Free radical-mediated postischemic injury in renal transplantation.

Oxygen free radicals are generated during reperfusion of ischemic organs. Studies employing several species of laboratory animal (rat, dog, pig, rabbit, mouse) have documented protective effects of a variety of free-radical scavengers and antioxidants when administered before or immediately preceding reperfusion of ischemic kidneys. These protective agents include superoxide dismutase, dimethylthiorea, dimethyl sulfoxide, alpha-tocopherol, glutathione, the iron chelator deferoxamine, probucol, allopurinol and oxypurinol, and the spin-trapping agent PBN. Furthermore, deficiency of antioxidants (selenium, alpha-tocopherol, or catalase) exacerbates postischemic renal injury. These findings have been applied to renal transplantation in an attempt to decrease the incidence of posttransplantation acute renal failure. This is important because acute renal failure results in morbidity, increases hospital stay and the cost of transplantation, and complicates the use of cyclosporine. In porcine and in canine kidney transplantation, superoxide dismutase and allopurinol have provided renal protection. Transplantation is complicated because there may be prolonged hypoperfusion before harvesting plus a brief period of total ischemia during harvesting, followed by a prolonged period of cold ischemia and/or reperfusion, then followed by another brief period of ischemia and reperfusion during transplantation. Injury may occur at each of these phases by different mechanisms.     

Leukocyte depletion ameliorates free radical-mediated lung injury after cardiopulmonary bypass

Activated leukocytes and oxygen free radicals have been implicated in the pathogenesis of lung injury associated with cardiopulmonary bypass. To determine whether leukocyte depletion could prevent this injury, we used a dog model simulating routine cardiac operations. Mongrel dogs (11 to 17 kg) were subjected to cardiopulmonary bypass with a bubble oxygenator and cooled to 27 degrees C. After aortic crossclamping and cardioplegic arrest for 90 minutes, control animals (n = 5) were rewarmed and weaned from bypass, and their condition was then stabilized for 90 minutes. Leukocyte-depleted animals (n = 5) had a leukocyte filter incorporated in the bypass circuit. During bypass, circulating leukocyte counts decreased by 60% in control dogs, and by 97% in leukocyte-depleted animals. Free radical generation (estimated by spectrophotometric assay of plasma conjugated dienes) was significantly reduced by leukocyte depletion during and after bypass. Total hemolytic complement activity and the titer of C5 decreased markedly immediately after the onset of bypass in both the control and leukocyte-depleted animals. Pulmonary function after bypass was better preserved in leukocyte-depleted animals. These data suggest that depletion of circulating leukocytes contributes to lung injury during cardiopulmonary bypass and is associated with increased oxygen radical activity, pulmonary edema, and vasoconstriction. Leukocyte depletion substantially reduced the pulmonary injury seen after cardiopulmonary bypass.     

Nitrotyrosine and 8-isoprostane formation indicate free radical-mediated injury in hearts of patients subjected to cardioplegia

OBJECTIVE: Myocardial ischemia and reperfusion induced by cardioplegic arrest subjects the heart to free radical-mediated stress. The purpose of our study was to investigate the effect of cardioplegia-induced ischemia and reperfusion on myocardial formation and distribution of (1) nitrotyrosine as an indicator for peroxynitrite-mediated tissue injury resulting from increased nitric oxide release and (2) 8-isoprostane as an indicator for oxygen-derived free radical-mediated lipid peroxidation. METHODS: In 10 patients undergoing coronary artery operations (64 +/- 6 [mean +/- SD] years, 3 women and 7 men) subjected to cardiopulmonary bypass and intermittent cold blood cardioplegia, we collected transmural left ventricular biopsy specimens before and at the end of cardiopulmonary bypass. Specimens were cut at 10 micro m and subjected to immunocytochemical staining against the nitric oxide-producing enzyme constitutive nitric oxide synthase, cyclic guanosine monophosphate (intracellular second messenger of nitric oxide), nitrotyrosine, and 8-isoprostane by using polyclonal antibodies. For global left ventricular function determination, we measured the fractional area of contraction using transesophageal echocardiography. RESULTS: Nitric oxide synthase activity in cardiac myocytes increased from 34 +/- 10 gray units before cardiopulmonary bypass to 47 +/- 12 gray units at the end of bypass (P =.015), and all hearts showed increased cyclic guanosine monophosphate content in both myocytes and endothelial cells at the end of bypass. The number of nitrotyrosine-positive capillaries increased from 36 +/- 29/mm(2) before bypass to 82 +/- 47/mm(2) at the end of bypass (P =.012), and 8-isoprostane-positive capillaries increased from 92 +/- 72/mm(2) before bypass to 209 +/- 108/mm(2) at the end of bypass (P =.005). The fractional area of contraction was 53% +/- 12% before bypass and 56% +/- 12% after bypass (P =.47) but was slightly decreased to 45% +/- 14% at 4 hours after bypass (P =.121). CONCLUSIONS: Our data show that cardioplegia-induced myocardial ischemia and reperfusion is associated with nitrotyrosine and 8-isoprostane formation mainly in the coronary endothelium, indicating injury mediated by both peroxynitrite and oxygen-derived free radicals. Because nitric oxide synthase activation was accompanied with increased cyclic guanosine monophosphate, these data suggest that direct effects of nitric oxide on cardiac myocytes, as well as nitric oxide-mediated coronary endothelial injury, might contribute to injury associated with cardioplegia and cardiopulmonary bypass.     

Therapeutic modulation of free radical-mediated reperfusion injury of the liver and its surgical implications

It is well known that ischemia causes functional and structural damage to liver cells, and that the status of energy metabolism provides an important means of assessing the functional viability of the ischemic organ. However, the specific sequence leading to ischemic liver cell injury is not yet fully understood; therefore, it is clinically and pathophysiologically important to elucidate the mechanism of cellular injury during hepatic ischemia and subsequent reperfusion. Whereas the conventional view attributes this injury process to the ischemia itself, recent studies have demonstrated that a variable but often substantial proportion of this injury is caused by reactive oxygen metabolites that are generated at the time of reperfusion. This article presents an outline of the mechanism of cellular injury caused during hepatic ischemia and subsequent reperfusion resulting from certain types of surgery, with special reference to the xanthine-xanthine oxidase system and the activation of neutrophils and macrophages.     

Inhibition of the compartment syndrome by the ablation of free radical-mediated reperfusion injury

Skeletal muscle edema secondary to an increase in capillary permeability after reflow is an important cause of the compartment syndrome after acute arterial revascularization. The purpose of this study was to investigate the possible role of oxygen free radicals, generated at reperfusion, in the pathogenesis of the compartment syndrome secondary to acute arterial ischemia/reperfusion. A reproducible model of this syndrome was produced in anesthetized rabbits by femoral artery occlusion after surgical devascularization of collateral branches from the aorta to the popliteal artery. Increasing periods of ischemia from 6 to 12 hours, followed by 2 hours of reperfusion, were associated with corresponding increases in the anterior muscle compartment hydrostatic pressure and inversely proportional decreases in tibialis anterior muscle blood flow within that compartment as assessed by xenon 133 washout (n = 46) (r = -0.62, p less than 0.001). Anterior compartment pressure increased from 5 +/- 1 to 48 +/- 5 mm Hg (n = 46) (p less than 0.001) after 7 hours of total arterial ischemia and 2 hours of reperfusion. Ablation of free radicals generated from xanthine oxidase with either allopurinol (n = 8) or oxypurinol (n = 8), by scavenging the superoxide radical at reperfusion with superoxide dismutase (n = 8), or by blocking secondary hydroxyl radical formation with deferoxamine (n = 8) significantly ameliorated the rise in compartment pressure (p less than 0.05) in each case; it also significantly improved muscle perfusion in the superoxide dismutase-, allopurinol-, and deferoxamine-treated animals (p less than 0.05). These findings indicate that development of the compartment syndrome after acute arterial revascularization may be due, at least in part, to microvascular injury mediated by oxygen-derived free radicals generated from xanthine oxidase at reperfusion.     

Oxygen free radicals and glutathione in hepatic ischemia/reperfusion injury

Oxygen free radicals have been implicated as mediators of ischemia/reperfusion injury in a variety of organs. We investigated the role of oxidative injury and endogenous hepatic glutathione (GSH) in liver cell injury associated with complete hepatic ischemia and reperfusion. Forty-five minutes of complete hepatic ischemia followed by reperfusion caused an increase in serum GPT and a fall in hepatic GSH but no increase in hepatic lipid peroxidation products. Chemical depletion of hepatic GSH with diethyl maleate did not cause hepatocellular injury but augmented hepatic ischemia/reperfusion-induced SGPT release and promoted lipid peroxidation. Pretreatment with the selective, membrane-permeable oxygen radical scavenger dimethyl sulfoxide protected against the ischemia/reperfusion-induced drop in hepatic GSH but did not reduce SGPT release in normal rats. In rats sensitized to oxidative injury by depletion of endogenous GSH with diethyl maleate the oxygen radical scavenger protected against ischemia/reperfusion-induced lipid peroxidation and reduced the release of SGPT. These findings suggest that the rich hepatic supply with endogenous GSH has a crucial role in the protection against oxygen radical injury following short periods of total hepatic ischemia. Oxygen radical injury only occurs after depletion of these endogenous GSH stores.     

Cold-blood potassium cardioplegia: evaluation of glutathione and postischemic cardioplegia

Potassium (34 mEq/L) cardioplegia was induced with cold blood (CBK) in three groups of six dogs undergoing 60 minutes of myocardial ischemia at a systemic temperature of 27 degrees +/- 2 degrees and a myocardial temperature of 7 degrees +/- 2 degrees C (crushed ice). Group 1 (CBK) animals were reperfused initially with 400 ml cold blood over 8 to 10 minutes at increasing pressures of up to 75 mm Hg. Group II (CBK-K) dogs were reperfused in the same manner as Group I with the addition of potassium chloride, 30 mEq/L. In Group III (CBKG-KG) glutathione, 30 mg/100 ml, was added to both the pre- and postischemic perfusions with CBK. After 30 minutes of reperfusion control studies were repeated. Heart rate, peak systolic pressure, rate of rise of left ventricular pressure, maximum velocity of contractile element, pressure-volume curves, coronary flow distribution, muscle stiffness, and heart water were not significantly different from control values. Total coronary flow and myocardial uptake of oxygen, lactate, and pyruvate did not serve to separate the three groups; the same was true for right ventricular creatine phosphate, adenosine triphosphate, and adenosine diphosphate during ischemia and recovery. Ultrastructural myofibrillar lesions were noted in all groups. thus, postischemic cardioplegia and use of a physiological reducing agent do not enhance CBK cardioplegia with topical and systemic hypothermia.     

Renal preservation after warm ischemia using oxygen free radical scavengers to prevent reperfusion injury

Oxygen free radicals damage kidneys and accumulate during the period of preservation prior to transplantation. We hypothesized that a perfusate containing either an oxygen free radical scavenger such as ceruloplasmin, or an iron-chelating agent such as deferoxamine, would improve kidney preservation. Thirty-eight mongrel dogs underwent autotransplantation of the left kidney after 30 min of warm ischemia and 48 hr of machine perfusion (MOX-100, Water Instruments, Rochester, MN) at 5 degrees C and pH of 7.4. The right kidney was removed at the time of autotransplantation. Four blind code-labeled preservation solutions were tested. SGF-I was used for the control group (Group 1, n = 13), and the remaining animals were transplanted with kidneys preserved with one of three solutions modified from the basic SGF-I solution: Group 2, SGF-I plus deferoxamine (656 mg/liter), n = 8; Group 3, SGF-I ceruloplasmin enriched (72 mg/dl), n = 8; and Group 4, SGF-I ceruloplasmin reduced (3.4 mg/dl), n = 9. Serum creatinine levels were measured daily for 2 weeks and survival curves for each of the four groups were estimated by the Kaplan-Meier method. Peak mean serum creatinine levels +/- standard errors in Groups 1 through 4 were 12.6 +/- 1.97, 7.8 +/- 0.90, 7.1 +/- 1.26, and 8.2 +/- 1.09, respectively. Repeated measures analysis of variance showed statistically significant differences between the groups with respect to their serum creatinine profiles (Wald's test x2 with 3 df = 22.39, P value less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

氟比洛芬酯预处理明显减轻冷水束缚应激大鼠氧自由基介导性急性胃黏膜损害

目的 探讨氟比洛芬酯预处理对冷水束缚应激大鼠急性胃黏膜损害及胃黏膜组织丙二醛(malondialdeyde,MDA)、超氧化物歧化酶(superexide dismuklse,SOD)及髓过氧化物酶(myelopemxidese,MPO)的影响.方法 30只成年雄性wistar大鼠随机分成3组,正常(A组,n=10),应激组(B组,n=10),氟比洛芬酯预处理组(c组,n=10).B、C组用浸水(水温20℃±1℃)束缚应激法复制急性胃黏膜病变模型.6 h后处死取胃组织标本,10×解剖显微镜下观察各组胃黏膜损伤指数(gastic mucosal lesion index,GI).光学显微镜下观察各组标本组织学改变,并用比色法测定各组胃组织中MDA、SOD与MPO的含量变化.结果 ①B组胃黏膜旱暗红色.有大量点状出血、线状出血或斑片状糜烂.黏膜下血管和皮区血管充血,光镜下上皮细胞黏液层变薄、缺失.细胞间隙变大.有不同程度的坏死;C组胃腔液体呈淡黄色,黏膜出血或糜烂程度较B组明显好转,光镜下可见上皮细胞黏液层有少许缺损,但基本完整.②与B组比较.C组急性胃黏膜损伤指数(gastic mucosal iesion index,GI)显著降低(P<0.05),MDA、MPO值显著降低.SOD值升高(P<0.05).结论 氟比洛芬酯预处理可明显减轻冷水束缚应激大鼠氧自由基介导性急性胃黏膜损害,其机制可能与减少脂质氧化及增加清除氧自由基的能力有关.     

谷胱甘肽S-转移酶基因多态性与膀胱癌易感性关系

目的 探讨谷胱甘肽S-转移酶(GST)基因多态性和环境危险因素与膀胱癌易感性的关系.方法 采用病例对照研究方法,应用多重聚合酶链反应和等位基因特异性聚合酶链反应基因分析技术,分别检测108例膀胱癌患者和112例非肿瘤患者GST基因型:GSTM1和GSTT1空白基因出现情况及GSTP1-105和GSTP1-114两个常见突变位点的多态性.采用x2检验和多因素、单因素非条件Logistic分析基因多态性与膀胱癌易感性的关系以及基因、环境交互作用对膀胱癌发病的影响.结果 膀胱癌组和对照组中GSTM1缺失型(GSTM1-null)、GSTT1缺失型(GSTT1-null)及GSTM1/GSTT1联合缺失型的分布频率分别为68.5%和54.5%,65.7%和51.8%,26.9%和15.2%,差异具有统计学意义(P=0.032,P=0.036,P=0.012).GSTP1-105和GSTP1-114突变纯合子(Val/Val)和突变杂合子(Ile/Val)基因型在2组中分布差异无统计学意义(P＞0.05),膀胱癌组中未检出GSTP1-114突变纯合子(Val/Val).GSTM1-null、GSTT1-null及GSTT1/GSTM1-null基因型在中、低分化(G2/G3)组中分布频率分别为58.3%、57.4%和24.1%,在临床分期T2～T4组中分布频率分别为47.2%、52.3%和16.7%,分别与高分化(G1)、临床分期Tis～T1组相比,差异有统计学意义(P＜0.05);吸烟、职业暴露、嗜食熟肉是膀胱癌发病的危险因素,OR值(95%可信区间)分别为7.510(2.899～19.454)、2.227(1.169～4.240)、6.147(2.451～15.419);女性膀胱癌发病危险性明显低于男性,OR值为0.273(0.133～0.557).吸烟、职业暴露、嗜食熟肉与GSTM1-null和GSTT1-null基因型存在不同程度的交互作用,使膀胱癌发病风险增高(P＜0.05). 结论 GSTM1-null和GSTT1-null基因型与膀胱癌易感性有关,且携带该基因型的膀胱癌患病群体肿瘤恶性程度较高、预后较差;吸烟、职业暴露、嗜食熟肉是膀胱癌发病危险因素,与GST易感基因型交互作用使膀胱癌发病风险增高.     

Return to work after foot and ankle injury

A surgeon faces important issues when returning a patient to the workplace, including workers' compensation issues and the need for physician guidance. These patients deserve appropriate diagnosis and treatment protocols, and the other members of the workers compensation treatment team can help maximize the effects of careful guidance by a physician. The speediest possible return to the workplace is usually best for all concerned, the injured worker, the employer and the physician. The treatment team within the workers' compensation system assists in the facilitation and coordination of the medical care and assists in bringing the patient back to work. The team includes the nurse case manager, the insurer, and physical and occupational therapists. Appropriate use of work hardening programs or functional capacity evaluations can be valuable in the return-to-work effort. Physicians who make use of these resources may find it easier to guide this process efficiently to achieve the desired outcome of return to work of the injured worker.     

Dupuytren's disease: relation to work and injury

The present status of adjudication for workers claiming compensation for Dupuytren's disease is inconsistent and, therefore, unfair to both workers and employers. In some Eastern European countries Dupuytren's disease is classified as an industrial disease, whereas in other countries it is considered to have no relation to manual work or hand injury. In jurisdictions that sometimes award compensation, the reasons for acceptance or rejection of a claim vary from case to case and are not necessarily based on our present knowledge of the disease. The purpose of this communication is to highlight the features of Dupuytren's disease that are pertinent to manual work and hand injury and to suggest guidelines that would provide some consistency in the adjudication process. It is hoped that these guidelines would be valuable to the individual surgeon, insurance agencies, and compensation boards.     

Return to work after spinal cord injury

OBJECTIVE: To describe predictors of return to work after spinal cord injury (SCI), in particular the physical intensity of the pre-injury job. STUDY DESIGN: Survey. SETTING: Patients' home. METHODS: We interviewed 234 persons with a spinal cord injury (SCI) between 18 and 65 years of age and who were gainfully employed at the time of the injury. Possible predictors were tested with logistic regression analyses. RESULTS: After the SCI, only 37% of the persons were gainfully employed. People with heavy and strenuous physical work pre-injury regained work in only 25%, respectively 21%. Being male (Odds Ratio (OR) 3.70), light to moderate physical job pre-injury (OR 3.16), high Barthel Index (OR 2.76), high educational level (OR 2.12) and education post-injury (OR 2.14) were significant predictors for returning to gainful work after injury. Many unemployed persons thought they were capable of working. CONCLUSIONS: Only a minority returned to gainful employment after SCI even to a physically less demanding job. In addition to intensive inpatient re-education, long-term support in job seeking is very important, including switching to a less demanding job.     

Resistance of ALR/Lt islets to free radical-mediated diabetogenic stress is inherited as a dominant trait.

ALS/Lt and ALR/Lt are inbred mouse strains selected for susceptibility and resistance to alloxan (AL)-induced diabetes. Within 24-h after AL administration in vivo, ALS/Lt islets were distinguished from ALR/Lt islets by more extensive necrotic changes. Within 7 days post-AL, ALS/Lt mice exhibited hyperglycemia and hypoinsulinemia, whereas ALR/Lt mice maintained normal plasma insulin and glucose levels. We have recently shown that resistance in ALR/Lt correlated with constitutively elevated systemic (and pancreatic) free radical defense status. In the present report, we examined whether ability to detoxify free radical stress extended to the level of ALR/Lt pancreatic islets. Cultured ALS/Lt islets exposed for 5 min to increasing (0-3 mmol/l) AL concentrations in vitro exhibited an 80% decline in numbers of intact islets after a subsequent 6-day culture period, as well as a 75% reduction in islet insulin content and a 94% decrease in glucose-stimulated insulin secretory capacity. In contrast, ALR/Lt islets remained viable and retained glucose-stimulated insulin secretory capacity as well as normal insulin content. This ALR/Lt islet resistance extended to hydrogen peroxide, a free radical generator whose entry into beta-cells is not dependent on glucose transporters. The elevated antioxidant defenses previously found in ALR/Lt pancreas were extended to isolated islets, which exhibited significantly higher glutathione and Cu-Zn superoxide dismutase 1 levels compared with ALS/Lt islets. A dominant genetic trait from ALR/Lt controlling this unusual AL resistance was indicated by the finding that reciprocal F1 mice of both sexes were resistant to AL administration in vivo. A backcross to ALS/Lt showed 1:1 segregation for susceptibility/resistance, indicative of a single gene controlling the phenotype. In conclusion, the ALR/Lt mouse may provide important insight into genetic mechanisms capable of rendering islets strongly resistant to free radical-mediated damage.     

Influence of postischemic administration of oxyradical antagonists on ischemic injury to rabbit skeletal muscle

The aim of this study was to determine whether the administration of free radical antagonists, immediately before and during the early minutes of reperfusion, improves muscle survival 24 hr after a period of ischemia. Rabbit rectus femoris muscles were isolated, made ischemic for 3½ hr and treated with either desferrioxamine (DFX), an Fe³⁺ chelator, superoxide dismutase and catalase (SOD & CAT), which quench superoxide and hydrogen peroxide, or allopurinol, an inhibitor of xanthine oxidase (XO). After 24 hr reperfusion, muscle viability (±s.e.m.), measured by the nitro blue tetrazolium (NBT) vital staining technique, was 41.6 ± 11.3% for saline-treated ischemic controls, 30.6 ± 7.6% for DFX-treated, 46.7 ± 10.3% for SOD & CAT-treated, and 43.3 ± 9.5% for allopurinol-treated muscles. None of the treated groups differed significantly from the ischemic control group. Tissue myeloperoxidase, ATP and reduced glutathione levels, and plasma lactate dehydrogenase (LDH) and aspartate transaminase (AST) levels were increased by ischemia and reperfusion in all groups, but the changes did not differ between the treatment groups. Levels of XO in the rabbit muscle were determined and found to be very low in both normal and postischemic muscle. As XO is the target enzyme of allopurinol, its absence provides a basis for the lack of effect of this agent. However, it is not clear why DFX and SOD & CAT had no protective effect © 1997 Wiley-Liss, Inc MICROSURGERY 17:517–523 1996     

Increased susceptibility of hypertrophied hearts to ischemic injury

Cardiac hypertrophy is an adaptive response that compensates for increased workload by normalizing wall stress and preserving cardiac contractile function. In advanced stages, however, clinical and experimental studies have shown that when the high workload is maintained, hypertrophy progresses to ventricular dilatation, contractile dysfunction, and decreased tolerance to ischemia/reperfusion. Development of hypertrophy is accompanied by distinct qualitative and quantitative changes in contractile protein expression and isoform switching, cytosolic calcium regulation, and substrate delivery and use. We have focused our investigations on changes in substrate delivery and capillary density in pressure overload hypertrophy and on the effects that these changes have on tolerance to ischemia/reperfusion. This report summarizes our work in this area using a model of aortic banding in 10-day-old rabbits, which exhibits the same pattern of concentric hypertrophy early, followed by ventricular dilatation and contractile dysfunction that is clinically apparent.     

Asanguineous reperfusion in a canine model of cardiopulmonary bypass and controlled postischemic work

This study was designed to test the hypothesis that asanguineous reperfusion with a standard crystalloid cardioplegic solution results in improved myocardial salvage after a period of global ischemia. Four groups of 6 dogs each were placed on cardiopulmonary bypass. Control group A (work only) performed two hours of controlled work by contracting against a saline-filled left intraventricular balloon. Control group B (ischemia only) underwent 45 minutes of global normothermic ischemia before simple blood reperfusion while supported on bypass. Groups C and D were subjected to ischemia and reperfusion as in group B, followed by controlled work stress as in group A. Group D, however, received 500 mL of St. Thomas' Hospital solution immediately before blood reperfusion. Morphological analysis showed no significant injury in groups A and B, whereas group C had 11.4% +/- 2.4% necrosis of heart mass versus 2.5% +/- 1.1% in group D (p less than 0.001). Biochemical data from left ventricular biopsies showed no significant differences between groups B, C, and D. Functional analyses showed deterioration of diastolic compliance in group C (p less than 0.05), although a significant difference in systolic functional indexes could not be detected. Myocardial protection and salvage was improved by initial reperfusion with an asanguineous cardioplegic solution versus reperfusion with blood alone.