Introduction to Synthetic Biology 423 2013 Herbert Sauro [email protected] .
Dalla ricerca di base alle ... - Erboristeria Sauro
Transcript of Dalla ricerca di base alle ... - Erboristeria Sauro
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Dalla ricerca di base alle applicazioni cliniche:
Cellfood TM migliora il metabolismo respiratorio delle cellule
endoteliali ed inibisce la generazione di ROS indotta da ipossia
Dr. Elisabetta Ferrero
DIBIT-Division of Molecular Oncology - San Raffaele Scientific Institute, Italy
WORKSHOP
"Inquinamento e danno da stress
ossidativo
NOVITA' dalla ricerca dal mondo
della nutraceutica»
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Endothelium is the inner cellular lining that covers all blood and lymphatic vessels
lumen
tissue
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3-dimensionality
growth factors, cytokines
chemical composition
physical characteristics
(stiffness, pH, O2 tension)
cell-cell interactions
cell-matrix interactions
mechanical stress fluid flow
pressure
AFFECT CELL BEHAVIOUR
AFFECT RESPONSE TO DRUG
Microenvironments
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Different microenvironments determine spatial EC heterogeneity
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Under physiological conditions, “quiescent” Endothelial Cells (EC) are as an always active input–output device
None
Hypoxia
+ Drug
HHV-8
+TNF
•contractile forces •pressure •drugs
TNF-a
CD-31 PECAM
NONE
ICAM-1 expression
NONE
Ferrero E, Belloni D
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to maintain a balance in the regulation of
Dilatation Constriction Growth inhibition Growth promotion Antithrombosis Prothrombosis Anti-inflammation Pro-inflammation
Endothelial Disfunction Hypoxia
ROS
TNF
VEGF
Dilatation Constriction Growth inhibition Growth promotion Antithrombosis Prothrombosis Anti-inflammation Pro-inflammation
Endothelial Disfunction Hypoxia
ROS
TNF
VEGF
Dilatation Constriction Growth inhibition Growth promotion Antithrombosis Prothrombosis Anti-inflammation Pro-inflammation
Endothelial Disfunction Hypoxia
ROS
TNF
VEGF
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need for compounds that tightly regulate endothelial
activation
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REVIEW ARTICLE
Cardiology Journal
2011, Vol. 18, No. 4, pp. 352–363
Novel therapeutic targets for preserving a healthy
endothelium: Strategies for reducing the risk
of vascular and cardiovascular disease Joseph Ramli Pedro CalderonArtero, Robert C. Block, Shaker A. Mousa1
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BE Leone, E.Ferrero
HUVEC ultrastructure
HUVEC model
Rat cerebral artery
(M Frontczak-Baniewiczi)
Normal cerebral artery
(JM Fain)
Heterotypic cells
(tumor cells)
GF, cytokines
Hypoxia Patients’sera E. Ferrero and B.E. Leone
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Hypoxia is common finding in tumors, pulmonary disorders, occlusive vascular disease, and septic shock
ROS production
Endothelial activation/dysfunction/death
Angiogenic switch
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Veschini L, Ferrero E. Blood 2007 Veschini l, Ferrero E, Faseb J 2011
EC are fine sensors of O2 activation
and are equipped with mechanisms that adjust metabolism to O2
fluctuation
HIF-1a/DAPI
Normoxia Hypoxia (2%O2)
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Specific aim
role of CellfoodTM on EC respiratory metabolism, ROS
generation and HIF-1alpha pathway
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Anti-oxidants limitations
poor solubility
inability to cross membrane barriers, poor delivery
rapid clearance from cells
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Cellfood TM does not affect EC viability and morphology
24 hrs
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CF increases O2 consumption and mitochondrial activity
Brief-time exposure Long-time exposure
T5 T1
T0
T0
T1 T5
NT
NT
Mit
oT
rak
er/
acri
din
e
ora
ng
e
Mit
oT
rak
er
NT
NT
1d
8d
8d
8d NT
O2
(%
)
CF single administration CF daily administration
…. without affecting EC viability.
to
t1 t5
nt
nt to
t1 t5
nt
1d
8d
nt 8d
nt 8d
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CellfoodTM sustains ATP production
But not LDH production:
Metabolic shift from glycolisis to mytochondrial pathway?
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Oxidative stress results from imbalance
Reductants
(antioxidants)
antioxidant supplementation could represent a common potential therapeutic strategy in conditions associated with
“increased oxidative stress”
Oxidants
(ROS)
Hormones, growth factors, proinflammatory cytokines
Proteins, lipids, DNA structural and functional
damage
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1h
r 2
4 h
rs
nox hypo nox+CF hypo+CF
nt
Time 24 hrs
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CellfoodTM down-regulates expression of hypoxia-induced HIF-1alpha
nox nox+CF hypo
nt hypo Hypo+CF nox nox+CF
and of glucose transporter Glut-1
thus CF interferes with the hypoxic response
and of glucose transporter Glut-1
thus CF interferes with the hypoxic response
HIF-1alpha/DAPI
nox nox+CF hypo+CF hypo
1 h
r
1 h
r
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1 - CellfoodTM preserves mitochondrial activity and increases O2 consumption 2 - CellfoodTM sustains ATP generation 3 - CellfoodTM inhibits hypoxia induced ROS generation 4 - CellfoodTM interferes with HIF1-a pathway 5 - CellfoodTM up-regulates MnSod expression as an adaptive mechanism
Specific Conclusions Specific Conclusions
hypo
Specific Conclusions
General Conclusion
CellfoodTM preserves EC from hypoxia-driven activation
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Endothelial Activation/ Disfunction
Relevance
CellfoodTM
Dilatation Constriction Growth inhibition Growth promotion Antithrombosis Prothrombosis Anti-inflammation Pro-inflammation
CFTM maintains proper EC
homeostasis ?
Dilatation Constriction Growth inhibition Growth promotion Antithrombosis Prothrombosis Anti-inflammation Pro-inflammation
CFTM maintains proper EC
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CellfoodTM could be useful against hypoxia induced angiogenesis?
HUVEC proliferation
0
2000
4000
6000
8000
10000
12000
48h 72h
NT
CF
Hyp
Hyp
+C
F
NT
CF
Hyp
Hyp
+C
F
n=2
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......and about other angiogenic hallmarks? -EC adhesion molecules expression and organization (VE-cadherin) -EC permeability -EC migration
perspectives
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EC response is not an all-or-nothing phenomenon
Spectrum of response depends on:
the stimulus
the spatio-temporal network
the concentration
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CONCLUSIONS
Specific
HIF-1alpha activation leads HUVEC activation/angiogenesis
General
The primordial function of HIF-1alpha was to mediate adaptive responses
that allow cells to survive oxygen deprivation
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ReactiveOxygenSpecies (ROS) are normal products of aerobic metabolism and participate in
physiological and pathophysiological processes
Bashan N, Physiol Rev, 89; 2009
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Antioxidants
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Mitochondrial hypothesis
Mitochondria must generate ATP If the set-point of a mitochondrion is low, it must work harder,
which will produce more ROS
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All these works are energy-dependent
Vander Heiden MG, SCIENCE 324 2009
Mitochondrial respiration
Anaerobic glycolysis
Aerobic glycolysis
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need for anti-oxidant and for compounds that tightly regulate endothelial
activation
interfering with HIF-1-alpha: HIF inhibitors (S.Galban, 2009; G. Melillo 2007)
down-modulating the cellular response to hypoxia recalibrating EC to their quiescent state
(P. Carmeliet, 2009)
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General AIM
treatments that can increase mitochondrial function and/or
eliminate ROS could be effective therapies?