Therapeutic interventions against reperfusion injury in skeletal muscle

Ischemia/reperfusion (I/R) injury in the skeletal muscle is inevitable in many vascular and musculoskeletal traumas, diseases, free tissue transfers, and during time-consuming reconstructive and transplantation surgeries. Although skeletal muscle has a higher tolerance to ischemia than other organs, prolonged ischemia can still result in significant complications, including muscle necrosis and apoptosis. One of the major goals in the treatment of ischemia is early restoration of blood flow (reperfusion) to the area at risk. However, reperfusion has led to a new pathophysiologic condition called "reperfusion injury," a phenomenon which actually provokes a distinct degree of tissue injury. The purpose of this review is to examine the current state of understanding of I/R injury as well as to highlight recent developments on I/R interventions including our own experience in this particular field. We expect, as our acquired experience and the increased knowledge of underlying mechanisms of I/R injury, more effective interventions aimed to reduce I/R injury will be developed to interfere with or modulate this particular pathophysiologic processes.     

Nitric oxide and skeletal muscle reperfusion injury: current controversies (research review)

Nitric oxide (NO) has been implicated in a large number of disease processes, including ischemia-reperfusion injury following the restoration of oxygenated blood to previously ischemic muscle, which is a recognized significant complication of vascular surgery. Altered metabolism of NO is implicated in the endothelial dysfunction that forms part of the pathophysiology of ischemia-reperfusion injury. However, NO can demonstrate either protective or cytotoxic effects during reperfusion injury. The use of transgenic mice, either NO synthase (NOS) gene knockout animals, or animals that over-express NOS isoforms, along with direct NO measurements and NO donor or inhibitor studies, have all demonstrated a role for NO in skeletal muscle reperfusion injury. There appears to be an initial stimulation of NO production in the first 20-min of ischemia, with a gradual decline through early reperfusion and a second higher peak of NO commencing in the later stages of reperfusion. The absolute levels of NO in the reperfused tissue and its regulation by the subtle interplay with superoxide and the subsequent production of the highly toxic peroxynitrite anion, are important factors in determining whether NO, in the context of ischemia-reperfusion injury, has damaging or protective effects in the body.     

The role of oxygen-derived free radicals and the effect of free radical scavengers on skeletal muscle ischemia/reperfusion injury

The aim of this study was to clarify the role of oxygen-derived free radicals and the effect of free radical scavengers on skeletal muscle ischemia/reperfusion injury. Male Wistar rats were divided into a complete ischemia group (C-group) and an incomplete ischemia group (IC-group) and each animal was subjected to 2h of ischemia and 1h of reperfusion. In an attempt to decrease reperfusion injury, the rats were given free radical scavengers either as allopurinol 50 mg/kg for 2 days or as superoxide dismutase 60,000 units/kg plus catalase 500,000 units/kg. Tissue malondialdehyde, a product of lipid peroxidation, was measured as an indicator of free radicals, with higher levels indicating higher concentrations of free radicals. The malondialdehyde level in the gastrocnemius muscle after 1h of reperfusion increased significantly in both groups when compared to the levels before and 2h after ischemia, although there was no significant difference between the two groups. The water content of the gastrocnemius muscle and serum creatinine phosphokinase MM isoenzyme (CPK-MM) in both groups, and GOT in the C-group, increased significantly after 1h of reperfusion when compared the values before and 2h after ischemia. In the C-group, these values were significantly higher than in the IC-group. The administration of free radical scavengers suppressed the increase in malondialdehyde in the gastrocnemius muscle after reperfusion in both groups. The increase in water content and CPK-MM after reperfusion was also suppressed by free radical scavengers in the IC-group, but not in the C-group. These findings suggest that ischemic damage predominates in complete severe ischemia/reperfusion injury, whereas reperfusion injury predominates in incomplete mild ischemia/reperfusion injury.     

Combined enzymatic and antioxidative treatment reduces ischemia-reperfusion injury in rabbit skeletal muscle

BACKGROUND: Ischemia/reperfusion (I/R) injury is characterized by the production of oxygen-free radicals leading to disturbances in vasomotility (microvascular constriction) and microvascular permeability (interstitial edema formation). The objective was to evaluate the effect of the combined antioxidative and enzymatic preparation Phlogenzym on I/R injury of skeletal muscle. MATERIALS AND METHODS: A rabbit hindlimb model of I/R (2.5/2 h) was used (IR group). Phlogenzym, containing rutin, trypsin, and bromelain, was applied enterally (60 mg/kg body weight) as a bolus 30 min prior to ischemia (Ph group). Sham-operated animals served as controls (CO group). Plasma malondialdehyde, potassium, and microvascular perfusion (monitored by laser flowmetry) were assessed. Histomorphometry and electron microscopy were performed from major adductor muscles. RESULTS: Two hours after reperfusion, potassium levels were significantly elevated in IR compared to Ph group (6.7 +/- 1.2 versus 4.9 +/- 0.9 mmol/l, P < 0.006). Enhanced lipid peroxidation, apparent by increased plasma malondialdehyde levels, was ameliorated in the Ph group (1.0 +/- 0.1 versus 0.7 +/- 0.1 nmol/ml, P < 0.0001). No-reflow (reduction of blood flow by 62% in IR group) was not observed in the Ph group (P < 0.004). Phlogenzym treatment prevented microvascular constriction (17.6 +/- 2.3 versus 12.6 +/- 1.1 microm(2), P < 0.0001) and mollified interstitial edema (21.5 +/- 2.0 versus 26.0 +/- 3.7%, P < 0.017), resulting in mild ultrastructural alterations in contrast to pronounced sarcolemmal and mitochondrial damage in untreated rabbits. CONCLUSIONS: Phlogenzym had a protective effect on skeletal muscle during I/R injury expressed by prevention of no-reflow and preservation of muscle tissue.     

Bradykinin Impairs and HOE 140 does not Protect Rat Hindlimb Skeletal Muscle Against Tourniquet-induced Reperfusion Injury

Background: Bradykinin (BK) is used in different tissues. Dose-dependent studies have demonstrated that low doses protect against ischemia/reperfusion (I/R) injury while higher doses lead to adverse effects. Although the beneficial effects of BK infusion were observed in myocardium, its role on the I/R impact in skeletal muscle (SM) has not been fully clarified. Objective: This study was carried out to evaluate the effects of BK, administered in the hindlimbs of rats subjected to I/R. Methods: The study design included three experimental groups: Group 1 control (saline), Group 2 (bradykinin), and Group 3 (HOE 140, a BK2 receptor blocker). In all three groups, rats were subjected to hindlimb ischemia for a total of 2 h followed by continuous 4 h of reperfusion with pharmacological interventions. The methods include analysis of enzymes (lactate dehydrogenase—LDH and creatinine phosphokinase—CPK), cell membrane marker of injury (malondialdeyde—MDA), recruitment of neutrophils (myeloperoxidase—MPO), and apoptosis index (immunohistochemistry TUNEL in situ peroxidase dead end). Results: Except for the apoptotic index, all parameters studied were shown to be elevated in the reperfusion group intervened with BK. The blocking of BK2 receptors by HOE 140 did not affect the I/R injury. Conclusion: After 2 h of total ischemia, infusion of bradykinin during 4 h of reperfusion, worsened the I/R injury in the hindlimb skeletal muscle.     

Ischemia/reperfusion‐induced necrosis and apoptosis in the cells isolated from rat skeletal muscle

Necrosis was considered to be the solo mechanism for ischemia/reperfusion (I/R)‐induced cell death. Recent evidence from I/R models of the heart, liver, kidney, and brain indicates that apoptosis is a major contributor to I/R‐induced cell death. However, evidence of I/R‐induced apoptosis in skeletal muscle is sparse and divided. The purpose for the present study was to investigate I/R‐induced necrosis and apoptosis in the cells isolated from rat skeletal muscle. A rat gracilis muscle model was used. After surgical preparation, clamps were applied on the vascular pedicle to create 4 h of ischemia and released for 24 h of reperfusion (I/R, n = 10). Clamping was omitted in sham I/R rats (sham I/R, n = 10). The muscle samples were harvested after 24 h of reperfusion for the process of cell isolation. Cells were stained by Propidium Iodide (PI) or Annexin V‐FITC or both. Twenty thousand cells from each muscle sample were scanned and analyzed by flow cytometry. The average percentage of live cells was 45 ± 2% in the I/R group versus 65 ± 3% in the sham I/R group (p < 0.01). The average percentage of necrotic cells was 18 ± 1% in I/R versus 12 ± 1% in sham I/R (p < 0.01). The average percentage of apoptotic cells was 40 ± 3% in I/R versus 27 ± 3% in sham I/R (p < 0.01). Our results clearly demonstrated that I/R not only causes necrosis, but also accelerates apoptosis in the cells isolated from rat skeletal muscle. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:351–356, 2008     

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     

Enoxaparin does not ameliorate limb ischemia-reperfusion injury

OBJECTIVE: Since low molecular weight heparin has greater bioavailability and sustained serum levels in vivo than unfractionated heparin, it has been used to supplant unfractionated heparin to achieve therapeutic anticoagulation in humans. These studies were designed to determine whether treatment with enoxaparin could protect murine skeletal muscle from ischemia reperfusion injury. METHODS: C57BL6 mice were divided into four groups. Sham control animals underwent 90 min of anesthesia alone. All other groups underwent 90 min of unilateral hindlimb ischemia. At the onset of reperfusion, animals received either normal saline (control and saline) or 4 mg/kg of enoxaparin subcutaneously twice daily. Groups were followed for 24 or 48 h reperfusion. Hindlimb skeletal muscle blood flow was measured by laser Doppler, and muscle was removed for histological and protein analysis. Tissue thrombosis was evaluated by thrombin antithrombin III (TAT III), local inflammation by measurement of proinflammatory cytokines (macrophage inflammatory protein-2: MIP-2, monocyte chemoattractant protein-1: MCP-1), and neutrophil infiltration by myeloperoxidase (MPO) using enzyme-linked immunosorbent assay. Plasma levels of Factor Xa were measured during reperfusion to confirm therapeutic levels of anticoagulation. Comparisons were calculated using analysis of variance. RESULTS: At 24 h reperfusion, there was increased expression of MIP-2, MCP-1, MPO, and TAT III in saline and enoxaparin treated mice compared with control (*P < 0.05). By 48 h reperfusion, all parameters measured remained greater than control except for the enoxaparin treated mice whose TAT III levels were significantly less than untreated mice (P < 0.05). Despite documented therapeutic anticoagulation and decreased levels of markers of thrombosis in enoxaparin treated mice, there was no difference in tissue cytokines, inflammatory markers, degree of muscle fiber injury (31% +/- 8% versus 30% +/- 5%) or muscle flow between ischemia-reperfusion groups (2447 +/- 141 versus 2475 +/- 74 flux units) at 48 h reperfusion. CONCLUSIONS: Post hoc administration of enoxaparin did not affect local tissue thrombosis, inflammatory markers, or muscle necrosis. This suggests that despite its potent in vivo activity, enoxaparin did not modulate skeletal muscle injury, thrombosis, or inflammatory following ischemia reperfusion. enoxaparin may not be useful in mediating skeletal muscle injury when administered in a clinically relevant scenario.     

Interleukin-1 and thromboxane release after skeletal muscle ischemia and reperfusion

Interleukin-1 and thromboxane are known to mediate the host response to sepsis, trauma, and myocardial ischemia. A well-established model of canine isolated gracilis muscle was used to evaluate whether cytokine (interleukin-1) played a role in skeletal muscle ischemia-reperfusion injury. Six adult mongrel dogs (25–30 kg) were subjected to six hours of muscle ischemia followed by reperfusion. Gracilis venous samples were collected pre-ischemia and at one hour of reperfusion. Systemic (arterial) blood samples were taken at one hour of reperfusion. Sera were analyzed for interleukin-1 by bioassay and thromboxane (B₂) by radio-immunoassay. The gracilis muscle of the operated limb was harvested in all the animals for assessment of the percentage of muscle necrosis. This was found to be 56.2±14.8% by serial transections, nitroblue tetrazolium staining, and computerized planimetry. Interleukin-1 levels in the gracilis venous effluent increased from 21.88±7.13 units/ml during pre-ischemic baseline to 50.42±9.12 units/ml after six hours of ischemia followed by one hour of reperfusion (p<0.04). Thromboxane B₂ levels were 2983±1083 pg/ml and 9483±2218 pg/ml at pre-ischemia and at one hour of reperfusion respectively (p<0.04). Systemic levels of both interleukin-1 and thromboxane B₂ at one hour of reperfusion were 0 units/ml and 1584±520 pg/ml respectively, which were significantly lower than the one hour reperfusion gracilis venous effluent levels (p<0.04). This is the first report in which cytokines have been implicated in skeletal muscle ischemia-reperfusion injury. Modulation of interleukin-1 may impact positively on muscle necrosis and systemic manifestations of reperfusion injury.Presented at the 16th Annual Meeting of the Peripheral Vascular Surgery Society, June 2, 1991, Boston, Massachusetts.     

Adiponectin protects skeletal muscle from ischaemia–reperfusion injury in mice through miR-21/PI3K/Akt signalling pathway

Previous studies have confirmed that adiponectin (APN) plays a protective role in myocardial ischaemia–reperfusion (IR) injury, and the aim of this study was to investigate its effect on skeletal muscle. ELISA was used to detect the levels of Creatinine Kinase (CK), LDH, SOD and MDA in the plasma of the lower limbs of mice, and the levels of IL-6, IL-1β and TNF-α in the gastrocnemius. Quantitative PCR was used to detect the expression level of miR-21. TUNEL staining was used to detect the apoptosis of the gastrocnemius. The expression levels of apoptosis proteins, autophagy marker proteins and downstream target genes of miR-21 in gastrocnemius were detected by Western Blot. The results of this study revealed that APN levels were significantly reduced in gastrocnemius of IR mice. The oxidative stress, inflammatory response, apoptosis and autophagy induced by IR were significantly ameliorated by APN injection. The above effects of APN may be achieved through miR-21/PI3K signalling pathway, as found by interfering gene expression levels with miRNA antagomir and lentiviral injection. Taken together, our study revealed that APN protects skeletal muscle from IR injury through miR-21 /PI3K/Akt signalling pathway through inhibiting inflammatory response, apoptosis and autophagy.     

Novel intracellular N‐terminal truncated matrix metalloproteinase‐2 isoform in skeletal muscle ischemia‐reperfusion injury

Ischemia‐reperfusion injury (IRI) occurs when blood returns to tissues following a period of ischemia. Reintroduction of blood flow results in the production of free radicals and reactive oxygen species that damage cells. Skeletal muscle IRI is commonly seen in orthopedic trauma patients. Experimental studies in other organ systems have elucidated the importance of extracellular and intracellular matrix metalloproteinase‐2 (MMP‐2) isoforms in regulating tissue damage in the setting of oxidant stress resulting from IRI. Although the extracellular full‐length isoform of MMP‐2 (FL‐MMP‐2) has been previously studied in the setting of skeletal muscle IRI, studies investigating the role of the N‐terminal truncated isoform (NTT‐MMP‐2) in this setting are lacking. In this study, we first demonstrated significant increases in FL‐ and NTT‐MMP‐2 gene expression in C2C12 myoblast cells responding to re‐oxygenation following hypoxia in vitro. We then evaluated the expression of FL‐ and NTT‐MMP‐2 in modulating skeletal muscle IRI using a previously validated murine model. NTT‐MMP‐2, but not FL‐MMP‐2 expression was significantly increased in skeletal muscle following IRI. Moreover, the expression of toll‐like receptors (TLRs) ‐2 and ‐4, IL‐6, OAS‐1A, and CXCL1 was also significantly up‐regulated following IRI. Treatment with the potent anti‐oxidant pyrrolidine dithiocarbamate (PDTC) significantly suppressed NTT‐MMP‐2, but not FL‐MMP‐2 expression and improved muscle viability following IRI. This data suggests that NTT‐MMP‐2, but not FL‐MMP‐2, is the major isoform of MMP‐2 involved in skeletal muscle IRI. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:502–509, 2016.     

Mitochondrial alterations in skeletal muscle submitted to total ischemia

BACKGROUND: The role of mitochondrial lesions in the pathogenesis of irreversible cellular ischemia is controversial. The inability to restore mitochondrial function is correlated with the inability to reverse cell damage in various tissues. The objective of the present study was to compare parameters associated with oxidative phosphorylation and the inner mitochondrial membrane potential of skeletal muscle of rats submitted to total ischemia in order to determine which mitochondrial alterations are mainly affected in this condition. MATERIAL AND METHODS: Wistar rats were submitted to 5 h total ischemia using the tourniquet method (ischemic limb group). The contralateral limb of each rat was used as control (control limb group). After the ischemic period, muscle biopsies were obtained for the isolation of the mitochondrial fractions, which were submitted to polarographic analysis for the determination of ADP-activated oxygen consumption (state 3), basal respiration (state 4), and the ratio of the two respiratory activities: the respiratory control ratio (RCR). The potential of the inner mitochondrial membrane was determined by measuring the fluorescence difference between coupled and uncoupled mitochondria using safranine O as indicator. RESULTS: After 5 h of ischemia, a significant reduction of all parameters studied was observed in skeletal muscle submitted to ischemia compared to the control limbs. CONCLUSIONS: Five-hour total ischemia applied to rat skeletal muscle led to the inhibition of the mitochondrial respiratory chain (represented by decreased state 3 and state 4 respiration rates). The percentage decrease in the electrical potential of the inner membrane was similar to the percentage reductions observed for state 4 respiration and the RCR. The parameter mostly affected by ischemia was ADP-activated respiration (state 3).     

缺血后处理对减轻骨骼肌缺血再灌注损伤作用的实验研究

目的：研究缺血后处理对鼠骨骼肌缺血再灌注损伤的保护影响,组织中凋亡和胀亡的存在情况。方法将54只SD大鼠随机分为空白对照组、缺血再灌注组、缺血后处理组,持续缺血4 h,再灌注6 h,24 h,48 h。检测血浆乳酸脱氢酶（LDH）、肌酸磷酸激酶（CPK）活性、肌肉内丙二醛（MDA）含量及总超氧化物歧化酶（SOD）活性,进行组织学、免疫组化、超微结构分析。结果相比缺血再灌注组,后处理组在再灌注6 h时,只SOD活性明显升高,而再灌注24 h,48 h时,在MDA含量下降、SOD活性升高、W/D值下降、组织学改变范围及免疫组化阳性范围方面,均较缺血再灌注组有明显差异。结论再灌注开始时应用后处理对于缺血再灌注损伤有明显的保护作用,主要体现在再灌注的稍后期阶段（再灌注24 h,48 h）。缺血再灌注过程中,凋亡和胀亡是并存的。     

Ischemic preconditioning of skeletal muscle mitigates remote injury and mortality

BACKGROUND: Ischemic preconditioning (IPC) mitigates ischemia-reperfusion (I/R) injury in experimental models. However, the clinical significance of this protection has been unclear and a mortality reduction has not been previously reported in noncardiac models. This study examined the local and remote protection afforded by skeletal muscle IPC and sought to determine the significance of this protection on mortality. METHODS: Mice subjected to 2 h hindlimb ischemia/24 h reperfusion (standard I/R injury) were compared with those undergoing a regimen of two 20-min cycles of IPC followed by standard I/R injury. Local injury was assessed via gastrocnemius histology, and remote injury was evaluated via intestinal histology and pulmonary neutrophil infiltration (n = 7). Mortality was compared in parallel groups for 1 week (n = 6). Groups were analyzed using an unpaired Student's t-test for gastrocnemius and pulmonary injury, and a Mann-Whitney rank sum test for intestinal injury. Mortality differences were interpreted through a hazard ratio. RESULTS: Significant protection was observed in preconditioned animals. There was a 35% local injury reduction in skeletal muscle (71.2% versus 46.0%, P < 0.01), a 50% reduction in remote intestinal injury (2.3 versus 1.1, P < 0.01), and a 43% reduction in remote pulmonary injury (14.9 versus 8.5, P < 0.01) compared with standard injury controls. Preconditioned animals were also significantly protected from mortality, demonstrating a 66.7% survival at 1 wk compared with 0% survival after standard injury alone (hazard ratio 0.20, 95% CI: 0.02-0.59). CONCLUSIONS: We have developed a murine model of IPC that demonstrates local and remote protection against I/R injury, and exhibits significant mortality reduction. This model demonstrates the powerful effect of IPC on local and remote tissues and will facilitate further study of potential mechanisms and therapies.     

利多卡因对骨骼肌缺血-再灌注损伤作用的实验研究

目的 阐明利多卡因对骨骼肌缺血-再关注损伤的作用.方法 采用MTT法测定骨骼肌组织活力,分别对缺血后、再灌注后加入利多卡因与对照组进行比较,并结合透射电镜,观察利多卡因对缺血再灌注损伤后的作用.结果 缺血后、再灌注后加入利多卡因与对照组进行比较,P＜0.01,差异有统计学意义;缺血后与再灌注后2组比较,P＞0.05,差...     

Artificial oxygen carrier improves fatigue resistance in slow muscle but not in fast muscle in a rat in situ model

The effects of liposome-encapsulated hemoglobin with high O₂ affinity (h-LEH), an artificial O₂ carrier in skeletal muscle, were studied by in situ fatigue resistance test in fast-type plantaris (PLT) and slow-type soleus (SOL) muscles with or without ischemia in the rat. The distal tendons of PLT and SOL muscles were isolated in situ and individually attached to the force transducers to record the developed tension in response to stimuli (80 Hz tetanus train, 1.5 minutes) to the ipsilateral sciatic nerve. The fatigue resistance test (five sets separated by 2-minute rests) was evaluated in terms of tension attenuation (fatigue) from the initial to the last tension (A) during each set, attenuation of the initial (B) or last tension (C) in each set, as compared to the first set in the presence or absence of ischemia or h-LEH (10 mL/kg). While ischemia significantly enhanced fatigue only in PLT, h-LEH showed no effect regardless of the perfusion pattern (normal/ischemia) or muscle-type (PLT/SOL) during each set (A). In parameter (B), set-by-set fatigue development was observed in PLT, whereas h-LEH-SOL showed a trend of advanced fatigue resistance. Such trends became clear in the parameter C (last tension), because h-LEH-SOL exerted, rather than decreased, the tension enhancement regardless of the presence or absence of ischemia, whereas there were no h-LEH effects in PLT. In addition, faster recovery of the nicotinamide adenine dinucleotide content in the muscle after 10 minutes of all fatigue tests was observed in h-LEH-SOL, while saline-SOL still showed a significantly higher value than that of control. These results suggested that additional O₂ supply by h-LEH may accelerate the tricarboxylic acid cycle/electron transport chain in slow-type aerobic SOL muscle containing abundant mitochondria and contribute to the faster removal of muscle fatigue substances such as lactate.     

间断缺血对犬骨骼肌存活时限影响的实验研究

目的 研究间断缺血延长离体骨骼肌存活时限。方法 以犬股薄肌为模型，利用“4C”法、琥珀酸脱氢酶（SDH）光镜组化、电镜超微结构观察等方法观察离体骨骼肌存活时限。结果 常温下犬股薄肌的缺血时限为7h，低温（4℃）可以延长骨骼肌的存活时限至15h，间断缺血可以延长骨骼肌的存活时限为25h。结论 间断缺血明显延长骨骼肌存活时限，从而可能为断肢保存提供了一种更为理想的方法。     

Striated muscle fiber compositions of human male urethral rhabdosphincter and levator ani

PURPOSE: We clarified the contractile properties of human male periurethral striated muscle fibers to better understand how the rhabdosphincter and the levator ani maintain urinary continence. MATERIALS AND METHODS: Muscle specimens were obtained from 52 male patients who underwent radical prostatectomy or radical cystectomy. The specimens were frozen in liquid nitrogen. Frozen sections (10 microm thick) were stained with myofibrillar ATPase at different pH values (pH 4.2, 4.6 and 10.6), and evaluated for quantitative parameters and fiber type distribution. Myosin heavy chain analysis was performed using SDS-PAGE. RESULTS: Of all 52 cases 37 provided specimens that could be divided into the 2 major fiber types, type 1 (slow twitch) and type 2 (fast twitch). Although type 1 muscle fibers were predominant in RS and LA muscle groups (RS 69.6 +/- 2.7%, LA 67.0 +/- 2.0%), mean muscle fiber size was significantly smaller in RS (mean area 906 +/- 86 microm(2)) than in LA (mean area 2,967 +/- 170 microm(2)) (p <0.0001). In 11 specimens type 2 muscle fibers could be subdivided into types 2A (fast fatigue resistant) and 2B (fast fatigable). Type 2A fibers were significantly more prevalent than type 2B fibers (p <0.05). Likewise, MHC analysis of these 11 specimens found a significantly higher percentage of fiber type 2A expression products (MHC 2A) than of fiber type 2B expression products (MHC 2X) (p <0.05). CONCLUSIONS: RS and LA contribute to urinary continence mechanism by slow contraction. Moreover, the smaller mean size of muscle fibers in RS suggests more fatigue resistance compared with muscle fibers in LA because small fibers have a shorter diffusion distance for metabolic substrates. These results should help contribute to a more detailed understanding of the function of periurethral striated muscles in the human male.     

Ischemia-reperfusion injury in rat skeletal muscle is attenuated by zinc aspartate

BACKGROUND: Oxygen-derived free radical-induced cell injury has been suggested to have a pivotal role in the etiology of ischemia-reperfusion injury. Thus, several lines of evidence indicate that antioxidant agents may be useful therapeutics in this condition. In this regard, the effect of zinc aspartate on ischemia-reperfusion injury was investigated in skeletal muscle. MATERIALS AND METHODS: Tourniquet ischemia-reperfusion injury method was applied to Sprague-Dawley rats. Experimental groups were as follows: 1) sham control, 2) rats received zinc aspartate, 3) rats received hind limb tourniquet operation (left side), and 4) rats received hind limb tourniquet operation and zinc aspartate. Viability of muscle was evaluated by triphenyltetrazolium chloride dye method by using a spectrophotometer. Malondialdehyde, superoxide dismutase, catalase, glutathione, and glutathione peroxidase were measured in muscle, heart, lung, and blood via a spectrophotometer. RESULTS: The viabilities of ischemic limbs, percentage of the contralateral control muscle, in group 1, 2, 3, and 4 were 114 +/- 12%, 87% +/- 5%, 20% +/- 2%, and 95 +/- 10%, respectively. In muscle, increased malondialdehyde and decreased superoxide dismutase, catalase, and glutathione levels in group 3 were normalized by zinc aspartate in both left and right limbs. While malondialdehyde levels in heart and blood increased in group 3, the levels of superoxide dismutase, catalase, glutathione, and glutathione peroxidase were lower in group 3 than those in group 1. All these alterations were prevented by zinc aspartate. Malondialdehyde level of lung in group 3 was significantly higher than group 1 and 2. However, this augmentation was halted by zinc aspartate. The decrease in superoxide dismutase levels in group 3 was statistically reversed by the administration of zinc aspartate. CONCLUSION: Zinc aspartate seems to be an effective treatment option against ischemia-reperfusion injury.