Revealing of cell death induced in different Ca2+ oscillations by optogenetics

Abstract

Calcium (Ca2+) plays an important and ubiquitous role of intracellular signaling as it involved in life and death signals of the cell for example fertilization transcription factors activation cell proliferation cell migration ATP synthesis cell apoptosis etc Previous studies mentioned that the homeostasis of Ca2+ concentrations in the cell is important to maintain cell life pathway; however if the concentration of Ca2+ was in the pathological level it will activate cell death pathway which leads to cell apoptosis Optogenetic technology a new approach with spatial and temporal precision have been developed and applied to control cell activity in many biomedical studies Therefore we set up optogenetic platforms with blue light (470 nm) illumination that can modulate the optical parameters (frequency power duty cycle and duration) The stable U2OS cell line overexpressing Ca2+ translocating channelrhodopsin (CatCh) a point-mutated form of channelrhodopsin 2 (ChR2) generates Ca2+ influx by illuminating 470 nm blue light In our study we investigated the patterns of Ca2+ oscillations in cell death by optogenetics The results showed that a higher frequency (1 Hz) induced cell swelling and round up of the morphological changes when the cells incubated over 6 hours in 10 30 and 60 minutes of light treatment In addition the sub G1 population increases during incubation after light treatment which represented cell necrosis On the other hand a lower frequency (0 1 Hz) induced cell shrinkage when the cells incubated over 48 and 12 hours in 30 and 60 minutes of light stimulation respectively Furthermore we found that the cell death in 0 1 Hz treatment was predominantly apoptotic cell as the sub G1 population was far lower than 1 Hz And in 0 1 Hz stimulation we found that the autophagy activity in the 30-minute group is higher than the 60-minute group which lowered down the percentage of cell death in the 30-minutes group

Date of Award	2018 Aug 21
Original language	English
Supervisor	Wen-Tai Chiu (Supervisor)