Dmzbb final mbl
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Transcript of Dmzbb final mbl
NQO1 modulates redox status and H2O2 levels in
pancreatic β-cells
Delaine Zayas-Bazán Burgos1 , Joshua Gray2,3, Emma Heart3
1University of Puerto Rico at Cayey2U.S. Coast Guard Academy3Marine Biology Laboratory
Pancreas: Islets of Langerhans
http://discovery.lifemapsc.com/in-vivo-development/pancreas/pancreatic-ducts
insulin
Glucose Metabolism is coupled to insulin secretion in pancreatic β-cell
The role of NQO1 in metabolism
Detoxification enzyme NQO1 knock-out mice
are sensitive to oxidant stress Joseph P et al (2007), Radjendirane V. (1998)
Intermediary metabolism NQO1 levels correlated
with fasting insulin levels Palming J et al (2007)
NQO1 knock-out mice have metabolic defects Gaikwad A et al (20o1)
NQO1 is cytosolic NAD(P)H-dependent Quinone Oxidoreductase 1
Quinones are? Class of organic compound
Exogenous Food: Thymoquinone Pollution: Athraquinone
Endogenous Ubiquinone (CoQ10) Electron carrier Mitochondrial: known function All membranes including Plasma membrane
▫PMET: Plasma Membrane Electron Transport
Hydroquinone form
Semi-quinone form: reactive
NQO1 reduces H2O2 production by quinones: might be especially important for β-cells, since they have a relatively low level of classical antioxidant enzymes.
NQO1 metabolizes Quinones
NQOI cytoprotection in β-cells
β-cells have relatively low levels of classical antioxidant enzymes SOD : Superoxide dismutases 1,2 and 3
Why?Oxidants are important for signaling in β-
cell under physiological situationPathological high doses of oxidants will
harm β-cell to greater extent than other cell types
O
O
O
O
O
O
OH
OH
NAD(P)H NAD(P)+
ubiquinone ubiquinonol
O2O2
extracellular space
cytosolnet
ele
ctro
n f
low
oxidoreductases
NQO1
Mediated by NQO1
Is the transfer of electrons (reduced equivalents)out of the cell: PMET is plasma membrane analog of the mitochondrial ETC
NQO1 is a member of PMET(Plasma Membrane Electron Transport)
Gray et al (2011)
Hypotheses to test:
Objective: to determine role of NQO1 in β-cell health and metabolism.
Hypotheses: (1) NQO1 reduces oxidative stress in β-cells by lowering quinone-dependent H2O2 production.
(2) NQO1 modulates NAD(P)H-to-NAD(P)+ ratio in β-cells.
Methods
INS-1 832/13 cells
• Over-expression of NQO1▫ Adenovirus with NQO1▫NQO1 activity measured
in cell lysate by decrease in absorbance of DCPIP (dichlorophenolindophenol)
▫Activity increased >10x
NQO1+
contro
l
NQO1 (31KD)
Western blot of NQO1 protein in lysates from INS-1 832/13 cells (adapted from Gray et al 2011)
3.3E-10 ± 5.16E -12 5.6E-9 ± 1.96E-12
mol DCPIP//min
Effect of NQO1 over-expression on menadione-dependent H2O2 production in INS-1 832/13 cells
Effect of NQO1 over-expression and knock-out (KO) on menadione-dependent H2O2 production in isolated
rodent islets
Over-expression Knock-out
Hypotheses: (1) NQO1 reduces oxidative stress in β-
cells by lowering quinone-dependent H2O2 production.
(2) NQO1 modulates NAD(P)H-to-NAD(P)+ ratio in β-cells. • NADH and NADPH are substrates for NQO1• Gaikwad et al (2001): NQO1 has been show
to modulate redox ratios in fat and liver
Hypotheses
NQO1 regulates NADH-to-NAD+ ratio in INS-1 832/13 cells and rodent islets
INS-1 832/13 Rodent Islets
Conclusions
NQO1
▫Protects β-cell from oxidant stress▫Modulates β-cell redox status by lowering the NADH-to-NAD+ ratio: effect on metabolism (GSIS)
Future Studies
Role of NQO1 under metabolic stress: high glucose + high fatty acids (glucolipotoxicicty)
Role of NQO1 in PMET
Role of NQO1 in Ubiquinone redox status
Acknowledgments