diet, microbiota and the immune system: a gut feeling about type … · diet, gut microbiota and...
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Diet, microbiota and the immune system: A gut
feeling about type 1 diabetes
Dr. Eliana Mariño
Monash University
Melbourne, Australia
Diet, gut microbiota and
Western lifestyle diseases
From Maslowski and Mackay Nat. Immunol 2011
Type 1 diabetes
Asthma
Food allergies
IBD
- Salt
- Articial sweetners
- Food emuldifiers
Fatty liver
Diet and Microbiota
Gut
Obesity
T1D
Kidney
Metabolic Sindrome
Cancer
The role of dietary metabolites in inflammatory diseases
Type 1 Diabetes
• Blood sugar regulation lost
• Loss of insulin production and Hyperglyceamia
• Insulin producing beta cells destroyed
• Immune attack - autoimmune disease
• Controlled with exogenous insulin
• Diabetic complications are a major issue
Images: thnx to Pete Campbell & Tom Kay SVI Melbourne
(x400) (x400)
Insulin labelling Insulin labelling - pancreas T1D subject
Epidemiology of T1D
Australia
UK USA
China
Venezuela
Diabetic Medicine Volume 23, Issue 8, pages 857-866, 26 JUN 2006 DOI: 10.1111/j.1464-5491.2006.01925.x
Findland Sweeden
Mexico
Diet and Inflammatory Disease
Increase in diabetes and inflammatory diseases = ↑ popularity of Western-style diets • Diet (↓ fibre ↑ fat)
Developing countries diets (↑ fibre) = ↓ inflammatory disease
• Mediterranean diet = ↓ cardiovascular disease, asthma
Gut microbiota and Short-Chain Fatty Acids (SCFAs)
• SCFAs are produced by the gut microbiota from fermentation of dietary fibre
• Most common are:
• SCFAs are absorbed across the gut epithelium • They pass through the portal vein to the liver – the primary site of
metabolism
Dietary fibre acetate
butyrate propionate
bacteria
Hypothesis
Diet alters gut microbiota and reduced production of microbial SCFAs affect immune tolerance that increase
T1D susceptibility
Aim
To study the cellular and molecular mechanisms by which diet and SCFAs influence the course of autoimmune diabetes
SCFAs protect from T1D
b
Marino et al. Fig. 2
(%
)D
iabe
tes-f
ree
0 (Clear)
1 (<25%)
2 (25-50%)
3 (50-75%)
4 (>75%)
cNOD 5 week-old NOD 15 week-old NOD diabetic HAMS-fed NOD
15 week-old
HAMSA-fed NOD 15 week-old
HAMSA-fed NOD 30 week-old
x400
HAMSB-fed NOD 15 week-old
HAMSB-fed NOD 30 week-old
5 w
ks- o
ld 1
5 wks
-old
15 w
ks-o
ld 15 w
ks-old
30
wks-
old
NP HAMS HAMSA
15 w
ks-old
30
wks-
old
HAMSB
Diab
etic
0
25
50
75
100
Age (weeks)
30wk
s-old
HAMSA+B
HAMSA #, **
HAMSB, *
HAMSA+B ##, ***
HAMS, NS
NP, NS
a Hepatic portal blood Cecal contentFeces Peripheral blood
HAM
S
HA
MSA
HAM
SB
HAM
SHAM
SA
HAM
SB
HAM
S
HAMS
A
HAMS
B0
20
40
60
80
100
Ace
tate
(m
M)
****
NS
0
20
40
60
80
100
Bu
tyra
te (
mM
) ****
NS
0
2
4
6
8
Pro
pio
na
te (
mM
)
*
NS
HAM
S
HAM
SA
HAM
SB
0
20
40
60
80
Ace
tate
(m
M)
**
NS
HAM
S
HAM
SA
HAM
SB
0
20
40
60
Bu
tyra
te (
mM
)
****
NS
HAM
S
HAM
SA
HAM
SB
0
5
10
15
20
Pro
pio
na
te (
mM
) ***
NS
HAM
S
HAM
SA
HAM
SB
0
500
1000
1500
2000
Ace
tate
(mM
)
*
NS
0
50
100
150
Bu
tyra
te (
mM
)
*
NS
0
50
100
150
200
250
Pro
pio
na
te (m
M)
NS
NS
HAM
S
HA
MSA
HAM
SB
HAM
S
HAM
SA
HAM
SB
HAM
S
HAM
SA
HAM
SB
HAM
S
HAM
SA
HAM
SB
0
100
200
300
400
Ace
tate
(mM
)
*
NS
0
5
10
15
Bu
tyra
te (
mM
)
NS
**
0
5
10
15
Pro
pio
na
te (
mM
)
*
NS
HAM
SHAM
SA
HAM
SB
0 5 10 15 20 25 30
0
20
40
60
80
100
200mm 200mm 200mm 200mm
200mm 200mm 200mm 200mm
Fre
qu
ency o
f is
let score
(%
)
b
Marino et al. Fig. 2 (%
)D
iabe
tes-f
ree
0 (Clear)
1 (<25%)
2 (25-50%)
3 (50-75%)
4 (>75%)
cNOD 5 week-old NOD 15 week-old NOD diabetic HAMS-fed NOD
15 week-old
HAMSA-fed NOD 15 week-old
HAMSA-fed NOD 30 week-old
x400
HAMSB-fed NOD 15 week-old
HAMSB-fed NOD 30 week-old
5 w
ks- o
ld 1
5 wks
-old
15 w
ks-o
ld 15 w
ks-old
30
wks-
old
NP HAMS HAMSA
15 w
ks-old
30
wks-
old
HAMSB
Diab
etic
0
25
50
75
100
Age (weeks)
30wk
s-old
HAMSA+B
HAMSA #, **
HAMSB, *
HAMSA+B ##, ***
HAMS, NS
NP, NS
a Hepatic portal blood Cecal contentFeces Peripheral blood
HAM
S
HA
MSA
HAM
SB
HAM
SHAM
SA
HAM
SB
HAM
S
HAMS
A
HAMS
B0
20
40
60
80
100
Ace
tate
(m
M)
****
NS
0
20
40
60
80
100
Bu
tyra
te (
mM
) ****
NS
0
2
4
6
8
Pro
pio
na
te (
mM
)
*
NS
HAM
S
HAM
SA
HAM
SB
0
20
40
60
80
Ace
tate
(m
M)
**
NS
HAM
S
HAM
SA
HAM
SB
0
20
40
60B
uty
rate
(m
M)
****
NS
HAM
S
HAM
SA
HAM
SB
0
5
10
15
20
Pro
pio
na
te (
mM
) ***
NS
HAM
S
HAM
SA
HAM
SB
0
500
1000
1500
2000
Ace
tate
(mM
)
*
NS
0
50
100
150
Bu
tyra
te (
mM
)
*
NS
0
50
100
150
200
250P
rop
ion
ate
(mM
)NS
NS
HAM
S
HA
MSA
HAM
SB
HAM
S
HAM
SA
HAM
SB
HAM
S
HAM
SA
HAM
SB
HAM
S
HAM
SA
HAM
SB
0
100
200
300
400
Ace
tate
(mM
)
*
NS
0
5
10
15
Bu
tyra
te (
mM
)
NS
**
0
5
10
15
Pro
pio
na
te (
mM
)
*
NS
HAM
SHAM
SA
HAM
SB
0 5 10 15 20 25 30
0
20
40
60
80
100
200mm 200mm 200mm 200mm
200mm 200mm 200mm 200mm
Fre
qu
ency o
f is
let score
(%
)
HAMSA diet increase levels of acetate In plasma from NOD mice
Protective bacteria Non-protective bacteria
Acetate markedly reduces auto-reactive effector T cell numbers in NOD8.3 mice
Diet controls peripheral Treg numbers
-
Bar
asto
c
Hig
h Fat
die
t (HFD
)
Hig
h Fib
re d
iet
HFD
/HF
HFD
/Bar
asto
c0.0
0.2
0.4
0.6
CD
4+
CD
25
+F
ox
P3
+ (x
10
^6
)
Spleen: C57BL/6 mice
**, P=0.0083****, P<0.0001
*, P=0.01
Marino et al. Fig. 5
% F
oxp
3+
[ga
ted
on
CD
4+
]
a
Num
be
r o
f F
oxp3
+ T
reg
cells
(x106
)
HAMS
HAMSA
HAMSB
Gata3 Gitr
Sell (CD62L)
Gata3Gitr
Sell (CD62L)
83
1 2
5
46
HAMS HAMSB g
0 0
00
0
0
f
HAMS
HAMSB
HAMSA
****,##
****
HAMSA
NP
HAMS
HAMSB
Weeks post injections
b
Dia
be
tes-f
ree
(%
)
NS
NP
NS
10
0
5
10
15
20
**
NS
HAMS
HAMSA
HAMSBNP
0.0
0.5
1.0
1.5
2.0 *** **
0 5 10 15 200
20
40
60
80
100
##
###### #### ##
c d
e
Spleen
NP
HAM
S
Nu
mbe
r o
f F
oxp3
+ T
reg
cells
(x10
3)
PLN
**
NSNS
NP
HAM
S
HAMS
A
HAMS
B
Nu
mb
er
of F
oxp
3+
Tre
g
ce
lls (
x10
3)
*NS
NS
0
20
40
60
80
0
5
10
15
HAMS
A
HAMS
B
##
HAM
S
HAM
SA
HAM
SB
0
10
20
30
40
50
% C
D4
+F
oxP
3+
/IL-1
0+
HE
LIO
S+
***
Spleen
0
20
40
60
80
HAM
S
HAM
SA
HAM
SBNP
NP
% C
D4
+F
oxP
3+
/IL-1
0+
HE
LIO
S+
PLN
NS
NS
**
NSNS
Spleen
Exp
ressio
n (
log
2E
x)
Gata3 Sell (CD62L)GitrGapdh Foxp3
0
5
10
15
0
5
10
15
0
5
10
15
0
5
10
15
##
0
5
10
15
HAMS
B
HAM
S
HAMS
A
HAMS
B
Acety
lation
at Foxp3
lo
ci (A
U) H3K9
IgG
**
Acety
lation
at
Foxp3 lo
ci (
AU
) H4 Penta
IgG
HAM
S
HAMS
A
HAMS
B
NS
0
1
2
3
4****
NS
0.0
0.1
0.2
0.310
15
20
HDAC inhibition/
epigenetic mechanisms
Foxp3
Single cell PCR
Diet (particularly acetate) changes MHC I and co-stimulatory molecules on B cells (and DCs)
Control diet
Acetate diet
Butyrate diet
Effector T cells don’t proliferate when transferred to HAMSA fed mice
NOD8.3 TCR Tg CD8+ cells transferred to NOD on different diets
autoantigen (IGRP) recognised by transgenic TCR
Pancreatic LN
Mesenteric LN
SCFAs improve barrier function
LPS in serum IL-22 (gut homeostasis) in serum
Microbiota shaped by different diets differs markedly contributes to disease
susceptibility/protection
Mice on different diets
HAMSA
HAMSB
chow
Dramtic changes in microbiota composition
High fat
Germ free Mice on the same chow diet
Chow microbiome
HAMSA shaped microbiome
HAMSB shaped microbiome
High fat shaped microbiome
Propionate
Acetate
Butyrate
Bacteroides
Lactobacillus
Allobaculum
Anaeroplasma
Unclassified
Oscillospira
Coprococcus
Eubacterium
Clostridium
Ruminococcus
Dehalobacterium
Akkermansia
Parabacteroides
Colours (phylum)
Bacteroidetes
Firmicutes
Tenericutes
Undefined
Verrucomicrobiota
Marino et al. Fig. 7
HAMSA.FT
HAMSB.FT
NP.FT
HAMS.FT
*
0
20
40
60
80
100
(%)
Dia
be
tes-f
ree
c
5 10 15 20 25 30
Age (weeks)
Unknown
B. acidifaciens Bacteroides Bacteroidaceae
Bacteroidales PorphyromonadaceaeUnknown Parabacteroides
Unknown FamilyUnknown Unknown Genus
Unknown Family Unknown Unknown Genus Clostridiales
Lactobacillaceae
Unknown Lactobacillus
Lactobacillales
Bacteroidia Bacteroidetes
Clostridia Firmicutes
Bacilli
Unknown Family
Unknown Unknown Genus Unknown order Alphaproteobacteria Proteobacteria
AnaeroplasmataceaeUnknown Anaeroplasma Anaeroplasmatales Mollicutes
Tenericutes
NP HAMSBHAMSAHAMS
a
Fe
ca
l con
cent
ratio
n (m
M)
eHAMS
HAMSA
HAMSB
Glu
tam
ate
Glu
tam
ine
Glu
tam
ate
Glu
tam
ine
Ce
cal c
once
ntra
tion
(mM
)
HAMS. FT
HAMSA. FT
HAMSB. FT
0
50
100
150
200
500
1000
1500
Port
al h
epa
tic v
ein
me
tabolit
es (uM
)
Acetate Butyrate Propionate
**
HAMSHAMSA
HAMSB
d
NS
0
50
100
150
200
250 ***
NS
b
*****
***
0
1000
2000
3000
Acetate Butyrate Propionate
Ce
cal m
eta
bo
lite
s (
mM
)
***
*
0
50
100
150
200
Glu
tam
ate
Glu
tam
ine
SCFA acetate change abundance of Bacteroidetes phyla in NOD mice associated with T1D protection
Propionate
Acetate
Butyrate
Bacteroides
Lactobacillus
Allobaculum
Anaeroplasma
Unclassified
Oscillospira
Coprococcus
Eubacterium
Clostridium
Ruminococcus
Dehalobacterium
Akkermansia
Parabacteroides
Colours (phylum)
Bacteroidetes
Firmicutes
Tenericutes
Undefined
Verrucomicrobiota
Marino et al. Fig. 7
HAMSA.FT
HAMSB.FT
NP.FT
HAMS.FT
*
0
20
40
60
80
100
(%)
Dia
be
tes-f
ree
c
5 10 15 20 25 30
Age (weeks)
Unknown
B. acidifaciens Bacteroides Bacteroidaceae
Bacteroidales PorphyromonadaceaeUnknown Parabacteroides
Unknown FamilyUnknown Unknown Genus
Unknown Family Unknown Unknown Genus Clostridiales
Lactobacillaceae
Unknown Lactobacillus
Lactobacillales
Bacteroidia Bacteroidetes
Clostridia Firmicutes
Bacilli
Unknown Family
Unknown Unknown Genus Unknown order Alphaproteobacteria Proteobacteria
AnaeroplasmataceaeUnknown Anaeroplasma Anaeroplasmatales Mollicutes
Tenericutes
NP HAMSBHAMSAHAMS
a
Fe
ca
l con
cent
ratio
n (m
M)
eHAMS
HAMSA
HAMSB
Glu
tamate
Glu
tam
ine
Glu
tamate
Glu
tam
ine
Ce
cal c
once
ntra
tion
(mM
)
HAMS. FT
HAMSA. FT
HAMSB. FT
0
50
100
150
200
500
1000
1500
Port
al h
epa
tic v
ein
me
tabolit
es (uM
)
Acetate Butyrate Propionate
**
HAMSHAMSA
HAMSB
d
NS
0
50
100
150
200
250 ***
NS
b
*****
***
0
1000
2000
3000
Acetate Butyrate Propionate
Ce
cal m
eta
bo
lite
s (
mM
)
***
*
0
50
100
150
200
Glu
tam
ate
Glu
tam
ine
• Improvements to gut homeostasis/integrity
• Effects on Treg biology
• Effects on co-stimulatory molecules on B cells/DCs
• Decreased autoimmune T effector numbers
• Changes in gut microbiota composition
Summary of mechanisms
Summary
acetate
butyrate
propionate
Omega-3 fatty acids
Cardiovascular
Neural conditions
Type 1 diabetes
Asthma
Food allergies
IBD
Fatty liver
Western lifestyle diet Hygiene? Antibiotic use? Microbiota composition
Dysbiosis Leaky gut LPS distribution
Metabolites and disease HDAC inhibition
MAP kinases
PKC
b-arrestin2 PI3K
TAK NF-kB Inhibition of inflammatory cytokines
mTOR
Ga Gg
Gb
Cell shape/ motility
Metabolite sensing by GPCRs
Inflammasome activation
Ah
R
AR
NT
• IL-22 CYP1 enzymes
Transcription factors
Tan et al Annual Review Immunol 2017
Acknowledgements
• Prof Charles Mackay
• James Richards
• Keiran McLeod
• Yu Anne Yap