5780447303

152 B. Rihn et al. I To.ricology 109 (1996) 147-156

Table 2

Cellular and TNF-st response to crocidolite fiber inhalation in bronchoaWeolar liquids

Number Yiable cells* Pulmonary alveolar macrophages # Polymorphonuclear Lymphocytes #TNF-**

cells*

N = 1    N~2 N > 2

Control    N - 3    93 ± 28    85.6 ± 6.6    0.6 ± 0.2    0    6.5 ± 6.5    5.6 ± 0.3    43.2 ± 52.0

Trcated    N - 9    118 ± 24    103.8 ± 13.0    4.8 ± 1.1    0.8    ± 0.4 7.1 ± 14.1    20.1 ± 8.3    36.5 ± 23.9

At least three determinations were carried out for each bronchoalveolar liquid. PAM N = 1, PAM N = 2, PAM N > 2 were mononucleated, binucleated and polynucleated macrophages, respectively. Total viable cells, PAM, polymorphonucleated cells, lymphocytes were expressed as x 10^_s/ml. TNF-a in unit/mg of protein. Statistical tests: dilTerences between means (smali samples), #P < 0.01; ^mP > 0.05.

similar increase was also noticed for mononucleated pulmonary alveolar macrophages (PAM): 103000 ± 13000 cells/ml in the intoxicated mice vs. 85 500 ± 6600 in the Controls. As for binucleated PAM (4800 ± 1100/ml in exposed vs. 600 ± 200 in control mice), the changes were significant in contrast with the ones observed for polymorphonucleated cells. Giant PAM (800 ± 400/ml) corresponding to multinucleated PAM as exemplified in Fig. 3B, were detected only in the crocidolite exposed group. This latter endpoint was introduced considering that these giant cells were specific of the fiber burden in the mouse lungs. It is noteworthy that even 5 weeks after a 5-day intoxication challenge, an inflam-matory response was still obvious showing mac-rophage distribution changes. The total lym-phocyte count was higher in the treated animals than in the control: 20100 + 8300 vs. 5600 ± 300. But surprisingly we did not observe any increase in the TNF-a levels in any group of mice: 36.5 ± 23.9 in treated bronchoalveolar fiuids vs. 43.2 ± 52.0 U/mg in control ones. As TNF-a is a multifunctional cytokine elaborated primarily by monocytes and activated macrophages (Rosenblum and Donato, 1989; Martinet et al, 1992; Warheit, 1994), one could expect a modified expression level. This in fact was not seen in our study.

Histologic examination of the lungs demon-strated the presence of macrophages in the lumen of the terminal bronchioles and alveoli. Perivas-cular and peribronchiolar inflammatory cells were observed in intoxicated mice. The inflammatory response corresponds to a Wagner scalę value of 2 as it was defined for studies involving rats (namely, presence of macrophages in the lumen of terminal bronchioles and alveoli, McConnell et al, 1984). By comparison control mice had no such changes. Asbestos bodies were seen in terminal bronchioles (Fig. 3A).

Very large macrophages as shown in Fig. 3B were seen on the May Grunewald Giemsa stained samples. Five weeks after the intoxica-tion, the long fibers seem to attract many macrophages with giant cellular polynucleated bodies limited by one membranę or limited each by their own membranę (Fig. 3B).

Ultrastructural studies have shown numerous macrophages internalizing long fibers measuring morę than 20 /im as shown in Fig. 3C.

4. Discussion

Rats are generally considered as a paradigm close to human diseases caused by fiber inhala-tions, sińce they develop lung tumors (adenocar-cinomas or mesotheliomas) and pulmonary asbestosis like humans (Davis and Donalson, 1993). Using the rat paradigm, it was well-estab-lished that short fibers do not produce pulmonary disease, medium length fibers are etiologie factors for mesotheliomas while the longer fibers lead to fibrosis and lung adenocarcinomas (Stan-ton et al. 1981; Barrett et al, 1989).

It is difficult to extrapolate results obtained with rodents to humans. Indeed rodents have a

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