Insulin dysregulation as a prominent driver of cell cycle re-entry in Alzheimer's disease.

Abstract

Cell Cycle Re-entry (CCR) is observed in several neurological disorders and contributes to cognitive impairment in the early stages. The CCR markers were monitored and correlated functionally with upregulation of Insulin Receptor (INSR) and Insulin like Growth factor 1 Receptor (IGF1R) in Alzheimer's Disease (AD) cell model and mouse brain tissue. The AD cell model was developed as per Majumder et al. (2019) and checked for Cyclin A, D and E Levels after 6 and 11 hours of model development in differentiated SHSY5Y cell line (with 10 µM RA and BDNF for cholinergic neuron like cells) by qPCR. We then checked for expression and activity of INSR and IGF1R in these cells and mouse brain (extracted from FFPE slides) through western blot and qPCR. To assess the correlation of the two events, we exploited overexpression based in-vitro studies for real-time assessment. Levels of INSR and IGF1R both increase on mRNA and protein levels in both cell and animal model. The cyclins A2, D and E however decline when the receptors are overexpressed in a serum free media for 6 hours and 11 hours. The time points were chosen to mimic the average mammalian cell cycle and showed that the upregulation of the receptors negatively impact and reduce cyclin D levels up to 4 fold whereas Cyclin A levels and Cyclin E levels are marginally reduced by 0.48 and 0.5 fold respectively. However, when stimulated with conventional ligands, Insulin and IGF1 or amyloid beta (1-42) the receptors positively correlate with the Cyclin levels. Western blots show that amyloid beta binding phosphorylates the pY1135/1136 of IGF1R, pY1150/1151 of INSR rather than conventional pY1158, 1162 and 1163. The reduction in conventional ligands, Insulin and IGF1 with ageing and build-up of amyloid beta which preferentially phosphorylates a different site could have implications that determines cellular fate after the re-entry. Reference: Majumder, P., Roy, K., Bagh, S. and Mukhopadhyay, D., 2019. Receptor tyrosine kinases (RTKs) consociate in regulatory clusters in Alzheimer's disease and type 2 diabetes. Molecular and cellular biochemistry, 459(1), pp.171-182.