Poster for the UCSD Faculty Mentor Program (FMP) Symposium Presented: May 24, 2016

1. Benjamin Yanga, Sung Sik Hura,b, Yingxiao Wanga,b, and Shu Chiena,b
aDepartment of Bioengineering, bThe Institute of Engineering in Medicine, University of California at San Diego, La Jolla, CA 92093
1. 3D Traction Force Microscopy (3D-TFM): To determine cell-ECM forces
INTRODUCTION
 Mechanical force can modulate
cell functions
 FAs mediate cell-ECM force
(traction force)
 AJs mediate cell-cell force
(intercellular force)
2. Glioblastoma (GBM), Doxorubicin
1. Cells and their environment can communicate through mechanical
interactions.
Force
Adherens
Junction
(AJ)
Mechanosensing
Extracellular Matrix
Force
Mechanosensing
Focal Adhesion
(FA)
Gene Expression
Cell Responses
Adherent Cells
SPECIFIC AIMS
1. To develop a method to simultaneously monitor epigenetic
modifications and intracellular stress in real-time
2. To quantify cell-ECM and cell-cell stresses of cancer cells in different
mechanical microenvironments
3. To investigate the molecular mechanisms by which cancer cells react to
chemotherapeutics
METHODS
Image
Displacement
Traction Force
Cells on PAA
Image Processing
Confocal Microscopy
Finite Element Analysis
TFM Sequence Confocal Microscopy
Hur SS et al. Cell Mol Bioeng. 2009
TS: Traction Stress
JT: Junctional Tension
IT: Intracellular Tension
Single Cell
Multiple Cells
TS
JT
IT
TS
TS
TS
TS
TS
∫∫ TS = 0
∫∫ TS + ∫ JT = 0
∫∫ TS + ∫ IT = 0
 When a cell is isolated as a single, total sum of cell-ECM force should
be balanced to be zero. TS can be determined from TFM.
 When a cell is in contact with other cells, total sum of cell-ECM force
and cell-cell force from neighboring cell should be balanced to be
zero. Junctional tension (JT) can be determined from TS.
 Intracellular tension (IT) can be determined similarly from TS by
splitting the domain along an intracellular section line.
Hur SS et al. PNAS. 2012
2. 3D Intracellular Force Microscopy (3D-IFM) : To determine cell-cell and
intracellular forces
CONCLUSIONS
RESULTS
Finite Element Analysis
Computing force from deformation
Image Processing
Tracking Substrate Deformation
Progression of glioblastoma cells and mechanical-epigenetic behaviors
3. FRET Biosensors: To monitor tri-methylation of H3K9
u87 u87vIII
Soft
u87 u87vIII
Hard
CT DOX CT DOX CT DOX CT DOX
Cell-ECMIntracellular
 Doxorubicin is a common
chemotherapeutic anti-mitotic drug
 GBM are usually highly malignant
cancerous astrocytes
 u87 and u87vIII are GBM cell lines;
u87vIII is a “sister” maligned line that
overexpresses the EGFRvIII gene
Figure 1. Intracellular and cell-ECM forces for u87 and u87vIII cells on
soft (1.7 kPa) and hard (21 kPa) gels.
CT = control, D=doxorubicin for 30 min
* p < 0.05, ** p<0.01, *** p < 0.001
u87 u87vIII
Soft
u87 u87vIII
Hard
CT DOX CT DOX CT DOX CT DOX
Figure 2: H3K9me FRET biosensor signal intensity for u87 and
u87vIII cells on soft (1.7 kPa) and hard (21 kPa) gels
a.u
Pa
CT DOX (30m)
u87
CT DOX (30m)
u87vIII
Figure 3: FRET imaging of u87 and u87vIII cells
REFERENCES
 u87 and u87vIII cells respond differently to treatment with DOX
 The more malignant u87vIII cells exhibit lower intracellular and cell-ECM
stresses than u87 cells for soft and hard microenvironments with DOX
 H3K9me trimethylation increases with DOX treatment on stiff and soft
gels for both u87 and u87vIII cells except for u87 cells on soft gels
 There is a correlation between H3K9 trimethylation and intraceullar
stress in nuclear regions
Parekh A & Weaver AM, 2009, Cell Adhesion & Migration
Jiang P, Mukthavavam R et al., 2014, Journal of Translational Medicine
Hur SS et al., 2009, Cellular and Molecular Bioengineering.
Hur SS et al., 2012, PNAS
Seong J et al. 2009, Chemistry & biology
Doxorubicin intercalating DNA
FUTURE WORK
 Develop method to directly quantify 3D intracellular forces
 Explore the effect of different drugs on the epigenetic and mechanical
responses of glioblastoma cells
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***
**
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20µm