Lymphatic endothelial cell RXRα is critical for 9-cis-retinoic acid-mediated lymphangiogenesis and prevention of secondary lymphedema.

Secondary lymphedema is a debilitating disease characterized by abnormal soft tissue swelling and caused by lymphatic system dysfunction. Despite a high prevalence of secondary lymphedema after cancer treatments, current management is supportive and there are no approved therapeutic agents that can thwart disease progression. We have previously demonstrated that 9-cis-retinoic acid (9-cisRA) has the potential to be repurposed for lymphedema as it mitigates disease by promoting lymphangiogenesis at the site of lymphatic injury. Although the efficacy of 9-cisRA has been demonstrated in previous studies, the mechanism of action is not completely understood. In this study, we demonstrate that when RXRα is specifically deleted in lymphatic endothelial cells, 9-cisRA fails to induce lymphangiogenesis in vitro and prevent pathologic progression of postsurgical lymphedema in vivo. These findings demonstrate that downstream nuclear receptor RXRα plays a critical role in the therapeutic efficacy of 9-cisRA in postsurgical lymphedema.

The BRD4 Inhibitor dBET57 Exerts Anticancer Effects by Targeting Superenhancer-Related Genes in Neuroblastoma.

Neuroblastoma (NB) is the most common solid tumor of the neural crest cell origin in children and has a poor prognosis in high-risk patients. The oncogene MYCN was found to be amplified at extremely high levels in approximately 20% of neuroblastoma cases. In recent years, research on the targeted hydrolysis of BRD4 to indirectly inhibit the transcription of the MYCN created by proteolysis targeting chimaera (PROTAC) technology has become very popular. dBET57 (S0137, Selleck, TX, USA) is a novel and potent heterobifunctional small molecule degrader based on PROTAC technology. The purpose of this study was to investigate the therapeutic effect of dBET57 in NB and its potential mechanism. In this study, we found that dBET57 can target BRD4 ubiquitination and disrupt the proliferation ability of NB cells. At the same time, dBET57 can also induce apoptosis, cell cycle arrest, and decrease migration. Furthermore, dBET57 also has a strong antiproliferation function in xenograft tumor models in vivo. In terms of mechanism, dBET57 targets the BET protein family and the MYCN protein family by associating with CRBN and destroys the SE landscape of NB cells. Combined with RNA-seq and ChIP-seq public database analysis, we identified the superenhancer-related genes TBX3 and ZMYND8 in NB as potential downstream targets of dBET57 and experimentally verified that they play an important role in the occurrence and development of NB. In conclusion, these results suggest that dBET57 may be an effective new therapeutic drug for the treatment of NB.

Piezo1-Regulated Mechanotransduction Controls Flow-Activated Lymphatic Expansion.

BACKGROUND: Mutations in PIEZO1 (Piezo type mechanosensitive ion channel component 1) cause human lymphatic malformations. We have previously uncovered an ORAI1 (ORAI calcium release-activated calcium modulator 1)-mediated mechanotransduction pathway that triggers lymphatic sprouting through Notch downregulation in response to fluid flow. However, the identity of its upstream mechanosensor remains unknown. This study aimed to identify and characterize the molecular sensor that translates the flow-mediated external signal to the Orai1-regulated lymphatic expansion. METHODS: Various mutant mouse models, cellular, biochemical, and molecular biology tools, and a mouse tail lymphedema model were employed to elucidate the role of Piezo1 in flow-induced lymphatic growth and regeneration. RESULTS: Piezo1 was found to be abundantly expressed in lymphatic endothelial cells. Piezo1 knockdown in cultured lymphatic endothelial cells inhibited the laminar flow-induced calcium influx and abrogated the flow-mediated regulation of the Orai1 downstream genes, such as KLF2 (Krüppel-like factor 2), DTX1 (Deltex E3 ubiquitin ligase 1), DTX3L (Deltex E3 ubiquitin ligase 3L,) and NOTCH1 (Notch receptor 1), which are involved in lymphatic sprouting. Conversely, stimulation of Piezo1 activated the Orai1-regulated mechanotransduction in the absence of fluid flow. Piezo1-mediated mechanotransduction was significantly blocked by Orai1 inhibition, establishing the epistatic relationship between Piezo1 and Orai1. Lymphatic-specific conditional Piezo1 knockout largely phenocopied sprouting defects shown in Orai1- or Klf2- knockout lymphatics during embryo development. Postnatal deletion of Piezo1 induced lymphatic regression in adults. Ectopic Dtx3L expression rescued the lymphatic defects caused by Piezo1 knockout, affirming that the Piezo1 promotes lymphatic sprouting through Notch downregulation. Consistently, transgenic Piezo1 expression or pharmacological Piezo1 activation enhanced lymphatic sprouting. Finally, we assessed a potential therapeutic value of Piezo1 activation in lymphatic regeneration and found that a Piezo1 agonist, Yoda1, effectively suppressed postsurgical lymphedema development. CONCLUSIONS: Piezo1 is an upstream mechanosensor for the lymphatic mechanotransduction pathway and regulates lymphatic growth in response to external physical stimuli. Piezo1 activation presents a novel therapeutic opportunity for preventing postsurgical lymphedema. The Piezo1-regulated lymphangiogenesis mechanism offers a molecular basis for Piezo1-associated lymphatic malformation in humans.

Urinary Bladder Matrix Improves Irradiated Wound Healing in a Murine Model.

ABSTRACT: Radiation skin damage is associated with chronic wounds and poor healing. Existing localized treatment modalities have limited benefit. Therefore, there has been increased interest in biologically based solutions. In this study, we aimed to determine the effect of topical urinary bladder matrix (UBM) on chronic irradiated skin wounds using an established murine model. Our findings demonstrated that topical urinary bladder matrix significantly accelerated the healing of irradiated wounds on day 7 (P = 0.0216), day 14 (P = 0.0140), and day 21 (P = 0.0393). Histologically, urinary bladder matrix treatment was associated with higher-quality reorganization and reepithelialization of wounds, an increased density of myofibroblasts (P = 0.0004), and increased collagen deposition (P < 0.0001). In addition, quantitative real-time polymerase chain reaction data demonstrated decreased expression of profibrotic mediators (P = 0.0049). We conclude that urinary bladder matrix may be a useful, noninvasive, adjunctive therapy for the treatment of chronic irradiated skin wounds.

BRD4 inhibitor GNE987 exerts anti-cancer effects by targeting super-enhancers in neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is a common extracranial malignancy with high mortality in children. Recently, super-enhancers (SEs) have been reported to play a critical role in the tumorigenesis and development of NB via regulating a wide range of oncogenes Thus, the synthesis and identification of chemical inhibitors specifically targeting SEs are of great urgency for the clinical therapy of NB. This study aimed to characterize the activity of the SEs inhibitor GNE987, which targets BRD4, in NB. RESULTS: In this study, we found that nanomolar concentrations of GNE987 markedly diminished NB cell proliferation and survival via degrading BRD4. Meanwhile, GNE987 significantly induced NB cell apoptosis and cell cycle arrest. Consistent with in vitro results, GNE987 administration (0.25 mg/kg) markedly decreased the tumor size in the xenograft model, with less toxicity, and induced similar BRD4 protein degradation to that observed in vitro. Mechanically, GNE987 led to significant downregulation of hallmark genes associated with MYC and the global disruption of the SEs landscape in NB cells. Moreover, a novel candidate oncogenic transcript, FAM163A, was identified through analysis of the RNA-seq and ChIP-seq data. FAM163A is abnormally transcribed by SEs, playing an important role in NB occurrence and development. CONCLUSION: GNE987 destroyed the abnormal transcriptional regulation of oncogenes in NB by downregulating BRD4, which could be a potential therapeutic candidate for NB.

Plasma Long Non-Coding RNA RP11-438N5.3 as a Novel Biomarker for Non-Small Cell Lung Cancer.

BACKGROUND: Lung cancer is one of the most common malignancies around the world. The lack of early diagnosis and effective treatment strategies contributes to the poor prognosis of patients with lung cancer. Recent studies have implied the role of long non-coding RNAs (lncRNAs) in oncogenesis. The purpose of our study was to identify specific lncRNAs which were correlated with non-small cell lung cancer (NSCLC) and their potential functions. MATERIALS AND METHODS: The global plasma lncRNA profiling was performed using LncPathTM Human Cancer Array, and 11 lncRNAs were then selected for quantitative reverse transcription PCR (qRT-PCR) validation in 138 plasma samples from 69 NSCLC patients and 69 healthy controls (HCs). A noteworthy lncRNA, RP11-438N5.3, the function of which was previously unknown, was further explored on the aspect of the correlation of its expression level with clinicopathological factors. RESULTS: The results revealed that plasma level of RP11-438N5.3 was significantly lower in NSCLCs than that in HCs (p <0.01). Receiver operating characteristic (ROC) analyses showed that the area under the ROC curve (AUC) for plasma RP11-438N5.3 was 0.814 (95% CI, 0.743-0.885; p<0.01). High expression of RP11-438N5.3 in plasma correlated with favorable prognosis for NSCLC patients (Hazard ratio = 2.827; 95% CI: 1.036 to 7.718; p = 0.024; Cox regression analysis). Moreover, we found that the plasma level of stromal interaction molecule 1 (STIM1) mRNA was remarkably higher in NSCLC compared with HC (p<0.01), and the AUC for STIM1 was 0.753 (95% CI, 0.673-0.833; p<0.01), RP11-438N5.3 and STIM1 were inversely correlated with each other. CONCLUSION: Our results indicated that RP11-438N5.3 and STIM1 might provide a new strategy for NSCLC diagnosis. Furthermore, increased circulating RP11-438N5.3 level holds great potential in indicating a beneficial prognosis in NSCLC patients.

Ischemia and reperfusion injury in superficial inferior epigastric artery-based vascularized lymph node flaps.

Vascularized lymph node transfer (VLNT) is a promising treatment modality for lymphedema; however, how lymphatic tissue responds to ischemia has not been well defined. This study investigates the cellular changes that occur in lymph nodes in response to ischemia and reperfusion. Lymph node containing superficial epigastric artery-based groin flaps were isolated in Prox-1 EGFP rats which permits real time identification of lymphatic tissue by green fluorescence during flap dissection. Flaps were subjected to ischemia for either 1, 2, 4, or 8 hours, by temporarily occluding the vascular pedicle. Flaps were harvested after 0 hours, 24 hours, or 5 days of reperfusion. Using EGFP signal guidance, lymph nodes were isolated from the flaps and tissue morphology, cell apoptosis, and inflammatory cytokines were quantified and analyzed via histology, immunostaining, and rtPCR. There was a significant increase in collagen deposition and tissue fibrosis in lymph nodes after 4 and 8 hours of ischemia compared to 1 and 2 hours, as assessed by picrosirius red staining. Cell apoptosis significantly increased after 4 hours of ischemia in all harvest times. In tissue subject to 4 hours of ischemia, longer reperfusion periods were associated with increased rates of CD3+ and CD45+ cell apoptosis. rtPCR analysis demonstrated significantly increased expression of CXCL1/GRO-α with 2 hours of ischemia and increased PECAM-1 and TNF-α expression with 1 hour of ischemia. Significant cell death and changes in tissue morphology do not occur until after 4 hours of ischemia; however, analysis of inflammatory biomarkers suggests that ischemia reperfusion injury can occur with as little as 2 hours of ischemia.

Prevention of postsurgical lymphedema via immediate delivery of sustained-release 9-cis retinoic acid to the lymphedenectomy site.

BACKGROUND AND OBJECTIVES: Previously, we have shown that 9-cis retinoic acid (9-cis RA) stimulates lymphangiogenesis and limits postsurgical lymphedema in animal models when administered via daily intraperitoneal injections. In this study, we investigate whether a single-use depot 9-cis RA drug delivery system (DDS) implanted at the site of lymphatic injury can mitigate the development of lymphedema in a clinically relevant mouse limb model. METHODS: Hind limb lymphedema was induced via surgical lymphadenectomy and irradiation. Animals were divided into two treatment groups: (1) 9-cis RA DDS, (2) placebo DDS. Outcomes measured included paw thickness, lymphatic clearance and density, epidermal thickness, and collagen deposition. RESULTS: Compared with control animals, 9-cis RA-treated animals had significantly less paw swelling from postoperative week 3 (P = .04) until the final timepoint at week 6 (P = .0007). Moreover, 9-cis RA-treated animals had significantly faster lymphatic clearance (P < .05), increased lymphatic density (P = .04), reduced lymphatic vessel size (P = .02), reduced epidermal hyperplasia (P = .04), and reduced collagen staining (P = .10). CONCLUSIONS: Animals receiving 9-cis RA sustained-release implants at the time of surgery had improved lymphatic function and structure, indicating reduced lymphedema progression. Thus, we demonstrate that 9-cis RA contained within a single-use depot DDS has favorable properties in limiting pathologic responses to lymphatic injury and may be an effective strategy against secondary lymphedema.

A Pre-clinical Animal Model of Secondary Head and Neck Lymphedema.

Head and neck lymphedema (HNL) is a disfiguring disease affecting over 90% of patients treated for head and neck cancer. Animal models of lymphedema are used to test pharmacologic and microsurgical therapies; however, no animal model for HNL is described in the literature to date. In this study we describe the first reproducible rat model for HNL. Animals were subjected to two surgical protocols: (1) lymphadenectomy plus irradiation; and (2) sham surgery and no irradiation. Head and neck expansion was measured on post-operative days 15, 30 and 60. Magnetic resonance imaging (MRI) was acquired at the same time points. Lymphatic drainage was measured at day 60 via indocyanine green (ICG) lymphography, after which animals were sacrificed for histological analysis. Postsurgical lymphedema was observed 100% of the time. Compared to sham-operated animals, lymphadenectomy animals experienced significantly more head and neck swelling at all timepoints (P < 0.01). Lymphadenectomy animals had significantly slower lymphatic drainage for 6 days post-ICG injection (P < 0.05). Histological analysis of lymphadenectomy animals revealed 83% greater subcutis thickness (P = 0.008), 22% greater collagen deposition (P = 0.001), 110% greater TGFß1+ cell density (P = 0.04), 1.7-fold increase in TGFß1 mRNA expression (P = 0.03), and 114% greater T-cell infiltration (P = 0.005) compared to sham-operated animals. In conclusion, animals subjected to complete lymph node dissection and irradiation developed changes consistent with human clinical postsurgical HNL. This was evidenced by significant increase in all head and neck measurements, slower lymphatic drainage, subcutaneous tissue expansion, increased fibrosis, and increased inflammation compared to sham-operated animals.

ShenFu Preparation Protects AML12 Cells Against Palmitic Acid-Induced Injury Through Inhibition of Both JNK/Nox4 and JNK/NFκB Pathways.

BACKGROUND/AIMS: Nonalcoholic steatohepatitis includes steatosis along with liver inflammation, hepatocyte injury and fibrosis. In this study, we investigated the protective role and the potential mechanisms of a traditional Chinese medicine ShenFu (SF) preparation in an in vitro hepatic steatosis model. METHODS: In palmitic acid (PA)-induced murine hepatic AML12 cell injury, effects of SF preparation on cellular apoptosis and intracellular triglyceride (iTG) level were assessed using TUNEL and TG Colorimetric Assay. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) levels were measured using DCF and JC-1 assay. Cytokine levels were evaluated using ELISA assay. Immunoblot was used to compare the activation level of c-Jun N terminal kinase (JNK), NADPH oxidase (Nox4), and NFκB pathways. RESULTS: Addition of SF preparation prevented PA-mediated increase of apoptosis and iTG as well as IL-8 and IL-6. In PA-treated cell, SF preparation reduced the level of Nox4 and ROS, while increasing the level of MMP and the expression of manganese superoxide dismutase (MnSOD) and catalase, indicating emendation of mitochondrial dysfunction. Nox4 inhibitor GKT137381 prevented PA-induced increase of ROS and apoptosis, while decreasing iTG slightly and not influencing the level of IL-8 and IL-6. SF preparation prevented PA-induced upregulation of phospho-JNK. JNK inhibitor SP600125 prevented PA-mediated increase of Nox4, IL-8, IL-6 and iTG. Nuclear translocation of NFκB/p65 was detected in PA-treated cells, which was prevented by SF preparation. An IκB degradation inhibitor, BAY11-7082, prevented PA-induced increase of IL-8 and IL-6 as well as iTG, whereas it only decreased ROS levels slightly and showed no influence on cellular apoptosis. CONCLUSION: SF preparation shows a beneficial role in prevention of hepatocyte injury by attenuating oxidative stress and cytokines production at least partially through inhibition of JNK/Nox4 and JNK/NFκB pathway, respectively.

Community Air Sensor Network (CAIRSENSE) project: evaluation of low-cost sensor performance in a suburban environment in the southeastern United States.

Advances in air pollution sensor technology have enabled the development of small and low-cost systems to measure outdoor air pollution. The deployment of a large number of sensors across a small geographic area would have potential benefits to supplement traditional monitoring networks with additional geographic and temporal measurement resolution, if the data quality were sufficient. To understand the capability of emerging air sensor technology, the Community Air Sensor Network (CAIRSENSE) project deployed low-cost, continuous, and commercially available air pollution sensors at a regulatory air monitoring site and as a local sensor network over a surrounding ∼ 2 km area in the southeastern United States. Collocation of sensors measuring oxides of nitrogen, ozone, carbon monoxide, sulfur dioxide, and particles revealed highly variable performance, both in terms of comparison to a reference monitor as well as the degree to which multiple identical sensors produced the same signal. Multiple ozone, nitrogen dioxide, and carbon monoxide sensors revealed low to very high correlation with a reference monitor, with Pearson sample correlation coefficient (r) ranging from 0.39 to 0.97, 0.25 to 0.76, and 0.40 to 0.82, respectively. The only sulfur dioxide sensor tested revealed no correlation (r < 0.5) with a reference monitor and erroneously high concentration values. A wide variety of particulate matter (PM) sensors were tested with variable results - some sensors had very high agreement (e.g., r = 0.99) between identical sensors but moderate agreement with a reference PM2.5 monitor (e.g., r = 0.65). For select sensors that had moderate to strong correlation with reference monitors (r > 0.5), step-wise multiple linear regression was performed to determine if ambient temperature, relative humidity (RH), or age of the sensor in number of sampling days could be used in a correction algorithm to improve the agreement. Maximum improvement in agreement with a reference, incorporating all factors, was observed for an NO2 sensor (multiple correlation coefficient R2 adj-orig = 0.57, R2 adj-final = 0.81); however, other sensors showed no apparent improvement in agreement. A four-node sensor network was successfully able to capture ozone (two nodes) and PM (four nodes) data for an 8-month period of time and show expected diurnal concentration patterns, as well as potential ozone titration due to nearby traffic emissions. Overall, this study demonstrates the performance of emerging air quality sensor technologies in a real-world setting; the variable agreement between sensors and reference monitors indicates that in situ testing of sensors against benchmark monitors should be a critical aspect of all field studies.

Performance of the Plusoptix A09 photoscreener in detecting amblyopia risk factors in Chinese children attending an eye clinic.

PURPOSE: To assess the accuracy of the Plusoptix A09 photoscreener in detecting amblyopia risk factors in children and determine referral criteria when using Plusoptix A09 for a large-scale vision screening. METHODS: Pediatric patients attending our eye clinic underwent a comprehensive ophthalmic examination that included photorefraction, orthoptic examination, anterior segment assessment, fundus examination and cycloplegic retinoscopy. The measurements were collected for statistical analyses. RESULTS: One hundred and seventy-eight children (mean age ± SD: 6.2±2.4 years, range: 2.2 to 14.1 years) were included in the study. The mean spherical equivalent (SE) obtained using Plusoptix A09 (PSE) was 0.57 D lower than that obtained from cycloplegic retinoscopy (CRSE) (P = 0.00). However, there was no statistically significant difference of Jackson cross cylinder J0 and J45 between Plusoptix A09 (PJ) and cycloplegic retinoscopy (CRJ) (P = 0.14, P = 0.26). The relationship of SE obtained from Plusoptix A09 and SE obtained from cycloplegic retinoscopy was presented as the equation: CRSE = 0.358 + 0.776 PSE + 0.064 PSE2 + 0.011 PSE3. Based on the Receiver Operating Characteristic (ROC) curve, the Plusoptix A09 had an overall sensitivity of 94.9% and specificity of 67.5% for detecting refractive amblyopia risk factors. The sensitivity and specificity of the Plusoptix A09 for detection of strabismus were 40.7% and 98.3%, respectively; detection of amblyopia and/or strabismus was 84.7% and 63.2%, respectively. CONCLUSIONS: The Plusoptix A09 photoscreener underestimated hyperopia and overestimated myopia according to SE when compared with cycloplegic retinoscopy. The accuracy of the Plusoptix A09 in detecting amblyopia risk factors in children could be improved by the regression equation and optimized criteria for refractive amblyopia risk factors developed in the present study. Moreover, the Plusoptix A09 photoscreener is not suitable for a large-scale strabismus screening when it is applied solely.

Field assessment of the Village Green Project: an autonomous community air quality monitoring system.

Continuous, long-term, and time-resolved measurement of outdoor air pollution has been limited by logistical hurdles and resource constraints. Measuring air pollution in more places is desired to address community concerns regarding local air quality impacts related to proximate sources, to provide data in areas lacking regional air monitoring altogether, or to support environmental awareness and education. This study integrated commercially available technologies to create the Village Green Project (VGP), a durable, solar-powered air monitoring park bench that measures real-time ozone, PM2.5, and meteorological parameters. The data are wirelessly transmitted via cellular modem to a server, where automated quality checks take place before data are provided to the public nearly instantaneously. Over 5500 h of data were successfully collected during the first ten months of pilot testing in Durham, North Carolina, with about 13 days (5.5%) of downtime because of low battery power. Additional data loss (4-14% depending on the measurement) was caused by infrequent wireless communication interruptions and instrument maintenance. The 94.5% operational time via solar power was within 1.5% of engineering calculations using historical solar data for the location. The performance of the VGP was evaluated by comparing the data to nearby air monitoring stations operating federal equivalent methods (FEM), which exhibited good agreement with the nearest benchmark FEMs for hourly ozone (r(2) = 0.79) and PM2.5 (r(2) = 0.76).

Comparing exposure metrics for the effects of fine particulate matter on emergency hospital admissions.

A crucial step in an epidemiological study of the effects of air pollution is to accurately quantify exposure of the population. In this paper, we investigate the sensitivity of the health effects estimates associated with short-term exposure to fine particulate matter with respect to three potential metrics for daily exposure: ambient monitor data, estimated values from a deterministic atmospheric chemistry model, and stochastic daily average human exposure simulation output. Each of these metrics has strengths and weaknesses when estimating the association between daily changes in ambient exposure to fine particulate matter and daily emergency hospital admissions. Monitor data is readily available, but is incomplete over space and time. The atmospheric chemistry model output is spatially and temporally complete but may be less accurate than monitor data. The stochastic human exposure estimates account for human activity patterns and variability in pollutant concentration across microenvironments, but requires extensive input information and computation time. To compare these metrics, we consider a case study of the association between fine particulate matter and emergency hospital admissions for respiratory cases for the Medicare population across three counties in New York. Of particular interest is to quantify the impact and/or benefit to using the stochastic human exposure output to measure ambient exposure to fine particulate matter. Results indicate that the stochastic human exposure simulation output indicates approximately the same increase in the relative risk associated with emergency admissions as using a chemistry model or monitoring data as exposure metrics. However, the stochastic human exposure simulation output and the atmospheric chemistry model both bring additional information, which helps to reduce the uncertainly in our estimated risk.

Assessment of inter-individual, geographic, and seasonal variability in estimated human exposure to fine particles.

Health effects associated with ambient fine particle (PM(2.5)) exposure are typically estimated based on concentration-response (C-R) functions using area-wide concentration as an exposure surrogate. Persons 65 and older are particularly susceptible to adverse effects from PM(2.5) exposure. Using a stochastic microenvironmental simulation model, distributions of daily PM(2.5) exposures were estimated based on ambient concentration, air exchange rate, penetration factor, deposition rate, indoor emission sources, census data, and activity diary data, and compared for selected regions and seasons. Even though the selected subpopulation spends an average of over 20 h per day indoors, the ratio of daily average estimated exposure to ambient concentration (E(a)/C) is approximately 0.5. The daily average E(a)/C ratio varies by a factor of 4-5 over a 95% frequency range among individuals, primarily from variability in air exchange rates. The mean E(a)/C varies by 6-36% among selected NC, TX, and NYC domains, and 15-34% among four seasons, as a result of regional differences in housing stock and seasonal differences in air exchange rates. Variability in E(a)/C is a key factor that may help explain heterogeneity in C-R functions across cities and seasons. Priorities for improving exposure estimates are discussed.

[Prevalence of trachoma in rural primary school children in Tengzhou City of Shandong Province in China].

OBJECTIVE: To investigate the prevalence of trachoma and its risk factors in rural primary school children in Tengzhou City of Shandong Province. METHODS: In this cross-sectional population-based study, children aged 5 to 14 years old in primary school were randomly selected by a cluster sampling in which school shift was the sampling unit. Out of 2742 students, 2676 were eligible. The examination rate was 97.60%. All selected students were assessed for trachoma using the simplified grading scheme proposed by the World Health Organization (WHO). Statistical significance was calculated using Chi-square tests. RESULTS: Out of 2676 eligible children, 593 cases of active trachoma were found, the prevalence of trachoma was 22.16% (95% CI:20.59%-23.73%). In 1606 boys, the prevalence of active trachoma was 19.74%, compared with 25.79% for girls. Girls were affected by active trachoma higher than boys (25.79% versus 19.74%, P=0.000). There was no significant difference among different age groups in term of the prevalence of trachoma (P=0.052). The prevalence of trachomatous follicle (TF), trachomatous inflammation (TI), and trachomatous scarring (TS) was 5.68% (152/2676), 19.21% (514/2676), 0.56% (15/2676), respectively. TI was more prevalent in girls than in boys (22.90% versus 16.75%, P=0.000). CONCLUSIONS: Trachoma is still endemic in children of primary schools in Tengzhou rural areas. Some interventions including mass treatment with antibiotics, improvement of hygienic conditions, and improvement of primary eye care are needed.

Wound trauma increases radiation-induced mortality by activation of iNOS pathway and elevation of cytokine concentrations and bacterial infection.

Abstract Although it is documented that concurrent wounding increases mortality from radiation injury, the molecular mechanism of combined injury is unknown. In this study, mice were exposed to gamma radiation followed by skin wounding. Wound trauma exacerbated radiation-induced mortality, reducing the LD(50/30) from 9.65 Gy to 8.95 Gy. Analyses of histopathology, inducible nitric oxide synthase (iNOS), and serum cytokines were performed on mouse ileum and skin at various times after 9.75 Gy and/or wounding. In the ileum, the villi were significantly shortened 3 days postirradiation but not after wounding; combined injury resulted in decreased villus width and tunica muscularis thickness. The skin of mice subjected to combined injury was less cellular and had a smaller healing bud than the skin of mice subjected to wounding alone. Combined injury significantly delayed wound closure times; it also prolonged the increased levels of iNOS protein in the skin and ileum. iNOS up-regulation was correlated with increases in transcription factors, including NF-kappaB and NF-IL6. The increase in NF-IL6 may be due to increases in cytokines, including IL-1beta, -6, -8, -9, -10 and -13, G-CSF, eotaxin, INF-gamma, MCP-1, MIP-1alpha and MIP-1beta. Combined injury resulted in early detection of bacteria in the blood of the heart and liver, whereas radiation alone resulted in later detection of bacteria; only a transient bacteremia occurred after wounding alone. Results suggest that enhancement of iNOS, cytokines and bacterial infection triggered by combined injury may contribute to mortality. Agents that inhibit these responses may prove to be therapeutic for combined injury and may reduce related mortality.

Nucleocytoplasmic shuttling of the retinoblastoma tumor suppressor protein via Cdk phosphorylation-dependent nuclear export.

The retinoblastoma (RB) tumor suppressor protein is a negative regulator of cell proliferation that is functionally inactivated in the majority of human tumors. Elevated Cdk activity via RB pathway mutations is observed in virtually every human cancer. Thus, Cdk inhibitors have tremendous promise as anticancer agents although detailed mechanistic knowledge of their effects on RB function is needed to harness their full potential. Here, we illustrate a novel function for Cdks in regulating the subcellular localization of RB. We present evidence of significant cytoplasmic mislocalization of ordinarily nuclear RB in cells harboring Cdk4 mutations. Our findings uncover a novel mechanism to circumvent RB-mediated growth suppression by altered nucleocytoplasmic trafficking via the Exportin1 pathway. Cytoplasmically mislocalized RB could be efficiently confined to the nucleus by inhibiting the Exportin1 pathway, reducing Cdk activity, or mutating the Cdk-dependent phosphorylation sites in RB that result in loss of RB-Exportin1 association. Thus RB-mediated tumor suppression can be subverted by phosphorylation-dependent enhancement of nuclear export. These results support the notion that tumor cells can modulate the protein transport machinery thereby making the protein transport process a viable therapeutic target.

E2F-dependent repression of topoisomerase II regulates heterochromatin formation and apoptosis in cells with melanoma-prone mutation.

RB pathway mutations, especially at the CDK4 and INK4A loci, are hallmarks of melanomagenesis. It is presently unclear what advantages these alterations confer during melanoma progression and how they influence melanoma therapy. Topoisomerase II inhibitors are widely used to treat human malignancies, including melanoma, although their variable success is attributable to a poor understanding of their mechanism of action. Using mouse and human cells harboring the melanoma-prone p16Ink4a-insensitive CDK4R24C mutation, we show here that topoisomerase II proteins are direct targets of E2F-mediated repression. Drug-treated cells fail to load repressor E2Fs on topoisomerase II promoters leading to elevated topoisomerase II levels and an enhanced sensitivity of cells to apoptosis. This is associated with the increased formation of heterochromatin domains enriched in structural heterochromatin proteins, methylated histones H3/H4, and topoisomerase II. We refer to these preapoptotic heterochromatin domains as apoptosis-associated heterochromatic foci. We suggest that cellular apoptosis is preceded by an intermediary chromatin remodeling state that involves alterations of DNA topology by topoisomerase II enzymes and gene silencing via formation of heterochromatin. These observations provide novel insight into the mechanism of drug action that influence treatment outcome: drug sensitivity or drug resistance.

Effects of E-cadherin transfection on gene expression of a gallbladder carcinoma cell line: repression of MTS1/S100A4 gene expression.

E-cadherin is important in cell-to-cell adhesion and controls cell polarity and tissue morphology. Loss of E-cadherin expression occurs in various human tumors and is the first step in cancer invasion and metastasis. We demonstrate that the exogenous expression of E-cadherin transfected into G-415 GB cells not only increases cell-to-cell adhesion but also reduces in vitro cell proliferation, motility and invasion. Our aim was to determine what genes are most affected by the exogenous expression of E-cadherin in GB cancer cells. We analyzed gene expression pertaining to cell proliferation, motility and invasion. Conventional RT-PCR was performed for these genes; quantitative RT-PCR was carried out on genes exhibiting altered expression. Conventional RT-PCR revealed that E-cadherin transfection suppressed expression of mts1 mRNA and increased that of c-myc and MT1-MMP. In quantitative RT-PCR analysis, levels of c-myc and MT1-MMP mRNA were elevated by to 2.56- and 2.22-fold, respectively, in the E-cadherin transfectant, whereas mts-1 was 7.14-fold suppressed compared to parental cells. These results indicated that expression of mts1 mRNA was most affected by E-cadherin transfection. Immunocytochemical analysis of transfectant and parental cells demonstrated an inverse correlation in E-cadherin and mts1 expression. Immunohistochemical analysis of 37 GB cancer specimens confirmed this observation in vivo. Loss of E-cadherin expression followed by expression of the mts1 gene may be an important event for increasing cell proliferation, motility and invasion activity in the progression of GB cancer.