Near-Wall μ-PIV Reveals a Hydrodynamically Relevant Endothelial Surface Layer in Venules In Vivo

Abstract

High-resolution near-wall fluorescent microparticle image velocimetry (micro-PIV) was used in mouse cremaster muscle venules in vivo to measure velocity profiles in the red cell-depleted plasma layer near the endothelial lining. micro-PIV data of the instantaneous translational speeds and radial positions of fluorescently labeled microspheres (0.47 microm) in an optical section through the midsagittal plane of each vessel were used to determine fluid particle translational speeds. Regression of a linear velocity distribution based on near-wall fluid-particle speeds consistently revealed a negative intercept when extrapolated to the vessel wall. Based on a detailed three-dimensional analysis of the local fluid dynamics, we estimate a mean effective thickness of approximately 0.33 micro m for an impermeable endothelial surface layer or approximately 0.44 micro m assuming the lowest hydraulic resistivity of the layer that is consistent with the observed particle motions. The extent of plasma flow retardation through the layer required to be consistent with our micro-PIV data results in near complete attenuation of fluid shear stress on the endothelial-cell surface. These findings confirm the presence of a hydrodynamically effective endothelial surface layer, and emphasize the need to revise previous concepts of leukocyte adhesion, stress transmission to vascular endothelium, permeability, and mechanotransduction mechanisms.

Microviscometry reveals reduced blood viscosity and altered shear rate and shear stress profiles in microvessels after hemodilution

“…Fluorescent -PIV (21, 22) was performed by using a method described in ref. 17. Fluoresbrite yellow-green microspheres (0.47 Ϯ 0.01 m, 1.05 g͞cm 3 ; Polysciences) were visualized by using stroboscopic doublef lash (5-to 16.67-ms apart; Strobex 11360, ChadwickHelmuth, El Monte, CA) epi-illumination, and recordings were made by using a VE-1000CD charge-coupled device camera (Dage-MTI, Michigan City, IN) on an S-VHS recorder (Panasonic, Secaucus, NJ).…”

“…The in vitro perfusion system consisted of a 3-ml feed reservoir and a horizontally mounted glass capillary tube (length, 27.5, 26.2, and 20 mm; i.d., 54.2, 50.7, and 81.0 m, respectively), which were connected by a microhematocrit tube (i.d., 1.2 mm) and silastic tubing (Dow-Corning), to a 10-ml downstream reservoir that could be manipulated vertically with a Vernier caliper (Nolan Supply, Syracuse, NY) (17,25). To account for the unavoidable mismatch of refractive indices between the perfusate and the inner tube wall, the radial position of every microsphere was corrected by using Snell's law (17).…”

“…To account for the unavoidable mismatch of refractive indices between the perfusate and the inner tube wall, the radial position of every microsphere was corrected by using Snell's law (17). Human blood samples were obtained from healthy volunteers by means of venipuncture, anticoagulated with heparin (final concentration, 10 units͞ml), and used within 2-4 h after withdrawal.…”

“…Measurements of H D have been attempted in microvessels in vivo either by micropipette aspiration and centrifugation (16), which is extremely cumbersome and feasible only in some tissues, or by a microphotometric method (13), which assumes the same Fåhraeus effect in glass tubes and microvessels of the same size. The accuracy of this assumption, however, is dubious in light of recent evidence of the influence of the endothelial surface layer (ESL) on plasma flow near the wall of microvessels in vivo (17,18). In addition to the difficulties associated with determining H T and H D in vivo, attempts (8,19) at measuring or estimating either axial pressure gradient or flow resistance accurately in microvessels have been unsuccessful in vivo.…”

“…The central tenet of our approach (20), which we hereafter shall refer to as the microviscometric method, is that distributions in the local viscosity, (r), as a function of the radial position, r, over the cross section of glass capillary tubes and microvessels, can be determined analytically from the crosssectional axial velocity distribution, v z (r), where v z (r) can be extracted from particle tracers in the flow by using intravital fluorescent microparticle image velocimetry ( -PIV) (17,21,22). In addition to the viscosity distribution, (r), we can use the microviscometric method to predict quantitatively various flow parameters in microvessels in vivo, including axial pressure gradient, dp͞dz; volume flow rate, Q; and the relative apparent blood viscosity, rel , defined as the ratio of steady volume-flow rates per unit pressure drop of blood plasma relative to whole blood.…”