Inflammation is a defensive process that allows immune cells to be mobilized to help with infection removal and tissue regeneration. Inflammasomes are multiprotein oligomers in the cytoplasm and components of the innate immune system that have a role in inflammation. Glycogen synthase kinase 3 (GSK3) is a critical molecule involved in a wide range of inflammatory reactions. It has been reported to suppress the production of pro-inflammatory mediators in response to LPS when it is inhibited. The aim of this project was to study the effect of GSK3 inhibition in a concentration-dependent manner on the production of IL-6 as well as ASC-speck formation in THP1 ASC GFP cells stimulated with LPS and activated using nigericin. Using the cell culture supernatant ELISA was performed to quantify the IL-6 protein secreted by THP-1 macrophages. Using reverse-transcribed cDNA, qPCR was performed to measure the IL-6 gene expression. Finally, live-cell imaging was done to visualize the ASC-speck formation. It was found that upon stimulation of THP-1 cells a remarkable increase in the production of IL-6 was observed, however, the inhibitor did not suppress the production of IL-6 as hypothesized. This could be primarily due to the presence of another NF-κB pathway which is not mediated by GSK3 and therefore could not be inhibited using the GKS3 inhibitor. Future studies could decrease the LPS concentration to see if the uninhibited pathway can be observed at lower stimulation. Another probable solution could be lowering the FBS percentage to avoid potential inhibition.
| Identifer | oai:union.ndltd.org:UPSALLA1/oai:DiVA.org:his-21450 |
| Date | January 2022 |
| Creators | Shunnar, Batoul |
| Publisher | Högskolan i Skövde, Institutionen för biovetenskap |
| Source Sets | DiVA Archive at Upsalla University |
| Language | English |
| Detected Language | English |
| Type | Student thesis, info:eu-repo/semantics/bachelorThesis, text |
| Format | application/pdf |
| Rights | info:eu-repo/semantics/openAccess |
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