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J Thorac Cardiovasc Surg 1998;115:200-204
© 1998 Mosby, Inc.
CARDIOPULMONARY SUPPORT AND PHYSIOLOGY |
From the Department of Cardiothoracic Surgery, Heart Science Centre,National Heart and Lung Institute at Harefield Hospital, Harefield, Middlesex,United Kingdom.
Received for publication April 7, 1997; revisions requested May 14,1997; revisions received July 9, 1997; accepted for publication August 13,1997. Address for reprints: Professor Sir Magdi Yacoub, FRCS, Departmentof Cardiothoracic Surgery Heart Science Centre, National Heart and Lung Instituteat Harefield Hospital, Harefield, Middlesex. UB9 6JH, United Kingdom.
Abstract
Objectives: Induction of the 70 kdheat shock protein in the heart is known to exert a protective effect againstpostischemic mechanical and endothelial dysfunction. However, the exact siteof induction and the mechanisms involved remain unknown. The aim of this studywas to investigate the relative capacity of endothelial and myocardial cellsto express the 70 kd heat shock protein in response to heat stress, as wellas their significance.
Methods: (1) Postischemic recoveryof cardiac mechanical and endothelial function was studied in isolated rathearts with and without endothelial denudation with saponin. (2) Semiquantitativedetermination of induction of 70 kd heat shock protein by Western immunoblottingwas performed in the whole cardiac homogenate, in isolated cardiac myocytes,and in coronary endothelial cells. (3) Immunocytochemistry was used to visualizethe distribution of induction of 70 kd heat shock protein in both cell types.
Results: Postischemic recovery (percentpreischemic value ± standard error of the mean) of cardiac outputin hearts from heat-stressed animals was significantly improved (66.7 ±6.9 vs 44.5 ± 4.5 in the control group, p < 0.01). In heat-stressed hearts treated with saponinno improvement in the recovery of cardiac output was noted (44.7 ±6.9 in heat-stressed hearts vs 38.0 ± 4.0 in heat-stressed, saponin-treatedhearts, p = not significant). Endothelialfunction (as assessed by the vasodilatory response to the endothelium-dependentvasodilator 5-hydroxytryptamine) improved from 31.0 ± 5.2 in thecontrol group to 65.8 ± 7.1 in heat-stressed hearts (p < 0.02 vs control) and dropped to –1.9 ±3.8 in heat-stressed hearts treated with saponin. Immunocytochemistry showedthat only sections of hearts from heat-treated rats showed a strong specificreaction with heat shock protein antibody. The positive staining was seenin endothelial cells. Induction of 70 kd heat shock protein content in thewhole cardiac homogenate from heat-stressed rats as measured by Western immunoblottingwas 5.2 ± 1.9 (vs 0.0 in non-heat–stressed rats, p < 0.0001) and dropped to 0.0 in heat-stressedhearts treated with saponin. The tentative amount of 70 kd heat shock proteinwas 18.1 ± 7.8 in isolated endothelial cells from heat-stressedhearts and 2.3 ± 2.3 in isolated cardiac myocytes (p < 0.01 vs endothelial cells).
Conclusions: Coronary endothelial cellsare the main site of induction of 70 kd heat shock protein in the heart andappear to contribute to the protective effects of heat stress on the recoveryof mechanical and endothelial function.
Heat-shock proteins (HSPs) are synthesized in most cells in responseto an increase in temperature or after exposure to a variety of physical orchemical stimuli.
1 Most areconstitutively expressed in normal, unstressed cells, where they play an importantphysiologic role by facilitating several aspects of protein maturation inthe cell, hence they have been called "molecular chaperones." 2 They also play a role in cellularhomeostasis in a variety of clinically relevant conditions, including malignancy,immune responses, infection, and ischemia. 3
Compelling evidence now exists that induction of 70 kd HSP (HSP70),the most abundant and the best studied HSP, is associated with an enhancementof cardiac mechanical and endothelial function after ischemia. 4 Endothelial function is known to be of major importancein the maintenance of basal coronary flow and normal mechanical function, 5 although the integrity of postischemiccoronary endothelial cells has become an important issue in the general strategyof myocardial protection. 6The contribution of cardiac myocytes and coronary endothelial cells in HSP70expression has not been investigated. The aim of this study was to investigatethe relative amount of HSP70 induced in these cellular components.
Material and methods
Experimental time course.
Five series of experiments were performed in this study. Series 1: fourgroups of hearts (n = 6 in each group)were studied to evaluate the preischemic and postischemic (4 hours of hypothermiccardioplegic arrest at 4° C) endothelial and mechanical function in controlhearts perfused with Krebs-Henseleit carbonate buffer (KH) (group 1) or saponin(group 2) and in heat-shocked hearts perfused with KH (group 3) or saponin(group 4). Series 2: four similar groups (n =6 in each group) were used to evaluate the relative amount of induction of70 kd HSP in the whole homogenate by Western immunoblotting. Series 3: Heartsfrom all four groups (n = 6 in each group)were used to assess the preischemic and postischemic wet weight. Hearts weredisconnected from the Langendorff apparatus, weighed, and reconnected. Thetime taken for the disconnection, weighing, and reconnecting ranged from 21to 33 seconds. Series 4: Hearts from heat-shocked (n = 4) and control (n =4) animals were used to evaluate induction of 70 kd HSP (iHSP70) in isolatedendothelial cells and myocytes. Series 5: Hearts from heat-shocked (n = 6) and control (n =4) animals were used to visualize iHSP70 in isolated endothelial and myocardialcells by immunocytochemistry.
Animals.
Sprague-Dawley rats weighing between 300 and 330 gm were used in allexperiments. Male rats were used to avoid a potential lack of homogeneityrelated to hormonal cycle. Six hearts were studied in each group. In all studies,animals received humane care in compliance with the "Principles of LaboratoryAnimal Care" formulated by the National Society for Medical Researchand the "Guide for the Care and Use of Laboratory Animals" preparedby the Institute of Laboratory Animal Resources and published by the NationalInstitutes of Health (NIH publication 86-23, revised 1985).
Induction of heat stress.
Rats were anesthetized with an intraperitoneal injection of sodium pentobarbitone(50 mg/kg), then placed on a temperature-controlled heating pad (IMS K-Tempcontrol unit; Congleton, Cheshire, United Kingdom) set at 45° C untilbody temperature reached 42° C. Body temperature was monitored with arectal temperature probe and maintained between 42° C and 42.5° Cfor 15 minutes as previously described. 1 The animals were left to recover for 24 hours before analysis of mechanicalfunction or determination of HSP70 content.
Isolated working rat heart preparation.
The isolated working rat heart preparation, which has been describedin detail elsewhere, 7 wasused in this study. In this left heart preparation oxygenated KH (NaCl, 118.5mmol/L; NaHCO3, 25 mmol/L; KCl, 4.75 mmol/L; MgSO4,1.19 mmol/L; KH2PO4, 1.18 mmol/L; CaCl2,2.5 mmol/L), pH 7.4, containing glucose (11.1 mmol/L) and gassed with 95%oxygen and 5% carbon dioxide at 37° C, enters the cannulated left atriumand passes into the left ventricle, from which it is spontaneously ejectedthrough an aortic cannula against a hydrostatic pressure of 100 cm H2O. The heart continues to eject as long as the pressure generated inthe left ventricle is greater than 100 cm H2O. Total cardiopulmonarybypass with maintained coronary perfusion can be simulated by clamping theleft atrial cannula and introducing perfusion fluid at 37° C into theaorta from a reservoir 100 cm above the heart. With this preparation, whichis essentially that described by Langendorff, the heart will continue to beatbut does not perform external work. Several parameters, including cardiacoutput (CO) and coronary flow, are measured to assess cardiac function. Ischemiccardiac arrest may be produced by clamping the aortic cannula. At this timea cardioplegic solution is infused into a sidearm of the aortic cannula. Duringthe ischemic period the heart is maintained under hypothermic conditions (4°C) by a cooling circuit.
Cardioplegic solution.
St. Thomas' Hospital cardioplegic solution No. 1, supplied as concentrate(David Bull Laboratories, Mulgrave, Victoria, Australia), was diluted in Ringer'ssolution (Travenol Laboratories, Thetford, Norfolk, United Kingdom) and passedthrough a 0.2 µm filter (Pall Biomedical, Glen Cove, N.Y.).
Endothelial function.
Endothelial function was assessed through observations of preischemicand postischemic coronary flow responses to 5-hydroxytryptamine (5-HT). Thisvasodilatory response is endothelium dependent. In the intact endothelium,5-HT causes vasodilatation through the release of NO, whereas in the presenceof endothelial damage it causes vasoconstriction by a direct effect on smoothmuscle. Our protocol for this test has been described by us in earlier studies. 8 We have previously shown that perfusionof the isolated rat heart by 5-HT leads to an increased secretion of nitricoxide in the coronary effluent. Endothelium-independent vasodilatation, asassessed by the response to glyceryl trinitrate, has been shown to be unchangedby a similar protocol (i.e., 4 hours hypothermic [4° C] cardioplegic arrest). 8 After excision of the heart andaortic cannulation, Langendorff perfusion was initiated at 37° C. Coronaryflow was monitored by an in-line electromagnetic flow probe (ECM2 20 ml, Scalar,Delft, Holland) proximal to the aortic cannula, connected to its compatibleflowmeter (MDL 140 l, Scalar). This provided an accurate (0.0 to 40.0ml/min) digital readout of mean coronary flow and simultaneous hard-copy recordingthrough a connection with a chart recorder (series 3000, Gould Electronics,Hainault, Essex, United Kingdom) that allowed accurate monitoring of steadystate conditions (<0.3 ml/min change in coronary flow over 3 minutes).The calibration of the flow meter is performed using the company manual andby collecting coronary flow in a measuring cylinder over 1 minute. After 9to 13 minutes, the initial baseline coronary flow was recorded. The Langendorffinfusion was switched to one containing additional 10–5 mmol/L5-HT (Sigma Chemical Co., Poole, Dorset, United Kingdom). The ensuing vasodilatorresponse was monitored and when the steady state had been reached (between5 and 7 minutes), the coronary flow was recorded. After this period, 5-HTwas washed out by switching back to ordinary KH until a steady state had beenreached (between 5 and 7 minutes).
The heart was then subjected to a 10 ml hypothermic (4° C) infusionwith the cardioplegic solution and maintained immersed in the same solutionfor 4 hours at 4° C. At the end of the ischemic period, the heart wasreperfused in the Langendorff mode at 37° C for at least 15 minutes. Whenthe baseline coronary flow had been reestablished, the heart was again subjectedto the same protocol of sequential infusion of 5-HT and KH as in the preischemicperiod. The rationale of using the steady baseline coronary flow has beendetailed in our previous study. 8
Endothelial cell removal with saponin.
To assess the importance of HSP70 induced in endothelial cells, endothelialfunction and mechanical function were assessed and compared in hearts perfusedwith or without saponin. This potent detergent agent has previously been shownto remove endothelial cells. 9Hearts were perfused with saponin (30 µg/ml) dissolved in KH solutionin three cycles, each consisting of a 2-minute perfusion period that precededthe ischemic interval.
Isolation of myocytes.
This technique has been described in detail elsewhere. 10 Hearts were perfused for 1 minute in the Langendorffmode with Krebs buffer to remove all traces of blood, and then a piece ofventricle was sliced quickly into pieces of approximately 1 mm3using razor blades. The pieces of myocardium were incubated for a total of12 minutes at 35° C in 25 ml of low-calcium medium containing the followingcomposition (in mmol/L): NaCl, 120; KCl, 5.4; MgSO4, 5; pyruvate,5; glucose, 20; taurine, 20; HEPES solution, 10; nitrilotriacetic acid, 5;pH, 6.96; containing 1 to 2 µm calcium. The medium was changed threetimes during this incubation and was stirred by bubbling with 100% oxygen.The low-calcium medium was removed by filtering through 300 µm gauze.The pieces of myocardium were then incubated at 35° C for 45 minutes inthe same solution but with nitrilotriacetic acid omitted, and 4 IU/ml typeXXIV Protease (Sigma) and 30 µm calcium added, followed by two 45-minuteincubations with the protease omitted and 400 IU/ml collagenase (BCL, BoehringerMannheim, United Kingdom Ltd.) added. The medium was shaken gently throughoutthe incubation and kept under 100% oxygen. At the end of each 45-minute periodthe solution containing the dispersed cells was filtered through 300 µmgauze and centrifuged at 1000 rpm for 1 to 2 minutes. The cells were thenwashed twice by centrifugation in KH. 10
Isolation of endothelial cells.
The method used to isolate endothelial cells has been described in detailelsewhere. 11 The fat, epicardium,and endocardium were removed from the ventricular tissue and diced up usingrazor blades. An amount of collagenase type II equivalent to twice the volumeof tissue was added, and this was incubated for 1 hour at 37° C with agitation.Hanks medium with 5% FCS (fetal calf serum) was added and the digest and spunat 1400 rpm for 10 minutes. The supernatant was gently poured off, and thepellet was washed in 10% bovine serum albumin (BSA) and spun at 1000 rpm for7 minutes. The pellet was again washed in Hanks medium with 5% FCS and spunat 1200 rpm for 6 minutes. The pellet was then incubated with twice its volumeof trypsin–ethylenediaminetetraacetic acid (EDTA) (0.25% in 1 mmol/LEDTA) at 37° C for 10 minutes with agitation. After this period, the tissuewas washed twice in Hanks medium with 5% FCS at 1200 rpm for 6 minutes andfiltered through 100 µm gauze. 11
Assessment of heat shock protein expression.
The induction of HSP70 was assessed by sodium dodecyl sulfate (SDS),polyacrylamide gel electrophoresis, and Western immunoblotting as previouslydescribed. 4 Whole heart homogenate,isolated endothelial cells, and isolated myocytes were solubilized in 1% wt/volSDS and assayed for total protein with the Bradford assay, denatured by heatingat 100° C in Laemmli buffer, and separated on 10% SDS gels until the bromophenolblue tracking dye reached the end of the gel. Gels were equilibrated for 30minutes in transfer buffer, and transfer of the proteins was performed for1 hour at 500 mA. Western blots were blocked using 3% wt/vol nonfat driedmilk (Marvel) in phosphate-buffered saline solution (PBS) containing 0.05%wt/vol Tween-20 (PBS-T) for 1 hour to block nonspecific binding sites. Theblots were then probed with mouse antibodies specific to inducible HSP70 (BioquoteLtd, United Kingdom) diluted to a final concentration of 1:1000 for 1 hour.Blots were washed three times and incubated with secondary horseradish-peroxidase–conjugatedrabbit anti-mouse antibody for 1 hour. The result was visualized using anenhanced chemiluminescence (ECL) detection system (Amersham). Hyperfilm MP(myoperoxidase) was exposed to blots treated with ECL for 30 seconds and developedin an automatic film processor. After ECL exposure, antibodies were removedfrom blots by incubation in a solution of 2% wt/vol SDS, 6.25% vol/vol 1 mmol/LTris-HCl, pH 6.8, and 0.7% vol/vol 2-mercaptoethanol. Proteins were then visualizedby staining with 0.01% Amido Black in a solution of methanol, water, and aceticacid (ratio of 45:45:10 vol/vol). Amido black–stained blots and ECLfilms were scanned with a Molecular Dynamics 300A laser densitometer and HSP70levels determined as a proportion of total protein loaded using the QuantityOne software package (PDI, Huntington, N.Y.).
Immunocytochemistry.
Rat hearts were sliced into pieces approximately 5 mm thick and snapfrozen in liquid nitrogen. Six hearts from rats subjected to heat shock andfour hearts from control groups were analyzed. For each heart, four serialfrozen sections, each 6 µm thick, were cut onto glass slides, allowedto dry for 30 to 120 minutes, and subsequently fixed for 15 minutes in acetonebefore the immunohistochemical procedure. Sections were incubated for 16 hoursat 4° C in either monoclonal mouse antibody to the inducible form of heatshock protein 72 (iHSP72) (Bioquote Ltd., United Kingdom) at a dilution of2 µg/ml or to endothelial cells with the antibody RECA-1 (Serotec, Oxford,United Kingdom) provided as a supernatant and diluted 1:150. Negative controlsincluded incubation of sections in antibody diluent only or in an irrelevantIgG1 monoclonal mouse antibody used at 2 µg/ml (Prod. no.X0931, Dako Ltd., High Wycombe, United Kingdom). This corresponds to the sameimmunoglobulin subclass as the iHSP72 and RECA-1 antibodies. All sectionswere subsequently incubated for 30 minutes at room temperature in biotinylatedrabbit anti-mouse F(ab)2 at a concentration of 0.002 µg/mland prepared at least 60 minutes before use in a 5% solution of rat serum.All antibodies were diluted in 0.005 mmol/L Tris-buffered saline solution,pH 7.6. After this, sections were immersed for 30 minutes at room temperaturein streptavidin biotin-horseradish peroxidase complexes (Dako Ltd.), whichhad been prepared at least 30 minutes before use by diluting the components1:200 in 0.05 mmol/L Tris buffer, pH 7.6. 12 Sections were then treated for 5 minutes with a diaminobenzidinehydrochloride/hydrogen peroxide substrate, prepared according to the manufacturer'sinstructions (Prod. no. D5905, Sigma, Poole), to visualize the antigenic sites.Each of the above stages was followed by three 5-minute washes in 0.005 mmol/LTris-buffered saline solution, pH 7.6. The sections were then rinsed in coldrunning tap water, counterstained with Mayer's hematoxylin, washed again incold running tap water, and then dehydrated through graded alcohols clearedin CNP30 (Merck, Lutterworth, United Kingdom) and mounted in DPX mountant(BDH, Poole, United Kingdom).
Statistical analysis.
Postischemic recoveries of mechanical function and coronary flows werecompared using two-way analysis of variance (ANOVA) with Scheffe's correctionfactor. Statistical significance of differences between groups was determinedwith a nonpaired Student's t test, and significancewas assumed when the p value was 0.05 or less.Values are given as means ± standard error of the mean (SEM).Preischemic and postischemic wet weights were compared with a paired t test.
Differences between postischemic wet weights were compared using ANOVAwith Scheffe's correction. Levels of HSP70 were compared using a nonpairedStudent's t test.
Results
Cardiac endothelial and mechanical function and coronary flow.
Table I shows preischemic and postischemic indices of cardiac mechanicalfunction CO and endothelial function as assessed by percentage change in coronaryflow in response to challenge with 5HT (En Fn), as well as recovery of functionas assessed by the same measurements after ischemia. In the heat-stressedgroup, the postischemic recovery (as percentage of the preischemic value ±SEM) of CO and En Fn were significantly improved when compared with the controlgroup and was: 66.9 (6.9) versus 49.5 (4.5) (p <0.01) and 65.8 (7.1) versus 31.0 (5.2) (p <0.02), respectively. The beneficial effect of heat stress was abolished bysaponin treatment: the postischemic recovery (as percentage of the preischemicvalue ± SEM) of CO and En Fn was significantly reduced to 44.7(6.9) versus 38.0 (4.0) (p < 0.05), –1.9(3.8) versus –3.6 (4.2) (p <0.001), respectively. Preischemic and postischemic coronary flow data areshown in Table II. After heat stress, no significantdifference was found in postischemic coronary flow compared with control hearts.However, saponin before treatment induced a significant reduction in postischemiccoronary flow compared with non-saponin–treated hearts, but no differencewas found between heat-stressed and control hearts after saponin treatment.
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Inducible HSP70.
In hearts perfused with saponin the relative amount of iHSP70 was 0in heat-shocked hearts (n = 6) and 0in control hearts (non-heat-shocked, n =6). Without saponin, iHSP70 was 5.2 (p <0.0001) in heat-shocked hearts (n = 6)and 0 in control hearts (non-heat-shocked hearts, n =6). This is graphically shown in Fig. 3.
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This study has demonstrated that after whole body heat stress, coronaryendothelial cells are the major site of iHSP70 in the heart. In addition,when endothelial cells were chemically removed, the protective effect of heatstress on postischemic recovery of mechanical function was abolished. Thedata obtained within this study are relative and depend on a direct comparisonof HSP levels; this is not an attempt to give absolute quantification. Whencompared with myocytes, the induction of HSP70 is seen to be predominant inendothelial cells. The very low basal level of HSP70 recorded in this studycould have been due to the sensitivity of the antibody and does not affectthe overall conclusion, which depends on comparison.
Over the past decade, a large number of in vivo and in vitro studieshave established the beneficial effect of HSP in myocardial ischemia in modelsof global and regional ischemia. 13,14 In a clinically relevant modelof myocardial protection for cardiac transplantation, we have previously shownthat both cardiac mechanical and endothelial function were improved by heatstress. 15 Endothelial functionhas been shown to influence myocardial performance. 12 Although endothelial cells are known to responddifferently to heat stress, 16and despite the fact that endothelial function has become an important componentof the general strategy in cardiac preservation, 17 to date no comparative data exist regarding iHSPin the different cellular components of the heart.
By the use of immunocytochemistry we have demonstrated that 24 hoursafter heat stress, when the maximal level of cardiac iHSP70 is known to bereached, 18 staining for iHSP70is only seen in coronary endothelial cells. Cardiac myocytes did not stainfor iHSP70 in any of the hearts. Whereas isolated endothelial cells showeda high level of iHSP70, isolated cardiac myocytes demonstrated a very weakconcentration of the protein. The discrete localization of HSP in endothelialcells is unlikely to be the result of redistribution of the antigen acrossthe section after leakage from the myocyte during immunohistochemical analysis.The method and subsequent detection of iHSP is similar to the method usedby Mestril and colleagues. 19By means of this process of fixing, no suggestion of leakage regarding theantigen from the cells was made. In addition, we have measured and comparedthe relative amount of iHSP70 in homogenates of whole hearts subjected tochemical denudation of endothelial cells by saponin. No trace of iHSP70 couldbe detected after this treatment.
To investigate the significance of these findings we have assessed theeffect of endothelial denudation by saponin on the recovery of mechanicaland endothelial function in control and heat-stressed hearts. Heat-stressedhearts displayed a better postischemic recovery of CO and endothelial functionas assessed by the endothelium-dependent vasodilatory effect of 5-HT. Notsurprisingly, pretreatment with saponin suppressed this beneficial effecton endothelial function. However, we have demonstrated that the protectiveeffect of heat stress on postischemic mechanical function was lost in heat-stressedhearts whose endothelium was removed. It could be argued that the latter findingsmight be related to the exacerbation of postischemic edema after endothelialremoval. This was made unlikely because no difference in cardiac wet weightwas found between hearts perfused with saponin and intact hearts. The mostlikely explanation for the loss of the protective effect of heat stress onpostischemic mechanical function could be the dramatic reduction in coronaryreflow after saponin treatment. This is in keeping with a previous study showinga correlation between the level of postischemic coronary flow and the recoveryof mechanical function. 17This is also supported by the fact that normal endothelial function is necessaryto sustain mechanical function. 5 To verify the relationship between coronary flow and postischemicfunction, experiments that manipulate coronary flow should be carried out.However, in the working mode this is technically not feasible because it wouldonly be possible through manipulation of afterload.
It has also been suggested that saponin can affect cardiac myocyte membraneand possibly alter mechanical function. However, several studies using similarconcentrations have demonstrated that saponin has no effect on cardiac systolicor diastolic function, nor does it affect endothelium-independent function. 20,21 In addition, electron microscopic studies show that even at ahigher concentration of 50 µg/ml, saponin only caused damage to theendothelium, whereas the underlying histologic layers were left intact. 21 Given the route of administration,saponin undoubtedly contacted the inner surface of the atria and ventricles,which could cause endocardial dysfunction. 22 However, at a concentration of 30 µg/ml, as used in this study,it has been shown that the effects of saponin are specific to the endotheliumand do not impair cardiac function. 20,21
Data from recent studies show that after mild ischemia and heat stress(43° C), isolated cardiomyocytes express enough HSP70 to induce cytoprotection. 23 These data are at odds with ourfindings that after endothelial denudation no HSP70 could be measured in thewhole cardiac homogenate. Although the exact explanation of our findings isunknown, it would appear that endothelial cells probably produce per cellmore iHSP70 than the myocardium. Because the endothelium only represents 3%of the total cardiac mass, 24the endothelial cells may not be the most important site of overall iHSP70expression. Considering the implications of our findings, experiments thatuse immunocytochemistry (which evaluate the level of HSP70 before and afterheat stress and before and after addition of saponin in endothelial cellsand myocytes) could further validate our results.
To the best of our knowledge, no study is investigating the effect ofHSP70 expression and the processes of endothelial cell and myocyte isolation;therefore we do not know whether these processes have a direct effect on theprotein synthesis of HSP70. However, we can conclude that this effect is minimalbecause results from both the immunocytochemical analysis of intact rat heartsand cell culture show similar findings.
The question as to whether endothelial cells are capable of producingmore HSP than other cells has been raised in different organs subjected tovarious stressful stimuli. 6Within the territory of an experimental infarct, cerebral endothelial cellshave been shown to produce HSP70, whereas astrocytes and neurones lying inthe infarcted region expressed no proteins. 25 This was associated with a protective effect regarding endothelialcell survival. Similar observations were made in endothelial cells from pulmonaryarteries subjected to oxidative stress in the form of hydrogen peroxidase. 6
To our knowledge, this study is the first to demonstrate the relativecontribution of cardiac endothelial cells and myocytes in HSP70 expression.This highlights the crucial role of the coronary endothelium in the endogenousmechanism of cellular protection against ischemia. It remains to be shownwhether these data can be confirmed in a higher species. These findings couldhave important physiologic and clinical implications. In conclusion, we havedemonstrated that the main site of induction of iHSP70 is the coronary endotheliumand that the protective effect of heat stress is mediated by the endothelium.
We thank the Harefield Heart Transplant Trust and the British HeartFoundation for financial assistance.
References
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 L. H. E. H. Snoeckx, R. N. Cornelussen, F. A. Van Nieuwenhoven, R. S. Reneman, and G. J. Van der Vusse Heat Shock Proteins and Cardiovascular Pathophysiology Physiol Rev, October 1, 2001; 81(4): 1461 - 1497. [Abstract] [Full Text] [PDF]
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