犬移植肺缺血再灌注实验动物模型的建立

目的为深入研究缺血再灌注对移植肺功能损害的发生机制,克服以往体外肺灌注模型存在的局限性,我们建立了模拟的犬肺自体原位移植模型。方法选用杂种犬１５只,建立左肺原位缺血模型,３小时后（热缺血１小时,冷缺血２小时）,开放左肺动、静脉,结扎对侧肺动脉,监测缺血３小时再灌注６小时过程中动脉血氧分压（ＰａＯ２）、动脉血二氧化碳分压（ＰａＣＯ２）、动脉血氧饱和度（ＳａＯ２）、平均肺动脉压（ＭＰＡＰ）的变化。结果再灌注前后的ＰａＯ２,ＭＰＡＰ有差异（Ｐ＜０．０５）。结论该动物模型能够较好地反映缺血再灌注对犬移植肺功能的损害,并具有一定的可复制性和扩展性     

Evaluation of Ischemia-Reperfusion Liver Injury by Near-Infrared Spectroscopy in an Experimental Swine Model: The Effect of Desferoxamine

ABSTRACT

Introduction: Ischemia-reperfusion (I-R) injury has long been regarded a primary factor for the physiological dysfunction that can occur following major liver resection performed under vascular control. The aim of our study was to assess the effect of treatment with desferoxamine (DFO), a potent antioxidative agent, monitoring the I-R injury on a porcine model of major hepatectomy. Materials and Methods: Twelve female pigs were allocated to control (n = 6) and DFO groups (n = 6) and underwent 30 min of liver ischemia, during which a ≥30% hepatectomy was performed, followed by six hours of postoperative monitoring. The DFO group animals were preconditioned with a continuous iv solution of DFO to a total dose of 100 mg/kg during their postoperative period. Liver remnants (≈70% of initial liver volume) were evaluated by means of infrared spectroscopy, serum lactate measurement of the systemic, portal and hepatic vein blood, and by immunohistochemical assessment of apoptosis in consecutive liver biopsies. Results: DFO group demonstrated considerably faster restoration of tissue oxygenation (92.33% vs. 80%, p < .05) and serum lactate values (1.23 mmol/l vs. 2.27 mmol/l, p < .05). Moreover, apoptosis as estimated by TUNEL and caspase-3 staining was significantly lower in the DFO group (0.06% vs. 1.17% and 1.17% vs. 2%, respectively, p < .05). The severity of the I-R injury showed a linear correlation to the restoration of tissue oxygenation, as estimated by infrared-spectroscopy (r² = 0.81, p < .01). Conclusion: Iron chelation with DFO appears to attenuate I-R injury of the liver remnant following hepatectomy, as reflected by faster restoration of tissue oxygenation and lower apoptotic activity.     

Skeletal Muscle Ischaemia-reperfusion Injury: Further Characterisation of a Rodent Model

BACKGROUND: Postischaemic damage in skeletal muscle may be reflected in changes to microvascular blood flow, vascular permeability, and subsequent tissue viability. Previous preclinical studies have not addressed all these parameters, and have not used periods of ischaemia and reperfusion relevant to the clinical setting. This study aimed to develop an animal model hindlimb ischaemia-reperfusion to simulate acute lower limb ischaemia. METHODS: A rodent model of hindlimb tourniquet-induced ischaemia-reperfusion was employed. Gastrocnemius muscle blood flow (GMBF; radio-labelled microspheres), oedema (GMO; using a wet:dry ratio method) and viability (GMV; histochemistry and computerised planimetry) were quantified. RESULTS: 6 h ischaemia per seresulted in neither muscle oedema nor loss of viability, but these changes were apparent following 4 h reperfusion. Early reperfusion at 10 min demonstrated low reflow, with GMBF improving at 120 min before declining sharply at 240 min. CONCLUSION: Prolonged hindlimb ischaemia followed by reperfusion in this rodent model caused significant reductions in gastrocnemius muscle blood flow, associated with muscle oedema and necrosis. These three parameters have not been previously reported together in the same model. This reproducible model could be used in the evaluation of potential therapeutic intervention strategies aimed at ameliorating skeletal muscle reperfusion injury.     

An Experimental Model of Intraoperative Venous Injury in the Rat

Intraoperative obliteration of cerebral veins occasionally causes unexpected severe complications, especially in elderly patients. However, very title information is available on the pathophysiology of cerebral venous circulation disturbance. Occlusion of cortical veins in rats by a photochemical thrombotic technique is a less invasive, clinically relevant and reproducible model that is suitable for the study of venous circulation disturbance. In the present study, 54 male Wistar rats were used. We examined changes of the cerebral venous flow pattern by fluorescence anglography and brain damage histologically in a one- or two-(cortical) vein occlusion model using a photochemical thrombotic technique. Approximately 30% (9 of 27) of animals in the single-vein occlusion group and 90% (15 of 17) of those in the two-vein occlusion group had microcirculation perturbation, which results very soon in the formation of venous thrombus accompanied by severe venous infarction. In addition, infarction size in the two-vein occlusion group (9.7 ± 3.2%) was significantly larger than in the single-vein group (2.9 ± 1.3%) (unpaired T-test, P < 0.01).

In conclusion, the photochemical dye technique of attaining cerebral venous occlusion is a worth while addition to the study of circulation perturbations of the brain.

     

Urinary Apolipoprotein M Could Be Used as a Biomarker of Acute Renal Injury: An Ischemia-Reperfusion Injury Model of Kidney in Rat

Background

It has been well documented that apolipoprotein M (apoM) is principally expressed in hepatocytes as well as renal tubular epithelial cells. The importance of apoM in the kidney is unknown. In the present study we examined urinary any apoM after short-term ischemia-reperfusion injury (IRI) of kidney in a rat model.

Methods

The kidneys of 11 male Sprague-Dawley rats were rendered ischemic for 45 minutes followed by different intervals of reperfusion. Serum and urine apoM concentrations were determined using a dot-blot analysis with specific rabbit anti-human apoM antibodies that cross-react with rat apoM. Serum concentrations of blood urea nitrogen (BUN) and creatinine (Cr) were determined using standard clinical automated analyses.

Results

BUN was significantly elevated after 45 minutes of ischemia followed by 24 hours of reperfusion; serum Cr concentrations were also significantly increased at 6 and 24 hours of reperfusion. Interestingly, similar to BUN and Cr, serum apoM concentrations were significantly increased after ischemia for 45 minutes alone and after 2 hours of reperfusion. Urinary apoM concentrations were obviously increased after 2 h as well as 6 hours of reperfusion.

Conclusion

apoM showed characteristics of an acute-phase reactive protein; its occurrence in urine may be considered to be a biomarker of acute renal injury.     

Axonal Sprouting Following Incomplete Spinal Cord Injury: An Experimental Model

Abstract

Recovery of function following incomplete spinal cord injury may in part result from growth of new connections by spared descending pathways. It has been difficult to demonstrate such anatomical reorganization with traditional anatomic techniques. This study utilizes an immunocytochemical method to demonstrate axonal growth cones within the lumbar spinal cord in rats recovering from an incomplete midthoracic spinal cord injury. Adult rats underwent subtotal section of the midthoracic cord sparing the left lateral funiculus and a portion of the left ventral funiculus. Light microscope immunocytochemistry was performed on sections of lumbar spinal cord with antibodies to identify sprouting axons. These antibodies were used to determine the distribution of growth cones on both sides of the lumbar spinal cord in experimental and control animals. Growth cones were first observed three days after the spinal cord lesion. Specific labeling, similar in appearance to previous reports of growth cone identification, was apparent within the intermediate gray and ventral horns on both sides of the cord. These data support the hypothesis of collateral sprouting distal to the lesion site following incomplete spinal cord injury. It further supports the idea that recovery of function following incomplete spinal cord injury is, in part, mediated by spared descending pathways. (J Spinal Cord Med 1997; 20:200-206)     

Sympathetic Nerves Do Not Affect Experimental Ischemia-Reperfusion Injury of Rat Liver

Background

We investigated whether sympathetic, noradrenergic nerves participate in experimental acute ischemia-reperfusion injury of the rat liver.

Methods

Female Wistar rats (200-250 g body weight) were anesthetized with pentobarbital. After tracheotomy, we cannulated a carotid artery and jugular vein. The rats were divided in 2 groups (n = 8 per group). The control group received NaCl IV and the test group received the sympatholytic agent, guanethidine (3 mg/kg, IV). After 30 minutes of drug equilibration, laparotomy was performed to arrange the liver for temporary occlusion (by a ligature) of its vascular supply, corresponding with 70% reduction in hepatic blood flow. The rats were then allowed 60 minutes of equilibration. Thereafter, regional ischemia was induced for 30 minutes. The animals were then monitored for 2 hours of reperfusion. Blood samples for alanine aminotransferase (ALT) estimation (as a measure of injury to the parenchyma) were drawn immediately before ischemia, as well as 60 and 120 minutes after reperfusion. Readings of mean arterial pressure were taken during these times.

Results

After 2 hours of reperfusion, there were no significant differences between the groups with regard to ALT or mean arterial pressure.

Conclusion

Sympathetic, noradrenergic nerves did not affect experimental ischemia-reperfusion injury of rat liver in the current model.     

In Situ Intestinal Ischemia-Reperfusion Injury in the Pig: A Model Using the First Jejunal Artery for Flushing

Background We describe a new surgical technique of in situ intestinal ischemia–reperfusion injury in the pig, which includes transection of the small bowel, extrinsic autonomic denervation, lymphatic disruption, and finally in-situ cold ischemia of the graft by flushing through the first jejunal artery. Material and methods Ten female pigs were used for the study. All neural and lymphatic connections to the jejunoileum were transected. The stripped superior mesenteric vessels remained as the only connections. The skeletonized mesenteric vessels were clamped and the superior mesenteric artery was cannulated through the first jejunal artery. The isolated jejunoileum was flushed with cold IGL-1 solution. A small incision on the superior mesenteric vein was made to allow outflow of the effluent. After the flushing process was complete, the small incision in the superior mesenteric vein was closed and the vascular clamps were removed. The proximal 70% of the graft was resected. Results The mean preoperative weight of the animals was 25.8 ±7.6 kg. The mean duration of the operation was 242.0 ± 28.6 min. The mean cold ischemia time was 47.6 ± 3.9 min. All animals survived the procedure and were sacrificed at day 8. At sacrifice, there were no adhesions. The small bowel appeared normal. On intestinal histology, there were no significant changes between specimens obtained from the animal immediately at the end of cold flushing (T0), 2 h after reperfusion (T1), and at sacrifice (T2). Conclusions This novel technique for intestinal ischemia–reperfusion injury in the pig provides an extremely useful model for experimental studies of immunological and cold ischemia–reperfusion injury of transplanted small bowels.     

Adverse Maternal and Fetal Outcomes in a Novel Experimental Model of Pregnancy after Recovery from Renal Ischemia-Reperfusion Injury

BackgroundRecent clinical studies report that women with a history of AKI have an increased incidence of maternal and fetal adverse outcomes during pregnancy, despite fully recovering renal function prior to conception. The mechanisms contributing to such adverse outcomes in pregnancy after AKI are not yet understood.MethodsTo develop a rodent model to investigate fetal and maternal outcomes in female animals with a history of AKI, we used ischemia-reperfusion injury as an experimental model of AKI in female Sprague Dawley rats. The 12-week-old animals underwent warm bilateral ischemia-reperfusion surgery involving clamping of both renal arteries for 45 minutes or sham surgery (control). Rats were allowed to recover for 1 month prior to mating. Recovery from ischemia-reperfusion injury was confirmed by measurements of plasma creatinine and urinary protein excretion. We assessed maternal and fetal outcomes during late pregnancy on gestational day 20.ResultsAfter recovery from ischemia-reperfusion injury, compared with healthy sham-surgery controls, dams exhibited pregnancy-induced renal insufficiency with increases in plasma creatinine and urea, along with increased urinary protein excretion. Additionally, recovered ischemia-reperfusion dams experienced worse fetal outcomes compared with controls, with intrauterine growth restriction leading to higher rates of fetal demise and smaller pups.ConclusionsIn this rat model, despite biochemical resolution of ischemia-reperfusion injury, subsequent pregnancy resulted in maternal renal insufficiency and significant impairments in fetal growth. This mirrors findings in recent reports in the clinical population, indicating that this model may be a useful tool to further explore the alterations in kidney function after AKI in women.     

The Effect of Octreotide on Hepatic Ischemia-Reperfusion Injury in a Rabbit Model

Background

Hepatic ischemic-reperfusion injury (HIRI) is a major cause of morbidity and mortality following liver surgery. Octreotide (Oct) has been reported to improve hepatocellular energy metabolism in a rat HIRI model. This study was designed to evaluate whether Oct could protect the liver of rabbits against ischemic-reperfusion (I/R) injury.

Methods

Twenty-four adult New Zealand rabbits were randomly divided into a sham operated group (Control), an ischemia/reperfusion group (I/R), and an ischemia/reperfusion + Oct pretreatment group (I/R + Oct). The hemodynamic (mean arterial pressure [MAP] and heart rate [HR]) changes, liver enzymes (alanine aminotransferase [ALT], aspartate aminotransferase [AST], and lactate dehydrogenase [LDH]) release, inflammatory cytokines (tumor necrosis factor [TNF]α and interleukin [IL]-1β) levels, and endotoxin (ETX) levels were measured during I/R.

Results

Compared with the Control group, the MAP decreased and HR increased in I/R and I/R + Oct groups at ischemia 15 minutes (P < .05) but were less in the I/R + Oct group relative to the I/R group (P < .05). ALT, AST, LDH, IL-1β, and ETX levels were increased in the I/R and I/R + Oct groups at ischemia 30 minutes (P < .05), however, the increase was lower in the I/R + Oct group relative to the I/R group (P < .05). Bcl-2 expression in the I/R + Oct group was higher compared with other groups (P < .05) and Bax expression in the I/R group was reduced compared with other groups (P < .05). Hepatocellular damage in the I/R + Oct group appeared to be less than in the I/R group by microscopy.

Conclusions

Oct pretreatment attenuated hemodynamic changes and decreased liver enzyme changes induced by HIRI in a rabbit model. The protection mechanisms of Oct may be related to reduced ETX levels, down-regulation of the inflammatory cytokines TNFα and IL-1β, and inhibition of hepatocellular apoptosis, as well as the modulation of the mitochondrion-mediated Bcl-2/Bax apoptosis pathway. Based on our study it appears that Oct may be useful in decreasing liver injury after liver surgery and/or transplantation and may serve as a promising agent against HIRI.