The choroid plexus may be an underestimated site of tumor invasion to the brain: an in vitro study using neuroblastoma cell lines

Background: The central nervous system (CNS) is protected by several barriers, including the blood–brain (BBB) and blood-cerebrospinal fluid (BCSFB) barriers. Understanding how cancer cells circumvent these protective barriers to invade the CNS is of crucial interest, since brain metastasis during cancer is often a fatal event in both children and adults. However, whereas much effort has been invested in elucidating the process of tumor cell transmigration across the BBB, the role of the BCSFB might still be underestimated considering the significant number of meningeal cancer involvement. Our work aimed to investigate the transmigration of neuroblastoma cells across the BCSFB in vitro. Methods: We used an inverted model of the human BCSFB presenting proper restrictive features including adequate expression of tight-junction proteins, low permeability to integrity markers, and high trans-epithelial electrical resistance. Two different human neuroblastoma cell lines (SH-SY5Y and IMR-32) were used to study the transmigration process by fluorescent microscopy analysis. Results: The results show that neuroblastoma cells are able to actively cross the tight human in vitro BCSFB model within 24 h. The presence and transmigration of neuroblastoma cancer cells did not affect the barrier integrity within the duration of the experiment. Conclusions: In conclusion, we presume that the choroid plexus might be an underestimated site of CNS invasion, since neuroblastoma cell lines are able to actively cross a choroid plexus epithelial cell layer. Further studies are warranted to elucidate the molecular mechanisms of tumor cell transmigration in vitro and in vivo

Postarrest stalling rather than crawling favors CD8(+) over CD4(+) T-cell migration across the blood-brain barrier under flow in vitro.

Although CD8(+) T cells have been implied in the pathogenesis of multiple sclerosis (MS), the molecular mechanisms mediating CD8(+) T-cell migration across the blood-brain barrier (BBB) into the central nervous system (CNS) are ill defined. Using in vitro live cell imaging, we directly compared the multistep extravasation of activated CD4(+) and CD8(+) T cells across primary mouse brain microvascular endothelial cells (pMBMECs) as a model for the BBB under physiological flow. Significantly higher numbers of CD8(+) than CD4(+) T cells arrested on pMBMECs under noninflammatory and inflammatory conditions. While CD4(+) T cells polarized and crawled prior to their diapedesis, the majority of CD8(+) T cells stalled and readily crossed the pMBMEC monolayer preferentially via a transcellular route. T-cell arrest and crawling were independent of G-protein-coupled receptor signaling. Rather, absence of endothelial ICAM-1 and ICAM-2 abolished increased arrest of CD8(+) over CD4(+) T cells and abrogated T-cell crawling, leading to the efficient reduction of CD4(+) , but to a lesser degree of CD8(+) , T-cell diapedesis across ICAM-1(null) /ICAM-2(-/-) pMBMECs. Thus, cellular and molecular mechanisms mediating the multistep extravasation of activated CD8(+) T cells across the BBB are distinguishable from those involved for CD4(+) T cells

Scandinavian trial of uncomplicated aortic dissection therapy : study protocol for a randomized controlled trial

Peer reviewe

Strain-dependent effects of clinical echovirus 30 outbreak isolates at the blood-CSF barrier

Background: Echovirus (E) 30 (E-30) meningitis is characterized by neuroinflammation involving immune cell pleocytosis at the protective barriers of the central nervous system (CNS). In this context, infection of the blood-cerebrospinal fluid barrier (BCSFB), which has been demonstrated to be involved in enteroviral CNS pathogenesis, may affect the tight junction (TJ) and adherens junction (AJ) function and morphology. Methods: We used an in vitro human choroid plexus epithelial (HIBCPP) cell model to investigate the effect of three clinical outbreak strains (13-311, 13-759, and 14-397) isolated in Germany in 2013, and compared them to E-30 Bastianni. Conducting transepithelial electrical resistance (TEER), paracellular dextran flux measurement, quantitative real-time polymerase chain reaction (qPCR), western blot, and immunofluorescence analysis, we investigated TJ and AJ function and morphology as well as strain-specific E-30 infection patterns. Additionally, transmission electron and focused ion beam microscopy electron microscopy (FIB-SEM) was used to evaluate the mode of leukocyte transmigration. Genome sequencing and phylogenetic analyses were performed to discriminate potential genetic differences among the outbreak strains. Results: We observed a significant strain-dependent decrease in TEER with strains E-30 Bastianni and 13-311, whereas paracellular dextran flux was only affected by E-30 Bastianni. Despite strong similarities among the outbreak strains in replication characteristics and particle distribution, strain 13-311 was the only outbreak isolate revealing comparable disruptive effects on TJ (Zonula Occludens (ZO) 1 and occludin) and AJ (E-cadherin) morphology to E-30 Bastianni. Notwithstanding significant junctional alterations upon E-30 infection, we observed both para- and transcellular leukocyte migration across HIBCPP cells. Complete genome sequencing revealed differences between the strains analyzed, but no explicit correlation with the observed strain-dependent effects on HIBCPP cells was possible. Conclusion: The findings revealed distinct E-30 strain-specific effects on barrier integrity and junctional morphology. Despite E-30-induced barrier alterations leukocyte trafficking did not exclusively occur via the paracellular route

Distinct migratory pattern of naive and effector T cells through the blood-CSF barrier following Echovirus 30 infection

AbstractBackgroundEchovirus 30 (E-30) is one of the most frequently isolated pathogens in aseptic meningitis worldwide. To gain access to the central nervous system (CNS), E-30 and immune cells have to cross one of the two main barriers of the CNS, the epithelial blood–cerebrospinal fluid barrier (BCSFB) or the endothelial blood–brain barrier (BBB). In an in vitro model of the BCSFB, it has been shown that E-30 can infect human immortalized brain choroid plexus papilloma (HIBCPP) cells.MethodsIn this study we investigated the migration of different T cell subpopulations, naive and effector T cells, through HIBCPP cells during E-30 infection. Effects of E-30 infection and the migration process were evaluated via immunofluorescence and flow cytometry analysis, as well as transepithelial resistance and dextran flux measurement.ResultsTh1 effector cells and enterovirus-specific effector T cells migrated through HIBCPP cells more efficiently than naive CD4+ T cells following E-30 infection of HIBCPP cells. Among the different naive T cell populations, CD8+ T cells crossed the E-30-infected HIBCPP cell layer in a significantly higher number than CD4+ T cells. A large amount of effector T cells also remained attached to the basolateral side of the HIBCPP cells compared with naive T cells. Analysis of HIBCPP barrier function showed significant alteration after E-30 infection and trans- as well as paracellular migration of T cells independent of the respective subpopulation. Morphologic analysis of migrating T cells revealed that a polarized phenotype was induced by the chemokine CXCL12, but reversed to a round phenotype after E-30 infection. Further characterization of migrating Th1 effector cells revealed a downregulation of surface adhesion proteins such as LFA-1 PSGL-1, CD44, and CD49d.ConclusionTaken together these results suggest that naive CD8+ and Th1 effector cells are highly efficient to migrate through the BCSFB in an inflammatory environment. The T cell phenotype is modified during the migration process through HIBCPP cells.</div

General Characteristics of Children with Single- and Co-Infections and Febrile Seizures with a Main Focus on Respiratory Pathogens: Preliminary Results

Febrile seizures (FS) affect up to 5% of children. The pathogen etiology in regard of viral loads has never been investigated. In a prospective cohort study we investigated the correlation between virus type and quantity in nasopharyngeal aspirates (NPAs) and the clinical characteristics in pediatric patients with a FS. From January 2014 to April 2016, 184 children with a FS were prospectively enrolled. The mean age of all included children was 26.7 ± 18.3 months with a male to female ratio of 1.4:1. Males with an acute disease and a short duration or absence of prior symptoms had a higher risk for complex FS. The majority of patients with FS presented with a generalized convulsion (180; 98%) and was admitted to hospital (178; 97%). Overall, 79 (43%) single and in 59 (32%) co-infections were detected. Human herpes virus 6 (HHV6), influenza, adenovirus (AV) and rhinovirus (RV) were the dominant pathogens, all detected with clinically significant high viral loads. HHV6 positive cases were significantly younger and less likely to have a positive family/personal history for FS. Influenza positives showed a higher rate of complex seizures, lower leukocyte and higher monocyte counts. AV positive cases were more likely to have a positive family history for FS and showed higher C-reactive protein values. In conclusion, a high viral load may contribute to the development of a FS in respiratory tract infections

Central nervous system mold infections in children with hematological malignancies: advances in diagnosis and treatment

The incidence of invasive mold disease (IMD) has significantly increased over the last decades, and IMD of the central nervous system (CNS) is a particularly severe form of this infection. Solid data on the incidence of CNS IMD in the pediatric setting are lacking, in which Aspergillus spp. is the most prevalent pathogen, followed by mucorales. CNS IMD is difficult to diagnose, and although imaging tools such as magnetic resonance imaging have considerably improved, these techniques are still unspecific. As microscopy and culture have a low sensitivity, non-culture-based assays such as the detection of fungal antigens (e.g., galactomannan or beta-D-glucan) or the detection of fungal nucleic acids by molecular assays need to be validated in children with suspected CNS IMD. New and potent antifungal compounds helped to improve outcome of CNS IMD, but not all agents are approved for children and a pediatric dosage has not been established. Therefore, studies have to rapidly evaluate dosage, safety and efficacy of antifungal compounds in the pediatric setting. This review will summarize the current knowledge on diagnostic tools and on the management of CNS IMD with a focus on pediatric patients