Effect of physical activity and BMI SDS on bone metabolism in children and adolescents.

OBJECTIVE: Children with obesity are known to have reduced bone density and are at a higher risk for fractures. This may be caused by decreased physical activity or a metabolic phenomenon. In this study, we evaluated associations of physical activity with bone metabolism in children and adolescents with and without obesity. METHODS: Results from 574 visits of 397 subjects, 191 girls and 206 boys aged five to 18 years (mean: 11.7 ± 2.8) representing 180 children with (mean BMI SDS 2.5 ± 0.4) and 217 without obesity (mean BMI SDS 0.2 ± 1.0) from the LIFE Child study, a population-based cohort of children/adolescents with normal weight and with obesity were analyzed for the impact of their daily physical activity (MET/day, SenseWear Accelerometer) on serum SDS levels for bone formation (alkaline phosphatase, osteocalcin, procollagen type I N propeptide [P1NP]), bone resorption (beta-crosslaps), and calcium homeostasis (parathormone, OH-25-vitamin D) by a linear regression model adjusted for gender- and age-based differences. RESULTS: For male subjects, BMI SDS significantly influenced the association of physical activity to PTH, vitamin D, and beta-crosslaps SDS levels. A higher physical activity was accompanied by increased PTH but decreased vitamin D SDS levels in children with normal weight. In males with obesity, all levels remained unaltered. In females, BMI SDS significantly impacted the association of physical activity to PTH, vitamin D, P1NP, beta-crosslaps, and osteocalcin SDS levels. In females with obesity, higher physical activity was related to higher SDS levels of vitamin D, P1NP, and beta-crosslaps. In contrast, in normal weight females, only PTH SDS was higher. CONCLUSIONS: The effect of daily physical activity on bone metabolic markers and calciotropic hormones depends significantly on gender and BMI SDS. However, higher levels of physical activity were associated with increased bone turnover for female subjects with obesity only. Thus, motivating especially girls with obesity to be physically active may help improve their bone health.

Publication Trends and Global Collaborations on Esophageal Atresia Research: A Bibliometric Study.

INTRODUCTION: Research on esophageal atresia (EA) has been heavily published over the past decades. Herein, we aimed to study the quantity and quality as well as key topics in EA research with regards to global collaborations among countries and authors. MATERIALS AND METHODS: Publications on EA from 1945 to 2018 were extracted from the Web of Science core collection database. Productivity (quantity) was assessed by the number of publications. Quality was estimated from the number of citations, citation rate per item and year, h-index, and impact index. Collaborative networks were evaluated using VOSviewer. All measures were analyzed for countries, authors, and journals. The 10 most cited original articles between 1969 and 2018 in 5-year intervals (n = 100) were manually screened to assess the key points of EA research. RESULTS: A total of 2,170 publications from 85 countries published in 388 journals were identified yielding 26,755 citations, both significantly increasing over time (p < 0.001). The most productive countries and authors also accounted for high-quality publications and benefited from an active global network. The most productive journals derived from the field of pediatric surgery but accounted only for one-third of EA papers. The best cited journals were unspecific for pediatric surgery. Long-term outcome remained the most important topic in EA research, followed by surgical techniques, epidemiology, associated anomalies, perioperative complications, and postnatal management. In contrast, basic science was underrepresented. CONCLUSION: Over the past seven decades, EA publications increased tremendously. Productiveness and quality benefited from global networking. Long-term outcome remains the key interest of EA research.

High resolution three-dimensional imaging and measurement of lung, heart, liver, and diaphragmatic development in the fetal rat based on micro-computed tomography (micro-CT).

Understanding of normal fetal organ development is crucial for the evaluation of the pathogenesis of congenital anomalies. Various techniques have been used to generate imaging of fetal rat organogenesis, such as histological dissection with 3-dimensional reconstruction and scanning electron microscopy. However, these techniques did not imply quantitative measurements of developing organs (volumes, surface areas of organs). Furthermore, a partial or total destruction of the embryos prior to analysis was inevitable. Recently, micro-computed tomography (micro-CT) has been established as a novel tool to investigate embryonic development in non-dissected embryos of rodents. In this study, we used the micro-CT technique to generate 4D datasets of rat embryos aged between embryonic day 15-22 and newborns. Lungs, hearts, diaphragms, and livers were digitally segmented in order to measure organ volumes and analyze organ development as well as generate high-resolution 3D images. These data provide objective values compiling a 4D atlas of pulmonary, cardiac, diaphragmatic, and hepatic development in the fetal rat.

CHL1 and NrCAM are Primarily Expressed in Low Grade Pediatric Neuroblastoma.

BACKGROUND: Neural cell adhesion molecules like close homolog of L1 protein (CHL1) and neuronal glia related cell adhesion molecule (NrCAM) play an important role in development and regeneration of the central nervous system. However, they are also associated with cancerogenesis and progression in adult malignancies, thus gain increasing importance in cancer research. We therefore studied the expression of CHL1 and NrCAM according to the course of disease in children with neuroblastoma. METHODS: CHL1 and NrCAM expression levels were histologically assessed by tissue microarrays from surgically resected neuroblastoma specimens of 56 children. Expression of both markers was correlated to demographics as well as clinical data including metastatic dissemination and survival. RESULTS: CHL1 was expressed in 9% and NrCAM in 51% of neuroblastoma tissue samples. Expression of CHL1 was higher in patients with low Hughes grade 1a/b (p=0.01). NrCAM was more often detected in patients with a low International Staging System (INSS) score 1/2 (p=0.04). CONCLUSION: CHL1 and NrCAM expression was associated with low-grade pediatric neuroblastoma. These adhesion molecules may play a role in early tumor development of neuroblastoma.

Prenatal treatment with rosiglitazone attenuates vascular remodeling and pulmonary monocyte influx in experimental congenital diaphragmatic hernia.

INTRODUCTION: Extensive vascular remodeling causing pulmonary hypertension (PH) represents a major cause of mortality in patients with congenital diaphragmatic hernia (CDH). The chemokine monocyte chemoattractant protein-1 (MCP-1) is a biomarker for the severity of PH and its activation is accompanied by pulmonary influx of monocytes and extensive vascular remodeling. MCP-1 activation can be reversed by application of rosiglitazone (thiazolidinedione). We performed this study to evaluate the role of MCP-1 for the pathogenesis of PH in experimental CDH. We hypothesized that vascular remodeling and MCP-1 activation is accompanied by pulmonary influx of fetal monocytes and can be attenuated by prenatal treatment with rosiglitazone. METHODS: In a first set of experiments pregnant rats were treated with either nitrofen or vehicle on gestational day 9 (D9). Fetal lungs were harvested on D21 and divided into CDH and control. Quantitative real-time polymerase chain reaction, Western blot (WB), and immunohistochemistry (IHC) were used to evaluate MCP-1 expression, activation, and localization. Quantification and localization of pulmonary monocytes/macrophages were carried out by IHC. In a second set of experiments nitrofen-exposed dams were randomly assigned to prenatal treatment with rosiglitazone or placebo on D18+D19. Fetal lungs were harvested on D21, divided into control, CDH+rosiglitazone, and CDH+placebo and evaluated by WB as well as IHC. RESULTS: Increased thickness of pulmonary arteries of CDH fetuses was accompanied by increased systemic and perivascular MCP-1 protein expression and significantly higher amounts of pulmonary monocytes/macrophages compared to controls (p<0.01). These effects were reversed by prenatal treatment with rosiglitazone (p<0.01 vs. CDH+P; control). CONCLUSION: Prenatal treatment with rosiglitazone has the potential to attenuate activation of pulmonary MCP-1, pulmonary monocyte influx, and vascular remodeling in experimental CDH. These results provide a basis for future research on prenatal immunomodulation as a novel treatment strategy to decrease secondary effects of PH in CDH.

Increased L1CAM (CD171) levels are associated with glioblastoma and metastatic brain tumors.

L1 cell adhesion molecule (L1CAM) is a member of the immunoglobulin-like cell-adhesion molecule family that was shown to be associated with a worse prognosis in several human cancers. L1 ectodomain shedding via vesicles or exosomes has been detected in extracellular fluids after cleavage from the cell surface by metalloproteases. We evaluated the presence of L1CAM in cyst fluid and tissue from glioblastomas or brain metastases.The amount of L1CAM in cyst fluid of 9 glioblastomas and 11 brain metastases was assessed using enzyme-linked immunosorbent assay (ELISA). Corresponding tumor tissue slices were stained immunohistochemically for L1CAM. Cerebrospinal fluid of 20 non-tumor patients served as controls.Mean levels of L1CAM in tumor cyst fluid were significantly higher in glioblastoma (6118 ±â€Š4095 ng/mL) and metastasis patients (8001 ±â€Š6535 ng/mL) than in CSF of control patients (714 ±â€Š22 ng/mL). The immunohistochemical expression of L1CAM in corresponding tissue was significantly higher in metastases than in glioblastomas.The present study demonstrates high levels of L1CAM in cyst fluid of glioblastoma and metastatic brain tumors. Soluble L1CAM may represent a motility promoting molecule in cancer progression, a co-factor for development of tumor cysts and a target for new treatment strategies.

Physiologic Changes in a Small Animal Model for Neonatal Minimally Invasive Surgery.

BACKGROUND: Minimally invasive surgery (MIS) has gained increasing importance in neonatal surgery but the effects on neonatal physiology remain unclear. We aimed to characterize the impact of capnoperitoneum on physiologic parameters in a small animal model for neonatal MIS. MATERIAL AND METHODS: Twenty-four 10-day-old Sprague Dawley rats underwent inhalative anesthesia (1% isoflurane in 100% O2 250 mL/minutes) and were allowed to breathe spontaneously. CO2 was insufflated into the abdominal cavity for 1 hour via a 24G cannula. Anesthetized litter mates without insufflation served as sham controls, those without any treatment as external controls. Continuous monitoring included O2-saturation, heart and respiration rate, pulse and breath distension. After euthanasia, blood gas analysis was performed. RESULTS: All animals survived the experiment. Capnoperitoneum was best tolerated at a pressure of 2 mmHg and a flow of 0.5 L/minutes. A significant decrease in heart rate was observed within the first 30 minutes of insufflation comparing the CO2 and sham group (P < .05). In both, the CO2 and sham group, postmortem pH-levels were lower and pCO2 levels were higher compared to external controls (P < .05). Additionally, levels of pCO2 were higher but pH levels remained unchanged in the CO2 compared to sham group (P < .05). CONCLUSION: We established a small animal model for neonatal laparoscopy. A pressure of 2 mmHg and flow of 0.5 L/minutes induced physiologic alterations but was well tolerated by the animals. These settings can be used in future studies on the impact of the capnoperitoneum in neonatal MIS.

The oral microbiome-the relevant reservoir for acute pediatric appendicitis?

PURPOSE: The oral microbiome has been related to numerous extra oral diseases. Recent studies detected a high abundance of oral bacteria in inflamed appendices in pediatric patients. To elucidate the role of oral bacteria in acute pediatric appendicitis, we studied the oral and appendiceal microbiome of affected children compared to healthy controls. METHODS: Between January and June 2015, 21 children undergoing appendectomy for acute appendicitis and 28 healthy controls were prospectively enrolled in the study. All individuals underwent thorough dental examination and laboratory for inflammatory parameters. Samples of inflamed appendices and the gingival sulcus were taken for 16S rDNA sequencing. RT-qPCR of Fusobacterium nucleatum, Peptostreptococcus stomatis, and Eikenella corrodens was performed and their viability was tested under acidic conditions to mimic gastric transfer. RESULTS: In phlegmonous appendices, Bacteroidetes and Porphyromonas were discovered as dominant phylum and genus. In sulcus samples, Firmicutes and Streptococcus were detected predominantly. P. stomatis, E. corrodens, and F. nucleatum were identified in each group. Viable amounts of P. stomatis were increased in sulci of children with acute appendicitis compared to sulci of healthy controls. In inflamed appendices, viable amounts of E. corrodens and F. nucleatum were decreased compared to sulci of children with appendicitis. Postprandial viability could be demonstrated for all tested bacteria. CONCLUSION: In children with acute appendicitis, we identified several oral bacterial pathogens. Based on postprandial viability of selected species, a viable migration from the oral cavity through the stomach to the appendix seems possible. Thus, the oral cavity could be a relevant reservoir for acute appendicitis.

Dextran sodium sulfate (DSS) induces necrotizing enterocolitis-like lesions in neonatal mice.

BACKGROUND: Necrotizing enterocolitis (NEC) is an inflammatory bowel disease of preterm human newborns with yet unresolved etiology. An established neonatal murine model for NEC employs oral administration of lipopolysaccharides (LPS) combined with hypoxia/hypothermia. In adult mice, feeding dextran sodium sulfate (DSS) represents a well-established model for experimental inflammatory bowel disease. Here we investigated the effect of DSS administration on the neonatal murine intestine in comparison with the established NEC model. METHODS: 3-day-old C57BL/6J mice were either fed formula containing DSS or LPS. LPS treated animals were additionally stressed by hypoxia/hypothermia twice daily. After 72 h, mice were euthanized, their intestinal tissue harvested and analyzed by histology, qRT-PCR and flow cytometry. For comparison, adult C57BL/6J mice were fed with DSS for 8 days and examined likewise. Untreated, age matched animals served as controls. RESULTS: Adult mice treated with DSS exhibited colonic inflammation with significantly increased Cxcl2 mRNA expression. In contrast, tissue inflammation in neonatal mice treated with DSS or LPS plus hypoxia/hypothermia was present in colon and small intestine as well. Comparative analysis of neonatal mice revealed a significantly increased lesion size and intestinal Cxcl2 mRNA expression after DSS exposure. Whereas LPS administration mainly induced local neutrophil recruitment, DSS treated animals displayed increased monocytes/macrophages infiltration. CONCLUSIONS: Our study demonstrates the potential of DSS to induce NEC-like lesions accompanied by a significant humoral and cellular immune response in the small and large intestine of neonatal mice. The new model therefore represents a good alternative to LPS plus hypoxia/hypothermia administration requiring no additional physical stress.

The intra- and extraluminal appendiceal microbiome in pediatric patients: A comparative study.

Intestinal microbiota is involved in metabolic processes and the pathophysiology of various gastrointestinal disorders. We aimed to characterize the microbiome of the appendix in acute pediatric appendicitis comparing extraluminal and intraluminal samples.Between January and June 2015, 29 children (3-17 years, mean age 10.7 ±â€Š3.4 years, sex M:F = 2.6:1) undergoing laparoscopic appendectomy for acute appendicitis were prospectively included in the study. Samples for bacterial cultures (n = 29) and 16S ribosomal desoxyribonucleic acid (rDNA) sequencing (randomly chosen n = 16/29) were taken intracorporeally from the appendiceal surface before preparation ("extraluminal") and from the appendiceal lumen after removal ("intraluminal"). The degree of inflammation was histologically classified into catarrhal, phlegmonous, and gangrenous appendicitis.Seventeen bacterial species were cultivated in 28 of 29 intraluminal samples and 4 species were cultivated in 2 of 29 extraluminal samples. Using 16S rDNA sequencing, 267 species were detected in intraluminal but none in extraluminal samples. Abundance and diversity of detected species differed significantly between histological groups of acute appendicitis in bacterial cultures (P = .001), but not after 16S rDNA sequencing.The appendiceal microbiome showed a high diversity in acute pediatric appendicitis. The intraluminal microbial composition differed significantly depending on the degree of inflammation. As bacteria were rarely found extraluminally by culture and not at all by sequencing, the inflammation in acute appendicitis may start inside the appendix and spread transmurally.

Abolished perineuronal nets and altered parvalbumin-immunoreactivity in the nucleus reticularis thalami of wildtype and 3xTg mice after experimental stroke.

Treatment strategies for ischemic stroke are still limited, since numerous attempts were successful only in preclinical research but failed under clinical condition. To overcome this translational roadblock, clinical relevant stroke models should consider co-morbidities, age-related effects and the complex neurovascular unit (NVU) concept. The NVU includes neurons, vessels and glial cells with astrocytic endfeet in close relation to the extracellular matrix (ECM). However, the role of the ECM after stroke-related tissue damage is poorly understood and mostly neglected for treatment strategies. This study is focused on alterations of perineuronal nets (PNs) as ECM constituents and parvalbumin-containing GABAergic neurons in mice with emphasis on the nucleus reticularis thalami (NRT) in close proximity to the ischemic lesion as induced by a filament-based stroke model. One day after ischemia onset, immunofluorescence-based quantitative analyses revealed drastically declined PNs in the ischemia-affected NRT from 3- and 12-month-old wildtype and co-morbid triple-transgenic (3xTg) mice with Alzheimer-like alterations. Parvalbumin-positive cells decreased numerically in the ischemia-affected NRT, while staining intensity did not differ between the affected and non-affected hemisphere. Additional qualitative analyses demonstrated ischemia-induced loss of PNs and allocated neuropil ECM immunoreactive for aggrecan and neurocan, and impaired immunoreactivity for calbindin, the potassium channel subunit Kv3.1b and the glutamate decarboxylase isoforms GAD65 and GAD67 in the NRT. In conclusion, these data confirm PNs as highly sensitive constituents of the ECM along with impaired neuronal integrity of GABAergic neurons. Therefore, specific targeting of ECM components might appear as a promising strategy for future treatment strategies in stroke.

Protective Properties of Neural Extracellular Matrix.

The extracellular matrix (ECM) of the central nervous system (CNS) occupies a large part of the neural tissue. It serves a variety of functions ranging from support of cell migration and regulating synaptic transmission and plasticity to the active modulation of the neural tissue after injury. In addition, evidence for neuroprotective properties of ECM components has accumulated more recently. In contrast to other connective tissues, the central nervous ECM is mainly composed of glycosaminoglycans, which can be present unbound in the form of hyaluronan or bound to proteins, thus forming proteoglycans. A subtype of this molecular family are the chondroitin sulphate proteoglycans (CSPGs), which are composed of a core protein that carries at least one covalently bound glycosaminoglycan side chain with a certain degree of sulphation. Several studies could show neuroprotective features of CSPGs against excitotoxicity, amyloid-ß toxicity, or oxidative stress. Recently, we could provide evidence for a neuroprotective function of a specialized form of ECM, the so-called perineuronal net ensheathing a subtype of neurons. Here, we will give an overview on recently emerging aspects of neuroprotective properties of CSPGs and perineuronal nets that might be relevant for our understanding on the distribution and progression of brain pathology and future perspectives toward modifying neurodegenerative diseases.

Hook proteins: association with Alzheimer pathology and regulatory role of hook3 in amyloid beta generation.

Defects in intracellular transport are implicated in the pathogenesis of Alzheimer's disease (AD). Hook proteins are a family of cytoplasmic linker proteins that participate in endosomal transport. In this study we show that Hook1 and Hook3 are expressed in neurons while Hook2 is predominantly expressed in astrocytes. Furthermore, Hook proteins are associated with pathological hallmarks in AD; Hook1 and Hook3 are localized to tau aggregates and Hook2 to glial components within amyloid plaques. Additionally, the expression of Hook3 is reduced in AD. Modelling of Hook3 deficiency in cultured cells leads to slowing of endosomal transport and increases ß-amyloid production. We propose that Hook3 plays a role in pathogenic events exacerbating AD.

RVG peptide as transfection reagent for specific cdk4 gene silencing in vitro and in vivo.

The rabies virus glycoprotein (RVG) peptide is known as a transfection reagent for systemic delivery of small interfering RNA (siRNA) into the brain. However, selective transfection of neuronal cells or specific brain regions remains a problem. In the present study, we show that the RVG peptide can efficiently be used as shuttle system to transfect neuronal cells with cdk4 siRNA leading to selective knockdown of cdk4 expression in vitro and in vivo. After transfection, cdk4 expression was reduced up to 75% in Neuro2A cells. Stereotactically injected RVG peptide delivered cdk4 siRNA specifically to neurons in the hippocampus, resulting in a specific knockdown of cdk4 expression up to 400 µm from the injection site. Further complexation studies of RVG peptide with larger molecules such as plasmid vectors or DNA fragments were also successfully performed and improved in vitro. Therefore, the peptide is not only a highly promising drug delivery system for siRNA and potentially other therapeutic molecules, but also a powerful tool to systematically analyze gene function in the brain under experimental settings in correlation to neurodegenerative disorders.

Neuroprotection against iron-induced cell death by perineuronal nets - an in vivo analysis of oxidative stress.

Perineuronal nets (PNs) are a specialized form of extracellular matrix, surrounding different types of neurons and mainly consist of chondroitin sulfate proteoglycans connected to hyaluronan, stabilized by link protein and cross-linked via tenascin-R. Due to their polyanionic character, caused by the highly charged chondroitin sulfate glycosaminoglycan and hyaluronan components, PNs might be involved in local ion homeostasis. They are able to scavenge and bind redox-active ions and thus reduce the local oxidative potential. We investigated whether netenwrapped neurons are less vulnerable against iron-induced oxidative processes. Oxidative stress is a key factor in the development and progression of neurodegenerative diseases like Alzheimer's and Parkinson's disease. Iron is believed to contribute to oxidative stress in Alzheimer brains by catalyzing the generation of free radicals. For examining potential neuroprotective effects of PNs, mice were microinjected with 0.2µl of a 20mM solution of FeCl3 into the barrel field while the control group received an equal volume of 0.9% NaCl. Brains were analyzed after time intervals of 24h and 72h. Neuronal degeneration was visualized using Fluoro-Jade B staining. The presence of PNs was assessed by Wisteria floribunda agglutinin histochemistry or aggrecan immunocytochemistry. The analysis showed a significant lower degeneration rate of net-ensheathed neurons in comparison to neurons without PNs. The results suggest a neuroprotective mechanism associated with the presence of PNs against iron-induced cell death.