Many studies that demonstrate an increase in airway smooth muscle in asthmatic patients rely on the assumption that bronchial internal perimeter (P_i) or basement membrane perimeter (P_bm) is a constant; i.e. not affected by fixation pressure or the degree of smooth muscle shortening. Since it is the basement membrane that has been purported to be the indistensible structure, this study examines the assumption that P_bm is not affected by fixation pressure. P_bm was determined for the same human airway segment (n=12) fixed at distending pressures of 0 cmH₂O and 21 cmH₂O in the absence of smooth muscle tone. P_bm for the segment fixed at 0 cmH₂O was determined morphometrically and the P_bm for the same segment, had the segment been fixed at 21 cmH₂O, was predicted from knowing the luminal volume and length of the airway when distended to 21 cmH₂O (organ bath-derived P_i). To ensure an accurate transformation of the organ bath-derived P_i value to a morphometry-derived P_bm value, had the segment been fixed at 21 cmH₂O, the relationship between organ bath-derived P_i and morphometry-derived P_bm was determined for 5 different bronchial segments distended to 21 cmH₂O and fixed at 21 cmH₂O (r² = 0.99, P < 0.0001). Mean P_bm for bronchial segments fixed at 0 cmH₂O was 9.4 ± 0.4 mm, whereas mean predicted P_bm had the segments been fixed at 21 cmH₂O was 14.1 ± 0.5 mm (P < 0.0001). This indicates that P_bm is not a constant when isolated airway segments without smooth muscle tone are fixed distended to 21 cmH₂O. The implication of these results is that the increase in smooth muscle mass in asthma may have been overestimated in some previous studies. Therefore further studies are required to examine the potential artifact using whole lungs with and without abolition of airway smooth muscle tone and/or inflation.