The prevalence of polycystic ovary syndrome (PCOS) has been gradually increasing among adult females worldwide. Laparoscopy drilling on ovary is the only available temporary solution with a high incidence of reoccurrence. S100A8 with S100A9 complex is believed to facilitate the cyst migration in PCOS condition. The high evident protein interaction network studies between PCOS biomarkers, cancer invasion markers, and the interactors of S100A8 confirm that this protein has strong interaction with other selective PCOS biomarkers, which may be associative in the immature cyst invasion process. Through the network studies, intensive structural and pathway analysis, S100A8 is identified as a targetable protein. In this research, the non-SELEX in silico method is adapted to construct RNA Library based on the consensus DNA sequence of Glucocorticoid Response Element (GRE) and screened the best nucleotide fragments which are bound within the active sites of the target protein. Selected sequences are joined as a single strand and screened the one which competitively binds with minimal energy. In vitro follow-up of this computational research, the designed RNA aptamer was used to infect the MCF7 cell line through Lipofectamine 2000 mediated delivery to study the anti-cell migration effect. Wound Scratch assay confirms that the synthesized 18-mer oligo has significant inhibition activity toward tumor cell migration at the cellular level.
Introduction:
Aptamers are emerging newer therapeutics and diagnostics can be designed to bind any kind of target proteins. Vascular endothelial damage by the excess amount of nitric oxide production in systemic circulation leads to the secretion of inflammatory chemoattractant and cell adhesion, which is the prime pro-atherogenic events in the formation of plagues at atrial intimal layers due to oxidation – sensitive mechanisms. Nitric oxide inhibition assay is one of the valuable qualitative anti-atherosclerosis matrices.
Methods:
In this research, Nitric oxide inhibition efficiency of a ssDNA aptamer on cell lines was studied and the respective targets of that aptamer were identified by network analysis. The aptamer used here was originally designed for Selectin P Ligand Protein to control atherogenic process. 20 nM of aptamer solution in LipofectamineTM 2000 shows the highest level of 70.5 % inhibition of nitric oxide liberation on 24 hours cultured medium of Lipopolysaccharide stimulated murine macrophage RAW 264.7 cell lines.
Results:
Protein interaction network analysis on the nitric oxide synthesis pathway interactors and the molecular docking analysis with network resulted proteins such as AKT Serine/Threonine Kinase 1, Calmodulin, Estrogen Receptor 1, and Nitric Oxide Synthase-3 confirms that the G – quadruplex Model of 18-mer sequence effectively binds on the active sites of Estrogen Receptor 1, and Nitric Oxide Synthase-3.
Conclusion :
The aptamer designed for atherosclerotic target have also exert significant nitric oxide inhibition to control the atherogenic events through the proteins, AKT1, NOS3 and ESR1.
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