Publications by authors named "Marla R Wolfson"

62 Publications

Perfluorochemical-facilitated plasminogen activator delivery to the airways: A novel treatment for inhalational smoke-induced acute lung injury.

Clin Transl Med 2020 Jan;10(1):258-274

Department of Thoracic Medicine & Surgery, Physiology & Pediatrics, and Temple Lung Center, Lewis Katz School of Medicine at Temple University, Philadelphia, Pennsylvania, USA.

Background: Effective clinical management of airway clot and fibrinous cast formation of severe inhalational smoke-induced acute lung injury (ISALI) is lacking. Aerosolized delivery of tissue plasminogen activator (tPA) is confounded by airway bleeding; single-chain urokinase plasminogen activator (scuPA) moderated this adverse effect and supported transient improvement in gas exchange and lung mechanics. However, neither aerosolized plasminogen activator (PA) yielded durable improvements in physiologic responses or reduction in cast burden. Here, we hypothesized that perfluorochemical (PFC) liquids would facilitate PA distribution and sustain improvements in physiologic outcomes in ISALI.

Methods: Spontaneously breathing adult sheep (n = 36) received anesthesia and analgesia and were instrumented, exposed to cotton smoke inhalation, and supported by mechanical ventilation for 48 h. Groups (n = 6/group) were studied without supplemental treatment, or, starting 4 h post injury, they received intratracheal low volume (8 mL) PFC liquid alone or a dose range of tPA/PFC or scuPA/PFC suspensions (4 or 8 mg in 8 mL PFC) every 8 h. Outcomes were evaluated by sequential measurements of cardiopulmonary parameters, lung histomorphology, and biochemical analyses of bronchoalveolar lavage fluid.

Results: Dose-response and PA-type comparisons of outcomes demonstrated sustained superiority with low-volume PFC suspensions of scuPA over tPA or PFC alone, favoring the highest dose of scuPA/PFC suspension over lower doses, without airway bleeding.

Conclusions: We propose that this improved profile over previously reported aerosolized delivery is likely related to improved dose distribution. Sustained salutary responses to scuPA/PFC suspension delivery in this translational model are encouraging and support the possibility that the observed outcomes could be of clinical importance.
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http://dx.doi.org/10.1002/ctm2.26DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7240845PMC
January 2020

HDAC inhibition improves cardiopulmonary function in a feline model of diastolic dysfunction.

Sci Transl Med 2020 01;12(525)

Cardiovascular Research Center, Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140, USA.

Heart failure with preserved ejection fraction (HFpEF) is a major health problem without effective therapies. This study assessed the effects of histone deacetylase (HDAC) inhibition on cardiopulmonary structure, function, and metabolism in a large mammalian model of pressure overload recapitulating features of diastolic dysfunction common to human HFpEF. Male domestic short-hair felines ( = 31, aged 2 months) underwent a sham procedure ( = 10) or loose aortic banding ( = 21), resulting in slow-progressive pressure overload. Two months after banding, animals were treated daily with suberoylanilide hydroxamic acid (b + SAHA, 10 mg/kg, = 8), a Food and Drug Administration-approved pan-HDAC inhibitor, or vehicle (b + veh, = 8) for 2 months. Echocardiography at 4 months after banding revealed that b + SAHA animals had significantly reduced left ventricular hypertrophy (LVH) ( < 0.0001) and left atrium size ( < 0.0001) versus b + veh animals. Left ventricular (LV) end-diastolic pressure and mean pulmonary arterial pressure were significantly reduced in b + SAHA ( < 0.01) versus b + veh. SAHA increased myofibril relaxation ex vivo, which correlated with in vivo improvements of LV relaxation. Furthermore, SAHA treatment preserved lung structure, compliance, blood oxygenation, and reduced perivascular fluid cuffs around extra-alveolar vessels, suggesting attenuated alveolar capillary stress failure. Acetylation proteomics revealed that SAHA altered lysine acetylation of mitochondrial metabolic enzymes. These results suggest that acetylation defects in hypertrophic stress can be reversed by HDAC inhibitors, with implications for improving cardiac structure and function in patients.
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http://dx.doi.org/10.1126/scitranslmed.aay7205DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7065257PMC
January 2020

Protein kinase C-delta inhibition is organ-protective, enhances pathogen clearance, and improves survival in sepsis.

FASEB J 2020 02 23;34(2):2497-2510. Epub 2019 Dec 23.

Center for Inflammation, Clinical and Translational Lung Research, Department of Thoracic Medicine and Surgery, Lewis Katz School of Medicine, Temple University, Philadelphia, PA.

Sepsis is a leading cause of morbidity and mortality in intensive care units. Previously, we identified Protein Kinase C-delta (PKCδ) as an important regulator of the inflammatory response in sepsis. An important issue in development of anti-inflammatory therapeutics is the risk of immunosuppression and inability to effectively clear pathogens. In this study, we investigated whether PKCδ inhibition prevented organ dysfunction and improved survival without compromising pathogen clearance. Sprague Dawley rats underwent sham surgery or cecal ligation and puncture (CLP) to induce sepsis. Post-surgery, PBS or a PKCδ inhibitor (200µg/kg) was administered intra-tracheally (IT). At 24 hours post-CLP, there was evidence of lung and kidney dysfunction. PKCδ inhibition decreased leukocyte influx in these organs, decreased endothelial permeability, improved gas exchange, and reduced blood urea nitrogen/creatinine ratios indicating organ protection. PKCδ inhibition significantly decreased bacterial levels in the peritoneal cavity, spleen and blood but did not exhibit direct bactericidal properties. Peritoneal chemokine levels, neutrophil numbers, or macrophage phenotypes were not altered by PKCδ inhibition. Peritoneal macrophages isolated from PKCδ inhibitor-treated septic rats demonstrated increased bacterial phagocytosis. Importantly, PKCδ inhibition increased survival. Thus, PKCδ inhibition improved survival and improved survival was associated with increased phagocytic activity, enhanced pathogen clearance, and decreased organ injury.
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http://dx.doi.org/10.1096/fj.201900897RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7568544PMC
February 2020

miR-200 family members reduce senescence and restore idiopathic pulmonary fibrosis type II alveolar epithelial cell transdifferentiation.

ERJ Open Res 2019 Oct 16;5(4). Epub 2019 Dec 16.

Pulmonology Dept, University Hospital of Cattinara, Trieste, Italy.

Rationale: Alveolar type II (ATII) cells act as adult stem cells contributing to alveolar type I (ATI) cell renewal and play a major role in idiopathic pulmonary fibrosis (IPF), as supported by familial cases harbouring mutations in genes specifically expressed by these cells. During IPF, ATII cells lose their regenerative potential and aberrantly express pathways contributing to epithelial-mesenchymal transition (EMT). The microRNA miR-200 family is downregulated in IPF, but its effect on human IPF ATII cells remains unproven. We wanted to 1) evaluate the characteristics and transdifferentiating ability of IPF ATII cells, and 2) test whether miR-200 family members can rescue the regenerative potential of fibrotic ATII cells.

Methods: ATII cells were isolated from control or IPF lungs and cultured in conditions promoting their transdifferentiation into ATI cells. Cells were either phenotypically monitored over time or transfected with miR-200 family members to evaluate the microRNA effect on the expression of transdifferentiation, senescence and EMT markers.

Results: IPF ATII cells show a senescent phenotype (p16 and p21), overexpression of EMT (ZEB1/2) and impaired expression of ATI cell markers (AQP5 and HOPX) after 6 days of culture in differentiating medium. Transfection with certain miR-200 family members (particularly miR-200b-3p and miR-200c-3p) reduced senescence marker expression and restored the ability to transdifferentiate into ATI cells.

Conclusions: We demonstrated that ATII cells from IPF patients express senescence and EMT markers, and display a reduced ability to transdifferentiate into ATI cells. Transfection with certain miR-200 family members rescues this phenotype, reducing senescence and restoring transdifferentiation marker expression.
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http://dx.doi.org/10.1183/23120541.00138-2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6911923PMC
October 2019

Yap/Taz regulate alveolar regeneration and resolution of lung inflammation.

J Clin Invest 2019 04 15;129(5):2107-2122. Epub 2019 Apr 15.

Department of Pharmacology, Center for Translational Medicine, Temple University Lewis Katz School of Medicine, Philadelphia, Pennsylvania, USA.

Alveolar epithelium plays a pivotal role in protecting the lungs from inhaled infectious agents. Therefore, the regenerative capacity of the alveolar epithelium is critical for recovery from these insults in order to rebuild the epithelial barrier and restore pulmonary functions. Here, we show that sublethal infection of mice with Streptococcus pneumoniae, the most common pathogen of community-acquired pneumonia, led to exclusive damage in lung alveoli, followed by alveolar epithelial regeneration and resolution of lung inflammation. We show that surfactant protein C-expressing (SPC-expressing) alveolar epithelial type II cells (AECIIs) underwent proliferation and differentiation after infection, which contributed to the newly formed alveolar epithelium. This increase in AECII activities was correlated with increased nuclear expression of Yap and Taz, the mediators of the Hippo pathway. Mice that lacked Yap/Taz in AECIIs exhibited prolonged inflammatory responses in the lung and were delayed in alveolar epithelial regeneration during bacterial pneumonia. This impaired alveolar epithelial regeneration was paralleled by a failure to upregulate IκBa, the molecule that terminates NF-κB-mediated inflammatory responses. These results demonstrate that signals governing resolution of lung inflammation were altered in Yap/Taz mutant mice, which prevented the development of a proper regenerative niche, delaying repair and regeneration of alveolar epithelium during bacterial pneumonia.
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http://dx.doi.org/10.1172/JCI125014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6486331PMC
April 2019

Regenerative therapy based on miRNA-302 mimics for enhancing host recovery from pneumonia caused by .

Proc Natl Acad Sci U S A 2019 04 10;116(17):8493-8498. Epub 2019 Apr 10.

Department of Microbiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104;

Bacterial pneumonia remains a leading cause of morbidity and mortality worldwide. A defining feature of pneumonia is lung injury, leading to protracted suffering and vulnerability long after bacterial clearance. Little is known about which cells are damaged during bacterial pneumonia and if the regenerative process can be harnessed to promote tissue repair and host recovery. Here, we show that infection of mice with () caused substantial damage to alveolar epithelial cells (AEC), followed by a slow process of regeneration. Concurrent with AEC regeneration, the expression of miRNA-302 is elevated in AEC. Treatment of -infected mice with miRNA-302 mimics improved lung functions, host recovery, and survival. miRNA-302 mediated its therapeutic effects, not by inhibiting apoptosis and preventing damage, but by promoting proliferation of local epithelial progenitor cells to regenerate AEC. These results demonstrate the ability of microRNA-based therapy to promote AEC regeneration and enhance host recovery from bacterial pneumonia.
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http://dx.doi.org/10.1073/pnas.1818522116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6486708PMC
April 2019

Neuroprotective Effects of Nasopharyngeal Perfluorochemical Cooling in a Rat Model of Subarachnoid Hemorrhage.

World Neurosurg 2019 Jan 26;121:e481-e492. Epub 2018 Sep 26.

Department of Physiology, Translational and Clinical Lung Research, Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania, USA; Department of Pediatrics, Translational and Clinical Lung Research, Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania, USA; Department of Thoracic Medicine and Surgery, Translational and Clinical Lung Research, Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania, USA; Center for Inflammation, Translational and Clinical Lung Research, Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania, USA.

Objective: Subarachnoid hemorrhage (SAH) frequently results in severe morbidity, even mortality. Hypothermia is known to have a neuroprotective effect in ischemic injuries. The aim of this study was to determine whether nasopharyngeal (NP) perfluorochemical (PFC) cooling could be used in a rat model of SAH model for neuroprotection.

Methods: SAH was induced in 16 male Sprague-Dawley rats by cisterna magna injection of 0.3 mL autologous blood. Vital signs, temperatures, cerebral blood flow (CBF), and brain histology were assessed. Brain cooling was performed on the treatment group using the NP-PFC method starting from 20 minutes after SAH.

Results: No SAH-related deaths were observed in either group. SAH caused an immediate decrease in mean arterial pressure (17.0% ± 4.90% below baseline values). SAH induction caused a significant and rapid decrease in CBF from baseline (approximately -65%, ranging from -32% to -85%) in both hemispheres. In the left hemisphere, cooling facilitated the return of CBF to baseline values within 20 minutes of treatment with further increase in CBF that stabilized by the 2 hours after injury time point. Quantitative immunohistochemistry showed that there were significantly more NeuN-positive cells in the cortex and significantly fewer IBA-1-positive microglia and glial fibrillary acidic protein-positive astrocytes cells in both cortex and hippocampus in the animals that received NP-PFC cooling compared with no treatment, reflecting preserved neuronal integrity and reduced inflammation.

Conclusions: The data from this study indicate that local hypothermia by NP-PFC cooling supports return of CBF and neuronal integrity and suppresses the inflammatory response in SAH, suggestive of a promising neuroprotective approach in management of SAH.
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http://dx.doi.org/10.1016/j.wneu.2018.09.142DOI Listing
January 2019

Nebulization of Single-Chain Tissue-Type and Single-Chain Urokinase Plasminogen Activator for Treatment of Inhalational Smoke-Induced Acute Lung Injury.

J Drug Deliv Sci Technol 2018 Aug 30;48:19-27. Epub 2018 Apr 30.

The University of Texas at Austin, College of Pharmacy, Division of Molecular Pharmaceutics and Drug Delivery, Austin, TX, USA.

Single-chain tissue-type plasminogen activator (sctPA) and single-chain urokinase plasminogen activator (scuPA) have attracted interest as enzymes for the treatment of inhalational smoke-induced acute lung injury (ISALI). In this study, the pulmonary delivery of commercial human sctPA and lyophilized scuPA and their reconstituted solution forms were demonstrated using vibrating mesh nebulizers (Aeroneb® Pro (active) and EZ Breathe® (passive)). Both the Aeroneb® Pro and EZ Breathe® vibrating mesh nebulizers produced atomized droplets of protein solution of similar size of less than about 5 μm, which is appropriate for pulmonary delivery. Enzymatic activities of scuPA and of sctPA were determined after nebulization and both remained stable (88.0% and 93.9%). Additionally, the enzymatic activities of sctPA and tcuPA were not significantly affected by excipients, lyophilization or reconstitution conditions. The results of these studies support further development of inhaled formulations of fibrinolysins for delivery to the lungs following smoke-induced acute pulmonary injury.
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http://dx.doi.org/10.1016/j.jddst.2018.04.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6095669PMC
August 2018

Lack of durable protection against cotton smoke-induced acute lung injury in sheep by nebulized single chain urokinase plasminogen activator or tissue plasminogen activator.

Clin Transl Med 2018 Jun 18;7(1):17. Epub 2018 Jun 18.

The Department of Cellular and Molecular Biology and the Texas Lung Institute, The University of Texas Health Science Center at Tyler, 11927 US HWY 271, Tyler, TX, 75708, USA.

Background: Airway fibrin casts are clinically important complications of severe inhalational smoke-induced acute lung injury (ISIALI) for which reliable evidence-based therapy is lacking. Nebulized anticoagulants or a tissue plasminogen activator; tPA, has been advocated, but airway bleeding is a known and lethal potential complication. We posited that nebulized delivery of single chain urokinase plasminogen activator, scuPA, is well-tolerated and improves physiologic outcomes in ISIALI. To test this hypothesis, we nebulized scuPA or tPA and delivered these agents every 4 h to sheep with cotton smoke induced ISIALI that were ventilated by either adaptive pressure ventilation/controlled mandatory ventilation (APVcmv; Group 1, n = 14) or synchronized controlled mandatory ventilation (SCMV)/limited suctioning; Group 2, n = 32). Physiologic readouts of acute lung injury included arterial blood gas analyses, PaO/FiO ratios, peak and plateau airway pressures, lung resistance and static lung compliance. Lung injury was further assessed by histologic scoring. Biochemical analyses included determination of antigenic and enzymographic uPA and tPA levels, plasminogen activator and plasminogen activator inhibitor-1 activities and D-dimer in bronchoalveolar lavage (BAL). Plasma levels of uPA, tPA antigens, D-dimers and α-macroglobulin-uPA complex levels were also assessed.

Results: In Group 1, tPA at the 2 mg dose was ineffective, but at 4 mg tPA or scuPA, the PaO/FiO ratios, peak/plateau pressures improved during evolving injury (p < 0.01) without significant differences at 48 h. To improve delivery of the interventions, the experiments were repeated in Group 2 with limited suctioning/SCMV, which generally increased PAs in (BAL). In Group 2, tPA was ineffective, but scuPA (4 or 8 mg) improved physiologic outcomes (p < 0.01) and plateau pressures remained lower at 48 h. Airway bleeding occurred at 8 mg tPA. BAL plasminogen activator (PA) levels positively correlated with physiologic outcomes at 48 h.

Conclusions: Physiologic outcomes improved in sheep in which better delivery of the PAs occurred. The benefits of nebulized scuPA were achieved without airway bleeding associated with tPA, but were transient and largely abrogated at 48 h, in part attributable to the progression and severity of ISIALI.
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http://dx.doi.org/10.1186/s40169-018-0196-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6006005PMC
June 2018

A Feline HFpEF Model with Pulmonary Hypertension and Compromised Pulmonary Function.

Sci Rep 2017 11 29;7(1):16587. Epub 2017 Nov 29.

Temple University Lewis Katz School of Medicine, Cardiovascular Research Center, Philadelphia, PA, United States.

Heart Failure with preserved Ejection Fraction (HFpEF) represents a major public health problem. The causative mechanisms are multifactorial and there are no effective treatments for HFpEF, partially attributable to the lack of well-established HFpEF animal models. We established a feline HFpEF model induced by slow-progressive pressure overload. Male domestic short hair cats (n = 20), underwent either sham procedures (n = 8) or aortic constriction (n = 12) with a customized pre-shaped band. Pulmonary function, gas exchange, and invasive hemodynamics were measured at 4-months post-banding. In banded cats, echocardiography at 4-months revealed concentric left ventricular (LV) hypertrophy, left atrial (LA) enlargement and dysfunction, and LV diastolic dysfunction with preserved systolic function, which subsequently led to elevated LV end-diastolic pressures and pulmonary hypertension. Furthermore, LV diastolic dysfunction was associated with increased LV fibrosis, cardiomyocyte hypertrophy, elevated NT-proBNP plasma levels, fluid and protein loss in pulmonary interstitium, impaired lung expansion, and alveolar-capillary membrane thickening. We report for the first time in HFpEF perivascular fluid cuff formation around extra-alveolar vessels with decreased respiratory compliance. Ultimately, these cardiopulmonary abnormalities resulted in impaired oxygenation. Our findings support the idea that this model can be used for testing novel therapeutic strategies to treat the ever growing HFpEF population.
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http://dx.doi.org/10.1038/s41598-017-15851-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5707379PMC
November 2017

Acute right heart failure after hemorrhagic shock and trauma pneumonectomy-a management approach: A blinded randomized controlled animal trial using inhaled nitric oxide.

J Trauma Acute Care Surg 2017 02;82(2):243-251

From the Department of Surgery at Temple University Hospital (A.L.L., L.O.S., A.G., A.P., T.S.), Cardiovascular Research Center (T.E.S., M.W., R.M.B.), Department of Physiology (T.E.S., M.W., R.M.B., L.A.P., J.W., M.R.W.), Department of Thoracic Medicine and Surgery (L.A.P., J.W., M.R.W.), Center for Inflammation, Translational and Clinical Lung Research (L.A.P., J.W., M.R.W.), CENTRe: Collaborative for Environmental and Neonatal Therapeutics Research (L.A.P., J.W., M.R.W.), and Temple Lung Center of Lewis Katz School of Medicine at Temple University Philadelphia, PA (L.A.P., J.W., M.R.W.).

Background: Hemorrhagic shock and pneumonectomy causes an acute increase in pulmonary vascular resistance (PVR). The increase in PVR and right ventricular (RV) afterload leads to acute RV failure, thus reducing left ventricular (LV) preload and output. Inhaled nitric oxide (iNO) lowers PVR by relaxing pulmonary arterial smooth muscle without remarkable systemic vascular effects. We hypothesized that with hemorrhagic shock and pneumonectomy, iNO can be used to decrease PVR and mitigate right heart failure.

Methods: A hemorrhagic shock and pneumonectomy model was developed using sheep. Sheep received lung protective ventilatory support and were instrumented to serially obtain measurements of hemodynamics, gas exchange, and blood chemistry. Heart function was assessed with echocardiography. After randomization to study gas of iNO 20 ppm (n = 9) or nitrogen as placebo (n = 9), baseline measurements were obtained. Hemorrhagic shock was initiated by exsanguination to a target of 50% of the baseline mean arterial pressure. The resuscitation phase was initiated, consisting of simultaneous left pulmonary hilum ligation, via median sternotomy, infusion of autologous blood and initiation of study gas. Animals were monitored for 4 hours.

Results: All animals had an initial increase in PVR. PVR remained elevated with placebo; with iNO, PVR decreased to baseline. Echo showed improved RV function in the iNO group while it remained impaired in the placebo group. After an initial increase in shunt and lactate and decrease in SvO2, all returned toward baseline in the iNO group but remained abnormal in the placebo group.

Conclusion: These data indicate that by decreasing PVR, iNO decreased RV afterload, preserved RV and LV function, and tissue oxygenation in this hemorrhagic shock and pneumonectomy model. This suggests that iNO may be a useful clinical adjunct to mitigate right heart failure and improve survival when trauma pneumonectomy is required.
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http://dx.doi.org/10.1097/TA.0000000000001325DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5315536PMC
February 2017

A Novel Approach for Ovine Primary Alveolar Epithelial Type II Cell Isolation and Culture from Fresh and Cryopreserved Tissue Obtained from Premature and Juvenile Animals.

PLoS One 2016 21;11(3):e0152027. Epub 2016 Mar 21.

Department of Thoracic Medicine and Surgery, Katz School of Medicine at Temple University, Philadelphia, PA, United States of America.

The in vivo ovine model provides a clinically relevant platform to study cardiopulmonary mechanisms and treatments of disease; however, a robust ovine primary alveolar epithelial type II (ATII) cell culture model is lacking. The objective of this study was to develop and optimize ovine lung tissue cryopreservation and primary ATII cell culture methodologies for the purposes of dissecting mechanisms at the cellular level to elucidate responses observed in vivo. To address this, we established in vitro submerged and air-liquid interface cultures of primary ovine ATII cells isolated from fresh or cryopreserved lung tissues obtained from mechanically ventilated sheep (128 days gestation-6 months of age). Presence, abundance, and mRNA expression of surfactant proteins was assessed by immunocytochemistry, Western Blot, and quantitative PCR respectively on the day of isolation, and throughout the 7 day cell culture study period. All biomarkers were significantly greater from cells isolated from fresh than cryopreserved tissue, and those cultured in air-liquid interface as compared to submerged culture conditions at all time points. Surfactant protein expression remained in the air-liquid interface culture system while that of cells cultured in the submerged system dissipated over time. Despite differences in biomarker magnitude between cells isolated from fresh and cryopreserved tissue, cells isolated from cryopreserved tissue remained metabolically active and demonstrated a similar response as cells from fresh tissue through 72 hr period of hyperoxia. These data demonstrate a cell culture methodology using fresh or cryopreserved tissue to support study of ovine primary ATII cell function and responses, to support expanded use of biobanked tissues, and to further understanding of mechanisms that contribute to in vivo function of the lung.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0152027PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4801353PMC
July 2016

Enhanced Re-Endothelialization of Decellularized Rat Lungs.

Tissue Eng Part C Methods 2016 05 1;22(5):439-50. Epub 2016 Apr 1.

1 Department of Bioengineering, College of Engineering, Temple University , Philadelphia, Pennsylvania.

Decellularized lung tissue has been recognized as a potential platform to engineer whole lung organs suitable for transplantation or for modeling a variety of lung diseases. However, many technical hurdles remain before this potential may be fully realized. Inability to efficiently re-endothelialize the pulmonary vasculature with a functional endothelium appears to be the primary cause of failure of recellularized lung scaffolds in early transplant studies. Here, we present an optimized approach for enhanced re-endothelialization of decellularized rodent lung scaffolds with rat lung microvascular endothelial cells (ECs). This was achieved by adjusting the posture of the lung to a supine position during cell seeding through the pulmonary artery. The supine position allowed for significantly more homogeneous seeding and better cell retention in the apex regions of all lobes than the traditional upright position, especially in the right upper and left lobes. Additionally, the supine position allowed for greater cell retention within large diameter vessels (proximal 100-5000 μm) than the upright position, with little to no difference in the small diameter distal vessels. EC adhesion in the proximal regions of the pulmonary vasculature in the decellularized lung was dependent on the binding of EC integrins, specifically α1β1, α2β1, and α5β1 integrins to, respectively, collagen type-I, type-IV, and fibronectin in the residual extracellular matrix. Following in vitro maturation of the seeded constructs under perfusion culture, the seeded ECs spread along the vascular wall, leading to a partial reestablishment of endothelial barrier function as inferred from a custom-designed leakage assay. Our results suggest that attention to cellular distribution within the whole organ is of paramount importance for restoring proper vascular function.
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http://dx.doi.org/10.1089/ten.TEC.2016.0012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4870653PMC
May 2016

The Effects of Lung Protective Ventilation or Hypercapnic Acidosis on Gas Exchange and Lung Injury in Surfactant Deficient Rabbits.

PLoS One 2016 3;11(2):e0147807. Epub 2016 Feb 3.

Division of Neonatology and Pediatric Critical Care, Department of Pediatrics, Children's Hospital, Ulm University, 89070 Ulm, Germany.

Background: Permissive hypercapnia has been shown to reduce lung injury in subjects with surfactant deficiency. Experimental studies suggest that hypercapnic acidosis by itself rather than decreased tidal volume may be a key protective factor.

Objectives: To study the differential effects of a lung protective ventilatory strategy or hypercapnic acidosis on gas exchange, hemodynamics and lung injury in an animal model of surfactant deficiency.

Methods: 30 anesthetized, surfactant-depleted rabbits were mechanically ventilated (FiO2 = 0.8, PEEP = 7cmH2O) and randomized into three groups: Normoventilation-Normocapnia (NN)-group: tidal volume (Vt) = 7.5 ml/kg, target PaCO2 = 40 mmHg; Normoventilation-Hypercapnia (NH)-group: Vt = 7.5 ml/kg, target PaCO2 = 80 mmHg by increasing FiCO2; and a Hypoventilation-Hypercapnia (HH)-group: Vt = 4.5 ml/kg, target PaCO2 = 80 mmHg. Plasma lactate and interleukin (IL)-8 were measured every 2 h. Animals were sacrificed after 6 h to perform bronchoalveolar lavage (BAL), to measure lung wet-to-dry weight, lung tissue IL-8, and to obtain lung histology.

Results: PaO2 was significantly higher in the HH-group compared to the NN-group (p<0.05), with values of the NH-group between the HH- and NN-groups. Other markers of lung injury (wet-dry-weight, BAL-Protein, histology-score, plasma-IL-8 and lung tissue IL-8) resulted in significantly lower values for the HH-group compared to the NN-group and trends for the NH-group towards lower values compared to the NN-group. Lactate was significantly lower in both hypercapnia groups compared to the NN-group.

Conclusion: Whereas hypercapnic acidosis may have some beneficial effects, a significant effect on lung injury and systemic inflammatory response is dependent upon a lower tidal volume rather than resultant arterial CO2 tensions and pH alone.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0147807PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4739580PMC
July 2016

Ex vivo ovine model for pediatric flexible endoscopy training.

Int J Pediatr Otorhinolaryngol 2015 Dec 23;79(12):2196-9. Epub 2015 Oct 23.

Department of Pediatrics, Temple University School of Medicine, United States; Department of Medicine, Temple University School of Medicine, United States; Department of Physiology, Temple University School of Medicine, United States; Department of Thoracic Medicine and Surgery, Temple University School of Medicine, United States; The Centers for Inflammation, Translation and Clinical Lung Research, Temple University School of Medicine, United States; CENTRe: Collaborative for Environmental and Neonatal Therapeutics Research, Temple University School of Medicine, United States.

Objectives: Medical students and residents in training have limited opportunities to develop pediatric endoscopy skills and would benefit from a realistic simulation model. We sought to develop such a model for flexible endoscopy using fresh head and neck tissue from young sheep.

Methods: Tissue was collected from pre-pubescent sheep (n=5; mean age: 4 months; mean mass: 28kg) following humane euthanasia at the end of an in vivo protocol. No live animals were used in this study. The head and neck of the sheep were disarticulated 4-6cm above the sternal notch and stored at 5°C for 1-5 days. With the preparation was supported in supine position, flexible nasopharyngolaryngoscopy and transnasal endoscopic intubation were performed with video recording.

Results: Five sheep were studied. Endoscopy was performed by a medical student under direct supervision by a pediatric otolaryngologist. Differences between ovine and human pediatric airway anatomy were defined.

Conclusions: Despite variations in proportion and structure, the experience of passing a flexible nasopharyngoscope through a sheep's airway is remarkably similar to pediatric endoscopy. The nasal anatomy is elongated, but very much like a child's in terms of anatomy, color and texture. The tactile feedback is nearly identical. Annoying secretions and their associated "whiteout" phenomena nicely simulate these challenges in pediatric endoscopy. When performing transnasal intubation, navigating to the larynx and advancing an endotracheal tube under guidance have the look and feel of the pediatric procedure. Issues of cost, availability, risk of zoonotic infection, and ethics are discussed.
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http://dx.doi.org/10.1016/j.ijporl.2015.10.002DOI Listing
December 2015

Biodistribution and Efficacy of Targeted Pulmonary Delivery of a Protein Kinase C-δ Inhibitory Peptide: Impact on Indirect Lung Injury.

J Pharmacol Exp Ther 2015 Oct 4;355(1):86-98. Epub 2015 Aug 4.

Center for Inflammation, Clinical and Translational Lung Research (M.J.M., P.A.K., J.W., M.K., S.T.B., M.R.W., L.E.K.), Department of Physiology (M.J.M., P.A.K., J.W., S.T.B., M.R.W., L.E.K.), Sol Sherry Thrombosis Research Center (M.J.M., L.C.K., L.E.K.), Departments of Pediatrics and Medicine (M.R.W.), and Department of Radiology (L.C.K.), Temple University School of Medicine, Philadelphia, Pennsylvania

Sepsis and sepsis-induced lung injury remain a leading cause of death in intensive care units. We identified protein kinase C-δ (PKCδ) as a critical regulator of the acute inflammatory response and demonstrated that PKCδ inhibition was lung-protective in a rodent sepsis model, suggesting that targeting PKCδ is a potential strategy for preserving pulmonary function in the setting of indirect lung injury. In this study, whole-body organ biodistribution and pulmonary cellular distribution of a transactivator of transcription (TAT)-conjugated PKCδ inhibitory peptide (PKCδ-TAT) was determined following intratracheal (IT) delivery in control and septic [cecal ligation and puncture (CLP)] rats to ascertain the impact of disease pathology on biodistribution and efficacy. There was negligible lung uptake of radiolabeled peptide upon intravenous delivery [<1% initial dose (ID)], whereas IT administration resulted in lung retention of >65% ID with minimal uptake in liver or kidney (<2% ID). IT delivery of a fluorescent-tagged (tetramethylrhodamine-PKCδ-TAT) peptide demonstrated uniform spatial distribution and cellular uptake throughout the peripheral lung. IT delivery of PKCδ-TAT at the time of CLP surgery significantly reduced PKCδ activation (tyrosine phosphorylation, nuclear translocation and cleavage) and acute lung inflammation, resulting in improved lung function and gas exchange. Importantly, peptide efficacy was similar when delivered at 4 hours post-CLP, demonstrating therapeutic relevance. Conversely, spatial lung distribution and efficacy were significantly impaired at 8 hours post-CLP, which corresponded to marked histopathological progression of lung injury. These studies establish a functional connection between peptide spatial distribution, inflammatory histopathology in the lung, and efficacy of this anti-inflammatory peptide.
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http://dx.doi.org/10.1124/jpet.115.224832DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4576664PMC
October 2015

The orl rat is more responsive to methacholine challenge than wild type.

Pulm Pharmacol Ther 2014 Dec 16;29(2):199-208. Epub 2014 Sep 16.

Alfred I. duPont Hospital for Children, Nemours Lung Center, Wilmington, DE 19803, United States; Temple University School of Medicine, Department of Physiology, Philadelphia, PA 19140, United States; Temple University School of Medicine, Department of Pediatrics, Philadelphia, PA 19140, United States.

Background: This study presents an animal model of native airway hyperresponsiveness (AHR). AHR is a fundamental aspect of asthma and reflects an abnormal response characterized by airway narrowing following exposure to a wide variety of non-immunological stimuli. Undescended testis (UDT) is one of the most common male congenital anomalies. The orl rat is a Long Evans substrain with inherited UDT. Since boys born with congenital UDT are more likely to manifest asthma symptoms, the main aim of this study was to investigate the alternative hypothesis that orl rats have greater AHR to a methacholine aerosol challenge than wild type rats.

Methods: Long Evans wild type (n = 9) and orl (n = 13) rats were anesthetized, tracheostomized, and mechanically ventilated at 4 weeks of age. Escalating concentrations of inhaled methacholine were delivered. The methacholine potency and efficacy in the strains were measured. Respiratory resistance was the primary endpoint. After the final methacholine aerosol challenge, the short-acting β2-adrenoceptor agonist albuterol was administered as an aerosol and lung/diaphragm tissues were assayed for interleukin (IL)-4, IL-6, and tumor necrosis factor (TNF)-α. Histological and histomorphometrical analyses were performed.

Results: The methacholine concentration-response curve in the orl group indicated increased sensitivity, hyperreactivity, and exaggerated maximal response in comparison with the wild type group, indicating that orl rats had abnormally greater AHR responses to methacholine. Histological findings in orl rats showed the presence of eosinophils, unlike wild type rats. β2-Adrenoceptor agonist intervention resulted in up-regulation of IL-4 diaphragmatic levels and down-regulation of IL-4 and IL-6 in the lungs of orl rats.

Conclusion: orl rats had greater AHR than wild type rats during methacholine challenge, with higher IL-4 levels in diaphragmatic tissue homogenates. Positive immunostaining for IL-4 was detected in lung and diaphragmatic tissue in both strains. This model offers advantages over other pre-clinical murine models for studying potential mechanistic links between cryptorchidism and asthma. This animal model may be useful for further testing of compounds/therapeutics options for treating AHR.
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http://dx.doi.org/10.1016/j.pupt.2014.09.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4323580PMC
December 2014

High flow nasal cannula (HFNC) with Heliox decreases diaphragmatic injury in a newborn porcine lung injury model.

Pediatr Pulmonol 2014 Dec 5;49(12):1214-22. Epub 2014 Feb 5.

Neonatology, Alfred I. duPont Hospital for Children, Wilmington, Delaware; Neonatology, Thomas Jefferson University Hospital, Philadelphia, Pennsylvania.

Background: High flow nasal cannula (HFNC) improves ventilation by washing out nasopharyngeal dead space while delivering oxygen. Heliox (helium-oxygen gas mixture), a low-density gas mixture, decreases resistance to airflow, reduces the work of breathing, and facilitates distribution of inspired gas. Excessive lung work and potential injury increases the workload on the immature diaphragm predisposing the muscle to fatigue, and can lead to inflammatory and oxidative stress, thereby contributing to impaired diaphragmatic function. We tested the hypothesis that HFNC with Heliox will decrease the work of breathing thereby unloading the neonatal diaphragm, and potentially reducing diaphragmatic injury.

Methods: Spontaneously breathing neonatal pigs were randomized to Nitrox (nitrogen-oxygen gas mixture) or Heliox, and studied over 4 hr following oleic acid injury. Gas exchange, pulmonary mechanics indices, and systemic markers of inflammation were measured serially. Diaphragm inflammation biomarkers and histology for muscle injury were assessed at termination.

Results: Heliox breathing animals demonstrated decreased respiratory load and work of breathing with lower pressure-rate product, lower labored breathing index, and lower levels of diaphragmatic inflammatory markers, and muscle injury score as compared to Nitrox.

Conclusion: These results suggest that HFNC with Heliox is a useful adjunct to attenuate diaphragmatic fatigue in the presence of lung injury by unloading the diaphragm, resulting in a more efficient breathing pattern, and decreased diaphragm injury.
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http://dx.doi.org/10.1002/ppul.23000DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4122654PMC
December 2014

Pulmonary endothelial protein kinase C-delta (PKCδ) regulates neutrophil migration in acute lung inflammation.

Am J Pathol 2014 Jan 6;184(1):200-13. Epub 2013 Nov 6.

Department of Physiology, Temple University School of Medicine, Philadelphia, Pennsylvania; Center for Inflammation, Translational and Clinical Lung Research, Temple University School of Medicine, Philadelphia, Pennsylvania; Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylvania. Electronic address:

Excessive neutrophil migration across the pulmonary endothelium into the lung and release of oxidants and proteases are key elements in pathogenesis of acute lung injury. Previously, we identified protein kinase C-delta (PKCδ) as an important regulator of proinflammatory signaling in human neutrophils and demonstrated that intratracheal instillation of a TAT-conjugated PKCδ inhibitory peptide (PKCδ-TAT) is lung protective in a rat model of sepsis-induced indirect pulmonary injury (cecal ligation and puncture). In the present study, intratracheal instillation of this PKCδ inhibitor resulted in peptide distribution throughout the lung parenchyma and pulmonary endothelium and decreased neutrophil influx, with concomitant attenuation of sepsis-induced endothelial ICAM-1 and VCAM-1 expression in this model. To further delineate the role of PKCδ in regulating neutrophil migration, we used an in vitro transmigration model with human pulmonary microvascular endothelial cells (PMVECs). Consistent with in vivo findings, inhibition of PMVEC PKCδ decreased IL-1β-mediated neutrophil transmigration. PKCδ regulation was stimulus-dependent; PKCδ was required for transmigration mediated by IL-1β and fMLP (integrin-dependent), but not IL-8 (integrin-independent). PKCδ was essential for IL-1β-mediated neutrophil adherence and NF-κB-dependent expression of ICAM-1 and VCAM-1. In PMVECs, IL-1β-mediated production of ROS and activation of redox-sensitive NF-κB were PKCδ dependent, suggesting an upstream signaling role. Thus, PKCδ has an important role in regulating neutrophil-endothelial cell interactions and recruitment to the inflamed lung.
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http://dx.doi.org/10.1016/j.ajpath.2013.09.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3873483PMC
January 2014

Aerosolized KL4 surfactant improves short-term survival and gas exchange in spontaneously breathing newborn pigs with hydrochloric acid-induced acute lung injury.

Pediatr Pulmonol 2014 May 3;49(5):482-9. Epub 2013 Sep 3.

Infant Diagnostic and Research Center, Children's Hospitals and Clinics of Minnesota, St. Paul, Minnesota; Department of Pediatrics-Neonatology, University of Minnesota, Minneapolis, Minnesota.

Background: Surfactant therapy may be beneficial in acute lung injury (ALI). In spontaneously breathing newborn pigs with ALI supported with continuous positive airway pressure (CPAP), we evaluated the hypothesis that aerosolized KL4 surfactant (AERO KL4 S) would provide a similar therapeutic effect as intratracheal KL4 surfactant (ETT KL4 S) when compared to controls.

Methods: We randomized pigs with HCl-induced ALI to: (1) 175 mg/kg KL4 surfactant via endotracheal tube (ETT); (2) AERO KL4 S (22.5 mg/min phospholipid) for 60 min via continuous positive airway pressure (CPAP); or (3) sham procedure on CPAP. We obtained physiologic data and arterial blood gases throughout the 3-hr study. At study end, lungs were excised for analysis of interleukin-8 (IL-8), myeloperoxidase (MPO) levels and histomorphometric data.

Results: Pigs treated with ETT KL4 S and AERO KL4 S had improved survival and sustained pO2 compared to controls. The AERO KL4 S group had higher pH compared to controls. Lung IL-8 levels were lower in the AERO KL4 S group compared to controls. Histomorphometric analysis showed less hemorrhage in the ETT and AERO KL4 S groups compared to controls. The AERO KL4 S group had more open lung units per fixed-field than the ETT KL4 S or controls.

Conclusions: AERO KL4 S produced similar improvements in survival, physiology, inflammatory markers, and morphology as ETT KL4 S in an ALI model.
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http://dx.doi.org/10.1002/ppul.22844DOI Listing
May 2014

Lucinactant attenuates pulmonary inflammatory response, preserves lung structure, and improves physiologic outcomes in a preterm lamb model of RDS.

Pediatr Res 2012 Oct 20;72(4):375-83. Epub 2012 Jul 20.

Department of Physiology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.

Background: Acute inflammatory responses to supplemental oxygen and mechanical ventilation have been implicated in the pathophysiological sequelae of respiratory distress syndrome (RDS). Although surfactant replacement therapy (SRT) has contributed to lung stability, the effect on lung inflammation is inconclusive. Lucinactant contains sinapultide (KL4), a novel synthetic peptide that functionally mimics surfactant protein B, a protein with anti-inflammatory properties. We tested the hypothesis that lucinactant may modulate lung inflammatory response to mechanical ventilation in the management of RDS and may confer greater protection than animal-derived surfactants.

Methods: Preterm lambs (126.8 ± 0.2 SD d gestation) were randomized to receive lucinactant, poractant alfa, beractant, or no surfactant and studied for 4 h. Gas exchange and pulmonary function were assessed serially. Lung inflammation biomarkers and lung histology were assessed at termination.

Results: SRT improved lung compliance relative to no SRT without significant difference between SRT groups. Lucinactant attenuated lung and systemic inflammatory response, supported oxygenation at lower ventilatory requirements, and preserved lung structural integrity to a greater degree than either no SRT or SRT with poractant alfa or beractant.

Conclusion: These data suggest that early intervention with lucinactant may more effectively mitigate pulmonary pathophysiological sequelae of RDS than the animal-derived surfactants poractant alfa or beractant.
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http://dx.doi.org/10.1038/pr.2012.96DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3888789PMC
October 2012

Neonatal non-invasive respiratory support: physiological implications.

Pediatr Pulmonol 2012 Sep 6;47(9):837-47. Epub 2012 Jul 6.

Nemours Center for Pediatric Lung Research, Nemours/Alfred I. duPont Hospital for Children, Wilmington, DE 19803, USA.

The introduction of assisted ventilation for neonatal pulmonary insufficiency has resulted in the successful treatment of many previously fatal diseases. During the past three decades, refinement of invasive mechanical ventilation techniques has dramatically improved survival of many high-risk neonates. However, as with many advances in medicine, while mortality has been reduced, morbidity has increased in the surviving high-risk neonate. In this regard, introduction of assisted ventilation has been associated with chronic lung injury, also known as bronchopulmonary dysplasia. This disease, unknown prior to the appearance of mechanical ventilation, has produced a population of patients characterized by ventilator or oxygen dependence with serious accompanying pulmonary and neurodevelopmental morbidity. The purpose of this article is to review non-invasive respiratory support methodologies to address the physiologic mechanisms by which these methods may prevent the pathophysiologic effects of invasive mechanical ventilation.
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http://dx.doi.org/10.1002/ppul.22610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3762325PMC
September 2012

Hyperoxia during one lung ventilation: inflammatory and oxidative responses.

Pediatr Pulmonol 2012 Oct 19;47(10):979-86. Epub 2012 Mar 19.

Nemours Biomedical Research, Nemours/Alfred I. duPont Hospital for Children, Wilmington, DE19803, USA.

Background: It is common practice during one lung ventilation (OLV) to use 100% oxygen, although this may cause hyperoxia- and oxidative stress-related lung injury. We hypothesized that lower oxygen (FiO(2) ) during OLV will result in less inflammatory and oxidative lung injury and improved lung function.

Methods: Twenty pigs (8.88 ± 0.84 kg; 38 ± 4.6 days) were assigned to either the hyperoxia group (n = 10; FiO(2)  = 100%) or the normoxia group (n = 10; FiO(2)  < 50%). Both groups were subjected to 3 hr of OLV. Blood samples were tested for pro-inflammatory cytokines and lung tissue was tested for these cytokines and oxidative biomarkers.

Results: There were no differences between groups for partial pressure of CO(2) , tidal volume, end-tidal CO(2) , plasma cytokines, or respiratory compliance. Total respiratory resistance was greater in the hyperoxia group (P = 0.02). There were higher levels of TNF-α, IL-1β, and IL-6 in the lung homogenates of the hyperoxia group than in the normoxia group (P ≤ 0.01, 0.001, and 0.001, respectively). Myeloperoxidase and protein carbonyls (PC) were higher (P = 0.03 and P = 0.01, respectively) and superoxide dismutase (SOD) was lower in the lung homogenates of the hyperoxia group (P ≤ 0.001).

Conclusion: Higher myeloperoxidase, PC, and cytokine levels, and lower SOD availability indicate a greater degree of injury in the lungs of the hyperoxia animals, possibly from using 100% oxygen. In this translational study using a pig model, FiO(2)  ≤ 50% during OLV reduced hyperoxic injury and improved function in the lungs.
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http://dx.doi.org/10.1002/ppul.22517DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3888791PMC
October 2012

Brief mechanical ventilation impacts airway cartilage properties in neonatal lambs.

Pediatr Pulmonol 2012 Aug 13;47(8):763-70. Epub 2011 Dec 13.

McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA.

Ultrasound imaging allows in vivo assessment of tracheal kinetics and cartilage structure. To date, the impact of mechanical ventilation (MV) on extracellular matrix (ECM) in airway cartilage is unclear, but an indication of its functional and structural change may support the development of protective therapies. The objective of this study was to characterize changes in mechanical properties of the neonatal airway during MV with alterations in cartilage ECM. Trachea segments were isolated in a neonatal lamb model; ultrasound dimensions and pressure-volume relationships were measured on sham (no MV; n = 6) and MV (n = 7) airways for 4 hr. Tracheal cross-sections were harvested at 4 hr, tissues were fixed and stained, and Fourier transform infrared imaging spectroscopy (FT-IRIS) was performed. Over 4 hr of MV, bulk modulus (28%) and elastic modulus (282%) increased. The MV tracheae showed higher collagen, proteoglycan content, and collagen integrity (new tissue formation); whereas no changes were seen in the controls. These data are clinically relevant in that airway properties can be correlated with MV and changes in cartilage ECM. MV increases the in vivo dimensions of the trachea and is associated with evidence of airway tissue remodeling. Injury to the neonatal airway from MV may have relevance for the development of tracheomalacia. We demonstrated active airway tissue remodeling during MV using an FT-IRIS technique which identifies changes in ECM.
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http://dx.doi.org/10.1002/ppul.21616DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3362668PMC
August 2012

Perfluorochemical liquid-adenovirus suspensions enhance gene delivery to the distal lung.

Pulm Med 2011 18;2011:918036. Epub 2011 Aug 18.

The Cardiopulmonary Research Institute and Departments of Medicine and Pediatrics, SUNY Stony Brook School of Medicine, Winthrop University Hospital, Mineola, NY 11507, USA.

WE COMPARED LUNG DELIVERY METHODS OF RECOMBINANT ADENOVIRUS (RAD): (1) rAd suspended in saline, (2) rAd suspended in saline followed by a pulse-chase of a perfluorochemical (PFC) liquid mixture, and (3) a PFC-rAd suspension. Cell uptake, distribution, and temporal expression of rAd were examined using A549 cells, a murine model using luciferase bioluminescence, and histological analyses. Relative to saline, a 4X increase in transduction efficiency was observed in A549 cells exposed to PFC-rAd for 2-4 h. rAd transgene expression was improved in alveolar epithelial cells, and the level and distribution of luciferase expression when delivered in PFC-rAd suspensions consistently peaked at 24 h. These results demonstrate that PFC-rAd suspensions improve distribution and enhance rAd-mediated gene expression which has important implications in improving lung function by gene therapy.
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http://dx.doi.org/10.1155/2011/918036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3159382PMC
November 2011

Pregnancy and outcome of uterine allotransplantation and assisted reproduction in sheep.

J Minim Invasive Gynecol 2011 Mar-Apr;18(2):238-45

St John's Regional Medical Center, Oxnard, California 93030, USA.

This pilot study was performed from March 2008 through February 2010 to demonstrate that pregnancy can be achieved in a uterine allograft in the sheep model with the guidance of assisted reproductive technology. Uterine allotransplantation was performed in 12 sexually mature African sheep (Sudanese and Ethiopian). All animals underwent uterine transplantation via a minilaparotomy incision using a Mobius retractor device. A control group of pregnant Romney Marsh sheep with nontransplanted uteri were used to compare fetal development, uterine and placental histologic findings, and blood samples of progeny of the uterine transplant recipient sheep. Fetal size was obtained from ultrasound measurements during the early (crown-rump length) and late (biparietal diameter and abdominal circumference) gestational periods. The primary end point variables included preoperative and postoperative management, embryo transfer protocol, intraoperative assessments, and physiologic cardiopulmonary changes in the lamb during the first 5 hours of life. Four months after the initial uterine transplantation, 5 of 12 uterine allografts were considered candidates for the embryo transfer procedure. Fresh and frozen blastocyst donors were transferred accordingly to the remaining 5 uterine allografts via a minilaparotomy incision. Three of these resulted in pregnancies. One was an ectopic gestation, 1 sheep carried the pregnancy to 105 days, and 1 delivered a fully developed lamb from the transplanted uterus that was delivered via cesarean section. Neonatal lamb blood gas values and chemistry, gross organ examination, and ventilation and respiratory compliance studies yielded results normal for gestational age. This first reported case demonstrates that pregnancy can be carried in an allotransplanted uterus, with the end result a successful delivery.
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http://dx.doi.org/10.1016/j.jmig.2010.11.006DOI Listing
July 2011

Prototype hybrid systems for neonatal warming: in vitro comparisons to standard of care devices.

Biomed Instrum Technol 2010 Nov-Dec;44(6):523-7

Department of Physiology, Temple University, Philadelphia, PA, USA.

Preterm infants lack necessary thermoregulation. An ideal incubator should maintain a uniform and constant thermal environment. We compared the effectiveness of a supplemental heating blanket to improve the heating characteristics of two different incubator warming devices using assessment of their respective function alone as controls. Device A and device B, with and without a heating blanket (Harvard Apparatus), were instrumented with a distribution matrix of multiple temperature (n = 11) and humidity probes. These data were serially measured during warm up to 37.5 °C and through a series of open-door perturbations. The time constant, temperature variation, and change in air temperature were calculated. Data were analyzed for significance by 2-factor ANOVA for each respective incubator either turned on or off with either the heating blanket turned on or off. Device A warms faster (33.87% ; p < 0.05) than device B, but has a greater (37.27% ; p < 0.05) temperature variation during warmup. The heating blanket enhances the thermal response of device A during warmup, but does not alter those of device B. With the side door open, device A shows a smaller (-16.5% ; p < 0.05) temperature variation than device B; the heating blanket attenuates the temperature change in both devices. These results demonstrate that the use of a supplemental heating blanket, as well as device-related differences, may impact clinical control of a thermal environment.
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http://dx.doi.org/10.2345/0899-8205-44.6.523DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3888790PMC
February 2011

Protection against sepsis-induced lung injury by selective inhibition of protein kinase C-δ (δ-PKC).

J Leukoc Biol 2011 Jan 19;89(1):3-10. Epub 2010 Aug 19.

Temple University School of Medicine, 3307 North Broad St., PAH-206, Philadelphia, PA 19140, USA.

Inflammation and proinflammatory mediators are activators of δ-PKC. In vitro, δ-PKC regulates proinflammatory signaling in neutrophils and endothelial and epithelial cells, cells that can contribute to lung tissue damage associated with inflammation. In this study, a specific δ-PKC TAT peptide inhibitor was used to test the hypothesis that inhibition of δ-PKC would attenuate lung injury in an animal model of ARDS. Experimental ARDS was induced in rats via 2CLP, a model of polymicrobial sepsis. Following 2CLP surgery, the δ-PKC TAT inhibitory peptide (2CLP+δ-PKC TAT in PBS) or PBS (2CLP+PBS) was administered intratracheally. Controls consisted of SO, where animals underwent a laparotomy without 2CLP. Twenty-four hours after SO or 2CLP, blood, BALF, and lung tissue were collected. 2CLP induced δ-PKC phosphorylation in the lung within 24 h. Treatment with the δ-PKC TAT inhibitory peptide significantly decreased pulmonary δ-PKC phosphorylation, indicating effective inhibition of δ-PKC activation. Plasma and BALF levels of the chemokines CINC-1 and MIP-2 were elevated in 2CLP + PBS rats as compared with SO rats. Treatment with δ-PKC TAT reduced 2CLP-induced elevations in chemokine levels in BALF and plasma, suggesting that δ-PKC modulated chemokine expression. Most importantly, intratracheal administration of δ-PKC TAT peptide significantly attenuated inflammatory cell infiltration, disruption of lung architecture, and pulmonary edema associated with 2CLP. Thus, δ-PKC is an important regulator of proinflammatory events in the lung. Targeted inhibition of δ-PKC exerted a lung-protective effect 24 h after 2CLP.
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http://dx.doi.org/10.1189/jlb.0510281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3004522PMC
January 2011

Improved gas exchange and survival after KL-4 surfactant in newborn pigs with severe acute lung injury.

Pediatr Pulmonol 2010 Aug;45(8):782-8

Department of Pediatrics, University of Minnesota, Minneapolis, MN 55110,USA.

Objective: To determine the effectiveness of artificial surfactant therapy using KL-4 surfactant in newborn pigs with hydrochloric acid (HCl)-induced acute lung injury (ALI).

Design: After induction of ALI via intratracheal HCl instillation, pigs were randomized to receive 5.8 ml/kg KL-4 surfactant or no surfactant prior to extubation to bubble CPAP.

Setting: Clinical laboratory.

Subjects: Spontaneously breathing newborn pigs (<1 week of age).

Interventions: Treatment with KL-4 surfactant on bubble CPAP with PEEP of 6 cmH(2)O for 3.5 hr after extubation compared with controls.

Measurements: Physiologic parameters and arterial blood gases were measured every 15 min. At the conclusion of the study, the lungs were excised for the analysis of histopathology and morphometric data.

Main Results: Pigs treated with KL-4 surfactant had arterial blood gases with less acidosis (P < 0.001), higher P(a)O(2) levels (P < 0.001), and lower P(a)CO(2) levels (P < 0.001). Pigs treated with KL-4 surfactant had improved survival compared with controls (6/12 KL-4, 2/12 control, P < 0.05). Postmortem morphometric data demonstrated that pigs treated with KL-4 surfactant had larger (P < 0.05) exchange units in the caudal-dorsal lung as compared to relatively atelectatic region in the control animals.

Conclusions: In newborn pigs with severe HCl-induced ALI, treatment with KL-4 surfactant resulted in improved respiratory parameters, less dependent atelectasis, and improved short-term survival.
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http://dx.doi.org/10.1002/ppul.21252DOI Listing
August 2010
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