bluuebunny/crossref_metadata_2025 huggingface dataset:
Lymphoid Tumors of Xenopus laevis with Different Capacities for Growth in Larvae and Adults
Authors: Robert Jacques , Guiet Chantal , Du Pasquier Louis
Published: June 2008

Three new lymphoid tumors offering an assortment of variants in terms of MHC class I
expressions, MHC class II expression, and Ig gene transcription have been discovered in the
amphibian Xenopus. One was developed in an individual of the isogenic LG15 clone
(LG15/0), one in a frog of the LG15/40 clone (derived from a small egg recombinant of
LG15), and one (ff‐2) in a male ff sib of the individual in which MAR1, the first lymphoid
tumor in Xenopus was found 2 years ago. These tumors developed primarily as thymus
outgrowths and were transplantable in histocompatible tadpoles but not in nonhistocompatible
hosts. Whereas LG15/0 and LG15/40 tumor cells also grow in adult LG15 frogs, the
ff‐2 tumor, like the MAR1 cell line, is rejected by adult ff animals. Using flow cytometry with
fluorescence‐labeled antibodies and immunoprecipitation analysis, we could demonstrate
that, like MAR1, these three new tumors express on their cell surface lymphopoietic
markers recognized by mAbs FIF6 and RC47, as well as T‐cell lineage markers recognized
by mAbs AM22 (CD8‐1ike) and X21.2, but not by immunologobulin (Ig) nor MHC class II
molecules. Another lymphocyte‐specific marker AM15 is expressed by 15/0 and 15/40 but
not ff‐2 tumor cells. The ff‐2 tumor cell expresses MHC class molecule in association with
β2‐microglobulin on the surface, 15/40 cells contain cytoplasmic I α chain that is
barely detected at the cell surface by fluocytometry, and 15/0 cells do not synthesize class
I α chain at all. The three new tumors all produce large amounts of IgM mRNA of two
different sizes but no Ig protein on the membrane nor in the cytoplasm. All tumor cell types
synthesize large amount of Myc mRNA and MHC class I‐like transcripts considered to be
non classical.
Effects of IgM Allotype Suppression on Serum IgMLevels, B‐1 and B‐2 Cells, and Antibody Responses irAllotype Heterozygous F1 Mice
Authors: Hamilton Ann Marie , Kearney John F.
Published: June 2008

IgM allotype heterozygous F1 mice were independently suppressed for Igh6a or Igh6b to
evaluate the contribution of B‐1 and B‐2 cells to natural serum IgM levels and Ab responses.
B‐2 B cells expressing IgM of the suppressed allotype were evident in the spleens of
suppressed mice 4 to 6 weeks after cessation of the suppression regimen, whereas B‐1 B
cells of the suppressed allotype were undetectable for up to 9 months. Although serum IgM
of the suppressed allotype was initially depleted in mice suppressed for either allotype, by
7 months of age, there were detectable levels of IgM of the suppressed allotype in the
serum; however, the levels were significantly below that found in nonsuppressed mice.
When mice were immunized with either the T‐independent or T‐dependent form of
phosphorylcholine, those suppressed for either allotype, and consequently depleted of B‐1
B cells of that allotype, did not respond with phosphorylcholine‐specific IgM of the
suppressed allotype. In contrast, when mice were immunized with α1‐3 dextran, the Igh6a
allotype‐suppressed mice were able to produce dextran‐specific IgM of that allotype. These
results show that allotype‐bearing B‐1 cells of both allotypes can be effectively suppressed
by this suppression protocol and this produces long‐lasting effects on B‐1 cell levels and
serum IgM of the suppressed allotype. These observations reflect the derivation of the
majority of B‐1 cells from fetal‐neonatal precursors, which cannot be replaced by newly
emerging B‐2 cells of adult origin. Their ablation by antibody treatment results in
permanent alterations to the adult B‐cell repertoire.
Reconstitution Properties of Thymus Stem Cells in Murine Fetal Liver
Authors: Mckenna Hilary J. , Birchall Nicholas M. , Watson James D.
Published: June 2008

Injection of day‐12 murine fetal liver cells into thymus lobes of Thy‐1 congenic adult
recipients results in a wave of thymocyte development. The kinetics of repopulation by
donor cells reaches a peak after 20–25 days. The frequency of thymic stem cells (TSC) in
day‐12 fetal liver was estimated, by limit dilution, as 1 in 4x104 cells. Within 8 hr of
injection into a thymus lobe, fetal liver TSC commit to T‐cell development, losing stem‐cell
activity. When fetal liver cells are maintained in culture for 7 days, with no exogenous
cytokines added, and then injected intra‐thymically (I.T.), thymus recolonization is not
observed. However, TSC can be maintained in culture for 7 days with IL‐1β, IL‐3, IL‐6, or
LIF added, alone or in combination, with steel factor (SLF). Poisson analysis of fetal liver
cells cultured with SLF and IL‐3 together revealed a precursor frequency of 1 in 1.8x 105
cells. In contrast, the frequency of TSC in adult bone marrow was estimated by limit
dilution as 1 in 12,000 cells.
Overusage of Mouse DH Gene Segment, DFL16.1, IsStrain‐Dependent and Determined by cis‐Acting Elements
Authors: Atkinson Michael J. , Chang Yenhui , Celler Jakub W. , Huang Carol , Paige Christopher J. , Wu Gillian E.
Published: June 2008

The DJH structure is of particular importance for diversity in the immunoglobulin heavy
chain because it encodes most of CDR3. Here, we investigate mechanisms responsible for
generating the DJH structure. We found DFL16.1 was used at a high frequency in normal
and transformed pre‐B cells (fetal liver > 50%, A‐MuLV lines ≅ 25%). One DFL16.1JH1
structure was found repeatedly and was also present in DJH and VDJH databases,
suggesting this structure may be conserved in the primary repertoire. Genetic analysis
demonstrated that C57BL/6 mice use DFL16.1 in DJH structures more frequently than
BALB/c. Examination of individual alleles in (C57BL/6 BALB/c)F1 A‐MuLV cell lines
revealed that the C57BL/6‐derived allele used DFL16.1 twice as often as the BALB/c. This
result indicates that part of the mechanism ensuring overusage of DFL16.1 gene segments
is cis‐acting.
Quantitative Analysis of the Efficiency of Clonal Deletion in the Thymus
Authors: Spain Lisa M. , Berg Leslie J.
Published: June 2008

One of the major mechanisms for establishing self‐tolerance is the clonal deletion of
self‐reactive T cells during their development in the thymus. Using a TCR transgenic mouse
model, we have established a quantitative ex vivo assay for examining the sensitivity and
specificity of negative selection. Thymic organ cultures established from mice of varying
MHC haplotypes were incubated with antigen, and the efficiency of clonal deletion
assessed. We show here that clonal deletion of CD4+8+ thymocytes is sensitive to both the
gene dosage and the allelic variation of MHC class II molecules expressed on thymic
antigen‐presenting cells. We also find that when epithelial cells in the thymic cortex are the
only antigen‐presenting cells expressing the appropriate MHC class II molecules, negative
selection of CD4+8+ cells is as efficient as when antigen is presented on all thymic
antigen‐presenting cells. These studies demonstrate that the induction of self‐tolerance via
clonal deletion in the thymus is a function not only of antigen concentration, but also of
MHC class II cell‐surface density. In addition, together with the reports of others, these
results confirm that cortical epithelial cells can mediate negative selection, and demonstrate
that they do so in the intact thymic microenvironment.
Role of β2 Integrins in the Binding of Thymocytes to RatThymic Macrophages
Authors: Čolić Miodrag , Ilić Vesna , Tamatani Takuya , Miyasaka Masayuki , Pavlović Miloš D.
Published: June 2008

A role of β2 integrins and one of their ligands, ICAM‐1, in thymic macrophage
(TMF)/thymocyte interactions was studied. TMF were isolated as adherent cells from 4‐day
old culture of thymic‐cell suspensions either from normal or hydrocortisone‐treated rats.
Adherent cells were 94‐98% positive with ED1 (a pan‐macrophage marker). The majority
of them (75‐95%) expressed the CD11b and CD18 molecules, and 60‐70% expressed CD54
(ICAM‐1). A low proportion of TMF (10‐20%) expressed CDlla (LFA‐1). The expression
of all these antigens was upregulated by IFN‐α and TNF‐α. The effect of these mAbs on
TMF/thymocyte binding was studied using a simple rosette assay by incubating unstimulated
or IFN‐γ or TNF‐α stimulated TMF, grown on microscopic slides with resting or
ConA +IL‐2 activated thymocytes. It was found that LFA‐1/CD18 and ICAM‐1 play a
significant role in the TMF/thymocyte adhesion. In addition, a LFA‐l‐dependent/ICAM‐
1‐independent adhesion pathway was observed, suggesting that LFA‐1 might use another
ligand. The inhibitory effect of anti‐CD18 mAb (WT‐3) was higher than the effect of
anti‐LFA‐1 mAb (WT‐1) and was a consequence of blocking the CD18 chain both on
thymocytes and TMF. No significant difference in the expression and function of adhesion
molecules was found between TMF obtained from normal or hydrocortisone‐treated rats.
The involvement of CD1 1b in these processes was of lesser importance than the role of the
CD11a molecule. By using mAbs to different epitopes of the CD11b molecule, such as
OX‐42 (anti‐CD11b/CD11c), ED7, and ED8 (anti‐CD11b), it was found that they were
either slightly or moderately inhibitory under certain experimental conditions or did not
significantly modulate TMF/thymocyte binding. Oχ‐42 was slightly stimulatory in some
experiments. Cumulatively, these results show that 2 integrins play a significant role in
TMF/thymocyte interactions and probably contribute to T‐cell development in vivo.
Thymic Microenvironment and Lymphoid Responses toSublethal Irradiation
Authors: Randle-Barrett Elise S. , Boyd Richard L.
Published: June 2008

Sublethal irradiation of the murine thymus has been a useful tool for depleting the thymus
of dividing immature thymocyte subsets, to sequence thymocyte differentiation events
occurring from radiation‐resistant precursors. This massive reduction in thymocytes also
represents a model in which the bidirectional interplay between the thymic stromal cells
and lymphocytes can be investigated. The purpose of this study was thus twofold: to
precisely map the initiation of thymopoiesis as a prelude to assessing the effects of injected
mAb to novel thymic antigens; and to use a panel of mAbs to determine the alterations in
the thymic stroma during the T‐cell depletion and reconstitution phases. The striking
finding from this study was that following T‐cell depletion, there was a marked upregulation
of specific stromal antigens, which retracted with the reappearance of T cells. Thus,
following sublethal irradiation, there are modifications in the thymic microenvironment that
may be necessary to support renewed thymopoiesis and the complete restoration of the
thymus involved the synchronous development of both the stromal and lymphocytic
components.
Differential Effects of X‐Irradiation and Cyclosporin‐AAdministration on the Thymus with Respect to theGeneration of Cyclosporin‐A‐Induced Autoimmunity
Authors: Beijleveld Leo J.J. , Damoiseaux Jan G.M.C. , Van Breda Vriesman Peter J.C.
Published: June 2008

Cyclosporin A (CsA), a potent inhibitor of T‐cell activation, has been shown to have several
effects on thymocyte maturation, thymic stromal cells, and the generation of autoreactive T
cells. In Lewis rats, the combination of lethal irradiation, syngeneic bone marrow
transplantation, and a 4‐week course of CsA administration causes the development of an
autoimmune disease (CsA‐AI) resembling allogeneic graft‐versus‐host disease. This occurs
upon withdrawal of CsA, provided the thymus receives irradiation and is present during
CsA treatment. In this study, the separate effects of irradiation or CsA treatment on thymic
stromal cells and thymocytes, compared to the combinatory effects, were examined using
immunohistochemistry and tricolor flow cytometric analysis.

CsA treatment causes an involution of the thymic medulla and a strong reduction of the
cell number of thymocytes and stromal cells residing in the medulla. However, within the
remaining medullary area, changes in cell distribution and antigen density on these cells
were not observed. Irradiation on the other hand causes a strong depletion of thymocytes.
The thymocyte population is recovered within 2 weeks and a cortical and medullary region
can be distinguished. CsA treatment in combination with irradiation results in a strongly
inhibited recovery of the medulla during CsA treatment, whereas the cortex recovers to
normal size and morphology. The presence of the medullary IDC and epithelial cells is
reduced proportionally to the small size of the medulla. However, the distribution of these
stromal cells is normal. During the CsA administration, the thymuses from irradiated and
CsA‐treated rats are very similar to thymuses from CsA‐treated rats. In conclusion, no
changes specific for irradiation plus CsA treatment have been observed. Regarding the
distribution and size of medullary stromal cells and residing thymocytes, thymuses from
irradiated and CsA‐treated rats hardly differ from the thymuses from rats treated only with
CsA. Therefore, irradiation seems essential in the generation of CsA‐AI by eliminating
suppressor‐cell circuits in the periphery.
A Common Stem Cell for Murine Cortical and MedullaryThymic Epithelial Cells?
Authors: Röpke Carsten , Van Soest Peter , Platenburg Peter Paul , Van Ewijk Willem
Published: June 2008

We have addressed the question whether the epithelial stroma in the thymus is derived
from a common stem cell or whether cortical and medullary epithelial cells are derived from
different embryonic stem cells emerging, for example, from endoderm and ectoderm. By the
use of rapidly expanding cultures of thymic epithelial cells (TEC) from 14 to 16 day‐old
murine fetuses and by specific antibodies against cortical and medullary epithelium,
respectively, we were able to demonstrate a small subpopulation of double‐labeled TEC in
the cultures. These cells were not present in TEC cultures initiated from thymuses of
neonatal mice. Double‐labeled TEC were also found in tissue sections from fetal thymuses.
These findings may indicate that TEC populations of the cortex and the medulla are derived
from a common stem cell, with potential for differentiation toward both cortical and
medullary TEC.
Antigen‐Independent Maturation of CD2, CD11a/CD18, CD44, and CD58 Expression on Thymic Emigrants in Fetal and Postnatal Sheep
Authors: Witherden Deborah A. , Abernethy Nevin J. , Kimpton Wayne G. , Cahill Ross N. P.
Published: June 2008

We have compared the expression of CD2, CD11a/CD18, CD44, and CD58 on αβ and γδ
T cells emigrating from the fetal and postnatal thymus. We report that both γδ and the
CD4+ CD8- and CD4-CD8+ subsets of αβ T cells express mature levels of the adhesion
molecules CD11a/CD18, CD44, and CD58 upon emigration from the thymus. Whereas
CD44 is up‐regulated on γδ+ thymocytes prior to export, down‐regulation of both
CD11a/CD18 and CD58 occurs prior to emigration from the thymus, suggesting that
down‐regulation of these molecules may be a final maturational step taken by developing
γδ T cells before their export from the thymus. In contrast, there is continued up‐regulation
of CD2 on αβ and γδ T cells upon emigration from the thymus and as they move into the
mature peripheral T‐cell pool. There was also a marked reduction in the number of CD2+γδ T cells exported during fetal development that was associated with a marked reduction
in the percentage of CD22+γδ thymocytes exported. The postthymic maturation of CD2 and
the other changes in adhesion‐molecule expression appear to be independent of extrinsic
antigen, as the same changes were observed in the antigen‐free environment of the fetus
as in the postnatal lamb, which has been exposed to extrinsic antigen. It thus appears that
these changes in adhesion‐molecule expression are as a result of the normal maturation
pathway from a developing thymocyte to a mature peripheral T cell.
Somatic Mutations During an Immune Response inXenopus Tadpoles
Authors: Wilson Melanie , Marcuz Anne , Du Pasquier Louis
Published: June 2008

The tadpole B‐cell repertoire is less diverse than that of the adult frog; their antibodies are
of lower affinity and are less heterogenous. In order to determine whether this difference
is due to a lack of or a reduced rate of somatic hypermutation, we analyzed and compared
cDNA sequences utilizing VH1 elements with germline counterparts in isogenic LG7
tadpoles during an immune response. Indeed, tadpole VH1 sequences contained somatic
mutations. There were zeo to 5 mutations per sequence, all single base‐point mutations,
with the high ratio of GC to AT base‐pair alterations similar to that observed in adult frogs.
Cytokine Production in the Serum and Spleen of Micefrom Day 6 to 14 of Gestation: Cytokines/Placenta/Spleen/Serum
Authors: Athanassakis Irene , Iconomidou Bessy
Published: June 2008

Pregnancy, like most biologic phenomena, involves the action of cytokines. These proteins
have a short half‐life and are believed to exert their effect close to their site of production,
where diagnostic tests cannot be easily performed. Here we show that the cytokine content
in the maternal serum reflects cytokine production and secretion from maternal spleen
cells, which also correlates with production from decidual cells. We show that GM‐CSF, IL‐
3, and IL‐10 are present in the serum at specific time intervals during the first half of murine
pregnancy, which correlates with their production from maternal spleen cells. Purified
GM‐CSF and IL‐3 from spleen‐cell‐culture supernatants are biologically active molecules,
able to stimulate placental‐cell proliferation. Furthermore, TNF‐0, which has been identified
in many cases of fetal rejection as well as in labor, is shown to be naturally produced
during the second half of pregnancy. Additionally, within the limits of the sensitivity of the
technique we have used, the detection of IL‐4 and the absence of detectable levels of IL‐
2 in the maternal serum strongly comforts the hypothesis that pregnancy is a Th2‐dependent
phenomenon. The results presented in this paper show that the cytokine profile during
pregnancy can be monitored by simple blood tests, which may be of relevance both in the
followup of a physiological human pregnancy and to the diagnosis of recurrent abortions
due to cytokine imbalance.
Two Distinct Pathways of B‐Cell Development inPeyer’s Patches
Authors: Griebel Philip J. , Kugelberg Birgit , Ferrari Giorgio
Published: June 2008

The developmental biology of sheep ileal and jejunal Peyer’s patches (PP) was investigated
using corticosteroids to deplete immature B lymphocytes. During a 7‐day treatment with
dexamethasone, ileal PP follicular (iPf)B‐cell proliferation was arrested and most iPfB‐cells
died. This resulted in follicular involution with the survival of mesenchymal cells. No
iPfB‐cell proliferation was detected in follicular remnants for 4 weeks postdexamethasone
treatment, and during a subsequent 3‐month period, there was limited iPfB‐cell proliferation
that resulted in a partial regeneration of follicles. Ileal PP involution was also associated
with a severe B lymphopenia that persisted for over 14 weeks and was characterized by the
survival of primarily isotype‐switched and CD5+ sIgM+ B‐cells in blood. In contrast, the
size of jejunal PP follicles was reduced following dexamethasone treatment, but intrafollicular
B‐cell proliferation was not arrested. Furthermore, within 4 weeks, the jejunal PP
follicles had recovered in size and cellularity and there was no disruption in IgA plasma‐cell
production. Thus, dexamethasone selectively depleted iPfB‐cells and revealed that the ileal
and jejunal PPs contain functionally distinct B‐cell populations. The partial regeneration of
the iPfB‐cell population indicated that either an intrafollicular, corticosteroid‐resistant
B‐stem cell existed or that ileal PP follicles can be repopulated by circulating B‐cells. Finally,
the association between ileal PP involution and the absence of circulating, CD5- B-cells
confirmed that this lymphoid tissue provides an essential environment for conventional
sIgM+ B‐cell development.
Development of Mouse Embryonic Stem (ES) Cells:IV Differentiation to Mature T and B Lymphocytes afterImplantation of Embryoid Bodies Into Nude Mice
Authors: Chen Una , Mok Hoyan
Published: June 2008

Mouse embryonic stem (ES) cells in culture can differentiate into late stages of many
lineage‐committed precursor cells. Under appropriate organ‐culture conditions, ES cels
differentiate into lymphoidlike cells at a stage equivalent to lymphoid cells found in fetal
liver. These hematopoietic precursors are located in cup‐shaped structures found in some
embryoid bodies; we called such embryoid bodies “ES fetuses.” In this study, we have
followed the maturation of hematopoietic cells after implantation of ES fetuses into nude
mice for 3 weeks. ES‐cell‐derived lymphoid cells‐pre‐B cells, mature B cells, and mature T
cells were found in all lymphoid organs. Interestingly, there was also an increase of T cells
of host origin. Because native nude mouse lack thymus, these T cells might be educated by
thymuslike epithelium generated from ES fetuses. Practical applications of this combined in vitro
and in vivo system are discussed.
Tolerance in TCR/Cognate Antigen Double‐TransgenicMice Mediated by Incomplete Thymic Deletion andPeripheral Receptor Downregulation
Authors: Mamalaki Clio , Murdjeva Marianna , Tolaini Mauro , Norton Trisha , Chandler Phillip , Townsend Alain , Simpson Elizabeth , Kioussis Dimitris
Published: June 2008

Influenza nucleoprotein (NP)‐specific T‐cell receptor transgenic mice (F5) were crossed
with transgenic mice expressing the cognate antigenic protein under the control of the H‐
2Kb promoter. Double‐transgenic mice show negative selection of thymocytes at the
CD4+8+TCR10 to CD4+8+TCRhi transition stage. A few CD8 T cells, however, escape clonal
deletion, and in the peripheral lymphoid organs of these mice, they exhibit low levels of
the transgenic receptor and upregulated levels of the CD44 memory marker. Such cells do
not proliferate upon exposure to antigen stimulation in vivo or ex vivo, however, they can
develop low but detectable levels of antigen‐specific cytotoxic function after stimulation
in vitro in the presence of IL‐2.
Kinetics of Thymocyte Subset Development and SelectionRevealed by Cyclosporin A Treatment
Authors: Huby Russell D.J. , Hicks Ray , Goff Lindsey K.
Published: June 2008

Cyclosporin A (CsA) inhibits the development of mature thymocytes from their
CD4+ CD8+ precursors, but may allow autoreactive cells to mature. Using 3‐color flow
cytometry, we have followed the progressive development of thymocytes, including
potentially autoreactive cells, during CsA treatment. Numbers of CD4+ CD8+ CD3high
thymocytes dropped immediately, suggesting that the generation of these mature thymocyte
precursors, normally dependent upon positive selection, was inhibited by CsA.
Numbers of CD4- CD8+ thymocytes also declined rapidly, but CD4- CD8+ thymocytes
were unaffected lfor 2 days, suggesting that the mature single‐positive subsets are not
symmetrically derived from a common GsA‐sensitive precursor. An exceptional subset of
CD8 SP thymocytes, expressing CD45RA, did not respond to CsA for about 10 days,
indicating that they are distantly derived from a CsA‐sensitive precursor. Apoptosis of
TCR‐Vβ3+ thymocytes caused by Mtυ‐6, quantified according to the down‐regulation of
CD4 and CD8 on immature thymocytes, was partially inhibited by CsA, to maximal effect
within 24 hours. This did not, however, facilitate their development into mature thymocytes.
In Vivo and In Vitro Expression of Tenascin by HumanThymic Microenvironmental Cells
Authors: Freitas Claudia Sondermann , Patricia O′Campo Lyra Jurandy Susana , Dalmau Sergio Ranto , Savino Wilson
Published: June 2008

Increasing evidence reveals that extracellular matrix components can be regarded as a group
of mediators in intrathymic T‐cell migration and/or differentiation. Yet, little is kown about
the expression and putative function of one particular extracellular matrix protein, namely,
tenascin in the thymus. Herein we investigated, by means of immunocytochemistry,
tenascin expression in normal infant and fetal human thymuses, as well as in cultures of
thymic microenvironmental cells.

In situ, tenascin distribution is restricted to the medulla and cortico‐medullary regions of
normal thymuses. This pattern thus differed from that of fibronectin, laminin and type IV
collagen, in which subseptal basement membranes were strongly labeled. Interestingly,
tenascin did not co‐localize with the cytokeratin‐defined thymic epithelial cell network. This
was in keeping with the in vitro data showing that tenascin‐bearing cells were nonepithelial
(and probably nonfibroblastic) microenvironmental elements.

Studies with fetal thymuses revealed a developmentally regulated expression of tenascin,
with a faint but consistent network labeling, in thymic rudiments as early as 12 weeks of
gestational age, that progressed to a strong TN expression at 18 weeks of fetal development,
which was similar to the distribution pattern observed thereafter, including postnatally.

Our results clearly indicated that tenascin is constitutively expressed in the human
thymus, since early stages of thymic ontogeny, and suggest that the cell type responsible for
its secretion is a nonepithelial microenvironmental cell.
Hemopoiesis in the Thymus
Authors: Kendall Marion D.
Published: June 2008

The presence in the thymus of hemopoietic cells other than thymocytes has been known for
many years, but the extent of the hemopoietic activity of the thymus and the possible
functional implications have only recently begun to receive much attention. This review
summarizes the literature in this field, especially in the light of current cytokine and
thymic‐factor knowledge, and includes clinical relevance where possible.

Corrigendum to “Hemopoiesis in the Thymus” dx.doi.org/10.1155/1995/12606
γ/δ Cells in Fetal, Neonatal, and Adult RatLymphoid Organs
Authors: Kühnlein P. , Vicente A. , Varas A. , Hünig T. , Zapata A.
Published: June 2008

In the present study, we have analyzed the appearance and maturation of γ/δ T cells,
recognized with a new mAb V65, in the central and peripheral lymphoid organs of fetal,
neonatal, and adult Wistar rats. Cytofluorometrical analysis demonstrated the first V65+γ/δ T cells in the thymus of 16‐17‐day embryonic rats, although by immunohistology, they
were identified only in 19‐day rat embryos in both the cortico‐medullary border and thymic
medulla. Phenotypically, γ/δ thymocytes from fetal and neonatal thymus expressed CD3,
CD2, and CD5, but only 60‐80% were CD8+ and approximately 40‐50% expressed the α
chain (p55) of the IL‐2R. In the periphery, the immunohistological study identified for the
first time ,γ/δ T cells in the splenic white pulp and the gut of 21‐day fetal rats, where they
occurred within the epithelium as well as in the lamina propria. After birth, γ/δ
lymphocytes appeared in the skin, where they were present as dendritic epidermal T cells
in increasing numbers during postnatal life. Whereas these γ/δ T cells formed the
predominant T‐cell population in the rat skin, γ/δ T cells in peripheral lymphoid organs,
BALT, or the gut only represented a minor T‐cell population. These results are discussed in
comparison to γ/δ T cells of other vertebrate species.
Frequency of T‐Cell Progenitors in Nude Mice
Authors: Crispe I. Nicholas
Published: June 2008

The hypothesis that prothymocytes are distinct from and regulated independently of
multilineage hemopoietic progenitors was tested by enumeration of these two cell populations
in normal versus congenitally athymic (nude) mice. The absence of a thymus and of
peripheral T cells in nude mice had no effect on the frequency of either multilineage
progenitors (day 12 CFU‐S) or prothymocytes (CFU‐T), suggesting that there is no feedback
regulation of CFU‐T frequency. Thymus seeding from the bone marrow is therefore likely
to be regulated by the availability of niches for prothymocyte maturation, rather than by
feedback control of prothymocyte production.
The Programmed Cell Death of an Immature ThymocyteCell Line Transgenic for an αβ TCR and the c‐mycProto‐Oncogene
Authors: Murdjeva Marianna , Tanaka Yujiro , Norton Trisha , Kioussis Dimitris
Published: June 2008

The c‐myc proto‐oncogene linked to the mouse Thy‐1 gene transcriptional unit predisposes
mice to development of thymic tumors consisting predominantly of immature CD4+ CD8+
cells. In an attempt to immortalize immature T cells expressing a known T‐cell antigen
receptor (TCR), Thy‐1/c‐myc transgenic mice were bred to αβ TCR transgenic mice (F5),
and CD4+ CD8+ cell lines were established from thymic tumors in double‐transgenic mice.
These cells expressed high‐level heat‐stable antigen (HSA) and were able to undergo
programmed cell death upon induction with steroids and CD3 cross‐linking, but not with
cognate peptide. In addition, one line had rearranged and transcribed endogenous TCR c
and genes, in spite of the fact that transgenic α and β genes were also expressed.
Furthermore, we show that Thy‐1/myc transgenic mice deficient in recombination activating
gene‐1 (RAG‐1) do not develop tumors, in contrast to RAG‐1-/- mice, which are also
transgenic for both Thy‐1/myc and the F5 TCR. This indicates that in order for thymocytes
to be transformed by the Thy‐myc transgene, they need to proceed to the double‐positive
stage.
Response of Leucocyte Populations in the Ileal Peyer′sPatch of Fetal Lambs Treated with Ferritin Per Os
Authors: Renström Lena H. M. , Press Charles Mcl. , Trevella Wendy , Landsverk Thor
Published: June 2008

A combination of immunohistochemical techniques, a panel of monoclonal antibodies, and
computer‐assisted morphometric analysis was used to examine the response of the ileal
Peyerr′s patch of fetal lambs 7 days after treatment with ferritin per os. Consistent with
previous studies in fetal lambs that have reported the ileal Peyer′s patch to be indifferent
to antigen, the present study did not find any significant changes in the size of the
predominantly B‐cell dome/follicle compartment or the predominantly T‐cell interfollicular
area, nor were differences identified in the distribution of IgM‐positive (+), CD4 +, and
CD8+ cells in these two compartments. However, both compartments showed a significant
increase (P < 0.05) in the percentage of area occupied by MHC II + cells and a significant
decrease (P < 0.05) in the percentage of area occupied by CD44+ and B5+ cells. These
changes show that the ileal Peyer’s patch of fetal lambs is not indifferent to antigen and may
represent the transition of a purely primary lymphoid organ to an organ that has both
primary and secondary lymphoid functions.
In Vitro and In Situ Characterization of Fish Thymic Nurse Cells
Authors: Flaño Emilio , Álvarez Francisco , López-Fierro Pilar , Razquin Blanca E. , Villena Alberto J. , Zapata Agustín G.
Published: June 2008

We present an enzyme‐ and immuno‐cytochemical, and ultrastructural characterization of
trout thymic nurse cells (TNCs). Our data suggest that isolated trout thymic multicellular
complexes are epithelial cells with acidic compartments that may be involved in the processing
of antigens and in the generation of the MHC‐II proteins that these cell express, and
also that isolated TNCs are the In Vitro equivalent of the pale and intermediate electronlucent
epithelial cells located in the inner zone of the trout thymus, constituting indirect
evidence of the phylogenetical relationships of the inner zone of the teleost thymus with
the thymic cortex of higher vertebrates.
A Novel Molecular Complex Expressed on Immature B Cells: A Possible Role in T Cell‐IndependentB Cell Development
Authors: Griebel Philip J. , Ghia Paolo , Grawunder Ulf , Ferrari Giorgio
Published: June 2008

To identify surface molecules that may play a role in regulating ileal Peyer′s patch (PP) B
cell growth, we generated monoclonal antibodies (mAbs) and then selected them for a
unique reactivity with ileal PP B cells. Flow cytometric analysis identified a mAb (SIC4.8R)
that labeled 97% of ileal and 50–60% of jejunal PP sIgM+B cells. SIC4.8R also labeled a
subpopulation of cortical thymocytes but few B or T cells in other lymphoid tissues, including
bone marrow. Immunohistochemistry revealed intense SIC4.8R staining of B cells
in the cortex of ileal PP follicles. SIC4.8R also labeled bovine PP B cells, a murine pro‐B
cell line, and pre‐B cells in human bone marrow. Protein chemistry revealed that a structurally
similar molecular complex was expressed on sheep ileal PP B cells and thymocytes
and murine pro‐B cells. Addition of soluble SIC4.8R to cultured ileal PP B cells reduced
apoptotic cell death, elevated proliferative responses, partially inhibited anti‐Ig‐induced
cell death, and induced IL‐4 responsiveness. In contrast, soluble SIC4.8R had an antiproliferative
effect on a mouse pro‐B cell line. Finally, SIC4.8R labeling declined following
the stimulation of ileal PP B cells with CD40 ligand. In conclusion, the present investigation
determined that SIC4.8R identified a novel molecular complex that is expressed at
several stages of T cell‐independent B cell development in a variety of mammalian species.
This observation confirmed that PP B cells are developmentally distinct from other
B cell populations in sheep and suggested that the bone marrow may not be a site of B
lymphopoiesis in young lambs.
Thymic Medulla Epithelial Cells Acquire Specific Markers by Post‐Mitotic Maturation
Authors: Penit Claude , Lucas Bruno , Vasseur Florence , Rieker Theresa , Boyd Richard L.
Published: June 2008

The development of thymocyte subsets and of the thymic epithelium in SCID and RAG‐2-/– mice was monitored after normal bone‐marrow‐cell transfer. The kinetics of thymic
reconstitution and their relationships with cell proliferation were investigated by using
bromodeoxyuridine to detect DNA‐synthesizing cells among lymphoid cells by 3‐color
flow cytometry, and in epithelial compartments by staining frozen sections. Thymocytes
started to express CD8 and CD4 10 days after transfer, simultaneously with extensive proliferation.
The first mature CD4+ single‐positive cells were generated, from resting CD4+CD8+
cells after day 15. During this day 10–15 period, many epithelial cells positive for cortexspecific
or panepithelial markers were labeled with BrdUrd after pulse‐injection. Organized
medullary epithelium also developed after day,15, that is, synchronously with the
appearance of mature thymocytes, but medullary cells were never found BrdUrd+. These
results suggest that, in these models, the reconstitution of the thymic epithelial network
proceeds through expansion of preexisting cortical or undifferentiated cells and by later
maturation (acquisition of specific markers) of medullary cells. This last process is dependent
of the presence of mature thymocytes.
Protein Expression During Murine Thymus Differentiation
Authors: Frey Johann Rudolf , Hartmann Klaus-Ulrich , Lefkovits Ivan
Published: June 2008

Driven by our long‐standing interest in identifying proteins of the immune system and in
characterizing processes involved in lymphocyte differentiation, we studied protein expression
in biosynthetically labeled fetal and newborn thymus by 2D gel electrophoresis.
Autoradiographs of the gels were scanned with a densitometer and image analysis was
performed using the Kepler system. Calibrated polypeptide spot abundances (volumes)
were compared to assesses qualitative and quantitative changes of the spot volumes. Among
over 300 proteins evaluated at GD (gestation day) 13,15, and 17, there were sets of proteins
that increased and others that decreased in intensity. We could in addition recognize proteins
that were completely absent at GD 13 and/or 15 and that appeared thereafter to
gradually increase in intensity. Conversely, various polypeptide spots present at early stages
(at GD 13 and 15) disappear later (at GD 17 or at birth). Among the proteins that increase
in intensity prevail molecules with masses less than 35 kD, whereas a considerable portion
of those that decrease in intensity are characterized by masses above 60 kD. Spots reported
in this communication were not defined beyond tagging them with numbers, which is a
prerequisite to follow them up in the proteinpaedia developed in our laboratory. The next
step will be to retrieve the coding sequences from the existing partitioned cDNA library
(BW 5147) as well as from thymocyte subtraction libraries. We predict that among those
polypeptides with varying intensity, important regulatory proteins in thymus development
will be found.
Requirements for Differentiation of an Immature CD4+8+ T‐Cell Line
Authors: Dekoning Jenefer , Kaye Jonathan G.
Published: June 2008

The CD3ɛ and ζ chains of the TCR have been shown to possess independent signaling capabilities.
Studies with chimeric molecules containing the cytoplasmic domains of either ζ or ɛ have
suggested that these two structurally distinct members of the TCR‐CD3 complex are able to
function autonomously and have redundant features in the context of TCR‐signal transduction
in mature T cells. Expression of a chimeric human IL‐2‐receptor‐ζ‐chain molecule in the
CD4+8+ T‐cell line, DPK, has enabled us to directly analyze responses initiated by the ζ‐chain‐signaling
module alone within the context of immature T‐cell differentiation. In this paper, we
show that antibody crosslinking of the chimeric ζ chain delivers only a limited activation signal
as measured by Ca[2+] flux, induction of low‐level CD5 expression, and minimal differentiation
as assessed by loss of cell‐surface CD8 expression. TCR‐induced activation through antibody
crosslinking of the endogenous CD3ɛ receptor in the absence of costimulation was also
relatively inefficient in initiating activation and differentiation. However, co‐crosslinking of the
CD4 coreceptor with CD3 resulted in a synergistic response, where as there was little effect of
co‐crosslinking of CD4 and the ζ‐chain chimera. Striking differences were also observed in the
substrate pattern of tyrosine phosphorylation, as well as lymphokine secretion following triggering
through the intact TCR versus the ζ chain alone. These results indicate that although the
ζ‐chain may possess some signaling capacities similar to that of the intact TCR, it appears to
have limited function as an autonomous subunit in initiating CD4+8+ T‐cell differentiation.
The Neonatal Development of Intraepithelial and LaminaPropria Lymphocytes in the Murine Small Intestine
Authors: Steege Jessica C. A. Ter , Buurman Wim A. , Forget Pierre-Philippe
Published: June 2008

During early neonatal life, important changes occur in the gut. The intestine is challenged by
both milk and a microbial flora. Later on, at weaning, the diet of mice changes from milk to
pelleted food leading to changes in microbial contents. This period seems essential for a complete
development of the mucosal immune system. We investigated the development of both
intraepithelial (IEL) and lamina propria lymphocytes (LPL), from day 5, and every 5 days, up
to day 30 after birth. IEL and LPL were isolated from the small intestine and the phenotype
was assessed by FACS analyses, using antibodies for detection of T‐cell markers CD3,
TCRαβ, TCRγδ, CD4, CD8α, CD8β, CD5, CD18, CD54, and CD49d. Our data show a clear
increase in the number of LPL just before weaning, while the number of IEL increased after
day 15. A more mature pattern of membrane antigen expression of both IEL and LPL was observed
at weaning. The adhesion molecules CD18, CD54, and CD49d, essential for cellular
communication of lymphocytes, showed an expression peak at weaning. In conclusion, the
mouse mucosal immune system develops during the first 3 weeks of neonatal life leading to
the formation of a more mature immune system at weaning.
Involvement of Thyroid Hormones in the Expression of MHC class I Antigens During Ontogeny in Xenopus
Authors: Rollins-Smith Louise A. , Flajnik Martin F. , Blair Patrick J. , Davis A. Tray , Green Wayne F.
Published: June 2008

The major histocompatibility complex (MHC) is a cluster of genes encoding products central
to all major functions of the vertebrate immune system. Evidence for an MHC can be found
in all vertebrate groups that have been examined except the jawless fishes. Expression of
MHC class I and class II antigens early in ontogeny is critically important for development
of T lymphocytes capable of discriminating self from nonself. Because of this essential role
in T‐cell development, the ontogeny of MHC expression in the South African clawed frog,
Xenopus laevis, was studied. Previous studies of MHC class I expression in Xenopus laevis suggested that class I antigens are virtually absent from tadpole tissues until climax of metamorphosis.
We therefore examined the possible role of thyroid hormones (TH) in the induction
of class I. By flow cytometry, a small amount of class I expression was detectable on
splenocytes and erythrocytes in untreated frogs at prometamorphic stages 55‐58, and the
amount increased significantly at the conclusion of metamorphic climax. Thus, metamorphosis
is associated with increased intensity of class I expression. Neither inhibition nor acceleration
of metamorphosis altered the timing of onset of class I expression. However, inhibition
of metamorphosis prevented the increase in class I expression characteristic of adult
cell populations. Because expression was not accelerated in TH‐treated frogs or delayed in
metamorphosis‐inhibited frogs, it is unlikely that TH are the direct developmental cues that
induce expression, although they seem to be required for the upregulation of class I expression
occurring at metamorphosis. Differences in the pattern of expression in different subpopulations
of cells suggest a complex pattern of regulation of expression of class I antigens
during ontogeny.
A Comparative Analysis of the Murine Thymic Microenvironment in Normal, Autoimmune, and Immunodeficiency States
Authors: Takeoka Yuichi , Chen Shao-Yuan , Boyd Richard L. , Tsuneyama Koichi , Taguchi Nobuhisa , Morita Shinji , Yago Hisashi , Suehiro Seishi , Ansari Aftab A. , Shultz Leonard D. , Gershwin M. Eric
Published: June 2008

It is widely accepted that the thymic microenvironment regulates normal thymopoiesis
through a highly coordinated and complex series of cellular and cytokine interactions. A direct
corollary of this is that abnormalities within the microenvironment could be of etiologic
significance in T‐cell‐based diseases. Our laboratory has developed a large panel of
monoclonal antibodies (mAbs) that react specifically with epithelial or nonepithelial
markers in the thymus. We have taken advantage of these reagents to characterize the
thymic microenvironment of several genetic strains of mice, including BALB/cJ,
C57BL/6J, NZB/BlnJ, SM/J, NOD/Ltz, NOD/Ltz‐scid/sz, C57BL/6J‐Hcphme/Hcphme, and
ALY/NscJcl‐aly/aly mice, and littermate control animals. We report herein that control
mice, including strains of several backgrounds, have a very consistent phenotypic profile
with this panel of monoclonal antibodies, including reactivity with thymic epithelial cells
in the cortex, the medulla and the corticomedullary junction, and the extracellular matrix.
In contrast, the disease‐prone strains studied have unique, abnormal staining of thymic cortex
and medulla at both the structural and cellular levels. These phenotypic data suggest
that abnormalities in interactions between developing thymocytes and stromal cells characterize
disease‐prone mice.
Both IFN‐γ and IL‐4 Induce MHC Class II Expression at the Surface of Mouse Pro‐B Cells
Authors: Lombard-Platet Suzanne , Meyer Valerie , Ceredig Rhodri
Published: June 2008

Pro‐B cells are early B‐cell progenitors that retain macrophage potential. We have studied
MHC class II molecules and invariant chain inducibility on four class II negative mouse pro‐
B‐cell clones. We analyzed the effects of IL‐4 and IFN‐γ, which represent the major inducers
of class II in the B‐lymphoid and monocytic/macrophage lineages, respectively. After 48 h of
treatment with either cytokine, three pro‐B‐cell clones (C2.13, A1.5, and F2.2) expressed intracellular
invariant chain and cell‐surface class II molecules. One clone (D2.1) remained negative.
As already reported, more differentiated 70Z/3 pre‐B cells were inducible by IL‐4 only.
These data suggest that the induction of class II and invariant‐chain genes are subject to regulation
throughout B‐cell differentiation.
Expression and Temperature‐Dependent Regulation of the Beta2‐Microglobulin (Cyca-B2m) Gene in a Cold‐Blooded Vertebrate, the Common Carp (Cyprinus carpio L.)
Authors: Rodrigues P. N. S. , Dixon B. , Roelofs J. , Rombout J. H. M. W. , Egberts E. , Pohajdak B. , Stet R. J. M.
Published: June 2008

Expression of beta2‐microglobulin (β2m) in the common carp was studied using a polyclonal antibody raised against a recombinant protein obtained from eukaryotic expression of the Cyca-B2m gene. β2m is expressed on peripheral blood Ig+ and Ig- lymphocytes, but not on
erythrocytes and thrombocytes. In spleen and pronephros, dull‐ and bright‐positive populations
could be identified correlating with the presence of erythrocytes, thrombocytes, and mature
leucocytes or immature and mature cells from the lympho‐myeloid lineage, respectively.
Thymocytes were shown to be comprised of a single bright‐positive population. The Cyca‐B2m
polyclonal antiserum was used in conjunction with a similarly produced polyclonal antiserum to
an MHC class I (Cyca-UA) α chain to investigate the expression of class I molecules on
peripheral blood leucocytes (PBL) at different permissive temperatures. At 12℃, a temporary
downregulation of class I molecules was demonstrated, which recovered to normal levels within
3 days. However, at 6℃, a lasting absence of class I cell‐surface expression was observed, which
could be restored slowly by transfer to 12C. The expression of immunoglobulin molecules on
B cells was unaffected by temperature changes. The absence of the class cell‐surface expression
was shown to be the result of a lack of sufficient Cyca-B2m gene transcription, although Cyca-UA mRNA was present at comparable levels at all temperatures. This suggests that class I
expression is regulated by a temperature‐sensitive transcription of the Cyca-B2m gene.
CD40 Ligand and Autoantigen Are Involved in the Pathogenesis of Low‐Grade B‐Cell Lymphomas of Mucosa‐Associated Lymphoid Tissue
Authors: Greiner A. , Knörr C. , Qin Y. , Schultz A. , Marx A. , Kroczek R. A. , Müller-Hermelink H. K.
Published: June 2008

Low‐grade MALT‐type lymphomas are malignancies of mucosal marginal‐zone B cells
and preceded by reactive inflammatory lymphoid tissue. Experimental observations suggest that antigen
and CD40 Ligand act during cognate T/B cell interaction and are crucial for germinal center B‐cell maturation
generating marginal‐zone B cells. To investigate the mechanisms underlying the development of extranodal MALT‐type lymphomas, the immunoglobulin receptor was sequenced and analyzed for antigen specificity using heterohybridoma technology.
Furthermore, CD40 ligand expression was evaluated by immunohistochemistry and by semiquantitative RT‐PCR,
and ligand binding to the CD40 of tumor B cells was studied using the CD40 system. Hypermutations were found in low‐grade
lymphomas throughout CDR1‐ CDR3 suggestive of positive selection through their antigen receptor.
Different VH families were used and more than 69% of tumor immunoglobulins bound different mucosal antigens.
CD40L expression was found in the tumor marginal zone in substantial amounts.
The in vitro proliferation response of all low‐grade MALT‐type lymphomas was dependent on
anti‐CD40‐ mediated signals and cytokines. Our data provide evidence that autoantigen as well as the CD40L
expressed by activated nonneoplastic T cells may drive the evolution of low‐grade MALT‐type lymphomas either
directly or by paracrine mechanisms and that antigen may contribute to lymphoma pathogenesis.