The authors would like to acknowledge the Cincinnati Children&#8217;s Hospital Division of Veterinary Services for their assistance. This manuscript was supported by the National Institutes of Health K12 HD028827. Anna Perl PhD taught the authors this surgical procedure.

הצהרת שימוש בבעלי חיים:: הערה כל הנהלים במחקר זה נערכו באישור וביצוע ההנחיות של בעלי החיים המוסדיים השימוש וטיפול הוועדה (IACUC) בבית החולים לילדים בסינסינטי. עכברי זכרים שמונה שבועות בן C57BL / 6J התקבלו מג&#39;קסון מעבדות (Bar Harbor, ME) ואפשרו להסתגל לשבוע לפני שימוש אחד. עד ניתוח, בעלי חיים שוכנו במתקן מחסום הפתוגן ללא וסיפקו אוכל autoclaved וlibdium מודעת מים מסונן. כל כלוב עכבר סופק עם אוויר ומים ייעודיים, וחדרים נשמרו במחזור היום-לילה 12 שעות. בעקבות התאוששות מניתוח, עכברים נשמרו בכלובים עם צמרות מסוננות, ובלבד libidum מודעת אוכל autoclaved, ובלבד שמים מסוננים מבקבוק מים. 1. הכנה של מכשירים <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> הפוך 6 מפשקי עור באמצעות אטבי נייר וסיכות. טוויסט יישר מהדקי נייר באנקס של סיכות נייר, leave תיל פלדה ישר 5 סנטימטר בקצה אחד ולעשות 0.5 סנטימטר בודד וו &quot;U&quot; בצורה בסוף החוט. </li><li style=";text-align:right;direction:rtl"> הפוך וילונות כירורגי מרובעים כ -15 x 15 סנטימטר באמצעות ניילון נצמד. הכן רוטב אחד לכל עכבר. שים מגדל נייר בין כל לעטוף. </li><li style=";text-align:right;direction:rtl"> לעקר את כל הכלים כירורגיים יחד עם ערימה של אריחי 12 x 12 אינץ &#39;פקק, גזה, וספוגיות כותנה שקצה. </li></ol> 2. עכבר הכנה <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לגרום להרדמה עם isoflurane 2%. לשקול בעלי חיים. </li><li style=";text-align:right;direction:rtl"> באזור הכנה כירורגית ייעודי לגלח בית חזה שמאלי ואזור צוואר עם מכונת גילוח חשמלי. </li><li style=";text-align:right;direction:rtl"> החל ירידה של המשחה הדמעה המלאכותית לעיניו של העכבר. </li><li style=";text-align:right;direction:rtl"> לטהר צוואר ובית חזה שמאלי עם chlorhexidine ואלכוהול איזופרופיל. חזור פעמיים יותר. </li></ol> אינטובציה 3. העכבר Oro-קנה הנשימה ואוורור מכאני <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> יש מקום טכנאי כירורגית שאינו סטרילי פרקדן העכבר באזור הניתוח המחומם מראש.</li><li style=";text-align:right;direction:rtl"> לאשר עומק ההרדמה על ידי תיעוד חוסר תגובה לקמצוץ כפה. </li><li style=";text-align:right;direction:rtl"> לאחר שטיפת ידיים ועוטים לבוש כירורגית, מסיכה, וכובע, דון כפפות מנתחים סטרילית. </li><li style=";text-align:right;direction:rtl"> לאחר כורך ובאמצעות טכניקה האספטי, לעשות חתך אנכי 1 סנטימטר מעל אמצע הצוואר-הקדמי כדי לחשוף את הגרון. קל לחזור שרירי הרצועה עם משוננים מלקחיים מעוקלים סנטימטר, 10 ולחשוף את הגרון וקנה נשימה על ידי הפצת שרירי הרצועה עם הקצה של מספריים ישר. </li><li style=";text-align:right;direction:rtl"> דרך הפה להכניס angiocatheter בוטה, קצה 22 G לאמצע קנה הנשימה (איור 1 א) וחזותי לאשר מיקום (איור 1). לשמור על ההרדמה ולאוורר באמצעות isoflurane 1-3% באמצעות מכונת הנשמה מכרסם (225 μl לשבץ; 200 סטוקס לדקה). להעסיק מגבלת לחץ של 15 סנטימטרים H 2 O. </li></ol> 4. עכבר ריאה <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> הנח את העכבר בעמדת פַּחֶסֶת רוחב תקין עם מול האחורי של העכברהמפעיל (בצד שמאל למעלה). השתמש בניילון נצמד איטום עצמי כוילון סטרילי. חיתוך דרך הווילון, להשתמש במספריים מעוגלים היטה בוטים לעשות מקביל ארוך קיצוץ של 2 סנטימטר לצלעות בחלל צלע 4 th וה -5. הכנס את המספריים קצה מעוקל בוטים ולנתח את העור מצלעות הבסיס ושרירים הבין-. </li><li style=";text-align:right;direction:rtl"> לחזור בו העור עם ארבעה מפשקים לחשוף מרובע 1.5 x 1.5 סנטימטר חלון כירורגית (איור 2 א). אבטח את המפשקים ללוח השעם. </li><li style=";text-align:right;direction:rtl"> לנתח את הצלעות באמצעות מלקחיים מעוקלים, ולהשתמש בקצה אחד של המלקחיים המעוקלים להיכנס לחלל בית החזה. </li><li style=";text-align:right;direction:rtl"> שימוש במייקרו-המספריים קצה הקהה, השתמש בלהב התחתון להיכנס לחלל בית החזה. ביצוע חתך 0.5 סנטימטר בין הצלעות וחזור על הפעולה בכיוון ההפוך. </li><li style=";text-align:right;direction:rtl"> באמצעות שני הכולים הכתיבה שנותרו, לפתוח את בית החזה בציר הקדמי-אחורית והמפשקים מאובטחים ללוח השעם (איור 2). </li> &lt;li&gt; שימוש שבקצו מלקחיים בוטים מעוקלים ביד השמאל, לתפוס את הריאה השמאלית ולעקור את החלק העליון של הריאה השמאלית רוחבי וinferiorly באמצעות פתיחת בית החזה עד עורק הריאה השמאלי והסמפונות נחשפים (איור 3 א &#39;, ב&#39;). </li><li style=";text-align:right;direction:rtl"> מחזיק המוליך microclip כלי הדם טיטניום הטעון ביד ימין עם הגוף של המוליך בכף וקצה מעוגל פונה אל כף היד (איור 3 ג), חלק את קצה המוליך לבית החזה לאורך העקמומיות של חלק האחורי של השמאל ריאות וקליפ הסמפונות עזבה ועורק ריאה (איור 3D). </li><li style=";text-align:right;direction:rtl"> הסר את המוליך אבל לשמור הריאה השמאלית חזר בו. אחוז במייקרו המספריים קצה הקהה ​​עם יד ימין וחתכתי את הסמפונות ודיסטלי עורק הריאה לקליפ ולהסיר ריאה שמאלית (איור 3E). </li><li style=";text-align:right;direction:rtl"> הסר את מפשקי צלעות. </li><li style=";text-align:right;direction:rtl"> השתמש במלקחיים בוטים המעוקלים לצבוט את 1 סנטימטר של עור נחות incisiבאבל מעל לרמה של הסרעפת והכנס angiocatheter G 24 דרך העור ולתוך חלל בית החזה השמאלי (איור 4 א &#39;, ב&#39;). </li><li style=";text-align:right;direction:rtl"> השתמש 5-0 prolene תפר למקום שני תפרים שהופסקו סביב ה 4 ו -5 צלעות ה לסגור את חלל בית החזה. </li><li style=";text-align:right;direction:rtl"> הסר את מפשקי העור. השתמש בשני סטים של מלקחיים כדי להתקרב לעור לאורך החתך ומדביק את העור סגור. </li><li style=";text-align:right;direction:rtl"> חבר מזרק luer נעילת 3 מיליליטר לangiocatheter ולהסיר אוויר שייר על ידי יישום שאיבה עדינה ונסיגת angiocatheter. </li><li style=";text-align:right;direction:rtl"> מדביק את החתך בצוואר נסגר באמצעות שני סטים של מלקחיים כמו קודם. </li></ol> 5. עכבר שאם ריאה <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לחשוף את הריאה השמאלית כפי שצוין בפרוטוקול &quot;עכבר ריאה&quot;. הרם את כלוב צלעות עם מלקחיים בוטים מעוקלים כדי לאפשר לאוויר לתוך חלל החזה השמאלי (איור 5 א &#39;, ב&#39;). </li><li style=";text-align:right;direction:rtl"> הנח angiocatheter G 24 לאף השמאלחלל racic כאמור לעיל נזהר שלא לפגוע הריאה השמאלית. </li><li style=";text-align:right;direction:rtl"> באמצעות 5-0 תפר prolene ונזהר שלא לנקב את הריאות (איור 5 ג), למקם את שני אורכים של חומר תפר לתוך 3 rd / 4 th וה -5 / 6 interspaces צלע ה (איור 5D). הנח את שני האורכים של חומר תפר לפני הקשירה כדי להפחית את הסיכונים של פריצת ריאה השמאלית. לקשור את חומר התפר לעשות שני תפרים שהופסקו (איור 5E). </li><li style=";text-align:right;direction:rtl"> מדביק את העור על החתך בבית החזה, להסיר אוויר שייר עם angiocatheter, ולהדביק את החתך בצוואר כאמור לעיל. </li></ol> 6. החייאה, שיכוך כאבים, ושחזור <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> כבה את isoflurane, ולנהל 0.1 מ&quot;ג / קילוגרם של עצירות ו0.5 מיליליטר של תת-עורית מלח הרגילה. </li><li style=";text-align:right;direction:rtl"> כאשר נשימה ספונטנית לחדש, להסיר את הטובוס. </li><li style=";text-align:right;direction:rtl"> שים לב עכבר עד שהוא שוב אמבולטורי. הליכה בדרך כלל resumes כמה דקות לאחר הסרת הטובוס. </li><li style=";text-align:right;direction:rtl"> מניחים את העכבר ב-27 מעלות צלזיוס חממה (חמצן humidified, 25%) כדי לשחזר O / N. הערה: אנו שמים כמה כדורים של האוכל רטוב עם מים על רצפת הכלוב ל24 שעות הראשונות שלאחר ניתוח. </li><li style=";text-align:right;direction:rtl"> לנהל 0.1 מ&quot;ג / קילוגרם של עצירות בזריקת intraperitoneal פעמיים ביום למשך שלושה ימים לאחר ניתוח. הקפד לא לפתוח את האתר כירורגית בעת טיפול בבעלי חיים. </li></ol> 7. ניטור עכבר <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לשקול עכברים בבית 1, 3, 5, ו -7 ימים לאחר ניתוח. </li></ol>

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The authors have nothing to disclose.

סיפקנו תיאור מפורט ביותר של נהלי PNX PNX העכבר ועכבר דמה שדווחו עד כה. אנחנו עשינו את הקורא מודע לכמה מהמכשולים הנפוצים שחוקרים לומדים ההליך נפוץ להיתקל, ויש לנו התווינו כמה טכניקות שפותחו על ידי המעבדה שלנו כדי להקטין נגד המלכודות הללו. מעבדות אחרות ניצול מודל זה עשויו?...

תפקידו העיקרי של הריאות הוא לספק לחמצן ופחמן דו-חמצני חילופי בין האורגניזם והאווירה. בבני אדם, שורה של תנאים מולדים ונרכשים להוביל לשטח ריאות מופחת וכתוצאה מכך תפקוד ריאות לקוי. למרות שורה של טיפולים כגון סטרואידים בשאיפה, מרחיבי סימפונות, חמצן, אוויר ואוורור מכאני כרוני משמשות כדי למתן את ההשלכות של תפקוד ריאות לקוי 1-3, הטיפול האידיאלי לתנאים אלה יקדם צמיחה מחודשת של רקמה תפקודית ריאות - כלומר, ריאות התחדשות. התחדשות רקמות של יונקים תועדה היטב. קוצני העכבר האפריקאי יכול להתחדש שטחים גדולים של עור ללא היווצרות צלקת 4. גדוד דיסטלי בבני אדם יכול להתחדש בעקבות פציעה או קטיעה 5-7. הריאה הבאה (PNX), צמיחת ריאות מפצה מתרחשת בעכברים 8, 9 חולדות, לעשותGS 10, ובני האדם 11. על פי הגדרה, צמיחת ריאות פיצוי כרוכה הרחבה לא רק של airspaces הקיים, אך מחדש septation של airspaces המוגדל אלה עם הרחבת זרימת הדם קשור 12. ניתוח ביטוי גנים הוכיח כי מודל זה משחזר רב של אירועי האיתות של התפתחות ריאות 13. ארבעה שבועות לאחר העכבר PNX, שטח פן alveolar הוא שווה ערך לזה של בעלי חיים דמה מופעלים 14. בכתב היד הזה, אנו מתארים את PNX העכבר ונהלי PNX דמה.

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mouse pneumonectomy model of compensatory lung growth

In humans, disrupted repair and remodeling of injured lung contributes to a host of acute and chronic lung disorders which may ultimately lead to disability or death. Injury-based animal models of lung repair and regeneration are limited by injury-specific responses making it difficult to differentiate changes related to the injury response and injury resolution from changes related to lung repair and lung regeneration. However, use of animal models to identify these repair and regeneration signaling pathways is critical to the development of new therapies aimed at improving pulmonary function following lung injury. The mouse pneumonectomy model utilizes compensatory lung growth to isolate those repair and regeneration signals in order to more clearly define mechanisms of alveolar re-septation. Here, we describe our technique for performing mouse pneumonectomy and sham pneumonectomy. This technique may be utilized in conjunction with lineage tracing or other transgenic mouse models to define molecular and cellular mechanism of lung repair and regeneration.

. חלקה של PNX ומשקולות עכבר מזויף פעל מסופקת באיור 6 בידיים שלנו, הישרדות היא באופן עקבי 95-100% עבור שתי pneumonectomy PNX והעמדת פן. לתיאורים של איך הריאה הימנית מחדש גדל במודל זה וכמובן הזמן הצפוי, אנחנו מפנים את הקורא לכתבי יד של גיבן et al. 15 וet al וואנג. 1...

Watch this Scientific Journal Video about Mouse Pneumonectomy Model of Compensatory Lung Growth at JoVE.com

Mouse Pneumonectomy Model of Compensatory Lung Growth

Mouse pneumonectomy is a commonly employed model of compensatory lung growth. This procedure can be used in conjunction with lineage tracing or transgenic mouse models to elucidate underlying mechanisms.

עכבר דגם ריאה של פיצוי ריאות צמיחה

In humans, disrupted repair and remodeling of injured lung contributes to a host of acute and chronic lung disorders which may ultimately lead to ...

mouse-pneumonectomy-model-of-compensatory-lung-growth

Research

JoVE Journal

Medicine

12.8K Views.  Cincinnati Children's Hospital Medical Center. Mouse pneumonectomy is a commonly employed model of compensatory lung growth. This procedure can be used in conjunction with lineage tracing or transgenic mouse models to elucidate underlying mechanisms.

Video: Mouse Pneumonectomy Model of Compensatory Lung Growth

Development of Obliterative Bronchiolitis in a Murine Model of Orthotopic Lung Transplantation

Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and standardized not only for the study of allo-rejection but also between syngeneic strains for examining mechanisms of ischemia-reperfusion injury after lung transplantation. Although the application of the rat and other large animal model 2 contributed significantly to the elucidation of these studies, the scope of those investigations is limited by the scarcity of knockout and transgenic rats. Due to no effective therapies for obliterative bronchiolitis, the leading cause of death in lung transplant patients, there has been an intensive search for pre-clinical models that replicate obliterative bronchiolitis. The tracheal allograft model is the most widely used and may reproduce some of the histopathologic features of obliterative bronchiolitis 3. However, the lack of an intact vasculature with no connection to the recipient's conducting airways, and incomplete pathologic features of obliterative bronchiolitis limit the utility of this model 4. Unlike transplantation of other solid organs, vascularized mouse lung transplants have only recently been reported by Okazaki and colleagues for the first time in 2007 5. Applying the basic principles of the rat lung transplant, our lab initiated the obliterative bronchiolitis model using minor histoincompatible antigen murine orthotopic single-left lung transplants which allows the further study of obliterative bronchiolitis immunopathogenesis6.

Obliterative bronchiolitis is the key impediment to the long-term survival of lung transplant recipients and the lack of a robust preclinical model precludes examining obliterative bronchiolitis immunopathogenesis. Unlike other solid organ transplants, vascularized mouse lung transplantation has only recently been developed. Here we show our independently developed obliterative bronchiolitis model after murine orthotopic single-lung transplantation.

פיתוח Bronchiolitis סותם במודל Murine של השתלת ריאות Orthotopic

Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and ...

A Standardized Method for Measuring Internal Lung Surface Area via Mouse Pneumonectomy and Prosthesis Implantation

Pulmonary morphology, physiology, and respiratory functions change in both physiological and pathological conditions. Internal lung surface area (ISA), representing the gas-exchange capacity of the lung, is a critical criterion to assess respiratory function. However, observer bias can significantly influence measured values for lung morphological parameters. The protocol that we describe here minimizes variations during measurements of two morphological parameters used for ISA calculation: internal lung volume (ILV) and mean linear intercept (MLI). Using ISA as a morphometric and functional parameter to determine the outcome of alveolar regeneration in both pneumonectomy (PNX) and prosthesis implantation mouse models, we found that the increased ISA following PNX treatment was significantly blocked by implantation of a prosthesis into the thoracic cavity1. The ability to accurately quantify ISA is not only expected to improve the reliability and reproducibility of lung function studies in injured-induced alveolar regeneration models, but also to promote mechanistic discoveries of multiple pulmonary diseases.

Internal lung surface area (ISA) is a critical criterion for assessing lung morphology and physiology in lung diseases and injury-induced alveolar regeneration. We describe here a standardized method that can minimize the measurement bias for ISA in both lung pneumonectomy and prosthesis implantation mouse models.

שיטה מתוקננת למדידת שטח הפנים של הריאות הפנימיות באמצעות דלקת ריאות של עכבר והשתלת תותב

Pulmonary morphology, physiology, and respiratory functions change in both physiological and pathological conditions. Internal lung surface area (ISA), ...

Developmental Biology

Measurement of the Pressure-volume Curve in Mouse Lungs

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential phenotypic changes are best assessed by measurement of the changes in lung elasticity. To best understand specific mechanisms underlying such pathologies in mice, it is essential to make functional measurements that can reflect the developing pathology. Although there are many ways to measure elasticity, the classical method is that of the total lung pressure-volume (PV) curve done over the whole range of lung volumes. This measurement has been made on adult lungs from nearly all mammalian species dating back almost 100 years, and such PV curves also played a major role in the discovery and understanding of the function of pulmonary surfactant in fetal lung development. Unfortunately, such total PV curves have not been widely reported in the mouse, despite the fact that they can provide useful information on the macroscopic effects of structural changes in the lung. Although partial PV curves measuring just the changes in lung volume are sometimes reported, without a measure of absolute volume, the nonlinear nature of the total PV curve makes these partial ones very difficult to interpret. In the present study, we describe a standardized way to measure the total PV curve. We have then tested the ability of these curves to detect changes in mouse lung structure in two common lung pathologies, emphysema and fibrosis. Results showed significant changes in several variables consistent with expected structural changes with these pathologies. This measurement of the lung PV curve in mice thus provides a straightforward means to monitor the progression of the pathophysiologic changes over time and the potential effect of therapeutic procedures.

Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung&#8217;s structural changes with developing pathology.

מדידה של עקומת לחץ-נפח ריאות עכבר

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential ...

Generation of a Chronic Obstructive Pulmonary Disease Model in Mice by Repeated Ozone Exposure

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung parenchymal destruction. It has a very high incidence in aging populations. The current conventional therapies for COPD focus mainly on symptom-modifying drugs; thus, the development of new therapies is urgently needed. Qualified animal models of COPD could help to characterize the underlying mechanisms and can be used for new drug screening. Current COPD models, such as lipopolysaccharide (LPS) or the porcine pancreatic elastase (PPE)-induced emphysema model, generate COPD-like lesions in the lungs and airways but do not otherwise resemble the pathogenesis of human COPD. A cigarette smoke (CS)-induced model remains one of the most popular because it not only simulates COPD-like lesions in the respiratory system, but it is also based on one of the main hazardous materials that causes COPD in humans. However, the time-consuming and labor-intensive aspects of the CS-induced model dramatically limit its application in new drug screening. In this study, we successfully generated a new COPD model by exposing mice to high levels of ozone. This model demonstrated the following: 1) decreased forced expiratory volume 25, 50, and 75/forced vital capacity (FEV25/FVC, FEV50/FVC, and FEV75/FVC), indicating the deterioration of lung function; 2) enlarged lung alveoli, with lung parenchymal destruction; 3) reduced fatigue time and distance; and 4) increased inflammation. Taken together, these data demonstrate that the ozone exposure (OE) model is a reliable animal model that is similar to humans because ozone overexposure is one of the etiological factors of COPD. Additionally, it only took 6 - 8 weeks, based on our previous work, to create an OE model, whereas it requires 3 - 12 months to induce the cigarette smoke model, indicating that the OE model might be a good choice for COPD research.

This study describes the successful generation of a new chronic obstructive pulmonary disease (COPD) animal model by repeatedly exposing mice to high concentrations of ozone.

דור של מחלת ריאות חסימתית כרונית מודל בעכברים על ידי חשיפה חוזרת ונשנית האוזון

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung parenchymal destruction. It has a very high ...

The Left Pneumonectomy Combined with Monocrotaline or Sugen as a Model of Pulmonary Hypertension in Rats

In this protocol, we detail the correct procedural steps and necessary precautions to successfully perform a left pneumonectomy and induce PAH in rats with the additional administration of monocrotaline (MCT) or SU5416 (Sugen). We also compare these two models to other PAH models commonly used in research. In the last few years, the focus of animal PAH models has moved towards studying the mechanism of angioproliferation of plexiform lesions, in which the role of increased pulmonary blood flow is considered as an important trigger in the development of severe pulmonary vascular remodeling. One of the most promising rodent models of increased pulmonary flow is the unilateral left pneumonectomy combined with a "second hit" of MCT or Sugen. The removal of the left lung leads to increased and turbulent pulmonary blood flow and vascular remodeling. Currently, there is no detailed procedure of the pneumonectomy surgery in rats. This article details a step-by-step protocol of the pneumonectomy surgical procedure and post-operative care in male Sprague-Dawley rats. Briefly, the animal is anesthetized and the chest is opened. Once the left pulmonary artery, pulmonary vein, and bronchus are visualized, they are ligated and the left lung is removed. The chest then closed and the animal recovered. Blood is forced to circulate only on the right lung. This increased vascular pressure leads to a progressive remodeling and occlusion of small pulmonary arteries. The second hit of MCT or Sugen is used one week post-surgery to induce endothelial dysfunction. The combination of increased blood flow in the lung and endothelial dysfunction produces severe PAH. The primary limitation of this procedure is that it requires general surgical skills.

The rodent left pneumonectomy is a valuable technique in pulmonary hypertension research. Here, we present a protocol to describe the rat pneumonectomy procedure and postoperative care to ensure minimal morbidity and mortality.

כריתת ריאה השמאלית בשילוב עם Monocrotaline או Sugen כמודל של יתר לחץ דם ריאתי אצל חולדות

In this protocol, we detail the correct procedural steps and necessary precautions to successfully perform a left pneumonectomy and induce PAH in rats ...

Experimental Model to Evaluate Resolution of Pneumonia

Acute respiratory distress syndrome (ARDS) causes acute lung injury, characterized by rapid alveolar damage and severe hypoxemia. This, in turn, leads to high morbidity and mortality. Currently, there are no pre-clinical models that recapitulate the complexity of human ARDS. However, infectious models of pneumonia (PNA) can replicate the main pathophysiological features of ARDS. Here, we describe a model of PNA induced by the intratracheal instillation of live Streptococcus pneumoniae and Klebsiella pneumoniae in C57BL6 mice. In order to evaluate and characterize the model, after inducing injury, we carried out serial measurements of body weight and bronchoalveolar lavage (BAL) for measuring markers of lung injury. Additionally, we harvested lungs for cell count and differentials, BAL protein quantification, cytospin, bacterial colony-forming unit counts, and histology. Lastly, high dimensional flow cytometry was performed. We propose this model as a tool to understand the immune landscape during the early and late resolution phases of lung injury.

This manuscript describes the establishment of an infectious model of pneumonia in mice and the respective characterization of injury resolution along with methods for growing bacteria and intratracheal instillation. A novel approach using high-dimensional flow cytometry to evaluate the immune landscape is also described.

מודל ניסיוני להערכת רזולוציה של דלקת ריאות

Acute respiratory distress syndrome (ARDS) causes acute lung injury, characterized by rapid alveolar damage and severe hypoxemia. This, in turn, leads to ...

Immunology and Infection

A Rat Lung Transplantation Model of Warm Ischemia/Reperfusion Injury: Optimizations to Improve Outcomes

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods for choosing appropriate cuff sizes for the pulmonary vein (PV), pulmonary artery (PA), or bronchus (Br) are varied, thus making the determination of proper cuff size during rat lung transplantation an exercise of trial and error. By standardizing the cuffing technique to use the smallest effective cuff appropriate for the size of the vessel or bronchus, one can make the transplantation procedure safer, faster, and more successful. Since diameters of the PV, PA, and Br are related to the body weight of the rat, we present a strategy to choosing an appropriate size using a weight-based guide. Since lung volume is also related to body weight, we recommend that this relationship should also be considered when choosing the proper volume of air for donor lung inflation during warm ischemia as well as for the proper volume of PBS to be instilled during bronchoalveolar lavage (BAL) fluid collection. We also describe methods for 4th intercostal space dissection, wound closure, and sample collection from both the native and transplanted lobes.

Here, we present optimizations to a rat lung transplantation model that serve to improve outcomes. We provide a size guide for cuffs based on body weight, a measurement strategy to ascertain the 4th intercostal space, and methods of wound closure and BAL (bronchoalveolar lavage) fluid and tissue collection.

מודל השתלת ריאה של חולדה של איסכמיה חמה / פגיעה ברפרפוזיה: אופטימיזציות לשיפור התוצאות

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods ...

Normothermic Negative Pressure Ventilation Ex Situ Lung Perfusion: Evaluation of Lung Function and Metabolism

Lung transplantation (LTx) remains the standard of care for end-stage lung disease. A shortage of suitable donor organs and concerns over donor organ quality exacerbated by excessive geographic transportation distance and stringent donor organ acceptance criteria pose limitations to current LTx efforts. Ex situ lung perfusion (ESLP) is an innovative technology that has shown promise in attenuating these limitations. The physiologic ventilation and perfusion of the lungs outside of the inflammatory milieu of the donor body affords ESLP several advantages over traditional cold static preservation (CSP). There is evidence that negative pressure ventilation (NPV) ESLP is superior to positive pressure ventilation (PPV) ESLP, with PPV inducing more significant ventilator-induced lung injury, pro-inflammatory cytokine production, pulmonary edema, and bullae formation. The NPV advantage is perhaps due to the homogenous distribution of intrathoracic pressure across the entire lung surface. The clinical safety and feasibility of a custom NPV-ESLP device have been demonstrated in a recent clinical trial involving extender criteria donor (ECD) human lungs. Herein, the use of this custom device is described in a juvenile porcine model of normothermic NPV-ESLP over a 12 h duration, paying particular attention to management techniques. Pre-surgical preparation, including ESLP software initialization, priming, and de-airing of the ESLP circuit, and the addition of anti-thrombotic, anti-microbial, and anti-inflammatory agents, is specified. The intraoperative techniques of central line insertion, lung biopsy, exsanguination, blood collection, cardiectomy, and pneumonectomy are described. Furthermore, particular focus is paid to anesthetic considerations, with anesthesia induction, maintenance, and dynamic modifications outlined. The protocol also specifies the custom device's initialization, maintenance, and termination of perfusion and ventilation. Dynamic organ management techniques, including alterations in ventilation and metabolic parameters to optimize organ function, are thoroughly described. Finally, the physiological and metabolic assessment of lung function is characterized and depicted in the representative results.

Normothermic Negative Pressure Ventilation Ex Situ Lung Perfusion: Evaluation of Lung Function and Metabolism

This paper describes a porcine model of negative pressure ventilation ex situ lung perfusion, including procurement, attachment, and management on the custom-made platform. Focus is made on anesthetic and surgical techniques, as well as troubleshooting.

אוורור לחץ שלילי נורממי זילוח ריאות Ex Situ : הערכת תפקוד הריאות וחילוף החומרים

Lung transplantation (LTx) remains the standard of care for end-stage lung disease. A shortage of suitable donor organs and concerns over donor organ ...

A Simple Cuff Technique for Murine Left Lung Transplantation

Over the past decade, our laboratory has made significant progress in developing and refining vascularized mouse lung transplantation models using an efficient and highly reliable "cuff technique" of transplantation. This article describes a sophisticated and comprehensive method for orthotopic lung transplantation in a vascularized orthotopic lung model, representing the most physiologic and clinically relevant model of mouse lung transplantation to date. The transplantation process consists of two distinct stages: donor harvest and subsequent implantation into the recipient. The method has been successfully mastered, and with several months of sufficient training, a skilled practitioner can perform the procedure in approximately 90 min from skin-to-skin. Surprisingly, once individuals overcome the initial learning curve, the survival rate during the perioperative period approaches nearly 100%. The mouse model allows for the use of multiple commercially available transgenic and mutant strains of mice, enabling the study of tolerance and rejection. Additionally, the unique features of this model make it a valuable tool for investigating tumor biology and immunology.

The present protocol describes a cuff technique for a mouse left lung transplantation model. This technique has been developed over several years and has performed well, serving effectively in immunological research.

Over the past decade, our laboratory has made significant progress in developing and refining vascularized mouse lung transplantation models using an ...

Ex Vivo Porcine Experimental Model for Studying and Teaching Lung Mechanics

Mechanical ventilation is widely used and requires specific knowledge for understanding and management. Health professionals in this field may feel insecure and lack knowledge because of inadequate training and teaching methods. Therefore, the objective of this article is to outline the steps involved in generating an ex vivo porcine lung model to be used in the future, to study and teach lung mechanics. To generate the model, five porcine lungs were carefully removed from the thorax following the guidelines of the Animal Research Ethics Committee with adequate care and were connected to the mechanical ventilator through a tracheal cannula. These lungs were then subjected to the alveolar recruitment maneuver. Respiratory mechanics parameters were recorded, and video cameras were used to obtain videos of the lungs during this process. This process was repeated for five consecutive days. When not used, the lungs were kept refrigerated. The model showed different lung mechanics after the alveolar recruitment maneuver every day; not being influenced by the days, only by the maneuver. Therefore, we conclude that the ex vivo lung model can provide a better understanding of lung mechanics and its effects, and even of the alveolar recruitment maneuver through visual feedback during all stages of the process.

Ex Vivo Porcine Experimental Model for Studying and Teaching Lung Mechanics

We present an ex vivo pig lung model for the demonstration of pulmonary mechanics and alveolar recruitment maneuvers for teaching purposes. The lungs can be used for more than one day (up to five days) with minimal changes in pulmonary mechanics variables.

אקס ויוו מודל ניסויי חזירי ללימוד והוראת מכניקת ריאות

Mechanical ventilation is widely used and requires specific knowledge for understanding and management. Health professionals in this field may feel ...