Microcirculation, endothelium, and lymphatics最新文献

Monolayer cultures of rat lymphatic endothelial cell were obtained from explants of rat thoracic ducts. The cells displayed a typical cobblestone morphology, expressed von Willebrand factor (factor VIII-associated antigen) and angiotensin converting enzyme, and took up acetylated-low density lipoprotein, being indistinguishable by these criteria from blood vessel endothelial cells. The availability of these cells will facilitate the use of rat experimental models for functional studies of lymphatic endothelium.

The effect of topically applied papaverine (PAP) and norepinephrine (NE), on the microvascular pressure distribution was studied in the cremaster muscle of anesthetized rats. The cremaster muscle was exteriorized into a tissue bath containing Krebs bicarbonate buffer, and microvessel diameters and pressures were measured using a video caliper and the resistance servo-null method, respectively. Pressures were measured in the 1st through 4th branch order arterioles (1A to 4A) and 1st through 4th branch order venules (1V to 4V). Resistances were calculated across each segment from pressure gradients and 1A blood flow which was estimated from red cell velocities. PAP (10 microM) produced approximately a 15% decrease in arteriolar pressures with a concomitant increase in venular pressures of between 15 and 50%. In contrast, NE (0.1 microM) significantly increased arteriolar pressure by approximately 30% and decreased venular pressure by about 25%. Changes in systemic pressure during treatment with PAP and NE were small or insignificant and could not account for the observed changes in microvascular pressure. Significant dilation was observed in 3A, 4A, and 3V vessels after papaverine. In comparison, NE caused significant constriction in all vessel orders except 4V. PAP decreased resistance across all segments between 1A and 4V by 22-42% and increased venular resistance by almost 400%. NE increased resistance in all microvascular segments with the largest changes occurring in 2A to 3A (+276%) and 4A to 4V (+277%) segments. These data demonstrate that PAP and NE induce significant and opposite changes in arteriolar and venular pressures. Such network alterations in microvascular pressure should be considered when evaluating microvascular reactivity and exchange in the presence of vasoactive agents.

In a group of 12 subjects with essential hypertension (EH), we evaluated the erythrocyte membrane fluidity and red cell membrane transverse fluidity gradient. We also evaluated the total red cell Ca content, the red cell cytosolic free calcium, the red cell membrane cholesterol/phospholipid ratio and the red cell membrane individual phospholipids. From the data obtained, it is evident that the erythrocyte membrane fluidity and red cell membrane transverse fluidity gradient discriminate normals from hypertensives. None of the red cell metabolic parameters is able, however, to differentiate normals from hypertensive subjects. Our data underline the abnormality of the red cell membrane dynamic properties in hypertension; this abnormality is not, however, related to the red cell metabolic parameters considered.

The role of leukocytes in the decreased perfusion following ischemia in skeletal muscle was examined in the microcirculation of the rat cremaster muscle. The isolated muscle was viewed with an intravital microscope. Diameters of A1 and A2 arterioles and collecting venules were determined hourly. The number of leukocytes rolling along the venular walls was determined from a videotape. Nonischemic (control) rats (n = 10) were observed for 6 hours. The ischemic group (n = 10) was observed for one hour, the iliac and femoral arteries and veins were then clamped for 4 hours, released, and the muscle was observed for another two hours. No change in arteriole or venule diameters occurred in the control group. The diameters of the arterioles in the ischemic group decreased significantly during reperfusion but, the venule diameters did not. There was a significant reduction during reperfusion but, perfused capillaries following ischemia compared to control. There was a small but not significant increase in the number of rolling leukocytes in the ischemic group. The extent of leukocyte rolling in postcapillary venules was found to not correlate with the decrease in capillary perfusion that occurs after ischemia and reperfusion. However, the decrease in capillary flow was associated with reduced arteriole diameters.

The suffused noneverted cheek pouch of pentobarbital anesthetized hamsters was used to study the effects of various inhibitors of receptor/cellular function on inflammatory mediator stimulated increases in vascular permeability. Fluorescein isothiocynate dextran (FITC-D, 70,000 Da) was utilized as a tracer, and intra-vital light microscopy was employed to monitor the formation of vascular leakage sites while direct measurement of plasma and suffusate tracer concentrations were used to monitor tracer clearance. Vascular permeability increases were triggered by suffusing the cheek pouch with histamine, bradykinin, or Compound 48/80 which stimulated the formation of focal FITC-D leakage sites in the postcapillary venules resulting in marked increases in [FITC-D]S, [FITC-D]S/[FITC-D]p. 10(-6), and FITC-D clearance. Saline, calmidazolium, and papaverine lacked intrinsic permeability increasing activity, and failed to alter histamine, bradykinin, and compound 48/80 stimulated formation of venular FITC-D leakage sites and increases in [FITC-D]S, [FITC-D]S/[FITC-D]p. 10(-6), and FITC-D clearance. In contrast, treatment with cytochalasin B, DDAVP, diphenhydramine, tubulazole C, or verapamil inhibited histamine and Compound 48/80 stimulated formation of venular FITC-D leakage sites and increases in [FITC-D]S, [FITC-D]S/[FITC-D]p. 10(-6), and FITC-D clearance. Bradykinin stimulated formation of venular FITC-D leakage sites and increases in [FITC-D]S, [FITC-D]S/[FITC-D]p. 10(-6), and FITC-D clearance were not affected by treatment with calmidazolium, cytochalasin B, DDAVP, diphenhydramine, tubulazole C, or verapamil. These findings demonstrate that inflammatory mediator stimulated increases in vascular permeability may be differentially affected by inhibitors of receptor/cellular function.

This study tested the hypothesis that the rarefaction of cutaneous capillaries with age is compensated by an increase in the response of remaining capillaries to heat stress. Experiments were performed on unanesthetized male Sprague Dawley rats of the following age groups: young (Y--age 4 mo.), middle aged (M--age 16 mo.) and old (O--age 27 mo.). The subepidermal vascular plexus of the tail was viewed by video microscopy from which measurements of blood cell velocity (BCV) in single plexus capillaries and the density of flowing capillaries (DFC) were made. The animal's body and tail were housed in separate chambers for respectively indirect and direct heat stress trials. At a neutral temperature of 25 degrees C there were no significant age differences in BCV, but DFC was decreased from the Y to the M to the O groups. During an indirect heat stress of 35 degrees C, there was a modest increase in DFC and the Y greater than M greater than O rank order persisted. By contrast, all groups experienced a marked increase in BCV during both indirect and direct heat stress, and the degree of the response was significantly greater for the O rats compared to the M or Y groups. These results support the hypothesis being tested.

Endothelium-derived relaxing factor, which is believed to be nitric oxide (NO), mediates vasodilation of arteries perfused with hypoxic solutions. The purpose of the present study was to determine if NO mediates the response of arterioles in the hamster cheek pouch to changes in superfusion solution PO2. This was accomplished by comparison of constriction of fourth order arterioles to increases in superfusate PO2 before and during superfusion with NG-nitro-L-arginine (L-NAG), a stereospecific inhibitor of NO synthesis. The efficacy of L-NAG was assessed by comparison of dilations induced by topical application of methacholine (MCH), an endothelium-dependent vasodilator. We found that 10-15 min superfusion with 30 microM L-NAG significantly inhibited MCH-induced arteriolar dilation. However, this concentration of L-NAG had no significant effect on resting arteriolar diameters, O2-induced constrictions, constrictions induced by phenylephrine or dilations induced by sodium nitroprusside (SNP). Increasing the concentration of L-NAG to 100 microM similarly inhibited MCH-induced dilations, but did not affect SNP reactivity and may have increased vasoconstriction induced by O2. Thus, effective inhibition of NO synthesis in the hamster cheek pouch does not inhibit responses to elevated oxygen. Therefore NO does not mediate arteriolar O2 reactivity in this tissue. Furthermore, there is little evidence for tonic modulation of arteriolar reactivity by NO in the microvessels observed in this study.

In a group of diabetics subdivided for type (12 of type 1 and 12 of type 2), we evaluated the total red cell Ca content, red cell cytosolic free calcium, erythrocyte membrane fluidity and erythrocyte membrane protein lateral mobility. From the results obtained, it is evident that the total red cell Ca content does not discriminate normals from type 1 and 2 diabetics, whereas the red cell cytosolic free calcium does differentiate between these diabetic types. Erythrocyte membrane fluidity and erythrocyte protein lateral mobility discriminate normals from type 1 and 2 diabetics. In normals and in diabetics of type 1 and 2, no relationship is evident between total red cell Ca content, membrane fluidity and membrane protein lateral mobility. A slight, but significant negative correlation between red cell cytosolic free calcium values and parameters reflecting the red cell dynamic properties is present in type 2 diabetics only.

In a group of subjects with essential obesity, in a group of obese subjects with non-insulin dependent diabetes mellitus (NIDDM), and in a group of obese subjects with impaired glucose tolerance (IGT), we evaluated whole-blood filtration, mean erythrocyte aggregation, erythrocyte membrane fluidity and red cell lipid pattern. From these data, it is evident that the macro- and microrheological determinants are able to discriminate normals from each group of obese subjects. Regarding the red cell lipids, few are the variations between each group of obese subjects and normal controls.

The suffused noneverted cheek pouch of pentobarbital anesthetized hamsters was used to study the effects of localized, selective mast cell degranulation on vascular permeability. Fluorescein isothiocynate dextran (FITC-D, 70,000 Da) was utilized as a tracer, and intra-vital light microscopy was employed to monitor the formation of vascular leakage sites while direct measurement of plasma and suffusate tracer concentrations were used to monitor tracer clearance. Varying the time at which the FITC-D tracer was injected i.v. relative to the start of the Compound 48/80 suffusion permitted direct determination of the duration of any observed increase in vascular permeability. Selective, local mast cell degranulation was triggered by suffusing the cheek pouch with Compound 48/80 for 10 minutes which stimulated the formation of focal FITC-D leakage sites in the postcapillary venules resulting in increases in [FITC-D]S, [FITC-D]S/[FITC-D]P. 10(-6), and FITC-D clearance. In contrast, suffusion of the cheek pouches with saline failed to trigger the formation of venular FITC-D leakage sites or to promote increases in [FITC-D]S, [FITC-D]S/[FITC-D]P. 10(-6), and FITC-D clearance. The increase in permeability produced by Compound 48/80 was marked but transient (duration less than 20 minutes), and subject to inhibition by treatment with either the H1 receptor antagonist diphenhydramine or the endothelial cell stabilizer isoproterenol. There was no evidence for for a non-histamine mediated or delayed-onset increase in vascular permeability to macromolecules during the course of these experiments.