1. Master Thesis By Judy Mutua
Phosphatidyl Serine Exposure in the Membrane of RBCs after inhibition of PKC and Scramblase
Master Thesis
By Judy Mutua

2. Outline Introduction Method Results Conclusion
After a clot forms, the actin and myosin in platelets start the contraction process and cause the clot to shrink to about one-third of its original size. This is an important step to stem bleeding, to decrease obstruction in blood vessels, and to provide a matrix for migration of cells involved in wound healing. Red blood cells are caught up in the contraction process, especially in the venous system, and get pulled by the platelets towards the interior of the clot, as the Blood study indicates.

3. Introduction Red Blood Cells: Role in blood clot formation.
Transport oxygen
120 days life span
Produced in Bone Marrow
Role in blood clot
formation.
qmul.ac.uk

4. Introduction Structure of RBC membrane
The human RBC membrane consists of: Lipids, Proteins
& Carbohydrates.
The main lipids are cholesterol and phospholipids Outer monolayer: (PC); (SM)
Inner monolayer: (PE); (PI); (PS)

5. Introduction
RBC Membrane
Modifiziert nach Zwaal et al., 2005

6. Introduction

7. Introduction

8. Introduction

9. Introduction
Calphostatin c
R5421

10. Method Wash 500 µl blood with physiological solution 3X
Incubate in 1 µM X-Rhod-1 (30 min)
Incubate with inhibitor
Add Annexin V-FITC
Measure in FACS
flourescent microscope

11. Results
Nguyen et al. 2011

12. Results
PS-Exposure of RBCs after 30 min activation with LPA, A23187 or PMA (Nguyen et al., 2011)

13. Results

14. Conclusion
Activation of RBCs by LPA ,A23187 or PMA leads to increase in the intracellular calcium.
Increasing of intracellular calcium activates the calcium pump, this leads to decrease in
intracellular calcium with time.
Cells with increased intracellular calcium content and showing PS exposure don’t always correlate together. Further experiments will solve this discrepancy!

15. References
L. Wagner, D.B. Nguyen, A. Jung, P. Steffen, C. Wagner, L. Kaestner, T. Mueller, I. Bernhardt, „Phosphatidylserine Exposure and Aggregation of Red Blood Cells“ (2011) Red Cell Club, Philadelphia
B. N. Yamaja Setty, S. Kulkarni, M. J. Stuart, “Role of erythrocyte phosphatidylserine in sickle red cell-endothelial adhesion” (2002) blood :
D. B. Nguyen, „Phosphatidylserine exposure in red blood cells: A suggestion for the active role of red blood cells in blood clot formation” (2010) Dissetation Saarbrücken
C. Le Closse. J. Dachary-Prigent, M. R. Boisseau, “Phosphatidylserine-related adhesion of human erythrocytes to vascular endothelium“ (1999) British Journal of Haematology, 1999, 107, 300±302
J. Connor, C. C. Pak, A. J. Schroit, “Exposure of Phosphatidylserine in the Outer Leafle of Human Red Blood Cells” (1993) THE JOLIRNAOFL BIOUXICACLHEMISTRY