Effects of Human Amniotic Fluid Mesenchymal Stem Cells Supernatant on Secretion of Enzymes Involved in Amyloid β Protein Degradation and Calpain 1 in SH-SY5Y Human Neuroblastoma Cell Line

Abstract

Alzheimer’s disease is commonly diagnosed by significant extracellular deposition of beta-amyloid and intracellular neurofibrillary tangles formation. Here, we examined the paracrine effect of amniotic fluid mesenchymal stem cells on Alzheimer’s changes in human SH-SY5Y cells. Methods and materials: SH-SY5Y cells were allocated into five groups as follows; Control, 0.1 µg/ml LPS, 10 µg/ml LPS, 0.1 µg/ml + Conditioned Medium and 10 µg/ml LPS + Conditioned Medium. Cells were incubated with 0.1 and 10 µg/ml LPS for 24 hours followed by incubation with the conditioned medium of amniotic fluid mesenchymal stem cells for the next 24 hours. The existence of beta-amyloid plaques was monitored using Congo-Red staining. Survival rate and apoptosis were assessed using MTT assay and flow cytometric analysis of Annexin-V. The levels of Neprilysin, Angiotensin-converting enzyme, and Matrix Metalloproteinase-9 were measured by ELISA. Using a PCR array, the expression of genes involved in neurogenesis was measured and Calpain 1 gene expression was assessed using real time PCR. Results: Bright-field imaging revealed the existence of beta-amyloid plaques in the group exposed to 10 µg/ml LPS. We found minimum effects in groups that received 0.1 µg/ml LPS. Amniotic fluid stem cell secretome increased the viability of LPS-treated SH-SY5Y cells (p<0.05) coincided with the reduction of apoptotic changes (p<0.05). According to data, Decreased Neprilysin, Angiotensin-converting enzyme, and Matrix Metalloproteinase-9 levels were blunted in LPS-treated cells upon incubation with stem cells secretome (p<0.05). We noted the modulation of different genes participating in the neurogenesis in the 10 µg/ml LPS + Conditioned Medium group compared to cells that received 10 µg/ml LPS.