The role of ribosylated-BSA in regulating PC12 cell viability

Abstract

Glycation, one of the post-translational modifications, is known to influence protein structure and biological function. Advanced glycation end products (AGEs) have been shown to cause pathologies of diabetes. Glycation levels in patients with Alzheimer's disease (AD) are higher than in normal people. However, whether the glycation of susceptible proteins is a triggering event for cell damage or simply a result remains to be elucidated. In this study, we demonstrated that ribose-conjugated BSA (Rib-BSA) directly induces PC12 cell death in a dose- and time-dependent manner. The IC(50) is 4.6 μM. Unlike glucose-incubated BSA, Rib-BSA rapidly forms cytotoxic AGEs. PC12 is vulnerable to Rib-BSA. However, fructose can induce AGE formation, although no effect on cell survival was observed. This effect of Rib-BSA is reversed by pretreatment of pioglitazone and rosiglitazone, which belongs to thiazolidinediones (TZDs) and are peroxisome proliferator-activated receptor (PPAR-γ) ligands. Moreover, Rib-BSA upregulates inducible nitric oxide synthase (iNOS), cycloxygenase 2 (COX-2) expression, and p-38 phosphorylation and leaves extracellular regulated protein1/2 (ERK1/2) phosphorylation unchanged. The Rib-BSA-induced signaling changes are blocked by rosiglitazone and confirmed by PPAR-γ small-interfering RNA transfection. The reduction of cell survival by Rib-BSA is blocked by the iNOS inhibitor and p38 inhibitor. No effect on cell survival was observed using the COX-2 inhibitor. Consequently, these results show that Rib-BSA directly inducing PC12 cell death is a triggering event and TZDs protect PC12 cell from Rib-BSA damage. Signaling molecules, such as PPAR-γ, P38, and iNOS, are involved in Rib-BSA-mediated cytotoxicity.

In Vitro Methodologies to Study the Role of Advanced Glycation End Products (AGEs) in Neurodegeneration

“…This line is easy to handle and homogenous [173], while the cells can undergo neuronal differentiation in response to nerve growth factors resulting in large numbers of post-mitotic cells [174]. Many researchers have worked with this line to study neurodegeneration [175,176] and neurotoxicity caused by AGEs [177][178][179][180]. PC12 cells express RAGE [179] and they were vulnerable to ribosylated-BSA induced cytotoxicity, while the exposure increased iNOS and COX-2 expression and phosphorylation of p38 [178].…”

“…Many researchers have worked with this line to study neurodegeneration [175,176] and neurotoxicity caused by AGEs [177][178][179][180]. PC12 cells express RAGE [179] and they were vulnerable to ribosylated-BSA induced cytotoxicity, while the exposure increased iNOS and COX-2 expression and phosphorylation of p38 [178]. Cell apoptosis and expression of RAGE and NF-κB were significantly increased in PC-12 cells exposed to glycated BSA [179].…”

NBM-T-L-BMX-OS01, Semisynthesized from Osthole, Is a Novel Inhibitor of Histone Deacetylase and Enhances Learning and Memory in Rats

“…Cell viability was evaluated by an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, Sigma-Aldrich) assay 24 h after SAHA and BMX treatment at concentrations of 0, 1, 2.5, 5, and 10  μ M. The protocol was performed as described previously [40]. Cells were incubated with an MTT (0.5 mg/mL) reagent (Sigma-Aldrich) at 37°C for 1 h. The MTT solution was removed, and DMSO was added to the wells shaken at room temperature for 1 h. The amount of MTT formazan product was quantified by measuring its absorbance at 570 and 630 nm by using an ELISA plate reader (SpectraMax M2 Microplate Readers, Molecular Devices, Sunnyvale, CA, USA).…”

Cell toxicity mechanism and biomarker

“…In diabetes and Alzheimer’s disease, the upregulates of NO synthase was also identified with the involved pathway of PPAR-γ, P38. These signaling changes are blocked by PPAR-γ small-interfering RNA transfection, and is also blocked by the NO inhibitor and p38 inhibitor [ 23 ].…”