1. The Interaction of Garlic With Mediators of Inflammation
Prof. A. M. Vollmar
Dr. V. M. Dirsch
H - P Keiß

2. Overview Interaction of garlic powders and constituents with NF-B
Assay setup
Results
Summary
Interaction of garlic powders and constituents with cytokines
Outlook

3. NF-B activation

4. Dual luciferase reporter gene assay
Induceable
Firefly luciferase
Not induceable
Renilla luciferase
I-B
Nucleus
TNF
NF-B
Luciferase

5. Transfection of Human Umbilical Vein Endothelial Cells
Transfection of HUVEC with Effectene®-transfection reagent
Low transfection efficiency (Max. - 10%)
No reliable results

6. Transfection of Human Embryonic Kidney 293 Cells
Transfection of HEK 293 cells with Ca3(PO4)2-method
High efficiency (~ %)
Reliable results

7. NF-B reporter gene assay

8. NF-B reporter gene assay

9. Examined garlic constituents and powders
Diallyldisulfide (Jacques Auger)
Allicin (Jacques Auger)
-Glutamylcysteine (Thomas Haffner)
Garlic powders
Morado 200 ( Hans Princen)
Morasol 0 (Remi Kahane)
Morasol 100 (Remi Kahane)
Printanor 0 (Remi Kahane)
Printanor 200 (Remi Kahane)

10. Garlic powders in DMEM

11. Garlic powders in DMSO

12. Summary
Garlic constituents showed no inhibition of TNF induced NF-B activation.
Only Printanor 0 solved in Media was able to inhibit TNF induced NF-B activation.

13. Interaction of garlic with cytokines
LPS
Stimulation
Il 1ß, Il 10
TNF 
Liberation
Human whole blood
+ complete media
+ garlic
Interleukin and TNF 
levels are measured with
ELISA - technique

14. Results of cytokine assay
Garlic constituents
Allicin and -Glutamylcysteine had no significant effect
DADS showed dose dependent reduction for Il-1 and Il-10

15. Il-1 

16. Il-1

17. TNF

18. TNF

19. Il-10

20. Il-10

21. Summary
Garlic extracts are able to reduce LPS induced liberation of the pro-inflammatory cytokines Il-1 and TNF.
Garlic extracts are able to increase the LPS induced liberation of anti-inflammatory cytokine Il-10.
It is noticeable that unfertilised garlic is less active than fertilised garlic.
DMSO garlic extracts show stronger inhibition of liberation of Il-1 and TNF than RPMI garlic extracts.
The liberation of cytokines through RPMI garlic extracts is most likely mediated by endotoxin contaminations.

22. Outlook NF-kB reporter gene assay Cytokine assay Optimise assay
Experiments with samples from 2001 field trail
Other transcription factors (AP-1)
Cytokine assay
Complete experiment
Check endotoxin contaminations with Limulus test