Transcript Slide 1 - Springer Static Content Server

Online Resource 3
S4S6
0.5
6x105
Control
1,25D3
BAF
CQ
5x105
0.4
Viable Cell Number
Absorbance (MTT)
S5
0.3
0.2
0.1
4x105
3x105
2x105
105
A
S7
D
C
o
R nt
1 r
C .71 ol
Q u
C 100 M
Q n
50 M
0
C nM
Q
C 1 uM
B Q5
A u
F
B 10 M
A n
F
B 25 M
A
B F 5 nM
A 0
F n
20 M
0n
M
0.0
0
1
2
3
4
5
6
7
Day
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40
30
##
#
20
10
5D
3+
I
R
+B
A
F
R
3+
I
5D
1,
2
1,
2
+B
A
F
IR
IR
A
F
B
on
tr
ol
0
C
Percent Positive AVO Staining
0
S5/S6 Sensitivity to chloroquine and bafilomycin A1 ZR-75-1 cells
were exposed to various doses of Bafilomycin A1 and Chloroquine and
toxicity was assessed by the MTT assay 72 hours following initial drug
treatment. Adriamycin was used as a positive control (S5). Cells were
exposed to 100nM 1,25D3, 200nM Bafilomycin A1 or 5µM
Chloroquiune and viable cell number was assessed by trypan blue
exclusion at days 3 and 6 post treatment (S6)
S7 Quantification of AVOs. Percentage of cells with positive AVO
staining was monitored using flow cytometry at 72 hours post
treatment. Values shown are from a representative experiment with
triplicate samples for each condition #p<0.05 compared to IR.
##p<0.05 compared to 1,25D3+IR
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S8
S9
S8/S9 Effects of pharmacologic autophagy inhibition on cell viability in IR and 1,25D3+IR treated cells ZR-75-1 cells were exposed to CQ, IR,
1,25D3+IR with and without CQ for 72 hours and cell viability was monitored by propidium iodide staining. Staining was quantified using flow
cytometry. Positive staining is indicated by cells gated in the M1 region (S8). ZR-75-1 cells were exposed to CQ, IR+ CQ or 1,25D3+IR +CQ for
72 hours and apoptosis was monitored by annexin v/propidium iodide staining by flow cytometry (S9).