INFLAMMATION AND HEART DISEASE:

1
INFLAMMATION AND HEART DISEASE Theres more to
heart disease than cholesterol and smoking
David J. Grainger, Ph.D. British Heart Foundation
Senior Research Fellow Department of Medicine,
Cambridge University

• Some statistics for the UK in 2002
• More than 100,000 deaths from coronary heart
  disease
• 40,000 premature deaths (age less than 75) - 20
  of the total
• More than twice as many deaths due to CHD than
  cancer

SourceBHF HeartStats www.heartstats.org

Cholesterol-lowering drugs and an aspirin-a-day
have helped, but this is still the number one
public health problem in the Western world today
2
ltpicture of dadgt
Ethelbert William Grainger 2nd March 1932 to 8th
April 1987 Age 55
3
Ethelbert William Grainger 2nd March 1932 to 8th
April 1987 Age 55
4
Ethelbert William Grainger 2nd March 1932 to 8th
April 1987 Age 55
5
Normal Coronary Artery
Diseased artery
Hard, fibrous cap Gk. Sclerosis hardening
Soft fatty deposits Gk. Athere gruel, porridge
6
Normal Coronary Artery
Diseased artery
Hard, fibrous cap Gk. Sclerosis hardening
Soft fatty deposits Gk. Athere gruel, porridge
ATHEROSCLEROSIS
7
Diseased artery

• HEART ATTACK
• The fibrous cap ruptures
• The fatty core comes into contact with the blood

8
Diseased artery

• HEART ATTACK
• The fibrous cap ruptures
• The fatty core comes into contact with the blood
• Within seconds a blood clot, or thrombus, forms
• The artery is totally occluded
• Oxygen delivery to the region of heart muscle
  served by the artery ceases
• If the vessel is not quickly re-opened, heart
  muscle, or myocardium, dies

9
Diseased artery

• HEART ATTACK
• The fibrous cap ruptures
• The fatty core comes into contact with the blood
• Within seconds a blood clot, or thrombus, forms
• The artery is totally occluded
• Oxygen delivery to the region of heart muscle
  served by the artery ceases
• If the vessel is not quickly re-opened, heart
  muscle, or myocardium, dies

IF TOO MUCH HEART MUSCLE DIES, THE ATTACK IS FATAL
10

THE ABC OF CORONARY HEART DISEASE
A
Fatty deposits, rich in cholesterol, form in the
arteries
B
Eventually, one of these plaques might rupture
C
A blood clot rapidly forms and blocks the blood
supply
A simplified three-step view of a very
complicated process!
11
So what causes heart disease, then ?
12
So what causes heart disease, then ?
HIGH RISK
LOW RISK
13
So what causes heart disease, then ?
Male gender
Smoking
High blood cholesterol
High blood pressure
Obesity
Fatty diet
Family history
Stress
14
So what causes heart disease, then ?
Male gender
Smoking
High blood cholesterol
High blood pressure
Obesity
Fatty diet
Family history
Stress
15
2004
16
Caleb Hillier Parry was performing an autopsy
when he found hardened coronary arteries and
glancing at the delapidated ceiling above
remarked that it appeared as if some plaster had
fallen down. He was the first doctor to realise
that changes in the coronary arteries caused
angina
1799
17
Joseph Hodgson describes the blockages in the
coronary arteries as an inflammatory reaction
1815
18
Carl Von Rokitansky champions the thrombotic
theory of heart disease, demonstrating the
presence of clotted blood in the atherosclerotic
plaque
1841
19
Johannes Vogel was the first to demonstrate the
presence of cholesterol and other fats in the
core of atherosclerotic plaques
1843
20
Sir Richard Quain expanded Vogels work and,
linking the fatty cholesterol-rich deposits in
the arteries to nutrition and circulating fats
1852
21
Rudolf Virchow, famous polish pathologist and
father of the cellular theory of human disease,
published a description of heart disease in which
he highlighted the INFLAMMATION, the deposition
of CHOLESTEROL and the importance of plaque
rupture and THROMBOSIS.
1856
22
INFLAMMATION
LIPID METABOLISM
THROMBOSIS
23
Starting in 1948, the Framingham Heart Study
remains the most extensive epidemiological study
ever conducted. By the early 1960s, it was clear
that elevated levels of cholesterol in the blood
were definitively associated with increased risk
of heart disease. Similar data for blood
clotting emerged from the same study
1950s
24
Joseph Goldstein and Michael Brown discovered
that a genetic defect in the gene encoding the
LDL-cholesterol receptor led to familial
hypercholesterolemia and a big increase in risk
of heart attack
1974
25
Jan Breslow and his colleagues systematically
deleted genes in mice which disturbed fat
metabolism and raised plasma cholesterol. In
almost every case, these deletions cause the
formation of fatty deposits in the arteries
1991
26
By the mid-1990s evidence from large, randomised
clinical trials proved beyond doubt that
treatment with cholesterol-lowering drugs called
statins could reduce the number of heart attacks
by as much as one third.
Low-dose aspirin (for two decades known to reduce
the risk of a second heart attack) was shown to
reduce the occurrence of a first heart attack
among men with angina by as much as one third.
1994
27
HIGH PLASMA CHOLESTEROL TREATED WITH DRUGS AND
DIET
EXCESSIVE BLOOD CLOTTING CONTROLLED WITH ASPIRIN
OR WARFARIN
1990s
28
Yet
40,000 premature deaths in 2002 were due to
coronary heart disease, which remains the number
one health problem in the Western world
HIGH PLASMA CHOLESTEROL TREATED WITH DRUGS AND
DIET
EXCESSIVE BLOOD CLOTTING CONTROLLED WITH ASPIRIN
OR WARFARIN
1990s
29
HIGH PLASMA CHOLESTEROL TREATED WITH DRUGS AND
DIET
UNCONTROLLED CHRONIC VASCULAR INFLAMMATION ?
EXCESSIVE BLOOD CLOTTING CONTROLLED WITH ASPIRIN
OR WARFARIN
1990s
30
Red blood cell
Numerous Oxygen transporters
White blood cell
Rarer Various different kinds Major
effectors of the immune system Also called
LEUKOCYTES
Picture of blood
x800
Picture of neutrophil
This leukocyte is engulfing and killing bacteria,
a process called PHAGOCYTOSIS. This is the major
mechanism that protects you from infection.
x11,750
Images taken from Functional Histology
Churchill Livingston 1979 with permission
31

THE IMMUNE SYSTEM ARMY
early

• Monocytes
• Surveillance
• Phagocytosis
• Neutrophils
• Release chemical weapons
• Phagocytosis
• Lymphocytes
• Produce antibody tags to mark out enemies
• Killer cells destroy marked enemies
• Generals, marshalling the other troops

minutes
hours
days
late
32

INFLAMMATION IS THE RESULT OF LEUKOCYTE
RECRUITMENT INTO TISSUE
This is acute inflammation of the lung, in
response to bacterial sepsis
33
Unfortunately, the leukocytes in an inflammatory
response can do almost as much damage to the host
as the invading pathogens

• Damage can include
• Oxidative damage
• Cellular death (necrosis)
• Formation of scar tissue (fibrosis)

34
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
35
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
36
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Framingham Study, 1960s
37
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Framingham Study, 1960s
Goldstein Brown, 1974 Breslow et al. 1991
38
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Framingham Study, 1960s
Goldstein Brown, 1974 Breslow et al. 1991
4S Trial, 1994
39
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Framingham Study, 1960s
Goldstein Brown, 1974 Breslow et al. 1991
4S Trial, 1994
40
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Hodgson, 1815
Framingham Study, 1960s
Goldstein Brown, 1974 Breslow et al. 1991
4S Trial, 1994
41
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Hodgson, 1815
Framingham Study, 1960s
Goldstein Brown, 1974 Breslow et al. 1991
4S Trial, 1994
42
Tissue damage
Monocytes
Myeloperoxidase
Macrophages
TNF-a
Neutrophils
Chemokines M-CSF
Chemokines
Interleukins
TGF-b
Immunoglobulins
Host tissue
Chemokines Interleukins TGF-b
CD4 T-cells
TNF-a
B-cells
LPS Antigen
Interleukins
Histamines
Chemokines
CD8 T-cells
Mast cells
Pathogens
Eosinophils
43
Tissue damage
Monocytes
Myeloperoxidase
Macrophages
TNF-a
Neutrophils
Chemokines M-CSF
Chemokines
Interleukins
TGF-b
Immunoglobulins
Host tissue
Chemokines Interleukins TGF-b
CD4 T-cells
TNF-a
B-cells
LPS Antigen
Interleukins
Histamines
Chemokines
CD8 T-cells
Mast cells
Pathogens
Eosinophils
44

• C-REACTIVE PROTEIN
• (CRP)
• Made in liver in response to IL-6
• Member of Pentraxin family, but its function
  still largely unknown
• Easily measured in blood
• Sensitive marker of inflammation
• Elevated during infections

LIVER
45

• C-REACTIVE PROTEIN
• (CRP)
• Made in liver in response to IL-6
• Member of Pentraxin family, but its function
  still largely unknown
• Easily measured in blood
• Sensitive marker of inflammation
• Elevated during infections

LIVER
46

PLASMA CRP PREDICTS FUTURE RISK OF MYOCARDIAL
INFARCTION
Dr. Paul M. Ridker

• Measure CRP using high sensitivity ELISA
• Determine levels in blood samples stored from
  healthy Framingham subjects
• Determine whether the risk of heart attack was
  higher in those with high CRP
• A pseudo-prospective nested case-control study
  design

47
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Hodgson, 1815
Framingham Study, 1960s
Ridker et al, 2000
Goldstein Brown, 1974 Breslow et al. 1991
4S Trial, 1994
48
Pro-inflammatory
Anti-inflammatory
Cytokines (TNFa, IL1, IL6) Chemokines
(MCP-1) Oxidised lipids Tissue damage Pathogens Hi
stamine Allergens etc
TGF-b IL-10
49

LOSS OF THE ANTI-INFLAMMATORY TGF-b SIGNAL CAUSES
VASCULAR INFLAMMATION
Genotype
Diet
Wild- type
Normal
tgfb1 /-
Normal

• ES technology to delete tgfb1 gene
• Homozygous deletion leads to rampant
  inflammation and death in days
• Heterozygous deletion reduces TGFb levels in
  adult arteries by approx 50
• Is there inflammation in the arteries?
• Do the animals develop atherosclerosis?

tgfb1 /-
Hi-Fat
ICAM-1
Control
Grainger et al. (2000). J. Cell Science
1132355-2361
50

LOSS OF THE ANTI-INFLAMMATORY TGF-b SIGNAL CAUSES
VASCULAR INFLAMMATION

AND ATHEROSCLEROSIS
Grainger et al. (2000). J. Cell Science
1132355-2361
51
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Hodgson, 1815
Framingham Study, 1960s
Ridker et al, 2000
Goldstein Brown, 1974 Breslow et al. 1991
Grainger et al., 2000
4S Trial, 1994
52
INFLAMMATION
LIPID METABOLISM
THROMBOSIS
53
INFLAMMATION
LIPID METABOLISM
Hypercholesterolemia
THROMBOSIS
54
INFLAMMATION
LIPID METABOLISM
Deep vein thrombosis
THROMBOSIS
55
INFLAMMATION
LIPID METABOLISM
THROMBOSIS
56
Analysis in apoE-/- mice
Treated for 24 weeks with TMX, aspirin, fish oil,
statins, EACA or warfarin
change in lipid lesions
change in lipid lesions
change in lipid lesions
change in LDL cholesterol
change in macrophage accumulation
change in fibrinogen deposition
LIPID METABOLISM
THROMBOSIS
INFLAMMATION
57
Analysis in apoE-/- mice
Treated for 24 weeks with TMX, aspirin, fish oil,
statins, EACA or warfarin
change in lipid lesions
change in lipid lesions
change in lipid lesions
change in LDL cholesterol
change in macrophage accumulation
change in fibrinogen deposition
LIPID METABOLISM
THROMBOSIS
INFLAMMATION
r 0.8
r 0.1
r 0.9
58
NEW SCIENTIST 11th January 2003 HEART
STOPPING! The conventional view of heart
disease is that there is something wrong with the
plumbing. During the 1980s researchers realised
that if there is too much cholesterol in the
bloodstream, it can build up inside the blood
vessel walls in the form of plaques. But since
the late 1980s it has become clear that this is
not the whole story inflammation also plays an
important role, according to Dr David Grainger,
an expert on inflammation from Cambridge
University.
59
NEW SCIENTIST 11th January 2003 HEART
STOPPING! The conventional view of heart
disease is that there is something wrong with the
plumbing. During the 1980s researchers realised
that if there is too much cholesterol in the
bloodstream, it can build up inside the blood
vessel walls in the form of plaques. But since
the late 1980s it has become clear that this is
not the whole story inflammation also plays an
important role, according to Dr David Grainger,
an expert on inflammation from Cambridge
University.
60
CHOLESTEROL
INFLAMMATION
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Is it present? Does an excess correlate with
disease incidence? Does an excess actually cause
the disease? Does treating the excess reduce
disease incidence?
Vogel, 1843
Hodgson, 1815
Framingham Study, 1960s
Ridker et al, 2000
Goldstein Brown, 1974 Breslow et al. 1991
Grainger et al., 2000
?
4S Trial, 1994
61
Pro-inflammatory
Anti-inflammatory
Cytokines (TNFa, IL1, IL6) Chemokines
(MCP-1) Oxidised lipids Tissue damage Pathogens Hi
stamine Allergens etc
TGF-b IL-10
62
Pro-inflammatory
Anti-inflammatory
Cytokines (TNFa, IL1, IL6) Chemokines
(MCP-1) Oxidised lipids Tissue damage Pathogens Hi
stamine Allergens etc
TGF-b IL-10
63

STRONG EVIDENCE FOR TGF-b AS A THERAPEUTIC TARGET

• Strong evidence for a constitutive, systemic
  anti-inflammatory role
• TGFb levels are perturbed in mouse models of
  atherosclerosis
• TGFb levels are perturbed among individuals with
  heart disease
• Polymorphisms of the tgfb1 gene may be associated
  with MI
• Heterozygous deletion of the tgfb1 gene causes
  vascular lipid lesion formation in mice

64
In vitro
Grainger et al. (1993) Biochem. J. 294109-112
In vivo

• Triphenylethylene class of drugs
• Breast cancer treatment and prevention
• Mechanism of action involves estrogen receptor
• Upregulate TGFb production by cancer cells

Grainger et al. (1995) Nature Med 11067-1076
65

TMX THERAPY PREVENTS ATHEROSCLEROSIS IN MICE,
RABBITS AND MONKEYS
MICE Grainger et al. (1995) Nature Med
11067 Lawn et al. (1996) J Biol Chem
27131367 Reckless et al. (1997) Circulation
951542
RABBIT Sugama et al. (2002) Jpn Heart J. 43545
MONKEY Williams et al. (1997) ATVB. 17403
66
(No Transcript)
67

TMX THERAPY REDUCES INCIDENCE OF MYOCARDIAL
INFARCTION IN WOMEN AT LOW RISK OF THE DISEASE
Meta-analysis of more than 13,000 women treated
with TMX Overall, risk of heart attack was cut by
almost 40
Braithwaite RS et al. (2003) J Gen Intern Med.
18937-47.
68

WILL TAMOXIFEN THERAPY BE A USEFUL WAY TO TREAT
HEART DISEASE SUFFERERS?
RUTH Raloxifene Use for the Heart

• Tamoxifen 20mg/day
• Men with unstable angina
• Multicentre study based at Papworth Hospital, UK
• Principal investigator Dr Peter Schofield
• Aim to begin in 2005

• Raloxifene 60mg/day
• Women at risk of MI
• Multicentre study based at Columbia University,
  NY
• Principal investigator Dr Elisabeth
  Barrett-Connor
• Began recruiting in 2002

69
ltpicture of dadgt
Jim Metcalfe Peter Weissberg Peter
Schofield David Mosedale Christine Witchell Jill
Reckless Sarah Hayns and all the rest of the
team
Ethelbert William Grainger 2nd March 1932 to 8th
April 1987 Age 55