bluuebunny/crossref_metadata_2025 huggingface dataset:
Transforming Growth Factor Beta (TGFβ) Is Produced by and Influences the Proliferative Response of Xenopus laevis Lymphocytes
Authors: Haynes Laura , Cohen Nicholas
Published: June 2008

Both TGF/β2 and 5 have been described in the South African clawed frog Xenopus laevis
and have been cloned from the tadpole‐derived fibroblast cell line, XTC. Because TGFβ
has such a profound inhibitory effect on the mammalian immune system, this study was
performed to determine whether TGFβ: (a) has any in vitro effects on the growth of
Xenopus lymphoblasts, and (b) is produced by mitogen‐activated Xenopus lymphocytes.

Following stimulation with mitogen or alloantigen, T lymphocytes from Xenopus
secrete a T‐cell growth factor (TCGF) that is functionally homologous to mammalian
interleukin‐2 (IL‐2). Both recombinant human TGFβ1 and Xenopus TGFβ5 inhibit TCGF‐induced
proliferation of Xenopus splenic blasts and this inhibition can be reversed with
anti‐pan TGFβ antiserum. The Xenopus mitogen‐induced saturated ammonium sulfate
precipitated TCGF‐containing supernatant (SAS TCGF SN) also contains latent TGFβ as
assayed on mink lung fibroblasts and Xenopus splenic blasts, and experiments utilizing
anti‐TGFβ antiserum showed that only TGFβ5 is present in this supernatant.
Further Characterization of an Interleukin‐2‐1Ike Cytokine Produced by Xenopus Laevis T Lymphocytes
Authors: Haynes Laura , Cohen Nicholas
Published: June 2008

A T‐cell growth factor (TCGF) is produced by antigen‐ or mitogen‐stimulated T
lymphocytes from the South African clawed frog Xenopus laevis. This study further
defines the physical and biological properties of this cytokine and demonstrates that
TCGF is biochemically similar to mammalian interleukin‐2 (IL‐2). Biologically active
TCGF eluted from SDS‐PAGE displays a Mr of 16 kD and lectin‐affinity chromatography
indicates that the three‐dimensionmal configuration of carbohydrates on TCGF and
human IL‐2 is similar. Secretion of TCGF is detectable 1 day after stimulation of
splenocytes with the T‐cell mitogen phytohemagglutinin (PHA) and peaks following 2
to 3 days of stimulation. Finally, despite the biological and physical similarities between
Xenopus TCGF and mammalian IL‐2, anti‐human IL‐2 monoclonal antibodies do not
recognize Xenopus TCGF.
Developmental and Anatomical Patterns Of IL‐2 Gene Expression in Vivo in The Murine Thymus
Authors: Yang-Snyder Julia A. , Rothenberg Ellen V.
Published: June 2008

Interleukin‐2 (IL‐2) is a potent growth factor that mature T lymphocytes synthesize and
use as a proliferation signal. Much controversy has arisen concerning whether it is used
to drive the extensive proliferation of immature pre‐T cells in the thymus. Immature
thymocytes acquire the competence to express IL‐2 at an early stage, but it has remained
uncertain whether they are activated to exercise this competence in vivo. Therefore, we
have used in situ hybridization and immunohistochemistry on serial sections obtained
from fetal and adult thymuses of normal C57BL/6 mice and of mice bearing the scid
defect to determine where, when, and whether IL‐2 is expressed in vivo. Our results
show a striking spatial and temporal pattern of IL‐2 expression in the normal fetal
thymus. We detected a burst of IL‐2 mRNA accumulation at day 14.5 of gestation, which
rapidly decreased by day 15. At day 15, we observed maximal IL‐2 protein production
that subsequently decreased by day 16 of gestation. Both in situ hybridization and
immunohistochemical staining revealed an unexpectedly strict localization of IL‐2
expressing cells to patches around the periphery of the fetal thymus, creating a
previously unrecognized compartment of high IL‐2 protein content. IL‐2 production in
the day‐15 fetal thymus appeared to be unaffected by the scid mutation, indicating that
this response is likely to be T‐cell receptor (TcR)‐independent. Several features
distinguish the IL‐2 induction pattern in the adult thymus from that in the fetal thymus.
In the normal adult thymus, IL‐2‐expressing cells are extremely rare (found at a
frequency of 10-7), but they are reproducibly detectable as isolated cells in the outer
cortex and subcapsular region of the thymus. Unlike the fetal thymic IL‐2 producers, the
IL‐2 producers in the adult thymus are completely eliminated in mice homozygous for
the scid mutation. This suggests that the IL‐2‐expressing cells in the normal adult
thymus are of a more mature phenotype than the immature, TcR‐negative cells that
accumulate in the scid adult thymus. Thus, our work demonstrates that two
developmentally distinct types of cell interactions induce IL‐2 expression in vivo: one, a
broadly localized interaction in day 14‐15 fetal thymus that is unaffected by the scid
mutation; the other, a rare event that occurs asynchronously from late fetal through
adult life, but which is completely eliminated by the scid defect. These results imply that
significant differences exist between the physiological processing of thymocytes in the
fetal and postnatal thymic microenvironments.
Modulation of Cytokeratin Expression in The Hamster Thymus: Evidence For a Plasticity of The Thymic Epithelium
Authors: De Souza Lucia Renata Meireles , Savino Wilson
Published: June 2008

Cytokeratin (CK) expression was investigated, by means of immunocytochemistry, in
the hamster thymic epithelium during ontogeny, as well as in primary cultures and
upon glucocorticoid hormone treatment in vivo. As compared to the distribution pattern
of distinct monoclonal antibody‐defined cytokeratins in the normal adult thymus, CK
modulation was evidenced in the three situations studied. During thymus ontogeny,
both cytokeratins of simple lining epithelia, as CK8 and CK18, as well as the CK1/CK10
pair (typical marker of terminal stage of keratinization), were expressed since early
stages of thymus development. They were located in the central region of thymic lobules
preceding the cortical‐medullary distinctions. This differed from what had been
previously shown for mouse thymus ontogeny, revealing that the interspecific diversity
in the distribution pattern of thymic cytokeratins occurred early in fetal life. A
modulation of CK expression was also detected when hamster thymic epithelial cells
(TEC) were led to grow in culture, with a down‐regulation of CK19 contrasting with an
enhancement of CK18 expression. This diverged from the maintenance of the in situ
pattern when human TEC were cultured. Last, in vivo hydrocortisone treatment, known
to increase the numbers of KL1+ cells in the mouse thymus medulla, promoted a cortical
expression of the CK1/CK10 pair in the hamster thymus. Taken together, our findings
demonstrate a continuous plasticity of the thymic epithelium, at least regarding
cytokeratin expression, and enlarge the concept of interspecific diversity of intrathymic
CK distribution in conditions as morphogenesis, in vitro system, and responsiveness to
glucocorticoid hormone treatment.
Differential Expression of Adhesion Molecules Within theHuman Thymus
Authors: Reza Julie Naima , Ritter Mary A.
Published: June 2008

Development of a diverse, MHC‐restricted yet self‐tolerant T‐cell repertoire occurs within
the thymus, and requires contact between developing T cells and their stromal microenvironment.
Such interactions are likely to depend on the combinatorial effect of specific
adhesion molecules. As a preliminary step to determining their role in T‐cell development,
we have studied the distribution of LFA‐1/ICAM‐1, CD2/LFA‐3, VLA‐4/VCAM‐1, and
HECA 452‐antigen/E‐Selectin ligand pairs on frozen sections of human thymus. Using two
color‐immunohistochemistry, and a variety of cell‐lineage markers that reveal the nature of
the cells on which these adhesion molecules are located, we find a differential distribution
of adhesion molecules, with some being shared by both endothelial and epithelial cells. We
also identify the VCAM‐1‐positive subpopulation as cortical macrophages. The relevance of
these findings to thymopoiesis is discussed.
Positive Selection by Purified MHC Class II / Thymic Epithelial Cells In Vitro: Costimulatory Signals Mediated by B7 Are Not Involved
Authors: Jenkinson Eric J. , Anderson Graham , Moore Nel C. , Smith Christopher A. , Owen John J. T.
Published: June 2008

We have investigated the possibility that the costimulatory signals required for activation of
mature T cells also play a role in providing differentiation signals for positive selection
during T‐cell development. We show that purified MHC Class II+ thymic epithelial cells are
able to support positive selection in vitro but lack both the functional capacity to deliver
costimulatory signals and expression of the costimulatory ligand B7. Our results suggest that
the additional signals provided by costimulatory ligands are not required for TCR‐mediated
positive selection, although other ancillary signals provided by thymic epithelial cells may
be involved.
Hemopoietic and Lymphoid Progenitor Cells in HumanUmbilical Cord Blood
Authors: Cicuttini F. M. , Boyd A. W.
Published: June 2008

Human umbilical cord blood, which in the past was discarded with the placental tissue,
provides a convenient source of fetal hemopoietic cells for scientific analysis and clinical use.
Cord blood cells are immature compared to analogous populations in adult peripheral
blood. Cord blood B lymphocytes display unique phenotypic and functional characteristics.
The antigens CD1C, CD38, CD5, and CD23, although normally expressed on only a small
percentage of circulating B cells in adults, are highly expressed on cord blood B cells. Recent
studies have demonstrated that whereas cord blood B cells are functionally naive, their
potential is similar to that of adult B cells if optimal T‐cell help is available. Thus, the failure
of B‐cell responses in cord blood is due to the T cells. The functional abnormalities of T cells
from newborns can be summarized as a dominance of the effects of TH0 cells. Thus, the
cytokines produced are immunosuppressive rather than mediating helper activity for B cells.
NK activity in cord blood is also depressed compared to that in adults. Cord blood is a very
rich source of hemopoietic progenitor cells. The spectrum of progenitors shows a predominance
of early progenitor cells when compared with bone marrow. These cells provide an
alternative source to adult bone marrow for stem cells to use for hemopoietic reconstitution
and as targets in the treatment of hereditary deficiencies by gene therapy. These features
make cord blood a unique research tool to investigate hemopoietic ontogeny and a unique
clinical tool for transplantation.
Expression of Functional MHC Class II Molecules By aMouse Pro‐B Cell Clone
Authors: Lombard-Platet Suzanne , Fisher Amanda G. , Meyer Valérie , Ceredig Rhodri
Published: June 2008

We describe here the G12 pro‐B cell clone that has been isolated from an IL‐7 transgenic
mouse. This clone has the phenotype B220+, BP‐1+ , HSA +, CD43+ λ5+ , and CD25-, and
has its Ig locus in a germline configuration. G12 cells spontaneously express cell‐surface
MHC class II molecules, although to a much lesser extent than the mature M12.4.1 B‐cell
lymphoma. G12 cells can process and present the native Hen Egg Lysozyme (HEL) to an
MHC class II‐restricted T‐cell hybridoma. The efficiency of presentation is inferior to that
obtained with M12.4.1 cells. This is the first report where a pro‐B cell can serve as APC in
an MHC class II‐restricted presentation.
The γδ T Lymphocytes of the Perinatal Murine Thymus
Authors: Zorbas Michelle I. , Scollay Roland
Published: June 2008

We have previously shown that the adult thymus contains three subsets of γδ T cells that
can be defined by the expression of Thy‐1 and heat‐stable antigen (HSA). In this study, the
number of cells in each of these thymic γδ populations was investigated at different stages
throughout life. In adult mice, the populations stayed relatively constant, however, in
contrast, there were major variations in them early in development. It was shown that only
two of the γδ populations were present in the prenatal thymus, a major population of
Thy‐1+ HSA- cells, and a smaller population of Thy‐1+ HSA- cells. However, after birth,
most of the Thy‐1+ HSA-cells appear to loose the Thy‐1 antigen, forming the third
population of HSA- Thy‐1-cells. The adult configuration of populations appeared to be
established within the first week after birth. Therefore, whereas the γδ populations stayed
relatively constant from this time point onwards, there were major variations early in
development. Throughout life, most γδ thymocytes are CD4- CD8-, however, in the
neonatal thymus, there are some CD4+ and CD8+ γδ thymocytes, and these are contained
in the Thy‐1+ HSA- population.
Cross‐Linking the TCR Complex Induces Apoptosis in CD4+8+ Thymocytes in the Presence of Cyclosporin A
Authors: Poetschke Heather L. , Klug David B. , Walker Dawn , Richie Ellen R.
Published: June 2008

Although it is generally agreed that TCR ligation is a minimal requirement for negative
selection in the CD+8+ double‐positive (DP) thymocyte subset, the costimulatory requirements
and specific signaling events necessary to induce apoptosis are not well defined. We
have explored the consequences of cross‐linking CD3/TCR complexes on thymocytes from
H‐Y TCR transgenic (Tg) mice. In agreement with previous reports, we demonstrate that
culturing DP thymocytes with plate‐bound anti‐TCR antibody induces downregulation of
CD4 and CD8 and upregulation of CD69 expression. Nevertheless, the activated cells did
not undergo apoptosis, as determined by viable cell recoveries and by quantitation of DNA
fragmentation using the TUNEL assay. However, specific depletion of the DP subset occurred
within 24 hr when thymocytes were incubated in the presence of both anti‐TCR and
the immunosuppressant cyclosporin A (CsA). CsA also induced depletion of anti‐CD3
stimulated normal DP thymocytes. Using mice homozygous for the lpr or gld mutation, we
also have shown that Fas/Fas ligand interactions are not involved in the CsA‐induced
death of TCR‐stimulated DP thymocytes. These data verify that TCR cross‐linking alone
is insufficient to induce apoptosis of DP thymocytes and further suggest that TCR stimulation
activates a CsA‐sensitive protective pathway that interferes with signaling events
leading to apoptosis in DP thymocytes.
A Protein Binding Specifically to the IgG2b Switch Region
Authors: Völk Heiner , Wabl Matthias
Published: June 2008

The Abelson‐virus‐transformed mouse pre‐B‐cell line 18‐81 switches almost exclusively from
μ to γ2b. From nuclear extracts of this cell line, we have isolated a factor that specifically binds
to Sγ2b. After an eight‐step purification scheme, in which different types of DNA‐affinity chromatography
were used as key elements, we obtained a preparation with two narrowly spaced
bands at approximately 69 kD on a silver‐stained SDS gel. Binding specificity of main‐peak
fractions of affinity‐purified proteins was analyzed by gel shift assays in which Sγ2b, but not
Sμ, competes. The results are consistent with this factor being part of the switch recombinase.
Involvement of Glucocorticoids in the Reorganization ofthe Amphibian Immune System at Metamorphosis
Authors: Rollins-Smith Louise A. , Barker Katherine S. , Davis A. Tray
Published: June 2008

In recent years, integrative animal biologists and behavioral scientists have begun to understand
the complex interactions between the immune system and the neuroendocrine system.
Amphibian metamorphosis offers a unique opportunity to study dramatic hormone‐driven
changes in the immune system in a compressed time frame. In the South African clawed frog,
Xenopus laevis, the larval pattern of immunity is distinct from that of the adult, and metamorphosis
marks the transition from one pattern to the other. Climax of metamorphosis is
characterized by significant elevations in thyroid hormones, glucocorticoid hormones, and the
pituitary hormones, prolactin and growth hormone. Previously, we and others have shown that
elevated levels of unbound glucocorticoid hormones found at climax of metamorphosis are associated
with a natural decline in lymphocyte numbers, lymphocyte viability, and mitogen‐induced
proliferation. Here we present evidence that the mechanism for loss of lymphocytes at
metamorphosis is glucocorticoid‐induced apoptosis. Inhibition of lymphocyte function and
loss of lymphocytes in the thymus and spleen are reversible by in vitro or in vivo treatment
with the glucocorticoid receptor antagonist, RU486, whereas the mineralocorticoid receptor
antagonist, RU26752, is poorly effective. These observations support the hypothesis that loss
of larval lymphocytes and changes in lymphocyte function are due to elevated concentrations
of glucocorticoids that remove unnecessary lymphocytes to allow for development of immunological
tolerance to the new adult‐specific antigens that appear as a result of metamorphosis.
An Adult Thymic Stromal‐Cell Suspension Model for in Vitro Positive Selection
Authors: Chidgey Ann P. , Pircher Hanspeter , Macdonald H. Robson , Boyd Richard L.
Published: June 2008

Presented here is a cell‐suspension model for positive selection using thymocytes from αβ‐TCR
(H‐2Db‐restricted) transgenic mice specific to the lymphocytic choriomeningitis virus (LCMV)
on a nonselecting MHC background (H‐2d or TAP‐1 –/–), cocultured with freshly isolated adult
thymus stromal cells of the selecting MHC type. The thymic stromal cells alone induced positive
selection of functional CD4-CD8+ cells whose kinetics and efficiency were enhanced by
nominal peptide. Fibroblasts expressing the selecting MHC alone did not induce positive
selection; however, together with nonselecting stroma and nominal peptide, there was inefficient
positive. These results suggest multiple signaling in positive selection with selection events able
to occur on multiple‐cell types. The ease with which this model can be manipulated should
greatly facilitate the resolution of the mechanisms of positive selection in normal and
pathological states.
Spontaneous Expression of Interleukin‐2 In Vivo in Specific Tissues of Young Mice
Authors: Yang-Snyder Julia A. , Rothenberg Ellen V.
Published: June 2008

In situ hybridization and immunohistochemistry were used to determine the spectrum of tissues
in which interleukin‐2 (IL‐2) mRNA and protein are found in healthy, normal young mice. In
neonatal animals, IL‐2 is expressed specifically by distinct, isolated cells at three major sites: the
thymus, skin, and gut. Based on morphology and distribution, the IL‐2‐expressing cells resemble
CD3ε+ T cells that are also present in all these locations. Within the thymus of postweanling
animals, both TcRαβ and TcRγδ lineage cells secrete "haloes" of the cytokine that diffuse over many cell diameters. Within the skin, isolated cells expressing IL‐2 are seen at birth in the
mesenchyme, and large numbers of IL‐2‐expressing cells are localized around hair follicles in
the epidermis in 3‐week‐old animals. At this age, a substantial subset of CD3ε+ cells is similarly
localized in the skin. Significantly, by 5 weeks of age and later when the CD3ε+ cells are evenly
distributed throughout the epidermis, IL‐2 RNA and protein expression are no longer detectable.
Finally, within the intestine, IL‐2 protein is first detected in association with a few discrete,
isolated cells at day 16 of gestation and the number of IL‐2 reactive cells increases in frequency
through El9 and remains abundant in adult life. In postnatal animals, the frequency of IL‐
2‐positive cells in villi exceeds by greater than fivefold that found in mesenteric lymph node or
Peyer′s patches. Overall, these temporal and spatial patterns of expression provide insight into
the regulation of IL‐2 in vivo and suggest a role for IL‐2 expression distinct from immunological
responses to antigen.
L1 Makes Immunological Progress by Expanding Its Relations
Authors: Kadmon Guni , Montgomery Anthony M. P. , Altevogt Peter
Published: June 2008

The cell‐adhesion molecule L1 was originally described in the nervous system.
It has recently been detected in CD4+ T lymphocytes, peripheral B lymphocytes, and granulocytes in the human
immune system and in similar leucocyte types in the murine immune system. L mediates neural recognition by
Ca+2, Mg+2‐independent homophilic binding. In the human and murine immune systems, L1 binds to the “classical”
vitronectin receptor, αVβ3, and fibronectin receptor, α5β1, respectively,
and abstains from homophilic binding. Homophilic L1 binding probably involves antiparallel alignment of several interactive
domains. Integrin binding is mediated by a short segment of immunoglobulinlike domain 6, which includes two
RGD repeats in rodent L1 and one RGD motif in human L1. L1 is modulated in activated leucocytes
in vitro in parallel to L‐selectin, and diverse cell types release intact L in vivo and in vitro.
Released L1 can bind to laminin and adheres to the extracellular matrix of sciatic nerve, M21 melanoma,
and possibly spleen and other tissues. It can support integrin‐dependent cell migration and preliminary data
implicate it in tumor development and transnodal lymphocyte migration.
K21‐Antigen: A Molecule Shared by the Microenvironments of the Human Thymus and Germinal Centers
Authors: Imami Nesrina , Ladyman Heather M. , Vincents Bjarne , Al-Tubuly Abdulhamid , Freysdóttir Jona , Sedibane Moditi L. , Taylor-Fishwick David A. , Foxwell Brian M.J. , Ritter Mary A.
Published: June 2008

The mouse IgG1 monoclonal antibody (mAb) K21 recognizes a 230‐kD molecule (K21‐Ag) on Hassall′s corpuscles in the human thymus. This mAb also stains cultured thymic epithelial cells as well as other epithelial cell lines, revealing a predominant intracellular localization. Further analysis with mAb K21 on other lymphoid tissues showed that it also stains cells within the germinal centers of human tonsils, both lymphoid (B) cells and some with the appearance of follicular dendritic cells. Double immunostaining of tonsil sections shows that K21‐Ag is not expressed by T cells, whereas staining with anti‐CD22 and ‐CD23 mAb revealed some doublepositive cells. A subpopulation of the lymphoid cells express the K21‐Ag much more strongly. This K21++/CD23++ subpopulation of cells is localized in the apical light zone of germinal centers, suggesting that K21‐Ag may be an important marker for the selected centrocytes within germinal centers and may play a role in B‐cell selection and/or development of B‐cell memory. Flow cytometric analysis showed that K21‐Ag is expressed on the surface of a very low percentage of thymocytes, tonsillar lymphocytes, and peripheral blood mononuclear cells. Analysis of purified/separated tonsillar T and B lymphocytes showed that T cells do not express the K21‐Ag; in contrast, B cells express low levels of the K21‐Ag, and this together with CD23 is upregulated after mitogenic stimulation. Our data therefore raise the possibility that the K2l‐ Ag may play a role in B‐lymphocyte activation/selection.
Studies on the Induction of Antigen‐Specific Antibody in Anti‐CD40 Cultured Human B Lymphocytes
Authors: Schilizzi B. M. , Harmsen M. C. , The T. H. , Leij L. De
Published: June 2008

Costimulatory signals provided by T cells are required for B cells to produce specific antibody to T‐dependent antigen. We have investigated the suitability of using the CD40 culture system for the proliferation and differentiation of Ag‐specific human B cells using cytomegalovirus (CMV) or tetanus toxoid (TT) as antigen. We modified the CD40 culture system (CD32‐ transfected L cells, anti‐CD40, and IL‐4) by applying a sequential cytokine stimulation and compared total B‐cell cultures with antigen‐specific B cells preselected by panning. The detection of specific antibody became possible when antigen‐selected B cells were cultured for 7 days in the CD40 system to induce clonal expansion, followed by the addition of IL‐2 and IL10 for an additional 7 days to induce plasma‐cell differentiation. We conclude that our intial inability to detect specific antibody in the CD40 system is due to overgrowth of nonspecific B cell clones and that selection of antigen‐specific B cells by panning overcomes this problem. Induction of antigen‐specific antibody production was found to be optimal when the initial
contact with antigen during panning was limited to between 1 to 24 hours.
Paraneoplastic Autoimmunity in Thymus Tumors
Authors: Marx Alexander , Schultz Anja , Wilisch Annette , Helmreich Markus , Nenninger Regina , Müller-Hermelink Hans Konrad
Published: June 2008

Autoimmune phenomena are more frequent in thymic epithelial tumors (TET) than in any other human tumor. Mysthenia gravis (MG) is by far the most common autoimmune disease in thymoma patients. MG is characterized by muscle weakness due to autoantibodies against the acetylcholine receptor (AChR), and CD4 +AChR‐specific T cells play a pivotal role for the production of these autoantibodies. About 10% of MG patients have a thymoma and, interestingly, only such thymomas exhibit an MG association that maintains thymuslike
morphological and functional features with respect to the homing and differentiation of immature T cells. Since AChR protein is not expressed in thymomas, the specificity of the autoimmunity in thymoma‐associated MG is thought to be determined by nonreceptor proteins with AChR epitopes. Such proteins are overexpressed in cortical‐type MG‐associated thymomas, and medullary thymomas express these proteins at barely detectable levels. Aside from this quantitative difference, the pathogenesis of anti‐AChR autoimmunity might be qualitatively
different in these thymoma subtypes. Our findings suggest that an antigen‐specific abnormal Tcell selection by cortical‐type TET may contribute to the pathogenesis of paraneoplastic MG. In contrast, an abnormal (intratumorous) activation of autoreactive T cells may be operative in medullary thymomas.
Sequence Analysis of the Mouse RAG Locus lntergenic Region
Authors: Bertrand F. E. , Olsona S. L. , Martin D. A. , Wu G. E.
Published: June 2008

The recombination activating genes RAG‐1 and RAG‐2 are highly conserved throughout
evolution and are necessary and essential for the DNA rearrangement of antigen‐receptor gene
segments. These convergently transcribed genes are expressed primarily by developing B and T
lineage cells. In addition, recent data suggest that the RAG locus can be reactivated in mouse
germinal center B cells. Despite these well‐defined patterns of expression, little is known about
mechanism(s) regulating transcription of the RAG locus. Experiments with a mouse fibroblast
line stably transfected with a genomic fragment of the RAG locus suggest that the intergenic
region between RAG‐1 and RAG‐2 may contain information modulating RAG transcription. In
order to begin testing this hypothesis, we have sequenced the 7.0‐kb RAG intergenic region of
the mouse. The sequence did not contain open reading frames larger than 60 amino acids.
Analysis with GCG software identified several potential transcription‐factor binding sequences
within this region. Many of these are associated with transcriptional regulation of the Ig locus.
Antigenic Stimuli do not Influence Thymic B Lymphocytes: A Morphological and Functional Study inGerm‐Free and Conventionally Reared Piglets
Authors: Cukrowska B. , Trebichavský I. , Rossmann P. , Řeháková Z. , ŠInkora J. , Haverson K. , Lodinová-Žádníková R. , Tlaskalová-Hogenová H.
Published: June 2008

We have recently reported that thymic B lymphocytes (TBL) are the first B‐cell subpopulation
undergoing isotype switching to IgG and IgA during embryonic life. The aim of this study is to
analyze the influence of antigenic stimulation on TBL location and activity using a germ‐free
(GF) newborn pig model, in which maternal antibodies and antigens do not affect B‐cell
development. Immunohistological analysis showed that TBL were disseminated mainly in the
thymic medulla. There were no differences in the distribution of TBL, both in GF newborn
piglets before and after colonization with Escherichia coli and in older conventionally reared
(CONV) piglets. The number of immunoglobulin (Ig)‐secreting cells measured by the ELISPOT
method was not influenced by microflora and food antigens. IgM‐positive cells secreting IgM
and CD45RC‐positive cells spontaneously producing IgM, IgG, and IgA were detected in
newborn thymus.

Our findings suggest that TBL differentiation and Ig switching to IgG and IgA‐secreting cells is
not influenced by external antigens and that the thymic microenviroment plays an important role
in this process.
Studies on the Tissue‐Related Phenotypic Heterogeneity of Murine B Cells
Authors: Balogh Péter , Kumánovics Attila , Juhasz Istvan
Published: June 2008

The development of B cells is accompanied by their ability to specifically enter
the peripheral lymphoid tissues. Recently, we described a novel rat monoclonal antibody
(IBL‐2; IgG2b/k reacting with a 26/29‐kD heterodimeric structure of the cell surface.
This mAb has been found to recognize differentially the peripheral B cells of mice depending on their tissue origin.
The majority of splenic B cells as well as the mature B cells in the bone marrow were stained with this mAb,
whereas the B lymphocytes isolated from LN or Peyer′s patches displayed only negligible reactivity. We extended
these observations by analyzing the relationship between the
expression of IBL‐2 antigen and L‐selection on the surface of B‐cell precursors in the bone marrow by multiparameter flow cytometry.
Within the B220 positive compartment, a significant difference of L‐selectin expression could be observed between the various IBL‐2‐reactive subsets. Furthermore, we investigated whether evidences
for the establishment of tissue‐associated phenotypic heterogeneity similar to that found in normal mice could be found upon the adoptive transfer of normal unselected splenic lymphocytes into SCID recipients (Spl‐SCID).
It has been found that a large part of the splenic B cells preserved their IBL‐2 reactivity, whereas the LN B cells had lost the IBL‐2 antigen in Spl‐SCID. These data indicate that the phenotypic difference within the SCID mice may be the result of the migration of B lymphocytes from the spleen toward the lymph nodes, and the altered expression of the IBL‐2 antigen correlates with this process.
Apoptosis‐Specific Protein (ASP) Identified in Apoptotic Xenopus Thymus Tumor Cells
Authors: Horton John , Milner Anne , Horton Trudy , Ritchie Pamela , Gascoyne Duncan , Hewson Tim , Hammond Ester , Gregory Christopher , Grand Roger
Published: June 2008

A novel apoptosis‐specific protein (ASP) has recently been identified in the cytoplasm of
apoptotic mammalian cells. This paper investigates whether ASP is found in Xenopus thymus
tumor‐derived lymphoid cell lines undergoing apoptosis and also in apoptotic, nontransformed
splenocytes. Cultured Xenopus tumor lymphoid cells induced to undergo, apoptosis by serum
deprivation or treatment with the calcium ionophore, ionomycin, displayed altered morphology
typical of apoptotic cells, as judged by flow cytometric light‐scatter characteristics and by
fluorescence microscopy of acridine‐orange‐stained cells. Flow cytometry of permeabilized cells
and fluorescence microscopy of acetone‐fixed cytospins revealed that apoptotic Xenopus tumor
cells, especially those displaying loss or condensation of DNA, displayed increased expression
of epitopes recognized by a rabbit polyclonal antibody against ASP. Flow cytometry confirmed
that ASP is also expressed in splenocytes induced to apoptose by culture in ionomycin or
following concanavalin A stimulation. No increased expression of ASP was seen when lymphoid
tumor cells or splenocytes were induced into necrosis by overdose with the antifungal agent
amphotericin B. Western blotting with antibody against ASP identified the emergence of several
protein bands in cell lysates from apoptotic, but not necrotic, Xenopus tumor cells. The new and
simple methodology for identifying apoptotic cells described here is likely to be of value to those
studying immune system development and associated programmed cell death in Xenopus.
Glucocorticoids Induce a TH2 Response In Vitro
Authors: Ramírez Francisco
Published: June 2008

Purified rat CD4+ T cells were activated in vitro, by the polyclonal mitogen Concanavalin A (Con A) or by
mixed lymphocyte reaction (MLR), in the presence or absence of the glucocorticoid dexamethasone (DEX).
They were then expanded in IL‐2 and subsequently restimulated, this time in the absence of the hormone.
The results indicate that the exposure of the cells to DEX in the primary stimulation changed the cytokine synthesis
induced by the secondary stimulation. IL‐4 production was increased by the pretreatment whereas synthesis of
IFN‐γ was diminished. Addition of DEX in the second activation suppressed all cytokine
production. In brief, the transient presence of glucocorticoids in the culture induces a change in the pattern of
cytokine production but the continuous presence causes inhibition of cytokine synthesis. Further studies in which
IL‐4 was used together with DEX showed that the cytokine potentiated the effect of the hormone.

The data here presented suggest that glucocorticoids and the neuroendocrine system may be expected
to have long‐term immunological effects as well as short‐lived immunosuppressive ones. High concentration of
glucocorticoids suppress cytokine production but when steroids return to basal levels the immune response is
directed in a way that favors Th2‐type reactions. Possible implications regarding the immune response to pathogens
and autoantigens are discussed.
Correction of the TNF‐LTα‐Deficient Phenotype by Bone Marrow Transplantation
Authors: Ryffel Bernhard , Hir Michel Le , Müller Mathias , Eugster Hans-Pietro
Published: June 2008

Mice with inactivated TNF‐LTα genes have profound abnormalities of the immune system with hypoimmunoglobulinaemia, lack of lymph nodes, undifferentiated spleen, and defective Ig class switch. Transplantation of bone marrow cells from wild‐type mice restored the synthesis of TNF, corrected the splenic microarchitecture, repopulated the lamina propria with IgA‐producing plasma cells, and normalized the serum immunoglobulin levels of TNF‐LTα deficient mice. Furthermore, the formation of germinal centers in the spleen and the defective Ig class switch in response to a T‐cell‐dependent antigen is corrected. These data demonstrate that most TNFproducing cells are bone‐marrow‐derived, and that the immunodeficiency due to TNF‐LTα deletion can be corrected to a large extent by normal bone marrow, cell transplantation.
Effector Mechanisms in Low‐Dose Streptozotocin‐induced Diabetes
Authors: Lukić Miodrag L. , Stošić-Grujičić Stanislava , Shahin Allen
Published: June 2008

The cellular and molecular requirements for β‐cell damages in an immune‐mediated toxininduced insulin‐dependent diabetes mellitus have been studied in the model of multiple low‐dose streptozotocin‐induced diabetes in rats and mice. It was found that strain‐related susceptibility to diabetes induction correlated with a higher level of IL‐2, IFN‐γ, and TNF‐α production, whereas such differences were not observed when IL‐1 and NO production by macrophages were analyzed; elimination of immunoregulatory RT6+T cells that increases IFN‐γ production, enhances susceptibility to MLD‐STZ‐induced diabetes; mercury‐induced Th‐2 cells downregulated
the disease; IFN‐γ‐mediated macrophage activation to produce proinflammatory cytokines rather than NO is an important event in early diabetogenic effects of invading macrophages; inhibition of IL‐1 activity downregulates diabetes induction; and generation of NO in β cells appears to be important for diabetogenic effects. Taken together, data indicate that MLD‐STZ diabetes is induced by Th‐1 lymphocytes that secrete soluble effector molecules that activate macrophages and promote destruction of β cells possibly by both nitric oxide and nonnitric oxide‐mediated mechanisms.
Increased Thymic B Cells but Maintenance of ThymicStructure, T Cell Differentiation and Negative Selectionin Lymphotoxin‐α and TNF Gene‐Targeted Mice
Authors: Grech Adrian P. , Riminton D. Sean , Gabor Melinda J. , Hardy Charles L. , Sedgwick Jonathon D. , Godfrey Dale I.
Published: June 2008

TNF, lymphotoxin (LT) and their receptors are expressed constitutively in the thymus. It
remains unclear whether these cytokines play a role in normal thymic structure or function.
We have investigated thymocyte differentiation, selection and thymic organogenesis in gene
targeted mice lacking LTα, TNF, or both (TNF/LTα-/-). The thymus was normal in
TNF/LTα-/- mice with regard to cell yields and stromal architecture. Detailed analysis of αβ
and γδ T cell‐lineage thymocyte subsets revealed no abnormalities, implying that neither TNF
nor LT play an essential role in T cell differentiation or positive selection. The number and
distribution of thymic CD11c+ dendritic cells was also normal in the absence of both TNF and
LTα. A three‐fold increase in B cell numbers was observed consistently in the TNF/LTα-/-
thymus. This phenotype was due entirely to the LTα deficiency and associated with changes
in the hemopoietic compartment, rather than the thymic stromal compartment of LTα-/- mice.
Finally, specific Vβ8+ T cell deletion within the thymus following intrathymic injection of
staphylococcal enterotoxin B (SEB) was TNF/LT independent. Thus, despite the presence of
these cytokines and their receptors in the normal thymus, there appears no essential role for
either TNF or LT in development of organ structure or for those processes associated with T
cell repertoire selection.
Extracellular Matrix and Cytokines: A Functional Unit
Authors: Schönherr Elke , Hausser Heinz-JüRgen
Published: June 2008

The extracellular matrix (ECM) as well as soluble mediators like cytokines can influence the
behavior of cells in very distinct as well as cooperative ways. One group of ECM molecules
which shows an especially broad cooperativety with cytokines and growth factors are the proteoglycans.
Proteoglycans can interact with their core proteins as well as their glycosaminoglycan
chains with cytokines. These interactions can modify the binding of
cytokines to their cell surface receptors or they can lead to the storage of the soluble factors in
the matrix. Proteoglycans themselves may even have cytokine activity. In this review we
describe different proteoglycans and their interactions and relationships with cytokines and
we discuss in more detail the extracellular regulation of the activity of transforming growth
factor‐β (TGF‐β) by proteoglycans and other ECM molecules. In the third part the interaction
of heparan sulfate chains with fibroblast growth factor‐2 (FGF‐2, basic FGF) as a prototype
example for the interaction of heparin‐binding cytokines with heparan sulfate proteoglycans
is presented to illustrate the different levels of mutual dependence of the cytokine network
and the ECM.
Role of Extracellular Matrix‐Mediated Interactionsin Thymocyte Migration
Authors: Savino Wilson , Dalmau Sérgio Ranto , Dealmeida Vinícius Cotta
Published: June 2008

Cell adhesion, migration, differentiation and survival or death is amongst a large spectrum of
biological responses that can be elicited by ligation of extracellular matrix components to
their corresponding receptors. As regards the physiology of the thymus, cell migration is a
crucial event in the general process of T cell differentiation. Studies on the intrathymic distribution
of ECM components revealed that fibronectin, laminin and type IV collagen, are not
restrictedly located at typical basement membrane sites, also forming a thick network in the
medullary region of the thymic lobules, whereas very thin ECM fibers are found within the
cortex. These ECM components are essentially produced by thymic microenvironmental
cells, which also drive thymocyte differentiation. Signals triggered by ECM are conveyed
into thymocytes or microenvironmental cells through specific membrane receptors, and most
of them belong to the integrin type, such as the VLA‐3, VLA‐4, VLA‐5 and VLA‐6. In vitro
studies revealed that adhesion of thymocytes to thymic microenvironmental cells is mediated
by extracellular matrix. Such an adhesion is preferentially done by immature thymocytes.
Importantly, ECM‐mediated interactions also govern the entrance and exit of thymocytes in
the lymphoepithelial complexes named thymic nurse cells. Lastly, pathological conditions,
including infectious and autoimmune diseases, in which changes of ECM ligands and receptors
are observed, course with alterations in thymocyte migration and death. In conclusion,
the fact that ECM can modulate traffic, differentiation, death and survival of normal thymocytes
adds clues for understanding how ECM‐mediated interactions behave in the thymus, not
only in normal, but also in pathological conditions.
Proteolytic Activity of Human Lymphoid Tumor Cells.Correlation with Tumor Progression
Authors: Vacca Angelo , Ribatti Domenico , Ria Roberto , Pellegrino Antonio , Bruno Michele , Merchionne Francesca , Dammacco Franco
Published: June 2008

Matrix metalloproteinase (MMP) expression and production are associated with
advanced‐stage tumor and contribute to tumor progression, invasion and metastases. The current
study was designed to determine the expression and production of MMP‐2 (gelatinase A)
and MMP‐9 (gelatinase B) by human lymphoid tumor cells. Changes in expression and production
were also investigated during tumor progression of multiple myeloma and mycosis
fungoides. In situ hybridization analysis revealed that lymphoblastic leukemia B cells (SB
cell line), multiple myeloma (MM) cells (U266 cell line) and lymphoblastic leukemia T cells
(CEM and Jurkat cell lines) express constitutively the mRNA for MMP‐2 and/or MMP‐9. We
demonstrated by gelatin‐zymography of cell culture medium that both enzymes were secreted
in their cleaved (activated) form. In situ hybridization of bone marrow plasma cells and gelatin‐
zymography of the medium showed that patients with active MM (diagnosis, relapse,
leukemic progression) express higher levels of MMP‐2 mRNA and protein than patients with
non‐active MM (complete/objective response, plateau) and with monoclonal gammopathies
of undetermined significance (MGUS). MMP‐9 expression and secretion was similar in all
patient groups. In patients with mycosis fungoides (MF), the expression of MMP‐2 and
MMP‐9 mRNAs was significantly upregulated with advancing stage, in terms of lesions both
positive for one of two mRNAs and with the greatest intensity of expression. Besides MF
cells, the MMP‐2 and/or MMP‐9 mRNAs were expressed by some stromal cell populations
(microvascular endothelial cells, fibroblasts, macrophages), suggesting that these cells cooperate
in the process of tumor invasion. Our studies identify MMPs as an important class of
proteinases involved in the extracellular matrix (ECM) degradation by human lymphoid
tumors, and suggest that MMPs inhibitors may lead to important new treatment for their control.
In vitro and in vivo Expression of InterstitialCollagenase/MMP‐1 by Human Mast Cells
Authors: Di Girolamo Nick , Wakefield Denis
Published: June 2008

Degradation of the extracellular matrix occurs under physiological and pathological conditions,
thought to be principally mediated by a family of neutral proteolytic enzymes termed
the matrix metalloproteinases (MMPs). The present study was initiated to determine whether
mast cells have the ability to produce these proteases in diseased and normal human tissue.
Immunohistochemistry and in situ hybridization was performed to localize interstitial collagenase
protein and mRNA transcripts in diseased human tissue. The human mast cell line
HMC‐1 was cultured under serum free conditions, stimulated with phorbol mystrate acetate
(PMA) and supernatants analyzed by Western blotting and zymography to determine the profile
of secreted MMPs. The dog mast cell line BR, known to secrete gelatinolytic enzymes,
was used in parallel studies. Total RNA was extracted and analyzed by RT‐PCR for the
expression of tissue inhibitors of MMP (TIMPs). Collagenase‐1 protein and mRNA were
expressed by tryptase and chymase positive human mast cells in all tissue analyzed. This proteinase
wa also detected in the cytoplasm and conditioned media of HMC‐1 cells. PMA
induced gelatinolytic activity in both mast cell lines examined. TIMP‐1 immunoreactivity
was detected and TIMP‐1, and‐2 (but not TIMP‐3) mRNA transcripts were amplified from
HMC‐1 cells. This is the first demonstration of the expression of collagenase‐1 by human
mast cells in both inflamed and normal tissues, and by a human mast cell line. MMPs secreted
by these cells could contribute to the extensive matrix lysis characteristic of diseases such as
rheumatoid arthritis and inflammatory ocular disorders. Alternatively collagenase‐1 production
by mast cells may play a critical role in cell invasion and migration into sites of inflammation.
From the ECM to the Cytoskeleton and Back: How Integrins Orchestrate T Cell Action
Authors: Epler Jennifer A. , Liu Rugao , Shimizu Yoji
Published: June 2008

T lymphocytes constitute a highly dynamic tissue type. During the course of their lives, they
travel through a variety of physiological environments and experience a multitude of interactions
with extracellular matrix components and other cells. In order to do this, they must
receive many environmental cues, and translate these signals into the appropriate biological
actions. Particularly dramatic are the cytoskeletal shape changes a T cell must undergo during
the processes of leaving the bloodstream, migrating through tissues, and encountering antigen.
In this review, we highlight the role of integrins in providing a link between the extracellular
environment and cytoskeletal regulation and how these receptors help to orchestrate T
cell migration and antigen recognition.
Transformation of Undifferentiated Thy‐1lo B220+ Thymic Lymphoid Cells by the Abelson Murine Leukemia Virus
Authors: Chen Eric Y. , Swanson Bradley J. , Clark Steven S.
Published: June 2008

Intrathymic injection of the Abelson murine leukemia virus (A‐MuLV) results in transformation
of immature T and B lymphoid cells. In this report we demonstrate that the concentration
of A‐MuLV injected into murine thymi influences the selection of the transformation target.
Thus, concentrated A‐MuLV gives rise to Thy‐1+ B220- thymomas. In contrast, dilute virus
induces B220+ thymomas that also express low levels of Thy‐ 1 (Thy‐1lo), a phenotype that is
similar to marrow‐derived progenitor B‐lymphoid cells (pro‐B cells) that are highly susceptible
to A‐MuLV transformation invitro. However, rare B220+ lymphoid cells isolated from
normal adult thymi were not transformed by A‐MuLV invitro, while B220+ cells isolated
from bone marrow were highly susceptible to transformation by A‐MuLV. The Thy‐1lo
B220+population in the primary thymomas had not rearranged TCRγ, TCRβ, or Igκ genes, but
contained subpopulations that assembled Ig DJH or VDJH genes and were therefore similar to
transformed pro‐ and pre‐B cells obtained from A‐MuLV infected fetal liver and adult bone
marrow, respectively. However, unlike A‐MuLV‐transformed pro‐ and pre‐B cells, many (40–
70%) of the Thy‐1lo B220+ transformed thymoma cells had not rearranged Igh genes, and
therefore appear to represent undifferentiated lymphoid cells. We conclude that A‐MuLV may
transform an undifferentiated lymphoid target in the thymus.