Propofol elicits apoptosis and attenuates cell growth in esophageal cancer cell lines

概要

主論文の要旨

Propofol elicits apoptosis and attenuates cell growth in
esophageal cancer cell lines
プロポフォールは食道がん細胞株のアポトーシスを
誘発し増殖を低下する

名古屋大学大学院医学系研究科
生体管理医学講座

総合医学専攻

麻酔・蘇生医学分野

（指導：西脇 公俊
周 睿

教授）

【Introduction】
The lipophilic intravenous anesthetic agent propofol is commonly used in surgical
operations and intensive care owing to its rapid and controllable sedative effects. Propofol has
diverse effects on cellular biological functions, including positive modulation of the γaminobutyric acid A (GABAA) receptor. Propofol also binds to cellular and mitochondrial
membranes, disrupts their lipid bilayer structure, and causes cellular and mitochondrial
dysfunctions. Propofol has been shown to confer negative effects on the proliferation and
invasion of cultured neoplastic cells and elicit cellular apoptosis in vitro. Conversely, evasion
from cell death and enhancement of proliferation in cancer cell lines caused by propofol
exposure have also been demonstrated in previous studies. Several reports suggest that
propofol likely exerts varied effects on the oncogenic properties of cancer cells, depending on
the cellular context or type of cancer. However, it is largely unknown how exposure to propofol
affects the proliferation, invasion, and apoptosis of neoplastic cells in esophageal cancer. This
study was conducted to elucidate the impact of propofol exposure on the growth properties of
human esophageal cancer cell lines in vitro.
【Material and Methods】
Two Japanese human esophageal cancer cell lines, KYSE30 and KYSE960, were treated
with up to 10 µg/mL propofol for 12‒36 hours (h). The cells were then analyzed by cell
proliferation assay. To explore the effect of propofol exposure on invasive tumorigenesis, a
Matrigel invasion assay using a Boyden chamber-based assay was performed. The quantification
of caspase-3/7 and -9 activities was evaluated using chemiluminescence-conjugated caspase3/7 and -9 antibodies. Cell staining with Annexin V and 7-aminoactinomycin D (7-AAD) was
performed to detect early apoptosis and cell death, respectively, via flow cytometry.
【Results】
To understand the impact of propofol exposure on esophageal cancer cell growth, two
proliferation assays were carried out using KYSE30 and KYSE960 cells that had been exposed
to 2‒5 µg/mL propofol in a regular medium for 12‒36 h. Cell counts demonstrated a significant
dose- and time-dependent attenuation of cell growth in the KYSE30 and KYSE960 cell lines.
Cell exposure to propofol resulted in a dose-dependent suppression of Matrigel invasion in the
KYSE30 and KYSE960 cell lines. To address whether propofol induces apoptosis in
esophageal cancer cell lines, caspase activity was examined in propofol-exposed KYSE30 and
KYSE960 cell lines. Propofol-exposed esophageal cell lines upregulated the activity of
apoptosis-inducing executioner caspases (caspase-3/7), whereas caspase-9 was not activated
by propofol exposure in either cell line. These data suggest that propofol triggers apoptosis in
esophageal cancer cell lines, but does not activate the intrinsic apoptotic signaling pathway.
To confirm the induction of cellular apoptosis by propofol exposure, cells were immunostained

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with Annexin V at 6 h or 24h after the completion of propofol exposure. Both cell lines
exhibited a statistically significant increase in Annexin V-positive cells after propofol exposure.
These results are consistent with elevated caspase-3/7 activity in KYSE30 and KYSE960 cells
exposed to propofol and provide further evidence that propofol promotes cellular apoptosis in
esophageal cancer cell lines. Further analysis was conducted to detect irreversible necrosis or
cell death by staining KYSE30 and KYSE960 cells exposed to propofol with 7-AAD. The
results demonstrated a significant increase in the frequency of 7-AAD-positive cells in the
KYSE30 cell line, whereas no significant change was detected in the KYSE960 cells. These
data collectively suggest that exposure of esophageal cancer cell lines to propofol enhances
early apoptosis as indicated by elevated Annexin V positivity, which results in an increase in
cell death as indicated by elevated 7-AAD positivity, at least in a subset of cell lines.
【Discussion】
In this study, cells exposed to propofol (3–5 µg/mL for 24 h) elicited a dose- and timedependent attenuation of cell growth and dose-dependent attenuation of Matrigel invasion in
the human esophageal cancer cell lines KYSE30 and KYSE960. In addition, propofol elicited
the activation of caspase-3/7, but not caspase-9, which is a key player in the intrinsic apoptosis
pathway. Finally, propofol exposure enhanced early apoptosis in both cell lines and accelerated
cell death in KYSE30 cells. Although previous studies have reported propofol-triggered
growth suppression, reduced invasion, caspase activation, and cellular apoptosis in other types
of cancer cell lines, to the best of my knowledge, this is the first report to provide a detailed
assessment of the impact of propofol exposure on the growth properties of esophageal cancer
cell lines.
This study suggests that cell exposure to propofol activates the caspase cascade and thereby
promotes cellular apoptosis, which results in an increase in cell death, at least in a subset of
esophageal cancer cell lines. It is further speculated that the observed attenuation of cell
proliferation in propofol-exposed esophageal cancer cell lines may not be entirely attributable
to the suppression of cell growth; it might be attributable, at least partly, to the activated
cellular apoptosis elicited by propofol exposure.
Propofol has generally been used within the concentration range of 1.6–8.9 μg/mL in most
in vitro studies exploring the biological effects of propofol on cultured cells including cancer
cell lines; this study was also conducted primarily using 3–5 μg/mL propofol. Thus, when
comparing the biological consequences of propofol exposure in multiple types of cancer, the
experimental data obtained in this study should be directly comparable to those obtained in
other in vitro studies. However, it has been established that the vast majority of propofol
injected into blood vessels is conjugated with erythrocytes or serum albumin; thus, it is unclear
which concentration of propofol should be used in in vitro cellular biological studies to
precisely recapitulate in vivo propofol administration in clinical settings. To circumvent this

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issue in addressing the pharmacological action of propofol on cancer cells, it will be beneficial
to conduct future studies using animal models or clinical specimens derived from patients
treated with propofol. Such efforts will eventually enable us to establish the safe usage of
propofol in patients suffering from neoplastic diseases, including esophageal cancer.
【Conclusion】
In summary, exposure to propofol at concentrations up to 5 µg/mL led to a reduction in cell
growth and Matrigel invasion as well as the augmentation of apoptosis in esophageal cancer
cell lines. These data will help define a methodology to safely utilize propofol, a common
general anesthetic and sedative, in patients with esophageal cancer.

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