In combination, these data indicate that L. rhamnosus and L.
reuteri colonization caused a cell cycle deceleration in ME-180 cells,
theoretically prolonging the cell cycle by 5–10 hours.
The Frequency of Cell Division Differs among the
Lactobacillus Strains
During the observations of 900 cells by live-cell microscopy, it
came to our attention that for certain strains, the number of cells
in late mitosis was not equally distributed over the 16 hours of the
assay. Subsequently, the number of completed cytokinesis events
that was observed in every timeframe of 10 minutes for the
duration of 16 hours was analyzed. For an average of 900 cells that
were followed during the 16-hour assay, the ME-180 cells alone
performed 1.7 cell divisions every 10 minutes (Fig. 2). Colonization
with L. rhamnosus, and L. reuteri produced lowered cell division
frequencies of 1.4 and 1.5 cell divisions every 10 minutes,
respectively, over the same time period (Fig. 2). By contrast, cells
that were colonized with L. crispatus shifted their modes of cell
division frequency between 7 and 11 hours; at those times, these
cells exhibited an average of 2.3 cell divisions every 10 minutes
with peaks of up to 5 cell divisions every 10 minutes (Fig. 2).
However, the increased number of cell divisions that was induced
by L. crispatus did not alter the overall average proliferation rate.
These data demonstrate the ability of L. crispatus to transiently
enhance the frequency of cell division.
S phase Progression is Reduced by L. rhamnosus and L.
reuteri but not L. crispatus
To further verify the finding that Lactobacillus colonization
reduces cell cycle progression, the number of cells in S phase at the
end of 24 hours of incubation was measured. Lactobacilli were
allowed to adhere to ME-180 cells for 24 hours. Bromodeoxyuridine
(BrdU) was added to cells two hours prior to the termination
of this assay, allowing the BrdU to be incorporated into the
replicating DNA of cells in S phase. The adherence by all of the
lactobacillus strains that were tested except for L. crispatus resulted
in a reduced number of ME-180 cells in S phase. L. rhamnosus and
L. reuteri reduced the level of BrdU incorporation to an average of
31% and 59%, respectively, relative to the BrdU incorporation
level of control ME-180 cells (Fig. 3). Thus, this finding once
again demonstrates that colonization with L. rhamnosus and L. reuteri
but not L. crispatus inhibits cell cycle progression in ME-180 cells