Altered processing of amyloid precursor protein in the human neuroblastoma SH‐SY5Y by chronic hypoxia

Abstract

Alzheimer's disease (AD) is more prevalent following an ischemic or hypoxic episode, such as stroke. Indeed, brain levels of amyloid precursor protein (APP) and the cytotoxic amyloid b peptide (Ab) fragment are enhanced in these patients and in animal models following experimental ischaemia. We have investigated the effect of chronic hypoxia (CH; 2.5% O 2 , 24 h) on processing of APP in the human neuroblastoma, SH-SY5Y. We demonstrate that constitutive and muscarinic-receptor-enhanced secretion of the a-secretase cleaved fragment of APP, sAPPa, was reduced by 60% in CH cells. The caspase inhibitor BOC-D(Ome)FMK did not reverse this effect of CH, and CH cells were as viable as controls, based on MTT assays. Thus, loss of sAPPa is not related to cell death or caspase processing of APP. Preincubation with antioxidants did not reverse the effect of CH, and the effect could not be mimicked by H 2 O 2 , discounting the involvement of reactive oxygen species in hypoxic loss of sAPPa. CH did not affect muscarinic activation of extracellular-signal regulated kinase. However, expression of ADAM 10 (widely believed to be a-secretase) was decreased approximately 50% following CH. Thus, CH selectively decreases processing of APP by the a-secretase pathway, most likely by decreasing levels of ADAM 10.

Hypoxia-inducible Factor 1α (HIF-1α)-mediated Hypoxia Increases BACE1 Expression and β-Amyloid Generation

“…Another study, however, using AD transgenic mice overexpressing mutant APP showed that neither expression nor activity of BACE was significantly affected by ischemic insult; instead ADAM10 was markedly increased in the early stage of ischemic insult and then down-regulated at later stages (41). Other studies report a drastic decrease in ADAM10 and TACE protein levels with unchanged BACE1 levels in human neuroblastoma SH-SY5Y cells subjected to chronic hypoxic treatments (42)(43)(44). While it is hard to reconcile these results, differing experimental procedures, such as cells/tissues examined, culture conditions, the percentage of O 2 utilized, or duration of hypoxic treatment, could all contribute to discrepant results.…”

Is Sleep Disruption a Risk Factor for Alzheimer’s Disease?

“…Chronic hypoxia leads to downregulation, in neurons, of the expression of ADAM10, a candidate protein for α-secretase which is essential to the processing of APP through the nonamyloidogenic pathway [85]. On the other hand, the level and activity of β-secretase (BACE-1), an enzyme that participates of the breakdown of APP through the amyloidogenic pathway, are both increased by the upregulation of hypoxia-inducible factor 1α (HIF-1-α) caused by hypoxia [86][87][88].…”

Photoluminescence intensity ratio of Eu‐conjugated lactates—A simple optical imaging technique for biomarker analysis for critical diseases

“…The cells were then transferred to 24-well plates filled with poly-L-lysine, rinsed with PBS and treated with Eu(NO 3 ) 3 . Cell viability was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT, Sigma, Thermo Fisher Scientific (Life Technologies) PaisleyPA4 9RF, UK) reduction assay as described previously [28][29][30]. Absorbance of the cells was measured using a spectrophotometer at a test wavelength of 570 nm and a reference wavelength of 630 nm.…”

“…A few studies focused on the toxicological effects of RE ions using human osteosarcoma cell line MG63, umbilical cord perivascular cell and mouse macrophages; reported large concentration or systemic accumulation of these ions could lead to acute pathology and cellular toxicity [29,30]. However, these studies produce variable results.…”