Abstract
Besides its insulinotropic actions on pancreatic β cells, neuroprotective activities of glucagon-like peptide-1 (GLP-1) have attracted attention. The efficacy of a GLP-1 receptor (GLP-1R) agonist exendin-4 (Ex-4) for functional repair after sciatic nerve injury and amelioration of diabetic peripheral neuropathy (DPN) has been reported; however, the underlying mechanisms remain unclear. In this study, the bioactivities of Ex-4 on immortalized adult rat Schwann cells IFRS1 and adult rat dorsal root ganglion (DRG) neuron–IFRS1 co-culture system were investigated. Localization of GLP-1R in both DRG neurons and IFRS1 cells were confirmed using knockout-validated monoclonal Mab7F38 antibody. Treatment with 100 nM Ex-4 significantly enhanced survival/proliferation and migration of IFRS1 cells, as well as stimulated the movement of IFRS1 cells toward neurites emerging from DRG neuron cell bodies in the co-culture with the upregulation of myelin protein 22 and myelin protein zero. Because Ex-4 induced phosphorylation of serine/threonine-specific protein kinase AKT in these cells and its effects on DRG neurons and IFRS1 cells were attenuated by phosphatidyl inositol-3′-phosphate-kinase (PI3K) inhibitor LY294002, Ex-4 might act on both cells to activate PI3K/AKT signaling pathway, thereby promoting myelination in the co-culture. These findings imply the potential efficacy of Ex-4 toward DPN and other peripheral nerve lesions.
Highlights
Glucagon-like peptide-1 (GLP-1) is an incretin hormone secreted from intestinal L cells in response to the oral nutrient ingestion and exhibits insulinotropic actions by stimulating specific GLP-1 receptors (GLP-1Rs) on the pancreatic β cells [1]
Because much smaller amount of RNA and protein was obtained from primary cultured dorsal root ganglion (DRG) neurons as compared with the lined cells, it was difficult to perform real-time Reverse Transcription-Polymerase Chain Reaction (RT-PCR) or quantitative Western blotting using these samples
RT-PCR analysis showed mRNA expression of GLP-1R and glyceraldehyde-3-phosphate dehydrogenase (GAPDH; a housekeeping gene used for confirmation of the proper reactions) in DRG neurons and IFRS1 cells, as well as NSC-34 cells [15] and ND7/23 mouse neuroblastoma/rat embryonic DRG neuron hybrid cells [18] (Figure 1a)
Summary
Glucagon-like peptide-1 (GLP-1) is an incretin hormone secreted from intestinal L cells in response to the oral nutrient ingestion and exhibits insulinotropic actions by stimulating specific GLP-1 receptors (GLP-1Rs) on the pancreatic β cells [1]. Himeno et al [9] reported that Ex-4 restored the decreases in motor nerve conduction velocities and other neurological abnormalities of streptozotocin (STZ)-induced diabetic mice without normalizing blood glucose levels These findings provide further evidence of the direct actions of Ex-4 on the peripheral nervous system; the underlying mechanisms remain largely unclear. Pre-treatment with LY294002 abolished Ex-4-induced suppression of the activity of RhoA, an inhibitory molecule of axonal regeneration and neuronal survival [11] These findings imply that Ex-4 promotes neurite outgrowth and survival of DRG neurons through the activation of PI3K signaling pathway, which negatively regulates RhoA activity. Direct evidence that Ex-4 activates PI3K pathway in DRG neurons has not been provided It remains unknown if GLP-1R agonists act on Schwann cells to promote axonal regeneration and remyelination after injury or protect neurons against diabetic and other peripheral neuropathies. We identified GLP-1R on DRG neurons and IFRS1 cells using a well-characterized and verified antibody, investigated the effects of exogenous Ex-4 on survival/proliferation and migration of IFRS1 cells, and myelination in the DRG neuron–IFRS1 co-culture system
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