Ferrostatin-1 specifically targets mitochondrial iron-sulfur clusters and aconitase to improve cardiac function in Sirtuin 3 cardiomyocyte knockout mice.

AIMS: Ferroptosis is a form of iron-regulated cell death implicated in ischemic heart disease. Our previous study revealed that Sirtuin 3 (SIRT3) is associated with ferroptosis and cardiac fibrosis. In this study, we tested whether the knockout of SIRT3 in cardiomyocytes (SIRT3cKO) promotes mitochondrial ferroptosis and whether the blockade of ferroptosis would ameliorate mitochondrial dysfunction. METHODS AND RESULTS: Mitochondrial and cytosolic fractions were isolated from the ventricles of mice. Cytosolic and mitochondrial ferroptosis were analyzed by comparison to SIRT3loxp mice. An echocardiography study showed that SIRT3cKO mice developed heart failure as evidenced by a reduction of EF% and FS% compared to SIRT3loxp mice. Comparison of mitochondrial and cytosolic fractions of SIRT3cKO and SIRT3loxp mice revealed that, upon loss of SIRT3, mitochondrial, but not cytosolic, total lysine acetylation was significantly increased. Similarly, acetylated p53 was significantly upregulated only in the mitochondria. These data demonstrate that SIRT3 is the primary mitochondrial deacetylase. Most importantly, loss of SIRT3 resulted in significant reductions of frataxin, aconitase, and glutathione peroxidase 4 (GPX4) in the mitochondria. This was accompanied by a significant increase in levels of mitochondrial 4-hydroxynonenal. Treatment of SIRT3cKO mice with the ferroptosis inhibitor ferrostatin-1 (Fer-1) for 14 days significantly improved preexisting heart failure. Mechanistically, Fer-1 treatment significantly increased GPX4 and aconitase expression/activity, increased mitochondrial iron­sulfur clusters, and improved mitochondrial membrane potential and Complex IV activity. CONCLUSIONS: Inhibition of ferroptosis ameliorated cardiac dysfunction by specifically targeting mitochondrial aconitase and iron­sulfur clusters. Blockade of mitochondrial ferroptosis may be a novel therapeutic target for mitochondrial cardiomyopathies.

Requesting air ambulance transport of patients with suspected appendicitis: The decision-making process through the eyes of the rural clinician.

OBJECTIVE: The primary aim is to explore rural clinicians' self-reported knowledge, skills and attitudes in the decision-making process for requesting aeromedical retrieval of patients with suspected appendicitis. A secondary aim is to understand the supports and barriers of rural clinicians experience in this clinical scenario. SETTING: Clinician interviews conducted face-to-face in three rural hospitals in Central Queensland. PARTICIPANTS: Rural doctors and nurses. DESIGN: A five-part qualitative content analysis. RESULTS: The majority of 44 participants identified the strong and effective teamwork. The decision to request aeromedical retrieval was a shared, joint process and identified a supportive collegial culture which supported the asking of questions and not expecting to have all the answers. Perceived barriers were lack of receiving clinicians understanding of transfer agreements, and data connectivity. Clinician pessimism was identified for perceived patient outcomes. DISCUSSION: Effective teamwork can nurture trust and collaboration across multiple health service roles. High job satisfaction may counter the physical isolation in some rural environments. Fragmentation of care is the unintended consequence of interhospital transfer and may impact rural clinicians' perception of patients' outcomes and hinder receiving clinicians' understanding of rural service limitations. CONCLUSION: Future work in the area of linked electronic medical records could remove a barrier for rural clinicians and improve their reflective practice by challenging their perception of definitive patient outcomes. Increased awareness by receiving clinicians of the limitation of rural services, may minimize communication barriers and thereby, improve timely patient care transfers.

Physician Burnout and Ethics Committees.

This article provides a brief background of key issues in physician burnout, a significant problem in the healthcare industry. The extent and severity of burnout are not well understood; and those seeking help are often stigmatized. A number of different approaches to alleviating burnout have been suggested, but the problem lacks any single or simple solution. We posit that an ethics committee may be well positioned to help address this issue because of its unique position within an institution. An ethics committee serves the entire hospital staff regardless of department. As such it may be able to identify common elements in the development of burnout, and can serve as a conduit to administration in identifying these. An ethics committee can obtain information about the extent of burnout by conducting surveys to assess the extent and severity of burnout in aninstitution, and serve as a central resource to help address and alleviate it. Finally, an ethics committee may be able to act as an intermediary between practitioners and the administration, in advising the administration of the extent of the problem and offer suggestions for alleviating it.

Computed Tomography Colonography vs Colonoscopy for Colorectal Cancer Surveillance After Surgery.

BACKGROUND & AIMS: Recommendations for surveillance after curative surgery for colorectal cancer (CRC) include a 1-year post-resection abdominal-pelvic computed tomography (CT) scan and optical colonoscopy (OC). CT colonography (CTC), when used in CRC screening, effectively identifies colorectal polyps ≥10 mm and cancers. We performed a prospective study to determine whether CTC, concurrent with CT, could substitute for OC in CRC surveillance. METHODS: Our study enrolled 231 patients with resected stage 0-III CRC, identified at 5 tertiary care academic centers. Approximately 1 year after surgery, participants underwent outpatient CTC plus CT, followed by same-day OC. CTC results were revealed after endoscopic visualization of sequential colonic segments, which were re-examined for discordant findings. The primary outcome was performance of CTC in the detection of colorectal adenomas and cancers using endoscopy as the reference standard. RESULTS: Of the 231 participants, 116 (50.2%) had polyps of any size or histology identified by OC, and 15.6% had conventional adenomas and/or serrated polyps ≥6 mm. No intra-luminal cancers were detected. CTC detected patients with polyps of ≥6 mm with 44.0% sensitivity (95% CI, 30.2-57.8) and 93.4% specificity (95% CI, 89.7-97.0). CTC detected polyps ≥10 mm with 76.9% sensitivity (95% CI, 54.0-99.8) and 89.0% specificity (95% CI, 84.8-93.1). Similar values were found when only adenomatous polyps were considered. The negative predictive value of CTC for adenomas ≥6 mm was 90.7% (95% CI, 86.7-94.5) and for adenomas ≥10 mm the negative predictive value was 98.6% (95% CI, 97.0-100). CONCLUSIONS: In a CRC surveillance population 1 year following resection, CTC was inferior to OC for detecting patients with polyps ≥6 mm. Clinical Trials.gov Registration Number: NCT02143115.

Non-contrast imaging characteristics of papillary renal cell carcinoma: implications for diagnosis and subtyping.

INTRODUCTION: Current radiographic guidelines suggest unenhanced renal lesions < 20 Hounsfield Units (HU) are overwhelmingly benign, requiring no further evaluation. We evaluate our experience with papillary renal cell carcinoma (pRCC) presenting with low pre-contrast attenuation and the relationship of attenuation with histologic pRCC subtype. MATERIALS AND METHODS: We reviewed our institutional kidney cancer database for patients with pT1 or pT2 pRCC between 2003-2017. Tumors were categorized by papillary subtype by expert uropathologists. Preoperative CT images were analyzed at six regional tumor locations. Low, presumably benign, unenhanced median attenuation was defined as ≤ 20 HU. We calculated the frequency of pRCC with low attenuation and assessed the relationship between attenuation and pRCC subtype using logistic regression. RESULTS: Sixty-one patients with evaluable imaging were included. Median tumor size was 6 cm (1.7 cm-15.3 cm) with 39% (n = 24) type-1 and 61% (n = 37) type-2. Half of all pRCC tumors (n = 30) exhibited very low pre-contrast attenuation (< 20 HU), risking misdiagnosis as benign using current guidelines. Of these, 80% (n = 24) were type-2 with significant biological potential. Overall, type-2 tumors demonstrated a lower pre-contrast attenuation than type-1 (median HU: 19.8 (1.5-42.3) versus 29.6 HU (10-45.8), p < 0.01; max HU: 25.3 versus 36.5 HU, p < 0.01). After adjustment, lower pre-contrast HU was an independent predictor of pRCC subtype associated with a 5.5-fold increase of being type-2 (OR = 5.47, p < 0.01). CONCLUSION: pRCCs may exhibit very low attenuation on pre-contrast CT. This appears more common among the more aggressive type-2 subtype. These data suggest that low attenuation (< 20 HU) alone on non-contrast CT imaging is insufficient as a single parameter to rule out malignancy.

A Program Profile of Air Medical Transport in Regional Central Queensland, Australia.

OBJECTIVE: The purpose of this study was to investigate the epidemiology of air medical patients and referral patterns in Central Queensland Hospital and Health Service (CQHHS). METHODS: Analysis of air medical transport from January 2010 to December 2014. Air medical tasks within the local health service boundary were included. All patients transported on rotor or fixed wing aircraft for medical purposes were included. Patterns of air medical tasks in and out of the region by referring and receiving location, aircraft type, flight priority, time of day, month, sex, age, illness, and referral indexes were analyzed. RESULTS: There were 11,456 air ambulance tasks in CQHHS region during the study period, an average of 2,291 retrievals per annum or 191 per month. Frequent referrals were to a tertiary facility, located 800 km across economic and political boundaries. Referral pattern indexes highlight a net patient flow of 1.2 to 1. Cardiology was the largest illness category (24%). Males represented 59% overall as well as patients 66 years and older (33%). Fixed wing aircraft carried out 87% of the tasks with a frequent response time of 6 to 24 hours. CONCLUSION: Air medical transports are an integral part of the health system in Central Queensland communities with vast geographic distances. Identifying regional referral pattern rates and ratios aid in the planning of resource allocation.

Uncoupling protein 3 deficiency impairs myocardial fatty acid oxidation and contractile recovery following ischemia/reperfusion.

Patients with insulin resistance and type 2 diabetes have poor cardiac outcomes following myocardial infarction (MI). The mitochondrial uncoupling protein 3 (UCP3) is down-regulated in the heart with insulin resistance. We hypothesized that decreased UCP3 levels contribute to poor cardiac recovery following ischemia/reperfusion (I/R). After confirming that myocardial UCP3 levels were systematically decreased by 20-49% in animal models of insulin resistance and type 2 diabetes, we genetically engineered Sprague-Dawley rats with partial loss of UCP3 (ucp3+/-). Wild-type littermates (ucp3+/+) were used as controls. Isolated working hearts from ucp3+/- rats were characterized by impaired recovery of cardiac power and decreased long-chain fatty acid (LCFA) oxidation following I/R. Mitochondria isolated from ucp3+/- hearts subjected to I/R in vivo displayed increased reactive oxygen species (ROS) generation and decreased respiratory complex I activity. Supplying ucp3+/- cardiac mitochondria with the medium-chain fatty acid (MCFA) octanoate slowed electron transport through the respiratory chain and reduced ROS generation. This was accompanied by improvement of cardiac LCFA oxidation and recovery of contractile function post ischemia. In conclusion, we demonstrated that normal cardiac UCP3 levels are essential to recovery of LCFA oxidation, mitochondrial respiratory capacity, and contractile function following I/R. These results reveal a potential mechanism for the poor prognosis of type 2 diabetic patients following MI and expose MCFA supplementation as a feasible metabolic intervention to improve recovery of these patients at reperfusion.

Metabolic Syndrome and Bone: Pharmacologically Induced Diabetes has Deleterious Effect on Bone in Growing Obese Rats.

Metabolic syndrome and osteoporosis share similar risk factors. Also, patients with diabetes have a higher risk of osteoporosis and fracture. Liver manifestations, such as non-alcoholic steatohepatitis (NASH), of metabolic syndrome are further aggravated in diabetics and often lead to liver failure. Our objective was to create a rat model of human metabolic syndrome and determine the long-term impact of early-onset T1D on bone structure and strength in obese growing rats. Male rats were given either standard chow and RO water (Controls) or a high-fat, high-cholesterol diet and sugar water containing 55% fructose and 45% glucose (HFD). A third group of rats received the HFD diet and a single dose of streptozotocin to induce type 1 diabetes (HFD/Sz). Body weight and glucose tolerance tests were conducted several times during the course of the study. Serum chemistry, liver enzymes, and biomarkers of bone metabolism were evaluated at 10 and 28 weeks. Shear wave elastography and histology were used to assess liver fibrosis. Cancellous bone structure and cortical bone geometry were evaluated by mCT and strength by the 3-point bending method. Body mass and fat accumulation was significantly higher in HFD and HFD/Sz rats compared to Controls. Rats in both the HFD and HFD/Sz groups developed NASH, although the change was more severe in diabetic rats. Although both groups of obese rats had larger bones, their cancellous structure and cortical thickness were reduced, resulting in diminished strength that was further aggravated by diabetes. The HFD and HFD/Sz rats recapitulate MeSy in humans with liver pathology consistent with NASH. Our data provide strong indication that obesity accompanied by type 1 diabetes significantly aggravates comorbidities of MeSy, including the development of osteopenia and weaker bones. The juvenile rat skeleton seems to be more vulnerable to damage imposed by obesity and diabetes and may offer a model to inform the underlying pathology associated with the unusually high fracture rates in obese adults with diabetes.

Prospective Trial Evaluating the Surgical Anastomosis at One-Year Colorectal Cancer Surveillance: CT Colonography Versus Optical Colonoscopy and Implications for Patient Care.

OBJECTIVE: The aim of this study was to compare the accuracy of CT colonography versus optical colonoscopy for neoplastic involvement at the surgical anastomosis 1 year after curative-intent colorectal cancer resection. DESIGN, SETTING, PATIENTS, AND INTERVENTIONS: Two hundred one patients (mean age, 58.6 years; 117 men, 84 women) underwent same-day contrast-enhanced CT colonography and colonoscopy approximately 1 year (mean, 12.1 months; median, 11.9 months) after colorectal cancer resection as part of a prospective, multicenter trial. All patients enrolled were without clinical evidence of disease and considered low risk for recurrence (stage I-III). MAIN OUTCOME MEASURES: Suspected neoplastic lesions within 5 cm of the colonic anastomosis were recorded at CT colonography, with subsequent colonoscopy performed for the same, with segmental unblinding of colonography findings. Anastomotic region biopsy or polypectomy was performed at the endoscopist's discretion. RESULTS: None of the 201 patients had intraluminal anastomotic cancer recurrence or advanced neoplasia (or metachronous cancers). CT colonography detected extramural perianastomotic recurrence in 2 patients (1.0%); neither was detected at colonoscopy. Only 2 patients (1.0%; 2/201) were called positive at CT colonography for intraluminal anastomotic nondiminutive lesions (7- to 8-mm polyps), which were confirmed at colonoscopy but nonneoplastic at histopathology. At optical colonoscopy, the anastomosis was deemed abnormal and/or biopsied in 10.0% (20/201), yielding only 1 nondiminutive benign neoplasm (7-mm tubular adenoma). LIMITATIONS: The lack of luminal cancer recurrence in our lower-risk cohort precludes assessment of sensitivity for detection, rendering the study underpowered in this regard. Potential cost savings of combined CT/CT colonography over the standard CT/colonoscopy approach were not assessed. CONCLUSIONS: Relevant intraluminal anastomotic pathology appears to be very uncommon 1 year after colorectal cancer resection in lower-risk cohorts. Unlike colonoscopy, diagnostic contrast-enhanced CT colonography effectively evaluates both the intra- and extraluminal aspects of the anastomosis. See Video Abstract at http://links.lww.com/DCR/A471.

Constructing a nursing identity within the clinical environment: The student nurse experience.

BACKGROUND: Nursing identity is an important element of being a nurse. Student nurses begin the construction of their nursing identity during their clinical placements. AIM: The aim of this research was to examine how the student nurses of a regional Australian university construct their identity when on off-campus clinical placement. METHODS/DESIGN: Using a constructivist approach an online survey was used to elicit data in response to the question 'What elements are needed during the work integrated learning experience to enable undergraduate nursing students to construct their nursing identity?' RESULTS/FINDINGS: Findings reveal five key elements to the construction of students' nursing identity; positive role models, belonging, peer support, critical thinking abilities and confidence. CONCLUSION: Such findings are important as they provide information for student nurses, preceptors and educators in guiding clinical placement experiences that are able to facilitate the development of the nursing identity.

Sex-specific colonic mitochondrial dysfunction in the indomethacin-induced rat model of inflammatory bowel disease.

Introduction: Inflammatory bowel disease (IBD) is characterized by chronic inflammation of the gastrointestinal tract and encompasses Crohn's Disease and Ulcerative Colitis. Women appear to have more severe and recurring symptoms of IBD compared to men, most likely due to hormonal fluctuations. Studies have shown that mitochondrial dysfunction plays a role in the development of inflammation and there is evidence of colon mitochondrial alterations in IBD models and patients. In this study we have identified the presence of sex-specific colon mitochondrial dysfunction in a rat model of IBD. Methods: Eight-week-old male and female rats were treated with indomethacin to induce IBD and mitoTEMPO was administered daily either after or before induction of IBD and until euthanasia. Colons were collected for histology and mitochondrial experiments. Intact mitochondrial respiration, reactive oxygen species (mtROS), the activities of the individual electron transport complexes and the activities of the antioxidant enzymes were measured to assess mitochondrial function. Results: IBD male rats showed a decrease in citrate synthase activity, cardiolipin levels, catalase activity and an increase in mtROS production. IBD females show a decrease in intact colon mitochondrial respiration, colon mitochondria respiratory control ratio (RCR), complex I activity, complex IV activity, and an increase in mtROS. Interestingly, control females showed a significantly higher rate of complex I and II-driven intact mitochondrial respiration, MCFA oxidation, complex II activity, complex III activity, and complex IV activity compared to control males. The use of a mitochondrial-targeted therapy, mitoTEMPO, improved the disease and colon mitochondrial function in female IBD rats. However, in the males there was no observed improvement, likely due to the decrease in catalase activity. Conclusion: Our study provides a better understanding of the role mitochondria in the development of IBD and highlights sex differences in colon mitochondrial function. It also opens an avenue for the development of strategies to re-establish normal mitochondrial function that could provide more options for preventive and therapeutic interventions for IBD.

Oxidation of hemoproteins by Streptococcus pneumoniae collapses the cell cytoskeleton and disrupts mitochondrial respiration leading to the cytotoxicity of human lung cells.

IMPORTANCE: Streptococcus pneumoniae (Spn) colonizes the lungs, killing millions every year. During its metabolism, Spn produces abundant amounts of hydrogen peroxide. When produced in the lung parenchyma, Spn-hydrogen peroxide (H2O2) causes the death of lung cells, and details of the mechanism are studied here. We found that Spn-H2O2 targets intracellular proteins, resulting in the contraction of the cell cytoskeleton and disruption of mitochondrial function, ultimately contributing to cell death. Intracellular proteins targeted by Spn-H2O2 included cytochrome c and, surprisingly, a protein of the cell cytoskeleton, beta-tubulin. To study the details of oxidative reactions, we used, as a surrogate model, the oxidation of another hemoprotein, hemoglobin. Using the surrogate model, we specifically identified a highly reactive radical whose creation was catalyzed by Spn-H2O2. In sum, we demonstrated that the oxidation of intracellular targets by Spn-H2O2 plays an important role in the cytotoxicity caused by Spn, thus providing new targets for interventions.

Increased platelet mitochondrial function correlates with clot strength in a rodent fracture model.

BACKGROUND: Thromboelastographic measures of clot strength increase early after injury, portending higher risks for thromboembolic complications during recovery. Understanding the specific role of platelets is challenging because of a lack of clinically relevant measures of platelet function. Platelet mitochondrial respirometry may provide insight to global platelet function but has not yet been correlated with functional coagulation studies. METHODS: Wistar rats underwent anesthesia and either immediate sacrifice for baseline values (n = 6) or (1) bilateral hindlimb orthopedic injury (n = 12), versus (2) sham anesthesia (n = 12) with terminal phlebotomy/hepatectomy after 24 hours. High-resolution respirometry was used to measure basal respiration, mitochondrial leak, maximal oxidative phosphorylation, and Complex IV activity in intact platelets; Complex I- and Complex II-driven respiration was measured in isolated liver mitochondria. Results were normalized to platelet number and protein mass, respectively. Citrated native thromboelastography (TEG) was performed in triplicate. RESULTS: Citrated native TEG maximal amplitude was significantly higher (81.0 ± 3.0 vs. 73.3 ± 3.5 mm, p < 0.001) in trauma compared with sham rats 24 hours after injury. Intact platelets from injured rats had higher basal oxygen consumption (17.7 ± 2.5 vs. 15.1 ± 3.2 pmol O 2 /[s × 10 8 cells], p = 0.045), with similar trends in mitochondrial leak rate ( p = 0.19) when compared with sham animals. Overall, platelet basal respiration significantly correlated with TEG maximal amplitude ( r = 0.44, p = 0.034). As a control for sex-dependent systemic mitochondrial differences, females displayed higher liver mitochondria Complex I-driven respiration (895.6 ± 123.7 vs. 622.1 ± 48.7 mmol e - /min/mg protein, p = 0.02); as a control for systemic mitochondrial effects of injury, no liver mitochondrial respiration differences were seen. CONCLUSION: Platelet mitochondrial basal respiration is increased after injury and correlates with clot strength in this rodent hindlimb fracture model. Several mitochondrial-targeted therapeutics exist in common use that are underexplored but hold promise as potential antithrombotic adjuncts that can be sensitively evaluated in this preclinical model.

Sex-specific colonic mitochondrial dysfunction in the indomethacin-induced inflammatory bowel disease model in rats.

Introduction: Inflammatory bowel disease (IBD) is characterized by chronic inflammation of the gastrointestinal tract and encompasses Crohn's disease and ulcerative colitis. Women appear to have more severe and recurring symptoms of IBD compared to men, most likely due to hormonal fluctuations. Studies have shown that mitochondrial dysfunction plays a role in the development of inflammation and there is evidence of colon mitochondrial alterations in IBD models and patients. In this study we have identified the presence of sex-specific colon mitochondrial dysfunction in a rat model of IBD. Methods: Eight-week-old male and female rats were treated with indomethacin to induce IBD and mitoTEMPO was administered daily either after or before induction of IBD and until euthanasia. Colons were collected for histology and mitochondrial experiments. Intact mitochondrial respiration, reactive oxygen species (mtROS), the activities of the individual electron transport complexes and the activities of the antioxidant enzymes were measured to assess mitochondrial function. Results: IBD male rats showed a decrease in citrate synthase activity, cardiolipin levels, catalase activity and an increase in mtROS production. IBD females show a decrease in intact colon mitochondrial respiration, colon mitochondria respiratory control ratio (RCR), complex I activity, complex IV activity, and an increase in mtROS. Interestingly, control females showed a significantly higher rate of complex I and II-driven intact mitochondrial respiration, MCFA oxidation, complex II activity, complex III activity, and complex IV activity compared to control males. The use of a mitochondrial-targeted therapy, mitoTEMPO, improved the disease and colon mitochondrial function in female IBD rats. However, in the males there was no observed improvement, likely due to the decrease in catalase activity. Conclusions: Our study provides a better understanding of the role mitochondria in the development of IBD and highlights sex differences in colon mitochondrial function. It also opens an avenue for the development of strategies to re-establish normal mitochondrial function that could provide more options for preventive and therapeutic interventions for IBD.

The cyclic lipopeptide micafungin induces rupture of isolated mitochondria by reprograming the mitochondrial inner membrane anion channel.

The antifungal activity of the drug micafungin, a cyclic lipopeptide that interacts with membrane proteins, may involve inhibition of fungal mitochondria. In humans, mitochondria are spared by the inability of micafungin to cross the cytoplasmic membrane. Using isolated mitochondria, we find that micafungin initiates the uptake of salts, causing rapid swelling and rupture of mitochondria with release of cytochrome c. The inner membrane anion channel (IMAC) is altered by micafungin to transfer both cations and anions. We propose that binding of anionic micafungin to IMAC attracts cations into the ion pore for the rapid transfer of ion pairs.

Analysis of oxygen consumption rates in zebrafish reveals differences based on sex, age and physical activity recovery.

Introduction: Mitochondrial dysfunction is linked to a variety of human diseases. Understanding the dynamic alterations in mitochondrial respiration at various stages of development is important to our understanding of disease progression. Zebrafish provide a system for investigating mitochondrial function and alterations during different life stages. The purpose of this study was to investigate our ability to measure mitochondrial oxygen consumption rates in zebrafish embryos, larvae, and adults as an indicator of mitochondrial function. Methods: Basal respiration of entire zebrafish embryos (5 dpf), larvae (0.6-0.9 cm), young adults (3-month-old), and old adults (12-month-old) was measured using an Oroboros Oxygraph, with a stirrer speed of 26 rpm. For embryos and larvae, "leak" respiration (plus oligomycin), maximum respiration (plus uncoupler), non-mitochondrial respiration (plus inhibitors), and complex IV activity were also measured. To induce physical activity in adult fish, the stirrer speed was increased to 200 rpm. Results and Discussion: We demonstrate the ability to accurately measure respiration rates in zebrafish at various ages using the Oroboros Oxygraph. When comparing zebrafish embryos to larvae, embryos have a higher maximum respiration. Three-month-old zebrafish males have higher basal respiration than females, while 12-month-old zebrafish females exhibit greater rates of respiration than males and younger females. When the stirrer speed was increased, respiration rates decrease, but with differences depending on sex. This study demonstrates a simple and accessible method to assess zebrafish physiology by mitochondrial oxygen consumption measurements in an unmodified Oroboros Oxygraph. The method should facilitate studies to understand the intricate interplay between mitochondrial function, development, and aging.

Air ambulance retrievals of patients with suspected appendicitis and acute abdominal pain: The patients' journeys, referral pathways and appendectomy outcomes using linked data in Central Queensland, Australia.

INTRODUCTION: Acute appendicitis is the most common cause of acute abdominal pain presentations to the ED and common air ambulance transfer. AIMS: describe how linked data can be used to explore patients' journeys, referral pathways and request-to-activation responsiveness of patients' appendectomy outcomes (minor vs major complexity). METHODS: Data sources were linked: aeromedical, hospital and death. Request-to-activation intervals showed strong right-tailed skewness. Quantile regression examined whether the longest request-to-activation intervals were associated with appendicitis complexity in patients who underwent an appendectomy. RESULTS: There were 684 patients in three referral pathways based on hospital capability levels. In total, 5.6 % patients were discharged from ED. 83.3 % of all rural origins entered via the ED. 3.8 % of appendicitis patients were triaged to tertiary hospitals. Appendectomy patients with major complexity outcomes were less likely to have longer request-to-activation wait times & had longer lengths of stay than patients with minor complexity outcomes. CONCLUSIONS: Linked data highlighted four aspects of a functioning referral system: appendectomy outcomes of major complexity were less likely to have longer request-to-activation intervals compared to minor (sicker patients were identified); few were discharged from EDs (validated transfer); few were triaged to tertiary hospitals (appropriate level for need), and no deaths relating to appendectomy.

Oxidation of hemoproteins by Streptococcus pneumoniae collapses the cell cytoskeleton and disrupts mitochondrial respiration leading to cytotoxicity of human lung cells.

Streptococcus pneumoniae (Spn) causes pneumonia that kills millions through acute toxicity and invasion of the lung parenchyma. During aerobic respiration, Spn releases hydrogen peroxide (Spn-H 2 O 2 ), as a by-product of enzymes SpxB and LctO, and causes cell death with signs of both apoptosis and pyroptosis by oxidizing unknown cell targets. Hemoproteins are molecules essential for life and prone to oxidation by H 2 O 2 . We recently demonstrated that during infection-mimicking conditions, Spn-H 2 O 2 oxidizes the hemoprotein hemoglobin (Hb), releasing toxic heme. In this study, we investigated details of the molecular mechanism(s) by which the oxidation of hemoproteins by Spn-H 2 O 2 causes human lung cell death. Spn strains, but not H 2 O 2 -deficient SpnΔ spxB Δ lctO strains caused time-dependent cell cytotoxicity characterized by the rearrangement of the actin, the loss of the microtubule cytoskeleton and nuclear contraction. Disruption of the cell cytoskeleton correlated with the presence of invasive pneumococci and an increase of intracellular reactive oxygen species. In cell culture, the oxidation of Hb or cytochrome c (Cyt c ) caused DNA degradation and mitochondrial dysfunction from inhibition of complex I-driven respiration, which was cytotoxic to human alveolar cells. Oxidation of hemoproteins resulted in the creation of a radical, which was identified as a protein derived side chain tyrosyl radical by using electron paramagnetic resonance (EPR). Thus, we demonstrate that Spn invades lung cells, releasing H 2 O 2 that oxidizes hemoproteins, including Cyt c , catalyzing the formation of a tyrosyl side chain radical on Hb and causing mitochondrial disruption, that ultimately leads to the collapse of the cell cytoskeleton.

RUPP Th17s cause hypertension and mitochondrial dysfunction in the kidney and placenta during pregnancy.

BACKGROUND: Preeclampsia (PE), new-onset hypertension (HTN), and organ dysfunction during the second half of pregnancy, is associated with an increase in inflammatory immune cells, including T helper 17 (Th17) cells. Studies have demonstrated that mitochondrial (mt) dysfunction is important in the pathogenesis of PE though causative factors have yet to be fully identified. Although Th17 cells, natural killer (NK) cells, and mt dysfunction contribute to HTN in the reduced uterine perfusion pressure (RUPP) rat model, the role of Th17 cells or IL-17 in mt dysfunction is unknown. Therefore, we hypothesize that RUPP stimulated Th17 cells cause HTN and mt dysfunction, which is alleviated with the blockade of IL-17. METHODS: On gestational day 12 (GD12), RUPP Th17 cells were transferred into normal pregnant (NP) Sprague Dawley rats. A subset of NP + RUPPTh17 rats received IL-17RC (100 pg/day) on GD14-19. Blood pressure (MAP), NK cells, and mt function were measured on GD19 in all groups. RESULTS: MAP increased in response to NP + RUPP Th17 compared to NP rats and was lowered with IL-17RC. Circulating and placental NK cells increased with NP + RUPP Th17 compared to NP and were lowered with IL-17RC. Renal mtROS increased in NP + RUPP Th17 compared to NP and was normalized with IL-17RC. Similar to PE women, placental mtROS decreased in NP + RUPP Th17 and was normalized with IL-17RC. CONCLUSION: Our results indicate that IL-17RC inhibition normalizes HTN, NK cell activation, and multi-organ mt dysfunction caused by Th17 cells stimulated in response to placental ischemia.

Oxidation of hemoglobin in the lung parenchyma facilitates the differentiation of pneumococci into encapsulated bacteria.

Pneumococcal pneumonia causes cytotoxicity in the lung parenchyma but the underlying mechanism involves multiple factors contributing to cell death. Here, we discovered that hydrogen peroxide produced by Streptococcus pneumoniae (Spn-H 2 O 2 ) plays a pivotal role by oxidizing hemoglobin, leading to its polymerization and subsequent release of labile heme. At physiologically relevant levels, heme selected a population of encapsulated pneumococci. In the absence of capsule and Spn-H 2 O 2 , host intracellular heme exhibited toxicity towards pneumococci, thus acting as an antibacterial mechanism. Further investigation revealed that heme-mediated toxicity required the ABC transporter GlnPQ. In vivo experiments demonstrated that pneumococci release H 2 O 2 to cause cytotoxicity in bronchi and alveoli through the non-proteolytic degradation of intracellular proteins such as actin, tubulin and GAPDH. Overall, our findings uncover a mechanism of lung toxicity mediated by oxidative stress that favor the growth of encapsulated pneumococci suggesting a therapeutic potential by targeting oxidative reactions. Highlights: Oxidation of hemoglobin by Streptococcus pneumoniae facilitates differentiation to encapsulated pneumococci in vivo Differentiated S. pneumoniae produces capsule and hydrogen peroxide (Spn-H 2 O 2 ) as defense mechanism against host heme-mediated toxicity. Spn-H 2 O 2 -induced lung toxicity causes the oxidation and non-proteolytic degradation of intracellular proteins tubulin, actin, and GAPDH. The ABC transporter GlnPQ is a heme-binding complex that makes Spn susceptible to heme toxicity.