PNNL-13501 - Pacific Northwest National Laboratory

Percent
80
60
40
20
0
Lymphocytes
60
31
Cell Type
% T Cells
%B cells
% Null Cells
9
Following BeSO4 or beryllium-oxide treatments, no
differences in the distribution of lymphocytes,
neutrophils, or eosinophils in the peripheral blood of mice
were observed, based on the differential blood counts
(data not shown). However, analysis of immune cells
isolated from the lavage fluid did suggest a temporal shift
in cell populations following beryllium treatments
(Figure 4). The percent of total cells expressing MAC-1
binding (alveolar macrophages) was about 20% at
2 weeks, decreased to about 5% at 4 weeks, but by
8 weeks post-dosing ranged from 40 to 50% of the total
cells. Overall the MAC-1 response was similar for both
the BeSO4 and beryllium-oxide treatments, although
beryllium-oxide was slightly higher by 8 weeks. A
similar temporal profile was seen for CD-69 binding cells
that represented activated T-cells, B-cells, natural killer
cells, and alveolar macrophages. These CD-69 binding
cells ranged from 5 to 20% of total cells from 2 to
4 weeks but were elevated from 40 to 60% by 8 weeks
post-treatment. In contrast, the CD-4 binding cells
(T-cells) did not show a similar profile of increasing with
time post-treatment. Although the pathological status of
the lungs were not evaluated in this project, the elevation
in MAC-1 and CD-69 binding cells at 8 weeks postdosing
may represent early cellular responses associated
with the inflammation process leading toward granuloma
formation.
Percent
80
60
40
20
284 FY 2000 Laboratory Directed Research and Development Annual Report
0
60
T Lymphocytes
% Helper cells
% Cytotoxic/
Suppressor Cells
Cell Type
Figure 3. Example of flow cytometric analysis of peripheral
blood cells obtained from naïve mice
31
Percent
Percent
Percent
60
50
40
30
20
10
0
12.5
10
7.5
5
2.5
0
80
60
40
20
0
BeSO4
BeO
2 4 8
Week
Summary and Conclusions
Mac-1 Binding Cells
CD 4 Binding Cells
2 4 8
Week
CD 69 Binding Cells
BeSO4
2 4 8
Week
Figure 4. Flow cytometric analysis of BALF cellular
distribution obtained from mice treated with BeSO 4 and
beryllium-oxide and analyzed at 2, 4, and 8 weeks posttreatment
BeO
This project investigated the early cellular dynamic
responses associated with the immune hypersensitivity to
beryllium that lead to a pulmonary granulomatous
reaction. The project established an in vivo mouse model
that may be used to evaluate early responses to beryllium
exposure. In addition, the project established the utility of
flow cytometry for assessing changes in peripheral blood
and broncial alveolar cell response to beryllium. Finally,
the results of these experiments suggest that by about
8 weeks post-exposure, there is an elevation in MAC-1
alveolar macrophages and in CD-69 activated T-cells,
B-cells, natural killer cells and alveolar macrophages in
response to beryllium exposure. This represents a
potential early indicator or biomarker of inflammatory
response to beryllium exposure.
BeSO4
BeO