In situ microscopic analysis of asbestos and synthetic vitreous fibers retained in hamster lungs following inhalation

Hamsters breathed, nose-only, for 13 weeks, 5 days/week, 6 hr/day, either m an-made vitreous fiber (MMVF)10a, MMVF33, or long amosite asbestos at appro ximately 300 World Health Organization (WHO) fibers/cc or long amosite at 2 5 WHO fibers/cc. [World Health Organization fibers are longer than 5 pm and thicker than 3 mu m, with aspect ratio >3.] After sacrifice, fiber burden was estimated (left lungs) by ashing and scanning electron microscopy (ashi ng/SEM) or (right middle lobes) by confocal laser scanning microscopy (CLSM ) in situ. In situ CLSM also provided three-dimensional views of fibers ret ained, undisturbed, in lung tissue. Fibers of each type were lodged in alve oli and small airways, especially at airway bifurcations, and were seen ful ly or partly engulfed by alveolar macrophages. Amosite fibers penetrated in to and through alveolar septa. Length densities of fibers in parenchyma (to tal length of fiber per unit volume of lungi were estimated stereologically from fiber transsections counted on two-dimensional optical sections and w ere 30.5, 25.3, 20.0, and 81.6 mm/mm(3) for MMVF10a, MMVF33, and low- and h igh-dose amosite, respectively. Lengths of individual fibers were measured in three dimensions by tracking individual fibers through series of optical sections. Length distributions of amosite fibers aerosolized, but before i nhalation versus after retention in the lung were similar, whether determin ed by ashing/SEM or in situ CLSM. In contrast, the fraction of short MMVF 1 0a and MMVF33 fibers increased and the geometric mean fiber lengths of both MMVFs decreased by approximately 60% during retention. Most likely due to fiber deposition pattern and differences in sampling, fiber burdens [MMVF10 a, MMVF33, and amosite thigh dose; 269 WHO fibers/cc)] determined by ashing /SEM were 1.4, 1.5, and 3.5 times greater, respectively, than those calcula ted from in situ CLSM data. in situ CLSM is able to provide detailed inform ation about the anatomic sites of fiber retention and also fiber lengths an d burdens in good agreement with ashing/SEM results.