Dr Brijesh Patel Clinical Research Fellow Wellcome Trust Clinical PhD Programme
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Imperial College Academic Trainees Annual Event The critical roles of TNF receptors in experimental acute lung injury Dr Brijesh Patel Clinical Research Fellow Wellcome Trust Clinical PhD Programme Anaesthetics, Pain Medicine, and Intensive Care
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Imperial College Academic Trainees Annual Event The critical roles of TNF receptors in experimental acute lung injury. Dr Brijesh Patel Clinical Research Fellow Wellcome Trust Clinical PhD Programme Anaesthetics, Pain Medicine, and Intensive Care. ALI/ARDS. ALI/ARDS is characterised by: - PowerPoint PPT Presentation
Transcript of Dr Brijesh Patel Clinical Research Fellow Wellcome Trust Clinical PhD Programme
Imperial College Academic Trainees Annual Event
The critical roles of TNF receptors in experimental acute lung injury
Dr Brijesh PatelClinical Research FellowWellcome Trust Clinical PhD ProgrammeAnaesthetics, Pain Medicine, and Intensive Care
ALI/ARDS
ALI/ARDS is characterised by:• acute onset• hypoxaemia• bilateral chest radiographic changes• normal left atrial pressures
Has a mortality of over 35% despite lung protective ventilation strategies.
Mulitiple aetiologies can lead to ALI/ARDS.• Direct – pneumonia; aspiration pneumonitis• Indirect – non-pulmonary sepsis; trauma; transfusion related; pancreatitis
Alveolar epithelial damage is a prerequisite to pulmonary oedema formation in all aetiologies of ALI/ARDS.
p55
TNFTNF
FADD
TRADD
Caspase 8/10
Apoptosis
Caspase 3
TRAF2
RIP
NEMO/IKK MKK3
p38
MAPKNFκB
cFLIP
MEKK1/7
JNK
AP1
Cell Survival
TNF receptor signalling
p75
Aims & Hypotheses
1. Do the two TNF receptors have distinct roles in experimental ALI/ARDS induced by acid aspiration?
2. What are the cellular mechanisms through which TNF receptor mediate their effects during experimental ALI/ARDS?
Experimental model of acid aspiration
Carotid LineTracheostomy
Intraperitoneal line for anaesthesia
Airway Pressure Transducer
Flow Transducer
Pneumotache
Flow Control
Inspiratory valveExpiratory valve
Setup for murine ventilation
Experimental Protocol
Baseline Respiratory Mechanics & Arterial Blood Gas
Acid Instillation
0 30 60 90 120 150 180
ABG
Mins
Respiratory Mechanics Measurements
GA + Setup for murine ventilation
Vt – 8mls/kgPEEP – 3cmH2O
p55KO shows improved respiratory physiology and p75KO show worse respiratory mechanics
Significant interaction P-value (P<0.0001) by 2-way ANOVA; *** P<0.001 vs WT Acid by 1-way ANOVA with Bonferroni test; Mean±SD; N=3-6
0
50
100
150
200
250
30 60 90 120 150 180
Elas
tanc
e C
hang
e (%
)
Time post instillation (minutes)Baseline
WT Saline
WT Acid
p55KO Acid
p75KO Acid
DKO Acid
0
100
200
300
400
500
PaO
2:FiO
2 R
atio
***
Acid +/- + + + +-
Mouse strain
WT
p55K
O
p75K
O
DK
O
WT
0
2
4
6
8
Wet
:Dry
R
atio
**
Acid +/- + + + +-
Mouse strain
WT
p55K
O
p75K
O
DK
O
WT
p55KO shows improved alveolar capillary barrier permeability
BA
LF P
rote
in
(mg/
ml)
0
1
2
3
*
Acid +/- + + + +-
Mouse strain
WT
p55K
O
p75K
O
DK
O
WT
• P<0.05; ** P<0.01 vs WT Acid by 1-way ANOVA with Bonferroni test; Mean±SD; N=3-6
Alveolar fluid clearance – an epithelial functional marker
Tight Junctions ENaC Na/K ATPase Aquaporins
Alveolus
Type 2 Alveolar Epithelial Cell
Type 1 Alveolar Epithelial Cell
Pulmonary capillary
Interstitium
Na+ H2O
Ex vivo measurement of alveolar fluid clearance
02468
101214161820
No
Dru
g C
ontr
ol
Am
ilorid
e
Isop
rote
reno
l
AFC
(%/3
0min
)
**
**
ΔVinstillate = AFC = ΔCAlb = ΔF = (F30-F0)/F30
** P<0.01 vs No Drug Control by 1-way ANOVA with Bonferroni test; Mean±SD; N=3-6
* P<0.05 vs WT Acid by 1-way ANOVA with Bonferroni test; Mean±SD; N=6-7
The p55 receptor mediates alveolar epithelial dysfunction in ALI
0
2
4
6
8
10
12
Alv
eola
r Flu
id C
lear
ance
(%
/30m
inut
es)
*
Acid +/-
Mouse strain
+
WT
+
p55K
O
-
WT
p75K
O
+
0
25
50
75
100
125
BA
LF R
AG
E (n
g/m
l)
Acid +/-
Mouse strain
+
WT
+
p55K
O
-
WT
p75K
O
+
*
Summary – Roles of TNF receptors in early ALI/ARDS
1. TNF receptors show opposing roles in pulmonary oedema formation in a model of direct ALI/ARDS induced by acid aspiration.
2. p75 TNF receptor deletion leads to worsening respiratory mechanics.
3. p55 TNF receptor deletion leads to reduced levels of alveolar epithelial injury as assessed by a functional marker i.e. AFC and a soluble marker i.e. RAGE.
4. This study provides a rationale for specific p55 TNF receptor blockade on the alveolar epithelium in ALI/ARDS states.
Acknowledgements
Professor Masao Takata – Magill Chair in AnaesthesiaDr Michael Wilson – Lecturer in Physiology
and the rest of the critical care group
0500
1000150020002500300035004000
BA
LF M
IP-2
(pg/
ml)
Acid +/- + + + +-Mouse strain W
T
p55K
O
p75K
O
DK
O
WT
0500
10001500200025003000350040004500
BA
LF K
C (p
g/m
l)
Acid +/- + + + +-Mouse strain W
T
p55K
O
p75K
O
DK
O
WT
p75 receptor attenuates alveolar inflammation
0500
100015002000250030003500400045005000
BA
LF IL
-6 (p
g/m
l)
Acid +/- + + + +-
Mouse strain WT
p55K
O
p75K
O
DK
O
WT
**
Future work
1. Early phase of ALI/ARDS:
• Investigate the influence of TNF p55 receptor death domain signalling in type 1 and type 2 AECs.
2. Extend the investigation into the longer term roles of TNF receptors in experimental ALI/ARDS:
• A resolving model of experimental acid aspiration.• The role of the p75 TNF receptor in promoting resolution and repair.
