Hypoxia is a prominent feature of malignant tumors that are characterized by angiogenesis and vascular hyperpermeability. Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) ...has been shown to be up-regulated in the vicinity of necrotic tumor areas, and hypoxia potently induces VPF/VEGF expression in several tumor cell lines in vitro. Here we report that hypoxia-induced VPF/VEGF expression is mediated by increased transcription and mRNA stability in human M21 melanoma cells. RNA-binding/electrophoretic mobility shift assays identified a single 125-bp AU-rich element in the 3' untranslated region that formed hypoxia-inducible RNA-protein complexes. Hypoxia-induced expression of chimeric luciferase reporter constructs containing this 125-bp AU-rich hypoxia stability region were significantly higher than constructs containing an adjacent 3' untranslated region element without RNA-binding activity. Using UV-cross-linking studies, we have identified a series of hypoxia-induced proteins of 90/88 kDa, 72 kDa, 60 kDa, 56 kDa, and 46 kDa that bound to the hypoxia stability region element. The 90/88-kDa and 60-kDa species were specifically competed by excess hypoxia stability region RNA. Thus, increased VPF/VEGF mRNA stability induced by hypoxia is mediated, at least in part, by specific interactions between a defined mRNA stability sequence in the 3' untranslated region and distinct mRNA-binding proteins in human tumor cells.
Angiogenesis is a key component of human cancer progression and metastasis. In an effort to recapitulate early events in tumor-induced angiogenesis, we have employed a subcutaneous Matrigel implant ...model using immunodeficient mice as hosts. Matrigel-containing fibroblast growth factor 2 (FGF-2; 1.2 μg/ml) induced stromal cell infiltration into the Matrigel/skin interface within 4 days and maximal neovascularization at 7 days. Cells staining positive for the endothelial cell marker, platelet-endothelial cell adhesion molecule 1 (PECAM-1), were present in neovessels and in isolated cells within the Matrigel matrix. Immunohistochemical analysis revealed high levels of vascular endothelial growth factor (VEGF) deposited in the stromal interface present only in the FGF-2–containing but not in control Matrigel implants. VEGF expression was confirmed with in situ hybridization. High VEGF mRNA levels were observed in the infiltrating stromal cells but not in endothelial or endothelial precursors as defined by PECAM-1 staining. In vitro analysis of FGF-2–treated embryonic fibroblasts, Balb/c 3T3 cells, showed an induction of VEGF transcription, mRNA synthesis, and protein secretion as defined by transcriptional reporter, Northern blot, and ELISA assays. The FGF-2–induced VEGF expression was not dependent on select matrix adherence or signaling components because VEGF mRNA expression induced by FGF-2 was equally activated on serum, basement membrane, and fibronectin matrix substrates. Systemic application of anti-VEGF antibodies significantly repressed FGF-2–induced angiogenesis over control antibody by 88% (p < 0.001). These data support an FGF-2 angiogenic model that is dependent on endothelial cell activation, stromal cell infiltration, and VEGF expression by the infiltrating stromal cell population.
Aberrant expression of the potent angiogenic cytokine, vascular endothelial growth factor (VEGF), has been demonstrated to be associated with most human solid tumors. Both transcriptional and ...post-transcriptional mechanisms have been shown to modulate VEGF expression in a multitude of cell types. Here we report that when protein kinase C (PKC) pathways were activated in human glioblastoma U373 cells by phorbol 12-myristate 13-acetate (PMA), VEGF mRNA expression was up-regulated via a post-transcriptional mRNA stabilization mechanism. PMA treatment exhibited no increase in VEGF-specific transcriptional activation as determined by run-off transcription assays and VEGF promoter-luciferase reporter assays. However, PMA increased VEGF mRNA half-life from 0.8 to 3.6 h which was blocked by PKC inhibitors but not by protein kinase A or cyclic nucleotide-dependent protein kinase inhibitors. When U373 cells were transfected with antisense oligonucleotide sequences to the translation start sites of PKC-α, -β, -γ, -δ, -ε, or -ζ isoforms, both PKC-α and -ζ antisense oligonucleotides showed substantial inhibition of PMA-induced VEGF mRNA. In addition, overexpression of PKC-ζ resulted in a strong constitutive up-regulation of VEGF mRNA expression. This study demonstrates for the first time that specific PKC isoforms regulate VEGF mRNA expression through post-transcriptional mechanisms.
Nicotinamide (
2) is a potent and selective inhibitor of the PDE4D isozyme and as a chemical tool selectively blocks eosinophil mediator release and chemotaxis thus linking the role of PDE4D to ...eosinophil function.
Nicotinamide (
2) is a potent and selective inhibitor of the PDE4D isozyme and as a chemical tool selectively blocks eosinophil mediator release and chemotaxis thus linking the role of PDE4D to eosinophil function.
Background Extracorporeal membrane oxygenation support (ECMO) typically requires multiple blood transfusions and is associated with frequent bleeding complications. Blood transfusions are known to ...increase morbidity and mortality in critically ill patients, which may extend to patients receiving ECMO. Aiming to reduce transfusion requirements, we implemented a blood conservation protocol in adults with severe acute respiratory distress syndrome (ARDS) receiving ECMO. Methods This was a retrospective study of adults receiving ECMO for ARDS after initiation of a blood conservation protocol that included a transfusion trigger of hemoglobin of less than 7.0 g/dL, use of low-dose anticoagulation targeting an activated partial thromboplastin time of 40 to 60 seconds, and autotransfusion of circuit blood during decannulation. The primary objective was to evaluate transfusion requirements during ECMO support. Clinical outcomes included survival, neurologic function, renal function, bleeding, and thrombotic complications. Results The analysis included 38 patients; of these, 24 (63.2%) received a transfusion while receiving ECMO. Median hemoglobin was 8.29 g/dL. A median of 1.0 units (range, 250 to 300 mL) was transfused during ECMO support over a median duration of 9.0 days, equivalent to 0.11 U/d (range, 27.5 to 33.3 mL/d). The median activated partial thromboplastin time was 46.5 seconds. Bleeding occurred in 10 patients (26.3%); severe bleeding occurred in 2 patients (5.3%). Twenty-eight patients (73.7%) survived to hospital discharge. Conclusions Implementation of a blood conservation protocol in adults receiving ECMO for ARDS resulted in lower transfusion requirements and bleeding complications than previously reported in the literature and was associated with comparable survival and organ recovery.
Extracorporeal membrane oxygenation (ECMO) can serve as a bridge to recovery in cases of acute reversible illness, a bridge to transplantation in circumstances of irreversible cardiac or respiratory ...failure, a bridge to ventricular assist device therapy in select cases of cardiac failure, or a bridge to decision when the prognosis remains uncertain. Recent advances in ECMO technology that allow for prolonged support with decreased complications, the development of mobile ECMO teams, the rapidity of initiation, and the growing body of evidence, much of which remains controversial, have led to a significant increase in the use of ECMO worldwide. This increasing use of a technology that is not a destination device in itself introduces many ethical dilemmas specific to this technology. In this article, we explore some of the ethical issues inherent in the decisions surrounding the initiation and withdrawal of ECMO by raising key questions and providing a framework for clinicians. We will address extracorporeal cardiopulmonary resuscitation, the inability to bridge a patient to transplant or recovery—the so-called “bridge to nowhere”—and the significance of resuscitation preferences in the setting of continual extracorporeal circulatory support.
The EGFR/PI3K/PTEN/Akt/mTORC1/GSK-3 pathway plays prominent roles in malignant transformation, prevention of apoptosis, drug resistance and metastasis. The expression of this pathway is frequently ...altered in breast cancer due to mutations at or aberrant expression of: HER2, ERalpha, BRCA1, BRCA2, EGFR1, PIK3CA, PTEN, TP53, RB as well as other oncogenes and tumor suppressor genes. In some breast cancer cases, mutations at certain components of this pathway (e.g., PIK3CA) are associated with a better prognosis than breast cancers lacking these mutations. The expression of this pathway and upstream HER2 has been associated with breast cancer initiating cells (CICs) and in some cases resistance to treatment. The anti-diabetes drug metformin can suppress the growth of breast CICs and herceptin-resistant HER2+ cells. This review will discuss the importance of the EGFR/PI3K/PTEN/Akt/mTORC1/GSK-3 pathway primarily in breast cancer but will also include relevant examples from other cancer types. The targeting of this pathway will be discussed as well as clinical trials with novel small molecule inhibitors. The targeting of the hormone receptor, HER2 and EGFR1 in breast cancer will be reviewed in association with suppression of the EGFR/PI3K/PTEN/Akt/mTORC1/GSK-3 pathway.
Two distinct subphenotypes have been identified in acute respiratory distress syndrome (ARDS), but the presence of subgroups in ARDS associated with coronavirus disease (COVID-19) is unknown.
To ...identify clinically relevant, novel subgroups in COVID-19-related ARDS and compare them with previously described ARDS subphenotypes.
Eligible participants were adults with COVID-19 and ARDS at Columbia University Irving Medical Center. Latent class analysis was used to identify subgroups with baseline clinical, respiratory, and laboratory data serving as partitioning variables. A previously developed machine learning model was used to classify patients as the hypoinflammatory and hyperinflammatory subphenotypes. Baseline characteristics and clinical outcomes were compared between subgroups. Heterogeneity of treatment effect for corticosteroid use in subgroups was tested.
From March 2, 2020, to April 30, 2020, 483 patients with COVID-19-related ARDS met study criteria. A two-class latent class analysis model best fit the population (
= 0.0075). Class 2 (23%) had higher proinflammatory markers, troponin, creatinine, and lactate, lower bicarbonate, and lower blood pressure than class 1 (77%). Ninety-day mortality was higher in class 2 versus class 1 (75% vs. 48%;
< 0.0001). Considerable overlap was observed between these subgroups and ARDS subphenotypes. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RT-PCR cycle threshold was associated with mortality in the hypoinflammatory but not the hyperinflammatory phenotype. Heterogeneity of treatment effect to corticosteroids was observed (
= 0.0295), with improved mortality in the hyperinflammatory phenotype and worse mortality in the hypoinflammatory phenotype, with the caveat that corticosteroid treatment was not randomized.
We identified two COVID-19-related ARDS subgroups with differential outcomes, similar to previously described ARDS subphenotypes. SARS-CoV-2 PCR cycle threshold had differential value for predicting mortality in the subphenotypes. The subphenotypes had differential treatment responses to corticosteroids.
Acute exacerbations of chronic obstructive pulmonary disease (COPD) requiring invasive mechanical ventilation (IMV) are associated with significant morbidity and mortality. Extracorporeal carbon ...dioxide removal (ECCO₂R) may facilitate extubation and ambulation in these patients and potentially improve outcomes.
We assessed the feasibility of achieving early extubation and ambulation in subjects requiring IMV for exacerbations of COPD using single-site ECCO₂R.
Five subjects with exacerbations of COPD with uncompensated hypercapnia requiring IMV were enrolled in this single-center, prospective, feasibility trial using a protocol of ECCO₂R, extubation, and physical rehabilitation. The primary endpoint was extubation within 72 hours of starting ECCO₂R.
Mean preintubation pH and PaCO₂ were 7.23 ± 0.05 and 81.6 ± 15.9 mm Hg, respectively. All subjects met the primary endpoint (median duration, 4 h; range, 1.5-21.5 h). Mean duration of extracorporeal support was 193.0 ± 76.5 hours. Mean time to ambulation after extracorporeal initiation was 29.4 ± 12.6 hours. Mean maximal ambulation on extracorporeal support was 302 feet (range, 70-600). Four subjects were discharged home, and one underwent planned lung transplantation. Two minor bleeding complications occurred. There were no complications from mobilization on extracorporeal support.
ECCO₂R facilitates early extubation and ambulation in exacerbations of COPD requiring IMV and has the potential to serve as a new paradigm for the management of a select group of patients. Rigorous clinical trials are needed to corroborate these results and to investigate the effect on long-term outcomes and cost effectiveness over conventional management.
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