ObjectiveTo study the protective effects of ischemia preconditioning (IPC) on cryopreservation injury of rat liver.MethodsThe model of isolated nonrecirculated perfusion rat liver was established. The grafts were treated with IPC in different time (ischemia preconditioning time in IPC1 group was 5 min; the time in IPC2 group was 10 min; while the time in IPC3 group was 15 min). The cryopreservation injury of the grafts in each group was determined and compared. ResultsThe levels of aspartate transaminase (AST) and alanine transaminase (ALT) in the effluent solutions in IPC1 group were (40.1±6.3) U/L and （17.1±0.5） U/L respectively, and IPC2 group (53.6±3.7) U/L, （19.7±0.5） U/L, which were much lower than those of nonpreconditioning (NPC) group 〔(64.5±8.2) U/L, （23.8±3.9） U/L〕 (P＜0.05). Those in IPC1 group was much lower than those in IPC2 group and IPC3 group 〔(63.8±7.2) U/L,（22.8±2.5） U/L〕 (P＜0.05). The level of lactic acid dehydrogenase (LDH) in NPC group (104.3±20.6) U/L, IPC1 group (84.1±19.7) U/L, IPC2 group (90.5±21.1) U/L, and IPC3 group (103.1±18.5) U/L were of no significant difference (Pgt;0.05). The contents of bile product and the hepatocellular contents of ATP in IPC1 group were (53.5±10.2) μl and (6.15±0.65) μmol/g respectively, and IPC2 group (41.5±8.1) μl, (4.77±0.21) μmol/g, which were much higher than those NPC group 〔(22.8±9.7) μl, (2.62±0.34) μmol/g〕 (P＜0.05). Those in IPC1 group were much higher than those in IPC2 group and IPC3 group 〔(27.5±2.8) μl, (2.61±0.29) μmol/g〕 (P＜0.05). The contents of malondialdehyde (MDA) in liver tissue in IPC1 group was (4.36±0.26) nmol/gand IPC2 group (5.51±0.13)
Objective To investigate the protective effects of ischemic postconditioning (IPo) on ischemiareperfusion (I/R) myocardium and the relationship with mitochondrial adenosine triphosphate (ATP) sensitive K+ channels (mitoKATP) and provide evidences to the development of druginduced postconditioning. Methods Langendorff models were established in 40 Wistar rats which were divided into 5 groups by random number table with 8 rats in each group. Normal control group(NC group): the rat hearts were continuously reperfused by KrebsHenseleit bicarbonate buffer (K-HB) for 100 min without any other treatment; I/R group: the rat hearts underwent a 40-min global ischemia followed by a 60-min reperfusion; IPo group: after a 40-min global ischemia, the process of 10-second reperfusion followed by a 10-second ischemia was repeated 6 times, then there was a continuous 58min reperfusion; 5-hydroxydecanoic acid(5-HD) group: after a 40min global ischemia, hearts with 5HD(100 μmol/L) K-HB were reperfused for 15min and then perfused without 5HD for 45min；IPo+5-HD group: after a 40-min global ischemia, the process that the isolated hearts with 5-HD(100 μmol/L) KHB were reperfused for 10second followed by a 10second ischemia was repeated 6 times, then the hearts with 5-HD(100 μmol/L) KHB were continuously [CM(159mm]perfused for 13-min followed by reperfusion without 5-HD(100 μmol/L) K-HB for 45-min. The cardiac function,coronary flow(CF), cardiac troponin I(cTnI) content in coronary effluent, the area of acute myocardial infarction (AMI) and myocardial ultrastructure were observed. Results Left ventricular developed pressure(74.3±3.3 mm Hg vs. 57.1±3.3 mm Hg,t=1300, P=0.000),+dp/dtmax(1 706.6±135.6 mm Hg/s vs. 1 313.3±96.2 mm Hg/s,t=6.28,P=0.000),－dp/dtmax(1 132.8±112.1 mm Hg/s vs. 575.7±67.7 mm Hg/s,t=13.48, P=0.000) and CF(6.49±0.30 ml/min vs. 3.70±0.24 ml/min,t=28.6,P=0.000) in IPo group were higher than those in I/R group. Left ventricular enddiastolic pressure(10.9±1.7mm Hg vs. 26.2±1.5 mm Hg,t=－19.21, P=0000)and cTnI content in coronary effluent (0.62±0.01 ng/ml vs. 0.71±0.01 ng/ml, t=-12.00,P=0.000) were lower than those in I/R group; the area of AMI decreased 20.8％ compared with that in I/R group (Plt;0.05). The myocardial protective effect in IPo+5HD group was similar with that in IPo group, but lower than that in IPo group. The electron microscope showed that IPo and IPo+5HD could reduce myocardial fiber damage and mitochondrial damage caused by I/R. Conclusion IPo can protect I/R myocardium, which is achieved mainly by activating mitoK-ATP channels.
Objective To investigate the relationship between graded spinal cord ischemia/reperfusion injury and somatosensory evoked potentials(SEP),neurologic function score（NFS）and the histopathological changes of spinal cord. Methods Forty rabbits were randomized and equally divided into 4 groups: shamoperation group, ischemia for 30min, 45min and 60min groups. The spinal cord ischemiareperfusion injury model was created by occlusion of the abdominal aorta in rabbits. SEP was monitored before ischemia,5,10minutes after ischemia, 15, 30 minutes, 1,2, 24 and 48 hours after reperfusion. NFS was evaluated at 6,12,24 and 48 hours after reperfusion.The pathological changes of spinal cord were observed after reperfusion 48 hours. Results The pathological characters with mild,moderate and severe spinal cord ischemia/reperfusion injury could be simulated by declamping after 30， 45 and 60 minutes infrarenal aorta crossclamping. SEP amplitude returned to normal after reperfusion 15 minutes（Pgt;0.05）and SEP latency returned to normal after reperfusion 30 minutes（Pgt;0.05）during mild spinal cord ischemia/reperfusion injury.SEP amplitude returned to normal after reperfusion 30 minutes（Pgt;0.05）and SEP latency returned to normal after reperfusion 60 minutes（Pgt;0.05）during moderate spinal cord ischemia/reperfusion injury. SEP latency increased and SEP amplitude decreased during severe spinal cord ischemia/reperfusion injury,compared with other groups, there were significant differences in SEP latency and SEP amplitude by clamping the infrarenal aorta for 60min（Plt;0.01）. With graded spinal cord ischemia/reperfusion injury, compared with shamoperation group, spinal cord ischemiareperfusion groups had significant differences in NFS（Plt;0.01）. Conclusion SEP is much quicker in the recovery of amplitude than latency during spinal cord ischemia/reperfusion. SEP is a sensitive and accurate index for spinal cord function during ischemia/reperfusion injury. SEP monitoring spinal cord ischemia/reperfusion injury during operation provides experimental basis for clinical application.
Objective To investigate the pathological changes in the neuromuscular junction during ischemiareperfusion(IR) in the skeletal muscle. Methods Forty-eight healthy adult Wistar rats (24 male, 24 female) were equally randomised into the following 6 groups: Group A (control group): no ischemiareperfusion; Group B: ischemia by clamping the blood vessels of the right hindlimb for 3 hours; Group C: ischemia by clamping for 4.5 hours;Group D: ischemia by the clamping for 4.5 hours followed by reperfusion for 1.5hours; Group E: ischemia for 4.5 hours followed by reperfusion for 24 hours; and Group F: ischemia for 4.5 hours followed by reperfusion for 2 weeks. Then, the medial head of the gastrocnemius muscle flap model was applied to the right hindlimb of each rat. The medial head of the gastrocnemius muscle was isolated completely,leaving only the major vascular pedicle, nerve and tendons intact.The proximal and distal ends (tendons) were ligated while the vessel pedicle was clamped. And then, Parameters of the muscle (performance,contraction index,colour,edema,bleeding) were observed. The muscle harvested was stained with gold chloride(AuCl3) and the enzymhistochemistry assay (succinate dehydrogenase combined with acetylcholine esterase) was performed. Morphology and configuration of the neuromuscular junction were observed during the ischemiareperfusion injury by means of the AuCl-3 staining. The result of the enzymhistochemical reactions was quantitatively analyzed with the computer imageanalysis system. And then, additional 5 rats were prepared for 3 different models identical with those in Groups A, C and E separately. The specimens were harvested from each rat and were stained with HE and AuCl-3, and they were examined under the light microscope. Results During the period of ischemia, the skeletal muscle of Group B showed the colour of purple and edema.The colour and edema became worse in Group ,while dysfunction of elasticity and contraction appeared obviously with plenty of dark red hemorrhagic effusion at the same time.After reperfusion,the color and edema of muscle in Group D became improved while the elasticity and function of contraction was not improved. Hemorrhagic effusion of Group D turned clearer and less than Group C.Group E was similar to Group D in these aspects of muscle except for much less hemorrhagic effusion. Skeletal muscle in Group F showed colour of red alternating with white, adhesion,contracture of muscle, exposure of necrotic yellow tissue and almost lost all its functions. The AuCl3 staining showed that during IR, necrosis of the myocytes was followed by degeneration of their neuromuscular junctions, and finally the nerve fibers attached to these neuromuscular junctions were disrupted like the withering of leaves. The enzymhistochemistry assay showed thatthere was no significant difference in the level of acetylcholine esterase between the ischemic group (Groups B and C) and the control group (Group A) (Pgt;0.05). However, the level of acetylcholine esterase in all the reperfused groups (Groups D, E and F) decreased significantly when compared with the control group（Group A）and the ischemic groups (Groups B and C) (Plt;0.01). Conclusion The distribution of the nerve fibers and the neuromuscular junctions in the mass of the muscles is almost like the shape of a tree. The neuromuscular junction seems to be more tolerant for ischemia than the myocyte. Survival ofthe neuromuscular junction depends on its myocytes alive. Therefore, an ischemiareperfusion injury will not be controlled unless an extensive debridement of the necrotic muscle is performed.
Objective To observe the protective role of the ectogenesis zinc on the cells in rat flap with ischemia reperfusion injury and study the mechanisms. Methods A right low abdominal island flap was created in Wistar rats. Fortyeight rats were randomly divded into 3 groups (n=16):the control group, the ischemia reperfusion group and adding zinc ischemia reperfusion group.The content of malondialdehyde(MDA) and the activity of myeloperoxidase(MPO) were measured by thiobarbituric acid methods and colorimetry. The location of expression of MT was observed,and the image analysis was performed. The quantity of MT was represented by the integratial optical density. The ultrastructure changes of skin flap with ischemia reperfusion injury and the flap viability were observed. Results In the ischemia reperfusion injury flaps, the content of MDA and MPO show no statistically significant difference among the control group,IR group and the adding-zinc-IR group (P＞0.05). Compared with the control group at 1 h and 24 h of reperfusion, the level of MDA increased 62.2% and 136.4%(P＜0.01) in the IR group, which increased 11.3% and 33.2%(P＜0.01) in the adding-zinc-IR group. The activity of MPO increased 238.4% and 503.4%(P＜0.01)in the IR group when compared with the control group, and increased 17.9%and 24.1%(P＜0.05) when compared with the adding-zinc-IR group. In the ischemia reperfusion injury falps, the content of MT in the control group and the IR group is too minimal to measure. While the content ofMT in the adding-zinc-IR group is 45.30±7.60. At 1 h and 24 h of reperfusiion, the content of MT in the adding-zinc-IR group increased 41.5% and 44.9% (P＜0.01) compared with the IR group, and increased 119.9% and 234.6% (P＜0.01) compared with the control group. The flap viability is 100% in the control group, 19.65%±4.38% in the IR group, and 24.99%±5.12% in the adding-zinc-IR group, which increased 27.2% (P＜0.05) compared with IR group. Conclusion Many kinds of cells in skin flap with ischemiareperfusion injury can be protected by ectogenesis zinc and the flap viability increases significantly.