DNA damage checkpoint activation contributes to the radio-resistance - - PowerPoint PPT Presentation

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Feb 05, 2024 740 likes •879 views

DNA damage checkpoint activation contributes to the radio-resistance of mouse mesenchymal stromal cells Tara Sugrue, & Rhodri Ceredig Regenerative Medicine Institute Centre for Chromosome Biology National University of ireland, Galway

SLIDE 1

DNA damage checkpoint activation contributes to the radio-resistance of mouse mesenchymal stromal cells

Tara Sugrue, & Rhodri Ceredig Regenerative Medicine Institute Centre for Chromosome Biology National University of ireland, Galway

SLIDE 2

The DNA Damage Response contributes to MSC radio-resistance

SLIDE 3

Haematopoietic stem cell (HSC) Mesenchymal stromal cell (MSC)

IR

Die Survive

Radiation Response of the Bone Marrow

Haematopoietic failure

Rieger K, Marinets O, Fietz T et al. Mesenchymal stem cells remain of host origin even a long time after allogeneic peripheral blood stem cells or bone marrow transplantation. Exp Hematol 2005;33:605-611. Dickhut A, Schwerdtfeger R, Kuklick L et al. Mesenchymal stem cells obtained after bone marrow transplantation or peripheral blood stem cells transplantation originate from host tissue. Ann Hematol 2005;84:722-727.

SLIDE 4

Isolation of mouse MSCs

Heterogenous population. “CFU-F” Clonal expansion Mouse MSC line Homogenous population

CD73 CD90 CD105 Sca-1 CD44

‘’Bulk MSCs’’

SLIDE 5

Role of the DNA Damage Response in mediating the radio- resistance of MSCs

V

Mouse haematopoietic cell line ST4.5

CD4+ CD8+ Thymocyte

Mouse mesenchymal stromal cell line MS5 Radio-sensitive Radio-resistant

SLIDE 6

DSB

The DNA Damage Response

Signalling to effector proteins Scaffolding proteins Effector kinases Regulatory proteins

Apoptosis Cell Cycle Arrest DNA Repair

IR

Chromatin

Activation of sensor and transducer proteins

Transcription

SLIDE 7

Tracking cell cycle progression

G1 S G2 M BrdU

G1 S G2+ M

PI BrdU Cell count

G1 S G2+M

DNA content (PI)

2n 4n

SLIDE 8

12 hrs 4 hrs 2 hrs

MSCs activate IR-induced DNA Damage checkpoints

MS5

G1 S G2/M

Time post irradiation (10 Gy) 0 hrs

BrdU PI

ST4.5

10 20 30 40 50 60 70 80 90 1 2 4 6 8 12 24 36 48 72

Time (Hrs) % S phase population

MS5 Control MS5 10 Gy ST4.5 10 Gy ST4.5 Control

SLIDE 9

MS5

G1 S G2/M

72 hrs Time post irradiation (10 Gy) 0 hrs 12 hrs

BrdU PI

ST4.5 24 hrs

MSCs recover from DNA damage checkpoint activaton

MS5 Control MS5 10 Gy ST4.5 10 Gy ST4.5 Control

10 20 30 40 50 60 70 80 1 2 4 6 8 12 24 36 48 72

Time (Hrs) % BrdU labelled G1 population

SLIDE 10

MSCs suppress IR-induced Apoptosis

20 40 60 80 100 2 4 8 12 24 36 Time post 10 Gy irradiation (Hrs) % Annexin V positive cells MS5 ST4.5

Time post irradiation (10 Gy) 2 hrs Untreated 12 hrs 36 hrs 4 hrs ST4.5 MS5

Annexin V PI

SLIDE 11

Conclusions

Ø MSCs survive long-term post high dose γ-irradiation. Ø MSCs activate a robust DNA Damage Response via rapid γ-H2AX formation. Ø MSCs recover from IR-induced DNA damage by: (i) Activating DNA damage checkpoint mechanisms. (ii) Activating DNA DSB repair. (iii) Preventing induction of apoptosis.

SLIDE 12

Immunology Group

Prof. Rhodri Ceredig
Prof. Matthew Griffin

Immunology and Transplant Biology Group

Dr. James Brown

Acknowledgements

Centre for Chromosome Biology

Prof. Noel Lowndes

Genome Stability Laboratory

Dr. Michael Rainey
Dr. Alessandro Natoni

Simona Moravcová Michelle Duffy Senthilkumar Alagesan

NUI Galway

Regenerative Medicine Institute