Simultaneous fluorescence imaging of hydrogen peroxide in mitochondria and endoplasmic reticulum during apoptosisElectronic supplementary information (ESI) available: Detailed experimental procedures, characterization of compounds, some fluorescence images. See DOI: 10.1039/c6sc01793b

• Main Authors: Xiao, Haibin, Li, Ping, Hu, Xiufen, Shi, Xiaohui, Zhang, Wen, Tang, Bo
• Format: Journal Article
• Published: 16-08-2016
• ISSN: 2041-6520; 2041-6539
• DOI: 10.1039/c6sc01793b

Cell apoptosis is a biochemical and molecular pathway essential for maintaining cellular homeostasis. It is an integrated process involving in a series of signal transduction cascades. Moreover, the apoptotic pathways may be initiated inside various subcellular organelles. Increasing evidence indicates that hydrogen peroxide (H
2
O
2
) is closely related to cell apoptosis, particularly in the mitochondria. However, during the apoptotic process, the synergetic variation of H
2
O
2
levels in different compartments is seldom explored, particularly in two important organelles: mitochondria and endoplasmic reticulum (ER). To solve this problem, we developed two new organelle-specific fluorescent probes termed
MI-H
2
O
2
and
ER-H
2
O
2
that can detect H
2
O
2
in mitochondria and ER, respectively or simultaneously. Experimental results demonstrated that
MI-H
2
O
2
and
ER-H
2
O
2
display distinguishable excitation and emission spectra, as well as excellent organelle targeting capabilities. Therefore, we used
MI-H
2
O
2
and
ER-H
2
O
2
to successfully image exogenous or endogenous hydrogen peroxide in the mitochondria and ER. Interestingly, during diverse apoptotic stimuli, dual-color fluorescence imaging results revealed that the changes of H
2
O
2
levels in mitochondria and ER are different. The H
2
O
2
levels are enhanced in both the mitochondria and ER during the
l
-buthionine sulfoximine (BSO)-treated cell apoptosis process. During mitochondria-oriented apoptosis induced by carbonyl cyanide
m
-chlorophenylhydrazone (CCCP) or rotenone, H
2
O
2
levels prominently and continuously increase in the mitochondria, whereas the ER H
2
O
2
levels were found to rise subsequently after a delay. Moreover, during ER-oriented apoptosis induced by tunicamycin, ER is the major site for overproduction of H
2
O
2
, and delayed elevation of the H
2
O
2
levels was found in the mitochondria. Altogether, this dual-probe and multicolor imaging approach may offer a proven methodology for studying molecular communication events on H
2
O
2
-related apoptosis and also other physiological and pathological processes within different subcellular organelles.
We have developed two new organelle-specific fluorescent probes for the simultaneous imaging of hydrogen peroxide in the mitochondria and the endoplasmic reticulum during apoptosis.