Prolonged recirculation is required to detect secondary ... - IOS Press

Clinical Hemorheology and Microcirculation 32 (2005) 1–12 IOS Press

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Prolonged recirculation is required to detect secondary metabolic and hemodynamic deterioration after superior mesenteric artery occlusion Michael Lauterbach a,b,∗ , Georg Horstick a,b , Nicola Plum a , Axel Heimann a , Dietmar Becker b , L.S. Weilemann b , Thomas Münzel b and Oliver Kempski a a

Institute for Neurosurgical Pathophysiology, Johannes Gutenberg-University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany b The 2nd Medical Clinic, Johannes Gutenberg-University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany Received 4 May 2004 Accepted 6 August 2004 Abstract. We evaluated late (4 hrs) effects of reperfusion on hemodynamics after 30 or 60 min occlusion of the superior mesenteric artery (SMA) in a rat model. Spontaneously breathing animals (n = 30) underwent occlusion of the SMA for 0 (sham), 30 (SMAO_30) or 60 min (SMAO_60) followed by reperfusion with normal saline. Abdominal blood flow (ABF), SMA blood flow (SBF), arterial blood pressure and heart rate were recorded continuously. Systemic vascular resistance (SVR) and SMA vascular resistance (MVR) were calculated at baseline and after 240 min reperfusion (240R). All animals survived in SMAO_30 and sham, two died in SMAO_60 after 120R. ABF remained constant in all groups. SVR increased in SMAO_30 and sham and decreased in SMAO_60 at 240R. SBF was significantly lower after reperfusion in ischemia groups as compared to sham. After 120R, SBF had increased significantly in SMAO_60 versus SMAO_30. MVR increased significantly in SMAO_30 but not in SMAO_60 and sham at 240R. 60 minutes SMA occlusion revealed early hemodynamic changes of septic circulation with increased blood flow in the SMA, decreased SVR, and pseudo-normalization of MVR. Prolonged observation periods are required to detect these significant changes which are overlooked when only studying 120 minutes of reperfusion as usually done.

1. Introduction Acute intestinal ischemia is a serious clinical problem with a high mortality (50–70%) [1,2]. Nonocclusive mesenteric ischemia in patients undergoing complicated cardiovascular surgery [3] or traumatic injury of the mesenteric arteries [1] are typical clinical examples. Furthermore, reduced splanchnic perfusion during and after hemorrhagic shock leads to gut injury [4]. These mechanisms result in * Corresponding author: Dr. M. Lauterbach, The 2nd Medical Clinic, Langenbeckstr. 1, 55131 Mainz, Germany. Tel.: +49 (0) 6131172741; Fax: +49 (0) 6131 176605; E-mail: [email protected].

1386-0291/05/$17.00  2005 – IOS Press and the authors. All rights reserved

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M. Lauterbach et al. / Detection of secondary metabolic and hemodynamic deterioration

inflammatory response following reestablishment of an adequate systemic perfusion and also to ‘collateral damages’ in various remote organs [5]. Well known in this context are the injury to the lung after gut ischemia and a suppressed bone marrow proliferation rendering the organism more susceptible to infections [4]. Reactive hyperemia is usually not seen during early reperfusion after ischemia (occlusion of the superior mesenteric artery) [6]. However, little is known about the late (4 hrs) effects of reperfusion on regional and systemic hemodynamics, metabolism and outcome. The significance of the duration of ischemia as well as reperfusion time on regional and systemic hemodynamics is not sufficiently explored. In current animal models of intestinal ischemia, superior mesenteric artery clamping time varies from 15 to 150 minutes, and reperfusion time with hemodynamic monitoring usually is not longer than one or two hours [7,8]. The severity of damage correlates with the duration of ischemia [9] and might also influence the efficiency of therapeutic agents. Additionally, initial therapeutic benefit might disappear with prolonged observation times. Our study evaluated the late (4 hrs) effects of reperfusion on regional and systemic hemodynamics and outcome after different occlusion periods of the superior mesenteric artery. Particular attention was paid to systemic blood flow, superior mesenteric artery blood flow, and changes in blood gases. The ischemia was set according to a previously published rat model [10,11] for 30 and 60 minutes followed by four hours of reperfusion. 2. Materials and methods Thirty male Sprague-Dawley rats (body wt. 350 ± 10 g) were maintained on a standard rat chow and water ad libitum before the experiment. After anesthesia with urethane (1.25 g/kg body wt. IM, single dose) the carotid artery and jugular vein were cannulated with a small PE-tube for arterial blood pressure recording, arterial blood gas analysis and volume substitution during the experiment via the central venous line. For each sample 250 µl of blood were drawn into heparinized syringes. Arterial blood gases (PaO2 , pH, SBE), hemoglobin concentration (HB) and hematocrit (Hct) as well as lactate, potassium and sodium were analyzed with Arterial Blood Gas Laboratory Radiometer Copenhagen 615. After a median laparatomy Doppler flow probes were placed around the abdominal aorta and the SMA near its origin from the abdominal aorta without impairing the normal blood flow in these vessels. A snare with a monofilament suture was placed around the SMA proximal to the flow probe for registration of complete occlusion and reestablishment of perfusion after release of the snare. The abdominal wound was covered with oxygen-impermeable plastic foil (Folio, Germany) to prevent evaporative loss of water [10]. Hemodynamic data [systolic, mean and diastolic arterial blood pressure (SAP, MAP, DAP), heart rate (HR), abdominal blood flow and superior mesenteric artery blood flow] R were recorded using System 6 (Triton Technology, Inc., San Diego, CA, USA), digitized and regisR tered on a beat-to-beat basis with a computer-based system (Dasylab , National Instruments Corporation, Austin, TX, USA). Stroke volume was calculated real-time from the area under the curve of the registered pulsatile velocity curves (abdominal stroke volume – ASV). Blood flow was calculated by multiplication of stroke volume and the simultaneously registered heart rate (abdominal blood flow – ABF, mesenteric blood flow – MBF). The Doppler flow values showed a linear correlation to electromagnetic blood flow sensors (Skalar-Medical b.v., Delft, The Netherlands) as previously described [10]. Central venous pressure (CVP) was obtained at baseline and at the end of the experiments for calculation of systemic (SVR) and superior mesenteric artery vascular resistance (MVR). SVR and MVR were calculated using the Bernoulli equation (SVR = 80(MAP−CVP)/ABF, MVR = 80(MAP−CVP)/MBF).


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An ECG in lead II was recorded during the entire duration of the experiment to detect morphological ECG-changes. The animals were placed on a heating pad and the rectal temperature was kept constant at 37.5 ± 0.5◦ C by means of a feedback controlled heating unit (Homeothermic Blanket Control, Harvard, South Natick, MA, USA). Rats received a basal infusion with albumin in physiological saline (0.3 ml/100 g body wt./h) for compensation of intra-operative albumin loss and evaporative water loss as previously published [11]. All investigative procedures and the animal facilities conformed to the Guide of Care and Use of Laboratory Animals published by the US National Institutes of Health. The regional animal care and use committee approved the protocol. 2.1. Experimental protocol Animals were randomized into three groups (10 animals per group): In SMAO_30 the SMA was occluded by tightening the snare for 30 minutes, in SMAO_60 the SMA was occluded for 60 minutes. Animals, who underwent the same operative procedure without tightening of the snare served as sham. Follow-up of SMAO_30 and SMAO_60 was four hours after release of the snare. Shams were followedup for a total of five hours. Mean arterial blood pressure was kept above 70 mm Hg by infusion of normal (0.9%) saline by central venous line infusion [10,11]. At baseline – prior to the occlusion – of the SMA, and at the beginning of reperfusion and 30, 60, 120, 180, 240 minutes of reperfusion (R, 30R, 60R, 120R, 180R, and 240R) arterial blood gases (ABG) were drawn in each group. Due to mechanical compression of the SMA by the Doppler flow probe in the eventration of the gut, parallel intravital microscopy of the mesenteric microcirculation could not be performed. After termination of the experiments, the SMA was dissected and morphologically analyzed to ensure the integrity of the vessel at the former position of the occluding snare. 2.2. Statistical analysis R (SPSS SciData are presented as means ± SEM. Statistical analysis was performed with Sigma Stat ence Inc., Chicago, IL). Statistical significance of changes from baseline values within each group was tested with analysis of variance (ANOVA) for repeated measures. For non-parametric values, an ANOVA on ranks was applied. Differences between groups were statistically analyzed by one way ANOVA comparing several groups. Likewise, ANOVA for non-parametric values was used (Kruskalis–Wallis test) with multiple comparison method (Student–Newman–Keuls test) where applicable. Statistical significance was accepted at p < 0.05.

3. Results Sham group animals remained at baseline levels in terms of all measured parameters throughout the whole observation period, as likewise previously published [11]. To increase readability, data from sham animals are provided in the figures as grey-colored line and scatter plots without highlighting statistically significant differences.

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3.1. Outcome All sham-operated animals and all animals from SMAO_30 (30 minutes SMA occlusion) survived until 240R. Two animals of SMAO_60 (60 minutes SMA occlusion) died prior to the end of the experiments, one at 165 minutes of reperfusion, the second at 180 minutes of reperfusion. Hemodynamics and blood gas values from non-survivors are included in the data. 3.2. Hemodynamics 3.2.1. Mean arterial blood pressure (MAP), central venous pressure (CVP), heart rate (HR) MAP was significantly lower at 60 minutes of reperfusion (60R) in SMAO_60 compared to SMAO_30 and controls (Fig. 1). From 120R to 240R, MAP remained on a similar level in ischemia groups (SMAO_30, SMAO_60). Decrease of MAP in SMAO_60 was significant from R to 30R, whereas the decrease in MAP in SMAO_30 was not significant in the first 30 minutes of reperfusion. 60 minutes of SMA occlusion led to a more pronounced decrease in MAP, which was most prominent 60 minutes after reopening of the SMA. MAP did not decrease below 70 mm Hg in all groups as intended in the study. CVP was comparable in all groups at baseline (SMAO_30: 1.8 ± 0.3 mm Hg, SMAO_60: 1.6 ± 0.4 mm Hg, sham: 1.7 ± 0.7 mm Hg), and 240R (SMAO_30: 2.4 ± 0.6 mm Hg, SMAO_60: 2.5 ± 0.4 mm Hg, sham: 2.2 ± 0.8 mm Hg). Heart rate increased in SMAO_30 and SMAO_60 till the end of the experiment without significant differences between SMAO_30 and SMAO_60 (Fig. 2). Morphological ECG changes were not seen. (Baseline; SMAO_30: 392 ± 12 bpm, SMAO_60: 389 ± 9 bpm, sham: 399 ± 12 bpm.) There was a trend towards higher heart rates in SMAO_60 (R; SMAO_30: 401 ± 15 bpm, SMAO_60: 425 ± 6 bpm, sham: 412 ± 13 bpm; 240R; SMAO_30: 489 ± 11 bpm, SMAO_60: 500 ± 9 bpm, sham: 427 ± 9 bpm, sham vs. SMAO_30, SMAO_60: p < 0.05).

Fig. 1. Mean arterial blood pressure (MAP). Data are presented as means ± SEM. SMAO_30: ischemia duration 30 min; SMAO_60: ischemia duration 60 min; Occ.: superior mesenteric artery occlusion; R, 30R, 60R, 120R, 180R, 240R: reperfusion (number indicates min); first arrow: Occ. + 10 min; second arrow: 10 min before R. ∗ Significant difference between the two groups, p < 0.05.


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Fig. 2. Heart rate (HR). Data are presented as means ± SEM. SMAO_30: ischemia duration 30 min; SMAO_60: ischemia duration 60 min; Occ.: superior mesenteric artery occlusion; R, 30R, 60R, 120R, 180R, 240R: reperfusion (number indicates min); first arrow: Occ. + 10 min; second arrow: 10 min before R.

Fig. 3. Abdominal stroke volume (ASV). Data are presented as means ± SEM. SMAO_30: ischemia duration 30 min; SMAO_60: ischemia duration 60 min; Occ.: superior mesenteric artery occlusion; R, 30R, 60R, 120R, 180R, 240R: reperfusion (number indicates min); first arrow: Occ. + 10 min, second arrow: 10 min before R. ∗ Significant difference between the two groups, p < 0.05.

3.2.2. Abdominal stroke volume (ASV), abdominal blood flow (ABF), systemic vascular resistance (SVR) The abdominal stroke volume significantly decreased in SMAO_60 (101 ± 13 µl) until R compared to SMAO_30 (134 ± 7 µl) and shams (149 ± 4 µl) (Fig. 3). The ASV remained lower in SMAO_60 compared to SMAO_30 until 180R without statistical significance between ischemia groups (sham vs.

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Fig. 4. Abdominal blood flow (ABF). Data are presented as means ± SEM. SMAO_30: ischemia duration 30 min; SMAO_60: ischemia duration 60 min; Occ.: superior mesenteric artery occlusion; R, 30R, 60R, 120R, 180R, 240R: reperfusion (number indicates min); first arrow: Occ. + 10 min; second arrow: 10 min before R. Table 1 Systemic vascular resistance (SVR) at baseline and 240R SVR [kPa sec/l] SMAO_30 SMAO_60 Sham

Baseline 14.3 ± 1.6 13.5 ± 1.3 12.5 ± 0.7

240R 15.1 ± 1.8 11.7 ± 0.3 13.2 ± 0.7

SMAO_60, p < 0.05). The decrease of the abdominal blood flow was less pronounced than abdominal stroke volume due to higher heart rates in SMAO_60 (Fig. 4). At 240R, ABF reached baseline values. There was trend towards a lower SVR in SMAO_60 compared to SMAO_30 and controls at 240R (see Table 1). 3.2.3. Superior mesenteric artery blood flow (SBF), superior mesenteric artery vascular resistance (MVR) Blood flow in the superior mesenteric artery did not differ between ischemia groups until 120R (Fig. 5). Reactive hyperemia was not observed after SMA occlusion. From 120R until end of observation, SBF remained reduced in SMAO_30 and was significantly less decreased in SMAO_60 versus SMAO_30. SBF in SMAO_60 progressively increased and had returned to baseline values at 240R (SMAO_30: 8.8 ± 1.3 ml/min, SMAO_60: 13.2 ± 1.2 ml/min, sham: 15.2 ± 1.1 ml/min, SMAO_60, sham vs. SMAO_30, p < 0.05). MVR was significantly higher in SMAO_30 compared to SMAO_60 and controls at 240R (p < 0.05) (see Table 2). 3.3. Arterial blood gas values Blood pH did not differ between the groups at the beginning of the experiments (SMAO_30: 7.36 ± 0.03, SMAO_60: 7.37 ± 0.04, and sham: 7.38 ± 0.01). In both ischemia groups, there was a significant


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Fig. 5. Superior mesenteric artery blood flow (SBF). Data are presented as means±SEM. SMAO_30: ischemia duration 30 min; SMAO_60: ischemia duration 60 min; Occ.: superior mesenteric artery occlusion; R, 30R, 60R, 120R, 180R, 240R: reperfusion (number indicates min); first arrow: Occ. + 10 min; second arrow: 10 min before R. ∗ Significant difference between the two groups, p < 0.05. Table 2 Superior mesenteric artery vascular resistance (MVR) at baseline and 240R MVR [kPa sec/l] Baseline SMAO_30 64.0 ± 4.8 SMAO_60 58.9 ± 2.0 Sham 60.8 ± 6.0 ∗ p < 0.05 SMAO_30 vs. SMAO_60, sham.

240R 77.4 ± 6.0∗ 50.8 ± 3.8 49.3 ± 3.3

decrease of blood pH after R (SMAO_30: 7.36 ± 0.03, SMAO_60: 7.38 ± 0.05, sham: 7.37 ± 0.02) until the end of the experiments (SMAO_30: 7.31 ± 0.02, SMAO_60: 7.29 ± 0.02, sham: 7.38 ± 0.01). Values between the two ischemia groups were not statistically significant (sham vs. SMAO_30, SMAO_60, p < 0.05). PaCO2 did not differ between the groups at the beginning of the experiments (SMAO_30: 42.6 ± 1.5 mm Hg, SMAO_60: 41.7 ± 1.2 mm Hg, sham: 40.9 ± 0.6 mm Hg), or at R (SMAO_30: 41.4 ± 1.5 mm Hg, SMAO_60: 40.5 ± 1.5 mm Hg, sham: 41.3 ± 1.5 mm Hg). PaCO2 decreased significantly in SMAO_30 and SMAO_60 at the beginning of reperfusion. PaCO2 was significantly lower in SMAO_60 compared to SMAO_30 and controls at 120R (SMAO_30: 33.6 ± 1.3 mm Hg, SMAO_60: 28.0 ± 1.7 mm Hg, sham: 38.1±0.9 mm Hg, sham, SMAO_30 vs. SMAO_60, p < 0.05) and 240R (SMAO_30: 32.3 ± 1.1 mm Hg, SMAO_60: 27.4 ± 1.7 mm Hg, sham: 36.7 ± 0.8 mm Hg, SMAO_60 vs. SMAO_30, shams p < 0.05). Base-excess values mirrored paCO2 values (Fig. 6). PaO2 values were neither different at baseline (SMAO_30: 71.1 ± 2.4 mm Hg, SMAO_60: 69.0 ± 1.9 mm Hg, sham: 73.3 ± 1.3 mm Hg) nor at R (SMAO_30: 76.2 ± 2.4 mm Hg, SMAO_60: 79.3 ± 1.1 mm Hg, sham: 77.9 ± 4.0 mm Hg), although paO2 significantly increased from baseline to R in SMAO_60. Until the end of the experiments, paO2 increased to 85.0 ± 7.5 mm Hg in SMAO_30 and

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Fig. 6. Base excess (BE). Data are presented as means ± SEM. SMAO_30: ischemia duration 30 min; SMAO_60: ischemia duration 60 min; Occ.: superior mesenteric artery occlusion; R, 30R, 60R, 120R, 180R, 240R: reperfusion (number indicates min). ∗ Significant difference between the two groups, p < 0.05.

96.8±6.4 mm Hg in SMAO_60, which was not significantly different, and remained at 75.5±1.4 mm Hg in controls (sham vs. SMAO_60, p < 0.05). Hematocrit was comparable at baseline (SMAO_30: 45.6 ± 1.3%, SMAO_60: 44.5 ± 1.3%, sham: 45.4 ± 1.5%), R and 30R. Due to the larger volume substituted in SMAO_60, hematocrit decreased significantly in this group until the end of the experiments (SMAO_60: 37.1 ± 1.4%). 3.4. Electrolytes, metabolites Lactate levels did not differ at baseline (SMAO_30: 1.6 ± 0.1 mmol/l, SMAO_60: 1.7 ± 0.1 mmol/l, sham: 1.8±1.8 mmol/l) or at R (SMAO_30: 1.6±0.1 mmol/l, SMAO_60: 1.7±0.1 mmol/l, sham: 1.3± 0.1 mmol/l). There was no increase in lactate levels at 30R (SMAO_30: 1.5 ± 0.2 mmol/l, SMAO_60: 1.8 ± 0.2 mmol/l, sham: 1.4 ± 0.2 mmol/l), but a significant decrease until 240R (SMAO_30: 0.8 ± 0.1 mmol/l, SMAO_60: 0.9 ± 0.1 mmol/l, sham: 0.5 ± 0.0 mmol/l, sham vs. SMAO_30, SMAO_60, p < 0.05). Serum potassium levels were 4.0 ± 0.1 mmol/l in SMAO_30, 3.9 ± 0.2 mmol/l in SMAO_60, and 4.1 ± 0.2 mmol/l in controls. Potassium levels increased to 4.7 ± 0.2 mmol/l (SMAO_30), 4.6 ± 0.2 mmol/l (SMAO_60), and 4.4 ± 0.1 mmol/l (sham) until the end of observation. There was no statistically significant difference between the groups. Sodium levels significantly increased from 139.0 ± 0.7 mmol/l (SMAO_30), 139.0 ± 1.7 mmol/l (SMAO_60), and 138.8 ± 1.0 mmol/l (sham) at baseline to 143.4 ± 0.8 mmol/l (SMAO_30), 143.5 ± 1.9 mmol/l (SMAO_60), and 142±0.5 mmol/l in controls at 240R. There were no statistically significant differences between the groups. Serum chloride levels likewise increased during reperfusion, with most prominent changes in SMAO_60. [R: 104.6±0.8 mmol/l (SMAO_30), 104.1±1.4 mmol/l (SMAO_60), 104.6 ± 1.7 mmol/l; 240R: 117.4 ± 1.7 mmol/l (SMAO_30), 121.2 ± 2.1 mmol/l (SMAO_60), and 109.6 ± 0.7 mmol/l (sham).]


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3.5. Volume substitution Sodium chloride 0.9% was infused to maintain MAP above 70 mm Hg. The infused volume during reperfusion was significantly larger in SMAO_60 (59.5 ± 13.7 ml) compared to SMAO_30 (30.9 ± 5.3 ml) (p < 0.05). Except for basal infusion, no additional volume replacement was needed in controls. 4. Discussion Our results clearly demonstrate the need for a superior mesenteric artery occlusion time of at least 60 minutes and prolonged observation (>120 minutes of reperfusion) to determine the early hemodynamic changes of ‘sepsis-like’ circulation in superior mesenteric artery ischemia and reperfusion. 4.1. Hemodynamics The abdominal stroke volume decreased significantly in SMAO_60 (60 minutes ischemia) compared to SMAO_30 (30 minutes ischemia) in the late occlusion period. Likewise, abdominal blood flow decreased in SMAO_60 in the late occlusion period. However, there was no significant difference between the groups due to higher heart rates in SMAO_60 as seen in other studies [12,13]. Animals were unaltered in hemodynamics and metabolism during ischemia except for the decrease of abdominal stroke volume in the late occlusion period. Similar observations have been made after clamping of other organs such as hepatic vascular exclusion in major liver resection [13]. SMA blood flow was significantly decreased in ischemia groups in early reperfusion compared to baseline values. Reactive hyperemia was not seen confirming an earlier pig study [14]. The presented hemodynamic data are in good agreement with previously published hemorrhagic shock experiments, although hemorrhagic shock creates a low-flow situation in the mesenteric microcirculation as compared to the no-flow situation in complete intestinal ischemia [15,16]. Under hemorrhagic shock with fluid resuscitation neither reactive hyperperfusion of global hemodynamics nor reactive hyperemia in the mesenteric microcirculation had been detected [16]. The reduced SMA blood flow in early reperfusion that has been reported under low-flow and no-flow conditions can be explained by a reduction of functional capillary density in the mesenteric microcirculation after ischemia and reperfusion [17]. Reduction of a relevant number of microvessels might increase the total resistance in the reperfused vascular bed and thereby reduce blood flow in the afferent vessels [18]. However, this does not explain the hemodynamic changes observed in late reperfusion in our experiments. SMA blood flow returned to baseline values in SMAO_60 (60 minutes ischemia) during late reperfusion. There was a trend to a lower SVR in SMAO_60 compared to SMAO_30 and controls, and a ‘normalization’ of MVR to the values calculated for controls in SMAO_60 compared to SMAO_30 at the end of the experiments. Since there were no significant changes in pressures in the Bernoulli equation, the ‘normalization’ of MVR solely reflects the increase in SBF at the end of the experiments and might therefore be termed ‘pseudo-normalization’. The observed hemodynamic pattern in late reperfusion might reflect an increased shunting in the mesentery, because most of the microvessels remain occluded after reperfusion [19–22]. Own pilot data from SMA occlusion experiments with intravital microscopy of the mesentery additionally support the hypothesis of shunts in the mesenteric microcirculation (unpublished). Further, the rise of SMA blood flow in SMAO_60 might be an increased circulatory demand of the inflamed tissue. In systemic circulation, functional shunts and increased circulatory demand are conditions most commonly observed in systemic inflammatory response syndrome and septic

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circulation [23,24]. These inflammatory states usually lead to a reduction in microcirculatory perfusion, that may even be more pronounced during systemic hypotension [25,26]. We hypothesize that a local inflammatory response triggers the ‘normalization’ of SMA blood flow and superior mesenteric artery vascular resistance [27]. A local inflammatory reaction, if covering a large proportion of the gut, leads to systemic inflammatory response syndrome with spillover of inflammatory mediators into systemic circulation [28]. The trend to a decrease in systemic vascular resistance in SMAO_60 calculated at 240R might be an early symptom of systemic inflammatory response in our study. The significant changes in SBF observed in SMAO_60 (60 minutes ischemia) at 240R are probably indicative of a more severe reperfusion injury, as they did not occur in SMAO_30 (30 minutes ischemia). Intravital microscopy studies are necessary for confirmation. Only experiments lasting long enough (240 minutes reperfusion) can provide the required data. Such studies are not available so far. 4.2. Fluid replacement SMAO_60 (60 minutes ischemia) required a significantly larger resuscitation volume as a sign of enhanced tissue injury with subsequent fluid loss (capillary leakage) [29]. Clinical studies have shown that a requirement of large volumes of crystalloid solutions in shock was associated with increased mortality [30]. However, normal saline is not considered an optimal fluid replacement in massive hemorrhagic shock [31], an issue which is still under discussion [32–34]. Hyperchloremic metabolic acidosis is a potential risk with large-volume isotonic saline resuscitation. In our study serum chloride levels increased together with the sodium levels, but exceeded physiological ranges only in SMAO_60 (60 minutes ischemia) with large volume replacement (mean: 170 ml/kg body weight). Other investigators likewise reported hyperchloremia; although metabolic acidosis caused by normal saline infusion is rare [32]. 4.3. Metabolism Lactate, as a biochemical marker for gut ischemia is not a reliable parameter due to several other possible causes of elevated lactate levels [35]. Lactate serum levels did not change in our experiments. The rapid metabolism of lactate might be one reason for the lack of increase in lactate levels during and after ischemia. Studies of other groups have shown that lactate levels have no prognostic value in predicting survival [36]. Arterial base excess is the only parameter, which correlates to the duration of ischemia in the current study. However, metabolic acidosis of any origin increases arterial base excess. As there is a good correlation between intestinal ischemia time and mortality [37,38], other markers for determination of ischemic gut damage and predicting survival have to be studied, i.e. I-FABP (intestinal fatty acid binding protein) [39,40]. 5. Conclusion In conclusion, 60 minutes of ischemia followed by reperfusion with normal saline led to a significant hemodynamic and metabolic deterioration with metabolic acidosis and need for large volume fluid replacement. Our experiments revealed hemodynamic changes of early local ‘sepsis-like’ circulation with an increased blood flow in the SMA and the need for aggressive fluid resuscitation after 60 minutes in contrast to 30 minutes of SMA ischemia followed by reperfusion. The delayed onset of these changes clearly showed that prolonged observation periods (>120 minutes) are required to study the hemodynamic sequelae of bowel ischemia.


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Acknowledgements We thank Mr. Kopacz and Mr. Malzahn for their excellent technical assistance. This work was supported by the University of Mainz (MAIFOR). The manuscript includes data from the doctoral thesis of Nicola Plum.

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