Critical Role of Methylglyoxal and AGE in Mycobacteria-Induced Macrophage Apoptosis and Activation

Apoptosis and activation of macrophages play an important role in the host response to mycobacterial infection involving TNF-α as a critical autocrine mediator. The underlying mechanisms are still ill-defined. Here, we demonstrate elevated levels of methylglyoxal (MG), a small and reactive molecule that is usually a physiological product of various metabolic pathways, and advanced glycation end products (AGE) during mycobacterial infection of macrophages, leading to apoptosis and activation of macrophages. Moreover, we demonstrate abundant AGE in pulmonary lesions of tuberculosis (TB) patients. Global gene expression profiling of MG-treated macrophages revealed a diverse spectrum of functions induced by MG, including apoptosis and immune response. Our results not only provide first evidence for the involvement of MG and AGE in TB, but also form a basis for novel intervention strategies against infectious diseases in which MG and AGE play critical roles

Representation of the Internet-based lecture in veterinary obstetrics and the investigation of this application regarding the acceptance and integration into the study of veterinary medicine

Die komplette Dissertation im pdf-Format (829.434 Bytes): rothe.pdfThe overall objective of the interactive Internet-based lecture in veterinary obstetrics was to make teaching more effective both for the lecturer and the students. Specific objectives were to save the time needed to deal with questions regarding factual knowledge and to gain time for dealing with current subjects. Through the interactive lecture we wanted to facilitate repetition and preparation of certain subjects at any time. The students have unrestricted access to the subject matter and to the slides and pictures used in the traditional lecture. Furthermore an attempt should be made to increase the motivation for self-study and to lay a foundation for a medical-oriented thinking. The interactive Internet-based lecture in veterinary obstetrics was founded on the structure and the contents of the traditional lecture in the auditorium. The scientific content is based on the lecture-notes and relevant literature. The lecture contains 600 pages and includes about 500 slides, pictures, graphs and animations. By the means of an evaluation the acceptance and usefulness of the interactive lecture was tested. 152 of 201 students of the third and fourth year took part in the survey and filled out an anonymous questionnaire. These two years were chosen specifically because the students in the third year attended the lecture in veterinary obstetrics and the fourth year students had to participate at the practical course. The students were required to use and to interact with the lecture at least once in the department´s computer laboratory. However they could choose the date. The questionnaire included general questions related to their attitudes towards computer and Internet. Another set of questions aimed to determine the perceptions of students about the interactive lecture in particular and the use of such tools in general. The evaluation showed that the veterinary students used the computer or considered it as a learning tool in a limited way even though they judged the interactive lecture as good and accepted it. The continual utilisation also for the revision for the exam was proven. For the students it was important to have the opportunity to choose between the lecture in the auditorium, the use of Internet, CD-ROM or the printed version. However, the exclusive presentation on the Internet was not accepted very well. Also the integration of the interactive lecture in the auditorium turned out to be complicated. The long term success of this concept depends on the level of rethinking of the lecturers and the students, new curriculum guidelines of the study and the technical equipment of the universities

EphrinA2 Receptor (EphA2) Is an Invasion and Intracellular Signaling Receptor for Chlamydia trachomatis

The obligate intracellular bacterium Chlamydia trachomatis invades into host cells to replicate inside a membrane-bound vacuole called inclusion. Multiple different host proteins are recruited to the inclusion and are functionally modulated to support chlamydial development. Invaded and replicating Chlamydia induces a long-lasting activation of the PI3 kinase signaling pathway that is required for efficient replication. We identified the cell surface tyrosine kinase EphrinA2 receptor (EphA2) as a chlamydial adherence and invasion receptor that induces PI3 kinase (PI3K) activation, promoting chlamydial replication. Interfering with binding of C. trachomatis serovar L2 (Ctr) to EphA2, downregulation of EphA2 expression or inhibition of EphA2 activity significantly reduced Ctr infection. Ctr interacts with and activates EphA2 on the cell surface resulting in Ctr and receptor internalization. During chlamydial replication, EphA2 remains active accumulating around the inclusion and interacts with the p85 regulatory subunit of PI3K to support the activation of the PI3K/Akt signaling pathway that is required for normal chlamydial development. Overexpression of full length EphA2, but not the mutant form lacking the intracellular cytoplasmic domain, enhanced PI3K activation and Ctr infection. Despite the depletion of EphA2 from the cell surface, Ctr infection induces upregulation of EphA2 through the activation of the ERK pathway, which keeps the infected cell in an apoptosis-resistant state. The significance of EphA2 as an entry and intracellular signaling receptor was also observed with the urogenital C. trachomatis-serovar D. Our findings provide the first evidence for a host cell surface receptor that is exploited for invasion as well as for receptor-mediated intracellular signaling to facilitate chlamydial replication. In addition, the engagement of a cell surface receptor at the inclusion membrane is a new mechanism by which Chlamydia subverts the host cell and induces apoptosis resistance

Treatment of high fat diet-induced obese pregnant mice with IL-6 receptor antibody does not ameliorate placental function and fetal growth restriction

Problem : Pregnancy complications and adverse birth outcomes are in part fueled by the rise in obesity and its associated co-morbidities in western societies. Fetal healthy development and placental function are disturbed by an obese, inflammatory environment associated with cytokines, such as interleukin-6, causing inadequate supply of nutrients to the fetus and perinatal programming with severe health consequences. Method of Study : Mice received high fat diet (HFD) before and during gestation to induce obesity. We performed an IL-6 receptor antibody (MR16-1) treatment in pregnant obese mice at embryonic days E0.5, E7.5 and E14.5 to investigate whether this could ameliorate HFD-induced and obesity-associated placental dysfunction, evaluated by stereology and western blot, and improve offspring outcome at E15.5 in obese dams. Results : We observed fewer fetuses below the 10th percentile and placental vascularization was less aggravated following MR16-1 treatment of obese dams, showing slight improvements in labyrinth zone (Lz) vascularization. However, placental dysfunction and fetal growth restriction were still apparent in MR16-1 dams compared to lean control dams. Molecular analysis showed significantly elevated IL-6 level in placentas of MR16-1 treated dams. Conclusion : Treatment with MR16-1 blocks IL-6 signaling in the placenta, but has only limited effects on preventing HFD-associated placental dysfunction and offspring outcomes in mice, suggesting further mechanisms in the deterioration of placental vascularization and fetal nutrient supply as a consequence of maternal obesity

A Potential Role for GSK3 beta in Glucose-Driven Intrauterine Catch-Up Growth in Maternal Obesity

Obesity and unhealthy nutrition are increasing and affect women of childbearing age and hence during pregnancy. Despite normal or even high birth weight, the offspring suffers from long-term metabolic risks. We hypothesized that fetal growth is disturbed during different intrauterine phases. Underlying molecular events remain elusive. Female mice were fed either a standard diet (SD) or a high-fat diet (HFD) after weaning until mating and during pregnancy. Pregnant mice were euthanized at gestational day (G) 15.5 and G18.5, and fetuses and placentas were removed for analysis. HFD fetuses displayed intrauterine growth restriction (IUGR) at G15.5, which disappeared until G18.5, indicating intrauterine catch-up growth during that time period. Main placental findings indicate decreased canonical Wnt-GSK3 beta signaling and lower proliferation rates at G18.5, which goes along with a smaller placental transfer zone. On the other hand, glucose depots (glycogen cluster) in HFD placentas decreased more strongly between G15.5 and G18.5 compared with placentas from SD mothers, and the glucose transporter protein GLUT-1 was increased at G18.5 in the HFD group. Maternal diet-induced obesity causes an IUGR phenotype at the beginning of the third week (G15.5) in our mouse model. This phenotype is reversed by the end of the third week (G18.5) despite a smaller placental transfer zone, probably based on GSK3 beta-mediated increased glucose mobilization in the placenta and hence an increased glucose supply to the fetus

Appel et al 2018_supplemental material

Supplemental material for publication 'A potential role for GSK3beta in glucose-driven intrauterine catch-up growth in maternal obesity'

Data from: A potential role for GSK3beta in glucose-driven intrauterine catch-up growth in maternal obesity

Obesity and an unhealthy nutrition are on the rise and affect also women in childbearing age and hence, during pregnancy. Despite normal or even high birth weight the offspring suffers from long term metabolic risks. We hypothesized that fetal growth is disturbed during different intrauterine phases. Underlying molecular events remain elusive. Female mice were fed either a control diet (SD) or a high fat diet (HFD) after weaning until mating and during pregnancy. Pregnant mice were sacrificed at gestational time points G15.5 and G18.5 and fetuses and placentas were removed for analysis. HFD fetuses displayed intrauterine growth restriction (IUGR) at G15.5, which disappeared until G18.5, indicating an intrauterine catch-up growth during that time period. Main placental findings indicate decreased canonical Wnt-GSK3beta signaling and lower proliferation rates at G18.5 which goes along with a smaller placental transfer zone. On the other hand, glucose depots (glycogen cluster) in HFD placentas decreased stronger between G15.5 and G18.5 compared to placentas from SD mothers, and the glucose transporter protein GLUT-1 was increased at G18.5 in the HFD group. Maternal diet-induced obesity causes an IUGR phenotype at the beginning of the third week (G15.5) in our mouse model. This phenotype is reversed by the end of the third week (G18.5) despite of a smaller placental transfer zone, probably based on GSK3beta-mediated increased glucose mobilization in the placenta and hence an increased glucose supply to the fetus

Chlamydia infection depends on a functional MdM2-p53 axis

Chlamydia, a major human bacterial pathogen, assumes effective strategies to protect infected cells against death-inducing stimuli, thereby ensuring completion of its developmental cycle. Paired with its capacity to cause extensive host DNA damage, this poses a potential risk of malignant transformation, consistent with circumstantial epidemiological evidence. Here we reveal a dramatic depletion of p53, a tumor suppressor deregulated in many cancers, during Chlamydia infection. Using biochemical approaches and live imaging of individual cells, we demonstrate that p53 diminution requires phosphorylation of Murine Double Minute 2 (MDM2; a ubiquitin ligase) and subsequent interaction of phospho-MDM2 with p53 before induced proteasomal degradation. Strikingly, inhibition of the p53-MDM2 interaction is sufficient to disrupt intracellular development of Chlamydia and interferes with the pathogen's anti-apoptotic effect on host cells. This highlights the dependency of the pathogen on a functional MDM2-p53 axis and lends support to a potentially pro-carcinogenic effect of chlamydial infection

Chlamydia infection depends on a functional MDM2-p53 axis.

Chlamydia, a major human bacterial pathogen, assumes effective strategies to protect infected cells against death-inducing stimuli, thereby ensuring completion of its developmental cycle. Paired with its capacity to cause extensive host DNA damage, this poses a potential risk of malignant transformation, consistent with circumstantial epidemiological evidence. Here we reveal a dramatic depletion of p53, a tumor suppressor deregulated in many cancers, during Chlamydia infection. Using biochemical approaches and live imaging of individual cells, we demonstrate that p53 diminution requires phosphorylation of Murine Double Minute 2 (MDM2; a ubiquitin ligase) and subsequent interaction of phospho-MDM2 with p53 before induced proteasomal degradation. Strikingly, inhibition of the p53-MDM2 interaction is sufficient to disrupt intracellular development of Chlamydia and interferes with the pathogen's anti-apoptotic effect on host cells. This highlights the dependency of the pathogen on a functional MDM2-p53 axis and lends support to a potentially pro-carcinogenic effect of chlamydial infection