a positive regulator of the high glucose-induced proliferation of VSMCs. However, the mechanisms remain to be determined.

Methods: The levels of cyclin/CDK expression in two cell models involving Irf-1 knockdown and overexpression were quantified to explore the relationship between Irf-1 and its downstream effectors under normal or high glucose conditions. Subsequently, cells were treated with high glucose/NAC, normal glucose/H₂O₂, high glucose/U0126 or normal glucose/H₂O₂/U0126 during an incubation period. Then proliferation, cyclin/CDK expression and cell cycle distribution assays were performed to determine whether ROS/Erk1/2 signaling pathway was involved in the Irf-1-induced regulation of VSMC growth under high glucose conditions.

Results: We found that Irf-1 overexpression led to down-regulation of cyclin D1/CDK4 and inhibited cell cycle progression in VSMCs under normal glucose conditions. In high glucose conditions, Irf-1 overexpression led to an up-regulation of cyclin E/CDK2 and an acceleration of cell cycle progression, whereas silencing of Irf-1 suppressed the expression of both proteins and inhibited the cell cycle during the high glucose-induced proliferation of VSMCs. Treatment of VSMCs with antioxidants prevented the Irf-1 overexpression-induced proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression in high glucose conditions. In contrast, under normal glucose conditions, H₂O₂ stimulation and Irf-1 overexpression induced cell proliferation, up-regulated cyclin E/CDK2 expression and promoted cell cycle acceleration. In addition, overexpression of Irf-1 promoted the activation of Erk1/2 and when VSMCs overexpressing Irf-1 were treated with U0126, the specific Erk1/2 inhibitor abolished the proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression under high glucose or normal glucose/H₂O₂ conditions.

Conclusions: These results demonstrate that the downstream effectors of Irf-1 are cyclin E/CDK2 during the high glucose-induced proliferation of VSMCs, whereas they are cyclin D1/CDK4 in normal glucose conditions. The Irf-1 overexpression-induced proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression are associated with ROS/Erk1/2 signaling pathway under high glucose conditions.

Show MeSH
Related in: MedlinePlus