Structural visualization of the p53/RNA polymerase II assembly.

The master tumor suppressor p53 activates transcription in response to various cellular stresses in part by facilitating recruitment of the transcription machinery to DNA. Recent studies have documented a direct yet poorly characterized interaction between p53 and RNA polymerase II (Pol II). Therefore, we dissected the human p53/Pol II interaction via single-particle cryo-electron microscopy, structural docking, and biochemical analyses. This study reveals that p53 binds Pol II via the Rpb1 and Rpb2 subunits, bridging the DNA-binding cleft of Pol II proximal to the upstream DNA entry site. In addition, the key DNA-binding surface of p53, frequently disrupted in various cancers, remains exposed within the assembly. Furthermore, the p53/Pol II cocomplex displays a closed conformation as defined by the position of the Pol II clamp domain. Notably, the interaction of p53 and Pol II leads to increased Pol II elongation activity. These findings indicate that p53 may structurally regulate DNA-binding functions of Pol II via the clamp domain, thereby providing insights into p53-regulated Pol II transcription.

Bromodomains regulate dynamic targeting of the PBAF chromatin-remodeling complex to chromatin hubs.

Chromatin remodelers actively target arrays of acetylated nucleosomes at select enhancers and promoters to facilitate or shut down the repeated recruitment of RNA polymerase II during transcriptional bursting. It is poorly understood how chromatin remodelers such as PBAF dynamically target different chromatin states inside a live cell. Our live-cell single-molecule fluorescence microscopy study reveals chromatin hubs throughout the nucleus where PBAF rapidly cycles on and off the genome. Deletion of PBAF's bromodomains impairs targeting and stable engagement of chromatin in hubs. Dual color imaging reveals that PBAF targets both euchromatic and heterochromatic hubs with distinct genome-binding kinetic profiles that mimic chromatin stability. Removal of PBAF's bromodomains stabilizes H3.3 binding within chromatin, indicating that bromodomains may play a direct role in remodeling of the nucleosome. Our data suggests that PBAF's dynamic bromodomain-mediated engagement of a nucleosome may reflect the chromatin-remodeling potential of differentially bound chromatin states.

Isolated partial anomalous origin of a branch pulmonary artery from the descending aorta.

The condition of partial anomalous origin of a branch pulmonary artery from the descending aorta could be found in several diseases and should be carefully differentiated. We report an unusual case of anomalous systemic arterial supply to normal basal segments of the left lower lung and another case of intralobar pulmonary sequestration. These two cases were treated successfully by transarterial embolisation using the Amplatzer Vascular Plug. We also set up a diagnostic algorithm to differentiate these diseases from anomalous systemic arterial supply to the pulmonary region. It is possible to make the correct diagnosis using the step-by-step diagnostic algorithm and careful interpretation of chest computed tomography angiography.

Initial fluid resuscitation (30 mL/kg) in patients with septic shock: More or less?

OBJECTIVES: The 2018 Surviving Sepsis Campaign (SSC) recommends rapid administration of 30 mL/kg crystalloid fluids for hypotension or lactate ≥4 mmol/L in patients with septic shock; however, there is limited evidence to support this recommendation. The purpose of this study was to examine the relationship between initial fluid resuscitation doses and prognosis in patients with septic shock. METHODS: This was a multicenter prospective observational study of adult patients with septic shock who were admitted to four intensive care units (ICUs) in a total of three Jiangsu Province teaching hospitals over a 3-year span from May 8, 2018, to June 15, 2021. Each enrolled patients with septic shock was categorized into the low-volume (below 20 mL/kg fluid), medium-volume (20-30 mL/kg fluid) or high-volume (above 30 mL/kg fluid) fluid group according to the initial infusion dose given for fluid resuscitation. Various demographic attributes and other variables were collected from medical records. Logistic regression and Kaplan-Meier curve analysis were used to determine the relationship between initial fluid resuscitation doses and patient outcomes. MEASUREMENTS AND MAIN RESULTS: A total of 302 patients who presented to the ICU were diagnosed with septic shock. The 28-day mortality was highest in the high-volume group (48.3%) and lowest in the medium-volume group (26.3%, P < 0.05). Patients who completed 30 mL/kg initial fluid resuscitation in the first 1-2 h had the lowest 28-day mortality rate (22.8%, P < 0.05). Logistic regression showed that a medium initial fluid volume dose was an independent protective factor, with the odds ratio (OR) indicating significantly decreased mortality (OR, 0.507; 95% confidence interval, 0.310-0.828; P = 0.007; P < 0.05). A Kaplan-Meier curve stratified by initial fluid resuscitation dose was constructed for the probability of 28-day mortality. The medium-volume fluid group showed a significantly lower 28-day mortality rate than the high-volume group or the low-volume group (log-rank test, P = 0.0016). CONCLUSION: In septic shock patients, an initial fluid resuscitation rate of 20-30 mL/kg within the first hour may be associated with reduced 28-day mortality; however, this result needs to be confirmed by further high-quality randomized controlled clinical trials. TRIAL REGISTRATION: Chinese Clinical Trial Registry, ChiCTR-OOC-17013223. Registered 2 November 2017, http://www.chictr.org.cn/showproj.aspx?proj=22674.

Ultrasonographic measurements of femoral vessel diameter in neonates weighing less than 2.5 kg.

BACKGROUND: Cannulation in low birth weight (LBW) neonates using larger sheaths could increase the risk of vascular injury. This study investigated the relationship between body weight (BW) and diameter of femoral vessels in LBW neonates and whether BW can be used to predict femoral vessel diameter. METHODS: The cohort included 100 neonates weighing < 2.5 kg (.57-2.42 kg) with a gestational age of 24-39 weeks. Vascular ultrasonography was used to measure diameters of the bilateral femoral arteries (FA) and veins (FV). The cohort was divided into four groups according to weight: group-A, 2-2.49 kg (n = 28); group-B, 1.5-1.99 kg (n = 38); group-C, 1-1.49 kg (n = 21); and group-D, < 1 kg (n = 13); or according to BSA: group-A, BSA > .16 m2 (n = 25); group-B, .13-.16 m2 (n = 40); group-C, .1-.13 m2 (n = 22); and group-D, < .1 m2 (n = 13). RESULTS: The median vessel diameters (mm) in groups A-D according to weight were FA, 1.96, 1.86, 1.78, and 1.53, and FV, 2.30, 2.28, 2.13, and 1.87, respectively. The median vessel diameters (mm) in groups A-D according to BSA were FA, 1.96, 1.86, 1.76, and 1.53, and FV, 2.30, 2.28, 2.05, and 1.87, respectively. There were positive correlations between BW and femoral vessel diameter (correlation coefficient: .56 and .55 between BW and FA and FV, respectively) (p < 0.001), and between BSA and femoral vessel diameter (correlation coefficient: .56 and .55 between BSA and FA and FV, respectively) (p < 0.001). CONCLUSIONS: BW is a predictor of femoral vessel diameter in LBW newborns. This finding may help to avoid using larger sheath in smaller vessels.

Hepatitis C virus infection induces endoplasmic reticulum stress and apoptosis in human fetal liver stem cells.

The cellular mechanisms by which hepatitis C virus (HCV) replication might mediate cytopathic effects are controversial and not entirely clear. In this study, we found that blood-borne HCV (bbHCV) infection could lead to endoplasmic reticulum (ER)-stress and mitochondria-related/caspase-dependent apoptosis at the early stages of infection based on use of the highly efficient bbHCV cell culture model established previously. Sections of bbHCV-infected human fetal liver stem cells (hFLSCs) revealed convolution and nonlinear ER, cell vacuolization, swelling of mitochondria, and numerous double membrane vesicles (DMVs). The percentage of apoptotic hFLSCs infected by bbHCV reached 29.8% at 16 h postinfection, and the amount of cytochrome c increased remarkably in the cytosolic protein fraction. However, over time, apoptosis was inhibited due to the activation of NF-κB. The expression of NF-κB-p65, Bcl-xL, XIAP, and c-FLIPL in hFLSCs was increased significantly 24 h after in infection by bbHCV. The accelerated cell death cycles involving apoptosis, regeneration and repair by bbHCV infection might give rise to the development of cirrhosis, and ultimately to hepatocellular carcinogenesis. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

[Risk factors for minimally invasive surfactant administration failure in preterm infants with respiratory distress syndrome].

OBJECTIVE: To identify risk factors for minimally invasive surfactant administration (MISA) failure in the treatment of preterm infants with respiratory distress syndrome (RDS) and the influence of MISA failure on neonatal outcome. METHODS: A retrospective analysis was performed for the clinical data of 148 preterm infants with a gestational age of ≤32 weeks and a clinical diagnosis of RDS, who were admitted to the neonatal intensive care unit of eight tertiary hospitals in Beijing, Tianjin and Hebei Province from July 1, 2017 to December 31, 2018 and were treated with MISA (bovine pulmonary surfactant, PS). According to whether MISA failure (defined as the need for mechanical ventilation within 72 hours after MISA) was observed, the infants were divided into two groups: MISA failure group (n=16) and MISA success (n=132). A logistic regression analysis was used to investigate the risk factors for MISA failure and its influence on neonatal outcome. RESULTS: The MISA failure rate was 10.8% (16/148). The logistic regression analysis showed that a high incidence rate of grade >II RDS before PS administration, low mean arterial pressure and high pulse pressure before administration, a low dose of initial PS administration, and long injection time and operation time were the risk factors for MISA failure (OR=5.983, 1.210, 1.183, 1.055, 1.036, and 1.058 respectively, P<0.05). After the control for the above risk factors, the logistic regression analysis showed that the MISA failure group had a significantly higher incidence rate of bronchopulmonary dysplasia (BPD) (OR=8.537, P<0.05). CONCLUSIONS: A high grade of RDS, a low mean arterial pressure, and a high pulse pressure before administration are independent risk factors for MISA failure, and a low dose of initial PS administration, a long injection time, and a long operation time may increase the risk of MISA failure. MISA failure may increase the incidence rate of BPD in preterm infants.

Leucine-Rich Repeat Neuronal Protein 1 Regulates Differentiation of Embryonic Stem Cells by Post-Translational Modifications of Pluripotency Factors.

Stem cell surface markers may facilitate a better understanding of stem cell biology through molecular function studies or serve as tools to monitor the differentiation status and behavior of stem cells in culture or tissue. Thus, it is important to identify additional novel stem cell markers. We used glycoproteomics to discover surface glycoproteins on human embryonic stem cells (hESCs) that may be useful stem cell markers. We found that a surface glycoprotein, leucine-rich repeat neuronal protein 1 (LRRN1), is expressed abundantly on the surface of hESCs before differentiation into embryoid bodies (EBs). Silencing of LRRN1 with short hairpin RNA (shLRRN1) in hESCs resulted in decreased capacity of self-renewal, and skewed differentiation toward endoderm/mesoderm lineages in vitro and in vivo. Meanwhile, the protein expression levels of the pluripotency factors OCT4, NANOG, and SOX2 were reduced. Interestingly, the mRNA levels of these pluripotency factors were not affected in LRRN1 silenced cells, but protein half-lives were substantially shortened. Furthermore, we found LRRN1 silencing led to nuclear export and proteasomal degradation of all three pluripotency factors. In addition, the effects on nuclear export were mediated by AKT phosphorylation. These results suggest that LRRN1 plays an important role in maintaining the protein stability of pluripotency factors through AKT phosphorylation, thus maintaining hESC self-renewal capacity and pluripotency. Overall, we found that LRRN1 contributes to pluripotency of hESC by preventing translocation of OCT4, NANOG, and SOX2 from nucleus to cytoplasm, thereby lessening their post-translational modification and degradation. Stem Cells 2018;36:1514-1524.

Analysis of small bowel angioectasia in asymptomatic individuals depending on patients' age and gender.

Objectives: Small bowel angioectasia (SBA) plays an important role in the etiologies of obscure gastrointestinal haemorrhage. But the exact prevalence of the disease is unknown, especially in asymptomatic populations. Therefore, we aimed to evaluate the prevalence of asymptomatic angioectasia in the small bowel (SB) with magnetically controlled capsule gastroscopy (MCCG).Methods: We retrospectively collected a multicentre clinical data of 508 asymptomatic patients who underwent MCCG from June 2018 to May 2019. Bowel cleanliness was rated as four grades according to the criteria, and the excellent or good preparation was classified as the adequate group. The detection rates of small bowel lesions were analysed according to the ages, genders and bowel preparations.Results: A total of 508 individuals have completed the examination. There were 316 men and 192 women with an average age of 44.5 years old. The prevalence of SBA was 11.8% (95% CI: 9.0-14.6%). 70.0% of them were over 40 years old and 73.3% were male although there was no obvious disparity found in age and gender for the SBA. Most findings were located in the proximal small bowel (jejunum). The incidence of small bowel lesions was not related to bowel preparations (p > .05).Conclusions: SBA is not uncommon in asymptomatic individuals. Age and gender may be risk factors for bleeding of angioectasia in the small bowel, but they seem to have little to do with the occurrence of it. MCCG showed no difference in ages, genders or bowel preparations of small bowel lesions among our study population.

Efficient replication of blood-borne hepatitis C virus in human fetal liver stem cells.

The development of pathogenic mechanisms, specific antiviral treatments and preventive vaccines for hepatitis C virus (HCV) infection has been limited due to lack of cell culture models that can naturally imitate the entire HCV life cycle. Here, we established an HCV cell culture model based on human fetal liver stem cells (hFLSCs) that supports the entire blood-borne hepatitis C virus (bbHCV) life cycle. More than 90% of cells remained infected by various genotypes. bbHCV was efficiently propagated, and progeny virus were infectious to hFLSCs. The virus could be passed efficiently between cells. The viral infectivity was partially blocked by specific antibodies or small interfering RNA against HCV entry factors, whereas HCV replication was inhibited by antiviral drugs. We observed viral particles of approximately 55 nm in diameter in both cell culture media and infected cells after bbHCV infection. CONCLUSION: Our data show that the entire bbHCV life cycle could be naturally imitated in hFLSCs. This model is expected to provide a powerful tool for exploring the process and the mechanism of bbHCV infection at the cellular level and for evaluating the treatment and preventive strategies of bbHCV infection. (Hepatology 2017;66:1045-1057).

Effects of summer internship and follow-up distance mentoring programs on middle and high school student perceptions and interest in health careers.

BACKGROUND: Minorities are underrepresented in health professions and efforts to recruit minority students into health careers are considered a way to reduce health disparities. There is little research about the effectiveness of these programs, other than satisfaction. This study aimed to measure program effects on student understanding of and interest in health careers. METHODS: Students took a career interest inventory, completed a scale measuring their self-reported understanding and interest in health careers, and wrote essays about health careers before and after completing a 1 week on campus internship on health careers and after a 9 month follow up distance mentoring program where they continued to interact with university faculty by videoconference about career options. Changes in inventory, scale, and essay scores were analyzed for changes over time using Wilcoxon and Mann-Whitney tests. RESULTS: Inventory scores were unchanged over time, but scale and essay scores trended upward significantly post internship and mentoring. CONCLUSION: Health career education and mentoring programs can positively affect student knowledge of health careers and their attitudes about them. The study's methods extend measures of program impact beyond satisfaction.

Structures of three distinct activator-TFIID complexes.

Sequence-specific DNA-binding activators, key regulators of gene expression, stimulate transcription in part by targeting the core promoter recognition TFIID complex and aiding in its recruitment to promoter DNA. Although it has been established that activators can interact with multiple components of TFIID, it is unknown whether common or distinct surfaces within TFIID are targeted by activators and what changes if any in the structure of TFIID may occur upon binding activators. As a first step toward structurally dissecting activator/TFIID interactions, we determined the three-dimensional structures of TFIID bound to three distinct activators (i.e., the tumor suppressor p53 protein, glutamine-rich Sp1 and the oncoprotein c-Jun) and compared their structures as determined by electron microscopy and single-particle reconstruction. By a combination of EM and biochemical mapping analysis, our results uncover distinct contact regions within TFIID bound by each activator. Unlike the coactivator CRSP/Mediator complex that undergoes drastic and global structural changes upon activator binding, instead, a rather confined set of local conserved structural changes were observed when each activator binds holo-TFIID. These results suggest that activator contact may induce unique structural features of TFIID, thus providing nanoscale information on activator-dependent TFIID assembly and transcription initiation.

Comparing High Definition Live Interactive and Store-and-Forward Consultations to In-Person Examinations.

BACKGROUND: There is little teledermatology research directly comparing remote methods, even less research with two in-person dermatologist agreement providing a baseline for comparing remote methods, and no research using high definition video as a live interactive method. OBJECTIVE: To compare in-person consultations with store-and-forward and live interactive methods, the latter having two levels of image quality. METHODS: A controlled study was conducted where patients were examined in-person, by high definition video, and by store-and-forward methods. The order patients experienced methods and residents assigned methods rotated, although an attending always saw patients in-person. The type of high definition video employed, lower resolution compressed or higher resolution uncompressed, was alternated between clinics. Primary and differential diagnoses, biopsy recommendations, and diagnostic and biopsy confidence ratings were recorded. RESULTS: Concordance and confidence were significantly better for in-person versus remote methods and biopsy recommendations were lower. Store-and-forward and higher resolution uncompressed video results were similar and better than those for lower resolution compressed video. LIMITATIONS: Dermatology residents took store-and-forward photos and their quality was likely superior to those normally taken in practice. There were variations in expertise between the attending and second and third year residents. CONCLUSION: The superiority of in-person consultations suggests the tendencies to order more biopsies or still see patients in-person are often justified in teledermatology and that high resolution uncompressed video can close the resolution gap between store-and-forward and live interactive methods.

Patient and Provider Satisfaction with Teledermatology.

BACKGROUND: There is little research comparing dermatologist and patient satisfaction with in-person, store-and-forward, and live interactive examinations. OBJECTIVE: To compare satisfaction with in-person examinations to store-and-forward and live interactive consultations having two types of video. METHODS: A controlled study was conducted where patients referred for dermatology consultations were examined in-person, by video, and by store-and-forward methods. Video changed between compressed and uncompressed on alternate clinics. Patients and dermatologists rated encounters after each examination. Dermatologists doing store-and-forward evaluations rated the quality of information provided. After experiencing all methods patients ranked their preferences. Dermatologists ranked their preferences at the end of the study. RESULTS: In-person examinations were preferred by both patients and dermatologists. Overall, satisfaction with teledermatology was still high. Patients were evenly divided in preferring store-and-forward workups or live interactive video. Dermatologists were also divided on store-and-forward and uncompressed video, but tended toward the latter. Compressed video was the least preferred method among dermatologists. LIMITATIONS: Dermatology residents took store-and-forward photos and their quality was likely superior to those normally taken in practice. CONCLUSIONS: Patients and dermatologists prefer in-person examinations and diverge on preferring store-and-forward and live interactive when video is not compressed. The amount of video compression that can be applied without noticeable image degradation is a question for future research.

Mobile Videoconferencing Apps for Telemedicine.

INTRODUCTION: The quality and performance of several videoconferencing applications (apps) tested on iOS (Apple, Cupertino, CA) and Android (Google, Mountain View, CA) mobile platforms using Wi-Fi (802.11), third-generation (3G), and fourth-generation (4G) cellular networks are described. MATERIALS AND METHODS: The tests were done to determine how well apps perform compared with videoconferencing software installed on computers or with more traditional videoconferencing using dedicated hardware. The rationale for app assessment and the testing methodology are described. RESULTS: Findings are discussed in relation to operating system platform (iOS or Android) for which the apps were designed and the type of network (Wi-Fi, 3G, or 4G) used. The platform, network, and apps interact, and it is impossible to discuss videoconferencing experienced on mobile devices in relation to one of these factors without referencing the others. CONCLUSIONS: Apps for mobile devices can vary significantly from other videoconferencing software or hardware. App performance increased over the testing period due to improvements in network infrastructure and how apps manage bandwidth.

Lessons learned from ten years of distance learning outreach.

OBJECTIVE: The study tested the efficacy of providing distance learning with real-time videoconferencing to broaden high school student knowledge of health careers. METHODS: A pilot program was tried out and extended over ten years to include other schools in four different time zones and the National Library of Medicine. Survey results, site visits, and continued school participation were used as effectiveness indicators. Student ratings, site visits, and ongoing discussions were used to evaluate critical factors in the program. RESULTS: Nine program factors contributed to success. CONCLUSIONS: Synchronous communication can be effective for outreach to special populations given appropriate infrastructure, technology, program design, and implementation.

Cloud and traditional videoconferencing technology for telemedicine and distance learning.

INTRODUCTION: Cloud-based videoconferencing versus traditional systems are described for possible use in telemedicine and distance learning. MATERIALS AND METHODS: Differences between traditional and cloud-based videoconferencing systems are examined, and the methods for identifying and testing systems are explained. Findings are presented characterizing the cloud conferencing genre and its attributes versus traditional H.323 conferencing. RESULTS: Because the technology is rapidly evolving and needs to be evaluated in reference to local needs, it is strongly recommended that this or other reviews not be considered substitutes for personal hands-on experience. CONCLUSIONS: This review identifies key attributes of the technology that can be used to appraise the relevance of cloud conferencing technology and to determine whether migration from traditional technology to a cloud environment is warranted. An evaluation template is provided for assessing systems appropriateness.

Isoquercitrin attenuates the osteoclast-mediated bone loss in rheumatoid arthritis via the Nrf2/ROS/NF-κB pathway.

An excess of osteoclastogenesis significantly contributes to the development of rheumatoid arthritis (RA). Activation of the nuclear factor erythroid-2 related factor 2 (Nrf2) and nuclear factor kappa B (NF-κB) ligand (RANKL)-induced reactive oxygen species (ROS)-to-NF-κB signaling cascade are important mechanisms regulating osteoclastogenesis; however, whether Nrf2 is involved in RANKL-induced NF-κB activation is controversial. Isoquercitrin, a natural flavonoid compound, has been shown to have Nrf2-dependent antioxidant effects inprevious studies. We sought to verify whether isoquercitrin could modulate RANKL-induced NF-κB activation by activating Nrf2, thereby affecting osteoclastogenesis. Tartrate-resistant acid phosphatase staining, F-actin ring staining and resorption pit assay suggested that isoquercitrin significantly inhibited osteoclastogenesis and osteolytic function. Mitosox staining showed that RANKL-induced ROS generation was significantly inhibited by isoquercitrin from day 3 of the osteoclast differentiation cycle. Quantitative real-time PCR, Western blot, and immunofluorescence indicated that isoquercitrin activated the Nrf2 signaling pathway and inhibited NF-κB expression. And when we used the Nrf2-specific inhibitor ML385, the inhibition of NF-κB by isoquercitrin disappeared. Moreover, we found that Nrf2 is not uninvolved in RANKL-induced NF-κB activation and may be related to the timing of ROS regulation. When we limited isoquercitrin administration to 2 days, Nrf2 remained activated and the inhibition of NF-κB disappeared. In vivo experiments suggested that isoquercitrin attenuated RA modeling-induced bone loss. Overall, isoquercitrin-activated Nrf2 blocked the RANKL-induced ROS-to-NF-κB signaling cascade response, thereby inhibiting osteoclastogenesis and bone loss. These findings provide new ideas for the treatment of RA.

Uncompressed high-definition videoconferencing tools for telemedicine and distance learning.

Uncompressed high-definition (HD) video image quality is superior to compressed HD video provided in most commercially available videoconferencing products. Uncompressed HD videoconferencing tools provide a more immersive experience because there is no reduction of image information and, in most cases, lower latency. Four open source uncompressed video applications are reviewed that have been tested at the National Library of Medicine: three transmitting uncompressed HD video and one transmitting loosely compressed standard-definition video. The technical requirements for implementing each are described, and test results in terms of image quality, latency, and application reliability are presented. Because the hardware and bandwidth requirements for uncompressed HD video are relatively high and most applications are still under development, they are generally not ready for mass deployment. Some are, however, ready for pilot testing and experimentation in clinical settings by either those who have or anticipate having bandwidth sufficient to support them or those interested in researching the effects higher-quality video may have on diagnostic and other clinical outcomes.