Our profile

What we do

The Sektion Experimentelle Anaesthesiologie of Prof. Dr. E. Marion Schneider pursues monocytes and immature dendritic cells in septic as well as in solid tumor patients which can be enriched by cell culture techniques from peripheral blood, bone marrow and cerebrospinal fluid. Such immature dendritic cells are resistant to maturation signals and manifest impressive hemophagocytic properties. Function and phenotype support the concept immunologic anergy in sepsis and malignancies.

Image 1: Hypothesis to explain the survival of iDC in states of inflammation and NK deficiencies

Since 2007, Eos-FP, a novel fluorescent dye, isolated from a marine organism by Joerg Wiedenmann (Univ. Souphampton, U.K., former Ulm University), is applied to study phagocytosis using Eos-FP-transfected bacteria. Early after ingestion, bacteria emit red and green fluorescent light. After fusion of the phasome with acidic lysosomes, the red fluorescence is lost and partially degraded bacteria emit only green fluorescent light.

Image 2: Degraded and non-degraded Bacteria, Schreiner et al. 2011

We hold an immunological cell, DNA and data bank on hemophagocytic lymphohistiocytosis (HLH) and hyperinflammatory diseases many of which are monogenetic, or manifest HLH during immune dysfunction and states of virus-re-activation. The biological material: mononuclear cells, bone marrow, cell lines, fibroblasts and endothelial cells, as well as sera and DNA are derived from > 2000 individual patients' samples and 200 - 300 families. Flow cytometric analyses, ex-vivo stimulation protocols, plasma cytokines and cellular cytolysis assays have been determined to be correlated with genetic testings of functional polymorphisms encoded in the gene for a purinergic ion channel receptor, P2RX7, the Toll-like-receptors 2 and 4, Perforin, and enzymes related to oxidative stress. All SNPs were determined by pyrosequencing technologies. A recently detected SNP haplotype appears to be relevant for the manifestation of severe sepsis. Phenotypically, P2RX7 is highly expressed in immature dendritic cells of patients. Upon activation by ATP in vivo, such as by trauma, P2RX7 generates a calcium signal and thus amplifies inflammatory signaling pathways by the secretion of IL-1β.

Image 3: P2RX7 stimulation by ATP in wild type dendritic cells.

Image 4: ATP stimulated IL-1ß secretion by LPS-primed sepsis-APC, Schneider et al. 2011

Another field of interest concentrates on ultrastructural studies of patients’ derived immature dendritic cells and their unique characteristic in autophagy. ATP stimulates the expression of  LC3, a component of the autophagosome, as determined by flow cytometry.

Image 5: Upregulation of LC3 expression by treatment with 1mM ATP for 3 h. Negative control (black line histogram); LC3-positive cells (red line histogram). Schneider et al. 2011

Schneider, E.M., Lorenz, M.R., Walther, P. Autophagy  as a hallmark of hemophagocytic diseases. In „Autophagy in Health and Diseases.“ (N. V. Gorbunov (ed.) NOVA Publishers N.Y. 2011.

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Image 6: Tomogram of a representative and autophagic vesicle-rich hemophagocyte.

We provide knowledge of immune interaction pathways leading to immune incompetence in major inflammatory diseases. With our team, we developed a mitochondrial SNP typing array for mitochondrial mutations within a European Research Project in FP5.  For patients with hemophagocytic diseases and inflammsome-realted macrophage activation syndromes, we provide short and long-term assays for Natural Killer Cell Functional Analysis. Our group participated in the STREP “GenOSept” of FP6 to establish a SNP platform for the identification of inflammatory genes related to the manifestation of severe sepsis in adult patients suffering from bacterial infections or non-infectious complications such as pancreatitis. In FP7, Marion Schneider and her team were responsible for the development of a single cell based multiplex PCR to detect Epstein Barr Virus (EBV) in immature dendritic cells, called QuAGSiC (quantitative analysis  of Genes in single cells). In collaboration with Alberto Pasquarelli (Institut für Elektronische Bauelemente und Schaltungen, Univ. Ulm), a diamond based chip has been set up to determine potentiometric as well as amperometric measurements of P2RX7 stimulated cells).

Image 7: ATP-induced P2RX7 activation on diamond chip . Ca++ translocation and subsequent generation of amperometric signals detected.

Recently, our group concentrated on the Calcium-dependent formation of neutrophil derived extracellular traps (NETs). Results suggest that increased NETosis by neutrophils is an excellent indicator to follow clinical changes in patients with sepsis. Infectious complications as well as traumatic injury lead to massive NETformation and correlate with biomarkers such as LBP (liposolysaccharide binding protein), IL-1β, IL-6, IL-8, or TNF-α, leukocyte increase, and CRP.

Image 8: NETosis of neutrophilic granulocytes and demonstration of neutrophil derived primary granules (arrows).

Contact

Office Tel.:

+49 - 731 - 50060080/81

Office Fax:

+49 - 731 - 50060082

Laboratory:

+49 - 731 - 50060083/84/85

Our team

Profilbild von Prof. Dr. rer. nat. Marion Schneider

Prof. Dr. rer. nat. Marion Schneider

Workgroup leader Experimental Anaesthesiology

Profilbild von PD Dr. Karl Föhr

PD Dr. Karl Föhr

Profilbild von Stefan Bäder

Stefan Bäder

Profilbild von Margot Autenrieth-Kronenthaler

Margot Autenrieth-Kronenthaler

Diseases

Hemophagocytic Lymphohistiocytosis

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Macrophage Activation Syndromes

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Pain Syndromes

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Trauma, Sepsis, Septic Shock

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Major Depression

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Malignancies

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Diagnostic parameters and procedures

Shipping Material to our Laboratory

If you wish to send us material to perform diagnostics, please use the request sheet below.

Request Sheet (English)

Request Sheet (German)

Biomarkers

Cytokine concentrations in Plasma / CSF / Ascites indicate state of disease.
Surface marker phenotypes (FACS analysis) on white cells assist diagnostic accuracy.
In HLH (Hemophagocytic Lymphohistiocytosis) and MAS (Macrophage activation syndrome), NK (Natural Killer) cell function is impaired or NK-effector cells are significantly diminished.

Plasma biomarkers for inflammatory diseases

Results sheet of a healthy donor (example)

Immune Phenotypes

Phenotype analysis for inflammatory diseases

Example of a healthy donor (flow cytometry)

Whole Blood Stimulation for inflammatory and infectious diseases

Procedure

Research Application

Patients / Clinical Study Application

Examples of a clinical study

Examples of results obtained by TLR stimulation using TruCultures. IL-1ß release by healthy donors differs from patients with trauma or sepsis. Results have been presented at the TSIS 2010 meeting1.

1 Bindja J., Weiss M.E., Schmolz M., Stein G.M., Mapes J., Schneiderhan-Marra N., Joos J.O., Schneider E.M. Synthetic ligands against TLR2-9 in TruCultureTM - whole blood assays distinguish clinical stages of SIRS (trauma) and sepsis. Medimont Press, Proceedings Trauma, Shock, Inflammation and Sepsis, (2010) 55-63.

Whole blood phagocytosis assay

Using Eos-FP transfected E. coli, we provide an in-vitro phagocytosis assay to quantify the phagocytic activities of granulocytes and monocytes in whole blood and simulatenously monitor the phagosome-lysosome fusion process as described, previously (Schreiner et al. 2011); for more information see also (Dreschers et al. 2013 and Wiedenmann et al. 2004).

Literature:

Schreiner L, Huber-Lang, M, Weiss, ME, Hohmann, H, Schmolz, M., Schneider, EM. Phagocytosis and digestion of pH-sensitive fluorescent dye (Eos-FP) transfected E. coli in whole blood assays from patients with severe sepsis and septic shock. Journal of Cell Communication and Signaling. 2011, 5:139-144.

Dreschers, S., et al. Infection-induced bystander-apoptosis of monocytes is TNF-alpha-mediated. PLoS One 8, e53589.

Wiedenmann, J., et al. EosFP, a fluorescent marker protein with UV-inducible green-to-red fluorescence conversion. Proc Natl Acad Sci U S A 101, 15905-15910 (2004).

Research

Hemophagocytosis by Antigen-Presenting Cells

Hemophagocytic Cells in HLH

The electron micrograph shows hemophagocytic cells during (A) and after (B) ingesting streptavidin-coated iron beads. Histiocytic progenitors were isolated from the lymphocyte fraction of the peripheral blood of a patient with acute HLH following Ficoll separation. Iron-containing beads of precisely 5 μm in diameter are rapidly phagocytosed (at a rate of roughly 10 min/bead). Histiocytes ingest a non-physiologic quantity of particles, causing massive cell enlargement and loss of surface membrane veils (arrow, A). No apoptosis occurs following phagocytosis, as can be identified from the structure of the nucleus in B. m, mitochondria.

(Schneider EM, Lorenz I, Walther P, Janka-Schaub GE. Natural killer deficiency: A minor or major factor in the manifestation of hemophagocytic lymphohistiocytosis? J Pediatr Hematol Oncol 25(9):680-683, 2003.)

In-Vitro Hemophagocytosis

Ficoll-isolated peripheral blood or bone marrow derived mononuclear cells from patients with HLH are depleted from non-adherent cells by plastic adherence.  Within 2-14 days these cultures give rise to a homogeneous population of large, floating cells with a pleomorphic morphology (Fig. 1), prominent signs of autophagy, and an immature dendritic cell phenotype1. Even after 21 days of in vitro culture, residual blood cells can be detected in cultured hemophagocytes1 (Fig. 2). Cultured hemophagocytes can be membrane-stained using fluorescent dyes and tested for their interaction with NK effectors such as NK-92 cells2. Figure 3 shows that such membrane-stained NK92 (red dye) interact with cultured hemophagocytes of a patient with HLH and shortly after cell-cell contact, the NK-effectors are phagocytozed by the hemophagocyte (stained with a green membrane-dye). The phagocytic activities of such cultured hemophagocytes derived from cultures of different patients with HLH vary:  Some hemophagocytes ingest NK-92 within minutes and accumulate more than 10 NK-92 cells per hemophagocyte.

Current research activites concentrate on the analysis of the hemophagocytic specifcities of hemophagocytes derived from individual patients with HLH.

Legends:

Figure 1:
Phase contrast of cultured hemophagocyte, floating, pleomorphic morphology and veiled surface structure.


Figure 2:
Structural features of hemophagocytes. Hemophagocytes with ingested and partially digested erythrocytes (arrows) located in a distal phagosome (HLH, chemical fixation). (A). Early hemophagocytosis of another autologous hemophagocyte with remarkable autophagous vacuoles (HLH, chemical fixation) (B). Hemophagocytosis of a hemophagocyte (HLH, chemical fixation) (C). More advanced state of erythrocyte digestion in a phagosome being engulfed by another autophagosome (HLH, chemical fixation) (D). As in C, a more advanced state of phagocytosis and digestion (sepsis, chemical fixation) (E).


Figure 3:
3a shows contact formation between NK-92 (red) and hemophagocyte (green)

3b shows early hemophagocytosis of NK-92 and another NK-92 forming contact

3c shows completed phagocytosis of one NK-92 and several other NK-92 cells forming contact

References:

  1. Schneider, E., Lorenz, M, Walther, P Autophagy as a hallmark of hemophagocytic diseases. Vol. Chapter  3 (ed. Gorbunov. Nikolai (eds.), i.A.P., Regulation and Roles in Disease" ) (Nova Science Publishers, Inc.,   Cell Biology Research Progress, e-book, 2012).
  2. Yan, Y., et al. Antileukemia activity of a natural killer cell line against human leukemias. Clin Cancer Res 4, 2859-2868 (1998).

Microparticles

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Cellular Cytotoxicity

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Some publications

Book Chapters

Schneider, E.M., Lorenz, M.R., Walther, P. Autophagy  as a hallmark of hemophagocytic diseases. In „Autophagy in Health and Diseases.“ (N. V. Gorbunov (ed.) NOVA Publishers N.Y. 2011.

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Bruecker UB, Schneider, EM. Microarrays in der experimentellen und klinischen Forschung. In: Chirurgische Forschung; 2005, p. 304-316. Krukemeyer, MG, Spiegel, H-U (eds);Thieme Verlag, Stuttgart, New York, ISBN: 3-13-133661-7

Janka-Schaub, G., Schneider, EM. Hämophagozytische Lymphohistiozytosen. In: Pädiatrische Hämatologie und Onkologie, 2005, p231-236. Gadner, H, Gaedicke, G, Niemeyer, C, Ritter, J (eds); Springer Verlag, Heidelberg, ISBN 13 978-3-540-03702-6
 

Selected Original Manuscripts (2008-2013)

Unnewehr H, Rittirsch D, Sarma JV, Zetoune F, Flierl MA, Perl M, Denk S, Weiss M, Schneider ME, Monk PN, et al. Changes and regulation of the c5a receptor on neutrophils during septic shock in humans. J Immunol 2013; 190(8):4215-4225.

Dreschers S, Gille C, Haas M, Grosse-Ophoff J, Schneider EM, Leiber A, Buhring HJ, Orlikowsky TW. Infection-induced bystander-apoptosis of monocytes is Tnf-alpha-mediated. PLoS One 2013; 8(1):e53589

Sahm, F, Capper D, Preusser M, Meyer J, Stenzinger A, Lasitschka F, Berghoff A-S, Habel A, Schneider EM, Kulozik A, AnagnostopoulosI, Müllauer L, Mechtersheimer G,von Deimling A. BRAFV600E mutant protein is expressed in cells of variable maturation in Langerhans Cell Histiocytosis. Blood. 2012; 120(12) e28-34.

Sunami Y, Leithauser F, Gul S, Fiedler K, Guldiken N, Espenlaub S, Holzmann KH, Hipp N, Sindrilaru A, Luedde T, et al. Hepatic activation of ikk/nfkappab signaling induces liver fibrosis via macrophage-mediated chronic inflammation. Hepatology 2012; 56(3):1117-1128.

Geistlinger J, Du, W., Groll, J, Liu, F, Hoegel, J, Foehr, KJ, Pasquarelli, A, Schneider, EM. P2RX7 genotype association in severe sepsis identified by a novel Multi-Individual Array for rapid screening and replication of risk SNPs. Clin Chim Acta. 2012, 413:39-47.

Hohn K, Sailer M, Wang L, Lorenz M, Schneider EM, Walther P. Preparation of cryofixed cells for improved 3d ultrastructure with scanning transmission electron tomography. Histochem Cell Biol 2011; 135(1):1-9

Schneider, EM, S. Flacke, F. Liu, M.R. Lorenz, P. Schilling, M.E. Nass, K.J. Foehr, M. Huber-Lang, and M.E. Weiss. Autophagy and ATP-induced anti-apoptosis in antigen presenting cells (APC) follows the cytokine storm in patients after major trauma. J Cell Commun Signal. 2011; 5:145-56.

Schreiner, L., M. Huber-Lang, M.E. Weiss, H. Hohmann, M. Schmolz, and E.M. Schneider. Phagocytosis and digestion of pH-sensitive fluorescent dye (Eos-FP) transfected E. coli in whole blood assays from patients with severe sepsis and septic shock. J Cell Commun Signal. 2011; 5:135-44.

Bindja, J, Weiss, ME, Schmolz, M, Stein,GM, Mapes, J, Schneiderhan-Marra, N, Joos, JO, Schneider, EM. Synthetic ligands against TLR2-9 in TruCultureTM - whole blood assays distinguish clinical stages of SIRS (trauma) and sepsis. Medimont Press, Proceedings Trauma, Shock, Inflammation and Sepsis, TSIS 2010, 55-63.

Schneider EM, Du W, Fiedler J, Högel J, Günther KP, Brenner H, Brenner RE. The (-765 G>C) promoter variant of the COX-2/PTGS2 gene is associated with a lower risk for end-stage hip and knee osteoarthritis. Ann Rheum Dis 2011, 70: 1458-1460.

Punyadeera C, Schneider, EM, Schaffer, D, Hsu, H-Y, Joos, TO, Kriebel, F, Weiss, ME, Verhaegh, W. A Biomarker Panel to Discriminate between SIRS and Sepsis and Sepsis Severity. Journal of Emergencies, Trauma, and Shock I 3:1 I Jan - Mar 2010

Weiss M, Taenzer M, Traeger K, Altherr J, Kron M, Hay B, Schneider M: Different patient case mix by applying the 2003 SCCM/ESICM/ACCP/ATS/SIS sepsis definitions instead of the 1992 ACCP/SCCM sepsis definitions in surgical patients: a retrospective observational study, BMC Med Inform Decis Mak 2009, 9:25

Buttenschoen K, Schneider EM, Utz K, Kornmann M, Beger HG, Carli Buttenschoen D. Effect of major abdominal surgery on endotoxin release and expression of Toll-like receptors 2/4. Langenbecks Arch Surg. 2009, 394:293-302.

Woehrle T, Du W, Goetz A, et al., Schneider, EM. Pathogen specific cytokine release reveals an effect of TLR2 Arg753Gln during Candida sepsis in humans. Cytokine. 2008, 41:322-329.

Hsu HY, Wittemann S, Schneider EM, Weiss M, Joos TO. Suspension microarrays for the identification of the response patterns in hyperinflammatory diseases. Med Eng Phys. 2008, 30:976-983.

Wittkowski, KM, Seybold, MP, Schneider, EM. Neue nichtparametrische Methoden und Tools zur Auswertung multivariater Daten in der klinischen Forschung und Diagnostik. Deutsche Zeitschrift für Klinische Forschung. 2008: 7/8:22-27

Collaborations

  • Dr. Thomas Joos, Dr. Nicole Schneiderhahn-Marra (Naturwissenschaftlich Medizinisches Institut Reutlingen, Universität Tübingen) „Clustering of Cytokinexpression patterns in patients with Trauma, Sepsis and Shock“. Peptide Ligand Assay develoments (Cathelicidines, defensins).
  • Dr. Manfred Schmolz (EDIgmbH, Reutlingen) Application and further developement of ex-vivo whole blood assays, TruCulture®.
  • Prof. Dr. Knut M. Wittkowski (Rockefeller University, Center for Clinical and Translational Science (CCTS), N.Y., USA) Biomarker-based clinical scores, genome wide association studies.
  • Kim, Hyung-Suk (NIH/NINR USA,  Bethesda, USA) GWAS in fibromyalgia.
  • Dr. Sandeep Kumar Vashist (IMTEK, Freiburg) Development of PoC devices.
  • Prof. Dr. Hsin-Yun HSU (Department of Applied Chemistry/Institute of Molecular Science National Chiao; Tung University,  Hsinchu 30010, Taiwan) Carbon nanotubes in immature dendritic cells.
Profilbild von Prof. Dr. rer. nat. Marion Schneider

Prof. Dr. rer. nat. Marion Schneider

Workgroup leader Experimental Anaesthesiology