Cigarette Smoke Negatively Affects Pulmonary Structure and Function in an Apoe - Mouse Model

The impact of cigarette smoke (CS) exposure on the respiratory system can best be captured by evaluating changes in lung function coupled with assessments of morphology and structural composition. These measurements are both quantitative and can capture the extent of disease severity such as degree, pattern, and location of tissue destruction. To date, the relationship between these structure-function outcomes have been only partially characterized. A recent publication by Matz, J., et al (2022) provides an extremely thorough series of studies to further uncover the relationship between the pulmonary changes in structure and function following chronic cigarette smoke exposure (CSE) in an apolipoprotein E-deficient (Apoe^-/-) mouse model.

SUBJECTS

Apoe^-/- mice were chosen as they are known to be an excellent model for CS-driven cardiopulmonary comorbidities.
Female subjects were selected as they are more suspectable to CS-induced cardiopulmonary dysfunction.

LUNG FUNCTION METHODS

Using the inExpose, Apoe^-/-female mice were exposed to chronic nose-only mainstream CS starting at 8-weeks of age for 5-days a week over 24-weeks. Air matched controls were exposed for 5-days a week for 24-weeks to fresh air. A full 24-hours after the last exposure (CS or air), detailed lung function parameters were measured using the flexiVent. As this chronic exposure experiment spanned 24-weeks, this team even went an additional step further to measure the natural effects of aging on lung structure and function. This was done by comparing flexiVent lung function and morphology measurements between 8-week-old mice (exposure onset) and 32-week-old mice (exposure endpoint).

FLEXIVENT MEASUREMENTS

Deep Inflation: recruitment manoeuvre and calculation of Inspiratory Capacity;
SnapShot: single-frequency manoeuvre matched to the subject’s breathing rate to give dynamic resistance (Rrs) and compliance (Crs);
QuickPrime: multiple frequency manoeuvre offering distinction between central (Newtonian Resistance; Rn) vs peripheral contribution (Tissue Damping & Elastance; G & H) in the airways;
Pressure-Volume Loop: step-wise manoeuvre to inflate the lungs to Total Lung Capacity, then backdown to Function Residual Capacity. P-V curves assess the distensibility of the respiratory system (quasi-static compliance; Cst) at rest over the entire inspiratory capacity;
Methacholine (Mch) dose response challenges: to asses airway hyperresponsiveness.

MORPHOMETRY AND MICROSTRUCTURE METHODS

Following flexiVent analysis, the subjects were perfused and fixed for histology. MOVAT and PSR stains were taken at three cross sections of the left lobe: above, below, and at the main bronchi. Next both the airways and parenchyma were analyzed for the following:

» Airways: thickness and tissue composition (cytoplasm+nucleus, fibrin, elastin, high- and low- density collagen)

» Parenchyma: thickness, tissue fraction, mean linear intercept (L_m), equivalent airspace diameter (D₀), heterogeneity, and tissue composition (cytoplasm+nucleus, fibrin, elastin, high- and low-density collagen)

RESULTS

	flexiVent Parameters	Natural Aging Effects	CS-Induced Effects
Lung Function	QuickPrime	Rn decreased in aging mice, particularly at Positive End Expiratory Pressures (PEEP) of 5 and 7 cmH₂O. G & H were decreased in aging mice at all PEEP levels (1, 3, 5, 7 cmH₂O).	Central Rn was not significantly different between the CSE and air control groups. However, the peripheral H and G were both elevated in the CSE group.
	PV-Loop	Aged mice displayed a left and upward PV-loop curve shift, with larger hysteresis and quasi-static compliance compared to young mice.	CS-exposed mice displayed a downward and rightward shift of the PV-Loop curve, indicating a structurally stiffer lung. C_st was significantly smaller as well in the CS-exposed mice.
	Mch Challenge	There was no increase in AHR in aged mice.	AHR was exacerbated at all concentrations (3-50 mg/mL) of Mch in the CSE mice.
Lung Structure	Collagen	Parenchyma: increase in high-density collagen Airways: Increase in low-density collagen	Parenchyma: increase in low-density collagen Airways: increase in low- and high- density collagen
	Airway Thickness	Decrease in airway thickness	Increase in airway thickness
	Parenchyma Airspace Morphology	L_m, D₀, and heterogeneity increased with age	D₀, airspace heterogeneity, and septal thickness increased with CSE (L_m was unchanged)

CONCLUSION

CSE over 24-weeks in female Apoe^-/- mice impaired lung function with global stiffening of the lung as evidenced by a downward-rightward shift of the PV-loop, decreased C_st, with elevated H and G at all levels of P_peep as compared to air exposed controls. These functional changes were corroborated with increased collagen deposition along with thickening of the airways and parenchyma. All of which highlight the ability of CSE in Apoe^-/- to replicate features of CS-induced COPD which is typically understated in wildtype mice.

Reference

Respiratory mechanics following chronic cigarette smoke exposure in the Apoe-/- mouse model. (2022). Matz, J., et al. Biomechanics and Modeling in Mechanobiology

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