Substance P

18 Pages·2001·0.506 MB·English

by  Marriott I., Bost K.L.

Substance P
Ian Marriott and Kenneth L. Bost*
Department of Biology, University of North Carolina at Charlotte, 9201 University City Blvd, Charlotte,
NC 28223-0001, USA
*corresponding author tel: [phone], fax: [phone], e-mail: [email protected]
DOI: 10.1006/rwcy.2001.13005.
SUMMARY inflammation. Taken together, these studies demon-
stratearoleforsubstancePinnormalandpathologic
Substance P is a member of the tachykinin family of immune responses.
peptides,andhasclassicallybeenconsideredaneuro-
peptide, secreted predominantly by neuronal cells of
BACKGROUND
thecentralandperipheralnervoussystems.Interaction
with tachykinin receptors on vascular endothelial or
Discovery
smoothmusclecellscancausevasodilationorcontrac-
tion, respectively. Furthermore, the presence of this
peptide in the nervous system can have a variety of The amino acid sequence of substance P was
effectsonbehaviorandthetransmissionofpain.These published (Chang et al., 1971) prior to the
effects of substance P have been reviewed (Pernow, determination of genes encoding the tachykinins
1983; Maggi, 1996; Holzer and Holzer-Petsche, 1997; (Nawaetal.,1983;Harmaretal.,1986;Krauseetal.,
Quartara and Maggi, 1998). 1987). The heteronuclear RNA encoding substance P
However, the following review will focus on obser- has been termed preprotachykinin A. This RNA
vations which suggest that substance P can also have species encodes several tachykinins, including sub-
an important influence on the immune response. The stanceP,neurokininA(substanceK),andneurokinin
mRNA encoding substance P can be found in some B.AtleastthreedifferentRNAspecies(designated(cid:11),
activated leukocytes, together with the secretion of (cid:12), or (cid:13) preprotachykinin A) can be processed and
thispeptide,suggestingthatthesecellsareapotential expressed from the single heteronuclear preprotachy-
non-neuronalsourceforsubstanceP(Weinstocketal., kinin A RNA. Expression of the (cid:12) preprotachykinin
1988; Pascual and Bost, 1990; Weinstock and Blum, mRNA species seems to be predominantly expressed
1990; Bost et al., 1992; Ho et al., 1997). While sub- in most tissues.
stance P may be released from leukocytes during cer-
tain immune responses, it is clear that peptidergic Alternative names
neurons innervate, and are intimately associated with
leukocytes, within primary and secondary lymphoid
Substance P is part of a family of tachykinins or
organs (Lorton et al., 1990; Lorton et al., 1991;
neurokinins, which includes substance P, neurokinin
Tabarowskietal.,1996;Gotoetal.,1998;Jurjusetal.,
A (substance K), and neurokinin B. Therefore
1998), within the skin (Ansel et al., 1996), and within
sometimes substance P is referred to in more general
the gut (Holzer and Holzer-Petsche, 1997). Further-
terms as a tachykinin, or as a neurokinin.
more, it is clear that leukocytes express specific,
functional receptors for substance P (Payan, 1989;
Structure
Bost and Pascual, 1992; Rameshwar and Gascon,
1997; Weinstock and Elliott, 1998). Antagonism of
substance P interacting with its receptor has been Substance P is an 11 amino acid peptide (Arg-Pro-
showntolimitthehostimmuneresponseandtolimit Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH ) which
2
CytokineReference Copyright#2001AcademicPress
2 Ian Marriott and Kenneth L. Bost
hasanamidatedC-terminus.Itbelongstoafamilyof Interestingly, substance P has been shown to limit
peptides called tachykinins, which also includes production of induced TGF(cid:12) secretion by macro-
neurokinin A (substance K) and neurokinin B. All phages (Marriott and Bost, 1998).
known vertebrate tachykinins share a common C-
terminal sequence motif, -Phe-XXX-Gly-Leu-Met,
with XXX representing the presence of a variable, GENE AND GENE REGULATION
hydrophobic amino acid.
Substance P has been described almost exclusively as
being a peptide of neuronal origin (Maggi, 1996;
Main activities and Nakanishi, 1987). In neurons, the peptide can be
transcribed from at least three distinctly different
pathophysiological roles
mRNAs (designated (cid:11), (cid:12), and (cid:13) preprotachykinin A)
which result from the differential processing of RNA
Substance P is a member of the tachykinin family of
transcribed from a single gene (Carter and Krause,
peptides,andhasclassicallybeenconsideredaneuro-
1990; Krause et al., 1987; Nakanishi, 1987; Nawa
peptide, secreted predominantly by neuronal cells of
et al., 1983).The geneencoding substance Phasbeen
thecentralandperipheralnervoussystems.Interaction
termed preprotachykinin A to indicate the possibility
with tachykinin receptors on vascular endothelial, or
that several tachykinin peptides (i.e. substance P,
smooth muscle cells can cause vasodilation or con-
neurokinin A, neuropeptide K, and neuropeptide (cid:13))
traction, respectively. Furthermore, the presence of
canbetranslatedfromthesamemessage.Translation
this peptide in the nervous system can have a variety
ofpreprotachykininmRNAbyneuronsisresponsible
of effects on behavior and the transmission of pain.
forthe presenceof substance Pin the central nervous
These effects of substance P have been reviewed
system as well as in peripheral tissues.
(Pernow, 1983; Maggi, 1996; Holzer and Holzer-
Petsche,1997;QuartaraandMaggi,1998).
However, this review will focus on the activities Accession numbers
which have been ascribed for substance P within the
immune response. Substance P can augment B lym-
The bovine preprotachykinin genes encoding sub-
phocyte responses, in particular the production of
stance P were the first cloned (Nawa et al., 1983),
antibody (Pascual et al., 1991a; Bost and Pascual,
accession number X00075, whereas the preprotachy-
1992; Braun et al., 1999). Substance P can also aug-
kinin genes encoding rat (Krause et al., 1987),
ment immune responses to pathogens which depend
accession numbers M15191 and AH002233, and
upon cell-mediated immunity (Croitoru et al., 1990;
human (Harmar et al., 1986), accession number
Kincy-Cain and Bost, 1996). Thus, it appears that
X54469, were subsequently cloned.
substance P has a dual role in augmenting both
humoral and cell-mediated immune responses.
SubstancePinteractingwithitsreceptorhasalsobeen Sequence
suggestedtocontributetotheinflammationasso-ciated
with infectious diseases or microbial products
See Figure 1.
(Kataeva et al., 1994; Mantyh et al., 1996a, 1996b;
Castagliuoloetal.,1998;WeinstockandElliott,1998;
Blum et al., 1999; Tripp et al., 2000), inflammation Chromosome location
associated with neurogenic input (Kataeva et al.,
1994; Bozic et al., 1996; Mantyh et al., 1996a, 1996b;
Mouse chromosome 6; human chromosome 2.
Castagliuoloetal.,1998;WeinstockandElliott,1998;
Blum et al., 1999; Saban et al., 2000; Tripp et al.,
2000),andinflammationassociatedwithautoimmune
PROTEIN
diseases(Levineetal.,1984;Lotzetal.,1987;Mantyh
et al., 1995). Substance P has also been documented
Accession numbers
toincreaseproductionofproinflammatorycytokines,
prostanoids, and reactive oxygen and nitrogen
intermediates by macrophages (Hartung and Toyka, Human (cid:12) preprotachykinin A-coding region:
1983; Hartung et al., 1986; Lotz et al., 1988; Murris- CAA38351
Espin et al., 1995; Zhu et al., 1996; Ho et al., 1996, Mouse(cid:12)preprotachykininA-codingregion:I52526
1998; Kincy-Cain and Bost, 1997; Jeon et al., 1999). Rat(cid:12)preprotachykininA-codingregion:AAA41928
Substance P 3
Figure 1 Nucleotide sequences for substance P.
4 Ian Marriott and Kenneth L. Bost
Figure 2 Amino acid sequences for human, mouse, and andthepresenceofpreprotachykininmRNAinthese
rat substance P. cells has been described (Weinstock et al., 1988;
Weinstock and Blum, 1990; De Giorgio et al., 1998).
ThedenovosynthesisandsecretionofsubstancePby
macrophageshasalsobeenreported(Bostetal.,1992;
De Giorgio et al., 1998; Ho et al., 1997; Pascual and
Bost, 1990). A recent study suggests that dendritic
cells can also express preprotachykinin mRNA and
secrete substance P (Lambrecht et al., 1999). Taken
together, these studies suggest an extraneuronal
source for substance P.
Since neurons containing substance P innervate a
variety of tissues, and since a variety of cells express
receptors for this peptide (Pernow, 1983; Maggi,
1996;HolzerandHolzer-Petsche,1997;Quartaraand
Maggi, 1998), it is not difficult to explain the many
Sequence
and varied effects of substance P. Especially relevant
for this chapter is the observation that substance P-
See Figure 2. containingneuronsareprevalentinthegutandinthe
skin (Ansel et al., 1996; Holzer and Holzer-Petsche,
1997; Maggi, 1996). The diversity of effects on
Important homologies
peripheral tissues, coupled with secretion by neurons,
suggests that tachykinins and their receptors repre-
The tachykinins or neurokinins, including substance sent an important mechanism by which the nervous
P,havesimilarC-terminalsequences.Specifically,the system can maintain homeostasis by affecting
C-terminal motif -Phe-XXX-Gly-Leu-Met is con- responses in the periphery.
served between all vertebrate tachykinins. XXX Table 1 summarizes the cellular sources of
representsavariablehydrophobicaminoacidresidue. substance P with a focus on leukocytes that have
been shown to produce this peptide.
Posttranslational modifications
Eliciting and inhibitory stimuli,
The preprotachykinin mRNA is translated into a including exogenous and
polyproteinwhichencodessubstanceP,neurokininA
endogenous modulators
(substance K), and neurokinin B. Posttranslational
processing cleaves these tachykinins from the poly-
protein. The C-terminal end of substance P is then For neuronal cells, excitatory or noxious stimuli can
amidated. induce secretion of substance P from nerve terminals
(Pernow,1983;Anseletal.,1996;Maggi,1996;Holzer
andHolzer-Petsche,1997).Forleukocytes,infectionor
CELLULAR SOURCES AND exposuretomicrobial productsis requiredforactiva-
tion and subsequent secretion of substance P (Levine
TISSUE EXPRESSION
etal.,1984;Weinstocketal.,1988;WeinstockandBlum,
1989,1990;Bostetal.,1992;Kataevaetal.,1994;Bost,
Cellular sources that produce
1995a; Kincy-Cain and Bost, 1996; Mantyh et al.,
1996b; Zhu et al., 1996; Castagliuolo et al., 1997;
Substance P can be secreted by neuronal cells within Garlandetal.,1997;Hoetal.,1997;MarriottandBost,
the central or peripheral nervous systems (Nakanishi, 1998;Sabanetal.,2000).
1987; Maggi, 1996). Substance P-containing neurons Elevated levels of substance P have been noted in
are especially prevalent in the gut and in the skin. areas of inflammation, though it is not altogether
Afferent sensory neurons contain high levels of this clearwhatparticularstimulusisrequiredtoelicitsuch
neuropeptide. While substance P present in tissues secretion.
can be neuronally derived, these cells may not be the Table 1 summarizes the stimuli that have been
onlyonescapableofsecretingthispeptide.Theability shown to induce release of substance P with a focus
of neutrophils to secrete immunoreactive substance P, on leukocyte-derived peptides.
Substance P 5
Capsaicin has been shown to induce release of andgranulocytes.Suchstudieshavedemonstratedthe
substance P from nerve terminals (Buck and Burks, presence of NK1 receptors on the surface of B and T
1986)aswellasfromhumanmonocytes(Hoetal.,1997). cells isolated from spleen and Peyer’s patches. The
binding characteristics of the substance P receptors
on leukocytes are similar to those for NK1 receptors
present on neural tissues. This conclusion is based on
RECEPTOR UTILIZATION
similar dissociation constants, and similar rank-order
displacementbyrelatedtachykinins.Insummary,itis
Three distinct neurokinin receptors (termed NK1,
clear that some leukocytes can express authentic
NK2, and NK3 receptors) have been cloned. These
substance P receptors.
receptors are members of the superfamily of G
protein-coupled receptors characterized by a seven
Signal Transduction
transmembrane domain motif. Effects of tachykinins
on peripheral tissues and cells are receptor mediated. The signaling mechanisms utilized following sub-
ThereceptorforsubstancePhasbeencloned(Hershey stance P/NK1 receptor interaction remain highly
and Krause, 1990; Yokota et al., 1989) and has also equivocal in cells of the immune sytem. A role for
been termed the neurokinin 1 (NK1) receptor. While elevations in intracellular calcium in the initiation of
each tachykinin can interact with each of the three immune responses by substance P has been widely
receptors, there is preferential binding of substance P reported in a variety of cell types including
tosubstanceP(NK1)receptors,neurokininAtoNK2 neutrophils (Serra et al., 1988; Tanabe et al., 1996),
receptors, and neurokinin B to NK3 receptors. Thus, human monocytes (Kavelaars et al., 1994), and
eachtachykininhasits‘own’receptorforwhichithas peripheral blood polymorphonuclear cells (PBMCs)
preferentialaffinity. (Kavelaars et al., 1993; Nowak et al., 1996). In
Recent observations have suggested that substance addition, elevations in cytosolic free calcium have
P and its receptor may play a significant role in the been observed in cell lines transfected with substance
modulation of immune responses. Specific receptors P receptors following exogenous treatment with
for this peptide have been detected on a variety of substance P (Christian et al., 1994; Seabrook and
immunocytes including lymphocytes, macrophages, Fong 1993; Sudduth-Klinger et al., 1992).Ithasbeen
neutrophils, and mast cells (Payan, 1989; Bost and suggestedfromsuchstudiesthatsubstancePinteracts
Pascual, 1992; Pascual et al., 1994, 1999; Rameshwar with NK1 receptors to promote G protein-linked IP
3
and Gascon, 1997; Weinstock and Elliott, 1998). In formation that subsequently mobilizes intracellular
manycases,substancePreceptorsonleukocyteshave stores and initiates capacitative, or store-mediated,
been shown to be identical to their neuronal counter- calcium influx across the plasma membrane.
parts at the mRNA and protein levels. Radiolabeled Incontrasttothesestudies,severalreportsindicate
ligand-binding studies have also been used to thatsubstancePfailstoelicitsignificantalterationsin
demonstratethe presenceofsubstancePreceptorson cytosolic free calcium in leukocytes (Haines et al.,
leukocytes, in particular, lymphocytes, macrophages, 1993), including a rat alveolar macrophage cell line
Table 1 Cellular sources and stimuli that induce release of substance P
Cell source Release stimulus Reference
Neuronal cells Excitatory or noxious stimuli Reviewed in Maggi, 1996
Capsaicin Buck and Burks, 1986
Major basic protein from Garland et al., 1997
activated eosinophils
Monocytes Capsaicin Ho et al., 1997
Macrophages LPS Bost, 1995a
C. difficile toxin A Castagliuolo et al., 1997
Capsaicin Ho et al., 1997
Eosinophils Calcium ionophores Weinstock and Blum, 1990
Histamine Weinstock and Blum, 1990
6 Ian Marriott and Kenneth L. Bost
(Zhangetal.,1997).Inaddition,inatleastonestudy, within the nervous system. Recent studies are con-
low concentrations of substance P (1–100nM) were sistent with this observation and demonstrate that
found to prime neutrophil responses via calcium substancePisapotentinducerofNF(cid:20)Bactivationin
independent mechanisms (Lloyds and Hallett, 1993). bothmurinemacrophagesanddendriticcells(Marriott
Suchfindingsareinagreementwithrecentstudiesthat and Bost, 2000). Targets of NF(cid:20)B include genes
showthatsubstancePhadnodemonstrableeffectupon encoding cytokines and their receptors (Baeuerle and
intracellularcalciumlevelsintwotypesofprofessional Henkel,1994).Importantly,NF(cid:20)Bactivationhasbeen
antigen-presentingcells,namely,murinemacrophages demonstratedtoplayakeyroleintheregulationofthe
and dendritic cells (Marriott and Bost, 2000). The proinflammatory cytokines TNF(cid:11) (Han and Beutler
apparent discrepancy between these results and those 1990; Shakhov et al., 1990; Wrighton et al., 1996;
studiesdemonstratingachangeinintracellularcalcium Carter et al., 1998; Bondeson et al., 1999), IL-1(cid:12)
followingsubstancePadditionmaybedue,inpart,to (Hiscott et al., 1993; Cogswell et al., 1994; Bondeson
the concentrations of substance P used. In several et al., 1999), IL-6 (Libermann and Baltimore, 1990;
studies in which substance P evoked an elevation in Carter et al., 1998; Bondeson et al., 1999), and IL-
intracellular calcium, the doses of substance P used 12p40(Murphyetal.,1995;Yoshimotoetal.,1997)by
wereinthemicromolarrange.Suchconcentrationsare a variety of cell types including macrophages. The
far in excess of those that evoke optimal immune potential importance of the involvement of NF(cid:20)B in
responsesinleukocyteswhichsignalthroughaclassical the regulation of these genes is underscored by
substance P receptor. Importantly, in at least one observations in situ that NF(cid:20)B is activated in macro-
study championing a role for calcium in substance P phages at sites of inflammation (Gveric et al., 1998;
signaling, substance P receptor antagonists failed to Rogleretal.,1998).Giventhepreviouslydocumented
attenuate the effect of substance P on intracellular roleforthistranscriptionalregulatorinthecontrolof
calcium (Kavelaars et al., 1993), suggesting the proinflammatory cytokine production by macro-
presenceofanonclassicalsubstancePreceptor.Addi- phages, the ability of substance P to activate NF(cid:20)B
tionally,inthesamestudy,anonfunctionalanalogof suggests an important mechanism by which this
substance P was also found to provoke increases in neuropeptidemayenhanceproinflammatorycytokine
calcium (Kavelaars et al., 1993). Such findings, in productionbyprofessionalantigen-presentingcells.
combination with the previously documented ability
of amphiphilic peptides such as substance P to insert
themselves into theplasma membraneand tointeract
INVITRO ACTIVITIES
directlywithintracellularproteins(RepkeandBienert,
1988; Rollandy et al., 1991), might be indicative of
substancePelicitingariseincytosolicfreecalciumvia The expression of specific substance P receptors on a
indirectmechanismsorviaanonclassicalsubstanceP widerangeofleukocyticcelltypes(Payan,1989;Bost
receptor. and Pascual, 1992; Pascual et al., 1994, 1999;
The disparate results concerning the signaling RameshwarandGascon,1997;WeinstockandElliott,
pathways utilized by substance P in cells transfected 1998) and the presence of peptidergic nerve fibers in
withsubstancePreceptorsmightarisefromtheability close proximity with immune cells suggests that the
of tachykinin receptors to interact with multiple G function of these cells might be modified by this
protein types. As a result, the signaling observed neuropeptide.Indeed,alargebodyofinvitroevidence
following stimulation of these receptors would has accumulated to support such a conjecture. The
probably be dependent on the nature of the host cell following section is devoted to descriptions of those
and its repertoire of signaling components. For in vitro studies that show definitive immune modula-
example, low concentrations that initiate neutrophil tion on B cells, T cells, and macrophages, and the
priming do not evoke elevations in intracellular effectsofsubstancePonimmunecellprogenitors.
calciumconcentration(LloydsandHallett,1993),but Table 2 summarizes the activities attributed to
havebeenshowntopromotetyrosinephosphorylation substance P upon interacting with leukocytes.
(Lloydsetal.,1995).
A recent study demonstrated the ability of nano-
In vitro findings
molar concentrations of substance P to activate the
transcriptionalactivatorNF(cid:20)Binanastrocytomacell
Substance P Involvement in Hematopoiesis
line resulting in the enhanced expression of IL-8, an
NF(cid:20)B-regulated gene (Lieb et al., 1997). This obser- Substance P has been demonstrated to play a role in
vation has led to the suggestion that NF(cid:20)B might be hematopoiesis using short-term bone marrow cul-
an important component controlling inflammation tures.SubstancePactsviaNK1receptorstoenhance
Substance P 7
Table 2 Substance P-induced activities reported for leukocytes
Cell type Activity References
Bone marrow stem cell Proliferation Rameshwar and Gascon, 1997
IL-1 and GM-CSF production Rameshwar et al., 1993, 1994
B lymphocytes Ig production from stimulated cells Stanisz et al., 1986; Pascual et al., 1991a, 1991b
Optimal Ig responses to antigenic challenge, Helme et al., 1987; Eglezos et al., 1990;
and adjuvant qualities in vivo Ijaz et al., 1990
Promotes isotype switching Stanisz et al., 1986; Braun et al., 1999
T lymphocytes T cell proliferation Payan et al., 1983; Stanisz et al., 1986;
Covas et al., 1995
Lymphokine production from stimulated
cells:
IL-2 Calvo et al., 1992; Nio et al., 1993;
Rameshwar et al., 1993
IL-4 Levite, 1998
IL-10 Kawamura et al., 1998
Macrophages Respiratory burst Hartung and Toyka, 1983
Hartung et al., 1986
Monokine production:
IL-1 Laurenzi et al., 1990
IL-6 Lotz et al., 1988
IL-12 Kincy-Cain and Bost, 1997
TNF(cid:11) Ho et al., 1996; 1998
Increased IL-6 production in vivo Zhu et al., 1996
Deceased TGF(cid:12)1 production from Marriott and Bost, 1998
stimulated macrophages
Prostanoid production Hartung and Toyka, 1983; Murris-Espin et al.,
1995
NO production Jeon et al., 1999
Increased phagocytic ability Bar-Shavit et al., 1980
Increased resistance to Salmonella infection Kincy-Cain and Bost, 1996
Neutrophils Activation and priming of oxidase response Serra et al., 1988
the proliferation of primative bone marrow stem cells Substance P as a B-cell Differentiation Factor
and progenitors (Rameshwar and Gascon, 1997) and
It has been shown that substance P can significantly
these effects correlate with the induction of produc-
influence both the isotype and the levels of Ig
tionofstimulatoryhematopoieticgrowthfactorssuch
production by B lymphocytes. Substance P-treated
as IL-1 and GM-CSF by bone marrow mononuclear
mononuclear cell fractions from murine spleen,
cells (Rameshwar et al., 1993, 1994). This production
mesentericlymphnodes,andPeyer’spatches,resulted
requires de novo synthesis and has been shown to be
in70%,40%,and300%increases,respectively,inIgA
inhibited by specific NK1antagonists (Rameshwaret
production (Stanisz et al., 1986). To a lesser extent,
al., 1994). Furthermore, cytokines that promote
IgMlevelsweresignificantlyalteredby20–40%,while
hematopoiesis upregulate the expression of NK1
IgG levels were unaltered. Such results suggest that
receptors in bone marrow stroma (Rameshwar et al.,
substance P may preferentially stimulate IgA secre-
1997). Taken together, these results suggest that
tion. Alternatively, these results may indicate that
cytokines and neuropeptides such as substance P can
substance P acts as an IgA switching factor.
interact to regulate hematopoiesis.
8 Ian Marriott and Kenneth L. Bost
Work with the CD5+ B lymphoma cell lines, Neuropeptides as a T cell Costimulation Factor
CH12.LX.C4.4F10 (an IgA producer) and
EarlystudieshaveshownthatsubstancePsupportsT
CH12.LX.C4.5F5 (an IgM producer), supports the
cell proliferation (Payan et al., 1983; Stanisz et al.,
contention that substance P behaves as a late-acting
1986; Covas et al., 1995). However, substance P was
Bcelldifferentiationfactor(Pascualetal.,1991b,1992;
unable to show increases in [3H]thymidine incorpora-
Bost and Pascual, 1992). These cell lines express
tion in costimulated, concanavalin A-treated lamina
between500and600NK1receptorspercell,levelsand
proprialymphocytes(64%CD3+and11%CD19+)
bindingaffinities(K (cid:24)0.69nM)thatarecompar-
d derivedfromhumancolonspecimens.Infact,adose-
abletonormalBlymphocytes(Staniszetal.,1987)and
dependentreductionincellproliferationwasobtained
IM-9 B lymphoblasts (Payan et al., 1984). Following
upon substance P addition (Elitsur and Luk, 1990).
direct substance P stimulation of CH12.LX.C4.4F10
Thus, the type of lymphocyte or the source of the
cells, IgA production is modestly increased, whereas
lymphocytemaydictateitssusceptibilitytosubstance
similarstimulationofCH12.LX.C4.5F5cellsresulted
P stimulation of proliferation.
innosignificantalterationsinIgMproduction.Inthese
Substance P has also been reported to induce
studies,substancePhadnoeffectoncellproliferation
lymphokine production. Studies have demonstrated
as measured by [3H]thymidine uptake. However, the
that substance P can augment production of the
addition of LPS in conjunction with subnanomolar
cytokineIL-2inhumanandmurineTcelllinesactivated
concentrations of substance P to CH12.LX.C4.5F5
with phorbol esters (Calvo et al., 1992) or a specific
cells resulted in optimal IgM production (172%
antigen (Nio et al., 1993). Such findings have been
increase). Modest increases in IgA production were
supported by studies in normal murine lymphocytes
seen in CH12.LX.C4.4F10 cells that were similarly
and purified CD4+ T cells costimulated with IL-1,
treated. In both cases, these effects were found to be
demonstratingthatsubstancePaugmentsIL-2produc-
mediatedbyNK1receptorsasevidencedbytheability
tion(Rameshwaretal.,1993).Inaddition,morerecent
of excess substance P antagonists to inhibit these
studies have shown that substance P can induce the
responses.
production of TH2-associated cytokines IL-4
Similar studies have been performed with purified
(Kawamura et al., 1998; Levite, 1998) and IL-10
Bcells isolatedfromthe spleen(Pascualetal.,1991a)
(Kawamuraetal.,1998),leadingtothesuggestionthat
and Peyer’s patches (Pascual et al., 1995). B
substancePcaninfluencethecommitmentofimmune
lymphocytes (> 99% sIg+) isolated from mouse
responses to a distinct TH phenotype and ultimately
spleensorPeyer’spatchesshowednodifferencesinIg
immunefunction.
production when cultured with varying concentra-
As describedabove, substance Phas been found to
tions of substance P. However, the addition of 10(cid:22)g/
augment Ig production by Con A-treated splenic and
mL of LPS to these cultures resulted in optimal Ig
Peyer’s patchmononuclearcells (Staniszet al., 1986).
production. Subnanomolar concentrations of sub-
Ithasbeensuggested(Pascual,1999)that,ratherthan
stance P in conjunction with LPS elicited increases in
exertingadirecteffectonBcells,thecostimulationof
IgM and IgG3 production of as much as 500% and
TcellsbysubstancePmayinducetheproductionofT
1000%, respectively (Pascual et al., 1991a). Peyer’s
cell-derived B cell-differentiating cytokines. Such an
patch Ig production was also enhanced when IL-6
hypothesis is supported by the observation that
was used as a costimulator (Pascual et al., 1995).
substance P-containing nerve fibers impinge on the T
Again, subnanomolar concentrations of substance P
cell zone but not the B cell follicles (Felten et al.,
induced production of IgA and IgG in Peyer’s patch
1987)suggestingthatsubstancePmayexertitseffects
cultures. The significance of these studies is 2-fold.
on Ig production via T cells.
First, substance P can stimulate B cells directly in the
absence of accessory cells, and secondly, physiologi-
cally relevant concentrations of substance P can
Substance P-mediated Modulation of Macrophage
enhance Ig synthesis by purified B cells. Such
Function
observations are in agreement with the previously
reported abilityof substance Pto modify Ig synthesis While numerous investigations have focused on the
(Staniszetal.,1986)intheabsenceofenhancedBcell ability of neuropeptides to significantly modulate
proliferation. Indeed, at the doses optimal for Ig antigen-specific T and B lymphocyte responses, a
production, substance P in combination with LPS growing body of work suggests that the innate
was found to be antiproliferative (Pascual et al., immune response can also be influenced by neuro-
1991a). Collectively, these in vitro studies suggest peptides, particularly macrophages.
substancePrequiresacoactivationsignaltoaugment The rationale for hypothesizing that substance
Ig production. P might play an important role in macrophage
Substance P 9
activation and the destruction of intracellular patho- macrophages (Bost, 1995a), and toxin A from
gens suchasSalmonella is basedon in vitro studiesof Clostridium difficile has been shown to increase sub-
the effects of substance P on macrophages (Hartung stancePmRNAmessageexpressionbylaminapropria
and Toyka, 1983; Hartung et al., 1986; Lotz et al., macrophages (Castagliuolo et al., 1997). In addition,
1988;Kincy-CainandBost,1996,1997)andfromthe capsaicinhasbeenshowntoreleasesubstancePfrom
demonstration of inducible NK1 receptor expression humanmonocytesandmacrophages(Hoetal.,1997).
by these cells (Marriott and Bost, 2000). Signaling Calciumionophoresorhistaminehavebeenshownto
through the substance P receptor can induce a elicit the release of substance P from eosinophils in
respiratory burst in macrophages, resulting in the granuloma elicited by murine Schistosoma mansoni
productionofreactiveoxygenintermediates(Hartung (Weinstock and Blum, 1990).
andToyka,1983;Hartungetal.,1986),andisknown
to be a potent stimulus for the production of
Bioassays used
proinflammatory molecules by these cells, such as
IL-1, IL-6, TNF(cid:11), and IL-12 (Lotz et al., 1988;
Laurenzi et al., 1990; Kincy-Cain and Bost, 1997; Ho There are currently no reliable bioassays for
et al., 1998). substance P in terms of altered immune function.
Furthermore, cultured macrophages exposed to Current bioassays for substance P are based upon its
Salmonella were found to rapidly upregulate expres- actions upon smooth muscle. By far, the most
sion of NK1 receptors (Kincy-Cain and Bost, 1996). common bioassay for substance P is the guinea pig
This finding has added significance given the ability ileumpreparationinwhichsubstancePelicitsspecific
of substance P to augment the inflammatory and dose-dependent contractions (Leban et al., 1979;
responses. Of particular interest is the finding that Salt et al., 1982; Chassaing et al., 1992).
substancePcansynergizewithLPSintheproduction Other bioassays for substance P include the vaso-
of bioactive IL-12 p70 (Kincy-Cain and Bost, 1997). relaxations induced in dog common carotid artery
Since the presence of IL-12 in vivo is an important preparation(Coutureetal.,1980;Snideretal.,1991),
step in resistance against salmonellosis (Kincy-Cain and the rabbit anterior mesenteric vein preparation
and Bost, 1996), substance P-induced production of (Berube et al., 1978).
this monokine would have significant impact on the
cell-mediatedimmuneresponseagainstthispathogen.
Inaddition,anovelmechanismbywhichsubstanceP IN VIVO BIOLOGICAL
might augment inflammatory responses has been
ACTIVITIES OF LIGANDS IN
suggested. Substance P was able to dramatically
ANIMAL MODELS
diminish the production of the immunosuppressive
cytokine TGF(cid:12)1 from macrophages stimulated with
LPS or IFN(cid:13) (Marriott and Bost, 1998). Normal physiological roles
Collectively, these in vitro results suggest mechan-
ismsbywhichsubstancePmayaugmentmacrophage The potential in vivo significance of substance P as a
function as well as mechanisms by which this neuroimmune modulator has largely been implied
neuropeptide may enhance cell-mediated immune from in vitro studies as described above. In contrast,
responses to intracellular pathogens such as in vivo studies on the importance of substance P-
Salmonella via production of monokines. mediated modulation of immune function are far less
numerous.Thefollowingsectionisdevotedtodescrip-
tions of those in vivo studies that show definitive
Regulatory molecules: Inhibitors immune modulation of B cells and macrophages.
Table 2 summarizes the in vivo affects that have
and enhancers
been attributed to the presence of substance P.
Activatedeosinophilsreleasemajorbasicproteinwhich
Substance P as a B Cell Differentiation Factor
maybethemediatorbywhichactivatedeosinophilscan
elicitrelease of substancePfrom cultured dorsalroot TheinvivorelevanceofsubstancePonantibodypro-
ganglionneurons(Garlandetal.,1997). ductionwasshownbyHelmeandcoworkers(1987).In
In in vitro studies of leukocytes, exposure to this study, peripheral stores of substance P were
microbial products have been demonstrated to cause depletedbytreatingneonatalratswiththeneurotoxin
thesecretion ofsubstancePfromthesecells. LPShas capsaicin, which destroys unmyelinated sensory neu-
been shown to elicit substance P release by rat rons present in peripheral tissues (Buck and Burks,
10 Ian Marriott and Kenneth L. Bost
1986). These neonatally capsaicin-treated rats were InadditiontotheabilityofsubstancePtomodulate
allowed to mature prior to antigenic challenge with immunecellfunctionasdescribedabove,classicalroles
sheep red blood cells. Capsaicin treatment resulted in of substance P include its role as a neurotransmitter.
a greater than 80% reduction in both IgM and IgG This molecule is released predominantly by neuronal
plaque-forming cell responses by popliteal lymph cellsofthecentralandperipheralnervoussystems.Itis
nodes compared with untreated sheep red blood cell- knowntoelicitvasodilationofbloodvesselsfollowing
challengedanimals.Importantly,thissuppressionwas interaction with NK1 receptors on vascular endothe-
found to be reversible following coadministration of lium or vasoconstriction of smooth muscle cells. In
exogenous substance P with sheep red blood cells. addition, the presence of substance P in the nervous
In a subsequent study (Eglezos et al., 1990), a sim- system mediates a variety of behavioral effects and
ilar magnitude of inhibition was observed in animals plays a key role in the transmission of pain signals.
treatedwiththesubstancePantagonistSpantideduring These effects are reviewed elsewhere (Holzer and
antigenpriming.Thesestudiesdemonstratethatendo- Holzer-Petsche, 1997; Maggi, 1996; Pernow, 1983;
genoussubstancePmayplayanimportantroleinthe QuartaraandMaggi,1998).
generation of optimal Ig responses. The ability of
substance P to augment Ig responses has been under-
scoredbythefindingthatinvivoinfusionofsubstance Species differences
P during immunization can act as an adjuvant.
Mice infused with substance P for 1 week via micro-
Significantspeciesdifferenceshavebeendemonstrated
osmotic pumps and simultaneously immunized with
betweentheoccurrenceanddistributionofsubstanceP-
UV-inactivated rotavirus demonstrated increased
containing nerve fibers both in the central nervous
levels of anti-rotavirus antibody in both serum and
system and in the periphery, and these findings are
in the milk of lactating females (Ijaz et al., 1990).
discussed elsewhere (Baker, 1986; Okado et al., 1991;
In summary, these studies demonstrate that the
Sundler et al., 1991; Smeets, 1992; Borhegyi and
in vivo addition or depletion of substance P can
Leranth,1997).Inaddition,markedspeciesdifferences
augment the development of Ig-producing cells.
have been reported in the actions of substance P on
urinarybladderfunction(Maggietal.,1987),pancreas
Substance P-Mediated Modulation of (Chiba et al., 1985), and eye (Tachado et al., 1991).
Macrophage Function Importantly, species differences in severity of neuro-
genicinflammatoryresponsesinairwaysexistbetween
Invivostudiesthathavefocusedontheimportanceof
rodentandhumanspecies(Lundberg,1995).
substance P-mediated modulation of macrophage
function are few in number. However, recent studies
have demonstrated an importantrole for substance P
Knockout mouse phenotypes
in mucosal immune responses to the intracellular
pathogen Salmonella. Surprisingly, oral inoculation
Arecentstudyhasreportedthedisruptionofthemouse
with Salmonella initiates rapid and dramatic upregu-
preprotachykinin A gene (PPT-A) that encodes
lation of the mRNAs encoding substance P (Kincy-
substance P and a related tachykinin, neurokinin A
CainandBost,1996)anditsreceptor(Bost,1995b)in
(Caoetal.,1998).Inthisstudy,itisreportedthatwhile
mucosal tissues (Kincy-Cain and Bost, 1996). This
the behavioral response to mildly painful stimuli is
resultsuggestedthatsubstancePanditsreceptorwere
intactintheseknockoutmice,theresponsetomoderate
involved in the initiation of the response to
to intense pain is significantly reduced (Cao et al.,
Salmonella. To address this possibility directly, mice
1998). In addition, it has been noted that neurogenic
were pretreated with the potent substance P
inflammation is almost completely absence in these
antagonist Spantide II prior to oral inoculation with
animals (Cao et al., 1998). Furthermore, mice with a
Salmonella. Mice pretreated with the substance P
disruptionofthePPT-Agenehavebeendemonstrated
antagonist could not resist Salmonella infection as
to be resistant to seizures induced by kainate or
wellascontrolmicetreatedwithanirrelevantpeptide
pentylenetetrazole(Liuetal.,1999).
(Kincy-Cain and Bost, 1996). Treatment with the
antagonist caused no apparent alterations in gut
function aside from a reduction in immune respon-
Transgenic overexpression
siveness. Therefore, in vivo antagonism of substance
P/NK1 interactions resulted in surprising and
dramatic reductions in the resistance against the Mice overexpressing substance P have not been
intracellular pathogen Salmonella. reported.