Name: postconditioning with lactate-enriched blood for cardioprotection in st-segment elevation myocardial infarction
Uploaded: 2019-05-28T14:00:00.000+00:00
Duration: 5 min 26 s
Description: Watch this Scientific Journal Video about Postconditioning with Lactate-enriched Blood for Cardioprotection in ST-segment Elevation Myocardial Infarction at JoVE.com

The authors have no acknowledgments.

This protocol was approved by the ethics review board of the Saitama Municipal Hospital. The patient, whose case study is presented, provided written informed consent for the publication of his case and accompanying case images.
1. Set-up for PCLeB
<ol>
	<li>When starting coronary angiography (CAG), prepare the manifold for PCLeB. Connect one inlet line of the manifold to a contrast medium bottle and another inlet line to a 500 mL bottle of lactated Ringer&#8217;s solution mixed with 500 U of heparin sodium (Figure 2).</li>
	<li>Perform CAG and pinpoint the occluded lesion. After finishing CAG, introduce a guiding catheter, which is connected to the manifold via a Y-connector, into the aorta and engage it in the ostium of the culprit coronary artery as usual.</li>
	<li>Start wiring procedures with a balloon catheter placed inside the guiding catheter close to its outlet.</li>
	<li>If the coronary flow is incidentally restarted with the passage of the wire through the occluded lesion, move the balloon catheter quickly toward the culprit lesion to re-occlude the lesion site. After re-occluding the lesion site, resume and finish the wiring procedures. 
	NOTE: Such incidental coronary reflow sometimes happens; therefore, the balloon catheter is placed inside the guiding catheter during the wiring procedures.</li>
	<li>After finishing the wiring procedures, move the balloon catheter forward and place the balloon at the occluded lesion. Check the position of the balloon by contrast medium injection, to see if the balloon is appropriately placed at the occluded lesion. 
	NOTE: After finishing the checking of the balloon position, naturally, the contrast medium fills all the lumen from the manifold to the tip of the guiding catheter.</li>
	<li>Disconnect the syringe injector from the manifold and connect a 30 mL syringe to the manifold instead, which is used for lactated Ringer&#8217;s solution injection. 
	NOTE: No special syringe is needed for replacing the syringe, but an ordinary syringe for normal usage is fine as long as it has a &#8805;30 mL volume and a lock connector, so it will not be disconnected from the manifold during rapid injections of lactated Ringer&#8217;s solution.</li>
	<li>Fill the 30 mL syringe with 20&#8211;30 mL of lactated Ringer&#8217;s solution by sucking it from the bottle of lactated Ringer&#8217;s solution through the line connected to the inlet of the manifold.</li>
	<li>Remove air bubbles inside the 30 mL syringe if present. Disconnect the syringe and push them out and re-connect it to the manifold. Suck lactated Ringer&#8217;s solution from the bottle to make up the amount used for pushing air bubbles out. 
	NOTE: Here, the default set-up of the injection system for PCLeB has been completed; that is, the contrast medium (approximately 4 mL) fills all the lumen from the manifold to the tip of the guiding catheter, and 20&#8211;30 mL of lactated Ringer&#8217;s solution fills the 30 mL syringe connected to the manifold.</li>
</ol>
2. Details of PCLeB Procedures
<ol>
	<li>Once the balloon catheter is placed at the occluded lesion and the default set-up of the injection system is completed, inflate the balloon for 20&#8211;30 s at low pressure.</li>
	<li>Deflate the balloon and start the initial 10 s brief reperfusion. Immediately push out all the contrast medium (approximately 4 mL) prefilled in the lumen from the manifold to the tip of the guiding catheter, by injecting &#8805;4 mL of 20&#8211;30 mL of the lactated Ringer&#8217;s solution prefilled in the 30 mL syringe into the guiding catheter, to see if the coronary flow is recovered. 
	NOTE: Approximately 4 mL of contrast medium is usually enough to confirm coronary flow recovery.</li>
	<li>If the coronary flow is recovered, replenish the syringe with the same amount of lactated Ringer&#8217;s solution used for pushing the contrast medium out by sucking the solution from the bottle connected to the manifold through an inlet line, to prepare for lactated Ringer&#8217;s solution injection at the end of reperfusion. Jump to step 2.7. 
	NOTE: If coronary flow recovery is not observed, the true occluded lesion may be located distal to the current location of the balloon. Go to step 2.4.</li>
	<li>Empty the syringe by injecting all the lactated Ringer&#8217;s solution into the coronary artery.</li>
	<li>Fill the syringe with 4 mL of the contrast medium by sucking it from the bottle connected to the manifold through an inlet line and inject it all to the guiding catheter to fill the lumen. Then, fill up the syringe with 20&#8211;30 mL of lactated Ringer&#8217;s solution by sucking it from the bottle of the lactated Ringer&#8217;s solution connected to the manifold through an inlet line. 
	NOTE: Here, the default set-up of the injection system for PCLeB has been re-established.</li>
	<li>Move the balloon distally to the true lesion, and repeat the same procedures described in 2.1&#8211;2.6.</li>
	<li>Once coronary flow recovery is observed, and the balloon is confirmed to be placed at the culprit lesion, keep the balloon there throughout the PCLeB procedures. 
	NOTE: The desirable balloon size may be the same as the lesion diameter. By selecting such balloon size, instead of a smaller-sized balloon, the coronary flow may not be hampered by the deflated balloon left at the lesion site during each brief repetitive reperfusion.</li>
	<li>Start lactated Ringer&#8217;s solution injection (20 mL for the right coronary artery and 30 mL for the left coronary artery) directly into the culprit coronary artery through the guiding catheter 4&#8211;5 s before the end of each brief reperfusion. 
	NOTE: Usually, it takes 5&#8211;7 s to inject 20&#8211;30 mL of lactated Ringer&#8217;s solution, which may overlap with the subsequent brief ischemic period; this is an intentional delay. The aim of this intentional delay is to ensure that the lactate is trapped inside the ischemic myocardium.</li>
	<li>In the middle of lactated Ringer&#8217;s solution injection, let the secondary operator start balloon inflation so that the balloon inflation is completed a little before completion of lactated Ringer&#8217;s solution injection. 
	NOTE: This maneuver is intended so that the final small amount of lactated Ringer&#8217;s solution (ideally 2&#8211;3 mL) injected into the culprit coronary artery is bounced back from the inflated balloon. Through this approach, the lactated Ringer&#8217;s solution can be injected into the ischemic myocardium until the last moment of the balloon inflation process. This procedure requires cooperation between the primary operator who injects the lactated Ringer&#8217;s solution and the secondary operator who inflates the balloon.</li>
	<li>Reestablish the default set-up of the injection system, as described in 2.5., during 60 s ischemia for the next brief reperfusion.</li>
	<li>Deflate the balloon and start the next 10 s longer reperfusion after 60 s ischemia. Push out all the contrast medium prefilled in the lumen from the manifold to the tip of the guiding catheter, to reconfirm coronary flow.</li>
	<li>If the balloon is found not to be placed at the center of the lesion site, adjust the balloon position. 
	NOTE: This might be done during the first 10 s reperfusion, but it can be postponed until the next 20 s reperfusion because the 10 s duration may be slightly very short to do it.</li>
	<li>Start lactated Ringer&#8217;s solution injection into the culprit coronary artery through the guiding catheter 4&#8211;5 s before the end of each brief reperfusion</li>
	<li>In the middle of lactated Ringer&#8217;s solution injection, let the secondary operator start balloon inflation so that the balloon inflation is completed a little before completion of lactated Ringer&#8217;s solution injection.</li>
	<li>Reestablish the default set-up of the injection system during 60 s ischemia.</li>
	<li>Repeat these brief ischemia and reperfusion sequences seven times until 60 s reperfusion is performed twice (Figure 1). 
	NOTE: These procedures can be performed without disconnecting the 30 mL syringe from the manifold. The balloon inflation pressure can be increased gradually throughout the procedures, but high pressure is not recommended.</li>
</ol>
3. Alternative Default Set-up
NOTE: There may be an alternative to the default set-up, where replacing the syringe injector with a 30 mL syringe is not needed.
<ol>
	<li>After finishing the wiring procedures, disconnect the inlet line connected to the lactated Ringer&#8217;s solution bottle from the manifold and connect the 30 mL syringe filled with lactated Ringer&#8217;s solution directly with the inlet of the manifold instead. 
	NOTE: This is the alternative default set-up.</li>
	<li>During each brief ischemia, disconnect the 30 mL syringe, which was emptied at the end of the preceding brief reperfusion, from the manifold. Refill the syringe with lactated Ringer&#8217;s solution by sucking it directly from the bottle. Reconnect the refilled 30 mL syringe to the inlet of the manifold to reestablish the alternative default set-up. 
	NOTE: With this set-up, it is possible to obtain more clear images of the culprit coronary artery by making a larger volume of contrast medium available. However, the aforementioned set-up may be more convenient because disconnecting the 30 mL syringe from the manifold is not necessary for each refilling of the syringe with lactated Ringer&#8217;s solution.</li>
</ol>

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E., Abdelnoor, M., Hoffmann, P., Eritsland, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+ischemic+postconditioning+on+infarct+size+in+patients+with+ST-elevation+myocardial+infarction+treated+by+primary+PCI+results+of+the+POSTEMI+(POstconditioning+in+ST-Elevation+Myocardial+Infarction)+randomized+trial.">Effect of ischemic postconditioning on infarct size in patients with ST-elevation myocardial infarction treated by primary PCI results of the POSTEMI (POstconditioning in ST-Elevation Myocardial Infarction) randomized trial.</a> Journal of American Heart Association. 3 (2), e000679 (2014).</li><li>Engstr&#248;m, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+ischemic+postconditioning+during+primary+percutaneous+coronary+intervention+for+patients+with+ST-segment+elevation+myocardial+infarction:+a+randomized+clinical+trial.">Effect of ischemic postconditioning during primary percutaneous coronary intervention for patients with ST-segment elevation myocardial infarction: a randomized clinical trial.</a> JAMA Cardiology. 2 (5), 490-497 (2017).</li><li>Inserte, J., Barba, I., Hernando, V., Garcia-Dorado, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Delayed+recovery+of+intracellular+acidosis+during+reperfusion+prevents+calpain+activation+and+determines+protection+in+postconditioned+myocardium.">Delayed recovery of intracellular acidosis during reperfusion prevents calpain activation and determines protection in postconditioned myocardium.</a> Cardiovascular Research. 81 (1), 116-122 (2009).</li><li>Koyama, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+postconditioning+with+lactate-enriched+blood+on+in-hospital+outcomes+of+patients+with+ST-segment+elevation+myocardial+infarction.">Impact of postconditioning with lactate-enriched blood on in-hospital outcomes of patients with ST-segment elevation myocardial infarction.</a> International Journal of Cardiology. 220, 146-148 (2016).</li><li>Koyama, T., Munakata, M., Akima, T., Miyamoto, K., Kanki, H., Ishikawa, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle+squeezing+immediately+after+coronary+reperfusion+therapy+using+postconditioning+with+lactate-enriched+blood.">Muscle squeezing immediately after coronary reperfusion therapy using postconditioning with lactate-enriched blood.</a> International Journal of Cardiology. 275, 36-38 (2019).</li><li>Hale, S. L., Mehra, A., Leeka, J., Kloner, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postconditioning+fails+to+improve+no+reflow+or+alter+infarct+size+in+an+open-chest+rabbit+model+of+myocardial+ischemia-reperfusion.">Postconditioning fails to improve no reflow or alter infarct size in an open-chest rabbit model of myocardial ischemia-reperfusion.</a> American Journal of Physiology-Heart and Circulatory Physiology. 294 (1), H421-H425 (2008).</li><li>Hasdai, D., Holmes, D. R., Garratt, K. N., Edwards, W. 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The authors have nothing to disclose.

A detailed description of the PCLeB procedures was provided with a representative case treated using PCLeB. PCLeB consists of intermittent reperfusion and timely coronary injections of lactated Ringer&#39;s solution to achieve controlled reperfusion with tissue oxygenation and minimal lactate washout7,8,9. Not only intermittent reperfusion but also supplementary lactate, administered as lactated Ringer&#39;s solution, may increase the delay in recovery from tissue acidosis produced during ischemia; in this supposed mechanism, PCLeB may potentiate the beneficial effects of the original postconditioning protocol that consists of intermittent reperfusion only10.
Several critical steps need to be pointed out to successfully perform PCLeB. First, before starting PCLeB, incidental coronary flow recovery should be prevented as much as possible because uncontrolled reperfusion before PCLeB ruins the subsequent&#160;controlled reperfusion achieved by PCLeB in terms of tissue oxygenation with minimal lactate washout. Spontaneous reperfusion before CAG cannot help. However, during the initial wiring procedures, coronary flow is sometimes unintentionally re-started before balloon delivery to the occluded coronary artery. This is an unwanted phenomenon. To minimize the effects of this phenomenon, the balloon catheter is recommended to be placed inside the guiding catheter beforehand, close to its outlet during the wiring procedures, so that the balloon can be quickly moved forward to the culprit lesion to re-occlude the lesion site once the coronary flow is unintentionally re-started.
Second, once PCLeB is started, confirming the restoration of coronary flow during each brief reperfusion is important because the failure to restore the coronary flow during PCLeB procedures makes PCLeB meaningless. Therefore, during each brief reperfusion, contrast medium injection into the culprit coronary artery should be performed to check if the coronary flow was restored. This can be achieved by injecting &#8805;4 mL of 20&#8211;30 mL of the lactated Ringer&#39;s solution prefilled in the 30 mL syringe into the guiding catheter, which pushes out approximately 4 mL of the contrast medium prefilled within the lumen from the manifold to the tip of the guiding catheter in the default set-up. Injections of both the contrast medium and 20&#8211;30 mL of lactated Ringer&#39; solution need to be performed even during the first 10 s reperfusion; thus, the busiest time during the entire procedures of PCLeB occurs at the very start of this protocol. Even if the initial brief reperfusion took 12&#8211;13 s instead of 10 s, it might be still acceptable. Since the reason for starting brief reperfusion with 10 s duration is to achieve minimal lactate washout during the very early phase of reperfusion, the initial brief reperfusion of 12&#8211;13 s duration can still achieve this goal.
Third, the size of the balloon used for PCLeB is important. Since the balloon is left at the lesion site throughout the procedures of PCLeB, if a small-sized balloon is selected, the lumen area gained after balloon dilation may be small and coronary flow may be hampered by the deflated balloon left at the lesion site during each brief reperfusion. Therefore, the balloon should ideally have the same size as the lumen diameter of the target lesion. However, one size smaller balloon may still be acceptable. Selection of balloons of such sizes is also beneficial in imposing adequate stretch stimuli on the vessel wall before stenting because of the reason mentioned later.
Fourth, the speed of lactated Ringer&#39;s solution injection and the timing of the balloon inflation are vital. The core objective of PCLeB is to keep tissue lactate concentrations high during the early reperfusion period. To achieve this objective, a larger amount of lactated Ringer&#39;s solution should be trapped inside the ischemic myocardium in a less diluted form. To make it possible, fast and continued injection of lactated Ringer&#39;s solution until the last moment of the balloon inflation process is needed. Therefore, 20&#8211;30 mL of lactated Ringer&#39;s solution needs to be injected within several seconds and the balloon inflation should be completed a little before completion of lactated Ringer&#39;s solution injection. To trap a larger amount of lactated Ringer&#39;s solution inside the ischemic myocardium, a larger amount of the solution, instead of 20&#8211;30 mL, can be used for each injection, but care should be taken to avoid volume overload.
Some modification of the PCLeB protocol might be possible if the modification adheres to the two critical components of PCLeB, i.e., starting with a very short period of reperfusion (i.e., 10&#8211;15 s) and trapping lactated Ringer&#39;s solution inside the ischemic myocardium in each brief repetitive ischemia. Reduction of the number of intermittent reperfusions and modification of the period of brief ischemia/reperfusion may be allowed. However, whether such modifications will reduce the beneficial effects of PCLeB is unknown.
Coronary flow recovery is generally very good after reperfusion therapy with PCLeB8,9,15. No-reflow phenomenon may not be experienced with this approach; this cannot be prevented by the original postconditioning protocol in animal experiments, reportedly17. However, at the end of the reperfusion therapy with PCLeB, if good coronary flow recovery cannot be achieved (i.e., TIMI grade flow II instead of III), there might be two possible explanations. First, insufficient stretch stimuli to the culprit lesion before stenting might reduce the beneficial effects of PCLeB. Stretch stimuli to the culprit lesion by balloon dilation or stenting procedures induces endothelin release from the endothelium of the lesion site18 and causes intracellular alkalization in myocardial cells19 distal to the lesion, which is an opposite effect of PCLeB20 and may reduce the beneficial effects of PCLeB. Therefore, it is recommended that supposed endothelin storage in the culprit lesion should be released as much as possible during the PCLeB procedures when tissue acidosis is maintained. If a smaller-sized balloon relative to the vessel size was used during the PCLeB procedures, stretch stimuli to the culprit lesion may be suboptimal and the endothelin inside the culprit lesion will be spared. Subsequent larger stimuli imposed by stenting procedures may induce an intense release of spared endothelin from the lesion site, possibly causing abrupt intracellular alkalization; this may attenuate the beneficial effects of PCLeB. Second, if a fairly longer stent, relative to the balloon used for PCLeB, is implanted, the similar phenomenon will ensue even if a sufficiently large balloon is used for the PCLeB procedures because no stretch stimuli is imposed on the coronary vessel wall excessively covered by the stent. Therefore, if possible, spot stenting using a shorter stent is preferable after the PCLeB procedures.
There might be no limitation in the application of PCLeB in patients with STEMI as long as reperfusion therapy is indicated. Cardiogenic shock is not a contraindication at all but rather a good indication for PCLeB. Increase in aortic pressure can be expected during PCI using PCLeB, as shown in the representative results. Simultaneous use of IABP may reduce the beneficial effects of PCLeB because IABP may enhance the washout of lactate via a mechanically driven force and may promote recovery from tissue acidosis produced during ischemia. Therefore, simultaneous use of IABP is not recommended even in severe cases, such as in patients with cardiogenic shock. Spontaneous reperfusion before CAG may reduce or remove the beneficial effects of PCLeB. However, spontaneous reperfusion before CAG does not necessarily preclude the application of PCLeB because coronary flow may still be insufficient and low-flow ischemia may still exist in the reperfused myocardium in such cases. Therefore, PCLeB may be worth trying as long as TIMI flow grade III is not achieved before PCI. Conversely, cases with spontaneous reperfusion with TIMI flow grade III achieved before PCI may be a clear limitation of the application of PCLeB.
The protocol of PCLeB appears complicated at a glance. However, once the initial busiest part of the protocol has finished, it can be realized that the patient&#39;s condition is becoming more stabilized as the procedures proceed to the less busy, later stages of the protocol. Before finishing the entire protocol, secondary operators often start to prepare for the next procedures after PCLeB, such as intravascular ultrasonography, because they know that nothing worse will happen, thereafter. Currently, myocardial reperfusion injury is generally left untreated during reperfusion therapy for STEMI. Despite the absence of firm evidence for the beneficial effects of PCLeB, considering its safety aspect and the current lack of alternative effective approaches, PCLeB is worth trying instead of leaving myocardial reperfusion injury as it occurs. Once the effectiveness of PCLeB is generally confirmed in the future, this technique might, hopefully, be applied to other arterial occlusive disorders, such as acute limb ischemia.

The widespread use of coronary reperfusion therapy has markedly improved the survival of patients with ST-segment elevation myocardial infarction (STEMI) over the past decades1,2. On the contrary, the proportion of patients with post-myocardial-infarction (post-MI) heart failure has increased, paradoxically3,4. To reduce the incidence of post-MI heart failure, further reduction of the infarct size is of paramount importance.
Timely restoration of the coronary blood flow is crucial for salvaging myocardial cells from the ischemic cell death. However, restoration of the coronary blood flow to the ischemic myocardium induces another type of cell death, which is myocardial reperfusion injury and reduces myocardial-salvaging effects of reperfusion therapy5,6. Therefore, to further reduce the infarct size and improve the outcomes of patients with STEMI, prevention of myocardial reperfusion injury is vital. However, despite notable efforts over the decades, no approach has been proven successful in preventing this injury in the clinical setting to date. Recently, a new approach for cardioprotection in patients with STEMI, i.e., postconditioning with lactate-enriched blood (PCLeB), has been reported7,8,9. PCLeB is a modification of the original protocol of postconditioning reported by Staat et al10. In the original postconditioning protocol, the culprit coronary artery is reopened and immediate application of four brief cycles of intermittent reperfusion is performed. Despite its initial success in a small pilot study10, the original postconditioning protocol has failed to improve the outcomes of patients with STEMI in large-scale clinical trials11,12,13. In&#160;the&#160;PCLeB protocol, the original postconditioning protocol&#160;was modified to increase the delay in recovery from tissue acidosis produced during ischemia because the delay in recovery from the tissue acidosis was thought to be responsible for the cardioprotective effects of postconditioning14. For this purpose, in addition to intermittent reperfusion, timely coronary injections of the lactated Ringer&#39;s solution were incorporated into the postconditioning protocol, aimed at achieving controlled reperfusion with tissue oxygenation and minimal lactate washout. Through this approach, the speed of the recovery from tissue acidosis relative to that of tissue re-oxygenation can be substantially reduced, thus making the delay in recovery from tissue acidosis definitive. The duration of each brief reperfusion, in this modified postconditioning protocol (Figure 1), is gradually increased in a stepwise manner from 10 to 60 s. This approach prevents the abrupt and rapid washout of lactate during the very early phase of reperfusion. Each brief ischemic period lasts for 60 s. Injection of lactated Ringer&#39;s solution is performed to supply lactate directly into the culprit coronary artery at the end of each brief reperfusion. 20 mL of lactated Ringer&#39;s solution is injected for the right coronary artery and 30 mL is injected for the left coronary artery. This is done immediately before the balloon inflation. During each brief repetitive ischemic period, to trap the lactate inside the ischemic myocardium, the balloon is quickly inflated at the lesion site. Full reperfusion is performed after seven cycles of balloon inflation and deflation. Stenting is performed thereafter, and the percutaneous coronary intervention (PCI) is completed. Excellent in-hospital15 and 6 month16 outcomes in a limited number of patients with STEMI treatment using PCLeB have already been reported. This method article provides a detailed description of each step of the PCLeB procedures, which is developed for preventing myocardial reperfusion injury in patients with STEMI.

<table><tbody><tr><td>Heparin Na (5,000 U/5 mL)</td><td>Mochida Pharmaceutical Company</td><td></td><td>Heparin sodium</td></tr><tr><td>Lactec Injection (500 mL)</td><td>Otsuka Pharmaceutical Factory</td><td></td><td>Lactated Ringer&#039;s solution</td></tr><tr><td>NAMIC CONVENIENCE KIT AKK-3435</td><td>NIPRO</td><td>6069410</td><td>a manifold-plus-syringe-injector kit</td></tr><tr><td>Y Connector</td><td>GOODMAN CO.,LTD.</td><td>YOL9A</td><td>Y-connector is connceted between a guiding catheter and a manifold, and enables both pressure momitoring and introduction of a balloon catheter into a guiding catheter simultaneously.</td></tr></tbody></table>

postconditioning with lactate-enriched blood for cardioprotection in st-segment elevation myocardial infarction

The beneficial effects of reperfusion therapy for ST-segment elevation myocardial infarction (STEMI) is attenuated by reperfusion injury. No approach has been proven successful in preventing this injury in the clinical setting to date. Meanwhile, a novel approach for cardioprotection in patients with STEMI, i.e., postconditioning with lactate-enriched blood (PCLeB), has recently been reported. PCLeB is a modification of the original protocol of postconditioning, aimed at increasing the delay in the recovery from tissue acidosis produced during ischemia. This was sought to achieve controlled reperfusion with tissue oxygenation and minimal lactate washout. In this modified postconditioning protocol, the duration of each brief reperfusion is gradually increased in a stepwise manner from 10 to 60 s. Each brief ischemic period lasts for 60 s. At the end of each brief reperfusion, injection of lactated Ringer&#39;s solution (20&#8211;30 mL) is performed directly into the culprit coronary artery immediately before the balloon inflation and the balloon is quickly inflated at the lesion site, so that the lactate is trapped inside the ischemic myocardium during each brief repetitive ischemic period. After seven cycles of balloon inflation and deflation, full reperfusion is performed. Stenting is performed thereafter, and the percutaneous coronary intervention is completed. Excellent in-hospital and 6 month outcomes in a limited number of patients with STEMI treated using PCLeB have already been reported. This method article provides a detailed description of each step of the PCLeB procedures.

A representative case treated using PCLeB is shown. A 48-year-old man presented at the emergency department of the Saitama Municipal Hospital for prolonged chest pain. Electrocardiography (ECG) revealed marked ST-segment elevation in precordial leads (Figure 3), and he was then diagnosed with anterior STEMI. Blood levels of glucose, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol were 111, 179, and 36 mg/dL, respectively. Glycated hemoglobin level was 5.9%. Creatine kinase (CK) and creatine kinase-MB (CK-MB) levels were 106 and 3 IU/L, respectively. The catheterization team was mobilized, and he was transferred to the catheterization laboratory immediately. He experienced frequent ventricular fibrillation before reperfusion therapy (Figure 4). DC cardioversion was performed three times at the emergency department and three times at the catheterization laboratory before CAG was performed.
CAG revealed proximal occlusion of the left anterior descending artery with thrombolysis-in-myocardial-infarction (TIMI) flow grade I (Figure 5A). No collateral circulation was observed. At this time, the aortic systolic pressure was approximately 70 mmHg (Figure 6A). However, intra-aortic balloon counterpulsation (IABP) was withheld because it enhances lactate washout via a mechanically driven force, which is opposite to what PCLeB does.
Reperfusion using PCLeB was started 60 min after symptom onset. No ventricular tachycardia/fibrillation was observed during and after PCLeB. The aortic systolic pressure increased to approximately 75 mmHg at the end of PCLeB (Figure 6B) and to 80 mmHg after the PCI (Figure 6C). Not only the systolic pressure but also the shape of the aortic pressure curve had improved; the apparent increase in the area under the curve of the aortic pressure suggested an increase in cardiac output. TIMI flow grade III was achieved after PCLeB. Stenting was performed, thereafter. TIMI flow grade III was still observed after stenting and the PCI was completed (Figure 5B). ECG recorded upon admission to the intensive care unit revealed complete ST resolution (Figure 7). The peak plasma CK and CK-MB levels were 4830 and 172 IU/L, respectively. The higher level of CK relative to that of CK-MB might be caused by a series of DC cardioversion before reperfusion therapy. No complication or adverse event was observed throughout his admission period. Resting thallium scintigraphy before discharge revealed well-preserved myocardial viability (Figure 8).
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/59672/59672fig1.jpg" /> 
Figure 1: Overview of the protocol for postconditioning with lactate-enriched blood. The duration of each brief reperfusion is gradually increased from 10 to 60 s in a stepwise manner. Each brief ischemic period lasts for 60 s. At the end of each brief reperfusion, lactate is supplied by injecting lactated Ringer&#8217;s solution directly into the culprit coronary artery immediately before balloon inflation and the balloon is quickly inflated at the lesion site so that the lactate is trapped inside the ischemic myocardium. After seven cycles of balloon inflation and deflation, full reperfusion is performed, followed by stenting. LCA = left coronary artery; RCA = right coronary artery. This figure has been modified from reference8. <a href="https://www.jove.com/files/ftp_upload/59672/59672fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/59672/59672fig2.jpg" /> 
Figure 2: Schematic representation of the manifold system for PCLeB. One inlet line of the manifold is connected to a bottle of contrast medium and another inlet line to a 500 mL bottle of lactated Ringer&#8217;s solution. The syringe injector is to be replaced by a 30 mL syringe equipped with a lock connector before starting PCLeB. PCLeB = postconditioning with lactate-enriched blood. <a href="https://www.jove.com/files/ftp_upload/59672/59672fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/59672/59672fig3.jpg" /> 
Figure 3: Electrocardiography in the emergency department. Marked elevation of ST-segment in precordial leads was observed. <a href="https://www.jove.com/files/ftp_upload/59672/59672fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/59672/59672fig4.jpg" /> 
Figure 4: An electrocardiography recording of ventricular fibrillation observed at the catheterization laboratory <a href="https://www.jove.com/files/ftp_upload/59672/59672fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/59672/59672fig5.jpg" /> 
Figure 5: Coronary angiography view before and after reperfusion therapy with PCLeB. (A) Coronary angiography view before reperfusion therapy. Subtotal occlusion of the proximal left anterior descending artery with thrombolysis-in-myocardial-infarction flow grade I was observed. (B) Final coronary angiography view after reperfusion therapy with PCLeB. Coronary flow in the left anterior descending artery was resumed. PCLeB, postconditioning with lactate-enriched blood. <a href="https://www.jove.com/files/ftp_upload/59672/59672fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/59672/59672fig6.jpg" /> 
Figure 6: Aortic pressure before and after reperfusion with PCLeB. (A) Aortic pressure before reperfusion. The systolic aortic pressure was around 70 mmHg. (B) The aortic pressure immediately after PCLeB. The systolic aortic pressure increased to around 75 mmHg and the shape of the aortic pressure curve had improved in view of an increase in the area under the curve. (C) Aortic pressure after PCI completion. The systolic aortic pressure increased to 80 mmHg. PCI = percutaneous coronary intervention; PCLeB = postconditioning with lactate-enriched blood. <a href="https://www.jove.com/files/ftp_upload/59672/59672fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/59672/59672fig7.jpg" /> 
Figure 7: Electrocardiography at the intensive care unit immediately after reperfusion therapy <a href="https://www.jove.com/files/ftp_upload/59672/59672fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Complete ST resolution was observed.
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/59672/59672fig8.jpg" /> 
Figure 8: Resting thallium scintigraphy before discharge. A bull&#8217;s eye imaging is shown. Myocardial viability was well preserved in the left anterior descending artery area. <a href="https://www.jove.com/files/ftp_upload/59672/59672fig8large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Postconditioning with Lactate-enriched Blood for Cardioprotection in ST-segment Elevation Myocardial Infarction at JoVE.com

Postconditioning with Lactate-enriched Blood for Cardioprotection in ST-segment Elevation Myocardial Infarction

Herein, we present a protocol for postconditioning with lactate-enriched blood, which is a novel approach for cardioprotection. This protocol comprises&#160;intermittent reperfusion and timely coronary injections of lactated Ringer&#39;s solution. This can be applied to ST-segment elevation myocardial infarction patients who undergo primary percutaneous coronary intervention.

The beneficial effects of reperfusion therapy for ST-segment elevation myocardial infarction (STEMI) is attenuated by reperfusion injury. No approach has ...

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9.2K Views.  Saitama Municipal Hospital. Herein, we present a protocol for postconditioning with lactate-enriched blood, which is a novel approach for cardioprotection. This protocol comprises intermittent reperfusion and timely coronary injections of lactated Ringer's solution. This can be applied to ST-segment elevation myocardial infarction patients who undergo primary percutaneous coronary intervention.

Video: Postconditioning with Lactate-enriched Blood for Cardioprotection in ST-segment Elevation Myocardial Infarction

LAD-Ligation: A Murine Model of Myocardial Infarction

Research models of infarction and myocardial ischemia are essential to investigate the acute and chronic pathobiological and pathophysiological processes in myocardial ischemia and to develop and optimize future treatment.
Two different methods of creating myocardial ischemia are performed in laboratory rodents. The first method is to create cryo infarction, a fast but inaccurate technique, where a cryo-pen is applied on the surface of the heart (1-3). Using this method the scientist can not guarantee that the cryo-scar leads to ischemia, also a vast myocardial injury is created that shows pathophysiological side effects that are not related to myocardial infarction. The second method is the permanent ligation of the left anterior descending artery (LAD). Here the LAD is ligated with one single stitch, forming an ischemia that can be seen almost immediately. By closing the LAD, no further blood flow is permitted in that area, while the surrounding myocardial tissue is nearly not affected. This surgical procedure imitates the pathobiological and pathophysiological aspects occurring in infarction-related myocardial ischemia.
The method introduced in this video demonstrates the surgical procedure of a mouse infarction model by ligating the LAD. This model is convenient for pathobiological and pathophysiological as well as immunobiological studies on cardiac infarction. The shown technique provides high accuracy and correlates well with histological sections.

This video demonstrates how to use a fast and reliable model to study pathobiological and pathophysiological processes of myocardial ischemia.

Research models of infarction and myocardial ischemia are essential to investigate the acute and chronic pathobiological and pathophysiological processes ...

Biology

A Murine Model of Myocardial Ischemia-reperfusion Injury through Ligation of the Left Anterior Descending Artery

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms at work during MI and developing effective therapeutic approaches requires methodology to reproducibly simulate the clinical incidence and reflect the pathophysiological changes associated with MI. Here, we describe a surgical method to induce MI in mouse models that can be used for short-term ischemia-reperfusion (I/R) injury as well as permanent ligation. The major advantage of this method is to facilitate location of the left anterior descending artery (LAD) to allow for accurate ligation of this artery to induce ischemia in the left ventricle of the mouse heart. Accurate positioning of the ligature on the LAD increases reproducibility of infarct size and thus produces more reliable results. Greater precision in placement of the ligature will improve the standard surgical approaches to simulate MI in mice, thus reducing the number of experimental animals necessary for statistically relevant studies and improving our understanding of the mechanisms producing cardiac dysfunction following MI. This mouse model of MI is also useful for the preclinical testing of treatments targeting myocardial damage following MI.

We introduce a surgical method to induce experimental ischemia/reperfusion (I/R) injury to simulate myocardial infarction (MI) in mouse models that allows for more clarity in positioning of the ligation on the left anterior descending artery (LAD) to increase the reproducibility of MI experiments in mice.

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms ...

Murine Isolated Heart Model of Myocardial Stunning Associated with Cardioplegic Arrest

The following protocol is of use to evaluate impaired cardiac function or myocardial stunning following moderate ischemic insults. The technique is useful for modeling ischemic injury associated with numerous clinically relevant phenomenon including cardiac surgery with cardioplegic arrest and cardiopulmonary bypass, off-pump CABG, transplant, angina, brief ischemia, etc. The protocol presents a general method to model hypothermic hyperkalemic cardioplegic arrest and reperfusion in rodent hearts focusing on measurement of myocardial contractile function. In brief, a mouse heart is perfused in langendorff mode, instrumented with an intraventricular balloon, and baseline cardiac functional parameters are recorded. Following stabilization, the heart is then subject to brief infusion of a cardioprotective hypothermic cardioplegia solution to initiate diastolic arrest. Cardioplegia is delivered intermittently over 2 hr. The heart is then reperfused and warmed to normothermic temperatures and recovery of myocardial function is monitored. Use of this protocol results in reliable depressed cardiac contractile function free from gross myocardial tissue damage in rodents.

The goal of this protocol to assess myocardial stunning following ischemic cardioplegic arrest in rodents.

The following protocol is of use to evaluate impaired cardiac function or myocardial stunning following moderate ischemic insults. The technique is useful ...

Induction and Assessment of Ischemia-reperfusion Injury in Langendorff-perfused Rat Hearts

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for myocardial infarction and cardiac complications of thoracic surgery. The ability of certain drugs to precondition the myocardium against ischemia reperfusion injury has led to multiple clinical trials, with little success. The isolated heart model allows acute observation of the functional effects of ischemia reperfusion injury in real time, including the effects of various pharmacological interventions administered at any time-point before or within the ischemia-reperfusion injury window. Since brief periods of ischemia can precondition the heart against ischemic injury, in situ aortic cannulation is performed to allow for functional assessment of non-preconditioned myocardium. A saline filled balloon is placed into the left ventricle to allow for real-time measurement of pressure generation. Ischemic injury is simulated by the cessation of perfusion buffer flow, followed by reperfusion. The duration of both ischemia and reperfusion can be modulated to examine biochemical events at any given time-point. Although the Langendorff isolated heart model does not allow for the consideration of systemic events affecting ischemia and reperfusion, it is an excellent model for the examination of acute functional and biochemical events within the window of ischemia reperfusion injury as well as the effect of pharmacological intervention on cardiac pre- and postconditioning. The goal of this protocol is to demonstrate how to perform in situ aortic cannulation and heart excision followed by ischemia/reperfusion injury in the Langendorff model.

The isolated rat heart is an enduring model for ischemia reperfusion injury. Here, we describe the process of harvesting the beating heart from a rat via in situ aortic cannulation, Langendorff perfusion of the heart, simulated ischemia-reperfusion injury, and infarct staining to confirm the extent of ischemic insult.

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for ...

Experimental Models to Study the Neuroprotection of Acidic Postconditioning Against Cerebral Ischemia

Stroke is one of the leading causes of mortality and disability worldwide, with limited therapeutic approaches. As an endogenous strategy for neuroprotection, postconditioning treatments have proven to be promising therapies against cerebral ischemia. However, complicated procedures and potential safety issues limit their clinical application. To overcome these disadvantages, we have developed acidic postconditioning (APC) as a therapy for experimental focal cerebral ischemia. APC refers to the mild acidosis treatment by inhaling CO2 during reperfusion following ischemia. Here we present two models to execute APC in vitro and in vivo, respectively. The oxygen-glucose deprivation (OGD) treatment of mice and the corticostriatal occlusion and middle cerebral artery occlusion (MCAO) of mice were employed to mimic cerebral ischemia. APC can be simply achieved by transferring brain slices to acidic buffer bubbled with 20% CO2, or by mice inhaling 20% CO2. APC showed significant protective effects against cerebral ischemia, as reflected by tissue viability and brain infarct volume.

Acidic postconditioning protects against cerebral ischemia. Here we present two models to execute APC. They are achieved respectively by transferring corticostriatal slices to acidic buffer after oxygen-glucose deprivation in vitro and by inhaling 20% CO2 after middle cerebral artery occlusion in vivo.

Stroke is one of the leading causes of mortality and disability worldwide, with limited therapeutic approaches. As an endogenous strategy for ...

Neuroscience

Improvement of a Closed Chest Porcine Myocardial Infarction Model by Standardization of Tissue and Blood Sampling Procedures

Myocardial ischemia reperfusion (I/R) injury contributes to almost half of the necrotic area after myocardial infarction. To date there is no approved drug to prevent or reduce myocardial I/R injury. The study and understanding of the pathophysiological mechanisms of myocardial I/R injury is essential to develop successful treatments. Large animal experiments are an important step in translational methods. The porcine model of acute myocardial infarction has been established and described by ourselves and others. We aimed to further improve the value of the model by focusing in detail on the sampling techniques for use in future experiments. Furthermore, we emphasize small but important steps that can affect the quality of the final results. To mimic the clinical situation of myocardial I/R injury, a percutaneous coronary intervention (PCI) catheter was inserted into the left anterior descending coronary artery (LAD) of an anesthetized pig. &#176;&#176;&#176;This model mimics acute myocardial infarction and PCI treatment in humans with the possibility of accurately determining the area at risk as well as the necrotic- and viable ischemic tissue. Here the model was used to investigate the effect of a bicyclic peptide inhibitor of FXIIa. The model can also be modified to allow longer reperfusion times to study later effects of myocardial infarction.

Here we demonstrate a protocol to standardize sampling procedures of an established porcine model of acute myocardial infarction in order to increase its translational value in the understanding of the pathophysiology of myocardial ischemia/reperfusion injury and to test novel drug candidates.

Myocardial ischemia reperfusion (I/R) injury contributes to almost half of the necrotic area after myocardial infarction. To date there is no approved ...

Pre-clinical Model of Cardiac Donation after Circulatory Death

Cardiac transplantation demand is on the rise; nevertheless, organ availability is limited due to a paucity of suitable donors. Organ donation after circulatory death (DCD) is a solution to address this limited availability, but due to a period of prolonged warm ischemia and the risk of tissue injury, its routine use in cardiac transplantation is seldom seen. In this manuscript we provide a detailed protocol closely mimicking current clinical practices in the context of DCD with continuous monitoring of heart function, allowing for the evaluation of novel cardioprotective strategies and interventions to decrease ischemia-reperfusion injury.
In this model, the DCD protocol is initiated in anesthetized Lewis rats by stopping ventilation to induce circulatory death. When systolic blood pressure drops below 30 mmHg, the warm ischemic time is initiated. After a pre-set warm ischemic period, hearts are flushed with a normothermic cardioplegic solution, procured, and mounted onto a Langendorff ex vivo heart perfusion system. Following 10 min of initial reperfusion and stabilization, cardiac reconditioning is continuously evaluated for 60 min using intraventricular pressure monitoring. A heart injury is assessed by measuring cardiac troponin T and the infarct size is quantified by histological staining. The warm ischemic time can be modulated and tailored to develop the desired amount of structural and functional damage. This simple protocol allows for the evaluation of different cardioprotective conditioning strategies introduced at the moment of cardioplegia, initial reperfusion and/or during ex vivo perfusion. Findings obtained from this protocol can be reproduced in large models, facilitating clinical translation.

This protocol shows a simple and flexible approach for the evaluation of new conditioning agents or strategies to increase the feasibility of cardiac donation after circulatory death.

Cardiac transplantation demand is on the rise; nevertheless, organ availability is limited due to a paucity of suitable donors. Organ donation after ...

Semi-Minimal Invasive Method to Induce Myocardial Infarction in Rats and the Assessment of Cardiac Function by an Isolated Working Heart System

Myocardial infarction (MI) remains the main contributor to morbidity and mortality worldwide. Therefore, research on this topic is mandatory. An easily and highly reproducible MI induction procedure is required to obtain further insight and better understanding of the underlying pathological changes. This procedure can also be used to evaluate the effects or potency of new and promising treatments (as drugs or interventions) in acute MI, subsequent remodeling and heart failure (HF). After intubation and pre-operative preparation of the animal, an anesthetic protocol with isoflurane was performed, and the surgical procedure was conducted&#160;quickly. Using a minimally invasive approach, the left anterior descending artery (LAD) was located and occluded by a ligature. The occlusion can be performed acutely for subsequent reperfusion (ischemia/reperfusion injury). Alternatively, the vessel can be ligated permanently to investigate the development of chronic MI, remodeling or HF. Despite common pitfalls, the drop-out rates are minimal. Various treatments such as remote ischemic conditioning can be examined for their cardioprotective potential pre-, peri- and post-operatively. The post-operative recovery was quick as the anesthesia was precisely controlled and the duration of the operation was short. Post-operative analgesia was administered for three days. The minimally invasive procedure reduces the risk of infection and inflammation. Furthermore, it facilitates rapid recovery. The &#8220;working heart&#8221; measurements were performed ex vivo and enabled precise control of preload, afterload and flow. This procedure requires specific equipment and training for adequate performance. This manuscript provides a detailed step-by-step introduction for conducting these measurements.

This article presents an efficient method to perform myocardial ischemia and subsequent chronic reperfusion in rats using a minimally invasive approach. In addition, left ventricular hemodynamic function of rats is assessed by echocardiography and isolated working heart methods.&#160;

Myocardial infarction (MI) remains the main contributor to morbidity and mortality worldwide. Therefore, research on this topic is mandatory. An easily ...

Delayed Intramyocardial Delivery of Stem Cells after Ischemia Reperfusion Injury in a Murine Model

There is significant interest in the use of stem cells (SCs) for the recovery of cardiac function in individuals with myocardial injuries. Most commonly, cardiac stem cell therapy is studied by delivering SCs concurrently with the induction of myocardial injury. However, this approach presents two significant limitations: the early hostile pro-inflammatory ischemic environment may affect the survival of transplanted SCs, and it does not represent the subacute infarction scenario where SCs will likely be used. Here we describe a two-part series of surgical procedures for the induction of ischemia-reperfusion injury and delivery of mesenchymal stem cells (MSCs). This method of stem cell administration may allow for the longer viability and retention around damaged tissue by circumventing the initial immune response. A model of ischemia reperfusion injury was induced in mice accompanied by the delivery of mesenchymal stem cells (3.0 x 105), stably expressing the reporter gene firefly luciferase under the constitutively expressed CMV promoter, intramyocardially 7 days later. The animals were imaged via ultrasound and bioluminescent imaging for confirmation of injury and injection of cells, respectively. Importantly, there was no added complication rate when performing this two-procedure approach for SC delivery. This method of stem cell administration, collectively with the utilization of state-of-the-art reporter genes, may allow for the in vivo study of viability and retention of transplanted SCs in a situation of chronic ischemia commonly seen clinically, while also circumventing the initial pro-inflammatory response. In summary, we established a protocol for the delayed delivery of stem cells into the myocardium, which can be used as a potential new approach in promoting regeneration of the damaged tissue.

Stems cells are continuously investigated as potential treatments for individuals with myocardial damage, however, their decreased viability and retention within injured tissue can impact their long-term efficacy. In this manuscript we describe an alternative method for stem cell delivery in a murine model of ischemia reperfusion injury.

There is significant interest in the use of stem cells (SCs) for the recovery of cardiac function in individuals with myocardial injuries. Most commonly, ...

Left Anterior Descending Coronary Artery Ligation for Ischemia-Reperfusion Research: Model Improvement via Technical Modifications and Quality Control

Coronary heart disease is the leading cause of death globally. Complete cessation of blood flow in coronary arteries causes ST-segment elevation myocardial infarction (STEMI), resulting in cardiogenic shock and fatal arrhythmia, which are associated with high mortality. Primary coronary intervention (PCI) for recanalizing the coronary artery significantly improves the outcomes of STEMI, but advancements made in shortening the door-to-balloon time have failed to reduce in-hospital mortality, suggesting that additional therapeutic strategies are required. Left anterior descending coronary artery (LAD) ligation in rats is an animal model for acute myocardial IR research that is comparable to the clinical scenario in which rapid coronary recanalization through PCI is used for STEMI; however, PCI-induced STEMI is a technically challenging and complicated operation associated with high mortality and great variation in infarction size. We identified the ideal position for LAD ligation, created a gadget to control a snare loop, and supported a modified surgical maneuver, thereby reducing tissue damage, to establish a reliable and reproducible acute myocardial ischemia-reperfusion (IR) research protocol for rats. This is a non-survival surgery. We also propose a method for validating the quality of study results, which is a critical step for determining the accuracy of subsequent biochemical analyses.

Herein, we present a protocol focusing on the quality control of the left anterior descending coronary artery ligation by technically modifying the traditional procedure in rats for acute myocardial ischemia-reperfusion research.