Department of Obstetrics and Gynecology, Medical University of Graz, Auenbruggerplatz 14, A-8036 Graz;
phone: +43-316-385 84605, fax: +43-316-385 12506, ✉ e-mail
Inflammation, obesity, trophoblast, oxidative stress, DNA damage, growth, cell cycle, placenta
Early human placental (and embryonic) development occurs in a physiologically low oxygen environment supported by histiotrophic secretions from endometrial glands. The low oxygen tension is the result of trophoblasts plugging the spiral arteries such that only a serum filtrate can reach the intervillous space. Under normal conditions these spiral arteries will only get opened after anti-oxidative defense systems have been adequately developed in the placenta. This will prevent that the oxidative stress associated with the gradual opening of the spiral arteries and the ensuing increase in oxygen tension in the intervillous space will compromise the development of the placenta. Conditions such as pre-eclampsia, maternal diabetes or obesity may put these anti-oxidative defense systems under excessive strain through the accompanying hyperglycemia and / or pro-inflammatory state. In pre-eclampsia the growth arrest and DNA damage-inducible 45 (GADD45) protein, a central stress sensor, is overexpressed and influences trophoblast activity through the p38 MAP kinase pathway (1). It may, thus, also be involved on placental growth impairment found in pre-eclampsia.
One focus of the laboratory has been to understand the effects of maternal diabetes and obesity on placental development and trophoblast proliferation (2). We analysed the first trimester trophoblast transcriptome under normo- and hyperglycemic conditions and found most dysregulated transcripts in the cell-cycle / apoptosis / transcription cluster (Maier A, Hiden U and Desoye G, unpublished). We have further demonstrated that the combination of hyperglycemia and high oxygen levels reduces proliferation of human first-trimester trophoblasts involving MAP kinase (3). This may account for reduced placental growth and, therefore, also for reduced embryonic growth, during the first-trimester of pregestational diabetic or hyperglycemic pregnancies, when the oxygen tension increases. Since inflammation and oxidative stress can also induce DNA damage, which may result in reduced proliferation, we hypothesize that obesity-associated enhanced inflammation early in pregnancy leads to DNA damage of the trophoblasts with ensuing cell cycle arrest until single-strand breaks have been repaired. We further hypothesize that GADD45 plays a crucial role in this process by integrating various input signals (e. g. inflammatory cytokines, changing oxygen levels, oxidative stress) and orchestrating the trophoblast response (e. g. cell cycle arrest, DNA repair) (4). We will dissect the sequence of these events and take into account the sex of the fetus, which may show different stress susceptibility and/or response to environmental stimuli.
Fig. 2: Fluorescence microscopy of mitochondrial superoxide levels in a first-trimester trophoblast model after 3 days under hyperglycemia (HG)
M. Gauster will instruct the students how to work with placental explants from first trimester of pregnancy.
A. Heinemann will help the students to carefully and comprehensively characterize the first trimester trophoblast isolations by FACS analysis.
DANONE nutrition, NL.
The laboratory of Gernot Desoye has more than 30 years expertise in the field of placental metabolism, growth and development in conditions of maternal overnutrition (diabetes, obesity). In addition to having established procedures to isolate primary trophoblasts from first-trimester human placentas, the laboratory is among the very few worldwide, in which several primary cells are available from term placentas (trophoblasts, macrophages, Tregs, endothelial cells). The laboratory has also generated and fully characterized a first-trimester trophoblast cell line, which is particularly suitable for studies into cell-cycle regulation and DNA damage using overexpression and silencing. Various techniques are being used for a detailed functional analysis of these cells in vitro including proliferation / cell cycle and DNA damage assays. All equipment, which is not available at the Department’s research laboratory, such as flow cytometry, real-time PCR systems, microarray and sequencing facilities, ECIS and multiplexing are available either at the collaboration partners or at the core facilities of the clinical research center.
The DP-iDP students will get background knowledge in early human development and will learn about the effect of diabetes and obesity on this process. They will be taught in placental functions relevant for adequate growth of the human fetus with special emphasis on nutrition and maintenance of placental integrity as well as the different growth and survival strategies of female and male fetuses. The students will learn a wide range of techniques beyond conventional molecular biology and biochemistry techniques. Cell culture work with human primary cells and tissue explants will be a strong focus allowing analyses of DNA repair, cell cycle, apoptosis and proliferation by various methods. The knowledge will be applied to develop strategies on how to delineate signaling pathways with emphasis on stress–response pathways (p38, JNK). The student will use immunohistochemistry combined with morphometry to determine changes associated with graded maternal overweight / obesity on structural components of the placenta as well as on target molecules relevant in stress response, DNA repair and cell-cycle regulation. Signalling pathways will be blocked in the tissue explants and trophoblasts using pharmacological inhibitors. Expression and protein levels GADD45 will be quantified in placental explants of mothers with various degrees of overweight / obesity and silenced / overexpressed to establish its key function as stress sensor and inducer of the DNA repair and cell cycle response.