Type 3 secretion system induced leukotriene B4 synthesis by leukocytes is actively inhibited by Yersinia pestis to evade early immune recognition.

Subverting the host immune response to inhibit inflammation is a key virulence strategy of Yersinia pestis. The inflammatory cascade is tightly controlled via the sequential action of lipid and protein mediators of inflammation. Because delayed inflammation is essential for Y. pestis to cause lethal infection, defining the Y. pestis mechanisms to manipulate the inflammatory cascade is necessary to understand this pathogen's virulence. While previous studies have established that Y. pestis actively inhibits the expression of host proteins that mediate inflammation, there is currently a gap in our understanding of the inflammatory lipid mediator response during plague. Here we used the murine model to define the kinetics of the synthesis of leukotriene B4 (LTB4), a pro-inflammatory lipid chemoattractant and immune cell activator, within the lungs during pneumonic plague. Furthermore, we demonstrated that exogenous administration of LTB4 prior to infection limited bacterial proliferation, suggesting that the absence of LTB4 synthesis during plague contributes to Y. pestis immune evasion. Using primary leukocytes from mice and humans further revealed that Y. pestis actively inhibits the synthesis of LTB4. Finally, using Y. pestis mutants in the Ysc type 3 secretion system (T3SS) and Yersinia outer protein (Yop) effectors, we demonstrate that leukocytes recognize the T3SS to initiate the rapid synthesis of LTB4. However, several Yop effectors secreted through the T3SS effectively inhibit this host response. Together, these data demonstrate that Y. pestis actively inhibits the synthesis of the inflammatory lipid LTB4 contributing to the delay in the inflammatory cascade required for rapid recruitment of leukocytes to sites of infection.

2021 AAPM Equity, Diversity, and Inclusion Climate Survey Executive Summary.

PURPOSE: The American Association of Physicists in Medicine (AAPM) shares the results, conclusions, and recommendations from the initial Equity, Diversity, and Inclusion Climate Survey conducted in 2021. METHODS AND MATERIALS: The climate survey targeted medical physicists who are full members of the AAPM and included demographic inquiries and questions intended to assess the working environmental climate in terms of a sense of belonging and inclusion, experiences of discrimination and harassment, and obstacles to participation within the AAPM. The survey invitation was sent to 5,500 members. Responses were collected from 1385 members (response rate of 25%) between January and February 2021. RESULTS: Overall, the medical physics workplace climate was positive. However, some demographic and professional subgroups reported lower levels of agreement with positive characteristics of their workplace climates. Compared with men, women ranked lower 7 of 8 categories that characterized the workplace climate. Other subgroups that also ranked the workplace climate descriptors lower included individuals not originally from the United States and Canada (3/8). Most respondents strongly agreed/agreed that the climate within the AAPM was welcoming. However, 17% of respondents reported personally experiencing or witnessing microaggressions within the AAPM. Overall, medical physicists reported low levels of agreement that opportunities within the AAPM were available to them, from 34% to 60% among 8 categories, including opportunities to volunteer, join committees, and compete for leadership positions within the AAPM. Several subgroups reported even lower levels of agreement that these opportunities are available. Asian and Asian American respondents (3/8) and physicists with origins in countries outside the United States and Canada (7/8) reported fewer opportunities to participate in the AAPM. Medical physicists reported their experiences of discrimination and sexual harassment in their workplaces and within the AAPM. For those who reported personal experiences of sexual harassment, only 24% (15/63) felt comfortable reporting when it occurred within their workplaces, and 35% (9/26) felt comfortable reporting when it occurred within the AAPM. CONCLUSIONS: The report concludes with several recommendations for action.

Exercise-induced specialized proresolving mediators stimulate AMPK phosphorylation to promote mitochondrial respiration in macrophages.

OBJECTIVE: Physical activity has been shown to reduce the risk of CVD mortality in large-cohort longitudinal studies; however, the mechanisms underpinning the beneficial effects of exercise remain incompletely understood. Emerging data suggest that the risk reducing effect of exercise extends beyond changes in traditional CVD risk factors alone and involves alterations in immunity and reductions in inflammatory mediator production. Our study aimed to determine whether exercise-enhanced production of proresolving lipid mediators contribute to alterations in macrophage intermediary metabolism, which may contribute to the anti-inflammatory effects of exercise. METHODS: Changes in lipid mediators and macrophage metabolism were assessed in C57Bl/6 mice following 4 weeks of voluntary exercise training. To investigate whether exercise-stimulated upregulation of specialized proresolving lipid mediators (SPMs) was sufficient to enhance mitochondrial respiration, both macrophages from control mice and human donors were incubated in vitro with SPMs and mitochondrial respiratory parameters were measured using extracellular flux analysis. Compound-C, an ATP-competitive inhibitor of AMPK kinase activity, was used to investigate the role of AMPK activity in SPM-induced mitochondrial metabolism. To assess the in vivo contribution of 5-lipoxygenase in AMPK activation and exercise-induced mitochondrial metabolism in macrophages, Alox5-/- mice were also subjected to exercise training. RESULTS: Four weeks of exercise training enhanced proresolving lipid mediator production, while also stimulating the catabolism of inflammatory lipid mediators (e.g., leukotrienes and prostaglandins). This shift in lipid mediator balance following exercise was associated with increased macrophage mitochondrial metabolism. We also find that treating human and murine macrophages in vitro with proresolving lipid mediators enhances mitochondrial respiratory parameters. The proresolving lipid mediators RvD1, RvE1, and MaR1, but not RvD2, stimulated mitochondrial respiration through an AMPK-dependent signaling mechanism. Additionally, in a subset of macrophages, exercise-induced mitochondrial activity in vivo was dependent upon 5-lipoxygenase activity. CONCLUSION: Collectively, these results suggest that exercise stimulates proresolving lipid mediator biosynthesis and mitochondrial metabolism in macrophages via AMPK, which might contribute to the anti-inflammatory and CVD risk reducing effect of exercise.

Risk perceptions of drinking bottled vs. tap water in a low-income community on the US-Mexico Border.

BACKGROUND: Previous studies have shown that low-income Latinos generally drink bottled water over tap water and might be at increased risks for cavities from unfluoridated bottled water. In order to better design interventions, it is important to understand the risk perceptions of this unique high-risk yet historically marginalized group. METHODS: We interviewed low-income Latino households (n = 90) from Nogales, Arizona who primarily drink bottled water and asked them to evaluate potential health risks of drinking tap water compared to 16 other voluntary activities. Unpaired t-tests were used to determine if statistically significant (α = 0.05) differences occurred in perceived risk by drinking-water source and differences among demographic groups in their level of (dis)agreement with statements regarding tap or bottled water safety. To assess significant differences (α = 0.05) in perceived risks and voluntariness to engage in a number of activities, including drinking local tap water and drinking water in different geographic regions, a one-way analysis of variance (ANOVA) followed by Scheffe's post-hoc test (a conservative post-hoc test) with adjustment for the number of pairwise comparisons was used. RESULTS: Participants viewed bottled water to be significantly safer to consume than tap water (p < 0.001). On a Likert scale from 1 (low risk) to 5 (high risk), "drinking tap water in Nogales, Arizona" received an average score of 4.7, which was significantly higher than the average perceived risk of drinking San Francisco, California tap water (µ = 3.4, p < 0.001), and as risky as drinking and driving (µ = 4.8, p = 1.00) and drinking Nogales, Sonora, Mexico tap water (µ = 4.8, p = 1.00). Ninety-eight percent of participants feared that drinking local tap water could result in illness, 79% did not drink their water because of fear of microbial and chemical contamination and 73% would drink their water if they knew it was safe regardless of taste. CONCLUSIONS: These results suggest that fear of illness from tap-water consumption is an important contributing factor to increased bottled water use. Future efforts should focus on the development of educational and outreach efforts to assess the safety and risks associated with tap-water consumption.

Measured and Modeled Comparisons of Chemical and Microbial Contaminants in Tap and Bottled Water in a US-Mexico Border Community.

Tap water quality concerns and advertisements often drive increased bottled water consumption, especially in communities with historical tap water quality problems (e.g., Nogales, Arizona). The study objective was to assess contamination of municipal tap and bottled water in Nogales, Arizona. Bottled (sealed, open/partially consumed bottles, and reusable containers for vended water) and tap water samples were collected from 30 homes and analyzed for chemical and microbial contaminants. Fisher exact tests and Wilcoxon rank sum tests were used to compare proportions of positive samples and contaminant concentrations between tap and bottled water samples. While none of the chemical contaminants were above MCLs, there were statistically significantly greater concentrations and proportions of positive samples for some contaminants, including arsenic, in tap vs. bottled water. E. coli concentrations were >0 CFU/100mL in some unsealed bottled water samples but not for sealed bottles. This study demonstrates that 1) the measured concentrations in tap and bottled water likely pose low risks, as they are below the MCLs, 2) more education in this community on hygiene maintenance of refillable or opened bottled water containers is needed, and 3) using tap water over bottled water is advantageous due to likely lower E. coli risk and lower cost.

Gamma spectrometry analysis of radioiodine in charcoal from high volume aerosol samples.

Following accidental releases, gamma spectrometry of impregnated charcoal filters is used to measure gaseous 131I contamination, but is subject to sampling inhomogeneity. In this study two germanium detectors are calibrated using a charcoal multi-gamma standard. Activities in samples spiked with a matrix of 131I aliquots are compared based on measurement, spike known activity, and monte-carlo simulation, and used to test a simple mixing method. Measurement efficiency, and removal of 11% inhomogeneity effect by mixing, was successfully reproduced in GESPECOR calculations.

Cohort Generation and Characterization of Patient-Specific Familial Hypercholesterolemia Induced Pluripotent Stem Cells.

Homozygous familial hypercholesterolemia (hoFH) is a rare disorder caused primarily by pathological mutations in the low-density lipoprotein receptor (LDLR), which disrupts LDL-cholesterol (LDL-C) metabolism homeostasis. hoFH patients are at extremely high risk for cardiovascular disease and are resistant to standard therapies. LDLR knockout animals and in vitro cell models overexpressing different mutations have proved useful, but may not fully recapitulate human LDLR mutation biology. We and others have generated induced pluripotent stem cells (iPSC) from hoFH patient's fibroblasts and T cells and demonstrated their ability to recapitulate hoFH biology. In this study, we present the generation and characterization of a cohort of seven hoFH-iPSC lines derived from peripheral blood mononuclear cells (PBMC) collected from four homozygous and three compound heterozygous patients. The hoFH-iPSC cohort demonstrated a wide range of LDLR expression and LDL-C internalization in response to rosuvastatin that correlated with the predicted pathogenicity of the mutation. We were able to confirm that hoFH-iPSC cohort were pluripotent by differentiation toward all three germ layers and specifically to hepatocyte-like cells (HLC), the cell with primary LDL-C metabolic regulatory control, by expression of hepatocyte markers. hoFH patient PBMC-derived iPSC recapitulate the LDLR dysfunction of their specific mutation. They were capable of differentiating to HLC and could be useful for early developmental studies, pharmacology/toxicology, and potentially autologous cell therapy.

Familial hypercholesterolemia class II low-density lipoprotein receptor response to statin treatment.

Low-density lipoprotein (LDL) receptor (LDLR) mutations are the primary cause of familial hypercholesterolemia (FH). Class II LDLR mutations result in a misfolded LDLR retained in the endoplasmic reticulum (ER). We have developed a model of FH class II and CRISPR-corrected induced pluripotent stem cells (iPSC) capable of replicating mutant and repaired LDLR functions. We show here that iPSC and derived hepatocyte-like cells (HLC) replicate misfolded LDLR accumulation and restoration of LDLR function in CRISPR-corrected cells. It was reported that model cells overexpressing class II LDLR mutants result in endoplasmic reticulum (ER) accumulation of immature LDLR and activation of the unfolded protein response (UPR). We show here that statins induce a similar accumulation of immature LDLR that is resolved with class II correction. We also demonstrate that, although capable of UPR induction with tunicamycin treatment, unlike overexpression models, statin-treated class II iPSC and derived HLC do not induce the common UPR markers Grp78 (also known as HSPA5) or spliced XBP1 [XBP1 (S)]. Because statins are reported to inhibit UPR, we utilized lipoprotein-deficient serum (LPDS) medium, but still did not detect UPR induction at the Grp78 and XBP1 (S) levels. Our study demonstrates the recapitulation of mutant and corrected class II LDLR function and suggests that overexpression models may not accurately predict statin-mediated class II protein biology.

Childhood adversity, mental health and suicide (CHASE): a methods protocol for a longitudinal case-control linked data study.

INTRODUCTION: Suicide is a tragic outcome with devastating consequences. In 2018, Scotland experienced a 15% increase in suicide from 680 to 784 deaths. This was marked among young people, with an increase of 53% in those aged 15-24, the highest since 2007. Early intervention in those most at risk is key, but identification of individuals at risk is complex, and efforts remain largely targeted towards universal suicide prevention strategies with little evidence of effectiveness.Recent evidence suggests childhood adversity is a predictor of subsequent poor social and health outcomes, including suicide. This protocol reports on methodology for harmonising lifespan hospital contacts for childhood adversity, mental health, and suicidal behaviour. This will inform where to 1) focus interventions, 2) prioritise trauma-informed approaches, and 3) adapt support avenues earlier in life for those most at risk. METHODS: This study will follow a case-control design. Scottish hospital data (physical health SMR01; mental health SMR04; maternity/birth record SMR02; mother's linked data SMR01, SMR04, death records) from 1981 to as recent as available will be extracted for people who died by suicide aged 10-34, and linked on Community Health Index unique identifier. A randomly selected control population matched on age and geography at death will be extracted in a 1:10 ratio. International Classification of Disease (ICD) codes will be harmonised between ICD9-CM, ICD9, ICD10-CM and ICD10 for childhood adversity, mental health, and suicidal behaviour. RESULTS: ICD codes for childhood adversity from four key studies are reported in two categories, 1) Maltreatment or violence-related codes, and 2) Codes suggestive of maltreatment. 'Clinical Classifications Software' ICD codes to operationalise mental health codes are also reported. Harmonised lifespan ICD categories were achieved semi-automatically, but required labour-intensive supplementary manual coding. Cross-mapped codes are reported. CONCLUSION: There is a dearth of evidence about touchpoints prior to suicide. This study reports methods and harmonised ICD codes along the lifespan to understand hospital contact patterns for childhood adversity, which come to the attention of hospital practitioners. KEY WORDS: Childhood Adversity, Adverse Childhood Experiences, Mental Health, Self-harm, Suicide, Suicidality, Violence, Hospital episodes, Routine Data, Data Linkage, Study Protocol.

WormBot, an open-source robotics platform for survival and behavior analysis in C. elegans.

Caenorhabditis elegans is a popular organism for aging research owing to its highly conserved molecular pathways, short lifespan, small size, and extensive genetic and reverse genetic resources. Here we describe the WormBot, an open-source robotic image capture platform capable of conducting 144 parallel C. elegans survival and behavioral phenotyping experiments. The WormBot uses standard 12-well tissue culture plates suitable for solid agar media and is built from commercially available robotics hardware. The WormBot is controlled by a web-based interface allowing control and monitoring of experiments from any internet connected device. The standard WormBot hardware features the ability to take both time-lapse bright field images and real-time video micrographs, allowing investigators to measure lifespan, as well as heathspan metrics as worms age. The open-source nature of the hardware and software will allow for users to extend the platform and implement new software and hardware features. This extensibility, coupled with the low cost and simplicity of the system, allows the automation of C. elegans survival analysis even in small laboratory settings with modest budgets.

Proactive multi-modality treatment of Pancreatic Neuroendocrine Tumours (PNETs): Potential survival benefits.

BACKGROUND/OBJECTIVES: Primary and metastatic pancreatic neuroendocrine tumours (PNET) can be treated with combination of surgery, locoregional and systemic therapy. Survival benefits from individual treatments have been well reported, however, the combined outcome from multimodal treatments are not well described in the literature. We report outcomes in a cohort of PNET patients treated with proactive, multimodality therapy. METHODS: 106 patients were identified from a single tertiary referral centre prospective database. Outcomes of treatment were studied, with the primary end point being death from any cause. RESULTS: Median follow-up was 71 months and overall 5-year survival of 62%. In patients with stage I-III disease (51 patients) estimated 5-year survival was 90%. Median survival in patients with stage IV disease was 51 months with an estimated 5-year survival of 40% in this group. A total of 80 patients (75%) had surgery of which 16% suffered complications requiring intervention. There was no perioperative mortality. CONCLUSIONS: This study demonstrates that proactive multimodal treatment is safe and may confer a survival benefit to patients in this cohort compared to historical data.

Assessment of neuronal autoantibodies in patients with small cell lung cancer treated with chemotherapy with or without ipilimumab.

Small-cell lung cancer (SCLC) is often associated with paraneoplastic syndromes. To assess the role of anti-neuronal autoantibodies (NAAs) as biomarkers of treatment outcome, we assessed NAAs in serial samples from SCLC patients treated with chemoimmunotherapy compared to chemotherapy alone. We evaluated 2 cohorts: in cohort 1 (C1), 47 patients received standard platinum/etoposide, and in cohort 2 (C2), 38 patients received ipilimumab, carboplatin and etoposide. Serum samples at baseline and subsequent time points were analyzed for the presence of NAAs. NAAs were detected at baseline in 25 patients (53.2%) in C1 and in 20 patients (52.6%) in C2 (most frequently anti-Sox1). NAA at baseline was associated with limited disease (75% vs 50%; p: 0.096) and better overall survival (15.1 m vs 11.7 m; p: 0.032) in C1. Thirteen patients (28.9%) showed 2 or more reactivities before treatment; this was associated with worse PFS (5.5 m vs 7.3 m; p: 0.005) in patients treated with chemoimmunotherapy. NAA titers decreased after therapy in 68.9% patients, with no differential patterns of change between cohorts. Patients whose NAA titer decreased after treatment, showed longer OS [18.5 m (95% CI: 15.8 - 21.2)] compared with those whose NAA increased [12.3 m (95% CI: 8.1 - 16.5; p 0.049)], suggesting that antibody levels correlate to tumor load. Our findings reinforce the role of NAAs as prognostic markers and tumor activity/burden in SCLC, warrant further investigation in their predictive role for immunotherapy and raise concern over the use of immunotherapy in patients with more than one anti-NAA reactivity.

The Adipose Stromal Vascular Fraction as a Complex Cellular Source for Tissue Engineering Applications.

A major challenge in tissue engineering is the generation of sufficient volumes of viable tissue for organ transplant. The development of a stable, mature vasculature is required to sustain the metabolic and functional activities of engineered tissues. Adipose stromal vascular fraction (SVF) cells are an easily accessible, heterogeneous cell system comprised of endothelial cells, macrophages, pericytes, and various stem cell populations. Collectively, SVF has been shown to spontaneously form vessel-like networks in vitro and robust, patent, and functional vasculatures in vivo. Capitalizing on this ability, we and others have demonstrated adipose SVF's utility in generating and augmenting engineered liver, cardiac, and vascular tissues, to name a few. This review highlights the scientific origins of SVF, the use of SVF as a clinically relevant vascular source, various SVF constituents and their roles, and practical considerations associated with isolating SVF for various tissue engineering applications.

Ribosomal stress and Tp53-mediated neuronal apoptosis in response to capsid protein of the Zika virus.

We report here that in rat and human neuroprogenitor cells as well as rat embryonic cortical neurons Zika virus (ZIKV) infection leads to ribosomal stress that is characterized by structural disruption of the nucleolus. The anti-nucleolar effects were most pronounced in postmitotic neurons. Moreover, in the latter system, nucleolar presence of ZIKV capsid protein (ZIKV-C) was associated with ribosomal stress and apoptosis. Deletion of 22 C-terminal residues of ZIKV-C prevented nucleolar localization, ribosomal stress and apoptosis. Consistent with a casual relationship between ZIKV-C-induced ribosomal stress and apoptosis, ZIKV-C-overexpressing neurons were protected by loss-of-function manipulations targeting the ribosomal stress effector Tp53 or knockdown of the ribosomal stress mediator RPL11. Finally, capsid protein of Dengue virus, but not West Nile virus, induced ribosomal stress and apoptosis. Thus, anti-nucleolar and pro-apoptotic effects of protein C are flavivirus-species specific. In the case of ZIKV, capsid protein-mediated ribosomal stress may contribute to neuronal death, neurodevelopmental disruption and microcephaly.

CRISPR Correction of a Homozygous Low-Density Lipoprotein Receptor Mutation in Familial Hypercholesterolemia Induced Pluripotent Stem Cells.

Familial hypercholesterolemia (FH) is a hereditary disease primarily due to mutations in the low-density lipoprotein receptor (LDLR) that lead to elevated cholesterol and premature development of cardiovascular disease. Homozygous FH patients (HoFH) with two dysfunctional LDLR alleles are not as successfully treated with standard hypercholesterol therapies, and more aggressive therapeutic approaches to control cholesterol levels must be considered. Liver transplant can resolve HoFH, and hepatocyte transplantation has shown promising results in animals and humans. However, demand for donated livers and high-quality hepatocytes overwhelm the supply. Human pluripotent stem cells can differentiate to hepatocyte-like cells (HLCs) with the potential for experimental and clinical use. To be of future clinical use as autologous cells, LDLR genetic mutations in derived FH-HLCs need to be corrected. Genome editing technology clustered-regularly-interspaced-short-palindromic-repeats/CRISPR-associated 9 (CRISPR/Cas9) can repair pathologic genetic mutations in human induced pluripotent stem cells. CONCLUSION: We used CRISPR/Cas9 genome editing to permanently correct a 3-base pair homozygous deletion in LDLR exon 4 of patient-derived HoFH induced pluripotent stem cells. The genetic correction restored LDLR-mediated endocytosis in FH-HLCs and demonstrates the proof-of-principle that CRISPR-mediated genetic modification can be successfully used to normalize HoFH cholesterol metabolism deficiency at the cellular level.

Comparison of Fluoride Levels in Tap and Bottled Water and Reported Use of Fluoride Supplementation in a United States-Mexico Border Community.

BACKGROUND: Compared to the general United States (U.S.) population, Arizona counties along the U.S.-Mexico border have a higher prevalence of dental caries, which can be reduced with adequate fluoride exposure. Because of concern regarding local tap water quality, fluoride-free bottled water consumption is common in this region, raising concern that families are not receiving adequate fluoride to promote dental health. OBJECTIVE: To evaluate the levels of fluoride in tap and bottled water as well as the use of fluoride supplements in an Arizona border community. METHODS: Low-income Latino households (n = 90) who report use of bottled water as their primary source of water intake were recruited. Participants completed a questionnaire about their and their children's dental histories and use of fluoride supplements. Water samples (bottled and tap) were collected from a subset of households (n = 30) for analysis of fluoride. RESULTS: Fluoride detection levels were significantly greater (p = 0.02, Fisher's exact test) in tap water (average = 0.49 mg/dL) than in bottled water, yet, the majority (22/30) were below the range for optimal dental health (0.7-1.2 mg/L). Concentration of fluoride in the majority (29/30) of bottled water samples was below the quantitative detection limit of 0.4 mg/L. Children were significantly less likely to have dental caries if they received fluoride varnishing treatments (p = 0.01, Fisher's exact test), lived in households that reported using fluoridated mouthwash (p < 0.001, Fisher's exact test), their parents received fluoride education (p = 0.01, Fisher's exact test), and their parents reported visiting a dentist yearly (p < 0.001, Fisher's exact test). Furthermore, none of the participants reported receiving recommendations from health-care providers about fluoride supplementation or variance in content by the type of water consumed. CONCLUSION: Although fluoride was significantly more likely to be detected in tap than bottled water, neither water source in this border community is likely to provide enough fluoride for optimal dental health. Low-income children in this region may benefit from regular access to fluoride varnishing treatments and/or use of fluoridated mouthwash, interventions that could be tested in future well-designed trials.

Dosimetric and radiobiologic comparison of 103Pd COMS plaque brachytherapy and Gamma Knife radiosurgery for choroidal melanoma.

PURPOSE: Plaque brachytherapy (BT) and Gamma Knife radiosurgery (GKRS) are highly conformal treatment options for choroidal melanoma. This study objectively compares physical dose and biologically effective dose (BED) distributions for these two modalities. METHODS AND MATERIALS: Tumor and organ-at-risk (OAR) dose distributions from a CT-defined reference right eye were compared between 103Pd COMS (Collaborative Ocular Melanoma Study Group) plaques delivering 70 Gy (plaque heterogeneity corrected) over 120 h to the tumor apex and GKRS plans delivering 22 Gy to the 40% isodose line for a representative sample of clinically relevant choroidal melanoma locations and sizes. Tumor and OAR biologically effective dose-volume histograms were generated using consensus radiobiologic parameters and modality-specific BED equations. RESULTS: Published institutional prescriptive practices generally lead to larger tumor and OAR physical doses from COMS BT vs. GKRS. Radiobiologic dose conversions, however, revealed variable BEDs. Medium and large tumors receive >1.3 times higher BEDs with COMS BT vs. GKRS. OAR BEDs have even greater dependence on tumor size, location, and treatment modality. For example, COMS BT maximum BEDs to the optic nerve are lower than from GKRS for large anterior and all posterior tumors but are higher for anterior small and medium tumors. CONCLUSIONS: BT and GKRS for choroidal melanoma have different physical dose and BED distributions with potentially unique clinical consequences. Using published institutional prescriptive practices, neither modality is uniformly favored, although COMS BT delivers higher physical doses and BEDs to tumors. These results suggest that lowering the physical prescription dose for COMS BT to more closely match the BED of GKRS might maintain equivalent tumor control with less potential morbidity.

Thymus transcriptome reveals novel pathways in response to avian pathogenic Escherichia coli infection.

Avian pathogenic Escherichia coli (APEC) can cause significant morbidity in chickens. The thymus provides the essential environment for T cell development; however, the thymus transcriptome has not been examined for gene expression in response to APEC infection. An improved understanding of the host genomic response to APEC infection could inform future breeding programs for disease resistance and APEC control. We therefore analyzed the transcriptome of the thymus of birds challenged with APEC, contrasting susceptible and resistant phenotypes. Thousands of genes were differentially expressed in birds of the 5-day post infection (dpi) challenged-susceptible group vs. 5 dpi non-challenged, in 5 dpi challenged-susceptible vs. 5 dpi challenged-resistant birds, as well as in 5 dpi vs. one dpi challenged-susceptible birds. The Toll-like receptor signaling pathway was the major innate immune response for birds to respond to APEC infection. Moreover, lysosome and cell adhesion molecules pathways were common mechanisms for chicken response to APEC infection. The T-cell receptor signaling pathway, cell cycle, and p53 signaling pathways were significantly activated in resistant birds to resist APEC infection. These results provide a comprehensive assessment of global gene networks and biological functionalities of differentially expressed genes in the thymus under APEC infection. These findings provide novel insights into key molecular genetic mechanisms that differentiate host resistance from susceptibility in this primary lymphoid tissue, the thymus.

Wnt5a Regulates the Assembly of Human Adipose Derived Stromal Vascular Fraction-Derived Microvasculatures.

Human adipose-derived stromal vascular fraction (hSVF) cells are an easily accessible, heterogeneous cell system that can spontaneously self-assemble into functional microvasculatures in vivo. However, the mechanisms underlying vascular self-assembly and maturation are poorly understood, therefore we utilized an in vitro model to identify potential in vivo regulatory mechanisms. We utilized passage one (P1) hSVF because of the rapid UEA1+ endothelium (EC) loss at even P2 culture. We exposed hSVF cells to a battery of angiogenesis inhibitors and found that the pan-Wnt inhibitor IWP2 produced the most significant hSVF-EC networking decrease (~25%). To determine which Wnt isoform(s) and receptor(s) may be involved, hSVF was screened by PCR for isoforms associated with angiogenesis, with only WNT5A and its receptor, FZD4, being expressed for all time points observed. Immunocytochemistry confirmed Wnt5a protein expression by hSVF. To see if Wnt5a alone could restore IWP2-induced EC network inhibition, recombinant human Wnt5a (0-150 ng/ml) was added to IWP2-treated cultures. The addition of rhWnt5a significantly increased EC network area and significantly decreased the ratio of total EC network length to EC network area compared to untreated controls. To determine if Wnt5a mediates in vivo microvascular self-assembly, 3D hSVF constructs containing an IgG isotype control, anti-Wnt5a neutralizing antibody or rhWnt5a were implanted subcutaneously for 2w in immune compromised mice. Compared to IgG controls, anti-Wnt5a treatment significantly reduced vessel length density by ~41%, while rhWnt5a significantly increased vessel length density by ~62%. However, anti-Wnt5a or rhWnt5a did not significantly affect the density of segments and nodes, both of which measure vascular complexity. Taken together, this data demonstrates that endogenous Wnt5a produced by hSVF plays a regulatory role in microvascular self-assembly in vivo. These findings also suggest that manipulating Wnt signaling could enhance control of hSVF vascularization in tissue engineering applications.

Brachytherapy vs. external beam radiotherapy for choroidal melanoma: Survival and patterns-of-care analyses.

PURPOSE: No modern randomized trials exist comparing external beam radiotherapy (EBRT) and plaque brachytherapy (BT) for choroidal melanoma, and the optimal treatment modality is currently unknown. This study compares the patterns of care and efficacy of EBRT vs. BT based on data in the Surveillance, Epidemiology, and End Results database. METHODS AND MATERIALS: The Surveillance, Epidemiology, and End Results database was queried for patients aged 20-79 diagnosed with choroidal melanoma from 2004 to 2011, treated with EBRT or BT; included patients were clinically T1-T4, N0, and M0. Overall survival and cause-specific survival curves were calculated by the Kaplan-Meier method. Univariate and multivariate analyses were performed in the survival and patterns-of-care analyses. RESULTS: A total of 1004 cases (380 EBRT and 624 BT) were included in the survival analysis. There was no difference in the 5-year overall survival (83.3% EBRT vs. 82.5% BT, p = 0.69) and 5-year cause-specific survival (88.3% EBRT vs. 88.3% BT, p = 0.92). In the survival analysis, older age and advanced tumor stage were predictors of increased risk of death. In the patterns-of-care analysis, later year of diagnosis and smaller tumor stage were predictors of BT use. CONCLUSIONS: Advanced tumor stage and older age seem to be independent predictors for risk of death from choroidal melanoma. The use of BT favors smaller tumors and later year of diagnosis. There is no difference in survival between those treated with EBRT or BT, and the utilization of BT is increasing.