M. OppertC. H. GleiterC. MüllerA. ReinickeN. von AhsenU. FreiK.-U. Eckardt

Kinetics and characteristics of an acutephase response following cardiac arrest

Received: 28 March 1999Final revision received: 10 June 1999Accepted: 1 September 1999

M. Oppert ´ A. Reinicke ´ N. von Ahsen ´U. Frei ´ K.-U.Eckardt ())Department of Nephrology andMedical Intensive Care,Campus Virchow Klinikum, CharitØ,Humboldt University,Augustenburger Platz 1, 13353 Berlin,Germanye-mail: kai-uwe.eckardt@charite.deTel.: + 49(30)45053132Fax: + 49(30)450539 09

C.MüllerDepartment of Clinical Chemistry,Campus Virchow-Klinikum, CharitØ,Humboldt University, Berlin, Germany

C. H. GleiterDepartment of Clinical Pharmacology,University of Göttingen, Germany

Abstract Objective: Inflammationand hypoxia are frequently associat-ed, but their interaction is poorlyunderstood. In vitro studies haveshown that hypoxia stimulates thegenes of acute phase proteins (APP)and cytokines known to induce APP.We decided to determine kineticsand potential determinants of anacute phase response after cardiacarrest and to assess whether isolatedmoderate hypoxia can induce APPin humans in vivo.Design: Prospective, observationalstudy in patients and human experi-ment.Setting: Tertiary care university hos-pital.Patients and participants: 22 patientsafter primarily successful cardiopul-monary resuscitation (CPR) and7 healthy volunteers.Interventions: None in patients; ex-posure of volunteers to simulatedaltitude (460 torr/6 h).Results: Following CPR, type-1 APP(C-reactive protein, a1-acidglyco-protein, serum amyloid A) and type-2 APP (haptoglobin, a1-antitrypsin)increased consistently within1±2 days and the `negative' APPtransferrin was downregulated. ThisAPP response occurred irrespective

of the cause of arrest, the estimatedtime of anoxia, clinical course or pa-tient outcome and was not differentin patients with and without infec-tious complications. Exposure ofhealthy volunteers to less severe butmore prolonged hypoxia did not in-duce APP, although a time depen-dent increase of serum erythropoi-etin (EPO) was measurable underthese conditions, indicating the acti-vation of oxygen dependent geneexpression.Conclusions: (i) A marked acutephase response occurs regularly af-ter cardiac arrest, but within thecomplexity of this situation the se-verity of hypoxia is not a predomi-nant determinant of this response.(ii) Despite in vitro evidence forsimilarities in the oxygen dependentregulation of APP and EPO pro-duction, the oxygen sensitivity ofthese proteins in vivo is different.(iii) Measurements of APP are notrevealing regarding infectious com-plications in the early phase afterCPR.

Key words Hypoxia ´ Ischemia ´Acute phase proteins ´ Cardiacarrest ´ Infections

Intensive Care Med (1999) 25: 1386±1394Ó Springer-Verlag 1999 ORIGINAL

Introduction

In critically ill patients, local or global inflammation fre-quently occurs simultaneously with tissue hypoxia.However, the interaction between reduced tissue oxy-genation and the induction and maintenance of inflam-matory processes and subsequent organ dysfunction ispoorly understood [1]. In patients surviving cardiac ar-rest, the most severe form of acute global hypoxia, a sys-temic inflammatory response may develop, which hasbeen termed ªpostresuscitation syndromeº and wasshown to have significant impact on patient morbidityand mortality [2±5]. Although hypoxia and/or subse-quent reoxygenation are believed to play a major rolein triggering the postresuscitation syndrome, direct ef-fects of changes in oxygenation on different componentsof this syndrome have not been characterized.

The synthesis of ªacute phase proteinsº (APP) in theliver changes significantly in the course of inflammatoryreactions and alterations in circulating APP levels aretherefore used to measure an inflammatory response[6]. The function of many APP remains incompletelyunderstood, but they are believed to play an importantrole in host defence and under certain conditions mayalso contribute to tissue injury [6,7]. The rate of synthe-sis of APP is determined at the level of gene expressionand is believed to be mainly regulated by cytokines. Ac-cording to their induction by different sets of cytokines,APP are subdivided into type-1 APP, which are stimu-lated by interleukin (IL)-1 type cytokines [e.g., IL-1a,IL-1b, tumor necrosis factor (TNF) a, TNFb) and type-2 APP, which are stimulated by IL-6 type cytokines(e. g., IL-6, IL-11) [6]. Moreover, certain proteins aredownregulated during the acute phase response andtherefore considered as ªnegativeº APP.

Interestingly, the genes for IL-1 and IL-6 have beenshown to be inducible by hypoxia in endothelial cells invitro [8,9], so that hypoxia may indirectly induce APPby stimulating these cytokines. In addition, it has recent-ly been reported that in vitro exposure of humanhepatoma cells to moderate hypoxia stimulates acutephase gene expression [7], which points toward a directlink between low oxygen supply and the induction ofan acute phase response. However, the effect of differ-ent types of hypoxia on APP production in humans invivo has not been determined.

We have therefore sequentially measured differenttype-1 and type-2 APP in patients after successful car-diopulmonary resuscitation (CPR) and have investigat-ed whether the APP response is related to the estimatedperiod of anoxia, neurological outcome or characteris-tics of the clinical course, including the development ofinfectious complications. We have also compared theAPP response to changes in plasma levels of the glyco-protein hormone erythropoietin (EPO), which is nor-mally produced in inverse relationship to oxygen supply

[10,11]. In addition, human volunteers were exposed tohypobaric hypoxia in a low pressure chamber in orderto test for a direct effect of more prolonged, moderatehypoxia on APP production.

Subjects and Methods

Patients

Twenty-two consecutive patients were studied after cardiac arrestwho survived for more than 72 h. They were admitted to the medi-cal intensive care unit of the Virchow-Klinikum in Berlin after suc-cessful CPR out of hospital. Patient characteristics, the cause ofcardiac arrest, the estimated collapse-to-first CPR attempt inter-val, infectious complications and final outcome are given in table 1.No patients were included in whom clinical and laboratory investi-gation or workup of the medical history suggested the presence ofa chronic inflammatory disease or a recent infection prior to cardi-ac arrest. In all patients the arrest was witnessed. Bystander CPRwas provided by medical professionals who happened to be at thescene in 4 cases (No 1, 6, 14, 16) and by laypersons in 3 cases (No8, 11, 22). Resuscitation was continued and in all other cases initiat-ed by ambulance crew of a mixed emergency medical service±fireservice, who are trained in basic life support and the use of auto-mated external defibrillators. Subsequent advanced cardiac lifesupport was provided by emergency physicians transported to thescene of cardiac arrest by special response units. Patients weretransported to hospital after restoration of spontaneous circulation(ROSC). Aliquots of plasma were obtained from blood samplesthat were drawn as part of the routine diagnostic tests after admis-sion and on each morning of the stay in the intensive care unit(ICU).

Patient status at the time of discharge from the ICU was re-corded using Glasgow-Pittsburgh cerebral performance categories,according to the Utstein recommendations [12]. In brief, the cate-gories are: 1 = conscious without or with minor psychological orneurological deficit, 2 = conscious with moderate cerebral disabili-ty, 3 = conscious with severe cerebral disability, 4 = unconsciousvegetative state and 5 = death.

Hypoxic exposure of healthy volunteers

Seven male volunteers aged 20±31 years participated in the studyafter being informed about the aim of the investigation and the ex-perimental protocol. All participants were healthy and, in particu-lar, free from any chronic disease or recent infection. Subjectswere exposed to a simulated altitude of 4000 m for 6 h in a decom-pression chamber as described previously [10]. Atmospheric pres-sure was reduced to 600 torr for 15 min and to 460 torr for the re-mainder of the time. Subjects remained in the chamber for a totalof 6 h. From each subject, 4 ml of venous blood was withdrawnfrom an indwelling cannula in the cubital vein before, every60 min during and immediately after the hypoxic exposure. In ad-dition, blood samples were taken 24, 48 and 72 h after the start ofhypoxic exposure. Previous measurements in volunteers of similarage had shown that this type of inspiratory hypoxia reduces themean alveolar partial pressure of oxygen from 100 to 48.6 torr [10].

The study protocol was approved by the local ethics committeeand the volunteers gave written informed consent.

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Analytical methods

Acute phase proteins

Plasma concentrations were determined of the three type-1 APPC-reactive protein (CRP), a1-acidglycoprotein (a1-AGP) and se-rum amyloid A (SAA), the two type-2 APP haptoglobin (HAP)and a1-antitrypsin (a1-AT) and the ªnegativeº APP transferrin(TRA). Plasma concentrations of these proteins were measuredby immunoturbidimetric methods: CRP using reagents from Grei-ner (Flacht Germany), adapted to a Dax 72 (Bayer, München, Ger-many) or a Hitachi 911 (Boehringer, Mannheim, Germany) ran-dom access clinical chemistry analyser; a1-AGP, a1-AT, HAP andTRA with reagents from Dako (Hamburg, Germany) adapted toHitachi 911 or Cobas Mira Plus (Roche, Grenzach, Germany) ran-dom access clinical chemistry analyzers. Calibrators from Dakoand Greiner were related to the CRM 470 reference material.SAA was determined with a microtiter plate sandwich enzyme-linked immunosorbent assay using monoclonal antibodies specificfor SAA (Biosource Cytoscreen, Ratingen, Germany) and a Dyna-tech reader.

Erythropoietin

Plasma EPO was measured by radioimmunoassay, as described[13] using a rabbit antiserum against recombinant human EPOand iodinated human EPO as tracer (Amersham International,Amersham, UK). The geometric mean EPO level for healthy non-anemic adults is 17.9 mU/ml; 5±95% percentile, 11±31 mU/ml(n = 84).

Statistics

Data are presented as median and interquartile range. To test forchanges in serially measured protein levels, the general linearmodel repeated measure test, which is part of SPSS (version 7.5)was used. For comparison of two groups, the Mann±Whitney Utest and for three groups the Kruskal±Wallis test was applied. A pvalue of < 0.05 was considered significant. To assess the correlationof different APPs, Spearman's rank correlation test was used.

Results

Time course of APP after cardiac arrest

Figure 1 (upper panel) shows the time course of the threetype-1 APP a1-AGP, CRP and SAA. Following CPR, a1-AGP increased in all patients and was significantly ele-vated on days 2 and 3, when the increase was about two-fold. Similarly, the levels of CRP and SAA also rose inall patients, but in contrast to a1-AGP they were alreadyelevated within one day in 14 (CRP) and 17 (SAA) of 22patients. The mean increase of SAA and CRP after3 days was 35-fold and 28-fold, respectively. One patientwith hepatic cirrhosis (No. 5) showed the least increasein both CRP (1.9 mg/dl and 2.2 mg/dl on day 2 and day3, respectively), SAA (7.0 and 13.8 mg/dl on day 2and day 3, respectively) and a1-AGP (0.4 mg/dl and0.52 mg/dl on day 2 and day 3, respectively).

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Table 1 Characteristics of patients with primarily successful resus-citation after cardiac arrest. AMI acute myocardial infarction, VFventricular fibrillation, CPC cerebral performance categories:

1 = no or minor deficit, 2 = moderate disability, 3 = severe disabili-ty, 4 = vegetative state (see Subjects and methods and [12])

No. Sex Age(years)

Cause ofcardiac arrest

Collapse-to-first CPRattempt interval(min)

Infectious compli-cations within 7 daysof resuscitation

Outcome Highest bodytemperature within72 h of resuscitation(� C)

1 M 50 AMI + VF < 2 None CPC 1 38.52 M 37 AMI + VF 5±10 None CPC 3 38.33 M 75 VF 10±15 Pneumonia Died day 16 39.44 F 55 AMI + VF 2±3 None CPC 2 38.45 F 46 Respiratory arrest 10±15 Pneumonia CPC 4 39.46 M 43 AMI + VF < 2 Pneumonia CPC 1 38.77 M 63 AMI + VF 10±15 Pneumonia Died day 29 38.38 F 69 AMI < 2 Pneumonia CPC 1 38.79 F 67 AMI + VF < 2 Pneumonia + sepsis CPC 2 38.9

10 M 46 AMI + VF < 2 None CPC 1 38.311 M 56 VF 10±15 Pneumonia + sepsis Died day 32 38.712 M 72 AMI + VF < 2 None CPC 1 38.113 M 55 Aspiration 2±5 Pneumonia + sepsis Died day 4 40.414 F 71 VF < 2 None CPC 1 38.815 F 78 AMI + VF 10 Pneumonia CPC 4 38.516 F 65 AMI + VF < 2 None CPC 1 37.517 M 16 Respiratory arrest < 2 Pneumonia CPC 1 39.618 M 57 VF 10 Pneumonia CPC 4 39.319 M 55 AMI + VF < 2 Pneumonia CPC 2 37.520 F 33 AMI + VF < 2 Pneumonia CPC 1 38.421 M 38 AMI + VF 5±10 Pneumonia Died day 25 38.822 M 64 AMI + VF 5 None CPC 2 37.5

The kinetics of the two type-2 APP HAP and a1-ATare illustrated in the middle panel of Fig 1. Similarly tothe type-1 APP, the circulating levels of these proteinsincreased in all patients. The time course resembledthe increase of a1-AGP in that it did not occur beforeday 2 and the maximal response was about twofold onday 3 for both HAP and a1-AT. Again the patient withliver cirrhosis had the lowest response (a1-AT 127 mg/dl and HAP 43 mg/dl on day 3).

Figure 1 (lower panel) shows a downregulation ofTRA levels that occurred after resuscitation, the meanplasma level after 3 days being 29 % lower than the levelimmediately after hospital admission. In 4 patients, thelevel at admission was below the normal range(200±400 mg/dl). After 3 days, 19 out of the 22 patientsshowed levels below 200 mg/dl.

Time course of plasma EPO and hemoglobin levelsafter cardiac arrest

During the ICU stay following CPR, plasma EPO lev-els increased slightly from 23.5 (17.3±38.8) to 26.5(17.5±38.0) (day 1), 25.5 (22.3±44.8) (day 2) and 31.0(22.5±39.8) mU/ml (day 3), but this increase was not sta-tistically significant. At the same time, the hemoglobinconcentrations fell gradually from a median of 14.0(12.9±15.5) g/dl on admission to 12.8 (11.2±13.6) (day 1),11.6 (10.1±12.7) (day 2) and 10.7 (8.9±11.5) g/dl on day 3.

APP and EPO levels in subgroups of patients aftercardiac arrest

In order to assess possible determinants of the APP re-sponse, patients were subdivided into different groupsaccording to the collapse-to-first CPR attempt interval[12], the clinical course and final outcome. The levelsof APP on days 1, 2 and 3 were not different in patientsin whom the estimated collapse-to-first CPR attempt in-terval was below 2, between 2 and 5 and above 5 min(Fig 2). Plasma EPO levels were also not different inthe three groups (data not shown).

As expected, the percentage of patients survivingwithout or with only minor neurological deficit corre-lated inversely with the estimated collapse-to-first CPRattempt interval: none of the 11 patients with an esti-mated anoxic time of < 2 min, 1 out of the 3 patientswith an estimated anoxic time of 2±5 min and all 8 pa-tients with an estimated anoxic time of > 5 min devel-oped severe disability or a vegetative state.

As indicated in Table 2 there was, however, no differ-ence between the course of APP in patient groups withdifferent outcome, with the only exception being thatthe a1-AT level in nonsurvivors was slightly higher onday 2. Plasma EPO concentrations in patients surviving

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Fig. 1 Time course of the type-1 APP a1-acid glycoprotein a1-(AGP), C-reactive protein (CRP) and serum amyloid A (SAA) up-per panel, of the type-2 APP haptoglobin (HAP) and a1-antitryp-sin a1-(AT) middle panel and of the ªnegativeº APP transferrin(TRA) lower panel in patients after CPR. Values are medianswith the 25th and 75th percentiles. * denotes significant increasecompared with level on admission

with or without neurologic deficit were not different,but in nonsurvivors EPO levels were significantly higheron day 3 than in both groups of survivors. Hemoglobinlevels were not different between the three groups(data not shown).

Furthermore, there was no difference in APP orEPO levels in patients requiring vasopressor supportfor more than 24 h after cardiac arrest (n = 14) as com-pared to those who did not (data not shown). Patientswith cardiac arrest due to ventricular fibrillation and si-

multaneous myocardial infarction showed no differencein the course and magnitude of APP compared to thosewith ventricular fibrillation in the absence of myocardialinfarction (data not shown).

As illustrated in Fig 3 the levels of APP were also notsignificantly different in patients developing clinicallyapparent infectious complications after cardiac arrestas compared to those who did not. The median maximalbody temperature measured within the study period of3 days was 38.8 �C (38.5±39.4) in patients with apparent

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Fig. 2 Comparison of differentAPP at day 3 in patients withcardiopulmonary resuscitationafter different collapse-to-firstCPR attempt intervals. Valuesare medians and 75th percen-tiles; n = 11 (< 2 min), n = 3(2±5 min), n = 8 (> 5 min). Thelevels of APP were not differ-ent between the three groups

Table 2 Time course of APP and EPO in patients with different outcomes after cardiopulmonary resuscitation. Values are medians(25th±75 percentile) (CPC cerebral performance categories)

APP Died (n = 5) CPC 3 or 4 (n = 4) CPC 1 or 2 (n = 13) p

Day 0 Day 1 Day 2 Day 3 Day 0 Day 1 Day 2 Day 3 Day 0 Day 1 Day 2 Day 3

a1-AGP(g/l)

0.9(0.8±0.9)

0.9(0.9±1.0)

1.4(1.2±1.6)

1.7(1.5±2.0)

0.9(0.4±1.1)

0.7(0.5±0.9)

1.0(0.4±1.5)

1.2(0.7±1.8)

1.0(0.7±1.1)

1.0(0.7±1.1)

1.3(1.1±1.7)

1.6(1.3±2.0)

NS

CRP(mg/dl)

0.9(< 0.6±2.6)

6.2(3.0±14.2)

13.9(10.3±25.2)

17.5(9.0±29.1)

< 0.6(0.6±0.8)

0.9(0.7±4.9)

7.0(1.9±9.3)

12.2(4.4±18.1)

< 0(0.6±< 0.6)

3.2(1.0±6.3)

14.4(10.0±17.8)

17.4(9.7±22.7)

NS

SAA(mg/dl)

2.2(0.3±15.6)

18.2(13.3±36)

43.8(22±46.9)

47(36.9±50.5)

0.9(0.5±2.7)

5.0(2.3±21.6)

29.1(4.1±44.9)

34(19.3±50.3)

1.3(0.6±4.5)

12.5(10.4±35)

43.4(42±49.3)

47.7(41±58.8)

NS

HAP(mg/dl)

87(60±110)

96(37±115)

138(113±195)

203(157±310)

108(23±214)

77(18±200)

184(28±281)

177(68±244)

115(90±222)

103(86±177)

142(108±217)

226(151±346)

NS

a1-AT(mg/dl)

149(125±189)

176(142±194)

231(209±286)

290(246±366)

159(122±210)

139(114±200)

216(144±230)

266(158±284)

132(120±154)

148(139±164)

198(183±213)

255(226±277)

*

TRA(mg/dl)

212(182±256)

181(168±192)

163(145±193)

170(132±183)

200(151±261)

171(111±238)

156(148±201)

126(112±165)

219(196±282)

205(188±241)

170(157±217)

158(141±192)

NS

EPO(U/l)

19(18±89)

27(16±40)

69(26±151)

56(43±137)

36(18±39)

33(27±61)

24(22±35)

27(21±33)

21(16±38)

23(16±36)

25(22±44)

26(19±39)

**

* On day 2 the level of a1-AT was significantly different between non-survivors and patients with CPC 3 or 4** the EPO level was significantly different on day 3 between patients who died vs patients with CPC 3 or 4 and between patients whodied vs those with CPC 1 or 2 (p < 0.021 and p < 0.017, respectively)

infection and 38.3 �C (37.5±38.4) in patients without(p < 0.01). Excluding the patient with hepatic cirrhosis(No. 5) with blunted APP response, subgroup analysisdid not reveal different results.

Correlation of the levels of different APPs and plasmaEPO

To test if the magnitude of increase of different APP, thereduction in TRA and changes in plasma EPO were cor-related, Spearman's rank correlation test was applied.However, there was no statistically significant correla-tion between any of the parameters determined.

Time course of APP and plasma EPO in healthyvolunteers exposed to hypobaric hypoxia

As illustrated in Fig 4, exposure to hypobaric hypoxiafor 6 h did not induce any significant change of type-1APP, type-2 APP or transferrin within the 3 day studyperiod. In contrast to all APP evaluated, serum EPOlevels increased following hypoxic exposure of healthyvolunteers (Fig 4, lower panel). This increase reachedstatistical significance 240 min after the start of expo-sure to hypoxia and persisted during the period of simu-lated altitude exposure, before EPO levels declinedagain and were no longer different from baseline after24 h.

Discussion

The changes in the blood levels of APP in patients in-vestigated after primarily successful CPR (Fig 1) indi-cate that an acute phase response occurs regularly aftercardiac arrest. Several lines of evidence suggest thatthis response is directly associated with the arrest andthe restoration of circulation rather than with complica-tions arising subsequently.

First, the response of APP was rapid with increases inCRP already demonstrable in the second set of samplestaken on the morning of day 1 after resuscitation(Fig 1). Second, despite variable clinical courses, thechanges in APP followed similar kinetics in all patients.Third, the magnitude of change in APP levels was inde-pendent of the clinical course, and in particular not dif-ferent in patients with and without clinically apparentbacterial infection (Fig 3). One patient, whose APP re-sponse was blunted compared with that of the other pa-tients had hepatic cirrhosis, which is in accordance withtheassumptionthatAPParemainlyproducedbytheliver.

To our knowledge an APP response after cardiac ar-rest has so far not been systematically investigated, butit is in line with the concept of a systemic inflammatoryresponse reaction following global ischemia, called theªpostresuscitation syndromeº [2±5]. The combinationof global ischemia/anoxia, resuscitation measures andsubsequent reperfusion are believed to induce this noso-logic entity, which shares many similarities with the mul-tiple organ dysfunction syndrome [14]. In contrast to thefull picture of postresuscitation syndrome, which is asso-ciated with poor prognosis and generally does not occur

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Fig. 3 Comparison of differentAPP at day 3 after resuscitationin patients with and without in-fectious complications (n = 14and 8, respectively). Values aremedians and 75th percentiles.The levels of APP were not dif-ferent between the two groups

in patients with optimal recovery [5], the changes inAPP were found in all patients irrespective of courseand outcome (Table 2, Figs 2, 3). In particular, the acutephase response was also found in patients with a verybrief collapse-to-ROSC interval, no subsequent compli-cations and excellent outcome.

There were considerable differences in the ampli-tude of changes in APP levels, but we were unable tofind a coordinated regulation of different APP in indi-vidual patients. Patients with particularly high levels ofCRP did not necessarily have particularly high levels ofa1-AGP or SAA, the other two type-1 APP investigat-ed, and vice versa. The amplitude of changes of thetype-1 APP was also not correlated with the extent ofchanges of type-2 APP.

Proinflammatory cytokines are major stimuli for theproduction of APP, and previous work has shown thatIL 1 and IL 6 are elevated after cardiac arrest [5]. Wedid not measure inflammatory cytokines in the presentstudy, but the lack of coordination in the rise of differentAPP suggests that they are partly regulated indepen-dently despite common mechanisms of induction, suchas the release of certain cytokines.

It is tempting to speculate that the sequence of hy-poxia and subsequent reoxygenation directly contrib-utes to the systemic inflammatory response that is trig-gered by cardiac arrest. However, two lines of evidencefrom our studies suggest that tissue hypoxia is probablynot a predominant stimulus of APP production in vivo.First, the APP response was not related to the collapse-to-first CPR attempt interval (Fig 2), although this timeperiod had a significant impact on neurologic outcome,indicating that it is a reliable estimate of the initial peri-od of hypoxia. Second, in a complementary approachwe found that more prolonged nonischemic hypoxia ofa severity that can be safely applied to volunteers didnot induce any change in the concentrations of type-1and type-2 APP (Fig 4). Similarly, 4 days at 4350 m wasrecently also found to have no effect on CRP concentra-tions, although there was a progressive, up to 7.3-fold in-crease in IL-6 levels [15]. Clearly, altitude exposure andcardiac arrest differ markedly with respect to the type,duration and severity of hypoxia that both are associat-ed with. It is noteworthy, however, that, as in a previousstudy [10], the conditions of hypobaric hypoxia em-ployed in the present protocol were sufficient to triggera moderate, but significant, time-dependent increase inthe erythropoietic hormone EPO (Fig 4). EPO is oneof the most obvious examples of an increasing numberof proteins which have been found to be regulated by awidespread system of oxygen-dependent gene expres-sion [16,17]. This system operates over a broad range ofoxygen tensions [18] and involves the activation of hy-poxia-inducible transcription factors. On the basis of invitro findings, it has been suggested that these mecha-nisms also determine the production of APP [7]. How-ever, since hypoxic exposure of the volunteers in thepresent study increased EPO but not APP, the oxygensensitivity of APP production in vivo is at least signifi-cantly lower than that of EPO production.

Similar considerations for a direct effect of hypoxiaon type-1 and type-2 APP in vivo hold true for the ef-

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Fig. 4 Time course of type-1 APP upper panel, type-2 APP, serumtransferrin middle panel and serum EPO levels lower panel in hu-man volunteers exposed to hypobaric hypoxia (corresponding to4000 m) for 6 h. * denotes significant difference compared to base-line values

fects of hypoxia on TRA production. Although in vitroin human hepatoma cells TRA was recently found tobe induced by hypoxia through mechanisms dependingon the transcription factor ªhypoxia-inducible factor 1º[19], which also activates EPO production, hypoxic ex-posure did not stimulate TRA levels in vivo, and follow-ing cardiac arrest TRA decreased rather than increased(Fig 2). This is compatible with downregulation as partof the acute phase response [6].

Although it appears that hypoxia per se is not a pre-dominant stimulus for APP production in vivo, the sud-den increase in oxygen availability that occurs duringand after resuscitation may play an important role. Wehave recently shown that another component of the in-flammatory response, the upregulation of endothelialadhesion molecules, is stimulated by reoxygenation butnot during hypoxia [20]. If similar mechanisms, whichare probably related to the generation of reactive oxy-gen species, were responsible for the induction of APP,this could explain why the APP response was indepen-dent of the duration of the hypoxic insult. In addition,it is also possible that mechanisms only indirectly relat-ed to circulatory arrest, such as endotoxemia or bactere-mia due to a temporary loss of gut barrier function, con-tribute to the induction of APP after resuscitation.

Finally, our data also confirm that EPO does not be-have like an APP, as it has been suggested previously instudies of patients after surgery [21]. At day 1 after car-diac arrest there was no change in EPO levels, and thesubsequent slight, albeit statistically nonsignificant, in-

crease in EPO levels paralleled a reduction in hemoglo-bin levels, suggesting that it is at least in part due to a re-duction in oxygen-carrying capacity. In animal experi-ments, it has been shown that 10 min of normobaric hy-poxia may be sufficient to trigger EPO production [22],but since we have not analyzed EPO levels at short in-tervals after cardiac arrest, a temporary stimulation dur-ing the period of arrest may have been missed, given aplasma half-life of EPO of about 5 h [10]. Interestingly,serum EPO levels were significantly elevated in nonsur-vivors at days 2 and 3 (Table 2), although the hemoglo-bin concentration was not lower in these patients. Ele-vations of EPO that are unrelated to hemoglobin haverecently also been reported in septic patients with pooroutcome [23]. The mechanisms of this response, howev-er, are unclear.

In conclusion, we have demonstrated that consistentand marked changes in the levels of circulating APP oc-cur regularly after cardiac arrest, indicating a systemicinflammatory response to global ischemia/reperfusioninjury and resuscitation measures. The magnitude ofthis response is unrelated to the underlying cause of ar-rest, the presumptive period of circulatory arrest, com-plications and outcome and could not be mimicked byexposure to more prolonged but less severe hypoxia.Since the APP response was unrevealing with respectto the development of infectious complications, the di-agnostic consideration of APP in the postresucitationperiod may be misleading.

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