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Mesenchymal stromal cells (MSCs) are released into the airways of preterm infants following lung injury. These cells display a proinflammatory phenotype and are associated with development of severe bronchopulmonary dysplasia (BPD). We aimed to characterize the functional properties of MSCs obtained from tracheal aspirates of 50 preterm infants who required invasive ventilation. Samples were separated by disease severity. The increased proliferative capacity of MSCs was associated with longer duration of mechanical ventilation and higher severity of BPD. Augmented growth depended on nuclear accumulation of NFκBp65 and was accompanied by reduced expression of cytosolic α-smooth muscle actin (α-SMA). The central role of NF-κB signaling was confirmed by inhibition of IκBα phosphorylation. The combined score of proliferative capacity, accumulation of NFκBp65, and expression of α-SMA was used to predict the development of severe BPD with an area under the curve (AUC) of 0.847. We mimicked the clinical situation in vitro, and stimulated MSCs with IL-1β and TNF-α. Both cytokines induced similar and persistent changes as was observed in MSCs obtained from preterm infants with severe BPD. RNA interference was employed to investigate the mechanistic link between NFκBp65 accumulation and alterations in phenotype. Our data indicate that determining the phenotype of resident pulmonary MSCs represents a promising biomarker-based approach. The persistent alterations in phenotype, observed in MSCs from preterm infants with severe BPD, were induced by the pulmonary inflammatory response. NFκBp65 accumulation was identified as a central regulatory mechanism. Future preclinical and clinical studies, aimed to prevent BPD, should focus on phenotype changes in pulmonary MSCs.

Preponderance of proinflammatory signals is a characteristic feature of all acute and resulting long-term morbidities of the preterm infant. The proinflammatory actions are best characterized for bronchopulmonary dysplasia (BPD) which is the chronic lung disease of the preterm infant with lifelong restrictions of pulmonary function and severe consequences for psychomotor development and quality of life. Besides BPD, the immature brain, eye, and gut are also exposed to inflammatory injuries provoked by infection, mechanical ventilation, and oxygen toxicity. Despite the tremendous progress in the understanding of disease pathologies, therapeutic interventions with proven efficiency remain restricted to a few drug therapies with restricted therapeutic benefit, partially considerable side effects, and missing option of applicability to the inflamed brain. The therapeutic potential of mesenchymal stromal cells (MSCs)-also known as mesenchymal stem cells-has attracted much attention during the recent years due to their anti-inflammatory activities and their secretion of growth and development-promoting factors. Based on a molecular understanding, this review summarizes the positive actions of exogenous umbilical cord-derived MSCs on the immature lung and brain and the therapeutic potential of reprogramming resident MSCs. The pathomechanistic understanding of MSC actions from the animal model is complemented by the promising results from the first phase I clinical trials testing allogenic MSC transplantation from umbilical cord blood. Despite all the enthusiasm towards this new therapeutic option, the caveats and outstanding issues have to be critically evaluated before a broad introduction of MSC-based therapies.

Neonatal chronic lung disease (nCLD) affects a significant number of neonates receiving mechanical ventilation with oxygen-rich gas (MV-O 2 ). Regardless, the primary molecular driver of the disease remains elusive. We discover significant enrichment for SNPs in the PDGF-Rα gene in preterms with nCLD and directly test the effect of PDGF-Rα haploinsufficiency on the development of nCLD using a preclinical mouse model of MV-O 2 In the context of MV-O 2 , attenuated PDGF signaling independently contributes to defective septation and endothelial cell apoptosis stemming from a PDGF-Rα-dependent reduction in lung VEGF-A. TGF-β contributes to the PDGF-Rα-dependent decrease in myofibroblast function. Remarkably, endotracheal treatment with exogenous PDGF-A rescues both the lung defects in haploinsufficient mice undergoing MV-O 2 Overall, our results establish attenuated PDGF signaling as an important driver of nCLD pathology with provision of PDGF-A as a protective strategy for newborns undergoing MV-O 2 ., (© 2017 Helmholtz Zentrum München Published under the terms of the CC BY 4.0 license.)

Background: An alloimmune response to red blood cell (RBC) transfusion in neonates is a rare event. Several guidelines recommend limited pretransfusion testing in neonates. The evidence for these recommendations is based on small studies with sample sizes of between 30 and 90 infants., Study Design and Methods: We conducted a retrospective cohort study among consecutive patients who received transfusions at a single university medical center. All non-alloimmunized patients who had received their first RBC transfusion between 1994 and 2013 and who underwent at least one antibody screening follow-up visit between 7 and 365 days after transfusion were included., Results: The incidence of alloimmunization in the control group of 17,084 adult patients age 45 years or older who had received a median of 5 RBC units (interquartile range, 2-12 RBC units) was 3.55% (n = 607 alloimmunized patients). After transfusion of 40 RBC units, the cumulative incidence of alloimmunization in adult controls was 10.24% (95% confidence interval, 7.71%-13.17%). In total, 1641 neonates and children up to age 3 years received a median of 4 RBC units (interquartile range, 2-7 RBC units) in a median of two RBC transfusion episodes (interquartile range, one to five RBC transfusion episodes). Two children developed anti-M and anti-E antibodies post-transfusion at the ages of 181 and 611 days, respectively., Conclusion: To our knowledge, this study presents the largest longitudinal cohort study of RBC alloimmunization in neonates. Antibodies against RBC antigens were not detected within the first 6 months of life. Repeat antibody screening and cross-matching during the first months of life can be safely omitted., (© 2017 AABB.)

Early onset infection (EOI) in preterm infants <32 weeks gestational age (GA) is associated with a high mortality rate and the development of severe acute and long-term complications. The pathophysiology of EOI is not fully understood and clinical and laboratory signs of early onset infections in this patient cohort are often not conclusive. Thus, the aim of this study was to identify signatures characterizing preterm infants with EOI by using genome-wide gene expression (GWGE) analyses from umbilical arterial blood of preterm infants. This prospective cohort study was conducted in preterm infants <32 weeks GA. GWGE analyses using CodeLink human microarrays were performed from umbilical arterial blood of preterm infants with and without EOI. GWGE analyses revealed differential expression of 292 genes in preterm infants with EOI as compared to infants without EOI. Infants with EOI could be further differentiated into two subclasses and were distinguished by the magnitude of the expression of genes involved in both neutrophil and T cell activation. A hallmark activity for both subclasses of EOI was a common suppression of genes involved in natural killer (NK) cell function, which was independent from NK cell numbers. Significant results were recapitulated in an independent validation cohort. Gene expression profiling may enable early and more precise diagnosis of EOI in preterm infants., Key Message: Gene expression (GE) profiling at birth characterizes preterm infants with EOI. GE analysis indicates dysregulation of NK cell activity. NK cell activity at birth may be a useful marker to improve early diagnosis of EOI., Competing Interests: Compliance with ethical standards Funding The present study was supported by the National Genome Network (NGFN), Germany (NGFN IE-S08T03) and by the Pneumonia Research Network on Genetic Resistance and Susceptibility for the Evolution of Severe Sepsis (PROGRESS), grant number 01KI07110. Conflict of interest The authors declare no conflict of interests related to this study.

Inflammation-induced FGF10 protein deficiency is associated with bronchopulmonary dysplasia (BPD), a chronic lung disease of prematurely born infants characterized by arrested alveolar development. So far, experimental evidence for a direct role of FGF10 in lung disease is lacking. Using the hyperoxia-induced neonatal lung injury as a mouse model of BPD, the impact of Fgf10 deficiency in Fgf10 +/- versus Fgf10 +/+ pups was investigated. In normoxia, no lethality of Fgf10 +/+ or Fgf10 +/- pups was observed. By contrast, all Fgf10 +/- pups died within 8 days of hyperoxic injury, with lethality starting at day 5, whereas Fgf10 +/+ pups were all alive. Lungs of pups from the two genotypes were collected on postnatal day 3 following normoxia or hyperoxia exposure for further analysis. In hyperoxia, Fgf10 +/- lungs exhibited increased hypoalveolarization. Analysis by FACS of the Fgf10 +/- versus control lungs in normoxia revealed a decreased ratio of alveolar epithelial type II (AECII) cells over total Epcam-positive cells. In addition, gene array analysis indicated reduced AECII and increased AECI transcriptome signatures in isolated AECII cells from Fgf10 +/- lungs. Such an imbalance in differentiation is also seen in hyperoxia and is associated with reduced mature surfactant protein B and C expression. Attenuation of the activity of Fgfr2b ligands postnatally in the context of hyperoxia also led to increased lethality with decreased surfactant expression. In summary, decreased Fgf10 mRNA levels lead to congenital lung defects, which are compatible with postnatal survival, but which compromise the ability of the lungs to cope with sub-lethal hyperoxic injury. Fgf10 deficiency affects quantitatively and qualitatively the formation of AECII cells. In addition, Fgfr2b ligands are also important for repair after hyperoxia exposure in neonates. Deficient AECII cells could be an additional complication for patients with BPD. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., Competing Interests: The authors disclose that they have no conflict of interest., (Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Bronchopulmonary dysplasia is a chronic lung disease of preterm infants. It is caused by the disturbance of physiologic lung development mainly in the saccular stage with lifelong restrictions of pulmonary function and an increased risk of abnormal somatic and psychomotor development. The contributors to this disease's entity are multifactorial with pre- and postnatal origin. Central to the pathogenesis of bronchopulmonary is the induction of a massive pulmonary inflammatory response due to mechanical ventilation and oxygen toxicity. The extent of the pro-inflammatory reaction and the disturbance of further alveolar growth and vasculogenesis vary largely and can be modified by prenatal infections, antenatal steroids, and surfactant application.This minireview summarizes the important recent research findings on the pulmonary inflammatory reaction obtained in patient cohorts and in experimental models. Unfortunately, recent changes in clinical practice based on these findings had only limited impact on the incidence of bronchopulmonary dysplasia.

Bronchopulmonary dysplasia (BPD), characterized by impaired alveolarization and vascularization in association with lung inflammation and apoptosis, often occurs after mechanical ventilation with oxygen-rich gas (MV-O2). As heightened expression of the proinflammatory cytokine TNF-α has been described in infants with BPD, we hypothesized that absence of TNF-α would reduce pulmonary inflammation, and attenuate structural changes in newborn mice undergoing MV-O2 Neonatal TNF-α null (TNF-α(-/-)) and wild type (TNF-α(+/+)) mice received MV-O2 for 8 h; controls spontaneously breathed 40% O2 Histologic, mRNA, and protein analysis in vivo were complemented by in vitro studies subjecting primary pulmonary myofibroblasts to mechanical stretch. Finally, TNF-α level in tracheal aspirates from preterm infants were determined by ELISA. Although MV-O2 induced larger and fewer alveoli in both, TNF-α(-/-) and TNF-α(+/+) mice, it caused enhanced lung apoptosis (TUNEL, caspase-3/-6/-8), infiltration of macrophages and neutrophils, and proinflammatory mediator expression (IL-1β, CXCL-1, MCP-1) in TNF-α(-/-) mice. These differences were associated with increased pulmonary transforming growth factor-β (TGF-β) signaling, decreased TGF-β inhibitor SMAD-7 expression, and reduced pulmonary NF-κB activity in ventilated TNF-α(-/-) mice. Preterm infants who went on to develop BPD showed significantly lower TNF-α levels at birth. Our results suggest a critical balance between TNF-α and TGF-β signaling in the developing lung, and underscore the critical importance of these key pathways in the pathogenesis of BPD. Future treatment strategies need to weigh the potential benefits of inhibiting pathologic cytokine expression against the potential of altering key developmental pathways., (Copyright © 2016 the American Physiological Society.)

Objective: To assess maternal morbidity and outcome in women undergoing minimal-access fetoscopic surgery for spina bifida aperta., Methods: This was a retrospective study of 51 women undergoing minimal-access fetoscopic surgery to improve postnatal neurological outcome of spina bifida aperta, at a mean gestational age of 24 weeks, at our center between July 2010 and June 2013. We analyzed various perioperative complications of surgery, namely: maternal and fetal death, need for maternal blood transfusion, placental abruption, pulmonary edema, spontaneous labor, oligohydramnios, chorioamnionitis, chorioamniotic membrane separation, duration of hospitalization, amniotic fluid leakage, gestational age at delivery and status of hysterotomy site., Results: In none of the 51 women was there maternal demise, spontaneous labor, placental abruption or a need for maternal blood transfusion in the perioperative period. Chorioamniotic membrane separation occurred in one patient, mild pulmonary edema occurred in one and oligohydramnios occurred in seven. All fetuses survived surgery, but there was one very early preterm delivery 1 week after the procedure and this neonate died immediately, from early postoperative chorioamnionitis. Amniotic fluid leakage occurred in 43 patients, at a mean gestational age of 29.7 (range, 22.6-37.3) weeks; two of these patients developed chorioamnionitis. Duration of maternal hospitalization after surgery was 7.2 (range, 4-12) days. Mean gestational age at delivery was 33 (range, 24.6-38.1) weeks. All abdominal and uterine trocar insertion sites healed well., Conclusion: Minimal-access fetoscopic surgery for spina bifida aperta is apparently safe for most maternal patients. Despite the common occurrence of amniotic leakage, the majority of women deliver beyond 32 weeks of gestation., (Copyright © 2014 ISUOG. Published by John Wiley & Sons Ltd.)

Neonatal chronic lung disease, also known as bronchopulmonary dysplasia (BPD), is the most common complication of premature birth, affecting up to 30% of very low birth weight infants. Improved medical care has allowed for the survival of the most premature infants and has significantly changed the pathology of BPD from a disease marked by severe lung injury to the "new" form characterized by alveolar hypoplasia and impaired vascular development. However, increased patient survival has led to a paucity of pathologic specimens available from infants with BPD. This, combined with the lack of a system to model alveolarization in vitro, has resulted in a great need for animal models that mimic key features of the disease. To this end, a number of animal models have been created by exposing the immature lung to injuries induced by hyperoxia, mechanical stretch, and inflammation and most recently by the genetic modification of mice. These animal studies have 1) allowed insight into the mechanisms that determine alveolar growth, 2) delineated factors central to the pathogenesis of neonatal chronic lung disease, and 3) informed the development of new therapies. In this review, we summarize the key findings and limitations of the most common animal models of BPD and discuss how knowledge obtained from these studies has informed clinical care. Future studies should aim to provide a more complete understanding of the pathways that preserve and repair alveolar growth during injury, which might be translated into novel strategies to treat lung diseases in infants and adults.

Objective: Evaluation of the effect of the train-the-trainer programme 'Fit for Shared Decision-Making' on internal (team) and external (patient) participation in medical rehabilitation from a patient and staff perspective., Design: A multicentre, cluster-randomized controlled study., Setting: Eleven medical rehabilitation clinics, divided into intervention and control groups., Subjects: A staff and a patient survey were conducted pre- and post-intervention, plus a further patient survey six months later., Intervention: Train-the-trainer programme 'Fit for Shared Decision-Making' for interprofessional settings., Main Measures: Each survey measured internal participation with a self-compiled six-item scale (Internal Participation Scale, IPS), and external participation by means of a nine-item Shared Decision-Making Questionnaire (SDM-Q-9) for the patients and for healthcare professionals., Results: Patient samples numbered 402 for the pre-, 463 for the post-intervention data collection period and 461 six months after the intervention. Patients' appraisal of external participation (Fperiod x group (2) = 0.256, p=0.774, η(2)=0.000) showed no change, whereas internal participation (Fperiod x group (2) = 3.785, p=0.023, η(2)=0.007) showed a significant increase. A total of 195 healthcare professionals participated in the pre- and 168 in the post-intervention staff survey. Here external participation was significantly enhanced in the intervention group (F(period x group) (1) = 4.893, p=0.028, η(2)=0.014)., Conclusions: The train-the-trainer approach can be recommended for implementing internal and external participation in interprofessional settings such as medical rehabilitation clinics. However, there is a need for more intensive staff training for internal participation and an additional intervention for patients to achieve success in all aspects.

Background/aim: Anthracyclines have been proven able to reduce the activity of vinca alkaloids by induction of cell-cycle arrest. The present study aims at identifying the critical initiation steps of signal transduction which transduce the inhibitory effects on the cytotoxicity of vinca alkaloids., Materials and Methods: Several new cytostatic drug classes were evaluated together with vincristine in tumor cell lines and patients' tumor cells. RNA interference was used for molecular analyses., Results: Inhibition of vincristine was observed by all cytostatic drugs, which induced cell-cycle arrest. Knockdown of proteins of the DNA damage response ascribed the inhibitory effect to a common pathway involving Chk-1, p53 and p21. Upstream of Chk-1 signal transduction depended on both ATM and ATR for all drugs except methotrexate., Conclusion: We have identified critical signaling steps of the DNA damage response system activated by cytostatic drugs, which reduce the anti-tumor activity of vinca alkaloids. The obtained results encourage the development of novel therapeutic strategies to prevent pathway interactions based on the molecular understanding of drug action and drug-drug interactions.

Resting tumor cells represent a huge challenge during anticancer therapy due to their increased treatment resistance. TNF-related apoptosis-inducing ligand (TRAIL) is a putative future anticancer drug, currently in phases I and II clinical studies. We recently showed that TRAIL is able to target leukemia stem cell surrogates. Here, we tested the ability of TRAIL to target cell cycle-arrested tumor cells. Cell cycle arrest was induced in tumor cell lines and xenografted tumor cells in G0, G1 or G2 using cytotoxic drugs, phase-specific inhibitors or RNA interference against cyclinB and E. Biochemical or molecular arrest at any point of the cell cycle increased TRAIL-induced apoptosis. Accordingly, when cell cycle arrest was disabled by addition of caffeine, the antitumor activity of TRAIL was reduced. Most important for clinical translation, tumor cells from three children with B precursor or T cell acute lymphoblastic leukemia showed increased TRAIL-induced apoptosis upon knockdown of either cyclinB or cyclinE, arresting the cell cycle in G2 or G1, respectively. Taken together and in contrast to most conventional cytotoxic drugs, TRAIL exerts enhanced antitumor activity against cell cycle-arrested tumor cells. Therefore, TRAIL might represent an interesting drug to treat static-tumor disease, for example, during minimal residual disease.

Background: The p53 protein is the best studied target in human cancer. For decades, p53 has been believed to act mainly as a tumor suppressor and by transcriptional regulation. Only recently, the complex and diverse function of p53 has attracted more attention. Using several molecular approaches, we studied the impact of different p53 variants on extrinsic and intrinsic apoptosis signaling., Results: We reproduced the previously published results within intrinsic apoptosis induction: while wild-type p53 promoted cell death, different p53 mutations reduced apoptosis sensitivity. The prediction of the impact of the p53 status on the extrinsic cell death induction was much more complex. The presence of p53 in tumor cell lines and primary xenograft tumor cells resulted in either augmented, unchanged or reduced cell death. The substitution of wild-type p53 by mutant p53 did not affect the extrinsic apoptosis inducing capacity., Conclusions: In summary, we have identified a non-expected impact of p53 on extrinsic cell death induction. We suggest that the impact of the p53 status of tumor cells on extrinsic apoptosis signaling should be studied in detail especially in the context of therapeutic approaches that aim to restore p53 function to facilitate cell death via the extrinsic apoptosis pathway.

Background and Purpose: In polychemotherapy protocols, that is for treatment of neuroblastoma and Ewing sarcoma, Vinca alkaloids and cell cycle-arresting drugs are usually administered on the same day. Here we studied whether this combination enables the optimal antitumour effects of Vinca alkaloids to be manifested., Experimental Approach: Vinca alkaloids were tested in a preclinical mouse model in vivo and in vitro in combination with cell cycle-arresting drugs. Signalling pathways were characterized using RNA interference., Key Results: In vitro, knockdown of cyclins significantly inhibited vincristine-induced cell death indicating, in accordance with previous findings, Vinca alkaloids require active cell cycling and M-phase transition for induction of cell death. In contrast, anthracyclines, irradiation and dexamethasone arrested the cell cycle and acted like cytostatic drugs. The combination of Vinca alkaloids with cytostatic therapeutics resulted in diminished cell death in 31 of 36 (86%) tumour cell lines. In a preclinical tumour model, anthracyclines significantly inhibited the antitumour effect of Vinca alkaloids in vivo. Antitumour effects of Vinca alkaloids in the presence of cytostatic drugs were restored by caffeine, which maintained active cell cycling, or by knockdown of p53, which prevented drug-induced cell cycle arrest. Therapeutically most important, optimal antitumour effects were obtained in vivo upon separating the application of Vinca alkaloids from cytostatic therapeutics., Conclusion and Implications: Clinical trials are required to prove whether Vinca alkaloids act more efficiently in cancer patients if they are applied uncoupled from cytostatic therapies. On a conceptual level, our data suggest the implementation of polychemotherapy protocols based on molecular mechanisms of drug-drug interactions., Linked Article: This article is commented on by Solary, pp 1555-1557 of this issue. To view this commentary visit http://dx.doi.org/10.1111/bph.12101., (© 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.)

For implementation of patient-centered treatment in interprofessional health care units, such as rehabilitation teams, external participation (interaction between patient and health care professionals) and internal participation (communication, coordination and cooperation in the interprofessional team) need to be considered. The aim of this study is to identify the preferences of patients and health care professionals concerning internal and external participation in rehabilitation clinics, in order to develop an interprofessional shared decision-making (SDM) training program for health care professionals to enhance both types of participation. Therefore, a cross-sectional mixed-methods study was implemented in four rehabilitation clinics. The study consists of two parts: focus groups with patients and a survey of experts (senior health care professionals from medicine, psychotherapy, physical therapy and nursing). More time, more respect from the health care professionals and the desire for more participation in decision-making processes were mentioned most frequently by patients (n = 36) in the focus groups. The health care professionals (n = 32) saw most deficits in internal participation, e.g. management of feedback, talking with difficult team members and moderate conflict discussion. The results of both assessments have been used to develop an interprofessional SDM training program for implementing internal and external participation in interprofessional teams in medical rehabilitation.

Objective: The aim of the study was to evaluate the interprofessional SDM training program "Fit for SDM" in medical rehabilitation, which was implemented in two steps: (1) university staff trained providers in executive positions as trainers and (2) the providers trained their staff., Methods: For the evaluation of the first step a questionnaire for shared decision-making (SDM) skills and satisfaction with the training was completed by the providers in executive positions. A staff survey was used in a cluster-randomized controlled study to determine the overall impact of the train-the-trainer program on internal and external participation in the team., Results: The providers in the six clinics evaluated their SDM competences and satisfaction very positively after training (step 1). External participation was enhanced by application of the training content, with significant changes recorded for females and nurses in particular. However, it had no direct influence on internal participation., Conclusions: This is the first interprofessional SDM train-the-trainer program in Germany to bridge interprofessionalism (internal participation) and SDM (external participation); it was implemented successfully and evaluated positively., Practice Implications: Establishing interprofessional SDM training programs should be encouraged for all health care professionals. Implementation in the interprofessional setting should consider interprofessional team factors., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Background: Glucocorticoids are widely used for cancer patients, although they can reduce the efficacy of anticancer treatment., Materials and Methods: We characterized non-apoptotic actions of glucocorticoids on tumor cell lines, primary tumor cells and an in vivo model, together with molecular signaling studies., Results: Glucocorticoids enhanced cell proliferation in 9/17 cell lines and significantly promoted tumor cell proliferation in a pre-clinical mouse model of lung carcinoma. 65/139 primary acute childhood leukemia samples were glucocorticoid-resistant. Both dexamethasone and prednisolone increased in vitro survival in 21/65 samples from glucocorticoid-resistant primary leukemias, revealing a completely new action of glucocorticoids. Dexamethasone-induced proliferation was mediated by glucocorticoid receptor and activated the proliferation signaling pathways of protein kinase B/AKT and p38 mitogen-activated protein kinase., Conclusion: Our data suggest that restriction of the use of glucocorticoids during anticancer treatment might improve the outcome of patients with solid tumors.

During polychemotherapy, cytotoxic drugs are given in combinations to enhance their anti-tumor effectiveness. For most drug combinations, underlying signaling mechanisms responsible for positive drug-drug interactions remain elusive. Here, we prove a decisive role for the Bcl-2 family member NOXA to mediate cell death by certain drug combinations, even if drugs were combined which acted independently from NOXA, when given alone. In proof-of-principle studies, betulinic acid, doxorubicin and vincristine induced cell death in a p53- and NOXA-independent pathway involving mitochondrial pore formation, release of cytochrome c and caspase activation. In contrast, when betulinic acid was combined with either doxorubicine or vincristine, cell death signaling changed considerably; the drug combinations clearly depended on both p53 and NOXA. Similarly and of high clinical relevance, in patient-derived childhood acute leukemia samples the drug combinations, but not the single drugs depended on p53 and NOXA, as shown by RNA interference studies in patient-derived cells. Our data emphasize that NOXA represents an important target molecule for combinations of drugs that alone do not target NOXA. NOXA might have a special role in regulating apoptosis sensitivity in the complex interplay of polychemotherapy. Deciphering the differences in signaling of single drugs and drug combinations might enable designing highly effective novel polychemotherapy regimens.

Background: Signaling studies in cell lines are hampered by non-physiological alterations obtained in vitro. Physiologic primary tumor cells from patients with leukemia require passaging through immune-compromised mice for amplification. The aim was to enable molecular work in patients' ALL cells by establishing siRNA transfection into cells amplified in mice., Results: We established delivering siRNA into these cells without affecting cell viability. Knockdown of single or multiple genes reduced constitutive or induced protein expression accompanied by marked signaling alterations., Conclusion: Our novel technique allows using patient-derived tumor cells instead of cell lines for signaling studies in leukemia.

Background: Xenograft mouse models represent helpful tools for preclinical studies on human tumors. For modeling the complexity of the human disease, primary tumor cells are by far superior to established cell lines. As qualified exemplary model, patients' acute lymphoblastic leukemia cells reliably engraft in mice inducing orthotopic disseminated leukemia closely resembling the disease in men. Unfortunately, disease monitoring of acute lymphoblastic leukemia in mice is hampered by lack of a suitable readout parameter., Design and Methods: Patients' acute lymphoblastic leukemia cells were lentivirally transduced to express the membrane-bound form of Gaussia luciferase. In vivo imaging was established in individual patients' leukemias and extensively validated., Results: Bioluminescence in vivo imaging enabled reliable and continuous follow-up of individual mice. Light emission strictly correlated to post mortem quantification of leukemic burden and revealed a logarithmic, time and cell number dependent growth pattern. Imaging conveniently quantified frequencies of leukemia initiating cells in limiting dilution transplantation assays. Upon detecting a single leukemia cell within more than 10,000 bone marrow cells, imaging enabled monitoring minimal residual disease, time to tumor re-growth and relapse. Imaging quantified therapy effects precisely and with low variances, discriminating treatment failure from partial and complete responses., Conclusions: For the first time, we characterized in detail how in vivo imaging reforms preclinical studies on patient-derived tumors upon increasing monitoring resolution. In the future, in vivo imaging will enable performing precise preclinical studies on a broad range of highly demanding clinical challenges, such as treatment failure, resistance in leukemia initiating cells, minimal residual disease and relapse.

Purpose: Sensitivity of tumor cells toward chemotherapy mainly determines the prognosis of patients suffering from acute lymphoblastic leukemia (ALL); nevertheless, underlying mechanisms regulating chemosensitivity remain poorly understood. Here, we aimed at characterizing the role of caspase-8 for chemosensitivity of B- and T-ALL cells., Experimental Design: Primary tumor cells from children with ALL were evaluated for expression levels of the caspase-8 protein, were amplified in nonobese diabetic/severe combined immunodeficient mice, transfected with siRNA, and evaluated for their chemosensitivity in vitro., Results: Effective cell death in B- and T-ALL cells depended on the presence of caspase-8 for the majority of cytotoxic drugs routinely used in antileukemia treatment. Caspase-8 was activated independently from extrinsic apoptosis signaling. Accordingly in primary ALL cells, the expression level of caspase-8 protein correlated with cell death sensitivity toward defined cytotoxic drugs in vitro. In the subgroup of primary ALL cells, with low expression of caspase-8, methotrexate (MTX) upregulated the expression of caspase-8 mediated by the transcription factor p53, suggesting epigenetic silencing of caspase-8. RNA interference in patient-derived B- and T-ALL cells revealed that effective cell death induction by most routine drug combinations involving MTX depended on the presence of caspase-8., Conclusion: Our results indicate that caspase-8 is crucial for the high antileukemic efficiency of numerous routine cytotoxic drugs. Reexpression of epigenetically downregulated caspase-8 represents a promising approach to increase efficiency of antileukemic therapy., (©2011 AACR.)

Application of anthracyclines and Vinca alkaloids on the same day represents a hallmark of polychemotherapy protocols for hematopoietic malignancies. Here we show, for the first time, that both drugs might act most efficiently if they are applied on different days. Proof-of-concept studies in 18 cell lines revealed that anthracyclines inhibited cell death by Vinca alkaloids in 83% of cell lines. Importantly, in a preclinical mouse model, doxorubicin reduced the anti-tumor effect of vincristine. Both drugs acted in a sequence-dependent manner and the strongest anti-tumor effect was obtained if both drugs were applied on different days. Most notably for clinical relevance, in 34% of 35 fresh primary childhood leukemia cells tested in vitro, doxorubicin reduced the anti-tumor effect of vincristine. As underlying mechanism, doxorubicin activated p53, p53 induced cell-cycle arrest, and cell-cycle arrest disabled inactivation of antiapoptotic Bcl-2 family members by vincristine; therefore, vincristine was unable to activate downstream apoptosis signaling. As molecular proof, antagonism was rescued by knockdown of p53, whereas knockdown of cyclin A inhibited vincristine-induced apoptosis. Our data suggest evaluating anthracyclines and Vinca alkaloids on different days in future trials. Selecting drug combinations based on mechanistic understanding represents a novel conceptional strategy for potent polychemotherapy protocols.

Background and Aims: Until now, training programmes on shared decision-making (SDM) have been designed exclusively for medical decision-making and predominantly for physicians. How-ever, interprofessional treatment, such as in medical rehabilitation, is very important in the treatment of chronic diseases. This requires an extended understanding of shared decision-making. Therefore the aim of the study is to develop an interprofessional training (IPT) for implementation of shared decision-making (SDM) in rehabilitation clinics., Methods: The needs and preferences of the persons undergoing rehabilitation were collected in 4 focus groups, with frequency of answers analyzed by means of inductive category formation. The providers' preferences and requirements concerning a training programme on shared decision-making were assessed through an expert survey and underwent a mainly descriptive-explorative evaluation as well as a partial content analysis., Results: 36 patients took part in the focus groups. Besides the wish for more participation in treatment decisions, they expressed further needs, such as more time and respect. The -experts of the 4 clinics (n=34, rate of response: 71%) also assessed these aspects of the patient-provider interaction as relevant. However, they saw the highest training need in the area of interdisciplinary team interactions., Conclusion: The interprofessional training programme "Fit for SDM" was developed on the basis of these results, and consists of 2 modules for the implementation of shared decision-making in medical rehabilitation. Module 1 focuses on external participation (provider-patient interaction), Module 2 on internal participation (team interaction). Module 2 was additionally used for preparing executives in their role as multipliers in the team. The training is currently being evaluated in a cluster-randomized multicentre study., (© Georg Thieme Verlag KG Stuttgart · New York.)

Purpose: Information about fetal and maternal outcome in pregnant women with mixed connective tissue disease (MCTD) is rare and in part contradictory. The purpose of this study is to review published literature about MCTD in pregnancy in the context of a rare case of peripartum bilateral uterine rupture in a patient with MCTD with favorable outcome for the severely asphyctic newborn after hypothermia., Method: The study included a selective literature review based on a PubMed search using the search terms MCTD, Sharp syndrome, uterine rupture and hypothermia, and a detailed report of our case with regard to the MCTD of the patient., Results: Rupture to the backside of the uterus during delivery, independent of prior cesarean section, was unpredictable and its cause remains unclear. The clinical outcome of the newborn was surprisingly favorable and there were no signs of neurodevelopmental sequelae in spite of the fact that the newborn was asphyctic and had a large excess of acids in the umbilical cord blood gas analysis. The favorable outcome is due to treatment with whole body hypothermia., Conclusions: Any type of prior surgery of the uterus puts the patient at risk during delivery. MCTD might be a risk factor during birth. These patients should be followed closely during pregnancy and should deliver at a center, which provides all options for immediate surgical and neonatological intervention.

Apoptosis resistance is crucially involved in cancer development and progression, represents the leading cause for failure of anticancer therapy and is caused, for example, by downregulation of proapoptotic intracellular signaling molecules such as caspase-8. We found that the cytotoxic drugs methotrexate (MTX) and 5-fluorouracil (5-FU) were both able to sensitize resistant tumor cells for induction of apoptosis by p53-mediated upregulation of caspase-8. Increase in caspase-8 messenger RNA and protein expression disabled tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced proliferation and restored sensitivity toward TRAIL-induced apoptosis which was inhibited by transfection of p53 decoy oligonucleotides, p53 shRNA and caspase-8 shRNA. Upregulation of caspase-8 and sensitization toward TRAIL-induced apoptosis was found both in a broad panel of tumor cell lines with downregulated caspase-8 and in TRAIL-resistant primary tumor cells of children with acute leukemia. Taken together, we have identified caspase-8 as an important p53 target gene regulated by cytotoxic drugs. These findings highlight a new drug-induced modulation of physiological apoptosis pathways, which may be involved in successful anticancer therapy using MTX and 5-FU in leukemia and solid tumors over decades.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) might represent a future cytotoxic drug to treat cancer as it induces apoptosis in tumor cells without toxicity in animal trials. We recently described that in contrast to apoptosis, TRAIL mediates tumor cell survival and proliferation in certain tumor cells. Here we studied the effect of TRAIL on 18 cell lines and 53 primary leukemia cells and classified these tumor cells into four groups: TRAIL, anti-DR4 or anti-DR5 induced apoptosis in group A cells, whereas they had no effect on group 0 cells and mediated proliferation in group P cells. To our surprise, TRAIL induced simultaneous apoptosis and proliferation in group AP cells. More than 20% of all cells tested belonged to group P and showed TRAIL-mediated proliferation even in the presence of certain cytotoxic drugs but not inhibitors of nuclear factor-kappaB. Transfection with B-cell leukemia/lymphoma protein 2 transformed group A cells into group 0 cells, whereas transfection with Fas-associated polypeptide with death domain (FADD)-like interleukin-1-converting enzyme-inhibitory protein (FLIP) transformed them into group AP cells. Loss of caspase-8 or transfection of dominant-negative FADD transformed group A cells into group P cells. Taken together, our data suggest that proliferation is a frequent effect of TRAIL on tumor cells, which is related to receptor-proximal apoptosis defects at the level of the death-inducing signaling complex and should be prevented during antitumor therapy with TRAIL.

Betulinic acid (BA), a natural component isolated from Birch trees, effectively induces apoptosis in neuroectodermal and epithelial tumor cells and exerts little toxicity in animal trials. Here, we show that BA-induced marked apoptosis in 65% of primary pediatric acute leukemia cells and all leukemia cell lines tested. When compared for in vitro efficiency with conventionally used cytotoxic drugs, BA was more potent than nine out of 10 standard therapeutics and especially efficient in tumor relapse. No crossresistances were found between BA and any cytotoxic drug. Intracellular apoptosis signaling in leukemia tumor cells paralleled the pathway found in neuroectodermal cells involving caspases, but not death receptors. In isolated mitochondria, BA induced release of both cytochrome c and Smac. Taken together, BA potently induces apoptosis in leukemia cells and should be further evaluated as a future drug to treat leukemia.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a potent inducer of apoptosis in cancer cells. Examining primary cells of children with untreated acute leukemia, TRAIL induced apoptosis in 50% of cells, but to our surprise attenuated spontaneous apoptosis in the remaining samples or, most importantly, even mediated proliferation. We therefore examined tumor cell lines of leukemic and nonleukemic origin with apoptosis resistance towards TRAIL because of absent Caspase-8 or dysfunctional FADD. In all cell lines tested, TRAIL treatment increased cell numbers in average to 163% within 4 days and accelerated doubling time from 24 to 19 h. TRAIL-mediated proliferation was completely abrogated by blockade of NF-kappaB activation using proteasome inhibitors or in RIP-negative, IKKgamma-negative cells or in cells overexpressing dominant-negative IkappaBalpha. Our data describe the biological significance of TRAIL-mediated activation of NF-kappaB in cancer cells resistant to TRAIL-mediated apoptosis: TRAIL leads to an increase in tumor cell count by a prosurvival and possibly mitogenic function. Given the promising therapeutic potential of TRAIL as a novel anticancer drug, TRAIL-mediated survival or proliferation of target cells may restrict its use to apoptosis-sensitive tumors.

Objective: To examine the effects of square wave, sinusoidal, and linear inspiratory pressure waveforms during pressure-controlled assist/control ventilation on the firing pattern of pulmonary stretch receptors and phrenic nerve activity., Design: Experimental, comparative study., Setting: Research laboratory at a university biomedical center., Subjects: Nine anesthetized, endotracheally intubated young cats (2.5-3.4 kg)., Intervention: With interposed periods of continuous positive airway pressure (0.2 kPa), each cat was exposed to periods of assist/control ventilation with three different pressure waveforms, where the peak inspiratory pressure (0.74 +/- 0.13 kPa), end-expiratory pressure (0.2 +/- 0.02 kPa), and tidal volume (14.9 +/- 5.22 mL/kg) were kept constant. Preset controlled ventilator rate was set below the rate of spontaneous breathing, and the mechanical inflation time equaled the inspiratory time during spontaneous breathing on continuous positive airway pressure., Measurements and Main Results: Respiratory rate and arterial blood gases did not change between the three pressure waveforms during assist/control ventilation. Peak pulmonary stretch receptor activity was lower and mean phrenic nerve activity higher during continuous positive airway pressure than during assist/control ventilation (p <.05). Peak inspiratory pulmonary stretch receptor activity was the same with all three pressure waveforms (82 +/- 17 impulses.sec-1) but occurred earlier with square wave than with sinusoidal or linear pressure waveforms (p <.05). The total number of impulses in the phrenic nerve activity burst was smaller with square wave than with the other two pressure waveforms (0.21 +/- 0.17 vs. 0.33 +/- 0.27 and 0.42 +/- 0.30 arbitrary units; p <.05), and the phrenic nerve activity burst duration was shorter with square wave (1.10 +/- 0.45 vs. 1.54 +/- 0.36 and 1.64 +/- 0.25 secs; p <.05)., Conclusion: Square wave pressure waveform during pressure-controlled assist/control ventilation strongly inhibits spontaneous inspiratory activity in cats. One mechanism for this inhibition is earlier and sustained peak pulmonary stretch receptor activity during inspiration. These findings show that differences in inspiratory pressure waveforms influence the spontaneous breathing effort during assist/control ventilation in cats.

In the period of 1958-1979 in the files of National Cancer Register (NCR) of GDR 1,313 patients with nasal cancer (ICD 160) were registered in seven districts of the GDR. In collaboration with NCR the occupational history of 1,160 patients with nasal cancer was recorded by a questionnaire. The results suggest an increased number of tumors in several professions. Compared to about 19 expected tumors in male woodworkers the number of the observed was 70. This is a statistical significant risk of 3.6.

68 mean values of airborne shift concentration of total dust were measured at 9 factories of woodworking industries. The values varied from 0.18 to 59.3 mg/m3 for area sampling. But at 63% of the woodworking machines with exhaustion the area sampling shift concentration of total dust reached fewer values than 5 mg/m3. This is a significant decrease of dust concentration in the last ten years.