What Does Increased Hematocrit Mean Blood Pressure
Am J Respir Cell Mol Biol. 2008 Feb; 38(2): 135–142.
Lowering of Blood Pressure level by Increasing Hematocrit with Non–Nitric Oxide–Scavenging Cerise Blood Cells
Beatriz Y. Salazar Vázquez
aneFaculty of Medicine, Universidad Juárez del Estado de Durango, Durango, Durango, Mexico; 2Section of Bioengineering, University of California, San Diego, La Jolla, California; and 3La Jolla Bioengineering Plant, La Jolla, California
Pedro Cabrales
iFaculty of Medicine, Universidad Juárez del Estado de Durango, Durango, Durango, Mexico; 2Department of Bioengineering, University of California, San Diego, La Jolla, California; and threeLa Jolla Bioengineering Institute, La Jolla, California
Amy Thou. Tsai
iFaculty of Medicine, Universidad Juárez del Estado de Durango, Durango, Durango, Mexico; 2Department of Bioengineering, Academy of California, San Diego, La Jolla, California; and 3La Jolla Bioengineering Constitute, La Jolla, California
Paul C. Johnson
1Kinesthesia of Medicine, Universidad Juárez del Estado de Durango, Durango, Durango, Mexico; 2Section of Bioengineering, University of California, San Diego, La Jolla, California; and 3La Jolla Bioengineering Institute, La Jolla, California
Marcos Intaglietta
1Faculty of Medicine, Universidad Juárez del Estado de Durango, Durango, Durango, United mexican states; iiDepartment of Bioengineering, University of California, San Diego, La Jolla, California; and 3La Jolla Bioengineering Institute, La Jolla, California
Received 2007 Mar eight; Accepted 2007 Jul 9.
Abstract
Isovolemic exchange transfusion of 40% of the claret volume in awake hamsters was used to replace native red blood cells (RBCs) with RBCs whose hemoglobin (Hb) was oxidized to methemoglobin (MetHb), MetRBCs. The substitution maintained abiding blood volume and produced different last hematocrits (Hcts), varying from 48 to 62% Hct. Mean arterial pressure (MAP) did non modify after exchange transfusion, in which 40% of the native RBCs were replaced with MetRBCs, without increasing Hct. Increasing Hct with MetRBCs lowered MAP by 12 mm Hg when Hct was increased 12% above baseline. Further increases of Hct with MetRBCs progressively returned MAP to baseline, which occurred at 62% Hct, a 30% increase in Hct from baseline. These observations show a parabolic "U" shaped distribution of MAP against the alter in Hct. Cardiac alphabetize, cardiac output divided by trunk weight, increased between 2 and 17% above baseline for the range of Hcts tested. Peripheral vascular resistance (VR) was decreased eighteen% from baseline when Hct was increased 12% from baseline. VR and MAP were higher up baseline for increases in Hct higher than 30%. Yet, vascular hindrance, VR normalized by claret viscosity (which reflects the contribution of vascular geometry), was lower than baseline for all the increases in Hct tested with MetRBC, indicating prevalence of vasodilation. These suggest that astute increases in Hct with MetRBCs increase endothelium shear stress and stimulate the product of vasoactive factors (e.g., nitric oxide [NO]). When MetRBCs were compared with functional RBCs, vasodilation was augmented for MetRBCs probably due to the lower NO scavenging of MetHb. Consequently, MetRBCs increased the viscosity related hypotension range compared with functional RBCs as NO shear stress vasodilation mediated responses are greater.
Keywords: blood pressure, shear stress, NO bioavailability, hematocrit, plasma layer
Increased hematocrit (Hct) above baseline is normally associated with top of systemic blood pressure level due to the increase in blood viscosity. These effects were found in studies in which Hct was increased 40% or more above baseline (i–three). However, a recent study by Martini and coworkers (4) showed that mean arterial pressure (MAP) increases merely when Hct is increased 20% above baseline using functional RBCs, and that MAP decreased when the change in Hct were less than xx%. In this range, cardiac output (CO) also increased and peripheral vascular resistance (VR) decreased (4, v).
Co-ordinate to Martini and colleagues (4), the cause of this paradoxical effect is that acute augmentation in Hct directly increases shear stress on the endothelium. This phenomenon may be due, in part, to the increase in viscosity and the decrease in cell-free layer (plasma layer) width, affecting blood properties and the endothelium interface, where shear signals are produced.
Increased endothelial shear stress promotes production of endothelium vasoactive and nonvasoactive factors (vi). Knockout mice deficient in endothelial nitric oxide (NO) synthases, and hamsters treated with Due north (G)-nitro-50-arginine methyl ester (l-NAME), did not lower MAP at Hct, whereas wild-blazon mice and untreated hamsters showed maximal reduction in MAP. These findings led to the conclusion that MAP decreased equally the production of vasodilation mediated by endothelial NO. This explains that the subtract in VR is proportional to the initial increase in Hct; however, this is eventually counteracted past the increase in VR due to the increase in blood viscosity. There is a point at which the vasodilator upshot due to the increase in shear stress no longer compensates for the enhance in viscosity, and VR and MAP are increased above baseline.
Evidence for a direct link between blood viscosity, shear stress, the product of NO, and vasodilation was reviewed by Smieško and Johnson (vii), who showed that increasing menstruum locally in arterioles (and therefore shear stress) caused "catamenia-dependent vasodilation." de Wit and coworkers (eight) showed that elevating plasma viscosity (and presumably shear stress) causes NO-mediated dilation in hamster skeletal muscle. Tsai and colleagues (9) showed that increasing plasma viscosity in extreme hemodilution in the hamster window chamber model increased shear stress, menses, and perivascular NO (measured with NO-specific microelectrodes). Shear stress also elicits the production of prostacyclin; withal, this mediator appears to provide residual vasodilatory effects compared with those that are NO mediated (10).
Changes in Hct may also affect NO bioavailability due to changes in NO scavenging by blood hemoglobin (Hb). The width of the plasma layer should decrease when Hct increases, bringing reddish blood cells (RBCs) closer to the endothelium, enhancing NO scavenging and counteracting effects of increased NO production (11, 12). Increasing Hct with not–oxygen-conveying (and therefore not–NO-scavenging) RBCs should extend the positive remainder of vasodilation and the range over which the increase in Hct lowers MAP compared with oxygen functional RBCs.
In this report, nosotros test the hypothesis that increasing the Hct using RBCs with decreased NO scavenging will extend the range in which increases in Hct cause hypotension when compared with similar increases in Hct using functional RBCs. NO scavenging by the native RBCs was reduced by isovolemic exchange transfusion of 40% of the animals' blood volume (BV) in the hamster window bedchamber model with MetRBCs, RBCs with Hb previously oxidized to methemoglobin (MetHb). MetHb has a v,000-fold decreased reaction rate with NO relative to either oxy or deoxyHb (13–xv). The substitution transfusion did non touch on BV, and reduced native oxygen carrying capacity by 40%, and to a higher place the level at which the systemic oxygen supply falls below the metabolic requirements of hamsters.
MATERIALS AND METHODS
Creature Preparation
Investigations were performed in 55- to 75-thou male person Golden Syrian Hamsters (Charles River Laboratories, Boston, MA). Animal handling and intendance were provided post-obit the procedures outlined in the Guide for the Care and Employ of Laboratory Animals (National Enquiry Council, 1996). The study was approved by the local Animal Subjects Committee. The window bedroom model is widely used for microvascular studies in the unanesthetized land, and the complete surgical technique is described in item elsewhere (16, 17). Briefly, hamsters were prepared for chamber implantation with a fifty mg/kg intraperitoneal injection of sodium pentobarbital anesthesia. Afterwards hair removal, sutures were used to lift the dorsal skin away from the creature, and 1 frame of the chamber was positioned on the brute's back. A bedroom consists of ii identical titanium frames with a xv-mm round window. With the aid of backlighting and a stereomicroscope, one side of the skinfold was removed, following the outline of the window, until only a thin layer of retractor muscle and the intact subcutaneous skin of the opposing side remained. Saline and then a cover glass were placed on the exposed skin held in place past the other part of the sleeping accommodation. The intact skin of the other side was exposed to the ambient environment.
The creature was allowed at to the lowest degree 2 days for recovery before catheter implantation. The animal was anesthetized again with sodium pentobarbital. Arterial and venous catheters (PE-fifty) were implanted in the carotid artery and jugular vein. The catheters were filled with a heparinized saline solution (30 IU/ml) to ensure their patency at the time of the experiment. Catheters were tunneled under the skin and exteriorized at the dorsal side of the neck, where they were attached to the sleeping accommodation frame with tape. The window chamber grooming was used in these experiments considering it provides a well-established animal model to compare results with those of a previous written report past Martini and colleagues (4). The window bedroom keeps the catheters out of attain of the fauna during the recovery menses. Experiments were performed three days afterward the initial surgery, and only with animals that passed established systemic inclusion criteria.
Inclusion Criteria
Animals were suitable for the experiments if systemic parameters were within normal limits, namely, heart rate (Hour) greater than 340 beats/minute, MAP greater than 80 mm Hg and less than 125 mm Hg, systemic Hct greater than 45%, and PaO2 greater than 50 mm Hg.
Preparation of Functional RBCs and RBCs Containing MetHb
Functional RBCs (oxygen functional RBCs).
RBCs were obtained every bit described by Martini and coworkers (4). Briefly, claret was nerveless from a donor in a heparinized syringe, centrifuged 5 minutes at ii,000 rpm, and buffy glaze was discarded and packed RBCs were stored. Packed cells Hct was adapted to the desired Hct past dilution using plasma from the donor.
MetRBCs (accelerated oxidation via nitrite).
Oxidized RBCs were prepared using the RBCs nerveless from donor. RBCs were resuspended in an equivalent amount of normal saline and mixed gently for 2 minutes with sodium nitrite (100 μl of i G sodium nitrite per 5 ml of resuspended RBCs). Cells were and then washed three times using heparinized saline and centrifuged at two,100 rpm. MetHb-loaded RBCs (MetRBC) were stored at iv°C. Aliquots of these cells were tested, and but those cells with 95 to 100% MetHb were used. To measure MetHb, nosotros nerveless approximately fifty μl of claret in microhematocrit tubes. MetHb was adamant using a Co-oximeter (IL 682; Instrumentation Laboratory, Lexington, MA). Calibration was ensured using standard levels at 5.2, 2.6, and i.ii% MetHb (RNA Medical, Bayer Diagnostics, Medfield, MA). In normal conditions, the concentration of cell-free Hb in the hamster is lower than 0.08 g/dl, and jail cell-free MetHb cannot exist detected. MetRBCs were re-suspended in fresh frozen plasma (FFP) taken from hamster donors to produce the desired Hct.
MetRBC (autoxidation).
A second method was used to gear up MetRBCs, without adding nitrite. Briefly, previously washed RBCs were incubated fully oxygenated at 37°C for 36 to 48 hours. MetRBCs produced this way were tested for MetHb levels, every bit described before, and only those with 85 to 100% MetHb were used. RBC manipulation (pipette, reaction, and transfer) was performed in a laminar flow hood for sterility.
Isovolemic Exchange Transfusion with Functional RBCs and MetRBCs
Exchange transfusion was 40% of the estimated animals' BV, 7% of the animals' body weight (BW). Since animate being weights ranged between 55 and 75 g, their BVs ranged between 3.9 and v.iii ml, and their bodily exchange transfusion volumes ranged between i.4 and 2.1 ml, and varied between the animals. The RBCs were infused using a dual syringe pump ("33" syringe pump; Harvard Apparatus Inc., Holliston, MA) into the jugular vein catheter at a rate of 100 μl/minute. Blood was simultaneously withdrawn from the carotid avenue catheter at the same rate.
Systemic Parameters and Blood Chemistry
MAP and 60 minutes were recorded continuously (MP 150; Biopac Organization, Santa Barbara, CA) except during the actual blood exchange. Hct was measured from centrifuged arterial blood samples taken in heparinized capillary tubes (Readacrit Centrifuge; Clay Adams, Segmentation of Becton-Dickinson, Parsippany, NJ). Hb content was adamant spectrophotometrically from a single drop of blood (B-Hemoglobin; Hemocue, Stockholm, Sweden). Arterial blood was collected in heparinized glass capillaries (0.05 ml) from the carotid catheter and immediately analyzed for PaO2 , PaCO2 , and pH (Blood Chemical science Analyzer 248; Bayer, Norwood, MA). The comparatively low PaO2 and loftier PaCOtwo of these animals is a result of their accommodation to a fossorial surround.
Measurements of Blood Viscosity
Claret was nerveless in heparinized syringes and analyzed for viscosity in a cone/plate viscometer (DV-II plus; Brookfield Engineering Laboratories, Middleboro, MA). Viscosities are reported at 150s−one shear rate.
Blood Rheological Changes Due to Nitrite-Accelerated Oxidation
Blood samples nerveless from donors were prepared as described before for exchange transfusion (functional RBCs and MetRBCs), then mixed with heparinized fresh blood. Last Hct of the samples was fixed to 52%. These samples consisted of either 50% fresh heparinized blood mixed with either 50% RBCs or 50% MetRBCs, and viscosity–shear rate relations were measured using a cone and plate viscometer (DV-Ii plus; Brookfield). Studied shear rates were in the range of x to 750 south−1.
Cardiac Output Measurements
CO was measured with the modified thermodilution technique described by Cabrales and coworkers (18). Due to volume improver with each CO measurement (150 μl of saline), measurements were fabricated twice in the same animal. Cardiac index (CI) is defined equally the measured CO divided past the BW, and used because it eliminates CO animal size dependence.
Measurement of Plasma Layer Width
The jail cell-free layer between the erythrocyte column and the vessel was continuously adamant from video images of microcirculatory vessels (xix). In this technique, the light intensity of a linear array of pixels perpendicular to the vessel axis is continuously recorded from a video image of a microcirculatory vessel (FASTCAM ultima SE, framing rate upward to 4,500/2d; Photron, San Diego, CA). An optical threshold level is used to found the interface between the cell-free layer and the erythrocyte column on the line intensity information forth a perpendicular cross-section of a claret vessel in the video image (nineteen). The distance between this optical threshold and the interface betwixt the endothelial surface and the plasma is measured continuously using image analysis software (MATLAB; Mathworks, Natick, MA).
Plasma Nitrite Concentration
Nitrite plasma concentration was measured at baseline, at xxx minutes, and at 1 60 minutes after the commutation, to ensure the absence of increased nitrite levels as a result of the method used to produce the MetRBCs. The Griess reactant was added to catechumen nitrites into a colorimetric compound (twenty). Optical absorbance was determined at 540 nm (Lambda 20 UV/VIS spectrometer; Perkin Elmer, Foster Metropolis, CA).
Experimental Protocol
Unanesthetized animals were placed in a restraining tube. They were given thirty minutes to adjust to the tube environment before baseline measurements (MAP, HR, Hct, Hb, and claret gases). After assay of baseline parameters, baseline CO was measured. Animals were substitution transfused and followed for 90 minutes after the substitution. MAP and Hour measurements were taken every 30 minutes; each measurement represents the average of MAP over iv minutes. 1 60 minutes subsequently the terminate of the exchange transfusion protocol CO was measured again. Assessment of the effects of changes in MAP as a function of Hct were fabricated i hour after the end of the exchange, to avert artefacts created by the exchange transfusion protocol (which, although advisedly implemented, resulted in cardiovascular in minor perturbations that subsided subsequently 30 min). Hct and Hb were measured again 1 hour after the stop of substitution protocol. Blood oxygen-carrying chapters was measured spectrophotometrically every bit the difference between total Hb and MetHb, from blood samples obtained i hour after the exchange protocol. Blood for viscosity determinations was collected at the end of the observation catamenia.
Experimental Groups
Identical substitution transfusion protocol was used in all groups. MetRBC: animals exchange transfused with MetRBCs; OxyRBC: animals exchange transfused with functional RBCs (data previously published [4]). Boosted experiments to consummate like range of Hct variation for both groups were added to previous results (4). Shut to 60% of the native RBCs remained in the animals afterwards the exchange protocol. The MetRBC grouping had approximately 60% of the native RBC (∼ ten g/dl), which maintained oxygen carrying capacity (Figure 1).
Concentration of reduced hemoglobin (Oxy and DeoxyHb) in blood after increasing hematocrit (Hct) by exchange transfusion with MetRBCs. The concentration of total Hb and MetHb was measured spectrophotometrically; reduced Hb was calculated as the difference between total Hb and MetHb.
Comparing after 30% Increment in Hct and 50-NAME Handling
Changes in MAP after a target 30% increment in baseline Hct (effective Hct ∼ 62%) were compared betwixt animals transfused with functional RBCs and MetRBCs. These ii groups were subsequently treated with l-Name (30 mg/Kg, 20 μl/min continuous infusion intravenously), and 10 minutes after stabilization MAP was recorded (21).
Statistical Analysis
Results are presented as hateful ± SD unless otherwise noted. MAP changes are reported in absolute values. Changes in Hct are presented equally percent changes from baseline Hct. CI is also reported equally a change from baseline. Data on VR and vascular hindrance (VR divided past claret viscosity) are presented as relative to levels at baseline. A ratio of 1.0 signifies no change from baseline, while lower or college ratios are indicative of changes proportionally higher or lower than baseline. Comparing of baseline values from the 3 unlike groups of animals was performed using one-style ANOVA, and mail service hoc analyses were performed with the Bonferroni'southward multiple comparison test. Data within each group were analyzed using ANOVA for repeated measures and followed by the Bonferroni'southward multiple comparison test. Changes were considered statistically pregnant if P < 0.05. The data were fitted to second-lodge polynomials and the resulting curves were compared by means of the F-test, and considered to be different if the F-test indicated a significantly smaller sum of squares for the deviations in each private fit compared with the divergence in the fit to the pooled data (22).
RESULTS
Experiments were performed in 40 animals; 16 were used for the measurement of MAP as a function of changes in Hct, 9 were used to measure CO and calculated VR, and 4 were used to make up one's mind the concentration of nitrite in plasma. Three animals were used to obtain a 30% increase in Hct with MetRBCs, and were compared with four animals whose Hct was increased by 30% using functional RBCs (data added to previously published study [4], OxyRBC group). 4 boosted animals were used to make up one's mind the upshot of Hb oxidation; they were exchange transfused with MetRBC (produced by autoxidation) to increment Hct 12% above baseline. The average weight of animals was 63 ± 15 g.
Systemic parameters were measured at baseline and at 1 hr later on exchange transfusion. Hct, Hb, MAP, Hr, blood gas parameter, and claret viscosity after substitution with MetRBC are given in Table 1. Comparisons between systemic parameters for both report groups (MetRBC and OxyRBC) after 30% increase in Hct are given in Tabular array 2. As in other studies, hamsters nowadays a natural Hct variability, which in the present report ranged from 42 to 48%. To account for the variability of results and reduce the number of animals required to detect effects in the range found in previous studies, results are presented equally pct changes from baseline Hct. This may be somewhat confusing for Hct changes, since Hct is itself a fraction (unremarkably reported as a percentage). In this context, Hct should be considered as a concentration of RBCs in the volume of blood, the difference existence that the amount of "solute" (i.e., RBCs) is expressed every bit a volume rather than mass weight.
Tabular array 1.
LABORATORY PARAMETERS
| Baseline | ane h after Substitution | |
|---|---|---|
| Hct, % | 47.9 ± 2.0 | 54.seven ± vi.2* |
| Hb, 1000/dl | 14.8 ± 0.six | sixteen.8 ± ane.9* |
| MetHb, g/dl | 7.5 ± ane.9* | |
| MAP, mm Hg | 102.four ± vi.5 | 96.four ± seven.5* |
| Heart rate, bpm | 432 ± 35 | 446 ± 32 |
| PaO2 , mm Hg | 56.4 ± 5.half dozen | 62.4 ± 5.6* |
| PaCO2 , mm Hg | 54.ii ± 4.two | 51.6 ± 6.0 |
| Arterial pH | 7.344 ± 0.007 | seven.346 ± 0.008 |
| BE, mmol/l | 3.0 ± ane.4 | 2.2 ± 1.6 |
| Viscosity, cp | ||
| Blood† | 4.2 ± 0.iv | iv.vii ± 0.five* |
| Plasma† | 1.2 ± 0.one | one.2 ± 0.1 |
| COP, mm Hg | 17.8 ± one.vi | 17.half dozen ± 1.viii |
TABLE 2.
LABORATORY PARAMETERS AFTER 30% Increase IN HCT
| MetRBC | OxyRBC | |||
|---|---|---|---|---|
| Baseline | i h | Baseline | 1 h | |
| Hct, % | 48.3 ± 0.vi | 62.1 ± i.4 | 47.8 ± i.3 | 61.2 ± ane.6 |
| Hb, g/dl | 14.7 ± 0.4 | eighteen.7 ± 0.ix | 14.5 ± 0.5 | 18.iv ± 0.viii |
| MetHb, thousand/dl | eight.iv ± 1.0 | |||
| MAP, mm Hg | 102.4 ± 6.five | 96.4 ± 7.5 | 105.0 ± 2.4 | 115.2 ± 2.4 |
| Middle rate, bpm | 441 ± 22 | 426 ± 21 | 447 ± 26 | 430 ± 24 |
| PaO2 , mm Hg | 66.5 ± 8.2 | 72.0 ± 2.9 | 52.5 ± 6.iv | 69.1 ± iv.4 |
| PaCO2 , mm Hg | 55.7 ± seven.3 | 52.1 ± two.3 | 48.2 ± five.half dozen | 51.seven ± 5.half dozen |
| Arterial pH | 7.371 ± 0.014 | seven.335 ± 0.017 | 7.355 ± 0.051 | seven.343 ± 0.037 |
| Exist, mmol/l | 6.0 ± 4.9 | i.3 ± 0.6 | 2.7 ± 2.6 | 2.3 ± 0.8 |
| Viscosity, cP | ||||
| Blood* | 6.vi ± 0.five | 6.5 ± 0.4 | ||
Claret Oxygen-Carrying Chapters after Exchange Transfusion
In the MetRBC grouping, blood maintained approximately sixty% of baseline oxygen-carrying chapters (9.iv ± 0.5 g/dl) for all the experiments. Nosotros strived to maintain similar oxygen-conveying capacity; the terminal consequence had some variability, and in two instances Hb was reduced slightly below 60% of baseline, every bit shown in Effigy ane.
Nitrite Levels
Nitrite levels were not dissimilar between baseline and 1 hour after the exchange with MetRBCs (baseline 3.six ± ane.2 μM, 1 hour later exchange 3.9 ± 1.6 μM).
MAP Changes later Increasing Hct with MetRBCs
MAP did not change from baseline when exchange transfusion did not modify baseline Hct, although, forty% of native RBCs were replaced with MetRBCs. Increasing the overall Hct above baseline using higher Hct in the infused MetRBCs during the exchange transfusion caused decreases in MAP proportional to the increases in Hct. MAP reached a minimum and and then increased every bit Hct was progressively increased. Increasing Hct betwixt 0 and 30% with MetRBCs (45 ± 3% at baseline versus 59 ± three% maximum attained after exchange transfusion) while keeping total BV abiding decreased MAP between 1 and 17 mm Hg of baseline, with the maximum interpolated pressure drop of 12 mm Hg based on a second-order polynomial occurring when Hct was increased to 12% of baseline (Figure 2). MAP increased to a higher place baseline when Hct was increased 30% above baseline (59% Hct). MAP changes occurred xxx to 60 minutes after exchange transfusion and remained stable for the whole observation menstruation. MAP and MetHb concentration were adamant at 1 hr subsequently the stop of the exchange transfusion. From previous studies in the same preparation, it was found that MetHb is reduced at the rate of 10%/hr (23). Changes in MAP versus percentage of Hct increase were fitted past a 2d-order polynomial (R 2 = 0.87). The maximal decrease in MAP was not dissimilar betwixt MetRBCs produced via nitrite-accelerated oxidation or autoxidation, suggesting no detectable upshot of the remaining nitrite contamination in the infused MetRBCs (Figure 2). MAP responses to small increases in Hct studied past Martini and colleagues using functional RBCs (OxyRBC group) are shown in Figure 3, where they are compared with the results of the present report (4).
Changes in hateful arterial claret pressure (MAP) and cardiac index (CI) every bit a function change in Hct using MetRBCs. In each example at least 40% of the native RBCs were exchanged with MetRBCs. Increased Hct above baseline values was attained using boosted MetRBCs. Solid circles: in the range between x and fifteen% increases in Hct from baseline were obtained with MetRBC produced by autoxidation of the Hb. No statistical difference was detected after increasing Hct by 10 to 15% using MetRBC, produced past accelerated oxidation via nitrite or autoxidation (incubation), P = 0.28. Normovolemia was maintained in all instances. The solid line shows the relation between MAP and Hct if blood viscosity is the only determinant of MAP.
Changes in MAP and CI as a function of the increment in Hct using MetRBCs. The information shown are compared with the information from the report of Martini and colleagues (iv, 5) using functional RBCs. Rectangular shadow expanse in the range between 27 and 30% increases in Hct from baseline shows the data used for direct comparing of the OxyRBC and MetRBC groups. Solid blackness thick line presents theoretical line in which MAP is merely a function of the variation in blood viscosity due to changes in Hct, causeless to be linear in this range as shown by Kameneva and colleagues (31). The maximal difference in blood pressure obtained using MetRBCs and functional RBCs is not statically significant (P < 0.10). The difference in CI is significant (P < 0.01). The shaded area shows the information obtained from exchange transfusing MetRBCs that were oxidized by exposure to ambient air, and the data used in Figure seven.
Cardiac Alphabetize
CI increased throughout the range of Hct changes, being approximately 12% higher than baseline. These data are fitted by a second-order polynomial (R two = 0.37) (Figures 2 and iii). CI and MAP returned to baseline values at approximately the same increment in Hct (∼ 30%).
Blood Rheological Changes
Viscosity of blood samples with Hct approximately 52% of either mix of 50/l functional RBCs and fresh claret or mix of 50/50 MetRBCs and fresh blood were non different betwixt shear rates of 10 and 450 south−1. At 750 s−1 the viscosities were 5.65 (50/fifty functional RBCs and fresh blood) and five.42 cp (50/50 MetRBCs and fresh blood), respectively. This divergence cannot be considered statistically significant, considering it is inside the 0.3-cp resolution limit of the viscometer.
Peripheral Vascular Resistance
VR obtained from the ratio MAP divided by CI was significantly decreased throughout the range of increased Hct. It reached a minimum at the Hct increase of 12% (Figure 4). Figure 5 shows the consequence of calculated vascular hindrance (VH), for both the MetRBC and OxyRBC groups. VH changes are mostly due to changes in vascular diameter and contained of changes in blood viscosity.
Changes in vascular resistance (VR) subsequently commutation transfusion with MetRBCs equally a office of the increase in Hct. Vascular resistance was computed from the data in Effigy two. Data obtained by increasing Hct with RBCs (OxyRBC) is reproduced from the report of Martini and coworkers (4, v). The deviation in VR between the 2 studies is statistically significant (P < 0.01). The solid line shows the relation between VR and Hct if blood viscosity is the but determinant of MAP.
VR independent of blood viscosity (Vascular hindrance, VH). This graph shows the overall conditions of vascular tone of the circulation indicating globally whether the apportionment is constricted or dilated. It provides an overall average, and may non correspond the perfusion of specific tissues. If blood viscosity were the only factor affecting VR, the relationship between VH and Hct should be a horizontal line.
Changes in MAP after 30% Increase in Hct with Functional RBCs and MetRBCs, and NO Synthase Effects (l-Proper name)
Independent experiments were fabricated with either functional RBCs or MetRBCs to increase baseline Hct by 30% (∼ 62% Hct). Equally shown in Effigy six, there was a significant deviation in MAP betwixt the two groups in support of the hypothesis that VR at the same Hct increase was significantly less for the MetRBC group. Treatment of each group with l-NAME increased MAP to identical levels in both groups. Changes in MAP elicited by l-Proper name advise that differences observed between groups were NO related. l-NAME eliminated the NO-vasodilatory responses observed in the OxyRBC group, although the change was more significant in the MetRBC grouping. This event can be interpreted to be indicative that in that location was less NO scavenging in the MetRBC group and therefore an increased vasodilatory reserve.
Comparison of the effects on MAP of increasing Hct past xxx% from baseline (final Hct ∼ 62%) using MetRBC according to the experimental protocol of this written report, and using functional RBCs (OxyRBC). 50-NAME was administered after cardiovascular conditions stabilized, and MAP was recorded equally information technology reached stable conditions.
Measurement of Plasma Layer Width
The plasma layer width was measured equally a part of arteriolar diameter at baseline Hct and when baseline Hct was increased by twenty%. As show in Figure vii, there is a statistical divergence in the measurement of the plasma layer width betwixt control and Hct augmented to twenty% of baseline. The dependence of plasma layer width on blood vessel bore was statistically significant for the two Hcts tested. The slopes of the two lines are not statistically significantly different; nonetheless, the difference in y-axis intercept is. Assuming that both lines have the same gradient, we estimate that increasing Hct past twenty% lowers the plasma layer width by near 0.3 μm in microvessels ranging from fifteen to 35 μm in diameter.
Plasma layer width equally a role of microvessel size (arterioles and venules) at baseline Hct (48% Hct), and when baseline Hct was increased by 20% (58% Hct) using RBCs. At that place is no statistical deviation between the slopes of the linear regressions; however, the correlation between plasma layer width versus vessel size is statistically pregnant for both Hcts. The boilerplate deviation betwixt plasma layer width betwixt baseline and 20% increase Hct is 0.3 μm, which is statistically significant (P < 0.05).
DISCUSSION
The main finding of this written report is that reducing oxygen-conveying chapters (without changing BV) by substituting 40% of native RBCs with non–oxygen-carrying RBCs, whose Hb was converted to MetHb (MetRBCs), did not change MAP or VR. Increasing Hct above baseline without changing BV with MetRBCs, thereby increasing the proportion of MetRBCs beyond the initial 40%, initially reduced MAP to the same extent equally in previous experiments using functional RBCs upwards to an increase in Hct of ∼ 12% (total Hct = 54%).
Augmenting Hct across 12% of baseline past increasing the Hct of the exchange transfused MetRBCs (while maintaining BV) reversed the decreased in MAP, which returned to baseline when the increase in Hct was around 30% in a higher place baseline (∼ 62% Hct), leading to a "U" shaped relationship between MAP and Hct. These furnishings were paralleled by a consequent elevation of CI; throughout the range of Hcts tested, CI returned to baseline values afterwards an increase in Hct of thirty% from baseline.
These results are qualitatively similar to those obtained by Martini and coworkers (4, 5) using functional RBCs, simply differ in some important quantitative features, since the range over which MAP is decreased versus the increment in Hct is significantly extended with the use of MetRBCs. In the studies of Martini and colleagues (four, 5) MAP decreased with an Hct increase up to 54% (an increase of 20% above baseline), while in the present report the range is extended to a 30% increase above baseline in Hct. The difference between results for MAP is not statistically meaning (P < 0.x), while that for CI and VR is pregnant (P < 0.01), every bit shown in Figures 3 and four.
Our aim was to increment Hct, and therefore blood viscosity, without increasing NO scavenging. We contemplated several approaches to hinder the intrinsic NO-scavenging chapters of Hb, and chose Hb oxidation equally the least probable to innovate artefacts, and suitable for short-duration experiments. We considered carboxyhemoglobin (COHb); however, carbon monoxide is released from Hb and the presence of CO in the circulation could exist potent vasodilator in the hamster model, as shown by Hangai-Hoger and colleagues (24). Another culling was inactivation of Hb gas transport carrying capacity using cyanide to form cyanomethemoglobin; nonetheless, this reaction affects RBCs membrane (increases fragility) and the cyanide cannot exist removed 100% from the treated blood.
When Hct was increased 12% above baseline (i.e., from 48% to 52% Hct) with additional MetRBCs (MetRBCs being now 53% of the total RBCs in circulation), the resulting decrease in MAP obtained with MetRBCs is identical to that obtained by increasing Hct with functional RBCs (OxyRBC group); however, CI does not change as much, causing a lesser modify in VR. Since Hct and therefore blood viscosity should exist the same in both atmospheric condition, it appears that MetRBCs do not produce an extra vasodilator effect beyond that induced by functional RBCs. In fact, in the range of Hcts leading to maximal MAP effects, the vasodilatory effect is reduced compared with that attained past functional RBCs.
Claret viscosity had a tendency to be somewhat lower (∼ 4%) for the claret samples that were l/50 MetRBCs/fresh claret when compared with samples of functional RBCs/fresh blood at the same Hct (∼ 52%), although the difference cannot considered significant due to resolution of the technique. Therefore, in principle, part of the deviation in response betwixt the two groups could be due to the lower viscosity due to the presence of MetRBCs. However, at the maximal Hct investigated when Hct was increased by 30% and l-Proper noun was administered, there was no difference in the final MAP. In this condition, NO-based vasodilatory mechanisms are not operational, and a deviation in blood viscosity should be evidenced by a change in MAP; however, this was not obtained (Figure 6).
Information technology could be argued that the fall in MAP over the extended range of Hct is due to the lowering of center function resulting from decreased oxygen commitment to the eye, as a consequence of the lowered intrinsic oxygen conveying RBCs in blood (ix.4 ± 0.5 thou/dl). This consequence is observed when claret oxygen-conveying chapters is lowered in hemodilution below the level that oxygen delivery is compensated by increased CO (3, 25). However, this may not be the full caption, since increasing Hct by thirty% with MetRBCs caused MAP and CI to be above baseline values, indicating that the heart was able to maintain MAP in the face of significantly increased VR due to the increased claret viscosity.
Increasing Hct more than xxx% from baseline may present as much as a 40 to 50% increase in blood viscosity, since in the high Hct range the relationship between blood viscosity and Hct is nonlinear (26). This increased viscosity should atomic number 82 to a proportional increase in VR and MAP; however, this was not observed in the MetRBC grouping. The absence of a major pressor response indicates that the increase in Hct leads to vasodilation that compensates for the related increase in viscosity.
The presence of pregnant vasodilation becomes evident when the increased blood viscosity is factored out from VR as shown in Effigy 5. This rendition of the data evidences the significant vasodilation caused by increasing Hct with functional RBCs and MetRBCs.
Lowering Hb concentration should lower NO scavenging co-ordinate to modeling analysis by Buerk (27). This effect would be important in the plasma free layer, which should reduce as Hct increases, thus bringing RBCs closer to the endothelial surface. Lowering the concentration of RBCs in the blood column would decrease NO scavenging, equally shown by the analysis of Chen and colleagues (28). We verified the changes of plasma free layer width due to the increase in Hct. Measurements were performed at baseline and later on an increase in Hct of 20% using functional RBCs. This value was chosen because this increase in Hct corresponded to levels at which MAP, CI, and VR return to baseline values (4, 5). Therefore, this could be defined as a situation in which the presumed NO bioavailability increases due to augmented shear stress and is balanced past NO scavenging and the increased blood viscosity. Changes in the plasma layer width are due to crowding of RBCs and should occur with the add-on of either functional RBCs or MetRBCs. We advise that decreasing the plasma layer width with RBCs increases NO scavenging. Conversely, if the plasma layer width is lowered past an increase in Hct by which approximately fifty% of RBCs do not scavenge NO, the resulting vasoactivity is diminished.
There was no effect on MAP or VR when native RBCs were decreased by 40% without changing Hct, a status in which, presumably, NO scavenging chapters should accept been contradistinct by the presence of MetRBCs. However, the event became apparent when Hct increased significantly. This result could exist explained if NO scavenging by RBCs is nonlinearly related to the size of the plasma layer, becoming prominent after a significant decrease of its width.
Results showing the decrease in VR and MAP as Hct increases should be directly related to the increase of shear stress in the blood/endothelium interface due to increased claret viscosity (29). This effect is directly related to catamenia-dependent vasodilatation (7), where increased shear stress is due to increased viscosity rather than blood flow velocity, with vasodilatation overcoming the increased viscosity. The subsequent increase of MAP and VR with the increase in Hct is due to viscosity eventually overcoming the effects of vasodilatation, and mayhap the increased NO scavenging due to the decreased plasma layer.
Comparison of Results with Functional RBCs and MetRBCs (OxyRBC and MetRBC Groups)
The difference in results between the present experiments and those previously made using RBCs (OxyRBC group) (4) would appear to be due to two effects: (1) the limitation on cardiac part due to increase Hct without an associated increase in oxygen-conveying capacity, and (2) reduction in NO scavenging due to lower concentration of reduced Hb (oxy and deoxy) as the plasma layer is reduced. These effects are observed as an increase in the extension of the Hct range which vasodilatation compensates for the increase in viscosity.
MAP and CI returned to baseline values at the Hct increase of thirty% with MetRBCs, corresponding to a blood viscosity increase of 30% (at Hct of 62% viscosity is ∼ six.58 cp). Therefore it can be causeless that the heart performance is not significantly limited by the reduction in oxygen-carrying chapters. Effects due to the increase in blood viscosity (product of the increase in Hct) should exist common to both groups (MetRBC and the OxyRBC), since endothelial role should be similarly afflicted in both experiments, unless there are other factors involved.
The increased intrinsic oxygen-conveying chapters via increased Hct has been shown to lead to an autoregulatory response, which is attributed to an oxygen-sensing mechanism that maintains oxygen supply at a constant level (30). Therefore, an caption is that the increment in MAP and VH observed in OxyRBC group was due to oxygen autoregulation, present to a lesser degree when Hct increases with functional RBCs that do not acquit oxygen. All the same, this machinery does not appear to exist operational for the initial small increases in Hct in the OxyRBC grouping, since oxygen transport capacity of the circulation (CI × ΔHct) increases to a maximum of approximately 45% before decreasing presumably due to increased viscosity and NO scavenging. Conversely, autoregulatory effects may non be operational in the MetRBC experiments, since oxygen ship capacity was abiding.
Our study provides indirect show for NO scavenging by RBCs in vivo in terms of changes in VR, a surrogate effect. An boosted finding in support of the influence of NO scavenging is that the OxyRBC group has a more pronounced increase in MAP than MetRBC, as evidenced by the steeper slope in the MAP and VR versus Hct plots (Figures 3 and 4). Both slopes are steeper than a theoretical line in which VR is only a function of the variation in blood viscosity due to changes in Hct, assumed to be linear in this range as shown by Kameneva and coworkers (31). An interpretation of this issue is that with RBCs there is an actress vasoconstrictor issue that increases VR beyond the effect due to blood viscosity. Furthermore, a reduced outcome, mostly due to the increase in viscosity and remaining NO scavenging from the native RBCs, was observed with MetRBCs.
These considerations are farther supported by the analysis of the trend of VH, which factors out the effect of blood viscosity on VR. Figure five shows that increasing Hct across that respective to the minimum of MAP for OxyRBC sharply increases VH, an effect due to vasoconstriction, while minimally impacting VH when the native RBC were replaced by MetRBCs. The significant deviation in the behavior of VH for both groups (OxyRBC and MetRBC) suggests the beingness of an extra vasoconstrictive effect due to increasing NO-scavenging capacity (blood Hb concentration) as plasma layer is decreased.
Nitrite concentration was measured to ensure the absence of increased nitrite concentration, a issue of the method used to produce MetRBCs. At that place was no statistical difference in this parameter between baseline and the time of measurement of MAP. Use of l-NAME showed no deviation in MAP between OxyRBC and MetRBC groups, after an Hct increase of 30% from baseline. An Hct increase of 10 to 15% from baseline using RBCs with Hb autoxidated to MetHb, produced similar decrease in MAP than the RBC, whose Hb was apace oxidized via nitrite. Thus, we conclude that at that place was no detectable outcome that resulted from the method of preparation of MetRBCs.
In summary, our findings prove that increasing Hct reduces VR, lowering MAP and increasing perfusion via the increase in CI. This effect takes place when Hct is increased with functional RBCs and using MetRBCs up to an Hct increase of approximately 12%, where information technology is maximal. MAP returns to baseline values when Hct is increased 20% for the OxyRBC group and 30% for the MetRBC grouping. Thus the lowering of MAP persists over a greater increase in Hct with MetRBCs than when RBCs were used. The effects up to these thresholds are due to increasing blood viscosity "enough" to cause shear stress–induced stimulation of vasodilation. In a higher place these thresholds, viscosity "swamps out" the vasodilator effects. Our findings on the occurrence of hypotension and reduction in VR at higher Hct with MetRBCs than with functional RBCs hemoconcentration are uniform with results predicted past mathematical modeling, where the effects are attributed to reduced NO scavenging in the plasma layer (11, 28). Furthermore, eliminating the effects of blood viscosity from the calculation of VR shows that at the Hct when MAP returns to baseline in that location is however meaning vasodilatation, showing that at the higher Hcts claret viscosity becomes the determinant factor in regulating VR. In decision, the relation between MAP, NO scavenging properties of Hb, blood viscosity, Hct, and blood flow appears to exist mediated inside the narrow claret tissue interface, the "plasma layer," suggesting that further analysis of the regulation of VR be directed at quantifying effects in this interface.
Acknowledgments
The authors give thanks Froilan P. Barra and Cynthia Walser for the surgical training of the animals.
Notes
This work was supported past the post-obit grants: R01-HL62354, R01-HL62318 and R01-HL64395 to M.I.; and R01-HL52684 to P.C.J.
Originally Published in Printing every bit DOI: 10.1165/rcmb.2007-0081OC on August xx, 2007
Conflict of Interest Statement: None of the authors has a financial relationship with a commercial entity that has an interest in the subject of this manuscript.
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What Does Increased Hematocrit Mean Blood Pressure,
Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2214674/#:~:text=Increased%20hematocrit%20(Hct)%20above%20baseline,baseline%20(1%E2%80%933).
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