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Cellular and Molecular Bioengineering

Effect of Receptor for Advanced Glycation End-Products Inflammatory Response in Alzheimer's Disease

(I-327) Effect of Receptor for Advanced Glycation End-Products Inflammatory Response in Alzheimer's Disease

Saturday, October 14, 2023

9:30 AM – 10:30 AM PDT

Location: Exhibit Hall - Row I - Poster # 327

Presenting Author(s)

HT

Hadley Taggart

Student
Benedictine College
Wakarusa, Kansas, United States

Co-Author(s)

MW

Mihyun Lim Waugh (she/her/hers)

Ph.D. Student
University of South Carolina
cayce, South Carolina, United States

Primary Investigator(s)

MM

Melissa Moss

Professor and Department Chair
University of South Carolina, United States

Introduction:: Alzheimer’s disease (AD) is the cause of 60-70% of the estimated 55 million dementia cases worldwide. This multifactorial disease has been characterized by amyloid-ꞵ (Aꞵ) plaques, neurofibrillary tangles, and chronic neuroinflammation. Studies have linked the receptor for advanced glycation end-products (RAGE) to development of the chronic inflammation observed in AD. RAGE is an innate immune receptor found in inflammation processes throughout the human body, and its ligands, including Aꞵ oligomers and S100B, are upregulated in AD. S100B, at low (nanomolar) levels, benefits brain health, but at higher levels contributes to the pathways for chronic inflammation. Another RAGE ligand, lipopolysaccharide (LPS), is an endotoxin that is used in AD inflammation models to study inflammatory response. These ligands, when bound to RAGE, trigger the NF-κB pathway, resulting in the release of inflammatory cytokines, such as IL-1ꞵ, and upregulation of RAGE. Chronic inflammation resulting from the upregulated ligands can lead to neuronal apoptosis and induce the brain shrinkage observed in AD. This study explores whether cytokines released by immune cells when stimulated by RAGE ligands could perpetuate neuronal death in the AD brain.

Materials and Methods::

THP-1 macrophages share similar function to microglial cells in that they are part of the innate immune system, which is the main method of immune response in AD. Both microglia and THP-1 macrophages release cytokines as a response to immune system stimuli. SH-SY5Y cells are neuroblastoma cells that can model the behavior of neurons. In recent in vitro studies, both undifferentiated and differentiated SHSY-5Y cells have been used when neuronal-like cell culture was required. The combined use of THP-1 macrophages and SHSY-5Y cells are used in this study to model the in vitro studies of inflammatory response in AD brain.

THP-1 monocytes were differentiated to macrophages via 72-h incubation with phorbol 12-myristate 13-acetate (PMA). The macrophages were then treated with either S100B (at 0, 0.05, 0.1, 0.5, 1, 5 µg/mL) or LPS (at 2, 4, 10, 20 ng/mL) for 72-h, and supernatants were collected.

Using an ELISA, levels of proinflammatory cytokine IL-1ꞵ were detected in supernatant from S100B and LPS treated cells. In addition, ELISA was used to evaluate levels of residual S100B in S100B-treated THP-1 macrophages.

To measure the effect of cytokines on neuron-like cell culture, SH-SY5Y cells were incubated for 24-h with the supernatants collected from S100B and LPS-treated THP-1 macrophages. An XTT assay was performed to measure cell viability at the end of the period of incubation.

Results, Conclusions, and Discussions::

Treatment of THP-1 macrophages with either LPS or S100B induces production of IL-1ꞵ with negligible cytokine production resulting from the untreated control (Figure 1). Moreover, the IL-1ꞵ concentration was higher in supernatants collected from THP-1 macrophages treated with LPS than from those treated with S100B. In addition, at the highest treatment concentrations, additional increases in IL-1ꞵ were not observed suggesting that the maximum production of IL-1ꞵ was induced. In contrast, residual concentrations of S100B after treatment displayed a general trend of increasing S100B concentration with increased concentration in the treatment phase. Treatment of SH-SY5Y cells with the supernatants from the LPS or S100B treated supernatants did not display a decrease in cell viability after 24-h (shown for S100B in Figure 2).

These results indicate IL-1ꞵ cytokine production did result from treatment with RAGE ligands S100B and LPS; however, these cytokines did not induce neurotoxicity following a 24-h exposure. Future work will explore longer treatment times and lower dilution ratios as well as treatment with cytokine-containing supernatants in the presence of Aꞵ. In particular, the latter is expected to potentiate Aꞵ toxicity.