Biomedical subjects

Thomas Hehlgans

Publications and source records attributed to Thomas Hehlgans.

13 recordsLinked to original sources

PubMed2006

Localization of the mouse defense lectin ficolin B in lysosomes of activated macrophages.

Ficolins are pattern-recognition molecules of the innate immune system able to trigger the lectin pathway of the complement activation upon binding to microbial surfaces. In humans, two plasma ficolins have been identified and characterized, whereas a third cell-associated ficolin (M-ficolin) was found on monocyte surfaces. The mouse homologue of M-ficolin is called ficolin B. Although the spatial-temporal expression patterns of mouse ficolins have been described recently, the subcellular localization of ficolin B protein is so far unknown. By using ficolin B-specific antibodies and confocal microscopy, we show that ficolin B is expressed within mouse peritoneal exudate macrophages and is co-localized with Lamp-1, a marker for lysosomes and late endosomes. In addition, the data indicate that ficolin B expression is up-regulated upon macrophage activation.

Animals↗

PubMed2005

Colocalization of endogenous TNF with a functional intracellular splice form of human TNF receptor type 2.

BACKGROUND: Tumor necrosis factor (TNF) is a pleiotropic cytokine involved in a broad spectrum of inflammatory and immune responses including proliferation, differentiation, and cell death. The biological effects of TNF are mediated via two cell surface TNF receptors: p55TNFR (TNFR1; CD120a) and p75TNFR (TNFR2; CD120b). Soluble forms of these two receptors consisting of the extracellular domains are proteolytically cleaved from the membrane and act as inhibitors. A novel p75TNFR isoform generated by the use of an additional transcriptional start site has been described and was termed hicp75TNFR. We focused on the characterization of this new isoform as this protein may be involved in chronic inflammatory processes. METHODS: Cell lines were retroviraly transduced with hp75TNFR isoforms. Subcellular localization and colocalization studies with TNF were performed using fluorescence microscopy including exhaustive photon reassignment software, flow cytometry, and receptosome isolation by magnetic means. Biochemical properties of the hicp75TNFR were determined by affinity chromatography, ELISA, and western blot techniques. RESULTS: We describe the localization and activation of a differentially spliced and mainly intracellularly expressed isoform of human p75TNFR, termed hicp75TNFR. Expression studies with hicp75TNFR cDNA in different cell types showed the resulting protein mostly retained in the trans-Golgi network and in endosomes and colocalizes with endogenous TNF. Surface expressed hicp75TNFR behaves like hp75TNFR demonstrating susceptibility for TACE-induced shedding and NFkappaB activation after TNF binding. CONCLUSION: Our data demonstrate that intracellular hicp75TNFR is not accessible for exogenously provided TNF but colocalizes with endogenously produced TNF. These findings suggest a possible intracellular activation mechanism of hicp75TNFR by endogenous TNF. Subsequent NFkappaB activation might induce anti-apoptotic mechanisms to protect TNF-producing cells from cytotoxic effects of TNF. In addition, the intracellular and not TACE-accessible splice form of the hp75TNFR could serve as a pool of preformed, functional hp75TNFR.

Journal Article↗

PubMed2005

Role of CREB1 and NF{kappa}B-p65 in the down-regulation of renin gene expression by tumor necrosis factor {alpha}.

Tumor necrosis factor-alpha (TNFalpha) is a potent inhibitor of renin gene expression in renal juxtaglomerular cells. We have found that TNFalpha suppresses renin transcription via transcription factor NFkappaB, which targets a cAMP responsive element (CRE) in the renin promoter. Here we aimed to further clarify the role of NFkappaB and the canonical CRE-binding proteins of the CRE-binding protein/activating transcription factor (CREB/ATF) family in the inhibition of renin gene expression by TNFalpha in the juxtaglomerular cell line As4.1. TNFalpha caused a moderate decrease in the binding of CREB1 to its cognate CRE DNA binding site. On the other hand, NFkappaB-p65 transcriptional activity was substantially reduced by TNFalpha, which targeted a trans-activation domain at the very C terminus of the p65 molecule. Our results suggest that TNFalpha inhibits renin gene expression by decreasing the transactivating capacity of NFkappaB-p65 and partially by attenuating CREB1 binding to CRE.

Animals↗

PubMed2005

The intriguing biology of the tumour necrosis factor/tumour necrosis factor receptor superfamily: players, rules and the games.

The members of the tumour necrosis factor (TNF)/tumour necrosis factor receptor (TNFR) superfamily are critically involved in the maintenance of homeostasis of the immune system. The biological functions of this system encompass beneficial and protective effects in inflammation and host defence as well as a crucial role in organogenesis. At the same time, members of this superfamily are responsible for host damaging effects in sepsis, cachexia, and autoimmune diseases. This review summarizes recent progress in the immunobiology of the TNF/TNFR superfamily focusing on results obtained from animal studies using gene targeted mice. The different modes of signalling pathways affecting cell proliferation, survival, differentiation, apoptosis, and immune organ development as well as host defence are reviewed. Molecular and cellular mechanisms that demonstrate a therapeutic potential by targeting individual receptors or ligands for the treatment of chronic inflammatory or autoimmune diseases are discussed.

Animals↗

PubMed2004

From "junk" to gene: curriculum vitae of a primate receptor isoform gene.

Exonization of Alu retroposons awakens public opinion, particularly when causing genetic diseases. However, often neglected, alternative "Alu-exons" also carry the potential to greatly enhance genetic diversity by increasing the transcriptome of primates chiefly via alternative splicing.Here, we report a 5' exon generated from one of the two alternative transcripts in human tumor necrosis factor receptor gene type 2 (p75TNFR) that contains an ancient Alu-SINE, which provides an alternative N-terminal protein-coding domain. We follow the primate evolution over the past 63 million years to reconstruct the key events that gave rise to a novel receptor isoform. The Alu integration and start codon formation occurred between 58 and 40 million years ago (MYA) in the common ancestor of anthropoid primates. Yet a functional gene product could not be generated until a novel splice site and an open reading frame were introduced between 40 and 25 MYA on the catarrhine lineage (Old World monkeys including apes).

Animals↗

PubMed2004

Lymphotoxin-beta receptor activation by activated T cells induces cytokine release from mouse bone marrow-derived mast cells.

Lymphotoxin-beta receptor (LTbetaR) signaling is known to play a key role in embryonic lymphoid organ formation as well as maintenance of lymphoid architecture. Activation of the LTbetaR is induced by either the heterotrimeric lymphotoxin-alpha(1)beta(2) (LTalpha(1)beta(2)) or the homotrimeric LIGHT (homologous to lymphotoxins, exhibits inducible expression, and competes with HSV gpD for herpes virus entry mediator, a receptor expressed by T lymphocyte). Both ligands are expressed on activated lymphocytes. As mast cells reside in close proximity to activated T cells in some inflammatory tissues, we examined the expression of LTbetaR on bone marrow-derived mast cells and asked whether the LTbetaR-ligand interaction would allow communication between mast cells and activated T cells. We found that mast cells express LTbetaR at the mRNA as well as at the protein level. To investigate LTbetaR-specific mast cell activation, the LTbetaR on BMMC from either wild-type or LTbetaR-deficient mice was stimulated with recombinant mouse LIGHT or agonistic mAbs in the presence of ionomycin. LTbetaR-specific release of the cytokines IL-4, IL-6, TNF, and the chemokines macrophage inflammatory protein 2 and RANTES was detected. Moreover, coculture of mast cells with T cells expressing the LTbetaR ligands also entailed the release of these cytokines. Interference with a specific LTbetaR inhibitor resulted in significant suppression of mast cell cytokine release. These data clearly show that LTbetaR expressed on mast cells can transduce a costimulatory signal in T cell-dependent mast cell activation.

Animals↗

PubMed2004

Involvement of the lectin pathway of complement activation in antimicrobial immune defense during experimental septic peritonitis.

A critical first line of defense against infection is constituted by the binding of natural antibodies to microbial surfaces, activating the complement system via the classical complement activation pathway. In this function, the classical activation pathway is supported and amplified by two antibody-independent complement activation routes, i.e., the lectin pathway and the alternative pathway. We studied the contribution of the different complement activation pathways in the host defense against experimental polymicrobial peritonitis induced by cecal ligation and puncture by using mice deficient in either C1q or factors B and C2. The C1q-deficient mice lack the classical complement activation pathway. While infection-induced mortality of wild-type mice was 27%, mortality of C1q-deficient mice was increased to 60%. Mice with a deficiency of both factors B and C2 lack complement activation via the classical, the alternative, and the lectin pathways and exhibit a mortality of 92%, indicating a significant contribution of the lectin and alternative pathways of complement activation to survival. For 14 days after infection, mannan-binding lectin (MBL)-dependent activation of C4 was compromised. Serum MBL-A and MBL-C levels were significantly reduced for 1 week, possibly due to consumption. mRNA expression profiles did not lend support for either of the two MBL genes to respond as typical acute-phase genes. Our results demonstrate a long-lasting depletion of MBL-A and MBL-C from serum during microbial infection and underline the importance of both the lectin and the alternative pathways for antimicrobial immune defense.

Animals↗

PubMed2003

Tumor necrosis factor-alpha activates NFkappaB to inhibit renin transcription by targeting cAMP-responsive element.

Tumor necrosis factor-alpha (TNFalpha) is known to inhibit renin gene expression in juxtaglomerular cells, which are the main source of renin in vivo. In the present study we aimed to characterize the intracellular mechanisms of TNFalpha signaling to renin gene in the mouse juxtaglomerular cell line As4.1. TNFalpha was found to activate NFkappaB, which is one of the principal intracellular mediators of TNFalpha signal transduction. Constitutive activation of NFkappaB suppressed renin gene transcription, but NFkappaB appeared not to target the NFkappaB binding sites in the renin promoter. Thus, NFkappaB, but not the canonical NFkappaB binding sequences in the renin promoter, seemed to be involved in the suppression of renin transcription by TNFalpha. Deletion/mutation analysis revealed that the effect of TNFalpha on renin gene is transmitted by a cAMP-responsive element (CRE) located at -2697 to -2690. Mobility shift/supershift assays evidenced for the presence of NFkappaB proteins in the complex that binds to mouse renin CRE. Our results strongly suggest that NFkappaB mediates the effect of TNFalpha on renin transcription targeting a CRE in the mouse renin promoter.

Animals↗

PubMed2003

The lymphotoxin beta receptor is critically involved in controlling infections with the intracellular pathogens Mycobacterium tuberculosis and Listeria monocytogenes.

Containment of intracellularly viable microorganisms requires an intricate cooperation between macrophages and T cells, the most potent mediators known to date being IFN-gamma and TNF. To identify novel mechanisms involved in combating intracellular infections, experiments were performed in mice with selective defects in the lymphotoxin (LT)/LT beta R pathway. When mice deficient in LT alpha or LT beta were challenged intranasally with Mycobacterium tuberculosis, they showed a significant increase in bacterial loads in lungs and livers compared with wild-type mice, suggesting a role for LT alpha beta heterotrimers in resistance to infection. Indeed, mice deficient in the receptor for LT alpha(1)beta(2) heterotrimers (LT beta R-knockout (KO) mice) also had significantly higher numbers of M. tuberculosis in infected lungs and exhibited widespread pulmonary necrosis already by day 35 after intranasal infection. Furthermore, LT beta R-KO mice were dramatically more susceptible than wild-type mice to i.p. infection with Listeria monocytogenes. Compared with wild-type mice, LT beta R-KO mice had similar transcript levels of TNF and IFN-gamma and recruited similar numbers of CD3(+) T cells inside granulomatous lesions in M. tuberculosis-infected lungs. Flow cytometry revealed that the LT beta R is expressed on pulmonary macrophages obtained after digestion of M. tuberculosis-infected lungs. LT beta R-KO mice showed delayed expression of inducible NO synthase protein in granuloma macrophages, implicating deficient macrophage activation as the most likely cause for enhanced susceptibility of these mice to intracellular infections. Since LIGHT-KO mice proved to be equally resistant to M. tuberculosis infection as wild-type mice, these data demonstrate that signaling of LT alpha(1)beta(2) heterotrimers via the LT beta R is an essential prerequisite for containment of intracellular pathogens.

Administration, Intranasal↗

PubMed2002

Lymphotoxin-beta receptor immune interaction promotes tumor growth by inducing angiogenesis.

Growth of solid fibrosarcoma tumors in mice was inhibited by the release of a solublelymphotoxin-beta receptor inhibitor (LTbetaR-immunoglobulin fusion protein) from the tumor cells. Tumor growth arrest in mice deficient in the ligand LTalpha1beta2 demonstrated the requirement for activation of the LTbetaR on the tumor cells by host cell-derived LTalpha1beta2. Activation of the LTbetaR resulted in enhanced release of macrophage inflammatory protein-2. Blocked angiogenesis was revealed in LTbetaR inhibitor-producing tumor nodules by immunohistochemistry and in vivo microscopy. The growth arrest of LTbetaR inhibitor-producing fibrosarcomas was overcome by forced MIP-2 expression in the tumor cells. Thus, LTbetaR activation on tumor cells by activated host lymphocytes can initiate a novel proangiogenic pathway leading to organized tumor tissue development.

Animals↗

PubMed2002

Requirement for tumor necrosis factor receptor 2 expression on vascular cells to induce experimental cerebral malaria.

Using tumor necrosis factor receptor type 2 (TNFR2)-deficient mice and generating bone marrow chimeras which express TNFR2 on either hematopoietic or nonhematopoietic cells, we demonstrated the requirement for TNFR2 expression on tissue cells to induce lethal cerebral malaria. Thus, TNFR2 on the brain vasculature mediates tumor necrosis factor-induced neurovascular lesions in experimental cerebral malaria.

Animals↗

PubMed2002

The TNF-TNF receptor system.

Different forms of tumor necrosis factor (TNF) interact with two specific receptors for TNF (TNFR) on the cell membrane to induce a variety of effects. While sharing structural similarities in their extracellular domains, the two TNFRs differ in their intracellular domain, their signal transduction, and consequently their function. In addition, one of the two TNFRs can be expressed in two differently located isoforms. This makes the TNF-TNFR system very complex. The dual TNF function for either cell death or survival upon interaction of members of the TNF ligand family with members of the TNF receptor family will be discussed.

Animals↗

PubMedDate not supplied

Activation of the lymphotoxin-beta receptor induces NFkappaB-dependent interleukin-6 and MIP-2 secretion in mouse fibrosarcoma cells.

Activation of the lymphotoxin beta-receptor (LTbetaR), a member of the tumor necrosis factor receptor family, plays a crucial role in lymphoid organogenesis and tumor development. Lymphotoxin alpha(1)beta(2) (LTalpha(1)beta(2)) and LIGHT have been identified as membrane anchored ligands for the LTbetaR. While LTbetaR is expressed on a wide range of cell types e.g. fibroblasts and monocytes, the ligands are expressed only on activated lymphocytes and NK cells. In order to characterize LTbetaR expression and the biological consequences of LTbetaR activation rat anti-mouse LTbetaR monoclonal antibodies were generated. These antibodies recognized a mouse LTbetaR-Ig fusion protein as well as endogenous LTbetaR on a variety of mouse fibroblast and fibrosarcoma cell lines. Specificity was demonstrated by the lack of binding to LTbetaR-deficient embryonic fibroblasts. Competitive binding studies revealed that three different epitopes were recognized by the monoclonal antibodies. Two of the monoclonals activated the LTbetaR and induced activation of NFkappaB and secretion of MIP-2 and IL-6 in L929 mouse fibroblast cells. MIP-2 and IL-6 secretion was NFkappaB-dependent because IkappaB-transfected cells released significantly reduced amounts of both mediators.

Animals↗