Intravascular thrombosis in skeletal muscle microcirculation after ischemia

This study was undertaken to elucidate the role of thrombosis in blood vessels less than 0.1 mm in diameter in skeletal muscle after an ischemic episode. Capillaries were examined after normothermic ischemia using intravital video microscopy of the cremaster muscle of an anesthetized rat. Histologic sections of the cremaster muscle and silicone rubber intravascular casts were also analyzed. Our model demonstrates that, after 3 hours of warm ischemia, up to two-thirds of capillaries are clotted, as is most of the venous system. Our findings also indicate that capillaries may reopen during 30 minutes of reperfusion. These findings suggest that ischemia may cause thrombosis in the microvasculature of skeletal muscle but the thrombosis appears to be partially reversible.     

Studies on microcirculation injury of skeletal muscle due to tourniquet-induced ischemia

Blood flow does not return to all areas after long period of ischemia produced by tourniquet application. The cause of this "no-reflow phenomenon" is still a matter of controversy. The purpose of this study was to investigate the relationship between thrombus formation and the no-reflow phenomenon by comparing the change of platelet function and the findings of vessel lumen observed by scanning electron microscopy. Through the results of a microangiogram, the author confirmed that the no-reflow phenomenon occurred on the first day and disappeared on the fifth day after the tourniquet was released. Mean volume, maximum aggregation and malondialdehyde of platelets decreased at two hours to one day after tourniquet release due to the consumption of larger platelets. There were irregular endothelia and a mural thrombus in the veins of the ischemic leg on the first day following release of the tourniquet. These findings suggest that thrombus formation plays an important role in the development of the no-reflow phenomenon.     

Combined effect of acute denervation and ischemia on the microcirculation of skeletal muscle.

Using direct in vivo videomicroscopy and a fluorescein dye technique, reperfusion injury after 3 h of ischemia was studied in the acutely denervated cremaster muscle of the rat. Compared with normally innervated controls, ischemia-induced reperfusion injury was more severe in the denervated group and included a delay of blood flow recovery, vortex formation, edema, hemorrhage, and vessel spasm. Vessel size was reduced at the arteriole and small artery level, and there was a decrease of reactive hyperemia. The injury mechanism may be related to a loss of active vasomotion and vascular response to vasoactive substances after denervation. The results suggest that shortening the ischemia time of denervated tissues may reduce ischemia-induced reperfusion injury. Similarly, given the same ischemia time, improved tissue reperfusion may be expected if the nerve supply is maintained.     

Histopathology of muscle flap microcirculation following prolonged ischemia

The purpose of this study was to evaluate histological changes occurring in the microcirculation of a muscle flap following prolonged ischemia and to correlate them with the flow hemodynamics. The cremaster muscle flap model was used for direct in vivo studies of the microcirculation. In 45 rats, vascular clamps were applied to the iliac and femoral vessels following flap isolation. Flaps were subjected to various periods of ischemia, ranging from 4 to 6 hours, and 2 hr of reperfusion. In vivo observations of the microcirculation and vessel diameter measurements were taken at 1 hour intervals for an 8 hr period. With prolonged ischemia, return of circulation to the flap was delayed and no flow was observed following 6 hr of ischemia. Morphologic changes at 6 hr revealed red cell and platelet thrombi formation within the capillaries, marked dilatation of postcapillary venules, endothelial swelling in the capillaries, and microhemorrhage formation around the venules. © 1995 Wiley-Liss, Inc.     

Reperfusion of skeletal muscle after warm ischemia

The effect of 2 to 10 hours of tourniquet ischemia on the microcirculation of the tenuissimus muscle in 29 cats was studied by in vivo microscopy and electron microscopy. After release of the tourniquet there was immediate reperfusion and hyperemia in all muscles. Arterioles of 20 micron increased in diameter by 50% and venules of 35 micron by 30%. If the ischemia had lasted for 8 hours or more, reocclusion of flow occurred after 3 to 55 minutes. Platelet thromboembolism, sticking of leukocytes, and red cell aggregation seem to cause the occlusion.     

Studies of ischemia-reperfusion injury in skeletal muscle: Efficacy of 21-aminosteroids on microcirculation and muscle contraction after an extended period of warm ischemia

The present study was conducted to elucidate the effects of tirilazad mesylate (U-74006F), a potent inhibitor of lipid peroxidation, on vessel diameter, capillary perfusion, and contractile function of rat cremaster muscle during a 90-minute reperfusion period that followed 4 hours of warm ischemia. Two groups of 32 animals were treated with either 3 mg/kg U-74006F or the vehicle (citrate buffer) alone 30 minutes before. ischemia, 90 minutes after ischemia, and immediately before reperfusion. With use of intravital videomicros-copy, the internal luminal diameters of preselected vessels were measured prior to ischemia and during reperfusion The area that filled with fluorescein was determined at 15-minute intervals for as long as 90 minutes of reperfusion, and contractile function was examined in vitro in an organ bath at that point. In the U-74006F group, after 90 minutes of reperfusion the vessel diameters returned completely to baseline and the diameters of all three categories of vessels at every time point from 10 to 90 minutes of reperfusion had significantly more rapid recovery than the controls. Although some evidence of more rapid fluorescence was noted in the U-74006F group, the two groups did not differ significantly at any time period of reperfusion. In response to tetanic stimulation, the muscles treated with U-74006F had a significantly greater contractile force at all stimulation frequencies than the control musclss. Our findings indicate that pretreatment with U-74006F can effectively decrease the rise of vascular resistance and preserve the contractile function of skeletal muscle during early reperfusion, thereby attenuating ischemia-reperfusion injury.     

Tourniquet-induced ischemia and reperfusion in human skeletal muscle

Microdialysis conceivably enables longitudinal and simultaneous investigation of several metabolites by repeated measurements in skeletal muscle. We used and evaluated microdialysis as an in vivo method to characterize the time-course and relative kinetics of pyruvate, glucose, lactate, glycerol, hypoxanthine, uric acid, and urea, in skeletal muscles, exposed to ischemia and reperfusion, in eight patients having arthroscopic-assisted anterior cruciate ligament reconstruction. A dialysis probe was implanted before surgery in the rectus femoris muscle. Dialysate samples were collected at 10-minute intervals at a flow rate of 1 microL/minute until 2 hours after tourniquet deflation. Ninety minutes of ischemia resulted in accumulation of lactate (234% +/- 38%), hypoxanthine (582% +/- 166%), and glycerol (146% +/- 46%), consumption of glucose (54% +/- 9%) and pyruvate (16% +/- 44%), and a slight decrease of urea (78% +/- 11%) compared with baseline (100%). Uric acid was unchanged (95% +/- 12%). Within 90 minutes after tourniquet deflation the concentrations were virtually normalized for all measured metabolites, suggesting that the duration of ischemia was well tolerated by the patients. The results indicate that the use of microdialysis for monitoring energy metabolic events during orthopaedic surgery that requires ischemia and reperfusion is feasible and safe.     

Effect of postischemic reperfusion on microcirculation and lipid metabolism of skeletal muscle

To study the effect of ischemia reperfusion injury on microvascular reactivity and tissue metabolism in skeletal muscle, a Sprague-Dawley rat cremaster muscle was prepared as a tourniquet ischemia model and subjected to 2 hr ischemia followed by 1 hr reperfusion to simulate the timing of ischemia during microvascular surgery. The dose-response curve of arteriolar reactivity to norepinephrine, lipid peroxidation, and ultrastructure of capillaries was determined in both the control and postischemic reperfusion stages. Judging from the results, we summarize our observations as follows: (1) Postischemic reperfusion significantly increased arteriolar reactivity to norepinephrine, in which the EC50 for vasoconstriction decreased in all three orders of arterioles. These results suggest that reperfusion could have impaired the vasodilation control mechanism, possibly being endothelium dependent. (2) Lipid peroxidation increased sixfold in the reperfusion group, suggesting that oxygen free radicals have produced significant tissue damage under the created conditions. (3) Significant endothelial damage in the capillaries shown by electron microscope observation supports these studies, indicating that ischemia/reperfusion in clinically transplanted skeletal muscles could cause significant damage to the tissue microcirculation both physiologically and metabolically. © 1995 Wiley-Liss, Inc.     

Changes in microcirculation after ischemic process in rat skeletal muscle

Laser Doppler flowmetry (LDF) is a good method to investigate tissue microcirculation, but it has many standardization and measuring problems. To exclude these effects, we performed a test using LDF on rat skeletal muscle. In 12 CD outbred anesthetized rats, bilateral femoral vessels and the quadriceps femoris muscle were exposed. The left femoral artery and vein were clamped for 1 h by microvascular clips (ischemic side). On the right side, no other intervention was made (control side). An LDF probe was applied on the medial vastus muscle. Short-term occlusions (2-3 s) were performed before and after the 1-h clamping and on the control side while LDF curves were registered. The halftimes of ascending curves on the ischemic side were significantly elongated vs. the condition before clamping (P = 0.0007) or the control side (P = 0.0017). Microcirculatory changes caused by 1-h ischemia were shown by this simple, quick, and well-reproducible test on rat skeletal muscle.     

Effect of increased compartment pressure on the microcirculation of skeletal muscle

To determine the changes in capillary perfusion, which occur with elevated tissue pressure, and to highlight the relationship between systemic blood pressure and compartment pressure, we designed an experiment that allowed direct observation of the microcirculation of skeletal muscle under normal and increased compartment pressures. In each of 10 anesthetized rats, the cremaster muscle was exposed and suspended in a transparent pressure chamber. In vivo videomicroscopy was then performed and blood pressure was monitored via left carotid artery cannulation. Two sets of data for each animal were obtained: ΔP (mean arterial pressure − compartment pressure) at which the muscle capillary blood flow was completely arrested, and the number of capillaries per 10,000 square micrometers of skeletal muscle with blood flowing at compartment pressures of 0, 15, 30, 45, and 60 mm Hg. Capillary blood flow stopped at a ΔP of 25.5 mm Hg ± 14.3 SD. We found that capillary blood flow, as measured by the number of capillaries with blood flow per 10,000 square micrometers, decreased significantly (P< 0.05) as compartment pressure reached 15, 30, 45, and 60 mm Hg, when compared to 0 mm Hg; there was no vessel collapse at these pressures. These data show that increasing compartment pressure reduces the number of perfused capillaries per unit area, and that there is complete cessation of muscle capillary blood flow when the compartment pressure is within about 25 mm Hg of the mean arterial pressure. © 1998 Wiley-Liss, Inc. MICROSURGERY 18:67-71 1998     

研究缺血对失神经支配骨骼肌萎缩的影响

目的　研究缺血对失神经支配骨骼肌萎缩的影响。方法　选用 SD大鼠 2 4只 ,建立右下肢缺血加腓肠肌失神经支配的动物模型 (实验组 ) ,左侧为对照组。术后 2、4周取双侧腓肠肌 ,另取 6只正常大鼠腓肠肌作正常对照 ,观察组织学、肌电图及酶组织化学的变化。结果　术后 2、4周 ,实验组肌细胞直径比正常对照组减少 5 1.1%和 6 3.6 % (t =2 .94,P <0 .0 5 ) ,截面积减少 5 3.6 %和 6 2 .0 % (t =2 .88,P <0 .0 5 )。实验组肌细胞线粒体呈空泡状、数量减少 ;肌质网明显扩张退变。肌肉出现纤颤电位 ,和对照组比 ,其波幅无明显差异 (t=2 .11,P<0 .0 5 )。术后 2周实验组的 Na,K- ATP酶活性比正常对照组增高 74.6 % ,术后 4周则下降至对照组的 88.0 % (t=7.6 4,2 .2 4,P<0 .0 1、<0 .0 5 )。术后2周 Ca- ATP酶活性比正常对照组下降 2 8.32 % ,术后 4周下降至 35 .8% (t=2 .98、2 .2 1,P<0 .0 5 )。结论　缺血可加速失神经支配肌肉萎缩的发生与发展     

Pravastatin attenuates tourniquet-induced skeletal muscle ischemia reperfusion injury

Background?Revascularization of a limb following prolonged ischemia results in substantial skeletal muscle injury. Statins play a well-understood role in the treatment of hypercholesterolemia but are also known to have anti-inflammatory properties. The purpose of this study was to examine the effects of pravastatin pre-treatment in the setting of skeletal muscle ischemia reperfusion injury (IRI).

Methods?Adult male Sprague Dawley rats (n = 27) were randomized into 3 groups: control group, I/R group, IR group pre-treated with pravastatin. Bilateral hind-limb ischemia was induced by rubber band application proximal to the level of the greater trochanters for 2.5 h. Treatment groups received normal saline in equal volumes prior to tourniquet release. Following 12 h reperfusion, the tibialis anterior muscle was dissected and muscle function assessed electrophysiologically by electrical field stimulation. The animals were then killed and skeletal muscle harvested for evaluation.

Results?We found that pre-treatment with pravastatin reduces the tissue oxidative damage and edema associated with skeletal muscle reperfusion injury. Skeletal muscle injury, measured by edema, leucose-questration and electrical properties were significantly lower with pravastatin pre-treatment compared to the non-treated group.

Interpretation?We feel that pravastatin pre-treatment may be a potential therapeutic intervention for skeletal muscle ischemia reperfusion injury in the clinical setting.??     

Pravastatin attenuates tourniquet-induced skeletal muscle ischemia reperfusion injury

Background Revascularization of a limb following prolonged ischemia results in substantial skeletal muscle injury. Statins play a well-understood role in the treatment of hypercholesterolemia but are also known to have anti-inflammatory properties. The purpose of this study was to examine the effects of pravastatin pre-treatment in the setting of skeletal muscle ischemia reperfusion injury (IRI).

Methods Adult male Sprague Dawley rats (n = 27) were randomized into 3 groups: control group, I/R group, IR group pre-treated with pravastatin. Bilateral hind-limb ischemia was induced by rubber band application proximal to the level of the greater trochanters for 2.5 h. Treatment groups received normal saline in equal volumes prior to tourniquet release. Following 12 h reperfusion, the tibialis anterior muscle was dissected and muscle function assessed electrophysiologically by electrical field stimulation. The animals were then killed and skeletal muscle harvested for evaluation.

Results We found that pre-treatment with pravastatin reduces the tissue oxidative damage and edema associated with skeletal muscle reperfusion injury. Skeletal muscle injury, measured by edema, leucose-questration and electrical properties were significantly lower with pravastatin pre-treatment compared to the non-treated group.

Interpretation We feel that pravastatin pre-treatment may be a potential therapeutic intervention for skeletal muscle ischemia reperfusion injury in the clinical setting. ▪     

Comparison between animal and human studies of skeletal muscle adaptation to chronic stimulation

Functional electrical stimulation (FES) has recently emerged as a clinical tool for treatment of neuromuscular disorders. Chronic muscle stimulation, however, has long been used by basic scientists studying the details of the muscular adaptation process. Biochemical, morphological, and functional changes occur in skeletal muscle secondary to chronic stimulation. Chronic stimulation (12-24 hours per day for six weeks) results in a well-defined progression of changes in which a "fast" muscle becomes a typical "slow" muscle with a large decrease in force-generating capacity. On the other hand, clinical studies of FES have demonstrated muscle strengthening following treatment. An attempt is made to reconcile the results obtained in the two fields.     

Quantitative histochemical evaluation of skeletal muscle ischemia and reperfusion injury

Acute arterial obstruction to the extremities is associated with significant morbidity and mortality. The evaluation of accompanying skeletal muscle injury has thus far been indirect and imprecise. Triphenyltetrazolium chloride (TTC) is an oxidation-reduction indicator which allows for the histochemical quantitation of skeletal muscle injury. In 21 anesthetized nonheparinized adult mongrel dogs, the isolated in vivo gracilis muscle underwent 4, 6, or 8 hr of ischemia with and without reperfusion. The muscles were excised and cut into 1-cm segments, representative muscle biopsies for electron microscopy were taken, each segment was stained in 1% TTC, and the total area of staining was measured with computerized planimetry. All control muscles stained completely with a dark red color. After 4, 6, or 8 hr of ischemia, quantitative measurements of muscle staining indicative of normal tissue were present in 98 +/- 1%, 59 +/- 5%, and 23 +/- 9% of the total muscle areas, respectively. Six hours of ischemia followed by reperfusion was associated with only 36 +/- 9% of the muscle being stained. Segmental TTC staining demonstrated that reperfusion was associated with greater injury, and less TTC staining, in the proximal portion of the gracilis muscle at the site of entry of the major arterial pedicle. The distal muscle did not demonstrate increased damage with reperfusion. It is hypothesized that protection of the distal muscle from reperfusion injury may be due to an absence of reflow farther away from the artery.     

缺血预处理减轻骨骼肌缺血再灌注损伤

目的 观察缺血预处理对骨骼肌缺血再灌注损伤的保护作用。方法 选择２４只健康兔,随机等分为实验组和对照组。实验组先进行缺血预处理,再持续阻断后肢血流４ｈ;对照组直接阻断后肢血流４ｈ,制作骨骼肌缺血再灌注损伤模型。测定再灌注期血清中肌酸磷酸激酶（ＣＰＫ）和天门冬氨酸氨基转移酶（ＡＳＴ）,镜下观察骨骼肌变化。结果 实验组血清中ＣＰＫ和ＡＳＴ的含量均明显低于对照组（Ｐ〈０．０５）。实验组骨骼肌线粒体空泡变     

Isovolemic hemodilution and skeletal muscle function during ischemia and reperfusion

Hemodilution has previously been shown to improve microcirculation in skeletal muscle after ischemia. We have studied the effects of isovolemic hemodilution with dextran on the function of anterior tibial muscle in the rabbit. Hemodiluted (hematocrit 28%) and nonhemodiluted animals were compared. Hemodilution led to an immediate increase in femoral blood flow. Flow normalized within 1–2 h, possibly due to flow redistribution. Hemodilution increased muscle force by 10%, which can reflect alterations in blood chemical composition or an improved microcirculation. Unilateral hindlimb ischemia induced by arterial occlusion inhibited muscle force to less than 15% in 150 min. Force and blood flow recovered almost completely after ischemia. After longer ischemia (170–300 min) when force was <5%, muscles did not recover. Hemodilution did not alter the muscle force or the extent or rate of force recovery after ischemia, which shows that the increased blood flow and improved microcirculation are not directly associated with changes in the sensitivity of muscle function to ischemia. © 1998 Wiley-Liss, Inc. MICROSURGERY 18:79-85 1998     

The effect of ischemia reperfusion injury on skeletal muscle

Ischemia reperfusion (IR) injury occurs when tissue is reperfused following a period of ischemia, and results from acute inflammation involving various mechanisms. IR injury can occur following a range of circumstances, ranging from a seemingly minor condition to major trauma. The intense inflammatory response has local as well as systemic effects because of the physiological, biochemical and immunological changes that occur during the ischemic and reperfusion periods. The sequellae of the cellular injury of IR may lead to the loss of organ or limb function, or even death. There are many factors which influence the outcome of these injuries, and it is important for clinicians to understand IR injury in order to minimize patient morbidity and mortality. In this paper, we review the pathophysiology, the effects of IR injury in skeletal muscle, and the associated clinical conditions; compartment syndrome, crush syndrome, and vascular injuries.     

Subpopulations of skeletal muscle mitochondria: response to ischemia

The respiratory activity of distinct populations of subsarcolemmal (SS) and interfibrillar (IF) skeletal muscle mitochondria was studied in rat hindlimb muscle subjected to 30 min of global ischemia at 37 C. State 3 (ADP-dependent) and state 4 (ADP-independent) rates of respiration were determined polarographically using glutamate as the substrate, in order to calculate the respiratory control index (RCI). The RCI is the ratio of state 3 to state 4 respirations and is a sensitive indicator of mitochondrial coupling. An approximately 20 per cent decline in RCI was noted in the SS mitochondria following 30 min of warm ischemia. This decrease was a direct result of a significant decline in the state 3 respiratory rates. IF RCIs following the ischemic interval were not different from control. These data support the concept of separate populations of mitochondria within the skeletal muscle cell and demonstrate a specific injury pattern in the response to ischemia.