Research

Funding: QOL Med LLC,Vero Beach, Florida, USA; Unrestricted Research Grant , 230.000 EUR

Project Details:
Current concepts of genetically-determined carbohydrate malabsorption have implicated homozygous and compound heterozygous mutations in the gene encoding SI as the molecular basis for the onset of this intestinal disorder (Naim et al., J Pediatr Gastroenterol Nutr. 2012 Nov;55 Suppl 2:S13-20. review). The major pathogenic variants in CSID are: V577G in the isomaltase subunit and G1073D, F1745C and R1124X in the sucrase subunit, whereby the G1073D is the mutation with the highest prevalence among CSID patients. We have recently suggested that the symptoms and clinical phenotype in CSID follow a gradient of severity that is directly associated with the cellular trafficking patterns of SI mutants between the ER, Golgi and cell surface (Gericke et al., Biochim Biophys Acta. 2017 Mar;1863(3):817-826). Interestingly, some of the mutations analysed occur as heterozygotes in these patients. Along similar lines is the finding that 5 out of 11 patients in our study by Sander et al. (Hum Mutat. 2006 Jan;27(1):119.) occur as heterozygotes towards mutations of the SI gene and harbor mutations such as G1073D, F1745C and T694P.

Funding: DFG, 215.000 EUR

Project Details:
The early steps of hepatitis C virus (HCV) cell entry involve a complex interplay between the incoming virion and host cell expressed factors. Key host factors in the early phase of HCV cell entry include scavenger receptor class B member 1 (SR-BI), low-density lipoprotein receptor (LDLR) and CD81. Work from the previous funding period has revealed how genetic variation in one host factor, the high-density lipoprotein (HDL) receptor SR-BI, can modulate HCV infection, while highlighting at the same time that to fully understand the mechanisms of early HCV-cell interactions a broader view of the host cell molecules involved is required. In particular, others and we demonstrated that HCV host factors interact with each other and with cholesterol and this critically determines susceptibility. Moreover, we have identified CD81-binding partners calpain-5 (CAPN5) and casitas B-lineage lymphoma proto-oncogene b (CBLB) as novel modulators of HCV entry.
The proposed project for the next funding period has the overarching goal to better characterize how the host factors involved in the early phase of HCV entry interact with the virus and each other, and how these interactions are modulated by genetic and pharmacological factors. The project has four parts: (1) We will evaluate coding SR-BI and CD81 variants in a LDLR/SR-BI negative or - respectively - CD81 negative context for their effect on the HCV replication cycle and especially the early phase of cell entry in vitro. (2) We will determine how selected CD81 variants influence the interaction between CD81 and SR-BI and to what extent this mechanistically explains cell entry phenotypes. Here we will cooperate with project A6 (Pietschmann) which focusesl conversely on variation in SR-BI and membrane cholesterol content. (3) We will investigate how statins and other lipid lowering drugs including the new and highly potent monoclonal antibodies directed at proprotein convertase subtilisin/kexin type 9 (PCSK9), a key regulator of LDLR subcellular localization and function, impact HCV infection. Specifically, we will address the impact on the expression level of SR-BI, LDLR and CD81 and on membrane and cellular cholesterol content of hepatocytes as well as the impact of statin therapy on viral load and hepatic inflammation in patients with chronic hepatitis C. (4) Finally, we will extend our analysis to newly identified CD81 interaction partners CAPN5 and CBLB and their role in HCV cell entry and receptor complex regulation beginning with the generation of human variants to probe whether they impact CD81 and SR-BI interactions as well as HCV infection. Taken together, this work holds the promise of a deeper understanding of the mechanistic role of established and newly discovered HCV entry factors during the early phase of HCV cell entry, its modulation by host genetic variants and non-genetic host factors such as exposure to lipid metabolism-modulating drugs and thus of interindividual variability in the course of chronic hepatitis C.

Cooperation Partners:

Prof. Dr. Thomas von Hahn

Klinik für Gastroenterologie, Hepatologie und Endokrinologie und Institut für Molekularbiologie - Medizinische Hochschule Hannover

Funding: DFG Sonderforschungsbereich 900 "Mirkobielle Persistenz und seine Kontrolle", 215.000 EUR

Project Details:
The early steps of hepatitis C virus (HCV) cell entry involve a complex interplay between the incoming virion and host cell expressed factors. Key host factors in the early phase of HCV cell entry include scavenger receptor class B member 1 (SR-BI), low-density lipoprotein receptor (LDLR) and CD81. Work from the previous funding period has revealed how genetic variation in one host factor, the high-density lipoprotein (HDL) receptor SR-BI, can modulate HCV infection, while highlighting at the same time that to fully understand the mechanisms of early HCV-cell interactions a broader view of the host cell molecules involved is required. In particular, others and we demonstrated that HCV host factors interact with each other and with cholesterol and this critically determines susceptibility. Moreover, we have identified CD81-binding partners calpain-5 (CAPN5) and casitas B-lineage lymphoma proto-oncogene b (CBLB) as novel modulators of HCV entry.
The proposed project for the next funding period has the overarching goal to better characterize how the host factors involved in the early phase of HCV entry interact with the virus and each other, and how these interactions are modulated by genetic and pharmacological factors. The project has four parts: (1) We will evaluate coding SR-BI and CD81 variants in a LDLR/SR-BI negative or - respectively - CD81 negative context for their effect on the HCV replication cycle and especially the early phase of cell entry in vitro. (2) We will determine how selected CD81 variants influence the interaction between CD81 and SR-BI and to what extent this mechanistically explains cell entry phenotypes. Here we will cooperate with project A6 (Pietschmann) which focusesl conversely on variation in SR-BI and membrane cholesterol content. (3) We will investigate how statins and other lipid lowering drugs including the new and highly potent monoclonal antibodies directed at proprotein convertase subtilisin/kexin type 9 (PCSK9), a key regulator of LDLR subcellular localization and function, impact HCV infection. Specifically, we will address the impact on the expression level of SR-BI, LDLR and CD81 and on membrane and cellular cholesterol content of hepatocytes as well as the impact of statin therapy on viral load and hepatic inflammation in patients with chronic hepatitis C. (4) Finally, we will extend our analysis to newly identified CD81 interaction partners CAPN5 and CBLB and their role in HCV cell entry and receptor complex regulation beginning with the generation of human variants to probe whether they impact CD81 and SR-BI interactions as well as HCV infection. Taken together, this work holds the promise of a deeper understanding of the mechanistic role of established and newly discovered HCV entry factors during the early phase of HCV cell entry, its modulation by host genetic variants and non-genetic host factors such as exposure to lipid metabolism-modulating drugs and thus of interindividual variability in the course of chronic hepatitis C.

Results:

https://www.sfb900.de/en/

Cooperation Partners:

Prof. Dr. Thomas von Hahn, Klinik für Gastroenterologie, Hepatologie und Endokrinologie und Institut für Molekularbiologie - Medizinische Hochschule Hannover

Funding: Bundesministerium für Ernährung und Landwirtschaft, 682.563 EUR

Project Details:
Unter standardisierten Versuchsbedingungen mit Einzeltierhaltung werden Akzeptanz und Verdaulichkeit sowie Auswirkungen auf die Magen-Darm-Gesundheit junger Mastschweine geprüft, in deren Mischfutter die Komponente Weizen (ca. 70 %) in zunehmenden Anteilen durch Roggen ersetzt wurde. Damit wird eine vergleichende Bewertung der beiden Schwergetreidearten in der Mastschweinefütterung ermöglicht.

Cooperation Partners:

Institut für Tierernährung, Freie Universität Berlin

Institut für Tierernährung, Universität Bonn

KWS LOCHOW GmbH, Bergen/Wohlde

Project Details:
Das Brain natriuretic Peptide (BNP), auch als b-Typ natriuretisches Peptid bezeichnet, ist ein Hormon, das in der Herzmuskulatur gebildet und bei Dehnung der Herzkammern ins Blut abgegeben wird. Dieses Hormon besitzt wichtige Aufgaben in der Herz-Kreislaufregulation und wird in der Human- und Kleintiermedizin zur Diagnostik von Herzerkrankungen verwendet.
Das Vorkommen dieses Hormonones im Herzen von Papageien soll untersucht und eine Verwendung in der Diagnostik von Herzerkrankungen dieser Voegelarten überprüft werden.

Results:

https://www.mdpi.com/2306-7381/9/2/64

Project Details:
Untersuchungen zur Verbesserung der kulturellen Re-Isolierung des Erregers Mycoplasma hyopneumoniae aus Lungengewebeproben des Schweins mittels polyklonaler Antikörper und Verifizierung der Modifikationen in einem standardisierten Infektionsmodell.

Funding: Niedersächsische Tierseuchenkasse, Hannover, 72.750 EUR

Project Details:
Vereinzelt wurden im Blut von Milchkühen aus Betrieben Niedersachsens Hemoplasmen (Mykoplasma wenyonii, Cand Mykoplasma hemobos) sowie Hepaciviren nachgewiesen. Bislang sind Prävalenz, Bedeutung für die Milchviehgesundheitheit und Übertragungswege von Hemoplasmen nicht hinreichend bekannt. Auch sind die Übertragungswege von Hepaciviren nicht untersucht. Ziel der Studie ist daher die Prävalenz von Hemoplasmen und deren Bedeutung für die Gesundheit von Milchkühen abzuschätzen. Ferner soll geprüft werden, ob bei infizierten Tieren intrauterine Übertragungen von Hepaciviren und Hemoplasmen auf die Nachkommen vorkommen.

Cooperation Partners:

Dr. Bernd Hoffmann, Institut für Virusdiagnostik, Friedrich Loeffler Institut, Insel Riems

Dr. Mark Holsteg, Rindergesundheitsdienst Nordrhein-Westalen, Bad Sassendorf

Prof. Dr. Wolfgang Hölzle,Fakultät Agrarwissenschaften der Universität Hohenheim, Infektions- und Umwelthygiene bei Nutztieren, Stuttgart/Hohenheim

Funding: Ministry of Science and Culture of Lower Saxony, 186.057 EUR

Project Details:
The worldwide occurrence of resistant bacteria limits the efficiency of antibiotic-based treatment concepts. Therefore, new promising therapeutic approaches are needed, such as the strengthening of the host's defense by stimulating the immune system. Since the complex host-pathogen interactions are still poorly understood, detailed knowledge is required to apply therapeutic strategies based on the innate immune system. However, animal-free in vitro model systems for infection and interaction studies as well as for drug screenings are only a real alternative if the results obtained can be reliably transferred to the in vivo situation. However, due to lack of complexity, incorrect cell differentiation status, and lack of physiological conditions, in vitro systems do not sufficiently accurately simulate the in vivo situation during infection or inflammatory response.
The project aims to characterize the innate immune response with focus on lung epithelial cells against respiratory bacteria such as the human pathogen Staphylococcus aureus or the zoonotic pathogen Streptococcus suis in vitro and ex vivo. The complexity of the infection model is significantly increased by physiologically relevant oxygen conditions (defined hypoxic conditions, <10% O2 by means of a hypoxia glove box) and by 3D co-cultivation of human and porcine pulmonary epithelial cells and neutrophils.
This study will help to minimize false negative results in screening potential protective immunomodulators or antibiotics that are found to be effective in vitro but ineffective in vivo. Increased complexity of the model system also allows the in vitro system to approach the in vivo situation and will help to reduce the number of animals.

Results:

Influence of Oxygen on Function and Cholesterol Composition of Murine Bone Marrow-Derived Neutrophils.

Branitzki-Heinemann K, Brogden G, von Köckritz-Blickwede M.

Methods Mol Biol. 2020;2087:223-233. doi: 10.1007/978-1-0716-0154-9_17.

 

Differentiation and Functionality of Bone Marrow-Derived Mast Cells Depend on Varying Physiologic Oxygen Conditions.

Möllerherm H, Meier K, Schmies K, Fuhrmann H, Naim HY, von Köckritz-Blickwede M, Branitzki-Heinemann K.

Front Immunol. 2017 Nov 30;8:1665. doi: 10.3389/fimmu.2017.01665. eCollection 2017.

Cooperation Partners:

Prof. A. Bleich, PhD - MHH, Institut für Versuchstierkunde

Prof. Dr. M. Stiesch - MHH, Klinik für Zahnärztliche Prothetik und Biomedizinische Werkstoffkunde

Prof. Dr. Gerhard Breves - TiHo, Physiologisches Institut

Prof. Dr. Pablo Steinberg - TiHo, Institut für Lebensmitteltoxikologie und Chemische Analytik

Prof. Dr. med. Tim Sparwasser - Dr. Luciana Berod

Twincore, Zentrum für Experimentelle und Klinische Infektionsforschung GmbH, Institut für Infektionsimmunologie,

Dr. Jörn Tongers - MHH, Klinik für Kardiologie und Angiologie

Prof. Dr. Jürgen Wienands, Dr. Niklas Engels -

Universitätsmedizin Göttingen, Institut für Zelluläre & Molekulare Immunologie

Prof. A. Bleich, PhD - MHH, Institut für Versuchstierkunde

Prof. Dr. Tobias Cantz - MHH, Exzellenzcluster REBIRTH, Klinik für Gas-troenterologie, Hepatologie und Endokrinologie

Dr. Tanja Hansen- Fraunhofer ITEM, Klinische Chemie und ADME

Dr. Andres Hilfiker - MHH, Klinik für Herz, Thorax-, Transplantations- und Gefäßchirurgie, LEBAO

Prof. Dr. Ulrich Martin, Dr. Ruth Olmer

MHH, Klinik für Herz, Thorax-, Transplantations- und Gefäßchirurgie, LEBAO

Prof. Dr. Axel Schambach, PhD, Dr. Michael Rothe - MHH, Institut für Experimentelle Hämatologie

Prof. Dr. Wolfgang Baumgartner - TiHo, Institut für Pathologie

Prof. Dr. Albert Osterhaus - TiHo, Research Center for Emerging Infections and Zoonoses (RIZ)

Prof. Dr. M. von Köckritz-Blickwede - TiHo, Research Center for Emerging Infections and Zoonoses (RIZ)/Institut für Physiologische Chemie

Prof. Dr. Dr. Daniel Strech - MHH, Institut für Geschichte, Ethik und Philosophie der Medizin

Prof. Dr. Nils Hoppe - Leibniz Universität Hannover, Leibniz Forschungsinitiative CELLS: Centre for Ethics and Law in the Life Sciences (CELLS-LUH)

Prof. A. Bleich, PhD - MHH, Institut für Versuchstierkunde

Funding: BMBF, 1.351.062 EUR

Project Details:
Zoonotic flaviviruses, such as TBEV and JEV, can infect a number of different vertebrate hosts, but cause clinical disease only in some species while others remain unaffected. Moreover, many flaviviruses that are pathogenic in humans cause clinical symptoms in only a small fraction of infected individuals, of which only a portion will go on to develop severe complications, such as encephalitis, hemorrhagic disease or auto-immune disorders. The determinants of these dramatically different disease outcomes between host species and individuals from the same species are currently not well understood. Molecular mechanisms at the cellular level could play a role in the observed differences between susceptible host species. Advanced techniques, such as mRNA-Seq, real-time RT-PCR, and surface plasmon resonance will be applied for the comparative analysis of virus-host interactions at molecular and cellular levels in different host species to identify host-dependent factors that restrict or facilitate flavivirus infection.
Virus-specific antibodies play a major role in the protection from infection by neutralization of infectious viral particles or mediation of immune effector functions. However, virus-specific antibodies can also lead to the enhancement of flavivirus infections, especially in the case of secondary infections, and antibody-induced immune effector functions can contribute to the often observed immunopathogenesis in viral infections. Second-generation serological assays and functional immune effector studies can be used to measure individual antibody profiles against the complete viral proteome. Antibody profiles may vary between asymptomatic and symptomatic individuals and across different host species, and can contribute to the identification of biomarkers and predictors of disease outcome.

Project Details:
S. aureus is a multifaceted commensal organism and a potentially harmful human pathogen. It can cause a broad spectrum of infections, which can involve any organ. Beside toxin mediated diseases, it can also play a role as an intracellular pathogen. In vivo studies concern the association of S. aureus and gastrointestinal disorders focussed on the colonization of the gastrointestinal tract by S. aureus, and on the influence of staphylococcal toxins. In this study the influence of S. aureus infections on the function of human intestinal cells were investigated in an in vitro colon carcinoma (Caco)-2 cell model. It could be shown that S. aureus Newman wild type (WT) and the mutant strain S. aureus Newman Äeap exhibiting a lack of the extracellular adherence protein (Eap), were able to invade day 7 post-confluent Caco-2 cells followed by intracellular survival, persistence and replication, even though substantial less S. aureus Newman Äeap bacteria were invasive. Staphylococcal infection did not induce any cytotoxic effect observed by a membrane integrity test showing lactate-dehydrogenase (LDH)-release of infected cells remained unchanged compared to uninfected cells. This was in addition visualized microscopically with LIVE/DEAD Viability/Cytotoxicity Kit for mammalian cells after infection with S. aureus Newman WT expressing the green fluorescence protein (GFP). However, the function of the infected cells was altered: a decrease in enriched specific activity of sucrose from the marker glycoprotein human sucrase-isomaltase (hSI) was observable in the apical membrane fraction 48h after infection. Specific SI-activity was increased in the basolateral membrane fraction as well as decreased in the apical membrane fraction with no alterations in the catalytic capacity. Results obtained from this study suggest that infections with S. aureus could influence the sorting of hSI, leading to intestinal disorders followed by symptoms like diarrhea. Further investigations based on these results will give new insights in S. aureus gut infections and the association with gastrointestinal diseases, which could possibly enable therapeutic steps because of shedding light on the causes of symptoms.