Microcirculation, endothelium, and lymphatics最新文献

The presence and distribution of glyceraldehyde-3-phosphate dehydrogenase (GPDH) in cultured bovine aortic endothelial cell (EC) and smooth muscle cell (SMC) were studied. For this purpose, we purified GPDH from human and bovine red blood cell (RBC) membranes and used it as antigen; anti-GPDH serum and affinity purified IgG were prepared. GPDH has been identified in the whole extracts of EC and SMC as a polypeptide having the same electrophoretic mobility as RBC protein. In addition, GPDH digested with V8 protease and analyzed by one dimensional peptide mapping presented the same pattern for the three cell types examined. Anti-RBC-GPDH cross-reacted with the polypeptide from EC and SMC. The intracellular localization of GPDH in EC and SMC was investigated by indirect immunofluorescence microscopy using affinity purified anti-GPDH. We found that the antigen exhibits a diffuse cytoplasmic distribution in both cell types; in addition, EC contained the antigen in the nucleus. The nuclear GPDH-like protein in EC has a characteristic pattern, suggesting yet unknown implications of GPDH in the cell metabolism.

The dose-dependents effects of topically applied verapamil (a calcium entry blocker) on vessel diameter and macromolecular clearance (as an index of permeability) were investigated in the hamster cheek pouch. Verapamil at concentrations between 10(-10)M and 10(-7)M produced a gradual and parallel increase in both vessel diameter and in macromolecular clearance. At concentrations between 10(-7)M and 10(-5)M verapamil greatly increased macromolecular transport, but only modestly increased vessel diameter. Our data indicate that low doses of verapamil influence microvascular transport by modulation of flow and associated changes in microvascular pressure, while doses of verapamil greater than or equal to 10(-7)M produce a more direct effect on microvascular permselectivity.

This study investigated effects of increased arterial carbon dioxide on the brain capillary perfusion pattern. Conscious rats were exposed to a 0%, 8% or 12% CO2 in air gas mixture. Arterial blood pressure, heart rate, arterial blood gases and pH were recorded, and either regional cerebral blood flow or the percent of capillary volume/mm3 or number/mm2 perfused were determined in cortical, hypothalamic, pontine or medullary regions of the brain. Arterial PCO2 increased from 37 +/- 1 in control to 74 +/- 1 torr in the high CO2 group. A position linear relationship was found between cerebral blood flow and arterial PCO2 in all examined regions. Approximately half of the capillaries in the examined regions were perfused under normocapnic conditions. Increasing arterial PCO2 had no effect on the percent of the capillary bed perfused in the cortex or hypothalamus. However, there was a significant linear relationship between carbon dioxide tension and the percent of the microvasculature perfused in the hindbrain. The percent of capillaries/mm2 perfused increased significantly in the medulla (to 60 +/- 5%) and pons (70 +/- 4%) with 12% CO2 in air. These data suggest that carbon dioxide may have differential effects on diffusion distances affecting the hindbrain to a greater extent than the forebrain.

In skeletal (cremaster) muscle of pentobarbital anesthetized rats we tested the hypothesis that blood flow-dependent regulation of vascular resistance exists in the microcirculation. During occlusion of an arteriole we found that the consequent increase in red blood cell (RBC) velocity in a proximal parallel arteriole was followed by a mean increase in diameter of 32 percent (mean control diameter: 21.5 +/- 0.5 microns) of the arteriole under study. The increase in arteriolar diameter always appeared with a delay (mean: 8.4 +/- 0.5 s) following the onset of changes in RBC velocity. Upon release of the occlusion RBC velocity decreased followed by a decline in diameter of the arteriole under study. Since the changes in arteriolar diameter during this experimental intervention cannot be explained on the basis of previously described blood flow-regulatory mechanisms in the microcirculation we conclude that changes in blood flow velocity (wall shear stress) per se induced the changes in arteriolar diameter. The existence of this phenomenon suggests a new, flow velocity-sensitive mechanism which can regulate - via changes in diameter - the supply and distribution of blood flow in the microcirculation in vivo.

The purpose of this study was to determine if microvascular reserve was recruited during atrial pacing in the hypertrophied myocardium. Hypertrophy was induced by one-kidney, one-clip (1K1C) renal hypertension and compared to uninephrectomized control rabbits 30 days after surgery. Coronary flow was determined by radioactive microspheres. Microvascular perfusion was determined by comparison of fluorescein isothiocyanate-dextran labeled vessels with an alkaline phosphatase stain. Heart rates were not different between groups and all animals were paced 35% above baseline. Baseline coronary flow (176 +/- 44 and 207 +/- 61 ml/min/100 g, control and 1K1C animals) was not altered by pacing in either group. The baseline percent of capillaries perfused was not different between groups (56 +/- 2 and 61 +/- 3%, sham and 1K1C) and the percent perfused increased significantly during pacing for both the non-hypertrophied and 1K1C myocardium (76 +/- 6 and 78 +/- 10%). The baseline percent of the arteriolar bed perfused, was higher in the 1K1C (86 +/- 5%) compared to non-hypertrophied (63 +/- 6%) myocardium. During pacing, the percent of the arteriolar bed perfused increased in non-hypertrophied (89 +/- 14%) but not in the 1K1C myocardium. Although the percent of arterioles perfused did not increase with pacing in the 1K1C myocardium, the capillary reserve was recruited, facilitating the transport of O2 in both the hypertrophied and non-hypertrophied myocardium.

This study investigated the contribution of cytotoxic oxygen-derived free radicals to the skeletal muscle injury seen in a rat hindlimb tourniquet model after ischemia and reperfusion. The free radical scavengers superoxide dismutase (SOD) and catalase (CAT) were used as biologic probes to detect free radical activity, while Ca2+ uptake by sarcoplasmic reticulum (SR) was used to measure subcellular muscle function. Anesthetized rats received SOD (2 mg/kg IV) plus CAT (3.5 mg/kg IV, n = 6 treated group) or saline alone (4 ml/kg, n = 6 control group) 5 min before unilateral hindlimb tourniquet ischemia of 3 hr duration. SOD and CAT were conjugated to polyethylene glycol to increase their plasma half-life. After 19 hr reperfusion, muscle from ischemic and non-ischemic lower legs of each rat was excised and homogenized. Skeletal muscle SR was isolated by differential centrifugation and ATP-dependent Ca2+ uptake by SR was then measured with dual wavelength spectrophotometry and a calcium-sensitive dye. In control rats, Ca2+ uptake velocity by SR from ischemic muscle was reduced by 48% compared with contralateral non-ischemic muscle (p less than .001). Rats pretreated with SOD + CAT showed a less severe (27%) reduction in Ca2+ uptake velocity by SR from ischemic muscle. Thus, SOD + CAT significantly (p less than .01) reduced the dysfunction of SR Ca2+ transport seen in this tourniquet ischemia model. These results strongly implicate the involvement of oxygen-derived free radicals in abnormal Ca2+ transport observed in skeletal muscle after ischemia and reperfusion.

Heparin continues to be recommended in the clinical management of limb ischemia to prevent extension of distal vascular thrombosis and increased rates of limb loss. However, heparin may also be responsible for reduced skeletal muscle injury. Although its mechanism of action has not been fully evaluated, we have investigated the ability of heparin to minimize skeletal muscle injury associated with the ischemia-reperfusion syndrome in an in vivo canine gracilis muscle model. Our findings demonstrated a significant reduction in the amount of skeletal muscle infarction, microvascular permeability, and H+ ion accumulation cumulation after preischemic heparinization. Diffuse intravascular coagulation also has been observed in observed in this model which may be prevented or reduced by the anticoagulant properties of heparin when administered prior to ischemia. However, heparin's protective effect may be independent of its anticoagulant activity. Heparin is a polycomponent drug with non-anticoagulant properties which may serve to reduce cellular injury during ischemia and reperfusion in several different ways. Microvascular injury is decreased by the restoration of normal intimal negative charge and through the binding and resultant inactivation of histamine, bradykinin and other vasoactive amines. Heparin inhibits the complement cascade which is known to determine ischemic infarct size. Other factors of importance in determining the extent of skeletal injury include neutrophil activation, chemotaxis, enzyme release, and free oxygen radical generation, all of which are decreased or modulated by heparin. Heparin is a complex substance and much more remains to be learned about its anticoagulant and nonanticoagulant properties as well as its protective effects on skeletal muscle injury in ischemia-reperfusion syndrome.

Oxygen-derived free radicals have been implicated as a mediator of the microvascular and parenchymal cell injury associated with reperfusion of ischemic tissues. Xanthine oxidase and neutrophilic NADPH oxidase are commonly invoked to explain reperfusion-induced production of oxygen radicals. The strengths and weaknesses of evidence used to suggest the involvement of both sources are discussed. Evidence is also presented which suggests that xanthine oxidase and neutrophils are redundant yet interactive mechanisms that play an important role in reperfusion injury.

Skeletal muscle is unique in its ability to tolerate relatively long periods of ischemia without demonstrable damage following reperfusion. Prolonged ischemia, however, has been associated with muscle necrosis and poor recovery of function. Using a rabbit model of hind limb ischemia, periods of ischemia of 1, 2, 3, and 5 hours were studied. Whereas almost complete recovery was seen after 1 or 2 hours of ischemia, a progressive loss of function is seen with increasing ischemic interval. In addition, within the 5 hour group, up to 40% of preparations did not recover function during reperfusion, with no Doppler signals audible over the pedicle. In these, microscopic thrombi was demonstrated histologically. Thus it appears that the "no reflow" phenomenon plays a major role after prolonged (greater than 4 hrs) ischemia. In order to evaluate the effect of fibrinolytic drugs on the "no reflow" phenomenon, urokinase was infused prior to reperfusion, and after 5 hours of ischemia, in a separate group of animals. All of these reperfused without any evidence of "no reflow". We conclude that reperfusion injury may have two major components: the "no reflow" phenomenon secondary to poor reperfusion, and cellular injury resulting from reperfusion itself. Infusion of fibrinolytic agents during the initial phases of reperfusion may have a salutory effect in preventing the "no reflow" phenomenon. It is likely, however, that attempts at effective and safe retrieval of ischemic tissue will necessarily have to address both mechanisms.