Effects of Gualou Guizhi Decoction Aqueous Extract on Axonal Regeneration in Organotypic Cortical Slice Culture after Oxygen-Glucose Deprivation.

Lihong Nan,Lan Yang,Mei Huang,Huang Li,Yanfang Zheng,Yibo He,Qingqing Xie,Zheming Chen

doi:10.1155/2017/5170538

Abstract

Gualou Guizhi decoction (GLGZD) is effective for the clinical treatment of limb spasms caused by ischemic stroke, but its underlying mechanism is unclear. Propidium iodide (PI) fluorescence staining, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), immunohistochemistry, western blot, and real-time qPCR were used to observe the axonal regeneration and neuroprotective effects of GLGZD aqueous extract on organotypic cortical slices exposed to oxygen-glucose deprivation (OGD) and further elucidate the potential mechanisms. Compared with the OGD group, the GLGZD aqueous extract decreased the red PI fluorescence intensity; inhibited neuronal apoptosis; improved the growth of slice axons; upregulated the protein expression of tau and growth-associated protein-43; and decreased protein and mRNA expression of neurite outgrowth inhibitor protein-A (Nogo-A), Nogo receptor 1 (NgR1), ras homolog gene family A (RhoA), rho-associated coiled-coil-containing protein kinase (ROCK), and phosphorylation of collapsin response mediator protein 2 (CRMP2). Our study found that GLGZD had a strong neuroprotective effect on brain slices after OGD injury. GLGZD plays a vital role in promoting axonal remodeling and functional remodeling, which may be related to regulation of the expression of Nogo-A and its receptor NgR1, near the injured axons, inhibition of the Rho-ROCK pathway, and reduction of CRMP2 phosphorylation.

Highlights

Stroke, an acute cerebrovascular condition, is the second most common cause of disease-related death
Organotypic brain slice culture, established by Stoppini et al [16], is a promising technique to avoid the effects of the blood-brain barrier, ion concentration, temperature, pH, and other individual differences and to maintain the structure of the cell and the complete neural circuits to simulate the physiological environment in vivo
Our findings suggest that the Rho-rho-associated coiled-coil-containing protein kinase (ROCK) pathway was activated by neurite outgrowth inhibitor protein-A (Nogo-A) after oxygenglucose deprivation (OGD), thereby decreasing microtubule stability and inhibiting axonal regeneration in the brain issue

Summary

Introduction

An acute cerebrovascular condition, is the second most common cause of disease-related death. Limb dysfunction often occurs after stroke and is the most common postapoplectic sequela associated with the upper motor neuron injury caused by various factors during the ischemic process. A previous study showed that reduction of fat molecules, separation of the axonal medullary sheath, and edema in axons were eventually observed in the injured neurons [1]. Various types of receptors exist on the surface of axonal growth cones. These receptors are highly sensitive to the surrounding microenvironment and identify specific signal factors in the extracellular matrix or other cells, affecting the orientation of axonal growth. The inhibition of the growth-inhibiting factors of the medullary sheath facilitates axonal regeneration and neural reorganization [2]. Regulation of the axonal microenvironment is critical to axonal regeneration [3]

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