ArticlePDF Available

Sulforaphane improves the bronchoprotective response in asthmatics through Nrf2-mediated gene pathways

Authors:

Abstract and Figures

Background: It is widely recognized that deep inspiration (DI), either before methacholine (MCh) challenge (Bronchoprotection, BP) or after MCh challenge (Bronchodilation, BD) protects against this challenge in healthy individuals, but not in asthmatics. Sulforaphane, a dietary antioxidant and antiinflammatory phytochemical derived from broccoli, may affect the pulmonary bronchoconstrictor responses to MCh and the responses to DI in asthmatic patients. Methods: Forty-five moderate asthmatics were administered sulforaphane (100 μmol daily for 14 days), BP, BD, lung volumes by body-plethsmography, and airway morphology by computed tomography (CT) were measured pre- and post sulforaphane consumption. Results: Sulforaphane ameliorated the bronchoconstrictor effects of MCh on FEV1 significantly (on average by 21 %; p = 0.01) in 60 % of these asthmatics. Interestingly, in 20 % of the asthmatics, sulforaphane aggravated the bronchoconstrictor effects of MCh and in a similar number was without effect, documenting the great heterogeneity of the responsiveness of these individuals to sulforaphane. Moreover, in individuals in whom the FEV1 response to MCh challenge decreased after sulforaphane administration, i.e., sulforaphane was protective, the activities of Nrf2-regulated antioxidant and anti-inflammatory genes decreased. In contrast, individuals in whom sulforaphane treatment enhanced the FEV1 response to MCh, had increased expression of the activities of these genes. High resolution CT scans disclosed that in asthmatics sulforaphane treatment resulted in a significant reduction in specific airway resistance and also increased small airway luminal area and airway trapping modestly but significantly. Conclusion: These findings suggest the potential value of blocking the bronchoconstrictor hyperresponsiveness in some types of asthmatics by phytochemicals such as sulforaphane.
This content is subject to copyright. Terms and conditions apply.
R E S E A R C H Open Access
Sulforaphane improves the bronchoprotective
response in asthmatics through Nrf2-mediated
gene pathways
Robert H. Brown
1,2,3,5*
, Curt Reynolds
5
, Allison Brooker
5
, Paul Talalay
4,6
and Jed W. Fahey
4,6
Abstract
Background: It is widely recognized that deep inspiration (DI), either before methacholine (MCh) challenge
(Bronchoprotection, BP) or after MCh challenge (Bronchodilation, BD) protects against this challenge in healthy
individuals, but not in asthmatics. Sulforaphane, a dietary antioxidant and antiinflammatory phytochemical derived from
broccoli, may affect the pulmonary bronchoconstrictor responses to MCh and the responses to DI in asthmatic patients.
Methods: Forty-five moderate asthmatics were administered sulforaphane (100 μmol daily for 14 days), BP, BD, lung
volumes by body-plethsmography, and airway morphology by computed tomography (CT) were measured pre- and
post sulforaphane consumption.
Results: Sulforaphane ameliorated the bronchoconstrictor effects of MCh on FEV
1
significantly (on average by 21 %;
p = 0.01) in 60 % of these asthmatics. Interestingly, in 20 % of the asthmatics, sulforaphane aggravated the
bronchoconstrictor effects of MCh and in a similar number was without effect, documenting the great heterogeneity
of the responsiveness of these individuals to sulforaphane. Moreover, in individuals in whom the FEV
1
response to MCh
challenge decreased after sulforaphane administration, i.e., sulforaphane was protective, the activities of Nrf2-regulated
antioxidant and anti-inflammatory genes decreased. In contrast, individuals in whom sulforaphane treatment enhanced
the FEV
1
response to MCh, had increased expression of the activities of these genes. High resolution CT scans disclosed
that in asthmatics sulforaphane treatment resulted in a significant reduction in specific airway resistance and also
increased small airway luminal area and airway trapping modestly but significantly.
Conclusion: These findings suggest the potential value of blocking the bronchoconstrictor hyperresponsiveness in
some types of asthmatics by phytochemicals such as sulforaphane.
Keywords: Asthma, Bronchodilation, Oxidative stress
Background
Although corticosteroids are currently the mainstay of
asthma treatment, some asthmatics do not respond to
corticosteroids even at high doses [1] at which adverse ef-
fects impose therapeutic limitations. Moreover, corticoste-
roids are almost universally effective in controlling many
types of asthma, and this obscures the heterogeneity of
the disease [2]. Although inflammation may be an
important inciting factor, other molecular mechanisms of
airway disease may play a crucial role in asthma.
Antioxidant deficiencies have been associated with
poor asthma control and accelerated decline of lung
function [35]. Studies in mice suggest that disruption
of the nuclear factor erythroid-related factor 2 (Nrf2)
signaling pathway is a possible cause of chronic inflam-
mation, such as that associated with asthma [6]. Nrf2 is
responsible for maintaining and restoring cellular
homeostasis through antioxidant, anti-inflammatory, and
other mechanisms. Thus Nrf2 regulates critical enzymes
concerned with the biosynthesis of glutathione (GSH),
the dominant, small molecule, intracellular antioxidant.
A protective effect of GSH on inflammatory pathologies
* Correspondence: rbrown@jhsph.edu
Supported by: Flight Attendant Medical Research Institute (FAMRI)
1
Department of Anesthesiology and Critical Care Medicine, Johns Hopkins
University School of Medicine, Baltimore, MD, USA
2
Division of Pulmonary Medicine and Critical Care, Department of Medicine,
Johns Hopkins University School of Medicine, Baltimore, MD, USA
Full list of author information is available at the end of the article
© 2016 Brown et al.
Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a
link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain
Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this
article, unless otherwise stated.
Brown et al. Respiratory Research (2015) 16:106
DOI 10.1186/s12931-015-0253-z
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
of the lung including asthma, has been demonstrated in
mice [7]. Another potentially critical role of the Nrf2 path-
way in asthma is highlighted by the dramatic increase in
eosinophilic airway inflammation, airway hyperreactivity,
and T helper cell 2 (Th2) cytokine production in Nrf2
knockout mice [6].
Sulforaphane, a phytochemical derived from cruciferous
vegetables, is a potent inducer of a variety of cytoprotective
(antioxidant and anti-inflammatory) enzymes (reviewed by
[8, 9]). It also inhibits the production of nitric oxide, coun-
teracts mitochondrial dysfunction and corrects low oxida-
tive phosphorylation, reduces lipid peroxidation, and
upregulates the heat shock response. In many tissues, sulfo-
raphane transcriptionally upregulates the genes encoding
cytoprotective enzymes primarily through the Keap1-Nrf2-
ARE system [9, 10]. Sulforaphane also potently inhibits in-
flammation by inhibiting the nuclear factor kappa-light-
chain-enhancer of activated B cells (NfκB) cascade [11], the
macrophage migration inhibitory factor (MIF), the p38
mitogen-activated protein kinase (MAPK) activation cas-
cade, and by modulating β-catenin signaling [1214].
Unlike direct antioxidants such as vitamin C, sulforaph-
ane is an indirect antioxidant and upregulates multiple
antioxidant pathways without being consumed in the anti-
oxidation process [15]. Sulforaphane boosts the antioxidant
capacity of cells by at least two major indirect mechanisms:
by induction of phase 2 cytoprotective enzymes and by
dramatically increasing cellular GSH levels. Antioxidant
deficiencies have been strongly associated with poor
asthma control and accelerated lung function decline, and
represent a reasonable therapeutic target [1618].
Nrf2-mediated signaling pathways limit airway eosino-
philia, mucus hypersecretion, and airway hyperresponsive-
ness to allergen challenge in a murine model of asthma
[6]. Genetic disruption of the Nrf2 gene (knock-out
models) leads to severe allergen-driven airway inflamma-
tion and hyper-responsiveness in mice [6]. An et al. [19]
recently showed that airway smooth muscle cells isolated
from Nrf2
/
mice exhibited significantly higher contract-
ile force compared to cells isolated from Nrf2
+/+
mice.
Thus, there is strong evidence for the role of oxidative
stress, mediated through dysfunction of the Nrf2 pathway,
as a mechanism of airway hyper-responsiveness character-
istic of asthma.
Sulforaphane can be safely and consistently administered
to humans by feeding a broccoli sprout extract in which its
precursor glucosinolate (glucoraphanin) has been enzymati-
cally hydrolyzed [9]. Sulforaphane has very low toxicity. Its
administration is well tolerated and it is extensively con-
sumed as a component of cruciferous vegetables. It may be
considered a food, a dietary supplement, or a drug, depend-
ing on dosage and its intended use [9, 20, 21].
It has been known for more than 20 years that deep
inspiration (DI) protects against MCh induced bronchial
constriction. This protective response is characteristic-
ally absent in even mild asthmatic patients [22, 23], and
indeed, DI can even exacerbate the bronchoconstrictor
effects of MCh in asthmatics [24]. The beneficial effects
of DIs occurring before or after MCh challenge have
been designated bronchoprotection (BP) and broncho-
dilation (BD), respectively [22, 25]. Elucidation of the
pathways leading to the loss of the BP and BD responses
in asthma may give further insight into the disease
process and reveal new treatments.
The current study was designed with the primary ob-
jective to test the hypothesis that sulforaphane could
augment the deep inspiration (DI)-induced bronchopro-
tection (BP) and bronchodilation (BD) responses in indi-
viduals with airways hyperresponsiveness. Secondary
objectives included testing the hypotheses that sulfo-
raphane would also affect pulmonary function and lung
morphology as shown by high resolution CT scans.
Methods
Screening
The protocol was approved by the Johns Hopkins
IRB and written informed consent was obtained
(NA_00011275) and registered at ClinicalTrials.gov
(NCT00994604). A total of 51 individuals with airway
hyperresponsiveness were screened who reported
upper or lower respiratory symptoms (or both) in the
absence of upper respiratory infections in the previous
12 months. Five subjects withdrew, and one subject
failed the screen. Subjects were also required to have
FEV
1
70 % of predicted values, and a positive con-
ventional multi-dose methacholine (MCh) inhalation
challenge (PC
20,
provocative concentration causing a
20 % drop in FEV
1
)of25 mg/ml, as well as a BP ef-
fect of DI of less than 40 %. Short-acting or long-
acting bronchodilators were withheld before MCh
challenge for 12 or 48 hours, respectively. The demo-
graphic and baseline pulmonary functions of smokers
(n = 15) and non-smokers (n = 30) were similar (Table 1).
Thirty-one subjects were taking albuterol, 13 were tak-
ing fluticasone/salmeterol, 8 were taking fluticasone
alone, 3 were taking montelukast, 1 was taking beclo-
methasone, 1 was taking budesonide/formoterol, 3
were taking fluticasone nasal spray, 4 were taking lora-
tadine, 1 was taking fexofenadine, 1 was taking cetiri-
zine, and 1 was taking benedryl. All participants were
at least 45 days past the end of their most recent URI
for all of their visits, and were asked to withhold inhaled
bronchodilators prior to each study visit: 12 hours for
short-acting and 48 hours for long-acting beta agonists.
Full analysis was possible in 44 individuals. Exclusion cri-
teria included persistent respiratory symptoms suggesting
uncontrolled asthma.
Brown et al. Respiratory Research (2015) 16:106 Page 2 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Pulmonary function measurements
All subjects underwent baseline spirometry, body-
plethysmography, diffusing capacity of the lung for carbon
monoxide, and exhaled nitric oxide (eNO) measurements.
For spirometry, we used Hankinson's (NHANES III) pre-
dicted values [26]. We further made the widely accepted
assumption that the reproducibility of FEV
1
values was
0.1 L [27, 28], so that x-axis changes in Fig. 2a and b be-
tween 0.1 and +0.1 L were considered as no effect. In
addition, blood was obtained before and immediately after
sulforaphane administration to monitor standard blood
chemistries for safety (n = 45) and to measures mRNA ex-
pression of Nrf2 target genes in peripheral blood mono-
nuclear cells (n = 35).
Evaluation of beneficial effects of deep inspiration (DI)
DI-induced bronchodilation was performed by using
multiple modified single dose MCh challenges on separ-
ate days [22, 25, 29]. Briefly, after baseline spirometry, at
every single dose challenge, study participants were
instructed to abstain from DI for 20 min. At the end of
this period, a single dose MCh challenge (starting at
0.025 mg/ml) was delivered with five tidal inspirations
from a deVilbiss 646 nebulizer attached to a model 2A
Rosenthal-French dosimeter (Laboratory for Applied Im-
munology, Fairfax, VA). Three minutes later, a single full
spirometric maneuver was performed and the degree of
airways obstruction was calculated by comparing base-
line to post MCh FEV
1
. If the MCh-induced reduction
in FEV
1
was less than 20 %, the participant was asked to
return on a separate day for another single dose MCh
challenge, using the next highest single dose of MCh
(e.g. 0.075 mg/ml). This process was continued with
additional single dose challenges (0.25, 0.75, 2.5,
7.5 mg/ml) on separate days, until the single dose in-
ducing 20 % reduction in FEV
1
was achieved. For the
challenge at this level, where 20 % or greater reduction
in FEV
1
was obtained, the participant was instructed
to continue the procedure by taking 4 DI immediately
after the single post-MCh spirometry. Another spiro-
metric maneuver was performed immediately after the
4 DI to calculate the degree to which the participant
was able to reverse the MCh-induced airway obstruc-
tion (Figure below).
Measuring the difference between the post-MCh FEV
1
and the FEV
1
obtained after the 4 DI, we calculated a
measure of bronchodilation induced by the DI, which
we termed the bronchodilation (BD) index. This meas-
ure is calculated as follows:
BD index ¼ð1ðð1ððFEV1af ter MCh and af ter DIÞ
ðFEV1baselineÞÞÞ  ð1ððFEV1af ter MChÞ
ðFEV1baselineÞÞÞÞÞ  100
Essentially, the BD index is derived from two compo-
nents, the reduction in FEV
1
from baseline after MCh
and after DI and the reduction in FEV
1
from baseline
after MCh, but before DI.
On a separate day, the same single MCh dose used to
achieve a 20 % or greater reduction in FEV
1
was again
Table 1 Baseline demographics and pulmonary functions (mean ± SD) of 45 asthmatic patients
Non-smoker Smoker All
Number of subjects (n) 30 15 45
Male/female 10/20 9/6 19/26
Age (mean ± SD) 38 ± 14 40 ± 13 38 ± 13
Smoking (pack-years) 0 11 ± 10 -
Race (black/white/asian) 20/9/1 10/5/0 30/14/1
PC
20
(provocative concentration causing a 20 % drop in FEV
1
, in mg/ml) 2.4 ± 4.2 2.6 ± 2.1 2.4 ± 3.6
FEV
1
(forced expiratory volume in 1-second, in L) 2.7 ± 0.7 2.8 ± 0.8 2.7 ± 0.7
FEV
1
% predicted (forced expiratory volume in 1-second) 87 ± 10 % 91 ± 13 % 88 ± 11 %
FVC % predicted (forced vital capacity) 90 ± 12 % 97 ± 15 % 92 ± 13 %
FEV
1
/FVC 0.80 ± 0.08 0.78 ± 0.01 0.79 ± 0.08
TLC (total lung capacity, in L) 5.0 ± 1.4 5.1 ± 1.2 5.0 ± 1.4
SVC (slow vital capacity, in L) 3.3 ± 1 3.5 ± 0.9 3.4 ± 1.0
FRC (functional residual capacity, in L) 3.0 ± 0.9 3.3 ± 1.5 3.1 ± 1.1
RV (residual volume, in L) 1.7 ± 0.7 1.7 ± 0.8 1.7 ± 0.7
RV/TLC 0.34 ± 0.09 0.33 ± 0.08 0.34 ± 0.09
Diffusing capacity of carbon monoxide (mL/min/mm Hg) 22.5 ± 5.9 22.9 ± 5.4 22.7 ± 5.6
Specific airway resistance, (in kiloPascals · s) 6.1 ± 3.4 6.0 ± 3.4 6.04 ± 3.3
Exhaled nitric oxide (ppb) 10.4 ± 11 6.15 9.1 ± 10
Brown et al. Respiratory Research (2015) 16:106 Page 3 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
administered after 20 min of quiet breathing, followed by 5
DI to prevent MCh-induced obstruction (Figure below).
Assessing the difference between the MCh-induced re-
duction in FEV
1
on the day no DI were taken to the day
on which 5 DI were taken prior to the challenge, we ob-
tained a measure of bronchoprotection induced by the
series of DI, which we termed the bronchoprotection
(BP) index which has been previously described [22, 29].
This measure is calculated as follows:
BP index ¼ð1ðð1ððFEV1af ter DI s and af ter MChBÞ
ðFEV1baselineBÞÞÞ  ð1ððFEV1af t er MChAÞ
ðFEV1baselineAÞÞÞÞÞ  100
The BP index is derived from two components, the
MCh-induced reduction in FEV
1
from baseline on the day
that 5 DI preceded the single-dose MCh challenge and the
MCh-induced reduction in FEV
1
from baseline on the day
no DI were taken before the single-dose MCh challenge.
Acquisition and analysis of CT scans
All scans were performed with a single spiral CT scanner
(Siemens, Definition 64) with settings of 120 Kilovolt peak.
mAs (milliampere · second) was based on body size (small
= 80 mAs, medium = 100 mAs, large = 145 mAs), with a ro-
tation time of 0.5 s, pitch of 1.0 mm, thickness of 0.75 mm,
and interval of 0.5 mm. Images were reconstructed using a
B35 and B31 algorithm. All subjects were coached and
practiced the breathing maneuvers before scanning. For the
total lung capacity scans, while in the scanner, all subjects
were instructed to take a deep breath and blow it out. This
maneuver was repeated three times. On the third deep in-
spiration, the subjects were instructed to hold their breath.
They were then coached to continue to keep holding their
breath for the <10 s duration of the scan. For the functional
residual capacity (FRC) scans, all subjects were similarly
instructed to take a breath in and blow it out. This maneu-
ver was repeated three times. On the third time, the subject
was instructed to expel their breath and then hold their
breath for the duration of the scan (<10 s). Lung volumes
and airway dimensions were calculated using PW software
(VIDA Diagnostics, Inc. Coralville, IA) based on the lung
CT scans. The PW software calculates the total lung volume,
the lung air volume, the lung tissue volume, the lung density
in Hounsfield Units (HU), air trapping (% voxels < 856
HU), the luminal diameter, the wall thickness and the wall
fraction (wall area/total airway area). In addition, the airways
were arbitrarily divided into three groups (small, medium,
and large) of a similar number of airways per group.
Preparation and administration of sulforaphane-rich
broccoli sprout extract
The sulforaphane-rich broccoli sprout extract was prepared
at the Lewis B. and Dorothy Cullman Chemoprotection
Brown et al. Respiratory Research (2015) 16:106 Page 4 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Center at Johns Hopkins University, essentially as previ-
ously described [30, 31]. We obtained IND approval from
the FDA for its use in this study (IND #79274). In brief,
specially selected BroccoSproutsseeds were surface-
disinfected and grown for 3 days in a commercial sprouting
facility under controlled light and moisture. A boiling water
extract was prepared, filtered, cooled, and treated with the
enzyme myrosinase (from daikon sprouts) in order to con-
vert precursor glucosinolates to isothiocyanates, and then
lyophilized at a food processing facility (Oregon Freeze Dry,
Albany, OR). The lyophilized powder (227 μmol sulforaph-
ane/g powder) was packaged in sterile plastic tubes that
were unitized for daily doses by ALFA Specialty Pharmacy
(Columbia, MD); each tube contained 100 μmol sulforaph-
ane (440 mg of BSE). The powders (bulk and individual
doses) were maintained at 20 °C, repeatedly checked for
microbial contaminants and sulforaphane titer before con-
veyance to the study site to be dispensed to patients.
Study subjects were given both verbal and written in-
structions to avoid eating any food products that contain
the following vegetables (either cooked or raw) until com-
pletion of the study: Cruciferous vegetables (such as broc-
coli, broccolini, broccoli raab, rapini, kale, cabbage, brussels
sprouts, cauliflower, arugula, turnips, radish, turnip, turnip
greens, kohlrabi, rutabaga, mustard greens, collard greens,
chinese cabbage, pak choi, bok choi, napa, watercress, broc-
coli sprouts, daikon, sauerkraut, coleslaw), vegetables in the
onion family (onions, leeks, garlic, or chives), and the fol-
lowing condiments: mustard, horseradish, wasabi, soy
sauce, or Worcestershire sauce.
mRNA gene expression
Nrf2regulated antioxidative genes were measured in per-
ipheral blood mononuclear cells with quantitative real-time
PCR (qRT-PCR). Total RNA was extracted from the cells
by use of the Qiagen RNeasy kit (Qiagen, Valencia, CA).
Total RNA was used for cDNA synthesis with random hex-
amers and MultiScribe reverse transcriptase, according to
the manufacturers recommendations (Applied Biosystems).
cDNA (100 ng) was used for quantitative PCR analyses of
selected genes: glutamate cysteine ligase catalytic (GCLC)
and modifier subunits (GCLM), NAD()H-quinone oxidore-
ductase 1 (NQO1), and glutathione S-transferase-1 (GST1)
by using primers and probe sets commercially available
from Applied Biosystems. Assays were performed by using
the ABI 7000 Taqman system (Applied Biosystems). β-
Actin was used for normalization.
After the baseline assessment of BP and BD, spirom-
etry, body-plethysmography, diffusion capacity of carbon
monoxide, exhaled NO, and CT scan measurements
were completed (Fig. 1), volunteers received a 1-week
supply of 20-ml bottles containing 100 μmol of sulfo-
raphane each dissolved in mango juice, which masks the
pungency of the BSE [20], to be stored in a freezer. Each
morning, a single bottle was allowed to thaw at room
temperature until consumption of the contents in the
evening. The subject then received a second 7-day sup-
ply from the laboratory, and the procedure was repeated.
At the end of this period, BP, BD, CT scan, spirometry,
body plethysmography, diffusion capacity of carbon
monoxide, and exhaled NO were re-assessed within
3 days of consumption of the last bottle (Fig. 1). In total,
the subjects consumed 100 μmol of sulforaphane per
day on 14 consecutive evenings.
Safety and adverse event monitoring
Adverse event monitoring and documentation by severity,
duration, and relatedness were performed by the study
Fig. 1 Time line of trial protocol
Brown et al. Respiratory Research (2015) 16:106 Page 5 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
physician at each visit. Standard laboratory chemistries
were drawn at baseline and after the two weeks of con-
suming sulforaphane. Study stopping rules were based on
the standard practice of our local IRB and the FDA. Study
drug safety and adherence to the protocol were monitored
annually by the IRB.
Statistics
Data analysis was performed with JMP 11.0.0 software
(SAS Institute, Cary, NC). To compare the demographic
and baseline pulmonary function data, one-way ANOVA
and Chi-squared tests were used. To compare the effects
of sulforaphane on changes in BP and BD, lung function,
CT measurements, oxidative stress genes, and effects of
smoking, matched-pairs comparisons (pre- and post-
sulforaphane) with non-parametric Wilcoxon Sign Rank
Analyses were performed. In addition, one-way ANOVA,
and simple linear regressions were used where indicated.
To examine the effects of specific genes and changes in
FEV
1
interaction on the BP, we constructed a multivariate
regression model. Significance was assumed at p 0.05.
Results
The time-line of the protocol for this study is presented
in Fig. 1.
Screening and selection of asthmatic subjects
A total of 51 individuals with airway hyperresponsive-
ness were screened. The demographic and baseline pul-
monary functions as shown in Table 1. Full analysis was
possible in 44 individuals.
Primary outcomes
The primary goal of these studies was to determine
whether sulforaphane administration to 44 asthmatic pa-
tients: (i) affected the magnitude of the bronchoconstric-
tor effect of single doses of MCh challenge, and (ii)
affected the protective effects of deep inspiration on the
magnitude of the bronchoconstrictor effect of MCh
challenge. The final metric in all these studies was FEV
1.
The mean reduction in FEV
1
resulting from a single
dose MCh challenge was 28.7 ± 7.2 % (mean ± SD)
(Table 2), but individual asthmatics varied markedly in
their responses to sulforaphane (Fig. 2a and b). Whereas
in 60 % of the asthmatics sulforaphane blocked the
bronchoconstrictor effects of MCh challenge, in 20 % of
these subjects sulforaphane aggravated the bronchocon-
strictor effect of MCh, and in a similar proportion (20 %)
it had no effect.
When the change in FEV
1
response to MCh was
analyzed post hoc as either a decrease or an increase
in response to sulforaphane administration, the result-
ing BP and BD responses were dramatically different.
Those subjects whose FEV
1
response to MCh de-
creased (13 ± 8 %), had a concomitant reduction in
BP response (p = 0.002; Table 3), whereas those sub-
jects whose FEV
1
response to MCh increased (11 ±
11 %), experienced an improvement in BP (p = 0.004,
Table 3). Changes in BD paralleled these changes in
BP, (p = 0.02 for the subjects whose FEV
1
response to
MCh decreased and p = 0.04 for the subjects whose
FEV
1
response to MCh increase; Table 3). The impli-
cation of these paradoxical responses is not yet clear
and the mechanism is not understood.
Secondary outcomes
Secondary outcomes examined changes in pulmonary
function (Table 4) and lung morphology as determined
by high-resolution CT scans (Table 5). We found a sig-
nificant reduction in specific airway resistance (p = 0.03,
Table 4), and a small but significant increase in the small
and medium airway luminal area (p = 0.03, Table 5).
Gene-dose effects
Peripheral blood mononuclear cells were obtained from
35 subjects for the measurement of Nrf2regulated anti-
oxidant genes, and we were able to compare 28 sample
Table 2 Effects of sulforaphane (SF) administration and deep inspiration on A. Changes in forced expiratory volume in 1-second
(FEV
1
) produced by the bronchoconstrictor MCh, and B. Magnitudes of bronchoprotection (BP) and bronchodilation (BD) effects
A. Treatment % Change in FEV
1
(mean ± SD)
a
Before SF After SF p for SF effect
MCh 28.7 ± 7.2 22.7 ± 12.7 0.006
MCh preceded by 4 deep inspirations (for BP)23.0 ± 13.4 19.1 ± 11.9 0.04
MCh followed by 4 deep inspirations (for BD)18.4 ± 7.7 16.3 ± 10.7 0.34
B. Magnitudes of deep inspiration effects Change in BP and BD (mean ± SD)
Before SF After SF p for SF effect
BP 15.5 ± 52.7 10.3 ± 57.1 0.83
BD 32.5 ± 32.7 26.7 ± 30.6
a
0.33
a
One value was outside the acceptable range (>2 S.D.) and has been censored
Brown et al. Respiratory Research (2015) 16:106 Page 6 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
pairs before and after sulforaphane treatment. When
gene activities were classified according to the changes
in FEV
1
(either a decrease or an increase response to
MCh after sulforaphane administration), the results were
very different. When the FEV
1
response to MCh
decreased after sulforaphane administration, the activ-
ities of all three Nrf2-regulated genes (NQO1, HO1,
GCLM) decreased. In contrast, when the FEV
1
response
to MCh increased after sulforaphane administration, the
gene activities of all three genes increased (Table 6).
Comparing the changes in each gene expression with
changes in FEV
1
response to sulforaphane treatment, the
differences in gene expression was significant for GCLM
(p = 0.03) and NQO1 (p = 0.047).
Bronchoprotection (BP), bronchodilation (BD), and gene
expression
The BP and BD responses were also analyzed according
to changes in gene expression (either a decrease or an
increase resulting from sulforaphane administration).
There was a significant relationship between the change
in NQO1 gene expression and BP (p = 0.03). When
NQO1 decreased after sulforaphane administration, the
BP worsened (54.4 ± 78.3 %) and when NQO1 in-
creased after sulforaphane administration, the BP im-
proved (20.6 ± 72.8 %). There were no statistically
significant relationships for the other genes or for BD.
We next examined the potential interactions among
these factors. A multivariate regression model was con-
structed using the change in BP as the outcome variable.
Independent variables were the changes in MCh-
induced decrease in FEV
1
(the difference between base-
line before and after sulforaphane treatment), and the
change in GCLM, GST1, and NQO1 gene expression.
The overall model was significant (r
2
= 0.67, p = 0.0003).
Controlling for the other variables, there was a signifi-
cant negative correlation between the change in BP and
the change in FEV
1
from baseline (p = 0.002), and a sig-
nificant positive correlation between the change in BP
and the change in NQO1 gene expression (p = 0.007).
Bronchoprotection, bronchodilation, and smoking history
When BD and BP responses in the non-smokers and
smokers were partitioned by the change in FEV
1
re-
sponse to MCh, they mirrored the overall changes. In
those subjects whose FEV
1
response to MCh de-
creased whether they were non-smokers or smokers,
there was a concomitant reduction in BP and BD re-
sponses (Table 7). Conversely, subjects whose FEV
1
re-
sponse to MCh increased whether they were non-
smokers or smokers, showed improvement in BP and
BD values (Table 7).
Safety and tolerance
Sulforaphane treatment was safe and well-tolerated. In-
spection of the laboratory results among all the subjects
showed that only one set of laboratory values in one
subject showed a significant liver function (transamin-
ase) elevation after sulforaphane administration. On
-1.5 -1.0 -0.5 0.0 0.5 1.0 1.5
-100
-50
0
50
100
150
Effects of SF on the change in FEV1 (L) caused by MCh
Effects of SF on BD (% change)
-1.5 -1.0 -0.5 0.0 0.5 1.0 1.5
-250
-200
-150
-100
-50
0
50
100
150
Effects of SF on the change in FEV1 (L) caused by MCh
Effects of SF on BP (% change)
b
Fig. 2 Negative correlation between the changes (%) resulting from
sulforaphane (SF) administration on: a. bronchodilation (BD) and on
b. bronchoprotection (BP) in asthmatic subjects, and the effects of
sulforaphane administration on the reduction of FEV
1
caused by
MCh challenge. a: There was a significant negative correlation
between the changes in BD and the changes in FEV
1
(r
2
= 0.13,
p = 0.01). As the decrease in FEV
1
with MCh challenge (airway
narrowing) became larger with administration of sulforaphane, the
BD response became smaller. b: There was a significant negative
correlation between the changes in BP and the changes in FEV
1
(r
2
= 0.26, p = 0.0005). As the decrease in FEV
1
with MCh challenge
(airway narrowing) became larger with administration of
sulforaphane, the BP response became smaller. The Mean control
FEV
1
was 2.7 ± 0.7 L, and a single MCh challenge caused a reduction
in FEV
1
by 0.78 ± 0.3 L. We further made the widely accepted
assumption that the reproducibility of FEV
1
values was 0.1 L, so that
x-axis changes in Fig. 2a and b between 0.1 and +0.1 L were
considered as no effect
Brown et al. Respiratory Research (2015) 16:106 Page 7 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
further questioning, the subject reported that he had run
a marathon the day before the blood draw after sulfo-
raphane treatment, and this may have been responsible
for changes in liver enzymes. On repeating the blood
chemistries, one week later, all blood chemistries were
within normal limits.
Discussion
To our knowledge, this is the first study that has ex-
amined the effects of the phytochemical sulforaphane
on the response to DI in individuals with asthma.
This study addressed the hypothesis that oxidative
stress and inflammation play roles in the mechanisms
of the beneficial effects of DI. While our a priori pri-
mary outcome was negative, post hoc analysis exam-
ining the groups according to whether they had an
increase or a decrease in FEV
1
response to Mch told
a very different story.
When sulforaphane attenuated the decline in FEV
1
fol-
lowing a single MCh challenge, BP worsened. In con-
trast, if the FEV
1
response to a single MCh challenge
remained the same or increased slightly after sulforaph-
ane treatment, BP improved dramatically. There were
similar changes for the BD responses. In addition when
sulforaphane increased NQO1 gene expression, there
was an associated increase in FEV
1
response to MCh
and independently associated improvement in the BP re-
sponse of DI in this cohort of asthmatics.
It is predictable that there should be a range of sulfo-
raphane responses among individuals, and that they be
related to some other biological phenomenon that we
have yet to understand. For example, we have observed
striking demonstrations of this effect in our clinical
studies of sulforaphane effects on autism spectrum dis-
order behavioral metrics in young men [31], and in clin-
ical evaluation of the effect of sulforaphane on both
aflatoxin and air pollutant metabolites [30, 32].
It is not yet fully explainable (although expected), that
multiple mechanisms could be at play in creating this
range of responses. The literature suggesting a bi-phasic
response, commonly referred to as the U-shaped or J-
shaped curve, or hormesis [33, 34] and well described in
the context of phytochemicals [35], points to many such
examples in which a compound may have a protective
or therapeutic effect at one concentration and a toxic or
detrimental effect at another higher level of exposure.
This is well demonstrated in the case of Nrf2-active
compounds, in particular in their effects on redox sig-
naling, and differential pro- and anti-oxidant activities
that are concentration dependent [36].
A limited number of animal and human studies have
examined the effects of sulforaphane on airway respon-
siveness. Park and colleagues showed that sulforaphane
Table 3 The BP and BD responses to sulforaphane treatment, classified according to initial increased or decreased forced expiratory
volume in 1-second (FEV
1
) response to MCh (mean ± SD). Wilcoxon sign rank test
BP BD
Before SF After SF p-for SF effect Before SF After SF p-for SF effect
Overall FEV
1
(from Table 2) 15.5 ± 52.7 10.3 ± 57 0.82 32.5 ± 32.7 28.2 ± 28 0.33
Decreased FEV
1
response to MCh
(n = 29)
37.6 ± 38.0 0.8 ± 64 0.002 38.3 ± 34.6 23.2 ± 35.1 0.02
Increased FEV
1
response to MCh
(n = 15)
27.2 ± 51.7 31.7 ± 33 0.004 19.6 ± 25.1 33.4 ± 18.6 0.04
Table 4 Change in pulmonary function with sulforaphane (SF) treatment (mean ± SD)
Before SF After SF p for SF effect
FEV
1
% predicted (forced expiratory volume in 1-second) 88 ± 11 % 88 ± 11 % 0.79
FVC % predicted (forced vital capacity) 92 ± 13 % 92 ± 11 % 0.95
FEV
1
/FVC 0.79 ± 0.08 0.77 ± 0.01 0.56
TLC (total lung capacity, in L) 5.0 ± 1.4 4.9 ± 13 0.10
SVC (slow vital capacity, in L) 3.4 ± 1.0 3.3 ± 0.9 0.38
FRC (functional residual capacity, in L) 3.1 ± 1.1 3.1 ± 1.0 0.56
RV (residual volume, in L) 1.7 ± 0.7 1.6 ± 0.7 0.56
RV/TLC 0.34 ± 0.09 0.34 ± 0.1 0.97
Diffusing capacity of carbon monoxide, (mL/min/mm Hg) 22.7 ± 5.6 22.4 ± 5.7 0.46
Specific airway resistance (kiloPascals · s) 6.04 ± 3.3 4.8 ± 2.5 0.008
Exhaled nitric oxide (ppb) 8.8 ± 10 9.3 ± 10.9 0.80
Brown et al. Respiratory Research (2015) 16:106 Page 8 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
inhibits ovalbumin-induced airway inflammation in a
murine model of asthma [37]. They further surmised
that this occurred though suppression of GATA-3 and
SOCS-3 and increased T-bet and SOCS-5 expression.
Whether this also occurs in humans has not been deter-
mined. Nrf2antioxidant response element binding was
reduced in cultured airway smooth muscle cells from pa-
tients with severe asthma compared to similar cells from
patients with moderate asthma or with those from nor-
mal subjects [38]. How this affects the asthmatic re-
sponse in vivo is unknown.
Sulforaphane effects on upper airway inflammation,
and on airway response to challenge, have also been
measured. Oral sulforaphane induced airway Phase 2
antioxidant enzyme expression in a dose-dependent
fashion in exfoliated human upper airway cells [39]. In a
follow-up study, when sulforaphane in the form of a
broccoli sprout extract was administered orally for 4 days
before a diesel exhaust particle challenge, total cell count
in the nasal lavage was reduced by more than 50 % [40].
These studies establish that the oral administration of
sulforaphane is an effective means of inducing cytopro-
tective enzymes in the upper airway and by inference,
probably also in the lower airway. In addition, the doses
in these published studies were similar to those used in
the current study and thus support our choice of dosing
regimen. We chose to administer the sulforaphane for a
longer period of time, 14 days rather than 4 days, in
order to be confident that enough time had elapsed for
substantial enzyme induction.
It is well recognized that pro-inflammatory cytokines,
chemokines, as well as other cell mediators, play import-
ant roles in the allergic inflammatory process in asthma.
Dysfunction of the Nrf2 pathway has also been linked to
severe allergen-driven airway inflammation and hyper-
responsiveness in mice [6]. Disruption of the Nrf2 sig-
naling pathway in mouse models has also been directly
linked to chronic inflammation such as that associated
with asthma [6]. As a potent inducer of cytoprotective
enzymes (antioxidant and anti-inflammatory) via Nrf2,
sulforaphane thus directly affects both the antioxidant
deficiencies that have been associated with poor asthma
control and accelerated lung function decline [1618],
and the inflammatory component of these conditions
[1214], both of which represent attractive therapeutic
targets.
Upregulation of Phase 2 enzymes and of other Nrf2
targets by sulforaphane has previously been shown in
vitro and in animal models [9, 39, 4143]. The use of
broccoli sprout extract as a vehicle for well-quantified
oral delivery of sulforaphane permitted us to examine
the role of oxidative stress mediated through the Nrf2-
signaling pathways on lung function in asthma. Edible
plants contain a wide variety of phytochemicals, some of
which are phase 2 enzyme inducers. Large quantities of
inducers of enzymes that are potent antioxidants can be
delivered in the diet by small quantities of broccoli
sprouts (e.g., three-day-old sprouts) that contain as
much inducer activity as 10100 times larger quantities
of mature broccoli. A well-characterized, stable, clean,
standardized, homogeneous, sulforaphane-rich food ex-
tract can be produced and utilized across a range of cell
culture, animal, and clinical studies, permitting direct
comparison of results. Thus, a large body of evidence
has accumulated around the world in a variety of experi-
mental systems suggesting that sulforaphane is the active
agent in broccoli sprout extracts responsible for almost
all of the phase 2 response that is induced following
treatment with broccoli sprout extracts. Quantifiable
and reproducible extraction of sulforaphane from BSE
has thus permitted us to target specifically the Nrf2
pathway in order to elucidate the mechanisms of oxida-
tive stress in patients with asthma.
Table 5 CT scan measurements of changes in volume, luminal
area and airway wall thickness with sulforaphane (SF) treatment
measured at either total lung capacity (TLC) or functional
residual capacity (FRC) (mean ± SD)
Before SF After SF p for SF effect
TLC lung volume (mL) 4175 4195 0.68
FRC lung volume (mL) 1470 1647 0.44
Mean lung density
at TLC (HU)
820 ± 34 823 ± 33 0.13
Air trapping (%) (FRC) 4.6 ± 8.3 7.9 ± 11.8 0.10
Airway luminal area
(TLC) mm
2
44.7 ± 10.3 45.1 ± 8.5 0.07
Large 103 ± 13 102.9 ± 16 0.73
Medium 30.9 ± 1.8 32 ± 4.5 0.04
Small 13.1 ± 1.6 14.3 ± 3.4 0.01
Airway wall thickness
(fraction luminal diameter)
(TLC)
0.57 ± 0.03 0.57 ± 0.02 0.82
Large 0.45 ± 0.02 0.46 ± 0.03 0.12
Medium 0.58 ± 0.02 0.58 ± 0.03 0.96
Small 0.67 ± 0.02 0.66 ± 0.04 0.03
Table 6 Change in oxidative stress gene expression related to
changes in Forced Expiratory Volume in 1 second (FEV
1
)in
response to single dose MCh challenge (mean ± SD). P-value for
pre vs. post sulforaphane
Gene Expression (% change from baseline)
GST1 NQO1 GCLM
Decreased FEV
1
response
to MCh
6.11 ± 27.0 73.1 ± 255.4 19.2 ± 40.6
p = 0.67 p = 0.23 p = 0.03
Increased FEV
1
response
to MCh
11.8 ± 56.0 141.9 ± 283.7 32.4 ± 75.0
p = 0.38 p = 0.047 p = 0.58
Brown et al. Respiratory Research (2015) 16:106 Page 9 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Few studies have examined the consumption of foods
rich in antioxidants in general and sulforaphane specific-
ally, on airway reactivity and asthma. Early studies sug-
gested that changes in the diets of children was a
determinant of the worldwide increases in asthma and
allergies [44], and that infrequent consumption of fresh
fruit was associated with impaired lung function [45].
More recently, intake of some vegetables was also asso-
ciated with reduced prevalence of adult asthma [46].
Commonly consumed fruits, vegetables, and nuts, and
high adherence to a traditional Mediterranean diet dur-
ing childhood were shown to have beneficial effects on
symptoms of asthma and rhinitis [47]. Adherence to a
Mediterranean diet during pregnancy also had a protect-
ive effect against asthma-like symptoms and atopy in
childhood [48]. An intervention that administered a food
plant, (Moringa oleifera) known to be rich in glucosino-
lates similar to the precursor of sulforaphane, to 20 asth-
matic subjects daily for 3 weeks, significantly improved
symptom severity, which was accompanied by improve-
ments in FEV
1
, FVC, and peak expiratory flow rate [49].
Most impressively, a new meta analysis of 12 cohort, 4
population-based casecontrol, and 26 cross-sectional
studies came to the conclusion that reduced risk of
asthma in adults and children was associated with higher
intake of fruit and vegetables (RR = 0.54; 95 % CI, 0.41-
0.69) [50].
The relationship between DI, airway mechanics, and air-
way hyperresponsiveness has been a focus of asthma re-
search for over 2 decades [24]. Several earlier studies have
shown that DI has beneficial effects on human airways
[51, 52]. In healthy individuals, DI taken before exposure to
MCh protected the airways from bronchoconstriction [53].
This property is largely lost in asthma, even in mild disease
(5). Deep inspirations can also reverse airway obstruction
that has been experimentally induced with a direct spasmo-
gen [54]. The bronchodilatory effect of DI is minimally af-
fected in mild asthmatics (5), but decreases with increasing
severity of asthma [55], raising the possibility that the
impairment of this physiologic function of the lung is one
of the determinants of severe obstructive disease.
Our findings are consistent with previous demonstra-
tions that BP and BD effects of DI become more prom-
inent when the MCh-induced bronchoconstriction in
the absence of DI is of substantial magnitude [25]. Our
findings are also consistent with the dependence of the
effects of DI on the magnitude of bronchoconstriction
(Fig. 2). When the responses of our subjects were evalu-
ated by group: (subjects with either a decrease or an in-
crease in response to MCh), the BD and BP outcomes
were dramatically different (Table 3). To our knowledge,
this is the first demonstration of a decrease in response
to MCh leading to a worsening of the BD and BP re-
sponses and not simply a diminution of the response
amplitude [25]. Why this occurred and whether the ef-
fect of sulforaphane on the oxidative stress balance was
involved will require further exploration.
Among nonsmokers, both BP and BD decreased in
those subjects with a decreased FEV
1
response to MCh,
(Table 7). In contrast, if they had no change, or any in-
crease in their FEV
1
responses to MCh, both BD and BP
improved significantly. There were similar trends for
smokers, but the changes in BD for subjects with a re-
duced FEV
1
response to MCh was significantly worse
than those who had non- or marginal FEV
1
responses to
MCh. Since changes were of similar magnitude in both
smokers and nonsmokers, the difference in the signifi-
cance of that effect may reflect the lower number of
smokers enrolled in the study. Based on animal studies,
a greater effect of sulforaphane on BD and BP would be
expected in smokers. Stimulation of Nrf2 pathways has
been shown to protect against the harmful effects of
cigarette smoke (which releases large quantities of free
radicals and increases lung levels of reactive oxygen and
nitrogen species (ROS and RNS) in murine models [56].
Among all the subjects, there was a significant de-
crease in specific airway resistance (Table 4) with the ad-
ministration of sulforaphane. This is likely explained by
Table 7 BP and BD responses to sulforaphane (SF) arranged according to changes in forced expiratory volume in 1-second (FEV
1
)to
MCh (mean ± SD). A. Non-smokers (n = 30), B. Smokers (n = 15).
A. Non-smokers
BP (% change) BD (% change)
Before SF After SF p for SF effect Before SF After SF p for SF effect
Decreased FEV
1
response to MCh 40.4 ± 40.4 3.8 ± 46.8 0.002 33.7 ± 39.4 23.8 ± 36.8 0.20
Increased FEV
1
response to MCh 49.3 ± 29.7 23.5 ± 35.8 0.004 17.7 ± 27.0 36.3 ± 17.7 0.04
B. Smokers
BP (% change) BD (% change)
Before SF After SF p for SF effect Before SF After SF p for SF effect
Decreased FEV
1
response to MCh 32.3 ± 34.2 5.1 ± 91.1 0.28 47.1 ± 21.8 22.1 ± 33.4 0.06
Increased FEV
1
response to MCh 17.1 ± 60.7 48.1 ± 20.6 0.44 22.3 ± 23.1 27.4 ± 20.1 0.63
Brown et al. Respiratory Research (2015) 16:106 Page 10 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
the concomitant increase in small and medium airway
luminal area (Table 5) after sulforaphane administration.
While these changes did not translate into an improve-
ment in the more commonly used lung function mea-
surements such as FEV
1
, FVC, and FEV
1
/FVC, it does
suggests an overall beneficial effect of sulforaphane on
the airways of asthmatic individuals.
In conclusion, administration of sulforaphane, a potent
upregulator of genes protecting against oxidative stress
and inflammation, may be useful both to determine the
mechanisms that lead to asthma, as well as suggesting a
potential therapy to treat asthma. Sulforaphane adminis-
tration improved the BP response in asthmatics who had
an increase in NQO1 gene expression and did not have a
decrease in their initial response to the MCh challenge.
Therefore, sulforaphane administration was able to im-
prove a major defect of even mild asthma. This should en-
courage further examination of this major cytoprotective
signaling pathway as a potential mechanistic approach to
the treatment or prevention of asthma. Furthermore, since
the administration of anti-inflammatory drugs do not
completely prevent the symptoms or progression of the
disease in a substantial subset of asthmatics, these findings
also raise the possibility of the use of sulforaphane, or
foods rich in sulforaphane, as potential adjuvant treat-
ments for asthma.
Abbreviations
BD: Bronchodilation; BP: Bronchoprotection; BSE: Broccoli sprout extract;
DI: Deep inspiration; ERV: Expiratory reserve volume; FEV
1
: Forced expiratory
volume in 1 second; FRC: Functional residual capacity; GST: Glutathione
transferase; GCLM: Glutamine-cysteine ligase, regulatory subunit; IC: Inspiratory
capacity; MAPK: Mitogen-activated protein kinase; MCh: Methacholine;
NfκB: Nuclear factor kappa-light-chain-enhancer of activated B cells;
NQO1: NAD(P)H, Nicotinamide-quinone oxidoreductase 1; Nrf2: Nuclear factor
erythroid-related factor 2; PBMC: Peripheral blood mononuclear cells;
PC
20
: Provocative concentration causing a 20 % drop in FEV
1
; RV: Residual
volume; sRAW: specific airway resistance; SVC: Slow vital capacity; TGV: Thoracic
gas volume; TLC: Total lung capacity; TVC: Total vital capacity.
Competing interests
The authors declare that they have no competing interests.
Authorscontributions
RHB designed the study and analyzed the data. RHB, PT, and JWF wrote the
paper. JWF and PT prepared the sulforaphane-rich broccoli sprout extract.
RHB, CR and AB carried out the clinical and laboratory studies. All authors
read and approved the final manuscript.
Author details
1
Department of Anesthesiology and Critical Care Medicine, Johns Hopkins
University School of Medicine, Baltimore, MD, USA.
2
Division of Pulmonary
Medicine and Critical Care, Department of Medicine, Johns Hopkins
University School of Medicine, Baltimore, MD, USA.
3
Department of
Radiology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
4
Lewis B. and Dorothy Cullman Chemoprotection Center, Department of
Pharmacology and Molecular Sciences, Johns Hopkins University School of
Medicine, Baltimore, MD, USA.
5
Department of Environmental Health
Sciences, Johns Hopkins University School of Public Health, Room E7614,
615 N. Wolfe Street, Baltimore, MD 21205, USA.
6
Center for Human Nutrition,
Department of International Health, Johns Hopkins University School of
Public Health, Baltimore, MD, USA.
Received: 19 May 2015 Accepted: 21 July 2015
References
1. Adcock IM, Barnes PJ. Molecular mechanisms of corticosteroid resistance.
Chest. 2008;134:394401.
2. Drazen JM. Asthma: the paradox of heterogeneity. J Allergy Clin Immunol.
2012;129:12001.
3. Allen S, Britton JR, Leonardi-Bee JA. Association between antioxidant
vitamins and asthma outcome measures: systematic review and
meta-analysis. Thorax. 2009;64:6109.
4. Comhair SA, Erzurum SC. Redox control of asthma: molecular mechanisms
and therapeutic opportunities. Antioxid Redox Signal. 2010;12:93124.
5. Patel BD, Welch AA, Bingham SA, Luben RN, Day NE, Khaw KT, et al. Dietary
antioxidants and asthma in adults. Thorax. 2006;61:38893.
6. Rangasamy T, Guo J, Mitzner WA, Roman J, Singh A, Fryer AD, et al.
Disruption of Nrf2 enhances susceptibility to severe airway inflammation
and asthma in mice. J Exp Med. 2005;202:4759.
7. Ghezzi P. Role of glutathione in immunity and inflammation in the lung.
Int J Gen Med. 2011;4:10513.
8. Hayes JD, Dinkova-Kostova AT. The Nrf2 regulatory network provides an
interface between redox and intermediary metabolism. Trends Biochem
Sci. 2014;39:199218.
9. Kensler TW, Egner PA, Agyeman AS, Visvanathan K, Groopman JD, Chen JG,
et al. Keap1-nrf2 signaling: a target for cancer prevention by sulforaphane.
Top Curr Chem. 2013;329:16377.
10. Baird L, Dinkova-Kostova AT. The cytoprotective role of the Keap1-Nrf2
pathway. Arch Toxicol. 2011;85:24172.
11. Liu H, Dinkova-Kostova AT, Talalay P. Coordinate regulation of enzyme
markers for inflammation and for protection against oxidants and
electrophiles. Proc Natl Acad Sci U S A. 2008;105:1592631.
12. Chen XL, Dodd G, Kunsch C. Sulforaphane inhibits TNF-alpha-induced
activation of p38 MAP kinase and VCAM-1 and MCP-1 expression in
endothelial cells. Inflamm Res. 2009;58:51321.
13. Kundu JK, Choi KY, Surh YJ. Beta-Catenin-mediated signaling: a novel
molecular target for chemoprevention with anti-inflammatory substances.
Biochim Biophys Acta. 2006;1765:1424.
14. Brown KK, Blaikie FH, Smith RA, Tyndall JD, Lue H, Bernhagen J, et al. Direct
modification of the proinflammatory cytokine macrophage migration
inhibitory factor by dietary isothiocyanates. J Biol Chem. 2009;284:3242533.
15. Dinkova-Kostova AT, Talalay P. Direct and indirect antioxidant properties of
inducers of cytoprotective proteins. Mol Nutr Food Res. 2008;52 Suppl 1:S12838.
16. Riedl MA, Nel AE. Importance of oxidative stress in the pathogenesis and
treatment of asthma. Curr Opin Allergy Clin Immunol. 2008;8:4956.
17. Nadeem A, Chhabra SK, Masood A, Raj HG. Increased oxidative stress and
altered levels of antioxidants in asthma. J Allergy Clin Immunol.
2003;111:728.
18. Ochs-Balcom HM, Grant BJ, Muti P, Sempos CT, Freudenheim JL, Browne
RW, et al. Antioxidants, oxidative stress, and pulmonary function in
individuals diagnosed with asthma or COPD. Eur J Clin Nutr. 2006;60:9919.
19. An SS, Kim J, Ahn K, Trepat X, Drake KJ, Kumar S, et al. Cell stiffness,
contractile stress and the role of extracellular matrix. Biochem Biophys
Res Commun. 2009;382:697703.
20. Fahey JW, Talalay P, Kensler TW. Notes from the field: green
chemoprevention as frugal medicine. Cancer Prev Res (Phila). 2012;5:17988.
21. Fahey JW, Kensler TW. Health span extension through green
chemoprevention. Virtual Mentor. 2013;15:3118.
22. Scichilone N, Permutt S, Togias A. The lack of the bronchoprotective
and not the bronchodilatory ability of deep inspiration is associated
with airway hyperresponsiveness. Am J Respir Crit Care Med.
2001;163:4139.
23. Crimi E, Pellegrino R, Milanese M, Brusasco V. Deep breaths, methacholine,
and airway narrowing in healthy and mild asthmatic subjects. J Appl
Physiol. 2002;93:138490.
24. Skloot G, Permutt S, Togias AG. Airway hyperresponsiveness in asthma: a
problem of limited smooth muscle relaxation with inspiration. J Clin Invest.
1995;96:2393403.
25. Scichilone N, Kapsali T, Permutt S, Togias A. Deep inspiration-induced
bronchoprotection is stronger than bronchodilation. Am J Respir Crit Care
Med. 2000;162:9106.
Brown et al. Respiratory Research (2015) 16:106 Page 11 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
26. Hankinson JL, Odencrantz JR, Fedan KB. Spirometric reference values from a
sample of the general U.S. population. Am J Respir Crit Care Med.
1999;159:17987.
27. Cooper B. Spirometry standards and FEV1/FVC repeatability. Prim Care
Respir J. 2010;19:2924.
28. Fletcher M, Loveridge C. Recommendations on repeatability of spirometry.
Prim Care Respir J. 2010;19:192. author reply 193.
29. Pyrgos G, Scichilone N, Togias A, Brown RH. Bronchodilation response to
deep inspirations in asthma is dependent on airway distensibility and air
trapping. J Appl Physiol. 2011;110:4729.
30. Egner PA, Chen JG, Zarth AT, Ng DK, Wang JB, Kensler KH, et al. Rapid and
sustainable detoxication of airborne pollutants by broccoli sprout beverage:
results of a randomized clinical trial in China. Cancer Prev Res (Phila).
2014;7:81323.
31. Singh K, Connors SL, Macklin EA, Smith KD, Fahey JW, Talalay P, et al.
Sulforaphane treatment of autism spectrum disorder (ASD). Proc Natl Acad
Sci U S A. 2014;111:155505.
32. Wan J, Diaz-Sanchez D. Phase II enzymes induction blocks the enhanced
IgE production in B cells by diesel exhaust particles. J Immunol.
2006;177:347783.
33. Kensler TW, Chen JG, Egner PA, Fahey JW, Jacobson LP, Stephenson KK,
et al. Effects of glucosinolate-rich broccoli sprouts on urinary levels of
aflatoxin-DNA adducts and phenanthrene tetraols in a randomized clinical
trial in He Zuo township, Qidong, People's Republic of China. Cancer
Epidemiol Biomarkers Prev. 2005;14:260513.
34. Fukushima S, Kinoshita A, Puatanachokchai R, Kushida M, Wanibuchi H,
Morimura K. Hormesis and doseresponse-mediated mechanisms in
carcinogenesis: evidence for a threshold in carcinogenicity of non-
genotoxic carcinogens. Carcinogenesis. 2005;26:183545.
35. Cook R, Calabrese EJ. The importance of hormesis to public health.
Environ Health Perspect. 2006;114:16315.
36. Son TG, Camandola S, Mattson MP. Hormetic dietary phytochemicals.
Neuromolecular Med. 2008;10:23646.
37. Filomeni G, Piccirillo S, Rotilio G, Ciriolo MR. p38(MAPK) and ERK1/2 dictate
cell death/survival response to different pro-oxidant stimuli via p53 and
Nrf2 in neuroblastoma cells SH-SY5Y. Biochem Pharmacol. 2012;83:134957.
38. Park JH, Kim JW, Lee CM, Kim YD, Chung SW, Jung ID, et al. Sulforaphane
inhibits the Th2 immune response in ovalbumin-induced asthma. BMB Rep.
2012;45:3116.
39. Michaeloudes C, Chang PJ, Petrou M, Chung KF. Transforming growth
factor-beta and nuclear factor E2-related factor 2 regulate antioxidant
responses in airway smooth muscle cells: role in asthma. Am J Respir Crit
Care Med. 2011;184:894903.
40. Riedl MA, Saxon A, Diaz-Sanchez D. Oral sulforaphane increases Phase II
antioxidant enzymes in the human upper airway. Clin Immunol.
2009;130:24451.
41. Heber D, Li Z, Garcia-Lloret M, Wong AM, Lee TY, Thames G, et al.
Sulforaphane-rich broccoli sprout extract attenuates nasal allergic response
to diesel exhaust particles. Food Funct. 2014;5:3541.
42. Ritz SA, Wan J, Diaz-Sanchez D. Sulforaphane-stimulated phase II enzyme
induction inhibits cytokine production by airway epithelial cells stimulated
with diesel extract. Am J Physiol Lung Cell Mol Physiol. 2007;292:L339.
43. Thimmulappa RK, Mai KH, Srisuma S, Kensler TW, Yamamoto M, Biswal S.
Identification of Nrf2-regulated genes induced by the chemopreventive
agent sulforaphane by oligonucleotide microarray. Cancer Res.
2002;62:5196203.
44. Hijazi N, Abalkhail B, Seaton A. Diet and childhood asthma in a society in
transition: a study in urban and rural Saudi Arabia. Thorax. 2000;55:7759.
45. Cook DG, Carey IM, Whincup PH, Papacosta O, Chirico S, Bruckdorfer KR,
et al. Effect of fresh fruit consumption on lung function and wheeze in
children. Thorax. 1997;52:62833.
46. Romieu I, Varraso R, Avenel V, Leynaert B, Kauffmann F, Clavel-Chapelon F.
Fruit and vegetable intakes and asthma in the E3N study. Thorax.
2006;61:20915.
47. Chatzi L, Apostolaki G, Bibakis I, Skypala I, Bibaki-Liakou V, Tzanakis N, et al.
Protective effect of fruits, vegetables and the Mediterranean diet on asthma
and allergies among children in Crete. Thorax. 2007;62:67783.
48. Chatzi L, Torrent M, Romieu I, Garcia-Esteban R, Ferrer C, Vioque J, et al.
Mediterranean diet in pregnancy is protective for wheeze and atopy in
childhood. Thorax. 2008;63:50713.
49. Agrawal B, Mehta A. Antiasthmatic activity of Moringa oleifera Lam: A
clinical study. Indian J Pharmacol. 2008;40:2831.
50. Seyedrezazadeh E, Moghaddam MP, Ansarin K, Vafa MR, Sharma S,
Kolahdooz F. Fruit and vegetable intake and risk of wheezing and asthma: a
systematic review and meta-analysis. Nutr Rev. 2014;72:41128.
51. Nadel JA, Tierney DF. Effect of a previous deep inspiration on airway
resistance in man. J Appl Physiol. 1961;16:7179.
52. Burns CB, Taylor WR, Ingram Jr RH. Effects of deep inhalation in asthma:
relative airway and parenchymal hysteresis. J Appl Physiol. 1985;59:15906.
53. Kapsali T, Permutt S, Laube B, Scichilone N, Togias A. The potent
bronchoprotective effect of deep inspiration and its absence in asthma.
J Appl Physiol. 2000;89:71120.
54. Skloot G, Togias A. Bronchodilation and bronchoprotection by deep
inspiration and their relationship to bronchial hyperresponsiveness. Clin Rev
Allergy Immunol. 2003;24:5572.
55. Scichilone N, Permutt S, Bellia V, Togias A. Inhaled corticosteroids and the
beneficial effect of deep inspiration in asthma. Am J Respir Crit Care Med.
2005;172:6939.
56. Sussan TE, Rangasamy T, Blake DJ, Malhotra D, El-Haddad H, Bedja D, et al.
Targeting Nrf2 with the triterpenoid CDDO-imidazolide attenuates cigarette
smoke-induced emphysema and cardiac dysfunction in mice. Proc Natl
Acad Sci U S A. 2009;106:2505.
Submit your next manuscript to BioMed Central
and take full advantage of:
Convenient online submission
Thorough peer review
No space constraints or color figure charges
Immediate publication on acceptance
Inclusion in PubMed, CAS, Scopus and Google Scholar
Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit
Brown et al. Respiratory Research (2015) 16:106 Page 12 of 12
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
1.
2.
3.
4.
5.
6.
Terms and Conditions
Springer Nature journal content, brought to you courtesy of Springer Nature Customer Service Center GmbH (“Springer Nature”).
Springer Nature supports a reasonable amount of sharing of research papers by authors, subscribers and authorised users (“Users”), for small-
scale personal, non-commercial use provided that all copyright, trade and service marks and other proprietary notices are maintained. By
accessing, sharing, receiving or otherwise using the Springer Nature journal content you agree to these terms of use (“Terms”). For these
purposes, Springer Nature considers academic use (by researchers and students) to be non-commercial.
These Terms are supplementary and will apply in addition to any applicable website terms and conditions, a relevant site licence or a personal
subscription. These Terms will prevail over any conflict or ambiguity with regards to the relevant terms, a site licence or a personal subscription
(to the extent of the conflict or ambiguity only). For Creative Commons-licensed articles, the terms of the Creative Commons license used will
apply.
We collect and use personal data to provide access to the Springer Nature journal content. We may also use these personal data internally within
ResearchGate and Springer Nature and as agreed share it, in an anonymised way, for purposes of tracking, analysis and reporting. We will not
otherwise disclose your personal data outside the ResearchGate or the Springer Nature group of companies unless we have your permission as
detailed in the Privacy Policy.
While Users may use the Springer Nature journal content for small scale, personal non-commercial use, it is important to note that Users may
not:
use such content for the purpose of providing other users with access on a regular or large scale basis or as a means to circumvent access
control;
use such content where to do so would be considered a criminal or statutory offence in any jurisdiction, or gives rise to civil liability, or is
otherwise unlawful;
falsely or misleadingly imply or suggest endorsement, approval , sponsorship, or association unless explicitly agreed to by Springer Nature in
writing;
use bots or other automated methods to access the content or redirect messages
override any security feature or exclusionary protocol; or
share the content in order to create substitute for Springer Nature products or services or a systematic database of Springer Nature journal
content.
In line with the restriction against commercial use, Springer Nature does not permit the creation of a product or service that creates revenue,
royalties, rent or income from our content or its inclusion as part of a paid for service or for other commercial gain. Springer Nature journal
content cannot be used for inter-library loans and librarians may not upload Springer Nature journal content on a large scale into their, or any
other, institutional repository.
These terms of use are reviewed regularly and may be amended at any time. Springer Nature is not obligated to publish any information or
content on this website and may remove it or features or functionality at our sole discretion, at any time with or without notice. Springer Nature
may revoke this licence to you at any time and remove access to any copies of the Springer Nature journal content which have been saved.
To the fullest extent permitted by law, Springer Nature makes no warranties, representations or guarantees to Users, either express or implied
with respect to the Springer nature journal content and all parties disclaim and waive any implied warranties or warranties imposed by law,
including merchantability or fitness for any particular purpose.
Please note that these rights do not automatically extend to content, data or other material published by Springer Nature that may be licensed
from third parties.
If you would like to use or distribute our Springer Nature journal content to a wider audience or on a regular basis or in any other manner not
expressly permitted by these Terms, please contact Springer Nature at
onlineservice@springernature.com
... Studies demonstrated that sulforaphane distributed to the breast epithelial cells in vivo and exerted pharmacodynamic action in these target cells, consistent with its mechanism of chemoprotective efficacy. Fahey et al. [172] showed that administration of sulforaphane (100 µmol/day on 14 days) improved the bronchoprotection response in asthmatics who had an increase in NQO1 gene expression and did not have a decrease in their initial response to the MCh challenge. Therefore, SFN administration was able to improve a major defect of even mild asthma. ...
... The Staudinger/aza-Wittig tandem reaction was used in the synthesis of enantiomerically pure (R)-SFN ((R)-24) [250] using diacetone-D-glucofuranose (171) (DAG)methodology [251]. The stereoselective synthesis of SFN was based on the reaction of 1-azidobutanesulfinyl chloride (172) with DAG (171), using Hünig's base as a catalyst, and affording the sulfinate ester (S)-173 in a 90% yield and in 94% diastereomeric excess. Reaction of methylmagnesium bromide with the sulfinate ester (S)-173 provided 4-azidobutyl methyl sulfoxide ((R)-174) with inversion of configuration, which, in turn, under the Staudinger/aza-Wittig reaction with triphenylphosphine and carbon disulfide, gave enantiopure (R)-SFN ((R)-24) in a 71% yield. ...
Article
Full-text available
For decades, various plants have been studied as sources of biologically active compounds. Compounds with anticancer and antimicrobial properties are the most frequently desired. Cruciferous plants, including Brussels sprouts, broccoli, and wasabi, have a special role in the research studies. Studies have shown that consumption of these plants reduce the risk of lung, breast, and prostate cancers. The high chemopreventive and anticancer potential of cruciferous plants results from the presence of a large amount of glucosinolates, which, under the influence of myrosinase, undergo an enzymatic transformation to biologically active isothiocyanates (ITCs). Natural isothiocyanates, such as benzyl isothiocyanate, phenethyl isothiocyanate, or the best-tested sulforaphane, possess anticancer activity at all stages of the carcinogenesis process, show antibacterial activity, and are used in organic synthesis. Methods of synthesis of sulforaphane, as well as its natural or synthetic bifunctional analogues with sulfinyl, sulfanyl, sulfonyl, phosphonate, phosphinate, phosphine oxide, carbonyl, ester, carboxamide, ether, or additional isothiocyanate functional groups, and with the unbranched alkyl chain containing 2–6 carbon atoms, are discussed in this review. The biological activity of these compounds are also reported. In the first section, glucosinolates, isothiocyanates, and mercapturic acids (their metabolites) are briefly characterized. Additionally, the most studied anticancer and antibacterial mechanisms of ITC actions are discussed.
... In the pathogenesis of allergic asthma, reduced expression of antioxidant genes is also observed, such as Nrf2. Clinical studies have shown that with the reduction of Nrf2 expression in the body, asthma becomes more serious (21,22), and the application of Nrf2 agonists significantly relaxes the bronchi and improves the symptoms (23), suggesting that the occurrence of allergic asthma is not only related to Th2 inflammatory reaction but also has a close relationship with oxidative stress. ...
Article
Full-text available
Thioredoxin-1 (Trx1) is an important regulator of cellular redox homeostasis that comprises a redox-active dithiol. Trx1 is induced in response to various stress conditions, such as oxidative damage, infection or inflammation, metabolic dysfunction, irradiation, and chemical exposure. It has shown excellent anti-inflammatory and immunomodulatory effects in the treatment of various human inflammatory disorders in animal models. This review focused on the protective roles and mechanisms of Trx1 in allergic diseases, such as allergic asthma, contact dermatitis, food allergies, allergic rhinitis, and drug allergies. Trx1 plays an important role in allergic diseases through processes, such as antioxidation, inhibiting macrophage migration inhibitory factor (MIF), regulating Th1/Th2 immune balance, modulating allergic inflammatory cells, and suppressing complement activation. The regulatory mechanism of Trx1 differs from that of glucocorticoids that regulates the inflammatory reactions associated with immune response suppression. Furthermore, Trx1 exerts a beneficial effect on glucocorticoid resistance of allergic inflammation by inhibiting the production and internalization of MIF. Our results suggest that Trx1 has the potential for future success in translational research.
... This effect could be due to attenuation of oxidative stress and activation of the peroxisome proliferator-activated receptors (PPAR), involved in glucidic and lipid metabolism (Melrose, 2019). Sulforaphane has also been suggested as a potential adjuvant treatment moderate asthmatics patients because of its bronchoprotective response through regulation of the Nrf2 signaling pathway (Brown, Reynolds, Brooker, Talalay & Fahey, 2015). This compound has even been shown to be useful in improving cognitive deficits in patients with mental disorder such as schizophrenia (Shiina et al., 2015). ...
Article
Full-text available
The intake of foods derived from plants has been proposed as an useful strategy in the prevention of several chronic diseases. However, plants also possess a group of substances known as antinutrients, which may be responsible for deleterious effects related to the absorption of nutrients and micronutrients, or exert beneficial health effects. This review compiles scientific evidence regarding the physiological impact of some antinutrients (lectins, goitrogens, phytates and oxalates) in the human health, their negative effects and the culinary and industrial procedures to reduce their presence in foods. It can be concluded that, the effects of antinutrients on human health could change when consumed in their natural food matrix, and after processing or culinary treatment. Accordingly, some of these compounds could have beneficial effects in different pathological conditions. Future research is required to understand the therapeutic potential of these compounds in humans.
... The Cytoprotective Activity of Nrf2 Is Regulated by Phytochemicals. . . Neonatal hypoxiaischemic (HI) brain injury rats Systemic pretreatment with SFN (5 mg / kg) to 7-day-old pups linked to the carotid artery and hypoxia -Reduced the infarct index, the number of TUNEL-positive neurons, the MDA indices, and the Cas-3 activity -Protection mediated by the expression of Nrf2 and HO-1 [70] Human bladder carcinoma RT4 cells and bladder tissue from C57BL/6 mice [wildtype (Nrf2 +/+) and Nrf2-deficient (Nrf2 À/À)] -Determination of the effect of SFN on inhibition of genetic damage induced by 4-aminobiphenyl (ABP, carcinogen present in tobacco smoke) -Were pretreated with SFN at 2 or 4 μm for 24 h and then exposed to ABP (0.5 mm, with S9) for 3 h -SFN induces the activation of Nrf2 to inhibit damage -Activation occurs preferentially in the epithelium -Consuming broccoli can prevent bladder cancer [71] (continued) [77] Hepa1c1c7 cells from murine hepatoma and CD-1 mice -Determine the potency of SFN to induce aldehyde dehydrogenase (ALDH) -Evaluate the effect of feeding CD-1 mice with SFN for 7 days before a single ethanol administration -SFN (5 or 20 μmol/day) for 7 days -The in vitro study confirms that the induction of ALDH is comparable to that of NQO1 dependent on SFN-Nrf2 -In vivo study induced tissue ALDH and doubled the rate of acetaldehyde elimination, reducing its toxicity [78] Mice with ethanol consumption and hepg2 E47 cells expressing the isoenzyme CYP2E1 -Analysis of the ability of SFN to mitigate CYP2E1-dependent ethanol-induced steatosis in vivo and in vitro -SFN at 0.05 g/kg injected daily -In vivo HO-1 and GSH levels increased and lipid peroxidation and 3-nitrotyrosine protein adducts decreased -In vitro, the activity of the CYP2E1 isoenzyme was not altered by treatment with SFN -SFN proved to be an effective agent against ethanol-induced acute fatty liver [79] MC3T3-E1 osteoblast cells -Effect of SFN on dexamethasone (Dex) induced apoptosis -SFN at 20 μm y 10 μm -SFN blocked Dex-induced Cas-3 / Cas-9 -Positive regulation on the expression and function of Nrf2 [80] (continued) Only half of the cases with SFN improved the bronchoconstrictor effect of methacholine (mch) and decreased the antioxidant activity of Nrf2 [84] (continued) ...
... Our new findings in the present study demonstrate that sulforaphane induced the expression of the tra-1 gene that in turn regulates DAF-16 signaling. These findings are consistent with previous studies showing a benefit of sulforaphane in cancer chemoprevention and human health, such as anti-inflammatory and anti-atopic allergic responses (Brown et al., 2015;Bayat Mokhtari et al., 2018;Sidhaye et al., 2019). Conversely, we observed that the loss of tra-1 expression or its function shortened the lifespan and decreased FIGURE 6 | Scheme of sulforaphane-promoted lifespan and healthspan by inducing TRA-1 and DAF-16 nuclear translocation. ...
Article
Full-text available
Broccoli-derived isothiocyanate sulforaphane inhibits inflammation and cancer. Sulforaphane may support healthy aging, but the underlying detailed mechanisms are unclear. We used the C. elegans nematode model to address this question. Wild-type and 4 mutant C. elegans worm strains were fed in the presence or absence of sulforaphane and E. coli food bacteria transfected with RNA interference gene constructs. Kaplan–Meier survival analysis, live imaging of mobility and pharyngeal pumping, fluorescence microscopy, RT–qPCR, and Western blotting were performed. In the wild type, sulforaphane prolonged lifespan and increased mobility and food intake because of sulforaphane-induced upregulation of the sex-determination transcription factor TRA-1, which is the ortholog of the human GLI mediator of sonic hedgehog signaling. In turn, the tra-1 target gene daf-16 , which is the ortholog of human FOXO and the major mediator of insulin/IGF-1 and aging signaling, was induced. By contrast, sulforaphane did not prolong lifespan and healthspan when tra-1 or daf-16 was inhibited by RNA interference or when worms with a loss-of-function mutation of the tra-1 or daf-16 genes were used. Conversely, the average lifespan of C. elegans with hyperactive TRA-1 increased by 8.9%, but this longer survival was abolished by RNAi-mediated inhibition of daf-16 . Our data suggest the involvement of sulforaphane in regulating healthy aging and prolonging lifespan by inducing the expression and nuclear translocation of TRA-1/GLI and its downstream target DAF-16/FOXO.
... Similarly, SFN administration, again using broccoli sprouts given on three consecutive days did not affect the fraction of exhaled nitric oxide or anti-oxidant enzymes in nasal epithelial cells in atopic asthmatics despite achieving substantial blood levels of SFN . However, in asthmatics, daily doses of SFN significantly ameliorated airway hyperresponsiveness to methacholine challenge in 60% of the subjects (Figure 4; Brown et al., 2015). SFN was also shown to reduce the nasal allergic response and the number of inflammatory cells in nasal lavage fluid following inhalation of diesel exhaust particles, a potent source of oxidative stress, in human subjects (Heber et al., 2014). ...
Article
Full-text available
Nuclear factor erythroid 2-related factor 2 (Nrf2) is a major transcription factor involved in redox homeostasis and in the response induced by oxidative injury. Nrf2 is present in an inactive state in the cytoplasm of cells. Its activation by internal or external stimuli, such as infections or pollution, leads to the transcription of more than 500 elements through its binding to the antioxidant response element. The lungs are particularly susceptible to factors that generate oxidative stress such as infections, allergens and hyperoxia. Nrf2 has a crucial protective role against these ROS. Oxidative stress and subsequent activation of Nrf2 have been demonstrated in many human respiratory diseases affecting the airways, including asthma and chronic obstructive pulmonary disease (COPD), or the pulmonary parenchyma such as acute respiratory distress syndrome (ARDS) and pulmonary fibrosis. Several compounds, both naturally occurring and synthetic, have been identified as Nrf2 inducers and enhance the activation of Nrf2 and expression of Nrf2-dependent genes. These inducers have proven particularly effective at reducing the severity of the oxidative stress-driven lung injury in various animal models. In humans, these compounds offer promise as potential therapeutic strategies for the management of respiratory pathologies associated with oxidative stress but there is thus far little evidence of efficacy through human trials. The purpose of this review is to summarize the involvement of Nrf2 and its inducers in ARDS, COPD, asthma and lung fibrosis in both human and in experimental models.
Article
The airway smooth muscle (ASM) surrounding the airways is dysfunctional in both asthma and chronic obstructive pulmonary disease (COPD), exhibiting; increased contraction, increased mass, increased inflammatory mediator release and decreased corticosteroid responsiveness. Due to this dysfunction, ASM is a key contributor to symptoms in patients that remain symptomatic despite optimal provision of currently available treatments. There is a significant body of research investigating the effects of oxidative stress/ROS on ASM behaviour, falling into the following categories; cigarette smoke and associated compounds, air pollutants, aero-allergens, asthma and COPD relevant mediators, and the anti-oxidant Nrf2/HO-1 signalling pathway. However, despite a number of recent reviews addressing the role of oxidative stress/ROS in asthma and COPD, the potential contribution of oxidative stress/ROS-related ASM dysfunction to asthma and COPD pathophysiology has not been comprehensively reviewed. We provide a thorough review of studies that have used primary airway, bronchial or tracheal smooth muscle cells to investigate the role of oxidative stress/ROS in ASM dysfunction and consider how they could contribute to the pathophysiology of asthma and COPD. We summarise the current state of play with regards to clinical trials/development of agents targeting oxidative stress and associated limitations, and the adverse effects of oxidative stress on the efficacy of current therapies, with reference to ASM related studies where appropriate. We also identify limitations in the current knowledge of the role of oxidative stress/ROS in ASM dysfunction and identify areas for future research.
Chapter
Over the last decades, high mortality indexes observed worldwide from noncommunicable diseases (NCDs), such as obesity or diabetes, have led to growing concerns about the implications of diet and physical activity in daily life. Although NCDs are caused by multiple factors, such as smoking and a sedentary life, it has been reported that diet plays a key role in their development, promoting their high prevalence. This interest in improving the quality of life, as well as lifespan, has led to nutritional intervention based on global health recommendations, supporting the message of eating at least 5 portions of fruits and vegetables every day. As a consequence, bioactive-enriched foods and functional ingredients have emerged as valuable sources of bioactives that contribute to the prevention and reduction of these chronic diseases. At the present time, it is unclear whether the intake of levels achievable through a normal diet is enough to have a therapeutic effect, or if the formulations enriched in bioactive compounds in the form of supplements would be needed. The intervention in the diet is more accessible and affordable from the public health care, but for this, much more research is needed to quantitatively describe the quality, bioavailability, metabolism, and biological activity of Isothiocyanates (ITCs) and other Glucosinolate (GSL) hydrolysis products from real foods. The influence of the consumption of cruciferous foods and derived ingredients enriched in bioactive compounds (GSL hydrolysis or breakdown products) on metabolic, cardiovascular, gastrointestinal, urogenital, and neurological problems is clear and challenges for the future of research on food and nutrition sciences, from plant-derived bioactives to foods and health are revised in this chapter.
Article
Innate immunity, particularly macrophages, is critical for intestinal homeostasis. Sulforaphane, a dietary isothiocyanate from cruciferous vegetables, has been reported to protect against intestinal inflammation. However, the role of macrophages in sulforaphane mediated intestinal inflammation and the underlying molecular mechanisms have not been studied yet. In this study, sulforaphane effectively attenuated dextran sodium sulphate (DSS) induced intestinal inflammation in murine model. Of note, sulforaphane skewed the switching from classically (M1) to alternatively (M2) activated phenotype both in intestinal and bone marrow-derived macrophages (BMDMs). The expression levels of M1 associated maker genes induced by DSS or lipopolysaccharide (LPS) plus interferon gamma-γ (IFN-γ) were suppressed by sulforaphane while M2 marker gene expression levels were improved. This resulted in alteration of inflammatory mediators, particularly interleukin-10 (IL-10), both in colon tissues and culture medium of BMDMs. Subsequently, IL-10 was found to mediate the sulforaphane induced M2 phenotype switching of BMDMs through the activation of STAT3 signaling. This was confirmed by immunofluorescence analysis with increased number of p-STAT3-positive cells in the colon sections. Moreover, anti-IL-10 neutralizing antibody significantly interfered M2 phenotyping of BMDMs induced by sulforaphane with reduced STAT3 phosphorylation. Findings here introduced a potential utilization of sulforaphane for intestinal inflammation treatment with macrophages as the therapeutic targets.
Article
Full-text available
Broccoli sprouts are a convenient and rich source of the glucosinolate, glucoraphanin, which can generate the chemopreventive agent, sulforaphane, an inducer of glutathione S-transferases (GSTs) and other cytoprotective enzymes. A broccoli sprout-derived beverage providing daily doses of 600 µmol glucoraphanin and 40 µmol sulforaphane was evaluated for magnitude and duration of pharmacodynamic action in a 12-week randomized clinical trial. Two hundred and ninety-one study participants were recruited from the rural He-He Township, Qidong, in the Yangtze River delta region of China, an area characterized by exposures to substantial levels of airborne pollutants. Exposure to air pollution has been associated with lung cancer and cardiopulmonary diseases. Urinary excretion of the mercapturic acids of the pollutants, benzene, acrolein, and crotonaldehyde, were measured before and during the intervention using liquid chromatography tandem mass spectrometry. Rapid and sustained, statistically significant (p ≤ 0.01) increases in the levels of excretion of the glutathione-derived conjugates of benzene (61%), acrolein (23%), but not crotonaldehyde were found in those receiving broccoli sprout beverage compared with placebo. Excretion of the benzene-derived mercapturic acid was higher in participants who were GSTT1-positive compared to the null genotype, irrespective of study arm assignment. Measures of sulforaphane metabolites in urine indicated that bioavailability did not decline over the 12-week daily dosing period. Thus, intervention with broccoli sprouts enhances the detoxication of some airborne pollutants and may provide a frugal means to attenuate their associated long-term health risks.
Article
Full-text available
Nuclear factor-erythroid 2 p45-related factor 2 (Nrf2, also called Nfe2l2) is a transcription factor that regulates the cellular redox status. Nrf2 is controlled through a complex transcriptional/epigenetic and post-translational network that ensures its activity increases during redox perturbation, inflammation, growth factor stimulation and nutrient/energy fluxes, thereby enabling the factor to orchestrate adaptive responses to diverse forms of stress. Besides mediating stress-stimulated induction of antioxidant and detoxification genes, Nrf2 contributes to adaptation by upregulating the repair and degradation of damaged macromolecules, and by modulating intermediary metabolism. In the latter case, Nrf2 inhibits lipogenesis, supports β-oxidation of fatty acids, facilitates flux through the pentose phosphate pathway, and increases NADPH regeneration and purine biosynthesis; observations that suggest it directs metabolic reprogramming during stress.
Article
Full-text available
The generation of oxidative stress by ambient air pollution particles contributes to the development of allergic sensitization and asthma, as demonstrated by intranasal challenge with well-characterized diesel exhaust particle (DEP) suspensions in humans. This effect is due to the presence of redox active organic chemicals in DEP, and can be suppressed by antioxidants and inducers of phase II enzymes in animals. In this communication, we determined whether the administration of a standardized broccoli sprout extract (BSE), which contains a reproducible amount of the sulforaphane (SFN) precursor, glucoraphanin, could be used to suppress the nasal inflammatory response in human subjects challenged with 300 μg of an aqueous DEP suspension (equivalent to daily PM exposure levels on a Los Angeles freeway). SFN is capable of inducing an antioxidant and phase II response via activation of the nuclear transcription factor (erythroid-derived 2)-like 2 (Nrf2). Previous studies have shown that 70-90% SFN delivered by BSE is absorbed, metabolized, and excreted in humans. An initial intranasal challenge with DEP in 29 human subjects was used to characterize the magnitude of the inflammatory response. Following a 4 week washout, a BSE that delivers a reproducible and standardized dose of 100 μmol SFN in mango juice was administered daily for four days. The nasal DEP challenge was repeated and lavage fluid collected to perform white blood cell (WBC) counts. The average nasal WBC increased by 66% over the initial screening levels and by 85% over the control levels 24 hours after DEP exposure. However, total cell counts decreased by 54% when DEP challenge was preceded by daily BSE administration for 4 days (p < 0.001). Since the SFN dose in these studies is equivalent to the consumption of 100-200 g broccoli, our study demonstrates the potential preventive and therapeutic potential of broccoli or broccoli sprouts rich in glucoraphanin for reducing the impact of particulate pollution on allergic disease and asthma.
Article
Full-text available
Sulforaphane (1-isothiocyanato-4-(methylsulfinyl)-butane), belonging to a family of natural compounds that are abundant in broccoli, has received significant therapeutic interest in recent years. However, the molecular basis of its effects remains to be elucidated. In this study, we attempt to determine whether sulforaphane regulates the inflammatory response in an ovalbumin (OVA)-induced murine asthma model. Mice were sensitized with OVA, treated with sulforaphane, and then challenged with OVA. Sulforaphane administration significantly alleviated the OVA-induced airway hyperresponsiveness to inhaled methacholine. Additionally, sulforaphane suppressed the increase in the levels of SOCS-3 and GATA-3 and IL-4 expression in the OVA-challenged mice. Collectively, our results demonstrate that sulforaphane regulates Th2 immune responses. This sutdy provides novel insights into the regulatory role of sulforaphane in allergen-induced Th2 inflammation and airway responses, which indicates its therapeutic potential for asthma and other allergic diseases.
Article
Autism spectrum disorder (ASD), characterized by both impaired communication and social interaction, and by stereotypic behavior, affects about 1 in 68, predominantly males. The medico-economic burdens of ASD are enormous, and no recognized treatment targets the core features of ASD. In a placebo-controlled, double-blind, randomized trial, young men (aged 13-27) with moderate to severe ASD received the phytochemical sulforaphane (n = 29)-derived from broccoli sprout extracts-or indistinguishable placebo (n = 15). The effects on behavior of daily oral doses of sulforaphane (50-150 µmol) for 18 wk, followed by 4 wk without treatment, were quantified by three widely accepted behavioral measures completed by parents/caregivers and physicians: the Aberrant Behavior Checklist (ABC), Social Responsiveness Scale (SRS), and Clinical Global Impression Improvement Scale (CGI-I). Initial scores for ABC and SRS were closely matched for participants assigned to placebo and sulforaphane. After 18 wk, participants receiving placebo experienced minimal change (<3.3%), whereas those receiving sulforaphane showed substantial declines (improvement of behavior): 34% for ABC (P < 0.001, comparing treatments) and 17% for SRS scores (P = 0.017). On CGI-I, a significantly greater number of participants receiving sulforaphane had improvement in social interaction, abnormal behavior, and verbal communication (P = 0.015-0.007). Upon discontinuation of sulforaphane, total scores on all scales rose toward pretreatment levels. Dietary sulforaphane, of recognized low toxicity, was selected for its capacity to reverse abnormalities that have been associated with ASD, including oxidative stress and lower antioxidant capacity, depressed glutathione synthesis, reduced mitochondrial function and oxidative phosphorylation, increased lipid peroxidation, and neuroinflammmation.
Article
Major bibliographic databases were searched for studies examining the relationship between fruit and vegetable consumption and the risk of wheezing and asthma. Random-effects models were used to pool study results. Subgroup analyses were conducted by fruit and vegetable categories, study design, and age group. Twelve cohorts, 4 population-based case-control studies, and 26 cross-sectional studies published between January 1990 and July 2013 were identified. For the meta-analysis of adults and children, the relative risk (RR) and confidence intervals (CI) when comparing the highest intake group with the lowest intake group were 0.78 (95%CI, 0.70-0.87) for fruit and 0.86 (95%CI, 0.75-0.98) for vegetables. High intake of fruit and vegetables (RR = 0.76; 95%CI, 0.68-0.86 and RR = 0.83; 95%CI, 0.72-0.96) reduced the risk of childhood wheezing. Total intake of fruit and vegetables had a negative association with risk of asthma in adults and children (RR = 0.54; 95%CI, 0.41-0.69). Consuming fruit and vegetables during pregnancy had no association with the risk of asthma in offspring. High intake of fruit and vegetables may reduce the risk of asthma and wheezing in adults and children.
Article
Cancer chemoprevention refers to the use of drugs or natural compounds (e.g., phytochemicals) to prevent cancer. Michael Sporn, who was instrumental in developing the field of chemoprevention, frames the development of cancer as a continuum, likening its latent, frequently invisible development to a smoldering barn full of hay that, before it bursts into flames, is not a safe place to be. In a recent discussion of chemoprevention he describes it as “the arrest or reversal of the progression of premalignant cells towards full malignancy, using physiological mechanisms that do not kill healthy cells” [15]. Stephen Hecht, another pioneer in the field, further suggests that we need to target susceptible individuals for interventions “including chemoprevention using nontoxic or dietary agents with demonstrated efficacy” and avoid “fleeting, flamboyant approaches” in favor of dealing “with lifestyle factors that link cause and prevention” [16]. The last 25 years have provided an abundance of quantifiable evidence to underpin a concept already supported by an ever-more-robust body of epidemiologic evidence—that specific diets can reduce the risks of and protect against cancer and other chronic diseases. The novelty of this approach is rooted in the concept that ingesting certain phytochemicals from specific plants can boost the intrinsic defensive mechanisms of cells that protect against oxidative damage, inflammation, and DNA-damaging chemicals—some of the fundamental causes of chronic disease and aging [17-18]. There are a number of questions specific to the target populations that will need to be addressed in delivering these interventions. We have discussed them previously [19-21] but they bear repeating: Are there countervailing health or ecological risks associated with the intervention? What is to be the delivery vehicle (fresh food or processed food or drink products)? Can the food product or intervention be manufactured or grown locally and sustainably? Can farmers or consumers afford the costs? Is the intervention culturally appropriate? Are there any contraindications? Can the actual cost effectiveness be determined as the intervention is implemented? And finally, will people comply? Adoption of healthier diets is of course an uphill battle, and the effect size will likely be small if efforts to encourage more healthy eating are not based on sound science, but progress is being made. Approaches were recently reviewed in a special issue of the journal Science [22-24].
Article
Sulforaphane is a promising agent under preclinical evaluation in many models of disease prevention. This bioactive phytochemical affects many molecular targets in cellular and animal models; however, amongst the most sensitive is Keap1, a key sensor for the adaptive stress response system regulated through the transcription factor Nrf2. Keap1 is a sulfhydryl-rich protein that represses Nrf2 signaling by facilitating the polyubiquitination of Nrf2, thereby enabling its subsequent proteasomal degradation. Interaction of sulforaphane with Keap1 disrupts this function and allows for nuclear accumulation of Nrf2 and activation of its transcriptional program. Enhanced transcription of Nrf2 target genes provokes a strong cytoprotective response that enhances resistance to carcinogenesis and other diseases mediated by exposures to electrophiles and oxidants. Clinical evaluation of sulforaphane has been largely conducted by utilizing preparations of broccoli or broccoli sprouts rich in either sulforaphane or its precursor form in plants, a stable thioglucose conjugate termed glucoraphanin. We have conducted a series of clinical trials in Qidong, China, a region where exposures to food- and air-borne carcinogens has been considerable, to evaluate the suitability of broccoli sprout beverages, rich in either glucoraphanin or sulforaphane or both, for their bioavailability, tolerability, and pharmacodynamic action in population-based interventions. Results from these clinical trials indicate that interventions with well characterized preparations of broccoli sprouts may enhance the detoxication of aflatoxins and air-borne toxins, which may in turn attenuate their associated health risks, including cancer, in exposed individuals.