Optimization of combined measures of airway physiology and cardiovascular hemodynamics in mice

Abstract

Pulmonary hypertension may arise as a complication of chronic lung disease typically associated with tissue hypoxia, as well as infectious agents or injury eliciting a type 2 immune response. The onset of pulmonary hypertension in this setting (classified as Group 3) often complicates treatment and worsens prognosis of chronic lung disease. Chronic lung diseases such as chronic obstructive lung disease (COPD), emphysema, and interstitial lung fibrosis impair airflow and alter lung elastance in addition to affecting pulmonary vascular hemodynamics that may culminate in right ventricle dysfunction. To date, functional endpoints in murine models of chronic lung disease have typically been limited to separately measuring airway and lung parenchyma physiology. These approaches may be lengthy and require a large number of animals per experiment. Here, we provide a detailed protocol for combined assessment of airway physiology with cardiovascular hemodynamics in mice. Ultimately, a comprehensive overview of pulmonary function in murine models of injury and disease will facilitate the integration of studies of the airway and vascular biology necessary to understand underlying pathophysiology of Group 3 pulmonary hypertension.

Keywords: Group 3 pulmonary hypertension; airway physiology; cardiovascular hemodynamics; vascular remodeling.

Fig. 1.

Baseline comparisons between treatment groups. Measurements of lung and hemodynamic function in C57Bl6J mice (18.5 weeks old) that were anesthetized with inhaled isoflurane and i.p. pancuronium followed by measurement of hemodynamics only (Hdx), lung function only (Flexi), or the two combined (Flexi + Hdx). (a) Body weight, (b) hematocrit (HCT) and (c) RV/LV + S. flexiVent analysis of (d) total resistance (Rrs), (e) system compliance (Crs), (f) elastance (Ers), (g) airway resistance (Rn) and (h) static lung compliance (Cst). Data presented as mean (±SEM). The mean is indicated by +. n = 7–9 mice/group.

Fig. 2.

Hemodynamic outcomes are not affected by flexiVent analysis. Hemodynamics (Hdx) measurement alone or following flexiVent analysis in C57Bl6J mice (18.5 weeks old) anesthetized with inhaled isoflurane and i.p. pancuronium. (a) A pressure transducer was placed in the right ventricle to measure RVSP or the (b) left ventricle to measure LVSP followed by (c) reinsertion into the RV and PA to measure PA pressure. (d) Cardiac output (CO), (e) heart rate, and (f) systemic pressure. The Hdx fell within a normal range and values were not significantly changed following flexivent analysis. Data presented as the mean (±SEM). The mean is indicated by +. n = 7–9 mice/group.

Fig. 3.

Aged mice demonstrate increased lung stiffness in the absence of hypertension. flexiVent analsysis with hemodynamics (Hdx) measurement in WT or BOE mice one year following induction. (a) Body weight, (b) hematocrit (HCT)m and (c) RV/LV+S. flexiVent analysis of (d) total resistance (Rrs), (e) system compliance (Crs), (f) elastance (Ers), (g) airway resistance (Rn), and (h) static lung compliance (Cst). (i) A pressure transducer was placed in the right ventricle to measure RVSP or the (j) left ventricle to measure LVSP followed by reinsertion into the RV to measure (k) Cardiac output (CO), (l) heart rate, and (m) systemic pressure. Data presented as mean (±SEM). The mean is indicated by +. n = 5–10 mice/group.

Fig. 4.

Coupling flexivent and hemodynamics does not affect tissue morphology. Lungs were agarose inflated using constant pressure to obtain tissue for histological H&E or trichrome staining. Representative images were presented for young WT mice: (a) Hdx only, (b) flexiVent only, and (c) flexivent + Hdx or flexivent + Hdx for aged (d) WT and (e) and (f). BOE mice. Scale bars = 100 μM. n = 7–9 mice/group.