Download Immunity against Mucosal Pathogens?

What Role Does the Route of Immunization
Play in the Generation of Protective
Immunity against Mucosal Pathogens?
This information is current as
of June 14, 2017.
Subscription
Permissions
Email Alerts
J Immunol 2009; 183:6883-6892; ;
doi: 10.4049/jimmunol.0901466
http://www.jimmunol.org/content/183/11/6883
This article cites 92 articles, 42 of which you can access for free at:
http://www.jimmunol.org/content/183/11/6883.full#ref-list-1
Information about subscribing to The Journal of Immunology is online at:
http://jimmunol.org/subscription
Submit copyright permission requests at:
http://www.aai.org/About/Publications/JI/copyright.html
Receive free email-alerts when new articles cite this article. Sign up at:
http://jimmunol.org/alerts
The Journal of Immunology is published twice each month by
The American Association of Immunologists, Inc.,
1451 Rockville Pike, Suite 650, Rockville, MD 20852
Copyright © 2009 by The American Association of
Immunologists, Inc. All rights reserved.
Print ISSN: 0022-1767 Online ISSN: 1550-6606.
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
References
Igor M. Belyakov and Jeffrey D. Ahlers
What Role Does the Route of Immunization Play in the
Generation of Protective Immunity against Mucosal
Pathogens?
Igor M. Belyakov1* and Jeffrey D. Ahlers†
T
he major entry point for many human pathogens occurs at gastrointestinal (e.g., polio virus, Escherichia
coli, Salmonella, Shigella, Vibrio cholerae, Helicobacter
pylori, and HIV-1), respiratory (e.g., influenza virus, Mycobacterium tuberculosis, adenovirus, coronavirus, rhinovirus, respiratory syncytial virus), or genital (HSV, human papillomavirus
(HPV),2 HIV-1, Chlamydia, and Neisseria gonorrhoeae) mucosal surfaces (1–11). The main function of the innate mucosal
immune system, to discriminate between dangerous and innocuous organisms, is determined by the recognition of specific
pathogen-associated molecular patterns via activation of TLRs,
NOD-like receptors, retinoic acid (RA)-inducible gene I-like
helicases, and C-type lectins (12). Mucosal pathogens may disseminate to distant systemic sites through entry into the blood
*Midwest Research Institute, Frederick, MD 21702; and †National Institute of Allergy and
Infectious Diseases, National Institutes of Health, Bethesda, MD 20817
Received for publication September 9, 2009. Accepted for publication October 14, 2009.
The costs of publication of this article were defrayed in part by the payment of page charges.
This article must therefore be hereby marked advertisement in accordance with 18 U.S.C.
Section 1734 solely to indicate this fact.
1
Address correspondence and reprint requests to Prof. Igor M. Belyakov, Midwest Research Institute, 110 Thomas Johnson Drive, Frederick, MD 21702. E-mail address:
[email protected]
www.jimmunol.org/cgi/doi/10.4049/jimmunol.0901466
stream, although some microorganisms are limited to the development of disease only at the site of initial mucosal invasion
(8, 13, 14). For HIV, numerous studies have demonstrated that
the gut mucosa is the predominant site of viral replication and
amplification (15).
Protective immunity against mucosal pathogens will require
novel vaccine strategies to induce mucosal immune responses
tailored to the anatomic location and the threat of the invading
pathogen (16 –32). However, the requirement for mucosal immunization to generate protective “frontline immunity” against
mucosal pathogens is highly controversial. On the one hand,
numerous studies in the literature have demonstrated that immune responses are readily detectable at mucosal sites following
systemic delivery of vaccines, and complete or partial protection
from mucosal challenge is attainable (2, 33– 40). Systemic immunization is adequate for successful vaccines for some mucosal
pathogens, notably the polio virus and the influenza virus,
where high titers of neutralizing Abs are capable of clearing cellfree virus and preventing disease (34, 41). On the other hand,
mucosal pathogens such as HIV-1, HPV, herpes viruses, Mycobacterium species, and other intracellular pathogens may require mucosal vaccine strategies that activate multiple arms of
the innate and adaptive immune systems (29, 30, 42– 45). We
as well as others have shown that protective mucosal immune
responses are most effectively induced by mucosal immunization through oral, intranasal (i.n.), intrarectal, or intravaginal
routes, and an optimized mucosal vaccination strategy may
have a much greater potential for generating local protective
mucosal immune responses (30, 46 –52). Studies evaluating
mucosal infection and immunization in humans and animals
have demonstrated the existence of a common mucosal immune system (CMIS) that consists of gastrointestinal, respiratory, and genital mucosa (3, 46, 53, 54). The CMIS implies the
ability of Ag-specific lymphocytes to home to mucosal effector
sites in addition to the site where initial Ag exposure occurred
(53). Different mucosal routes of immunization such as oral,
nasal, or rectal routes can induce generalized mucosal immune
responses not only at the portals of entry of infectious agents but
2
Abbreviations used in this paper: HPV, human papillomavirus; ASC, Ab-secreting cell;
BCG, bacillus Calmette-Guérin; CMIR, compartmentalized mucosal immune response;
CMIS, common mucosal immune system; DC, dendritic cell; i.n., intranasal; IPV, inactive
poliovirus vaccine; iTreg, induced Treg; LP, lamina propria; MLN, mesenteric lymph
node; OPV, oral poliovirus vaccine; RA, retinoic acid; SHIV, simian/human immunodeficiency virus; sIg, secretory Ig; TB, tuberculosis; Treg, regulatory T cell; VLP, virus-like
particle.
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
The route of vaccination is important in influencing immune responses at the initial site of pathogen invasion
where protection is most effective. Immune responses
required for mucosal protection can differ vastly depending on the individual pathogen. For some mucosal
pathogens, including acute self-limiting infections,
high-titer neutralizing Abs that enter tissue parenchyma
or transude into the mucosal lumen are sufficient for
clearing cell-free virus. However, for pathogens causing
chronic infections such as HIV, hepatitis C virus, herpes
viruses, mycobacteria, and fungal and parasitic infections, a single arm of the immune response generated by
systemic vaccination may be insufficient for protection.
Induction of the mucosal innate and adaptive immune
systems, including CD4ⴙ T help, Th17, high avidity
CD8ⴙ CTL, and secretory IgA and IgG1 neutralizing
Abs, at the site of pathogen entry may be required for
effective protection against highly invasive pathogens
that lead to chronic infection and may be generated predominantly by mucosal vaccination. The Journal of
Immunology, 2009, 183: 6883– 6892.
6884
BRIEF REVIEWS: COMPARTMENTALIZED MUCOSAL IMMUNE RESPONSE AND PROTECTION
in distant mucosal effector sites as well, although additional
constraints on mucosal compartmentalization are evidently dependent upon the adjuvant and the Ag delivery vehicle. The
term mucosal compartmentalization refers to the process of Ag
uptake, migration, and processing by local APCs and the magnitude and phenotype of T cells or predominant Ab isotype(s)
induced in relationship to the route of immunization and mucosal site(s) analyzed (3, 54). Cytokine and costimulatory molecule expression by locally activated dendritic cells (DCs), epithelial cells, and stromal cells account for the differences in T
cell function induced in proximal mucosal sites vs distal tissue
or lymph nodes. Thus, the generation of mucosal immunity after systemic vaccination may not recapitulate the immune signatures generated after mucosal vaccination with the same Ag.
In general, compartmentalized mucosal immune responses
(CMIRs) that consist of innate responses, mucosal Abs (secretory Ig (sIg) A and sIgG) and CD8⫹ CTLs localized to tissues
proximal to the mucosal site of immunization are necessary for
protection from mucosal pathogens (Fig. 1). Optimum mucosal vaccination leading to compartmentalized mucosal immune
responses might ensure that the appropriate cells are armed and
ready to respond immediately to infection and to confer protection not achieved following natural infection (Fig. 1). Vaccine strategies that induce both Ag-specific CD4⫹ Th cells in
draining lymph nodes to provide help during recall and effec-
tor/memory CD8⫹ cells in mucosal effector sites (lamina propria (LP)) will be crucial in containing initial viral replication
and subsequent disease course following HIV infection. Induction of central memory CD8⫹ T cells rapidly recruited to sites
of initial virus replication could provide a concerted second
wave attack to limit spread of the virus systemically.
Only a few mucosal vaccines have been approved for human
vaccination, including poliovirus, influenza, Salmonella typhi,
and V. cholerae vaccines (16, 20, 21, 24). Improved mucosal
vaccines are needed against a wide range of respiratory, gastrointestinal, and urogenital tract mucosal pathogens. The main
challenge for mucosal vaccine development is to design vaccines
that are sufficiently immunogenic and can be safely delivered by
mucosal routes of vaccination to prevent (or reduce) transmission of the mucosal pathogen through the mucosal barrier and
dissemination into the systemic circulation. This will require
vaccine strategies that induce immune responses tailored to the
individual pathogen. In this review we posit mechanisms of
protection for existing mucosal vaccines currently in use and
provide analysis of experimental studies comparing mucosal vs
systemic vaccination strategies. This knowledge may help to
provide unifying principles toward the design of better vaccines
with mucosal delivery strategies that can be effective against diverse pathogens associated with chronic mucosal infection and
disease.
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
FIGURE 1. Schema of compartmentalized immunity (functional CD8⫹ CTL activity and avidity and neutralizing IgG and sIgA antibodies) according to the site
of immunization. CMIR contributes to the protection of macaques against SIV depletion of mucosal CD4⫹ T cell after optimum mucosal vaccination (46). CTL
that can recognize peptide/MHC only at high Ag density are termed low avidity CTLs, whereas those that can recognize their cognate Ag at low densities are termed
high avidity CTLs (93). High avidity CTLs are essential for the effective clearance of viral infections and for the elimination of tumors. The high avidity CD8⫹ T
cells are predominately found compartmentalized at the site of vaccination. The simple application of the Ag on a mucosal surface does not guarantee the success of
mucosal vaccination. Non-optimal mucosal vaccination (upper panel) may provide limited mucosal immunity (both functional CD8⫹ CTL and neutralizing antibodies) and limited protection against mucosal challenge. Optimal mucosal immunization (lower panel) depends on multiple factors including type of Ag, targeting
and delivery of Ag to Dcs, mucosal adjuvant, and frequency of immunization. Optimal mucosal vaccination is most effective in the generation of mucosal immunity
(natural antibodies (NAb) IgG and IgA, high-avidity CD8⫹ CTL, and CD4⫹ T cells) and complete protection against mucosal infection. Systemic immunization
is less effective in generation of high-avidity CTL and NAb at mucosal sites and may result in only partial protection from mucosal challenge. Green arrows indicate
cell migration toward mucosal tissue and the red arrows indicate cell migration to systemic lymphoid tissue. Thick arrows indicate a higher frequency of functionally
active Ag-specific T and B cells in the tissue proximal to the site of vaccination, whereas the thin arrows indicate limited immune responses at the distant sites.
The Journal of Immunology
A timeless debate: the oral poliovirus vaccine (OPV) vs inactivated
poliovirus vaccine (IPV)
The prospect of mucosal vaccines for influenza
Systemic delivery of inactivated influenza vaccines has proven
effective in providing protection against the specific influenza
subtypes included in the vaccine. These vaccines are capable of
eliciting high-titer, subtype-specific neutralizing Abs that prevent disease but are insufficient to prevent infection at mucosal
surfaces. Mucosal delivery strategies that mimic the process of
natural infection to elicit effective innate and adaptive immune
responses are currently being explored in an effort to broaden
the protective capabilities of the influenza vaccines in current
use. The innate immune response to influenza virus involves
activation of multiple pathways of pattern-recognition recep-
tors in immune and nonimmune cells, including TLRs, RAinducible gene I-like receptors (RLR), and the NALP3 inflammasome (59). Currently licensed inactivated influenza vaccines
administered by the i.m. route induce systemic IgG Abs but fail
to induce highly cross-reactive mucosal IgA or heterosubtypic
CD8⫹ CTL. Therefore, yearly influenza vaccines are developed
based upon prediction of the most common subtypes prevalent
in the coming flu season. Licensed attenuated influenza vaccine
in man can induce mucosal IgA and IgG Abs and CTL against
virus subtypes, but their use is precluded due to adverse effects
in immunocompromised individuals.
A recent study found that mice who received two i.n. immunizations of H1N1 influenza virus-like particles (VLPs) were
protected against a lethal challenge with both the 1918 H1N1
virus and the heterotypic H5N1 virus (60). In addition, mucosal immunization was able to induce a higher level of cross-reactive IgG and IgA Abs compared with parenteral vaccination
with the same vaccine (60). Mice immunized i.m. with two
doses of 1918 VLPs were only protected against the homologous virus challenge. Mucosal vaccination with H1N1 VLPs in
ferrets also provided complete protection against lethal challenge with the H5N1 virus, whereas only 50% of ferrets survived infection after i.m. vaccination (60). Pulmonary immunization with extremely low doses (375-fold reduction in Ag
dose) of influenza vaccine with the ISCOMATRIX adjuvant
induced significantly higher titer serum Abs, and neutralizing
anti-hemagglutinin Abs were only present in the lung after vaccination via the respiratory tract (61). In the cotton rat model,
partial protection against different influenza A subtypes has
been achieved by i.n. immunization with a live virus or by i.m.
vaccination with an inactivated virus (62). Another study found
that parenteral immunization with influenza vaccine in humans
could induce both systemic immunity (anti-influenza Ab in serum and Ag-specific CD4⫹T cells in PBMC) as well as IgA Abs
in saliva and IFN-␥-secreting T cells in palatine tonsils (63).
FluMist is the first live attenuated influenza vaccine and also the
first nasally administered vaccine to be marketed in the United
States for the prevention of flu in individuals 2– 49 years of age
(64). FluMist is administered as an i.n. spray and has been
shown to provide high level of protection against both matched
and mismatched strains in children and adults. In a direct comparison of FluMist and inactivated vaccine in children, individuals receiving FluMist expressed 35–53% fewer cases of flu illness caused by antigenically matched strains (64).
Live oral attenuated vaccine against typhoid fever (S. typhi Ty21a)
Salmonella infections transmitted by contaminated poultry and
eggs represent a major global health burden. The Ty21a vaccine is
available as enteric coated capsules or as a “liquid formulation” (lyophilized vaccine organisms reconstituted with buffer and water
into a vaccine mixture). A recent study demonstrated that the inductive site of immunization rather than the isotype commitment
of Ab-secreting cells (ASCs) was the determining factor in cytokine
responsiveness and migration of human effector B cells (18). Oral
immunization with a Salmonella vaccine induced a robust migration of specific IgA and IgM Ab-secreting cells that coexpressed
CCR9 and CCR10 toward the mucosal chemokines CCL25
(TECK) in the small bowel and CCL28 (MEC) in the colon (18).
After oral Ty21a vaccination, all circulating ASCs in peripheral
blood expressed the ␣4␤7 integrin and migrated to the mucosal
compartment, whereas only a portion of ASCs after parenteral
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
The first effective polio vaccine was developed in 1952 by Jonas
Salk at the University of Pittsburgh (Pittsburgh, PA). The Salk
or inactivated poliovirus vaccine is based on three wild-type reference strains (Mahoney, MEF-1, and Saukett) inactivated
with formalin. The oral poliovirus vaccine was developed by
Albert Sabin using attenuated live poliovirus that he had received from Hilary Koprowski (16, 55). Sabin’s vaccine was licensed in 1962 and the World Health Organization adopted a
policy for the exclusive use of OPV in developing countries.
The advantage attributed to the live attenuated vaccine is that it
persisted in the gastrointestinal tract for 4 – 6 wk and induced
both mucosal and serum-neutralizing anti-poliovirus Abs
against poliovirus types 1, 2, and 3. In contrast, the IPV is also
highly immunogenic, confers low titers of mucosal neutralizing
Abs, and affords protection, all qualities of a good vaccine.
However, parenteral vaccination with IPV does not appear to
be highly effective in inducing secretory Ab response in the nasopharynx or intestinal mucosa during primary immunization
(16). In some cases, IPV alone was shown to provide inadequate
protection against poliovirus, especially type 1 (56). A recent
study (1) characterized mucosal immunity in a polio-free community immunized with a combined enhanced inactivated/oral
polio vaccine. The percentage of excreted virus was inversely
correlated with prechallenged intestinal neutralizing Ab titers.
The authors concluded that the main factor in the interruption
of disease and virus transmission in the community was a memory Ab response induced by the vaccine. In the United States,
⬍10 cases of paralytic poliomyelitis are reported each year, and
all cases are associated with the vaccine (57). In developing
countries, the incidence of poliomyelitis is due to poor seroconversion rates after vaccination with IPV (57). In the United
States OPV is recommended for routine vaccination in part because of its ability to generate intestinal and pharyngeal immunity to enhance protection during reinfection (17). It was demonstrated that even enhanced potency IPV was less effective
than OPV in preventing and limiting intestinal infection, even
though the vaccine generated higher serum Ab titers (17). According to a recent publication, healthcare officials in Germany
and Switzerland recommended vaccination with IPV instead of
OPV (58) in an effort to achieve complete global eradication of
the virus. However, travelers vaccinated with IPV only might
become infected with vaccine-derived virus, shedding it in large
quantities and increasing the probability of transmission to unvaccinated individuals. It is therefore recommended to include
at least one dose of OPV in a polio vaccination schedule to generate strong intestinal immunity (58).
6885
6886
BRIEF REVIEWS: COMPARTMENTALIZED MUCOSAL IMMUNE RESPONSE AND PROTECTION
Live oral attenuated vaccine against cholera (V. cholerae CVD
103-HgR)
The CVD 103-HgR is constructed by deleting 94% of the enzymatically active A subunit of cholera toxin from the classical
Inaba V. cholerae O1 569B. This vaccine, administered as a “liquid formulation,” is highly immunogenic following a single oral
dose. In experimental challenge studies in volunteers, a single
dose of CVD 103-HgR resulted in intestinal colonization and
rapid immune responses that conferred protection for up to 6
mo against challenge with either the wild-type V. cholerae O1 or
the El Tor biotype and either the Inaba or the Ogawa serotype.
The best correlate of protection against cholera is the titer of
serum vibriocidal Abs directed against the O Ag of V. cholerae
O1. However, one study found no correlation between protection and secondary Ab responses following wild-type homologous challenge in individuals vaccinated with CVD 103-HgR
(68). It is possible that pre-existing immunity may limit the efficacy of the live oral vaccine among partially immune individuals. A safety concern associated with CVD 103-HgR is the
possibility of horizontal gene transfer and recombination, leading to reversion to virulence. In volunteer studies, CVD 103HgR provides 62–100% protection against cholera caused by
V. cholerae for at least 6 mo; however, results of a large-scale
field trial in Indonesia demonstrated lower protective efficacy.
A single dose of CVD 103-HgR was found to be immunogenic
and without adverse effects in HIV-infected adults in Mali (24).
The live attenuated vaccine strains V. cholerae CVD 103-HgR
and S. typhi Ty21a can be combined into an oral bivalent vaccine
without compromising the immunogenicity of the individual vaccine strains. Seroconversion rates of 87–94% for Inaba vibriocidal
Abs and 72–91% for anti-S. typhi LPS Abs were reported in
healthy volunteers receiving the bivalent vaccine.
The live attenuated bacillus Calmette-Guérin (BCG) vaccine against
tuberculosis (TB)
Since 1923, BCG vaccines have been used worldwide and, to date,
are the only licensed TB vaccines administered in humans. Over 3
billion people have been vaccinated with BCG, and over 100 million newborn children are vaccinated with BCG every year. BCG
induces protection against childhood TB but fails to protect
against pulmonary TB in adults. Because pulmonary TB represents ⬎85% of the total TB burden, new-generation TB vaccines
are urgently needed. A mucosally administered vaccine could provide better protection against M. tuberculosis infection in the respiratory mucosa. A recent study demonstrated that i.m. immunization with a M. tuberculosis DNA vaccine induced robust T cell
responses in the spleen but failed to generate Ag-specific T cells in
the airway lumen (28). Pulmonary protection against M. tuberculosis after systemic vaccination is critically determined by T cell geography and depends on whether Ag-specific CD8⫹ T cells are
located within or outside of the mucosal lumen. Airway boosting
immunizations with soluble Ag were shown to effectively mobilize
systemically activated T cells into the airway lumen. Ag-specific local airway T cells can provide significant protection against airway
M. tuberculosis challenge. Adoptive transfer of either local airway T
cells of i.n. vaccinated animals or splenic Ag-specific T cells of systemically immunized mice into the lungs of naive SCID mice
could confer protection against pulmonary M. tuberculosis exposure (69). Protection against pulmonary M. tuberculosis challenge
correlated with the generation of Ag 85A-specific, IFN-␥-producing T cells in draining lung lymph nodes. In vivo neutralization
experiments demonstrated that the accumulation of T cells in the
airway lumen was mediated by chemokines, including MIP-1␣ or
IFN-inducible protein-10 (28). A recombinant vesicular stomatitis
virus (VSVAg85A) was shown to be immunogenic in mice by either the i.n. or the i.m. route (70). However, only i.n. vaccination
with VSVAg85A generated a mucosal T cell response that was protective against pulmonary M. tuberculosis exposure. A heterologous
VSVAg85A vaccine boost following recombinant adenoviral
prime immunization translated into significantly better protective
efficacy against pulmonary challenge with M. tuberculosis than either vaccine used alone (70). For protection following a BCG
prime immunization, an i.n. AdAg85A boost was protective
whereas i.m. boosting with the same recombinant vector was not
effective. A recent study showed that i.n. administration of the antigenic fusion protein Ag85B-EAST-6 mixed with the mucosal
combined adjuvant vector CTA1-DD/ISCOM could significantly boost IFN-␥-secreting CD4⫹ T cells in the lung elicited by
prior BCG immunization and improved protection against M. tuberculosis challenge (25).
Candidate mucosal vaccines against HIV/AIDS
Gastrointestinal and vaginal mucosa are the primary sites of
natural HIV transmission and a major reservoir for virus replication. Approximately 30 – 40% of all new HIV-1 infections
occur in women through the vaginal mucosa. It is clear today
that an effective HIV vaccine must be capable of eliciting local
mucosal immune responses (both mucosal Ab and CD8⫹
CTL) to impact initial virus replication and prevent CD4⫹ depletion, enteropathy, immune dysregulation, and the rapid dissemination of virus from the mucosa into the systemic lymphoid tissue. HIV-1 envelope-specific mucosal IgA Abs have
been shown to block mucosal HIV entry in vitro as well as transcytosis of the virus through epithelial barriers. Highly exposed
seronegative women who have unprotected sex with HIV-positive individuals have been shown to secrete ␤-defensins and
have sIgA in their cervicovaginal secretions that could contribute to a temporary protective state (71). In addition, passive infusion of IgG1 neutralizing Abs has been shown to be effective
in the protection of macaques against vaginal transmission of
pathogenic simian HIV (SHIV). A recent study in macaques
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
Ty21a vaccination expressed mucosal homing markers (65). Oral
vaccination with Ty21a predominantly elicited immune response
in saliva and vaginal secretion, whereas rectal immunization was
most effective in inducing responses in nasal secretion, rectum, and
tears (53). Mucosal application of Salmonella Dublin flagellin containing microparticles has been shown to enhance transepithelial
transport by the follicle-associated epithelium (FAE) in mouse
Peyer’s patches (PPs) and rapid migration of subepithelial DCs
into the FAE (66). Thus, innate responses elicited by Salmonella
flagellin in mucosal epithelial cells are important in determining an
effective adaptive immune response (66).
In human clinical trials a two-dose regimen with the Ty21a
vaccine has proven to provide statistically significant protection
for the first 2 years, whereas a three dose-regimen provided protection for up to four years (67). The purified bacterial capsule
(Vi vaccine) given parenterally appears to be less toxic and
equally effective for up to 2 years compared with a single-dose
parenteral immunization with the whole cell vaccine. Two dose
regimens provided significant protection for 5 years, yet whole
cell vaccines were associated with higher toxicity (67).
The Journal of Immunology
CMIS vs CMIR and “long-lived” mucosal memory T and B cells
Evidence for a CMIS is based on the concept that primed B cells
from the gut subsequently enter the circulation and migrate to
local (intestinal LP) and distant mucosal tissues (the respiratory
and urogenital mucosae) and to glands where terminal differentiation occurs. For example, it is well known that intrarectal
immunization can elicit specific Abs not only in local rectal secretions but can also lead to Ab production at distant sites via
the CMIS. Intrarectal immunization with the Ty21a typhoid
vaccine generated Ag-specific IgA in saliva. However, local exposure to Ag can result in much higher levels of specific IgA in
the region of exposure than at distant sites. Intrarectal vaccination induces the most optimum mucosal immune responses in
the rectum and yet was ineffective for eliciting cholera toxin Bspecific Abs in the genital mucosa in women (45). The generation of vigorous and protective HIV immune responses in both
the rectum and the genital mucosa will likely require a combination of rectal and vaginal immunization. It is the authors’
opinion that compartmentalized mucosal immune responses in
the majority of cases play a much more significant role for mucosal vaccine protection compared with mucosal responses in
distant sites based on the concept of a common mucosal immune system.
Anatomic location directly impacts memory T cell formation
and functional programs. Mucosal memory T cell phenotypes
differ substantially depending on the regimen of immunization,
and secondary responses result in preferential accumulation of
memory T cells in the LP following mucosal immunization.
Thus, heterologous prime-boost immunization has been shown
to have a major effect on CD8 T cells’ functional quality and
surveillance within mucosal effector sites (74). Effector memory
cells at mucosal sites appear to be “long-lived” and can proliferate and exercise a similar function as that of central memory in
protection from disease (74). Furthermore, a direct correlation
between the magnitude and quality of effector CD8⫹ T cells at
mucosal sites and viral load was found during acute SIV infection (75). In our opinion, mucosal immunity and compartmentalized mucosal protection benefit the most from prime-boost
strategies, and such strategies should be widely applied for future mucosal vaccine development against infectious diseases
and cancer. However, to prevent viral transmission through a
mucosal surface, immune cells should be substantially activatable because the presence of memory T and B cells in LP may
not be sufficient for protection. Recently, a number of studies
have reported on the immunogenicity and safety of plant-derived, cold chain-free oral vaccines that effectively activate T
and B cell immunity at mucosal sites.
Role of mucosal DCs in the generation of protective immunity in mucosa
Mucosal DCs play a central role in both the induction and
maintenance of tolerance toward the normal microflora and the
foods we ingest as well as the induction of protective immunity
against invasive pathogens. Unique DC subsets are responsible
for Ag presentation following mucosal vaccination, and systemic immunization may have a limited effect on the delivery of
Ag to mucosal DCs (46, 76 – 81). Other nonhematopoietic cell
types, such as intestinal epithelial cells, that encounter a mucosal pathogen (e.g., invasive Salmonella typhimurium) serve important roles in the induction of adaptive immune responses
and can produce CXCL8, which rapidly attracts neutrophils to
the site of infection followed by inflammatory monocytes and
DC precursors, with eventual infiltration of lymphoid cells into
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
evaluated whether a plasma concentration of Ab corresponding
to modest neutralization titers in vitro could protect animals
against intravaginal challenge with low doses of SHIV that uses
the CCR5 coreceptor (72). It was demonstrated that lower titers of neutralizing Ab could provide a benefit in the context
of typical human exposure to HIV-1 (72). However, for protective T cell responses there is no consensus within the field as
to whether parenteral immunization can generate mucosal immunity sufficient to protect against mucosal exposure to
HIV-1, or whether a mucosal route of vaccination is capable of
generating the quality of compartmentalized local immunity
necessary for an effective HIV vaccine. A recent study by Kaufman et al., found that i.m. immunization with recombinant adenovirus generated CD8⫹ CTL responses at multiple mucosal
sites in both mice and rhesus macaques (39). The authors concluded that i.m. immunization with a replication-defective recombinant adenovirus was sufficient for induction of both systemic and long-lived tissue resident mucosal CD8⫹ T cell
memory, thereby bypassing the necessity of a mucosal route of
administration for the establishment of Ag-specific T cells at
mucosal effector sites (39). Another study by Pal et al., showed
that systemic immunization with an ALVAC-HIV recombinant vaccine decreased the viral set point in blood as well as in
mucosal sites and protected macaques from peripheral CD4⫹ T
cell depletion (36). It is important to note that in these two
studies a mucosal immunization regimen was not evaluated for
comparison of the relative efficacy of systemic vs mucosal delivery. Although results from a number of studies have demonstrated that mucosal immune responses are detected after systemic vaccination, comparative studies in macaques have
shown that a vaccine delivered to mucosal inductive sites can be
more effective in eliciting high avidity protective mucosal immune responses than the same vaccine given systemically. Results from our studies showed that a mucosal vaccine can induce
mucosal immune responses and protect macaques against mucosal infection with SHIV much more effectively than the same
vaccine given s.c. (42). In intrarecatally immunized macaques,
viral load was more effectively controlled in intestinal tissues,
which are major reservoirs for virus replication and persistence
(42). Although s.c. immunization could induce tetramer-positive CD8⫹ T cells in gut mucosa, functionally active CD8⫹
CTLs in mucosal compartments were best generated after intrarectal immunization (46). Subcutaneous vaccination generated higher avidity CD8⫹ T cells in systemic lymphoid tissue,
whereas mucosal immunization induced a higher proportion of
functionally active CD8⫹ CTLs in the GALT. Generation of
high-avidity CD8⫹ CTLs by the mucosally delivered vaccine
inversely correlated with viral load in the gut mucosa and directly correlated with the preservation of CD4⫹ T help in the
colonic LP after intrarectal challenge of macaques with SHIV
(46). Furthermore, we showed that a single systemic immunization is capable of inducing high avidity CTLs in regional
lymph nodes, whereas a mucosal heterologous prime-boost
strategy is critical for the generation of high-avidity CD8⫹ CTL
responses in mucosal tissue, at least in a small animal model test
system (73).
6887
6888
BRIEF REVIEWS: COMPARTMENTALIZED MUCOSAL IMMUNE RESPONSE AND PROTECTION
the inflammatory site (82). DC precursors recruited to the mesenteric lymph nodes (MLNs) during inflammation can undergo activation in situ before exposure to stromal and epithelial conditioning; they are fully capable of secreting IL-12 and
are potent inducers of Th1 IFN-␥ responses. During infection
with S. typhimurium, activation of a protective T cell response
requires the recruitment of CCR6⫹ DCs from the subepithelial
dome of PPs to the follicle-associated epithelium (83) (Fig. 2).
CX3CR1⫹ DCs also reside in the follicle-associated epithelium, where they can extend dendrites into the intestinal lumen
to sample Ags (Fig. 2). DCs normally resident in the MLNs
(CD103-MLN DCs) are capable of producing higher levels of
proinflammatory cytokines than intestine-derived CD103⫹
DCs and driving IFN-␥ production by CD4⫹ T cells (76) (Fig.
2). Although the PPs and MLNs are hotbeds of regulatory T
cells (Tregs) during normal homeostasis, innate signals delivered to DCs and activated T cells can temporarily suspend Treg
suppressive function and shift the balance from an antiinflammatory milieu characterized by cytokines IL-10 and
TGF-␤ toward a proinflammatory milieu of Il-1␤, IL-1␣, IL-6,
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
FIGURE 2. Overview of the compartmentalization of GALT. Left side, Cells responsible for converting vitamins A and D to their active metabolites play key roles
in maintaining homeostasis and oral tolerance. Epithelial cells, macrophages, and monocyte-derived DCs express the two sets of enzymes, CYP27A and CYP27B1
(25-hydroxyvitamin D3-1␣ hydroxylase) necessary for synthesis and autocrine/paracrine activity of 1,25(OH)2D3. The net effect is inhibition of differentiation and
maturation of DCs and an anti-inflammatory response through the induction of Th2 cells and IL-10-producing T regulatory type 1 (Tr1) cells. Specialized ␣E␤7
(CD103) DCs, and epithelial cells expressing retinal dehydrogenase (RALDH) convert vitamin A to its active metabolite, retinoic acid (RA). CD103⫹ RALDH⫹
DCs that line the subepithelial dome of Peyers’s patches and are activated following detachment from E cadherin carry antigenic peptides to the MLN via afferent
lymphatics, where they secrete TGF-␤ and RA to induce FoxP3 Tregs. RA produced by mucosal DCs is essential for both induction of local regulatory immune
responses and T cell and B cell expression of the homing markers ␣4␤7 and CCR9, which confer migration into the gastrointestinal mucosa. Additional cell types
and innate mechanisms responsible for maintaining intestinal mucosal homeostasis include Paneth cells located in intestinal crypts, invariant NKT (iNKT) cells, and
IL-22 secreting ROR␥t NKp46 cells found in cryptopatches within the LP. Commensal bacteria influence the context of immune cells in the small bowel and
steady-state differentiation of regulatory Th17 cells is dependent upon the presence of the cytophaga-flavobacter-bacteriodes (CFB) phylum. Adaptive immune
responses can be readily induced in MLNs by shifting the balance between DC subsets regulating homeostasis and those activated by pathogens and signals delivered
through TLRs, NOD-like receptors (NLRs), retinoic acid-inducible gene I-like helicases (RLHs), and C-type lectins. CD8␣⫹ DCs located in interfollicular regions
of PPs are capable of IL-12 production and cross-presentation of Ag to CD8 T cells. Ligation of TLR5 on CD103⫹ DCs by flagellin on pathogenic bacteria can
induce Th1 responses in MLNs. Inflammatory monocytes can capture Ag and differentiate into DCs secreting TNF-␣ and IL-12, skewing CD4⫹
Th1differentiation. DCs activated by extracellular bacteria and fungi secrete cytokines responsible for skewing Th17 differentiation and inhibiting iTregs. Constitutive expression of chemokines and chemokine receptors induced on T and B cells dictate the localization and retention of memory cells in discrete mucosal niches.
Homing of memory T and B cells requires expression of ␣4␤7 and CCR9 to interact with their ligands, mucosal addressin cell adhesion molecule 1 (MAdCam-1),
expressed on high endothelial venules in PPs, and CCL25, constitutively expressed in the small intestine. Tissue-resident memory T cells also express CCR10 and
interact with the ligand CCL28, expressed in colonic mucosal tissues. Right side, Commensal bacteria sampled by DCs in the PPs induce local IgA responses. CD103,
RALDH, inducible NO synthase DCs generate NO, which is essential for both T-dependent and T-independent IgA responses. IgA class switching occurs predominantly in the B cell follicles in PPs. APRIL secreted by NO-stimulated DCs is primarily responsible for in situ IgA ASC and sIgA. TGF-␤, in addition to CD4⫹
Th provided through CD40 ligand (CD40L), is required for T-dependent IgA and IgG responses, whereas IL-5 or IL-6 is needed to differentiate into ASCs.
IgA-secreting cells exhibit a short half-life, and it is unclear whether long-lived IgA plasma cells exist in PPs. Two subsets of memory B cells, conventional CD27⫹
memory B cells and FCRL4⫹ CD27⫺ B cells, undergo affinity maturation and somatic hypermutation in germinal centers. The different functional properties of
these two subsets and the question of whether FCRL4⫹ memory B cells preferentially localize in mucosal tissue have yet to be determined. Tfh, T follicular helper;
TSLP, thymic stromal lymphopoietin.
The Journal of Immunology
Th17 cells and mucosal protection
Th17 cells have emerged as a unique mucosal CD4⫹ effector
cell population responsible for protection from extracellular
bacteria and fungi. Effector function is attributed to the interaction of its signature cytokine IL-17 with IL-17R expressed on
fibroblasts and epithelial cells to induce chemokines such as
MCP-2 and G-CSF and CXC chemokines. Enhanced recruitment of neutrophils and granulopoiesis leads to pathogen clearance, whereas CXCR-3 ligating chemokines can activate macrophages and recruit additional immune cells, including CD4⫹
Th1 cells. IL-22 is another important effector cytokine produced by Th17 cells for inducing the secretion of antimicrobial
peptides and ␤-defensin-2 by epithelial cells and for contributing to barrier function and tissue repair (Fig. 2). In the gut,
Th17 cells are the main sources of IL-17, whereas in the lung
␥␦T cells, NKT, NK, and ROR␥t NKp46⫹ cells are the major
producers of this cytokine. The rules governing Th17 differentiation in vivo reflect tissue localization and the balance of inflammatory and anti-inflammatory cytokines induced by
pathogen-activated DCs. In the PPs and MLNs, DC production of TGF-␤ and RA plus IL-2 leads to the preferential induction of induced Treg (iTreg). TGF-␤ and IL-10 are pivotal
cytokines in the control of inflammation induced by TLR activation, suppressing the MyD88 signaling pathway and type I
IFN production. Pathogens that induce robust tissue inflammation and proinflammatory cytokines can shift this balance
toward differentiation of Th17 cells (Fig. 1). Human CD4 cells
most efficiently differentiate into Th17 cells by a combination
of TGF-␤ and IL-6 plus IL-1␤ or IL-␤ plus IL-23 or IL-21.
IL-6-STAT3 signaling is critical for Th17 differentiation, inducing expression of two transcription factors, ROR␥␶ and
ROR␣, and the IL-23R. IL-21 and IL-23 also signal through
STAT3, stabilizing the Th17 phenotype and amplifying effector function by decreasing IL-10 production and increasing
trafficking. Patients with an autosomal dominant STAT3
mutation (hyper-IgE syndrome/Jobs syndrome) are deficient in
Th17 cell differentiation and suffer mucocutaneous Candida
and recurrent pyogenic infections.
Although Th17 inflammatory responses have been shown to
play a key role in both protection and pathology to Gram-negative extracellular bacteria and fungi as well as autoimmune diseases, their role in enforcing mucosal barrier function and protective immunity to intracellular bacteria and viral infections is
beginning to be appreciated. A recent study showed that strong
influenza-specific Th17 responses develop in IL-10-deficient
mice and can provide protection from lethal challenge by mechanisms independent of Th1 helper functions (perforin-mediated cytotoxicity and IFN-␥) or IL-17A (91). Another study
suggests that CCR6 ␣4␤7high Th17 central memory cells in gut
mucosa are the initial population of cells infected following SIV
infection. This result is not surprising considering that tissueresident Th17 cells in the small intestine LP comprise ⬃80% of
the total CD4 cells and express CCR6 and high levels of CCR5.
Furthermore, depletion of this cell population in the gut mucosa is predictive of subsequent disease pathology (92). It will be
important to determine whether mucosal vaccine strategies that
induce HIV specific-Th17/Th1 effector memory responses can
reverse the outcome of CD4⫹ Th17 ␣4␤7highCCR5high cells as
targets of HIV infection into aggressive effector cells capable of
contributing to a protective response.
Similarly as in the reciprocal transcriptional regulation of
iTregs and Th17 differentiation by TGF␤, Th17 cells subsequently share a common branch point with T follicular helper
cells (Fig. 2). Although both subsets rely on autocrine IL-21 for
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
IL-12, IFN-␥, and IFN-␣ leading to the induction of adaptive
immune responses calibrated to the pathogenic threat (Fig. 2).
A recent study showed that IL-10-secreting Tr1 regulatory T
cells lose their suppressive functions and convert to IFN-␥- and
IL-4 secreting T cells following CpG ligation of TLR9 up-regulated on Tr1 cells by 1␣,25-dihydroxyvitamin D3 (84). In addition, TLR8 ligation expressed on natural FoxP3⫹CD4⫹
CD25⫹ cells abrogates Treg suppression, whereas TLR2
ligation results in temporary loss of suppressor activity (85, 86).
A recent study linked the sphingosine 1-phosphate receptor
type 1-Akt-mammalian targets of rapamycin kinase pathway
(S1P1-Akt-mTOR) to suppression of the FoxP3 Treg function,
suggesting one mechanism for the differential regulation of
Treg function and the activation of naive T cells for induction
of adaptive immune responses in MLNs (87). In addition,
CD4⫹ T effector cells stimulated through TLR5 are refractory
to the suppressive effects of Tregs. Understanding the mechanisms that influence the balance between mucosal homeostasis
and the induction of adaptive immune responses has important
implications for mucosal vaccine design and delivery strategies.
Diverse populations of respiratory DCs have been characterized
in both normal and inflamed lungs (88). A recent study demonstrated that in response to influenza infection, two distinct
subsets of migrant respiratory DCs expressing CD103⫹ and
CD11b⫹ were involved in presenting Ag to naive CD4⫹ and
CD8⫹ T cells in the draining lymph nodes (88). However, airway-derived migrant CD103⫹ DCs in the inflamed MLNs
were superior at cross-presenting influenza virus Ags to naive
CD8⫹ T lymphocytes (88). Similarly, only the migratory subtype of CD103⫹ skin-derived DCs efficiently presented herpes
simplex virus type 1 Ags for the induction of naive CD8⫹ T
cells (89). DCs can be recruited to the intestinal LP as a result of
local inflammation.
A recent study identified a population of CD11chigh
CD11bhigh LP DCs that express TLR5 and produce proinflammatory cytokines such as IL-6 and IL-12, but not IL-23 or IL10, in response to flagellin (90) (Fig. 2). Different DC
populations are also responsible for priming T cells dependent
upon the route of HSV-1 infection (78). Following vaginal infection, HSV-1 Ags were largely presented by tissue-derived migratory DCs, whereas after needle injection HSV-1 was rapidly
presented by lymph node-resident DCs to rare Ag-specific T
cells (78). Understanding the unique properties of mucosal
DCs and the mucosal milieu in regulating local T and B cell
responses and “mucosal memory” will be critical in determining
how best to deliver vaccines for protection from mucosal infection. Studies in diverse infection models involving different
mucosal compartments have clearly shown that targeting local
mucosal DCs and the induction of local immune responses
by mucosal vaccination is more effective in containing mucosal
infections, as our understanding of the magnitude and quality
of responses and the tissue residency of cells migrating to mucosal sites following systemic immunization needs further investigation. Developing mucosal immunization strategies will
depend upon further understanding of how best to induce a
compartmentalized immune response tailored to the specific
pathogen and calibrated to the potential pathogenic threat.
6889
6890
BRIEF REVIEWS: COMPARTMENTALIZED MUCOSAL IMMUNE RESPONSE AND PROTECTION
their differentiation, lineage commitment might be dependent
on additional signals delivered by DCs such as IL-1␤ and IL-23
or ICOSL to determine CCR6 Th17 or CXCR5 T follicular
helper cell fate. Thus, Th17 and Treg cells are the main modulators of the balance between tolerance and inflammation in
mucosal tissue and are involved both in primary infection and
the recall responses seen in vaccine studies. These cells might be
exploited to develop the most effective mucosal vaccines against
pathogens causing chronic infections such as HIV, HCV, herpes viruses, mycobacteria, and fungi.
Concluding remarks
Acknowledgments
We thank Drs. David Franz, Niveen Mulholland, Kenton Lohman, and John
Lednicky (Midwest Research Institute, Frederick, MD) for critical reading of
the manuscript and helpful suggestions. We apologize to the authors whose
work we were unable to include in our citations due to space limitations.
Disclosures
The authors have no financial conflict of interest.
References
1. Swartz, T. A., M. S. Green, R. Handscher, D. Sofer, M. Cohen-Dar, T. Shohat,
S. Habib, E. Barak, Z. Dror, E. Somekh, T. Peled-Leviathan, et al. 2008. Intestinal
immunity following a combined enhanced inactivated polio vaccine/oral polio vaccine
programme in Israel. Vaccine 26: 1083–1090.
2. John, T. J. 2001. Anomalous observations on IPV and OPV vaccination. Dev. Biol.
(Basel) 105: 197–208.
3. Kaul, D., and P. L. Ogra. (1998) Mucosal responses to parenteral and mucosal vaccines. Dev. Biol. Stand. 95: 141–146.
4. Jang, M. H., M. N. Kweon, K. Iwatani, M. Yamamoto, K. Terahara, C. Sasakawa,
T. Suzuki, T. Nochi, Y. Yokota, P. D. Rennert, et al. 2004. Intestinal villous M cells:
an antigen entry site in the mucosal epithelium. Proc. Natl. Acad. Sci. USA 101:
6110 – 6115.
5. Meng, G., X. Wei, X. Wu, M. T. Sellers, J. M. Decker, Z. Moldoveanu,
J. M. Orenstein, M. F. Graham, J. C. Kappes, J. Mestecky, et al. 2002. Primary intestinal epithelial cells selectively transfer R5 HIV-1 to CCR5⫹ cells. Nat. Med. 8:
150 –156.
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
New design and delivery strategies for eliciting local mucosal
immunity are urgently needed for developing effective prophylactic and therapeutic vaccines against mucosal pathogens such
as HIV, HPV, influenza, herpes viruses, and M. tuberculosis, as
well as pathogenic enteric and fungal infections. Many vaccinologists feel that parenteral immunization is sufficient, given the
history of successful vaccination in humans. However, numerous studies in human clinical trial and experimental settings
have demonstrated that mucosal vaccine strategies are more effective in the prevention of pathogen transmission and the resolution of infection at the site of initial pathogen entry. An optimum mucosal vaccine strategy can activate local innate
immune responses that translate into the generation of neutralizing sIgA Abs, polyfunctional CD8⫹ CTLs in effector target
sites, and CD4⫹ T cell effector memory responses, whereas systemic vaccine strategies are much less effective in eliciting the
compartmentalized mucosal immunity necessary for these protective responses. Targeting Ags to mucosal DC that imprint
gut-homing receptors on pathogen-specific T and B cells and
the induction of Th17/Th1 responses will require novel mucosal vaccine strategies, including effective adjuvants and immunomodulatory molecules. Compared with adjuvants for parenteral vaccination, mucosal adjuvants today are much less
effective. Advances in new design and delivery strategies and
novel mucosal adjuvants will greatly improve the efficacy of
mucosal vaccination and contribute to the broad implementation of effective mucosal vaccines in the future.
6. Stahl-Hennig, C., R. M. Steinman, K. Tenner-Racz, M. Pope, N. Stolte,
K. Mätz-Rensing, G. Grobschupff, B. Raschdorff, G. Hunsmann, and P. Racz. 1999.
Rapid infection of oral mucosal-associated lymphoid tissue with simian immunodeficiency virus. Science 285: 1261–1265.
7. Geijtenbeek, T. B., D. S. Kwon, R. Torensma, S. J. van Vliet, G. C. van Duijnhoven,
J. Middel, I. L. Cornelissen, H. S. Nottet, V. N. KewalRamani, D. R. Littman, et al.
2000. DC-SIGN, a dendritic cell-specific HIV-1-binding protein that enhances transinfection of T cells. Cell 100: 587–597.
8. Abel, K., B. Pahar, K. K. Van Rompay, L. Fritts, C. Sin, K. Schmidt, R. Colón,
M. McChesney, and M. L. Marthas. 2006. Rapid virus dissemination in infant macaques after oral simian immunodeficiency virus exposure in the presence of local innate immune responses. J. Virol. 80: 6357– 6367.
9. Hladik, F., P. Sakchalathorn, L. Ballweber, G. Lentz, M. Fialkow, D. Eschenbach, and
M. J. McElrath. 2007. Initial events in establishing vaginal entry and infection by
human immunodeficiency virus type-1. Immunity 26: 257–270.
10. Belyakov, I. M., J. D. Ahlers, and J. A. Berzofsky. 2004 Mucosal AIDS vaccines: current status and future directions. Expert Rev. Vaccines 3(4 Suppl.): S65–S73.
11. Berzofsky, J. A., J. D. Ahlers, and I. M. Belyakov. 2001. Strategies for designing and
optimizing new generation vaccines. Nat. Rev. Immunol. 1: 209 –219.
12. Ichinohe, T., H. K. Lee, Y. Ogura, R. Flavell, and A. Iwasaki. 2009. Inflammasome
recognition of influenza virus is essential for adaptive immune responses. J. Exp. Med.
206: 79 – 87.
13. Spira, A. I., P. A. Marx, B. K. Patterson, J. Mahoney, R. A. Koup, S. M. Wolinsky, and
D. D. Ho. 1996. Cellular targets of infection and route of viral dissemination after an
intravaginal inoculation of simian immunodeficiency virus into rhesus macaques.
J. Exp. Med. 183: 215–225.
14. Belyakov, I. M., V. A. Kuznetsov, B. Kelsall, D. Klinman, M. Moniuszko, M. Lemon,
P. D. Markham, R. Pal, J. D. Clements, M. G. Lewis, et al. 2006. Impact of vaccineinduced mucosal high avidity CD8⫹ CTLs in delay of AIDS-viral dissemination from
mucosa. Blood 107: 3258 –3264.
15. Veazey, R. S., M. DeMaria, L. V. Chalifoux, D. E. Shvetz, D. R. Pauley, H. L. Knight,
M. Rosenzweig, R. P. Johnson, R. C. Derosiers, and A. A. Lackner. 1998. Gastrointestinal tract as a major site of CD4⫹ T cell depletion and viral replication in SIV
infection. Science 280: 427– 431.
16. Ogra, P. L. 1984. Mucosal immune response to poliovirus vaccines in childhood. Rev.
Infect. Dis. 6(Suppl. 2): S361–S368.
17. Onorato, I. M., J. F. Modlin, A. M. McBean, M. L. Thomas, G. A. Losonsky, and
R. H. Bernier. 1991. Mucosal immunity induced by enhance-potency inactivated and
oral polio vaccines. J. Infect. Dis. 163: 1– 6.
18. Sundström, P., S. B. Lundin, L. A. Nilsson, and M. Quiding-Järbrink. 2008. Human
IgA-secreting cells induced by intestinal, but not systemic, immunization respond to
CCL25 (TECK) and CCL28 (MEC). Eur. J. Immunol. 38: 3327–3338.
19. Stephenson, I., M. C. Zambon, A. Rudin, A. Colegate, A. Podda, R. Bugarini,
G. Del Giudice, A. Minutello, S. Bonnington, J. Holmgren, et al. 2006. Phase I evaluation of intranasal trivalent inactivated influenza vaccine with nontoxigenic Escherichia coli enterotoxin and novel biovector as mucosal adjuvants, using adult volunteers. J. Virol. 80: 4962– 4970.
20. Viret, J. F., D. Favre, B. Wegmüller, C. Herzog, J. U. Que, S. J. Cryz, Jr., and
A. B. Lang. 1999. Mucosal and systemic immune responses in humans after primary
and booster immunizations with orally administered invasive and noninvasive live attenuated bacteria. Infect. Immun. 67: 3680 –3685.
21. Belshe, R. B., P. M. Mendlelman, J. Treanor, J. King, W. C. Gruber, P. Piedra,
D. I. Bernstein, F. G. Hayden, K. Kotloff, K. Zangwill, et al. 1998. The efficacy of live
attenuated, cold-adapted, trivalent, intranasal influenzavirus vaccine in children.
N. Engl. J. Med. 338: 1405–1412.
22. Nichol, K. L., P. M. Mendelman, K. P. Mallon, L. A. Jackson, G. J. Gorse,
R. B. Belshe, W. P. Glezen, and J. Wittes. 1999. Effectiveness of live, attenuated intranasal influenza virus vaccine in healthy, working adults: a randomized controlled
trial. J. Am. Med. Assoc. 282: 137–144.
23. Agnello, D., C. A. Hervé, A. Lavaux, M. Darniot, P. Guillon, A. Charpilienne, and
P. Pothier. 2006. Intrarectal immunization with rotavirus 2/6 virus-like particles induces an antirotavirus immune response localized in the intestinal mucosa and protects against rotavirus infection in mice. J. Virol. 80: 3823–3832.
24. Perry, R. T., C. V. Plowe, B. Koumaré, F. Bougoudogo, K. L. Kotloff, G. A. Losonsky,
S. S. Wasserman, and M. M. Levine. 1998. A single dose of live oral cholera vaccine
CVD 103-HgR is safe and immunogenic in HIV-infected and HIV-noninfected
adults in Mali. Bull. World Health Organ. 76: 63–71.
25. Andersen, C. S., J. Dietrich, E. M. Agger, N. Y. Lycke, K. Lövgren, and P. Andersen.
2007. The combined CTA1-DD/ISCOMs vector is an effective intranasal adjuvant
for boosting prior Mycobacterium bovis BCG immunity to Mycobacterium tuberculosis.
Infect. Immun. 75: 408 – 416.
26. McNeal, M. M., M. Basu, J. A. Bean, J. D. Clements, N. Y. Lycke, A. Ramne,
B. Löwenadler, A. H. Choi, and R. L. Ward. 2007. Intrarectal immunization of mice
with VP6 and either LT(R192G) or CTA1-DD as adjuvant protects against fecal rotavirus shedding after EDIM challenge. Vaccine 25: 6224 – 6231.
27. Goonetilleke, N. P., H. McShane, C. M. Hannan, R. J. Anderson, R. H. Brookes, and
A. V. S. Hill. 2003. Enhanced immunogenicity and protective efficacy against Mycobacterium tuberculosis of bacille Calmette-Guerin vaccine using mucosal administration and boosting with a recombinant modified vaccinia virus Ankara. J. Immunol.
171: 1602–1609.
28. Jeyanathan, M., J. Mu, K. Kugathasan, X. Zhang, D. Damjanovic, C. Small,
M. Divangahi, B. J. Petrof, C. Hogaboam, and Z. Xing. 2008. Airway delivery of
soluble mycobacterial antigens restores protective mucosal immunity by single intramuscular plasmid DNA tuberculosis vaccination: role of proinflammatory signals in
the lung. J. Immunol. 181: 5618 –5626.
29. Wang, J., L. Thorson, R. W. Stokes, M. Santosuosso, K. Huygen, A. Zganiacz,
M. Hitt, and Z. Xing. 2004. Single mucosal, but not parenteral, immunization with
The Journal of Immunology
30.
31.
32.
33.
34.
35.
36.
38.
39.
40.
41.
42.
43.
44.
45.
46.
47.
48.
49.
50.
51.
52.
53.
54.
55.
56.
57.
58.
59.
60.
61.
62.
63.
64.
65.
66.
67.
68.
69.
70.
71.
72.
73.
74.
75.
76.
77.
78.
79.
and enhancement of resistance by local administration of IL-12. J. Clin. Invest. 102:
2072–2081.
Kantele, A., M. Häkkinen, Z. Moldoveanu, A. Lu, E. Savilahti, R. D. Alvarez,
S. Michalek, and J. Mestecky. 1998. Differences in immune responses induced by oral
and rectal immunizations with Salmonella typhi Ty21a: evidence for compartmentalization within the common mucosal immune system in humans. Infect. Immun. 66:
5630 –5635.
Bienenstock, J., A. D. Befus, M. McDermott, S. Mirski, K. Rosenthal, and
A. Tagliabue. 1983. The mucosal immunological network: compartmentalization of
lymphocytes, natural killer cells, and mast cells. Ann. NY Acad. Sci. 409: 164 –170.
Koprowski, H. 2006. First decade (1950 –1960) of studies and trials with the polio
vaccine. Biologicals 34: 81– 86.
World health organization collaborative study group on oral and inactivated poliovirus vaccines. 1996. Combined immunization of infants with oral and inactivated poliovirus vaccines: results of a randomized trial in The Gambia, Oman, and Thailand.
Bull. World Health Organ. 74: 253–268.
Faden, H. 1993. Poliovirus vaccination: a trilogy. J. Infect. Dis. 168: 25–28.
Knolle, H. T., A. Egli, and U. Candrian. 2004. [The perspective of global eradication
of poliomyelitis]. Gesundheitswesen 66: 1– 6.
Ichinohe, A. Iwasaki, and H. Hasegawa. 2008. Innate sensors of influenza virus: clues
to developing better intranasal vaccines. Expert Rev. Vaccines. 7: 1435–1445.
Perrone, L. A., A. Ahmad, A. Vequilla, X. Lu, G. Smith, J. M. Katz, P. Pushko, and
T. M. Tumpey. 2009. Intranasal vaccination with 1918 influenza virus-like particles
protects mice and ferrets from lethal 1918 and H5N1 influenza virus challenge. J. Virol. 83: 5726 –5734.
Wee, J. L., J. P. Scheerlinck, K. J. Snibson, S. Edwards, M. Pearse, C. Quinn, and
P. Sutton. 2008. Pulmonary delivery of ISCOMATRIX influenza vaccine induces
both systemic and mucosal immunity with antigen dose sparing. Mucosal Immunol. 1:
489 – 496.
Straight, T. M., M. G. Ottolini, G. A. Prince, and M. C. Eichelberger. 2008. Antibody contributes to heterosubtypic protection against influenza A-induced tachypnea
in cotton rats. Virol. J. 5: 44.
Guthrie, T., C. G. Hobbs, V. Davenport, R. E. Horton, R. S. Heyderman, and
N. A. Williams. 2004. Parenteral influenza vaccination influences mucosal and systemic T cell-mediated immunity in healthy adults. J. Infect. Dis. 190: 1927–1935.
Ambrose, C. S., C. Luke, and K. Coelingh. 2008. Current status of live attenuated
influenza vaccine in the United States for seasonal and pandemic influenza. Influenza
Other Respi. Viruses 2: 193–202.
Kantele, A., J. M. Kantele, E. Savilahti, M. Westerholm, H. Arvilommi, A. Lazarovits,
E. C. Butcher, and P. H. Makela. 1997. Homing potentials of circulating lymphocytes
in humans depend on the site of activation: oral, but not parenteral, typhoid vaccination induces circulating antibody-secreting cells that all bear homing receptors directing them to the gut. J. Immunol. 158: 574 –579.
Chabot, S. M., M. Shawi, T. Eaves-Pyles, and M. R. Neutra. 2008. Effects of flagellin
on the functions of follicle-associated epithelium. J. Infect. Dis. 198: 907–910.
Engels, E. A., and J. Lau. 2000. Vaccines for preventing typhoid fever. Cochrane Database Syst Rev. CD001261.
Losonsky, G. A., C. O. Tacket, S. S. Wasserman, J. B. Kaper, and M. M. Levine. 1993.
Secondary Vibrio cholerae-specific cellular antibody responses following wild-type homologous challenge in people vaccinated with CVD 103-HgR live oral cholera vaccine: changes with time and lack of correlation with protection. Infect. Immun. 61:
729 –733.
Santosuosso, M., X. Zhang, S. McCormick, J. Wang, M. Hitt, and Z. Xing. 2005.
Mechanisms of mucosal and parenteral tuberculosis vaccinations: adenoviral-based
mucosal immunization preferentially elicits sustained accumulation of immune protective CD4 and CD8 T cells within the airway lumen. J. Immunol. 174: 7986 –7994.
Roediger, E. K., K. Kugathasan, X. Zhang, B. D. Lichty, and Z. Xing. 2008. Heterologous boosting of recombinant adenoviral prime immunization with a novel vesicular stomatitis virus-vectored tuberculosis vaccine. Mol. Ther. 16: 1161–1169.
Kaul, R., F. Plummer, M. Clerici, M. Bomsel, L. Lopalco, and K. Broliden. 2001.
Mucosal IgA in exposed, uninfected subjects: evidence for a role in protection against
HIV infection. AIDS 15: 431– 432.
Hessell, A. J., P. Poignard, M. Hunter, L. Hangartner, D. M. Tehrani, W. K. Bleeker,
P. W. Parren, P. A. Marx, and D. R. Burton. 2009. Effective, low-titer antibody protection against low-dose repeated mucosal SHIV challenge in macaques. Nat. Med. 15:
951–954.
Belyakov, I. M., J. D. Ahlers, G. J. Nabel, B. Moss, and J. A. Berzofsky. 2008. Generation of functionally active HIV-1 specific CD8⫹ CTL in intestinal mucosa following mucosal, systemic or mixed prime-boost immunization. Virology 381: 106 –115.
Masopust, D., S.-J. Ha, V. Vezys, and R. Ahmed. 2006. Stimulation history dictates
memory CD8 T cell phenotype: implications for prime-boost vaccination. J. Immunol. 177: 831– 839.
Li, Q., P. J. Skinner, S. J. Ha, L. Duan, T. L. Mattila, A. Hage, C. White, D. L. Barber,
L. O’Mara, P. J. Southern, et al. 2009. Visualizing antigen-specific and infected cells
in situ predicts outcomes in early viral infection. Science 323: 1726 –1729.
Coombes, J. L., and F. Powrie. 2008. Dendritic cells in intestinal immune regulation.
Nat. Rev. Immunol. 8: 435– 446.
Johansson-Lindbom, B., M. Svensson, O. Pabst, C. Palmqvist, G. Marquez,
R. Förster, and W. W. Agace. 2005. Functional specialization of gut CD103⫹ dendritic cells in the regulation of tissue-selective T cell homing. J. Exp. Med. 202:
1063–1073.
Lee, H. K., M. Zamora, M. M. Linehan, N. Iijima, D. Gonzalez, A. Haberman, and
A. Iwasaki. 2009. Differential roles of migratory and resident DCs in T cell priming
after mucosal or skin HSV-1 infection. J. Exp. Med. 206: 359 –370.
Kelsall, B. 2008. Recent progress in understanding the phenotype and function of
intestinal dendritic cells and macrophages. Mucosal Immunol. 1: 460 – 469.
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017
37.
recombinant adenoviral-based vaccine provides potent protection from pulmonary tuberculosis. J. Immunol. 173: 6357– 6365.
Neutra, M. R., and P. A. Kozlowski. 2006 Mucosal vaccines: the promise and the
challenge. Nat. Rev. Immunol. 6: 148 –158.
Belyakov, I. M., and J. D. Ahlers. 2008. Functional CD8⫹ CTLs in mucosal sites and
HIV infection: moving forward toward a mucosal AIDS vaccine. Trends Immunol. 29:
574 –585.
Ahlers, J. D., and I. M. Belyakov. 2009. Strategies for optimizing targeting and delivery of mucosal HIV vaccines. Eur. J. Immunol. 39: 2657–2669.
Suzich, J. A., S. J. Ghim, F. J. Palmer-Hill, W. I. White, J. K. Tamura, J. A. Bell,
J. A. Newsome, A. B. Jenson, and R. Schlegel. 1995. Systemic immunization with
papillomavirus L1 protein completely prevents the development of viral mucosal papillomas. Proc. Nat. Acad. Sci. USA 92: 11553–11557.
Haan, L., W. R. Verweij, M. Holtrop, R. Brands, G. J. van Scharrenburg,
A. M. Palache, E. Agsteribbe, and J. Wilschut. 2001. Nasal or intramuscular immunization of mice with influenza subunit antigen and the B subunit of Escherichia coli
heat-labile toxin induces IgA- or IgG-mediated protective mucosal immunity. Vaccine
19: 2898 –2907.
Morello, C. S., M. Ye, S. Hung, L. A. Kelley, and D. H. Spector. 2005. Systemic
priming-boosting immunization with a trivalent plasmid DNA and inactivated murine cytomegalovirus (MCMV) vaccine provides long-term protection against viral
replication following systemic or mucosal MCMV challenge. J. Virol. 79: 159 –175.
Pal, R., D. Venzon, S. Santra, V. S. Kalyanaraman, D. C. Montefiori, L. Hocker,
L. Hudacik, N. Rose, J. Nacsa, Y. Edghill-Smith, et al. 2006. Systemic immunization
with an ALVAC-HIV-1/protein boost vaccine strategy protects rhesus macaques from
CD4⫹ T-cell loss and reduces both systemic and mucosal SHIVKU2 RNA levels.
J. Virol. 80: 3732–3742.
Tatsis, N., S. W. Lin, K. Harris-McCoy, D. A. Garber, M. B. Feinberg, and H. C. Ertl.
2007. Multiple immunizations with adenovirus and MVA vectors improve CD8⫹ T
cell functionality and mucosal homing. Virology 367: 156 –167.
Lin, S. W., A. S. Cun, K. Harris-McCoy, and H. C. Ertl. 2007. Intramuscular rather
than oral administration of replication-defective adenoviral vaccine vector induces
specific CD8⫹ T cell responses in the gut. Vaccine 25: 2187–2193.
Kaufman, D. R., J. Liu, A. Carville, K. G. Mansfield, M. J. E. Havenga, J. Goudsmit,
and D. H. Barouch. 2008. Trafficking of antigen-specific CD8⫹ T lymphocytes to
mucosal surfaces following intramuscular vaccination. J. Immunol. 181: 4188 – 4198.
Hansen, S. G., C. Vieville, N. Whizin, L. Coyne-Johnson, D. C. Siess,
D. D. Drummond, A. W. Legasse, M. K. Axthelm, K. Oswald, C. M. Trubey, et al.
2009. Effector memory T cell responses are associated with protection of rhesus monkeys from mucosal simian immunodeficiency virus challenge. Nat. Med. 15: 293–299.
Tineke, M., P. T. Herremans, J. H. J. Reimerink, A. M. Buisman, T. G. Kimman, and
M. P. G. Koopmans. 1999. Induction of mucosal immunity by inactivated poliovirus
vaccine is dependent on previous mucosal contact with live virus. J. Immunol. 162:
5011–5018.
Belyakov, I. M., Z. Hel, B. Kelsall, V. A. Kuznetsov, J. D. Ahlers, J. Nacsa,
D. I. Watkins, T. M. Allen, A. Sette, J. Altman, et al. 2001. Mucosal AIDS vaccine
reduces disease and viral load in gut reservoir and blood after mucosal infection of
macaques. Nat. Med. 7: 1320 –1326.
Gallichan, W. S., and K. L. Rosenthal. 1996. Long-lived cytotoxic T lymphocyte
memory in mucosal tissues after mucosal but not systemic immunization. J. Exp. Med.
184: 1879 –1890.
Revaz, V., J. Benyacoub, W. M. Kast, J. T. Schiller, P. De Grandi, and
D. Nardelli-Haefliger. 2001. Mucosal vaccination with a recombinant Salmonella typhimurium expressing human papillomavirus type 16 (HPV16) L1 virus-like particles
(VLPs) or HPV16 VLPs purified from insect cells inhibits the growth of HPV16expressing tumor cells in mice. Virology 279: 354 –360.
Kozlowski, P. A., S. Cu-Uvin, M. R. Neutra, and T. P. Flanigan. 1997. Comparison
of the oral, rectal, and vaginal immunization routes for induction of antibodies in
rectal and genital tract secretions of women. Infect. Immun. 65: 1387–1394.
Belyakov, I. M., D. Isakov, Q. Zhu, A. Dzutsev, and J. A. Berzofsky. 2007. A novel
functional CTL avidity/activity compartmentalization to the site of mucosal immunization contributes to protection of macaques against simian/human immunodeficiency viral depletion of mucosal CD4⫹ T cells. J. Immunol. 178: 7211–7221.
Li, Z., M. Zhang, C. Zhou, X. Zhao, N. Iijima, and F. R. Frankel. 2008. Novel vaccination protocol with two live mucosal vectors elicits strong cell-mediated immunity
in the vagina and protects against vaginal virus challenge. J. Immunol. 180:
2504 –2513.
Ranasinghe, C., S. J. Turner, C. McArthur, D. B. Sutherland, J.-H. Kim,
P. C. Doherty, and I. A. Ramshaw. 2007. Mucosal HIV-1 pox virus prime-boost immunization induces high-avidity CD8⫹ T cells with regime-dependent cytokine/
granzyme B profiles. J. Immunol. 178: 2370 –2379.
Amorij, J. P., V. Saluja, A. H. Petersen, W. L. Hinrichs, A. Huckriede, and
H. W. Frijlink. 2007. Pulmonary delivery of an inulin-stabilized influenza subunit
vaccine prepared by spray-freeze drying induces systemic, mucosal humoral as well as
cell-mediated immune responses in BALB/c mice. Vaccine 25: 8707– 8717.
Simmons, C. P., T. Hussell, T. Sparer, G. Walzl, P. Openshaw, and G. Dougan.
2001. Mucosal delivery of a respiratory syncytial virus CTL peptide with enterotoxinbased adjuvants elicits protective, immunopathogenic, and immunoregulatory antiviral CD8⫹ T cell responses. J. Immunol. 166: 1106 –1113.
Belyakov, I. M., M. A. Derby, J. D. Ahlers, B. L. Kelsall, P. Earl, B. Moss, W. Strober,
and J. F. Berzofsky. 1998. Mucosal immunization with HIV-1 peptide vaccine induces mucosal and systemic cytotoxic T lymphocytes and protective immunity in mice
against intrarectal recombinant HIV-vaccinia challenge. Proc. Natl. Acad. Sci. USA 95:
1709 –1714.
Belyakov, I. M., J. D. Ahlers, B. Y. Brandwein, P. Earl, B. L. Kelsall, B. Moss,
W. Strober, and J. A. Berzofsky. 1998. The importance of local mucosal HIV-specific
CD8⫹ cytotoxic T lymphocytes for resistance to mucosal-viral transmission in mice
6891
6892
BRIEF REVIEWS: COMPARTMENTALIZED MUCOSAL IMMUNE RESPONSE AND PROTECTION
80. Belyakov, I. M., S. A. Hammond, J. D. Ahlers, G. M. Glenn, and J. A. Berzofsky.
2004. Transcutaneous immunization induces mucosal CTL and protective immunity
by migration of primed skin dendritic cells. J. Clin. Invest. 113: 998 –1007.
81. Ahlers, J. D., and I. M. Belyakov. 2009 Strategies for recruiting and targeting dendritic
cells for optimizing HIV vaccines. Trends Mol. Med. 15: 263–274.
82. Rimoldi, M., M. Chieppa, P. Larghi, M. Vulcano, P. Allavena, and P. Rescigno. 2005.
Monocyte-derived dendritic cells activated by bacteria or by bacteria-stimulated epithelial cells are functionally different. Blood 106: 2818 –2826.
83. Salazar-Gonzalez, R. M., J. H. Niess, D. J. Zammit, R. Ravindran, A. Srinivasan,
J. R. Maxwell, T. Stoklasek, R. Yadav, I. R. Williams, X. Gu, et al. 2006. CCR6mediated dendritic cell activation of pathogen-specific T cells in Peyer’s patches. Immunity. 24: 623– 632.
84. Urry, Z., E. Xystrakis, D. F. Richards, J. McDonald, Z. Sattar, D. J. Cousins,
C. J. Corrigan, E. Hickman, Z. Brown, and C. M. Hawrylowicz. 2009. Ligation of
TLR9 induced on human IL-10-secreting Tregs by 1␣,25-dihydroxyvitamin D3 abrogates regulatory function. J. Clin. Invest. 119: 387–398.
85. Sutmuller, R. P., M. H. den Brok, M. Kramer, E. J. Bennink, L. W. Toonen,
B. J. Kullberg, L. A. Joosten, S. Akira, M. G. Netea, and G. J. Adema. 2006. Toll-like
receptor 2 controls expansion and function of regulatory T cells. J. Clin. Invest. 116:
485– 494.
86. Peng, G., Z. Guo, Y. Kiniwa, K. S. Voo, W. Peng, T. Fu, D. Y. Wang, Y. Li,
H. Y. Wang, and R. F. Wang. 2005. Toll-like receptor 8-mediated reversal of CD4⫹
regulatory T cell function. Science 309: 1380 –1384.
87. Liu, G., S. Burns, G. Huang, K. Boyd, R. L. Proia, R. A. Flavell, and H. Chi. 2009.
The receptor S1P1 overrides regulatory T cell-mediated immune suppression through
Akt-mTOR. Nat. Immunol. 10: 769 –777.
88. Kim, T. S., and T. J. Braciale. 2009. Respiratory dendritic cell subsets differ in their
capacity to support the induction of virus-specific cytotoxic CD8⫹ T cell responses.
PLoS One 4: e4204.
89. Bedoui, S., P. G. Whitney, J. Waithman, L. Eidsmo, L. Wakim, I. Caminschi,
R. S. Allan, J. Wojtasiak, K. Shortman, F. R. Carbone, et al. 2009. Cross-presentation
of viral and self antigens by skin-derived CD103⫹ dendritic cells. Nat. Immunol. 10:
488 – 495.
90. Uematsu, S., J. Fujimoto, M. H. Jang, B. G. Yang, Y. J. Jung, M. Nishiyama, S. Sato,
T. Tsujimura, M. Yamamoto, Y. Yokota, et al. 2008. Regulation of humoral and cellular gut immunity by lamina propria dendritic cells expressing Toll-like receptor 5.
Nat. Immunol. 9: 769 –776.
91. McKinstry, K. K., et al. 2009. IL-10 deficiency unleashes an influenza-specific Th17
response and enhances survival against high-dose challenge. J. Immunol. 182:
7353–7363.
92. Kader, M., X. Wang, M. Piatak, J. Lifson, M. Roederer, R. Veazey, and
J. J. Mattapallil. 2009. ␣4⫹␤7hiCD4⫹ memory T cells harbor most Th-17 cells and
are preferentially infected during acute SIV infection. Mucosal Immunol. 2: 439 – 449.
93. Snyder, J. T., M. A. Alexander-Miller, J. A. Berzofsky, and I. M. Belyakov. 2003.
Molecular mechanisms and biological significance of CTL avidity. Current HIV Res. 1:
287–294.
Downloaded from http://www.jimmunol.org/ by guest on June 14, 2017