RasGRF Couples Nox4-Dependent Endoplasmic Reticulum Signaling to Ras.

OBJECTIVES: In response to endoplasmic reticulum (ER) stress, endothelial cells initiate corrective pathways such as the unfolded protein response. Recent studies suggest that reactive oxygen species produced on the ER may participate in homeostatic signaling through Ras in response to ER stress. We sought to identify mechanisms responsible for this focal signaling pathway. APPROACH AND RESULTS: In endothelial cells, we found that ER stress induced by tunicamycin activates the NADPH (nicotinamide adenine dinucleotide phosphate) oxidase Nox4 focally on the ER surface but not on the plasma membrane. Ras activation is also restricted to the ER, occurs downstream of Nox4, and is required for activation of the unfolded protein response. In contrast, treatment with the growth factor VEGF (vascular endothelial growth factor) results in Ras activation and reactive oxygen species production confined instead to the plasma membrane and not to the ER, demonstrating local coupling of reactive oxygen species and Ras signals. We further identify the calcium-responsive, ER-resident guanyl exchange factors RasGRF1 and RasGRF2 as novel upstream mediators linking Nox4 with Ras activation in response to ER stress. Oxidation of the sarcoendoplasmic reticulum calcium ATPase and increases in cytosolic calcium caused by ER stress are blocked by Nox4 knockdown, and reduction in cytosolic free calcium prevents both Ras activation and the unfolded protein response. CONCLUSIONS: ER stress triggers a localized signaling module on the ER surface involving Nox4-dependent calcium mobilization, which directs local Ras activation through ER-associated, calcium-responsive RasGRF.

The SF-36 and 6-Minute Walk Test are Significant Predictors of Complications After Major Surgery.

BACKGROUND: Major surgeries are associated with postoperative morbidity and mortality. Current preoperative evaluation fails to identify patients at increased risk of postoperative complications. This study is aimed to determine whether the Short Form-36 health survey (SF-36) and the 6-minute walk test (6-MWT) are useful predictors of postoperative complications after major surgery. METHODS: All patients scheduled to undergo major surgery were eligible for the study. Major surgeries include patients undergoing thoracotomy, sternotomy, or upper abdominal laparotomy. The SF-36 health survey and 6-MWT were administered prior to surgery. Spirometry and other preoperative testing, ordered by the surgeon, like echocardiography were included in the study. Patients were then followed-up for postoperative complications for 30 days. RESULTS: One-hundred and seventeen subjects undergoing major surgery were recruited to the study. The mean age was 58 years and 66 (56.4%) were male. Physical Functioning as a component of the SF-36 positively correlated with decreased length of hospital stay (LOS). The 6-MWT had a negative correlation with LOS (p < 0.0001) and with severity of postoperative complications (p < 0.0001). Spirometry and echocardiography did not correlate with LOS or grade of complications. CONCLUSIONS: SF-36 (Physical Functioning) and 6-MWT are useful indicators for predicting postoperative complications and LOS. Patients undergoing major surgery answered SF-36 and performed 6-MWT. Physical Functioning as a component of the SF-36 correlated with LOS. The 6-MWT had a negative correlation with LOS and with complication grade. SF-36 and 6-MWT are useful predictors of postoperative complications.

A Case Series of Secondary Spontaneous Pneumomediastinum and Pneumothorax in Severe COVID-19 Pneumonia.

Introduction Pneumomediastinum and pneumothorax are uncommon complications in COVID-19 patients. The exact prevalence, etiology, and outcomes are not well known. We report a case series of patients in our institution with COVID-19 related pneumomediastinum and pneumothorax and address these questions. Methods We conducted a single-center retrospective chart review of patients admitted at our institution with a positive polymerase chain reaction (PCR) confirming the diagnosis of COVID-19. A cohort of 500 potential study candidates was identified, of whom eight were investigated. Demographic data, hospital course, patient co-morbidities, and outcome data were collected. Results Eight patients were included in our study who were identified as having an event (i.e., pneumomediastinum and/or pneumothorax) during the specified timeframe. Overall, 62% of patients were on high-flow nasal cannula with an average FiO2 of >70%. The average oxygen saturation//fraction of inspired oxygen (SpO2/FiO2) ratio leading up to an event was 113.7286 (range: 101.11-130.66), and all of the patients not on mechanical ventilation met the criteria for acute respiratory distress syndrome (ARDS) based on the Kigali definition with SpO2/FiO2 < 315. The three patients who developed an event while requiring mechanical ventilation both had PaO2/FiO2 < 100, consistent with severe ARDS at the time of an event. The mean time in days, counted from the day of hospital admission until an event, was 10 days (range: 3-23 days). None of the cases had documented pulmonary parenchymal disease prior to developing COVID-19. To the best of our knowledge, these events were not iatrogenic in nature. Conclusion Secondary spontaneous pneumomediastinum and pneumothorax are rare albeit well-documented phenomena in hospitalized patients with COVID-19 infection. Interestingly, the majority of patients in our study were on high-flow nasal cannula at the time of an event. The majority of previously published data on this topic are on those who required positive pressure ventilation; however, there have been more recent papers that also describe these events in non-mechanically ventilated patients. The exact pathophysiology remains unknown, but it is likely multifactorial, and additional studies are needed to further evaluate this phenomenon.

Mechanisms of lung endothelial barrier disruption induced by cigarette smoke: role of oxidative stress and ceramides.

The epithelial and endothelial cells lining the alveolus form a barrier essential for the preservation of the lung respiratory function, which is, however, vulnerable to excessive oxidative, inflammatory, and apoptotic insults. Whereas profound breaches in this barrier function cause pulmonary edema, more subtle changes may contribute to inflammation. The mechanisms by which cigarette smoke (CS) exposure induce lung inflammation are not fully understood, but an early alteration in the epithelial barrier function has been documented. We sought to investigate the occurrence and mechanisms by which soluble components of mainstream CS disrupt the lung endothelial cell barrier function. Using cultured primary rat microvascular cell monolayers, we report that CS induces endothelial cell barrier disruption in a dose- and time-dependent manner of similar magnitude to that of the epithelial cell barrier. CS exposure triggered a mechanism of neutral sphingomyelinase-mediated ceramide upregulation and p38 MAPK and JNK activation that were oxidative stress dependent and that, along with Rho kinase activation, mediated the endothelial barrier dysfunction. The morphological changes in endothelial cell monolayers induced by CS included actin cytoskeletal rearrangement, junctional protein zonula occludens-1 loss, and intercellular gap formation, which were abolished by the glutathione modulator N-acetylcysteine and ameliorated by neutral sphingomyelinase inhibition. The direct application of ceramide recapitulated the effects of CS, by disrupting both endothelial and epithelial cells barrier, by a mechanism that was redox and apoptosis independent and required Rho kinase activation. Furthermore, ceramide induced dose-dependent alterations of alveolar microcirculatory barrier in vivo, measured by two-photon excitation microscopy in the intact rat. In conclusion, soluble components of CS have direct endothelial barrier-disruptive effects that could be ameliorated by glutathione modulators or by inhibitors of neutral sphingomyelinase, p38 MAPK, JNK, and Rho kinase. Amelioration of endothelial permeability may alleviate lung and systemic vascular dysfunction associated with smoking-related chronic obstructive lung diseases.

Hypertriglyceridemia and massive rhabdomyolysis in a patient with disseminated legionella.

Legionella most commonly presents as pneumonia but can have disseminated involvement, presenting as extra-pulmonary disease involving gastrointestinal, neurological, cardiac, renal, and musculoskeletal systems, and skin and soft tissues. We present a case of a patient with pneumonia, rhabdomyolysis, renal failure, hypertriglyceridemia, pancreatitis, and cutaneous involvement. This case highlights the breath of involvement legionella can have, including the never previously documented manifestation of hypertriglyceridemia and severe rhabdomyolysis with the highest creatinine kinase recorded.

Malignant Müllerian Adenocarcinoma Manifesting With Cardiac Tamponade and Pleural Effusion.

A 54-year-old woman with a past medical history of untreated stage IV Müllerian adenocarcinoma presented for dyspnea. She was found to have a large right-sided pleural effusion through basic radiology and clinically improved after a CT-guided therapeutic thoracocentesis. However, the patient rapidly deteriorated shortly afterward. A broader workup that included echocardiography revealed a large pericardial effusion with tamponade physiology. The patient underwent an emergent pericardiocentesis, which briefly improved hemodynamics, but her clinical status kept declining until she eventually expired. Subsequent cytology of the pleural and pericardial fluid revealed malignant cells of Müllerian origin.

Activated protein C attenuates acute lung injury and apoptosis in a hyperoxic animal model.

Evidence suggests that activated protein C (APC) attenuates acute lung injury (ALI) through antithrombotic and anti-inflammatory mechanisms. The aim of this study was to determine the effects of APC on ALI in adult rats exposed to hyperoxic environment. Rats were divided into control, hyperoxia, hyperoxia + APC, and APC. Hyperoxia and hyperoxia + APC were exposed to 1, 3, and 5 days of hyperoxia. Hyperoxia + APC and APC were injected with APC (5 mg/kg, i.p.) every 12 h. Control and hyperoxia received isotonic sodium chloride solution injection. Measurement of wet to dry ratio and albumin leak demonstrated significant improvement in hyperoxia + APC when compared with hyperoxia. Apoptosis, as measured by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, was significantly reduced in hyperoxia + APC when compared with hyperoxia. Histological evaluation of lung sections showed significant reduction in inflammation, edema, and in the number of marginating neutrophils in hyperoxia + APC as compared with hyperoxia. Transcriptional expression of lung inflammatory mediators demonstrated a time-dependent surge in the levels TNF-alpha, IL-1beta, and IL-6 in response to hyperoxia that was attenuated with APC administration in the presence of hyperoxia. In this rat model, APC attenuates lung injury and the expression of inflammatory mediators in ALI secondary to hyperoxia.