Amyloid-beta mediates the receptor of advanced glycation end product-induced pro-inflammatory response via toll-like receptor 4 signaling pathway in retinal ganglion cell line RGC-5

Abstract

Patients with diabetes mellitus have an increased risk of developing Alzheimer's disease. Amyloid-β, a product of amyloid precursor protein, is associated with neuro-inflammation in patients with Alzheimer's diseases. The correlation between amyloid-beta and advanced glycation end products, which accumulate in tissue of diabetic patients, is not clear. The aims of this study were to determine the effect of advanced glycation end product on the expression of amyloid precursor protein/amyloid-beta and associated pro-inflammatory responses in retinal ganglion cell line RGC-5. Treatment with advanced glycation end product produced upregulation of amyloid precursor protein and increased secretion of amyloid-β(1–40). Additionally, amyloid-β(1–40) induced toll-like receptor 4-dependent phosphorylation of tyrosine in myeloid differentiation primary response gene (88). We found that N-[N-(3,5-Difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester, a γ-secretase inhibitor, reduced the secretion of amyloid-β(1–40) and inhibited the advanced glycation end product-induced activation of myeloid differentiation primary response gene (88). Amyloid-β(1–40) induced the activation of NF-κB and the expression of TNFα mRNA. Knockdown of toll-like receptor 4 inhibited the amyloid-β(1–40)-induced phosphorylation of p65 in NF-κB. Additionally, the nuclear translocation of p65 and transcriptions of TNFα were inhibited by siRNA knockdown of receptor of advanced glycation end product or toll-like receptor 4. The advanced glycation end product-induced secretion of VEGF-A was also reduced by knockdown of toll-like receptor 4. Taken together, our data suggested that amyloid-β(1–40) mediates the interaction between receptor of advanced glycation end product and toll-like receptor 4. Inhibition of the toll-like receptor 4 is an effective method for suppressing the amyloid-β(1–40)-induced pro-inflammatory responses in RGC-5 cells.

Introduction

The presence of amyloid-beta (amyloid-β or Aβ) plaques formed by Aβ aggregation and neurofibrillary tangles containing the microtubule-stabilizing protein tau is a characteristic feature of Alzheimer's disease (AD) (Huang and Jiang, 2009, Sisodia and Price, 1995). The correlation between AD and diabetes mellitus (DM) has gained increasing attention. Epidemiological studies show that diabetic patients have an increased risk of developing AD (Maher and Schubert, 2009, Ott et al., 1996, Ott et al., 1999). The two are likely linked because impaired insulin signaling in Type 2 DM disrupts both the processing of amyloid precursor protein (APP) and the clearing of Aβ (Messier and Teutenberg, 2005).

The receptor for advanced glycation end products (RAGE), which mediates oxidative stress and vascular inflammation in DM (Nogueira-Machado and Chaves, 2008, Schmidt et al., 1999), is a putative receptor for Aβ peptide and plays a role in AD (Arancio et al., 2004, Deane et al., 2003, Donahue et al., 2006, Yan et al., 1996). RAGE is also involved in the translocation of Aβ from the extracellular to intracellular space and is correlated with neuronal cytotoxicity in the central nervous system (Takuma et al., 2009). Therefore, cerebrovascular alteration, which is a common pathological change in both AD and DM, could be the mechanism linking the two diseases (Takeda et al., 2011).

RAGE and toll-like receptors (TLRs) share common ligands and show crosstalk in signaling pathways that mediate the host immune response (Ibrahim et al., 2013). The TLR family is the best characterized of the pattern recognition receptor families, which detect conserved molecular patterns invariant among entire classes of pathogens and thereby trigger an innate immune response (Kawai and Akira, 2010). TLRs are activated in patients with DM (Dasu et al., 2010). Nutrients or other ligands can trigger TLRs to activate the inflammasome in cells, which releases cytokines, or to induce insulin resistance (Reynolds et al., 2012). Evidence from in vitro and in vivo studies suggests that TLR signaling pathway, including TLR2, 4 or 9 may be involved in clearance of Aβ-deposits in the brain and, therefore, that the TLR family could be a therapeutic target for AD (Tahara et al., 2006). Evidence for TLR4 signaling has been demonstrated in the mouse model of AD, in which it was associated with an increase expression in inflammatory cytokines (Jin et al., 2008).

In the eye, Aβ deposition disrupts retinal structure and may contribute to the visual deficits and retinal inflammation observed in the mouse model of AD (Ning et al., 2008, Perez et al., 2009). Aβ is also a known component of drusen, and it plays a proinflammatory role in the retinal pigment epithelium and neuroretina (Liu et al., 2013). However, the role played by APP and Aβ in retinal cells during DM is currently unclear. Therefore, we conducted an in vitro study using RGC-5 cells, which are identical to the retinal photoreceptor cell line 661W (Krishnamoorthy et al., 2013), to represent a neuronal cell in the retina. We used RGC-5 cells to investigate the effect of advanced glycation end products (AGEs), components that are associated with hyperglycemia, hyperlipidemia, enhanced oxidative stress and obesity (Gaens et al., 2013) on APP expression. Additionally, we studied the role of RAGE and TLR signaling in mediating Aβ-induced innate immune responses.