Studies
on
Blood Rheology in Patients with
Primary Pulmonary Hypertension Sylvi Ulrika Persson, B.SC. Carl Gunner Gustavsson, M.D.* Hans Larsson,
M.D., Ph.D.
and
Stig Persson, M.D., Ph.D.* LUND,SWEDEN
Abstract The
rheologic properties of blood were studied in 6 patients with primary pulmonary hypertension (PPH) and compared with those of a control group of 10 healthy subjects. Blood viscosity was studied with a rotational viscometer and blood cell deformability with a filtrometer giving values for clogging particles (CP) and red cell transit time (RCTT). Blood viscosity at varying shear rates was found to be increased both at natural (p < 0.025-0.005) and standardized -1 in patients with PPH. Red cell deformabils hematocrit, 45% (p < 0.05 at 40 ) ity was reduced as indicated by a significant increase of RCTT (p 20 mmHg or systolic pulmonary artery pressure > 30 mmHg at rest. Excluded were patients with any disease that might cause pulmonary hypertension, eg, congenital or valvular heart disease, pulmonary embolism, collagenosis, parenchymal pulmonary disease, or a history of drug abuse. Seven consecutive patients, satisfying these criteria, were regarded as having PPH. One of them died shortly after admission. The other 6 patients gave their informed consent to participate in this study. The study group comprised 2 men and 4 women, aged thirty to seventy (mean forty-four years). One patient had stopped smoking ten years earlier, whereas the other 5 were nonsmokers. In case number 5 the diagnosis of pulmonary hypertension was based upon echocardiography showing dilatation and hypertrophy of the right ventricle with inverse septal movement and with Doppler a peak systolic, right ventricular to right atrial, pressure gradient of 92 mmHg. In this case pulmonary hypertension had earlier occurred in a first-degree relative. In the other 5 patients the diagnoses was based upon right heart catheterization. Clinical and hemodynamic data are detailed in Table I. Five of our patients were receiving treatment with calcium channel blockers, 3 with anticoagulants, and 1 each with an ACE inhibitor and a diuretic.
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838 The control group in the hemorheologic study comprised 10 healthy nonsmoking subjects without medication, 5 men and 5 women, aged twenty to sixty-two (mean forty-one
years). Blood Rheology Measurements Blood samples were drawn in the morning from an antecubital vein with the subjects supine and fasting overnight. Anticoagulation was performed with EDTA, except for the samples bound for fibrinogen analysis where citrate was used. Plasma viscosity was measured in a capillary U-tube at 37 °C and expressed as the ratio between the passage times of plasma and water, ie, as plasma viscosity relative to the 2 viscosity of water of the same temperature . Whole blood viscosity was determined at natural hematocrit, as well as at a standardized hematocrit of 45070. All measurements were performed at 37 °C. A computer-controlled rotational viscometer was used, measuring at stepwise increasing shear rates from 0.8 s-’ to 40.0 s-’ and then with decreasing shear rates back to 0.8 S-1 as previously described.’ For measuring rec cell deformability, whole blood was centrifuged for ten minutes at 2500 g. The plasma and buffy coat were removed and discarded. Then 0.55 mL of the packed red cells were aspirated from the middle of the centrifuged red call column and washed twice in phosphate-buffered saline (PBS) of pH 7.4 and containing bovine serum albumin, 2.5 g/L. Before use the buffer was filtered through a 45 micrometer filter. TABLE II Blood
Rheology Parameters Controls (n 10) PPH patients (n 6) (n
(n =
Downloaded from ang.sagepub.com at PRINCETON UNIV LIBRARY on July 16, 2015
839
Finally the red blood cells were resuspended in the PBS to a hematocrit of 1 0to . Residual leukocytes in the erythrocyte suspensions were counted under the microscope. In all suspensions the leukocyte count was < 0.025 x 109/L. Blood cell filtration studies were finally performed with a filtrometer according to Dormandy et a14 with a negative pressure of 3 cm of water. Filters with a 5 micrometer pore diameter were used. In all filtration studies we used filters from the same batch. The time from blood sampling to finishing 5 the measurements did not exceed three hours in any case.5 For determining white cell deformability, centrifugation of whole blood was performed as for red cells. The buffy coat was aspirated completely from the cell column and suspended in 2 mL of plasma in a 5 mL disposable plastic tube with cap and placed in a holder at a horizontal position for fifteen minutes. During this time the red blood cells aggregated and sedimented to the tube wall. The tube was then tilted carefully to about 45 degrees and the plasma supernatant was removed and washed twice in phosphatebuffered saline (PBS) containing 1 mL plasma to 19 mL of PBS. Leukocytes were counted under the microscope and finally suspended to a concentration of 5 x 109/L. Filtration studies were performed in the same way as for red blood cells but with filters with a pore size of 8 micrometers. Plasma fibrinogen concentration was determined by a kit method. Total cholesterol concentration in plasma was measured by an enzymatic colorimetric method and HDL cholesterol concentration by the same method after chylomicrons, VLDL, and LDL were precipitated by addition of phosphotungstic acid and magnesium ions to the sample. Erythrocyte mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), hematocrit, hemoglobin concentration, and counts of erythrocytes and leukocytes were determined by routine methods. Heart Catheterization The indication for right heart catheterization was entirely clinical and in case affected the by viscosity studies. A balloon-tipped thermodilution catheter was introduced percutaneously via an antecubital vein and positioned in a pulmonary artery under fluoroscopic control. For measurements of systemic blood pressure an indwelling plastic cannula was placed in a brachial artery, usually on the contralateral side. Both catheters were connected to pressure transducers at the level of the anterior axillary line. Leads V 1, V4, and V6 were continuously monitored and recorded together with the pressures. Cardiac output was measured with a standard thermodilution equipment, and all cardiac output determinations represented the average of three or more measurements.
Right
Echocardiography Two-dimensional and M-mode echocardiography was performed with the patients in the left semilateral decubitus position. Tricuspid regurgitation flow was examined from the apex by a separate continuous wave Doppler transducer. Peak systolic right ventricular to right atrial pressure gradients were calculated from the peak systolic regurgitation 6 velocities as described elsewhere .
Downloaded from ang.sagepub.com at PRINCETON UNIV LIBRARY on July 16, 2015
840
Results The blood viscosity (Table II) at 37 °C was increased in patients with PPH as compared with the control subjects. At natural hematocrit (Fig. 1) the differences were statistically significant at all shear rates studied, 0.8, 2.3, 19.6, and 40.0 s-’ (p
on
Blood Rheology in Patients with
Primary Pulmonary Hypertension Sylvi Ulrika Persson, B.SC. Carl Gunner Gustavsson, M.D.* Hans Larsson,
M.D., Ph.D.
and
Stig Persson, M.D., Ph.D.* LUND,SWEDEN
Abstract The
rheologic properties of blood were studied in 6 patients with primary pulmonary hypertension (PPH) and compared with those of a control group of 10 healthy subjects. Blood viscosity was studied with a rotational viscometer and blood cell deformability with a filtrometer giving values for clogging particles (CP) and red cell transit time (RCTT). Blood viscosity at varying shear rates was found to be increased both at natural (p < 0.025-0.005) and standardized -1 in patients with PPH. Red cell deformabils hematocrit, 45% (p < 0.05 at 40 ) ity was reduced as indicated by a significant increase of RCTT (p 20 mmHg or systolic pulmonary artery pressure > 30 mmHg at rest. Excluded were patients with any disease that might cause pulmonary hypertension, eg, congenital or valvular heart disease, pulmonary embolism, collagenosis, parenchymal pulmonary disease, or a history of drug abuse. Seven consecutive patients, satisfying these criteria, were regarded as having PPH. One of them died shortly after admission. The other 6 patients gave their informed consent to participate in this study. The study group comprised 2 men and 4 women, aged thirty to seventy (mean forty-four years). One patient had stopped smoking ten years earlier, whereas the other 5 were nonsmokers. In case number 5 the diagnosis of pulmonary hypertension was based upon echocardiography showing dilatation and hypertrophy of the right ventricle with inverse septal movement and with Doppler a peak systolic, right ventricular to right atrial, pressure gradient of 92 mmHg. In this case pulmonary hypertension had earlier occurred in a first-degree relative. In the other 5 patients the diagnoses was based upon right heart catheterization. Clinical and hemodynamic data are detailed in Table I. Five of our patients were receiving treatment with calcium channel blockers, 3 with anticoagulants, and 1 each with an ACE inhibitor and a diuretic.
Downloaded from ang.sagepub.com at PRINCETON UNIV LIBRARY on July 16, 2015
838 The control group in the hemorheologic study comprised 10 healthy nonsmoking subjects without medication, 5 men and 5 women, aged twenty to sixty-two (mean forty-one
years). Blood Rheology Measurements Blood samples were drawn in the morning from an antecubital vein with the subjects supine and fasting overnight. Anticoagulation was performed with EDTA, except for the samples bound for fibrinogen analysis where citrate was used. Plasma viscosity was measured in a capillary U-tube at 37 °C and expressed as the ratio between the passage times of plasma and water, ie, as plasma viscosity relative to the 2 viscosity of water of the same temperature . Whole blood viscosity was determined at natural hematocrit, as well as at a standardized hematocrit of 45070. All measurements were performed at 37 °C. A computer-controlled rotational viscometer was used, measuring at stepwise increasing shear rates from 0.8 s-’ to 40.0 s-’ and then with decreasing shear rates back to 0.8 S-1 as previously described.’ For measuring rec cell deformability, whole blood was centrifuged for ten minutes at 2500 g. The plasma and buffy coat were removed and discarded. Then 0.55 mL of the packed red cells were aspirated from the middle of the centrifuged red call column and washed twice in phosphate-buffered saline (PBS) of pH 7.4 and containing bovine serum albumin, 2.5 g/L. Before use the buffer was filtered through a 45 micrometer filter. TABLE II Blood
Rheology Parameters Controls (n 10) PPH patients (n 6) (n
(n =
Downloaded from ang.sagepub.com at PRINCETON UNIV LIBRARY on July 16, 2015
839
Finally the red blood cells were resuspended in the PBS to a hematocrit of 1 0to . Residual leukocytes in the erythrocyte suspensions were counted under the microscope. In all suspensions the leukocyte count was < 0.025 x 109/L. Blood cell filtration studies were finally performed with a filtrometer according to Dormandy et a14 with a negative pressure of 3 cm of water. Filters with a 5 micrometer pore diameter were used. In all filtration studies we used filters from the same batch. The time from blood sampling to finishing 5 the measurements did not exceed three hours in any case.5 For determining white cell deformability, centrifugation of whole blood was performed as for red cells. The buffy coat was aspirated completely from the cell column and suspended in 2 mL of plasma in a 5 mL disposable plastic tube with cap and placed in a holder at a horizontal position for fifteen minutes. During this time the red blood cells aggregated and sedimented to the tube wall. The tube was then tilted carefully to about 45 degrees and the plasma supernatant was removed and washed twice in phosphatebuffered saline (PBS) containing 1 mL plasma to 19 mL of PBS. Leukocytes were counted under the microscope and finally suspended to a concentration of 5 x 109/L. Filtration studies were performed in the same way as for red blood cells but with filters with a pore size of 8 micrometers. Plasma fibrinogen concentration was determined by a kit method. Total cholesterol concentration in plasma was measured by an enzymatic colorimetric method and HDL cholesterol concentration by the same method after chylomicrons, VLDL, and LDL were precipitated by addition of phosphotungstic acid and magnesium ions to the sample. Erythrocyte mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), hematocrit, hemoglobin concentration, and counts of erythrocytes and leukocytes were determined by routine methods. Heart Catheterization The indication for right heart catheterization was entirely clinical and in case affected the by viscosity studies. A balloon-tipped thermodilution catheter was introduced percutaneously via an antecubital vein and positioned in a pulmonary artery under fluoroscopic control. For measurements of systemic blood pressure an indwelling plastic cannula was placed in a brachial artery, usually on the contralateral side. Both catheters were connected to pressure transducers at the level of the anterior axillary line. Leads V 1, V4, and V6 were continuously monitored and recorded together with the pressures. Cardiac output was measured with a standard thermodilution equipment, and all cardiac output determinations represented the average of three or more measurements.
Right
Echocardiography Two-dimensional and M-mode echocardiography was performed with the patients in the left semilateral decubitus position. Tricuspid regurgitation flow was examined from the apex by a separate continuous wave Doppler transducer. Peak systolic right ventricular to right atrial pressure gradients were calculated from the peak systolic regurgitation 6 velocities as described elsewhere .
Downloaded from ang.sagepub.com at PRINCETON UNIV LIBRARY on July 16, 2015
840
Results The blood viscosity (Table II) at 37 °C was increased in patients with PPH as compared with the control subjects. At natural hematocrit (Fig. 1) the differences were statistically significant at all shear rates studied, 0.8, 2.3, 19.6, and 40.0 s-’ (p
