Download Paneth cell phenotype is associated with clinical outcome

Paneth Cell Phenotype Correlates with
Genetics, Transcriptome Profile, Pathologic
Hallmark and Predicts Clinical Outcome in
Patients with Crohn's Disease
Ta-Chiang Liu, M.D., Ph.D.
Assistant Professor
Department of Pathology and Immunology
Washington University in St. Louis
Disclosures
Nothing to disclose
Goal: Identification of a cellular readout that
synthesizes genetic and environmental
factors and subclassifies Crohn’s Disease
Genetic
Susceptibility
Cellular readout
Disease
subclassification
Environment
Paneth Cells: Critical Roles in Innate Immunity
Antimicrobials
Lysozyme
α-Defensins
Risk Loci
Predicted to
Affect Paneth
Cell Function
ATG16L1
NOD2
XBP1
ITLN1
Secretion defect in
Atg16L1-deficient Paneth cells
WT
Atg16l1 deficient
Cadwell et al., Nature 2008.
NOD2 Variants and Disease Susceptibility
Allele carriage rate: <1%
Rare CD-associated
variants (6 known)
Relative risk for
development of CD:
NOD2 Heterozygous = 2-3
NOD2 Homozygous = ~20
Common CD-associated
variants
Allele carriage rate: <5%
Hypothesis
NOD2 risk variants may be associated with an abnormal granule
phenotype in the Paneth cells of CD patients
Materials and Methods
 De-identified ileocolectomy tissue from CD patients
 Inclusion criteria:
 Access to proximal margin of ileocolic resection
 Minimum of 100 well-oriented crypts
 No active or chronic inflammatory disease in the ileal
sections used for lysozyme stain
NOD2 Risk Variants are Associated with
Abnormal Paneth cell Phenotype
Scale bars: 10 µM
Immunofluorescence
Lysozyme
Hoescht
Gastroenterology, in press
diminished
NOD2 Risk Variants are Associated with
Abnormal Paneth cell Phenotype
*p<0.05 compared to 0 risk variants
Gastroenterology, in press
NOD2 and ATG16L1 Risk Alleles are Additive
for Abnormal Paneth Cell Phenotypes
Activated Immune Response Profile is
Associated with Abnormal Paneth Cells
Phenotype
GO term analysis performed using DAVID
Gastroenterology, in press
Paneth cell phenotype is associated with
clinical outcome
CD patients from WU and
CSMC (n=143) underwent
resection and received
post-op prophylactic
therapy
End point: endoscopic
evidence of disease
recurrence after surgery
Gastroenterology, in press
Paneth cell phenotype as
predictive cellular biomarker for
CD – practical issues
80
in in v o lv e d a r e a
% A b n o r m a l P a n e t h c e lls
Paneth cell phenotypes in involved areas
correlate with that of the uninvolved areas
60
40
20
0
0
10
20
30
40
50
% A b o r m a l P a n e t h c e lls
in u n in v o lv e d a r e a
P<0.0001
40
in s e c o n d r e s e c t io n
% A b n o r m a l P a n e t h c e lls
Paneth cell phenotype remains stable over
the years
30
20
10
0
0
10
20
30
40
% A b o r m a l P a n e t h c e lls
in f ir s t r e s e c t io n
50
(n=30)
P<0.0001
Paneth cell defects may be patchy: how many
crypts are needed to generate readout equivalent
to resection specimens?
“Virtual biopsy”
At least 40-50 crypts are needed to generate
equivalent results as with resection specimens.
50
40
in b io p s y
% A b n o r m a l P a n e t h c e lls
Paneth cell phenotypes in matched biopsy
and resection specimens are consistent
30
20
10
0
0
20
40
60
80
% A b o r m a l P a n e t h c e lls
in r e s e c t io n
100
(n=20)
P=0.0004
E
e
h
rg
d
e
e
s
d
d
e
d
d
e
re
fu
lu
if
is
la
c
n
x
D
in
rd
Adult (n=124)
E
im
o
40
D
is
l
0
a
20
T o t a l P a n e t h c e lls ( % )
40
rm
d
d
e
d
15%
o
e
e
s
e
d
l
80
N
rg
d
fu
lu
la
c
n
x
if
h
re
a
100
D
E
E
D
e
rm
is
rd
in
o
im
is
o
60
D
D
N
T o t a l P a n e t h c e lls ( % )
Bad Paneth cell phenotype is more prevalent
in pediatric CD
(Collaboration with Nita Salzman)
Pediatric (n=77)
100
80
60
47%
20
0
P< 0.0001
Significant increase in bad Paneth cell
phenotype over the past 30 years
1980-1982
(n=52)
1980-1982 (n=52)
2011-2013 (n=124)
T o t a l P a n e t h c e lls ( % )
100
80
3%
60
40
20
0
80
60
15%
40
20
n
E
c
x
la
lu
rg
d
e
e
d
d
e
s
fu
if
D
E
im
D
o
is
D
in
rd
is
e
h
re
e
d
d
l
a
rm
o
N
En
la
rg
ed
Ex
cl
ud
ed
iff
us
e
D
he
d
is
in
im
D
is
or
de
re
d
D
or
m
al
0
N
Total Paneth cells (%)
100
Paneth Cell Phenotype
P=0.0328
Summary
• Bad Paneth cell phenotype is associated with CDassociated risk alleles, distinct gene expression
profile, pathology hallmark, and clinical outcome.
• Paneth cell phenotype is spatially and temporally
stable.
• Paneth cell phenotype analysis can be performed
using routine biopsy material.
• Bad Paneth cell phenotype is commonly seen in
pediatric patients, and appears to be a new form
of disease that have emerged over the last 30
years.
Acknowledgement
Washington University
Cedars-Sinai
Kelli VanDussen
Robi D. Mitra
Rich Head
Rodney D. Newberry
Feng Gao
Thaddeus S. Stappenbeck
Dalin Li
Fadi Towfic
Nir Modiano
Rachel Winter
Talin Haritunains
Deepti Dhall
Stephan Targan
Dermot P. B. McGovern
Medical College of
Wisconsin
Harvard/MGH
Nita Salzman
Ramnik Xavier