What Can We Do Together To Understand DBA?...Kelly O’Brien DBA CELLS PROLIFERATE SLOWLY D Control...
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What Can We Do Together To Understand DBA?
Transcript of What Can We Do Together To Understand DBA?...Kelly O’Brien DBA CELLS PROLIFERATE SLOWLY D Control...
What Can We Do Together To Understand DBA?
CULTURE OF DBA CELLS
Ficoll gradient
Magnetic
labeling of CD34+ cells
Selection of CD34+ cells
~15,000
Day 0-7 Day 8-14 SCF, TPO, IL-3, SCF, TPO, IL-3, IL-6, EPO (0.5U) IL-6, EPO (3U)
CULTURE OF DBA CELLS
Kelly O’Brien
DBA CELLS PROLIFERATE SLOWLY
D
Control
DBA
0
1,000,000
2,000,000
3,000,000
4,000,000
5,000,000
6,000,000
7,000,000
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16
Num
ber o
f Cel
ls
cells051415_control.fcsEvent Count: 44982
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<PE
-A>:
CD
41
23.3 13.7
17.245.9
cells051415_0649.fcsEvent Count: 42750
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<PE
-A>:
CD
41
22.8 18.9
30.727.6
cells051415_0721.fcsEvent Count: 40972
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<PE
-A>:
CD
41
25.4 26.9
28.519.2
cells051415_0649.fcsEvent Count: 42750
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<Per
CP
-Cy5
-5-A
>: C
D44
32.2 53.1
6.198.55
cells051415_control.fcsEvent Count: 44982
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105<P
erC
P-C
y5-5
-A>:
CD
4433.7 62.9
1.631.78
cells051415_0721.fcsEvent Count: 40972
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<Per
CP
-Cy5
-5-A
>: C
D44
30.1 60.3
7.891.73
CD235
CD
44
CONTROL DBA
CD235
CD
44
STEROID RESPONSIVNESS IN A DBA PATIENT
EPIGENETIC REGULATION OF GENE EXPRESSION Chromatin Accessibility
closed open
DNA Methylation
Methylation site Methylation site
cells051415_control.fcsEvent Count: 44982
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<PE
-A>:
CD
41
23.3 13.7
17.245.9
cells051415_0649.fcsEvent Count: 42750
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<PE
-A>:
CD
41
22.8 18.9
30.727.6
cells051415_0721.fcsEvent Count: 40972
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<PE
-A>:
CD
41
25.4 26.9
28.519.2
cells051415_0649.fcsEvent Count: 42750
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<Per
CP
-Cy5
-5-A
>: C
D44
32.2 53.1
6.198.55
cells051415_control.fcsEvent Count: 44982
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105<P
erC
P-C
y5-5
-A>:
CD
4433.7 62.9
1.631.78
cells051415_0721.fcsEvent Count: 40972
0 102 103 104 105
<FITC-A>: CD235
0
102
103
104
105
<Per
CP
-Cy5
-5-A
>: C
D44
30.1 60.3
7.891.73
CD235
CD
44
CONTROL DBA
CD235
CD
44
RNA DNA methylation Chromatin Accessibility
RNA DNA methylation Chromatin Accessibility
RNA DNA methylation Chromatin Accessibility
RNA DNA methylation Chromatin Accessibility
EPIGENETIC ANALYSIS OF DBA PATIENT CELLS
Next Steps: Establish a baseline of transfusion dependent DBA patients and controls. Analyze steroid responsive DBA patients. Analyze DBA patient in remission. Correlate epigenetic changes with changes in transcriptional profiles.
Goal: To identify genes or pathways that are altered in steroid responsive or remission DBA patient cells
