1. The Role of Inflammation and Immune Responses
Robert L Wilensky, MD
University of Pennsylvania

2. I have no real or apparent conflicts of interest to report.
Robert L. Wilensky, MD
I have no real or apparent conflicts of interest to report.

3. Wilensky, Hamamdzic Curr Opin Cardiol 2007;22:545
Immunomodulatory cells and development of unstable coronary artery disease
Wilensky, Hamamdzic Curr Opin Cardiol 2007;22:545

4. Role of mast cells in plaque instability
Kovanen PT Immunol Rev 2007;217:105

5. Selective inhibition of Lp-PLA2, an important inflammatory mediator, reduces necrotic core area
DM/HC Control DM/HC Darapladib
Wilensky et al Nat Med 2008;14:1059

6. Upregulated with DM/HC (fold) Influence of Darapladib
Gene Name
Human
Entrez GeneID
Upregulated with DM/HC (fold)
Influence of Darapladib
% Change
P value
b-Arrestin 2
409
3.0
-47
0.004
HLA-DMA
3108
3.6
-38
0.007
PTAFR
5724
5.8
-49
0.009
BIN2
51411
7.1
-53
Lp-PLA2
7941
14.6
-55
0.012
CXCR3
2833
4.0
-68
0.015
GM2A
2760
5.4
-50
0.016
LAIR1
3903
4.8
-42
IL1RN
3557
21.1
-56
0.019
Cat. S
1520
16.0
0.020
gp91phox
1536
19.4
0.024
p47phox
653361
10.4
-54
0.025
CCR2
1231
1.9
-86
0.026
PLAUR
5329
6.7
-39
CD18
3689
14.3
0.029
UCP2
7351
6.1
0.030
HMOX1
3162
6.4
-44
0.033
DENND2D
79961
6.9
-45
0.034
CD68
968
16.7
-48
0.040
EVI2A
2123
3.7
0.041
EVI2B
2124
7.2
-41
0.043
CCL5
6352
0.046
SLC27A4
10999
3.2
0.047
NPL
80896
10.7
0.048
All coronary genes altered by Lp-PLA2 inhibition
Monocyte/T cell recruitment
Oxidative stress
Markers of unstable human atheroma

7. Hypothesis: Subsequent development of high-risk coronary artery lesions can be predicted using invasive coronary imaging.

8. Angiography/IVUS/NIRS Angiography/IVUS/NIRS Angiography/IVUS/NIRS
Experimental approach
Induction
DM/HC
n=21
Tissue
harvest
n=13
5 sudden deaths 1m 2m 3m 4m 5m 6m 7m 8m 9m
Angiography/IVUS/NIRS
Angiography/IVUS/NIRS
Angiography/IVUS/NIRS
Morphometrics
Histology (Movat’s)
Fibrous cap thickness (Picro-sirius Red)
Activated macrophages (Cathepsin S)
Cellular proliferation (Ki-67)
Apoptosis (TUNEL)
Serum glucose- 304 ± 49 mg/dl
Serum cholesterol- 511 ± 64 mg/dl

9. IVUS distribution of plaques: n(%)
IVUS results
Months
IVUS distribution of plaques: n(%)
No advanced disease
Pathologic intimal thickening
Fibroatheromas 3
388 (96.3%)
15 (3.7%) 6
214 (52.3%)
89 (21.8%)
106 (25.9%) 9
144 (31.5%)
177 (38.7%)
136 (29.8%)
P value
<0.001
0.019

10. NIRS positivity predicts lesion type by histology
NIRS pos (n= 66)
NIRS neg (n= 238)
P value
TCFA n (%)
29 (43.9)
34 (14.3)
0.0001
Fibroatheroma n(%)
38 (16.0)
PIT n (%)
2 (3.0)
64 (26.9)
Intimal hyperplasia n(%)
6 (9.2)
61 (25.6)
0.004
Non-atherosclerotic n(%)
0 (0)
41 (17.2)
Plaque area (mm2)
6.3 ± 4.9
2.7 ± 3.0
<
Plaque + Media area (mm2)
8.8 ± 5.3
4.6 ± 3.4
Necrotic core area (mm2)
4.5 ± 8.8
0.93 ± 2.1
0.002
% necrotic core area
40 ± 64
11 ± 19
0.0004
Fibrous cap thickness (μ)
145.9 ± 131.5
206.7 ± 199.9
0.04

11. NIRS positivity is associated with increased inflammation

12. NIRS positive
NIRS negative
macrophages
proliferation
apoptosis

13. Increased coronary artery lipid deposition (by NIRS) predicts increased necrotic core area at 9 months (by IVUS) 9
P< 0.01 8 7
3m Pos/6m Pos 6
P=0.38 5
3m Neg/6m Pos
P=0.002
P<0.0001
NC area (mm2) 4
P= 0.09
P=0.008
P<0.0001 3
3m Pos/6m Neg 2
3m Neg/6m Neg 1
3 month
6 month
9 month
NIRS POS at 3 months
NIRS NEG at 3 months

14. Correlation of NIRS and histology: A) Plaque area 2 4 6 8 10 12
Plaque area (mm2)
NEG
POS
3 mth
6 mth
9 mth
Correlation of NIRS and histology:
A) Plaque area
B) Necrotic core area 1 2 3 4 5 6 7 8
NC area (mm2)
NEG
POS
3 mth
6 mth
9 mth
NIRS at 3, 6 and 9 month

16. Antecedent NIRS positivity is associated with increased incidence of fibroatheromas at 9 months by histology
For both 3 and 6 month positive NIRS
p= 0.03 for TCFA
p= for fibroatheromas
For either 3 or 6 month positive NIRS
p= for TCFA
p= for fibroatheromas

17. Predictors of TCFA and fibroatheroma at 9-m
Thin cap fibroatheromas
Fibroatheromas
Months
Univariate
Multivariate
9-m
NIRS positive
P+M area
Necrotic core area
CSN >40%
Positive RI
(All p<0.001)
Lumen area
(p=0.01)
NIRS positive (p=0.000,
OR= 3.97, )
CSN>40% (p=0.046,
OR= 2.76, )
Positive RI (p= 0.002,
OR=3.81, )
(P=0.015)
OR= 13.37, )
CSN>40% (p=0.001,
OR= 5.14, )
6-m
NC area
(All p=0.001)
NIRS positive (p=0.143,
OR=1.71, )
NIRS positive (p=0.005,
(OR=2.71, )
3-m
None
NIRS positive (p=0.048)
NIRS positive (p=0.137,
OR=1.63, )

18. A
NIRS, IVUS, histology of culprit lesion in animal suffering sudden death, 132 days after DM/HC induction and 42 days after NIRS/IVUS evaluation C B

19. Summary
In DM/HC pigs inflammation is an important driving factor for the development of high-risk lesions. Inhibition of inflammation reduces necrotic core area.
In-vivo anatomic and biochemical evaluation predicted the presence of and progression of atherosclerotic lesions with a high-risk phenotype, by histology.
NIRS positive fibroatheromas had increased inflammation, cellular proliferation and apoptosis within a thinned fibrous cap; all characteristics of unstable lesions.
The persistence of a lipid core plaque within the arterial wall predicted subsequent progressive plaque development, larger necrotic core size and the presence of a TCFA.
In this model focal inflammation is dynamic as some lesions, despite persistent high- cholesterol levels did not maintain NIRS positivity or transformed from a NIRS negative to NIRS positive state during the study period.

20. Summary
In DM/HC pigs inflammation is an important driving factor for the development of high-risk lesions. Inhibition of inflammation reduces necrotic core area.
In-vivo anatomic and biochemical evaluation predicted the presence of and progression of atherosclerotic lesions with a high-risk phenotype, by histology.
NIRS positive fibroatheromas had increased inflammation, cellular proliferation and apoptosis within a thinned fibrous cap; all characteristics of unstable lesions.
The persistence of a lipid core plaque within the arterial wall predicted subsequent progressive plaque development, larger necrotic core size and the presence of a TCFA.
In this model focal inflammation is dynamic as some lesions, despite persistent high- cholesterol levels did not maintain NIRS positivity or transformed from a NIRS negative to NIRS positive state during the study period.

21. Summary
In DM/HC pigs inflammation is an important driving factor for the development of high-risk lesions. Inhibition of inflammation reduces necrotic core area.
In-vivo anatomic and biochemical evaluation predicted the presence of and progression of atherosclerotic lesions with a high-risk phenotype, by histology.
NIRS positive fibroatheromas had increased inflammation, cellular proliferation and apoptosis within a thinned fibrous cap; all characteristics of unstable lesions.
The persistence of a lipid core plaque within the arterial wall predicted subsequent progressive plaque development, larger necrotic core size and the presence of a TCFA.
In this model focal inflammation is dynamic as some lesions, despite persistent high- cholesterol levels did not maintain NIRS positivity or transformed from a NIRS negative to NIRS positive state during the study period.

22. Summary
In DM/HC pigs inflammation is an important driving factor for the development of high-risk lesions. Inhibition of inflammation reduces necrotic core area.
In-vivo anatomic and biochemical evaluation predicted the presence of and progression of atherosclerotic lesions with a high-risk phenotype, by histology.
NIRS positive fibroatheromas had increased inflammation, cellular proliferation and apoptosis within a thinned fibrous cap; all characteristics of unstable lesions.
The persistence of a lipid core plaque within the arterial wall predicted subsequent progressive plaque development, larger necrotic core size and the presence of a TCFA.
In this model focal inflammation is dynamic as some lesions, despite persistent high- cholesterol levels did not maintain NIRS positivity or transformed from a NIRS negative to NIRS positive state during the study period.

23. Summary
In DM/HC pigs inflammation is an important driving factor for the development of high-risk lesions. Inhibition of inflammation reduces necrotic core area.
In-vivo anatomic and biochemical evaluation predicted the presence of and progression of atherosclerotic lesions with a high-risk phenotype, by histology.
NIRS positive fibroatheromas had increased inflammation, cellular proliferation and apoptosis within a thinned fibrous cap; all characteristics of unstable lesions.
The persistence of a lipid core plaque within the arterial wall predicted subsequent progressive plaque development, larger necrotic core size and the presence of a TCFA.
In this model focal inflammation is dynamic as some lesions, despite persistent high- cholesterol levels did not maintain NIRS positivity or transformed from a NIRS negative to NIRS positive state during the study period.

25. Clinical results
21 pigs demonstrated consistently elevated glucose and cholesterol levels.
5 animals died unexpectedly due to ischemic sudden death
4 of 5 had multi-vessel disease with thrombus
1 of 5 had single TCFA but no thrombus
2 animals died of procedurally related complications- both with 3 vessel disease.
1 animal of non-cardiac causes
13 animals remained in the study

26. Excellent correlation of P + M areas by histology and IVUS
IVUS: Plaque + media area (mm2)
R= 0.87
P<0.001
Histology: Plaque + media area (mm2)

27. Progression of IVUS/NIRS over timeA 1 4 2 3
Progression of IVUS/NIRS over time B 1 4 2 3
A: 3 months
B: 6 months
C: 9 months 1 2 3 4 C 1 2 3 4 1 2 3 4

28. IVUS results (2) Months P+M area (mm2) NC area (mm2) EEL area (mm2)
Lumen
area (mm2)
Cross sectional narrowing (%) 3
1.94 ± 0.4
0 ± 0
9.35 ± 2.2
8.1 ± 1.5
21.3 ± 2.1 6
4.29 ± 1.9
1.50 ± 2.88
12.87 ± 6.5
8.8 ± 2.6
32.8 ± 11.8 9
8.19 ± 4.2
2.65 ± 4.88
16.76 ± 6.5
7.9 ± 1.9
40.9 ±15.5
P value
<0.001
0.423

29. NIRS results 303 (82.1) 30 (8.1) 19 (5.2) 17 (4.6) 311 (75.3)
Months
Block chemogram color threshold
Number (%) of sections positive for NIRS
Red
Dark orange
Light orange
Yellow 3
303 (82.1)
30 (8.1)
19 (5.2)
17 (4.6) 6
311 (75.3)
44 (10.6)
35 (8.5)
23 (5.6) 9
365 (79.3)
39 (8.5)
35 (7.6)
21 (4.6)
P value
0.405
0.841
0.680
0.887