Elsevier

Neuroscience

Volume 240, 14 June 2013, Pages 54-62
Neuroscience

Phosphatidylinositol 3 kinase/protein kinase B is responsible for the protection of paeoniflorin upon H2O2-induced neural progenitor cell injury

https://doi.org/10.1016/j.neuroscience.2013.02.037Get rights and content

Abstract

Promoting neural stem/progenitor cell (NSC/NPC) survival in the pro-apoptotic environment is critical to stem cell replacement for neurodegenerative disease therapy. Paeoniflorin (PF), one of the principal bioactive components in Paeoniae Radix, has been used widely in central nervous system (CNS) diseases treatment and serves as an antioxidant to protect neurons against oxidative stress. The present study investigated the protective effects of PF on NPC injury induced by hydrogen peroxide (H2O2). After challenge with 200 μM H2O2 for 2 h, loss of cell viability and excessive apoptotic cell death were observed in cultured NPC, PF treatment conferred protective effects against the loss of cellular viability in a concentration-dependent manner. PF pretreatment also inhibited NPC apoptosis induced by H2O2 by reversing the decreased level of Procaspase-3 and balancing Bcl-2 and Bax expression. Furthermore, PF-mediated NPC protection was associated with an increase in phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt-1) phosphorylation in a time- and concentration-dependent manner. Selective inhibition of PI3K using LY294002 abolished PF-mediated phosphorylation of Akt-1 and NPC protection upon oxidative stress. These data suggest that PF-mediated NPC protection on H2O2 injury is reliant on the activation of the PI3K/Akt-1 pathway, giving insight to an essential role of PF in NPC protection.

Highlights

Paeoniflorin confers protective effects to NPCs against oxidative stress-induced apoptosis. ► PF inhibited H2O2-induced NPC apoptosis by increasing Procaspase-3 and balancing Bcl-2/Bax expression. ► Paeoniflorin-mediated protection of NPCs is reliant upon the PI3K/Akt-1 signaling pathway.

Introduction

There is growing evidence that neural stem/progenitor cell (NSC/NPC) exist throughout life and these progenitors have the ability to produce new neurons and glia in the adult central nervous system (CNS). Adult progenitors are known to be abundant in the subventricular zone (SVZ), subgranular zone (SGZ) and olfactory bulb (OB) (Palmer et al., 1999, Ming and Song, 2011) and give birth to new neurons which are critical for learning and memory. Impaired neurogenesis has been evidenced in neural degenerative diseases. Decreased number of NPCs was found and altered division rates happened in Alzheimer’s disease (AD) patients (Ziabreva et al., 2006). Postmortem brains of Parkinson’s disease (PD) patients also showed the reduced numbers of NPC in the subependymal zone and SGZ (Hoglinger et al., 2004). Oxidative stress plays an important role in neurodegenerative disorders including AD (Akiyama et al., 2000, Tamagno et al., 2012), and is very likely involved in the cascade of events leading to neural cell death in PD patients (Hunot et al., 1996). Therefore, it is very important to promote the survival of NSC/NPC on oxidative injury for the treatment of degenerative diseases by neural transplantation.

Paeoniflorin (PF) is one of the principal bioactive components derived from the root of P. lactiflora Palls/Paeoniae Radix, a traditional Chinese herbal which has been widely used in the treatment of CNS diseases including cognitive disorder (Ohta et al., 1993, Ohta et al., 1994, Watanabe, 1997), Parkinson’s disease (Liu et al., 2007), ischemia (Liu et al., 2005, Cao et al., 2006) and Alzheimer’s disease (Zhong et al., 2009). It has been reported PF has various pharmacological activities including antioxidant, anti-inflammation and neuroprotective effects on various types of cells (Liu et al., 2006a, Zhang et al., 2008). As a natural antioxidant, PF penetrates through the blood–brain barrier (Cao et al., 2006) and the neuroprotective effects of PF have been evidenced both in vivo and in vitro (Tabata et al., 2001). PF significantly reduced the infarct volume and ameliorated the deficits in cognition and neurological symptoms caused by transient MCAO (Xiao et al., 2005). Further study showed that PF attenuated cognitive deficits and neuron damage caused by chronic cerebral hypoperfusion in rats (Liu et al., 2006b). However, whether PF protects NPC from oxidative-induced apoptosis and related signaling pathways involved in this process is still unclear. In the present study, we used cultured NPC to characterize hydrogen peroxide (H2O2)-induced NPC apoptosis and related cell signaling pathways. Our data showed that PF protected NPC from oxidative stress-induced apoptosis by activating PI3K/Akt-1, upregulating Procaspase-3, as well as regulating the Bcl-2 family, including Bcl-2 and Bax expression.

Section snippets

Reagents

Dulbecco’s Modified Eagle’s Medium/F12 (DMEM/F12) was purchased from Hyclone (Logan, UT, USA). Basic fibroblast growth factor (bFGF), epidermal growth factor (EGF) and B27 supplement were obtained from Invitrogen (Carlsbad, CA, USA). LY294002 was obtained from Calbiochem (San Diego, CA, USA) and anti-nestin antibody was purchased from Millipore/Chemicon (Billerica, MA, USA). Anti-phospho-Akt-1, anti-Akt-1 and anti-phospho-PI3K antibodies were purchased from Cell Signaling (Danvers, MA, USA).

PF increased NPC viability upon H2O2 injury

Oxidative stress caused by the cellular accumulation of reactive oxygen species (ROS) is a major contributor to diseases and cell death. The production of intracellular ROS caused by exposure to external oxidant is destructive to normal cells and plays critical roles in diverse cellular processes from tumorigenesis to aging (Finkel, 2003). H2O2 has been used as a well-established model to investigate the mechanisms of oxidative stress in NPCs by many investigators because of its rapid membrane

Discussion

In the present study, we revealed that PF, one of the principal bioactive components in Paeoniae Radix, attenuated mouse NPC apoptosis induced by H2O2 and the neuroprotective effects of PF might be due to the activation of PI3K/Akt-1. Furthermore, PF treatment inhibited NPC apoptosis induced by H2O2 by reversing the decreased level of Procaspase-3 and balancing Bcl-2 and Bax expression. These results provided evidence that PF may play an important role in NPC protection.

Neuroregeneration is a

Conclusion

In summary, the present study showed that PF could protect mouse NPC against H2O2-induced oxidative stress, as measured by cell viability, apoptosis, Procaspase-3 level, and the PI3K/Akt-1 signaling pathway. Notably, the inhibitors of PI3K, LY294002 abolished PF-mediated NPC survival upon H2O2 stress, indicating the important roles of Akt-1 in the regulation of NPC survival. As a whole, our data show that PI3K/Akt-1 signaling pathway is responsible for PF-mediated NPC protection. Therefore, PF

Conflict of interest

The authors declare that they have no competing interests.

Acknowledgments

This work was supported in part by research grants from the National Natural Science Foundation of China, No. 81101457, Traditional Chinese Medicine Administration of Shaanxi Province Grant No. JC045 and Scientific Research Foundation for the Returned Overseas Chinese Scholars No. HG3402.

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    These authors contributed equally to this study.

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