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Recent reports postulate that the dual oxidase (DUOX) proteins function as part of a multicomponent oxidative pathway used by the respiratory mucosa to kill bacteria. The other components include epithelial ion transporters, which mediate the secretion of the oxidizable anion thiocyanate (SCN(-)) into airway surface liquid, and lactoperoxidase (LPO), which catalyzes the H(2)O(2)-dependent oxidation of the pseudohalide SCN(-) to yield the antimicrobial molecule hypothiocyanite (OSCN(-)). We hypothesized that this oxidative host defense system is also active against respiratory viruses. We evaluated the activity of oxidized LPO substrates against encapsidated and enveloped viruses. When tested for antiviral properties, the LPO-dependent production of OSCN(-) did not inactivate adenovirus or respiratory syncytial virus (RSV). However, substituting SCN(-) with the alternative LPO substrate iodide (I(-)) resulted in a marked reduction of both adenovirus transduction and RSV titer. Importantly, well-differentiated primary airway epithelia generated sufficient H(2)O(2) to inactivate adenovirus or RSV when LPO and I(-) were supplied. The administration of a single dose of 130 mg of oral potassium iodide to human subjects increased serum I(-) concentrations, and resulted in the accumulation of I(-) in upper airway secretions. These results suggest that the LPO/I(-)/H(2)O(2) system can contribute to airway antiviral defenses. Furthermore, the delivery of I(-) to the airway mucosa may augment innate antiviral immunity.
(A) Rates of hypoiodous acid (HOI) production by ATPstimulated (100 mM) porcine airway epithelial cells in the presence of apical LPO (6.5 mg/ml) and the indicated I 2 concentrations (open squares, mean 6 SE; n 5 3). The production of HOI was determined by adding nicotinamide adenine dinucleotide phosphate reduced (NADPH) (100 mM) to the apical buffer, and measuring changes in absorbance at 282 and 340 nm. The apical buffer of negative control cultures lacked LPO (solid square), or was supplemented with 200 U/ml of catalase (solid triangle). (B) Cell-dependent inactivation of adenovirus by the generation of HOI. Porcine airway epithelia were stimulated with ATP to generate H 2 O 2 and were supplemented with 0 mM, 5 mM, or 500 mM NaI, as indicated. LPO and Ad5–CMV–eGFP were added to the apical surface after 30 minutes of stimulation, and were allowed to adsorb overnight. Representative micrographs from three independent specimens are presented from 48 hours after transduction. The addition of 5 mM or 500 mM I 2 reduced the transduction of adenovirus, compared with no I 2 . Scale bar 5 500 mm. (C) Cell-dependent inactivation of adenovirus by the generation of HOI (quantitation of data presented in B). Porcine airway epithelia were stimulated with ATP and supplemented with 0 mM, 5 mM, or 500 mM NaI. LPO and Ad5– CMV–eGFP were added to the apical surface after 30 minutes of stimulation, and allowed to adsorb overnight. Error bars represent standard error for three independent porcine airway epithelial cell cultures. *P , 0.05, **P , 0.005, Student t test with Bonferroni correction for multiple comparisons.
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Enhancement of Respiratory Mucosal Antiviral Defenses
by the Oxidation of Iodide
Anthony J. Fischer
, Nicholas J. Lennemann
, Sateesh Krishnamurthy
´ter Po
, Lakshmi Durairaj
Janice L. Launspach
, Bethany A. Rhein
, Christine Wohlford-Lenane
, Daniel Lorentzen
, Botond Ba
and Paul B. McCray, Jr.
Interdisciplinary PhD Program in Genetics,
Medical Scientist Training Program,
Department of Pediatrics,
Department of Microbiology,
Department of Anatomy and Cell Biology, and
Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, Iowa
Recent reports postulate that the dual oxidase (DUOX) proteins
function as part of a multicomponent oxidative pathway used by the
respiratory mucosa to kill bacteria. The other components include
epithelial ion transporters, which mediate the secretion of the
oxidizable anion thiocyanate (SCN
) into airway surface liquid,
and lactoperoxidase (LPO), which catalyzes the H
oxidation of the pseudohalide SCN
to yield the antimicrobial
molecule hypothiocyanite (OSCN
). We hypothesized that this
oxidative host defense system is also active against respiratory
viruses. We evaluated the activity of oxidized LPO substrates against
encapsidated and enveloped viruses. When tested for antiviral
properties, the LPO-dependent production of OSCN
did not in-
activate adenovirus or respiratory syncytial virus (RSV). However,
substituting SCN
with the alternative LPO substrate iodide (I
resulted in a marked reduction of both adenovirus transduction and
RSV titer. Importantly, well-differentiated primary airway epithelia
generated sufficient H
to inactivate adenovirus or RSV when LPO
and I
were supplied. The administration of a single dose of 130 mg
of oral potassium iodide to human subjects increased serum I
concentrations, and resulted in the accumulation of I
in upper
airway secretions. These results suggest that the LPO/I
system can contribute to airway antiviral defenses. Furthermore,
the delivery of I
to the airway mucosa may augment innate antiviral
Keywords: respiratory syncytial virus; adenovirus; lactoperoxidase;
thiocyanate; iodide
Acute viral lower respiratory tract infections represent enor-
mous disease burdens for infants, children, and adults through-
out the world. In children, acute lower respiratory infections are
the most frequent cause of mortality worldwide (1, 2). Despite
the prevalence and severity of these infections, few specific
prophylactic or therapeutic interventions are available for many
of these common diseases, including effective vaccines or anti-
viral treatments. For example, Group B and C adenoviruses are
frequent causes of bronchiolitis and pneumonia, and occasion-
ally cause postinfectious bronchiolitis obliterans (3, 4). Although
respiratory syncytial virus (RSV) is the most common cause of
acute lower respiratory infections throughout the world (5), no
effective vaccine or specific, widely used antiviral therapy is
In evaluating novel approaches to antiviral defenses, we
considered the redundant layers of innate immunity at play in
the airways. We questioned whether an existing host defense
mechanism might be induced or augmented to achieve en-
hanced antiviral protection in the lung. Both nonoxidative and
oxidative host defense systems contribute to the antimicrobial
activity of airway surface liquid (ASL) (6). Although the host
defense roles of many secreted proteins and peptides are well-
characterized, recognition that the airway epithelium orches-
trates an oxidative extracellular microbicidal system is recent
(7–10). This reactive oxygen species–producing system consists
of the airway epithelial proteins dual oxidase (DUOX) 1 and 2,
lactoperoxidase (LPO; secreted by submucosal glands), and the
pseudohalide ion (thiocyanate, SCN
; secreted by epithelia)
(11). However, the efficacy of this LPO/halide/H
mucosal defense system has not been investigated for viral
respiratory pathogens.
The LPO-catalyzed oxidation of SCN
produces hypothio-
cyanite (OSCN
) (12), whereas the oxidation of I
hypoiodous acid (HOI) (13). Both OSCN
and HOI are short-
lived, reactive intermediates that are known to be bactericidal.
We hypothesized that this oxidative antimicrobial system might
possess functional relevance for preventing respiratory viral
infections, through the generation of OSCN
or HOI. Here we
show that the delivery of I
to respiratory epithelial cells
supports the generation of HOI in ASL. HOI, but not OSCN
readily inactivates adenovirus and RSV. Supplementation with
may serve to augment mucosal antiviral defenses.
Chemicals and Enzymes
Sodium iodide (catalogue number S324–500; Fisher, Pittsburgh, PA)
and sodium thiocyanate (catalogue number 251410–500G; Sigma, St.
Louis, MO) were dissolved in deionized, distilled water, sterile-filtered,
and stored at 2208C. Thirty percent H
(catalogue number H325–
500) was purchased from Fisher. Bovine milk LPO was obtained from
Sigma (catalogue number L2005–25MG). The entire sample was
resuspended in deionized, distilled water to a final concentration of 1
This study shows that the lactoperoxidase/I
can contribute to airway antiviral defenses. Furthermore,
the delivery of I
to airway mucosa may augment innate
antiviral immunity. The identification of a new prophylac-
tic or therapeutic approach to prevent or ameliorate
respiratory viral infections could have broad implications
for human health.
(Received in original form September 30, 2010 and in final form March 18, 2011)
This study was supported by National Institutes of Health grants P50 HL-61234
(P.B.M.), PO1 HL-091842 (P.B.M. and B.B.), N01 AI-30040 (P.B.M.), and R01 HL-
090830 (B.B.), the Roy J. Carver Charitable Trust (P.B.M.), and Cystic Fibrosis
Foundation Therapeutics Grant BANFI07A0 (B.B.). This study was also supported
by the Cell and Tissues and Cell Morphology Cores, which is partly supported by
the Center for Gene Therapy for Cystic Fibrosis (through National Institutes of
Health grant P30 DK-54759) and the Cystic Fibrosis Foundation.
Correspondence and requests for reprints should be addressed to Paul B.
McCray, Jr., M.D., Department of Pediatrics, University of Iowa, 240F EMRB,
Iowa City, IA 52242. E-mail:
This article has an online supplement, which is accessible from this issue’s table of
contents at
Am J Respir Cell Mol Biol Vol 45. pp 874–881, 2011
Originally Published in Press as DOI: 10.1165/rcmb.2010-0329OC on March 25, 2011
Internet address:
mg/ml (39 U/ml). Bovine liver catalase was obtained from Sigma
(catalogue number C1345).
Culture of Cell Lines
Vero cells (ATCC, Manassas, VA) were grown in Dulbecco’s minimum
essential medium (DMEM; Invitrogen, Carlsbad, CA), supplemented
with 10% FBS and 1% penicillin–streptomycin (PS). A549 and HEp-2
cells (ATCC) were grown in DMEM/F12 with 10% FBS and 1% PS.
Primary Culture of Airway Epithelia
Primary human or porcine airway epithelial cells grown at the air–
liquid interface were prepared at the In Vitro Models Core Facility at
the University of Iowa, using previously described methods (14). The
use of human tissue samples in this study was approved by the
Institutional Review Board of the University of Iowa. The use of
porcine tissue was approved by the Institutional Animal Care and Use
Committee of the University of Iowa. All primary cells used in this
study were well-differentiated (.2 weeks of culture), and were
maintained in DMEM/F12 medium containing 1% PS, 50 mg/ml
gentamicin sulfate, and 2% Ultroser G (BioSepra, Villeneuve, La
Garenne, France).
Replication-deficient recombinant adenovirus expressing enhanced
green fluorescent protein (eGFP) (Ad5–CMV–eGFP) was produced
and tittered, using previously described methods (15). Sucrose gradi-
ent–purified virus preparations were obtained from the Gene Transfer
Vector Core at the University of Iowa. The virus was titered at 1 310
particles/ml (z1310
transducing units (TU)/ml).
Respiratory Syncytial Virus
RSV strain A2 (provided by Barney Graham at the National Institutes
of Health and Dr. Steven Varga at the University of Iowa) was grown
in HEp-2 cells and prepared as a clarified crude lysate, with a titer of
approximately 2 310
plaque forming units (PFU)/ml. Virus was
titered via syncytia titration on Vero cells (16). Red fluorescent protein
(RFP)–expressing virus (rrRSV), provided by Dr. Steven Varga
(University of Iowa), was previously described (17).
In Vitro Virus Inactivation Assays
See the online supplement for details.
Cell-Dependent Viral Inactivation
For experimental details, see the online supplement.
Measurement of HOI
The production of HOI was detected based on the iodination of
nicotinamide adenine dinucleotide phosphate reduced (NADPH), as
described previously (18). For additional details, see the online
Cell Toxicity Assay
The release of lactate dehydrogenase (LDH) was measured in well-
differentiated human airway epithelia, using a commercially available kit
according to the manufacturer’s instructions (LDH-Cytotoxicity Assay
Kit, catalogue number K311–400; BioVision, Mountain View, CA).
pH Dependence of Oxidative Antiviral Activity
For experimental details, see the online supplement.
RSV Syncytia Titration
RSV was titrated by a variation of the microtiter assay originally
described by Trepanier and colleagues (16). For experimental details,
see the online supplement.
Administration of Iodide to Human Subjects, and Analysis of
Nasal Airway Surface Liquid Composition
For experimental details, see the online supplement.
Oxidized Forms of I
, but Not SCN
, Are Virucidal
against Adenovirus
We first asked whether the oxidized halide and pseudohalide
substrates of LPO-catalyzed oxidation were active against ade-
novirus. We selected adenovirus, a protein-encapsidated DNA
virus, because it is an important respiratory pathogen. To test
whether reactions generating OSCN
or HOI inhibit infection
with adenovirus, we mixed both complete and incomplete LPO
mixtures with recombinant Ad5–CMV–eGFP under cell-free
conditions, and used the expression of eGFP to monitor the
virus transduction of target cells. In Figure 1A, we show that the
addition of the replete HOI-generating reaction completely
abrogated transduction by Ad5–CMV–eGFP. By contrast,
a comparable OSCN
-generating reaction failed to block trans-
duction. None of the incomplete LPO reactions possessed
discernable antiviral activity. H
alone did not inhibit in-
fection by adenovirus.
Inactivation of Adenovirus by Hypoiodous
Acid Is pH-Dependent
The pH of airway surface liquid is slightly acidic, with pH values
ranging from 6.57–7.18 (19–21). To determine whether or not
the inactivation of adenovirus by HOI was pH-dependent, we
monitored adenovirus titers after exposure to HOI across pH
ranges from 5–8. At a pH between 6 and 7, the application of
, and H
was significantly more virucidal than at
physiologic pH (Figure 1B). At pH 8, the oxidation of I
significantly reduced the adenovirus titer, compared with the
PBS control.
Airway Epithelia Produce Hypoiodous Acid in an Iodide
Concentration–Dependent Manner
We previously reported that well-differentiated airway epithelia
generate H
apically, in a DUOX-dependent manner (8).
Here we asked whether airway epithelia produced sufficient
to support the production of HOI in the presence of the
physiological LPO concentration of airway surface liquid and
. Well-differentiated primary airway epithelia were cultured
at the air–liquid interface and stimulated by the apical addition
of ATP, which maximizes the DUOX-mediated production of
(22). The apical buffer (PBS) also contained a range of I
concentrations and LPO. Although a high concentration of
LPO is maintained in the airway surface fluid by the LPO-
secreting submucosal glands (23), the cultured surface airway
epithelia do not secrete LPO (data not shown) (9). The
production of HOI by airway epithelia was determined accord-
ing to the iodination of extracellular NADPH, as previously
described (18). As shown in Figure 2A, under these conditions,
well-differentiated porcine airway epithelia supported the gen-
eration of HOI at a rate similar to that of previously reported
production (8). Similar results were observed using
primary human airway epithelia (data not shown). The addition
of the H
scavenger catalase to the apical buffer, or the
omission of either LPO or I
, inhibited the generation of HOI,
confirming that the production of HOI was dependent on apical
, LPO, and I
Production of HOI by Well-Differentiated Airway Epithelia
Inhibits Adenovirus Transduction without Evidence
of Cytotoxicity
To test whether airway epithelia could generate enough reactive
product to inactivate adenovirus, Ad5–CMV–eGFP (multiplic-
ity of infection [MOI], 12.5 TU/cell) was inoculated apically
Fischer, Lennemann, Krishnamurthy, et al.: Mucosal Oxidative Virucidal Defense System 875
onto well-differentiated primary porcine airway epithelial cells.
Because the Coxsackie virus and adenovirus receptor is
expressed basolaterally (24), adenovirus requires prolonged
incubation to enter when applied to the apical surface (25).
Therefore, the virus was allowed to adsorb overnight to allow
for transduction. As shown in Figure 2B, supplementing epithe-
lia with LPO and I
at 5 mMor500mM significantly reduced the
number of eGFP-expressing cells compared with no I
indicating that viral transduction was inhibited. Quantification of
the foci of infection indicated that the inactivation of adenovirus
was I
dose–dependent (Figure 2C). The addition of 5 mMI
was sufficient to decrease the foci of infection significantly.
Figure 1. (A) Cell-free inactivation of adenovirus by hypoiodous acid.
Replication-deficient recombinant adenovirus expressing enhanced green
fluorescent protein (eGFP) (Ad5–CMV–eGFP) was mixed with combina-
tions of H
,NaSCN,NaI,andlactoperoxidase(LPO)(20mg/ml), as
described in the text, and incubated for 10 minutes. The virus was then
applied to monolayers of A549 cells. Cell monolayers were imaged by
fluorescence microscopy at 48 hours after transduction to monitor the
expression of GFP, and photomicrographs were captured. Representative
en face views are shown. The LPO-catalyzed oxidation of I
resulted in
a complete block of adenovirus transduction. Scale bars,500mm. Results
are representative of three independent experiments. (B) Dependence on
pH of adenovirus inactivation by HOI. Cell-free inactivation reactions for
Ad5–CMV–eGFP were prepared with incomplete or complete (5 mMNaI,
20 mg/ml LPO, and 100 mMH
) LPO reactions at different pH
conditions and incubated for 10 minutes. Samples were then diluted
and applied to A549 cells at approximately 50 multiplicity of infection
(MOI), and 48 hours later, numbers of eGFP-positive cells were counted
by fluorescence microscopy. The percentage of eGFP-positive cells was the
average counted from three separate fields. Results are representative of
three independent experiments. Values shown represent the mean 6SE.
*P,0.0001, two-way ANOVA with Tukey post hoc test.
Figure 2. (A) Rates of hypoiodous acid (HOI) production by ATP-
stimulated (100 mM) porcine airway epithelial cells in the presence of
apical LPO (6.5 mg/ml) and the indicated I
concentrations (open
squares, mean 6SE; n53). The production of HOI was determined by
adding nicotinamide adenine dinucleotide phosphate reduced
(NADPH) (100 mM) to the apical buffer, and measuring changes in
absorbance at 282 and 340 nm. The apical buffer of negative control
cultures lacked LPO (solid square), or was supplemented with 200 U/ml
of catalase (solid triangle). (B) Cell-dependent inactivation of adenovirus
by the generation of HOI. Porcine airway epithelia were stimulated with
ATP to generate H
and were supplemented with 0 mM, 5 mM, or
500 mM NaI, as indicated. LPO and Ad5–CMV–eGFP were added to the
apical surface after 30 minutes of stimulation, and were allowed to
adsorb overnight. Representative micrographs from three independent
specimens are presented from 48 hours after transduction. The
addition of 5 mM or 500 mMI
reduced the transduction of adenovirus,
compared with no I
.Scale bar 5500 mm. (C) Cell-dependent
inactivation of adenovirus by the generation of HOI (quantitation of
data presented in B). Porcine airway epithelia were stimulated with ATP
and supplemented with 0 mM, 5 mM, or 500 mM NaI. LPO and Ad5–
CMV–eGFP were added to the apical surface after 30 minutes of
stimulation, and allowed to adsorb overnight. Error bars represent
standard error for three independent porcine airway epithelial cell
cultures. *P,0.05, **P,0.005, Student ttest with Bonferroni
correction for multiple comparisons.
We previously showed that OSCN
was not toxic to airway
epithelia (8). We asked whether the production of HOI by
airway epithelia was associated with evidence of cell injury by
measuring the release of LDH. The addition of a complete
mixture of LPO, NaI, and H
to well-differentiated epithelia
did not result in significant cell toxicity (Figure 3).
Oxidized Forms of I
, but Not SCN
, Are Virucidal
against RSV
We extended the observations with adenovirus to an enveloped
respiratory virus, RSV. We hypothesized that OSCN
and HOI
might exhibit antiviral activity against RSV. We incubated wild-
type RSV for up to 2 hours with 100 mMH
at pH 7.4. Under
these conditions, the virus titer was unchanged compared with
control PBS (Figure 4A). To test whether oxidized or unoxidized
LPO substrates are virucidal, we assembled various mixtures of
, and approximately 1 310
PFU/ml of
RSV (Figure 4A). We found that neither SCN
and I
nor LPO delivered with either I
or SCN
, decreased the
recovery of RSV, compared with PBS. These controls demon-
strated that neither LPO nor its substrates were virucidal under
these conditions. When a complete reaction mixture of LPO, I
and H
was assembled to yield HOI, the RSV titer was rapidly
reduced to undetectable levels. Therefore, the generation of HOI
is significantly virucidal against RSV, whereas the oxidation of
failed to reduce the RSV titer.
Inactivation of RSV by Hypoiodous Acid Is Most Effective
at Low pH
We evaluated the pH dependence of RSV inactivation by
applying either 5 mM NaI or 500 mM NaSCN in cell-free LPO
reactions. Because the application of 5 mM NaI results in a
suboptimal reduction in RSV titer at pH 7.4 (Figure 4A), we
used this concentration to monitor the dependence of buffer pH
on virus inactivation in hypoiodous acid–generating reactions.
We found that at a pH between 6 and 6.5, the application of
, and H
was significantly more virucidal than at
physiologic pH, with at least a 10-fold reduction in RSV titer
compared with pH 7 (Figure 4B). Under high pH conditions
(pH 8.0), the oxidation of I
did not substantially reduce the
virus titer. The oxidation products of SCN
failed to show sig-
nificant antiviral activity across a range of pH conditions.
Airway Epithelia Produce Sufficient Hypoiodous Acid for the
Inactivation of RSV
We next asked whether airway epithelia produce enough HOI
to inactivate viruses in situ via the LPO and I
system. Well-differentiated primary human airway epithelia,
cultured at the air–liquid interface, were stimulated to produce
by an apical addition of ATP, and I
was supplied to the
cells by addition to the basolateral and apical media. LPO was
added to the apical surface at a concentration of 6.5 mg/ml (8).
We added rrRSV to the apical surface in a total volume of 50 ml,
with combinations of ATP, LPO, catalase, and varying concen-
trations of I
. If the epithelia produced sufficient H
significant loss of RFP expression should occur. Indeed, when
was present in the reaction, a dose-dependent decrease in
foci of infection was evident, 48 hours after infection (Figure 5).
When I
was present at a concentration of 5 mM, the number of
foci of infection decreased, but this trend did not reach sta-
tistical significance. Increasing the concentration of I
to 500 mM
resulted in a marked and significant decrease in the number of
foci of infection. The addition of the enzyme catalase rescued
RSV infection. Based on these results, we conclude that human
airway epithelia can generate sufficient H
to inactivate RSV
when supplemented with pharmacologic concentrations of I
the presence of physiologic concentrations of LPO.
After Bolus Intake of I
, the Concentration of I
in ASL
Exceeds That of SCN
in Humans
The secretion of SCN
by the airway epithelium is thought to be
mediated by the sodium–iodide symporter (NIS) in the basolateral
plasma membrane, and by cystic fibrosis transmembrane conduc-
tance regulator (CFTR) and pendrin in the apical plasma mem-
brane (9, 10). Because these SCN
-transporting proteins are also
capable of I
transport, we hypothesized that I
is absent from
ASL only because of its low serum concentration (10–100 nM). To
examine the I
-secreting capacity of the airway, we performed
a study with nine human subjects. After the verification of normal
thyroid function (based on blood thyroid-stimulating hormone
[TSH] and free thyroxine [T4] levels; data not shown), subjects
took a potassium iodide tablet orally (130 mg KI, Iosat; Anbex
Inc., Williamsburg, VA). Twenty-four hours before the intake of
KI, and 2, 6, and 24 hours later, nasal ASL was harvested using
microsampling probes (26), and blood was drawn from arm veins
for inorganic ion analysis by anion-exchange chromatography (27).
Before the intake of KI, the I
content of ASL was below the
detection limit of our assay (,0.25 mM in 50-fold diluted ASL).
However, 2 hours after the intake of KI, I
approximately 500 mM (Figures 6A and 6B). The concentration of
in ASL remained in the range of several hundred micromolar
for many hours, and exceeded the concentration of serum I
more than 50-fold at 6 hours (Figures 6B). Interestingly, the
concentration of SCN
was reduced in ASL after the intake of KI
(Figures 6A and 6C), whereas the serum concentration of SCN
remained stable. These results suggests that I
and SCN
for transepithelial secretion in the respiratory tract. Our data also
show that the oral intake of a single KI dose (130 mg) leads to the
accumulation of I
in ASL at concentrations that (based on our
cell-culture assays) can support antiviral activities.
Viral respiratory-tract infections are significant causes of mor-
bidity and mortality throughout the world (1). Here we show that
Figure 3. Airway epithelium–dependent production of HOI is not toxic
to epithelial cells. Cytotoxicity was monitored by lactate dehydrogenase
release assay, as described in MATERIALS AND METHODS. Well-differentiated
primary human airway epithelial cells were treated overnight under the
indicated conditions. These included assay buffer alone (PBS, pH 6.6),
and assay buffer containing the indicated combinations of catalase
(750 U/ml), ATP (100 mM), LPO (20 mg/ml), and I
(400 mM). Media
from both apical and basolateral cell surfaces were harvested and
pooled. The positive control represents cells exposed to 1% Triton X-
100 for 30 minutes at 378C. Results shown represent mean 6SE, and
are from three independent experiments with cells from three different
human donors. *P,0.05, Student ttest.
Fischer, Lennemann, Krishnamurthy, et al.: Mucosal Oxidative Virucidal Defense System 877
the LPO/I
oxidative host defense system demonstrated
robust activity against two major respiratory viral pathogens,
adenovirus and RSV. Although OSCN
was not effective
against either adenovirus or RSV, we discovered that the delivery
of the alternative halide substrate I
to the airway epithelium
converted the DUOX/LPO enzymes into a HOI-producing
antiviral system. Remarkably, HOI exhibited virucidal activity
against both encapsidated and enveloped respiratory viruses.
Importantly, the oral administration of KI in human subjects
increased serum I
concentrations and yielded I
in the upper respiratory secretions that supported the antiviral
activity of airway epithelial cells ex vivo. Because the concentra-
tion of LPO in our cell-based study was similar to that of in vivo
airways (28), we speculate that a topical or systemic supplemen-
tation of I
alone might be sufficient to inactivate virus.
The LPO/halide/H
antimicrobial system is a host defense
mechanism previously proposed to serve as a nonspecific anti-
viral defense (12, 29), particularly in the saliva (30, 31), tears
(32), and milk (33). The discovery of DUOX gene products
revealed their distribution in human airways (34). The expres-
sion of DUOX in epithelia stimulated interest in the role of the
system in airway mucosal immunity, because
it identified a mechanism for the production of epithelial H
(11). The two DUOX isoforms, DUOX1 and DUOX2, are
encoded by closely spaced genes on human chromosome 15,
and are transcribed in opposite directions (35, 36). DUOX uses
cytoplasmic NADPH as an electron donor to transfer two
electrons to oxygen, which is reduced to H
and released
extracellularly. The functions of DUOX1 and DUOX2 are also
dependent on the co-expression of the regulatory proteins
DUOXa1 and DUOXa2, respectively (37). Although they are
functionally indistinguishable, DUOX1 and DUOX2 are differ-
entially regulated. Harper and colleagues previously reported
that DUOX2 is inducible by IFN-g, as well as by rhinovirus
infection and polyinosinic:polycytidylic acid (poly[I:C]) (38).
DUOX1 is induced by IL-4 and IL-13 (10, 38), as well as by
bacterial products such as Pseudomonas aeruginosa endotoxin,
flagellum, and Type III secretion system products (39).
Three reports showed that airway epithelial cells supple-
mented with both SCN
and LPO can kill two types of bacteria,
Pseudomonas aeruginosa and Staphylococcus aureus (8, 9, 40).
This provides evidence that the airway LPO/halide/H
Figure 4. (A) Cell-free inactivation of respiratory syncytial
virus (RSV) by the generation of HOI. RSV (1.7 310
plaque forming units [PFU]/ml) was incubated for 5 min-
utes with various combinations of H
(100 mM), NaSCN
(500 mM), NaI (500 nM, 5 mM, and 500 mM), and LPO (20
mg/ml). Treatment categories are indicated on the x-axis,
and the resultant virus titer is presented on the y-axis. SFU,
syncytia-forming units. The limit of detection for this assay
is 10 SFU/ml. Values represent mean 6SD for one of three
independent experiments. *P,0.001, one-way ANOVA
with Tukey post hoc test. (B) Inactivation of RSV by HOI is
dependent on pH. RSV was mixed with complete or
incomplete LPO reactions (5 mMNaI,100mMH
incubated for 10 minutes. Samples were then diluted and
applied to Vero cell monolayers for syncytia titration. At
a pH less than 7, the I
-dependent inactivation of virus
was significantly greater than at physiologic pH (open
diamonds). No other conditions caused dramatic shifts in
RSV titer, including buffer pH, hydrogen peroxide, or LPO
substrates alone. Values represent the mean 6SD for n53
replicate wells. Results are representative of three indepen-
dent experiments. *P,0.001, two-way ANOVA with
Tukey post hoc test.
Figure 5. Cell-dependent inactivation of RSV by the generation of HOI.
Human airway epithelia were stimulated with ATP and supplemented
with 0 mM, 5 mM, or 500 mM NaI, as indicated. LPO and red
fluorescent protein–expressing virus were added to the apical surface
after 30 minutes of stimulation, and allowed to adsorb overnight. Foci
of infection were identified by the expression of red fluorescent protein.
Data are presented as the percentage of foci formed in the absence of
. Values shown represent mean 6SE for results from three separate
human donor cultures. * P,0.05, ** P,0.005, one-way ANOVA with
Tukey post hoc test.
is active against bacteria. The recognition that the CFTR anion
channel conducts SCN
raised the possibility that patients with
cystic fibrosis (CF) might manifest impaired LPO/halide/H
microbicidal activity. Indeed, airway epithelia from patients with
CF exhibited defects in CFTR-dependent SCN
transport and
defective bacterial killing (8–10). These studies suggest a possible
mechanistic link between CF lung disease and altered function of
the LPO/halide/H
In vitro, LPO has a high affinity for both SCN
and I
this study, the I
concentration in ASL had not (to the best of
our knowledge) been reported, but SCN
is present in ASL at
approximately 450 mM (41). Therefore, SCN
is thought to be
the physiologic LPO substrate in the respiratory tract. What
mechanism favors the accumulation of SCN
over I
in the
airways? Animal cells cannot synthesize SCN
. Thus, the diet is
the only source of both SCN
and I
(8, 11, 42, 43). The average
concentration of SCN
in serum is approximately 20 mM(44),
whereas the average concentration of I
in serum is only 10
100 nM (45). With a serum concentration of SCN
fold higher than that of I
, the secretion of SCN
Airway epithelial cells generate an approximately 20-fold serosal-
to-mucosal concentration gradient for the secretion of SCN
importing SCN
on the basolateral side through NIS (46), and
exporting it on the apical side via CFTR and perhaps the anion
transporter pendrin (SLC26A4) (9, 10). I
can follow the same
secretory pathways. In fact, the NIS favors I
threefold over
(47). Thus, if serum I
reaches micromolar concentra-
tions, I
can also be secreted into the respiratory tract. The data
from our investigations of human subjects (Figure 6) support
this concept. Therefore, supplementation with I
may allow
sufficient I
concentrations in ASL to support the production of
HOI. Although chronic treatment with I
may be associated
with toxicity, short-term daily administration to humans is safe
(48, 49). The topical delivery of I
may further reduce any risk
of systemic toxicity.
The mechanism by which hypohalides eliminate bacteria is
not yet known, but OSCN
can oxidize thiol groups in surface
proteins, thereby mediating conformational changes (13, 50).
Although we do not yet know how HOI inactivates adenovirus
and RSV, similar mechanisms are likely in play. Adenovirus
and RSV represent two different classes of viruses. Adenovirus is
a nonenveloped, protein-encapsidated double-stranded DNA
virus that enters through endocytosis, whereas RSV is an
enveloped, single-stranded RNA virus that enters the cell
through receptor-mediated membrane fusion. The results of
our cell-free virus inactivation assays are consistent with
observations by Belding and colleagues that the peroxidase-
mediated oxidation of I
is strongly antiviral (29). Belding and
colleagues further reported that HOI was active against polio
and vaccinia viruses, particularly at a low pH (29). Interestingly,
the LPO/I
system was most virucidal against both adeno-
virus and RSV at slightly acidic pH. Because the pH of ASL is
slightly acidic (19–21), these conditions are expected to favor HOI-
dependent virucidal activity. We speculate that HOI modifies viral
surface proteins and prevents the binding or the entry of
adenovirus and RSV into the epithelium. Alternatively, HOI
may inhibit the synthesis or assembly of viral nucleic acids and
proteins, or prevent the release of virus from cells. Our results
suggest that the innate oxidative antiviral system present in the
airways is a nonspecific mechanism to inactivate invading viruses.
To test the in vivo efficacy of the antiviral activity of this
HOI-generating mechanism, large animal models of viral in-
fections are needed, because murine airways lack both LPO and
DUOX (8), and rat airways contain few LPO-secreting cells
(51). In contrast, the respiratory tracts of larger mammals such
as sheep and pigs are similar to those of humans in regard to the
expression of LPO and DUOX, anatomy, physiology, innate
and adaptive immunity, and susceptibility to viral infection (23,
52). An important question for future study involves whether
the inactivation of virus in vivo is limited by the generation of
, the concentration of I
, or both.
In conclusion, the airway LPO/I
system exhibits ro-
bust antiviral activity against relevant human pathogens. The
identification of a new prophylactic or therapeutic approach to
Figure 6. Accumulation of I
in airway surface liquid (ASL) after bolus
intake of KI. (A) A representative result of nasal ASL analysis before (blue
trace) and 2 hours after (red trace) intake of KI (130 mg) by a human
subject. (Band C) Summarized data show concentrations of I
(B) and
oxidizable anion thiocyanate (SCN
)(C) in nasal ASL (triangles) and
serum (closed squares) at indicated time points before and after intake
of KI (0 hour). Results are presented as mean 6SE (n59). For ASL
, repeated-measures ANOVA, P50.0032; post hoc Dunnett test,
control time point at 224 hours, *P,0.05 and **P,0.01. For ASL I
repeated-measures ANOVA, P,0.0001; post hoc Dunnett test, control
time point at 224 hours, *P,0.05 and **P,0.01.
Fischer, Lennemann, Krishnamurthy, et al.: Mucosal Oxidative Virucidal Defense System 879
prevent or ameliorate respiratory viral infections could have
broad implications for human health.
Author Disclosure:None of the authors have a financial relationship with a
commercial entity that has an interest in the subject of this manuscript.
Acknowledgments: The authors thank Jennifer Bartlett and Jeydith Gutierrez for
critically reviewing and commenting on the manuscript, and Dr. Steven Varga for
providing stocks of RSV and technical assistance.
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Fischer, Lennemann, Krishnamurthy, et al.: Mucosal Oxidative Virucidal Defense System 881
... LPO is secreted in mucosal surfaces or exocrine secretions such as saliva, milk, or tears and is essential for the innate immune defense system [28,47]. The sole action of LPO is bacteriostatic; however, in conjunction with I − , it becomes bactericidal and antiviral [62][63][64]. ...
... The immune response can be influenced indirectly by iodine through the regulation of oxidative stress, which could be a consequence of infections and inflammation processes, or via direct participation, such as its role in the LPO system, part of the innate immune response [62]. The enzymatic activity of LPO yields HIO and other ions such as I 3 − and I 2 in the presence of I − [28,64,169]. LPO is secreted in mucosal surfaces or exocrine secretions such as saliva, milk, or tears and is essential for the innate immune defense system [28,47]. ...
Full-text available
Iodine is a highly reactive element with a single natural and stable isotopic form (127I). In the biosphere, it is one of the 30 essential elements for life, and its chemical speciation defines its availability and biological activities. The most relevant chemical species are iodate (IO3−) and iodide (I−) as the major sources of iodine, with molecular iodine (I2) and hypoiodous acid (HIO) as the most reactive species, and thyroid hormones (THs) as the representative organic compounds. In human biology, THs are master regulators of metabolism, while inorganic species serve for the iodination of organic molecules and contribute to the innate immune system and the antioxidant cellular defense. Additionally, I−, I2, δ-lactone (6-IL), and α-iodohexadecanal (α-IHDA) have shown therapeutic potential in counteracting oxidative stress, cancer, and inflammation. Both inorganic and organic species have applications in the health science industry, from the manufacturing of disinfection and wound care products to supplements, medicines, and contrast media for radiography. Even after nuclear disasters, intake of high doses of iodine prevents the accumulation of radioactive iodine in the body. Conversely, the controlled production of iodine radioisotopes such as 123I, 124I, 125I, and 131I is exploited in nuclear medicine for radiotherapy and diagnostics.
... Iodine supplementation may augment mucosal antiviral defense. According to a study, it has been demonstrated that iodide when added with LPO enzyme exhibited robust antiviral activity in cultured cells against adenovirus (nonenveloped, dsDNA) and RSV (enveloped, −ve sense ssRNA) (Fischer et al., 2011). Since the mechanism is effective against two different types of viruses, it is most likely to be effective against SARS-CoV-2 (enveloped, +ve sense ssRNA) virus also. ...
Full-text available
The global pandemic of COVID‐19 is considered one of the most catastrophic events on earth. During the pandemic, food ingredients may play crucial roles in preventing infectious diseases and sustaining people's general health and well‐being. Animal milk acts as a super food since it has the capacity to minimize the occurrence of viral infections due to inherent antiviral properties of its ingredients. SARS‐CoV‐2 virus infection can be prevented by immune‐enhancing and antiviral properties of caseins, α‐lactalbumin, β‐lactoglobulin, mucin, lactoferrin, lysozyme, lactoperoxidase, oligosaccharides, glycosaminoglycans, and glycerol monolaurate. Some of the milk proteins (i.e., lactoferrin) may work synergistically with antiviral medications (e.g., remdesivir), and enhance the effectiveness of treatment in this disease. Cytokine storm during COVID‐19 can be managed by casein hydrolyzates, lactoferrin, lysozyme, and lactoperoxidase. Thrombus formation can be prevented by casoplatelins as these can inhibit human platelet aggregation. Milk vitamins (i.e., A, D, E, and B complexes) and minerals (i.e., Ca, P, Mg, Zn, and Se) can have significantly positive effects on boosting the immunity and health status of individuals. In addition, certain vitamins and minerals can also act as antioxidants, anti‐inflammatory, and antivirals. Thus, the overall effect of milk might be a result of synergistic antiviral effects and host immunomodulator activities from multiple components. Due to multiple overlapping functions of milk ingredients, they can play vital and synergistic roles in prevention as well as supportive agents during principle therapy of COVID‐19.
... The presence of the LPO system and its role in the host defense has been investigated in human airways and tissue secretions [75]. LPO causes the oxidation of iodine, which is reported to increase the antiviral defense of respiratory mucosal surfaces [76]. Next, LPO-produced hypothiocyanite and hypoiodite were shown to have anti-influenza activity and suggested the development of an LPO-based antiviral system to protect against airway infections [77]. ...
Full-text available
The vast surface area of the respiratory system acts as an initial site of contact for microbes and foreign particles. The whole respiratory epithelium is covered with a thin layer of the airway and alveolar secretions. Respiratory secretions contain host defense peptides (HDPs), such as defensins and cathelicidins, which are the best-studied antimicrobial components expressed in the respiratory tract. HDPs have an important role in the human body’s initial line of defense against pathogenic microbes. Epithelial and immunological cells produce HDPs in the surface fluids of the lungs, which act as endogenous antibiotics in the respiratory tract. The production and action of these antimicrobial peptides (AMPs) are critical in the host’s defense against respiratory infections. In this study, we have described all the HDPs secreted in the respiratory tract as well as how their expression is regulated during respiratory disorders. We focused on the transcriptional expression and regulation mechanisms of respiratory tract HDPs. Understanding how HDPs are controlled throughout infections might provide an alternative to relying on the host’s innate immunity to combat respiratory viral infections.
... The Caspian Sea of the Republic of Kazakhstan belongs to the regions in which the iodine content in the soil is reduced, and iodine deficiency conditions are pretty common [65,66]. Iodine may support the innate immune system to fight bacterial and viral infections [67,68]. Early in the COVID-19 pandemic, it was suggested that Japan's relatively low COVID-19 mortality rate was due to high iodine intake, despite Japan having the oldest population in the world [69,70]. ...
Full-text available
This study aimed to investigate the content of essential elements in the hair of unvaccinated residents of the Caspian region who recovered from COVID-19. This cross-sectional study involved 260 unvaccinated permanent residents of Mangistau oblast aged 18–60. The diagnosis and severity of COVID-19 were based on clinical signs and symptoms, laboratory data, R-graph results, and oxygen saturation by the Clinical Protocol of the Ministry of Health of the Republic of Kazakhstan. Inductively coupled plasma mass spectrometry determined the content of trace elements cobalt (Co), chromium (Cr), copper (Cu), iron (Fe), iodine (I), manganese (Mn), selenium (Se), and zinc (Zn). The content of Cr (p < 0.05), Cu (p < 0.05), Fe (p < 0.001), I (p < 0.05), Mn (p < 0.001), and Zn (p < 0.05) in the hair of individuals who had a coronavirus infection was lower than those who did not have this infection. There were significantly higher levels of Cu (p < 0.05) in the hair of participants who had moderate or severe COVID-19 compared to those with mild forms. The results of multiple regression analysis showed that in the presence of a COVID-19 infection in a subject’s history, the content of Cr (0.871 (95% CI: 0.811; 0.936)), Cu (0.875 (95% CI: 0.803; 0.955)), Fe (0.745 (95% CI: 0.636; 0.873)), and Mn (0.642 (95%CI: 00.518; 0.795)) decreased in the hair. The data obtained indicate that past COVID-19 infections affect the trace element status of the inhabitants of the Caspian region of Kazakhstan.
... Увеличение концентрации йодида показало выраженную активность против ДНК-содержащего аденовируса и РНК-содержащего респираторно-синцитиального вируса, являющихся основными респираторными вирусными патогенами. A.J. Fischer et al. предполагают, что противовирусная защита реализуется за счет активации системы «лактопероксидаза/I 2 /H 2 O 2 », что способствует повышению врожденного иммунитета [36]. Также в литературе есть данные о таких биологических эффектах йодидов, как регулирование воспаления и улучшение фагоцитоза бактерий иммунными клетками [37]. ...
Despite the development and implementation of vaccines in various countries of the world, COVID-19 remains a significant medical and social problem. This is directly related to the characteristic feature of SARS-CoV-2 to form new strains, which reduces the effectiveness of vaccination. In connection with the foregoing, the priority in the prevention of COVID-19 is to improve and maintain the normal functioning of the human immune system. Recently, more and more scientists have noted the significant role of micronutrients in ensuring immune function. However, most research focuses on micronutrients such as zinc, selenium, iron and copper, while it is known that the balance of micronutrients depends on all its constituents. Consequently, any change in the content of one mineral substance can affect the level of others, leading to an imbalance of trace elements in the body. The aim of this work was to analyze literature data on less studied microelements in the context of the COVID-19 pandemic, both essential and toxic, that can affect the state of the immune system and, as a result, the incidence and risk of complications and adverse outcomes in COVID-19. An analysis of the literature on the effect of manganese, chromium, iodine, cadmium, mercury, lead, arsenic and lithium on human antiviral protection, including in the case of a disease caused by SARS-CoV-2, showed that the determination of the microelement status, taking into account the above microelements and, with necessary, the appointment of preparations containing minerals is promising for the purpose of prevention and as an additional therapy for COVID-19.
... Iodine possesses a significant activity as a scavenger of ROS; therefore, it is an effective antioxidant in living organisms [65,66]. As iodide is oxidized to hypoiodite anion (IO − ) it reveals strong bactericidal, antiviral, and antifungal effects [67][68][69][70][71][72]. It is documented to be an apoptotic, antiproliferative, and differentiation factor in various tissues [65,66]. ...
Full-text available
Purpose Iodine is an essential micronutrient required for thyroid hormone biosynthesis. However, overtreatment with iodine can unfavorably affect thyroid physiology. The aim of this review is to present the evidence that iodine—when in excess—can interfere with thyroid hormone synthesis and, therefore, can act as a potential endocrine-disrupting chemical (EDC), and that this action, as well as other abnormalities in the thyroid, occurs—at least partially—via oxidative stress. Methods We reviewed published studies on iodine as a potential EDC, with particular emphasis on the phenomenon of oxidative stress. Results This paper summarizes current knowledge on iodine excess in the context of its properties as an EDC and its effects on oxidative processes. Conclusion Iodine does fulfill the criteria of an EDC because it is an exogenous chemical that interferes—when in excess—with thyroid hormone synthesis. However, this statement cannot change general rules regarding iodine supply, which means that iodine deficiency should be still eliminated worldwide and, at the same time, iodine excess should be avoided. Universal awareness that iodine is a potential EDC would make consumers more careful regarding their diet and what they supplement in tablets, and—what is of great importance—it would make caregivers choose iodine-containing medications (or other chemicals) more prudently. It should be stressed that compared to iodine deficiency, iodine in excess (acting either as a potential EDC or via other mechanisms) is much less harmful in such a sense that it affects only a small percentage of sensitive individuals, whereas the former affects whole populations; therefore, it causes endemic consequences.
We draw the attention of readers and governments to the death rate from coronavirus disease 2019 in Japan, continuing as a fraction of that experienced by many other developed nations. We think this is due to the activity of the powerful, protective lactoperoxidase system (LPO) which prevents serious airborne infections. The LPO system requires iodine, which is liberally provided by the typical Japanese diet but lacking in many others. One might consider the Japanese experience an incredibly large, open‐label study exhibiting the preventative power of a high‐iodine diet. We predict this favourable trend will continue for Japan because deadly variants of the severe, acute respiratory syndrome coronavirus 2 will be with us, forever.
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The lactoperoxidase (LPO)-hydrogen peroxide-halides reaction (LPO system) converts iodide and thiocyanate (SCN⁻) into hypoiodous acid (HOI) and hypothiocyanite (OSCN⁻), respectively. Since this system has been implicated in defense of the airways and oropharynx from microbial invasion, in this proof-of-concept study we measured the concentrations of these analytes in human saliva from a convenience clinical sample of 40 qualifying subjects before and after acute iodine administration via the iodinated contrast medium used in coronary angiography to test the hypothesis that an iodide load increases salivary iodide and HOI concentrations. Saliva was collected and salivary iodide, SCN⁻, HOI and OSCN⁻ were measured using standard methodology. The large iodine load delivered by the angiographic dye, several 100-fold in excess of the U.S. Recommended Daily Allowance for iodine (150 µg/day), significantly increased salivary iodide and HOI levels compared with baseline levels, whereas there was no significant change in salivary SCN⁻ and OSCN⁻ levels. Iodine load and changes of salivary iodide and HOI levels were positively correlated, suggesting that higher iodide in the circulation increases iodide output and salivary HOI production. This first of its kind study suggests that a sufficient but safe iodide supplementation less than the Tolerable Upper Limit for iodine set by the U.S. Institute of Medicine (1,100 µg/day) may augment the generation of antimicrobial HOI by the salivary LPO system in concentrations sufficient to at least in theory protect the host against susceptible airborne microbial pathogens, including enveloped viruses such as coronaviruses and influenza viruses.
The domestic cat is the most susceptible host to Sporothrix infection, developing severe clinical forms. Few effective antifungal agents are available for treating feline sporotrichosis, and cases of treatment failure are common. Treatment success depends on cat health status, therapy-related factors, as well as social/economic issues, but it is mainly contingent upon the host–fungus interaction. The owner's adherence is critical and should be reinforced throughout the treatment to increase the chances of a successful outcome. The antifungal agents described for feline sporotrichosis are most often used in monotherapy regimens. Due to cases in which the treatment with itraconazole failed, the use of antifungal agents in combination should be considered to achieve synergy. The combination of itraconazole and potassium iodide represents an important option for the treatment of naïve cats presenting multiple cutaneous lesions, nasal mucosal lesions and/or respiratory signs, as well as for refractory cases. However, the therapeutic options for unsuccessfully treated cases are scarce. Therefore new options are needed, even more taking into account that there are many in vitro potential molecules not available for use in cats yet. More studies are necessary to correlate in vitro antifungal susceptibility tests results and the outcome of cats treated due to sporotrichosis. This review will briefly discuss both the antifungal drugs and treatment protocols used in cats with sporotrichosis, as well as the determinants of treatment failure.
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Hydrogen peroxide is the final electron acceptor for the biosynthesis of thyroid hormone catalyzed by thyroperoxidase at the apical surface of thyrocytes. Pig and human thyroid plasma membrane contain a Ca(2+)-dependent NAD(P)H oxidase that generates H(2)O(2) by transferring electrons from NAD(P)H to molecular oxygen. We purified from pig thyroid plasma membrane a flavoprotein which constitutes the main, if not the sole, component of the thyroid NAD(P)H oxidase. Microsequences permitted the cloning of porcine and human full-length cDNAs encoding, respectively, 1207- and 1210-amino acid proteins with a predicted molecular mass of 138 kDa (p138(Tox)). Human and porcine p138(Tox) have 86.7% identity. The strongest similarity was to a predicted polypeptide encoded by a Caenorhabditis cDNA and with rbohA, a protein involved in the Arabidopsis NADPH oxidase. p138(Tox) shows also similarity to the p65(Mox) and to the gp91(Phox) in their C-terminal region and have consensus sequences for FAD- and NADPH-binding sites. Compared with gp91(Phox), p138(Tox) shows an extended N-terminal containing two EF-hand motifs that may account for its calcium-dependent activity, whereas three of four sequences implicated in the interaction of gp91(Phox) with the p47(Phox) cytosolic factor are absent in p138(Tox). The expression of porcine p138(Tox) mRNA analyzed by Northern blot is specific of thyroid tissue and induced by cyclic AMP showing that p138(Tox) is a differentiation marker of thyrocytes. The gene of human p138(Tox) has been localized on chromosome 15q15.
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SCN(-) (thiocyanate) is an important physiological anion involved in innate defense of mucosal surfaces. SCN(-) is oxidized by H(2)O(2), a reaction catalyzed by lactoperoxidase, to produce OSCN(-) (hypothiocyanite), a molecule with antimicrobial activity. Given the importance of the availability of SCN(-) in the airway surface fluid, we studied transepithelial SCN(-) transport in the human bronchial epithelium. We found evidence for at least three mechanisms for basolateral to apical SCN(-) flux. cAMP and Ca(2+) regulatory pathways controlled SCN(-) transport through cystic fibrosis transmembrane conductance regulator and Ca(2+)-activated Cl(-) channels, respectively, the latter mechanism being significantly increased by treatment with IL-4. Stimulation with IL-4 also induced the strong up-regulation of an electroneutral SCN(-)/Cl(-) exchange. Global gene expression analysis with microarrays and functional studies indicated pendrin (SLC26A4) as the protein responsible for this SCN(-) transport. Measurements of H(2)O(2) production at the apical surface of bronchial cells indicated that the extent of SCN(-) transport is important to modulate the conversion of this oxidant molecule by the lactoperoxidase system. Our studies indicate that the human bronchial epithelium expresses various SCN(-) transport mechanisms under resting and stimulated conditions. Defects in SCN(-) transport in the airways may be responsible for susceptibility to infections and/or decreased ability to scavenge oxidants.
Bovine tissues were analyzed for the presence of lactoperoxidase by immunodiffusion analysis. A protein antigenically identical to lactoperoxidase was shown to be present in the sublingual, submaxillary, and parotid glands. An antigenically related but not identical protein was present in pig salivary glands. As the enzyme could not be detected in any of the other tissues employed, it could be concluded that if present, it represented less than 0.04% of the extracted protein. Most notable among the tissues which did not contain the enzyme, are the thyroid, uterus, and lymph node.
La pneumonie est la principale cause simple de mortalite chez les enfants de moins de 5 ans. L'incidence dans cette tranche d'âge est estimee a 0,29 episode/enfant/an dans les pays en developpement et a 0,05 episode/enfant/an dans les pays developpes. Il en resulte environ 156 millions de nouveaux episodes de pneumonie chaque annee dans le monde, dont 151 millions dans les pays en developpement. La plupart des cas se produisent en Inde (43 millions), en Chine (21 millions), au Pakistan (10 millions) et egalement en grands nombres au Bengladesh, en Indonesie et au Nigeria (6 million pour chacun de ces pays). Parmi l'ensemble des cas communautaires, 7 a 13 % sont assez graves pour menacer le pronostic vital et necessiter une hospitalisation. De nombreux elements ont fait apparaitre comme facteurs de risque principaux pour l'incidence de la pneumonie l'absence d'allaitement au sein exclusif, la denutrition, la pollution de l'air interieur, le petit poids a la naissance, le surpeuplement et le manque de couverture par la vaccination antirougeoleuse. La pneumonie est responsable d'environ 19 % des deces d'enfants de moins de 5 ans, dont plus de 70 % se produisent en Afrique sub-saharienne et en Asie du Sud-est. Bien que reposant sur les donnees disponibles limitees, les etudes recentes ont identifie Streptococcus pneumonia, Haemophilus influenzae et le virus respiratoire syncytial comme les principaux agents pathogenes associes a la pneumonie de l'enfant.
The human heme peroxidases myeloperoxidase (MPO),eosinophil peroxidase (EPO) and lactoperoxidase (LPO) are able to oxidise (pseudo)halides and nitrite to reactive species that participate in host defence against foreign microorganisms as well as in immunomodulation and tissue degradation in certain pathologies. The heme in EPO and LPO is covalently linked to the apoprotein by two ester bonds, whereas in MPO it is additionally linked by a unique sulfonium ion bond to a methionine residue. As a consequence, the prosthetic group in MPO is significantly distorted from a planar conformation. These structural differences are reflected by distinct spectral and redox properties as well as reactivities toward chloride, bromide, iodide, thiocyanate and nitrite, which function as endogenous two- and one-electron donors for these enzymes in vivo. Standard reduction potentials at pH 7 have been determined for all redox couples involved in the halogenation and peroxidase cycle of MPO and LPO and partially of EPO. A detailed thermodynamic analysis of the formation of reactive halide species by MPO and EPO was also performed. Thus, for the first time, a comprehensive analysis of reactions catalysed by human heme peroxidases is presented that allows a better understanding of their role in physiological and pathophysiological processes. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)
A recently discovered enzyme system produces antibacterial hypothiocyanite (OSCN(-)) in the airway lumen by oxidizing the secreted precursor thiocyanate (SCN(-)). Airway epithelial cultures have been shown to secrete SCN(-) in a CFTR-dependent manner. Thus, reduced SCN(-) availability in the airway might contribute to the pathogenesis of cystic fibrosis (CF), a disease caused by mutations in the CFTR gene and characterized by an airway host defense defect. We tested this hypothesis by analyzing the SCN(-) concentration in the nasal airway surface liquid (ASL) of CF patients and non-CF subjects and in the tracheobronchial ASL of CFTR-ΔF508 homozygous pigs and control littermates. In the nasal ASL, the SCN(-) concentration was ~30-fold higher than in serum independent of the CFTR mutation status of the human subject. In the tracheobronchial ASL of CF pigs, the SCN(-) concentration was somewhat reduced. Among human subjects, SCN(-) concentrations in the ASL varied from person to person independent of CFTR expression, and CF patients with high SCN(-) levels had better lung function than those with low SCN(-) levels. Thus, although CFTR can contribute to SCN(-) transport, it is not indispensable for the high SCN(-) concentration in ASL. The correlation between lung function and SCN(-) concentration in CF patients may reflect a beneficial role for SCN(-).
The global burden of disease attributable to respiratory syncytial virus (RSV) remains unknown. We aimed to estimate the global incidence of and mortality from episodes of acute lower respiratory infection (ALRI) due to RSV in children younger than 5 years in 2005. We estimated the incidence of RSV-associated ALRI in children younger than 5 years, stratified by age, using data from a systematic review of studies published between January, 1995, and June, 2009, and ten unpublished population-based studies. We estimated possible boundaries for RSV-associated ALRI mortality by combining case fatality ratios with incidence estimates from hospital-based reports from published and unpublished studies and identifying studies with population-based data for RSV seasonality and monthly ALRI mortality. In 2005, an estimated 33.8 (95% CI 19.3-46.2) million new episodes of RSV-associated ALRI occurred worldwide in children younger than 5 years (22% of ALRI episodes), with at least 3.4 (2.8-4.3) million episodes representing severe RSV-associated ALRI necessitating hospital admission. We estimated that 66 000-199 000 children younger than 5 years died from RSV-associated ALRI in 2005, with 99% of these deaths occurring in developing countries. Incidence and mortality can vary substantially from year to year in any one setting. Globally, RSV is the most common cause of childhood ALRI and a major cause of admission to hospital as a result of severe ALRI. Mortality data suggest that RSV is an important cause of death in childhood from ALRI, after pneumococcal pneumonia and Haemophilus influenzae type b. The development of novel prevention and treatment strategies should be accelerated as a priority. WHO; Bill & Melinda Gates Foundation.
Hydrogen peroxide production by the NADPH oxidase Duox1 occurs during activation of respiratory epithelial cells stimulated by purified bacterial ligands, such as lipopolysaccharide. Here, we characterize Duox activation using intact bacterial cells of several airway pathogens. We found that only Pseudomonas aeruginosa, not Burkholderia cepacia or Staphylococcus aureus, triggers H2O2 production in bronchial epithelial cells in a calcium-dependent but predominantly ATP-independent manner. Moreover, by comparing mutant Pseudomonas strains, we identify several virulence factors that participate in Duox activation, including the type-three secretion system. These data provide insight on Duox activation by mechanisms unique to P. aeruginosa.
Combined treatment with anti-thyroid drugs (ATDs) and potassium iodide (KI) has been used only for severe thyrotoxicosis or as a pretreatment before urgent thyroidectomy in patients with Graves' disease. We compared methimazole (MMI) treatment with MMI + KI treatment in terms of rapid normalization of thyroid hormones during the early phase and examined the later induction of disease remission. A total of 134 untreated patients with Graves' disease were randomly assigned to one of four regimens: Group 1, MMI 30 mg; Group 2, MMI 30 mg + KI; Group 3, MMI 15 mg and Group 4, MMI 15 mg + KI. For easy handling, KI tablets were used instead of saturated solution of KI. KI was discontinued when patients showed normal free thyroxine (FT4) levels but MMI was continued with a tapering dosage until remission. Remission rate was examined during a 4- to 5-year observation. Serum FT4, FT3 and TSH were measured by chemiluminescent immunoassays. TSH receptor antibody (TRAb) was assayed with TRAb-ELISA. Goitre size was estimated by ultrasonography. After 2 weeks of treatment, normal FT4 was observed in 29% of patients in Group 1 and 59% (P < 0.05) of patients in Group 2. Furthermore, normal FT4 after 2 weeks of treatment was observed in 27% of patients in Group 3 and 54% (P < 0.05) of patients in Group 4. Similarly, FT3 normalized more rapidly in Groups 2 and 4 than in Groups 1 and 3. None of the patients showed an increase in thyroid hormones or aggravation of disease during combined treatment with MMI and KI. The remission rates in Groups 1, 2, 3 and 4 were 34%, 44%, 33% and 51%, respectively, and were higher in the groups receiving combined therapy but differences among four groups did not reach significance. Combined treatment with MMI and KI improved the short-term control of Graves' hyperthyroidism and was not associated with worsening hyperthyroidism or induction of thionamide resistance.