Research projects

Project Details:
The aim of the work is to investigate relevant anatomical structures of the middle and inner ear of the peregrine falcon, hawk and common buzzard with the help of the µCT.

Project Details:
Gap junctions (Gj) are composed of two hemichannels called connexons, which are responsible for direct intercellular communication between adjoining cells. Each cell contains one connexon, composed of six connexin (cx) proteins. The so far discovered functions of Gj are mostly limited to the transport of molecules and ions (< 1 kDa). Nevertheless these junctions tend to be selective and specialize in cellular growth and differentiation (Bruzzone et al., 1996; Kumar and Gilula, 1996). As of today there are at least twenty different cx genes coding for Gj in humans and mice. One of the most researched Gj proteins is Cx43. In the testis, Cx43 is located between two Sertoli cells (SC) and between SC and germ cells (GC), it is involved in testicular development, GC and SC differentiation and spermatogenesis. SC nurture the developing GC and aid in their translocation and development from the basal to the adluminal compartment of the seminiferous tubule. Cx43 is further participating in blood-testis barrier formation and composition together with tight and adherens junctions as member of the "SC-SC junctional complexes" (Carrette et al., 2010).
A conditional SC-specific knockout (KO) of the Gja1 gene (SCCx43KO) revealed Cx43 expression in SC as an absolute requirement for normal testicular development and initiation of spermatogenesis (Brehm et al., 2007; Sridharan et al., 2007). Adult SCCx43KO mice showed normal testis descent, but testis size and weight was drastically lower when compared with heterozygous and WT littermates. Histological analysis revealed that SC specific deletion of Cx43 mostly results in an arrest of spermatogenesis at the level of spermatogonia or SC-only syndrome, intratubular cell clusters, abnormal SC cytoplasmic vacuoles, increased SC numbers and reduced number of spermatogonia per seminiferous tubule. Furthermore, as SCs were found to be still proliferating in adult mice (Sridharan et al., 2007), it was postulated that lack of Cx43 expression in SCs caused these somatic cells to remain in an apparently permanent proliferative state. These results emphasize the critical contribution of Cx43 to the normal maturational progression of SCs which normally results in the cessation of SC mitogenesis during the pubertal period.
Thus, as of today only one mouse stem with SCCx43KO-/- has been established in vivo (Brehm et al., 2007; Sridharan et al., 2007), yet no successful in vitro culturization of a &#61508;Cx43 SC line has been published. Although Carette et al. (2010) were successful in a partial inhibition of the Gj through siRNAs, a complete in vitro knockout may provide beneficial results in understanding the roles of Cx43 for normal spermatogenesis and may help to develop a mechanistic hypothesis in understanding the altered functions of Cx43 in SC leading to impaired spermatogenesis.

Project Details:
In diesem Projekt werden in einem transgenen Mausmodell die Folgen einer Sertoli Zell-spezifischen Deletion des Connexin43-Gens auf die Regulation der Spermatogenese u.a. mittels Microarray und qRT-PCR untersucht. Ausgewählte signifikant veränderte Kandidatengene aus den Mausversuchen werden zudem mit Hodenbiopsien von infertilen Männern verglichen.
Das Projekt bildet die Grundlage verschiedener Forschungsprojekte und Dissertationen sowie künftiger Drittmittelprojekte.
Geldgeber:
DFG, Klinische Forschergruppe KFO181, Male factor infertility due to impaired spermatogenesis
Volumen: 250000 ?

Results:

https://www.sciencedirect.com/science/article/pii/S1642431X17302371?via%3Dihub

Cooperation Partners:

Dr. Daniela Fietz, JLU Giessen

Klinische Forschergruppe Giessen + Marburg

http://www.uni-giessen.de/cms/fbz/fb11/forschung/forschergruppen/kfo_181/home

Funding: DFG (Heisenberg), 256.200 EUR

Project Details:
The objective of this research work is to evaluate the genetic architecture of body size using a pig model in a multi-omics approach. It particularly aims at in-depth investigations of interrelations of size determining variants, transcriptional variation among miniature and large size as well as the identification of topologically associated domains (TADs) and putative enhancers involved in body size determination. This study is particularly focusing on the elucidation of the regulatory landscape in pigs and its essential role in the determination of body size.Initial analyses of whole genome sequencing data of miniature versus standard sized breeds/populations are supposed to identify potential signatures of selection reflecting high selection pressures in both directions- miniature and large size- harboring variants causative for the miniaturization across-breeds. Subsequently, I aim at detecting chromatin interactions defined as TADs and putative enhancer elements in the region of these variants involved in size-determination by targeting high intensity peaks of chromatin interactions from Hi-C analysis as well as high histone modification levels associated with active enhancers (H3K27ac and H3K4me1) from ChIP-seq in the growth plates of the long bones. These marks of active DNA sequences will then be linked to transcriptional variation in-between miniature and large pigs. An in vitro model will be established for further functional validation.This will be the first study investigating genomic and functional effects on body size specifically targeting growth plates in pigs. Based on these data, we aim at increasing the knowledge of biological processes in mammals regulating growth and determining the size of a body. This profound understanding of body size development will not only be of high importance for livestock breeding but will also provide better understanding of growth biology, developmental genetics and disturbances in growth processes.

Results:

https://link.springer.com/article/10.1186/s12864-022-08801-4

Cooperation Partners:

Prof. Dr. Stefan Mundlos, Max-Planck-Institut für Molekulare Genetik, Berlin

Funding: DFG, 466.350 EUR

Project Details:
The objective of this research work is to evaluate the genetic architecture of body size using a pig model in a multi-omics approach. It particularly aims at in-depth investigations of interrelations of size determining variants, transcriptional variation among miniature and large size as well as the identification of topologically associated domains (TADs) and putative enhancers involved in body size determination. This study is particularly focusing on the elucidation of the regulatory landscape in pigs and its essential role in the determination of body size.Initial analyses of whole genome sequencing data of miniature versus standard sized breeds/populations are supposed to identify potential signatures of selection reflecting high selection pressures in both directions- miniature and large size- harboring variants causative for the miniaturization across-breeds. Subsequently, I aim at detecting chromatin interactions defined as TADs and putative enhancer elements in the region of these variants involved in size-determination by targeting high intensity peaks of chromatin interactions from Hi-C analysis as well as high histone modification levels associated with active enhancers (H3K27ac and H3K4me1) from ChIP-seq in the growth plates of the long bones. These marks of active DNA sequences will then be linked to transcriptional variation in-between miniature and large pigs. An in vitro model will be established for further functional validation.This will be the first study investigating genomic and functional effects on body size specifically targeting growth plates in pigs. Based on these data, we aim at increasing the knowledge of biological processes in mammals regulating growth and determining the size of a body. This profound understanding of body size development will not only be of high importance for livestock breeding but will also provide better understanding of growth biology, developmental genetics and disturbances in growth processes.

Results:

https://link.springer.com/article/10.1186/s12864-022-08801-4

Cooperation Partners:

Prof. Stefan Mundlos, Max-Planck-Institut für Molekulare Genetik, Berlin

Project Details:
In avian medicine, there are only a few methods available to assess peripheral blood flow. This study aims to evaluate passive IR thermography and µCT to assess blood flow in the skin in the area of &#8203;&#8203;the feet of birds of prey.

Results:

http://doi.org/10.1111/ahe.12369

Project Details:
In vorangegangenen Studien wurde die essentielle Bedeutung von Connexin 43 (Cx43) in Sertolizellen für die Spermatogenese bereits hervorgehoben. Da Untersuchungen darauf hindeuten, dass eine reziproke Regulation zwischen Keim- und Sertolizellen besteht und auch Keimzellen die Sertolizellfunktion beeinflussen können, stellt sich nun die Frage, ob ähnliche Effekte auftreten, wenn das Cx43-Gen nur in den Keimzellen ausgeknockt wird. Funktionelle Cx43-gap-junctions wurden u.a. in Spermatogonien und Spermatozyten nachgewiesen.
In diesem Projekt sollen die Auswirkungen der Deletion von Cx43 in Keimzellen auf die Hodenentwicklung und die Spermatogenese in transgenen Mäusen erforscht werden. Zu diesem Zweck wurden mittels des Cre/LoxP-Rekombinasesystems zwei konditionelle KO-Mauslinien generiert, denen das Cx43 entweder in den prämeoitischen oder in den postmeiotischen Keimzellpopulationen fehlt. Es werden Untersuchungen zur Morphologie/Ultrastruktur (Histologie/Elektronenmikroskopie) und Fertilität, zu Zell-Zell-Verbindungen und assoziierten Proteinen und Genen (IHC; WB, PCR) an männlichen KO-Tieren im Vergleich zum WT verschiedener Altersstufen durchgeführt und ggf. mit Fertilitätsstörungen des Mannes korreliert.

Results:

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8348783/

Cooperation Partners:

Prof. Dr. Martin Bergmann, JLU Gießen

Prof. Dr. Marc Mesnil, Universität Poitiers, Frankreich

Prof. Dr. Anaclet Ngezahayo, Leibniz Universität Hannover

Jon Oatley, PhD, College of Veterinary Medicine Washington State University, USA

Dr. Joachim Wistuba, Universität Münster

Project Details:
Für Erkrankungen der Fußsohle von Greifvögeln und auch Eulen scheint die Durchblutung eine entscheidende Rolle zu spielen. In diesen Untersuchungen sollen mit Hilfe von Korrosionspräparaten der Gefäße mit einer makroskopischer Auswertung mit Hilfe der Fotographie und Rasterelekronenmikroskopie sowie mit Hilfe von histologischen Untersuchungen die Mikrovascularisation der Fußsohle verschiedener Greifvögel- und Eulenarten untersucht und verglichen werden.

Project Details:
Connexin43 (Cx43) ist das dominierende Cx im Hoden und man findet es intratubulär in Gap junctions zwischen benachbarten Sertoli Zellen sowie zwischen Sertoli Zellen und einigen Keimzellen (Spermatogonien und primären Spermatozyten), aber auch zwischen interstitiellen Leydig-Zellen. Da ein generalisierter Knockout von Cx43 in Mäusen aufgrund einer Herz-Kreislauf-Missbildung zum perinatalen Tod der Tiere führt, wurde eine konditionale Cx43-Knockout-Mauslinie (SCCx43KO) mithilfe des Cre/LoxP-Rekombinasesystems generiert, bei der sich die Deletion des Cx43-Gens nur auf die Sertoli Zellen beschränkt. Dieser Sertoli Zell-spezifische Knockout (KO) hat gezeigt, dass die Tiere zwar lebensfähig, die männlichen Nachkommen (homozygote Männchen) aber infertil sind, da in 95% der Keimtubuli ein Arrest der Spermatogenese auf der Stufe der Spermatogonien oder ein Sertoli Cell-Only(SCO)-Syndrom ausgebildet ist. Weiterhin fiel in den KO-Tieren eine signifikante Erhöhung der Sertoli Zellzahl pro Keimtubulus auf, was auf eine veränderte Proliferationstätigkeit dieser Zellen hinweist. Weiterhin wurde vermutet, dass die eigentlich ab der Pubertät als postmitotische Zellen angesehenen Sertoli Zellen in dem transgenen Tiermodell auch nach Ende ihrer "terminalen Differenzierung" noch proliferieren können. Dies konnte bislang aber noch nicht mit den herkömmlichen Nachweisverfahren (z.B. Ki67 oder PCNA-Immunfärbungen) nachgeweisen werden. Es sind u.a. intraperitoneale Injektionen von BrdU geplant, um eine Prolifertaion der adulten Sertoli Zellen sicher nachzuweisen.

Project Details:
In diesem Projekt werden in einem transgenen Mausmodell die Folgen einer Sertoli Zell-spezifischen Deletion des Connexin45-Gens auf die Regulation der Spermatogenese u.a. mittels IHC und WB untersucht. Die zelluläre Lokalisation von Connexin45 im Hoden adulter Mäuse soll mittels Cx45EGFP-Mäusen bestätigt werden.

Cooperation Partners:

Frau PD Dr. Karin Dedek, Universität Oldenburg.