Clinical Hemorheology and Microcirculation 61 (2015) 591–597 DOI 10.3233/CH-141919 IOS Press
591
Acute carbon monoxide poisoning alters hemorheological parameters in human Baris Ozturka,∗ , Okan Arihanb , Figen Coskunc and Neslihan H. Dikmenoglu-Falkmarkenb a
Ankara Numune Education and Research Hospital, Emergency Clinic, Ankara, Turkey Hacettepe University, Faculty of Medicine, Department of Physiology, Ankara, Turkey c Ankara Education and Research Hospital, Emergency Clinic, Ankara, Turkey b
Abstract. Acute carbon monoxide (CO) poisoning seriously hinders oxygen delivery to tissues. This harmful effect of CO may be aggravated by accompanying changes in the viscosity of blood. We had previously reported increased plasma viscosity in people chronically exposed to CO. This study was planned to test our hypothesis that acute CO poisoning increases blood viscosity. For this purpose four main parameters contributing to blood viscosity –hematocrit, erythrocyte deformability, erythrocyte aggregation and plasma viscosity – were determined in patients with acute CO poisoning and compared with healthy controls. Plasma viscosity and erythrocyte aggregation tendency were lower in the CO group (p < 0.05). Erythrocyte deformability was also lower in CO group (p < 0.05). Our results indicate that acute CO poisoning has diverse effects on hemorheological parameters such as attenuating hematocrit value, plasma viscosity, erythrocyte aggregation tendency and erythrocyte deformability. Keywords: Hemorheology, plasma viscosity, erythrocyte deformability, erythrocyte aggregation, hematocrit, acute carbon monoxide poisoning
1. Introduction Carbon monoxide (CO) is a chemical asphyxiant that is produced during the combustion of organic matters such as gasoline, coal, natural gas, tobacco, wood [21]. Humans may accidentally encounter toxic levels of CO mainly by inhalation of smoke or exhaust in closed environment [7, 13]. CO not only competes with and replaces oxygen from hemoglobin to form carboxy-hemoglobin (CO-Hb) but it also shifts the hemoglobin-oxygen dissociation curve to left, at the sites which are still available for oxygen binding, resulting in a decrease in the ability of hemoglobin to release oxygen [20]. Consequent tissue hypoxia is the main means by which CO exerts its toxic effect in the body. The severe hypoxia of CO poisoning has adverse effects on mainly the cardiac and nervous systems [32]. The blood level of CO is expressed in terms of a percentage of the total hemoglobin that is in the form of carboxy-hemoglobin. When carboxy-hemoglobin constitutes less than 10% of the hemoglobin available in blood usually no symptoms appear. At 10–30%, neurological symptoms such as headache, dizziness, weakness, nausea, confusion, disorientation and visual disturbances appear [31]. Ischemic ECG changes, rise in cardiac biomarkers have been reported [23]. Carboxy-hemoglobin levels of 50–60% are often lethal. However, levels as low as approximately 20% was reported to be lethal, due to coronary events, in patients with severe coronary artery disease [31]. ∗ Corresponding author: Baris Ozturk, Ankara Numune Education and Research Hospital, Emergency Clinic, Ankara, Turkey. Tel.: +90 5055251954; Fax: +90 4662123423; E-mail: dr [email protected].
1386-0291/15/$35.00 © 2015 – IOS Press and the authors. All rights reserved
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B. Ozturk et al. / Acute carbon monoxide poisoning alters hemorheological parameters in human
Alteration in the rheological properties of blood (or in other words an increase in the viscosity of it) has been associated with ischemic heart diseases and stroke [14, 30]. Blood viscosity is determined mainly by four properties: hematocrit, plasma viscosity, erythrocyte aggregation, and erythrocyte deformability. An increase in hematocrit, plasma viscosity and erythrocyte aggregation or a fall in erythrocyte deformability results in higher blood viscosity. High blood viscosity in turn slows down the blood flow and results in occlusion. However it should be kept in mind that this “Poiseuille-paradigm” is based on the experiments in rigid tubes which doesn’t fully model natural conditions since it underestimates the contribution of vasoreactivity. Moderate increases in viscosity factors were shown to augment blood flow and tissue perfusion, displaying a bell-shaped curve with an impairment in flow only at higher values of viscosity [22]. Chronic CO poisoning in human [4], and acute CO poisoning in laboratory animals [3, 8, 28] were shown to alter rheological properties of blood. We hypothesized that a rise in the viscosity of blood during acute CO poisoning may contribute to the untoward effects of the gas. We ended up with data indicating a dual effect of acute CO poisoning on hemorheological parameters: a fall in hematocrit which lowers blood viscosity and an attenuation of erythrocyte deformability which has an opposite impact. 2. Materials and methods 2.1. Subjects The study protocol was approved by the Ethics Committee of Ankara Numune Education and Research Hospital (No: 2010-011) and all subjects provided written consent before participating in the study. This study was performed on 2 groups: acute CO poisoning and healthy control. Thirty-two patients who applied to the Emergency Service of Ankara Numune Education and Research Hospital due to acute CO poisoning during November 2010 – March 2011 and who had a carboxy-hemoglobin level above 10% were included in the acute CO group. Thirty-two healthy volunteers from among the hospital personnel were included in the control group. A history of systemic diseases (such as dyslipidemia, hematologic diseases, hypertension, diabetes mellitus, cardiovascular disease), smoking, being on any type of medication were the exclusion criteria for both acute CO and control groups. 2.2. CO measurements Venous blood samples obtained at the time of arrival at the emergency service were used for spectrophotometric analysis of CO level by the CO-Oximeter module of Roche OMNI® S Blood Gas Analyzer (Roche Diagnostics, Basel, Switzerland). 2.3. Hemorheological measurements Venous blood was withdrawn into heparinized tubes (Li Heparin 215 I.U. Vacutest - 9 ml), at the time of arrival, prior to any intervention including the administration of an i.v. solution. Since temperature and time delay may affect the measurements, blood was carried to the laboratory in containers at 4◦ C and all measurements were performed within 2 hours after the blood was collected. Plasma viscosity was measured by a cone-plate viscometer (Wells-Brookfield LVT, USA) at 37◦ C and at 120 rpm (900s–1). For each sample, the average of the 3 measurements was taken.
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Erythrocyte deformability was determined by LORCA (laser-assisted optical rotational cell analyzer) (Mechatronics, Holland) at 37◦ C and at different shear stress values in order to experimentally model the response of erythrocytes in different regions of circulatory system modeled by different shear stresses. The deformation is expressed by the elongation index (EI), a larger EI indicating greater deformation [1, 10]. Erythrocyte aggregation was determined by LORCA at 37◦ C. The amplitude value (AMP) represents the total extent of aggregation, a higher value indicating greater aggregation. t ½ represents the time that elapses until the peak intensity is reduced by half, reflecting the kinetics of aggregation. It is the time required to go half the way to maximum aggregation, a higher value indicating slower aggregation. The aggregation index (AI) represents an overall effect of amplitude and speed, a larger index indicating quicker and/or greater aggregation [9, 11]. 2.4. Other measurements Venous blood was collected into EDTA-containing tubes. A complete blood count including hemoglobin (Hb), hematocrit (Htc), white blood cell (WBC) and red blood cell (RBC) counts, mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC) was done by an electronic blood counter (Coulter, USA). 2.5. Statistics Data were evaluated by the statistics program SPSS 15.0 for Windows. To compare the groups, Student’s t test was used. Data were presented as mean ± SEM. P < 0.05 was considered statistically significant. Pearson correlation test was used to analyze correlations between various parameters. 3. Results Mean age of the subjects in the control group was 35.5 ± 12.2, mean age of the subjects in the acute CO group was 32.8 ± 14.3. Nineteen subjects in the acute CO group were women (59.4%), 13 were men (40.6%); 21 subjects in the control group were women (65.6%), 11 were men (34.4%). The mean CO level of the patients in the acute CO group at the time of arrival was 25.63 ± 9.92%, mean exposure duration was 4 hr 7 min ± 2 hr 29 min (1–8 hr). Mean plasma viscosity (1.32 ± 0.08 mPa·sn) and aggregation index (54.44 ± 10.03%) of the acute CO group were significantly lower than the mean plasma viscosity (1.37 ± 0.07 mPa·sn) and aggregation index (61.12 ± 5.79%) of the control group (P < 0.05). Mean aggregation half-life (t ½ ) value of the acute CO group was significantly higher than mean t ½ of the control group. Aggregation amplitudes of the groups were similar. Erythrocyte deformability, measured as elongation index, for shear stresses 5.33, 9.49, 16.87 and 30 Pa were significantly lower in the acute CO poisoning group (0.418 ± 0.002, 0.494 ± 0.002, 0.551 ± 0.002, 0.590 ± 0.002) compared with the controls (0.426 ± 0.002, 0.502 ± 0.002, 0.559 ± 0.001, 0.598 ± 0.001) (Table 2). There was no significant difference between albumin levels of the acute CO and control groups (4.1 ± 0.1 and 4.2 ± 0.1 g/dL respectively). However, total protein level of the acute CO group was significantly lower than that of the control group (6.7 ± 0.1 and 6.9 ± 0.1 respectively) (P < 0.05).
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B. Ozturk et al. / Acute carbon monoxide poisoning alters hemorheological parameters in human Table 1 Hemorheological parameters and hematocrit values of the groups Plasma viscosity (mPa·sn)
Amplitude (au)
t ½ (s)
AI (%)
Hematocrit (%)
61.12 ± 1.09 54.44 ± 1.77∗
43.3 ± 0.9 40.5 ± 0.9∗
Erythrocyte aggregation Control Acute CO
1.37 ± 0.01 1.33 ± 0.01∗
22.10 ± 0.46 21.47 ± 0.42
2.49 ± 0.13 3.56 ± 0.29∗
Results of plasma viscosity, erythrocyte aggregation and hematocrit for control and acute carbon monoxide poisoning groups. Data was statistically evaluated by Student’s t-test. ∗ P < 0.05 vs. control group. Data is presented mean ± standard error of mean.
Table 2 Erythrocyte deformability of the groups Shear Stress Control Acute CO
5.33 0.426 ± 0.002 0.418 ± 0.002∗
9.49 0.502 ± 0.002 0.494 ± 0.002∗
16.87 0.559 ± 0.001 0.551 ± 0.002∗
30 0.598 ± 0.001 0.590 ± 0.002∗
Results of erythrocyte deformability measured in different shear stress values for control and acute carbon monoxide poisoning groups. Data was statistically evaluated by Student’s t-test. ∗ P < 0.05 vs. control group. Data is presented mean ± standard error of mean.
Hematocrit level of the acute CO group (40.5 ± 0.9%) was also significantly lower than that of the control group (43.3 ± 0.9%) (P < 0.05). Hemoglobin level of the acute CO group was slightly lower (13.5 ± 0.3 g/dL) than that of the control group (14.4 ± 0.3 g/dL). However, this difference did not reach statistical significance. Hemorheological findings proved to be correlated with the duration of exposure to CO. Duration of exposure was negatively correlated (P < 0.05) with AI and plasma viscosity, whereas it was positively correlated (p < 0.05) with t ½ . There was no correlation between the hemorheological findings and carboxy-hemoglobin level. 4. Discussion The present study was undertaken to evaluate the effects of acute CO poisoning on the rheological properties of blood. Our results revealed that acute CO poisoning was associated with decreased plasma viscosity and decreased erythrocyte deformability as well as slowing down of erythrocyte aggregation as indicated by a lower aggregation index and a longer aggregation half-life. These changes were accompanied by a lower level of hematocrit. A lower aggregation index can be due to a decrease either in the amount of aggregation (which will be reflected by a fall in the amplitude value), or in the speed of aggregation (which will be reflected by a rise in t ½ value). Since our results did not reveal a significant change in the amplitude, the decrease in the aggregation index was attributed to the rise in t ½ value. Plasma viscosity is determined mainly by its protein content [16] however lipid content is also influential [12]. The lower plasma viscosity observed in the present study in acute CO poisoning group was accompanied by lower level of total plasma proteins in comparison with the control group. Since the changes in plasma viscosity and erythrocyte aggregation were accompanied by a lower level of
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hematocrit, the results suggest that these changes may be related with hemodilution. The relatively short time needed for all the changes to appear is also in favor of a water shift into the vascular compartment. A decrease in hematocrit and plasma viscosity by a shift of interstitious fluids into the vascular lumen was described as “endogenous dilution” [29]. The hematocrit response observed in this study in human subjects is in accordance with the results of the study by Ramsey on rats [18]. Ramsey has reported an immediate fall of approximately 9% (from 46.2 to 41.9) after acute CO exposure. However, Ramsey has also reported an accompanying rise in the hemoglobin level which made it difficult to explain his results by hemodilution. On the other hand, our results reveal an accompanying fall in the hemoglobin level, more strongly implying hemodilution. In the study by Ramsey hematocrit level was reported to rise half the way back towards normal in 48 hours in animals with intact spleen whereas no rise was observed in splenectomized animals. The results of that study by Ramsey suggest that spleen plays important role in such changes. On the other hand, hematocrit levels dropped both in intact and splenectomized animals, implying additional mechanisms [18]. Our results are contradictory with that of other studies that have reported a rise in hematocrit and hemoglobin levels in acute CO poisoning in humans [15, 19]. The variability of findings may be explained by dose and/or time dependency. The changes in hematocrit may be dose dependent. Dodds et al. [6] have reported a rise in hematocrit in their “high” CO group 45–90 minutes after exposure. However hematocrit slightly decreased in the “low” CO group in 2 hours. The changes may as well be time dependent. Dodds et al, in the same article, have also reported that the rise in hematocrit “45–90 minutes” after exposure in their high CO group returned to the initial value “4 hours” after exposure. Similarly Penney et al. [17] have reported that hematocrit was elevated at the termination of acute CO exposure; however hematocrit declined toward or “below” control levels 4 hours later. In the studies of both Dodds and Penney CO was administered for 90 minutes. In another study by Wang et al. [28] CO was administered intraperitoneally during a 24 hour period. Red blood cell count, hemoglobin and hematocrit levels at 30 minutes after acute CO poisoning were reported to be below the control values before exposure. All 3 levels rose above control values during the following 5 days. These results seem confusing but they may be representing a compensatory mechanism of the body in response to various needs during the course of intoxication and recovery. But it should be kept in mind that rat and rabbit erythrocytes used in these 3 studies differ from human erythrocytes in morphology, deformability and reaction on various stimuli. This is probably the first report about the hemorheological effects of acute CO poisoning in humans. There are only a few previous reports on such acute effects, all of which are from animal studies. Wang et al. [28] have reported that red blood cell count, hematocrit, hemoglobin, whole blood viscosity (at high, low and medium shear rates), aggregation index, deformability index all decreased at 30 minutes after acute intoxication in rabbits. All these results are in accordance with ours except for the rise in plasma viscosity in their study. In the present study plasma viscosity of the acute CO group was lower than the controls. Guan et al. [8] have reported that blood viscosity and hematocrit decreased immediately after acute CO administration following ischemia-reperfusion in rats, which is also in accordance with our results. Significantly lower elongation index values for different shear stresses 5.33, 9.49, 16.87 and 30 Pa in the acute CO poisoning group indicate an attenuation in erythrocyte deformability with exposure to CO. This result is in accordance with Shperling et al. [24] who have also reported a decrease of deformability being most evident at the end of the first day after CO administration. However they have also reported an increase of aggregative capacity of erythrocytes which is on the contrary of ours. On the other hand Bor-Kucukatay et al. [3] have reported no statistically significant changes in erythrocyte aggregation, whole blood viscosity or plasma viscosity after acute intoxication in rats. The only change they detected
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was in erythrocyte deformability. They have reported higher deformability indices at shear stress levels at and below 5.33 Pa in intoxicated animals. It has been reported that oxidative damage results in lower erythrocyte deformability [26]. Wang et al. have claimed that the decrease in erythrocyte deformability in response to acute CO might be due to oxidative damage. They supported this claim by a parallel change in MDA levels: MDA levels rose and deformability indices fall at 30 minutes after CO poisoning. On the other hand, pretreatment of erythrocytes in vitro with CO protects them from oxidative damage thus preserving erythrocyte deformability [2, 27]. Our results as well as that of Wang et al. reveal that the acute in vivo effect of CO is on the opposite to its in vitro effect. It is well known that the signs and prognosis of acute CO poisoning does not correlate well with the level of carboxy-hemoglobin at the time of admission [31]. It was suggested that differences in exposure duration might be responsible for the lack of correlation. In accordance with this knowledge our hemorheological findings were not correlated with the carboxy-hemoglobin levels. However there was correlation between the hemorheological findings and the duration of exposure. Acute CO poisoning seriously hinders oxygen delivery to the tissues. We hypothesized that the harmful effect of resultant hypoxia may be aggravated by accompanying changes in the viscosity of blood. However the results of the present study revealed lower plasma viscosity and erythrocyte aggregation, which proved the opposite. The lower level of hematocrit in the patient group suggests dilution as the cause of these changes. However, since the blood samples for hemorheological evaluation were obtained before any medical intervention, dilution can only be due to unexpected but beneficial compensation of the body by itself. Formation of carboxyhemoglobin seems to have a negative impact on the contribution of red blood cells to blood viscosity parameters which was compensated with a hemodilution suggesting a “viscoregulation” postulated by Dintenfass and supported with the studies of Reinhardt and his colleagues [5, 25]. As summarized here, there exists a lot of contradiction between the results of acute CO experiments. The variation of results may be due to differences in exposure dose and duration, the lapse of time between exposure and hemorheological evaluation as well as species differences. The variations may as well be representing a compensatory mechanism of the body in response to various needs during the course of intoxication and recovery. Further studies taking these possibilities into account are needed to throw light on the whole process. References [1] O.K. Baskurt, M.R. Hardeman, M. Uyuklu, P. Ulker, M. Cengiz, N. Nemeth, S. Shin, T. Alexy and H.J. Meiselman, Comparison of three commercially available ektacytometers with different shearing geometries, Biorheol 46 (2009), 251–264. [2] M.W. Bitensky, Safe Extension of Red Blood Cell Storage Life at 4(C, DOE Office of Scientific and Technical Information (OSTI). LA-UR-96-793. US. 1995. [3] M. Bor-Kucukatay, H. Atalay, N. Karagenc, G. Erken and V. Kucukatay, The effect of carbon monoxide poisoning on hemorheological parameters in rats and the alterations in these parameters in response to three kinds of treatments, Clin Hemorheol Microcirc 44 (2010), 87–96. [4] N. Dikmeno˘glu and N. Seringec¸, Effects of work place carbon monoxide exposure on blood viscosity, Archives of Environmental & Occupational Health 65 (2010), 49–53. [5] L. Dintenfass, Blood Viscosity, Hyperviscosity & Hyperviscosaemia, MTP Press, Melbourne, 1985, p. 482. [6] R.G. Dodds, D.G. Penney and B.B. Sutariya, Cardiovascular, metabolic and neurologic effects of carbon monoxide and cyanide in the rat, Toxicoloy Letters 61 (1992), 243–254.
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[7] M. Goldstein, Carbon monoxide poisoning, J Emergency Nursing 34 (2008), 538–542. [8] L. Guan, Z.Y. Li, J.Y. Zhao, X.X. Xu, T. Wen and Y.L. Zhang, Dynamic changes of hemorheology in rats after carbon monoxide poisoning, Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za Zhi 28 (2010), 885–890. [9] M.R. Hardeman, J.G.G. Dobbe and C. Ince, The Laser-assisted optical rotational cell analyzer (LORCA) as red blood cell aggregometer, Clin Hemorheol Microcirc 25 (2001), 1–11. [10] M.R. Hardeman, P.T. Goedhart and N.H. Schut, Laser-assisted optical rotational cell analyzer (L.O.R.C.A.); II red blood cell deformability: Elongation index versus cell transit time, Clin Hemorheol Microcirc 14 (1994), 619–630. [11] M.R. Hardeman, M. Levitus, A. Pelliccia and A.A. Bouman, Test 1 analyser for determination of ESR. 2. Experimental evaluation and comparison with RBC aggregometry, Scand J Clin Lab Invest 70 (2010), 26–32. [12] C. Irace, C. Carallo, F. Scavelli, T. Esposito, M.S. De Franceschi, C. Tripolino and A. Gnasso, Influence of blood lipids on plasma and blood viscosity, Clin Hemorheol Microcirc 57 (2014), 283–290. [13] L.W. Kao and K.A. Nanagas, Carbon monoxide poisoning, Emerg Med Clin N Am 22 (2004), 985–1018. [14] K.R. Kensey, The mechanistic relationship between hemorheological characteristics and cardiovascular disease, Cur Med Res Opin 19 (2003), 587–596. [15] S. Lee, I. Choi and K. Song, Hematological changes in acute carbon monoxide intoxication, Yonsei Medical J 35 (1994), 245–251. [16] G.D.O. Lowe, T.C. Pearson, J. Stuart, D.J. Thomas and C.H.M. Walker, Clinical Blood Rheology, Volume I, CRC Press, Boca Raton- Florida, 1998. [17] D.G. Penney, K. Verma and J.A. Hull, Cardiovascular, metabolic and neurologic effects of acute carbon monoxide poisoning in the rat, Toxicology Letters 45 (1989), 207–213. [18] J.M. Ramsey, The immediate haematological response in the rat to experimental exposures of carbon monoxide, J Physiol 202 (1969), 297–304. [19] J.M. Ramsey, The time course of hematological response to experimental exposures of carbon monoxide, Arch Environ Health 18 (1969), 323–329. [20] F.J.W. Roughton and R.C. Darling, The effect of carbon monoxide on the oxyhemoglobin dissociation curve, Am J Physiol 141 (1944), 17–31. [21] S.W. Ryter and L.E. Otterbein, Carbon monoxide in biology and medicine, Bio Essays 26 (2004), 270–280. [22] B.Y. Salazar V´azquez, P. Cabrales, A.G. Tsai and M. Intaglietta, Nonlinear cardiovascular regulation consequent to changes in blood viscosity, Clin Hemorheol Microcirc 49 (2011), 29–36. [23] D. Satran, C.R. Henry, C. Adkinson, C.I. Nicholson, Y. Bracha and T.D. Henry, Cardiovascular manifestation of moderate to severe carbon monoxide poisoning, J Am College of Cardiology 45 (2005), 1513–1516. [24] I.A. Shperling, V.V. Novitskii, N.V. Riazantseva, O.A. Rogov, S.B. Tkachenko, O.N. Filippova, E.V. Saprykina, O.I. Aliev and M.B. Plotnikov, Mechanisms underlying changes in functional properties of red cells in acute action of carbon monoxide, Patol Fiziol Eksp Ter 1 (2008), 18–20. [25] A. Singh, A., K.U. Eckardt, A. Zimmermann, K.H. G¨otz, M. Hamann, P.J. Ratcliffe, A. Kurtz and W.H. Reinhart, Increased plasma viscosity as a reason for inappropriate erythropoietin formation, J Clin Invest 91 (1993), 251–256. [26] L.M. Snyder, N.N. Fortier, J. Trainor, J. Jacobs, L. Leb, B. Lubin, D. Chiu, S. Shohet and N. Mohandas, Effect of hydrogen peroxide exposure on normal human erythrocyte deformability, morphology, surface characteristics and spectrinhemoglobin cross-linking, J Clin Invest 76 (1985), 1971–1977. [27] M.A. Srour, Y.Y. Bilto, M. Juma and M.R. Irhimeh, Exposure of human erythrocytes to oxygen radicals causes loss of deformability, increased osmotic fragility, lipid peroxidation and protein degradation, Clin Hemorheol Microcirc 23 (2000), 13–21. [28] X. Wang, X. Wang, T. Wen, L. Guan, Y. Zhang, M. Zhu and J. Zhao, Hemorheological changes in cerebral circulation of rabbits with acute carbon monoxide poisoning, Clin Hemorheol Microcirc 43 (2009), 271–282. [29] S. Wolf, M. Reim and F. Jung, Effect of garlic on conjunctival vessels: A randomised, placebo-controlled, double-blind trial, Br J Clin Pract Suppl 69 (1990), 36–39. [30] M. Woodward, A. Rumley, H. Tunstall-Pedoe and G. Lowe, Does sticky blood predict a sticky end? Association of blood viscosity, haematocrit and fibrinogen with mortality in the West of Scotland, Br J Haematol 122 (2003), 645–650. [31] World Health Organization (WHO). Environmental Health Criteria report 213: Carbon monoxide poisoning, Geneva, 1999, pp. 307–328. [32] B. Yelken, B. Tanriverdi, F. C¸etinbas¸, D. Memis¸ and N. S¨ut, The assessment of QT intervals in acute carbon monoxide poisoning, Anadolu Kardiyol Derg 9 (2009), 397–400.
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Acute carbon monoxide poisoning alters hemorheological parameters in human Baris Ozturka,∗ , Okan Arihanb , Figen Coskunc and Neslihan H. Dikmenoglu-Falkmarkenb a
Ankara Numune Education and Research Hospital, Emergency Clinic, Ankara, Turkey Hacettepe University, Faculty of Medicine, Department of Physiology, Ankara, Turkey c Ankara Education and Research Hospital, Emergency Clinic, Ankara, Turkey b
Abstract. Acute carbon monoxide (CO) poisoning seriously hinders oxygen delivery to tissues. This harmful effect of CO may be aggravated by accompanying changes in the viscosity of blood. We had previously reported increased plasma viscosity in people chronically exposed to CO. This study was planned to test our hypothesis that acute CO poisoning increases blood viscosity. For this purpose four main parameters contributing to blood viscosity –hematocrit, erythrocyte deformability, erythrocyte aggregation and plasma viscosity – were determined in patients with acute CO poisoning and compared with healthy controls. Plasma viscosity and erythrocyte aggregation tendency were lower in the CO group (p < 0.05). Erythrocyte deformability was also lower in CO group (p < 0.05). Our results indicate that acute CO poisoning has diverse effects on hemorheological parameters such as attenuating hematocrit value, plasma viscosity, erythrocyte aggregation tendency and erythrocyte deformability. Keywords: Hemorheology, plasma viscosity, erythrocyte deformability, erythrocyte aggregation, hematocrit, acute carbon monoxide poisoning
1. Introduction Carbon monoxide (CO) is a chemical asphyxiant that is produced during the combustion of organic matters such as gasoline, coal, natural gas, tobacco, wood [21]. Humans may accidentally encounter toxic levels of CO mainly by inhalation of smoke or exhaust in closed environment [7, 13]. CO not only competes with and replaces oxygen from hemoglobin to form carboxy-hemoglobin (CO-Hb) but it also shifts the hemoglobin-oxygen dissociation curve to left, at the sites which are still available for oxygen binding, resulting in a decrease in the ability of hemoglobin to release oxygen [20]. Consequent tissue hypoxia is the main means by which CO exerts its toxic effect in the body. The severe hypoxia of CO poisoning has adverse effects on mainly the cardiac and nervous systems [32]. The blood level of CO is expressed in terms of a percentage of the total hemoglobin that is in the form of carboxy-hemoglobin. When carboxy-hemoglobin constitutes less than 10% of the hemoglobin available in blood usually no symptoms appear. At 10–30%, neurological symptoms such as headache, dizziness, weakness, nausea, confusion, disorientation and visual disturbances appear [31]. Ischemic ECG changes, rise in cardiac biomarkers have been reported [23]. Carboxy-hemoglobin levels of 50–60% are often lethal. However, levels as low as approximately 20% was reported to be lethal, due to coronary events, in patients with severe coronary artery disease [31]. ∗ Corresponding author: Baris Ozturk, Ankara Numune Education and Research Hospital, Emergency Clinic, Ankara, Turkey. Tel.: +90 5055251954; Fax: +90 4662123423; E-mail: dr [email protected].
1386-0291/15/$35.00 © 2015 – IOS Press and the authors. All rights reserved
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B. Ozturk et al. / Acute carbon monoxide poisoning alters hemorheological parameters in human
Alteration in the rheological properties of blood (or in other words an increase in the viscosity of it) has been associated with ischemic heart diseases and stroke [14, 30]. Blood viscosity is determined mainly by four properties: hematocrit, plasma viscosity, erythrocyte aggregation, and erythrocyte deformability. An increase in hematocrit, plasma viscosity and erythrocyte aggregation or a fall in erythrocyte deformability results in higher blood viscosity. High blood viscosity in turn slows down the blood flow and results in occlusion. However it should be kept in mind that this “Poiseuille-paradigm” is based on the experiments in rigid tubes which doesn’t fully model natural conditions since it underestimates the contribution of vasoreactivity. Moderate increases in viscosity factors were shown to augment blood flow and tissue perfusion, displaying a bell-shaped curve with an impairment in flow only at higher values of viscosity [22]. Chronic CO poisoning in human [4], and acute CO poisoning in laboratory animals [3, 8, 28] were shown to alter rheological properties of blood. We hypothesized that a rise in the viscosity of blood during acute CO poisoning may contribute to the untoward effects of the gas. We ended up with data indicating a dual effect of acute CO poisoning on hemorheological parameters: a fall in hematocrit which lowers blood viscosity and an attenuation of erythrocyte deformability which has an opposite impact. 2. Materials and methods 2.1. Subjects The study protocol was approved by the Ethics Committee of Ankara Numune Education and Research Hospital (No: 2010-011) and all subjects provided written consent before participating in the study. This study was performed on 2 groups: acute CO poisoning and healthy control. Thirty-two patients who applied to the Emergency Service of Ankara Numune Education and Research Hospital due to acute CO poisoning during November 2010 – March 2011 and who had a carboxy-hemoglobin level above 10% were included in the acute CO group. Thirty-two healthy volunteers from among the hospital personnel were included in the control group. A history of systemic diseases (such as dyslipidemia, hematologic diseases, hypertension, diabetes mellitus, cardiovascular disease), smoking, being on any type of medication were the exclusion criteria for both acute CO and control groups. 2.2. CO measurements Venous blood samples obtained at the time of arrival at the emergency service were used for spectrophotometric analysis of CO level by the CO-Oximeter module of Roche OMNI® S Blood Gas Analyzer (Roche Diagnostics, Basel, Switzerland). 2.3. Hemorheological measurements Venous blood was withdrawn into heparinized tubes (Li Heparin 215 I.U. Vacutest - 9 ml), at the time of arrival, prior to any intervention including the administration of an i.v. solution. Since temperature and time delay may affect the measurements, blood was carried to the laboratory in containers at 4◦ C and all measurements were performed within 2 hours after the blood was collected. Plasma viscosity was measured by a cone-plate viscometer (Wells-Brookfield LVT, USA) at 37◦ C and at 120 rpm (900s–1). For each sample, the average of the 3 measurements was taken.
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Erythrocyte deformability was determined by LORCA (laser-assisted optical rotational cell analyzer) (Mechatronics, Holland) at 37◦ C and at different shear stress values in order to experimentally model the response of erythrocytes in different regions of circulatory system modeled by different shear stresses. The deformation is expressed by the elongation index (EI), a larger EI indicating greater deformation [1, 10]. Erythrocyte aggregation was determined by LORCA at 37◦ C. The amplitude value (AMP) represents the total extent of aggregation, a higher value indicating greater aggregation. t ½ represents the time that elapses until the peak intensity is reduced by half, reflecting the kinetics of aggregation. It is the time required to go half the way to maximum aggregation, a higher value indicating slower aggregation. The aggregation index (AI) represents an overall effect of amplitude and speed, a larger index indicating quicker and/or greater aggregation [9, 11]. 2.4. Other measurements Venous blood was collected into EDTA-containing tubes. A complete blood count including hemoglobin (Hb), hematocrit (Htc), white blood cell (WBC) and red blood cell (RBC) counts, mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC) was done by an electronic blood counter (Coulter, USA). 2.5. Statistics Data were evaluated by the statistics program SPSS 15.0 for Windows. To compare the groups, Student’s t test was used. Data were presented as mean ± SEM. P < 0.05 was considered statistically significant. Pearson correlation test was used to analyze correlations between various parameters. 3. Results Mean age of the subjects in the control group was 35.5 ± 12.2, mean age of the subjects in the acute CO group was 32.8 ± 14.3. Nineteen subjects in the acute CO group were women (59.4%), 13 were men (40.6%); 21 subjects in the control group were women (65.6%), 11 were men (34.4%). The mean CO level of the patients in the acute CO group at the time of arrival was 25.63 ± 9.92%, mean exposure duration was 4 hr 7 min ± 2 hr 29 min (1–8 hr). Mean plasma viscosity (1.32 ± 0.08 mPa·sn) and aggregation index (54.44 ± 10.03%) of the acute CO group were significantly lower than the mean plasma viscosity (1.37 ± 0.07 mPa·sn) and aggregation index (61.12 ± 5.79%) of the control group (P < 0.05). Mean aggregation half-life (t ½ ) value of the acute CO group was significantly higher than mean t ½ of the control group. Aggregation amplitudes of the groups were similar. Erythrocyte deformability, measured as elongation index, for shear stresses 5.33, 9.49, 16.87 and 30 Pa were significantly lower in the acute CO poisoning group (0.418 ± 0.002, 0.494 ± 0.002, 0.551 ± 0.002, 0.590 ± 0.002) compared with the controls (0.426 ± 0.002, 0.502 ± 0.002, 0.559 ± 0.001, 0.598 ± 0.001) (Table 2). There was no significant difference between albumin levels of the acute CO and control groups (4.1 ± 0.1 and 4.2 ± 0.1 g/dL respectively). However, total protein level of the acute CO group was significantly lower than that of the control group (6.7 ± 0.1 and 6.9 ± 0.1 respectively) (P < 0.05).
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B. Ozturk et al. / Acute carbon monoxide poisoning alters hemorheological parameters in human Table 1 Hemorheological parameters and hematocrit values of the groups Plasma viscosity (mPa·sn)
Amplitude (au)
t ½ (s)
AI (%)
Hematocrit (%)
61.12 ± 1.09 54.44 ± 1.77∗
43.3 ± 0.9 40.5 ± 0.9∗
Erythrocyte aggregation Control Acute CO
1.37 ± 0.01 1.33 ± 0.01∗
22.10 ± 0.46 21.47 ± 0.42
2.49 ± 0.13 3.56 ± 0.29∗
Results of plasma viscosity, erythrocyte aggregation and hematocrit for control and acute carbon monoxide poisoning groups. Data was statistically evaluated by Student’s t-test. ∗ P < 0.05 vs. control group. Data is presented mean ± standard error of mean.
Table 2 Erythrocyte deformability of the groups Shear Stress Control Acute CO
5.33 0.426 ± 0.002 0.418 ± 0.002∗
9.49 0.502 ± 0.002 0.494 ± 0.002∗
16.87 0.559 ± 0.001 0.551 ± 0.002∗
30 0.598 ± 0.001 0.590 ± 0.002∗
Results of erythrocyte deformability measured in different shear stress values for control and acute carbon monoxide poisoning groups. Data was statistically evaluated by Student’s t-test. ∗ P < 0.05 vs. control group. Data is presented mean ± standard error of mean.
Hematocrit level of the acute CO group (40.5 ± 0.9%) was also significantly lower than that of the control group (43.3 ± 0.9%) (P < 0.05). Hemoglobin level of the acute CO group was slightly lower (13.5 ± 0.3 g/dL) than that of the control group (14.4 ± 0.3 g/dL). However, this difference did not reach statistical significance. Hemorheological findings proved to be correlated with the duration of exposure to CO. Duration of exposure was negatively correlated (P < 0.05) with AI and plasma viscosity, whereas it was positively correlated (p < 0.05) with t ½ . There was no correlation between the hemorheological findings and carboxy-hemoglobin level. 4. Discussion The present study was undertaken to evaluate the effects of acute CO poisoning on the rheological properties of blood. Our results revealed that acute CO poisoning was associated with decreased plasma viscosity and decreased erythrocyte deformability as well as slowing down of erythrocyte aggregation as indicated by a lower aggregation index and a longer aggregation half-life. These changes were accompanied by a lower level of hematocrit. A lower aggregation index can be due to a decrease either in the amount of aggregation (which will be reflected by a fall in the amplitude value), or in the speed of aggregation (which will be reflected by a rise in t ½ value). Since our results did not reveal a significant change in the amplitude, the decrease in the aggregation index was attributed to the rise in t ½ value. Plasma viscosity is determined mainly by its protein content [16] however lipid content is also influential [12]. The lower plasma viscosity observed in the present study in acute CO poisoning group was accompanied by lower level of total plasma proteins in comparison with the control group. Since the changes in plasma viscosity and erythrocyte aggregation were accompanied by a lower level of
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hematocrit, the results suggest that these changes may be related with hemodilution. The relatively short time needed for all the changes to appear is also in favor of a water shift into the vascular compartment. A decrease in hematocrit and plasma viscosity by a shift of interstitious fluids into the vascular lumen was described as “endogenous dilution” [29]. The hematocrit response observed in this study in human subjects is in accordance with the results of the study by Ramsey on rats [18]. Ramsey has reported an immediate fall of approximately 9% (from 46.2 to 41.9) after acute CO exposure. However, Ramsey has also reported an accompanying rise in the hemoglobin level which made it difficult to explain his results by hemodilution. On the other hand, our results reveal an accompanying fall in the hemoglobin level, more strongly implying hemodilution. In the study by Ramsey hematocrit level was reported to rise half the way back towards normal in 48 hours in animals with intact spleen whereas no rise was observed in splenectomized animals. The results of that study by Ramsey suggest that spleen plays important role in such changes. On the other hand, hematocrit levels dropped both in intact and splenectomized animals, implying additional mechanisms [18]. Our results are contradictory with that of other studies that have reported a rise in hematocrit and hemoglobin levels in acute CO poisoning in humans [15, 19]. The variability of findings may be explained by dose and/or time dependency. The changes in hematocrit may be dose dependent. Dodds et al. [6] have reported a rise in hematocrit in their “high” CO group 45–90 minutes after exposure. However hematocrit slightly decreased in the “low” CO group in 2 hours. The changes may as well be time dependent. Dodds et al, in the same article, have also reported that the rise in hematocrit “45–90 minutes” after exposure in their high CO group returned to the initial value “4 hours” after exposure. Similarly Penney et al. [17] have reported that hematocrit was elevated at the termination of acute CO exposure; however hematocrit declined toward or “below” control levels 4 hours later. In the studies of both Dodds and Penney CO was administered for 90 minutes. In another study by Wang et al. [28] CO was administered intraperitoneally during a 24 hour period. Red blood cell count, hemoglobin and hematocrit levels at 30 minutes after acute CO poisoning were reported to be below the control values before exposure. All 3 levels rose above control values during the following 5 days. These results seem confusing but they may be representing a compensatory mechanism of the body in response to various needs during the course of intoxication and recovery. But it should be kept in mind that rat and rabbit erythrocytes used in these 3 studies differ from human erythrocytes in morphology, deformability and reaction on various stimuli. This is probably the first report about the hemorheological effects of acute CO poisoning in humans. There are only a few previous reports on such acute effects, all of which are from animal studies. Wang et al. [28] have reported that red blood cell count, hematocrit, hemoglobin, whole blood viscosity (at high, low and medium shear rates), aggregation index, deformability index all decreased at 30 minutes after acute intoxication in rabbits. All these results are in accordance with ours except for the rise in plasma viscosity in their study. In the present study plasma viscosity of the acute CO group was lower than the controls. Guan et al. [8] have reported that blood viscosity and hematocrit decreased immediately after acute CO administration following ischemia-reperfusion in rats, which is also in accordance with our results. Significantly lower elongation index values for different shear stresses 5.33, 9.49, 16.87 and 30 Pa in the acute CO poisoning group indicate an attenuation in erythrocyte deformability with exposure to CO. This result is in accordance with Shperling et al. [24] who have also reported a decrease of deformability being most evident at the end of the first day after CO administration. However they have also reported an increase of aggregative capacity of erythrocytes which is on the contrary of ours. On the other hand Bor-Kucukatay et al. [3] have reported no statistically significant changes in erythrocyte aggregation, whole blood viscosity or plasma viscosity after acute intoxication in rats. The only change they detected
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was in erythrocyte deformability. They have reported higher deformability indices at shear stress levels at and below 5.33 Pa in intoxicated animals. It has been reported that oxidative damage results in lower erythrocyte deformability [26]. Wang et al. have claimed that the decrease in erythrocyte deformability in response to acute CO might be due to oxidative damage. They supported this claim by a parallel change in MDA levels: MDA levels rose and deformability indices fall at 30 minutes after CO poisoning. On the other hand, pretreatment of erythrocytes in vitro with CO protects them from oxidative damage thus preserving erythrocyte deformability [2, 27]. Our results as well as that of Wang et al. reveal that the acute in vivo effect of CO is on the opposite to its in vitro effect. It is well known that the signs and prognosis of acute CO poisoning does not correlate well with the level of carboxy-hemoglobin at the time of admission [31]. It was suggested that differences in exposure duration might be responsible for the lack of correlation. In accordance with this knowledge our hemorheological findings were not correlated with the carboxy-hemoglobin levels. However there was correlation between the hemorheological findings and the duration of exposure. Acute CO poisoning seriously hinders oxygen delivery to the tissues. We hypothesized that the harmful effect of resultant hypoxia may be aggravated by accompanying changes in the viscosity of blood. However the results of the present study revealed lower plasma viscosity and erythrocyte aggregation, which proved the opposite. The lower level of hematocrit in the patient group suggests dilution as the cause of these changes. However, since the blood samples for hemorheological evaluation were obtained before any medical intervention, dilution can only be due to unexpected but beneficial compensation of the body by itself. Formation of carboxyhemoglobin seems to have a negative impact on the contribution of red blood cells to blood viscosity parameters which was compensated with a hemodilution suggesting a “viscoregulation” postulated by Dintenfass and supported with the studies of Reinhardt and his colleagues [5, 25]. As summarized here, there exists a lot of contradiction between the results of acute CO experiments. The variation of results may be due to differences in exposure dose and duration, the lapse of time between exposure and hemorheological evaluation as well as species differences. The variations may as well be representing a compensatory mechanism of the body in response to various needs during the course of intoxication and recovery. Further studies taking these possibilities into account are needed to throw light on the whole process. References [1] O.K. Baskurt, M.R. Hardeman, M. Uyuklu, P. Ulker, M. Cengiz, N. Nemeth, S. Shin, T. Alexy and H.J. Meiselman, Comparison of three commercially available ektacytometers with different shearing geometries, Biorheol 46 (2009), 251–264. [2] M.W. Bitensky, Safe Extension of Red Blood Cell Storage Life at 4(C, DOE Office of Scientific and Technical Information (OSTI). LA-UR-96-793. US. 1995. [3] M. Bor-Kucukatay, H. Atalay, N. Karagenc, G. Erken and V. Kucukatay, The effect of carbon monoxide poisoning on hemorheological parameters in rats and the alterations in these parameters in response to three kinds of treatments, Clin Hemorheol Microcirc 44 (2010), 87–96. [4] N. Dikmeno˘glu and N. Seringec¸, Effects of work place carbon monoxide exposure on blood viscosity, Archives of Environmental & Occupational Health 65 (2010), 49–53. [5] L. Dintenfass, Blood Viscosity, Hyperviscosity & Hyperviscosaemia, MTP Press, Melbourne, 1985, p. 482. [6] R.G. Dodds, D.G. Penney and B.B. Sutariya, Cardiovascular, metabolic and neurologic effects of carbon monoxide and cyanide in the rat, Toxicoloy Letters 61 (1992), 243–254.
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