U.S. patent application number 14/750500 was filed with the patent office on 2016-02-25 for use of an ang-(1-7) receptor agonist in acute lung injury.
The applicant listed for this patent is Charite-Universitatsmedizin Berlin. Invention is credited to Wolfgang Kuebler, Thomas Walther.
Application Number | 20160051621 14/750500 |
Document ID | / |
Family ID | 40430691 |
Filed Date | 2016-02-25 |
United States Patent
Application |
20160051621 |
Kind Code |
A1 |
Walther; Thomas ; et
al. |
February 25, 2016 |
USE OF AN ANG-(1-7) RECEPTOR AGONIST IN ACUTE LUNG INJURY
Abstract
The present invention refers to a peptidic or non-peptidic
angiotensin(1-7) (Ang-(1-7)) receptor agonist, preferably a Mas
receptor agonist, for the prevention and/or treatment of acute lung
injury, preferably acute respiratory distress syndrome.
Inventors: |
Walther; Thomas;
(Angermunde, DE) ; Kuebler; Wolfgang; (Berlin,
DE) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Charite-Universitatsmedizin Berlin |
Berlin |
|
DE |
|
|
Family ID: |
40430691 |
Appl. No.: |
14/750500 |
Filed: |
June 25, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13931168 |
Jun 28, 2013 |
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14750500 |
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13063685 |
May 5, 2011 |
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PCT/EP2009/006619 |
Sep 11, 2009 |
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13931168 |
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Current U.S.
Class: |
514/1.5 ;
514/9.7 |
Current CPC
Class: |
A61K 31/4178 20130101;
C07K 7/14 20130101; A61P 11/00 20180101; A61P 17/00 20180101; A61P
43/00 20180101; A61P 7/02 20180101; A61P 25/30 20180101; A61P 1/18
20180101; A61P 25/32 20180101; C07D 409/10 20130101; A61P 17/02
20180101; A61P 31/04 20180101; A61K 38/085 20130101 |
International
Class: |
A61K 38/08 20060101
A61K038/08 |
Foreign Application Data
Date |
Code |
Application Number |
Sep 12, 2008 |
EP |
08016142.5 |
Claims
1.-15. (canceled)
16. A method of treating acute lung injury in a subject comprising
administering to a subject who is suffering from or susceptible to
acute lung injury an Ang-(1-7) peptide comprising an amino acid
sequence according to one of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO:
3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, or SEQ ID NO: 7.
17. The method of claim 16, wherein the acute lung injury is an
acute respiratory distress syndrome.
18. The method of claim 16, wherein the acute lung injury is
related to a pulmonary or an extrapulmonary lung injury.
19. The method of claim 18, wherein the pulmonary lung injury is
selected from the group consisting of inhalation trauma, aspiration
trauma, toxic lung oedema, lung infection, preferably pneumonia,
lung contusion, and embolism.
20. The method of claim 18, wherein the extrapulmonary lung injury
is associated with a disorder selected from the group consisting of
sepsis, polytrauma, shock, burn, acute pancreatitis, drug
intoxication, alcohol abuse, chronic lung disease, mass
transfusion, disseminated intravascular coagulation, erythema, and
autoimmune lung disease.
21. The method of claim 16, wherein the subject is a mammal,
preferably a human, most preferably an adult human.
22. A method of treating acute lung injury in a subject comprising
administering to a subject who is suffering from or susceptible to
acute lung injury a pharmaceutical composition comprising an
Ang-(1-7) peptide comprising an amino acid sequence according to
one of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ
ID NO: 5, SEQ ID NO: 6, or SEQ ID NO: 7, and a pharmaceutically
acceptable excipient.
23. The method of claim 22, wherein the acute lung injury is an
acute respiratory distress syndrome.
24. The method of claim 22, wherein the acute lung injury is
related to a pulmonary or an extrapulmonary lung injury.
25. The method of claim 24, wherein the pulmonary lung injury is
selected from the group consisting of inhalation trauma, aspiration
trauma, toxic lung oedema, lung infection, preferably pneumonia,
lung contusion, and embolism.
26. The method of claim 24, wherein the extrapulmonary lung injury
is associated with a disorder selected from the group consisting of
sepsis, polytrauma, shock, burn, acute pancreatitis, drug
intoxication, alcohol abuse, chronic lung disease, mass
transfusion, disseminated intravascular coagulation, erythema, and
autoimmune lung disease.
27. The method of claim 22, wherein the subject is a mammal,
preferably a human, most preferably an adult human.
28. The method of claim 22, wherein the pharmaceutical composition
is formulated for oral, intramuscular, intravenous, subcutaneous,
topical, transdermal, rectal, vaginal, pulmonary, intranasal,
intrabuccal, or sublingual administration.
29. The method of claim 28, wherein, wherein the pharmaceutical
composition is formulated as a tablet, pill, capsule, granules, a
syrup, a spray, an aerosol, a liposomal composition, an ointment, a
suppository, an implant, a plaster, or a slow release
formulation.
30. The method of claim 28, wherein the pharmaceutical composition
further comprises one or more pharmacologically inert and
pharmacologically acceptable excipients such as a polymer carrier,
a distintegration agent, a lubricant, a solvent, or a swelling
agent.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This present invention is a continuation of U.S. patent
application Ser. No. 13/063,685, filed May 5, 2011, which is a
national phase entry of International Application No.
PCT/EP2009/006619, filed Sep. 11, 2009, and refers to a peptidic or
non-peptidic angiotensin-(1-7) (Ang-(1-7)) receptor agonist,
preferably a Mas receptor agonist, for the prevention and/or
treatment of acute lung injury, preferably acute respiratory
distress syndrome, the entire contents of both of which are
incorporated herein by reference.
[0002] The present invention refers to a peptidic or non-peptidic
angiotensin-(1-7) (Ang-(1-7)) receptor agonist, preferably a Mas
receptor agonist, for the prevention and/or treatment of acute lung
injury, preferably acute respiratory distress syndrome.
BACKGROUND OF THE INVENTION
[0003] With age-adjusted incidences of 86.2 per 100,000
person-years and overall mortality rates of .about.43%, acute lung
injury (ALI) and its most severe form, the acute respiratory
distress syndrome (ARDS), remain a major cause of death in
intensive care (1,2). The pathological hallmarks of the disease
comprise diffuse alveolo-capillary injury and an increased lung
permeability associated with a strong inflammatory response (3,4).
These changes underlie the clinical presentation which is
characterized by an acute onset, severe hypoxemia and a
proteinaceous lung oedema. Despite a multitude of large
multi-centric clinical trials to explore the potential of various
therapeutic strategies including the use of glucocorticoids,
ketoconazole, lisofylline, alprostadil, inhaled NO or supplemented
surfactant (5-7), no therapeutic pharmacological intervention could
so far improve the clinical outcome of ALI/ARDS. So far, the only
evident improvement for the survival of ARDS patients has been
achieved by the implementation of minimal invasive ventilation
strategies with low tidal volumes as compared to the previously
used high tidal volumes (8).
[0004] In a recent experimental study, Imai and co-workers could
demonstrate that angiotensin converting enzyme 2 (ACE2), which
converts Ang II to Ang-(1-7) by cleavage of one amino acid,
protects mice from severe acute lung injury induced by acid
aspiration or sepsis (9). The authors attributed this finding to
the fact that ACE2 will decrease Ang II concentration and thus,
reduce the activation of the Ang II type I receptor (AT1). This
notion has triggered a series of studies demonstrating the
effectiveness of AT1 receptor blockers or ACE inhibitors for the
treatment of various forms of experimental acute lung injury
(10-13).
[0005] Importantly however, the cleavage product of Ang II by ACE2,
Ang-(1-7), is not an inert waste product of the
angiotensin-pathway, but may exert active biological functions.
Ang-(1-7) binds to the G protein-coupled receptor Mas (14) which
appears to be a physiological antagonist of the AT1a receptor (15),
and potentially to other receptors. Binding of Ang-(1-7) to its
receptor(s) may thus contribute critically to the previously
demonstrated beneficial effects of interventions in the angiotensin
pathway on the pathology of ALI/ARDS.
[0006] The U.S. Pat. No. 6,235,766 refers to non-peptidic agonists
of Ang-(1-7) receptors, and particularly discloses
1-(p-thienylbenzyl)imidazoles having a marked action on Ang-(1-7)
receptors and mimicking the biological action of the effector
hormone Ang-(1-7).
[0007] The international patent application WO 2006/128266 refers
to the interaction between the Mas receptor and Ang-(1-7) or its
analogues in the context of controlling the functions of the
reproductive system.
[0008] The international patent application WO 2007/000036 refers
to the use of peptidic or non-peptidic Mas receptor agonists and
antagonists as apoptotic activity modulators.
[0009] The international patent application WO 2007/121546 refers
to the use of peptidic or non-peptidic Mas receptor agonists for
modulating metabolic activities related to the clinical
manifestation of the metabolic syndrome or its complications.
[0010] We speculated that Ang-(1-7) or related agonists may
represent a new and promising strategy for the treatment of
ALI/ARDS. Thus, the object of the present invention is to provide
means and methods for a pharmacological intervention in the
patho-physiologic events underlying ALI/ARDS.
SUMMARY OF THE INVENTION
[0011] The object of the present invention is solved by an
Ang-(1-7) receptor agonist for use in the prevention and/or
treatment of an acute lung injury in a subject.
[0012] The object of the present invention is solved by a method
for the prevention and/or treatment of an acute lung injury in a
subject using an Ang-(1-7) receptor agonist.
[0013] The object of the present invention is further solved by a
use of an Ang-(1-7) receptor agonist for the preparation of a
pharmaceutical composition for the prevention and/or treatment of
an acute lung injury in a subject.
[0014] The object of the present invention is further solved by a
method of prevention and/or treatment of an acute lung injury by
administering an Ang-(1-7) receptor agonist to a subject.
[0015] In one embodiment, the Ang-(1-7) receptor agonist is a Mas
receptor agonist. In one embodiment, the Ang-(1-7) receptor agonist
interacts with a Mas receptor or a receptor associated with a Mas
receptor.
[0016] In one embodiment, the Ang-(1-7) receptor agonist stimulates
a receptor that physically interacts with a Mas receptor.
[0017] In one embodiment, the Ang-(1-7) receptor agonist stimulates
a receptor that shares pharmacological similarities with a Mas
receptor.
[0018] In one embodiment, the Ang-(1-7) receptor agonist is an Ang
II metabolite sharing structural similarities with the Ang-(1-7)
peptide.
[0019] In one embodiment, the receptor agonist is a peptidic or
non-peptidic agonist. In one embodiment, the peptidic agonist is an
exogenous or endogenous Ang-(1-7) peptide comprising an amino acid
sequence
Asp.sup.1-Arg.sup.2-Val.sup.3-Tyr.sup.4-Ile.sup.5-His.sup.6-Pro.-
sup.7 according to SEQ ID NO: 1 or is a derivative or analogue
thereof.
[0020] In one embodiment, the peptidic agonist is a derivative or
analogue of the Ang-(1-7) peptide, the derivative or analogue
comprising an amino acid exchange, deletion or insertion.
Preferably, the derivative or analogue has conserved or better
agonistic properties.
[0021] In one embodiment, the peptidic agonist is a derivative or
analogue of the Ang-(1-7) peptide, the derivative or analogue
comprising an amino acid sequence
Asp.sup.1-Arg.sup.2-Val.sup.3-Ser.sup.4-Ile.sup.5-His.sup.6-Pro.sup.7
according to SEQ ID NO: 2,
Asp.sup.1-Arg.sup.2-Val.sup.3-Tyr.sup.4-Ile.sup.5-His.sup.6-Cys.sup.7
according to SEQ ID NO: 3 or
Asp.sup.1-Arg.sup.2-Val.sup.3-Ser.sup.4-Ile.sup.5-His.sup.6-Cys.sup.7
according to SEQ ID NO: 4.
[0022] In one embodiment, the peptidic agonist is a peptide
comprising an amino acid sequence according to SEQ ID NO: 2, SEQ ID
NO: 3 or SEQ ID NO: 4.
[0023] In one embodiment, the peptidic agonist is an exogenous or
endogenous NorLeu3-Ang-(1-7) peptide comprising an amino acid
sequence
Asp.sup.1-Arg.sup.2-NorLeu.sup.3-Tyr.sup.4-Ile.sup.5-His.sup.6-Pro.sup.7
according to SEQ ID NO: 5 or is a derivative or analogue
thereof.
[0024] In one embodiment, the peptidic agonist is an exogenous or
endogenous Ang IV peptide comprising an amino acid sequence
Val.sup.1-Tyr.sup.2-Ile.sup.3-His.sup.4-Pro.sup.5-Phe.sup.6
according to SEQ ID NO: 6 or is a derivative or analogue
thereof.
[0025] In one embodiment, the peptidic agonist is an exogenous or
endogenous Ang III peptide comprising an amino acid sequence
Arg.sup.1-Val.sup.2-Tyr.sup.3-Ile.sup.4-His.sup.5-Pro.sup.6-Phe.sup.7
according to SEQ ID NO: 7 or is a derivative or analogue
thereof.
[0026] In one embodiment, the non-peptidic agonist is selected from
the group of 1-(p-thienylbenzyl)imidazole compounds, and preferably
is Ave 0991 (i.e.
5-formyl-4-methoxy-2-phenyl-1[[4-[2-(ethylaminocarbonylsulfonamido)-5-iso-
butyl-3-thienyl]phenyl]methyl]-imidazole).
[0027] In one embodiment, the acute lung injury is an acute
respiratory distress syndrome.
[0028] In one embodiment, the acute lung injury is related to a
pulmonary (direct) or an extrapulmonary (indirect) lung injury.
[0029] In one embodiment, the pulmonary lung injury is selected
from the group consisting of inhalation trauma, aspiration trauma,
toxic lung oedema, lung infection, preferably pneumonia, lung
contusion, and embolism.
[0030] In one embodiment, the extrapulmonary lung damage is
associated with a disorder selected from the group consisting of
sepsis, systemic inflammatory response syndrome (SIRS), polytrauma,
shock, burn, acute pancreatitis, drug intoxication, alcohol abuse,
chronic lung disease, mass transfusion, disseminated intravascular
coagulation, erythema, and autoimmune lung disease.
[0031] In one embodiment, the subject is a mammal, preferably a
human, most preferably an adult human.
[0032] The object of the present invention is further solved by a
pharmaceutical composition comprising an Ang-(1-7) receptor agonist
for use in the prevention and/or treatment of an acute lung injury
in a subject.
[0033] The object of the present invention is further solved by a
method for the prevention and/or treatment of an acute lung injury
in a subject using a pharmaceutical composition comprising an
Ang-(1-7) receptor agonist.
[0034] In one embodiment of the pharmaceutical composition, the
Ang-(1-7) receptor agonist is a Mas receptor agonist.
[0035] In one embodiment of the pharmaceutical composition, the
receptor agonist is a peptidic or non-peptidic agonist.
[0036] In one embodiment of the pharmaceutical composition, the
peptidic agonist is an Ang-(1-7) peptide comprising an amino acid
sequence according to SEQ ID NO: 1 or is a derivative or analogue
thereof.
[0037] In one embodiment of the pharmaceutical composition, the
peptidic agonist is a peptide comprising an amino acid sequence
according to SEQ ID NO: 2, SEQ ID NO: 3 or SEQ ID NO: 4.
[0038] In one embodiment of the pharmaceutical composition, the
peptidic agonist is a peptide comprising an amino acid sequence
according to SEQ ID NO: 5 or is a derivative or analogue
thereof.
[0039] In one embodiment of the pharmaceutical composition, the
peptidic agonist is an Ang IV peptide comprising an amino acid
sequence according to SEQ ID NO: 6 or is a derivative or analogue
thereof.
[0040] In one embodiment of the pharmaceutical composition, the
peptidic agonist is an Ang III peptide comprising an amino acid
sequence according to SEQ ID NO: 7 or is a derivative or analogue
thereof.
[0041] In one embodiment of the pharmaceutical composition, the
non-peptidic agonist is selected from the group of
1-(p-thienylbenzyl)imidazole compounds, and preferably is Ave 0991
(i.e.
5-formyl-4-methoxy-2-phenyl-1-[[4-[2-(ethylaminocarbonylsulfonamido)-5-is-
obutyl-3-thienyl]phenyl]methyl]imidazole).
[0042] In one embodiment of the pharmaceutical composition, the
acute lung injury is an acute respiratory distress syndrome.
[0043] In one embodiment of the pharmaceutical composition, the
acute lung injury is related to a pulmonary (direct) or an
extrapulmonary (indirect) lung injury.
[0044] In one embodiment of the pharmaceutical composition, the
pulmonary lung injury is selected from the group consisting of
inhalation trauma, aspiration trauma, toxic lung oedema, lung
infection, preferably pneumonia, lung contusion, and embolism.
[0045] In one embodiment of the pharmaceutical composition, the
extrapulmonary lung damage is associated with a disorder selected
from the group consisting of sepsis, polytrauma, shock, burn, acute
pancreatitis, drug intoxication, alcohol abuse, chronic lung
disease, mass transfusion, disseminated intravascular coagulation,
erythema, and autoimmune lung disease.
[0046] In one embodiment of the pharmaceutical composition, the
subject is a mammal, preferably a human, most preferably an adult
human.
[0047] In one embodiment, the pharmaceutical composition is
formulated for a parenteral or enteral administration, preferably
for a parenteral administration by the route of inhalation,
infusion or injection. A pharmaceutical composition formulated for
an oral, an intramuscular, an intravenous, a subcutaneous, a
topical, a transdermal, a rectal, a vaginal, a pulmonary, an
intranasal, an intrabuccal, or a sublingual administration is also
considered.
[0048] In one embodiment, the pharmaceutical composition is
formulated as a tablet, a pill, a capsule, granules, a syrup, a
spray, an aerosol, a liposomal composition, an ointment, a
suppository, an implant, a plaster, or a slow release
formulation.
[0049] In one embodiment, the pharmaceutical composition further
comprises one or more pharmacologically inert and pharmaceutically
acceptable excipients such as a polymer carrier, a disintegration
agent, a lubricant, a solvent, or a swelling agent.
[0050] The term "receptor agonist" refers to an agent being capable
of activating a receptor, i.e. eliciting a receptor response. The
term "Ang-(1-7) receptor agonist" refers to an agent being capable
of activating a receptor that is also activated by Ang-(1-7). The
term "Mas receptor agonist" refers to an agent being capable of
activating the G protein-coupled Mas receptor. As an adequate
receptor response, a direct or indirect (due to a receptor
physically interacting with Mas) G protein-mediated signalling
cascade downstream from the receptor is initiated resulting in e.g.
arachidonic acid release, PGI.sub.2 formation, NO formation, and/or
cGMP generation.
[0051] Since Ang-(1-7) signalling is blocked by A779
([D-Ala.sup.7-Ang-(1-7);
Asp.sup.1-Arg.sup.2-Val.sup.3-Tyr.sup.4-Ile.sup.5-His.sup.6-D-Ala.sup.7,
SEQ ID NO: 8) and/or D-Pro.sup.7-Ang-(1-7)
(Asp.sup.1-Arg.sup.2-Val.sup.3-Tyr.sup.4-Ile.sup.5-His.sup.6-D-Pro.sup.7,
SEQ ID NO: 9), a further characteristic of an Ang-(1-7) agonist is
the inhibition of its effects by A779 and/or D-Pro.sup.7-Ang-(1-7).
Further non-competitive or competitive inhibitors are also
considered.
[0052] The term "peptidic agonist" refers to a compound comprising
one or more peptide bonds. The term encompasses compounds
consisting of a pure peptidic structure, i.e. a peptide composed of
two or more amino acids, as well as compounds comprising peptidic
and non-peptidic structures.
[0053] The term "non-peptidic agonist" refers to a compound not
comprising a peptide bond. Preferably, such a compound is of low
molecular weight, i.e. a small molecule. The term "derivative"
means a compound differing from another compound by a structural
modification, for example by replacement of one atom or a group of
atoms or a functional group with another atom or group of atoms or
functional group.
[0054] The term "analogue" means a compound which is similar in
structure or function to another compound.
[0055] The term "exogenous Ang-(I-7)" means Ang-(1-7) that is
produced outside of the subject's body to be treated and is
exogenously applied. This, however, does not exclude that Ang-(1-7)
is produced in e.g. a transgenic animal not to be treated.
Generally considered is Ang-(1-7) produced by biosynthesis or
conventional lab synthesis, e.g. solid-phase synthesis.
[0056] The term "endogenous Ang-(1-7)" means that Ang-(1-7) is
endogenously produced by the subject's body to be treated. An
(increased) endogenous production of Ang-(1-7) can be the result of
a stimulation of its generation from Ang II, e.g. by
pharmacologically activating ACE2, by blocking the AT1a receptor,
or by inhibiting the degradation of Ang II to Ang III by APA.
Similarly, endogenous Ang-(1-7) production can be increased by
stimulation of its generation from Ang I by NEP. An (increased)
endogenous production can also be the result of a gene therapeutic
intervention, e.g. by overexpressing ACE2 or a construct that
generates directly Ang-(1-7) or one of its precursors. An increased
concentration of endogenously produced Ang-(1-7) can also result
from a reduced degradation of Ang-(1-7) e.g. by pharmacological
inhibition of ACE which degrades Ang-(1-7) to Ang-(1-5).
[0057] Recombinant or overexpressed ACE2 will increase the
concentration of Ang-(1-7) directly by stimulating its conversion
from Ang II. ACE inhibitors will reduce Ang-(1-7) degradation to
Ang-(1-5) while AT1a receptor blockers will increase Ang-(1-7)
levels by elevation of ACE2 substrate availability.
[0058] Further considered are molecules, e.g. peptides or proteins,
comprising the Ang-(1-7) peptide sequence or chemical
structure.
[0059] "Acute lung injury" (ALI) and "acute respiratory distress
syndrome" (ARDS) are inflammatory disorders of the lung most
commonly caused by sepsis, pneumonia, trauma, and/or aspiration.
Inflammation can be locally restricted to the lung, or the
pulmonary inflammation can be part of a systemic inflammatory
process. ALI and ARDS are characterized by hypoxemia and diffuse
infiltrates on chest x-ray in the absence of elevated left atrial
pressure. ALI and ARDS differ only in the degree of hypoxemia in
that ALI is defined as a ratio of arterial oxygen partial pressure
over inspiratory oxygen fraction (PaO.sub.2/FiO.sub.2)<300 and
ARDS as a PaO.sub.2/FiO2<200 (16). Diagnosis is by clinical
presentation, ABGs (arterial blood gas analyses) and imaging
studies. Treatment is with lung-protective, low tidal volume
mechanical ventilation, supportive therapy, and treatment of
underlying causes.
DETAILED DESCRIPTION OF THE INVENTION
[0060] FIG. 1(A)-FIG. 1(D) shows the effects of Ang-(1-7) and the
non-peptidic Ang-(1-7) receptor agonist AVE0991 on lung MPO
activity (FIG. 1(A)), wet-to-dry weight ratio (FIG. 1(B)), mean
arterial pressure (FIG. 1(C)) and pulmonary vascular resistance
(FIG. 1(D)) in oleic acid induced acute lung injury. All data are
mean.+-.SEM from n=6 animals each; *p<0.05 vs. control;
#p<0.05 vs. OA.
EXAMPLE
[0061] Animals.
[0062] Experiments were performed in male Sprague-Dawley rats
(Charles River Wiga GmbH, Sulzfeld, Germany) with a body weight
(bw) of 330-360 g. Animals received care in accordance with the
Guide for the Care and Use of Laboratory Animals (Institute of
Laboratory Animal Resources, 7th edition 1996). The study was
approved by the local animal care and use committee.
[0063] Surgical Preparation and Hemodynamic Monitoring.
[0064] Rats were anesthetized by intraperitoneal injection of
medetomidine (0.5 mg/kg bw, Domitor.RTM., Dr. E. Graeub AG, Basel,
Switzerland), fentanyl (0.05 mg/kg bw, JanssenCilag, Neuss,
Germany) and midazolam (5 mg/kg bw, Dormicum.RTM., Roche, Basel,
Switzerland) as previously described (17). Following tracheotomy,
the trachea was cannulated and ventilation was established
(Advanced Animal Respirator, TSE Systems GmbH, Bad Homburg,
Germany) with a tidal volume of 6 ml/kg bw at 80 breaths/min.
Catheters (internal diameter 0.58 mm; Sims Portex Ltd., Hythe,
United Kingdom) were introduced into the left carotid artery and
the right internal jugular vein for monitoring of arterial blood
pressure (AP), fluid replacement and drug delivery as previously
described (18). An ultrasonic flowprobe (Transonic.RTM., Transonic
Systems Inc., Ithaca, N.Y.) was placed around the ascending aorta
distal to the branching of the coronary arteries for continuous
monitoring of cardiac output (CO). After median thoracotomy, a
catheter was introduced via the right ventricle into the pulmonary
artery for measurement of pulmonary artery pressure (PAP). AP, PAP
and CO were continuously recorded by the software package DasyLab
32 (DasyLab, Moenchengladbach, Germany). Pulmonary vascular
resistance (PVR) was calculated as arteriovenous pressure
differences over flow under the assumption of a constant left
atrial pressure of 2 mmHg.
[0065] Experimental Groups and Protocol.
[0066] Rats were randomly assigned to four groups of six animals
each: Animals in group 1 (control) did not receive any
pharmacological interventions. In group 2 (OA), ALI was induced by
intravenous infusion of 0.2 mg/kg oleic acid (Sigma, Munich,
Germany) over 30 min in the absence of any treatment. In group 3
(OA+Ang-(1-7)), ALI was induced as in group II, and infusion of
Ang-(1-7) at 5 pmol/kg per min was initiated immediately after ALI
induction. In group 4 (OA+AVE0991), ALI was induced as in group II,
and infusion of AVE0991 at 500 pmol/kg per min was initiated
immediately after ALI induction. In two additional groups of n=6
each, in which only myeloperoxidase (MPO) activity was measured,
infusion of the Ang-(1-7) receptor blocker A779 (10 pmol/kg per
min) was initiated either alone (group 5) or in combination with
Ang-(1-7) (5 pmol/kg per min; group 6) immediately after induction
of ALI.
[0067] After surgical preparation and hemodynamic stabilization
over at least 15 min, baseline hemodynamics were recorded and
arterial blood gases analyzed (RapidLab 348; Chiron Diagnostics
GmbH, Fernwald, Germany). Removed blood volume was replaced by
hydroxyethyl starch (6% hydroxyethyl starch 200/0,6; Fresenius, Bad
Homburg, Germany). Immediately after baseline recordings, 0.2 mg/kg
oleic acid was infused intravenously over 30 min in groups II-VI or
an equal volume of 0.9% NaCl in group I. In all groups,
measurements were repeated in 60 min intervals up to a total of 4 h
at which time animals were sacrificed by exsanguination. After in
situ ligation of the right main bronchus, lungs were excised and
processed for determination of wet-to-dry weight ratio and
myeloperoxidase (MPO) activity as described below.
[0068] Assessment of Lung Water and Inflammatory Response.
[0069] For determination of lung water content, wet-to-dry weight
ratio was measured by use of the microwave drying technique (18).
Recruitment of inflammatory cells was analyzed by measurements of
MPO activity in lung homogenates as previously described (19). In
brief, MPO activity was determined by a
3,3'-5,5'-tetramethylbenzidine (TMB)-based photometric assay,
compared to appropriate standard curves, and expressed as units per
gram lung tissue (U/g).
[0070] Statistical Analysis.
[0071] All data are presented as mean.+-.SEM. Data were tested by
Kruskal-Wallis test for differences between groups. Statistical
significance was assumed at P<0.05.
[0072] Results.
[0073] Oleic acid induced ALI as characterized by an increase in
MPO activity (FIG. 1(A)), lung wet-to-dry weight ratio (FIG. 1
(B)), and pulmonary vascular resistance (FIG. 1(D)), while systemic
arterial pressure was reduced after 4 h as compared to control
(FIG. 1(C)). Ang-(1-7) completely prevented the development of
OA-induced ALI, as demonstrated by the fact that Ang-(1-7) infusion
abrogated OA-induced changes in lung wet-to-dry weight ratio, MPO
activity, and pulmonary vascular resistance. The protective effect
of Ang-(1-7) was apparently not attributable to its described
vasodilatory effect in systemic blood vessels, since Ang-(1-7)
infusion attenuated systemic hypotension in OA-infused rats. The
non-peptidic Ang-(1-7) receptor agonist AVE0991 similarly
attenuated OA-induced ALI. Additional MPO analyses in groups 4 and
5 indicate that blockade of the Mas receptor by A779 aggravates
OA-induced ALI, and blocks the rescue effect of Ang-(1-7)
infusion.
CONCLUSIONS
[0074] The present findings show that infusion of Ang-(1-7) or a
non-peptidic Ang-(1-7) receptor agonist completely prevents lung
oedema and inflammation in an experimental model of oleic-acid
induced ALI. This protective effect is mediated via the G-protein
coupled receptor Mas, since it is lost after Mas blockade by A779.
Endogenously formed Ang-(1-7) serves as an intrinsic protection
mechanism against ALI, which is aggravated after Mas blockade by
A779.
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Sequence CWU 1
1
917PRTArtificial SequenceChemically synthesized peptide - Ang-(1-7)
1Asp Arg Val Tyr Ile His Pro 1 5 27PRTArtificial SequenceChemically
synthesized peptide - Ang-(1-7) derivative 2Asp Arg Val Ser Ile His
Pro 1 5 37PRTArtificial SequenceChemically synthesized peptide -
Ang-(1-7) derivative 3Asp Arg Val Tyr Ile His Cys 1 5
47PRTArtificial SequenceChemically synthesized peptide - Ang-(1-7)
derivative 4Asp Arg Val Ser Ile His Cys 1 5 57PRTArtificial
SequenceChemically synthesized peptide - NorLeu3-Ang-(1-7)peptide
5Asp Arg Leu Tyr Ile His Pro 1 5 66PRTArtificial SequenceChemically
synthesized peptide - Ang IV 6Val Tyr Ile His Pro Phe 1 5
77PRTArtificial SequenceChemically synthesized peptide - Ang III
7Arg Val Tyr Ile His Pro Phe 1 5 87PRTArtificial SequenceChemically
synthesized peptide - A779 8Asp Arg Val Tyr Ile His Ala 1 5
97PRTArtificial SequenceChemically synthesized peptide -
D-Pro7-Ang-(1-7) 9Asp Arg Val Tyr Ile His Pro 1 5
* * * * *