BDNF-induced Migration Research Articles

Phosphacan acts as a repulsive cue in murine and rat cerebellar granule cells in a TAG-1/GD3 rafts-dependent manner.

Phosphacan, a chondroitin sulfate proteoglycan, is a repulsive cue of cerebellar granule cells. This study aims to explore the molecular mechanism. The glycosylphosphatidylinositol-anchored neural adhesion molecule TAG-1 is a binding partner of phosphacan, suggesting that the repulsive effect of phosphacan is possibly because of its interaction with TAG-1. The repulsive effect was greatly reduced on primary cerebellar granule cells of TAG-1-deficient mice. Surface plasmon resonance analysis confirmed the direct interaction of TAG-1 with chondroitin sulfate C. On postnatal days 1, 4, 7, 11, 15, and 20 and in adulthood, phosphacan was present in the molecular layer and internal granular layer, but not in the external granular layer. In contrast, transient TAG-1 expression was observed exclusively within the premigratory zone of the external granular layer on postnatal days 1, 4, 7, and 11. Boyden chamber cell migration assay demonstrated that phosphacan exerted its repulsive effect on the spontaneous and brain-derived neurotrophic factor (BDNF)-induced migration of cerebellar granule cells. The BDNF-induced migration was inhibited by MK-2206, an Akt inhibitor. The pre-treatment with a raft-disrupting agent, methyl-β-cyclodextrin, also inhibited the BDNF-induced migration, suggesting that lipid rafts are involved in the migration of cerebellar granule cells. In primary cerebellar granule cells obtained on postnatal day 7 and cultured for 7 days, the ganglioside GD3 and TAG-1 preferentially localized in the cell body, whereas the ganglioside GD1b and NB-3 localized in not only the cell body but also neurites. Pre-treatment with the anti-GD3 antibody R24, but not the anti-GD1b antibody GGR12, inhibited the spontaneous and BDNF-induced migration, and attenuated BDNF-induced Akt activation. These findings suggest that phosphacan is responsible for the repulsion of TAG-1-expressing cerebellar granule cells via GD3 rafts to attenuate BDNF-induced migration signaling.

BDNF promoted osteoblast migration and fracture healing by up-regulating integrin β1 via TrkB-mediated ERK1/2 and AKT signalling.

Brain‐derived neurotrophic factor (BDNF) has been reported to participate in fracture healing, whereas the mechanism is still unclear. Since osteoblast migration is important for fracture healing, investigating effects of BDNF on osteoblasts migration may help to reveal its mechanism. Here, MC3T3‐E1 cells were used in vitro while closed femur fracture mice were applied in vivo. Cells migration was assessed with Transwell assay. The protein expression was analysed by immunoblotting. X‐ray and Micro‐CT were performed at different time after fracture. Our results showed that BDNF promoted MC3T3‐E1 cells migration, integrin β1 expression and ERK1/2 and AKT phosphorylation. K252a, a specific inhibitor for TrkB, suppressed BDNF‐induced migration, integrin β1 expression and activation of ERK1/2 and AKT. PD98059 (an ERK1/2 inhibitor) and LY294002 (an AKT inhibitor) both inhibited BDNF‐induced migration and integrin β1 expression while integrin β1 blocking antibody only suppressed cell migration. X‐ray and Micro‐CT analyses showed that the adenoviral carried integrin β1 shRNA group had slower fracture healing at 7 and 21 days, but not 35 days compared to the control group. Thus, we proposed that BDNF stimulated MC3T3‐E1 cells migration by up‐regulating integrin β1 via TrkB mediated ERK1/2 and AKT signalling, and this may help to enhance the fracture healing.

Delphinidin inhibits BDNF-induced migration and invasion in SKOV3 ovarian cancer cells

Brain-derived neurotrophic factor (BDNF), the TrkB ligand, is associated with aggressive malignant behavior, including migration and invasion, in tumor cells and a poor prognosis in patients with various types of cancer. Delphinidin is a diphenylpropane-based polyphenolic ring structure-harboring compound, which exhibits a wide range of pharmacological activities, anti-tumor, anti-oxidant, anti-inflammatory, anti-angiogenic and anti-mutagenic activity. However, the possible role of delphinidin in the cancer migration and invasion is unclear. We investigated the suppressive effect of delphinidin on the cancer migration and invasion. Thus, we found that BDNF enhanced cancer migration and invasion in SKOV3 ovarian cancer cell. To exam the inhibitory role of delphinidin in SKOV3 ovarian cancer migration and invasion, we investigated the use of delphinidin as inhibitors of BDNF-induced motility and invasiveness in SKOV3 ovarian cancer cells in vitro. Here, we found that delphinidin prominently inhibited the BDNF-induced increase in cell migration and invasion of SKOV3 ovarian cancer cells. Furthermore, delphinidin remarkably inhibited BDNF-stimulated expression of MMP-2 and MMP-9. Also, delphinidin antagonized the phosphorylation of Akt and nuclear translocation of NF-κB permitted by the BDNF in SKOV3 ovarian cancer cells. Taken together, our findings provide new evidence that delphinidin suppressed the BDNF-induced ovarian cancer migration and invasion through decreasing of Akt activation.

Brain‐derived neurotrophic factor induces migration of endothelial cells through a TrkB–ERK–integrin αVβ3–FAK cascade

Brain-derived neurotrophic factor (BDNF) promotes the regeneration of periodontal tissue. Since angiogenesis is important for tissue regeneration, investigating effect of BDNF on endothelial cell function may help to reveal its mechanism, whereby, BDNF promotes periodontal tissue regeneration. In this study, we examined the influence of BDNF on migration in human microvascular endothelial cells (HMVECs), focusing on the effects on extracellular signal-regulated kinase (ERK), integrin α(V)β(3), and focal adhesion kinase (FAK). The migration of endothelial cells was assessed with a modified Boyden chamber and a wound healing assay. The expression of integrin α(V)β(3) and the phosphorylation of ERK and FAK were analyzed by immunoblotting and immunofluorescence microscopy. BDNF (25 ng/ml) induced cell migration. PD98059, an ERK inhibitor, K252a, a specific inhibitor for TrkB, a high affinity receptor of BDNF, and an anti-integrin α(V)β(3) antibody suppressed the BDNF-induced migration. BDNF increased the levels of integrin α(V)β(3) and phosphorylated ERK1/2 and FAK. The ERK inhibitor and TrkB inhibitor also reduced levels of integrin α(V)β(3) and phosphorylated FAK. We propose that BDNF stimulates endothelial cell migration by a process involving TrkB/ERK/integrin α(V)β(3)/FAK, and this may help to enhance the regeneration of periodontal tissue.

K252a induces anoikis-sensitization with suppression of cellular migration in Epstein-Barr Virus (EBV)—associated nasopharyngeal carcinoma cells

Recent studies revealed an unexpected role of the neurotrophin receptor pathway, BDNF/TrkB signaling, in cancer metastasis and anoikis (i.e. detachment-induced cell death). Survival of cancer cells in detached state (known as anoikis-resistance) is known to be pre-requisite for metastasis. Nasopharyngeal carcinoma (NPC), an endemic head and neck cancer in Southeast Asia, is highly invasive, metastatic, and etiologically associated with Epstein-Barr virus (EBV, an oncovirus) infection. Mechanistic studies on the invasive/metastatic nature of NPC can facilitate the development of anti-metastatic therapy in NPC. Thus far, the role of BDNF/TrkB signaling in virus-associated human cancer is unclear. Here, using multiple cell line models of NPC with EBV-association (HONE-1-EBV, HK1-LMP1 and C666-1), we investigated the potential involvement of BDNF/TrkB signaling in cellular migration and anoikis-resistant characteristics of NPC. We found that all three EBV-associated NPC cell lines tested were intrinsically anoikis-resistant (i.e. survived in detached state) and expressed both BDNF and TrkB. BDNF stimulation induced cellular migration, but not proliferation of these cells. Further, we examined if pharmacologic targeting of anoikis-resistance of NPC cells can be achievable by a proof-of-concept Trk inhibitor, K252a, in these EBV-associated NPC models. Our results demonstrated that K252a, was able to attenuate BDNF-induced migration and proliferation of NPC cells. More importantly, we demonstrated for the first time that K252a harbored potent anoikis-sensitization activity (i.e. sensitizing cancer cells to detachment-induced cell death) against EBV-associated human cancer cells, namely NPC cells. This proof-of-concept study demonstrated that K252a, a Trk inhibitor, can potentially be used as an anoikis-sensitizing agent in NPC.

Signaling Endosomes Point the Way

Brain-derived neurotrophic factor (BDNF) stimulates the migration of cerebellar granule cell precursors (GCPs) from the external granule layer to the internal granule layer. The movement of the GCPs is impaired in BDNF knockout mice. Zhou et al . isolated GCPs from postnatal day 6 (P6) mice and monitored their migration in culture in response to BDNF, applied uniformly or as a gradient. BDNF promoted the directional chemotaxis of the GCPs only when presented as a gradient. Although exogenously applied BDNF stimulated movement of GCPs from bdnf −/− mice, these cells lacked directional migration even in response to a gradient of BDNF, which suggests that an autocrine function for BDNF is also important for proper chemotaxis. To understand how autocrine BDNF may contribute to directional movement, cultured GCPs were stimulated with a related neurotrophin NT4, which also stimulates the TrkB receptor, and BDNF was found to be secreted, which could amplify the BDNF gradient and thus contribute to directional migration. GCPs have two receptors for BDNF: TrkB and p75 NTR . BDNF-induced chemotaxis (in cultured cells and organotypic slices) was blocked if the TrkB receptor activity was inhibited but was unaffected in cells defective for p75 NTR signaling. In the cultured GCPs, TrkB receptors colocalized with early endosomal markers at the leading edge toward the BDNF gradient, suggesting internalization of the receptor. In the P6 mouse cerebellum, TrkB was found in punctate structures that often colocalized with an endosomal marker; however, only in the wild-type mouse brains was TrkB consistently oriented toward the internal granule layer. Pharmacological inhibition of endocytosis inhibited cultured GCP migration toward a BDNF gradient, and impairment of dynamin function, which is required for clathrin-mediated endocytosis, blocked movement of GCPs from the external granule layer in organotypic slices. BDNF-induced GCP migration involved phosphoinositide 3-kinase (PI3K) signaling and activation of the Rac-specific guanine-nucleotide exchange factor Tiam1 (based on pharmacological inhibition and siRNA knockdown experiments). Thus, BDNF-induced migration appears to involve both the formation of oriented signaling endosomes and endogenous production of BDNF, which may amplify the BDNF gradient to promote motility (see Kaplan and Miller for commentary). P. Zhou, M. Porcionatto, M. Pilapil, Y. Chen, Y. Choi, K. F. Tolias, J. B. Bikoff, E. J. Hong, M. E. Greenberg, R. A. Segal, Polarized signaling endosomes coordinate BDNF-induced chemotaxis of cerebellar precursors. Neuron 55 , 53-68 (2007). [PubMed] D. R. Kaplan, F. D. Miller, Developing with BDNF: A moving experience. Neuron 55 , 1-2 (2007). [PubMed]