Conditioning the whole heart—not just the cardiomyocyte

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Review article

Conditioning the whole heartmdashnot just the cardiomyocyte

Robert M Bell Derek M Yellon

Hatter Cardiovascular Institute Institute of Cardiovascular Science University College London Hospital and Medical School 67 Chenies Mews London WC1E 6HX UK

a b s t r a c ta r t i c l e i n f o

Article history

Received 5 September 2011

Received in revised form 5 March 2012

Accepted 4 April 2012

Available online 11 April 2012

Keywords

Preconditioning

Remote conditioning

Postconditioning

Cardiac myocyte

Endothelium

Fibroblast

Conditioning the recruitment of endogenous cytoprotective pathways that protect the myocardium against in-

jurious ischaemiareperfusion injury has developed into a range of modalities that can be applied before (pre-

conditioning) during (perconditioning) or after the injurious ischaemic insult (postconditioning) either

directly to the heart or in a distal tissue (remote preconditioning) A wide range of triggers signaling pathwaysand potential end-effector mechanisms have been identi1047297edwhich appear commonto all forms of conditioning

Interestingly conditioningapplies to notonly thecardiac myocyte butto allthe constitutive cell types within the

myocardium As our understanding of conditioning mechanisms continue to develop and we start to realise

some of the dif 1047297culties in translating these phenomena to clinical treatments it may be time to take a more in-

tegrative approach to conditioning considering the many cellular and tissue types within the heart and how

they contribute to cytoprotective adaptations In this review we shall look at the conditioning phenomena

how different cell types contribute to the conditioned phenotype and where novel cardioprotective modalities

may be developed

copy 2012 Published by Elsevier Ltd

Contents

1 Introduction 2411 Ischaemiareperfusion injury 25

2 Cardiac conditioning 25

3 Composition of the myocardium targets for conditioning 25

31 Endothelium 27

311 Endothelium as part of conditioning signaling 27

312 Endothelium as a target for conditioning 28

32 Fibroblasts 28

321 Fibroblasts as a paracrine organ 28

322 Fibroblasts as a target for conditioning transformation to myo1047297broblasts 29

33 Connective tissue the extracellular matrix and matrix metalloproteinases 29

331 MMPs and receptor transactivation 29

332 MMPs and mobilisation of signaling molecules 29

333 MMPs as mediators of injury 29

34 Cardiac myocyte 30

4 Conclusions 30

Disclosure statement 30Acknowledgements 30

References 30

1 Introduction

Ischaemic heart disease is a signi1047297cant world-wide challenge for

healthcare services and will doubtless remain so for the foreseeable

future ischaemic heart disease in recent times has rapidly become the

leading cause of death globally with data from 2004 estimating 72

Journal of Molecular and Cellular Cardiology 53 (2012) 24ndash32

Corresponding author at The Hatter Cardiovascular Institute University College

London Hospital and Medical School 67 Chenies Mews London WC1E 6HX UK

Tel +44 20 3447 9888 fax +44 20 3447 9505

E-mail address hatter-instituteuclacuk (DM Yellon)

0022-2828$ ndash see front matter copy 2012 Published by Elsevier Ltd

doi101016jyjmcc201204001

Contents lists available at SciVerse ScienceDirect

Journal of Molecular and Cellular Cardiology

j o u r n a l h o m e p a g e w w w e l s e v i e r c o m l o c a t e y j m c c



million deathsper annumdirectly attributable to myocardial ischaemia

and is projected to increase still further [1] The gold standard manage-

ment of acute coronary syndromes remains expeditious revascularisa-

tion achieved with percutaneous coronary intervention (PCI) While

rapid restoration of blood 1047298ow to the ischaemic area of myocardium is

essential to stave off necrosis within the area at risk the question

remains as to whether the myocardial salvage achieved by timely

revascularisation can be augmented by manipulation of endogenous

cytoprotective mechanisms or exogenous inhibition of destructive pro-cesses activated by the combination of injurious ischaemia and reperfu-

sion In this review we shall undertake to discuss the modalities of

cardiac conditioning and how cardiac conditioning relates to the three

predominant cell populations (cardiomyocyte endothelium and 1047297bro-

blast) and the extracellular matrix of the heart

11 Ischaemiareperfusion injury

Non-re-vascularised occlusion of an epicardial coronary artery will

inevitably result in necrosis of the supplied myocardium (the area at

risk) If the occlusion is relieved through percutaneous intervention

the result of reestablishing 1047298ow down the re-opened coronary artery

is often rapidly appreciated with prompt brisk 1047298ow down the distal

vessel However this is not uniformly achieved 1047298ow may be sluggish

or even non-existant in 30ndash40 of cases presenting as a primary PCI

for acute ST-elevation myocardial infarction (the no-re1047298ow phenome-

non) [2] This may be the result of distal embolisation of the ruptured

atherosclerotic plaque and clot that had initially occluded the coronary

artery mdash or may in fact be an early angiographic manifestation of what

has been termed ldquoreperfusion injuryrdquo re1047298ecting marked endothelial in-

jury manifest by endothelial oedemaand discontinuity arising as a con-

sequence of ischaemia and reperfusion (review [34]) The no re1047298ow

phenomenon resulting from microvascular obstruction (MVO) is in it-

self associated with a worse clinical outcome [5] and thus amelioration

of microvascular dysfunction resulting from ischaemiareperfusion in-

jury may be a key target in the clinical management of reperfusion inju-

ryIn theabsence of MVO we know from basic science investigationsin

a wide variety of animal models that re-opening of the coronary artery

results in reperfusion injurya componentof myocardialdeath thatmaybe amenable to pharmacological manipulation or cardiac conditioning

up to 40ndash50 of the infarct observed following re-vascularisation is

thought to be potentially salvagable through conditioning modalities

[67]

2 Cardiac conditioning

Since the original description of the recruitment of innate cardiopro-

tection by a transient ischaemia and reperfusion prior to a lethal ischae-

mic insult by Murry Reimer and Jennings in 1986 [8] the basic

understanding of the modes of cellular injury (necrosis apoptosis

autophagy) and the signaling and mechanisms that increase resistance

to cell death has expanded dramatically Moreover these innate cardio-

protective mechanisms can not only be recruited by non-lethal ischae-miaof theheart but also throughpharmacological manipulation or even

remote-organ ischaemia prior (preconditioning) during (per-condi-

tioning) and immediately following (postconditioning) the ischaemic

insult The reader is encouraged to read one of the many excellent re-

cent reviews on this subject that cover the mechanisms by which con-

ditioning manifests protection and how these are currently being

translated into clinical practice [910]

At the mechanistic level recruitment of cardioprotection by condi-

tioning can be subdivided into triggers mediators and end-effectors

The mitochondrial permeability transition pore (mPTP) is currently the

strongest candidate for an end-effector of cardiac conditioning As

reviewed elsewhere modulation of the mPTP is thought to have a signif-

icant impact upon the myocardium in the reperfusion phase of injurious

ischaemiareperfusion injury [9] The up-stream signaling that impacts

upon the open probability of the mPTP mdash the mediators of conditioning

protection mdash have been subject of intense research and include variously

PI3K Akt Erkand Stat (reviewed [11ndash13]) (Fig 1) The activators of these

signaling cascades are termed ldquotriggersrdquo and are broadly extracellular

receptorligand interactions with autocoid endocrine or paracrine signal-

ing molecules A signi1047297cant number of receptors have been identi1047297ed

which when stimulated (so-called preconditioning-mimetics) may result

in recruitment of an ischaemiareperfusion resistant phenotype It has

been long recognised that the ischaemic conditioning signal is a summa-tion of multiple signals derived from multiple disparate receptorligand

interactions that once a suf 1047297cient combined signal is generatedbreeches

a lsquothresholdrsquo for triggering the conditioning response [14] What has

not been explored in great detail to date is how the various cellular

components within the myocardium contribute to this lsquoconditioning

souprsquo or how the various cellular populations interact to propagate the

conditioning signal Extracellular receptors offer considerable opportuni-

ty for inter and intracellular interaction and the study of these may

offer new insights into mechanisms underlying conditioning the heart

(Fig 2)

3 Composition of the myocardium targets for conditioning

Broadly the data available in the study of the conditioning phe-

nomenon have not endeavored to specify the cell type involved in

the protective pathway nor which is the cellular target for end-

effector protection since it is frequently assumed that conditioning

applies homogenously or that the speci1047297c target is the cardiac myo-

cyte There may be some justi1047297cation in this pragmatic view but it

is clearly a simpli1047297cation Indeed it is perfectly possible to trigger

protection in the isolated myocyte but in in-vitro whole tissue prep-

arations or in-vivo cardiac myocytes are unlikely to be conditioned in

isolation In studies of cardiac conditioning the archetypal primary

end point has been myocardial viability within the region at risk

Typically this is assessed by a number of methods that ultimately

leads to expression of infarct size as a ratiometric determination of

infarct to risk zone volumes By volumetric determination cardiac

myocytes are the predominant cell type estimated to contribute

around 75ndash80 of the total myocardium volume [1516] The nextmost prominent cell populations endothelium (3) and 1047297broblasts

(2) appear contribute relatively little by this measure [15] Macro-

phages (01) and pericytes (02) contribute a negligible amount

to the total volume and the remainder of the myocardial volume is

non-cellular comprising of connective tissue (6) and intracapillary

luminal volume (8ndash9) [15] However while cardiac myocytes are

the largest cell by volume within the myocardium they are not nec-

essarily the most numerate cell type depending on the methodology

the myocyte population contributes between 35 and 55 of total cell

number with variability between species age of the animal and re-

gion from where the myocardium is isolated [1718] The principal

non-myocyte cell populations consist of 1047297broblasts and endothelium

[16] but also include vascular smooth muscle macrophages and cir-

culating blood cells [1920] From Banerjees work using FACS analy-sis to count differing cell populations in adult murine myocardium

myocytes were found to contribute 56 of cell number followed by

1047297broblasts (27) endothelial cells (7) and vascular smooth muscle

cells (10) [18] Thus it can be argued that when ischaemic condition-

ing is triggered it is not an obligate phenomenon of cardiomyocytes

alone but rather a combination and interaction between the many

cell types found within the heart as illustrated in Fig 2 Interestingly

there are data available to support the hypothesis of cellular interac-

tion in response to a conditioning stimulus within the myocardium

Erythropoietin (EPO) is a well recognised preconditioning mimetic

[2122] with receptors found on a wide number of cell populations

within the myocardium including cardiomyocytes [23] Exposure to

EPO results in conditioning of the myocardium against lethal ischae-

mic insult Interestingly Lefers group developed a transgenic mouse

25RM Bell DM Yellon Journal of Molecular and Cellular Cardiology 53 (2012) 24ndash 32



RISK pathwayactivation

GPCR TKLR

1a

2

1b

3

Fig 1 Highlysimpli1047297ed schemaof conditioning demonstrating the threephases of conditioningadaptation 1mdashthe Trigger phaseusually characterised by receptorligandinteraction be

that derived from an autocrine source (1amdashfor example adenosine) or an exogenous or paracrine source (1bmdashfor example bradykinin) This is typically related to G-protein coupled

receptors (GPCR) or tyrosine kinase linked receptors (TKLR) and the two may interact as discussed in Section 31 Following receptor activation there is initiation of the mediator

phase that involves recruitment of a range of signaling kinases of the reperfusion injury salvage kinase pathway (2) and described in greater detail in Section 32 Finally the mediators

impact upon end-effector mechanisms (3) which include the mitochondrial permeability transition pore (mPTP) that increase resistance to ischaemiareperfusion mediated cell injury

CardiomyocyteEndothelial cell Fibroblast

Adenosine opiateH2S growth factors

ET-1 BradykininATII PGE2

NO CO H2SGrowth factors

AT II CT-1 ET-1NO CO

FGF2 EGF

Growth factors

Extracellular matrix

MMP

TGF-βMMP ndashProteaseactivatedreceptor

Achendorthins

PDGFGrowth factors

cytokins

Circulatoryfactors


Fig 2 A speculative cartoon of potential interactions between cell types and different compartments within the myocardium revealing some of the paracrine communication be-

tween cell types Following initiation of a conditioning stimulus there is increasing evidence of a complex interaction between the cell populations within the myocardium that may

signal increased resistance to injury and improve measures of cellular function in the reperfusion period following an injurious insult (Section 4)

26 RM Bell DM Yellon Journal of Molecular and Cellular Cardiology 53 (2012) 24ndash 32



model de1047297cient in EPO receptors in all but endothelial and haemato-

poietic cell lines [24] That hearts from these mice can still be condi-

tioned by exposure to EPO suggests that non-endothelial myocardial

cell lines areprotected indirectly through activationof theendothelium

potentially through the recruitment of endothelialnitric oxide synthase

[24]

In the following sections we have attempted to summarise the cur-

rent literature with respect to the predominant sources of many of the

endogenous autocrineparacrine signaling triggers that have a potentialrole to play in both conditioning and ischaemiareperfusion injury and

speculate where inter-cellular interaction may potentially contribute

to the evolution of cardioprotection in response to conditioning

31 Endothelium

The heart is a highly metabolically active organ with an extensive

microvascular blood supply Within the myocardium capillary length

density (the length of capillary per unit volume of myocardium) is esti-

mated at approximately 3300 mmmm3 [25] with a mean distance

between capillaries of approximately 14 μ m [26] Endothelium is there-

fore not only highly numerate within the myocardium but also ideally

placed to interact and modify cardiac myocyte function

In the context of myocardial conditioning the endothelium can be

postulated to have multiple roles a lsquoreceptorrsquo for blood-borne condi-

tioning moieties a sensor for hypoxic stress a paracrine organ involved

with recruitment of protectionin distal non-endothelial myocardialcell

populations and as a potential target for protection itself necessary to

preserve vital microvascular function

311 Endothelium as part of conditioning signaling

The endothelium could be postulated to be a lsquostepping stonersquo in the

pathway to cardioprotection following a conditioning stimulus Form-

ing the lining for the vasculature the endothelium is the 1047297rst point of

contact between the myocardium and humoral factors which may not

have direct access to non-vascular compartments of the heart due to a

competent ldquobloodndashheart barrierrdquo [27] Therefore for a vascular-borne

conditioning signal to be propagated beyond the endothelium and

into the myocardium the signal would necessarily have to be processedby and transmitted from the endothelium of the microvasculature That

such a process potentially exists in the context of conditioning has al-

ready been alluded to by Teng et al in their endothelial speci1047297c EPO re-

ceptor transgenic model [24] The hypothesis is further corroborated by

data demonstrating that eluent from conditioned isolated endothelial

cells is capable to imbuing increased resistance to ischaemic injury of

isolated naiumlve myocytes preservingfunction following injurious ischae-

mia [28]

3111 Endothelium as a paracrine organ ligand generation As a para-

crine organ the endothelium is capable of generating a wide range of

signals that have the potential to trigger protection in other cell types

and principally in cardiomyocytes Endothelin-1 (ET1) receptor ago-

nists were recognised early on as a pharmacological trigger of precon-ditioning [29] with evidence of ET1 receptors on the myocardial cell

surface [29] However while ET1 administered as an exogenous bolus

to trigger conditioning is effective [29] the endothelial release of ET1

during transient ischaemiareperfusion is probably insuf 1047297cient to trig-

ger ischaemic preconditioning on its own and ET1 receptor antagonists

insuf 1047297cient to attenuate ischaemic preconditioning [30]

Endothelial-derived kinins however have the potential play a far

more important role in the preconditioning trigger stimulus [31] Bra-

dykinin B(2) receptors are widely expressed throughout the myocar-

dium including both vascular smooth muscle and cardiac myocytes

[32] and the use of bradykinin B(2) receptor antagonists such as

HOE140 abrogate the conditioning response when used to bracket the

preconditioning stimulus [33] Moreover exogenously administered

bradykinin triggers both preconditioning [33] and postconditioning

[34] Interestingly the preconditioning stimulus does not appear to

signi1047297cantly increase the release of bradykinin into the coronary circu-

lation following the ischaemic preconditioning stimulus [3536]

perhaps because the released kinin is interstitial and does not leak

back into the coronary circulation unless the bloodndashheart barrier is

rendered incompetentmdashand certainly there is a signi1047297cant rise of

measured bradykinin in the coronary ef 1047298uent following the restoration

of 1047298ow after prolonged injurious ischaemia [35] While bradykinin

may not be measurably increased by ischaemic conditioning stimulikallidin-like peptide is released by ischaemic preconditioning protocols

increasing near six-fold and kallidin-like peptide induced protection is

abrogated by bradykinin B(2) receptor blockade [37] A problem with

identifyingkininsmdashor indeed anyother signalingmoleculemdashas a speci1047297c

endothelial paracrine agent is that it remains unclear as to the capacity

of other cell types to synthesise or release them into the intercellular

milieu It appears possible to protect isolated myocytes using an angio-

tensin converting enzyme inhibitor which results in accumulation of

bradykinin which raises the possibility that kinins may be synthesised

by cardiomyocytes forexampleand clearly more work is required to re-

solve this problem

Other endothelial products also have the potential to be involved

in the conditioning response including prostagladins such as PGE2

PGE2 levels are not only measurably augmented by ischaemic pre-

conditioning and the protection is abrogated by indomethacin but

exogenous administration of PGE2 is cardioprotective [38]

Endothelial cells are also a source of growth factors that impact

upon myocyte organisation and differentiation One recent example is

neuregulin a member of the epidermal growth factor family that plays

an important role in embryonic heart development [38] Neuregulin

hasbeen shown to be released from endothelial cells subjected to a hyp-

oxic stress to increase ischaemic tolerance of the co-cultured myocytes

[38] and moreover targeted deletion of the myocyte receptor (erbB)

increases cellular death in response to simulated ischaemiareperfusion

in a co-culture model [39]

3112 Endothelium as a paracrine organ gaseotransmitters Paracrine

signaling by endothelial cells appears not to be restricted to recep-

torligand interaction at least three gaseous signals have been associ-ated with endothelial interaction with cardiac myocytes in induction

of conditioning protection nitric oxide (NO) carbon monoxide (CO)

and hydrogen sulphide (H2S)

Nitric oxide has long been associated with ischaemic conditioning

Whereas inducible isoforms of nitric oxide may be found in myocytes

the endothelial isoform of nitric oxide synthase (eNOS) is mostly

although not exclusively distributed in the endothelium And yet

eNOS is essential for ischaemic [40] and pharmacological conditioning

[3441] as demonstrated in a seriesof experimentsusingeNOS knock-

out mice In the normal rat heart the capillary spacing between adja-

cent vessels is estimated being between 14 and 16 μ m and 20 μ m in

papillary [1526] and ventricular [42] muscle respectively Given that

nitric oxide may diffuse over 150ndash300 μ m within 4ndash15 seconds [43]

the proximity of capillary endothelium and the myocyte enablesendothelial-derived nitric oxide to effectively signal to subtended car-

diac myocytes and thus impact upon downstream myocyte signaling

and even directly upon myocyte end-effector mechanisms such as the

mPTP [44] Nitric oxide may do more than simply signal to adjacent

myocytes however NO may also prevent the adherence of platelets

and neutrophils thought to be play an important role in preservation

of microvascular function and prevention of microvascular obstruc-

tion (and is covered in more detail in Section 312) [4546]

CO may have a similar role to that of NO in the vasculature CO is

formed by the enzymatic degradation of heme by heme oxygenase

(HO) There are three known isoforms of heme oxidase HO-1 which

is inducible under cellular stress HO-2 the homeostatic form and

HO-3 which was originally discovered in rat brain [47] Induction of

HO-1 through pharmacological agents has been shown to signi1047297cantly




reduce myocardial infarct size in vivo [48] and cardiac-speci1047297c over-

expression reveals greater resistance to ischaemiareperfusion injury

[49] Moreover exogenous administration of CO (using CO releasing

molecule (CORM-3)) results in similar protection [50] but the down-

stream mechanisms of this protection are still to be determined (recent

comprehensive review [51])

Hydrogen sulphide the third of the gaseotransmitters has also been

demonstrated to be cardioprotective [52] eliciting a preconditioning-

like protection when administered 24 hours prior to injurious ischae-mia [53] Indeed H2S administration has been shown to trigger both

pre- and postconditioning pathways (review [54]) A number of poten-

tial mechanisms potentially underlie H2S mediated protection includ-

ing the involvement of RISK pathway components ERK and Akt [55]

and mediating protection through GSK-3β and mPTP [56] (reviewed

[57])

There is therefore a signi1047297cant body of evidence suggesting that

the endothelium plays an important role in induction of protection

as a paracrine organ cross-talking to other cell populations in the

heart but signi1047297cantly the endothelium may itself be a vital target

for conditioning protection

312 Endothelium as a target for conditioning

The signaling pathways and end-effector mechanisms are certain-

ly not speci1047297c to cardiomyocytes and are evident in endothelial cells

and other myocardial cell populations While there is some data in

isolated cell lines to suggest that the endothelium may be more sus-

ceptible to ischaemia yet more resistant to preconditioning [58] ad-

ministration of pharmacological preconditioning-mimetics such as

muscarinic acetylcholine receptor agonists not only attenuate infarct

size but also preserve endothelial function [59] Similar preservation

of endothelium-dependent vasodilation is seen following administra-

tion of adenosine receptor agonists (A1 and A3) [6061] bradykinin

(B(2) receptor mediated) [60] and angiotensin II [61] mdash a group of

pharmacological agents that clearly overlap with known triggers of

conditioning-mediated protection of the whole heart Targeting

these receptors to preserve the myocardium will impact not only

upon the myocyte but also to the endothelium mdash and in doing so

not only preserve cardiomyocyte viability but also conserve micro-vascular function

3121 No re- 1047298ow and microvascular obstruction Endothelium forms

the arteriolarcapillary conduits that channel and regulate the supply

of oxygenated blood to the myocardium playing a critical role under

both physiological conditions and the pathophysiological circum-

stances found following ischaemiareperfusion injury Endothelial

damage is thought to be an important component of the microvascu-

lar obstruction (MVO) characterised by the no-re1047298ow phenomenon

an often overlooked and perhaps sometimes forgotten form of reper-

fusion injury Endothelial injury is characterised by endothelial cellu-

lar oedema and failure of the bloodndashheart barrier [62] promoting not

only porosity of the microvasculature but also vasoconstriction and

activation of extrinsic coagulation cascades within the vessels soaf 1047298icted [63] Endothelial injury can therefore result in persistent lim-

itation of coronary microvascular 1047298ow resulting in enduring myocar-

dial ischaemia and inevitable cardiomyocyte necrosis a conditioning

regime selectively targeting the cardiomyocyte would be predicted to

be unsuccessful should the endothelial compartment be neglected

and the microvasculature fails

31211 Conditioning against No re- 1047298ow and MicrovascularObstruction

The no-re1047298ow phenomena in animal models tends to require pro-

longed injurious ischaemia the extent of no-re1047298ow dependent upon

the duration and severity of the injurious ischaemic insult [6264] and

is often associated with necrosis of the subtended myocardium [3]

Given the severity of the ischaemic insult required endothelial injury

is likely to be inevitable Preserving endothelial viability and function

might attenuate the severity of the MVO and the no-re1047298ow seen and

therefore one would postulate that conditioning the endothelium

would attenuate the severity of the no-re1047298ow phenomenon While

early studies failed to show a signi1047297cant reduction in no-re1047298ow follow-

ing preconditioning in dog [65] more recent studies have shown

ischaemic preconditioning [66] postconditioning [67] and remote con-

ditioning [68] to be effective in attenuating no-re1047298ow in various animal

models The hypothesis that conditioning may ameliorate endothelial

dysfunction is also supported by the clinical observation that pre-

infarct angina is associated with attenuation of no-re1047298

ow in the contextof primary percutaneous intervention (PPCI) in acute ST-elevation

myocardial infarction [64]

While comparatively little is known about the mechanisms of MVO

and no-re1047298ow in the setting of PPCI it is accepted to be a multifactorial

problem that includes not only distal embolisation fromthe atheroscle-

rotic plaque but also endothelial injury and dysfunction Interestingly

many of the agents that have been used clinically to attenuate the no-

re1047298ow phenomenon in the context of PPCI also impact upon condition-

ing signaling (adenosine and nitric oxide donors for example) [69] and

there are also data linking KATP channels and Rho kinases to the mech-

anism of protection against no-re1047298ow in the context of remote per-

conditioning [68]

The no-re1047298ow phenomenon and MVO may be of particular inter-

est in our attempts to translate conditioning through to a practical

therapy in the clinical arena Certainly in our recent study looking

at the clinical application of exogenous erythropoietin (EPO) a condi-

tioning mimetic with a robust basic science literature suggesting pro-

tection [70] and clinical data suggesting that lower endogenous levels

of EPO are associated with greater no-relow [71] we not only failed to

1047297nd evidence of infarct size limitation when administered as an ad-

junct to PPCI in the setting of acute ST-elevation MI but revealed an

alarming increase in MVO as suggested by gadolinium-enhanced car-

diac magnetic resonance imaging [72] A question therefore arises

might MVO be contributing to the failure of exogenous EPO to protect

the ischaemicreperfused myocardium The role of endothelial func-

tion and a greater understanding of the pathophysiology of MVO de-

serves further investigation in order to optimise outcomes following

reperfusion of ischaemically jeopardised myocardium

32 Fibroblasts

Cardiac 1047297broblastsplay an important role in cardiacphysiology pro-

viding the three-dimensional structure and milieu in which cardiac

myocytes and other myocardial cell types reside maintaining the

dynamic equilibrium of synthesis and degradation of the extracellular

matrix (ECM) and interstitium thatconsists of multiple collagens 1047297bro-

nectin proteoglycans and glycoproteins Unsurprisingly 1047297broblasts

play a vital role in embryonic development (review [73]) Commensu-

rate with their role in providing the myocardial scaffold they are also

one of the single largest population of cells within the heart [2073]

But 1047297broblasts possess much more than just an ECM sentinel function

1047297broblasts respond to chemical and mechanical changes [73] and ap-

pear to have the potential to interact electrically with the cardiac myo-cytes with many 1047297broblasts located in close proximity to inter-myocyte

gap junctions essential for conduction and propagation of action poten-

tials (reviews [7374])

321 Fibroblasts as a paracrine organ

Like endothelial cells 1047297broblasts possess an important paracrine

function with the potential to release conditioning-mimetic peptides

such as angiotensin II [75] cardiotrophin-1 [76] and ET1 [77] which

have the potential to induce a conditioned state in surrounding myo-

cardial cells Moreover like endothelium 1047297broblasts are also capable

of releasing various gaseotransmitters particularly NO [78] and CO

[79] although the role of 1047297broblast-derived gaseotransmitters in the

context of ischaemiareperfusion injury is unclear and perhaps the

role of 1047297broblast synthesised NO in particular is debatable Fibroblast




NO is derived from the inducible isoform of nitric oxide synthase

(iNOS) [78] and since the amount of NO generated is related to the

transcription of iNOS it is perhaps unlikely to play an immediate

role in early ischaemic preconditioning preconditioning or postcon-

ditioning although one may speculate a role for 1047297broblast iNOS in

the second window of protection following ischaemic precondition-

ing [80]

Impacting upon tyrosine kinase receptors with the potential to in-

teract witha wide range of myocardialcell types1047297

broblasts are respon-sible for the release of a range of growth factors that have the potential

to contribute to the conditioning stimulus including acid and basic

1047297broblast growth factors (aFGFFGF-1 [81] and bFGFFGF-2 [82]) and

tumour necrosis factor-α [83] It is not known however whether

conditioning of the myocardium results in suf 1047297cient release of these

growth factors to contribute to the conditioning signal although data

does exist to indicate that these growth factors when administered

exogenously can result in signi1047297cant attenuation of myocardial injury

However myocardial ischaemia does result in a measurable increase

in FGF-2 release into the coronary ef 1047298uent [84] over-expression of

FGF-2 receptors confers greater resistance to injurious ischaemic insults

[84] and exogenous administration of FGF-2 promotes pharmacological

postconditioning via the reperfusion injury salvage kinase (RISK) path-

way recruiting Akt and p70s6 kinase [82] Therefore FGF-2 has the

potential to contribute to the triggering of cardioprotective signaling

(review [85])

That 1047297broblasts do have a paracrine function in the induction of

cardioprotection is strongly suggested by data demonstrating that

the media of conditioned 1047297broblasts applied to naiumlve myocytes in-

creases their resistance against injurious ischaemia [86] Similarly

the media of conditioned 1047297broblasts also attenuates post-ischaemic

myocardial dysfunction when perfused through isolated heart on the

Langendorff rig [86] Moreover not only does FGF-2 impact upon

myocytes but are also implicated in preserving endothelial function

in an autocrine fashion Matsunaga et al demonstrate that over-

expressing FGF-2 receptor on the endothelium attenuates infarct

size and myocyte apoptosis compared to wild type mice [87]

To suppose that1047297broblasts in conditioningfunction solelyproviding

a paracrine signal to conditioning induction is an over-simpli1047297cationlike endothelium 1047297broblasts too can be a target for conditioning-like

protection and their recruitment having the potential to impact upon

myocardial function and viability

322 Fibroblasts as a target for conditioning transformation

to myo 1047297broblasts

The pathophysiology of ischaemiareperfusion injury appears to

signi1047297cantly alter 1047297broblast function and the constituent make up of

the ECM The consequence of ischaemiareperfusion is cytokine re-

lease (for example IL-1β) in1047298ammasome activation and in1047298amma-

tory cell recruitment [88] Moreover there is a shift from 1047297broblast

to myo1047297broblast phenotype with the myo1047297broblast implicated in tis-

sue repair granuloma formation and 1047297brosis [89] The importance of

this phenotype shift in acute ischaemiareperfusion injury is currentlyunclear and warrants further investigation but conditioning appears

to preserve 1047297broblastmyo1047297broblast gap junction functionality [90]

and the myo1047297broblasts can themselves be preconditioned against ne-

crotic cell death following lethal ischaemic injury by the signaling

mechanisms already well characterised in myocardial conditioning

[91] Given that cardiac conditioning attenuates cardiac arrhythmia

[9293] and that conditioning appears to preserve gap junction con-

nectivity it is tempting to hypothesise that preservation and recruit-

ment of 1047297broblasts myo1047297broblasts may be an essential component to

the anti-arrhythmic properties of cardioprotection through preserva-

tion of gap junction function but to date there is surprisingly little

evidence to support such a link Indeed at the present time there is

little data available to ascertain the impact of conditioning upon

1047297broblast myo1047297broblast function ischaemiareperfusion injury and

given the importance of this cell type to cardiac remodeling this is

an area that requires further study

33 Connective tissue the extracellular matrix and

matrix metalloproteinases

As indicated in the previous section 1047297broblasts are essential to the

synthesis and maintenance of the extracellular matrix A key protease

involved in the cycling of the collagens and 1047297

bronectin found withinthe ECM are the matrix metalloproteinases (MMPs) and their regula-

tors tissue inhibitors of metalloproteinases (TIMPs) an area that has

been the subject of considerable research and is reviewed elsewhere

[9495] ECM turnover in the context of early myocardial necrosis and

scar formation is an important part of the evolution of the infarct and

its repair with data demonstrating that MMP-2 increases by 25 dur-

ing injurious ischaemia and remains elevated in reperfusion (40

over baseline) for days afterwards [96] and interestingly elevated

MMP activity appears correlated to myocardial dysfunction following

acute ST elevation myocardial infarction in patients [97] Therefore

the impact of conditioning upon MMP activity and the ECM is of par-

ticular interest

331 MMPs and receptor transactivation

Intriguingly MMPs may play an important role in up-stream sig-

naling of the conditioning response with recent data to suggest that

MMPs are essential to the induction of preconditioning and postcon-

ditioning pathways MMPs appear to play an essential role in epider-

mal growth factor receptor (EGFR) transactivation a pivotal signaling

step in both adenosine triggered preconditioning [98] and bradykinin

postconditioning [99]

332 MMPs and mobilisation of signaling molecules

MMPs are associated with the mobilisation of TGF-β from the ECM

[100] which in turn may trigger a conditioning response (review

[101]) and in so doing so reveals another characteristic of MMPsmdashthe

ability to mobilise potential extracellular ligands at the time of ischae-

mia and reperfusion that can contribute to the conditioning trigger re-

quired for endogenous cardioprotection Interestingly there are alsofamilies of G-protein coupled protease-activated receptors on the endo-

thelium that not only are activated by plasma-borne proteases such as

tissue plasminogen activator but also by MMPs which has been pro-

posed as one potential mechanism in heart failure [102] but so far little

work has been done to ascertain whether there is a role for these recep-

tors in conditioning-mediated cardioprotection

333 MMPs as mediators of injury

MMPs are increasingly associated with adverse aspects of ischae-

miareperfusion injury MMP inhibitors have been found to attenuate

myocardial dysfunction following injurious ischaemia [103] and this

has been associated with intracellular cardiomyocyte targets of degra-

dation including myocin light chain-1 [104] alpha actinin [105] and

titin [106] Moreover preconditioning has been shown to attenuateMMP activity [107108] and MMP inhibitors have been shown to

attenuate myocardial necrosis for example minocycline a tetracycline

antibiotic with MMP inhibitoryaction administered to bracket theinju-

rious ischaemic insult 48 hours before and after was found to be pro-

tective signi1047297cantly attenuating infarct size by 33 [109]

Given that MMPs appear important for EGFR transactivation in

conditioning triggering these 1047297ndings are perhaps surprising but

clearly MMPs have a multifaceted role in both conditioning signaling

and the mediation of the pathophysiology of cellular injury following

injurious myocardial ischaemia Therefore the question is whether

MMPs may represent a new target for cardioprotection a RISK-

independent IschaemiaReperfusion Injury Salvage (IRIS) paradigm

which is separate from traditional conditioning that warrants further

investigation




34 Cardiac myocyte

In studies of myocardial viability the cardiomyocyte is perceived

as the cell that only receives the conditioning signalmdashbut this may

not be entirely the case While it is certainly the case that the myocyte

expresses a wide range of receptors including the majority associated

with ischaemic preconditioning (for example the lsquoclassicalrsquo G-protein

coupled receptors adenosine A1 A2A A3 bradykinin and δ-opioid)

can be identi1047297

ed on the surface of cardiomyocytes [110] the cardio-myocyte also has autocrine and therefore the potential for paracrine

signaling to adjacent cells The isolated cardiomyocyte model is ideal

for identifying such signaling candidates and while our knowledge

can probably be regarded as being incomplete a surprising number

of autocoids have thus been identi1047297ed These include adenosine

[111] opiates [112] the gaseotransmitter H2S [113] and growth fac-

tors including transforming growth factor-beta superfamily member

growth-differentiation factor-15 (GDF-15) [114] stromal cell derived

factor-1 alpha (SDF-1α) [115] and adiponectin (APN) [116] Each has

been demonstrated to attenuate cardiomyocyte injury but not all are

necessarily linked to paracrine effects upon epithelial or 1047297broblasts

although it is attractive to postulate a link between them NO synthe-

sis is an important component of conditioning signaling within the

myocyte [344041] but like endothelium eNOS derived NO may

not necessarily be restricted to intramyocyte signaling iNOS derived

NO is thought to be synthesised primarily in the cardiomyocyte con-

tributing to the protection observed following preconditioning (with

comparatively little generated by other populations within the myo-

cardium [117]) with generation of nitric oxide and its ability to per-

meate throughout the myocardium NO may have the potential to

impact upon other myocardial cell populations although such a para-

crine link has yet to be de1047297nitively proven

4 Conclusions

The expansion of our knowledge of cardiac conditioning since pre-

conditionings original description a quarter of a century ago has been

dramatic but for all this expansion our understanding of the mecha-

nisms of cell death and cardioprotection remain incomplete A widerange of autocoid and paracrine preconditioning-mimetics have been

identi1047297ed withcell surfacereceptors identi1047297ed on many of themyocar-

dial cellular populations which would clearly suggest the potential for

intercellular signaling Elucidating the interaction between the many

cell types and differing tissue compartments within the myocardium

may yield further clues to further extend our understanding of the con-

ditioning phenomena and their successful export to clinical application

Disclosure statement

None

Acknowledgements

We thank the British Heart Foundation (RG0801526411) for

their continued support Dr RM Bell is an Academic Clinical Lecturer

supported by the National Institute of Health Research (NIHR) This

work was undertaken at UCLHUCL who received a proportion of

funding from the Department of Healths NIHR Biomedical Research

Centres funding scheme

References

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[2] Butler MJ Chan W Taylor AJ Dart AM Duffy SJ Management of the no-re1047298owphenomenon Pharmacol Ther Oct 2011132(1)72ndash85

[3] Kloner RA No-re1047298ow phenomenon maintaining vascular integrity J CardiovascPharmacol Ther Sep 201116(3ndash4)244ndash50

[4] Schwartz BG Kloner RA Coronary no re1047298ow J Mol Cell Cardiol 201252873ndash82[5] Rezkalla SH Dharmashankar KC Abdalrahman IB Kloner RA No-re1047298ow phe-

nomenon following percutaneous coronary intervention for acute myocardial in-farction incidence outcome and effect of pharmacologic therapy J IntervCardiol Oct 201123(5)429ndash36

[6] Hausenloy DJ Lecour S Yellon DM Reperfusion injury salvage kinase and survivoractivatingfactor enhancement prosurvival signaling pathways in ischemic postcon-ditioning two sides of the same coin Antioxid Redox Signal Mar 1 201114(5)893ndash907

[7] Yellon DM Hausenloy DJ Myocardial reperfusion injury N Engl J Med Sep 132007357(11)1121ndash35

[8] Murry CE Jennings RB Reimer KA Preconditioning with ischemia a delay of le-thal cell injury in ischemic myocardium Circulation Nov 198674(5)1124ndash36[9] Shi W Vinten-Johansen J Endogenous Cardioprotection by Ischaemic Postcondi-

tioning and Remote Conditioning Cardiovasc Res 201294206ndash16[10] Hausenloy DJ Yellon DM The therapeutic potential of ischemic conditioning an

update Nat Rev Nov 20118(11)619ndash29[11] Yang X Cohen MV Downey JM Mechanism of cardioprotection by early ische-

mic preconditioning Cardiovasc Drugs Ther Jun 201024(3)225ndash34[12] Cohen MV Downey JM Ischemic postconditioning from receptor to end-

effector Antioxid Redox Signal Mar 1 201114(5)821ndash31[13] Boengler K Hil1047297ker-Kleiner D Drexler H Heusch G Schulz R The myocardial

JAKSTAT pathway from protection to failure Pharmacol Ther Nov 2008120(2)172ndash85

[14] Goto M Liu Y Yang XM Ardell JL Cohen MV Downey JM Role of bradykinin inprotection of ischemic preconditioning in rabbit hearts Circ Res Sep 199577(3)611ndash21

[15] Anversa P Olivetti G Melissari M Loud AV Stereological measurement of cellu-lar and subcellular hypertrophy and hyperplasia in the papillary muscle of adultrat J Mol Cell Cardiol Aug 198012(8)781ndash95

[16] Vliegen HW van der Laarse A Cornelisse CJ Eulderink F Myocardial changes inpressure overload-induced left ventricular hypertrophy A study on tissue com-position polyploidization and multinucleation Eur Heart J 199112(4)488ndash94

[17] Nag AC Zak R Dissociation of adult mammalian heart into single cell suspen-sion an ultrastructural study J Anat Oct 1979129(Pt 3)541ndash59

[18] Banerjee I Fuseler JW Price RL Borg TK Baudino TA Determination of cell typesand numbers during cardiac development in the neonatal and adult rat andmouse Am J Physiol Sep 2007293(3)H1883ndash91

[19] Brutsaert DL Cardiac endothelial-myocardial signaling its role in cardiac growthcontractile performance and rhythmicity Physiol Rev Jan 200383(1)59ndash115

[20] Nag AC Study of non-muscle cells of the adult mammalian heart a 1047297ne structur-al analysis and distribution Cytobios 198028(109)41ndash61

[21] Cai Z Manalo DJ Wei G Rodriguez ER Fox-Talbot K Lu H et al Hearts from ro-dents exposed to intermittent hypoxia or erythropoietin are protected againstischemiandashreperfusion injury Circulation Jul 8 2003108(1)79ndash85

[22] Bullard AJ Govewalla P Yellon DM Erythropoietin protects the myocardiumagainst reperfusion injury in vitro and in vivo Basic Res Cardiol Sep 2005100(5)397ndash403

[23] Parsa CJ Matsumoto A Kim J Riel RU Pascal LS Walton GB et al A novel protec-tive effect of erythropoietin in the infarcted heart J Clin Invest Oct 2003112(7)999ndash1007

[24] Teng R Calvert JW Sibmooh N Piknova B Suzuki N Sun J et al Acute erythro-poietin cardioprotection is mediated by endothelial response Basic Res CardiolMay 2011106(3)343ndash54

[25] Amann K Wiest G Zimmer G Gretz N Ritz E Mall G Reduced capillary densityin the myocardium of uremic ratsmdasha stereological study Kidney Int Nov199242(5)1079ndash85

[26] Henquell L Odoroff CL Honig CR Intercapillary distance and capillary reserve inhypertrophied rat hearts beating in situ Circ Res Sep 197741(3)400ndash8

[27] Brutsaert DL De Keulenaer GW Fransen P Mohan P Kaluza GL Andries LJ et alThe cardiac endothelium functional morphology development and physiologyProg Cardiovasc Dis Nov-Dec 199639(3)239ndash62

[28] SJ O Zellner JL Cox MH Hebbar L Brothers TE Mukherjee R et al Contributorymechanisms for the bene1047297cial effects of myocyte preconditioning during cardio-plegic arrest Circulation Nov 1 199694(9 Suppl)II389ndash97

[29] Wang P Gallagher KP Downey JM Cohen MV Pretreatment with endothelin-1mimics ischemic preconditioning against infarction in isolated rabbit heart

J Mol Cell Cardiol Mar 199628(3)579ndash

88[30] Erikson JM Velasco CE Endothelin-1 and myocardial preconditioning Am Heart

J Jul 1996132(1 Pt 1)84ndash90[31] Wirth KJLinz W WiemerG ScholkensBA Kininsand cardioprotection Pharmacol

Res Jun 199735(6)527ndash30[32] Minshall RD Nakamura F Becker RP Rabito SF Characterization of bradykinin

B2 receptors in adult myocardium and neonatal rat cardiomyocytes Circ ResMay 199576(5)773ndash80

[33] Wall TM Sheehy R Hartman JC Role of bradykinin in myocardial precondition-ing J Pharmacol Exp Ther Aug 1994270(2)681ndash9

[34] Bell RM Yellon DM Bradykinin limits infarction when administered as an adjunct to reperfusion in mouse heart the role of PI3K Akt and eNOS J Mol CellCardiol Feb 200335(2)185ndash93

[35] Parratt JR Vegh A Zeitlin IJ Ahmad M Oldroyd K Kaszala K et al Bradykinin andendothelial-cardiacmyocyte interactions in ischemic preconditioning Am J CardiolAug 4 199780(3A)124Andash31A

[36] Liu X Lukasova M Zubakova R Lewicka S Hilgenfeldt U Kallidin-like peptide me-diates the cardioprotective effect of the ACE inhibitor captopril against ischaemicreperfusion injury of rat heart Br J Pharmacol Jul 2006148(6)825ndash32




[37] Liu X Lukasova M Zubakova R Lewicka S Hilgenfeldt U A kallidin-like peptideis a protective cardiac kinin released by ischaemic preconditioning of rat heartBr J Pharmacol Dec 2005146(7)952ndash7

[38] Bouchard JF Chouinard J Lamontagne D Participation of prostaglandin E2 in theendothelial protective effect of ischaemic preconditioning in isolated rat heartCardiovasc Res Jan 14 200045(2)418ndash27

[39] Hedhli N Huang Q Kalinowski A Palmeri M Hu X Russell RR et al Endotheli-um-derived neuregulin protects the heart against ischemic injury CirculationMay 24 2011123(20)2254ndash62

[40] Bell RM Yellon DM The contribution of endothelial nitric oxide synthase to earlyischaemic preconditioning the lowering of the preconditioning threshold An

investigation in eNOS knockout mice Cardiovasc Res Nov 200152(2)274ndash

80[41] Bell RM Yellon DM Atorvastatin administered at the onset of reperfusion andindependent of lipid lowering protects the myocardium by up-regulating apro-survival pathway J Am Coll Cardiol Feb 5 200341(3)508ndash15

[42] Turek Z Grandtner M Kubat K Ringnalda BE Kreuzer F Arterial blood gases mus-cle1047297berdiameterand intercapillary distance in cardiachypertrophyof rats with anold myocardial infarction P1047298ugers Arch Sep 29 1978376(3)209ndash15

[43] Lancaster Jr JR Diffusion of free nitric oxide Methods Enzymol 199626831ndash50[44] Ohtani H Katoh H Tanaka T Saotome M Urushida T Satoh H et al Effects of

nitric oxide on mitochondrial permeability transition pore and thiol-mediatedresponses in cardiac myocytes Nitric Oxide Dec 30 201126(2)95ndash101

[45] Hansen PR In1047298ammatory alterations in the myocardial microcirculation J MolCell Cardiol Dec 199830(12)2555ndash9

[46] Seal JB Gewertz BL Vascular dysfunction in ischemiandashreperfusion injury AnnVasc Surg Jul 200519(4)572ndash84

[47] Kikuchi G Yoshida T Noguchi M Heme oxygenase and hemedegradation BiochemBiophys Res Commun Dec 9 2005338(1)558ndash67

[48] Hangaishi M Ishizaka N Aizawa T Kurihara Y Taguchi J Nagai R et al Inductionof heme oxygenase-1 can act protectively against cardiac ischemiareperfusion

in vivo Biochem Biophys Res Commun Dec 20 2000279(2)582ndash8[49] YetSF Tian R LayneMD Wang ZYMaemura K SolovyevaM et alCardiac-speci1047297c

expression of heme oxygenase-1 protects against ischemia and reperfusion injuryin transgenic mice Circ Res Jul 20 200189(2)168ndash73

[50] Wang G Hamid T Keith RJ Zhou G Partridge CR Xiang X et al Cardioprotectiveand antiapoptotic effects of heme oxygenase-1 in the failing heart Circulation2010121(17)1912ndash25

[51] Peers C Steele DS Carbon monoxide a vital signalling molecule and potent toxinin the myocardium J Mol Cell Cardiol 201252(2)359ndash65

[52] Elrod JW Calvert JW Morrison J Doeller JE Kraus DW Tao L et al Hydrogensul1047297de attenuates myocardial ischemiandashreperfusion injury by preservation of mitochondrial function Proc Natl Acad Sci U S A Sep 25 2007104(39)15560ndash5

[53] Calvert JW Jha S Gundewar S Elrod JW Ramachandran A Pattillo CB et al Hy-drogen sul1047297de mediates cardioprotection through Nrf2 signaling Circ Res Aug14 2009105(4)365ndash74

[54] Predmore BL Lefer DJ Hydrogen sul1047297de-mediated myocardial pre- and post-conditioning Expert Rev Clin Pharmacol 20114(1)83ndash96

[55] Hu Y Chen X Pan TT Neo KL Lee SW Khin ES et al Cardioprotection induced byhydrogen sul1047297de preconditioning involves activation of ERK and PI3KAkt path-ways P1047298ugers Arch Jan 2008455(4)607ndash16

[56] Yao LL Huang XW Wang YG Cao YX Zhang CC Zhu YC Hydrogen sul 1047297de pro-tects cardiomyocytes from hypoxiareoxygenation-induced apoptosis by pre-venting GSK-3beta-dependent opening of mPTP Am J Physiol May2010298(5)H1310ndash9

[57] Nicholson CK Calvert JW Hydrogen sul1047297de and ischemiandashreperfusion injuryPharmacol Res Oct 201062(4)289ndash97

[58] Shirai T Rao V Weisel RD Ikonomidis JS Li RK Tumiati LC et al Preconditioninghuman cardiomyocytes and endothelial cells J Thorac Cardiovasc Surg Jan1998115(1)210ndash9

[59] Richard V Blanc T Kaeffer N Tron C Thuillez C Myocardial and coronary endo-thelial protective effects of acetylcholine after myocardial ischaemia and reper-fusion in rats role of nitric oxide Br J Pharmacol Aug 1995115(8)1532ndash8

[60] Giannella E Mochmann HC Levi R Ischemic preconditioning prevents the im-pairment of hypoxic coronary vasodilatation caused by ischemiareperfusionrole of adenosine A1A3 and bradykinin B2 receptor activation Circ Res Sep199781(3)415ndash22

[61] Kuzner J Drevensek G Gersak B Budihna M Hypoxic and pharmacological pre-

conditioning preserves vasomotor response of porcine coronary artery Pol JPharmacol Nov-Dec 200456(6)789ndash97

[62] Kloner RA Ganote CE Jennings RB The ldquono-re1047298owrdquo phenomenon after tempo-rary coronary occlusion in the dog J Clin Invest Dec 197454(6)1496 ndash508

[63] Kleinbongard P Konorza T Bose D Baars T Haude M Erbel R et al Lessons fromhuman coronary aspirate J Mol Cell Cardiol 201252(4)890ndash6

[64] Iwakura K Ito H Kawano S Shintani Y Yamamoto K Kato A et al Predictive fac-tors for development of the no-re1047298ow phenomenon in patients with reperfusedanterior wall acute myocardial infarction J Am Coll Cardiol Aug 200138(2)472ndash7

[65] Bauer B Simkhovich BZ Kloner RA Przyklenk K Does preconditioning protectthecoronaryvasculaturefrom subsequentischemiareperfusion injury CirculationAug 199388(2)659ndash72

[66] Jerome SN Akimitsu T Gute DC Korthuis RJ Ischemic preconditioning attenu-ates capillary no-re1047298ow induced by prolonged ischemia and reperfusion Am JPhysiol May 1995268(5 Pt 2)H2063ndash7

[67] Zhao JL Yang YJ You SJ Cui CJ Gao RL Different effects of postconditioning onmyocardial no-re1047298ow in the normal and hypercholesterolemic mini-swinesMicrovasc Res Mar 200773(2)137ndash42

[68] Zhao JL Yang YJ Pei WD Sun YH You SJ Gao RL Remote periconditioning re-duces myocardial no-re1047298ow by the activation of K ATP channel via inhibitionof Rho-kinase Int J Cardiol Apr 3 2009133(2)179ndash84

[69] Jaffe R Dick A Strauss BH Prevention and treatment of microvascularobstruction-related myocardial injury and coronary no-re1047298ow following percu-taneous coronary intervention a systematic approach Jacc Jul 20113(7)695ndash704

[70] Riksen NP Hausenloy DJ Yellon DM Erythropoietin ready for prime-time cardi-oprotection Trends Pharmacol Sci May 200829(5)258ndash67

[71] Niccoli G Andreotti F Marzo F Cecchetti S Santucci E DAmario D et al Endoge-nous serum erythropoietin and no-re1047298ow in patients with ST-elevation myocardial

infarction Eur J Clin Invest 2011411210ndash

9[72] Ludman AJ Yellon DM Hasleton J Ariti C Babu GG Boston-Grif 1047297ths E et al Ef-fect of erthropoietin as an adjunct to primary percutaneous coronary interven-tion a randomised controlled clinical trial Heart 2011971560ndash5

[73] Souders CA Bowers SL Baudino TA Cardiac 1047297broblast the renaissance cell CircRes Dec 4 2009105(12)1164ndash76

[74] Camelliti P Green CR Kohl P Structural and functional coupling of cardiac myo-cytes and 1047297broblasts Adv Cardiol 200642132ndash49

[75] Dostal DE Rothblum KN Chernin MI Cooper GR Baker KM Intracardiac detec-tion of angiotensinogen and renin a localized reninndashangiotensin system in neo-natal rat heart Am J Physiol Oct 1992263(4 Pt 1)C838ndash50

[76] Kuwahara K Saito Y Harada M Ishikawa M Ogawa E Miyamoto Y et al Involve-ment of cardiotrophin-1 in cardiac myocytendashnonmyocyte interactions duringhypertrophy of rat cardiac myocytes in vitro Circulation Sep 7 1999100(10)1116ndash24

[77] Tomoda Y Kikumoto K Isumi Y Katafuchi T Tanaka A Kangawa K et al Cardiac1047297broblasts are major production and target cells of adrenomedullin in the heartin vitro Cardiovasc Res Mar 200149(4)721ndash30

[78] Farivar RS Chobanian AV Brecher P Salicylate or aspirin inhibits the induction

of the inducible nitric oxide synthase in rat cardiac 1047297broblasts Circ Res May199678(5)759ndash68

[79] Lakkisto P Palojoki E Backlund T Saraste A Tikkanen I Voipio-Pulkki LM et alExpression of heme oxygenase-1 in response to myocardial infarction in rats

J Mol Cell Cardiol Oct 200234(10)1357ndash65[80] Bolli R Cardioprotective function of inducible nitric oxide synthase and role of

nitric oxide in myocardial ischemia and preconditioning an overview of a de-cade of research J Mol Cell Cardiol Nov 200133(11)1897ndash918

[81] Htun P Ito WD Hoefer IE Schaper J Schaper W Intramyocardial infusion of FGF-1 mimics ischemic preconditioning in pig myocardium J Mol Cell Cardiol Apr199830(4)867ndash77

[82] Jiang ZSWen GB Tang ZH Srisakuldee W Fandrich RR Kardami E Highmolecularweight FGF-2 promotes postconditioning-like cardioprotection linked to activationof protein kinase C isoforms as well as Akt and p70 S6 kinases [corrected] Can JPhysiol Pharmacol Oct 200987(10)798ndash804

[83] Martire A Fernandez B Buehler A Strohm C Schaper J Zimmermann R et alCardiac overexpression of monocyte chemoattractant protein-1 in transgenicmicemimics ischemic preconditioningthroughSAPKJNK12 activationCardiovascRes Feb 200357(2)523ndash34

[84] House SL Bolte C Zhou M Doetschman T Klevitsky R Newman G et al Cardiac-speci1047297c overexpression of 1047297broblast growth factor-2 protects against myocardialdysfunction and infarction in a murine model of low-1047298ow ischemia CirculationDec 23 2003108(25)3140ndash8

[85] Kardami E Detillieux K Ma X Jiang Z Santiago JJ Jimenez SK et al Fibroblastgrowth factor-2 and cardioprotection Heart Fail Rev Dec 200712(3ndash4)267ndash77

[86] Nakazato K Naganuma W Ogawa K Yaoita H Mizuno S Nakamura T et al At-tenuation of ischemic myocardial injury and dysfunction by cardiac 1047297broblast-derived factor(s) Fukushima J Med Sci Jun 201056(1)1ndash16

[87] Matsunaga S Okigaki M Takeda M Matsui A Honsho S Katsume A et alEndothelium-targeted overexpression of constitutively active FGF receptor in-duces cardioprotection in mice myocardial infarction J Mol Cell Cardiol May200946(5)663ndash73

[88] Kawaguchi M Takahashi M Hata T Kashima Y Usui F Morimoto H et al In1047298am-masome activation of cardiac 1047297broblasts is essential for myocardial ischemia reperfusion injury Circulation Feb 15 2011123(6)594ndash604

[89] Baum J Duffy HS Fibroblasts and myo1047297broblasts what are we talking about J Cardiovasc Pharmacol Apr 201157(4)376ndash9

[90] Sundset R Cooper M Mikalsen SO Ytrehus K Ischemic preconditioning protectsagainst gapjunctional uncoupling in cardiac myo1047297broblasts Cell Commun AdhesMar-Aug 200411(2ndash4)51ndash66

[91] Cooper M Ytrehus K Cell survival signalling in heart derived myo1047297broblasts in-duced by preconditioning and bradykinin the role of p38 MAP kinase Mol CellBiochem Apr 2004259(1ndash2)83ndash90

[92] Hagar JM Hale SL Kloner RA Effect of preconditioning ischemia on reperfusionarrhythmias after coronary artery occlusion and reperfusion in the rat Circ Res

Jan 199168(1)61ndash8[93] Shiki K Hearse DJ Preconditioning of ischemic myocardium reperfusion-

induced arrhythmias Am J Physiol Dec 1987253(6 Pt 2)H1470ndash6[94] Vanhoutte D Heymans S TIMPs and cardiac remodeling embracing the MMP-

independent-side of the family J Mol Cell Cardiol 201048(3)445ndash53[95] M Dobaczewski C Gonzalez-uesada NG Frangogiannis The extracellular ma-

trix as a modulator of the in1047298ammatory and reparative response following myo-cardial infarction J Mol Cell Cardiol Mar 201048(3)504ndash11

[96] Vilahur G Juan-Babot O Pena E Onate B Casani L Badimon L Molecular and cel-lular mechanisms involved in cardiac remodeling after acute myocardial infarc-tion J Mol Cell Cardiol Mar 201150(3)522ndash33




[97] Nilsson L Hallen J Atar D Jonasson L Swahn E Early measurements of plasmamatrix metalloproteinase-2 predict infarct size and ventricular dysfunction inST-elevation myocardial infarction Heart 20129831ndash6

[98] Williams-Pritchard G Knight M Hoe LS Headrick JP Peart JN Essential role of EGFR in cardioprotection and signaling responses to A1 adenosine receptorsand ischemic preconditioning Am J Physiol Jun 2011300(6)H2161ndash8

[99] Methner C Donat U Felix SB Krieg T Cardioprotection of bradykinin at reperfu-sion involves transactivation of the epidermal growth factor receptor via matrixmetalloproteinase-8 Acta Physiol (Oxf) Dec 2009197(4)265ndash71

[100] Yu Q Stamenkovic I Cell surface-localized matrix metalloproteinase-9 proteo-lytically activates TGF-beta and promotes tumor invasion and angiogenesis

Genes Dev Jan 15 200014(2)163ndash

76[101] Hausenloy DJ Yellon DM Cardioprotective growth factors Cardiovasc Res Jul 15200983(2)179ndash94

[102] MoshalKS Tyagi N HendersonB Ovechkin AVTyagi SCProtease-activated recep-tor and endothelial-myocyte uncoupling in chronic heart failure Am J Physiol Jun2005288(6)H2770ndash7

[103] Cheung PY Sawicki G Wozniak M Wang W Radomski MW Schulz R Matrixmetalloproteinase-2 contributes to ischemiandashreperfusion injury in the heartCirculation Apr 18 2000101(15)1833ndash9

[104] Sawicki G Leon H Sawicka J Sariahmetoglu M Schulze CJ Scott PG et al Degra-dation of myosin light chain in isolated rat hearts subjected to ischemia-reperfusion injury a new intracellular target for matrix metalloproteinase-2Circulation Jul 26 2005112(4)544ndash52

[105] Sung MM Schulz CG Wang W Sawicki G Bautista-Lopez NL Schulz R Matrixmetalloproteinase-2 degrades the cytoskeletal protein alpha-actinin in peroxy-nitrite mediated myocardial injury J Mol Cell Cardiol Oct 200743(4)429ndash36

[106] Ali MA Cho WJ Hudson B Kassiri Z Granzier H Schulz R Titin is a target of ma-trix metalloproteinase-2 implications in myocardial ischemiareperfusion inju-ry Circulation Nov 16 2010122(20)2039ndash47

[107] Bencsik P Kupai K Giricz Z Gorbe A Pipis J Murlasits Z et al Role of iNOS andperoxynitrite-matrix metalloproteinase-2 signaling in myocardial late precondi-tioning in rats Am J Physiol Aug 2010299(2)H512ndash8

[108] Giricz Z Lalu MM Csonka C Bencsik P Schulz R Ferdinandy P Hyperlipidemiaattenuates the infarct size-limiting effect of ischemic preconditioning role of matrix metalloproteinase-2 inhibition J Pharmacol Exp Ther Jan 2006316(1)154ndash61

[109] Romero-Perez D Fricovsky E Yamasaki KG Grif 1047297n M Barraza-Hidalgo MDillmann W et al Cardiac uptake of minocycline and mechanisms for in vivocardioprotection J Am Coll Cardiol Sep 23 200852(13)1086ndash94

[110] Xin W Yang X Rich TC Krieg T Barrington R Cohen MV et al Allpreconditioning-related G protein-coupled receptors can be demonstrated inthe rabbit cardiomyocyte J Cardiovasc Pharmacol Ther 201217190ndash8

[111] Safran N Shneyvays V Balas N Jacobson KA Nawrath H Shainberg A Cardiopro-

tective effects of adenosine A1 and A3 receptor activation during hypoxia in iso-lated rat cardiac myocytes Mol Cell Biochem Jan 2001217(1 ndash2)143ndash52[112] Wu S Li HY Wong TM Cardioprotection of preconditioning by metabolic inhibi-

tion in the rat ventricular myocyte Involvement of kappa-opioid receptor CircRes Jun 25 199984(12)1388ndash95

[113] Pan TT Feng ZN Lee SW Moore PK Bian JS Endogenous hydrogen sul1047297de con-tributes to the cardioprotection by metabolic inhibition preconditioning in therat ventricular myocytes J Mol Cell Cardiol Jan 200640(1)119ndash30

[114] Kempf T Eden M Strelau J Naguib M Willenbockel C Tongers J et al The trans-forming growth factor-beta superfamily member growth-differentiation factor-15protects the heart from ischemiareperfusion injury Circ Res Feb 17 200698(3)351ndash60

[115] Hu X Dai S Wu WJ Tan W Zhu X Mu J et al Stromal cell derived factor-1 alphaconfers protection against myocardial ischemiareperfusion injury role of thecardiac stromal cell derived factor-1 alpha CXCR4 axis Circulation Aug 72007116(6)654ndash63

[116] Wang Y LauWB GaoE TaoL YuanY Li R et al Cardiomyocyte-derived adiponec-tin is biologically active in protecting against myocardial ischemiandashreperfusion in-

jury Am J Physiol Endocrinol Metab Mar 2011298(3)E663ndash70

[117] Wang Y Guo Y Zhang SX Wu WJ WangJ Bao W et al Ischemic preconditioningupregulates inducible nitric oxide synthase in cardiac myocyte J Mol Cell Cardiol

Jan 200234(1)5ndash15




million deathsper annumdirectly attributable to myocardial ischaemia

and is projected to increase still further [1] The gold standard manage-

ment of acute coronary syndromes remains expeditious revascularisa-

tion achieved with percutaneous coronary intervention (PCI) While

rapid restoration of blood 1047298ow to the ischaemic area of myocardium is

essential to stave off necrosis within the area at risk the question

remains as to whether the myocardial salvage achieved by timely

revascularisation can be augmented by manipulation of endogenous

cytoprotective mechanisms or exogenous inhibition of destructive pro-cesses activated by the combination of injurious ischaemia and reperfu-

sion In this review we shall undertake to discuss the modalities of

cardiac conditioning and how cardiac conditioning relates to the three

predominant cell populations (cardiomyocyte endothelium and 1047297bro-

blast) and the extracellular matrix of the heart

11 Ischaemiareperfusion injury

Non-re-vascularised occlusion of an epicardial coronary artery will

inevitably result in necrosis of the supplied myocardium (the area at

risk) If the occlusion is relieved through percutaneous intervention

the result of reestablishing 1047298ow down the re-opened coronary artery

is often rapidly appreciated with prompt brisk 1047298ow down the distal

vessel However this is not uniformly achieved 1047298ow may be sluggish

or even non-existant in 30ndash40 of cases presenting as a primary PCI

for acute ST-elevation myocardial infarction (the no-re1047298ow phenome-

non) [2] This may be the result of distal embolisation of the ruptured

atherosclerotic plaque and clot that had initially occluded the coronary

artery mdash or may in fact be an early angiographic manifestation of what

has been termed ldquoreperfusion injuryrdquo re1047298ecting marked endothelial in-

jury manifest by endothelial oedemaand discontinuity arising as a con-

sequence of ischaemia and reperfusion (review [34]) The no re1047298ow

phenomenon resulting from microvascular obstruction (MVO) is in it-

self associated with a worse clinical outcome [5] and thus amelioration

of microvascular dysfunction resulting from ischaemiareperfusion in-

jury may be a key target in the clinical management of reperfusion inju-

ryIn theabsence of MVO we know from basic science investigationsin

a wide variety of animal models that re-opening of the coronary artery

results in reperfusion injurya componentof myocardialdeath thatmaybe amenable to pharmacological manipulation or cardiac conditioning

up to 40ndash50 of the infarct observed following re-vascularisation is

thought to be potentially salvagable through conditioning modalities

[67]

2 Cardiac conditioning

Since the original description of the recruitment of innate cardiopro-

tection by a transient ischaemia and reperfusion prior to a lethal ischae-

mic insult by Murry Reimer and Jennings in 1986 [8] the basic

understanding of the modes of cellular injury (necrosis apoptosis

autophagy) and the signaling and mechanisms that increase resistance

to cell death has expanded dramatically Moreover these innate cardio-

protective mechanisms can not only be recruited by non-lethal ischae-miaof theheart but also throughpharmacological manipulation or even

remote-organ ischaemia prior (preconditioning) during (per-condi-

tioning) and immediately following (postconditioning) the ischaemic

insult The reader is encouraged to read one of the many excellent re-

cent reviews on this subject that cover the mechanisms by which con-

ditioning manifests protection and how these are currently being

translated into clinical practice [910]

At the mechanistic level recruitment of cardioprotection by condi-

tioning can be subdivided into triggers mediators and end-effectors

The mitochondrial permeability transition pore (mPTP) is currently the

strongest candidate for an end-effector of cardiac conditioning As

reviewed elsewhere modulation of the mPTP is thought to have a signif-

icant impact upon the myocardium in the reperfusion phase of injurious

ischaemiareperfusion injury [9] The up-stream signaling that impacts

upon the open probability of the mPTP mdash the mediators of conditioning

protection mdash have been subject of intense research and include variously

PI3K Akt Erkand Stat (reviewed [11ndash13]) (Fig 1) The activators of these

signaling cascades are termed ldquotriggersrdquo and are broadly extracellular

receptorligand interactions with autocoid endocrine or paracrine signal-

ing molecules A signi1047297cant number of receptors have been identi1047297ed

which when stimulated (so-called preconditioning-mimetics) may result

in recruitment of an ischaemiareperfusion resistant phenotype It has

been long recognised that the ischaemic conditioning signal is a summa-tion of multiple signals derived from multiple disparate receptorligand

interactions that once a suf 1047297cient combined signal is generatedbreeches

a lsquothresholdrsquo for triggering the conditioning response [14] What has

not been explored in great detail to date is how the various cellular

components within the myocardium contribute to this lsquoconditioning

souprsquo or how the various cellular populations interact to propagate the

conditioning signal Extracellular receptors offer considerable opportuni-

ty for inter and intracellular interaction and the study of these may

offer new insights into mechanisms underlying conditioning the heart

(Fig 2)

3 Composition of the myocardium targets for conditioning

Broadly the data available in the study of the conditioning phe-

nomenon have not endeavored to specify the cell type involved in

the protective pathway nor which is the cellular target for end-

effector protection since it is frequently assumed that conditioning

applies homogenously or that the speci1047297c target is the cardiac myo-

cyte There may be some justi1047297cation in this pragmatic view but it

is clearly a simpli1047297cation Indeed it is perfectly possible to trigger

protection in the isolated myocyte but in in-vitro whole tissue prep-

arations or in-vivo cardiac myocytes are unlikely to be conditioned in

isolation In studies of cardiac conditioning the archetypal primary

end point has been myocardial viability within the region at risk

Typically this is assessed by a number of methods that ultimately

leads to expression of infarct size as a ratiometric determination of

infarct to risk zone volumes By volumetric determination cardiac

myocytes are the predominant cell type estimated to contribute

around 75ndash80 of the total myocardium volume [1516] The nextmost prominent cell populations endothelium (3) and 1047297broblasts

(2) appear contribute relatively little by this measure [15] Macro-

phages (01) and pericytes (02) contribute a negligible amount

to the total volume and the remainder of the myocardial volume is

non-cellular comprising of connective tissue (6) and intracapillary

luminal volume (8ndash9) [15] However while cardiac myocytes are

the largest cell by volume within the myocardium they are not nec-

essarily the most numerate cell type depending on the methodology

the myocyte population contributes between 35 and 55 of total cell

number with variability between species age of the animal and re-

gion from where the myocardium is isolated [1718] The principal

non-myocyte cell populations consist of 1047297broblasts and endothelium

[16] but also include vascular smooth muscle macrophages and cir-

culating blood cells [1920] From Banerjees work using FACS analy-sis to count differing cell populations in adult murine myocardium

myocytes were found to contribute 56 of cell number followed by

1047297broblasts (27) endothelial cells (7) and vascular smooth muscle

cells (10) [18] Thus it can be argued that when ischaemic condition-

ing is triggered it is not an obligate phenomenon of cardiomyocytes

alone but rather a combination and interaction between the many

cell types found within the heart as illustrated in Fig 2 Interestingly

there are data available to support the hypothesis of cellular interac-

tion in response to a conditioning stimulus within the myocardium

Erythropoietin (EPO) is a well recognised preconditioning mimetic

[2122] with receptors found on a wide number of cell populations

within the myocardium including cardiomyocytes [23] Exposure to

EPO results in conditioning of the myocardium against lethal ischae-

mic insult Interestingly Lefers group developed a transgenic mouse





GPCR TKLR

1a

2

1b

3











FGF2 EGF

Growth factors


MMP


Achendorthins

PDGFGrowth factors

cytokins

Circulatoryfactors













[24]






31 Endothelium





























mia [28]





































solve this problem




























































[57])




































































conditioning [68]

















32 Fibroblasts


































ing [80]


















(review [85])

























































ticular interest








































investigation




34 Cardiac myocyte






























4 Conclusions












None

Acknowledgements







References








































































































































Jan 200234(1)5ndash15





GPCR TKLR

1a

2

1b

3











FGF2 EGF

Growth factors


MMP


Achendorthins

PDGFGrowth factors

cytokins

Circulatoryfactors













[24]






31 Endothelium





























mia [28]





































solve this problem




























































[57])




































































conditioning [68]

















32 Fibroblasts


































ing [80]


















(review [85])

























































ticular interest








































investigation




34 Cardiac myocyte






























4 Conclusions












None

Acknowledgements







References








































































































































Jan 200234(1)5ndash15









[24]






31 Endothelium





























mia [28]





































solve this problem




























































[57])




































































conditioning [68]

















32 Fibroblasts


































ing [80]


















(review [85])

























































ticular interest








































investigation




34 Cardiac myocyte






























4 Conclusions












None

Acknowledgements







References








































































































































Jan 200234(1)5ndash15

















[57])




































































conditioning [68]

















32 Fibroblasts


































ing [80]


















(review [85])

























































ticular interest








































investigation




34 Cardiac myocyte






























4 Conclusions












None

Acknowledgements







References








































































































































Jan 200234(1)5ndash15










ing [80]


















(review [85])

























































ticular interest








































investigation




34 Cardiac myocyte






























4 Conclusions












None

Acknowledgements







References








































































































































Jan 200234(1)5ndash15




34 Cardiac myocyte






























4 Conclusions












None

Acknowledgements







References








































































































































Jan 200234(1)5ndash15


































































































Jan 200234(1)5ndash15


Conditioning the whole heart—not just the cardiomyocyte

Documents

Transcript of Conditioning the whole heart—not just the cardiomyocyte