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Um kritisch kranke Kinder erfolgreich versorgen zu konnen, ist es notwendig, diese so rasch wie moglich zu identifizieren. Hierbei konnen Algorithmen und standardisierte Vorgehensweisen helfen. Ist ein Kind kritisch krank, muss umgehend eine adaquate Therapie erfolgen. Vermittlung einer strukturierten und standardisierten Untersuchungstechnik zur Erkennung des kritisch kranken Kindes und Darstellung der notwendigen Masnahmen der Erstversorgung. Die aktuelle Literatur und Empfehlungen zur Versorgung des kritisch kranken Kindes werden ausgewertet und diskutiert. Zudem erfolgt eine Darstellung der geschilderten Untersuchungstechniken und der Vorgehensweise in der Erstversorgung des kritisch kranken Kindes anhand eines online verfugbaren Videos. Die dargestellten Hilfsmittel (z. B. das padiatrische Diagnosedreieck) und der ABCDE-Algorithmus helfen, kritisch kranke Kinder standardisiert, strukturiert und rasch zu identifizieren. Eine adaquate und dem aufgezeigten Schema folgende Erstversorgung hilft bei der zielfuhrenden Therapie. Regelmasige Teamtrainings der Notfallversorgung tragen dazu bei, das erlernte Wissen praktisch umzusetzen.

Management and monitoring of pain and sedation to reduce discomfort as well as side effects, such as over- and under-sedation, withdrawal syndrome and delirium, is an integral part of pediatric intensive care practice. However, the current state of management and monitoring of analgosedation across European pediatric intensive care units (PICUs) remains unknown. The aim of this survey was to describe current practices across European PICUs regarding the management and monitoring of pain and sedation.An online survey was distributed among 357 European PICUs assessing demographic features, drug choices and dosing, as well as usage of instruments for monitoring pain and sedation. We also compared low- and high-volume PICUs practices. Responses were collected from January to April 2021.A total of 215 (60% response rate) PICUs from 27 European countries responded. Seventy-one percent of PICUs stated to use protocols for analgosedation management, more frequently in high-volume PICUs (77% vs 63%, p = 0.028). First-choice drug combination was an opioid with a benzodiazepine, namely fentanyl (51%) and midazolam (71%) being the preferred drugs. The starting doses differed between PICUs from 0.1 to 5 mcg/kg/h for fentanyl, and 0.01 to 0.5 mg/kg/h for midazolam. Daily assessment and documentation for pain (81%) and sedation (87%) was reported by most of the PICUs, using the preferred validated FLACC scale (54%) and the COMFORT Behavioural scale (48%), respectively. Both analgesia and sedation were mainly monitored by nurses (92% and 84%, respectively). Eighty-six percent of the responding PICUs stated to use neuromuscular blocking agents in some scenarios. Monitoring of paralysed patients was preferably done by observation of vital signs with electronic devices support.This survey provides an overview of current analgosedation practices among European PICUs. Drugs of choice, dosing and assessment strategies were shown to differ widely. Further research and development of evidence-based guidelines for optimal drug dosing and analgosedation assessment are needed.

Background Isolated myelosarcoma of infancy is a rare presentation of acute myelogenous leukaemia (AML). Because of its rarity and early onset in infancy underlying genetic predisposition is potentially relevant in disease initiation. Methods and Results We report an oncologic emergency in an infant with thoracic and intraspinal aleukaemic myeloid sarcoma causing acute myelon compression and lower leg palsy. Whole‐exome sequencing of the patient's germline DNA identified a rare PALB2 (OMIM 610355) variant (p.A1079S), which is located in a domain critical for the gene's proper function within the homology‐directed repair pathway. In line with potential DNA damage repair defects mediated by the PALB2 deregulation, the patient's fibroblasts showed increased sensitivity towards radiation and DNA intercalating agents. Conclusion Therefore, we suggest PALB2 p.A1079S as a pathogenic variant potentially contributing to the here observed patient phenotype., We present the finding of an infant with thoracic and intraspinal aleukemic myeloid sarcoma with a rare germline PALB2 variant (p.A1079S). The identified variant is located in a domain critical for the gene's proper function within the homology‐directed repair pathway. A functional role of the variant is further supported by increased sensitivity towards irradiation and DNA intercalating agents observed in primary cells from the patient carrying PALB2 p.A1079S.

Inflammatory conditions are critically influenced by neuroimmune crosstalk. Cytokines and neurotrophic factors shape the responses of both nervous and immune systems. Although much progress has been made, most findings to date are based on expression of recombinant (tagged) proteins. The examination of receptor interactions by immunoprecipitation (IP) at endogenous levels provides further insight into the more subtle regulations of immune responses. Here, we present a comprehensive workflow and an optimized IP protocol that provide step-by-step instructions to investigate neurotrophin receptor p75NTR at endogenous, low abundance levels: from lysate preparation and confirmation of receptor expression to antibody validation and successful detection of protein-protein interactions. We employ human melanoma cell line A375 to validate specific antibodies and IP conditions, and apply these methods to explore p75NTR interactions in human leukemic plasmacytoid dendritic cell line PMDC05 detecting 14-3-3ϵ:p75NTR interaction in this cell type. With p75NTR as an exemplary protein, our approach provides a strategy to detect specific interaction partners even under endogenous, low abundance expression conditions.

Fur das neurologische Outcome der Kinder hat das Temperaturmanagement eine entscheidende Bedeutung. Moderate (28–32 °C) und schwere Hypothermien (

Storungen der Atmung dominieren einen Grosteil padiatrischer Notfallsituationen und stellen den haufigsten Grund fur einen Kreislaufstillstand im Kindesalter dar. Anatomische und funktionelle Besonderheiten im Kindes- und Sauglingsalter fuhren, verglichen mit Erwachsenen, zu einer reduzierten Apnoetoleranz und einer Neigung zu obstruktivem Ventilationsversagen. Im Rahmen dieses Kapitels werden Besonderheiten des Respirationstraktes im Sauglings- und Kindesalter erlautert und Ursachen und Therapie der akuten respiratorischen Insuffizienz systematisch dargestellt.

Wahrend einer kardiopulmonalen Reanimation mussen reversible Ursachen des Atem-Kreislauf-Stillstands erkannt und behandelt werden. Die Uberprufung der 4H und HITS erlaubt ein strukturiertes Vorgehen. Eine fruhzeitige Korrektur der reversiblen Ursachen verbessert die Chancen auf die Wiederherstellung eines Spontankreislaufs.

Der Sommerurlaub war vorbei und Dr. Levis war froh, dass sein erster Tag auf der padiatrischen Intensivstation ruhig verlief. Er verbrachte die Zeit damit, sich mit den Patientenverlaufen vertraut zu machen. – In diesem Kapitel geht es um die Qualifikation Intensivtransport, das Monitoring beim Intensivtransport, Ubergaben, PPV, SSV, DOPES, Apnoetoleranz, Blutdruck, Propofol, invasive Blutdruckmessung sowie Planungsoptimismus.

Chronic granulomatous disease (CGD) is an inherited immunodeficiency, caused by the inability of neutrophils to produce functional NADPH oxidase required for fighting microbial infections. The X-linked form of CGD (X-CGD), which is due to mutations in the CYBB (gp91phox) gene, a component of NADPH oxidase, accounts for about two-thirds of CGD cases. We derived induced pluripotent stem cells (iPSCs) from X-CGD patient keratinocytes using a Flp recombinase excisable lentiviral reprogramming vector. For restoring gp91phox function, we applied two strategies: transposon-mediated bacterial artificial chromosome (BAC) transgenesis and gene targeting using vectors with a fixed 5′ homology arm (HA) of 8 kb and 3′HA varying in size from 30 to 80 kb. High efficiency of homologous recombination (up to 22%) was observed with increased size of the 3′HA. Both, BAC transgenesis and gene targeting resulted in functional restoration of the gp91phox measured by an oxidase activity assay in X-CGD iPSCs differentiated into the myeloid lineage. In conclusion, we delivered an important milestone towards the use of genetically corrected autologous cells for the treatment of X-CGD and monogenic diseases in general.

OBJECTIVE The aim of this study was to identify the causes and incidences of neonatal diseases and deaths in five provincial hospitals in People's Democratic Republic of Laos retrospectively for the years 2010-12. METHODS Data of neonatal patients were collected before a 3-year-training program for medical and nursing staff involved in the care of newborn infants in the provincial and associated district hospitals. RESULTS In the years 2010-12, a total of 1673 neonatal patients were treated in the provincial hospitals. The reasons of treatment were as follows: 48% infections, 17% complications of prematurity, 14% intrapartum-related complications and 9% other, not categorized diseases. The average mortality rate in all hospitals was 6.5%. The main causes of death were complications because of prematurity, infectious diseases and asphyxia. CONCLUSION These data could be the basis for any teaching program aimed at reducing neonatal mortality. Furthermore, they enable an evaluation of the ongoing teaching program.

This study investigates the attitudes towards and social impacts of Edward Snowden's revelations in Germany through a questionnaire survey with German youngsters as part of the worldwide cross-cultural analyses. However due to Snowden's revelations a continuing discussion about privacy, safety, security and data protection was unleashed in Germany. The results show interesting values and settings of young people in Germany. For example: The majority 69.41% of the surveyed persons feel that the usage of the Internet threatens their privacy.

The X-linked histone demethylase UTX has a pivotal role in cellular and developmental processes including embryogenesis, hematopoiesis and cancer. UTX removes di- and trimethyl groups on histone H3 lysine 27, thereby regulating gene expression. But there is growing evidence that UTX displays biological functions independent of its histone demethylase activity. To elucidate these novel functions, it is of great interest to define subcellular localizations of UTX. Here we show for the first time that native UTX is primarily localized in the cytoplasm whereas ectopic GFP and Flag-tagged UTX display nuclear and cytoplasmic localization. While its epigenetic function is exerted in the nucleus, its cytoplasmic localization points to a novel function.

This consensus- based S1 Guideline for perioperative infusion therapy in children is focused on safety and efficacy. The objective is to maintain or re-establish the child's normal physiological state (normovolemia, normal tissue perfusion, normal metabolic function, normal acid- base- electrolyte status). Therefore, the perioperative fasting times should be as short as possible to prevent patient discomfort, dehydration, and ketoacidosis. A physiologically composed balanced isotonic electrolyte solution (BS) with 1-2.5% glucose is recommended for the intraoperative background infusion to maintain normal glucose concentrations and to avoid hyponatremia, hyperchloremia, and lipolysis. Additional BS without glucose can be used in patients with circulatory instability until the desired effect is achieved. The additional use of colloids (albumin, gelatin, hydroxyethyl starch) is recommended to recover normovolemia and to avoid fluid overload when crystalloids alone are not sufficient and blood products are not indicated. Monitoring should be extended in cases with major surgery, and autotransfusion maneuvers should be performed to assess fluid responsiveness.

Mesenchymal stromal cells (MSC) and factors secreted by them are essential components of the hematopoietic stem cell (HSC) niche within the bone marrow microenvironment. It has been shown that the extracellular matrix (ECM) can influence HSC-supportive potential of MSC and is a prerequisite for the proper signaling of morphogens. Therefore, we aimed at the identification of ECM components and candidate morphogens capable of enhancing the expression of HSC-supportive proteins in human MSC, namely, angiopoietin-1 (Ang-1) and stromal cell-derived factor 1 (SDF-1). For this purpose, highly sensitive secreted dual reporter constructs for Ang-1 and SDF-1 were established. These newly designed dual reporter systems enable continuous monitoring of the Ang-1 and SDF-1 promoter activity in an immortalized human MSC line cultured on ECM/morphogen microarrays. Reporter arrays showed that Ang-1 and SDF-1 expression can be induced by different ECM/morphogen combinations. In addition, continuous monitoring of promoter activity allows delineating time-dependent effects of the ECM and morphogens. Thus, we identified that collagen I and vitronectin in combination with Wnt3a favored SDF-1 expression over time, while only transiently inducing the expression of Ang-1. Taken together, the newly developed reporter systems allow for the monitoring of Ang-1 and SDF-1 promoter activity induced by morphogens and the ECM in a combinatorial and high-throughput manner. This technology might therefore be helpful to optimize culture conditions, which favor the activity of MSC as feeder cells for various types of stem and progenitor cells.

The proinflammatory enzyme caspase-1 plays an important role in the innate immune system and is involved in a variety of inflammatory conditions. Rare naturally occurring human variants of the caspase-1 gene (CASP1) lead to different protein expression and structure and to decreased or absent enzymatic activity. Paradoxically, a significant number of patients with such variants suffer from febrile episodes despite decreased IL-1β production and secretion. In this study, we investigate how variant (pro)caspase-1 can possibly contribute to inflammation. In a transfection model, such variant procaspase-1 binds receptor interacting protein kinase 2 (RIP2) via Caspase activation and recruitment domain (CARD)/CARD interaction and thereby activates NF-κB, whereas wild-type procaspase-1 reduces intracellular RIP2 levels by enzymatic cleavage and release into the supernatant. We approach the protein interactions by coimmunoprecipitation and confocal microscopy and show that NF-κB activation is inhibited by anti–RIP2-short hairpin RNA and by the expression of a RIP2 CARD-only protein. In conclusion, variant procaspase-1 binds RIP2 and thereby activates NF-κB. This pathway could possibly contribute to proinflammatory signaling.

Mesenchymal stromal cells (MSC) are an important component of the bone marrow microenvironment. Notch ligands expressed by MSC are known to play a regulatory role for hematopoietic stem and progenitor cells (HSPC) and in support of bone marrow homeostasis. While the role of Notch signaling in HSPC, their progeny, and MSC has been relatively well studied, little is known about the Notch-independent regulatory impact of Notch ligands on MSC themselves. In the present study, we used genetically engineered bone marrow-derived human MSC to study the function role of Jagged-1 and the Jagged-1 intracellular domain (JICD) with regard to the interaction with HSPC. We demonstrate that Jagged-1 in human MSC undergoes cleavage to produce an intracellular domain that translocates into the nucleus. JICD but not Jagged-1 overexpression was associated with an increased expression of stromal cell-derived factor-1. Short-term co-culture (7 days) of HSPC with JICD-overexpressing MSC, but not with Jagged-1-overexpressing MSC, led to increased proliferation of CD34(+) progenitors. In contrast, long-term co-culture of HSPC with Jagged-1-overexpressing MSC (up to 6 weeks) led to a significantly better support of cobblestone area-forming cells and long-term culture-initiating cells (LTC-ICs) compared with JICD-overexpressing MSC. Taken together, results of this study indicate that full-length Jagged-1 and JICD have differential effects on MSC and on their interaction with HSPC ex-vivo. JICD-overexpressing MSC induce proliferation of HSPCs in short-term culture at the expense of immature precursors (LTC-ICs), whereas Jagged-1 overexpressing MSC favor LTC-IC formation.

The melanoma cell adhesion molecule defines mesenchymal stromal cells in the human bone marrow that regenerate bone and establish a hematopoietic microenvironment in vivo. The role of the melanoma cell adhesion molecule in primary human mesenchymal stromal cells and the maintenance of hematopoietic stem and progenitor cells during ex vivo culture has not yet been demonstrated. We applied RNA interference or ectopic overexpression of the melanoma cell adhesion molecule in human mesenchymal stromal cells to evaluate the effect of the melanoma cell adhesion molecule on their proliferation and differentiation as well as its influence on co-cultivated hematopoietic stem and progenitor cells. Knockdown and overexpression of the melanoma cell adhesion molecule affected several characteristics of human mesenchymal stromal cells related to osteogenic differentiation, proliferation, and migration. Furthermore, knockdown of the melanoma cell adhesion molecule in human mesenchymal stromal cells stimulated the proliferation of hematopoietic stem and progenitor cells, and strongly reduced the formation of long-term culture-initiating cells. In contrast, melanoma cell adhesion molecule-overexpressing human mesenchymal stromal cells provided a supportive microenvironment for hematopoietic stem and progenitor cells. Expression of the melanoma cell adhesion molecule increased the adhesion of hematopoietic stem and progenitor cells to human mesenchymal stromal cells and their migration beneath the monolayer of human mesenchymal stromal cells. Our results demonstrate that the expression of the melanoma cell adhesion molecule in human mesenchymal stromal cells determines their fate and regulates the maintenance of hematopoietic stem and progenitor cells through direct cell-cell contact.

Background Monogenic autoinflammatory disorders are characterized by dysregulation of the innate immune system, for example by gain-of-function mutations in inflammasome-forming proteins, such as NOD-like receptor family CARD-containing 4 protein (NLRC4). Objective Here we investigate the mechanism by which a novel mutation in the leucine-rich repeat (LRR) domain of NLRC4 (c.G1965C, p.W655C) contributes to autoinflammatory disease. Methods: We studied 2 unrelated patients with early-onset macrophage activation syndrome harboring the same de novo mutation in NLRC4. In vitro inflammasome complex formation was quantified by using flow cytometric analysis of apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) specks. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 techniques and lentiviral transduction were used to generate THP-1 cells with either wild-type or mutant NLRC4 cDNA. Cell death and release of IL-1β/IL-18 were quantified by using flow cytometry and ELISA, respectively. Results The p.W655C NLRC4 mutation caused increased ASC speck formation, caspase-1–dependent cell death, and IL-1β/IL-18 production. ASC contributed to p.W655C NLRC4–mediated cytokine release but not cell death. Mutation of p.W655 activated the NLRC4 inflammasome complex by engaging with 2 interfaces on the opposing LRR domain of the oligomer. One key set of residues (p.D1010, p.D1011, p.L1012, and p.I1015) participated in LRR-LRR oligomerization when triggered by mutant NLRC4 or type 3 secretion system effector (PrgI) stimulation of the NLRC4 inflammasome complex. Conclusion This is the first report of a mutation in the LRR domain of NLRC4 causing autoinflammatory disease. c.G1965C/p.W655C NLRC4 increased inflammasome activation in vitro. Data generated from various NLRC4 mutations provides evidence that the LRR-LRR interface has an important and previously unrecognized role in oligomerization of the NLRC4 inflammasome complex., Graphical abstract

Extracellular matrices determine cellular fate decisions through the regulation of intracellular force and stress. Previous studies suggest that matrix stiffness and ligand anchorage cause distinct signaling effects. We show herein how defined noncovalent anchorage of adhesion ligands to elastic substrates allows for dissection of intracellular adhesion signaling pathways related to matrix stiffness and receptor forces. Quantitative analysis of the mechanical balance in cell adhesion using traction force microscopy revealed distinct scalings of the strain energy imparted by the cells on the substrates dependent either on matrix stiffness or on receptor force. Those scalings suggested the applicability of a linear elastic theoretical framework for the description of cell adhesion in a certain parameter range, which is cell-type-dependent. Besides the deconvolution of biophysical adhesion signaling, site-specific phosphorylation of focal adhesion kinase, dependent either on matrix stiffness or on receptor force, also demonstrated the dissection of biochemical signaling events in our approach. Moreover, the net contractile moment of the adherent cells and their strain energy exerted on the elastic substrate was found to be a robust measure of cell adhesion with a unifying power-law scaling exponent of 1.5 independent of matrix stiffness.

We present a general, entirely PCR-based strategy to construct mRNAs coding for green fluorescent protein (GFP) fusion proteins from a cDNA pool. We exemplify our approach for the chemokine receptor CXCR4. mRNA transfection of the PCR-generated fusion of CXCR4-GFP into K562 cells or primary mesenchymal stem cells (MSCs) resulted in excellent viability (90%) with more than 90% of target cells expressing easily detectable CXCR4-GFP for72 h. The fusion protein was localized in the plasma membrane and was rapidly internalized upon incubation with the CXCR4 ligand stromal cell-derived factor-1 (SDF-1). Transwell migration experiments showed significantly increased migration of CXCR4-GFP mRNA-transfected MSCs toward a gradient of SDF-1, demonstrating that mRNA-mediated chemokine receptor overexpression allows for transient initiation of chemotaxis. The presented strategy to construct a PCR-based fluorescent fusion protein can be generally applied to other genes of interest to study their function by simple overexpression and easy detection in primary cells.

Case Study: Hoyeraal-Hreidarsson syndrome (HHS; MIM #305000, Xq28) is a rare X-linked recessive multisystem disorder characterized by intrauterine growth retardation, microcephaly, severe developmental delay, cerebellar hypoplasia, and bone marrow failure. We report the clinical course of a male neonatal floppy infant with progressive bone marrow failure. The infant was born after 33 weeks' of gestation by secondary cesarean after severe intrauterine growth retardation and anhydramnios. Because of the primary asphyxia (APGAR score 1/6/6) and a neonatal infection, the infant was ventilated for 5 days. A thrombocytopenia was present from the beginning, while a progressive pancytopenia developed over the following months. At the age of 3 months, the bone marrow was aplastic with scattered hemophagocytosis raising the suspicion of a connatal primary hemophagocytic lymphohistiocytosis. Immunomodulatory therapy with corticosteroids and cyclosporine A had no effect on the progressive bone marrow failure. Neurologically, the child was born with a severe microcephaly with primary cerebellar hypoplasia. Clinically, he demonstrated a developmental delay and progressive dysphagia. The brain MRI at the age of 4 and 8 months revealed severe brain atrophy with predominant pontocerebellar hypoplasia, pronounced delay in myelination, and multiple small infection-like subcortical white matter calcifications in the occipital region. Because of the radiomorphological phenotype, the differential diagnosis of a pseudotorch syndrome was discussed. At the age of 7 months a leukoplakia was seen on the tongue, characteristic of Hoyeraal-Hreidarsson syndrome. The child died at the age of 8 months in acute respiratory failure. Mutational analyses revealed a novel hemizygous variant c.1072 T> C, p.C358G in exon 11 of the DKC1 gene. Offering the correct diagnosis in this rare disease with a fulminant clinical course is important to offer genetic counseling to the family. Hoyeraal-Hreidarsson syndrome is a rare but important differential diagnosis in neonates with severe neurological and hematological symptoms. Keywords: neonatal Hoyeraal-Hreidarsson syndrome, severe developmental delay, progressive bone marrow failure.