Complex Patterns of Genomic Heterogeneity Identified in 42 Tumor Samples and ctDNA of a Pulmonary Atypical Carcinoid Patient.

Tumor evolution underlies many challenges facing precision oncology, and improving our understanding has the potential to improve clinical care. This study represents a rare opportunity to study tumor heterogeneity and evolution in a patient with an understudied cancer type. A patient with pulmonary atypical carcinoid, a neuroendocrine tumor, metastatic to 90 sites, requested and consented to donate tissues for research. 42 tumor samples collected at rapid autopsy from 14 anatomically distinct sites were analyzed through DNA whole-exome sequencing and RNA sequencing, and five analyzed through linked-read sequencing. Targeted DNA sequencing was completed on two clinical tissue biopsies and one blood plasma sample. Chromosomal alterations and gene variants accumulated over time, and specific chromosomal alterations preceded the single predicted gene driver variant (ARID1A). At the time of autopsy, all sites shared the gain of one copy of Chr 5, loss of one copy of Chr 6 and 21, chromothripsis of one copy of Chr 11, and 39 small variants. Two tumor clones (carrying additional variants) were detected at metastatic sites, and occasionally in different regions of the same organ (e.g., within the pancreas). Circulating tumor DNA (ctDNA) sequencing detected shared tumor variants in the blood plasma and captured marked genomic heterogeneity, including all metastatic clones but few private tumor variants. This study describes genomic tumor evolution and dissemination of a pulmonary atypical carcinoid donated by a single generous patient. It highlights the critical role of chromosomal alterations in tumor initiation and explores the potential of ctDNA analysis to represent genomically heterogeneous disease. Significance: DNA sequencing data from tumor samples and blood plasma from a single patient highlighted the critical early role of chromosomal alterations in atypical carcinoid tumor development. Common tumor variants were readily detected in the blood plasma, unlike emerging tumor variants, which has implications for using ctDNA to capture cancer evolution.

Chromosomal Aberrations Accumulate during Metastasis of Virus-Negative Merkel Cell Carcinoma.

Merkel cell carcinoma is a rare, aggressive skin tumor initiated by polyomavirus integration or UV light DNA damage. In New Zealand, there is a propensity toward the UV-driven form (31 of 107, 29% virus positive). Using archival formalin-fixed, paraffin-embedded tissues, we report targeted DNA sequencing covering 246 cancer genes on 71 tumor tissues and 38 nonmalignant tissues from 37 individuals, with 33 of 37 being negative for the virus. Somatic variants of New Zealand virus-negative Merkel cell carcinomas partially overlapped with those reported overseas, including TP53 variants in all tumors and RB1, LRP1B, NOTCH1, and EPHA3/7 variants each found in over half of the cohort. Variants in genes not analyzed or reported in previous studies were also found. Cataloging variants in TP53 and RB1 from published datasets revealed a broad distribution across these genes. Chr 1p gain and Chr 3p loss were identified in around 50% of New Zealand virus-negative Merkel cell carcinomas, and RB1 loss of heterozygosity was found in 90% of cases. Copy number variants accumulate in most metastases. Virus-negative Merkel cell carcinomas have complex combinations of somatic DNA-sequence variants and copy number variants. They likely carry the small genomic changes permissive for metastasis from early tumor development; however, chromosomal alterations may contribute to driving metastatic progression.

Interleukin 1 Receptor 1 Knockout and Maternal High Fat Diet Exposure Induces Sex-Specific Effects on Adipose Tissue Adipogenic and Inflammatory Gene Expression in Adult Mouse Offspring.

Background: The global incidence of obesity continues to rise, increasing the prevalence of metabolic diseases such as insulin resistance, dyslipidemia, and type 2 diabetes mellitus. Low-grade chronic inflammation, associated with the obese state, also contributes to the development of these metabolic comorbidities. Interleukin-1-receptor-1 (IL-1R1), a pro-inflammatory mediator, bridges the metabolic and inflammatory systems. In young male mice, deficiency of IL-1R1 (IL-1R1-/-) paired with a high-fat diet (HFD) offered beneficial metabolic effects, however in female mice, the same pairing led to metabolic dysfunction. Therefore, we examined the contribution of maternal HFD in combination with IL1R1-/- to metabolic health in adult offspring. Methods: Female C57BL/6 and IL-1R1-/- mice were randomly assigned to a control diet (10% kcal from fat) or HFD (45% kcal from fat) 10 days prior to mating and throughout gestation and lactation. Male and female offspring were housed in same-sex pairs post-weaning and maintained on control diets until 16 weeks old. At 15 weeks, an oral glucose tolerance test (OGTT) was performed to assess glucose tolerance. Histological analysis was carried out to assess adipocyte size and gene expression of adipogenic and inflammatory markers were examined. Results: IL-1R1-/- contributed to increased body weight in male and female adult offspring, irrespective of maternal diet. IL-1R1-/- and maternal HFD increased adipocyte size in the gonadal fat depot of female, but not male offspring. In female offspring, there was reduced expression of genes involved in adipogenesis and lipid metabolism in response to IL1R1-/- and maternal HFD. While there was an increase in inflammatory gene expression in response to maternal HFD, this appeared to be reversed in IL1R1-/- female offspring. In male offspring, there was no significant impact on adipogenic or lipid metabolism pathways. There was an increase in inflammatory gene expression in IL1R1-/- male offspring from HFD-fed mothers. Conclusion: This study suggests that IL-1R1 plays a complex and important role in the metabolic health of offspring, impacting adipogenesis, lipogenesis, and inflammation in a sex-specific manner.

Fixed dose rivaroxaban can be used in extremes of bodyweight: A population pharmacokinetic analysis.

BACKGROUND: Emerging safety and efficacy data for rivaroxaban suggest traditional therapy and rivaroxaban are comparable in the morbidly obese. However, real-world data that indicate pharmacokinetic (PK) parameters are comparable at the extremes of body size are lacking. The International Society of Thrombosis and Haemostasis Scientific and Standardisation Committee (ISTH SSC) suggests avoiding the use of direct oral anticoagulants (DOACs) in patients weighing >120 kg or with a body mass index >40 kg/m2 and gives no recommendation on the use of DOACs in those <50 kg. OBJECTIVES: To generate a population PK model to understand the influence of bodyweight on rivaroxaban exposure from clinical practice data. METHOD: Rivaroxaban plasma concentrations and patient characteristics were collated between 2013 and 2018 at King's College Hospital anticoagulation clinic. A population PK model was developed using a nonlinear mixed effects approach and then used to simulate rivaroxaban concentrations at the extremes of bodyweight. RESULTS: A robust population PK model derived from 913 patients weighing between 39 kg and 172 kg was developed. The model included data from n = 86 >120 kg, n = 74 BMI >40 kg/m2 , and n = 30 <50 kg. A one-compartment model with between-subject variability on clearance and a proportional error model best described the data. Creatinine clearance calculated by Cockcroft-Gault, with lean bodyweight as the weight descriptor in this equation, was the most significant covariate influencing rivaroxaban exposure. CONCLUSIONS: Our work demonstrates rivaroxaban can be used at extremes of bodyweight provided renal function is satisfactory. We recommend that the ISTH SSC revises the current guidance with respect to rivaroxaban at extremes of body size.

Th17 Cells in Helicobacter pylori Infection: a Dichotomy of Help and Harm.

Helicobacter pylori is a Gram-negative bacterium that infects the gastric epithelia of its human host. Everyone who is colonized with these pathogenic bacteria can develop gastric inflammation, termed gastritis. Additionally, a small proportion of colonized people develop more adverse outcomes, including gastric ulcer disease, gastric adenocarcinoma, or gastric mucosa-associated lymphoid tissue lymphoma. The development of these adverse outcomes is dependent on the establishment of a chronic inflammatory response. The development and control of this chronic inflammatory response are significantly impacted by CD4+ T helper cell activity. Noteworthy, T helper 17 (Th17) cells, a proinflammatory subset of CD4+ T cells, produce several proinflammatory cytokines that activate innate immune cell antimicrobial activity, drive a pathogenic immune response, regulate B cell responses, and participate in wound healing. Therefore, this review was written to take an intricate look at the involvement of Th17 cells and their affiliated cytokines (interleukin-17A [IL-17A], IL-17F, IL-21, IL-22, and IL-26) in regulating the immune response to H. pylori colonization and carcinogenesis.

Polysialic acid masks neural cell adhesion molecule antigenicity.

The neural cell adhesion molecule (NCAM) is a transmembrane protein involved in major cellular processes. The addition of polysialic acid (PSA), a post-translational modification (PTM) almost exclusively carried by NCAM, alters NCAM properties and functions and is therefore tightly regulated. Changes in NCAM and PSA-NCAM take place during development and ageing and occur in various diseases. The presence of PTMs can reduce the accessibility of antibodies to their epitopes and lead to false negative results. Thus, it is vital to identify antibodies that can specifically detect their target regardless of the presence of PTMs. In the present study, four commercially available NCAM antibodies were characterized by western blot and immunocytochemistry. Antibody specificity was determined by decreasing NCAM expression with small interfering RNA and subsequently determining whether the antibodies still produced a signal. In addition, PSA was digested with endoneuraminidase N to assess whether removing PSA improves NCAM detection with these antibodies. Our study revealed that the presence of PSA on NCAM reduced antibody accessibility to the epitope and consequently masked NCAM antigenicity for both techniques investigated. Moreover, three of the four antibodies tested were specific for the detection of NCAM by western blot and by immunocytochemistry. Altogether, this study demonstrates the importance of choosing the correct antibody to study NCAM depending on the technique of interest and underlines the importance of taking PTMs into account when using antibody-based techniques for the study of NCAM.

Characterization of the hepatosplenic and portal venous findings in patients with Proteus syndrome.

Proteus syndrome (PS) is a rare disorder caused by a mosaic AKT1 variant that comprises patchy overgrowth of tissues derived from all three germinal layers affecting multiple viscera. We sought to delineate the extent of hepatoportal manifestations in patients with PS. We identified patients with PS who had abdominal imaging from 1989 to 2015 in a natural history study. Imaging was characterized for evidence of focal findings in the liver, spleen, and portal vasculature and for organomegaly. Relevant clinical and laboratory data were compared among those with or without organomegaly. Abdominal imaging was available on 38 patients including 20 who had serial studies. Nine patients had focal hepatic lesions including vascular malformations (VMs). Focal splenic abnormalities were noted in seven patients. Patients without cutaneous VMs did not have visceral VMs. Nine patients had splenomegaly, 12 had portal vein dilation, and 4 had hepatomegaly. There was a weak correlation of portal vein dilation to spleen height ratio (r2 = 0.18, p < .05). On laboratory evaluation, hepatic function was normal but there was thrombocytopenia in those with splenomegaly; platelet counts were 179 ± 87K/µL compared to those with normal spleen size at 253 ± 57K/µL (p < .05). Overall, focal hepatosplenic abnormalities occurred in 11 of 38 (29%) patients with PS. Splenomegaly and portal venous dilation were both found in 8 of 38 (21%) patients; however, other than relative thrombocytopenia, there was no evidence of portal hypertension. Although the AKT1-E17K somatic variant is a suspected oncogene, there were no malignant lesions identified in this study.

Maternal High-Fat and High-Salt Diets Have Differential Programming Effects on Metabolism in Adult Male Rat Offspring.

Maternal high-fat or high-salt diets can independently program adverse cardiometabolic outcomes in offspring. However, there is a paucity of evidence examining their effects in combination on metabolic function in adult offspring. Female Sprague Dawley rats were randomly assigned to either: control (CD; 10% kcal from fat, 1% NaCl), high-salt (SD; 10% kcal from fat, 4% NaCl), high-fat (HF; 45% kcal from fat, 1% NaCl) or high-fat and salt (HFSD; 45% kcal from fat, 4% NaCl) diets 21 days prior to mating and throughout pregnancy and lactation. Male offspring were weaned onto a standard chow diet and were culled on postnatal day 130 for plasma and tissue collection. Adipocyte histology and adipose tissue, liver, and gut gene expression were examined in adult male offspring. HF offspring had significantly greater body weight, impaired insulin sensitivity and hyperleptinemia compared to CD offspring, but these increases were blunted in HFSD offspring. HF offspring had moderate adipocyte hypertrophy and increased expression of the pre-adipocyte marker Dlk1. There was a significant effect of maternal salt with increased hepatic expression of Dgat1 and Igfb2. Gut expression of inflammatory (Il1r1, Tnfα, Il6, and Il6r) and renin-angiotensin system (Agtr1a, Agtr1b) markers was significantly reduced in HFSD offspring compared to HF offspring. Therefore, salt mitigates some adverse offspring outcomes associated with a maternal HF diet, which may be mediated by altered adipose tissue morphology and gut inflammatory and renin-angiotensin regulation.

Pathogenetic insights from quantification of the cerebriform connective tissue nevus in Proteus syndrome.

BACKGROUND: The plantar cerebriform connective tissue nevus (CCTN) is the most common and problematic cutaneous manifestation of Proteus syndrome. OBJECTIVE: To gain insights into CCTN pathogenesis and natural history. METHODS: The size and location of plantar CCTN was measured on 152 images from 22 individuals with Proteus syndrome by 2 independent, blinded reviewers. Average measures of plantar CCTN were transformed into a linear mixed model to estimate proportionate change in size with age. RESULTS: Median patient age was 6.9 years at study onset. The intraclass correlation coefficient between 2 blinded reviewers was 0.946 for CCTN single measures. The CCTN relative area increased with age in children (n = 18, P < .0001) by 5.6% per year. Confluent papules and nodules extending beyond the boundaries of CCTNs were gradually replaced by typical CCTN over time. The location of CCTN in different individuals overlapped near the ball of the foot. A positive relationship between CCTN growth rate and AKT1 mutant allele frequency was observed (0.62, P = .10, n = 8). LIMITATIONS: This was a retrospective review using photographs. CONCLUSION: CCTN growth is affected by age and extent of the CCTN precursor lesion. Monitoring of CCTN size might prove useful for evaluating drug response in the treatment of Proteus syndrome.

Maternal plasma miRNAs as biomarkers during mid-pregnancy to predict later spontaneous preterm birth: a pilot study.

More than 10% of babies are born too early resulting in over 15 million preterm births and more than one million new-born deaths globally. Although women with a previous spontaneous preterm birth (SPTB) are considered at high risk for recurrence, the majority occur in women without prior history. Prediction of SPTB risk allows for improved care and potential for targeting novel and existing therapeutics to prevent SPTB, which may result in improved outcomes for infant and mother. In this pilot study, a miRNA array was used to analyse plasma from healthy women in their first pregnancy at 20 weeks of gestation who then went on to deliver either at term or experience SPTB at 28-32 weeks. We identified specific miRNA expression profiles that differentiated between those mothers who delivered at term or delivered following SPTB. miR302b, miR1253 and a clustering of miR548 miRNAs were underexpressed in SPTB cases compared to term controls. Conversely, miR223 was elevated in mothers that later experienced a SPTB. The circulating miRNAs identified in the present study may therefore be attractive candidates as non-invasive biomarkers for the early prediction of SPTB. Further larger studies are now warranted to investigate the potential clinical utility of these markers.

TLR9 activation suppresses inflammation in response to Helicobacter pylori infection.

Helicobacter pylori (H. pylori) induces chronic gastritis in humans, and infection can persist for decades. One H. pylori strain-specific constituent that augments disease risk is the cag pathogenicity island. The cag island encodes a type IV secretion system (T4SS) that translocates DNA into host cells. Toll-like receptor 9 (TLR9) is an innate immune receptor that detects hypo-methylated CpG DNA motifs. In this study, we sought to define the role of the H. pylori cag T4SS on TLR9-mediated responses in vivo. H. pylori strain PMSS1 or its cagE- mutant, which fails to assemble a T4SS, were used to infect wild-type or Tlr9-/- C57BL/6 mice. PMSS1-infected Tlr9-/- mice developed significantly higher levels of inflammation, despite similar levels of colonization density, compared with PMSS1-infected wild-type mice. These changes were cag dependent, as both mouse genotypes infected with the cagE- mutant only developed minimal inflammation. Tlr9-/- genotypes did not alter the microbial phenotypes of in vivo-adapted H. pylori strains; therefore, we examined host immunological responses. There were no differences in levels of TH1 or TH2 cytokines in infected mice when stratified by host genotype. However, gastric mucosal levels of IL-17 were significantly increased in infected Tlr9-/- mice compared with infected wild-type mice, and H. pylori infection of IL-17A-/- mice concordantly led to significantly decreased levels of gastritis. Thus loss of Tlr9 selectively augments the intensity of IL-17-driven immune responses to H. pylori in a cag T4SS-dependent manner. These results suggest that H. pylori utilizes the cag T4SS to manipulate the intensity of the host immune response.

New approaches for bacteriotherapy: prebiotics, new-generation probiotics, and synbiotics.

The gut microbiota has a significant role in human health and disease. Dysbiosis of the intestinal ecosystem contributes to the development of certain illnesses that can be reversed by favorable alterations by probiotics. The published literature was reviewed to identify scientific data showing a relationship between imbalance of gut bacteria and development of diseases that can be improved by biologic products. The medical conditions vary from infectious and antibiotic-associated diarrhea to obesity to chronic neurologic disorders. A number of controlled clinical trials have been performed to show important biologic effects in a number of these conditions through administration of prebiotics, probiotics, and synbiotics. Controlled clinical trials have identified a limited number of prebiotics, probiotic strains, and synbiotics that favorably prevent or improve the symptoms of various disorders including inflammatory bowel disease, irritable bowel syndrome, infectious and antibiotic-associated diarrhea, diabetes, nonalcoholic fatty liver disease, necrotizing enterocolitis in very low birth weight infants, and hepatic encephalopathy. Studies have shown that probiotics alter gut flora and lead to elaboration of flora metabolites that influence health through 1 of 3 general mechanisms: direct antimicrobial effects, enhancement of mucosal barrier integrity, and immune modulation. Restoring the balance of intestinal flora by introducing probiotics for disease prevention and treatment could be beneficial to human health. It is also clear that significant differences exist between different probiotic species. Metagenomics and metatranscriptomics together with bioinformatics have allowed us to study the cross-talk between the gut microbiota and the host, furthering insight into the next generation of biologic products.

The impact of orthodontic treatment on quality of life and self-esteem in adult patients.

OBJECTIVES: To assess the impact of fixed orthodontic treatment on oral health related quality of life (OHRQoL) and self-esteem in adults. SUBJECTS AND METHODS: A prospective study design was applied, within private practice. Sample size estimation revealed a minimum of 52 subjects, allowing for drop outs. All participants completed a set of validated questionnaires at baseline (T0), 1- (T1), 3- (T2), and 6-months (T3) and post-treatment (T4). These included the Rosenberg Self-esteem scale, the Oral Health Impact Profile (OHIP-14) and a socioeconomic status questionnaire. The Dental Health Component of the Index of Orthodontic Treatment Need (IOTN) was used to assess malocclusion severity. RESULTS: Sixty-one subjects were recruited, with only one subject lost to follow-up. A statistically significant difference in OHRQoL scores was seen between: T0 and T1 (P = 0.001); T0 and T2 (P = 0.020). There was no statistical difference between T0 and T3 (P = 0.078) or T4 (P = 0.565), where OHRQoL improved to pre-treatment scores. A significant difference in self-esteem scores was observed between baseline and end of treatment (P = 0.002). CONCLUSIONS: Undergoing fixed orthodontic therapy had a negative impact on the overall OHRQoL, during the first 3 months of treatment, which then improved to pre-treatment scores, whilst a significant increase was observed in self-esteem as a result of treatment.

Cost-effectiveness analysis of nebivolol and metoprolol in essential hypertension: a pharmacoeconomic comparison of antihypertensive efficacy of beta blockers.

OBJECTIVE: To estimate and compare the cost-effectiveness and safety of nebivolol with sustained-release metoprolol in reducing blood pressure by 1 mm of Hg per day in hypertensive patients. MATERIALS AND METHODS: This was a prospective, randomized, open label, observational analysis of cost-effectiveness, in a questionnaire-based fashion to compare the cost of nebivolol (2.5 mg, 5 mg, 10 mg) and sustained released metoprolol succinate (25 mg, 50 mg, 100 mg) in hypertensive patients using either of the two drugs. A total of 60 newly detected drug naïve hypertensive patients were considered for the comparison, of which 30 patients were prescribed nebivolol and the other 30 were prescribed metoprolol succinate as per the recommended dosage. Based on the data, statistical analysis was carried out using GraphPad Prism 5 and MS Excel Spreadsheet 2007. RESULT: The cost of reducing 1 mm of Hg blood pressure per day with nebivolol was 0.60, 0.70, and 1.06 INR, whereas that of metoprolol succinate was 0.93, 1.18, and 1.25 INR at their respective equivalent doses, hence significantly lower with the nebivolol group as compared to the metoprolol group (P < 0.05). CONCLUSION: This pharmacoeconomic analysis shows that nebivolol is more cost-effective as compared to metoprolol when the cost per reduction in blood pressure per day is considered. This may affect the patients economically during their long-term use of these molecules for the treatment of hypertension.

Let-7 miRNA profiles are associated with the reversal of left ventricular hypertrophy and hypertension in adult male offspring from mothers undernourished during pregnancy after preweaning growth hormone treatment.

Maternal undernutrition (UN) is known to cause cardiac hypertrophy, elevated blood pressure, and endothelial dysfunction in adult offspring. Maternal UN may also lead to disturbances in GH regulation in offspring. Because GH plays a key role in cardiac development, we used a model of maternal UN to examine the effects of neonatal GH treatment on cardiac hypertrophy, cardiac micro RNA (miRNA) profiles, and associated gene regulation in adult offspring. Female Sprague-Dawley rats were fed either a standard control diet (CON) or 50% of CON intake throughout pregnancy (UN). From neonatal day 3 until weaning (d 21), CON and UN pups received either saline (S) (CON-S, UN-S) or GH (2.5 µg/g·d) (CON-GH, UN-GH). Heart structure was determined by hematoxylin and eosin staining, and miRNA was isolated from cardiac tissue and miRNA expression analyzed using Cardiovascular miRNA gene Arrays (SABiosciences Ltd). Maternal UN caused marked increases in cardiac hypertrophy and left ventricular cardiomyocyte area, which were reversed by preweaning GH treatment. Systolic blood pressure was increased in UN-S groups and normalized in UN-GH groups (CON-S 121 ± 2 mmHg, CON-GH 115 ± 3 mm Hg, UN-S 146 ± 3 mmHg, and UN-GH 127 ± 2 mmHg). GH treatment during early development facilitated a reversal of pathological changes in offspring hearts caused by UN during pregnancy. Specific cardiac miRNA profiles were exhibited in response to maternal UN, accompanied by up-regulation of the lethal-7 (LET-7) miRNA family in GH-treated offspring. miRNA target analysis revealed a number of genes associated with inflammation and cardiovascular development, which may be involved in the altered cardiac function of these offspring. Up-regulation of the LET-7 family of miRNAs observed in GH groups may mediate the reversal of cardiac hypertrophy observed in adult offspring males of UN mothers.

Early life exposure to fructose and offspring phenotype: implications for long term metabolic homeostasis.

The consumption of artificially sweetened processed foods, particularly high in fructose or high fructose corn syrup, has increased significantly in the past few decades. As such, interest into the long term outcomes of consuming high levels of fructose has increased significantly, particularly when the exposure is early in life. Epidemiological and experimental evidence has linked fructose consumption to the metabolic syndrome and associated comorbidities-implicating fructose as a potential factor in the obesity epidemic. Yet, despite the widespread consumption of fructose-containing foods and beverages and the rising incidence of maternal obesity, little attention has been paid to the possible adverse effects of maternal fructose consumption on the developing fetus and long term effects on offspring. In this paper we review studies investigating the effects of fructose intake on metabolic outcomes in both mother and offspring using human and experimental studies.

End-of-life care decisions in the PICU: roles professionals play.

OBJECTIVE: Describe the roles and respective responsibilities of PICU healthcare professionals in end-of-life care decisions faced by PICU parents. DESIGN: Retrospective qualitative study. SETTING: University-based tertiary care children's hospital. PARTICIPANTS: Eighteen parents of children who died in the pediatric ICU and 48 PICU healthcare professionals (physicians, nurses, social workers, child-life specialists, chaplains, and case managers). INTERVENTIONS: In depth, semi-structured focus groups and one-on-one interviews designed to explore experiences in end-of-life care decision making. MEASUREMENTS AND MAIN RESULTS: We identified end-of-life care decisions that parents face based on descriptions by parents and healthcare professionals. Participants described medical and nonmedical decisions addressed toward the end of a child's life. From the descriptions, we identified seven roles healthcare professionals play in end-of-life care decisions. The family supporter addresses emotional, spiritual, environmental, relational, and informational family needs in a nondirective way. The family advocate helps families articulate their views and needs to healthcare professionals. The information giver provides parents with medical information, identifies decisions or describes available options, and clarifies parents' understanding. The general care coordinator helps facilitate interactions among healthcare professionals in the PICU, among healthcare professionals from different subspecialty teams, and between healthcare professionals and parents. The decision maker makes or directly influences the defined plan of action. The end-of-life care coordinator organizes and executes functions occurring directly before, during, and after dying/death. The point person develops a unique trusting relationship with parents. CONCLUSIONS: Our results describe a framework for healthcare professionals' roles in parental end-of-life care decision making in the pediatric ICU that includes directive, value-neutral, and organizational roles. More research is needed to validate these roles. Actively ensuring attention to these roles during the decision-making process could improve parents' experiences at the end of a child's life.

Toxicodendron pubescens retains its anti-arthritic efficacy at 1M, 10M and CM homeopathic dilutions.

BACKGROUND: Our previous studies of Toxicodendron pubescens (Rhus tox) in homeopathic dilutions have shown anti-inflammatory activity in line with the principle of similia. The present study aimed to evaluate its anti-inflammatory activity in 1M, 10M and CM dilutions in rats. METHOD: Arthritis was induced by subplantar injection of 0.1 ml of Complete Freund's Adjuvant (CFA) in the right hind paws of rats. The severity of inflammatory lesions was measured plethysmometrically on 21st day post CFA injection. The intensity of pain was measured using digital Von Frey apparatus. Other estimations included serum C-reactive protein (CRP), hematological parameters, body weight changes, arthritic pain score and radiological analysis of the arthritic paws. RESULT: The 1M, 10M and CM homeopathic dilutions of Rhus tox reduced primary and secondary arthritic lesions, improved body weight gain and protected rats against CFA-induced hematological and radiological perturbations. A significant reduction in the serum levels of CRP and an improvement in pain threshold of injected paws was observed in the groups treated with the Rhus tox dilutions. CONCLUSION: The anti-arthritic potential of Rhus tox is retained at 1M, 10M and CM dilutions.