Specification of CNS macrophage subsets occurs postnatally in defined niches.

All tissue-resident macrophages of the central nervous system (CNS)-including parenchymal microglia, as well as CNS-associated macrophages (CAMs1) such as meningeal and perivascular macrophages2-7-are part of the CNS endogenous innate immune system that acts as the first line of defence during infections or trauma2,8-10. It has been suggested that microglia and all subsets of CAMs are derived from prenatal cellular sources in the yolk sac that were defined as early erythromyeloid progenitors11-15. However, the precise ontogenetic relationships, the underlying transcriptional programs and the molecular signals that drive the development of distinct CAM subsets in situ are poorly understood. Here we show, using fate-mapping systems, single-cell profiling and cell-specific mutants, that only meningeal macrophages and microglia share a common prenatal progenitor. By contrast, perivascular macrophages originate from perinatal meningeal macrophages only after birth in an integrin-dependent manner. The establishment of perivascular macrophages critically requires the presence of arterial vascular smooth muscle cells. Together, our data reveal a precisely timed process in distinct anatomical niches for the establishment of macrophage subsets in the CNS.

NRG1 type I dependent autoparacrine stimulation of Schwann cells in onion bulbs of peripheral neuropathies.

In contrast to acute peripheral nerve injury, the molecular response of Schwann cells in chronic neuropathies remains poorly understood. Onion bulb structures are a pathological hallmark of demyelinating neuropathies, but the nature of these formations is unknown. Here, we show that Schwann cells induce the expression of Neuregulin-1 type I (NRG1-I), a paracrine growth factor, in various chronic demyelinating diseases. Genetic disruption of Schwann cell-derived NRG1 signalling in a mouse model of Charcot-Marie-Tooth Disease 1A (CMT1A), suppresses hypermyelination and the formation of onion bulbs. Transgenic overexpression of NRG1-I in Schwann cells on a wildtype background is sufficient to mediate an interaction between Schwann cells via an ErbB2 receptor-MEK/ERK signaling axis, which causes onion bulb formations and results in a peripheral neuropathy reminiscent of CMT1A. We suggest that diseased Schwann cells mount a regeneration program that is beneficial in acute nerve injury, but that overstimulation of Schwann cells in chronic neuropathies is detrimental.

Fatty Acid Oxidation Compensates for Lipopolysaccharide-Induced Warburg Effect in Glucose-Deprived Monocytes.

Monocytes enter sites of microbial or sterile inflammation as the first line of defense of the immune system and initiate pro-inflammatory effector mechanisms. We show that activation with bacterial lipopolysaccharide (LPS) induces them to undergo a metabolic shift toward aerobic glycolysis, similar to the Warburg effect observed in cancer cells. At sites of inflammation, however, glucose concentrations are often drastically decreased, which prompted us to study monocyte function under conditions of glucose deprivation and abrogated Warburg effect. Experiments using the Seahorse Extracellular Flux Analyzer revealed that limited glucose supply shifts monocyte metabolism toward oxidative phosphorylation, fueled largely by fatty acid oxidation at the expense of lipid droplets. While this metabolic state appears to provide sufficient energy to sustain functional properties like cytokine secretion, migration, and phagocytosis, it cannot prevent a rise in the AMP/ATP ratio and a decreased respiratory burst. The molecular trigger mediating the metabolic shift and the functional consequences is activation of AMP-activated protein kinase (AMPK). Taken together, our results indicate that monocytes are sufficiently metabolically flexible to perform pro-inflammatory functions at sites of inflammation despite glucose deprivation and inhibition of the LPS-induced Warburg effect. AMPK seems to play a pivotal role in orchestrating these processes during glucose deprivation in monocytes.

Origin, fate and dynamics of macrophages at central nervous system interfaces.

Perivascular, subdural meningeal and choroid plexus macrophages are non-parenchymal macrophages that mediate immune responses at brain boundaries. Although the origin of parenchymal microglia has recently been elucidated, much less is known about the precursors, the underlying transcriptional program and the dynamics of the other macrophages in the central nervous system (CNS). It was assumed that they have a high turnover from blood-borne monocytes. However, using parabiosis and fate-mapping approaches in mice, we found that CNS macrophages arose from hematopoietic precursors during embryonic development and established stable populations, with the notable exception of choroid plexus macrophages, which had dual origins and a shorter life span. The generation of CNS macrophages relied on the transcription factor PU.1, whereas the MYB, BATF3 and NR4A1 transcription factors were not required.

Effector T-cell trafficking between the leptomeninges and the cerebrospinal fluid.

In multiple sclerosis, brain-reactive T cells invade the central nervous system (CNS) and induce a self-destructive inflammatory process. T-cell infiltrates are not only found within the parenchyma and the meninges, but also in the cerebrospinal fluid (CSF) that bathes the entire CNS tissue. How the T cells reach the CSF, their functionality, and whether they traffic between the CSF and other CNS compartments remains hypothetical. Here we show that effector T cells enter the CSF from the leptomeninges during Lewis rat experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. While moving through the three-dimensional leptomeningeal network of collagen fibres in a random Brownian walk, T cells were flushed from the surface by the flow of the CSF. The detached cells displayed significantly lower activation levels compared to T cells from the leptomeninges and CNS parenchyma. However, they did not represent a specialized non-pathogenic cellular sub-fraction, as their gene expression profile strongly resembled that of tissue-derived T cells and they fully retained their encephalitogenic potential. T-cell detachment from the leptomeninges was counteracted by integrins VLA-4 and LFA-1 binding to their respective ligands produced by resident macrophages. Chemokine signalling via CCR5/CXCR3 and antigenic stimulation of T cells in contact with the leptomeningeal macrophages enforced their adhesiveness. T cells floating in the CSF were able to reattach to the leptomeninges through steps reminiscent of vascular adhesion in CNS blood vessels, and invade the parenchyma. The molecular/cellular conditions for T-cell reattachment were the same as the requirements for detachment from the leptomeningeal milieu. Our data indicate that the leptomeninges represent a checkpoint at which activated T cells are licensed to enter the CNS parenchyma and non-activated T cells are preferentially released into the CSF, from where they can reach areas of antigen availability and tissue damage.

Defective podocyte insulin signalling through p85-XBP1 promotes ATF6-dependent maladaptive ER-stress response in diabetic nephropathy.

Endoplasmic reticulum (ER) stress is associated with diabetic nephropathy (DN), but its pathophysiological relevance and the mechanisms that compromise adaptive ER signalling in podocytes remain unknown. Here we show that nuclear translocation of the transcription factor spliced X-box binding protein-1 (sXBP1) is selectively impaired in DN, inducing activating transcription factor-6 (ATF6) and C/EBP homology protein (CHOP). Podocyte-specific genetic ablation of XBP1 or inducible expression of ATF6 in mice aggravates DN. sXBP1 lies downstream of insulin signalling and attenuating podocyte insulin signalling by genetic ablation of the insulin receptor or the regulatory subunits phosphatidylinositol 3-kinase (PI3K) p85α or p85ß impairs sXBP1 nuclear translocation and exacerbates DN. Corroborating our findings from murine DN, the interaction of sXBP1 with p85α and p85ß is markedly impaired in the glomerular compartment of human DN. Thus, signalling via the insulin receptor, p85, and XBP1 maintains podocyte homeostasis, while disruption of this pathway impairs podocyte function in DN.

CD11c-positive cells from brain, spleen, lung, and liver exhibit site-specific immune phenotypes and plastically adapt to new environments.

The brain's immune privilege has been also attributed to the lack of dendritic cells (DC) within its parenchyma and the adjacent meninges, an assumption, which implies maintenance of antigens rather than their presentation in lymphoid organs. Using mice transcribing the green fluorescent protein under the promoter of the DC marker CD11c (itgax), we identified a juxtavascular population of cells expressing this DC marker and demonstrated their origin from bone marrow and local microglia. We now phenotypically compared this population with CD11c/CD45 double-positive cells from lung, liver, and spleen in healthy mice using seven-color flow cytometry. We identified unique, site-specific expression patterns of F4/80, CD80, CD86, CX3CR1, CCR2, FLT3, CD103, and MHC-II. Furthermore, we observed the two known CD45-positive populations (CD45(high) and CD45(int) ) in the brain, whereas liver, lung, and spleen exhibited a homogeneous CD45(high) population. CD11c-positive microglia lacked MHC-II expression and CD45(high) /CD11c-positive cells from the brain have a lower percentage of MHC-II-positive cells. To test whether phenotypical differences are fixed by origin or specifically develop due to environmental factors, we transplanted brain and spleen mononuclear cells on organotypic slice cultures from brain (OHSC) and spleen (OSSC). We demonstrate that adaption and ramification of MHC-II-positive splenocytes is paralleled by down-regulation of MHC-II, whereas brain-derived mononuclear cells neither ramified nor up-regulated MHC-II in OSSCs. Thus, brain-derived mononuclear cells maintain their MHC-II-negative phenotype within the environment of an immune organ. Intraparenchymal CD11c-positive cells share immunophenotypical characteristics of DCs from other organs but remain unique for their low MHC-II expression.

Personalization of atrial anatomy and electrophysiology as a basis for clinical modeling of radio-frequency ablation of atrial fibrillation.

Multiscale cardiac modeling has made great advances over the last decade. Highly detailed atrial models were created and used for the investigation of initiation and perpetuation of atrial fibrillation. The next challenge is the use of personalized atrial models in clinical practice. In this study, a framework of simple and robust tools is presented, which enables the generation and validation of patient-specific anatomical and electrophysiological atrial models. Introduction of rule-based atrial fiber orientation produced a realistic excitation sequence and a better correlation to the measured electrocardiograms. Personalization of the global conduction velocity lead to a precise match of the measured P-wave duration. The use of a virtual cohort of nine patient and volunteer models averaged out possible model-specific errors. Intra-atrial excitation conduction was personalized manually from left atrial local activation time maps. Inclusion of LE-MRI data into the simulations revealed possible gaps in ablation lesions. A fast marching level set approach to compute atrial depolarization was extended to incorporate anisotropy and conduction velocity heterogeneities and reproduced the monodomain solution. The presented chain of tools is an important step towards the use of atrial models for the patient-specific AF diagnosis and ablation therapy planing.

Structure of a methyl-coenzyme M reductase from Black Sea mats that oxidize methane anaerobically.

The anaerobic oxidation of methane (AOM) with sulphate, an area currently generating great interest in microbiology, is accomplished by consortia of methanotrophic archaea (ANME) and sulphate-reducing bacteria. The enzyme activating methane in methanotrophic archaea has tentatively been identified as a homologue of methyl-coenzyme M reductase (MCR) that catalyses the methane-forming step in methanogenic archaea. Here we report an X-ray structure of the 280 kDa heterohexameric ANME-1 MCR complex. It was crystallized uniquely from a protein ensemble purified from consortia of microorganisms collected with a submersible from a Black Sea mat catalysing AOM with sulphate. Crystals grown from the heterogeneous sample diffract to 2.1 Å resolution and consist of a single ANME-1 MCR population, demonstrating the strong selective power of crystallization. The structure revealed ANME-1 MCR in complex with coenzyme M and coenzyme B, indicating the same substrates for MCR from methanotrophic and methanogenic archaea. Differences between the highly similar structures of ANME-1 MCR and methanogenic MCR include a F(430) modification, a cysteine-rich patch and an altered post-translational amino acid modification pattern, which may tune the enzymes for their functions in different biological contexts.

Model-based assessment of tissue perfusion and temperature in deep hypothermic patients.

Deep hypothermic circulatory arrest is necessary for some types of cardiac and aortic surgery. Perfusion of the brain can be maintained using a heart-lung machine and unilateral antegrade cerebral perfusion. Cooling rates during extracorporeal circulation depend on local perfusion. A core temperature of 24 degrees C-25 degrees C is aimed at to extend ischemic tolerance of tissues. Information on cerebral perfusion and temperature is important for the safety of patients, but hardly accessible to measurement. A combined simulation model of hemodynamics and temperature is presented in this paper. The hemodynamics model employs the transmission-line approach and integrates the Circle of Willis (CoW). This allows for parameterization of individual aberrations. Simulation results of cerebral perfusion are shown for two configurations of the CoW. The temperature model provides spatial information on temperature fields. It considers heat transfer in the various tissues retrieving data of local tissue perfusion from the hemodynamics model. The combined model is evaluated by retrospective simulation of two aortic operations.

Intrahepatic long-term persistence of parvovirus B19 and its role in chronic viral hepatitis.

Parvovirus B19 (B19V) has been detected in the liver of Asian patients infected with HBV and may contribute to acute and chronic liver disease. This study aimed to investigate the impact of B19V infection in European patients with viral hepatitis. B19V DNA was detected in 1/91 and 0/50 serum samples from patients with chronic hepatitis C and B, respectively. In contrast, B19V DNA was amplified frequently from explanted end-stage liver tissues (37/50, 74%) and from routine biopsy samples (14/32, 44%) (P < 0.05). However, there was no significant difference in B19V copy number per cell between these two groups. B19V-specific CD4(+) T-cell responses to two dominant MHC-class-restricted epitopes were detected in a similar frequency in healthy anti-B19V-positive individuals (3/19; 16%) and patients with chronic hepatitis C (3/13; 23%). These results indicate that B19V can persist in the liver. However, there is no evidence that B19V is a "hepatitis virus" worsening liver disease in European patients with chronic hepatitis C.