@article{HaarmannZimmermannBieberetal.2022, author = {Haarmann, Axel and Zimmermann, Lena and Bieber, Michael and Silwedel, Christine and Stoll, Guido and Schuhmann, Michael K.}, title = {Regulation and release of vasoactive endoglin by brain endothelium in response to hypoxia/reoxygenation in stroke}, series = {International Journal of Molecular Sciences}, volume = {23}, journal = {International Journal of Molecular Sciences}, number = {13}, issn = {1422-0067}, doi = {10.3390/ijms23137085}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-284361}, year = {2022}, abstract = {In large vessel occlusion stroke, recanalization to restore cerebral perfusion is essential but not necessarily sufficient for a favorable outcome. Paradoxically, in some patients, reperfusion carries the risk of increased tissue damage and cerebral hemorrhage. Experimental and clinical data suggest that endothelial cells, representing the interface for detrimental platelet and leukocyte responses, likely play a crucial role in the phenomenon referred to as ischemia/reperfusion (I/R)-injury, but the mechanisms are unknown. We aimed to determine the role of endoglin in cerebral I/R-injury; endoglin is a membrane-bound protein abundantly expressed by endothelial cells that has previously been shown to be involved in the maintenance of vascular homeostasis. We investigated the expression of membranous endoglin (using Western blotting and RT-PCR) and the generation of soluble endoglin (using an enzyme-linked immunosorbent assay of cell culture supernatants) after hypoxia and subsequent reoxygenation in human non-immortalized brain endothelial cells. To validate these in vitro data, we additionally examined endoglin expression in an intraluminal monofilament model of permanent and transient middle cerebral artery occlusion in mice. Subsequently, the effects of recombinant human soluble endoglin were assessed by label-free impedance-based measurement of endothelial monolayer integrity (using the xCELLigence DP system) and immunocytochemistry. Endoglin expression is highly inducible by hypoxia in human brain endothelial monolayers in vitro, and subsequent reoxygenation induced its shedding. These findings were corroborated in mice during MCAO; an upregulation of endoglin was displayed in the infarcted hemispheres under occlusion, whereas endoglin expression was significantly diminished after transient MCAO, which is indicative of shedding. Of note is the finding that soluble endoglin induced an inflammatory phenotype in endothelial monolayers. The treatment of HBMEC with endoglin resulted in a decrease in transendothelial resistance and the downregulation of VE-cadherin. Our data establish a novel mechanism in which hypoxia triggers the initial endothelial upregulation of endoglin and subsequent reoxygenation triggers its release as a vasoactive mediator that, when rinsed into adjacent vascular beds after recanalization, can contribute to cerebral reperfusion injury.}, language = {en} } @article{BellutRaimondiHaarmannetal.2022, author = {Bellut, Maximilian and Raimondi, Anthony T. and Haarmann, Axel and Zimmermann, Lena and Stoll, Guido and Schuhmann, Michael K.}, title = {NLRP3 inhibition reduces rt-PA induced endothelial dysfunction under ischemic conditions}, series = {Biomedicines}, volume = {10}, journal = {Biomedicines}, number = {4}, issn = {2227-9059}, doi = {10.3390/biomedicines10040762}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-267261}, year = {2022}, abstract = {Thrombolysis with recombinant tissue plasminogen activator (rt-PA) is a mainstay of acute ischemic stroke treatment but is associated with bleeding complications, especially after prolonged large vessel occlusion. Recently, inhibition of the NLRP3 inflammasome led to preserved blood-brain barrier (BBB) integrity in experimental stroke in vivo. To further address the potential of NLRP3 inflammasome inhibition as adjunct stroke treatment we used immortalized brain derived endothelial cells (bEnd5) as an in vitro model of the BBB. We treated bEnd5 with rt-PA in combination with the NLRP3 specific inhibitor MCC950 or vehicle under normoxic as well as ischemic (OGD) conditions. We found that rt-PA exerted a cytotoxic effect on bEnd5 cells under OGD confirming that rt-PA is harmful to the BBB. This detrimental effect could be significantly reduced by MCC950 treatment. Moreover, under ischemic conditions, the Cell Index — a sensible indicator for a patent BBB — and the protein expression of Zonula occludens 1 stabilized after MCC950 treatment. At the same time, the extent of endothelial cell death and NLRP3 expression decreased. In conclusion, NLRP3 inhibition can protect the BBB from rt-PA-induced damage and thereby potentially increase the narrow time window for safe thrombolysis in stroke.}, language = {en} } @article{GoebVollZimmermannetal.2021, author = {G{\"o}b, Vanessa and Voll, Maximilian G. and Zimmermann, Lena and Hemmen, Katharina and Stoll, Guido and Nieswandt, Bernhard and Schuhmann, Michael K. and Heinze, Katrin G. and Stegner, David}, title = {Infarct growth precedes cerebral thrombosis following experimental stroke in mice}, series = {Scientific Reports}, volume = {11}, journal = {Scientific Reports}, number = {1}, doi = {10.1038/s41598-021-02360-6}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-265791}, year = {2021}, abstract = {Ischemic stroke is among the leading causes of disability and death worldwide. In acute ischemic stroke, successful recanalization of occluded vessels is the primary therapeutic aim, but even if it is achieved, not all patients benefit. Although blockade of platelet aggregation did not prevent infarct progression, cerebral thrombosis as cause of secondary infarct growth has remained a matter of debate. As cerebral thrombi are frequently observed after experimental stroke, a thrombus-induced impairment of the brain microcirculation is considered to contribute to tissue damage. Here, we combine the model of transient middle cerebral artery occlusion (tMCAO) with light sheet fluorescence microscopy and immunohistochemistry of brain slices to investigate the kinetics of thrombus formation and infarct progression. Our data reveal that tissue damage already peaks after 8 h of reperfusion following 60 min MCAO, while cerebral thrombi are only observed at later time points. Thus, cerebral thrombosis is not causative for secondary infarct growth during ischemic stroke.}, language = {en} } @article{KuhtzSchneiderElHajjetal.2014, author = {Kuhtz, Juliane and Schneider, Eberhard and El Hajj, Nady and Zimmermann, Lena and Fust, Olga and Linek, Bartosz and Seufert, Rudolf and Hahn, Thomas and Schorsch, Martin and Haaf, Thomas}, title = {Epigenetic heterogeneity of developmentally important genes in human sperm: Implications for assisted reproduction outcome}, series = {Epigenetics}, volume = {9}, journal = {Epigenetics}, number = {12}, doi = {10.4161/15592294.2014.988063}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-150261}, pages = {1648-1658}, year = {2014}, abstract = {The molecular basis of male infertility is poorly understood, the majority of cases remaining unsolved. The association of aberrant sperm DNA methylation patterns and compromised semen parameters suggests that disturbances in male germline epigenetic reprogramming contribute to this problem. So far there are only few data on the epigenetic heterogeneity of sperm within a given sample and how to select the best sperm for successful infertility treatment. Limiting dilution bisulfite sequencing of small pools of sperm from fertile donors did not reveal significant differences in the occurrence of abnormal methylation imprints between sperm with and without morphological abnormalities. Intracytoplasmic morphologically selected sperm injection was not associated with an improved epigenetic quality, compared to standard intracytoplasmatic sperm injection. Deep bisulfite sequencing (DBS) of 2 imprinted and 2 pluripotency genes in sperm from men attending a fertility center showed that in both samples with normozoospermia and oligoasthenoteratozoospermia (OAT) the vast majority of sperm alleles was normally (de)methylated and the percentage of epimutations (allele methylation errors) was generally low (<1\%). However, DBS allowed one to identify and quantify these rare epimutations with high accuracy. Sperm samples not leading to a pregnancy, in particular in the OAT group, had significantly more epimutations in the paternally methylated GTL2 gene than samples leading to a live birth. All 13 normozoospermic and 13 OAT samples leading to a child had <1\% GTL2 epimutations, whereas one (7\%) of 14 normozoospermic and 7 (50\%) of 14 OAT samples without pregnancy displayed 1-14\% GTL2 epimutations.}, language = {en} } @article{SchuhmannLanghauserZimmermannetal.2023, author = {Schuhmann, Michael K. and Langhauser, Friederike and Zimmermann, Lena and Bellut, Maximilian and Kleinschnitz, Christoph and Fluri, Felix}, title = {Dimethyl fumarate attenuates lymphocyte infiltration and reduces infarct size in experimental stroke}, series = {International journal of molecular sciences}, volume = {24}, journal = {International journal of molecular sciences}, number = {21}, doi = {10.3390/ijms242115540}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-357731}, year = {2023}, abstract = {Ischemic stroke is associated with exacerbated tissue damage caused by the activation of immune cells and the initiation of other inflammatory processes. Dimethyl fumarate (DMF) is known to modulate the immune response, activate antioxidative pathways, and improve the blood-brain barrier (BBB) after stroke. However, the specific impact of DMF on immune cells after cerebral ischemia remains unclear. In our study, male mice underwent transient middle cerebral artery occlusion (tMCAO) for 30 min and received oral DMF (15 mg/kg) or a vehicle immediately after tMCAO, followed by twice-daily administrations for 7 days. Infarct volume was assessed on T2-weighted magnetic resonance images on days 1 and 7 after tMCAO. Brain-infiltrating immune cells (lymphocytes, monocytes) and microglia were quantified using fluorescence-activated cell sorting. DMF treatment significantly reduced infarct volumes and brain edema. On day 1 after tMCAO, DMF-treated mice showed reduced lymphocyte infiltration compared to controls, which was not observed on day 7. Monocyte and microglial cell counts did not differ between groups on either day. In the acute phase of stroke, DMF administration attenuated lymphocyte infiltration, probably due to its stabilizing effect on the BBB. This highlights the potential of DMF as a therapeutic candidate for mitigating immune cell-driven damage in stroke.}, language = {en} }