The authors thank Guangdong Laboratory Animals Monitoring Institute for technical support, animal care, and sample collection. They also thank Shenzhen Mindray Bio-Medical Electronics Co., Ltd, for technical support in the ultrasonic examination. This work was supported by Guangdong Science and Technology Program, China, and Jinan University Central Universities Basic Scientific Research Business Expenses Project (2017A020215076, 2008A08003, and 21621409).

The procedures for the care and use of laboratory animals were approved by the Institutional Animal Care and Use Committee of the Guangdong Laboratory Animals Monitoring Institute. All experiments were conducted in accordance with the Guide for the Care and Use of Laboratory Animals (8th Ed., 2011, National Research Council, USA). The surgical procedure is shown in Figure 1.
1. Preoperative preparation of animals
<ol>
	<li>Randomly divide 10 3-month-old male minipigs weighing 30-35 kg into the sham group (n = 5) and VGD group (n = 5).</li>
	<li>Evaluate the preoperative and postoperative health conditions of the pigs using the body mass index (BMI). Calculate the BMI as follows: 
	BMI = body weight (kg)/(body length [cm] &#215; body length [cm]) 
	NOTE: Body length is measured from the pig&#39;s nose to the base of the tail.</li>
	<li>Fast the animals for 12 h before surgery to avoid aspiration after anesthesia. Prepare anesthesia appliances and surgical instruments, including an anesthesia machine, gas, anesthetic drugs, an anesthesia pipeline, a special laryngoscope, and surgical instruments, a rib retainer, sutures, a thyroid retractor, surgical forceps, etc. Sterilize all instruments to be used in the surgery.</li>
</ol>
2. Preparing the animals for surgery
<ol>
	<li>Weigh the animals and calculate the anesthetic dose. Administer intramuscularly the anesthetic mixture comprising&#160;2 mg/kg of 1:1 Tiletamine and Zolazepam, 0.2 mg/kg diazepam, and 0.02 mg/kg atropine17.&#160;Use fentanyl (50 mg/kg) for intraoperative pain relief30.</li>
	<li>Ensure that a proper anesthetic plane is achieved and insert an indwelling venous catheter (20G) into the marginal ear vein to establish ear access. Transfer the pig onto the operation table and place it in the supine position. Immobilize the limbs with bandages and elevate the head with a sterile drape. 
	NOTE: The state of anesthesia was monitored by central fixation of the eyeball, miosis, loss of pupillary reflex, and loss of pain reflex.&#160; The heart rate and blood pressure were maintained at a lower level than the baseline. The surgeon should monitor HR, BP, and other parameters under paralysis and increase the anesthetic dose if HR increases &#62; 20% above baseline.</li>
	<li>Expose the epiglottis and glottis using a veterinary laryngoscope. Perform tracheal intubation with a 7.0-7.5Fr tube and connect it to the anesthesia breathing circuit. 
	NOTE: The ventilator is used for continuous positive pressure ventilation with tidal volume 280 mL, inspiratory/expiratory ratio 1:2, respiratory rate 20 times/min, and positive end-expiratory pressure (5 cm H2O).</li>
	<li>Intravenously inject vecuronium bromide (0.1 mg/kg) to relax the muscles during the surgical procedures and use 2% isoflurane to maintain anesthesia at a respiratory rate of 16-20 bpm and a tidal volume of 10 mL/kg. 
	NOTE: Vecuronium is given to ensure adequate depth of anesthesia in paralyzed animals, especially since the dose of induction drug and isoflurane is on the lower end of recommended.</li>
	<li>Use vet ointment on the pig&#39;s eyes to prevent dryness while under anesthesia. Use electric blankets to maintain the pig&#39;s body temperature at 38 &#176;C &#177; 5 &#176;C.</li>
	<li>Use an electrocardiogram to monitor heart rate, blood oxygen levels, and body temperature.</li>
</ol>
3. Surgical procedures
<ol>
	<li>Shave the left chest wall and apply three alternating rounds of 0.7% iodine and 75% alcohol to aseptically prepare the surgical area up to the left mandibular angle, down to the umbilical cord, left to the posterior axillary line, and right to the axillary front. Place a sterile surgical drape around the surgical area.</li>
	<li>Make a 7-10 cm transverse incision with an electric knife in the third left intercostal space and separate the subcutaneous tissues layer by layer (Figure 2A). Remove a 5-6 cm segment of the third rib with bone scissors and expose the internal mammary vein by a retractor after exposing the third rib-sternal joint (Figure 2B).</li>
	<li>Locate the internal mammary vein along with the left internal mammary artery on the left side of the sternum. Perform a blunt dissection of the internal mammary vein with vascular forceps.</li>
	<li>Perform hemostasis by electrocoagulation of the branches of the left internal mammary vein with an electric knife. If hemostasis is incomplete, use cotton thread ligation for hemostasis. Ligate and mark the two ends of the vein while it is being harvested (Figure 2C).</li>
	<li>Prepare heparin normal saline by adding 2 mL of heparin sodium solution and 98 mL of normal saline. After removing the vein, inject heparin normal saline into the vein for pretreatment (Figure 2D). Then, put the vein into normal saline and keep it for backup.</li>
	<li>Make a similar incision as described above and remove the internal mammary vein in the sham group. Open the pericardium, then close the chest wall in the sham group. Use the internal mammary vein of the sham group for pathological control without coronary artery bypass grafting.</li>
	<li>Make a ~7 cm incision with an electric knife on the pericardium to expose the right coronary artery trunk. Suspend the pericardium and sew on the skin on the ipsilateral side with the 1-0 surgical sutures (Figure 2E). Separate the right coronary artery trunk from the surrounding tissues (Figure 2E).</li>
	<li>Bypass the blocking band under the proximal end of the isolated right coronary artery near the aorta with a wire hook and treat the myocardium with three cycles of 2 min ischemia and 5 min reperfusion by tightening and relaxing the blocking band (Figure 2F). Monitor the heart&#39;s electrical activity with the electrocardiogram monitor during the ischemia/reperfusion preconditioning (Figure 2G). 
	NOTE: When the right coronary artery is blocked, the electrocardiogram shows an increased heart rate and ST-segment elevation.</li>
	<li>Tighten the band to block the right coronary blood flow. Cut the epicardium covering the blood vessels. Expose the coronary artery wall, and cut longitudinally with the tip of a surgical blade against the center of the anterior wall of the blood vessels.</li>
	<li>After cutting the lumen, enlarge the incision with scissors and place a coronary shunt. Insert one end of the shunt with a coil into the distal coronary artery through the tear. Shunt the blood in the coronary arteries into the hollow coronary shunt to ensure a clear operative field (Figure 2H).</li>
	<li>Perform an end-to-side continuous suture between the internal mammary vein and the right coronary trunk with the 7-0 polypropylene suture (Figure 2I). In the middle of the ascending aorta, occlude the left anterolateral wall of the ascending aorta with a semi-occlusion clamp.</li>
	<li>Use a surgical blade to make a small incision in the aortic wall where the adventitia has been cut, insert the head end of the sliding shaft at the head end of the punch into the aortic cavity through this incision, contract the sliding shaft outward, and the circular knife above it cuts off a piece of the arterial wall. The tissue block cut out by the punch is about 3 mm in diameter (Figure 2J).</li>
	<li>Pull out the shunt. Perform an end-to-side continuous suture between the internal mammary vein and the the aortic wall with the 6-0 polypropylene suture (Figure 2K). Open the semi-occlusion clamp.</li>
	<li>Record the bypass flow of the right coronary artery trunk proximal to the anastomosis site using ultrasound. Monitor the heart&#39;s electrical activity using the electrocardiogram (Figure 2L).</li>
	<li>Indwell a temporary drainage tube (Fr: 16) into the chest cavity to allow the blood and fluids to drain. Sew the pericardium incision using a 1-0 cotton thread and close the chest layer by layer (from the inside to the outside: Pleura layer, muscle layer, subcutaneous tissue layer, skin layer) while placing penicillin (about 0.5 g) powder onto each layer. Remove the drainage tube after sewing the skin incision using a 1-0 cotton thread.</li>
</ol>
4. Post-surgery care
<ol>
	<li>Remove the endotracheal tube after the animals returned to spontaneous breathing. The anesthesiologist should assess the animal&#8217;s vitals (e.g., respiratory rate, heart rate, oxygen saturation, etc.) and remove the ECG after the animals wake up and return to spontaneous activity. Send the animals back to the feeding room and place the sham animals in another pen in the breeding room. Keep the animals warm with an electric blanket. Observe the animals every hour after surgery (at least 4 times).</li>
	<li>Feed the animal the day after surgery. Add aspirin (200 mg) 2x daily for 7 days to the animal feed to prevent post-operative thrombosis and reduce wound pain. 
	NOTE: Avoid feeding animals on the day of surgery to prevent aspiration.&#160;</li>
	<li>Administer the animal with an intramuscular injection of penicillin 1x daily for 7 consecutive days to prevent post-operative infection (14,000 units per kg).</li>
</ol>
5. Ultrasonic examination
<ol>
	<li>After CABG surgery, use a sterile ultrasonic probe sleeve to wrap the high-frequency linear array probe. Place the probe on the surface of the venous graft.</li>
	<li>Display the outline of the graft in the two-dimensional ultrasound mode, then shift to the color Doppler mode to detect blood flow in the graft.</li>
</ol>
6. Venous graft tissue collection
<ol>
	<li>Collect 10 mL of blood sample from the venous circuit of the ear vein for biochemical testing. (Table 1). Centrifuge the blood sample at 1,000 x g for 5 min and perform biochemical tests with an automatic biochemical analyzer.</li>
	<li>Anesthetize the animal as described earlier. Upon confirmation of anesthesia depth, inject 10% potassium chloride 0.5mL/kg body weight from the ear marginal vein or forelimb vein. Then, make a 10 cm median sternal incision with an electric knife to harvest the vein graft 30 days after surgery. Fix the body position as in step 2.2., and after sterilization and placement of a drape, make a median sternum incision to split the sternum. During the separation, avoid the main blood vessels and the heart, and separate the grafted blood vessels layer by layer.</li>
	<li>Quickly cut off the large blood vessels connecting to the heart, place the heart and ascending aorta on ice chips, and remove the graft vascular bridge, the connected aorta, and the right coronary artery. Rinse all the samples with normal saline at 4 &#176;C.</li>
	<li>Take the entire graft vessel of about 3-4 cm in size, divide it into 4-5 equal parts, and transfer to cryopreservation tubes. Quickly put the tubes into liquid nitrogen to flash freeze and move to a &#8722;80 &#176;C ultra-low temperature freezer for storage.</li>
	<li>For analysis, rinse the graft with ice-cold 0.9% saline and fix it in 4% paraformaldehyde solution. Maintain a ratio of tissue block size to fixative solution of 1:10 and fix the tissue for more than 12 h.</li>
	<li>Stain the sections in 50 mL of aqueous solution of hematoxylin for 3 min. Separate the sections by washing with 50 mL of 0.5% hydrochloric acid ethanol and 50 mL of 0.2% ammonia water for 10 s each.</li>
	<li>Rinse with running water for 1 h and then clean in distilled water by soaking for 3 min. Dehydrate in 70% and 90% ethanol for 10 min each. Place in 50 mL of 0.5% alcohol eosin staining solution for 2-3 min.</li>
	<li>Dehydrate the stained sections with pure ethanol for 10 min and then soak in pure xylene for 10 min to make the samples transparent. Drip the transparent sections with neutral glue and cover with a coverslip. Observe pathological sections under a light microscope at 40x magnification.</li>
</ol>

<ol>
	<li>Lloyd-Jones, D., 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=Executive+summary:+Heart+disease+and+stroke+statistics--2010+update:+A+report+from+the+American+Heart+Association.">Executive summary: Heart disease and stroke statistics--2010 update: A report from the American Heart Association.</a> Circulation. 121 (7), 948-954 (2010).</li><li>Taggart, D. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contemporary+coronary+artery+bypass+grafting.">Contemporary coronary artery bypass grafting.</a> Frontiers of Medicine. 8, 395-398 (2014).</li><li>Wolny, R., Mintz, G. S., Pregowski, J., Witkowski, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms,+prevention+and+treatment+of+saphenous+vein+graft+disease.">Mechanisms, prevention and treatment of saphenous vein graft disease.</a> The American Journal of Cardiology. 154, 41-47 (2021).</li><li>Schachner, T., Laufer, G., Bonatti, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+(animal)+models+of+vein+graft+disease.">In vivo (animal) models of vein graft disease.</a> European Journal of Cardio-thoracic Surgery: Official Journal of the European Association for Cardio-thoracic Surgery. 30 (3), 451-463 (2006).</li><li>Suggs, W. D., 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=Antisense+oligonucleotides+to+c-fos+and+c-jun+inhibit+intimal+thickening+in+a+rat+vein+graft+model.">Antisense oligonucleotides to c-fos and c-jun inhibit intimal thickening in a rat vein graft model.</a> Surgery. 126 (2), 443-449 (1999).</li><li>Jiang, Z., 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=A+novel+vein+graft+model:+Adaptation+to+differential+flow+environments.">A novel vein graft model: Adaptation to differential flow environments.</a> American Journal of Physiology. Heart and Circulatory Physiology. 286 (1), H240-H245 (2004).</li><li>O'Brien, J. E., 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=Early+injury+to+the+media+after+saphenous+vein+grafting.">Early injury to the media after saphenous vein grafting.</a> The Annals of Thoracic Surgery. 65 (5), 1273-1278 (1998).</li><li>Zou, Y., 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=Mouse+model+of+venous+bypass+graft+arteriosclerosis.">Mouse model of venous bypass graft arteriosclerosis.</a> The American Journal of Pathology. 153 (4), 1301-1310 (1998).</li><li>Klyachkin, M. L., 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=Postoperative+reduction+of+high+serum+cholesterol+concentrations+and+experimental+vein+bypass+grafts.+Effect+on+the+development+of+intimal+hyperplasia+and+abnormal+vasomotor+function.">Postoperative reduction of high serum cholesterol concentrations and experimental vein bypass grafts. Effect on the development of intimal hyperplasia and abnormal vasomotor function.</a> The Journal of Thoracic and Cardiovascular Surgery. 108 (3), 556-566 (1994).</li><li>Tan, W., 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=A+porcine+model+of+heart+failure+with+preserved+ejection+fraction+induced+by+chronic+pressure+overload+characterized+by+cardiac+fibrosis+and+remodeling.">A porcine model of heart failure with preserved ejection fraction induced by chronic pressure overload characterized by cardiac fibrosis and remodeling.</a> Frontiers in Cardiovascular Medicine. 8, 677727 (2021).</li><li>Hocum Stone, ., L, L., 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=Magnetic+resonance+imaging+assessment+of+cardiac+function+in+a+swine+model+of+hibernating+myocardium+3+months+following+bypass+surgery.">Magnetic resonance imaging assessment of cardiac function in a swine model of hibernating myocardium 3 months following bypass surgery.</a> The Journal of Thoracic and Cardiovascular Surgery. 153 (3), 582-590 (2017).</li><li>Gedik, N., 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=Proteomics/phosphoproteomics+of+left+ventricular+biopsies+from+patients+with+surgical+coronary+revascularization+and+pigs+with+coronary+occlusion/reperfusion:+Remote+ischemic+preconditioning.">Proteomics/phosphoproteomics of left ventricular biopsies from patients with surgical coronary revascularization and pigs with coronary occlusion/reperfusion: Remote ischemic preconditioning.</a> Scientific Reports. 7 (1), 7629 (2017).</li><li>Tsirikos Karapanos, ., N, , 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=The+impact+of+competitive+flow+on+distal+coronary+flow+and+on+graft+flow+during+coronary+artery+bypass+surgery.">The impact of competitive flow on distal coronary flow and on graft flow during coronary artery bypass surgery.</a> Interactive Cardiovascular and Thoracic Surgery. 12 (6), 993-997 (2011).</li><li>Meng, X., 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=Competitive+flow+arising+from+varying+degrees+of+coronary+artery+stenosis+affects+the+blood+flow+and+the+production+of+nitric+oxide+and+endothelin+in+the+internal+mammary+artery+graft.">Competitive flow arising from varying degrees of coronary artery stenosis affects the blood flow and the production of nitric oxide and endothelin in the internal mammary artery graft.</a> European Journal of Cardio-thoracic Surgery: Official Journal of the. 43 (5), 1022-1027 (2013).</li><li>Thankam, F. G., 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=Association+of+hypoxia+and+mitochondrial+damage+associated+molecular+patterns+in+the+pathogenesis+of+vein+graft+failure:+A+pilot+study.">Association of hypoxia and mitochondrial damage associated molecular patterns in the pathogenesis of vein graft failure: A pilot study.</a> Translational Research: The Journal of Laboratory and Clinical. , 38-52 (2021).</li><li>Li, X., 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=A+surgical+model+of+heart+failure+with+preserved+ejection+fraction+in+Tibetan+minipigs.">A surgical model of heart failure with preserved ejection fraction in Tibetan minipigs.</a> Journal of Visualized Experiments: JoVE. 180 (180), 63526 (2022).</li><li>Rueda, A. l. c. a. l. &. #. 2. 2. 5. ;., I, , 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=A+live+porcine+model+for+surgical+training+in+tracheostomy,+neck+dissection,+and+total+laryngectomy.">A live porcine model for surgical training in tracheostomy, neck dissection, and total laryngectomy.</a> European Archives of Oto-Rhino-Laryngology: Official Journal of the European Federation of Oto-Rhino-Laryngological Societies (EUFOS): Affiliated with the German Society for Oto-Rhino-Laryngology - Head and Neck Surgery. 278 (8), 3081-3090 (2021).</li><li>Proudfit, W. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prognostic+value+of+coronary+arteriography.">Prognostic value of coronary arteriography.</a> Cardiovascular Clinics. 12 (2), 1-8 (1981).</li><li>Clark, 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=Regulation+of+fibroplasia+in+cutaneous+wound+repair.">Regulation of fibroplasia in cutaneous wound repair.</a> TheAmerican Journal of the Medical Sciences. 306 (1), 42-48 (1993).</li><li>Darby, I., Skalli, O., Gabbiani, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alpha-smooth+muscle+actin+is+transiently+expressed+by+myofibroblasts+during+experimental+wound+healing.">Alpha-smooth muscle actin is transiently expressed by myofibroblasts during experimental wound healing.</a> Laboratory Investigation. 63 (1), 21-29 (1990).</li><li>Sterpetti, A. V., 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=Formation+of+myointimal+hyperplasia+and+cytokine+production+in+experimental+vein+grafts.">Formation of myointimal hyperplasia and cytokine production in experimental vein grafts.</a> Surgery. 123 (4), 461-469 (1998).</li><li>Shannon, A. H., 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=Porcine+model+of+infrarenal+abdominal+aortic+aneurysm.">Porcine model of infrarenal abdominal aortic aneurysm.</a> Journal of Visualized Experiments: JoVE. 153 (153), (2019).</li><li>Langille, B. L., O'Donnell, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reductions+in+arterial+diameter+produced+by+chronic+decreases+in+blood+flow+are+endothelium-dependent.">Reductions in arterial diameter produced by chronic decreases in blood flow are endothelium-dependent.</a> Science. 231 (4736), 405-407 (1986).</li><li>Zwolak, R. M., Adams, M. C., Clowes, A. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinetics+of+vein+graft+hyperplasia:+Association+with+tangential+stress.">Kinetics of vein graft hyperplasia: Association with tangential stress.</a> Journal of Vascular Surgery. 5 (1), 126-136 (1987).</li><li>Kotani, K., 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=A+subacute+hypoxic+model+using+a+pig.">A subacute hypoxic model using a pig.</a> Surgery Today. 35 (11), 951-954 (2005).</li><li>Liu, D., 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=Comparison+of+ketamine-pentobarbital+anesthesia+and+fentanyl-pentobarbital+anesthesia+for+open-heart+surgery+in+minipigs.">Comparison of ketamine-pentobarbital anesthesia and fentanyl-pentobarbital anesthesia for open-heart surgery in minipigs.</a> Lab Animal. 38 (7), 234-240 (2009).</li><li>Geovanini, G. R., Pinna, F. R., Prado, F. A., Tamaki, W. T., Marques, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Standardization+of+anesthesia+in+swine+for+experimental+cardiovascular+surgeries.">Standardization of anesthesia in swine for experimental cardiovascular surgeries.</a> Revista Brasileira de Anestesiologia. 58 (4), 363-370 (2008).</li><li>Alhomary, M., Ramadan, E., Curran, E., Walsh, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Videolaryngoscopy+vs.+fibreoptic+bronchoscopy+for+awake+tracheal+intubation:+A+systematic+review+and+meta-analysis.">Videolaryngoscopy vs. fibreoptic bronchoscopy for awake tracheal intubation: A systematic review and meta-analysis.</a> Anaesthesia. 73 (9), 1151-1161 (2018).</li><li>Parang, P., Arora, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coronary+vein+graft+disease:+Pathogenesis+and+prevention.">Coronary vein graft disease: Pathogenesis and prevention.</a> Canadian Journal of Cardiology. 25 (2), e57-e62 (2009).</li><li>Egan, T. D., 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=Fentanyl+pharmacokinetics+in+hemorrhagic+shock:+a+porcine+model.">Fentanyl pharmacokinetics in hemorrhagic shock: a porcine model.</a> Anesthesiology. 91 (1), 156-166 (1999).</li></ol>

The authors have no conflicts of interest to disclose.

In this study, we described in detail the protocol for animal selection, instrument preparation, surgical procedures, and post-operative evaluation when developing a CABG-induced VGD model. We performed ultrasonic examination of the venous graft before and after CABG surgery and histological examination of the graft 30 days after the surgery. The blood flow in the internal mammary vein was normal before the CABG surgery, while retrograde flow was observed in the graft of the internal mammary vein. Compared with the sham ...

Although coronary heart disease mortality has declined significantly in recent years, half of middle-aged adults in the United States develop ischemic heart-related symptoms each year, and one-third of older adults die from coronary heart disease1. Coronary artery bypass grafting (CABG) is an effective surgical modality to improve myocardial ischemia, and more importantly, it is an irreplaceable surgical modality for the treatment of multivessel coronary artery disease2. Over time, however, vascular grafts develop inflammation, intimal hyperplasia, and progressive atherosclerosis, which is known to lead to vein graft failure or vein graft disease (VGD)3. In patients after CABG, if restenosis occurs, only the diseased blood vessel can be replaced in some cases2. Older patients and added comorbidities make redoing coronary artery bypass grafting quite challenging. Delaying or controlling the pathological problems associated with grafted blood vessels is an urgent problem to be solved. Large animal models of CABG-VGD are needed for the investigation of disease mechanisms and the development of therapeutic strategies. Researchers have successfully established animal VGD models in small and large animals such as mice4, rats5, rabbits6, and pigs7. Compared with small animals, large animals such as pigs have anatomical structures and physiological characteristics similar to humans and have longer lifespans8,9. Thus, large animals are more suitable for exploring long-term pathological changes in venous graft disease and for preclinical testing of drugs or devices. We and our collaborating team have successfully applied surgical techniques to establish a porcine heart failure model and described the cardiac pathological changes in this model10.
CABG surgery has been standardized in clinical practice, but when it is applied to the establishment of VGD animal models, the differences between species, the acquisition of animal equipment and facilities, animal surgical operations, and animal feeding and nursing are huge challenges for researchers. As in clinical practice, the approaches for CABG surgery used to establish VGD animal models include midline sternotomy11 and left lateral thoracotomy12. Midline sternotomy is more commonly used13,14. However, this approach has high risks for both humans and animals. In the study reported by Thankam et al., two of the six pigs used for modeling died during surgery15. High model mortality increases study costs and affects the accuracy of results. A study showed earlier that a left chest wall incision was feasible to establish CABG-induced VGD in pigs11. Here, this study aims to describe a step-by-step protocol to establish a reproducible surgery for a CABG-induced VGD model in minipigs and to evaluate the phenotype of this model. The experimental protocol was jointly designed by the cardiac surgery and anesthesia teams. The surgical approach for the left third intercostal space was determined according to the cadavers of other minipigs in the laboratory before surgery, and the anesthesia method was performed according to the method used at the center16. Blood biochemical tests, ultrasonic examination, and histology examination were conducted to evaluate animal models.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Aortic Punch</td>
			<td>Medtronic Inc. , America</td>
			<td>3.0mm, 3.5mm, 4.0mm</td>
			<td>Used for proximal coronary bridge anastomosis</td>
		</tr>
		<tr>
			<td>Automatic biochemical analyzer</td>
			<td>IDEXX Laboratories, Inc. America</td>
			<td>Catalyst One</td>
			<td></td>
		</tr>
		<tr>
			<td>Cardiac coronary artery bypass grafting instrument kit</td>
			<td>LANDANGER, France</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Cardiogram monitor</td>
			<td>Shenzhen Mindray Bio-Medical Electronics Co, Ltd</td>
			<td>MEC-1000</td>
			<td></td>
		</tr>
		<tr>
			<td>Coronary Shunt</td>
			<td>AXIUS&#160;</td>
			<td>OF-1500, OF-2500, OF-3000</td>
			<td>The product temporarily blocks the coronary artery during arteriotomy to reduce the amount of bleeding in the surgical field and provide blood flow to the distal end during anastomosis.&#160; The Axius shunt plug is not an implant and should be removed prior to completion of the anastomosis.&#160;&#160;</td>
		</tr>
		<tr>
			<td>Defibrillator</td>
			<td>MEDIANA</td>
			<td>Mediana D500</td>
			<td></td>
		</tr>
		<tr>
			<td>Diazepam</td>
			<td>Nanguo pharmaceutical Co. LTD, Guangdong, China</td>
			<td>H37023039&#160;</td>
			<td>Narcotic inducer</td>
		</tr>
		<tr>
			<td>Disposable manual electric knife</td>
			<td>Covidien, America</td>
			<td>E2516H</td>
			<td></td>
		</tr>
		<tr>
			<td>Electric negative pressure suction machine</td>
			<td>Shanghai Baojia Medical Instrument Co, Ltd</td>
			<td>YX932D</td>
			<td></td>
		</tr>
		<tr>
			<td>Esmolol</td>
			<td>Guangzhou Wanzheng Pharmaceutical Co. LTD</td>
			<td>H20055990</td>
			<td>Emergency drugs</td>
		</tr>
		<tr>
			<td>Ice machine&#160;</td>
			<td>Local suppliers, Guangzhou, China</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Lidocaine&#160;</td>
			<td>Chengdu First Pharmaceutical Co. LTD</td>
			<td>H51021662</td>
			<td>Emergency drugs</td>
		</tr>
		<tr>
			<td>Luxtec headlight system</td>
			<td>Luxtec, America</td>
			<td>AX-1375-BIF</td>
			<td>Used for lighting fine parts during operation</td>
		</tr>
		<tr>
			<td>Medical operation magnifier (glasses)</td>
			<td>Germany Lista co, LTD</td>
			<td>SuperVu Galilean 3.5&#215;</td>
			<td>Used for fine site operation during operation</td>
		</tr>
		<tr>
			<td>Multi-function high-frequency electrotome</td>
			<td>Shanghai Hutong Electronics Co, Ltd</td>
			<td>GD350-B</td>
			<td></td>
		</tr>
		<tr>
			<td>Nitrogen canister</td>
			<td>Local suppliers, Guangzhou, China</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Nonabsorbable surgical suture (polypropylene suture)</td>
			<td>Johnson &#38; Johnson, America</td>
			<td>6-0, 7-0</td>
			<td>Used to suture blood vessels.</td>
		</tr>
		<tr>
			<td>Nonabsorbable suture (cotton thread)</td>
			<td>Covidien, America</td>
			<td>1-0</td>
			<td>Used for skin and muscle tissue tugging</td>
		</tr>
		<tr>
			<td>Open heart surgery instrument kit</td>
			<td>Shanghai Medical Instrument (Group) Co., LTD</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Propofol injection</td>
			<td>Xi &#39;an Libang Pharmaceutical Co. LTD</td>
			<td>H19990282</td>
			<td>Anesthetic sedative</td>
		</tr>
		<tr>
			<td>Refrigerator</td>
			<td>Local suppliers, Guangzhou, China</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Respiratory anesthesia machine for animal</td>
			<td>Shenzhen Reward Life Technology Co, Ltd, China</td>
			<td>R620-S1</td>
			<td></td>
		</tr>
		<tr>
			<td>Semi-occlusion clamp</td>
			<td>Xinhua Surgical Instrument Co., Ltd.</td>
			<td>ZL1701RB</td>
			<td>Temporarily cut off the aortic flow</td>
		</tr>
		<tr>
			<td>vecuronium bromide</td>
			<td>Richter, Hungary&#160;</td>
			<td>JX20090127</td>
			<td>Muscle relaxant</td>
		</tr>
		<tr>
			<td>Veterinary ultrasound system&#160;</td>
			<td>Royal Philips, Netherlands</td>
			<td>CX50</td>
			<td></td>
		</tr>
		<tr>
			<td>Zoletil</td>
			<td>Virbac, France</td>
			<td>Zoletil 50&#160;</td>
			<td>Animal narcotic</td>
		</tr>
	</tbody>
</table>

establishment and evaluation of a porcine vein graft disease model

Venous graft disease (VGD) is the leading cause of coronary artery bypass graft (CABG) failure. Large animal models of CABG-VGD are needed for the investigation of disease mechanisms and the development of therapeutic strategies. 
To perform the surgery, we enter the cardiac chamber through the third intercostal space and carefully dissect the internal mammary vein and immerse it in normal saline. The right main coronary artery is then treated for ischemia. The target vessel is incised, a shunt plug is placed, and the distal end of the graft vein is anastomosed. The ascending aorta is partially blocked, and the proximal end of the graft vein is anastomosed after perforation. The graft vein is checked for patency, and the proximal right coronary artery is ligated. 
CABG surgery is performed in minipigs to harvest the left internal mammary vein for its use as a vascular graft. Serum biochemical tests are used to evaluate the physiological status of the animals after surgery. Ultrasound examination shows that the proximal, middle, and distal end of the graft vessel are unobstructed. In the surgical model, turbulent blood flow in the graft is observed upon histological examination after the CABG surgery, and venous graft stenosis associated with intimal hyperplasia is observed in the graft. The study here provides detailed surgical procedures for the establishment of a repeatable CABG-induced VGD model.

BMI and serum biochemical indices 
The BMI between the sham and VGD groups was not significantly different (sham vs. VGD, 22.05 kg/cm2 &#177; 0.46 kg/cm2 vs. 21.14 kg/cm2 &#177; 0.39 kg/cm2, p = 0.46). The serum biochemical results&#160;are listed in Table 1. Statistically significant changes between the groups were found in four biochemical indexes, including aspartate aminotransferase (AST, sham vs. VGD, 25.25 IU/L &#177; 1.88 I...

Watch this Scientific Journal Video about Establishment and Evaluation of a Porcine Vein Graft Disease Model at JoVE.com

Establishment and Evaluation of a Porcine Vein Graft Disease Model

In this protocol, novel pig vein bypass grafting was performed through a small incision in the left chest wall without cardiopulmonary bypass. A postoperative pathology study was done, which showed intimal thickening.

Venous graft disease (VGD) is the leading cause of coronary artery bypass graft (CABG) failure. Large animal models of CABG-VGD are needed for the ...

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1.6K Views.  the First Affiliated Hospital of Jinan University. In this protocol, novel pig vein bypass grafting was performed through a small incision in the left chest wall without cardiopulmonary bypass. A postoperative pathology study was done, which showed intimal thickening.

Video: Establishment and Evaluation of a Porcine Vein Graft Disease Model

Evaluation of a Novel Laser-assisted Coronary Anastomotic Connector - the Trinity Clip - in a Porcine Off-pump Bypass Model

To simplify and facilitate beating heart (i.e., off-pump), minimally invasive coronary artery bypass surgery, a new coronary anastomotic connector, the Trinity Clip, is developed based on the excimer laser-assisted nonocclusive anastomosis technique. The Trinity Clip connector enables simplified, sutureless, and nonocclusive connection of the graft to the coronary artery, and an excimer laser catheter laser-punches the opening of the anastomosis. Consequently, owing to the complete nonocclusive anastomosis construction, coronary conditioning (i.e., occluding or shunting) is not necessary, in contrast to the conventional anastomotic technique, hence simplifying the off-pump bypass procedure. Prior to clinical application in coronary artery bypass grafting, the safety and quality of this novel connector will be evaluated in a long-term experimental porcine off-pump coronary artery bypass (OPCAB) study. In this paper, we describe how to evaluate the coronary anastomosis in the porcine OPCAB model using various techniques to assess its quality. Representative results are summarized and visually demonstrated.

This paper describes a novel nonocclusive coronary anastomotic connector in a porcine off-pump coronary artery bypass (OPCAB) model. This easy-to-use coronary connector has intrinsic potential to facilitate minimally invasive OPCAB surgery.

To simplify and facilitate beating heart (i.e., off-pump), minimally invasive coronary artery bypass surgery, a new coronary anastomotic connector, the ...

A Multicenter MRI Protocol for the Evaluation and Quantification of Deep Vein Thrombosis

We evaluated a magnetic resonance venography (MRV) approach with gadofosveset to quantify total thrombus volume changes as the principal criterion for treatment efficacy&#160;in a multicenter randomized study comparing edoxaban monotherapy with a heparin/warfarin regimen for acute, symptomatic lower extremities deep vein thrombosis (DVT) treatment. We also used a direct thrombus imaging approach (DTHI, without the use of a contrast agent) to quantify fresh thrombus. We then sought to evaluate the reproducibility of the analysis methodology and applicability of using 3D magnetic resonance venography and direct thrombus imaging for the quantification of DVT in a multicenter trial setting. From 10 randomly selected subjects participating in the edoxaban Thrombus Reduction Imaging Study (eTRIS), total thrombus volume in the entire lower extremity deep venous system was quantified bilaterally. Subjects were imaged using 3D-T1W gradient echo sequences before (direct thrombus imaging, DTHI) and 5 min after injection of 0.03 mmol/kg of gadofosveset trisodium (magnetic resonance venography, MRV). The margins of the DVT on corresponding axial, curved multi-planar reformatted images were manually delineated by two observers to obtain volumetric measurements of the venous thrombi. MRV was used to compute total DVT volume, whereas DTHI was used to compute volume of fresh thrombus. Intra-class correlation (ICC) and Bland Altman analysis were performed to compare inter and intra-observer variability of the analysis. The ICC for inter and intra-observer variability was excellent (0.99 and 0.98, p &#60;0.001, respectively) with no bias on Bland-Altman analysis for MRV images. For DTHI images, the results were slightly lower (ICC = 0.88 and 0.95 respectively, p &#60;0.001), with bias for inter-observer results on Bland-Altman plots. This study showed feasibility of thrombus volume estimation in DVT using MRV with gadofosveset trisodium, with good intra- and inter-observer reproducibility in a multicenter setting.

The goal of this study is to use magnetic resonance venography with long-circulating gadolinium-based contrast agent and direct thrombus imaging for quantitative evaluation of DVT volume in a multicenter, clinical trial setting. Inter- and intra-observer variability assessments were conducted, and reproducibility of the protocol was determined.

We evaluated a magnetic resonance venography (MRV) approach with gadofosveset to quantify total thrombus volume changes as the principal criterion for ...

Establishment and Characterization of UTI and CAUTI in a Mouse Model

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. Females are disproportionately afflicted by UTI: 50% of all women will have a UTI in their lifetime. Additionally, 20-40% of these women who have an initial UTI will suffer a recurrence with some suffering frequent recurrences with serious deterioration in the quality of life, pain and discomfort, disruption of daily activities, increased healthcare costs, and few treatment options other than long-term antibiotic prophylaxis. Uropathogenic Escherichia coli (UPEC) is the primary causative agent of community acquired UTI. Catheter-associated UTI (CAUTI) is the most common hospital acquired infection accounting for a million occurrences in the US annually and dramatic healthcare costs. While UPEC is also the primary cause of CAUTI, other causative agents are of increased significance including Enterococcus faecalis. Here we utilize two well-established mouse models that recapitulate many of the clinical characteristics of these human diseases. For UTI, a C3H/HeN model recapitulates many of the features of UPEC virulence observed in humans including host responses, IBC formation and filamentation. For CAUTI, a model using C57BL/6 mice, which retain catheter bladder implants, has been shown to be susceptible to E. faecalis bladder infection. These representative models are being used to gain striking new insights into the pathogenesis of UTI disease, which is leading to the development of novel therapeutics and management or prevention strategies.

The ability to model urinary tract infections (UTI) is crucial in order to be able to understand bacterial pathogenesis and spawn the development of novel therapeutics. This work&#8217;s goal is to demonstrate mouse models of experimental UTI and catheter associated UTI that recapitulate and predict findings seen in humans.

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. ...

Vein Interposition Model: A Suitable Model to Study Bypass Graft Patency

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. Research models for bypass graft failure are essential for a better understanding of pathobiological and pathophysiological processes during graft patency loss. Large animal models, such as pigs or sheep, resemble human anatomical structures but require special facilities and equipment. This video describes a rat vein interposition model to investigate vein graft patency loss. Rats are inexpensive and easy to handle. Compared to mouse models, the convenient size of rats permits better operability and enables a sufficient amount of material to be obtained for further diverse analysis. In brief, the inferior epigastric vein of a donor rat is harvested and used to replace a segment of the femoral artery. Anastomosis is conducted via single stitches and sealed with fibrin glue. Graft patency can be monitored non-invasively using duplex sonography. Myointimal hyperplasia, which is the main cause for graft patency loss, develops progressively over time and can be calculated from histological cross sections.

This video demonstrates a model to study the development of myointimal hyperplasia after venous interposition surgery in rats.

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. ...

Improved Method for the Establishment of an In Vitro Blood-Brain Barrier Model Based on Porcine Brain Endothelial Cells

The aim of this protocol presents an optimized procedure for the purification and cultivation of pBECs and to establish in vitro blood-brain barrier (BBB) models based on pBECs in mono-culture (MC), MC with astrocyte-conditioned medium (ACM), and non-contact co-culture (NCC) with astrocytes of porcine or rat origin. pBECs were isolated and cultured from fragments of capillaries from the brain cortices of domestic pigs 5-6 months old. These fragments were purified by careful removal of meninges, isolation and homogenization of grey matter, filtration, enzymatic digestion, and centrifugation. To further eliminate contaminating cells, the capillary fragments were cultured with puromycin-containing medium. When 60-95% confluent, pBECs growing from the capillary fragments were passaged to permeable membrane filter inserts and established in the models. To increase barrier tightness and BBB characteristic phenotype of pBECs, the cells were treated with the following differentiation factors: membrane permeant 8-CPT-cAMP (here abbreviated cAMP), hydrocortisone, and a phosphodiesterase inhibitor, RO-20-1724 (RO). The procedure was carried out over a period of 9-11 days, and when establishing the NCC model, the astrocytes were cultured 2-8 weeks in advance. Adherence to the described procedures in the protocol has allowed the establishment of endothelial layers with highly restricted paracellular permeability, with the NCC model showing an average transendothelial electrical resistance (TEER) of 1249 &#177; 80 &#937; cm2, and paracellular permeability (Papp) for Lucifer Yellow of 0.90 10-6 &#177; 0.13 10-6 cm sec-1 (mean &#177; SEM, n=55). Further evaluation of this pBEC phenotype showed good expression of the tight junctional proteins claudin 5, ZO-1, occludin and adherens junction protein p120 catenin. The model presented can be used for a range of studies of the BBB in health and disease and, with the highly restrictive paracellular permeability, this model is suitable for studies of transport and intracellular trafficking.

Improved Method for the Establishment of an In Vitro Blood-Brain Barrier Model Based on Porcine Brain Endothelial Cells

The aim of the protocol is to present an optimized procedure for the establishment of an in vitro blood-brain barrier (BBB) model based on primary porcine brain endothelial cells (pBECs). The model shows high reproducibility, high tightness, and is suitable for studies of transport and intracellular trafficking in drug discovery.

The aim of this protocol presents an optimized procedure for the purification and cultivation of pBECs and to establish in vitro blood-brain barrier (BBB) ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Establishment of a Mouse Severe Acute Pancreatitis Model using Retrograde Injection of Sodium Taurocholate into the Biliopancreatic Duct

The prevalence of acute pancreatitis (AP), especially severe acute pancreatitis (SAP), is increasing in younger age groups annually. However, there is a lack of effective treatments in the current clinical practice. With the easy accessibility of transgenic and knockout strains and their small size, which allows minimal doses of drugs required for in vivo evaluation, a well-established experimental model in mice is preferred for AP research. Moreover, SAP induced through sodium taurocholate (TC) is currently one of the most widely used and best characterized models. This model has been investigated for novel therapies and possible molecular events during the process of AP. Here, we present the generation of an AP mouse model using sodium taurocholate and a simple homemade microsyringe. Moreover, we also provide the methodology for the subsequent histology and serological testing.

A mouse model of severe acute pancreatitis is described herein. The procedure presented here is very rapid, simple, and accessible, thereby potentially allowing the study of the molecular mechanisms and different therapeutic interventions in acute pancreatitis in a convenient way.

The prevalence of acute pancreatitis (AP), especially severe acute pancreatitis (SAP), is increasing in younger age groups annually. However, there is a ...

Evaluation of a Point-of-Care Testing Analyzer for Measuring Peripheral Blood Leukocytes

White blood cell (WBC) is an important indicator of inflammation in the body, and it can help distinguish between bacterial and viral infections. At present, most primary medical institutions in China have a poor percentage of adoption of blood-testing technology, and a hematology detection system with a high price to performance ratio and easy operation is urgently needed in primary healthcare centers. This paper introduces the principle and operation procedures of a point-of-care testing (POCT) card-based leukocyte analyzer (evaluated system), which was used to detect WBC indexes such as neutrophils, lymphocytes, and intermediate group cells (including eosinophils, basophils, and monocytes) in whole blood. The results from the evaluated system were compared to those from two commercial automatic hematology analyzers (reference system). The correlation and consistency between the evaluated system and the commercial reference systems were analyzed. The results showed that WBC count and number of granulocytes detected by the evaluated and reference systems showed a strong positive correlation (rs = 0.972 and 0.973, respectively), while the number of lymphocytes showed a relatively low correlation (rs = 0.851). A Bland-Altman plot showed that the major difference between the values detected by the evaluated system and the reference systems is within 95% limits of agreement (LoA), indicating that the two systems are in good agreement. In conclusion, the evaluated system has an excellent correlation, robust consistency, and a reliable comparison with the results of the widely used automatic hematology analyzers. It is ideal for WBC detection in primary medical institutions where a full-automatic five-category hematology analyzer is unavailable, especially during the COVID-19 normalized prevention and control period.

A protocol to measure peripheral blood leukocytes using a POCT card-based leukocyte analyzer is presented here. Same blood samples were tested by two automated hematology analyzers to evaluate the consistency and accuracy of the results. The results showed that the evaluated analyzer had a good correlation with the reference system.

White blood cell (WBC) is an important indicator of inflammation in the body, and it can help distinguish between bacterial and viral infections. At ...

Repetitive Blood Sampling from the Subclavian Vein of Conscious Rat

Rats are widely used in pharmacokinetics (PK) and toxicokinetic (TK) studies that need to collect a certain amount of blood at specific time points to detect drug exposure. The rat blood sampling method determines the quality of the plasma and further affects the precision of the test results. The subclavian vein blood collection method described in this protocol collects blood samples repeatedly in the conscious state of animals to meet the needs of PK and TK tests. The skills of restraint handling and appropriate procedure of needle incision ensure the success rate of blood collection. It is easy to operate while ensuring the quality of plasma and at the same time catering to animal welfare. However, this method requires skilled operation, and an improper one may cause animal weakness, pain, lameness, and even mortality. The current method has been used in the testing facility for a 4-week oral toxicity study in Sprague Dawley (SD) rats with TK. The maximum amount of blood collected within 24 h did not exceed 20% of the animal's total blood. The animals' body weight was more than 200 g for males and females. The data showed that the animals' bodyweight increased steadily every week, and the clinical observation was normal after repetitive sample collection.

The present protocol describes a simple and efficient method for collecting blood from the subclavian vein in rats. It enables rapid, timely, and easily identifiable sampling without anesthesia and obtains high-quality blood through repetitive sample collection.

Rats are widely used in pharmacokinetics (PK) and toxicokinetic (TK) studies that need to collect a certain amount of blood at specific time points to ...

Modified Arteriovenous Fistula Surgery to Establish the RV Volume Overload Mouse Model

Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model

Right ventricular (RV) volume overload (VO) is common in children with congenital heart disease. In view of distinct developmental stages,the RV myocardium may respond differently to VO in children compared to adults. The present study aims to establish a postnatal RV VO model in mice using a modified abdominal arteriovenous fistula. To confirm the creation of VO and the following morphological and hemodynamic changes of the RV, abdominal ultrasound, echocardiography, and histochemical staining were performed for 3 months. As a result, the procedure in postnatal mice showed an acceptable survival and fistula success rate. In VO mice, the RV cavity was enlarged with a thickened free wall, and the stroke volume was increased by about 30%-40% within 2 months after surgery. Thereafter, the RV systolic pressure increased, corresponding pulmonary valve regurgitation was observed, and small pulmonary artery remodeling appeared. In conclusion, modified arteriovenous fistula (AVF) surgery is feasible to establish the RV VO model in postnatal mice. Considering the probability of fistula closure and elevated pulmonary artery resistance, abdominal ultrasound and echocardiography must be performed to confirm the model status before application.

Author Spotlight: Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model  

This protocol presents the establishment and confirmation of a postnatal right ventricular volume overload (VO) model in mice with abdominal arteriovenous fistula (AVF), which can be applied to investigate how VO contributes to postnatal heart development.