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How Artificial Sweeteners Reduce Diabetes-Associated Blood Vessel Injury in
the Eye
BARS conference, 27 September 2019, 11:50-12:15
Crown Plaza Hotel, Liverpool@DrHaviChichger
Dr Havovi [email protected]
Diabetic Eye Disease
• Diabetes – 4.5 million UK, ~10% global community
• Diabetic eye disease comprises a group of eye conditions including diabetic retinopathy (DR), diabetic macular edema (DME), cataract, and glaucoma.
• All forms of diabetic eye disease have the potential to cause severe vision impairment, vision loss and blindness.
• In 2014, 54.6% patients with type I diabetes had DR; and 30% patients with type II diabetes had DR
Mathur R et al, 2017, RNIB; Diabetes UK 2018; Eschenbach Optik 2019; Mayo Clinic 2019
Clinical signs of diabetic retinopathy on fundoscopic examination
Mild, non-proliferative DR
Proliferative DR
Moderative, non-proliferative DR
Diabetic macular oedema
Wong TY, 2016, Nature Reviews Disease Primers
Pathogenic events in DR
Wong TY, 2016, Nature Reviews Disease Primers
1 2
3 4
Macular oedema (fluid leak) Proliferative retinopathy (new blood vessel formation)
Diabetes
High blood glucose
Physiological stress
Release of factors(e.g. VEGF)
Breakdown of junctions between cells
Cells divideCells become stickyCells move
Cells form tubes
Clapp C, 2009, Physiological Reviews
• Afilbercept (Eylea)• Ranibizumab (Lucentis)• Bevacizumab (Avastin)• Pegaptanib (Macugen)
Methods to study DR processes in the lab
Proliferative
Moderate
Fluid leak
Tube formation
+ VEGF
Cell migration
Chichger H, 2015, FASEB, 2016, AJRCMB
Paths
Tube
formation
Joints
paths
tubes
joints
Retinal microvascular endothelial cells
Endothelial cells
Matrigel
Fluorescent dextran
Endothelial cells
+ VEGF
Cell proliferation
-400
-350
-300
-250
-200
-150
-100
-50
0
De
cre
ase
in e
nd
oth
elia
l re
sist
ance
at
10
hr
(oh
ms)
- + - +- - + +
Sucralose:LPS:
*
*#
Harrington EO & Chichger H, 2017, AJP LungIP3, inositol triphosphate; PKC, protein kinase C; LPS. Lipopolysaccharide, *p<0.05 vs LPS vehicle, #p<0.05 vs Suc vehicle
Fluid leak
Healthy barrier
Ca2+PKC
IP3
Lung endothelial cells
Lessons from the lung – the sweet taste receptor
Sweet taste receptor
• Receptor which has remained unchangedthrough evolution - important
• Oral and extra-oral localisation
• Activated by high concentration glucose or lowconcentration of acutely-sweet artificialsweeteners
Behrens, Physiol Behav, 2011; Hass, Cell Tissue Res, 2010; Dyer J, Biochem SocTrans; 2005; Henquin, Sci Signal, 2012; Masubuchi, PloS One, 2013; Rozengurt, AJP
Gastro, 2006; Ren, Front Integr Neurosci, 2009; Foster, Plos One, 2013; Elliott, J Urol, 2011; Mosinger, PNAS, 2013; Simon, PloS One, 2014; Max, Nat Genet, 2001
Sweet taste molecules
Commercialname
Sweetmolecule
Sweet taste perception relative
to table sugar
Equal Cyclamate 30-50
Equal Aspartame 160-200
Sunett Acesulfame K 200
Sweet n Low Saccharin 300-450
Splenda Sucralose 600
NHDC Neohesperidin 1500 - 1800
Aclame Alitame 2000
E961 Neotame 7000-8000
Natural sweetmolecule
Sweet taste perception relative
to table sugar
Mabinlin 400
Pentadin 500
Brazzein 500
Monelin 800-1000
Thaumatin 3000
Does the sweet taste receptor play a role in fluid leak and new vessel formation in the retina?
+ Artificial sweeteners
+ lactisole
Saccharin (0.1 µM); Sucralose (0.1 µM)
Proliferative
Moderate
Fluid leak
Tube formation
+ VEGF
Cell migration
Chichger H, 2015, FASEB, 2016, AJRCMB
Paths
Tube
formation
Joints
paths
tubes
joints
Retinal microvascular endothelial cells
Endothelial cells
Matrigel
Fluorescent dextran
Endothelial cells
+ VEGF
Cell proliferationVEGF – vascular endothelial growth factor
Sucralose reduces VEGF-induced leak
VEGF: - -+ +Vehicle Sucralose
- -+ +Vehicle Sucralose
Vehicle Lactisole
* * *
0
2
4
6
8
10
12
14
16
% p
erm
ea
bili
ty
Fluid leak
Healthy barrier
Lizunkova P, 2019, Graefe's Clinical and Experimental Ophthalmology*p<0.05 vs vehicle for VEGF; n = 6
+ Artificial sweeteners
+ lactisole
Fluorescent dextran
Endothelial cells
- dependent on sweet taste receptor
+-VEGF
0 hr
6 hr
0
10
20
30
40
50
60
70
Cel
l Mig
rati
on
(µ
m/h
r)
* * *
VEGF: - -+ +Vehicle Saccharin
- -+ +Vehicle Saccharin
Vehicle Lactisole
*p<0.05 vs vehicle for VEGF; n = 4-6. x 10 magnification, scale bar 20 µm
+ Artificial sweeteners
+ lactisole
0
2
4
6
8
10
12
14
Nu
mb
er o
f tu
bes
* * *
VEGF: - -+ +Vehicle Sucralose
- -+ +Vehicle Sucralose
Vehicle Lactisole
Artificial sweeteners reduce angiogenesis
+-
VEGF
Lizunkova P, 2019, Graefe's Clinical and Experimental Ophthalmology
Src
RhoA
VEGFR2
VEGF
Akt
AdhesionProliferation
Migration
Tube formationProliferative (tube formation)
Diabetes
Mean ± SD, *p<0.05 vs vehicle for VEGF
*
#
# #
*
V e h i c l e V e h i c l eS u c r a lo s e S u c r a lo s e
Veh
icle
VE
GF
Veh
icle
VE
GF
Veh
icle
VE
GF
Veh
icle
VE
GF
0
5
1 0
1 5
2 0
Tu
be
fo
rm
ati
on
(nu
mb
er
of
tub
es
)
S C 7 9V e h ic le
*
V e h ic le V E G F V e h ic le V E G F
0 .0
0 .5
1 .0
1 .5
Ph
os
ph
ory
late
d A
kt
(ra
tio
of
un
ph
os
ph
ory
late
d A
kt)
S u c ra lo s eV e h ic le
:Sucralose
P-Akt
Akt
:VEGF- +- +
- - + +
T1R3
SC79 – Akt activator; Akt – protein kinase B; Src – kinase; VEGFR2 – VEGF receptor Lizunkova P, 2019, Graefe's Clinical and Experimental Ophthalmology
Src
VEGFR2
PROTECTION
T1R3
VEGF
Akt
AdhesionProliferation
Migration
Tube formation
Proliferative (tube formation)
Diabetes
???
PAK VE-cadherin
PI3K p110α
Pyk2 RhoA
P
Actin remodelling
Cell contact disassembly
MLC2
Moderate (fluid leak)
Cell junction
Suez, Nature, 2014
The controversy
If people with diabetes eat more Splenda, will it stop their proliferative retinopathy getting worse?
Artificial sweeteners in the diet –negative consequences
*
0
0.0
10.1 1
10
100
1000
10000
50000
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
A rtif ic ia l s w e e te n e r (µ M )
Ca
co
-2 c
ell
via
bil
ity
(%
)
S a c c h a r in
S u c ra lo s e
A s p a rta m e
N e o ta m e
IC 5 0
*
*
**
*
**
** *
Mean ± SEM, *p<0.05 vs 0 µM Invasion index = CFU/Caco2 cell # Steinberg, AJP Cell, 2006
0 1 0 0 0
0
1
2
3
4
A rtif ic ia l s w e e te n e r (µ M )
E.f
ae
ca
lis
in
va
sio
n i
nd
ex
(no
rma
lis
ed
to
0
M)
S a c c h a r in
S u c ra lo s e
A s p a r ta m e
N e o ta m e
*
0 1 0 0 0
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
S a c c h a r in (µ M )
RO
S p
ro
du
cti
on
(% n
orm
ali
se
d t
o 0
M)
*
The future: The therapeutic implications of taste sensors in the microvasculature
Acknowledgements
• Prof Elizabeth Harrington
• Alex Vang
• Julie Braza
Related publications:
Thank you for your attention