Background:
Tuberculosis (TB) is a major global health problem, and there is an association between tobacco
smoke and TB. Water pipe smoking has become an increasing problem not only in Middle Eastern countries but
also globally because users consider it as safer than cigarettes. The presence of high levels of toxic substances in
water-pipe smoke may be a predisposing factor that enhances the incidence of pulmonary disorders. For example,
uncontrolled macropinocytosis in alveolar epithelial cells following exposure to water-pipe smoke may predispose
subjects to pulmonary infection. Here, we studied the effects of water-pipe condense (WPC) on the internalization
of Mycobacterium Bovis BCG by macropinocytosis in the alveolar epithelial cell line A549.
Methods:
A549 cells were exposed to WPC (4 mg/ml) for 24, 48, 72 and 96 h. Cell viability was studied using the
methyl thiazolyldipenyl-tetrazolium bromide (MTT) reduction assay and proliferation by bromodeoxyUridine (BrdU)
incorporation. Cells were exposed to FITC-Dextran (1 mg/ml) (as a control) and FITC-BCG (MOI = 10) for 20 min at
37 °
Cbeforecellswere
collected and the uptake of BCG-FITC determined by flow cytometry. Similar experiments
were performed at 4 °
Casacontrol
. The Rho-associated protein kinase (ROCK) inhibitor Y-27632 (1
μ
M) was used to
assess the mechanism by which WPC enhanced BCG uptake.
Results:
WPC (4 mg/ml) increased the uptake of BCG-FITC after 72 (1.3 ± 0.1 fold,
p
< 0.05) and 96 (1.4 ± 0.05 fold,
p
< 0.05) hours. No effect on BCG-FITC uptake was observed at 24 or 48 h. WPC also significantly increased the
uptake of FITC-Dextran (2.9 ± 0.3 fold,
p
< 0.05) after 24 h. WPC significantly decreased cell viability after 24 (84 ± 2%,
p
< 0.05), 48 (78±, 3%,
p
< 0.05), 72 (64 ± 2%,
p
< 0.05) and 96 h (45 ± 2%,
p
< 0.05). Y-27632 completely attenuated
the increased uptake of BCG by WPC. Cell proliferation showed a decreasing trend in a time-dependent manner
with WPC exposure.
Conclusion:
WPC exposure increased epithelial cell endocytosis activity and death as well as enhancing their
capacity for macropinocytosis. Our in vitro data indicates possible harmful effects of WPC on the ability of lung
epithelial cells to phagocytose mycobacterium.