As the growing global population continues to age, and the social and economic impacts grow, there will be an ever-growing need for Alzheimer’s Disease (AD) treatments. Neuroinflammation one of the hallmarks of AD, plays a major role in disease progression causing damaged neurons and activated microglia to release proinflammatory cytokines and chemokines. Somatostatin and its receptors have previously been implicated in AD. In this study the Somatostatin receptor subtype-4 (SST4) selective agonist SM-I-26 was investigated as a possible mitigator of neuroinflammation by moving microglia from a neurotoxic M1 state to a neuroprotective M2 state. Immortalized BV2 microglial cells were activated using the proinflammatory mediator lipopolysaccharide (LPS, 0 ng/mL, 10 ng/mL, and 100 ng/mL) and cells were treated with SM-I-26 (0 nM, 10 nM, 1000 nM) for 6 hours and 24 hours. Cells were then used to complete AlamarBlue assays, RT-qPCR, and nitrite assays. The AlamarBlue assay is a cell viability assay that measures the reduction capability of a cell. It was found that with a 6-hour treatment there was no decrease in viability with LPS or SM-I-26, and with a 24-hour treatment there was a significant decrease in cell viability in SM-I-26 across all LPS treatments. This eludes to SM-I-26 damaging these cells, but more work must be done to know for sure. RT-qPCR measures relative changes in mRNA expression. Within the 6-hour treatment there is not significant change with SM-I-26 treatment although there is a trend that iNOS and AIF-1 mRNA expression decreases with treatment. Interestingly with 6-hour treatment SST4 mRNA expression is significantly decreased with LPS, while without LPS treatment SM-I-26 treatment causes an increase in SM-I-26 expression. With the 24-hour treatment there is a significant increase in expression with SM-I-26 treatment for all LPS treatments. Comparing 6-hour and 24-hour treatments, it is likely that LPS overshadows SM-I-26 effects but as LPS effects fade over time the changes in mRNA expression caused by SM-I-26 begin to show at 24 hours. Within the 24-hour treatment there were large increases in mRNA expression with 1000 nM SM-I-26 across all LPS treatments of TLR4, CD14, MYD88, and NF-κB which are all a part of the same inflammatory pathway. This gives reason to think that SM-I-26 would result in the production of proinflammatory cytokines but data showed that there was a decrease in proinflammatory cytokines TNFα, IL-1β, IL-6, and iNOS with SM-I-26 treatment and an increase in the anti-inflammatory cytokine IL-10. With this indication that the microglia are moving to an anti-inflammatory M2 state it is likely that another inflammatory pathway is also being affected or one of the proteins in the TLR4 cascade is being inhibited. As the RNS nitric oxide has a half-life of around 2 minutes it must be measured indirectly through its product nitrite. The nitrite data showed no change with SM-I-26 at 6-hour but there was a decrease in nitrite levels with SM-I-26 treatment in the 24-hour treatment. Overall, it appears that the microglia are moving to an M2 neuroprotective state with the decrease in proinflammatory cytokines and increase in anti-inflammatory cytokines.
|Commitee:||Sandoval, Karen, Schober, Joseph|
|School:||Southern Illinois University at Edwardsville|
|School Location:||United States -- Illinois|
|Source:||MAI 82/1(E), Masters Abstracts International|
|Keywords:||Mitigation, Activated microglia, Associated pathways , SST4 Agonist SMI-26|
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