Fatigue-induced changes to the biaxial mechanical properties of glutaraldehyde-fixed porcine aortic valve leaflets.

Circumferential extension is a direct measure of the preservation of functional collagen crimp in the fibrosal layer of aortic valve leaflets. The aim of this study was to determine whether the elastic properties of zero-pressure, glutaraldehyde-fixed leaflets are changed by mechanical fatigue. Nine Medtronic Freestyle bioprostheses were subjected to 200x10(6) cycles of accelerated fatigue and then biaxially tested to quantify the elastic properties of the leaflets. At physiological load (60 Nm(-1)) the radial extensibility was approximately halved relative to controls (P<10(-4)); there were also lesser reductions in the circumferential extensions (P<.01). The pulsatile regurgitant volume showed no change relative to the control leaflets. The natural corrugations of the fibrosal layer were flattened by the fatigue cycling, but this was not related to an increase in the radial size of the leaflets. Valve competency was maintained.

Analysis of the effects of oxygen supply and demand on hypoxic fraction in tumors.

The extent of hypoxic regions in a tumor tissue depends on the arrangement, blood flow rate and blood oxygen content of microvessels, and on the tissue's oxygen consumption rate. Here, the effects of blood flow rate, blood oxygen content and oxygen consumption on hypoxic fraction are simulated theoretically, for a region whose microvascular geometry was derived from observations of a transplanted mammary andenocarcinoma (R3230AC) in a rat dorsal skin flap preparation. In the control state, arterial PO2 is 100 mmHg, consumption rate is 2.4 cm3 O2/100 g/min, and hypoxic fraction (tissue with PO2 < 3 mmHg) is 30%. Hypoxia is abolished by a reduction in consumption rate of at least 30%, relative to control, or an increase in flow rate by a factor of 4 or more, or an increase in arterial PO2 by a factor of 11 or more. These results suggest that reducing oxygen consumption rate may be more effective than elevating blood flow or oxygen content as a method to reduce tumor hypoxia.

Resistance to blood flow in microvessels in vivo.

Resistance to blood flow through peripheral vascular beds strongly influences cardiovascular function and transport to tissue. For a given vascular architecture, flow resistance is determined by the rheological behavior of blood flowing through microvessels. A new approach for calculating the contribution of blood rheology to microvascular flow resistance is presented. Morphology (diameter and length), flow velocity, hematocrit, and topological position were determined for all vessel segments (up to 913) of terminal microcirculatory networks in the rat mesentery by intravital microscopy. Flow velocity and hematocrit were also predicted from mathematical flow simulations, in which the assumed dependence of flow resistance on diameter, hematocrit, and shear rate was optimized to minimize the deviation between measured and predicted values. For microvessels with diameters below approximately 40 microns, the resulting flow resistances are markedly higher and show a stronger dependence on hematocrit than previously estimated from measurements of blood flow in narrow glass tubes. For example, flow resistance in 10-microns microvessels at normal hematocrit is found to exceed that of a corresponding glass tube by a factor of approximately 4. In separate experiments, flow resistance of microvascular networks was estimated from direct measurements of total pressure drop and volume flow, at systemic hematocrits intentionally varied from 0.08 to 0.68. The results agree closely with predictions based on the above-optimized resistance but not with predictions based on glass-tube data. The unexpectedly high flow resistance in small microvessels may be related to interactions between blood components and the inner vessel surface that do not occur in smooth-walled tubes.

Determination of local oxygen consumption rates in tumors.

At any location in a respiring tissue, partial pressure of oxygen (PO2) is influenced by the local oxygen consumption rate. Consumption rates in vascular tumor tissues have previously been estimated for macroscopic regions. Using oxygen electrodes, we measured PO2 profiles across microregions (87 microns to 286 microns wide) of tumors (R3230AC mammary adenocarcinoma) in a rat dorsal skin flap preparation and mapped adjacent microvessels. By comparing measured PO2 values with theoretical simulations, we deduced local consumption rates. Results for six profiles ranged from 0.83 to 2.22 cm3 O2/100 g/min. The mean (+/- SD) was 1.52 +/- 0.51 cm3 O2/100 g/min. This technique permits investigation of variations in consumption at a microregional level.

Analysis of oxygen transport to tumor tissue by microvascular networks.

We present theoretical simulations of oxygen delivery to tumor tissues by networks of microvessels, based on in vivo observations of vascular geometry and blood flow in the tumor microcirculation. The aim of these studies is to investigate the impact of vascular geometry on the occurrence of tissue hypoxia. The observations were made in the tissue (thickness 200 microns) contained between two glass plates in a dorsal skin flap preparation in the rat. Mammary adenocarcinomas (R3230 AC) were introduced and allowed to grow, and networks of microvessels in the tumors were mapped, providing data on length, geometric orientation, diameter and blood velocity in each segment. Based on these data, simulations were made of a 1 mm x 1 mm region containing five unbranched vascular segments and a 0.25 mm x 0.35 mm region containing 22 segments. Generally, vessels were assumed to lie in the plane midway between the glass plates, at 100 microns depth. Flow rates in the vessels were based on measured velocities and diameters. The assumed rate of oxygen consumption in the tissue was varied over a range of values. Using a Green's function method, partial pressure of oxygen (PO2) was computed at each point in the tissue region. As oxygen consumption is increased, tissue PO2 falls, with hypoxia first appearing at points relatively distant from the nearest blood vessel. The width of the well-oxygenated region is comparable to that predicted by simpler analyses. Cumulative frequency distributions of tissue PO2 were compared with predictions of a Krogh-type model with the same vascular densities, and it was found that the latter approach, which assumes a uniform spacing of vessels, may underestimate the extent of the hypoxic tissue. Our estimates of the maximum consumption rate that can be sustained without tissue hypoxia were substantially lower than those obtained from the Krogh-type model. We conclude that the heterogeneous structure of tumor microcirculation can have a substantial effect on the occurrence of hypoxic micro-regions.

Mathematical model of the filtration in the vascular network of the ophidian glomerulus.

The present work is a mathematical model of the fluid filtration in the glomerular network occurring in snakes. The model is based on the differential form of Starling's hypothesis and takes into account the angioarchitecture of the network and the behaviour on the microrheology of blood with nucleated red cells. The model predicts the hemodynamics and the transvascular fluxes in each vascular segment within the network. The model is applied to a vascular network of the glomerulus of the garter snake. A value of 0.593 microns/(s.mmHg) was determined for the hydraulic conductivity of the glomerular capillaries using the geometrical data of the network together with experimental data for the pressures and the blood flow rate reported in the literature. The analysis shows that the local filtration rates cover a wide range. In some of the vascular segments, the filtration leads to such a high increase in colloid-osmotic pressure that the level of the transvascular hydrostatic pressure difference is reached. Mathematical simulations of the variation of the glomerular blood flow rate due to vasoactivity of preglomerular arterioles show the effect on the filtration rate and the hemorheologic parameters.

Effects of bradykinin on the hemodynamics of tumor and granulating normal tissue microvasculature.

Bradykinin (BK) is an important endogenous mediator of microvascular flow modulation. Since the structure of the microcirculation is very different in tumor tissues than in normal tissues, bradykinin may elicit different responses in tumors. This study was designed to test the hypothesis that local administration of bradykinin increases blood flow preferentially in normal tissue relative to adjacent tumor tissue, resulting in a "vascular steal" phenomenon. Microvessel diameters (D), velocities (Vc), length densities, shear rates, and intermittent flow frequencies were measured every 10 min before, during, and after 40 min exposure to BK in rats with dorsal flap window chambers 9 days after tumor implantation. Separate studies were made of normal vessels outside the tumor margin, the hypervascular tumor periphery, and the tumor center. Bradykinin was administered with a suffusion medium flowing over the tissue at 1-2 ml/min with a BK concentration of 1.6 x 10(7) M. Administration of BK created five distinct changes in normal and tumor vessel function that varied over time, but coincidentally reached a maximum effect after 20 min exposure to BK. In normal vessels, increased Vc and D led to increased flow, which reached a peak 20 min after onset of suffusion with BK. In contrast, in centrally located tumor vessels, decreased D and Vc were observed in most vessels during the initial 10-20 min of suffusion. In addition, there was a significant increase in intermittent flow frequency in tumor central vessels, which peaked after 20 min of suffusion with BK. These five separate observations that coincided at 20 min of suffusion are consistent with a "vascular steal" phenomenon. The increase in normal microvessel D and Vc at 20 min suggests that BK causes vasodilation in arterioles. The coincident decrease in tumor microvessel D and Vc suggests that tumor feeding vessels are less able to respond to BK by vasodilating. The concomitant increase in intermittent flow frequency in tumor vessels suggests that a reduction in pressure drop occurred after 20 min exposure to BK, which is also consistent with "vascular steal." Since BK is also known to increase vascular permeability, it is possible that increases in interstitial fluid pressure brought on by exposure to BK contributed to the observed reduction in tumor blood flow. In normal vessels, reduced D and Vc, relative to peak values, were noted after 40 min suffusion with BK. Adherence of leukocytes to the vessel walls was prominent and microthrombi were also observed during this period. No evidence of such adhesion was seen in tumor vessels, although microthrombi were observed.(ABSTRACT TRUNCATED AT 400 WORDS)

Perivascular oxygen tensions in a transplantable mammary tumor growing in a dorsal flap window chamber.

Fischer 344 rats with R3230 Ac mammary carcinomas implanted in dorsal flap window chambers served as a model to obtain measurements of perivascular and stromal oxygen tension in normal and tumor tissues using Whalen recessed-tip microelectrodes (3- to 6-microns tip). Perivascular measurements were made adjacent to vessels with continuous blood flow. Thus the measurements and models provided are reflective of conditions leading to chronic hypoxia. Perivascular oxygen tensions averaged 72 +/- 13 mmHg in normal tissue vessels adjacent to tumor, 26 +/- 5 mmHg in tumor periphery, and 12 +/- 3 mmHg in tumor central vessels. There was a significant trend toward lower perivascular oxygen tensions in the tumor center (Kruskal-Wallis test, P = 0.002). A similar tendency was seen with a limited number of stromal measurements. Krogh cylinder models, which incorporate these data for perivascular oxygen tension, along with morphometric data obtained from the same tumor model suggest that hypoxic regions will exist between tumor vessels in the tumor center unless O2 consumption rates are well below 0.6 ml/100 g/min. The low perivascular measurements observed near the tumor center combined with the theoretical considerations suggest, for this model at least, that tissue oxygenation may best be improved by increasing red cell velocity and input pO2 and reducing oxygen consumption. The low perivascular oxygen tensions observed near the center also suggest that conditions conducive to increased red cell rigidity exist, that drugs which can decrease red cell rigidity could improve tumor blood flow and oxygenation, and that the endothelium of those vessels may be susceptible to hypoxia-reoxygenation injury.

Blood flow in microvascular networks. Experiments and simulation.

A theoretical model has been developed to simulate blood flow through large microcirculatory networks. The model takes into account the dependence of apparent viscosity of blood on vessel diameter and hematocrit (the Fahraeus-Lindqvist effect), the reduction of intravascular hematocrit relative to the inflow hematocrit of a vessel (the Fahraeus effect), and the disproportionate distribution of red blood cells and plasma at arteriolar bifurcations (phase separation). The model was used to simulate flow in three microvascular networks in the rat mesentery with 436,583, and 913 vessel segments, respectively, using experimental data (length, diameter, and topological organization) obtained from the same networks. Measurements of hematocrit and flow direction in all vessel segments of these networks tested the validity of model results. These tests demonstrate that the prediction of parameters for individual vessel segments in large networks exhibits a high degree of uncertainty; for example, the squared coefficient of correlation between predicted and measured hematocrit of single vessel segments ranges only between 0.15 and 0.33. In contrast, the simulation of integrated characteristics of the network hemodynamics, such as the mean segment hematocrit or the distribution of blood flow velocities, is very precise. In addition, the following conclusions were derived from the comparison of predicted and measured values: 1) The low capillary hematocrits found in mesenteric microcirculatory networks as well as their heterogeneity can be explained on the basis of the Fahraeus effect and phase-separation phenomena. 2) The apparent viscosity of blood in vessels of the investigated tissue with diameters less than 15 microns is substantially higher than expected compared with measurements in glass tubes with the same diameter.

Heterogeneity in tumor microvascular response to radiation.

Viable hypoxic cells have reduced radiosensitivity and could be a potential cause for treatment failure with radiotherapy. The process of reoxygenation, which may occur after radiation exposure, could increase the probability for control. However, incomplete or insufficient reoxygenation may still be a potential cause for local treatment failure. One mechanism that has been thought to be responsible for reoxygenation is an increase in vascular prominence after radiation. However, the effect is known to be heterogeneous. In this study, tumor microvascular hemodynamics and morphologies were studied using the R3230 Ac mammary adenocarcinoma transplanted in a dorsal flap window chamber of the Fischer-344 rat. Measurements were made before and after (at 24 and 72 hr) 5-Gy radiation exposure to assess microvascular changes and to explore possible explanations for the heterogeneity of the effect. There was considerable heterogeneity between tumors prior to radiation. Vascular densities ranged from 67 to 3000 vessels/mm3 and median vessel diameters from 22 to 85 microns. Pretreatment perfusion values varied by a factor of six. In irradiated tumors, conjoint increases in both vascular density and perfusion occurred in most tumors, although the degree of change was variable from one individual to the next. The degree of change in density was inversely related to median pretreatment diameter. Relative change in flow, as predicted by morphometric measurements, overestimated observed changes in flow measured hemodynamically. These results support that heterogeneity in tumor vascular effects from radiation are somewhat dependent on pretreatment morphology as well as relative change in morphology. Since changes in flow could not be completely explained by morphometric measurements, however, it is likely that radiation induced changes in pressure and/or viscosity contribute to the overall effect. Further work in this laboratory will investigate these hypotheses.

Morphologic and hemodynamic comparison of tumor and healing normal tissue microvasculature.

The purpose of this study was to compare microvascular morphometric and hemodynamic characteristics of a tumor and granulating normal tissue to develop quantitative data that could be used to predict microvascular characteristics which would be most likely associated with hypoxia. The dorsal flap window chamber of the Fisher 344 rat was used to visualize the microvasculature of 10 granulating and 12 tumor (R3230 AC adenocarcinoma) tissues at 2 weeks following surgical implantation of the chamber. Morphometric measurements were made from photomontages and video techniques were used to assess red cell velocities in individual vessels. The percent vascular volume of both tissues was close to 20%, but significant differences were noted in other morphometric and hemodynamic measurements. Individual vessel dimensions (length and diameter) in tumors averaged twice as large as those in granulating tissues. Furthermore, red cell velocities were twice as high in tumors as in granulating tissues. In addition to these large differences in average values, there was significant heterogeneity in tumor microvascular morphometry, indicating spatial nonuniformity compared with the granulating tissue. Approximations of vessel spacing, indicated an average of 257 and 118 microns in tumors and granulating tissues, respectively. Vessel densities were four times greater in granulating tissues than in tumor tissues. These results indicated that intervessel distances were more likely to result in hypoxia in tumors, especially considering the wide variability in that tissue. Analysis of flow branching patterns showed that vascular shunts occurred frequently in vessels ranging from 10 to 90 microns in diameter. The results of this study indicate, in this tumor model, that conditions such as low vascular density, vascular shunts, excessive vascular length and/or low red cell velocity exist to a greater extent than the granulating tissue control. These conditions are likely to be conductive to the development of hypoxia.

Temporal effects of 5.0 Gy radiation in healing subcutaneous microvasculature of a dorsal flap window chamber.

The temporal effects of 5.0 Gy of radiation on healing subcutaneous microvasculature were studied using a window chamber in the dorsal flap of the Fischer-344 rat. Microvascular function was assessed by morphometric and dynamic flow measurements which were made prior to and at 24 and 72 h after exposure. A comparison was made between chamber preps that were 3 and 14 days postsurgery. The hypothesis of the study was that the older preparation would be more refractory to damage by radiation. Both unirradiated preparations showed an increase in capillary numbers over the period of observation, while irradiated preps had a reduction, especially in vessels less than 50 microns in diameter. Red cells velocities increased by 20-100% in those vessels which survived the radiation exposure, indicating that tissue oxygen tensions might be preserved in spite of a loss of vasculature. These results explain the need for both morphologic and dynamic flow measurements when assessing the effect of therapeutic intervention on microcirculatory function. Further studies are underway to identify a fully mature capillary bed in this model, since it is apparent that capillary growth is continuing in the 14-day preparation.

Transvascular exchange of fluid and plasma proteins.

A theoretical model of transvascular exchange of fluid and plasma proteins in the microcirculation is developed based on fundamental laws of the fluid mechanics and on phenomenological transport equations of the irreversible thermodynamics. Intravascular axial changes of the pressure, flow and plasma protein concentration are taken into account as well as axial gradients of vascular permeability. Proper nondimensionalization of the resulting equations leads to the identification of dimensionless parameters which combine the transport characteristics of the endothelial wall and the intravascular flow resistance. In the theory, the dependence of the reflection coefficient on the transport coefficients of the vascular wall and on the plasma protein concentration is established. The model is applied to the cat mesentery and the rat intestinal muscle. The numerical simulations indicate that taking into account vascular protein permeability yields considerable differences in the axial distribution of the plasma protein concentration and transvascular fluxes in comparison with the case of protein impermeability of the endothelial wall. The results show that the maximum of the transvascular fluid and plasma protein movement resides at the site of the small venules while a minimum of the exchange occurs at the site of the midcapillaries.

Vasomotion and transvascular exchange of fluid and plasma proteins.

The effect of spontaneous arteriolar vasomotion on the transvascular exchange of fluid and plasma proteins has been studied theoretically. The model combines fluid dynamic principles with a phenomenological approach to transvascular exchange on the basis of irreversible thermodynamics. The analytical treatment of the intravascular flow together with consideration of local changes of the morphology and vascular membrane characteristics makes it possible to determine the spatial and temporal variation of transvascular fluid and plasma protein fluxes. Since the undulations of the arteriolar diameter have low frequencies (3 to 10 per cycle), the transvascular fluxes are in phase with the vasomotion. The model combines hemodynamics, microhemorheology and mass transfer at the vessel wall and provides pressures, velocities, plasma protein concentration, hematocrit and apparent blood viscosity at any position in the microvascular bed at any time. In addition, global parameters as the total filtration rate and the total transvascular mass flow rate of plasma proteins are determined as functions of time. Numerical results are obtained for the cat mesentery with the terminal arteriole exhibiting vasomotion. The vascular arrangement studied comprised a terminal arteriole, capillaries and a venule together with appropriate side branches. The results show dramatic temporal changes in the hemodynamic parameters. Fluid filtration occurs along the entire vascular length at all times except during vasoconstriction when small absorption rates occur on the venous side. The transvascular fluxes achieve maximum values either on the arteriolar or on the venular side depending on the moment of the vasomotion cycle. The transvascular exchange rates in the mid-capillaries are generally low.

Disposition of 1,2,3-trichloropropane in the Fischer 344 rat: conventional and physiological pharmacokinetics.

To investigate the disposition of 1,2,3-trichloropropane (TCP), [14C]-TCP was administered iv to male Fischer 344 rats. Unchanged TCP and total radiolabel were determined in tissues and excreta at varying intervals after administration. The compound was distributed and eliminated rapidly. Initial and terminal half-lives of unchanged TCP in the blood were 0.29 and 23 hr. Adipose tissue accumulated 37% of the dose within 15 min and retained more of the dose than any other tissue until 4 hr; most (69%) of the radiolabel in adipose tissue through 4 hr was unchanged TCP. After 4 hr, the liver contained the largest fraction of the dose, primarily as metabolites. Thus TCP disappeared from adipose tissue while metabolites appeared in liver and other tissues. Excretion was nearly complete (90% of the dose) in 24 hr and was predominantly via the urine (47% of the dose). Expiration was the only route by which unchanged TCP (5% of the dose) was excreted. In addition, 25% of the dose was expired as carbon dioxide. There were numerous other metabolites, none accounting for more than 10% of the dose. Nonvolatile metabolites were longer lived than the parent compound. On the basis of high water solubility, reaction with 2,4-dinitrofluorobenzene, and diminished radiolabel in bile of glycidol-treated rats, glutathione conjugation is suggested as an important metabolic route for TCP. A physiological pharmacokinetic model was developed to describe the time course of trichloropropane concentration in tissues. The model demonstrates the possibility of using physiological and pharmacokinetic data to predict concentration-time relations for toxic compounds.

The effect of rate of heating or cooling prior to heating on tumor and normal tissue microcirculatory blood flow.

Single vessel responses to hyperthermia were studied in tumor and normal tissues using a transparent access window chamber. Rates of heating less than or equal to .68 degrees C/minute preserved relatively better vascular function in normal than tumor tissue. A rate of heating of 1.0 degrees C/minute lowered normal tissue statis temperatures so they were no different from tumor. Cooling to 30 degrees C prior to heating slowed normal arteriolar flows to less than 5% of 38 degrees C controls. Heating resulted in increased flow in those vessels, but maximum flows never exceeded 5% of flows achieved in similar vessels which were not cooled first. The implications of this work are that rate of heating and cooling prior to heating can alter normal tissue vascular response to heat in a way that could prove deleterious to maintaining efficient vascular function in that tissue relative to tumor.

Analysis of transcapillary exchange and intraluminal transport in the microocclusion of single capillaries.

A theoretical model, appropriate for data interpretation, is presented for the transcapillary fluid exchange and associated intraluminal hydrodynamic and solute transport occurring in the microocclusion of single capillaries. This analysis describes the spatial and temporal behavior of the intraluminal flow. A comparison of the theory with in vivo data suggests good qualitative agreement with that data and, further, the specification of filtration parameters in the model leads to good quantitative agreement regarding the displacement histories of erythrocytes. The model includes both natural and tagged colloidal osmotic influences and can, therefore, be used to represent experiments wherein the concentration of a dyed colloid is measured and related to fluid filtration. The relative merits of measuring colloidal concentration or erythrocyte displacement are discussed within the context of the model.

Flow of red blood cells in narrow capillaries: role of membrane tension.

A theoretical model is developed to describe blood flow in narrow capillaries, with inside diameters 3 microns to 6 microns. Each red blood cell is assumed to have axisymmetric geometry, and fixed surface area and volume. Cell velocities in the range 1 mm s-1 or higher are assumed, and the stress in the cell membrane is approximated by an isotropic tension. This tension is assumed to fall to zero at the concave trailing end of the cell, except in vessels whose diameter is near the minimum for passage of the cell. In the latter case, a separate analysis is used, in which the cell is effectively rigid and fully distended at each end. Lubrication theory is used to describe the plasma flow in the narrow gap between the cell and the vessel wall. Good agreement is obtained between predicted values of the tube hematocrit and apparent viscosity and published experimental values for these parameters.