bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mitochondrial Dysfunction in Acute Lung Injury is Characterized By Loss of Mitochondrial Calcium Uniporter (MCU)
Authors: Islam Mohammad N. , Gusarova Galina A , Das Shonit , Bhattacharya Jahar 
Published: November 2023

Acute lung injury (ALI) associates with loss of lung mitochondrial function. Oxidant damage by mitochondrial H2O2 may cause mitochondrial dysfunction. Since the mitochondrial calcium uniporter (MCU) determines mitochondrial Ca2+ levels, hence function, we considered the possibility that oxidant‐induced MCU impairment might underlie mitochondrial dysfunction in ALI. To induce ALI, we intra‐nasally instilled lipopolysaccharide (LPS, 1mg/kg) or PBS (control). To determine H2O2, we expressed a mitochondrial matrix‐targeted probe, matrix‐roGFP, in alveolar type 2 (AT2) cells in situ. By alveolar micropuncture we loaded mitochondrial Ca2+ dye, rhod 2, or the lamellar body (LB)‐staining dye LysoTracker Red (LTR). In control lungs, H2O2 fluorescence was 145±7 grey levels in AT2 cells, indicating successful probe expression. In LPS lungs, AT2 mitochondrial H2O2 fluorescence was 75±8 (P<0.05), indicating increased H2O2 generation. Immunoblots in AT2 cell mitochondria show that LPS downregulated MCU. However, LPS‐induced MCU downregulation could be reversed in mice over‐expressing human catalase (hmCAT) in mitochondria of AT2 cells. In addition, hmCAT over‐expressing mice reversed LPS‐induced inhibition of mitochondrial Ca2+ entry and surfactant secretion in response to a 15s lung hyperinflation. These data are the first direct evidence that lung exposure to LPS causes mitochondrial dysfunction marked by oxidant‐induced loss of MCU. Taken together, our findings indicate that alveolar MCU is a major determinant of ALI. Strategies aimed at restoring alveolar MCU expression might lead to novel therapy for ALI.Support or Funding InformationPBF fellowship for MNI and HL122730 for JB
Claudin‐5 Decreases Alveolar Barrier Function in Alcoholic Lung Syndrome by Displacing Claudin‐18 from Tight Junctions
Authors: Schlingmann Barbara Lisa , White StevenClaude Dorsainvil , Molina Samuel , Lynn K. Sabrina , Capaldo Christopher T. , Koval Michael 
Published: November 2023

We found that alcohol abuse increases the susceptibility to develop Acute Respiratory Distress Syndrome (ARDS) by impairing alveolar epithelial cell tight junctions. Understanding how tight junction proteins are regulated on a molecular level may identify potential new therapeutic targets for the treatment and prevention of ARDS. We used a rodent model of chronic alcohol ingestion and cultured primary alveolar epithelial cells (AEC) to investigate the influence of alcohol on tight junction formation and function. We found that AECs from alcohol fed rats had disrupted tight junction morphology and were leakier than control AECs. Alcohol‐induced changes to AEC tight junctions were accompanied by increased expression of a claudin family tight junction protein, claudin‐5. Immunofluorescence analysis showed that impaired barrier function of AECs from alcohol fed rats was accompanied by disruption of tight junction morphology, spikes, containing claudin‐18. Quantification of cells containing claudin‐18 spikes showed that alcohol increases spike formation and this was also associated with increased claudin‐5 expression. Claudin‐5 knockdown experiments in alcohol exposed AECs showed that decreasing claudin‐5 improved barrier function and reduced the number of cells containing claudin‐18 spikes. Conversely, increasing claudin‐5 expression in control AECs mimicked the effects of alcohol by increasing claudin‐18 spikes and paracellular leak. Live cell imaging experiments revealed that claudin‐18 spikes are sites where claudin‐containing vesicles fuse and form reflecting enhanced claudin turnover which was associated with decreased barrier function. To elucidate the molecular mechanisms of alcohol dependent changes in tight junction proteins, we performed Stochastic optical reconstruction microscopy (STORM), a form of super resolution immunofluorescence microscopy. The data revealed that alcohol ingestion lead to an increased colocalization between claudin‐18 and claudin‐5 which in turn was associated with a decreased colocalization between claudin‐18 and the scaffold protein ZO‐1. When YFP‐tagged claudin‐5 was expressed in normal AECs there was a decreased co‐localization between claudin‐18 and ZO‐1 suggesting that the interaction between claudin‐5 and claudin‐18 impairs its ability to interact with ZO‐1, thus destabilizing incorporation of claudin‐18 in tight junctions. Moreover, agents that inhibit claudin‐5 function in alcohol exposed AECs increased barrier function and stabilized tight junction morphology. The data suggest that targeting claudin‐5 may provide a therapeutic strategy to improve barrier function in ARDS.Support or Funding InformationSupport: Emory URC, NIH‐NIAAA, NIH‐NHLBI. DAAD
MORPHINE PROTECTS AGAINST CARDIAC DYSFUNCTION DURING TYPE 2 DIABETES BY IMPROVING MITOCHONDRIAL STRUCTURE AND FUNCTION
Authors: Zemljic‐Harpf Alice Eugenie , Hoe Louise See , Schilling Jan Mirko , Nguyen Alexander , Vainshav Yash J. , Peart Jason N. , Mahata Sushil K. , Patel Hemal H. , Headrick John P. 
Published: November 2023

Diabetes promotes ischemic heart disease, cardiomyopathy, intolerance to ischemia‐reperfusion (I‐R) injury, and refractoriness to cardioprotection. Evidence suggests that cardiac abnormalities during diabetes involve inflammation, lipotoxicity, posttranslational‐modifications as well as mitochondrial dysfunction and impaired opioid receptor (OR) signaling. We test the hypothesis that a novel sustained OR stimulus may overcome diabetes induced cardiac dysfunction via mitochondrial dependent protection.In a murine type 2 diabetes mellitus (T2DM) model we investigated effects of morphine on cardiac function, structure and I‐R tolerance, mitochondrial function and ultrastructure. Streptozotozin (75mg/kg i.p.) was administered once in non‐fasted 12‐week old mice, and normal chow was switched to 60% kcal high‐fat diet (HFD). 12 weeks of HFD induced 25% weight gain, hyperglycemia, hyperinsulinemia, glucose intolerance, cardiac hypertrophy, moderate cardiac depression, exaggerated post‐ischemic myocardial dysfunction and death, and abnormalities in mitochondrial respiration, ultrastructure and Ca2+‐induced swelling. Morphine administration for 5 days: 1) improved glucose homeostasis and body weight; 2) reversed cardiac depression; 3) enhanced I‐R tolerance; 4) restored mitochondrial ultrastructure; and 5) improved mitochondrial function. A treatment‐matched caloric restriction regime modestly improved I‐R tolerance in T2DM mice without improving in vivo cardiac function.These data reveal that morphine treatment restores contractile function and ischemic tolerance in diabetic hearts via improvements in mitochondrial structure and function, implicating OR activation as a potential novel target in managing T2DM‐induced myocardial abnormalities.Support or Funding InformationThis study was supported by grants from the National Institutes of Health, USA (HL091071, HL107200, HL115933, and HL066941), a VA Merit grant (BX001963 and BX000783), and the National Health and Medical Research Council of Australia (481922).
Evaluation of the vasopressor responses of metformin and one analogue of metformin in insulin resistance rats
Authors: Gutierrez‐Lara Erika J , Sanchez‐Lopez Araceli , Navarrete‐Vazquez Gabriel , Centurion David 
Published: November 2023

Cardiovascular diseases and diabetes are the leading causes of mortality in the world; both of them are associated with insulin resistance. Metformin improves insulin sensitivity and it is associated with cardioprotection and vasculoprotection. In vitro studies have demonstrated that metformin improves vasorelaxation and normalization of endothelial function during diabetes. Thus, in this study the vasopressor responses induced by sympathetic stimulation or by i.v. bolus injections of the agonists noradrenaline (endogenous ligand), methoxamine (α1) and UK 14,304 (α2) were determined in rats with insulin resistance or control rats pretreated with: (1) metformin; (2) one analogue of metformin (N‐benzylimidodicarbonimidic diamide; EGL‐6M); or (3) its vehicle. For this purpose, rats were treated with fructose (solution 15%) or its vehicle (water; control) for four months. Next, both groups were divided into three subgroups which were administrated with: (i) metformin (50 mg Kg−1day−1), (ii) EGL‐6M (50 mg Kg−1day−1) or its (iii) vehicle (water; 1 mL Kg−1day−1) during 28 days. Tolerance glucose curve (TGC) and insulin levels were determined after administration of glucose (1 g Kg−1, p.o.) in the above subgroups. Animals treated with fructose showed hyperinsulinaemia during TGC, which were reverted by metformin or EGL‐6M. Then, under anesthesia with isoflurane, animals were pithed and prepared to measure blood pressure and heart rate. In animals with insulin resistance, the vasopressor responses to sympathetic stimulation were decreased, while responses to noradrenaline were increased and those induced by methoxamine and UK 14,304 were not modified. In control animals, metformin failed to modify the vasopressor responses induced by sympathetic stimulation or by noradrenaline, methoxamine and UK‐14,304. In contrast, EGL‐6M significantly increased the vasopressor responses to sympathetic stimulation; the same results were showed in the insulin resistance rats. In animals with insulin resistance, metformin decreased vasopressor response to noradrenaline but did not modify the sympathetic stimulation responses. These results suggest that insulin resistance modified the vasopressor responses by inhibiting sympathetic tone and probably as a compensatory mechanism it may increase α‐adrenoceptor expression. Moreover, during insulin resistance, the analogue, but not metformin, improved sympathetic neurotransmission in the systemic vasculature.Support or Funding InformationThe authors acknowledge to Conacyt for their financial support.
A Novel Treatment for Late‐Stage Metastatic Carcinoma: Targeted Osmotic Lysis by Simultaneous Blockade of Sodium Pumps and Stimulation of Voltage‐Gated Sodium Channels
Authors: Paul Dennis , Gould Harry J. 
Published: November 2023

About 80% metastatic carcinomas (MCs) overexpress voltage‐gated sodium channels (VGSCs) by 50‐fold or more. We hypothesized that electrical stimulation of VGSCs with concomitant blockade of Na+, K+‐ATPase (sodium pumps) would produce a rapid increase in intracellular sodium, causing an influx of water, leading to lysis of MC cells (Targeted Osmotic Lysis, US Patent #8921320). To test this hypothesis in vitro, we cultured highly metastatic MDA‐MB‐231 cells, which overexpress NaV1.5 channels. For single cell experiments, treatment with the sodium pump blocker ouabain and electrical stimulation of the cells produced lysis in as little as 16 sec. Ouabain or electric current alone did not lyse the cells. Treatment did not lyseMCF‐10a or MCF‐12a (normal breast) cells. For experiments in 24‐well plates, entire wells of MDA‐MB‐231 cells were lysed in 10 min. To test the hypothesis in vivo, we established MDA‐MB‐231allographs in Nu/J nude mice. When tumor diameter reached approximately 1 cm, we administered 10 mg/kg ouabain s.c. or saline. Thirty and 60 min later, tumors received 60 sec of 10V direct current delivered through direct contact stimulation. This single‐treatment produced 70% cell lysis in tumors treated with both ouabain and stimulation. Tumors treated with saline alone and saline plus stimulation averaged 10% lysis. Ouabain alone produced 15% cell death. Treatment did not affect non cancerous muscle tissue surrounding the tumor. In survival studies, single day treatment attenuated tumor growth by 45%. Three treatments separated by 3 days attenuated growth by 60–100%. Thus, targeted osmotic lysis is a promising option as an effective treatment of MCs that overexpress VGSCs.Support or Funding InformationThis research was supported by the LSU Depts. of Pharmacology and Neurology, and Oleander Medical Technologies, LLC.
Glucose‐Mediated Regulation of UQCRFS1 and the Cardiac Metabolome
Authors: Williams Lamario J , Brahma Manoja K , McCrory Mark M , Wende Adam R 
Published: November 2023

Diabetes, among other metabolic disorders, is associated with harmful changes to the heart. One potential mechanism for this is reduced flexibility in mitochondrial substrate utilization leading to heart failure. The goal of this project is to define the molecular mechanisms by which diabetes contributes to the development of mitochondrial dysfunction. Accumulating evidence suggests that increased flux of glucose through non‐oxidative pathways such as the hexosamine biosynthetic pathway (HBP) may lead to cardiovascular complications in diabetic patients. HBP produces UDP‐N‐acetylglucosamine (UDP‐GlcNAc) that provides the sugar molecule for O‐linked attachment of GlcNAc moieties to specific serine/threonine residues of nuclear, cytoplasmic, and mitochondrial proteins. We hypothesized that hyperglycemia‐induced mitochondrial protein O‐GlcNAcylation may contribute to the impaired substrate utilization resulting in increased reactive oxygen species (ROS) production in the diabetic myocardium. Excess ROS production has been widely implicated in both the onset of diabetes and many of its complications. To test this hypothesis, we used the streptozotocin‐induced diabetic mouse model. In addition, to investigate the glucose‐specific contribution, an inducible transgenic mouse model overexpressing glucose transporter 4 (GLUT4; mG4H) specifically in cardiomyocytes was used. We found that increased flux of glucose leads to enhanced posttranslational modification of mitochondrial proteins by O‐GlcNAcylation in diabetic and mG4H mice. From wheat germ agglutinin enrichment and immunoprecipitation assays, we have identified that UQCRFS1, an OXPHOS complex III protein, is O‐GlcNAcylated in diabetic and mG4H mice heart. In addition, microarray and qPCR analysis confirmed that Uqcrfs1 mRNA levels were unchanged suggesting that this regulation does not involve transcriptional pathways and is specific to O‐GlcNAc of UQCRFS1 in the development of mitochondrial dysfunction. Future goals for this project are to identify diabetic signatures that define cardiac mitochondrial dysfunction related to decrease in OXPHOS complex activity. To test this we have also developed an in vitro model using immortalized human cardiomyocytes that can be tested using the Seahorse Flux Analyzer to directly interrogate each OXPHOS complex. Further characterization of GlcNAc‐mediated regulation of UQCRFS1 in response to increased glucose delivery and substrate overload will be confirmed using immunoprecipitation followed by mass spectrometry to define this mechanism in disease progression in a diabetic myocardium. Together successful completion of these studies will provide novel insights into the molecular mechanisms by which excess glucose in diabetes contributes to mitochondrial dysfunction.Support or Funding InformationNIH R00 HL111322 and HL111322‐04S1 to A.R.W.NIH R25 DK078381 to L.J.W.
Regression of pressure overload‐induced cardiac hypertrophy by trientine‐induced copper redistribution to the heart in rats
Authors: Liu Jiaming , Liu Yinjie , Fu Chunyan , Xiao Ying , Feng Qipu , Fan Suzhen , Ding Xueqin , Qiu Liying , Han Pengfei , Kang Y. James 
Published: November 2023

Sustained pressure overload leads to pathological cardiac hypertrophy, and copper efflux from the heart with an increase in plasma copper concentrations. Dietary copper supplementation replenishes copper content and reverses the pathogenesis in mice with cardiac hypertrophy. We recently observed that copper‐trientine complex increased copper uptake by primary cultures of neonatal rat cardiomyocytes. The present study was undertaken to test the hypothesis that trientine can redistribute copper from plasma to the copper‐depleted heart tissue. Adult male Sprague‐Dawley rats were subjected to ascending aortic constriction (AAC). Sixteen weeks after the operation, cardiac hypertrophy was observed by echocardiography and trientine at a dose of 8 mg/kg, twice a day, was administered orally for 6 weeks. At the end of the treatment, copper concentrations were significantly increased in the heart and significantly reduced in the plasma. Along with copper redistribution, cardiac hypertrophy was reversed almost back to the normal as judged by IVSD and LVPWD, and cardiac function was improved as indicated by EF and FS. In addition, the level of cardiac fibrosis was reduced as reflected by the decrease in the content of collagens type I and III and the reduced cross linking of the collagens. This study demonstrates that trientine, although traditionally considered as a copper chelator, is capable of redistributing copper from plasma to the heart leading to regression of pathological cardiac hypertrophy and improved cardiac function.Support or Funding InformationNational Science Foundation of China (81230004 and 81300109).
Optical cryo‐imaging reveals a heterogeneous distribution of mitochondrial redox state in the ex‐vivo guinea pig heart and its alteration during ischemia and reperfusion
Authors: Camara Amadou K.S. , Salehpour Fahimeh , Motlagh Mohammad M , Sepehr Reyhaneh , Heisner James S , Dash Ranjan K , Ranji Mahsa 
Published: November 2023

RationaleOxidation of substrates to generate ATP in mitochondria is mediated through the redox reactions of reducing equivalents, NADH and FADH2, via the mitochondrial electron transport chain. Cardiac ischemia and reperfusion (IR) injury compromises mitochondrial oxidative phosphorylation (OxPhos), in part, by altering the mitochondrial redox states. We have shown previously using a fiber optic probe online (spectroflurometrically) in the ex‐vivo heart changes in the mitochondrial redox states before, during, and after IR. However, this approach is limited in that it only shows the changes in the redox states in a patch of cardiac tissue in the left ventricular free wall. The heart is a complex and heterogeneous organ anatomically and physiologically, and alteration of the metabolic heterogeneity can portend the state of its health. We hypothesize that IR alters the metabolic heterogeneity of the mitochondrial redox state of the heart that is only evident in the 3‐dimensonal (3‐D) optical cryo‐imaging of the perfused heart before, during, and after IR.MethodsThe study involved four groups of hearts: time control (TC: heart perfusion without IR), perfusion followed by global ischemia only (Isch), perfusion followed by IR, and TC with PCP (a mitochondrial uncoupler) perfusion. Mitochondrial NADH and FAD autofluorescence signals were recorded spectrofluorometrically online (using a PTI system) in guinea pig ex‐vivo perfused hearts in the Langendorff mode before, during, and after IR. At the end of each specified protocol, hearts were rapidly removed and snap frozen in liquid N2 for later 3‐D optical cryo‐imaging of the mitochondrial NADH, FAD, and NADH/FAD redox ratio (RR).ResultsThe TC hearts revealed a heterogeneous spatial distribution of NADH, FAD and RR. Ischemia and IR altered the spatial distribution and caused an overall increase and decrease in the RR by 55% and 64%, respectively. During ischemia, the RR distribution appeared to be similar to the TC and the distribution of the RR in the IR group displayed a shift that is different from the TC group. The spectrofluorometric data of the RR paralleled our previous results in the changes in the redox state during IR. Uncoupling with PCP resulted in the lowest level of the RR (73% oxidation) compared with control (TC).ConclusionThe 3‐D optical cryo‐imaging of the heart provides novel insights into the heterogeneous distribution of the mitochondrial RR and metabolism from the base to the apex during ischemia and IR. This 3‐D information of the mitochondrial redox state in the normal and ischemic heart was not apparent in the dynamic spectrofluorometric data.Support or Funding InformationThe work was supported by the following grants: NIH P01 GM 066730 (ZB PI/PD); NIH R01 HL095122 (RKD and AKSC); CTSI and RGI grants (MR).
Determination of Regional Flow Utilizing Microspheres in ex vivo Myocardial Ischemia/Reperfusion (MI/R) Injury
Authors: Kuhn Tarah F. , Clark Sabrina , Kuo Daniel , Liu Stephanie , Chen Qian , Barsotti Robert , Young Lindon H. 
Published: November 2023

MI/R results in marked cardiac contractile dysfunction and cell death. We previously discovered that protein kinase C epsilon peptide inhibitor (PKC ɛ‐) robustly restored postreperfused cardiac function, and reduced infarct size, oxidative stress, and leukocyte endothelial interactions in coronary, hind limb, renal, and mesenteric vascular inflammation models. The mechanisms of these effects in part are due to attenuating uncoupled endothelial nitric oxide (NO) synthase activity and increasing endothelial NO bioavailability. Therefore, we hypothesize that PKC ɛ‐will increase regional flow during reperfusion. We determined regional flow to the right ventricle, left ventricle, and septum at baseline and after 10 and 45 min reperfusion using fluorescent microspheres in isolated perfused rat hearts subjected to global I(30min)/R(45min). A cell permeable PKC ɛ‐(myr‐EAVSLKPT, MW=1054 g/mol, 10μM, n=8) was given at beginning of reperfusion for 5 min. We found that final left ventricular developed pressure (LVDP) recovered to 70 ± 7%, maximal rate of left ventricular contraction, (+dP/dtmax) to 57 ± 7% and maximal rate of left ventricular relaxation, (−dP/dtmin) to 58 ± 6% of baseline values. These parameters were significantly improved compared to untreated control I/R hearts (n=7) that only recovered to 30 ± 5% in LVDP, 21 ± 5% in +dP/dtmax and 25 ± 5% in −dP/dtmin relative to baseline values (all p < 0.01). Moreover, our preliminary data suggest that PKC ɛ‐treatment increased regional flow by 190 ± 40% in the right ventricle, 140 ± 30% in the left ventricle, and 120 ± 20% in the septum at 10 min postreperfusion compared to untreated control I/R hearts. These values were maintained throughout the remaining 35 min of reperfusion. In summary, the data indicates that PKC ɛ‐improves postreperfused cardiac function in part by restoration of regional flow.Support or Funding InformationThis study was supported by the Department of Bio‐Medical Sciences, Division of Research and the Center for Chronic Disorders of Aging at Philadelphia College of Osteopathic Medicine and the Pennsylvania Department of Health.