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Lab Metting On Friday 2011.03.11
Lab Metting On Friday 2011.03.11
MTT assay:
Baicalein reduced cell death
in adose dependent manner
Flow cytrometry:
- Absence of Baicalein apoptotic
DNA sub-G1 increase from 6.95%
(control) to 63.84%(TG) and 55.27%
(BFA).
-Baicalein alone increase 6.95 % to
9.79%
- Combine Baicalein +TG reduces
63.84 to 28.96% and Baicalein+
BFA reduces 55.27 to 26.20%
Baicalein inhibits the cleavage of caspase-12/-3 and
poly(ADP-ribose) polymerase
Western blot:
- Alone TG and BFA increase the expression of caspase-12, -3 and cleavage of
PARP
- Treatment with Baicalein reduce their expression and cleavage
Baicalein regulates unfolded protein responses
Western blot:
- TG and BFA alone
increase the expression
of CHOP, GRP78,
cleavage of XBP-1, ATF6α
and phosphorylation of
elF2α
Western blot:
- Alone TG and BFA
increase the phosphorylation
of p38, JNK, and ERK
Result: Baicalein reduces the cellular ROS level therefore it has the
anti-oxidant activity
Anti-oxidants:
N-acetyl cysteine (NAC)
C. NAC reduce the TG and BFA induced
Catalase
apoptotic death
Results:
B. NAC and catalase reduce the ROS
level more effectively than Baicalein
Results:
D+ E. NAC block the expression of CHOP and cleavage of caspase-12 in TG and
BFA induced cell
Results:
- MMP is an indicator of mitochondrial damage due to ER-stress examined by
3,3’-dihexyloxacarbocyanine iodide (DiOC 6)
- NAC reduce the MMP reduction more effectively than Baicalein
- Rotenone used as an inhibitor of mitochondrial complex –I as positive control
Conclusion
• Baicalein reduc e TG and BFA induced unfolded
protein response such as expression of CHOP and
GRP78, spliced form of XBP-1, cleavage of ATF-6, and
phosphorylation of MAPKS and elF2α
• Baicalein alone slightly induce MMP reduction and
apoptosis in HT22 cells
• Pretreatment with baicalein attenuated TG and BFA
induce ROS accumulation
• So, Baicalein exerts an anti-oxidative effect against
TG and BFA induced mitochondrial dysfunction and
apoptosis
Caspase activity assay