Circulating markers of inflammation and angiogenesis and clinical outcomes across subtypes of pulmonary arterial hypertension.

BACKGROUND: Subtypes of pulmonary arterial hypertension (PAH) differ in both fundamental disease features and clinical outcomes. Angiogenesis and inflammation represent disease features that may differ across subtypes and are of special interest in connective tissue disease-associated PAH (CTD-PAH). We compared inflammatory and angiogenic biomarker profiles across different etiologies of PAH and related them to clinical outcomes. METHODS: Participants with idiopathic PAH, CTD-PAH, toxin-associated PAH (tox-PAH), or congenital heart disease-associated PAH (CHD-PAH) were enrolled into a prospective observational cohort. Baseline serum concentrations of 33 biomarkers were related to 3-year mortality, echocardiogram, REVEAL score, and 6-minute walk distance (6MWD). Findings were validated using plasma proteomic data from the UK PAH Cohort Study. RESULTS: One hundred twelve patients were enrolled: 45 idiopathic, 27 CTD-PAH, 20 tox-PAH, and 20 CHD-PAH. Angiogenic and inflammatory biomarkers were distinctly elevated within the CTD-PAH cohort. Six biomarkers were associated with mortality within the entire PAH cohort: interleukin-6 (IL-6, HR:1.6, 95% CI:1.18-2.18), soluble fms-like tyrosine kinase 1 (sFlt-1, HR:1.35, 95% CI:1.02-1.80), placental growth factor (PlGF, HR:1.55, 95% CI:1.07-2.25), interferon gamma-induced protein 10 (IP-10, HR:1.44, 95% CI:1.04-1.99), tumor necrosis factor-beta (TNF-ß, HR:1.81, 95% CI:1.11-2.95), and NT-proBNP (HR:2.19, 95% CI:1.52-3.14). Only IL-6 and NT-proBNP remained significant after controlling for multiple comparisons. IL-6, IP-10, and sFlt-1 significantly associated with mortality in CTD-PAH, but not non-CTD-PAH subgroups. In the UK cohort, IP-10, PlGF, TNF-ß, and NT-proBNP significantly associated with 5-year survival. CONCLUSION: Levels of angiogenic and inflammatory biomarkers are elevated in CTD-PAH, compared with other etiologies of PAH, and may correlate with clinical outcomes including mortality.

Spontaneous coronary artery dissection and exogenous estrogen in a transgender female.

We report the case of a 37-year-old transgender patient with a history of orchiectomy on gender-affirming estrogen therapy who was hospitalized with an acute onset of chest pain with a resolution shortly after the presentation. On presentation, the patient had a rapid rise in troponin level and was urgently taken to the cardiac catheterization lab where spontaneous coronary artery dissection was diagnosed and treated with cutting balloon angioplasty and medical management. After a multidisciplinary discussion, the only notable risk factor was estrogen supplementation after gender reassignment surgery. To our knowledge, this is the only report of a male to female transgender patient reported to have spontaneous coronary artery dissection.

The Predictive Potential of Elevated Serum Inflammatory Markers in Determining the Need for Intubation in CoVID-19 Patients.

Introduction: The predictive potential of demographics, clinical characteristics, and inflammatory markers at admission to determine future intubation needs of hospitalised CoVID-19 patients is unknown. The study aimed to determine the predictive potential of elevated serum inflammatory markers in determining the need for intubation in CoVID-19 Patients. Methods: In a retrospective cohort study of hospitalised SARS-CoV2 positive patients, single and multivariable regression analyses were used to determine covariate effects on intubation odds, and a minimax concave penalty regularised logistic regression was used to build a predictive model. A second prospective independent cohort tested the model. Results: Systemic inflammatory markers obtained at admission were higher in patients that required subsequent intubation, and adjusted odds of intubation increased for every standard deviation above the mean for c-reactive protein (CRP) OR:2.8 (95% CI 1.8-4.5, p<0.001) and lactate dehydrogenase OR:2.1 (95% CI 1.33.3, p=0.002). A predictive model incorporating C-reactive protein, lactate dehydrogenase, and diabetes status at the time of admission predicted intubation status with an area under the curve (AUC) of 0.78 with corresponding sensitivity of 86%, specificity of 63%. This predictive model achieved an AUC of 0.83, 91% sensitivity, and 41% specificity on the validation cohort. Conclusion: In patients hospitalised with CoVID-19, elevated serum inflammatory markers measured within the first twenty-four hours of admission are associated with an increased need for intubation. Additionally, a model of C-reactive protein, lactate dehydrogenase, and the presence of diabetes may play a predictive role in determining the future need for intubation.

Progressive Pulmonary Fibrosis After Non-Critical COVID-19: A Case Report.

BACKGROUND The COVID-19 global pandemic is ongoing, and despite vaccination efforts, SARS-CoV-2 continues to circulate worldwide. The spectrum of COVID-19 illness is broad, from asymptomatic infection to respiratory failure and acute respiratory distress syndrome (ARDS), and the long-term sequelae of infection are unclear. COVID-19-related pulmonary fibrosis has been previously described in the setting of critical illness and ARDS but has not been well described in cases requiring minimal supplemental oxygen. CASE REPORT We present the case of a 42-year-old man hospitalized with coronavirus disease 2019 (COVID-19) who initially required minimal supplemental oxygen but weeks later developed progressive pulmonary fibrosis requiring high-flow nasal cannula and ICU admission. Using novel computed tomography (CT) imaging processing techniques, we demonstrate progression from initial ground-glass opacities to pulmonary fibrosis and traction bronchiectasis over several months. Additionally, we describe clinical responsiveness to an extended course of corticosteroids. CONCLUSIONS Although pulmonary fibrosis is a known complication of severe COVID-19-related ARDS requiring mechanical ventilation, our report suggests that patients with milder forms of COVID-19 infection may develop post-acute pulmonary fibrosis.

Perinatal Hypoxia-Inducible Factor Stabilization Preserves Lung Alveolar and Vascular Growth in Experimental Bronchopulmonary Dysplasia.

Rationale: Antenatal inflammation with placental dysfunction is strongly associated with high bronchopulmonary dysplasia (BPD) risk in preterm infants. Whether antenatal or postnatal HIF (hypoxia-inducible factor) augmentation can preserve lung structure and function and prevent pulmonary hypertension after intrauterine inflammation is controversial.Objectives: To determine whether antenatal or postnatal prolyl-hydroxylase inhibitor (PHi) therapy increases lung HIF expression, preserves lung growth and function, and prevents pulmonary hypertension in a rat model of chorioamnionitis-induced BPD caused by antenatal inflammation.Methods: Endotoxin (ETX) was administered to pregnant rats by intraamniotic injection at Embryonic Day 20, and pups were delivered by cesarean section at Embryonic Day 22. Selective PHi drugs, dimethyloxalylglycine or GSK360A, were administered into the amniotic space at Embryonic Day 20 or after birth by intraperitoneal injection for 2 weeks. Placentas and lung tissue were collected at birth for morphometric and Western blot measurements of HIF-1a, HIF-2a, VEGF (vascular endothelial growth factor), and eNOS (endothelial nitric oxide synthase) protein contents. At Day 14, lung function was assessed, and tissues were harvested to determine alveolarization by radial alveolar counts, pulmonary vessel density, and right ventricle hypertrophy (RVH).Measurements and Main Results: Antenatal PHi therapy preserves lung alveolar and vascular growth and lung function and prevents RVH after intrauterine ETX exposure. Antenatal administration of PHi markedly upregulates lung HIF-1a, HIF-2a, VEGF, and eNOS expression after ETX exposure.Conclusions: HIF augmentation improves lung structure and function, prevents RVH, and improves placental structure following antenatal ETX exposure. We speculate that antenatal or postnatal PHi therapy may provide novel strategies to prevent BPD due to antenatal inflammation.

Combined EphB2 receptor knockdown with radiation decreases cell viability and invasion in medulloblastoma.

BACKGROUND: Medulloblastoma is one of the most common types of pediatric brain tumor characterized by the subpopulation of cells that exhibit high invasive potential and radioresistant properties. In addition, dysregulated function and signaling by Eph family of receptors have been shown to impart pro-tumorigenic characteristics in this brain malignancy. In the current study, we investigated whether EphB2 knockdown in combination with radiation can alter invasiveness and decrease medulloblastoma tumor growth or viability in vitro. METHODS: The expression of EphB2 receptor was analyzed by immunohistochemistry and Western blotting. Microarray analysis and mRNA analysis was performed on medulloblastoma patient datasets and compared to the normal cerebellum. The radiosensitization effect following EphB2 knockdown was determined by clonogenic assay in human medulloblastoma cells. Effects of EphB2-siRNA in absence or presence of radiation on cell cycle distribution, cell viability, and invasion were analyzed by flow cytometry, MTT assay, trypan blue exclusion assay, xcelligence system, and Western blotting. RESULTS: We observed that EphB2 is expressed in both medulloblastoma cell lines and patient samples and its downregulation sensitized these cells to radiation as evident by decreased clonogenic survival fractions. EphB2 expression was also high across different medulloblastoma subgroups compared to normal cerebellum. The radiosensitization effect observed following EphB2 knockdown was in part mediated by enhanced G2/M cell cycle arrest. We also found that the combined approach of EphB2 knockdown and radiation exposure significantly reduced overall cell viability in medulloblastoma cells compared to control groups. Similar results were obtained in the xcelligence-based invasion assay. Western blot analysis also demonstrated changes in the protein expression of cell proliferation, cell survival, and invasion molecules in the combination group versus others. CONCLUSIONS: Overall, our findings indicate that specific targeting of EphB2 receptor in combination with radiation may serve as an effective therapeutic strategy in medulloblastoma. Future studies are warranted to test the efficacy of this approach in in vivo preclinical models.

Ephrin-B2 overexpression predicts for poor prognosis and response to therapy in solid tumors.

Ephrin B2 is variably expressed on tumor cells and its blockade has been shown to inhibit angiogenesis in animal models of pancreatic, colorectal, lung and head, and neck squamous cell carcinomas. However, the implications of ephrinB2 expression in cancer patients have remained elusive. In this study, we analyzed the cancer genome atlas (TCGA) for ephrinB2 expression. We report significant correlations between EFNB2 expression, overall survival and disease-free survival in head and neck squamous cell carcinoma (HNSCC, n = 519), pancreatic adenocarcinoma (n = 186), and bladder urothelial carcinoma (n = 410). In HNSCC patients, high-EFNB2 mRNA expression was associated with tumor HPV negativity, oral cavity location, alcohol intake, higher TP53 mutation, and EGFR amplification. EphrinB2 overexpression also correlated with worse response to chemotherapy and radiotherapy. The therapeutic potential of blocking ephrinB2 was validated in HNSCC patient-derived tumor xenografts and showed significant improvement in survival and tumor growth delay. Our data shows that ephrinB2 overexpression can serve as a critical biomarker for patient prognosis and response to therapy. These results should guide design of future clinical trials exploring EphrinB2 inhibition in cancer patients. © 2016 Wiley Periodicals, Inc.

Enhancing radiosensitization in EphB4 receptor-expressing Head and Neck Squamous Cell Carcinomas.

Members of the Eph family of receptor tyrosine kinases have been implicated in a wide array of human cancers. The EphB4 receptor is ubiquitously expressed in head and neck squamous cell carcinoma (HNSCC) and has been shown to impart tumorigenic and invasive characteristics to these cancers. In this study, we investigated whether EphB4 receptor targeting can enhance the radiosensitization of HNSCC. Our data show that EphB4 is expressed at high to moderate levels in HNSCC cell lines and patient-derived xenograft (PDX) tumors. We observed decreased survival fractions in HNSCC cells following EphB4 knockdown in clonogenic assays. An enhanced G2 cell cycle arrest with activation of DNA damage response pathway and increased apoptosis was evident in HNSCC cells following combined EphB4 downregulation and radiation compared to EphB4 knockdown and radiation alone. Data using HNSCC PDX models showed significant reduction in tumor volume and enhanced delay in tumor regrowth following sEphB4-HSA administration with radiation compared to single agent treatment. sEphB4-HSA is a protein known to block the interaction between the EphB4 receptor and its ephrin-B2 ligand. Overall, our findings emphasize the therapeutic relevance of EphB4 targeting as a radiosensitizer that can be exploited for the treatment of human head and neck carcinomas.

Effects of altered ephrin-A5 and EphA4/EphA7 expression on tumor growth in a medulloblastoma mouse model.

BACKGROUND: Members of the Eph/ephrin gene families act as key regulators of cerebellar development during embryogenesis. Aberrant signaling of Eph family of receptor tyrosine kinases and their ephrin ligands has also been implicated in human cancers. Medulloblastoma is an aggressive primitive neuroectodermal tumor that originates from granule neuron precursors in the cerebellum. Previous studies have suggested a role for the ephrin-A5 ligand and its receptors, EphA4 and EphA7, in granule cell-precursor formation and in guiding cell migration. In the present study, we investigated the effects of genetic loss of ephrin-A5, EphA4, and EphA7 on the spatiotemporal development of medulloblastoma tumors in the context of the smoothened transgenic mouse model system. FINDINGS: Radiographic magnetic resonance imaging (MRI) was performed to monitor tumor growth in a genetically engineered mouse model of medulloblastoma. Tumor tissue was harvested to determine changes in the expression of phosphorylated Akt by Western blotting. This helped to establish a correlation between genotype and/or tumor size and survival. Our in vivo data establish that in ND2-SmoA1 transgenic mice, the homozygous deletion of ephrin-A5 resulted in a consistent pattern of tumor growth inhibition compared to their ephrin-A5 wild-type littermate controls, while the loss of EphA4/EphA7 failed to produce consistent effects versus EphA4/EphA7 wild-type mice. A positive correlation was evident between tumor size, p-Akt, and proliferating cell nuclear antigen (PCNA) expression in our transgenic mouse model system, regardless of genotype. CONCLUSIONS: Taken together, our findings underscore the importance of targeting specific members of the Eph/ephrin families in conjunction with the Akt pathway in order to inhibit medulloblastoma tumor growth and progression.

Knockdown of EphB1 receptor decreases medulloblastoma cell growth and migration and increases cellular radiosensitization.

The expression of members of the Eph family of receptor tyrosine kinases and their ephrin ligands is frequently dysregulated in medulloblastomas. We assessed the expression and functional role of EphB1 in medulloblastoma cell lines and engineered mouse models. mRNA and protein expression profiling showed expression of EphB1 receptor in the human medulloblastoma cell lines DAOY and UW228. EphB1 downregulation reduced cell growth and viability, decreased the expression of important cell cycle regulators, and increased the percentage of cells in G1 phase of the cell cycle. It also modulated the expression of proliferation, and cell survival markers. In addition, EphB1 knockdown in DAOY cells resulted in significant decrease in migration, which correlated with decreased ß1-integrin expression and levels of phosphorylated Src. Furthermore, EphB1 knockdown enhanced cellular radiosensitization of medulloblastoma cells in culture and in a genetically engineered mouse medulloblastoma model. Using genetically engineered mouse models, we established that genetic loss of EphB1 resulted in a significant delay in tumor recurrence following irradiation compared to EphB1-expressing control tumors. Taken together, our findings establish that EphB1 plays a key role in medulloblastoma cell growth, viability, migration, and radiation sensitivity, making EphB1 a promising therapeutic target.