NYU School of Medicine
METABOLIC ALTERATIONS TO THE MUCOSAL
MICROBIOTA OF INFLAMMATORY BOWEL
DISEASE PATIENTS ARE ASSOCIATED WITH
CD4+ T CELL HOMEOSTASIS
Michael Davenport, Jordan Poles, Jacqueline Leung,
Martin J. Wolff, Wasif M. Abidi, Thomas Ullman,
Lloyd Mayer, Ilseung Cho, and P’ng Loke.
12/13/13
DISCLOSURES
• Nothing to disclose
Presentation Title Goes Here
2
Background
•
Pathogenesis of IBD is likely related to abnormal interactions
between the immune system and gut microbiota
•
Studies using 16S sequencing have shown that IBD is associated
with dysbiosis of the gut microbiota:
• Decreased microbial diversity
• Alterations in specific bacterial taxa
•
The mucosal microbiota is distinct from the luminal microbiota
•
Inflammation may alter nutrient availability to mucosal bacteria and
impact their metabolic function.
•
Microbial metabolites may also regulate intestinal CD4+ T cell
homeostasis.
3
Methods
•
Paired pinch biopsy samples of known inflammation
states were collected and analyzed from patients with UC
(n = 23) and CD (n = 21) at Mt Sinai
•
Active inflammation was defined histopathologically by
neutrophil infiltration into the epithelium
•
Previously, we found that TH22 cells were depleted in
actively inflamed tissue only from these UC patients and
not CD patients. (Leung et. al. Mucosal Immunology
2013)
4
Methods
•
16S sequencing can be used identify changes to the
mucosal microbiota from pinch biopsies of known
inflammation states
•
However, there is not enough bacterial DNA to determine
microbial function by shotgun metagenomic sequencing
analyses
•
PICRUSt is a new technique for inferring the
metagenome through the closest available bacterial
whole genome sequences in available databases using
the 16S data. (Langille et al. Nature Biotechnology 2013)
5
Metabolic pathways are relatively stable despite
variations in bacterial community structures
CD
Healthy
UC
1.0
0.5
0.0
Actinobacteria
Bacteroidetes
Cyanobacteria
Firmicutes
Fusobacteria
Planctomycetes
Proteobacteria
Tenericutes
1.0
0.5
C
0.0
Poorly Characterized
Unclassified Metabolism
Genetic Information Processing
Cellular Processes and Signaling
Xenobiotics Metabolism
Nucleotide Metabolism
Metabolism of Terpenoids and Polyketides
Metabolism of Other Amino Acids
Metabolism of Cofactors and Vitamins
Lipid Metabolism
Glycan Biosynthesis and Metabolism
Enzyme Families
Energy Metabolism
Carbohydrate Metabolism
Amino Acid Metabolism
Translation
Replication and Repair
Folding, Sorting and Degradation
Signal Transduction
Cell Motility
Membrane Transport
6
Microbial function is significantly altered in
inflamed tissue of UC patients
B
0.2
0.1
-0.2
0.2
-0.1
PC1
Normal
0.1
-0.1
PC2
A
-0.2
Inflamed
7
Reduction in carbohydrate and nucleotide metabolism and
increased lipid and amino acid metabolism with inflammation
0.035
0.030
0.12
0.11
0.045
0.040
0.035
0.030
0.025
0.10
No
rm
al
Inf
lam
ed
No
rm
al
0.025
0.13
**
No
rm
al
0.040
0.050
***
Inf
lam
ed
0.08
0.14
Lipid
Metabolism
Relative abundance
0.09
**
No
rm
al
0.10
Amino acid
Metabolism
Relative abundance
0.11
0.045
Relative abundance
**
0.12
Inf
lam
ed
Relative abundance
0.13
Nucleotide
Metabolism
Inf
lam
ed
Carbohydrate
Metabolism
8
Aqua
CD3
Foxp3
CD4
SSC-A
FSC-A
B
CD8
FSC-H
SSC-A
CD56
SSC-A
Characterization of CD4+ cells in LPMCs by Flow Cytometry
IL22
9
CD4+IL-22+ cells are associated with lipid, carbohydrate and
amino acid metabolism KEGG pathways in UC.
0.050
Relative abundance
15
10
5
0
N
I
0.13
0.045
0.040
0.035
0.030
R2=0.4067
P = 0.0105
0.025
I
%CD4+IL22+
*
Relative abundance
20
0
5
10
15
%CD4+IL22+
Carbohydrate
Metabolism
0.12
0.11
0.10
0.09
R2=0.2693
P = 0.0475
0.08
20
0
5
10
15
Amino acid
Metabolism
0.14
20
Relative abundance
Lipid
Metabolism
UC
0.13
0.12
0.11
R2=0.4010
P = 0.0041
0.10
0
5
10
15
20
%CD4+IL22+
Presentation Title Goes Here%CD4+IL22+
10
Foxp3+ cells are associated with lipid, carbohydrate and
amino acid metabolism KEGG pathways in CD.
10
0.045
0.040
0.035
0.030
R2=0.5163
0.025
P = 0.0008
0.020
N
I
0
0
10
20
30
0.12
0.11
0.10
0.09
0.08
40
%CD4+Foxp3+
50
Amino acid
Metabolism
0.15
Relative abundance
20
Carbohydrate
Metabolism
0.13
Relative abundance
30
Lipid
Metabolism
0.050
Relative abundance
40
I
%CD4+Foxp3+
50
CD
ns
R2=0.3332
P = 0.0121
0
10
20
30
40
%CD4+Foxp3+
50
0.14
0.13
0.12
0.11
R2=0.2901
P = 0.0211
0.10
0.09
0
10
20
30
40
50
%CD4+Foxp3+
11
Summary
•
Metabolic pathways are relatively stable despite variations in
bacterial community structures
•
Microbial function is significantly altered in inflamed tissue of UC
patients, but not in CD
•
Reduction in carbohydrate and nucleotide metabolism and
increased lipid and amino acid metabolism with inflammation
•
CD4+IL-22+ cells are associated with lipid, carbohydrate, and amino
acid metabolism KEGG pathways in UC patients
•
Foxp3+ cells are associated with lipid, carbohydrate and amino acid
metabolism KEGG pathways in CD
12
Conclusions
1. Metabolic pathways of the mucosal microbiota in CD do
not vary as much as UC with inflammation state,
indicating a more systemic perturbation of host-bacteria
interactions in CD compared to more localized
dysfunction in UC.
2. The alterations in metabolic pathways correlate
specifically with frequency of Tregs during CD, but with
TH22 cells during UC.
3. Alterations to metabolic pathways of the mucosal
microbiota may affect the production of metabolites that
can regulate intestinal CD4+ T cell populations and
inflammatory responses of the gut.
13
Acknowledgements:
Principal Investigator:
P’ng Loke
Labmates:
Jordan Poles
Jacqueline Leung
Martin Wolff
Collaborators:
Ilseung Cho (NYU)
Mike Poles (NYU)
Marty Blaser (NYU)
Lloyd Mayer (Mt. Sinai)
Thomas Ullman (Mt. Sinai)
Wasif Abidi (Mt. Sinai)
Funding:
Broad Foundation BMRP
NIH: NIAID, NCRR
NYU CTSI
AGA-Eli and Edythe Broad Student
Research Fellowship
Acknowledgements:
Knight Lab (QIIME)
Huttenhower Lab (PICRUSt and
LEFse)