By
From the Department of Molecular Genetics, Hellenic Pasteur Institute, 115 21 Athens, Greece
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Abstract |
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Lymphotoxin (LT) knockout mice, as well as double LT
/tumor necrosis factor (TNF)
knockout mice, show a severe splenic disorganization with nonsegregating T/B cell zones and
complete absence of primary B cell follicles, follicular dendritic cell (FDC) networks, and germinal centers. In contrast, as shown previously and confirmed in this study, LT
-deficient
mice show much more conserved T/B cell areas and a reduced but preserved capacity to form
germinal centers and FDC networks. We show here that similar to the splenic phenotype of
LT
-deficient mice, complementation of LT
knockout mice with TNF-expressing transgenes leads to a p55 TNF receptor-dependent restoration of B/T cell zone segregation and a
partial preservation of primary B cell follicles, FDC networks, and germinal centers. Notably,
upon lipopolysaccharide challenge, LT
knockout mice fail to produce physiological levels of
TNF both in peritoneal macrophage supernatants and in their serum, indicating a coinciding deficiency in TNF expression. These findings suggest that defective TNF expression contributes to the complex phenotype of the LT
knockout mice, and uncover a predominant role
for TNF and its p55 TNF receptor in supporting, even in the absence of LT
, the development and maintenance of splenic B cell follicles, FDC networks, and germinal centers.
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Introduction |
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Tumor necrosis factor (TNF), lymphotoxin (LT
),1
and LT
are structurally homologous cytokines, and
their genes are closely clustered within the MHC (1, 2). TNF
is expressed mainly by macrophages and T cells as either a
transmembrane protein or a soluble homotrimeric molecule
(3). LT
exists as a soluble homotrimer (LT
3) but also on
the membrane of activated lymphocytes in heterotrimeric
complexes with LT
(1). TNF and LT
3 use the same cell
surface receptors, the p55 and p75 TNFRs, which are expressed on a wide variety of cells (4), whereas the predominant surface LT
1
2 heterotrimer binds to the LT
R,
which is expressed on cells of nonhematopoietic origin (5).
Recent studies in gene-targeted mice have revealed essential roles for TNF, LT, and LT
in secondary lymphoid organ structure and function. TNF was found to be
essential for the formation of primary B cell follicles, follicular dendritic cell (FDC) networks, and germinal centers in
all secondary lymphoid organs (6, 7). LT
knockout mice
lack lymph nodes and Peyer's patches and show a severe
disorganization of splenic architecture where B/T cell areas
do not segregate, marginal zones are absent, and FDC networks and germinal centers do not form (8, 9). Interestingly, mice deficient in both the LT
and TNF genes show
a phenotype identical to the LT
knockout phenotype
(10). Remarkably, however, these phenotypes were not
fully reproduced in LT
knockout mice, in which mesenteric and cervical lymph nodes do develop, splenic white
pulp lymphocytes segregate into B/T cell zones (11, 12), and
some capacity to form FDC networks and germinal centers
is preserved (12). These discrepancies led to the hypothesis
that LT
should have additional functions independent of
LT
in the organogenesis of mesenteric and cervical lymph
nodes and in the formation of distinct B/T cell areas, FDC
networks, and germinal centers in the spleen.
The apparently similar phenotypes of LT and double
TNF/LT
(10) knockout mice and the differences these
mice show when compared with LT
knockout mice (11-
13) have led us to search for functional redundancies in the
TNF/LT system by complementing LT
knockout mice
with TNF-expressing transgenes. Surprisingly, although
the lack of mesenteric and peripheral lymph nodes and Peyer's patches could not be rescued by transgenic expression of TNF, TNF-complemented LT
knockout mice
displayed intact T/B cell segregation and retained a suboptimal capacity to develop primary B cell follicles that contained FDC networks and could support the formation
of germinal centers. TNF-mediated restoration of LT
knockout splenic architecture was dependent on the presence of the p55TNFR. These observations suggested that
altered TNF expression may contribute to the complex
splenic phenotype of the LT
knockout mice. Indeed, defective TNF production in the LT
knockout mice could
be documented by measuring TNF accumulation in LPS-stimulated sera and in the supernatants of macrophages from LT
knockout mice. Our studies support a model
where LTs function to provide the optimal reticular/stromal splenic architecture for the efficient formation of B cell
follicles and FDC networks, phenomena primarily dependent on TNF/p55TNFR interactions.
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Materials and Methods |
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Mice.
LTImmunocytochemical Analysis of Splenic Structure.
Spleens were harvested, embedded in O.C.T. compound (BDH Laboratory Supplies, Dorset, UK), and frozen in liquid nitrogen. Cryostat sections were cut at 7 µm, thaw-mounted on gelatinized slides, air dried, and stored desiccated atImmunization Protocol.
Indicated groups of mice were immunized intraperitoneally with 108 SRBC in PBS on days 0 and 21. Mice were bled on day 28 for measurement of anti-SRBC serum antibodies.ELISA for SRBC-specific Serum Antibodies.
Sera from immunized mice were assayed using SRBC-specific ELISA for IgG1 antibodies as described previously (6). In brief, 96-well Immuno-Maxisorp plates (Nunc, Roskilde, Denmark) were coated with a solubilized extract from SRBC (100 µl at 5 µg/ml [21]) suspended in carbonate buffer, pH 9.6. Plates were washed with 0.05% Tween 20 in PBS and blocked with 1% BSA in PBS. Serum samples diluted in PBS containing 0.05% Tween 20, 1% BSA, and 1 M NaCl were incubated overnight at 4°C. Horseradish peroxidase-conjugated goat anti-mouse IgG1 (Southern Biotechnology Associates, Inc.) was diluted 1:5,000 in PBS containing 1% BSA, and 100 µl/well was added and incubated for 1 h at room temperature. ELISAs were developed with 0.4 mg/ml o-phenyldiamine dihydrochloride (Sigma Chemical Co.) in 0.05 M phosphate-citrate buffer, pH 5.0, containing 0.03% H2O2, stopped with 2 M H2SO4, and OD490 from duplicate wells was measured using a microplate reader (MRX; Dynatech Laboratories, Inc., Chantilly, VA).Measurement of TNF Production after LPS Stimulation.
Levels of TNF in macrophage supernatants and sera were determined as described previously (16). In brief, thioglycollate-elicited peritoneal macrophages were seeded at 5 × 105 cells/ml and incubated in the presence of 1 µg/ml LPS (Salmonella enteritidis, L6011; Sigma Chemical Co.) at 37°C, 5% CO2 for 24 h. Mice were injected intraperitoneally with 100 µg LPS in 0.5 ml PBS, and 90 min later blood samples were collected by cardiac puncture. The ELISA assay for murine TNF was provided by Dr. Wim Buurman (University of Limburg, Maastricht, The Netherlands) and performed as described previously (22). ![]() |
Results |
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LT knockout
mice (8, 9) as well as double LT
/TNF knockout mice (10)
show a more severe disorganization of their splenic architecture compared with TNF (6) or LT
knockout mice
(11, 12). To determine whether the additional lymphoid abnormalities seen in the LT
knockout mice may be rescued by TNF-specific signals, we complemented LT
knockout mice with TNF-expressing transgenes (TgTNF/
LT
/
). Two previously characterized TNF transgenic
lines were used: Tg1278 mice expressing a human wild-type TNF transgene (15) and TgA86 mice expressing a mutant transmembrane form of murine TNF from a TNF
1-12/
globin hybrid gene construct (reference 16, and see Materials and Methods). Tg1278 mice are free of pathology and
express a low level of wild-type human TNF mRNA in
several tissues, including thioglycollate-elicited peritoneal macrophages, thymus, lung, spleen, kidney, brain, skin, and
joints. Similar to endogenous murine TNF, low-level
mRNA expression of the wild-type human TNF transgene
does not result in a detectable level of TNF protein secretion in either sera or supernatants from thioglycollate-
elicited peritoneal macrophages as assessed by ELISA or
cytotoxicity assays (not shown). However, after LPS stimulation of ex vivo peritoneal macrophages, correct upregulation of transgene mRNA (15) and of protein production
(not shown) could be demonstrated. On the other hand,
TgA86 mice express a constitutively high level of the
TNF
1-12/globin mRNA in several tissues, including thioglycollate-elicited peritoneal macrophages, thymus, lung,
spleen, mesenteric lymph nodes, kidney, heart, brain, skin,
and joints (16). Constitutive overproduction of a bioactive
transmembrane TNF protein is suggested by the development of chronic inflammatory arthritis in these mice (16),
indicating aberrant regulation probably resulting from the
absence of the putatively suppressive TNF 3'-UTR from
the mRNA of this specific transgene.
Macroscopic examination of TNF-complemented LT
knockout mice (n >7 per transgenic line) showed that they
lack mesenteric and peripheral lymph nodes and Peyer's
patches, confirming a TNF-independent role for LTs in
the organogenesis of these lymphoid tissues. However, in
both TgTNF/LT
/
mouse lines, a substantial preservation of splenic structure could be observed (see below), indicating a composite nature of the LT
null mutation.
Double immunostaining with anti-B220 and anti-CD3 antibodies in
spleen sections of Tg1278/LT/
and TgA86/LT
/
mice revealed a conserved segregation of B and T cell
zones, with T cells clustered around the central arterioles
and B cells located peripheral to the T cell zone (Fig. 1).
This structural feature is directly comparable to the appearance of B/T cell areas in the spleens of LT
/
mice (Fig.
1), but is lacking completely in the spleens of LT
knockout mice (reference 8, and Fig. 1). These results show that
even in the absence of LT
, TNF is sufficient to maintain correct segregation of B/T cell areas in the spleen.
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To assess whether the observed involvement of TNF in
rescuing B/T cell segregation in LT knockout mice is
p55TNFR dependent, double TgA86/LT
/
mice were
bred into a p55TNFR knockout background (14). Splenic architecture in triple TgA86/LT
/
/p55TNFR
/
mice
remained indistinguishable from control LT
/
mice
(Fig. 1) and LT
/
/p55TNFR
/
mice (not shown),
demonstrating that in the absence of the p55TNFR, transgenic TNF expression could not rescue the splenic phenotype in LT
knockout mice. Therefore, restoration of
splenic structure in LT
knockout mice by TNF is mediated via the p55TNFR.
To determine whether transgenic TNF expression could reconstitute deficient follicular organization in LT knockout
mice, we analyzed splenic sections from TgTNF/LT
/
mice using double immunostaining with anti-IgM and
anti-IgD antibodies. By this staining, B cell follicles containing IgM+/IgD+ B cells can be clearly discriminated
from the IgMhigh/IgDlow marginal zone B cell population in
splenic sections from wild-type mice (Fig. 1). Consistent
with previous studies (23), double IgM/IgD staining revealed
the absence of follicular organization in spleens from LT
knockout mice (Fig. 1). However, similar analyses in spleens
from Tg1278/LT
/
, TgA86/LT
/
, and LT
/
mice
(Fig. 1) revealed the presence of organized primary B cell follicles although at reduced size and numbers compared
with wild-type controls. Double immunostaining with
anti-B220 as a marker for B cells, and an antibody to CR1
(mAb 8C12) as a marker for FDCs (18), showed that the
follicular structures observed in TgTNF/LT
/
and LT
/
mice contain networks of CR1+ FDCs (Fig. 2). These
FDC networks were greatly diminished in number and
size; however, similar to wild-type FDCs, they showed a
typical network organization and follicular localization.
Thus, even in the absence of LT
, TNF has the capacity to
support, albeit suboptimally, the development and maintenance of organized B cell follicles and FDC networks.
|
To examine the structure of the splenic marginal zone in
TgTNF/LT/
mice, we performed immunocytochemical analysis of splenic sections using markers specific for the
specialized macrophage populations of the marginal zone.
Double immunostaining with ER-TR9, an mAb recognizing marginal zone macrophages (19), and the 1C2 mAb against mouse sialoadhesin (20), which stains specifically the metallophilic macrophages in the spleen (24), revealed the
characteristic concentric organization of these macrophage
subsets peripheral to the white pulp in spleen sections from
wild-type mice (Fig. 2). However, similar to spleen sections
from LT
knockout mice (Fig. 2), double ER-TR9/antisialoadhesin staining of sections from TgTNF/LT
/
and
LT
/
spleens showed the absence of these macrophage
populations of the marginal zone (Fig. 2). Absence of metallophilic macrophages was also shown using the MOMA-1
mAb (data not shown; provided by G. Kraal, Vrije Universiteit, Amsterdam, The Netherlands), and staining using the
R3-3C12C7 anti-mucosal addressin cell adhesion molecule 1 mAb (provided by B. Holzmann, Technical University of Munich, Munich, Germany) demonstrated the complete absence of marginal zone mucosal addressin cell
adhesion molecule 1 expression from the spleens of LT
/
,
Tg1278/LT
/
, TgA86/LT
/
, and LT
/
mice (not
shown). Deficient marginal zone formation in these mice
was also confirmed by the absence of the characteristic
Ig-Mhigh/IgDlow marginal zone B cell population as assessed
by double IgM/IgD staining (Fig. 1). Taken together, these
data confirm the requirement for LT function in the development of the splenic marginal zone.
To investigate whether the rectified follicular organization in TgTNF/LT/
mice could
support development of germinal centers, we analyzed germinal center formation in the spleens of wild-type, LT
/
,
LT
/
, and TgTNF/LT
/
mice 10 d after immunization with the T cell-dependent (TD) antigen SRBC. Double immunocytochemical analysis using anti-B220 antibodies as a B cell marker and PNA as a marker for germinal
center B cells demonstrated the presence of typical germinal
centers forming within B cell follicles in wild-type mice
(Fig. 2). Consistent with previous studies (9), control LT
/
mice did not form typical germinal centers, although rare
aggregates of PNA+ cells could be detected around central
arterioles (reference 12, and Fig. 2). Similar analyses of
spleen sections from immunized Tg1278/LT
/
, TgA86/
LT
/
, and LT
/
mice revealed the presence of PNA+
germinal centers forming within B cell follicles (Fig. 2).
Although the number and size of these germinal centers
are reduced compared with wild-type mice, their formation within B cell follicles, their typical appearance as
B220+IgD
areas surrounded by IgD+ follicular mantle B
cells (not shown), and the finding that they contain FDC
networks distinguish them from the PNA+ patches often
observed to form around central arterioles in LT
(12) and
TNF knockout mice (6).
To investigate the ability of the TNF-complemented
LT knockout mice to respond to a TD immunization, we
tested the secondary antibody responses of TgTNF/LT
/
mice to the TD antigen SRBC. Wild-type, Tg1278/
LT
/
, and LT
/
mice were immunized intraperitoneally with SRBC on days 0 and 21, and anti-SRBC IgG1
antibody responses were measured on day 28 using an antigen-specific ELISA. Tg1278/LT
/
mice showed increased levels of IgG1 anti-SRBC antibodies compared with LT
/
mice, although they could not reach the levels produced by wild-type mice (Fig. 3). Similar results
were observed using TgA86/LT
/
mice (data not shown).
Thus, the observed germinal centers in the TNF-complemented LT
/
mice appear functional and contribute
positively, albeit suboptimally, to the development of a TD
humoral immune response.
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Given the surprising result of partial reconstitution
of the splenic phenotype of the LT knockout mice with
TNF-expressing transgenes, and the striking similarities in
the splenic architecture of TgTNF/LT
/
and LT
/
mice, it would seem likely that the splenic phenotype of
the LT
knockout mice could actually result from a coexistent deficiency in TNF production in these mice. To address this question, we measured levels of TNF protein
production in LPS-stimulated LT
knockout mouse sera
or ex vivo peritoneal macrophage supernatants. Compared
with background-matched wild-type control mice, LT
knockout mice show severely reduced accumulation of
TNF protein in both sera and macrophage exudates (Fig.
4), indicating that in addition to the LT
null mutation,
this strain of mice may show further complications due to
inherent defects in the neighboring TNF gene expression.
Further analyses should address the underlying mechanism producing the additional defect on TNF expression and assess the impact of this phenomenon in the interpretation
of different phenotypes occurring in the LT
knockout
mouse strain.
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![]() |
Discussion |
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Targeted disruption of genes encoding ligands and receptors in the TNF/LT family have clearly established the
important roles these molecules play in regulating the development and function of secondary lymphoid tissues (13,
25, 26). From these studies, a role has been proposed for
LT heterotrimers, presumably signaling through the
LT
R, in the organogenesis of lymph nodes and Peyer's patches and in the regulation of splenic structural organization. However, apparent differences between the lymphoid
phenotypes of LT
and LT
knockout mice suggested that
LT
should have additional biological activities independent of LT
(11). For example, defects in splenic T/B
cell organization and complete absence of FDC networks
and typical germinal centers in LT
or double LT
/TNF
knockout mice (8) could not be fully reproduced in LT
-deficient mice (12), which show more conserved B/T
cell organization and retain some capacity to form FDC
networks and germinal centers (Figs. 1 and 2). Using complementation analysis, we show in this study that many of
the phenotypic complications of LT
knockout mice in
the spleen may be compensated by the reintroduction of
functional TNF transgenes. Complementation of LT
knockout mice with TNF-encoding transgenes leads to a
p55TNFR-dependent restoration of B/T cell zone segregation and to the partial preservation of B cell follicles,
FDC networks, and germinal centers. Interestingly, the
TNF-complemented LT
-deficient phenotype is strikingly
similar to the splenic phenotype of LT
-deficient mice, as
demonstrated in this study (Figs. 1 and 2). These observations suggest that defective TNF expression may contribute
to the complex phenotype of the LT
knockout mice. Alternatively, it may be suggested that complementation of
the LT
knockout phenotype by TNF is due to a substitution for LT function rather than restoration of defective TNF expression, for example by transgenic overexpression
of TNF. However, this seems unlikely, since phenotypic
complementation by TNF occurs similarly with three independent transgenic lines carrying either correctly regulated or overexpressing TNF transgenes (i.e., Tg1278 mice
[15] expressing correctly regulated levels of human wild-type TNF, TgA86 mice [16] overexpressing a bioactive murine transmembrane TNF, or Tg6079 mice [our unpublished data and not shown here] expressing wild-type murine
TNF). In addition, transgenic expression of LT
in LT
knockout mice was reported recently as not sufficient to
rescue defective splenic architecture, although it could restore lymph node organogenesis (27). Interestingly, the observed splenic phenotype of TgLT
/LT
/
mice seems
to be similar to the phenotype of TNF-deficient mice (6),
with B cells organized in ring-like structures around the
periarteriolar lymphoid sheath, and PNA+ patches forming
around central arterioles (see Fig. 4 C in reference 27).
Most importantly, decreased expression of the endogenous TNF gene in LT knockout mice could be documented in this study by TNF-specific quantitative assays
(Fig. 4). It is not clear, however, whether this defect in
TNF expression occurs at the level of gene transcription,
mRNA translation, or protein processing. Although this
certainly awaits further detailed characterization, it is tempting to speculate that defective expression of the TNF gene in the LT
knockout mouse strain is due to transcriptional
interference caused by retention of a phosphoglycarate kinase
(PGK)-neo selection cassette within the targeted LT
locus.
This is now well documented in several other cases where retention of the PGK-neo cassette in targeted loci has yielded
unexpected phenotypes due to the altered expression of
neighboring genes (28, 29). In light of the evidence presented
here, interpretation of the different phenotypes occurring in
the LT
knockout mice should be carefully readdressed.
Interestingly, although expression of TNF is sufficient,
even in the absence of LT, to drive white pulp organization into distinct B and T cell areas and to partially support
follicular structure and germinal center formation, the function of LTs is clearly required for the development of the
marginal zone, as documented in this study but also as suggested previously by the complete absence of marginal
zone structures in LT
-deficient mice (11, 12). In this context, it is perhaps not very surprising that humoral responses to SRBC, as measured in the TNF-complemented LT
knockout system, although partially restored could
not reach normal levels. This may be due to the presence
of a suboptimal number of germinal centers in the spleen of
these mice, but it may also be due to the documented complete absence of marginal zones in this system. Indeed,
marginal zones are thought to play an important role in
processing particulate antigens such as SRBC (30).
Taken together with the presence of mesenteric and cervical lymph nodes in LT-deficient mice (11, 12), our evidence that TNF-complemented LT
knockout mice still
lack their mesenteric and peripheral lymphoid organs supports previous suggestions that LT
should have additional
lymphoid organogenetic functions, independent of LT
(11). It is perhaps interesting to note that correct B/T cell
segregation, and also primary follicular organization and
germinal center reactivity, appear in this study to be tightly regulated phenomena occurring in the absence of preserved
marginal zone structures, and seemingly uninfluenced by a
coinciding complete absence of secondary lymphoid organs. Therefore, it seems that they represent primary phenomena directly dependent on the local functioning of
TNF and LTs.
Inhibition of LT signaling either in transgenic mice
expressing a soluble LT
R-IgG1 fusion protein (31), or by
administration of soluble LT
R-Ig fusion proteins in normal adult mice (32), was shown to have profound effects
on splenic organization. Similar effects were not observed
when a p55TNFR-Ig fusion protein was administered to
adult mice, suggesting that at least within the 3-4-wk time
frame of inhibition, the TNF/p55TNFR system is not required for the maintenance of splenic architecture (32). We and others have previously suggested that a function for
TNF/p55TNFR in splenic organization may be in the development and differentiation of FDC networks which
would then support follicular organization and germinal
center reactivity. If this is true, then given the long-living
character of the FDCs, one would expect that only long-term inhibition of TNF signaling would be revealing for a role in the maintenance of splenic organization in the adult. The role of the TNF/p55TNFR system may also seem dispensable for correct B/T cell segregation, since this phenomenon is not affected in either TNF (6) or p55TNFR
knockout mice (14). However, our present evidence suggests that even in the absence of LTs, TNF is sufficient to
suboptimally support development and maintenance of follicular organization, indicating that the LT system shares a
redundant role with TNF in regulating the conserved appearance of splenic white pulp areas. In addition to the
splenic defects, administration of LT
R-Ig during gestation disrupted lymph node development (33), suggesting a
basic organogenetic role for LT
/
in these processes.
However, a differential role for TNF/p55TNFR in these
phenomena is suggested by the presence of secondary lymphoid organs with clear B/T cell zone segregation in the
TNF- (6) or p55TNFR-deficient strains of mice (14). As
discussed above, a function for TNF/p55TNFR in splenic
organization may be in the development and differentiation
of FDC networks which would then support follicular organization and germinal center reactivity (6, 7, 34). In support
of this hypothesis, recent bone marrow transplantation experiments in LT
- and p55TNFR-deficient mice showed
that FDC clustering induced by wild-type bone marrow transfers is dependent on the presence of the p55TNFR on
nonhematopoietic cells (35, 36). On the other hand, the
LT/LT
R system is expected to function on hematopoietic lineage cell interactions with nonlymphocytic stroma elements (37, 38), and it may be that such interactions control the basic architecture, which appears essential for the organogenesis of the lymph nodes but also for the fine
structural organization of the spleen. Formation of FDC
networks should be a composite phenomenon requiring an
optimal splenic infrastructure, perhaps provided by LTs,
and a TNF/p55TNFR-specific signal which leads to the
maturation/differentiation and/or follicular localization of
FDCs. The capacity to form FDC networks and germinal
centers even in the absence of LTs, as documented in both
TNF-complemented LT
knockout mice and LT
knockout mice, supports this hypothesis. Therefore, it is
likely that the role of LTs is functionally distinct from that
of TNF, and that both pathways need to cooperate for the
optimal development and maintenance of splenic structure.
![]() |
Footnotes |
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Address correspondence to George Kollias, Department of Molecular Genetics, Hellenic Pasteur Institute, 127 Vas. Sophias Ave., 115 21 Athens, Greece. Phone: 30-1-6455071; Fax: 30-1-6456547; E-mail: giorgos_kollias{at}hol.gr M. Pasparakis's current address is Institute for Genetics, University of Cologne, 50931 Cologne, Germany.
Received for publication 15 April 1998 and in revised form 8 June 1998.
This project was supported in part by the Hellenic Secretariat for Research and Technology, and by European Commission grants BIO-CT96-0077 and BIO-CT96-0174.We wish to thank Klaus Pfeffer and Sergei Nedospasov for providing LT knockout mice and for useful discussions, Horst Bluethmann for providing p55TNFR knockout mice, Wim Buurman for the murine TNF-specific ELISA, Paul Crocker for antisialoadhesin antibody, Steve Cobbold for the KT3 anti-CD3 antibody,
Peter Leenen for the ER-TR9 antibody, George Kraal for the MOMA-1 antibody, Dimitris Kontoyiannis
for sharing unpublished data, and Anna Kefalaki for technical assistance with the histological analyses.
Abbreviations used in this paper FDC, follicular dendritic cell; LT, lymphotoxin; PNA, peanut agglutinin; TD, T cell-dependent; Tg, transgenic; UTR, untranslated region.
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References |
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---|
1. | Browning, J.L., A. Ngam-ek, P. Lawton, J. DeMarinis, R. Tizard, E.P. Chow, C. Hession, B. O'Brine-Greco, S.F. Foley, and C.F. Ware. 1993. Lymphotoxin beta, a novel member of the TNF family that forms a heteromeric complex with lymphotoxin on the cell surface. Cell. 72: 847-856 [Medline]. |
2. | Pokholok, D.K., I.G. Maroulakou, D.V. Kuprash, M.B. Alimzhanov, S.V. Kozlov, T.I. Novobrantseva, R.L. Turetskaya, J.E. Green, and S.A. Nedospasov. 1995. Cloning and expression analysis of the murine lymphotoxin beta gene. Proc. Natl. Acad. Sci. USA. 92: 674-678 [Abstract]. |
3. | Kriegler, M., C. Perez, K. DeFay, I. Albert, and S.D. Lu. 1988. A novel form of TNF/cachectin is a cell surface cytotoxic transmembrane protein: ramifications for the complex physiology of TNF. Cell. 53: 45-53 [Medline]. |
4. | Vandenabeele, P., W. Declercq, R. Beyaert, and W. Fiers. 1995. Two tumour necrosis factor receptors: structure and function. Trends Cell Biol. 5: 392-399 . |
5. | Crowe, P.D., T.L. VanArsdale, B.N. Walter, C.F. Ware, C. Hession, B. Ehrenfels, J.L. Browning, W.S. Din, R.G. Goodwin, and C.A. Smith. 1994. A lymphotoxin-beta-specific receptor. Science. 264: 707-710 [Medline]. |
6. | Pasparakis, M., L. Alexopoulou, V. Episkopou, and G. Kollias. 1996. Immune and inflammatory responses in TNF alpha-deficient mice: a critical requirement for TNF alpha in the formation of primary B cell follicles, follicular dendritic cell networks and germinal centers, and in the maturation of the humoral immune response. J. Exp. Med. 184: 1397-1411 [Abstract]. |
7. |
Pasparakis, M.,
L. Alexopoulou,
M. Grell,
K. Pfizenmaier,
H. Bluethmann, and
G. Kollias.
1997.
Peyer's patch organogenesis is intact yet formation of B lymphocyte follicles is defective in peripheral lymphoid organs of mice deficient for tumor necrosis factor and its 55-kDa receptor.
Proc. Natl. Acad.
Sci. USA.
94:
6319-6323
|
8. | De Togni, P., J. Goellner, N.H. Ruddle, P.R. Streeter, A. Fick, S. Mariathasan, S.C. Smith, R. Carlson, L.P. Shornick, J. Strauss-Schoenberger, et al . 1994. Abnormal development of peripheral lymphoid organs in mice deficient in lymphotoxin. Science. 264: 703-707 [Medline]. |
9. | Matsumoto, M., S. Mariathasan, M.H. Nahm, F. Baranyay, J.J. Peschon, and D.D. Chaplin. 1996. Role of lymphotoxin and the type I TNF receptor in the formation of germinal centers. Science. 271: 1289-1291 [Abstract]. |
10. | Eugster, H.P., M. Muller, U. Karrer, B.D. Car, B. Schnyder, V.M. Eng, G. Woerly, M. Le Hir, F. di Padova, M. Aguet, et al . 1996. Multiple immune abnormalities in tumor necrosis factor and lymphotoxin-alpha double-deficient mice. Int. Immunol. 8: 23-36 [Abstract]. |
11. | Koni, P.A., R. Sacca, P. Lawton, J.L. Browning, N.H. Ruddle, and R.A. Flavell. 1997. Distinct roles in lymphoid organogenesis for lymphotoxins alpha and beta revealed in lymphotoxin beta-deficient mice. Immunity. 6: 491-500 [Medline]. |
12. |
Alimzhanov, M.B.,
D.V. Kuprash,
M.H. Kosco-Vilbois,
A. Luz,
R.L. Turetskaya,
A. Tarakhovsky,
K. Rajewsky,
S.A. Nedospasov, and
K. Pfeffer.
1997.
Abnormal development of
secondary lymphoid tissues in lymphotoxin beta-deficient
mice.
Proc. Natl. Acad. Sci. USA.
94:
9302-9307
|
13. |
von Boehmer, H..
1997.
Lymphotoxins: from cytotoxicity to
lymphoid organogenesis.
Proc. Natl. Acad. Sci. USA.
94:
8926-8927
|
14. | Rothe, J., W. Lesslauer, H. Lotscher, Y. Lang, P. Koebel, F. Kontgen, A. Althage, R. Zinkernagel, M. Steinmetz, and H. Bluethmann. 1993. Mice lacking the tumour necrosis factor receptor 1 are resistant to TNF-mediated toxicity but highly susceptible to infection by Listeria monocytogenes. Nature. 364: 798-802 [Medline]. |
15. | Keffer, J., L. Probert, H. Cazlaris, S. Georgopoulos, E. Kaslaris, D. Kioussis, and G. Kollias. 1991. Transgenic mice expressing human tumour necrosis factor: a predictive genetic model of arthritis. EMBO (Eur. Mol. Biol. Organ.) J. 10: 4025-4031 [Abstract]. |
16. | Alexopoulou, L., M. Pasparakis, and G. Kollias. 1997. A murine transmembrane tumour necrosis factor transgene induces arthritis by cooperative p55/p75 tumour necrosis factor receptor signaling. Eur. J. Immunol. 27: 2588-2592 [Medline]. |
17. | Tomonari, K.. 1988. A rat antibody against a structure functionally related to the mouse T-cell receptor/T3 complex. Immunogenetics. 28: 455-458 [Medline]. |
18. |
Kinoshita, T.,
J. Takeda,
K. Hong,
H. Kozono,
H. Sakai, and
K. Inoue.
1988.
Monoclonal antibodies to mouse complement receptor type 1 (CR1). Their use in a distribution study
showing that mouse erythrocytes and platelets are CR1-negative.
J. Immunol.
140:
3066-3072
|
19. | van Vliet, E., M. Melis, and W. van Ewijk. 1985. Marginal zone macrophages in the mouse spleen identified by a monoclonal antibody. Anatomical correlation with a B cell subpopulation. J. Histochem. Cytochem. 33: 40-44 [Abstract]. |
20. | Crocker, P.R., S. Freeman, S. Gordon, and S. Kelm. 1995. Sialoadhesin binds preferentially to cells of the granulocytic lineage. J. Clin. Invest. 95: 635-643 [Medline]. |
21. | Kelly, B.S., J.G. Levy, and L. Sikora. 1979. The use of the enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of specific antibody from cell cultures. Immunology. 37: 45-52 [Medline]. |
22. |
Bemelmans, M.H.A,
D.J. Gouma, and
W.A. Buurman.
1993.
LPS-induced sTNF-receptor release in vivo in a murine
model. Investigation of the role of tumor necrosis factor, IL-1,
leukemia inhibiting factor, and IFN-gamma.
J. Immunol.
151:
5554-5562
|
23. |
Fu, Y.X.,
G. Huang,
M. Matsumoto,
H. Molina, and
D.D. Chaplin.
1997.
Independent signals regulate development of
primary and secondary follicle structure in spleen and mesenteric lymph node.
Proc. Natl. Acad. Sci. USA.
94:
5739-5743
|
24. | Crocker, P.R., and S. Gordon. 1989. Mouse macrophage hemagglutinin (sheep erythrocyte receptor) with specificity for sialylated glycoconjugates characterized by a monoclonal antibody. J. Exp. Med. 169: 1333-1346 [Abstract]. |
25. | Pasparakis, M., L. Alexopoulou, E. Douni, and G. Kollias. 1996. Tumour necrosis factors in immune regulation: everything that's interesting is . . . new! Cytokine Growth Factor Rev. 7: 223-229 . [Medline] |
26. | Liu, Y.J., and J. Banchereau. 1996. Mutant mice without B lymphocyte follicles. J. Exp. Med. 184: 1207-1211 [Medline]. |
27. |
Sacca, R.,
S. Turley,
L. Soong,
I. Mellman, and
N.H. Ruddle.
1997.
Transgenic expression of lymphotoxin restores
lymph nodes to lymphotoxin-![]() |
28. | Olson, E.N., H.H. Arnold, P.W. Rigby, and B.J. Wold. 1996. Know your neighbors: three phenotypes in null mutants of the myogenic bHLH gene MRF4. Cell. 85: 1-4 [Medline]. |
29. |
Pham, C.T.,
D.M. MacIvor,
B.A. Hug,
J.W. Heusel, and
T.J. Ley.
1996.
Long-range disruption of gene expression by
a selectable marker cassette.
Proc. Natl. Acad. Sci. USA.
93:
13090-13095
|
30. | Kraal, G.. 1992. Cells in the marginal zone of the spleen. Int. Rev. Cytol. 132: 31-74 [Medline]. |
31. |
Ettinger, R.,
J.L. Browning,
S.A. Michie,
W. van Ewijk, and
H.O. McDevitt.
1996.
Disrupted splenic architecture, but
normal lymph node development in mice expressing a soluble lymphotoxin-beta receptor-IgG1 fusion protein.
Proc.
Natl. Acad. Sci. USA.
93:
13102-13107
|
32. | Mackay, F., G.R. Majeau, P. Lawton, P.S. Hochman, and J.L. Browning. 1997. Lymphotoxin but not tumor necrosis factor functions to maintain splenic architecture and humoral responsiveness in adult mice. Eur. J. Immunol. 27: 2033-2042 [Medline]. |
33. |
Rennert, P.D.,
J.L. Browning,
R. Mebius,
F. Mackay, and
P.S. Hochman.
1996.
Surface lymphotoxin ![]() ![]() |
34. | Le Hir, M., H. Bluethmann, M.H. Kosco-Vilbois, M. Muller, F. di Padova, M. Moore, B. Ryffel, and H.P. Eugster. 1996. Differentiation of follicular dendritic cells and full antibody responses require tumor necrosis factor receptor-1 signaling. J. Exp. Med. 183: 2367-2372 [Abstract]. |
35. |
Matsumoto, M.,
Y.X. Fu,
H. Molina,
G. Huang,
J. Kim,
D.A. Thomas,
M.H. Nahm, and
D.D. Chaplin.
1997.
Distinct roles of lymphotoxin alpha and the type I tumor necrosis factor (TNF) receptor in the establishment of follicular
dendritic cells from non-bone marrow-derived cells.
J. Exp.
Med.
186:
1997-2004
|
36. |
Tkachuk, M.,
S. Bolliger,
B. Ryffel,
G. Pluschke,
T.A. Banks,
S. Herren,
R.H. Gisler, and
M.H. Kosco-Vilbois.
1998.
Crucial role of tumor necrosis factor receptor 1 expression on nonhematopoietic cells for B cell localization within
the splenic white pulp.
J. Exp. Med.
187:
469-477
|
37. | Ware, C.F., T.L. VanArsdale, P.D. Crowe, and J.L. Browning. 1995. The ligands and receptors of the lymphotoxin system. Curr. Top. Microbiol. Immunol. 198: 175-218 [Medline]. |
38. |
Mebius, R.E.,
P. Rennert, and
I.L. Weissman.
1997.
Developing lymph nodes collect CD4+CD3![]() ![]() |