Therese Eldh

Formation and maintenance of Alzheimer's disease beta-amyloid plaques in the absence of microglia

In Alzheimers disease, microglia cluster around β-amyloid deposits, suggesting that these cells are important for amyloid plaque formation, maintenance and/or clearance. We crossed two distinct APP transgenic mouse strains with CD11b-HSVTK mice, in which nearly complete ablation of microglia was achieved for up to 4 weeks after ganciclovir application. Neither amyloid plaque formation and maintenance nor amyloid-associated neuritic dystrophy depended on the presence of microglia.

New insights into the molecular pathology of radiation-induced pneumopathy.

BACKGROUND AND PURPOSE Pneumonitis and fibrosis constitute dose-limiting side effects of thorax or total body irradiation. An improved understanding of the underlying mechanisms is a prerequisite for the development of effective radioprotective strategies. Here we characterized the behavior of resident and immune cells in a murine model of radiation-induced pneumopathy. MATERIALS AND METHODS Wild type (WT) or RAG-2 deficient C57BL/6 mice received 15 Gray of (hemi)-thorax irradiation in a single dose. Bronchoalveolar lavage fluid (BALF) and lung tissue were collected at defined time points post-irradiation for the determination of apoptosis, microvascular injury, and histological and immunohistochemical analyses. RESULTS Higher albumin levels and increased apoptosis were detected in the BALF 21 days after irradiation, indicative for delayed damage to resident cells. Irradiation also induced time-dependent changes in the BALF cytokine profile, the recruitment of activated T-cells into the lung and the formation of lipid-loaded resident cells. Lung fibrosis occurred earlier in RAG-2(-/-) mice, which lack mature T and B cells, compared to WT mice. CONCLUSIONS Thorax irradiation triggers a delayed disturbance of tissue integrity and lipid metabolism in the lung. Activated T-lymphocytes infiltrating the lung tissue upon thorax irradiation participate in the protection of the lung from radiation-induced fibrosis.

Inflammation-associated repression of vasodilator-stimulated phosphoprotein (VASP) reduces alveolar-capillary barrier function during acute lung injury

Acute lung injury (ALI) is an inflammatory disorder associated with reduced alveolar‐capillary barrier function, increased pulmonary vascular permeability, and infiltration of leukocytes into the alveolar space. Pulmonary function might be compromised, its most severe form being the acute respiratory distress syndrome. A protein central to physiological barrier properties is vasodilator‐stimulated phosphoprotein (VASP). Given the fact that VASP expression is reduced during periods of cellular hypoxia, we investigated the role of VASP during ALI. Initial studies revealed reduced VASP expressional levels through cyto‐kines in vitro. Studies in the putative human VASP promoter identified NF‐κB as a key regulator of VASP transcription. This VASP repression results in increased paracellular permeability and migration of neutrophils in vitro. In a model of LPS‐induced ALI, VASP −/− mice demonstrated increased pulmonary damage compared with wild‐type animals. These findings were confirmed in a second model of ventilator‐induced lung injury. Studies employing bone marrow chimeric animals identified tissue‐specific repression of VASP as the underlying cause of decreased barrier properties of the alveolar‐capillary barrier during ALI. Taken together these studies identify tissue‐specific VASP as a central protein in the control of the alveolar‐capillary barrier properties during ALI.—Henes, J., Schmit, M. A., Morote‐Garcia, J. C., Mirakaj, V., Kohler, D., Glover, L., Eldh, T., Walter, U., Karhausen, J., Colgan, S. P., Rosenberger, P. Inflammation‐associated repression of vasodilator‐stimulated phosphoprotein (VASP) reduces alveolar‐capillary barrier function during acute lung injury. FASEB J. 23, 4244‐4255 (2009). www.fasebj.org

Gαi2- and Gαi3-Deficient Mice Display Opposite Severity of Myocardial Ischemia Reperfusion Injury

G-protein-coupled receptors (GPCRs) are the most abundant receptors in the heart and therefore are common targets for cardiovascular therapeutics. The activated GPCRs transduce their signals via heterotrimeric G-proteins. The four major families of G-proteins identified so far are specified through their α-subunit: Gαi, Gαs, Gαq and G12/13. Gαi-proteins have been reported to protect hearts from ischemia reperfusion injury. However, determining the individual impact of Gαi2 or Gαi3 on myocardial ischemia injury has not been clarified yet. Here, we first investigated expression of Gαi2 and Gαi3 on transcriptional level by quantitative PCR and on protein level by immunoblot analysis as well as by immunofluorescence in cardiac tissues of wild-type, Gαi2-, and Gαi3-deficient mice. Gαi2 was expressed at higher levels than Gαi3 in murine hearts, and irrespective of the isoform being knocked out we observed an up regulation of the remaining Gαi-protein. Myocardial ischemia promptly regulated cardiac mRNA and with a slight delay protein levels of both Gαi2 and Gαi3, indicating important roles for both Gαi isoforms. Furthermore, ischemia reperfusion injury in Gαi2- and Gαi3-deficient mice exhibited opposite outcomes. Whereas the absence of Gαi2 significantly increased the infarct size in the heart, the absence of Gαi3 or the concomitant upregulation of Gαi2 dramatically reduced cardiac infarction. In conclusion, we demonstrate for the first time that the genetic ablation of Gαi proteins has protective or deleterious effects on cardiac ischemia reperfusion injury depending on the isoform being absent.

Phosphorylation of Vasodilator-Stimulated Phosphoprotein (VASP) Dampens Hepatic Ischemia-Reperfusion Injury

Recent work has demonstrated that the formation of platelet neutrophil complexes (PNCs) affects inflammatory tissue injury. Vasodilator-stimulated phosphoprotein (VASP) is crucially involved into the control of PNC formation and myocardial reperfusion injury. Given the clinical importance of hepatic IR injury we pursued the role of VASP during hepatic ischemia followed by reperfusion. We report here that VASP −/− animals demonstrate reduced hepatic IR injury compared to wildtype (WT) controls. This correlated with serum levels of lactate dehydrogenase (LDH), aspartate (AST) and alanine (ALT) aminotransferase and the presence of PNCs within ischemic hepatic tissue and could be confirmed using repression of VASP through siRNA. In studies employing bone marrow chimeric mice we identified hematopoietic VASP to be of crucial importance for the extent of hepatic injury. Phosphorylation of VASP on Ser153 through Prostaglandin E1 or on Ser235 through atrial natriuretic peptide resulted in a significant reduction of hepatic IR injury. This was associated with a reduced presence of PNCs in ischemic hepatic tissue. Taken together, these studies identified VASP and VASP phosphorylation as crucial target for future hepatoprotective strategies.

Repression of the Equilibrative Nucleoside Transporters Dampens Inflammatory Lung Injury

Acute lung injury (ALI) is a devastating disorder of the lung that is characterized by hypoxemia, overwhelming pulmonary inflammation, and a high mortality in the critically ill. Adenosine has been implicated as an anti-inflammatory signaling molecule, and previous studies showed that extracellular adenosine concentrations are increased in inflamed tissues. Adenosine signaling is terminated by the uptake of adenosine from the extracellular into the intracellular compartment via equilibrative nucleoside transporters (ENTs). However, their role in controlling adenosine signaling during pulmonary inflammation remains unknown. After inflammatory in vitro experiments, we observed a repression of ENT1 and ENT2 that was associated with an attenuation of extracellular adenosine uptake. Experiments using short, interfering RNA silencing confirmed a significant contribution of ENT repression in elevating extracellular adenosine concentrations during inflammation. Furthermore, an examination of the ENT2 promoter implicated NF-κB as a key regulator for the observed ENT repression. Additional in vivo experiments using a murine model of inflammatory lung injury showed that the pharmacological inhibition of ENT1 and ENT2 resulted in improved pulmonary barrier function and reduced signs of acute inflammation of the lung. Whereas experiments on Ent1(-/-) or Ent2(-/-) mice revealed lung protection in LPS-induced lung injury, an examination of bone marrow chimeras for ENTs pointed to the nonhematopoetic expression of ENTs as the underlying cause of dampened pulmonary inflammation during ALI. Taken together, these findings reveal the transcriptional repression of ENTs as an innate protective response during acute pulmonary inflammation. The inhibition of ENTs could be pursued as a therapeutic option to ameliorate inflammatory lung injury.

Abstract 79: Immunomodulation in response to radiation-induced lung injury.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Introduction. Pneumonitis and fibrosis constitute dose-limiting side effects of thorax and whole body irradiation. Although several novel disease biomarkers have been described during recent years, the cellular and molecular factors responsible for disease initiation and progression are still not sufficiently understood. Here, we characterized local and systemic changes in the immune cell compartment in response to radiation-induced lung injury and defined the role of lymphocytes for fibrosis development in a murine model. Material and methods A single dose of hemithorax or whole thorax irradiation (15 Gy) was applied on C57BL/6 wild type mice (WT) or immunodeficient RAG-2-/- mice. Mice were sacrificed at defined time points and lungs were isolated for histological and immunohistochemical analysis as well as immune cell phenotyping. Moreover, immune cells were isolated from spleen, blood and cervical lymph nodes and also characterized by flow cytometry. Bronchioalveolar lavage fluid was collected for determination of cytokine levels. Results and discussion. Whole thorax irradiation triggered time-dependent changes in the composition and activation state of immune cells isolated from the lung tissue. These changes were paralleled by specific alterations in the cytokines present in the bronchoalveolar lavage fluid. In WT mice whole thorax irradiation with 15 Gy led to a time-dependent increase in collagen deposition and lung fibrosis. Irradiated RAG-2-/- mice showed an earlier onset of collagen deposition and an increased number of fibrotic foci in the lung when compared to WT mice. Conclusion. Thorax irradiation triggers local and systemic changes in the composition and the activation state of immune cells. The increased sensitivity of RAG-2-/- mice to radiation-induced fibrosis suggests a role of mature lymphocytes in the suppression of fibrotic changes. An improved understanding of the contribution of specific immune cells to the pathogenic process in the lung may open novel routes to prevent or treat radiation-induced pneumopathy. Citation Format: Federica Cappuccini, Florian Wirsdorfer, Malagorzata Drabczyk, Therese Eldh, Ali Sak, Verena Jendrossek. Immunomodulation in response to radiation-induced lung injury. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 79. doi:10.1158/1538-7445.AM2013-79