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Objective: Evaluate popular explanation methods using heatmap visualizations to explain the predictions of deep neural networks for electrocardiogram (ECG) analysis and provide recommendations for selection of explanations methods., Materials and Methods: A residual deep neural network was trained on ECGs to predict intervals and amplitudes. Nine commonly used explanation methods (Saliency, Deconvolution, Guided backpropagation, Gradient SHAP, SmoothGrad, Input × gradient, DeepLIFT, Integrated gradients, GradCAM) were qualitatively evaluated by medical experts and objectively evaluated using a perturbation-based method., Results: No single explanation method consistently outperformed the other methods, but some methods were clearly inferior. We found considerable disagreement between the human expert evaluation and the objective evaluation by perturbation., Discussion: The best explanation method depended on the ECG measure. To ensure that future explanations of deep neural networks for medical data analyses are useful to medical experts, data scientists developing new explanation methods should collaborate tightly with domain experts. Because there is no explanation method that performs best in all use cases, several methods should be applied., Conclusion: Several explanation methods should be used to determine the most suitable approach., (© The Author(s) 2024. Published by Oxford University Press on behalf of the American Medical Informatics Association. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Gluteal augmentation with autologous fat transfer is one of the fastest growing aesthetic surgical procedures worldwide over the past decade. However, this procedure can be associated with high mortality from fatal pulmonary fat embolism events caused by intramuscular injection of fat. Ultrasound-guided fat grafting allows visualization of the transfer in the subcutaneous space, avoiding intramuscular injection., Objectives: The aim of this study was to assess the safety and efficacy of gluteal fat grafting performed with ultrasound-guided cannulation., Methods: A retrospective chart review of all patients undergoing ultrasound-guided gluteal fat grafting at the authors' center between 2019 and 2022 was performed. All cases were performed by board-certified and board-eligible plastic surgeons under general anesthesia in ASA Class I or II patients. Fat was only transferred to the subcutaneous plane when over the gluteal muscle. Patients underwent postoperative follow-up from a minimum of 3 months up to 2 years. Results were analyzed with standard statistical tests., Results: The study encompassed 1815 female patients with a median age of 34 years. Controlled medical comorbidities were present in 14%, with the most frequent being hypothyroidism (0.7%), polycystic ovarian syndrome (0.7%), anxiety (0.6%), and asthma (0.6%). Postoperative complications occurred in 4% of the total cohort, with the most common being seroma (1.2%), local skin ischemia (1.2%), and surgical site infection (0.8%). There were no macroscopic fat emboli complications or mortalities., Conclusions: These data suggest that direct visualization of anatomic plane injection through ultrasound guidance is associated with a low rate of complications. Ultrasound guidance is an efficacious adjunct to gluteal fat grafting and is associated with an improved safety profile that should be considered by every surgeon performing this procedure., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Aesthetic Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Secreted and membrane proteins represent a third of all cellular proteins and contain N-terminal signal peptides that are required for protein targeting to endoplasmic reticulum (ER). Mutations in signal peptides affect protein targeting, translocation, processing, and stability, and are associated with human diseases. However, only a few of them have been identified or characterized. In this report, we identified pathogenic signal peptide variants across the human genome using bioinformatic analyses and predicted the molecular mechanisms of their pathology. We recovered more than 65 thousand signal peptide mutations, over 11 thousand we classified as pathogenic, and proposed framework for distinction of their molecular mechanisms. The pathogenic mutations affect over 3.3 thousand genes coding for secreted and membrane proteins. Most pathogenic mutations alter the signal peptide hydrophobic core, a critical recognition region for the signal recognition particle, potentially activating the Regulation of Aberrant Protein Production (RAPP) quality control and specific mRNA degradation. The remaining pathogenic variants (about 25%) alter either the N-terminal region or signal peptidase processing site that can result in translocation deficiencies at the ER membrane or inhibit protein processing. This work provides a conceptual framework for the identification of mutations across the genome and their connection with human disease., (© The Author(s) 2023. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Objectives: Colloid oncotic pressure (COP) is an important factor in cardiac surgery, owing to its role in haemodilution. The effect of cardiopulmonary bypass prime fluids on the COP is unknown. In this study, the effect of crystalloid and colloid prime fluids, with or without retrograde autologous priming (RAP), on the COP during elective cardiac surgery was evaluated., Methods: Randomized controlled trials and prospective clinical trials comparing crystalloid and colloid priming fluids or with RAP were selected. The primary outcome was the COP; secondary outcomes were fluid balance, fluid requirements, weight gain, blood loss, platelet count and transfusion requirements., Results: From 1582 records, 29 eligible studies were identified. COPs were comparable between gelofusine and hydroxyethyl starch (HES) during bypass [mean difference (MD): 0.69; 95% confidence interval (CI): -2.05, 3.43; P = 0.621], after bypass (MD: -0.11; 95% CI: -2.54, 2.32; P = 0.930) and postoperative (MD: -0.61; 95% CI: -1.60, 0.38; P = 0.228). Fluid balance was lower with HES than with crystalloids. RAP reduced transfusion requirements compared with crystalloids. Blood loss was comparable between groups., Conclusions: COPs did not differ between crystalloids and colloids. As a result of increased transcapillary fluid movement, fluid balance was lower with HES than with crystalloids. Haematocrit and transfusion requirements were comparable between groups. However, the latter was lower when RAP was applied to crystalloid priming compared with crystalloids alone. Finally, no differences in blood loss were observed between the groups., (© The Author(s) 2022. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery.)

Buttock augmentation is a commonly performed aesthetic surgery. Several methods have been described, but only the use of implants or autologous fat is consensually deemed safe and effective. Synthetic fillers in gluteal augmentation have been described despite potential severe long-term complications, both medical and aesthetic. The aim of this study is to report a series of 2 consecutive cases who underwent buttock and hip augmentation with large volumes of permanent copolyamide filler requiring surgical removal due to significant complications. Based on these cases and a review of recent literature, a management algorithm is proposed. The authors conducted a retrospective chart review of 2 consecutive cases of failed copolyamide filler augmentations in the gluteal and inguinal regions. The authors conducted a literature overview using PubMed (National Institutes of Health, Bethesda, MD) and Google Scholar (Google, Mountain View, CA) to include all articles concerning the removal of large quantities of permanent copolyamide fillers. Based on the physical properties of copolyamide, resection of the filler was performed by percutaneous aspiration with liposuction cannulas, with varying infiltration protocols. Both cases showed successful removal of major parts of the filler; however, residual material tended toward migration, requiring a secondary intervention. Hydrated low-pressure aspiration can manage nonintegrated gluteal copolyamide filler but will achieve only partial resection. Literature shows that radical excision is possible, however, with major drawbacks in function and aesthetics. Moreover, in acute inflammation and infection, an open approach should be preferred., (© 2022 The Aesthetic Society.)

During the first wave of the COVID-19 pandemic, global measures preventing the spread of the new coronavirus required most of the population to lockdown at home. This sudden halt to collective life meant that non-essential services were closed and many health promoting activities (i.e. physical activity, school) were stopped in their tracks. To curb the negative health impacts of lockdown measures, activities adapting to this new reality were urgently developed. One form of activity promoted indoor physical activity to prevent the adverse physical and psychological effects of the lockdown. Another form of activity included the rapid development of online learning tools to keep children and youth engaged academically while not attending school. While these health promoting efforts were meant to benefit the general population, we argue that these interventions may have unintended consequences and inadvertently increase health inequalities affecting marginalized youth in particular, as they may not reap the same benefits, both social and physical, from the interventions promoting at-home physical activities or distance learning measures. We elaborate on several interventions and their possible unintended consequences for marginalized youth and suggest several strategies that may mitigate their impact., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Context: In primary aldosteronism, cosecretion of cortisol may alter cortisol-derived adrenal venous sampling indices., Objective: To identify whether cortisol cosecretion in primary aldosteronism alters adrenal venous sampling parameters and interpretation., Design: Retrospective case-control study., Setting: A tertiary referral center., Patients: 144 adult patients with primary aldosteronism who had undergone both adrenocorticotropic hormone-stimulated adrenal venous sampling and dexamethasone suppression testing between 2004 and 2018., Main Outcome Measures: Adrenal venous sampling indices including adrenal vein aldosterone/cortisol ratios and the selectivity, lateralization, and contralateral suppression indices., Results: 21 (14.6%) patients had evidence of cortisol cosecretion (defined as a failure to suppress cortisol to ≤50 nmol/L post dexamethasone). Patients with evidence of cortisol cosecretion had a higher inferior vena cava cortisol concentration (P = .01) than those without. No difference was observed between the groups in terms of selectivity index, lateralization index, lateralization of aldosterone excess, or adrenal vein cannulation rate., Conclusions: Cortisol cosecretion alters some parameters in adrenocorticotrophic hormone-stimulated adrenal venous sampling but does not result in alterations in patient management., (© Endocrine Society 2020. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objectives: Superficial pleomorphic liposarcoma (PL) has a favorable prognosis compared to deeply seated PL. Given developments in the classification of lipomatous neoplasms, we reappraised a series of cases., Methods: Retrospective clinicopathologic evaluation and genome-wide single-nucleotide polymorphism (SNP) microarray studies were performed for cases previously designated superficial PL., Results: Four cases were identified (age, 48-70 years). Two were dermally confined, whereas two were superficial subcutaneous; no recurrences or metastases were reported. Tumors demonstrated pleomorphic spindled morphology with variable cellularity. Multivacuolated atypical lipoblasts were focal in 3 and abundant in 1. Dermal tumors demonstrated atypical cells within sclerotic collagen. Genome-wide SNP microarray studies revealed consistent gains and losses, including losses at the 13q14.2 locus encompassing RB1 and DLEU2 and deletion/disruption of the TP53 locus. Although subcutaneous examples showed genomic changes similar to deep PL, the dermal examples showed fewer genetic alterations, including changes reported in the spectrum of atypical spindle cell/pleomorphic lipomatous tumors (ASPLT). All lacked MDM2 amplification., Conclusions: Careful integration of histologic and genetic features may improve classification of lipomatous neoplasms with atypia, allowing reclassification of some superficial PL as ASPLT., (© American Society for Clinical Pathology, 2020. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

To date, only few data exist on mechanisms underlying the human papillomavirus (HPV)-associated irradiation response. It has been suggested, that the viral E2 gene plays an important role in that context. The aim of the current study is to compare the effect of photon- and carbon-ion (12C)-radiation therapy (RT) on cells with different HPV and E2 gene status. We hypothesized that 12C-RT might overcome the radioresistance of E2 gene-disrupted cells. We analyzed four different cell lines that differed in HPV status or E2 gene status. Cells were irradiated with either photons or 12C. Clonogenic survival, cell cycle and expression of Rb and p53 were analyzed. Radiosensitivity seemed to be dependent on E2 gene status and type of RT. 12C-RT led to lower surviving fractions, indicating higher radiosensitivity even in cells with disrupted E2 gene. The observed relative biological effectiveness (RBE) of 12C-RT for C33a/Caski and W12/S12 was 1.3/4 and 2.7/2.5, respectively. Cell cycle regulation after both photon- and 12C-RT was dependent on HPV status and on E2 gene status. Furthermore, the effect of RT on expression of p53 and Rb seemed to be dependent on E2 gene status and type of RT. We showed that 12C-RT overcomes HPV-integration induced radioresistance. The effect of RT on cell cycle regulation as well as on expression of p53 and Rb seemed to be dependent on HPV status, E2 gene status and type of RT. Differences in Rb expression and cell cycle regulation may play a role for enhanced radiosensitivity to 12C-RT of cells with disrupted E2 gene., (© The Author(s) 2019. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.)

Little has been published regarding intravenous (IV) ketamine for burn wound care in adult patients. Ketamine may serve as a safe alternative to provide conscious sedation and limit opioid administration to patients. The purpose of this study was to characterize IV ketamine use during burn wound care and establish its potential role as a safe adjunct to opioid and benzodiazepine medications. This is a retrospective review of adult patients admitted to a regional burn center who received IV ketamine for burn wound care. Patient demographics, medications, and ketamine-related adverse effects including hypertension and dysphoric reactions were recorded. Cardiopulmonary complications were also tracked. Thirty-six patients met inclusion criteria; fifty total cases were performed. The median patient age was 37 (interquartile range [IQR]: 28-55] years with a median burn size of 9.5 (IQR: 4.0-52) %TBSA. The median ketamine dose administered was 1.2 (IQR: 0.8-2.1) mg/kg. IV midazolam was administered in almost all cases (98%) at a median dose of 3.0 (IQR: 2.0-5.0) mg. Opioids were administered in 13 of 50 cases (26%) at a median morphine equivalent dose of 10 (IQR: 5.0-18) mg. In 46 cases (92%), patients denied unpleasant recall of medication. Dysphoric reactions were observed in three cases (6%). Ketamine-induced hypertension occurred in three cases (6%) and all immediately responded to IV labetalol. There were no cardiopulmonary complications. These findings suggest that IV ketamine provides a safe analgesia and sedative option for burn wound care. Given these findings, IV ketamine for burn wound care warrants further study., (© American Burn Association 2018. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

With the advent of social media platforms such as Facebook and YouTube, online dissemination of exhibitionist videos has gained popularity. One recent disturbing trend is the "fire challenge" wherein a participant douses his or herself in a household accelerant such as isopropyl alcohol or acetone, sets him or herself ablaze, and attempts to extinguish the flames before serious burns are incurred. As expected, participants in the "fire challenge" often accidentally suffer serious burns. A 17-year-old white male was recently treated at our burn center after participating in the "fire challenge." He suffered 15% TBSA full and partial thickness burns requiring split thickness skin grafting to his abdomen. He reported lighting himself on fire because he had seen this stunt performed on the internet. A search for "fire challenge" and similar terms was conducted on YouTube (www.youtube.com). Gender and ethnicity of each participant were documented. Burn size, burn depth, and age of video participant were estimated by two attending burn surgeons evaluating YouTube videos. Results were reported with descriptive statistics. The search yielded thousands of hits, mostly home videos, compilations of stunts, and commentaries. After omitting duplicate and irrelevant videos, 50 videos were selected for the study. Of these, 13 videos included postburn footage demonstrating burn wounds of various location, size, and severity. Of these burns, the median TBSA burned was 4 ± 2.7% with a maximum size of 10%. Superficial and partial thickness burns were sustained on the torso (10/13, 77%), face (4/13, 31%), and extremities (2/13, 15%). Full thickness burns were seen in 2/13 videos. Some burn wounds were obscured by dressings. Of the 50 videos reviewed, 45/50 participants (90%) were male and 32/50 (64%) were African American with 29/50 participants (58%) estimated to be under age 20. The "fire challenge" is a popular social media phenomenon, but it can result in severe injury as seen with the patient at our institution. The lure of a challenge and potential for a shocking video to "go viral" might entice people to mimic this risky behavior. This study shows a disturbing trend, but undoubtedly only reflects a small portion of actual participants. A disproportionate number of videos featured young African American males, making this a target population for education and prevention efforts. Our patient's TBSA exceeded the maximum found on YouTube, suggesting that less severe burns may be posted online while larger burns are not, diminishing perceived risk and encouraging this behavior.

Concerns over dwindling play opportunities for children have recently become a preoccupation for health promotion in western industrialized countries. The emerging discussions of play seem to be shaped by the urgency to address the children's obesity epidemic and by societal concerns around risk. Accordingly, the promotion of play from within the field appears to have adopted the following principles: (i) particular forms of play are critical for increasing children's levels of physical activity; and (ii) play should be limited to activities that are not risky. In this article, we argue that these emerging principles may begin to re-shape children's play: play is predominantly instrumentalized as a means to promote children's physical health, which may result in a reduction of possibilities for children to play freely and a restriction of the kinds of play designated as appropriate for physical health. We argue that within this context some of the social and emotional elements of health and well-being that children gain through diverse forms of playing are neglected. This has implications for health promotion because it suggests a narrowing of the conception of health that was originally advocated for within the field. Additionally, this reveals a curious paradox; despite the urgency to promote physical activity through play, this position may limit the range of opportunities for children to freely engage in play, in effect reducing their activity levels. We propose an example that promotes play for children and better aligns with the conception of health as outlined in the Ottawa Charter.

Fentanyl lacks the antiinflammatory properties of morphine. Morphine attenuates the inflammatory response through differential stimulation of μ-receptor subtypes. Patients who receive morphine during coronary artery bypass graft have been shown to experience less postoperative fever than those who receive fentanyl. Patients who receive continuous fentanyl infusions in increased room temperatures after thermal injury may be at increased risk to experience higher body temperature than those who receive morphine. The records of 28 patients with >20%TBSA burn in 30 intensive care unit rooms (13 received fentanyl and 15 received morphine or hydromorphone) and 12 trauma patients who received fentanyl in 22°C intensive care unit rooms were reviewed. Mean maximum core temperature and percentage of temperature recordings > 39°C in the first 48 hours of admission were compared between burn patients who received fentanyl, those who did not, and with trauma patients. Burn patients exposed to fentanyl experienced significantly higher temperatures (40.1 ± 0.9°C) compared with those given morphine (38.7 ± 0.8°C) and compared with trauma patients (37.5 ± 2.4°C), P < .01 and P < .001, respectively. Burn patients on fentanyl had temperatures > 39°C for a higher percentage of time (33 ± 27%) than those without fentanyl (7.2 ± 13%) and trauma patients (1 ± 2.8%), P < .01 and P < .001, respectively. No differences in other medications administered, acute physiology and chronic health evaluation II scores, or the number of infections that could account for temperature disparities between groups existed. Burn patients who receive fentanyl in 30°C rooms experience higher body temperatures and are febrile for a higher percentage of time than those receiving morphine only. Morphine has well-established antiinflammatory properties and likely attenuates the postburn inflammatory response more than fentanyl, resulting in lower body temperatures. This phenomenon needs to be further investigated in additional studies.

Dermal templates are well established in the treatment of burn wounds and acute nonburn wounds. However, the literature regarding their use for reconstruction of chronic, nonhealing wounds is limited. This study describes a series of patients with chronic wounds reconstructed with a commercially available bilayer, acellular dermal replacement (ADR) containing a collagen-glycosaminoglycan dermal template and a silicone outer layer. A retrospective review was performed of 10 patients treated for chronic wounds with ADR and negative pressure dressing followed by split-thickness skin graft between July 2006 and January 2009. Data collected included age, gender, comorbidities, medications, wound type or location, wound size, the number of applications of ADR, the amount of ADR applied (in square centimeter), the amount of time between ADR placement and grafting, complications, need for reoperation, and percentage of graft take after 5 and 14 days. The mean age of study subjects was 44 years. All patients in the study had comorbidities that interfere with wound healing and were treated for lower extremity wounds (four to legs, five to ankles, and one to foot). The wounds had a variety of causative factors including venostasis ulcers (6, 60%), trauma in diabetic patients (2, 20%), brown recluse bite (1, 10%), and a wound caused from purpura fulminans (1, 10%). The average wound size and amount of ADR applied was 162±182 cm². Each patient required only one application of ADR. The average time between ADR placement and skin grafting was 36.5 days. The mean percentage of graft take at 5 days was 89.55%, 14 days was 90%, and 21 days was 87.3%. Only two patients required regrafting, and one of these grafts was lost because of patient noncompliance. ADR can be used successfully in the treatment of chronic wounds. ADR provides direct wound coverage and can conform to a variety of anatomical sites. This study demonstrates that the use of ADR in treating chronic wounds results in high rates of skin graft take. Favorable results were obtained despite the majority of patients having comorbidities that would normally interfere with wound healing.

The nation is faced with a shortage of subspecialty physicians, including burn surgeons. Exposure to a specialty in medical school has been shown to influence students' career choices. The authors postulate that exposure to burn surgery increases their interest in the field. Students from a medical school with an American Burn Association-verified burn center and from a school without a burn center were anonymously surveyed and asked to report their interest and knowledge regarding burn surgery using a 5-point Likert scale. They were asked about their current year in school, gender, overall interest in surgery, and any prior exposure to burn surgery (eg, preceptorship or rotation). Students were asked whether exposure to burn surgery or to a strong mentor might increase their interest in the field. Finally, they were asked to pick the most important factor in a list of deterrents to pursuing a career in burn surgery. Predictors of interest in burn surgery were determined with regression analyses. A total of 380 of 662 students responded to the survey (57.4%). Significant predictors of interest in burn surgery were an interest in surgery (P < .001, odds ratio [OR] = 56.3), prior exposure to burn surgery (P = .02, OR = 5.7), and year in school (P = .006, OR = 1.7). First- and second-year students were more likely to report interest in burn surgery (P < .001). Gender and medical school attended were not significant predictors. Prior exposure to burn surgery became a stronger predictor in subgroup analysis of the fourth-year students (P < .001, OR = 24.5). The majority of students reported that exposure to burn surgery (76%) and a strong mentor (87%) would make them more likely to consider burn surgery as a career. "Not interested in surgery" was the most important deterrent to pursuing a career in burn surgery, which was selected by 33% of students. However, 25% of students chose "I don't know anything about burn surgery" as the most important deterrent. Factors specific to burn surgery were less frequently selected (eg, wound care and hot operating room). The majority of students reported ignorance of the field of burns: 64% disagreed that they understood what a burn surgeon does on a daily basis, and 74% agreed they did not know enough about the field to consider it as a career. Exposure to burn surgery in the form of a clinical rotation fosters medical student interest. However, the majority of medical students lack knowledge about the field of burn surgery. Many consider this a deterrent to selecting it as a career. More exposure during medical school and strong mentorship may influence more students to become burn surgeons. Mentorship and recruiting efforts should be focused on students with a general interest in surgery.

Resuscitation of burn victims with high-dose ascorbic acid (vitamin C [VC]) was reported in Japan in the year 2000. Benefits of VC include reduction in fluid requirements, resulting in less tissue edema and body weight gain. In turn, these patients suffer less respiratory impairment and reduced requirement for mechanical ventilation. Despite these results, few burn centers resuscitate patients with VC in fear that it may increase the risk of renal failure. A retrospective review of 40 patients with greater than 20% TBSA between 2007 and 2009 was performed. Patients were divided into two groups: one received only lactated Ringer's (LR) solution and another received LR solution plus 66 mg/kg/hr VC. Both groups were resuscitated with the Parkland formula to maintain stable hemodynamics and adequate urine output (>0.5 ml/kg/hr). Patients with >10-hour delay in transfer to the burn center were excluded. Data collected included age, gender, weight, %TBSA, fluid administered in the first 24 hours, urine output in the first 24 hours, and Acute Physiology and Chronic Health Evaluation II score. PaO2 in millimeters mercury:%FIO2 ratio and positive end-expiratory pressure were measured at 12-hour intervals, and hematocrit was measured at 6-hour intervals. Comorbidities, mortality, pneumonia, fasciotomies, and renal failure were also noted. After 7 patients were excluded, 17 patients were included in the VC group and 16 in the LR group. VC and LR were matched for age (42 ± 16 years vs 50 ± 20 years, P = .2), burn size (45 ± 21%TBSA vs 39 ± 15%TBSA, P = .45), Acute Physiology and Chronic Health Evaluation II (17 ± 7 vs 18 ± 8, P = .8), and gender. Fluid requirements in the first 24 hours were 5.3 ± 1 ml/kg/%TBSA for VC and 7.1 ± 1 ml/kg/%TBSA for LR (P < .05). Urine output was 1.5 ± 0.4 ml/kg/hr for VC and 1 ± 0.5 ml/kg/hr for LR (P < .05). Vasopressors were needed in four VC patients and nine LR patients (P = .07). VC patients required vasopressors to maintain mean arterial pressure for a mean of 6 hours, but LR needed vasopressors for 11 hours (P = .2). No significant differences in PaO2 in millimeters mercury:%FIO2 ratio, positive end-expiratory pressure, frequency of pneumonia, renal failure, or inhalation injury were found. VC group had four mortalities, and LR group had three mortalities (P = 1). VC is associated with a decrease in fluid requirements and an increase in urine output during resuscitation after thermal injury. Although this study did not find a difference in outcomes with VC administration, it demonstrates that VC can be safely used without an increased risk of renal failure. The effects of VC should be further studied in a large-scale, prospective, randomized trial.

Ecosystem simulation models are designed to assess the flux of energy, water, carbon and nitrogen according to a given vegetation type. The reliability of the modeled results is determined by model validations. Model validations are typically done using classical statistical methods like regression analysis of predicted versus observed values, paired t statistics and error assessment procedures to characterize the quality of current and future model predictions. All these validation efforts concentrate on static aspects of the model but fail to describe the model dynamics. In this paper, we introduce methods from ergodic theory to analyze the dynamic behavior of ecosystem models. We describe (1) how the attractor representation of model behavior can be reconstructed from a time series of model outputs, and (2) what we can learn from the attractor to assess the model dynamics. As an application example, we provide simulation results for two important pine forest ecosystems in Austria, i.e., 23 Cembran pine and 16 Scots pine stands. These stands were simulated with three model parameterizations: one representing a generic, evergreen needle-leaf forest, and two species- specific parameter sets, one for Cembran pine and one for Scots pine. First, we applied standard validation methods to get static measures of model accuracy and precision. Next, we used ergodic theory to assess model dynamics. A comparison of both analyses reveals important issues related to model dynamics, such as the finding that the occurrence of instabilities in model behavior may not be detected by standard validation methods. Using ergodic theory, we were able to reconstruct the attractors of model behavior. In the attractor describing model dynamics for Cembran pine, simulated with the generic, evergreen needle-leaf forest parameter set, we detected instability in model behavior. We identified this instability as a riddled basin configuration, which is a strong indicator for the occurrence of a chaotic model behavior that may result in random predictions. Our results suggest that ergodic theory is a useful tool for assessing inconsistencies in the dynamics of ecosystem model simulations that have not been detected by standard statistical validation methods.

Pluripotent stem cell lines can be induced to differentiate into a range of somatic cell types in response to various stimuli. Such cell-based systems provide powerful tools for the investigation of molecules that modulate cellular development. For instance, the formation of the nervous system is a highly regulated process, controlled by molecular pathways that determine the expression of specific proteins involved in cell differentiation. To begin to decipher this mechanism in humans, we used oligonucleotide microarrays to profile the complex patterns of gene expression during the differentiation of neurons from pluripotent human stem cells. Samples of mRNA were isolated from cultured NTERA2 human embryonal carcinoma stem cells and their retinoic-acid-induced derivatives and were prepared for hybridization on custom microarrays designed to detect the expression of genes primarily associated with the neural lineage. In response to retinoic acid, human NTERA2 cells coordinately regulate the expression of large numbers of neural transcripts simultaneously. Transcriptional profiles of many individual genes aligned closely with expression patterns previously recorded by developing neural cells in vitro and in vivo, demonstrating that cultured human pluripotent stem cells appear to form neurons in a conserved manner. These experiments have produced many new expression data concerning neuronal differentiation from human stem cells in vitro. Of particular interest was the regulated expression of Pax6 and Nkx6.1 mRNA and the absence of Pax7 transcription, indicating that neurons derived from NTERA2 pluripotent stem cells are characteristic of neuroectodermal cells of the ventral phenotype.

There are few reliable cell systems available to study the process of human neural development. Embryonal carcinoma (EC) cells are pluripotent stem cells derived from teratocarcinomas and offer a robust culture system to research cell differentiation in a manner pertinent to embryogenesis. Here, we describe the recent development of a series of culture procedures that together can be used to induce the differentiation of human EC stem cells, resulting in the formation of either pure populations of differentiated neurons, populations of differentiated astrocytes, or populations of immature neuronal cell types. Cell-type-specific markers were used to examine the induction of EC stem cell differentiation by retinoic acid. In direct response to manipulation of the culture environment, the expression of cell type markers correlated with the differentiation and appearance of distinct neural cell types, including neurons and astrocytes. These experiments demonstrate that cultured human EC stem cells provide a robust model cell system capable of reproducibly forming neural subtypes for research purposes.