The anti-cancer components of Ganoderma lucidum possesses cardiovascular protective effect by regulating circular RNA expression

Abstract

To examine the role of oral Ganoderma spore oil in cardiovascular disease, we used transverse aortic constriction (TAC) in mice to model pressure overload-induced cardiomyopathy. Our preliminary results demonstrated a potential cardioprotective role for spore oil extracted from Ganoderma. We found that Ganoderma treatment normalized ejection fraction and corrected the fractional shortening generated by TAC. We also found evidence of reduced left ventricular hypertrophy as assessed by left ventricular end diastolic diameter. Analysis of total RNA expression using cardiac tissue samples from these mice corroborated our findings. We found reduced expression of genes associated with heart failure, including a novel circular RNA circ-Foxo3. Thus our data provides evidence for Ganoderma lucidum as a potential cardioprotective agent, warranting further preclinical exploration.

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www.impactjournals.com/oncoscience Oncoscience, Vol. 3 (7-8), July 2016
The anti-cancer components of Ganoderma lucidum possesses
cardiovascular protective effect by regulating circular RNA
expression
Yi-Zhen Xie1,2,*, Fenghua Yang3,*, Weijiang Tan3, Xiangmin Li1,4, Chunwei Jiao2,
Ren Huang3, Burton B. Yang4,5
1 State Key Laboratory of Applied Microbiology Southern China, Guangdong Institute of Microbiology, Guangzhou, China
2 Guangdong Yuewei Edible Fungi Technology Co. Ltd., Guangzhou, China
3 Guangdong Laboratory Animals Monitoring Institute, Guangdong Provincial Key Laboratory of Laboratory Animals,
Guangzhou, Guangdong, China
4 Sunnybrook Research Institute, Sunnybrook Health Sciences Centre, Toronto, ON, Canada
5 Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada
* These authors contributed equally
Correspondence to: Burton B. Yang, email: byang@sri.utoronto.ca
Keywords: herbal medicine, Ganoderma lucidum, anti-cancer, heart function, circular RNA
Received: June 14, 2016 Accepted: August 12, 2016 Published: August 28, 2016
This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use,
distribution, and reproduction in any medium, provided the original author and source are credited.
ABSTRACT
To examine the role of oral Ganoderma spore oil in cardiovascular disease, we
used transverse aortic constriction (TAC) in mice to model pressure overload-induced
cardiomyopathy. Our preliminary results demonstrated a potential cardioprotective
role for spore oil extracted from Ganoderma. We found that Ganoderma treatment
normalized ejection fraction and corrected the fractional shortening generated by
TAC. We also found evidence of reduced left ventricular hypertrophy as assessed
by left ventricular end diastolic diameter. Analysis of total RNA expression using
cardiac tissue samples from these mice corroborated our ndings. We found reduced
expression of genes associated with heart failure, including a novel circular RNA
circ-Foxo3. Thus our data provides evidence for Ganoderma lucidum as a potential
cardioprotective agent, warranting further preclinical exploration.
INTRODUCTION
Complementary and alternative medicines have
attracted increasing attention as disease treatments,
adjuvants, and alternative supplements [1-4]. Medicinal
mushrooms comprise a large proportion of these
alternative products, among which Ganoderma lucidum
is the most highly studied [5-8]. Preclinical studies
have demonstrated anti-tumorigenic roles in a range
of medicinal mushrooms [9-11]. A Cochrane meta-
analysis showed that patients who had been administered
Ganoderma lucidum alongside chemo/radiotherapy
were more likely to respond positively compared to
chemo/radiotherapy alone. These trials demonstrated
improved immune functions as measured by increased
CD3, CD4, and CD8 immune response cells [12]. In
vitro, Ganoderma lucidum was also found to inhibit
proliferation and induce apoptosis in ovarian, colon, and
gastric cancer cell lines [13-15]. Ganoderma lucidum
contains beta glucans and other polysaccharides which
stimulate innate immunity and activate host dendritic cells
[16, 17]. Ganoderma lucidum also produces a group of
ganoderic acids, which have molecular structures that are
similar to steroids [18, 19].
DISCUSSION
An exploratory trial of 26 patients with hypertension
and/or dyslipidemia demonstrated that Ganoderma
lucidum treatment reduced total triglycerides and increased
HDL-cholesterol, implicating a cardio-protective role
[20]. We employed a transverse aortic constriction
(TAC) mouse model of pressure overload-induced
cardiomyopathy and heart failure to examine the role of

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Ganoderma spore oil administration. The TAC model
induces an initial compensatory cardiac remodeling which
enhances cardiac contractility. Gradually, the response
to chronic overload leads to cardiac dilatation and heart
failure. The murine TAC model has been extensively used
to study cardiovascular disease and to elucidate signaling
pathways involved in cardiac hypertrophy and heart
failure.
TAC mice were administered oral Ganoderma spore
oil every other day for 14 days. The control group were
administered vegetable oil and an anti-hypertensive β2
adrenergic receptor antagonist. Mice were anesthetized
with 2% isourane inhalation for transthoracic
echocardiography and invasive hemodynamic assessment.
Transthoracic echocardiography was performed to
measure left ventricular ejection fraction (LVEF), left
ventricular fractional shortening (LVFS), left ventricular
end diastolic diameter, and cardiac output. Data analysis
was conducted in a blinded manner.
Relative to the 65.23% ejection fraction in
healthy sham mice, TAC mice were found to have a
43.26% ejection fraction (Fig 1A). This was below the
normal range of 55-75%, validating the TAC model
we employed. Treatment with the anti-hypertensive
medication approximated the normal range, while delivery
of Ganoderma spore oil recovered the stroke volume to
normal ranges. Consistent with these results, TAC mice
displayed 21.7% fractional shortening (Fig 1B). This
was within the mildly abnormal range of 20-25%, while
treatment with Ganoderma spore oil brought it to the
normal range of 25-45%. The TAC mice also showed
increased left ventricular end diastolic diameters, while
mice treated with Ganoderma spore oil did not exhibit the
same levels of left ventricular hypertrophy (Fig 2A). As a
result, mice treated with Ganoderma spore oil recovered to
physiologic cardiac output levels at 24.1 ml/min (Fig 2B).
This led to improved vascular perfusion within the mice
(Fig 2B). This suggests that Ganoderma spore oil increases
the heart function to meet the demands of the body.
After functional analyses, mice were sacriced and
heart tissue was harvested. Total RNA was extracted with
an RNeasy Mini Kit (Qiagen), followed by real-time PCR
measurement with miScriptSYBR GreenPCR Kit (Qiagen)
as described [21] to analyze levels of a circular RNA circ-
Fig 1. Ganoderma spore oil improves ejection fraction and fractional shortening. TAC mice showed decreased levels of LVEF (A)
and LVFS (B). Treatment with Ganoderma spore oil increased the levels of LVEF and LVFS.

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Foxo3. We recently demonstrated that expression of circ-
Foxo3 RNA could inhibit tumor cell cycle progression
[22, 23] and promote cardiac senescence [24]. In this
study, mice injected with the chemotherapeutic agent
Doxorubicin (Dox) for induction of cardiomyopathy were
analyzed for RNA expression levels. Mice with reduced
cardiac function had increased expression of circ-Foxo3
RNA [24]. In the current study, we found that treatment
with Ganoderma spore oil decreased levels of circ-Foxo3
(Fig 3A).
To corroborate these results, we cultured mouse
cardiac broblasts and treated the cells with Ganoderma
spore oil, following hydrogen peroxide induced oxidative
stress. Relative to control groups, treatment with
Ganoderma spore oil decreased circ-Foxo3 levels in a
concentration- (Fig 3B) and time-dependent manner (Fig
3C).
Our previous studies have shown that the oil
fraction of Ganoderma spores could induce death in
versican-transformed cancer cells [25]. Further study
found that the Ganoderma spore oil could induce death
of cancer stem-like cells [6], potentially mediated by the
molecule ergosterol peroxide [19]. Our preclinical results
here show that Ganoderma spore oil has a protective role
within the cardiovascular system. Treatment in TAC mice
was found to normalize ejection fraction and correct the
fractional shortening generated by this model. We also
found evidence of reduced left ventricular hypertrophy
as assessed by left ventricular end diastolic diameter.
Thus in our TAC model, cardiac output was improved by
oral administration of Ganoderma spore oil. This data
provides rationale for further preclinical exploration of
Ganoderma lucidum as a cardioprotective agent.
ACKNOWLEDGEMENTS
This work was supported by the High-Level
Leading Talent Introduction Program of GDAS
(2016GDASRC-0102), The Introduction of Leading
Talent Project of Guangdong Province, and Open Fund of
the Guangdong Provincial Key Laboratory of Laboratory
Animals (No. 2013A061402001).
Fig 2. Ganoderma spore oil improves cardiac output. (A) TAC mice had increased left ventricular end diastolic diameters, while
treatment with Ganoderma spore oil decreased the end diastolic diameters. (B) Treatment with Ganoderma spore oil increased cardiac
output.

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CONFLICTS OF INTEREST
No conicts of interest were disclosed.
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