Morphometric study of the ultrastructural organization of the endotheliocytes of blood capillaries in the iliac lymph nodes of pregnant rats

The ultrastructure of blood capillary endotheliocytes of iliac lymph nodes during pregnancy was investigated in rats. It was found that changes in microcirculation during pregnancy are accompanied by the appearance of the adaptive changes at all levels of lymph node structural organization. They are: increased lumen of arterial and venous part of the capillaries, increased intermediate sinus volume, increased fenestration of both parts of capillaries, a sharp increase in the number of micropinocytous vesicles and activation of transendothelial channel formation.     

Early ultrastructural changes in the myocardium following thyroxine-induced hypertrophy

The model of myocardial hypertrophy induced by thyroxine was studied with particular regard to the early ultrastructural changes in fractional volume of the mitochondria and myofibrils, and capillary distribution. Following injections of L-thyroxine (25 mg/kg IP) for 9 consecutive days, rats were sacrificed by vascular perfusion and cardiac tissue samples from the mid-wall zone of the left ventricle were processed routinely for electron microscopy. Heart weight/body weight ratios of thyroxine treated (T) rats showed a significant increase (P less than 0.001) over the ratios in control (C) rats. Likewise, the fractional volume of mitochondria (42%) was significantly increased (P less than 0.001) in the myocardium of T rats when compared with C rats (31%). However, the fractional volume of myofibrils was significantly decreased in the myocardium of T rats (P less than 0.001) and there was no significant difference between the hearts of T and C rats with respect to capillary luminal area/myocyte area. The mitochondria/myofibril ratio was increased in the hearts of T rats (0.82) over that found in control hearts (0.52). These results suggest that in the early stages of thyroxine-induced myocardial hypertrophy there is not an immediate increase in capillary area which may account for the ischemia and significant increase in mitochondrial volume which characterized myocardial hypertrophy in this model.     

Myocardial capillaries in the fetal and the neonatal rat: a morphometric analysis of the maturing myocardial capillary bed

Developing myocardial capillaries from 16-day-gestation fetus to adult undergo several morphological changes including a thinning of the lateral extensions of the capillary endothelial cells, the formation of a basal lamina, and an increase in the number of plasmalemmal vesicles. A decrease in the extracellular space, an increase in the number of capillaries, and a decrease in the capillary diameter were also observed during the developmental period. In view of these ultrastructural changes, a morphometric analysis was made on the developing myocardial wall to demonstrate specific quantitative changes. The volumes which were occupied by capillary endothelial cells, capillary lumina, extracellular space, and myocardial myocytes within a reference volume of myocardium were measured; and we found that 8% of the reference myocardial volume was occupied by capillary endothelial cells, 85% was occupied by myocardial myocytes, 4% was occupied by capillary lumina, and, except for a significant change in extracellular space at 16 days gestation, 3% was occupied by extracellular space. Each volume ratio was found to be nearly constant throughout the studied period. In contrast to this constancy in the volume ratios, other parameters which were measured demonstrated significant changes during the developmental period studied. These overall changes include a 135% increase in capillary density, a 63% increase in luminal surface area of capillary endothelial cells, a 24% decrease in capillary diameter, a 12% decrease in diffusion distance, and a 35% decrease in the diameter of the erythrocyte population.(ABSTRACT TRUNCATED AT 250 WORDS)     

Testicular blood flow and microcirculation in rats after treatment with ethane dimethyl sulfonate

The effects of ethane dimethyl sulfonate (EDS) on total testicular blood flow, microcirculation, and the testicular interstitial fluid volume (IFV) in rats were studied. In agreement with previous studies, treatment of control rats with human chorionic gonadotropin (hCG) induced an increase in IFV and total testicular blood flow as measured with radioactive microspheres. These effects of hCG were completely abolished in rats pretreated with EDS; in EDS-treated rats not receiving any hCG, there were decreases in IFV when compared with untreated control rats. Furthermore, the pulsatile pattern of testicular microcirculation registered with laser-Doppler flowmetry was abolished after EDS treatment, and this effect was not influenced by hCG treatment. The hCG-induced increase in IFV is associated with an increased accumulation of polymorphonuclear leukocytes locally in the testis, but this accumulation of leukocytes was not observed in rats pretreated with EDS. It was concluded from the present study that hCG-induced changes in total testicular blood flow and testicular microcirculation require functionally intact Leydig cells.     

Improvement of the microcirculation in the acute ischemic rat limb during intravenous infusion of drag-reducing polymers

Drag-reducing polymers (DRPs) are blood-soluble macromolecules that can increase blood flow and reduce vascular resistance. The purpose of the present study is to examine the effects of DRPs on microcirculation in rat hind limb during acute femoral artery occlusion. Two groups of 20 male Wistar rats were subjected to either hemodynamic measurement or contrast enhanced ultrasound (CEU) imaging during peripheral ischemia. Both groups were further subdivided into a DRP-treated group or a saline-treated group. Polyethylene oxide (PEO) was chosen as the test DRP, and rats were injected with either 10 ppm PEO solution or saline through the caudal vein at a constant rate of 5 ml/h for 20 min. Abdominal aortic flow, iliac artery pressure, iliac vein pressure, heart rate, carotid artery pressure and central venous pressure (CVP) were monitored, and vascular resistance was calculated by (iliac artery pressure-iliac vein pressure)/abdominal aortic blood flow. Flow perfusion and capillary volume of skeletal muscle were measured by CEU. During PEO infusion, abdominal aortic blood flow increased (p<0.001) and vascular resistance decreased (p<0.001) compared to rats that received saline during peripheral ischemia. There was no significant change in ischemic skeletal capillary volume (A) with DRP treatment (p>0.05), but red blood cell velocity (β) and capillary blood flow (A×β) increased significantly (p<0.05) during PEO infusion. In addition, A, β and A×β all increased (p<0.05) in the contralateral hind limb muscle. In contrast, PEO had no significant influence on heart rate, mean carotid artery blood pressure or CVP. Intravenous infusion of drag reducing polymers may offer a novel hydrodynamic approach for improving microcirculation during acute peripheral ischemia.     

Metabolic models of microcirculatory regulation.

The functions and integrity of body tissues are critically dependent on an adequate oxygen supply. Because the transport of oxygen to the cells is intimately linked to the microcirculation, the concept of microcirculation-metabolism coupling has received much attention. In essence, the metabolic theory of intrinsic control of the microcirculation states that microvascular tone is locally modulated to maintain adequate oxygen levels in the parenchymal cells. We propose a two-component control system for the regulation of tissue O2 delivery in accordance with metabolic needs. A precapillary sphincter control mechanism maintains tissue PO2 by governing the number of perfused capillaries. Functional capillary density in turn determines surface area available for diffusion and capillary-to-cell diffusion distance. On the other hand, the arteriolar control system modulates local blood flow in accordance with parenchymal O2 utilization and thereby minimizes changes in capillary PO2 when the O2 availability/demand ratio is decreased. We propose that the precapillary sphincters are more sensitive to changes in tissue PO2 than are the flow-regulating arterioles. Consequently, for mild stresses, adequate tissue oxygenation is maintained mainly by precapillary sphincter control of diffusion parameters without the need for changes in blood flow. However, as metabolic stresses become greater, blood flow regulation becomes the dominant factor in the control of tissue O2 delivery. Thus, by working in concert, the local mechanisms regulating microvascular resistance and effective capillary density provide a wide margin of safety against the development of cellular hypoxia.     

Plasmodium coatneyi: alterations of transcapillary escape rate and capillary permeability to fibrinogen in rhesus monkeys

The transcapillary escape rate and plasma clearance of fibrinogen were studied in six rhesus monkeys infected with Plasmodium coatneyi as well as in six control monkeys using 131I-fibrinogen as a tracer. The mean transcapillary escape rate of fibrinogen in the infected group was significantly higher than that of the control group. Both plasma volume and plasma fibrinogen concentration were also elevated in the infected group, these resulting in a significantly higher intravascular fibrinogen mass, plasma clearance rate, and outflux of fibrinogen from the intravascular to the extravascular compartments. Both effective capillary pore area/unit path length available for restricted diffusion and the specific permeability coefficient of plasma fibrinogen in the infected monkeys were also found to be significantly higher than those of the control group. These findings indicated that there was an increased leakage of plasma fibrinogen from the circulation into the extravascular space which was due either to increased capillary surface area and/or to an increased capillary permeability in rhesus monkeys infected with P. coatneyi.     

Effects of hetastarch and mannitol on prolonging survival in stable hypothermia in rats

In rats, prolonged stable hypothermia ( approximately 24 h at body temperature of 19 degrees C) is characterized by a time-dependent increase in hematocrit, plasma osmolality, and red blood cell fragility and a decrease in plasma volume. These changes impede tissue microcirculation and could limit survival. As a countermeasure, we used plasma volume expanders of both long (hetastarch)- and short-lasting (mannitol) characteristics to improve microcirculation and hopefully hypothermia survival. Infusion of 6% hetastarch at hour 3 in hypothermia significantly (P < 0.05) enhanced survival over saline control (33.5 vs. 23.8 h); a significant delay in the increases of hematocrit and cell fragility was also observed compared with those in saline controls. Treating the animal with 6% hetastarch at hour 20 during hypothermia caused a similar but less-effective improvement in survival. In contrast, treating the rats with 6% mannitol at hour 3 or 20 during hypothermia failed to enhance survival over saline control, although transient improvement in plasma volume was observed. Our results indicate that by using a long-lasting volume expander, which tends to better maintain plasma volume and rheological parameters governing microcirculation than does saline or a short-lasting volume expander, hypothermia survival can be significantly improved.     

Microcirculatory disturbances following the passage of emboli in an experimental free-flap model.

Following completion of arterial repair in an experimental free-flap model, platelet emboli have been observed passing through the microcirculation downstream. The purpose of this experimental study was to observe and quantitate changes in capillary perfusion occurring subsequent to these events. The isolated rat cremaster model was used. For 6 hours subsequent to surgical injury of the main artery in this model, the number of emboli and the number of perfused capillaries downstream were counted. In eight rats having an intentional arterial wall injury, emboli were consistently seen during the first hour of reflow. In the nine control animals having no arterial injury, no emboli were seen. The presence of emboli in the cremaster muscle, resulting from the arterial injury, was associated with a significant reduction in the number of perfused capillaries. We suggest that the observed decrease in capillary perfusion was associated with microemboli that produced an adverse effect for several hours after their initial presence in the circulation.     

肠系膜微血管通透速度的图象分析

本文应用微循环活体观察和荧光示踪技术,通过计算机数字图象处理对荧光素钠（ＦＩＮａ）和ＦＩＴＣ－Ｄｅｘｔｒａｎｓ（ＦＤ－４,ＭＷ４０００,ＦＤ－１５０,ＭＷ１５００００）在肠系膜微血管的通透过程进行了定量研究,建立了正常大鼠肠系膜微血管对不同分子量荧光物质的通透方程,得到了通透系数及对通透系数评价的定量方法。结果证实：颈动脉注射荧光素钠后４ｓ即可在微血管中见到,扩散速度很快,通透系数为３．３６２×１０－８ｃｍ２／ｓ。注射ＦＤ－４后１５ｓ可在微血管中见到,扩散速度比荧光素钠慢１倍左右,通透系数为１．７１８×１０－８ｃｍ２／ｓ。注射ＦＤ－１５０后８ｓ可在微血管中见到,扩散速度很慢,约为荧光素钠扩散速度的１／７０,通透系数为０．０４８６×１０－８ｃｍ２／ｓ。荧光物质的渗出部位主要在毛细血管及细静脉,在细动脉则极少见到。证实我们计算得出的通透系数能够较好地反映肠系膜微血管通透的实际情况,所采用的评价方法及得到的通透方程可以用于微血管物质交换参数的定量评价。     

Exercise-induced focal skeletal muscle fiber degeneration and capillary morphology

The relationship between exercise-induced focal muscle fiber degeneration and changes in capillary morphology was investigated in male Wistar rats. Untrained animals ran on a treadmill for 1 h at submaximal intensity and were killed 0, 6, or 24 h after running. Nonexercised rats served as controls. In situ perfused soleus muscles were prepared for electron microscopy. Micrographed cross sections were quantitatively analyzed for parameters indicative of capillary blood flow or transcapillary exchange. Capillary lumina were ovally rather than circularly shaped, and no indications for obstruction of blood flow at the capillary level were found. Endothelial cells and their organelles had a normal appearance in all groups. However, immediately after exercise, capillaries showed a decreased thickness of their endothelium and basal membrane, probably caused by dehydration. Six hours after exercise, muscle fibers were swollen (28% increase in cross-sectional area), resulting in a slightly increased diffusion distance. This fiber swelling was not associated with an increase in muscle water content, a finding for which no explanation could be found. Twenty-four hours after the animals ran, capillaries located near degenerated muscle fibers had an increased cross-sectional luminal area and an increased luminal circumference. This effect decreased gradually with increasing distance from the degenerated fiber area. The present morphometric results do not support the hypothesis that changes in capillary morphology primarily contribute to exercise-induced focal muscle fiber degeneration.     

Ketamine does not affect intestinal microcirculation in pentobarbital-anaesthetized rats during experimental endotoxaemia

The objective of the study was to evaluate the effects of ketamine on intestinal microcirculation in pentobarbital-anaesthetized rats during experimental endotoxaemia. A prospective, randomized, controlled study was carried out using 32 male Lewis rats. The animals were divided into four groups (n = 8 each). All animals were initially anaesthetized with 60 mg/kg pentobarbital (i.p.). Group 1 served as a control (18.5 mg/kg/h pentobarbital i.v.). Groups 2 and 4 received an endotoxin intravenous infusion of 15 mg/kg lipopolysaccharide (LPS) from Escherichia coli. Groups 3 and 4 also received 10 mg/kg/h ketamine (i.v.). After 2 h of observation, the animals were examined for intestinal functional capillary density (FCD) and leukocyte adherence to the venular endothelium by means of intravital fluorescence microscopy (IVM). Subsequent to this examination, blood samples were collected to determine release of the cytokines tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6 and IL-10. Endotoxaemia tended to decrease intestinal FCD (mucosa: -10.1%, muscularis longitudinalis: -2%, muscularis circularis: -9.8%) and significantly increase leukocyte adherence within submucosal venules (collecting venules: +133%, postcapillary venules: +207%; P<0.05). TNF-alpha, IL-1beta, IL-6 and IL-10 levels were significantly elevated following endotoxin challenge. The addition of ketamine to pentobarbital anaesthesia did not significantly affect FCD, leukocyte behaviour or cytokine levels. In conclusion, intravenous pentobarbital anaesthesia with the additional administration of ketamine did not cause alterations within the microcirculation or changes in cytokine release during endotoxaemia. In rats, the combination of pentobarbital and ketamine is suitable for use during the study of intestinal microcirculation in experimental endotoxaemia.     

Coronary capillary network remodeling and hypofibrinolysis in aged obese diabetic rats: Implications for increased myocardial vulnerability to ischemia

Despite the known abnormalities of cardiac function in patients with overt non-insulin dependent diabetes mellitus (NIDDM) the temporal changes of coronary capillary network remodeling leading to potential microcirculatory dysfunction have not been elucidated. To this end, left ventricular subendocardial capillary network of Otsuka Long-Evans Tokushima Fatty (OLETF) rats, characterized by hypertension, obesity, hyperglycemia, hyperinsulinemia and mild NIDDM, and control Long-Evans Tokushima (LETO) rats were investigated. Total capillary density in OLETF was significantly higher than that in LETO at 20 weeks, suggesting compensatory improvement of O₂ transport at early stages of NIDDM. The increase in capillary density in OLETF was lost at 40 and 60 weeks due to the decreases of intermediate capillary portions and venular capillary portions. Although capillary domain area (area innervated by single capillary) in OLETF was lower than that in LETO at 20 weeks, the values were similar between OLETF and LETO at 40 and 60 weeks, suggesting that adaptive improvement in the capacity for O₂ transport with a high perfusion was lost in late stages of NIDDM. Activity of plasma plasminogen activator inhibitor-1 (PAI-1), the major physiologic inhibitor of proteo(fibrino)lysis, in OLETF was higher than that in LETO at 40 and 60 weeks, suggesting that increase of PAI-1 may downregulate compensatory adaptive capillary network remodeling by inhibiting proteolysis and angiogenesis in the cardiac interstitium. Loss of adaptive myocardial microcirculation may therefore contribute to increased vulnerability in ischemic injury and to cardiac dysfunction in NIDDM.     

Temperature and angiogenesis: the possible role of mechanical factors in capillary growth

This review examines the effect of prolonged cold exposure on muscle capillary supply in mammals and fishes. In rats and hamsters, the response to a simulated onset of winter is to conserve the microcirculation and maintain a constant capillary to fibre ratio (C:F), implying either an unaltered vacular bed or angiogenesis matched by muscle hyperplasia, while chronic acclimation to low environmental temperature induces a variable degree of muscle atrophy, which in turn increases capillary density (CD). In striped bass and rainbow trout, cold-induced angiogenesis results in an increase in C:F, but also a cold-induced fibre hypertrophy that is accompanied by a powerful angiogenic response such that CD is much less sensitive to changes in fibre size. Endothelial cells can act as mechanotransducers such that angiogenesis may be initiated by changes in their physical environment. It is hypothesised that in mammals, the metabolic consequences of cold exposure increases the luminal shear stress, while in fishes the stimulus for angiogenesis is abluminal stretch following an increase in fibre size.     

A quantitative investigation of microvascular changes in the thyroid gland after infrared (IR) laser radiation.

We present an ultrastructural study of thyroid capillaries in which 50-day-old rats Wistar rats, were irradiated with an infrared (IR) laser, (total dose, 46.80 J/cm2), the tissue quantified 1 day after ending treatment and a quantitative capillary analysis carried out by light and electron microscopy. Light microscopy was used to calculate capillary volume density revealing a significant increase in the irradiated rats. The quantitative measurement of parameters by electron microscopy required a two stage analysis: Level I, Electron Microscopy (Magnification x5,000); and Level II, Electron Microscopy (Magnification x26,000). At Level I, the following parameters were measured in each capillary: capillary area, capillary diameter, luminal area, luminal diameter, endothelial area, nuclear area and mean endothelial thickness. At Level II, pinocytotic vesicle diameter and their numerical density in endothelial cells were evaluated. Electron microscopic analysis revealed an increased luminal area in the capillaries of the irradiated rats. They also presented a decrease in endothelial cell thickness and vesicular diameter and an increase in vesicle numerical density. This latter increase is indicative of presumptive changes in capillary permeability, but the possible functional significance of these morphological changes in the endothelial cells requires further investigation.     

Pulmonary injury in rats following continuous exposure to 60% O2 for 7 days

Morphological, biochemical, and physiological studies were done on rats exposed to 60% O2 for 7 days. This exposure did not induce O2 tolerance but instead caused a significant decrease in survival time of animals subsequently exposed to pure O2. The activity of lung superoxide dismutases and glucose-6-phosphate dehydrogenase were unchanged after exposure to 60% O2. A decrease in lung compliance was suggested by changes in the total lung capacity and in the pressure-volume curves of excised lungs. Ventilation of these animals with large tidal excursion resulted in pulmonary edema. Morphometric analyses revealed a significant decrease in alveolar air volume and an increase in the number of alveolar macrophages. The most significant lesions involved the pulmonary vascular bed. The volume and thickness of the capillary endothelium was decreased. There were focal areas of pericapillary fluid accumulations, and a number of the smaller vessels had perivascular edema. These findings suggest that significant pulmonary injury occurs in rats exposed to 60% O2 and that the primary site of injury is the pulmonary capillary endothelium.     

Influence of lung volume on pulmonary microvascular pressure-volume characteristics.

The pressure-volume (P-V) characteristics of the lung microcirculation are important determinants of the pattern of pulmonary perfusion and of red and white cell transit times. Using diffuse light scattering, we measured capillary P-V loops in seven excised perfused dog lobes at four lung volumes, from functional residual capacity (FRC) to total lung capacity (TLC), over a wide range of vascular transmural pressures (Ptm). At Ptm 5 cmH(2)O, specific compliance of the microvasculature was 8.6%/cmH(2)O near FRC, decreasing to 2.7%/cmH(2)O as lung volume increased to TLC. At low lung volumes, the vasculature showed signs of strain stiffening (specific compliance fell as Ptm rose), but stiffening decreased as lung volume increased and was essentially absent at TLC. The P-V loops were smooth without sharp transitions, consistent with vascular distension as the primary mode of changes in vascular volume with changes in Ptm. Hysteresis was small (0.013) at all lung volumes, suggesting that, although surface tension may set basal capillary shape, it does not strongly affect capillary compliance.     

Photooptical and ultrastructural changes in the microcirculatory bed of the kidney after functional vasoconstriction

In experiments on rats it was found that at the early stages (5 to 15 minutes) after vasoconstriction of the kidney caused by adrenaline solution there occurred sharp narrowing of the intraorganic arterial bed lumen, particularly that of the afferent arterioles. Ultrastructural changes in the glomerular renal capillary components observed were morphological expression of the effect of angiospasm and circulatory hypoxia. Residual phenomena of constriction of the renal microcirculatory bed still persisted at later stages--in 3, 7 days. These changes characterized the stage of peculiar mobility and contractile properties of their endothelial cells, caused by spasmogenic disturbances of microcirculation.     

Estimation of hematocrit profile symmetry recovery length downstream from a bifurcation

Downstream from a microvascular bifurcation the distribution of blood cells in the vessel lumen is not symmetric. A diffusion process is used to model the rearrangement of red cells as blood flows between junctions in the microcirculation. A Fourier series approach is used to solve the model diffusion convection equation in slit geometry. Both flat and parabolic velocity profiles are considered. The eigenvalues, found using the Rayleigh-Ritz method, are used to find an upper bound on distance required for a symmetric red cell distribution to be obtained. The method has also been applied to cylindrical geometry and the computed symmetry recovery lengths are compared to distances between bifurcations measured in vivo. These estimates indicate that red cell distributions are frequently asymmetric in the microcirculation. Such asymmetries can have a strong effect on plasma skimming and material balance calculations.     

Quantitative study of the microvascular pattern in tourniquet-induced muscle ischemia

Experiments were carried out on 40 adult Wistar rats to determine the effect of the application of a pneumatic tourniquet on the capillary network of the tibialis anterior muscles. The tourniquet was applied for four hours to the right hind limb above the knee. During the degeneration processes (6 to 24 hours postischemia), the capillary density and capillary-to-fiber ratio decreased significantly with respect to a control group of untreated animals. In the subsequent regenerative processes, there was a greater increase of fiber density than of capillary density, resulting in a capillary-to-fiber ratio lower than that of the control group. In the last period (30 to 90 days postischemia), both capillary density and fiber density returned to normal values.