Ischemic stroke causes severe neuronal damage. Chlorogenic acid is a phenolic substance present in fruits and coffee. It also exerts neuroprotective effects against various brain injuries. The 14-3-3 family protein perform a variety of functions including metabolism, signal transduction, cell differentiation, and apoptosis. The purpose of this study is to investigate whether chlorogenic acid regulates the expression of 14-3-3 protein in stroke animal models. Ischemic stroke was induced by middle cerebral artery occlusion (MCAO) surgery. Phosphate buffered saline (PBS) or chlorogenic acid (30 mg/kg) were intraperitoneally injected to adult male rats 2 h before MCAO surgery. Adhesive-removal test was performed 24 h after MCAO surgery and cerebral cortical tissues were collected for further study. MCAO damage caused severe neurological impairment and chlorogenic acid treatment ameliorated this disorder. Our proteomic approach showed a decrease in 14-3-3 expression in MCAO animals with PBS. The decrease in 14-3-3 expression alleviated in MCAO animal with chlorogenic acid. We confirmed changes in various 14-3-3 protein isoforms, including beta/alpha, zeta/delta, gamma, epsilon, eta, and tau through reverse transcription-PCR. These results explained that chlorogenic acid regulates the expression of 14-3-3 protein in MCAO-induced cerebral ischemia. 14-3-3 is considered to be an important protein for cell survival through binding to pro-apoptotic proteins. The maintenance of 14-3-3 levels is an important event in neuroprotection against ischemic injury. Therefore, we can demonstrate that the 14-3-3 protein contributes to the neuroprotective effect of chlorogenic acid in stroke animal models.

INTRODUCTION
MATERIALS AND METHODS
    Experimental animal and drug treatment
    Middle cerebral artery occlusion (MCAO)
    Adhesive removal test
    Two-dimensional gel electrophoresis
    Reverse transcription-polymerase chain reaction
    Western blot analysis
    Statistical analysis
RESULTS
DISCUSSION
Acknowledgments

• Murad-Ali Shah(Department of Veterinary Anatomy, College of Veterinary Medicine, Research Institute of Life Science)
• Yeung Bae Jin(Department of Laboratory Animal Medicine, College of Veterinary Medicine)
• Myeong-Ok Kim(Division of Life Science and Applied Life Science, College of Natural Sciences, Gyeongsang National University, Jinju 52828, Korea)
• Phil-Ok Koh(Department of Veterinary Anatomy, College of Veterinary Medicine, Research Institute of Life Science) Corresponding Author