Migration of CD11b+ accessory cells during murine lung regeneration

In many mammalian species, the removal of one lung leads to growth of the remaining lung to near-baseline levels. In studying post-pneumonectomy mice, we used morphometric measures to demonstrate neoalveolarization within 21 days of pneumonectomy. Of note, the detailed histology during this period demonstrated no significant pulmonary inflammation. To identify occult blood-borne cells, we used a parabiotic model (wild-type/GFP) of post-pneumonectomy lung growth. Flow cytometry of post-pneumonectomy lung digests demonstrated a rapid increase in the number of cells expressing the hematopoietic membrane molecule CD11b; 64.5% of the entire GFP(+) population were CD11b(+). Fluorescence microscop…

Mechanostructural adaptations preceding postpneumonectomy lung growth

In many species, pneumonectomy results in compensatory growth in the remaining lung. Although the late mechanical consequences of murine pneumonectomy are known, little is known about the anatomic adaptations and respiratory mechanics during compensatory lung growth. To investigate the structural and mechanical changes during compensatory growth, mice were studied for 21 days after left pneumonectomy using microCT and respiratory system impedance (FlexiVent). Anatomic changes after left pneumonectomy included minimal mediastinal shift or chestwall remodeling, but significant displacement of the heart and cardiac lobe. Mean displacement of the cardiac lobe centroid was 5.2 ± 0.8 mm. Lung imp…

Vascular Microarchitecture of Murine Colitis-Associated Lymphoid Angiogenesis

In permissive tissues, such as the gut and synovium, chronic inflammation can result in the ectopic development of anatomic structures that resemble lymph nodes. These inflammation-induced structures, termed lymphoid neogenesis or tertiary lymphoid organs, may reflect differential stromal responsiveness to the process of lymphoid neogenesis. To investigate the structural reorganization of the microcirculation involved in colonic lymphoid neogenesis, we studied a murine model of dextran sodium sulfate (DSS)-induced colitis. Standard 2-dimensional histology demonstrated both submucosal and intramucosal lymphoid structures in DSS-induced colitis. A spatial frequency analysis of serial histolog…

Computational flow dynamics in a geometric model of intussusceptive angiogenesis.

Intussusceptive angiogenesis is a process that forms new blood vessels by the intraluminal division of a single blood vessel into two lumens. Referred to as nonsprouting or intussusceptive angiogenesis, this angiogenic process has been described in morphogenesis and chronic inflammation. Mechanical forces are relevant to the structural changes associated with intussusceptive angiogenesis because of the growing evidence that physiologic forces influence gene transcription. To provide a detailed analysis of the spatial distribution of physiologic shear stresses, we developed a 3D finite element model of the intraluminal intussusceptive pillar. Based on geometries observed in adult intussuscep…

Alveolar Epithelial Dynamics in Postpneumonectomy Lung Growth

The intimate anatomic and functional relationship between epithelial cells and endothelial cells within the alveolus suggests the likelihood of a coordinated response during postpneumonectomy lung growth. To define the population dynamics and potential contribution of alveolar epithelial cells to alveolar angiogenesis, we studied alveolar Type II and I cells during the 21 days after pneumonectomy. Alveolar Type II cells were defined and isolated by flow cytometry using a CD45(-) , MHC class II(+) , phosphine(+) phenotype. These phenotypically defined alveolar Type II cells demonstrated an increase in cell number after pneumonectomy; the increase in cell number preceded the increase in Type …

Alveolar macrophage dynamics in murine lung regeneration

In most mammalian species, the removal of one lung results in dramatic compensatory growth of the remaining lung. To investigate the contribution of alveolar macrophages (AMs) to murine post-pneumonectomy lung growth, we studied bronchoalveolar lavage (BAL)-derived AM on 3, 7, 14 and 21 days after left pneumonectomy. BAL demonstrated a 3.0-fold increase in AM (CD45(+), CD11b(-), CD11c(+), F4/80(+), Gr-1(-)) by 14 days after pneumonectomy. Cell cycle flow cytometry of the BAL-derived cells demonstrated an increase in S + G2 phase cells on days 3 (11.3 ± 2.7%) and 7 (12.1 ± 1.8%) after pneumonectomy. Correspondingly, AM demonstrated increased expression of VEGFR1 and MHC class II between days…

Pulmonary Mechanics Suggest Mechanical Forces Trigger Neoalveolarization In A Murine Model Of Compensatory Lung Growth

Dendritic Cell Migration To The Post Pneumonectomy Lung

Selective Laser Photocoagulation Manipulates Blood Flow Dynamics in Microcirculation

Blood flow patterns spatially associated with platelet aggregates in murine colitis.

In the normal murine mucosal plexus, blood flow is generally smooth and continuous. In inflammatory conditions, such as chemically-induced murine colitis, the mucosal plexus demonstrates markedly abnormal flow patterns. The inflamed mucosal plexus is associated with widely variable blood flow velocity as well as discontinuous and even bidirectional flow. To investigate the mechanisms responsible for these blood flow patterns, we used intravital microscopic examination of blood flow within the murine mucosal plexus during dextran sodium sulphate-and trinitrobenzenesulfonic acid-induced colitis. The blood flow patterns within the mucosal plexus demonstrated flow exclusion in 18% of the vessel…

QS280. Mesoscopic Traffic Flow Theory Characterizes Microhemodynamics in Chemically-Induced Murine Colitis

Spatial calibration of structured illumination fluorescence microscopy using capillary tissue phantoms.

Quantitative assessment of microvascular structure is relevant to the investigations of ischemic injury, reparative angiogenesis and tumor revascularization. In light microscopy applications, thick tissue specimens are necessary to characterize microvascular networks; however, thick tissue leads to image distortions due to out-of-focus light. Structured illumination confocal microscopy is an optical sectioning technique that improves contrast and resolution by using a grid pattern to identify the plane-of-focus within the specimen. Because structured illumination can be applied to wide-field (nonscanning) microscopes, the microcirculation can be studied by sequential intravital and confocal…

Blood flow shapes intravascular pillar geometry in the chick chorioallantoic membrane.

The relative contribution of blood flow to vessel structure remains a fundamental question in biology. To define the influence of intravascular flow fields, we studied tissue islands--here defined as intravascular pillars--in the chick chorioallantoic membrane. Pillars comprised 0.02 to 0.5% of the vascular system in 2-dimensional projection and were predominantly observed at vessel bifurcations. The bifurcation angle was generally inversely related to the length of the pillar (R = -0.47, P .05). 3-dimensional computational flow simulations indicated that the intravascular pillars were located in regions of low shear stress. Both wide-angle and acute-angle models mapped the pillars to regio…

Intravascular pillars and pruning in the extraembryonic vessels of chick embryos.

To investigate the local mechanical forces associated with intravascular pillars and vessel pruning, we studied the conducting vessels in the extraembryonic circulation of the chick embryo. During the development days 13-17, intravascular pillars and blood flow parameters were identified using fluorescent vascular tracers and digital time-series video reconstructions. The geometry of selected vessels was confirmed by corrosion casting and scanning electron microscopy. Computational simulations of pruning vessels suggested that serial pillars form along pre-existing velocity streamlines; blood pressure demonstrated no obvious spatial relationship with the intravascular pillars. Modeling a Re…

CD34+ progenitor to endothelial cell transition in post-pneumonectomy angiogenesis.

In many species, pneumonectomy triggers compensatory lung growth that results in an increase not only in lung volume, but also in alveolar number. Whether the associated alveolar angiogenesis involves the contribution of blood-borne progenitor cells is unknown. To identify and characterize blood-borne progenitor cells contributing to lung growth after pneumonectomy in mice, we studied wild-type and wild-type/green fluorescence protein (GFP) parabiotic mice after left pneumonectomy. Within 21 days of pneumonectomy, a 3.2-fold increase occurred in the number of lung endothelial cells. This increase in total endothelial cells was temporally associated with a 7.3-fold increase in the number of …

Cross-circulation and Cell Distribution Kinetics in Parabiotic Mice

Blood-borne nucleated cells participate not only in inflammation, but in tissue repair and regeneration. Because progenitor and stem cell populations have a low concentration in the blood, the circulation kinetics and tissue distribution of these cells is largely unknown. An important approach to tracking cell lineage is the use of fluorescent tracers and parabiotic models of cross-circulation. Here, we investigated the cross-circulation and cell distribution kinetics of C57/B6 GFP(+)/wild-type parabionts. Flow cytometry analysis of the peripheral blood after parabiosis demonstrated no evidence for a "parabiotic barrier" based on cell size or surface characterstics; all peripheral blood cel…

Dynamic determination of oxygenation and lung compliance in murine pneumonectomy.

Thoracic surgical procedures in mice have been applied to a wide range of investigations, but little is known about the murine physiologic response to pulmonary surgery. Using continuous arterial oximetry monitoring and the FlexiVent murine ventilator, the authors investigated the effect of anesthesia and pneumonectomy on mouse oxygen saturation and lung mechanics. Sedation resulted in a dose-dependent decline of oxygen saturation that ranged from 55% to 82%. Oxygen saturation was restored by mechanical ventilation with increased rate and tidal volumes. In the mouse strain studied, optimal ventilatory rates were a rate of 200/minute and a tidal volume of 10 mL/kg. Sustained inflation pressu…