Biology subjects

Steffens, S.

Publications and source records attributed to Steffens, S..

3 recordsLinked to original sources

bioRxiv2022

Physiological alterations in microglial morphology associate with the sleep-wake cycle in a brain region-specific manner

Long-term total sleep deprivation induces changes in cortical - and hippocampal microglial morphology that closely resemble the microglial response to the gram-negative bacterial cell wall component lipopolysaccharide (LPS). A recent study found evidence that microglia could modify vigilance-states/sleep, but only few studies investigated microglial throughout the diurnal behavioral inactivity/activity pattern or the naturally occurring sleep-wake cycle, and those who have, only concentrated on the cortical or hippocampal microglia. As microglia demonstrate regional heterogeneity, we compared microglial diurnal morphological alterations in the somatosensory cortex (SC) and dorsal hippocampus (HC) to the basal forebrain (BF), which is a subcortical brain area involved in the regulation of vigilance states. We collected mouse brain samples every 3h throughout the 24h light-dark-cycle and applied a 3D reconstruction method for the acquired confocal microscopy images for each brain area separately. While microglial regional heterogeneity was evident, stimulation of microglia with LPS caused comparable microglial responses in all brain areas. When comparing microglial features between the 12h light- and dark periods, regional heterogeneity re-appeared. As most of the morphological alterations occurred during the light period-the habitual sleeping period of the mice, we performed polysomnography to study the possible interaction of microglial morphology and sleep. We found that cortical-, but not HC- or BF microglial territory and volume negatively correlated with sleep slow wave activity (SWA), an electroencephalic feature of non-REM sleep (NREMS). Since microglia are sensitive to neuronal activity, we propose that the regional differences reflect vigilance-state specific neuronal activity patterns. Table of contents image O_FIG O_LINKSMALLFIG WIDTH=184 HEIGHT=200 SRC="FIGDIR/small/482976v1_ufig1.gif" ALT="Figure 1"> View larger version (71K): org.highwire.dtl.DTLVardef@112d346org.highwire.dtl.DTLVardef@ecc138org.highwire.dtl.DTLVardef@1817331org.highwire.dtl.DTLVardef@1b1b47c_HPS_FORMAT_FIGEXP M_FIG C_FIG Main PointsMicroglia show morphological differences between the somatosensory cortex (SC), hippocampus (HC) and basal forebrain (BF) under physiological conditions. Cortical-, but not HC- or BF microglial cell volume negatively correlates with non-REM sleep slow wave activity.

neuroscience↗

bioRxiv2021

GPR55 in B cells limits atherosclerosis development and regulates plasma cell maturation

Identifying novel pathways regulating the adaptive immune response in chronic inflammatory diseases such as atherosclerosis is of particular interest in view of developing new therapeutic drugs. Here we report that the lipid receptor GPR55 is highly expressed by splenic B cells and inversely correlates with atheroma plaque size in mice. In human carotid endarterectomy specimen, GPR55 transcript levels were significantly lower in unstable compared to stable carotid plaques. To study the impact of GPR55 deficiency in atherosclerosis, we crossed Gpr55 knockout mice with apolipoprotein E (ApoE) knockout mice and subjected the mice to Western diet for 4 to 16 weeks. Compared to ApoE-/- controls, ApoE-/-Gpr55-/- mice developed larger plaques with increased necrotic core size, associated with elevated circulating and aortic leukocyte counts. Flow cytometry, immunofluorescence and RNA-sequencing analysis of splenic B cells in these mice revealed a hyperactivated B cell phenotype with disturbed plasma cell maturation and immunoglobulin (Ig)G antibody overproduction. The specific contribution of B cell GPR55 in atherosclerosis was further studied in mixed Gpr55-/-/{micro}MT bone marrow chimeras on low density receptor deficiency (Ldlr-/-) background, revealing that B-cell specific depletion of Gpr55 was sufficient to promote plaque development. Conversely, adoptive transfer of wildtype B cells into ApoE-/-Gpr55-/- mice blunted the proatherogenic phenotype. In vitro stimulation of splenocytes with the endogenous GPR55 ligand LPI promoted plasma cell proliferation and enhanced B cell activation marker expression, which was inhibited by the GPR55 antagonist CID16020046. Collectively, these discoveries provide new evidence for GPR55 as key modulator of the adaptive immune response in atherosclerosis. Targeting GPR55 could be useful to limit inflammation and plaque progression in patients suffering from atherosclerosis.

immunology↗

bioRxiv2021

Direct addition of poly-lysine or poly-ethylenimine to the medium, a simple alternative to plate pre-coating

For most cell culture experiments, it is indispensable that the cells are firmly anchored to the culture plates, tolerating several rinsing steps, and withstanding shear forces or temperature changes without detaching. For semi-adherent cells such as the very common HEK 293 cells, this could so far be obtained only by time-consuming plate pre-coating with cationic polymer solutions. We report here, that i) pre-coating with the cheaper poly-ethylenimine (PEI) works as well as the commonly used poly-D-lysine (PDL), but more importantly and novel ii) that simple direct addition of either PEI (1.5 g/ml) or PDL (2 g/ml) to the cell culture medium results in strongly anchored HEK 293 cells, indistinguishable from ones seeded on pre-coated plates. Therefore, the replacement of plate pre-coating by direct addition of either PEI or PDL gives comparable excellent results, but is highly labour-, time-, and cost-efficient. Interestingly, additional experiments in this context showed that strong cell attachment requires only cationic polymers but not fetal calf serum added to the medium. Fetal calf serum is, however, of course required for further maintenance and growth of the cells.

cell biology↗