Indicator Of Tissue Perfusion Research Articles

Comparison between central venous oxygen saturation and tissue oxygenation and perfusion: a case report

Comparison between central venous oxygen saturation and tissue oxygenation and perfusion: a case report

Martin Kelly, F.R.C.S., 1965 to 2008

Martin Kelly, F.R.C.S., 1965 to 2008

Central venous-to-arterial carbon dioxide difference: an additional target for goal-directed therapy in septic shock?

To test the hypothesis that, in resuscitated septic shock patients, central venous-to-arterial carbon dioxide difference [P(cv-a)CO(2)] may serve as a global index of tissue perfusion when the central venous oxygen saturation (ScvO(2)) goal value has already been reached. Prospective observational study. A 22-bed intensive care unit (ICU). After early resuscitation in the emergency unit, 50 consecutive septic shock patients with ScvO(2) > 70% were included immediately after their admission into the ICU (T0). Patients were separated in Low P(cv-a)CO(2) group (Low gap; n = 26) and High P(cv-a)CO(2) group (High gap; n = 24) according to a threshold of 6 mmHg at T0. Measurements were performed every 6 h over 12 h (T0, T6, T12). At T0, there was a significant difference between Low gap patients and High gap patients for cardiac index (CI) (4.3 +/- 1.6 vs. 2.7 +/- 0.8 l/min/m(2), P < 0.0001) but not for ScvO(2) values (78 +/- 5 vs. 75 +/- 5%, P = 0.07). From T0 to T12, the clearance of lactate was significantly larger for the Low gap group than for the High gap group (P < 0.05) as well as the decrease of SOFA score at T24 (P < 0.01). At T0, T6 and T12, CI and P(cv-a)CO(2) values were inversely correlated (P < 0.0001). In ICU-resuscitated patients, targeting only ScvO(2) may not be sufficient to guide therapy. When the 70% ScvO(2) goal-value is reached, the presence of a P(cv-a)CO(2) larger than 6 mmHg might be a useful tool to identify patients who still remain inadequately resuscitated.

Relationship between the sublingual microcirculation and lactate levels in patients with heart failure

Treatment of patients with severe heart failure aims at normalizing hemodynamic and metabolic parameters. We tested whether the sublingual functional capillary density (FCD), an indicator of tissue perfusion at the microvascular level, correlates with lactate levels in heart failure patients.

Haemodynamic changes during positive‐pressure ventilation in children

Positive-pressure ventilation may alter cardiac function. Our objective was to determine with the use of impedance cardiography (ICG) whether altering airway pressure modifies the central haemodynamics in mechanically ventilated children with no pulmonary pathology. Central venous saturation (S(cv)O(2)) was measured as an indicator of tissue perfusion. Twelve children between 7 and 65 months of age, requiring mechanical ventilation as a consequence of a non-pulmonary disease, were enrolled in the study. All patients had a central venous line as a part of their routine management. Using pressure controlled ventilation (PCV) the baseline PEEP value of 5 cmH(2)O (P(b5)) was increased to 10 cmH(2)O (P(i10)) and then to 15 cmH(2)O (P(i15)). After P(i15), PEEP was decreased to 10 (P(d10)) and then to 5 cmH(2)O (P(d5)). Each time period lasted 5 min heart rate (HR), mean arterial blood pressure (MABP), central venous pressure (CVP), end-tidal carbon dioxide (ETCO(2)), mean airway pressure (P(aw)), stroke volume index (SVI), cardiac index (CI) and central venous oxygen saturation (S(cv)O(2)) were recorded at the end of the five periods. The values of CI did not change when 10 and 15 cmH(2)O of PEEP were applied. Elevation of PEEP and thus P(aw) caused slight but not significant reductions in SVI and S(cv)O(2) as compared to the baseline (T(b5)). After reducing PEEP in T(d5) we found statistically significant elevations of SVI and CI, as compared to T(i15) heart rate, ETCO(2) and MABP remained unchanged. We did not find significant haemodynamic changes following PEEP elevation in ventilated children, as measured using impedance cardiography. Reducing the value of PEEP to 5 cmH(2)O resulted in statistically significant SVI elevations. The values of S(cv)O(2) remained unaffected.

Analysis of Combined Treatment of Embolic Stroke in Rat with r-tPA and a GPIIb/IIIa Inhibitor

Suppression of platelet activation improves the efficacy of thrombolytic therapy for stroke. Thus, combination treatment with recombinant tissue plasminogen activator (r-tPA) and 7E3 F(ab')2, a GPIIb/IIIa inhibitor that binds the platelet to fibrin, may improve the efficacy of thrombolytic therapy in embolic stroke. Magnetic resonance imaging (MRI) was used to monitor treatment response in rats subjected to embolic middle cerebral artery (MCA) occlusion (MCAo). Animals were randomized into treated (n=12) and control (n=10) groups and received intravenous combination therapy or saline, respectively, 4 hours after MCAo. Magnetic resonance imaging (MRI) measurements performed 1 hour after MCAo showed no difference between groups. However, an increased incidence (50%) of MCA recanalization was found in the treated group at 24 hours compared with 20% in the control group. The area of low cerebral blood flow at 24 and 48 hours was significantly smaller in the combination treatment group, and the lesion size, as indicated from the T2 and T1 maps, differed significantly between groups. Fluorescence microscopy measurements of cerebral microvessels perfused with fluorescein isothiocyanate-dextran and measurements of infarct volume revealed that the combination treatment significantly increased microvascular patency and reduced infarct volume, respectively, compared with the control rats. The efficacy of combination treatment 4 hours after ischemia is reflected by MRI indices of tissue perfusion, MCA recanalization, and reduction of lesion volume. The treatment also reduced secondary microvascular perfusion deficits.

Contrast-enhanced ultrasound assessment of tissue response to high-intensity focused ultrasound

We report the use of contrast-enhanced ultrasonography as an immediate means of assessing the clinical response to high-intensity focused ultrasound (US) or HIFU treatment of liver tumours. HIFU is a noninvasive transcutaneous technique for the ablation of tumours that has been shown to destroy tumour vasculature, as well as to cause coagulative necrosis of tumour cells. As a dynamic indicator of tissue perfusion, microbubble contrast agents have already been reported to increase the diagnostic sensitivity of ultrasonography in the detection of liver tumours. This report documents the ability of one IV microbubble contrast agent (SonoVue®, Bracco, Italy) to delineate the extent of HIFU ablation by comparison of pre- and immediately posttreatment perfusion within the target tumour. Observed changes were seen to correlate well with the ablated volume on histologic evaluation of the treated volume. This is the first time that this imaging technique has been reported in this setting. (E-mail: hifus.oxford@utlltd.com)

Quantification of perfusion using bolus tracking magnetic resonance imaging in stroke: assumptions, limitations, and potential implications for clinical use.

MR techniques have been very powerful in providing indicators of tissue perfusion, particularly in studies of cerebral ischemia. There is considerable interest in performing absolute perfusion measurements, with the aim of improving the characterization of tissue "at risk" of stroke. However, some important caveats relating to absolute measurements need to be taken into account. The purpose of this article is to discuss some of the issues involved and the potential implications for absolute cerebral blood flow measurements in clinical use. In bolus tracking MRI, deconvolution of the concentration-time course can in theory provide accurate quantification. However, there are several important assumptions in the tracer kinetic model used, some of which may be invalid in cerebral ischemia. These can introduce significant errors in perfusion quantification. Although we believe that bolus tracking MRI is a powerful technique for the evaluation of perfusion in cerebral ischemia, interpretation of perfusion maps requires caution; this is particularly true when absolute quantification is attempted. Work is currently under way in a number of centers to address these problems, and with appropriate modeling they may be overcome in the future. In the interim, we believe that it is necessary for users of bolus tracking perfusion data to be aware of the current technical limitations if they are to avoid misinterpretation or overinterpretation of their findings.

NIRS Measurement of Peripheral Fractional Oxygen Extraction (FOE) after Cardiopulmonary Bypass

Objectives: To compare peripheral fractional oxygen extraction (FOE), as measured by near infrared spectroscopy (NIRS), with conventional indicators of tissue perfusion in haemodynamically stable and unstable children after cardiopulmonary bypass. Design: Observational study. Setting: Paediatric Intensive Care Unit of a large teaching hospital. Patients: 17 children immediately after cardiopulmonary bypass. Male : female = 9 : 8, median age 7 months (range, newborn to 16 years). Methods: On admission, children were classified as “stable” or “unstable” based on the haemodynamic support they needed. Peripheral venous oxyhaemoglobin saturation (SvO2) was measured non‐invasively using NIRS with venous occlusion. FOE was calculated from SvO2 and arterial saturation measured by pulse oximetry. Repeated measurements of peripheral SvO2 were made for up to 8 hours. In 5 children who had pulmonary artery catheters, simultaneous mixed SvO2 measurements were recorded. Results: Median FOE was 7.9% higher in the unstable group than in the stable group (p = 0.013). Peripheral SvO2 and mixed SvO2 were correlated (R2 = 0.65, p &lt; 0.0001). Conclusions: Peripheral FOE is higher in unstable children. Changes in peripheral SvO2 are related to changes in mixed SvO2. These measurements may provide useful information about haemodynamic status in critically ill children. Further evaluation of the technique is warranted.

Monitoring tissue oxygenation during resuscitation of major burns.

Because subcutaneous and splanchnic oxygenation indices are sensitive indicators of evolving hemorrhagic shock and adequacy of resuscitation, we postulated that these indices might have an equivalent role in the monitoring of severely burned patients. This observational study was undertaken to examine changes in tissue oxygenation indices during burn resuscitation. Seven patients with major burns (54 +/- 21% total body surface area) were studied during the first 36 hours of fluid resuscitation. Silastic tubing was placed in the subcutaneous tissue just beneath both normal skin and deep partial thickness burn. Fiberoptic sensors inserted into the tubing measured subcutaneous oxygen and carbon dioxide tensions in the burnt skin (PO2scb and PCO2scb) and normal skin (PO2scn and PCO2scn) continuously. Gastric intramucosal pH (pHi) and the mucosal CO2 (PCO2m) gap were calculated using gastric tonometers. Mean arterial pressure, arterial pH, lactate, and pHi measurements were obtained for 36 hours. There were no significant differences in mean arterial pressure, arterial pH, or lactate concentrations throughout the study period, whereas indices of tissue oxygenation showed deterioration: pHi decreased from 7.2 +/- 0.1 to 6.7 +/- 0.3 (p = 0.06), the PCO2m gap increased from 12 +/- 17 to 108 +/- 123 mm Hg (p < 0.01), PO2scn decreased from 112 +/- 18 to 50 +/- 11 mm Hg (p < 0.01), PO2scb decreased from 62 +/- 23 to 29 +/- 16 mm Hg (p < 0.01), PCO2scn increased from 42 +/- 4 to 46 +/- 10 mm Hg (p = 0.2), and PCO2scb increased from 42 +/- 10 to 52 +/- 5 mm Hg (p = 0.05). Despite adequate global indices of tissue perfusion after 36 hours of resuscitation, tissue monitoring indicated significant deterioration in the splanchnic circulation and in the normal and burnt skin.

A novel approach to monitor tissue perfusion: Bladder mucosal P co2, P o2, and pHi during ischemia and reperfusion

Purpose: The purpose of this study is to determine if monitoring urinary bladder P co 2, P o 2, and calculated intramucosal pH would be a reliable index of tissue perfusion. Materials and Methods: This nonrandomized controlled study was conducted in a laboratory at a university medical center. Eight immature female Yorkshire pigs were studied with T-9 aortic cross-clamping for 30 minutes followed by a 60-minute period of reperfusion. Cystotomy was performed for placement of a Foley catheter and Paratrend 7 o 2 co 2 sensor. Results: Baseline hemodynamic and metabolic measurements were obtained along with measurements of bladder mucosal Pot and P o 2 (mean ± SEM). Blood flow measured with microspheres confirmed absence of blood flow during occlusion and hyperemia during reperfusion. Bladder mucosal P o 2 decreased from 42 ± 14.0 mm Hg (5.6 kPa)to 1.3 ± 1.3 mm Hg(1.4 kPa) during the 30-minute interval of ischemia. This was followed by an increase of bladder P o 2 to greater than baseline values at the end of the reperfusion period. Bladder mucosal P co 2 increased from 57 ± 4.7 mm Hg (7.6 kPa) to 117 ± 7.1 mm Hg (15.6 kPa) ( P < .05) during ischemia. During reperfusion the P co 2 returned to baseline levels (55 ± 4.0 mm Hg [7.3 kPa]). Calculated bladder mucosal pHi declined from 7.31 ± 0.04 to 7.08 ± 0.05 ( P < .05) during the ischemic period and after reperfusion pHi was 7.17 ± 0.03. Conclusions: Monitoring urinary bladder P o 2, P co 2, or calculating pHi may provide a simple and reliable means of monitoring tissue perfusion.

Evaluation of the Capabilities of a Hemoglobin Vesicle as an Artificial Oxygen Carrier in a Rat Exchange Transfusion Model

Encapsulation of hemoglobin within a liposome is one of the strategies in the development of artificial oxygen carriers. It maintains the oxygen transporting properties of hemoglobin and, at the same time, eliminates the side effects of cell free hemoglobin. Hemoglobin vesicles (HbV) are a type of liposome encapsulated hemoglobin. They have a particle size of approximately 250 nm, a hemoglobin concentration of 10 g/dl, and the oxygen affinity, P50, is regulated to 32 Torr. In this study the authors examined the oxygen transporting capability of HbV in vivo, by performing exchange transfusions in rats. Exchange transfusion (90% of the estimated circulatory volume) with HbV suspended in 5% albumin (containing 160 mEq/L, sodium and 107 mEq/L, chloride) was carried out in male Wistar rats. Mean arterial pressure and heart rate were monitored through the arterial catheter. Arterial blood samples for gas analyses were also obtained from the arterial catheter. Abdominal aortic blood flow was measured by an ultrasonic pulsed Doppler flowmeter as an indicator of cardiac output. The oxygen tension of blood withdrawn from the right atrium was measured as an indicator of mixed venous oxygen tension. These values were employed to calculate oxygen delivery and consumption. Renal cortical and skeletal muscle tissue oxygen tensions were monitored as indicators of tissue perfusion. Five percent albumin and washed rat red blood cells suspended in 5% albumin containing 10 g/dl of hemoglobin; were employed as controls. At the completion of a 90% exchange transfusion, renal cortical and skeletal muscle tissue oxygen tensions, along with oxygen delivery and consumption, were sustained almost equally well with the HbV suspension compared to the washed rat red blood cell suspension, but declined significantly with the albumin suspension. The results indicate that the oxygen transporting capability of HbV was almost equivalent to that of rat red blood cells.

Continuous tissue oxygen tension measurement as a monitor of free-flap viability.

Early recognition of vascular compromise within microvascular free-tissue transfers is essential if reexploration is to prove successful. Tissue oxygen tension is increasingly recognized to be a sensitive and reliable index of tissue perfusion, and preliminary studies suggest that it may be of value in the assessment of free-flap viability. We describe our investigation into the application of an implantable microcatheter oxygen sensor in the monitoring of free flaps used in head and neck and extremity reconstruction. In a preliminary study using the rabbit model, we sought to evaluate the response of oxygen tension as an index of tissue perfusion in myocutaneous (n = 20) and osteomyocutaneous flaps (n = 5) under conditions of arterial and venous occlusion. A clinical study was then undertaken to evaluate the role of this method in the monitoring of surface and buried free flaps. In 30 heterogeneous free-tissue transfers, sensors placed intraoperatively were used to provide continuous information about flap oxygen tension (mean monitoring period 3.2 +/- 0.8 days). The data generated were correlated with changes in clinical parameters and routine flap observations. Results for experimental and clinical data have confirmed the efficacy of continuous tissue oxygen measurements using this device as a method that provides an objective, recordable index of free-tissue transfer viability in a variety of circumstances and vascular events. Tissue oxygen tension is a suitable index by which to evaluate flap viability with the probe placed in muscle or bone but is unreliable when used for the monitoring of revascularized cutaneous flaps.

Functional Microcirculatory Impairment: A Possible Source of Reduced Skin Oxygen Tension in Human Diabetes Mellitus

Lower extremity transcutaneous oxygen tension (TcPO2) is used as a diagnostic and prognostic indicator of tissue perfusion and is reduced in diabetes mellitus. Since cardiac output, leg blood flow and microvascular perfusion each can singly or jointly effect tissue oxygenation, the relative importance of macro- vs microvascular factors has not been resolved. To clarify this issue we compared TcPO2 levels in diabetic and nondiabetic subjects in whom cardiac output, leg pulsatile blood flow, and microcirculatory perfusion parameters were noninvasively measured. In 60 diabetic and 60 nondiabetic subjects the following measurements were done on both legs during a single session evaluation: foot dorsum TcPO2 at 45°, skin microvascular perfusion reserve (MVR) in response to local thermal provocation from 35 to 45° using laser-Doppler, ankle-brachial index using Doppler ultrasound (ABI), and pulsatile leg blood flow using magnetic resonance flowmetry; cardiac output was determined using transthoracic bioimpedance. The diabetic and nondiabetic groups were determined to have nonsignificant differences (mean ± SEM, DM vs NODM) with respect to age (63.3 ± 1.1 vs 60.1 ± 1.5 years), cardiac output (5.5 ± 0.2 vs 5.5 ± 0.2 l/min), leg blood flow (1.6 ± 0.05 vs 1.7 ± 0.06 ml/min/100 cc) and ABI. Although macrocirculatory values were equivalent, microvascular function indicators were significantly reduced in the diabetic group: TcPO2 (51.9 ± 1.4 vs 62.9 ± 1.3 mmHg); MVR (76.7 ± 1.5 vs 84.9 ± 0.9%) and were correlated only in diabetics (r2= 0.48,P< 0.001). The findings suggest a primary linkage between the diabetic TcPO2 deficit and the microcirculatory submaximal vasodilatory response, with little if any role of macrocirculatory factors.

Functional capillary density: an indicator of tissue perfusion?

Functional capillary density (FCD) is one of the parameters obtained by intravital microscopy using epi-illumination of the tissue surface or trans-illumination of thin tissue layers. FCD, defined as the length of red cell-perfused capillaries per observation area (cm-1), has been used as an indicator of the quality of tissue perfusion in various animal models. Quantitative analysis of FCD in randomly selected regions of the tissue is performed by means of a computer-assisted video analysis system which allows calculation of the length of RBC-perfused capillaries. Basically, two different mathematical approaches can be employed: the first approach is based on the addition of the distances between two neighboring points (pixels) on the video screen (Pythagorean principle). The second approach uses the superimposition of a grid system that allows estimation of the capillary length by counting the number of intersections between the capillaries and the grid lines (stereological approach). The immanent error has been calculated in our laboratory to be +/- 1% with the Pythagorean and +/- 5% with the stereological method. Beside these systematic errors of computerized measurement, the individual (user-dependent) errors occurring during recognition and redrawing of the capillaries on the video image with use of a digitizing tablet are in the range of +/- 10% (intraindividual) and +/- 70% (interindividual) for the recognition and +/- 3% (interindividual) for the redrawing procedure. Our studies indicate that the errors resulting from the use of a computer-assisted calculation (Pythagorean or stereological approach) or the user-assisted redrawing of the capillaries are negligible when compared to the errors made during recognition of the capillaries on the video screen. The methods are applied for assessment of FCD in two different microcirculation models of skin muscle and pancreas yielding highly reproducible, user-independent results under physiologic conditions and the pathophysiologic conditions of ischemia-reperfusion.

Capillary perfusion during ischemia-reperfusion in subcutaneous connective tissue and skin muscle.

Ischemia-reperfusion injury was investigated in terms of functional capillary density (FCD), capillary red blood cell velocity (cRBCv), and arteriolar and venular diameter after 4-h ischemia in the unanesthetized hamster skin-fold preparation. Animals in group 1 were studied by transillumination. Group 2 received a bolus injection of fluorescein isothiocyanate (FITC)-dextran (mol wt 150,000) and was studied by transillumination (zone 1) and epi-illumination (zone 2). In group 1, FCD decreased after ischemia (92% of baseline, 30 min), returning to control up to 24 h. cRBCv increased after reperfusion, being 175% of baseline at 24 h. Arterioles and venules dilated for 24 h after reperfusion. In group 2/zone 2, FCD progressively decreased to 11% of control, arteriolar dilation was inhibited, and cRBCv increased 30 min and 2 h after reperfusion. Tissue perfusion index (FCD x cRBCv) increased 158% in group 1 at 24 h, did not change in group 2/zone 1, and was 9% of control at 24 h in group 2/zone 2 (P < or = 0.05). We conclude that increased perfusion is a normal reaction to ischemia-reperfusion injury in this model, and previously observed capillary no reflow is due to FITC-dextran phototoxicity.

Capillary Refill

We are writing to comment on the article by Gorelick et al, "Effective Ambient Temperature and Capillary Refill in Healthy Children" (Pediatrics 1993;92:699-702) and, in particular, the accompanying editorial by Barth (Pediatrics 1993;92:723-724). It is hardly surprising that capillary refill time is affected by ambient temperature or that it is an imperfect indicator of tissue perfusion. We hope that their intention was not to imply that clinical assessment of perfusion is a useless endeavor, however.

Effect of carbon monoxide intoxication on skin perfusion in anaesthetized rats.

Carbon monoxide intoxication decreases systemic blood pressure and peripheral resistance. In order to assess the role of the skin in this process, we measured the perfusion of hind limb shaven skin in anaesthetized rats during acute moderate carbon monoxide intoxication. At a steady blood level of 25% carboxyhaemoglobin, the red blood cell flux was measured as an index of tissue perfusion, using laser Doppler fluxmetry. The mean blood pressure decreased by 30% during carbon monoxide exposure, but there was no change in mean red blood cell flux of the hind limb skin microvessel bed. Thus, rat hind limb skin perfusion was not affected by acute moderate steady state carbon monoxide intoxication.

Assessment of Myocardial Perfusion during CABG Surgery with Two-dimensional Transesophageal Contrast Echocardiography

No reliable, quantifiable index of tissue perfusion is currently available to assess the efforts of coronary artery bypass graft (CABG) surgery. We used two-dimensional transesophageal contrast echocardiography with sonicated Renografin-76 microbubbles to determine the distribution of myocardial blood flow during coronary artery bypass graft surgery in 15 patients. Sonicated Renografin-76 contrast agent was injected into the aortic root of all patients after institution of cardiopulmonary bypass and application of the aortic occlusive clamp. Eight patients had contrast agent injected directly into the free proximal end of the vein-CABG anastomosis. All patients again received aortic root injections during reperfusion after anastomosis of the proximal aortovein and distal coronary artery. Echocardiographic images of the left ventricle short axis at the level of the papillary muscles were obtained in real time and analyzed retrospectively from videotape. Injection of contrast provided information about the magnitude and geometric distribution of coronary artery-vein bypass run-off and enabled identification of poorly perfused myocardial regions. When predicted myocardial perfusion patterns, based on preoperative evaluation of epicardial vessel distribution derived from coronary angiography, were compared to actual perfusion patterns assessed with intraoperative echocardiography, contrast regional myocardial perfusion patterns were predicted 84% of the time (71-97%, 95% confidence limit). Regional myocardial perfusion deficits detected after coronary bypass grafting were associated with regional wall motion abnormalities detected after separation from cardiopulmonary bypass. Our technique makes possible on-line visualization of changes in regional blood flow in the heart before, during, and after CABG.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood lactate and mixed venous-arterial PCO2 gradient as indices of poor peripheral perfusion following cardiopulmonary bypass surgry

Conventional indices of tissue perfusion after surgery involving cardiopulmonary bypass (CPB) may not accurately reflect disordered cell metabolism. Venous hypercarbia leading to an increased veno-arterial difference in CO2 tensions (V-aCO2 gradient) has been shown to reflect critical reductions in systemic and pulmonary blood flow that occur during cardiorespiratory arrest and septic shock. We therefore measured plasma lactate levels and V-aCO2 gradients in 10 patients (mean age 57.2 years) following CPB and compared them with conventional indices of tissue perfusion. Plasma lactate levels, cardiac index (CI) and oxygen uptake (VO2) all increased significantly (p less than 0.05 vs baseline levels) up to 3 h following surgery. Oxygen delivery (DO2) did not change. Plasma lactate levels correlated significantly with CI (r = 0.47, p less than 0.01). V-aCO2 fell significantly with time (p less than 0.01 vs baseline). There was an inverse relationship between V-aCO2 and cardiac index and V-aCO2 and lactate (r = -0.37, p less than 0.05; r = -0.3, p less than 0.05 respectively). We conclude that blood lactate, CI and VO2 increase progressively following CPB. An increase in lactate was associated with a decrease in V-aCO2. An increase in V-aCO2 was not therefore associated with evidence of inadequate tissue perfusion as indicated by an increased blood lactate concentration.