Abstract
Axonal damage is recognized as an important pathological feature in the chronic progressive neurological disorder multiple sclerosis (MS). Promoting axonal regeneration is a critical strategy for the treatment of MS. Our clinical and experimental studies have shown that the Bu Shen Yi Sui formula (BSYS) promotes axonal regeneration in MS and experimental autoimmune encephalomyelitis (EAE), an animal model of MS, but the exact mechanism has not been thoroughly elucidated to date. In this study, we investigated the effects of BSYS and its two decomposed formulas—the Bu Shen formula (BS) and the Hua Tan Huo Xue formula (HTHX)—on brain-derived neurotrophic factor (BDNF)/TrkB and related signaling pathways to explore the mechanism by which axonal regeneration is promoted in vitro and in vivo. Damaged SH-SY5Y cells incubated with low serum were treated with BSYS-, BS-, and HTHX-containing serum, and EAE mice induced by the myelin oligodendrocyte glycoprotein (MOG)35–55 peptide were treated with BSYS. The results showed that the BSYS-containing serum markedly increased cell viability and increased the levels of growth associated protein (GAP)-43, phosphorylated (p)-cAMP-response element binding protein (CREB), BDNF, TrkB, and p-PI3K. The BS and HTHX treatments also induced the protein expression of GAP-43 and p-extracellular signal-regulated kinase (ERK) in the cells. Furthermore, the effects of BSYS on cell viability, GAP-43, p-CREB, and neurite outgrowth were clearly inhibited by LY294002, a specific antagonist of the PI3K signaling pathways. The addition of U0126 and U73122, antagonists of the ERK and PLCγ pathway, respectively, significantly inhibited cell viability and GAP-43 protein expression. Moreover, BSYS treatment significantly increased the expression of the 68-, 160-, and 200-kDa neurofilaments (NFs) of proteins and the BDNF, TrkB, PI3K, and Akt mRNA and proteins in the brain or spinal cord of mice at different stages. These results indicated that BSYS promotes nerve regeneration, and its mechanism is mainly related to the upregulation of the BDNF/TrkB and PI3K/Akt signaling pathways. BS and HTHX also promoted nerve regeneration, and this effect involved the ERK pathway.
Highlights
Multiple sclerosis (MS) is an autoimmune response-mediated inflammatory neurodegenerative disease of the central nervous system (CNS) (Herz et al, 2010; Bowles et al, 2017)
Antibodies specific for β-tubulin-III, NF200, growth associated protein (GAP)-43, phosphorylated (p)-cAMP-response element binding protein (CREB), extracellular signal-regulated kinase (ERK), p-ERK, PLCγ, TrkB, PI3K, p-PI3K, Akt, and p-Akt were purchased from Cell Signaling Technology (Danvers, MA, USA); antibodies specific for CREB, NF68, and NF160 were purchased from Abcam (Cambridge, MA, UK), and the antibody specific for brain-derived neurotrophic factor (BDNF) antibody was purchased from Epitomics (Burlingame, USA)
The possible involvement of Bu Shen Yi Sui formula (BSYS) and its decomposed Bu Shen formula (BS) and Hua Tan Huo Xue formula (HTHX) formulas in the PI3K, ERK, and PLCγ signaling pathways was investigated in SH-SY5Y cells
Summary
Multiple sclerosis (MS) is an autoimmune response-mediated inflammatory neurodegenerative disease of the central nervous system (CNS) (Herz et al, 2010; Bowles et al, 2017). Brain-derived neurotrophic factor (BDNF) is a small molecular peptide that plays an important role in the growth and development of neurons, the repair of damaged nerves, and the induction of axonal regeneration (Zhu et al, 2012; Wang et al, 2016). BDNF activates three main signal transduction pathways, PI3K, extracellular signal-regulated kinases (ERK), and PLCγ, after binding to its receptor TrkB in presynaptic terminals (Zhu et al, 2012). Experimental studies found decreased expression of BDNF in the brain and spinal cord in mice with experimental autoimmune encephalomyelitis (EAE), an animal model of MS (Wang et al, 2014). Activation of the PI3K/Akt signaling pathway helped to stabilize the internal milieu in the brain and protect neurons, which could accelerate angiogenesis and resistance to oxidative stress in MS patients. ERK was markedly decreased after U0126 treatment (Zhang et al, 2015b), and activation of PLCγ was significantly decreased by U73122 treatment (Liu et al, 2015), which inhibited the pathway
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