Is something other than clonal selection required to account for T-cell regulation? Inga Melchers and Klaus Eichmann show by carefully quantified studies that T-helper cells may be seen to react in a "monogamous" fashion whereas frequency analyses indicate that suppressor cells are at least polygamous if not promiscuous. Do we have a paradox here within the fundamental framework of the clonal selection theory?

During the first decade of the twentieth century, the German bacteriologist Fred Neufeld, later Director of the Robert Koch-Institute in Berlin, first described the differentiation of pneumococci into serotypes on the basis of type-specific antisera. This finding was essential for subsequent research at the Rockefeller Institute of Medical Research (RIMR) in New York, and elsewhere, aiming for the conquest of human pneumococcal pneumonia, including antiserum therapy, the discovery that the type-specific antigens were carbohydrates, and the development of effective multivalent pneumococcal polysaccharide vaccines. Moreover, on the basis of pneumococcal serotypes Fred Griffith, in 1928 in London, discovered pneumococcal transformation, and Oswald T. Avery and coworkers, in 1944 at RIMR, identified DNA as the transforming substance. This sequence of events, leading to today's knowledge that genes consist of DNA, was initiated by a farsighted move of Simon Flexner, first Director of the RIMR, who asked Neufeld to send his pneumococcal typing strains, thus setting the stage for pneumococcal research at RIMR. Here, we describe Fred Neufeld's contributions in this development, which have remained largely unknown.

The giant cytosolic protease tripeptidyl peptidase II (TPPII) has been implicated in the regulation of proliferation and survival of malignant cells, particularly lymphoma cells. To address its functions in normal cellular and systemic physiology we have generated TPPII-deficient mice. TPPII deficiency activates cell type-specific death programs, including proliferative apoptosis in several T lineage subsets and premature cellular senescence in fibroblasts and CD8(+) T cells. This coincides with up-regulation of p53 and dysregulation of NF-kappaB. Prominent degenerative alterations at the organismic level were a decreased lifespan and symptoms characteristic of immunohematopoietic senescence. These symptoms include accelerated thymic involution, lymphopenia, impaired proliferative T cell responses, extramedullary hematopoiesis, and inflammation. Thus, TPPII is important for maintaining normal cellular and systemic physiology, which may be relevant for potential therapeutic applications of TPPII inhibitors.

Tripeptidyl peptidase II (TPPII) is an oligopeptidase forming giant complexes in the cytosol that have high exo-, but also, endoproteolytic activity. Immunohistochemically, the complexes appear as distinct foci in the cytosol. In part controversial biochemical and functional studies have suggested that TPPII contributes, on the one hand, positively to Ag processing by generating epitope carboxyl termini or by trimming epitope precursors, and, on the other, negatively by destroying potentially antigenic peptides. To clarify which of these roles is predominant, we generated and analyzed TPPII-deficient mice. Cell surface levels of MHC class I peptide complexes tended to be increased on most cell types of these mice. Although presentation of three individual epitopes derived from lymphocytic choriomeningitis virus was not elevated on TPPII-/- cells, that of the immunodominant OVA epitope SIINFEKL was significantly enhanced. Consistent with this, degradation of a synthetic peptide corresponding to the OVA epitope and of another corresponding to a precursor thereof, both being proteasomally generated OVA fragments, was delayed in TPPII-deficient cytosolic extracts. In addition, dendritic cell cross-presentation of phagocytosed OVA and of OVA internalized as an immune complex was increased to about the same level as direct presentation of the Ag. The data suggest a moderate, predominantly destructive role of TPPII in class I Ag processing, in line with our finding that TPPII is not induced by IFN-gamma, which up-regulates numerous, predominantly constructive components of the Ag processing and presentation machinery.

Background: As deficiency of innate immunity was found by us earlier in elderly (> 60) and also in leukemia patients, the innate immunity of blood leukocytes obtained from tumorous patients (carcinoma ovarii and adenocarcinoma endometrium), Alzheimer disease (AD) patients and control group in the same age was compared. Two of the mechanisms engaged in innate immunity were studied: — resistance of leukocytes ex vivo to viral infection, — production of cytokines (TNFα, IFNs, IL-10, IL-12) Methods: The resistance of leukocytes was designated just after isolation by infection with vesicular stomatitis virus (VSV). Titer of VSV 0-1 log TCID50 indicate for complete resistance, titer 2–3 log for partial resistance, > 4 log indicate for very low or lack of resistance. Cytokines were assayed with ELISA test. Results: Results of experiments showed that leukocytes of the three groups are very sensitive to VSV infection. In contrary, the basic differences in cytokines production by leukocytes of these groups. The leukocytes of cancer group (n = 15) produced more TNFα than control (n = 10) and AD group(n = 35), less IFNs and IL-10, and not et all IL-12. The leukocytes of AD patients produce high level of early IL-12 and IL-10 (spontaneous and VSV-induced) and less than control TNFα and IFNs. Conclusion: The sensitivity of leukocytes to VSV in these three groups is caused by deficiency of innate antiviral immunity. Different panel of cytokines produced by leukocytes of patients with senile diseases may participate in their development: TNFα in cancer and IL12 in AD. O2

The development of T cells in the thymus is dependent on interactions between thymocytes and thymic stromal cells, on stimulation by growth factors, and on the binding to and migration along extracellular matrix (ECM) components. As metalloproteinases (MP) are involved in processes such as growth factor release and ECM modelling, we assessed the effect of MP inhibitors on T-cell development using fetal thymic organ culture systems. MP inhibitors significantly reduced the numbers of CD4/CD8 double-positive (DP) and mature single-positive thymocytes generated, correlated with a reduced number of cell cycles between the double-negative (DN)3 and DP stages. The progression of early thymocyte progenitors through the DN1-4 stages of development was also severely affected, including incomplete upregulation of CD25, decreased DN3 cell numbers, reduced rearrangement of the T-cell receptor (TCR)-beta locus and expression of intracellular TCR-beta by fewer DN3 cells. When purified DN1 cells were utilized as donor cells in reaggregate thymic organ cultures, essentially no DP thymocytes were produced in the presence of MP inhibitors. The results suggest that MP inhibitors affect the differentiation of developing thymocytes before, and reduce proliferation after, pre-TCR-mediated selection.

The Ebeta enhancer has been shown to be dispensable for germline transcription of nonrearranged TCRbeta segments but appears to be required for TCRbeta V to DJ rearrangement. Ebeta dependency of the subsequent expression of VDJ-rearranged TCRbeta genes in thymic subpopulations has so far not been analyzed. We generated transgenic mice, using a Vbeta8.2Dbeta1Jbeta1.3-rearranged TCRbeta bacterial artificial chromosome, which lacked Ebeta, and monitored transgene expression by flow cytometry using Vbeta-specific mAbs and an IRES-eGFP reporter. Transgene expression was found in double negative (DN)2 and DN3 but not at later stages of thymopoesis. There was no toxicity associated with the transgene given that apoptosis in DN3, DN4 was not increased, and the number of DN4 cells generated from DN3 cells in reaggregate thymic organ cultures was not diminished. The transgenic TCRbeta gave rise to a pre-TCR, as suggested by its ability to suppress endogenous TCRbeta rearrangement, to facilitate beta-selection on a TCRbeta-deficient background and to inhibit gammadelta T cell lineage development. The results suggest that the Vbeta8.2 promoter is sufficient to drive expression of rearranged TCRbeta VDJ genes Ebeta independently in DN2/DN3 but not at later stages.

Otto Westphal, the founding Editor of the EJI, has died at 91 years of age at his home in Montreux, Switzerland. He was born in Berlin-Charlottenburg at a time which he himself referred to as the “Belle Epoque” of Science in Germany. His parents had been part of the intellectual elite in Berlin. His father, Wilhelm, was a professor in the Physics Faculty of Berlin University, which included scientists as prominent as Max Planck, Albert Einstein, Max von Laue and Gustav Hertz, who were regular guests at the Westphal's home. Westphal later wrote “I had no idea how privileged I was to share their company”. His mother, Olga, a gifted pianist, played four hands with Max Planck and accompanied Einstein who played the violin. She is quoted to have written in her diary “I joined in the singing (of Brahms’ Liebeslieder performed at her home by a choir directed by Max Planck) so intensely that Otto was born 10 days early”. Westphal studied chemistry, the leading science at the time, and became fascinated by the concepts of stereochemistry introduced by Jacobus Van't Hoff, Emil Fischer, Paul Ehrlich, and Svante Arrhenius. In 1937 he obtained his PhD with a study on blood group serology, and thereafter worked as scientific assistant in the Kaiser Wilhelm Institute of Medical Research, in Heidelberg, on the synthesis of vitamins and studied immunology and serology. As a chemist with experience in immunology, Westphal was one of the first to understand the modern concepts of immunochemistry. At age 29 he was appointed head of the department of biochemistry at the medical faculty of the University of Göttingen. There he met Otto Lüderitz and Botho Kickhöfen, who stayed with Westphal for their entire scientific careers. The group collaborated with F. Schütz, professor of hygiene in Göttingen, who was working on bacterial vaccines. Together they extracted Proteus OX19, which cross-reacted with Salmonella typhi and seemed a promising typhus vaccine candidate. A laboratory worker got contaminated and developed a high but transient fever, an incident leading to the identification of the first purified bacterial pyrogen. Fever therapy was an important medical regime at the time and the discovery soon made Westphal prominent in the field. He started to work on LPS, initially described as endotoxin, in 1947 after he had accepted an offer of the Swiss pharmaceutical company, Dr. Wander AG, to head a new institute in Bad Säckingen, near the Swiss border. Feeling that Bad Säckingen was a somewhat isolated place for an international research institute, Wander and Westphal agreed in 1956 to move the institute to Freiburg, near a major university. It soon became evident that the future of the institute was in basic research, and in 1961 the decision was made to incorporate the institute into the Max Planck Society, the follower of the Kaiser-Wilhelm-Gesellschaft. It was named Max-Planck-Institut für Immunbiologie (MPIIB) and Westphal was its first director. Under his directorship the structure of Lipid A, the moiety that accounts for the biological activity of LPS, was elucidated, and variations in the polysaccharide side chains were shown to form the structural basis of the serological specificities. Under his direction, the MPIIB became a nationally and internationally leading center of immunological research, and a breeding ground for generations of biomedical investigators who later held key positions in research and medicine in Germany. He was the main initiator in the foundation of the German Society of Immunology in Germany (DGfI) in 1967, and was elected first president, an office he held for 10 years. He played a key role in the foundation of the EJI in 1971 and appointed his long-time collaborator Botho Kickhöfen as Editor in Chief. The MPIIB has been home to its editorial office ever since. In the Max-Planck-Society and in the Deutsche Forschungsgemeinschaft, he chaired at one time or another nearly all important decision-making bodies and committees. He was a member of multiple learned academies in Germany and abroad, and received numerous honors and prizes, including the Emil von Behring Prize (1964), the Paul Ehrlich Prize (1968) and Robert Koch Medal (1983). After Westphal retired in 1981, he served for a year as interim Chairman of the German Cancer Research Center in Heidelberg. Westphal`s charismatic personality held many facettes beyond that of a prominent scientist. He was a skilled player of the flute and loved to organize classical music recitals at his institute where he performed together with professional musicians. At symposia termed “Reflexions”, where Westphal invited his friends to listen to music and discuss the meaning of life, eminent scientists, including among others Sir Peter Medawar, Lewis Thomas, Manfred Eigen, met with outstanding artists such as the cembalists Fritz Neumeyer, who pioneered performance on historical keyboard instruments, and Edith Picht-Axenfeld. He paved the career path for many of us, regardless of whether one had trained in the MPIIB or not. He was not secretive about his juvenile membership in the Nazi SA, a mistake which he regretted during his entire life, and it is significant that many Israeli scientists were in friendly contact with Westphal, who had been a driving force in the early post-war relationships between the Weizmann Institute of Science and the Max Planck Society. For the 40th anniversary of the MPIIB in 2001, the late Charles Janeway Jr began his lecture with the remark: “I should particularly like to thank Otto Westphal who spent years in purifying my favorite molecule, which is LPS”. The encounter between these two outstanding scientists, Westphal the chemist who elucidated the structure of the molecule and Janeway the biologist who, almost half a century later, defined its crucial role in immunity, was to remain a one-time event. At this symposium, 20 years after his retirement, Westphal gave a lucid speech commemorating the foundation of the institute, and was honored by standing ovations. The MPIIB and the EJI will always take pride in being brainchildren of this remarkable man.

The balance of arginine metabolism via nitric oxide synthase (NOS) or arginase is an important determinant of the inflammatory response of murine macrophages and dendritic cells. Here we analyzed the expression of the isoform arginase I in human myeloid cells. Using healthy donors and patients with arginase I deficiency, we found that in human leukocytes arginase I is constitutively expressed only in granulocytes and is not modulated by a variety of proinflammatory and anti-inflammatory stimuli in vitro. We demonstrate that arginase I is localized in azurophil granules of neutrophils and constitutes a novel antimicrobial effector pathway, likely through arginine depletion in the phagolysosome. Our findings demonstrate important differences between murine and human leukocytes with respect to regulation and function of arginine metabolism via arginase.

Human in vivo -activated CD45R0 + CD4 + Tcells are susceptible to spontaneous apoptosis that can be inhibited by the chemokine CXCL12 and IL-2, -6, -7, and -15

The expression of housekeeping and/or immunoproteasomes in isolated thymic stroma subsets has so far not been analyzed but may have important consequences for self peptide repertoires presented by MHC class I molecules during positive and negative thymic selection. Here we determined the expression of housekeeping and immunoproteasome beta subunits and of PA28 in positively and negatively selecting stroma subsets. Positively selecting cortical thymic epithelial cells (cTEC) expressed only housekeeping but no immunoproteasome beta subunit mRNA and proteins. However, immunoproteasome beta subunits could be induced in cTEC by infection with Listeria monocytogenes or injection of IFN-gamma. In negatively selecting stroma including medullary epithelial cells and dendritic cells, incomplete and low representation of housekeeping beta subunit proteins but high and complete expression of immunoproteasome beta subunit proteins suggests absence of proper housekeeping proteasomes and predominance of immunoproteasomes. Expression of immunoproteasome beta subunits in negatively selecting stroma was independent of IFN-gamma receptor as shown in knockout (KO) mice. Absence of LMP2 altered thymic selection of the MHC class I-restricted transgenic P14 TCR in KO mice. The data suggest that negative selection may primarily involve immunoproteasome peptide repertoires and that peripheral infection may influence peptide repertoires involved in positive selection.

Development of alphabeta and gammadelta T cells depends on productive rearrangement of the appropriate TCR genes and their subsequent expression as proteins. TCRbeta and TCRgammadelta proteins first appear in DN3 and DN4 thymocytes, respectively. So far, it is not clear whether this is due to a delayed expression of TCRgammadelta proteins or to a more rapid progression to DN4 of thymocytes expressing TCRgammadelta. The answer to this question bears on the distinction between instructive and stochastic models of alphabeta/gammadelta lineage decision. To study this question, we first monitored initial TCR protein expression in wild-type and TCR transgenic mice in reaggregate thymic organ cultures. A TCRbeta transgene was expressed in nearly all DN3 and DN4 cells, accelerated DN3 to DN4 transition, and strongly diminished the number of cells that express TCRgammadelta proteins. In contrast, TCRgammadelta transgenes were expressed only in a fraction of DN4 cells, did not accelerate DN3 to DN4 transition, and did not reduce the number of DN4 cells expressing TCRbeta proteins. The TCRbeta transgene partially inhibited endogenous TCRgamma rearrangements, whereas the TCRgammadelta transgenes did not inhibit endogenous TCRbeta rearrangements. Second, we analyzed frequencies of productive TCRbeta and TCRgammadelta V(D)J junctions in DN3 and DN4 subsets. Most importantly, frequencies of productive TCRgammadelta rearrangements (Vdelta5, Vgamma1.1, and Vgamma2) appeared unselected in DN3. The results suggest a late and restricted expression of the corresponding gammadeltaTCR, severely limiting their putative instructional opportunities in alphabeta/gammadelta divergence.