Long noncoding RNA GATA2-AS1 augments endothelial hypoxia inducible factor 1-α induction and regulates hypoxic signaling.

Vascular endothelial cells form the inner cellular lining of blood vessels and have myriad physiologic functions including angiogenesis and response to hypoxia. We recently identified a set of endothelial cell (EC)-enriched long noncoding RNAs (lncRNAs) in differentiated human primary cell types and described the role of the STEEL lncRNA in angiogenic patterning. We sought to further understand the role of EC-enriched lncRNAs in physiologic adaptation of the vascular endothelium. In this work, we describe an abundant, cytoplasmic, and EC-enriched lncRNA, GATA2-AS1, that is divergently transcribed from the EC-enriched developmental regulator, GATA2. While GATA2-AS1 is largely coexpressed with GATA2 in ECs, GATA2-AS1 and GATA2 appear to be complementary rather than synergistic as they have mostly distinct target genes. Common single nucleotide variants in GATA2-AS1 exons are associated with early-onset coronary artery disease and decreased expression of GATA2-AS1 in endothelial cell lines. In most cells, HIF1-α is central to the transcriptional response to hypoxia, while in ECs, both HIF1-α and HIF2-α are required to coordinate an acute and chronic response, respectively. In this setting, GATA2-AS1 contributes to the "HIF switch" and augments HIF1-α induction in acute hypoxia to regulate HIF1-α/HIF2-α balance. In hypoxia, GATA2-AS1 orchestrates HIF1-α-dependent induction of the glycolytic pathway and HIF1-α-independent maintenance of mitochondrial biogenesis. Similarly, GATA2-AS1 coordinates both metabolism and "tip/stalk" cell signaling to regulate angiogenesis in hypoxic ECs. Furthermore, we find that GATA2-AS1 expression patterns are perturbed in atherosclerotic disease. Together, these results define a role for GATA2-AS1 in the EC-specific response to hypoxia.

Prognostic Value of EMT Gene Signature in Malignant Mesothelioma.

Malignant mesothelioma (MESO) consists of epithelioid, biphasic, and sarcomatoid subtypes with different epithelial-mesenchymal transition (EMT) phenotypes. We previously identified a panel of four MESO EMT genes correlating with an immunosuppressive tumor microenvironment and poor survival. In this study, we investigated the correlation between these MESO EMT genes, the immune profile, and the genomic and epigenomic alterations to identify potential therapeutic targets to prevent or reverse the EMT process. Using multiomic analysis, we observed that the MESO EMT genes were positively correlated with hypermethylation of epigenetic genes and loss of CDKN2A/B expression. MESO EMT genes such as COL5A2, ITGAV, SERPINH1, CALD1, SPARC, and ACTA2 were associated with upregulation of TGF-ß signaling, hedgehog signaling, and IL-2-STAT5 signaling and downregulation of the IFN-α and IFN-γ response. Immune checkpoints such as CTLA4, CD274 (PD-L1), PDCD1LG2 (PD-L2), PDCD1 (PD-1), and TIGIT were upregulated, while LAG3, LGALS9, and VTCN1 were downregulated with the expression of MESO EMT genes. CD160, KIR2DL1, and KIR2DL3 were also broadly downregulated with the expression of MESO EMT genes. In conclusion, we observed that the expression of a panel of MESO EMT genes was associated with hypermethylation of epigenetic genes and loss of expression of CDKN2A and CDKN2B. Expression of MESO EMT genes was associated with downregulation of the type I and type II IFN response, loss of cytotoxicity and NK cell activity, and upregulation of specific immune checkpoints, as well as upregulation of the TGF-ß1/TGFBR1 pathway.

Angiogenic patterning by STEEL, an endothelial-enriched long noncoding RNA.

Endothelial cell (EC)-enriched protein coding genes, such as endothelial nitric oxide synthase (eNOS), define quintessential EC-specific physiologic functions. It is not clear whether long noncoding RNAs (lncRNAs) also define cardiovascular cell type-specific phenotypes, especially in the vascular endothelium. Here, we report the existence of a set of EC-enriched lncRNAs and define a role for spliced-transcript endothelial-enriched lncRNA (STEEL) in angiogenic potential, macrovascular/microvascular identity, and shear stress responsiveness. STEEL is expressed from the terminus of the HOXD locus and is transcribed antisense to HOXD transcription factors. STEEL RNA increases the number and integrity of de novo perfused microvessels in an in vivo model and augments angiogenesis in vitro. The STEEL RNA is polyadenylated, nuclear enriched, and has microvascular predominance. Functionally, STEEL regulates a number of genes in diverse ECs. Of interest, STEEL up-regulates both eNOS and the transcription factor Kruppel-like factor 2 (KLF2), and is subject to feedback inhibition by both eNOS and shear-augmented KLF2. Mechanistically, STEEL up-regulation of eNOS and KLF2 is transcriptionally mediated, in part, via interaction of chromatin-associated STEEL with the poly-ADP ribosylase, PARP1. For instance, STEEL recruits PARP1 to the KLF2 promoter. This work identifies a role for EC-enriched lncRNAs in the phenotypic adaptation of ECs to both body position and hemodynamic forces and establishes a newer role for lncRNAs in the transcriptional regulation of EC identity.

A Newborn With Skin Lesions, Thrombocytopenia, and Gastrointestinal Bleeding.

A term infant girl was admitted for evaluation of severe thrombocytopenia. She also had purpura-like skin lesions. A complete blood count showed a platelet count of 40×10/L (normal value: 150 to 400×10/L). She received random donor platelet transfusions and intravenous immunoglobulin therapy; however, thrombocytopenia persisted. She developed bloody stools on day 5 of life and hematemesis on day 9. Upper gastrointestinal endoscopy revealed multiple small, 2 to 5 mm, vascular lesions throughout the stomach body and proximal duodenum. Our multidisciplinary team will discuss an approach towards a term infant with thrombocytopenia and gastrointestinal bleeding, the diagnostic challenges, and patient management.

The mucinous domain of pancreatic carboxyl-ester lipase (CEL) contains core 1/core 2 O-glycans that can be modified by ABO blood group determinants.

Carboxyl-ester lipase (CEL) is a pancreatic fat-digesting enzyme associated with human disease. Rare mutations in the CEL gene cause a syndrome of pancreatic exocrine and endocrine dysfunction denoted MODY8, whereas a recombined CEL allele increases the risk for chronic pancreatitis. Moreover, CEL has been linked to pancreatic ductal adenocarcinoma (PDAC) through a postulated oncofetal CEL variant termed feto-acinar pancreatic protein (FAPP). The monoclonal antibody mAb16D10 was previously reported to detect a glycotope in the highly O-glycosylated, mucin-like C terminus of CEL/FAPP. We here assessed the expression of human CEL in malignant pancreatic lesions and cell lines. CEL was not detectably expressed in neoplastic cells, implying that FAPP is unlikely to be a glycoisoform of CEL in pancreatic cancer. Testing of the mAb16D10 antibody in glycan microarrays then demonstrated that it recognized structures containing terminal GalNAc-α1,3(Fuc-α1,2)Gal (blood group A antigen) and also repeated protein sequences containing GalNAc residues linked to Ser/Thr (Tn antigen), findings that were supported by immunostainings of human pancreatic tissue. To examine whether the CEL glycoprotein might be modified by blood group antigens, we used high-sensitivity MALDI-TOF MS to characterize the released O-glycan pool of CEL immunoprecipitated from human pancreatic juice. We found that the O-glycome of CEL consisted mainly of core 1/core 2 structures with a composition depending on the subject's FUT2 and ABO gene polymorphisms. Thus, among digestive enzymes secreted by the pancreas, CEL is a glycoprotein with some unique characteristics, supporting the view that it could serve additional biological functions to its cholesteryl esterase activity in the duodenum.

USR-VS: a web server for large-scale prospective virtual screening using ultrafast shape recognition techniques.

Ligand-based Virtual Screening (VS) methods aim at identifying molecules with a similar activity profile across phenotypic and macromolecular targets to that of a query molecule used as search template. VS using 3D similarity methods have the advantage of biasing this search toward active molecules with innovative chemical scaffolds, which are highly sought after in drug design to provide novel leads with improved properties over the query molecule (e.g. patentable, of lower toxicity or increased potency). Ultrafast Shape Recognition (USR) has demonstrated excellent performance in the discovery of molecules with previously-unknown phenotypic or target activity, with retrospective studies suggesting that its pharmacophoric extension (USRCAT) should obtain even better hit rates once it is used prospectively. Here we present USR-VS (http://usr.marseille.inserm.fr/), the first web server using these two validated ligand-based 3D methods for large-scale prospective VS. In about 2 s, 93.9 million 3D conformers, expanded from 23.1 million purchasable molecules, are screened and the 100 most similar molecules among them in terms of 3D shape and pharmacophoric properties are shown. USR-VS functionality also provides interactive visualization of the similarity of the query molecule against the hit molecules as well as vendor information to purchase selected hits in order to be experimentally tested.

A systematic review of evidence for fitness-to-drive among people with the mental health conditions of schizophrenia, stress/anxiety disorder, depression, personality disorder and obsessive compulsive disorder.

BACKGROUND: Limited evidence exists regarding fitness-to-drive for people with the mental health conditions of schizophrenia, stress/anxiety disorder, depression, personality disorder and obsessive compulsive disorder (herein simply referred to as 'mental health conditions'). The aim of this paper was to systematically search and classify all published studies regarding driving for this population, and then critically appraise papers addressing assessment of fitness-to-drive where the focus was not on the impact of medication on driving. METHODS: A systematic search of three databases (CINAHL, PSYCHINFO, EMBASE) was completed from inception to May 2016 to identify all articles on driving and mental health conditions. Papers meeting the eligibility criteria of including data relating to assessment of fitness-to-drive were critically appraised using the American Academy of Neurology and Centre for Evidence-Based Medicine protocols. RESULTS: A total of 58 articles met the inclusion criteria of driving among people with mental health conditions studied, and of these, 16 contained data and an explicit focus on assessment of fitness-to-drive. Assessment of fitness-to-drive was reported in three ways: 1) factors impacting on the ability to drive safely among people with mental health conditions, 2) capability and perception of health professionals assessing fitness-to-drive of people with mental health conditions, and 3) crash rates. The level of evidence of the published studies was low due to the absence of controls, and the inability to pool data from different diagnostic groups. Evidence supporting fitness-to-drive is conflicting. CONCLUSIONS: There is a relatively small literature in the area of driving with mental health conditions, and the overall quality of studies examining fitness-to-drive is low. Large-scale longitudinal studies with age-matched controls are urgently needed in order to determine the effects of different conditions on fitness-to-drive.

Gastrointestinal, influenza-like illness and dermatological complaints following exposure to floodwater: a cross-sectional survey in The Netherlands.

Extreme rainfall events may cause pluvial flooding, increasing the transmission of several waterborne pathogens. However, the risk of experiencing clinically overt infections following exposure to pluvial floodwater is poorly estimated. A retrospective cross-sectional survey was performed to quantify the occurrence of self-reported gastrointestinal, influenza-like illness (ILI) and dermatological complaints, and the frequency of visits to the general practitioner (GP), during a 4-week observation period following pluvial flooding at seven locations in The Netherlands. Questionnaires were sent to 817 flooded households, 149 (17%) of which returned the questionnaire reporting information for 199 participants. Contact with floodwater was significantly associated with increased occurrence of gastrointestinal [odds ratio (OR 4·44)], ILI (OR 2·75) and dermatological (OR 6·67) complaints, and GP visits (OR 2·72). Having hand contact with floodwater was associated with gastrointestinal and dermatological complaints, whereas ILI complaints were associated with being engaged in post-flooding cleaning operations and having walked/cycled through floodwater. This study shows that floodwater-associated diseases occur in urban settings following extreme rainfall events in a high-income country. As pluvial floods are expected to escalate in the future due to global climate change, further research is warranted to determine the disease burden of pluvial flooding and to assess the effect of different interventions, including raising awareness among stakeholders.

Decreased microRNA-198 expression and its prognostic significance in human glioma.

Previous evidence has shown the association of aberrant miR-198 expression with tumorigenesis and progression of many human malignancies. However, its involvement in human glioma is still unclear. Therefore, the aim of the current study was to investigate the expression and function of miR-198 in human gliomas. Using real-time quantitative RT-PCR, we examined miR-198 expression in 122 pairs of human gliomas and matched non-neoplastic brain tissues. The association of miR-198 expression with clinicopathological factors was also analyzed. Then, the effects of miR-198 on the biological behavior of glioma cells in vitro were evaluated. Our results showed that miR-198 expression was significantly downregulated in gliomas compared with corresponding non-neoplastic brain tissues (P < 0.001). Furthermore, low levels of miR-198 were associated with a higher WHO grade and lower Karnofsky performance status (KPS) score. A multivariate Cox regression analysis identified decreased miR-198 expression as an independent factor predicting poor prognosis for glioma patients. Lastly, in vitro functional analysis revealed that overexpression of miR- 198 in U87 cells reduced cell proliferation, promoted cell apoptosis, and inhibited cell invasion and migration. Taken together, these findings indicate that miR-198 may act as a tumor suppressor in human glioma, and may serve as a novel target for molecular therapies of this disease.

Proposed management algorithm for severe hypoxemia after liver transplantation in the hepatopulmonary syndrome.

The hepatopulmonary syndrome (HPS) is defined as the triad of liver disease, intrapulmonary vascular dilatation, and abnormal gas exchange, and is found in 10-32% of patients with liver disease. Liver transplantation is the only known cure for HPS, but patients can develop severe posttransplant hypoxemia, defined as a need for 100% inspired oxygen to maintain a saturation of ≥85%. This complication is seen in 6-21% of patients and carries a 45% mortality. Its management requires the application of specific strategies targeting the underlying physiologic abnormalities in HPS, but awareness of these strategies and knowledge on their optimal use is limited. We reviewed existing literature to identify strategies that can be used for this complication, and developed a clinical management algorithm based on best evidence and expert opinion. Evidence was limited to case reports and case series, and we determined which treatments to include in the algorithm and their recommended sequence based on their relative likelihood of success, invasiveness, and risk. Recommended therapies include: Trendelenburg positioning, inhaled epoprostenol or nitric oxide, methylene blue, embolization of abnormal pulmonary vessels, and extracorporeal life support. Availability and use of this pragmatic algorithm may improve management of this complication, and will benefit from prospective validation.

Defining and characterizing severe hypoxemia after liver transplantation in hepatopulmonary syndrome.

Hepatopulmonary syndrome is defined as a triad of liver disease, intrapulmonary vascular dilatations, and abnormal gas exchange, and it carries a poor prognosis. Liver transplantation is the only known cure for this syndrome. Severe hypoxemia in the early postoperative period has been reported to be a major complication and often leads to death in this population, but it has been poorly characterized. We sought to propose an objective definition for this complication and to describe its risk factors, incidence, and outcomes. We performed a systematic literature search and reviewed our single-center experience to characterize this complication. On the basis of the most commonly applied definition in 27 identified studies, we objectively defined severe postoperative hypoxemia as hypoxemia requiring a 100% fraction of inhaled oxygen to maintain a saturation ≥ 85% and out of proportion to any concurrent lung process. Nineteen of the 27 reports (70%) fulfilled this definition, as did 4 of the 21 patients (19%) at our center. We determined the prevalence and mortality of this complication from reports including 10 or more consecutive patients and providing sufficient postoperative details to determine whether this complication had occurred. In these reports, the prevalence of this complication was 12% (25/209). For the 11 cases with reported outcomes, the posttransplant mortality rate was 45% (5/11). There was a trend toward an increased risk of developing this complication in patients with very severe preoperative hypoxemia, defined as a partial pressure of arterial oxygen ≤ 50 mm Hg (8/41 with very severe hypoxemia versus 3/49 without severe hypoxemia, P = 0.053), and there was a significantly increased risk for patients with anatomic shunting ≥ 20% (7/25 with anatomic shunting ≥ 20% versus 1/25 without anatomic shunting ≥ 20%, P = 0.049). In conclusion, increased preoperative vigilance for this common complication is required among high-risk patients, and further research is required to identify the best management strategies.

Risk factors and monitoring for water quality to determine best management practices for splash parks.

Splash parks have been associated with infectious disease outbreaks as a result of exposure to poor water quality. To be able to protect public health, risk factors were identified that determine poor water quality. Samples were taken at seven splash parks where operators were willing to participate in the study. Higher concentrations of Escherichia coli were measured in water of splash parks filled with rainwater or surface water as compared with sites filled with tap water, independent of routine inspection intervals and employed disinfection. Management practices to prevent fecal contamination and guarantee maintaining good water quality at splash parks should include selection of source water of acceptable quality.

[Jisuikang formula promotes spinal cord injury repair in rats by activating the YAP/PKM2 signaling axis in astrocytes].

OBJECTIVE: To investigate the effect of Jisuikang formula-medicated serum for promoting spinal cord injury (SCI) repair in rats and explore the possible mechanism. METHODS: Thirty adult SD rats were randomized into sham-operated group, SCI (induced using a modified Allen method) model group, and Jisuikang formula-medicated serum treatment group. After the operations, the rats were treated with normal saline or Jisuikang by gavage on a daily basis for 14 days, and the changes in hindlimb motor function of the rats was assessed with Basso-Beattie-Bresnahan (BBB) scores and inclined-plate test. The injured spinal cord tissues were sampled from the SCI rat models for single-cell RNA sequencing, and bioinformatics analysis was performed to identify the target genes of Jisuikang, spinal cord injury and glycolysis. In the cell experiment, cultured astrocytes from neonatal SD rat cortex were treated with SOX2 alone or in combination with Jisuikang-medicated serum for 21 days, and the protein expressions of PKM2, p-PKM2 and YAP and colocalization of PKM2 and YAP in the cells were analyzed with Western blotting and immunofluorescence staining, respectively. RESULTS: The SCI rats with Jisuikang treatment showed significantly improved BBB scores and performance in inclined-plate test. At the injury site, high PKM2 expression was detected in various cell types. Bioinformatic analysis identified the HIPPO-YAP signaling pathway as the target pathway of Jisuikang. In cultured astrocytes, SOX2 combined with the mediated serum, as compared with SOX2 alone, significantly increased PKM2, p-PKM2 and YAP expressions and entry of phosphorylated PKM2 into the nucleus, and promoted PKM2 and YAP co-localization in the cells. CONCLUSION: Jisuikang formula accelerates SCI repair in rats possibly by promoting aerobic glycolysis of the astrocytes via activating the PKM2/YAP axis to induce reprogramming of the astrocytes into neurons.

Functional importance of Dicer protein in the adaptive cellular response to hypoxia.

The processes by which cells sense and respond to ambient oxygen concentration are fundamental to cell survival and function, and they commonly target gene regulatory events. To date, however, little is known about the link between the microRNA pathway and hypoxia signaling. Here, we show in vitro and in vivo that chronic hypoxia impairs Dicer (DICER1) expression and activity, resulting in global consequences on microRNA biogenesis. We show that von Hippel-Lindau-dependent down-regulation of Dicer is key to the expression and function of hypoxia-inducible factor α (HIF-α) subunits. Specifically, we show that EPAS1/HIF-2α is regulated by the Dicer-dependent microRNA miR-185, which is down-regulated by hypoxia. Full expression of hypoxia-responsive/HIF target genes in chronic hypoxia (e.g. VEGFA, FLT1/VEGFR1, KDR/VEGFR2, BNIP3L, and SLC2A1/GLUT1), the function of which is to regulate various adaptive responses to compromised oxygen availability, is also dependent on hypoxia-mediated down-regulation of Dicer function and changes in post-transcriptional gene regulation. Therefore, functional deficiency of Dicer in chronic hypoxia is relevant to both HIF-α isoforms and hypoxia-responsive/HIF target genes, especially in the vascular endothelium. These findings have relevance to emerging therapies given that we show that the efficacy of RNA interference under chronic hypoxia, but not normal oxygen availability, is Dicer-dependent. Collectively, these findings show that the down-regulation of Dicer under chronic hypoxia is an adaptive mechanism that serves to maintain the cellular hypoxic response through HIF-α- and microRNA-dependent mechanisms, thereby providing an essential mechanistic insight into the oxygen-dependent microRNA regulatory pathway.

Unlocking the potential of RNA-based therapeutics in the lung: current status and future directions.

Awareness of RNA-based therapies has increased after the widespread adoption of mRNA vaccines against SARS-CoV-2 during the COVID-19 pandemic. These mRNA vaccines had a significant impact on reducing lung disease and mortality. They highlighted the potential for rapid development of RNA-based therapies and advances in nanoparticle delivery systems. Along with the rapid advancement in RNA biology, including the description of noncoding RNAs as major products of the genome, this success presents an opportunity to highlight the potential of RNA as a therapeutic modality. Here, we review the expanding compendium of RNA-based therapies, their mechanisms of action and examples of application in the lung. The airways provide a convenient conduit for drug delivery to the lungs with decreased systemic exposure. This review will also describe other delivery methods, including local delivery to the pleura and delivery vehicles that can target the lung after systemic administration, each providing access options that are advantageous for a specific application. We present clinical trials of RNA-based therapy in lung disease and potential areas for future directions. This review aims to provide an overview that will bring together researchers and clinicians to advance this burgeoning field.

Impact of sex on outcome after pulmonary endarterectomy for chronic thromboembolic pulmonary hypertension.

BACKGROUND: The impact of sex on long-term outcomes after pulmonary endarterectomy (PEA) for chronic thromboembolic pulmonary hypertension (PH) remains unclear. We therefore examined the early and long-term outcome after PEA to determine whether sex had an impact on the risk of residual PH and need for targeted PH medical therapy. METHODS: Retrospective study of 401 consecutive patients undergoing PEA at our institution between August 2005 and March 2020 was performed. Primary outcome was the need for targeted PH medical therapy postoperatively. Secondary outcomes included survival and measures of hemodynamic improvement. RESULTS: Females (N = 203, 51%) were more likely to have preoperative home oxygen therapy (29.6% vs 11.6%, p < 0.01), and to present with segmental and subsegmental disease compared to males (49.2% vs 21.2%, p < 0.01). Despite similar preoperative values, females had higher postoperative pulmonary vascular resistance (final total pulmonary vascular resistance after PEA, 437 Dynes∙s∙cm-5 vs 324 Dynes∙s∙cm-5 in males, p < 0.01). Although survival at 10 years was not significantly different between sexes (73% in females vs 84% in males, p = 0.08), freedom from targeted PH medical therapy was lower in females (72.9% vs 89.9% in males at 5 years, p < 0.001). Female sex remained an independent factor affecting the need for targeted PH medical therapy after PEA in multivariate analysis (HR 2.03, 95%CI 1.03-3.98, p = 0.04). CONCLUSIONS: Although outcomes are excellent for both sexes, females had greater need for targeted PH medical therapy in the long-term. Early reassessment and long-term follow-up of these patients are important. Further investigations into possible mechanisms to explain the differences are warranted.

Isolation of Legionella pneumophila from pluvial floods by amoebal coculture.

Viable Legionella pneumophila bacteria were isolated by amoebal coculture from pluvial floods after intense rainfall and from water collected at sewage treatment plants. Several isolated L. pneumophila strains belonged to sequence types that have been previously identified in patients.

Optimization of subcritical water pre-treatment for biogas enhancement on co-digestion of pineapple waste and cow dung using the response surface methodology.

The optimal pre-treatment method and conditions depend on the types of lignocellulose present due to the complexity and the variability of biomass chemical structures. This study optimized subcritical water pre-treatment to ensure maximum methane production from pineapple waste prior to anaerobic co-digestion with cow dung using the response surface methodology. A central composite design was achieved with three different factors and one response. A total of 20 pre-treatment runs were performed at different temperatures, reaction times and water to solid ratios suggesting optimum values for subcritical water pre-treatment at 128.52℃ for 5 min with 5.67 to 1 water to solid ratio. Under these conditions, methane yield increased from 59.09 to 85.05 mL CH4/g VS with an increase of 23% biogas yield and 44% methane yield from the untreated. All pre-treatments above 200℃ showed reductions in biogas yield. Compositional analysis showed slight reduction of lignin and increase in α-cellulose content after the pre-treatment. Analysis using Fourier transform infrared spectroscopy and thermogravimetric analysis verified the presence of cellulosic material in pre-treated pineapple waste. Most of the hemicellulose was solubilized in the liquid samples after SCW pre-treatment. The crystallinity index of pineapple waste was reduced from 57.58% (untreated) to 54.29% (pre-treated). Scanning electron microscopy confirmed the structural modification of pre-treated pineapple waste for better microbial attack. Subcritical water pre-treatment is feasible as a promising method to enhance the anaerobic co-digestion process. Further study should be conducted to assess the scale-up of the process from pre-treatment to anaerobic digestion at the pilot plant level.