Abstract
The purpose of this study was to examine the differences which exist between murine, human, and rat Siglec-1, then to research novel inhibitors and assess applicability of HAPI cells to study Siglec-1. Using molecular modelling software, amino acid sequences were analysed for differences across orthologs. Compound library screening of human Siglec-1 was performed, 9 compounds of interest were identified, and 4 were included additionally out of clinical interest. Rational structure activity analysis was conducted on orthologs and chosen compounds. Competitive inhibition enzyme-linked immunosorbent assay (ELISA) using Siglec-1 was used on these 13 chosen compounds. This was followed by lipopolysaccharide (LPS) treatment of HAPI cells and Siglec-1 detection ELISA to confirm upregulation of Siglec-1. All 13 compounds were analysed using resazurin in HAPI cells with and without treatment of LPS. The N-terminal of murine Siglec-1 varies by approximately by 22% and 11% compared to human and rat orthologs respectively. Murine Siglec-1 includes Leu107 while rat and human share the more polar Ser107, despite a highly similar structure overall. Four compounds showed definable IC(50) values. LPS stimulation compared to normal growth conditions provided significant Siglec-1 upregulation. Analysis of chosen compounds was carried out using resazurin and there was no significant cytotoxicity in Siglec-1 upregulated cells. Siglec-1 is not as highly conserved across species as previously thought, but human and rat maybe be more similar than either to mouse. HAPI cells were shown as an appropriate microglial cell model, with capable of showing upregulation of Siglec-1 under LPS conditions.