I would like to thank Professor Long Tian (Department of Urology, Beijing Chaoyang Hospital) for the technical instruction on vasoepididymostomy. This modified procedure I designed is inspired by his artery-sparing microsurgical vasoepididymostomy. I would also like to thank Dr. Moqi Lv (Medical School, Xi&#39;an Jiaotong University) for the help with polishing the present paper.

This study was approved by the Ethics Committee of Northwest Women&#39;s and Children&#39;s Hospital (No. 2021002).
1. Preparations
<ol>
	<li>Place the patient supine on the operating table and perform anesthesia as per the anesthetist&#39;s recommendations.</li>
	<li>Insert a 16Fr Foley catheter around the incision site after the field is disinfected and covered with surgical drapes.</li>
</ol>
2. Modified vessel-sparing LIVE
<ol>
	<li>Create a 3 cm vertical incision in the middle of the scrotum and deliver the testis from the incision.</li>
	<li>Open the tunica vaginalis. Carefully dissociate the vas deferens and adjacent vessels. The dissociated length is ~5 cm (Figure 1).</li>
	<li>Place an operating microscope and ready the micro instruments: a micro needle holder, micro scissors, and micro forceps. Execute the following steps using the operating microscope.</li>
	<li>Create a buttonhole of a diameter of 5 mm at the tunica of cauda epididymis using micro scissors. Identify a dilated epididymal tubule and carefully dissociate it using micro forceps. After that, keep it for anastomosis.</li>
	<li>Place two needles of double-armed 11-0 microsutures longitudinally in the selected epididymal tubule, and open the epididymal tubule longitudinally between the two needles using a 15&#176; ophthalmic knife. Ensure that the opening length does not exceed the length between the entry point and the exit point of the suture on the epididymal tubule (Figure 2).</li>
	<li>Sample some epididymal fluid flowing from the incision in the tubule with a glass slide and hand it to an examiner to check for sperm. Gently pull out two needles in the epididymal tubule separately if sperm are found.</li>
	<li>Make a puncture in the gap between the vas deferens and adjacent vessels using deferens separating forceps. The puncturing site is approximately 1 cm away from the junction of the cauda epididymis and the vas deferens. Completely transect the vas deferens above the deferens separating forceps with a knife, and preserve the vasal vessels below the deferens separating forceps (Figure 3).</li>
	<li>Cannulate the 24-G angiocatheter sheath into the vas deferens lumen on the distal testicular side, followed by a bolus injection of diluted methylene blue. Observe the color of the urine. Blue staining of the urine indicates distal patency.</li>
	<li>Dissociate the vas deferens on the proximal testicular side using micro scissors to separate it from the vasal vessels. Ensure that the dissociation is executed along the vas deferens side to avoid damage to the vasal vessels. The dissociated length is ~2-3 cm. Then, remove the dissociated vas deferens and ligate the broken end of the vas deferens. During the dissociation, stop the bleeding using bipolar electrocoagulation or by ligating some small branch vessels using a 5-0 silk thread. This operation is called retrograde dissociation (Figure 4 and Figure 5).</li>
	<li>Carefully separate the vas deferens on the distal testicular side from the vasal vessels using deferens separating forceps combined with micro scissors. The dissociated length is ~0.5 cm. Separate the vas deferens and the vasal vessels without damaging the vas deferens or the vasal vessels. This operation is called anterograde dissociation (Figure 4 and Figure 6).</li>
	<li>Fix the vas deferens to be anastomosed with the tunica vaginalis using 8-0 microsutures at the included angle between the vas deferens and adjacent vessels. After fixation, ensure that the lumen of the vas deferens reaches the anastomosis site with no significant tension. This is regarded as the first-stage tension reduction.</li>
	<li>Fix the muscularis edge of the vas deferens and epididymis tunica using 3-4 interrupted 8-0 microsutures. This is regarded as the second-stage tension reduction.</li>
	<li>Pass the four needles of 11-0 microsutures in the epididymal tubule through four points (Figure 7) on the cutting surface of the vas deferens in an inside-out fashion. Then, perform the intussusception of the epididymal tubule into the vas deferens8.</li>
	<li>Suture the muscularis edge of the vas deferens and the epididymal tunica using 10-12 interrupted sutures of 8-0 microsutures.</li>
	<li>Fix the broken end of the vas deferens on the tunica vaginalis using 3-4 interrupted sutures of 8-0 microsutures (Figure 4 and Figure 8). Ensure that the free vasal vessels are fixed and cannot move. This operation is called fixation of the vasal vessels.</li>
	<li>Check whether there is tension in the vas deferens and whether there is tension and bleeding in the preserved vasal vessels. Then, suture each layer of tissue sequentially and terminate the operation.</li>
</ol>
3. Postoperative management
<ol>
	<li>Avoid sexual intercourse for 1 month after surgery.</li>
	<li>Wear a scrotal supporter or tight underpants for 1 week after surgery, and avoid standing or walking for a long time.</li>
	<li>Take antibiotics for 1 week to prevent infection.</li>
</ol>

<ol>
	<li>Wagenknecht, L. V., Klosterhalfen, H., Schirren, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microsurgery+in+andrologic+urology.+I.Refertilization.">Microsurgery in andrologic urology. I.Refertilization.</a> Journal of Microsurgery. 1 (5), 370-376 (1980).</li><li>Thomas, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vasoepididymostomy.">Vasoepididymostomy.</a> Urologic Clinics of North America. 14 (3), 527-538 (1987).</li><li>Berger, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Triangulation+end-to-side+vasoepididymostomy.">Triangulation end-to-side vasoepididymostomy.</a> The Journal of Urology. 159 (6), 1951-1953 (1998).</li><li>Marmar, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modified+vasoepididymostomy+with+simultaneous+double+needle+placement,+tubulotomy+and+tubular+invagination.">Modified vasoepididymostomy with simultaneous double needle placement, tubulotomy and tubular invagination.</a> The Journal of Urology. 163 (2), 483-486 (2000).</li><li>Chan, P. T., Li, P. S., Goldstein, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microsurgical+vasoepididymostomy:+A+prospective+randomized+study+of+3+intussusception+techniques+in+rats.">Microsurgical vasoepididymostomy: A prospective randomized study of 3 intussusception techniques in rats.</a> The Journal of Urology. 169 (5), 1924-1929 (2003).</li><li>Chan, P. T., Graham, S. D., Keane, T. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vasoepididymostomy.">Vasoepididymostomy.</a> Glenn's Urologic Surgery, 7th ed. , 379-386 (2009).</li><li>Chan, P. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+evolution+and+refinement+of+vasoepididymostomy+techniques.">The evolution and refinement of vasoepididymostomy techniques.</a> Asian Journal of Andrology. 15 (1), 49-55 (2013).</li><li>Mostafa, T., Labib, I., El-Khayat, Y., El-Rahman El-Shahat, A., Gadallah, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+testicular+arterial+supply:+Gross+anatomy,+corrosion+cast,+and+radiologic+study.">Human testicular arterial supply: Gross anatomy, corrosion cast, and radiologic study.</a> Fertility and Sterility. 90 (6), 2226-2230 (2008).</li><li>Zhang, Y., Wu, X., Yang, X. J., Zhang, H., Zhang, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vasal+vessels+preserving+microsurgical+vasoepididymostomy+in+cases+of+previous+varicocelectomy:+A+case+report+and+literature+review.">Vasal vessels preserving microsurgical vasoepididymostomy in cases of previous varicocelectomy: A case report and literature review.</a> Asian Journal of Andrology. 18 (1), 154-156 (2016).</li><li>Lyu, K. 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=A+novel+experience+of+deferential+vessel-sparing+microsurgical+vasoepididymostomy.">A novel experience of deferential vessel-sparing microsurgical vasoepididymostomy.</a> Asian Journal of Andrology. 20 (6), 576-580 (2018).</li><li>Li, P., 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=Vasal+vessel-sparing+microsurgical+single-armed+vasoepididymostomy+to+epididymal+obstructive+azoospermia:+A+retrospective+control+study.">Vasal vessel-sparing microsurgical single-armed vasoepididymostomy to epididymal obstructive azoospermia: A retrospective control study.</a> Andrologia. 53 (8), 14133 (2021).</li></ol>

The authors have nothing to disclose.

Vessel-sparing microsurgical vasoepididymostomy might have some clinical significance. Although it is not confirmed that this approach can improve the patency rate, it might better simulate the normal physiological structure and has certain significance for patients who have undergone varicocelectomy. This can be attributed to the fact that preservation of the deferential artery and vas deferens vein have positive effects on the blood supply to the testis after varicocelectomy and the venous return of the testis, respect...

Epididymal obstruction is the most common cause of obstructive azoospermia. Microsurgical vasoepididymostomy (MVE) is the main surgical treatment for epididymal obstruction. Although various MVE techniques1,2,3,4 have been described previously, a two-needle longitudinal intussusception vasoepididymostomy (LIVE) technique described by Chan et al.5,6 has been recognized as the gold standard for achieving a superior patency rate7. The vasal vessels are typically ligated in LIVE. Ligation of the vasal vessels simplifies the procedure and reduces the tension of anastomosis, and a tension-free anastomosis is critical to achieving a successful outcome.
The vas artery primarily supplies the epididymis and vas deferens8. Whether preservation of the vas artery improves the postoperative patency rate remains unclear. However, preserving the vasal vessels during MVE might better simulate the normal physiological structure. Also, for patients who have undergone varicocelectomy, it may be meaningful to perform the vessel-sparing MVE, because the integrity of vasal vasculature plays a vital role in post-varicocelectomy blood supply and the venous return of the testis. A few studies9,10,11 have attempted to preserve the vasal vessels during MVE and suggested that the procedure may have certain benefits. However, preserving the vasal vessels makes the operation of anastomosis more difficult and increases the tension of anastomosis. In addition, the tension of the vas deferens may be transmitted to the vasal vessels and cause postoperative bleeding of the little branches of the vessels. Besides, the existing vessel-sparing procedure may affect the blood supply of the vas deferens. Therefore, we improved the surgical technique of vessel-sparing MVE to make it safer. We called this new procedure microsurgical vasoepididymostomy by preserving the vasal vessels using a retrograde-anterograde dissociation.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>11-0 microsutures</td>
			<td>Ningbo Medical Needle Co.,Ltd</td>
			<td>211115</td>
			<td>Double-armed microsurgical nylon suture length: 5 cm</td>
		</tr>
		<tr>
			<td>15&#176; ophthalmic knife</td>
			<td>pearsalls limited</td>
			<td>72-1501</td>
			<td>&#160;open the epididymal tubule</td>
		</tr>
		<tr>
			<td>2-0 silk braided non-absorbable suture</td>
			<td>Coated</td>
			<td>1604-51</td>
			<td>Ligation of the vas deferens</td>
		</tr>
		<tr>
			<td>24-Gangiocatheter sheath</td>
			<td>Melsungen AG</td>
			<td>4253523-03</td>
			<td>injection</td>
		</tr>
		<tr>
			<td>5-0 silk braided non-absorbable suture</td>
			<td>Johnson &#38; Johnson</td>
			<td>SA82G</td>
			<td>Ligation of blood vessels</td>
		</tr>
		<tr>
			<td>8-0 microsutures</td>
			<td>Johnson &#38; Johnson</td>
			<td>W2908</td>
			<td>Single-armed microsurgical nylon suture length: 13 cm</td>
		</tr>
		<tr>
			<td>Deferens separating forceps</td>
			<td>Shanghai Medical Instrument Co., Ltd</td>
			<td>JCZ210</td>
			<td>Separation of vas deferens</td>
		</tr>
		<tr>
			<td>Micro scissors</td>
			<td>Shanghai Medical Instrument Co., Ltd</td>
			<td>WA1040</td>
			<td>Microsurgical operation</td>
		</tr>
		<tr>
			<td>Microforceps</td>
			<td>Shanghai Medical Instrument Co., Ltd</td>
			<td>WA3090</td>
			<td>Microsurgical operation</td>
		</tr>
		<tr>
			<td>Microneedle holder</td>
			<td>Shanghai Medical Instrument Co., Ltd</td>
			<td>WA2040</td>
			<td>Microsurgical operation</td>
		</tr>
		<tr>
			<td>Operating microscope</td>
			<td>Leica Microsystems(Sch weiz) AG</td>
			<td>M525MS3</td>
			<td>Microsurgical operation</td>
		</tr>
	</tbody>
</table>

a modified vessel-sparing microsurgical vasoepididymostomy

Microsurgical vasoepididymostomy (MVE) is the main surgical treatment for epididymal obstruction. The vasal vessels are ligated during MVE. However, preserving the vasal vessels during MVE might better simulate the normal physiological structure and be meaningful for patients who have undergone varicocelectomy. Nevertheless, preserving the vasal vessels might elevate the risk of increasing the tension of anastomosis, affecting the patency rate and leading to delayed postoperative bleeding. Therefore, we developed a novel vessel-sparing MVE to make it safer. Here is a summary of the improvements to the procedure. 1) The retrograde dissociation of the vasal vessels on the proximal testicular side was adopted as the main method, and the anterograde dissociation of the vasal vessels on the distal testicular side was adopted as a supplement to dissociate the vasal vessels to be preserved. This improvement ensures the blood supply to the vas deferens that will be used for anastomosis and also provides longer vasal vessels, which reduces the tension of anastomosis. 2) By fixing the vas deferens to be anastomosed and the broken end of the vas deferens, the free vasal vessels get fixed, which resolves the problem of transmission of vas tension to the vasal vessels and reduces the risk of vasal vessel hemorrhage. 3) Dissociation of the vas deferens after opening the tunica vaginalis increases the mobilization of the vas deferens, which also makes the new procedure easier to complete. The evaluation of the outcomes of this new procedure showed that no significant postoperative complications occurred in the patients, and the patency rate was no different from that of the conventional procedure. Therefore, this new, improved procedure can be considered safe, with satisfactory postoperative results.

A total of 51 patients who underwent vasoepididymostomy at our center between February 2018 and November 2020 were retrospectively analyzed. Considering the anastomotic tension, the modified procedure was performed only in patients with obstruction at the corpus or caudal epididymal, and these patients were included in the current study. Semen examination was performed 1.5 months after the surgery, and the patients were followed up for more than 1 year. Relevant data of patients with the results of at least one semen tes...

Watch this Scientific Journal Video about A Modified Vessel-Sparing Microsurgical Vasoepididymostomy at JoVE.com

A Modified Vessel-Sparing Microsurgical Vasoepididymostomy

Here, we present a protocol to preserve the vasal vessels in microsurgical vasoepididymostomy. The surgical security is enhanced by preserving the vasal vessels using a retrograde-anterograde dissociation and fixing the vasal vessels.

Microsurgical vasoepididymostomy (MVE) is the main surgical treatment for epididymal obstruction. The vasal vessels are ligated during MVE. However, ...

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1.8K Views.  Northwest Women’s and Children’s Hospital. Here, we present a protocol to preserve the vasal vessels in microsurgical vasoepididymostomy. The surgical security is enhanced by preserving the vasal vessels using a retrograde-anterograde dissociation and fixing the vasal vessels.

Video: A Modified Vessel-Sparing Microsurgical Vasoepididymostomy

Microsurgical Venous Pouch Arterial-Bifurcation Aneurysms in the Rabbit Model: Technical Aspects

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials have shown disappointing long-term results with unacceptable high rates of aneurysm recanalization and delayed aneurysm rupture.2 To overcome these problems, animal experimental studies are crucial for the development of better endovascular devices.3-5
Several animal models in rats, rabbits, canines and swine are available.6-8 Comparisons of the different animal models showed the superiority of the rabbit model with regard to hemodynamics and comparability of the coagulation system and cost-effectiveness.9-11 
The venous pouch arterial bifurcation model in rabbits is formed by a venous pouch sutured into an artificially created true bifurcation of both common carotid arteries (CCA). The main advantage of this model are true bifurcational hemodynamics.12 The major drawbacks are the sofar high microsurgical technical demands and high morbidity and mortality rates of up to 50%.13 These limitations have resulted in less frequent use of this aneurysm model in the recent years. These shortcomings could be overcome with improved surgical procedures and modified peri- and postoperative analgetic management and anticoagulation.14-16 Our techniques reported in this paper demonstrate this optimized technique for microsurgical creation of arterial bifurcation aneurysms.

An optimized technique for the microsurgical creation of arterial bifurcation aneurysms mimicking bifurcation human cerebral aneurysms is described. A venous pouch is sutured into an artificially created true bifurcation of both common carotid arteries. Facilitated microsurgical techniques and aggressive postoperative anticoagulation and analgesia lead to minimized morbidity rates and high aneurysm patency rates.

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials ...

Training a Sophisticated Microsurgical Technique: Interposition of External Jugular Vein Graft in the Common Carotid Artery in Rats

Neointimal hyperplasia is one the primary causes of stenosis in arterialized veins that are of great importance in arterial coronary bypass surgery, in peripheral arterial bypass surgery as well as in arteriovenous fistulas.1-5 The experimental procedure of vein graft interposition in the common carotid artery by using the cuff-technique has been applied in several research projects to examine the aetiology of neointimal hyperplasia and therapeutic options to address it. 6-8 The cuff prevents vessel anastomotic remodeling and induces turbulence within the graft and thereby the development of neointimal hyperplasia.
Using the superior caval vein graft is an established small-animal model for venous arterialization experiment.9-11 This current protocol refers to an established jugular vein graft interposition technique first described by Zou et al., 9 as well as others.12-14 Nevertheless, these cited small animal protocols are complicated.
To simplify the procedure and to minimize the number of experimental animals needed, a detailed operation protocol by video training is presented. This video should help the novice surgeon to learn both the cuff-technique and the vein graft interposition. Hereby, the right external jugular vein was grafted in cuff-technique in the common carotid artery of 21 female Sprague Dawley rats categorized in three equal groups that were sacrificed on day 21, 42 and 84, respectively. Notably, no donor animals were needed, because auto-transplantations were performed. The survival rate was 100 % at the time point of sacrifice. In addition, the graft patency rate was 60 % for the first 10 operated animals and 82 % for the remaining 11 animals. The blood flow at the time of sacrifice was 8&plusmn;3 ml/min. In conclusion, this surgical protocol considerably simplifies, optimizes and standardizes this complicated procedure. It gives novice surgeons easy, step-by-step instruction, explaining possible pitfalls, thereby helping them to gain expertise fast and avoid useless sacrifice of experimental animals.

Neointimal hyperplasia is the primary cause of stenosis in arterialized veins. We propose a new protocol whereby the right external jugular vein is grafted using the cuff technique in the common carotid artery of Sprague Dawley rats. The survival rate was 100 % at the time point of sacrifice.

Neointimal hyperplasia is one the primary causes of stenosis in arterialized veins that are of great importance in arterial coronary bypass surgery, in ...

Murine Cervical Heart Transplantation Model Using a Modified Cuff Technique

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are available to perform mechanistic in vivo studies. While heart transplantation models using a suture technique were first successfully developed in rats, the translation into an equally widespread used murine equivalent was never achieved due the technical complexity of the microsurgical procedure. In contrast, non-suture cuff techniques, also developed initially in rats, were successfully adapted for use in mice1-3. This technique for revascularization involves two major steps I) everting the recipient vessel over a polyethylene cuff; II) pulling the donor vessel over the formerly everted recipient vessel and holding it in place with a circumferential tie. This ensures a continuity of the endothelial layer, short operating time and very high patency rates4.
Using this technique for vascular anastomosis we performed more than 1,000 cervical heart transplants with an overall success rate of 95%. For arterial inflow the common carotid artery and the proximal aortic arch were anastomosed resulting in a retrograde perfusion of the transplanted heart. For venous drainage the pulmonary artery of the graft was anastomosed with the external jugular vein of the recipient5.
Herein, we provide additional details of this technique to supplement the video.

The murine cervical heart transplantation model is well suited for immunological as well as ischemia reperfusion injury studies. We modified the procedure using a non-suture cuff technique and performed more than 1,000 successful transplants with this approach.
Herein, we provide additional details of this technique to supplement the video.

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are ...

The Helsinki Rat Microsurgical Sidewall Aneurysm Model

Experimental saccular aneurysm models are necessary for testing novel surgical and endovascular treatment options and devices before they are introduced into clinical practice. Furthermore, experimental models are needed to elucidate the complex aneurysm biology leading to rupture of saccular aneurysms.
Several different kinds of experimental models for saccular aneurysms have been established in different species. Many of them, however, require special skills, expensive equipment, or special environments, which limits their widespread use. A simple, robust, and inexpensive experimental model is needed as a standardized tool that can be used in a standardized manner in various institutions.
The microsurgical rat abdominal aortic sidewall aneurysm model combines the possibility to study both novel endovascular treatment strategies and the molecular basis of aneurysm biology in a standardized and inexpensive manner. Standardized grafts by means of shape, size, and geometry are harvested from a donor rat&#39;s descending thoracic aorta and then transplanted to a syngenic recipient rat. The aneurysms are sutured end-to-side with continuous or interrupted 9-0 nylon sutures to the infrarenal abdominal aorta.
We present step-by-step procedural instructions, information on necessary equipment, and discuss important anatomical and surgical details for successful microsurgical creation of an abdominal aortic sidewall aneurysm in the rat.

Microsurgical sidewall aneurysms in rats are created by end-to-side anastomosis of an aortic graft to the abdominal aorta. We present step-by-step instructions and discuss anatomical and surgical details for successful experimental saccular aneurysm creation.

Experimental saccular aneurysm models are necessary for testing novel surgical and endovascular treatment options and devices before they are introduced ...

A Modified Precipitation Method to Isolate Urinary Exosomes

Identification of biomarkers that allow early detection of kidney diseases in urine and plasma has been an area of active interest for several years. Urinary exosome vesicles, 40-100 nm in size, are released into the urine under normal conditions by cells from all nephron segments and may contain protein, mRNA and microRNA representative of their cell type of origin. Under conditions of renal dysfunction or injury, exosomes may contain altered proportions of these components, which may serve as biomarkers for disease. There are currently several methods available for isolation of urinary exosomes, and we have previously conducted an experimental comparison of each of these approaches, including three based on ultracentrifugation, one using a nanomembrane ultrafiltration concentrator, one using a commercial precipitation reagent and one using a modification of the precipitation technique using ExoQuick reagent that we developed in our laboratory. We found the modified precipitation method produced the highest yield of exosome particles, miRNA, and mRNA, making this approach suitable for the isolation of exosomes for subsequent RNA profiling. We conclude that the modified exosome precipitation method offers a quick, scalable, and effective alternative for the isolation of exosomes from urine. In this report, we describe our modified precipitation technique using ExoQuick reagent for isolating exosomes from human urine.

This manuscript describes a protocol for isolating exosomes from human urine using a modified precipitation technique.

Identification of biomarkers that allow early detection of kidney diseases in urine and plasma has been an area of active interest for several years. ...

A Modified Method for Heterotopic Mouse Heart Transplantion

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents available. A difficulty is presented due to the small size of the animal and the considerable technical challenges of the microsurgery involved in heart transplantation. In particular, a high rate of technical failure early after transplantation may result from recipient death and post-operative complications such as hind limb paralysis or a non-beating heart. Here, the complete technique for heterotopic mouse heart transplantation is demonstrated, involving harvesting the donor heart and its subsequent implantation into a recipient mouse. The donor heart is harvested immediately following in situ perfusion with cold heparinized saline and transection of the ascending aorta and pulmonary artery. The recipient operation involves preparation of the abdominal aorta and inferior vena cava (IVC), followed by end-to-side anastomosis of the donor aorta with the recipient aorta using a single running 10-0 microsuture and a similar anastomosis of the donor pulmonary artery with the recipient IVC. Following the operation the animal is injected with 0.6 ml normal saline subcutaneously and allowed to recover on a 37&#160;&#176;C heating pad. The results from 227 mouse heart transplants are summarized with a success rate at 48 hr of 86.8%. Of the 13.2% failures within 48 hr, 5 (2.2%) experienced hind limb paralysis, 10 (4.4%) had a non-beating heart due to graft ischemic injury and/or thrombosis, while 15 (6.6%) died within 48 hr.

A technique is demonstrated for the microsurgical procedure for heterotopic transplantation of hearts in mice, including simplified methods for donor harvesting and recipient vessel anastomosis.

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents ...

Human Skeletal Muscle Biopsy Procedures Using the Modified Bergstr&#246;m Technique

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly effective. This video describes the percutaneous biopsy technique using a modified Bergstr&#246;m needle to obtain skeletal muscle tissue samples from the vastus lateralis of human subjects. The Bergstr&#246;m needle consists of an outer cannula with a small opening (&#8216;window&#8217;) at the side of the tip and an inner trocar with a cutting blade at the distal end. Under local anesthesia and aseptic conditions, the needle is advanced into the skeletal muscle through an incision in the skin, subcutaneous tissue, and fascia. Next, suction is applied to the inner trocar, the outer trocar is pulled back, skeletal muscle tissue is drawn into the window of the outer cannula by the suction, and the inner trocar is rapidly closed, thus cutting or clipping the skeletal muscle tissue sample. The needle is rotated 90&#176;&#160;and another cut is made. This process may be repeated three more times. This multiple cutting technique typically produces a sample of 100-200 mg or more in healthy subjects and can be done immediately before, during, and after a bout of exercise or other intervention. Following post-biopsy dressing of the incision site, subjects typically resume their activities of daily living right away and can fully participate in vigorous physical activity within 48-72 hr. Subjects should avoid heavy resistance exercise for 48 hr to reduce the risk of herniation of the muscle through the incision in the fascia.

Human Skeletal Muscle Biopsy Procedures Using the Modified Bergstr&amp;#246;m Technique

The purpose of this video is to present the modified Bergstr&#246;m skeletal muscle biopsy technique on human subjects.

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly ...

A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression must be pursued. Employing vascularized bone marrow transplantation and co-transplantation of the thymus have shown promise in this regard in various animal models.1-11 Vascularized bone marrow transplantation allows for the uninterrupted transfer of donor bone marrow cells within the preserved donor microenvironment, and the incorporation of thymus tissue with vascularized bone marrow transplantation has shown to increase T-cell chimerism ultimately playing a supportive role in the induction of immune regulation. The combination of solid organ and vascularized composite allotransplantation can uniquely combine these strategies in the form of a novel transplant model. Murine models serve as an excellent paradigm to explore the mechanisms of acute and chronic rejection, chimerism, and tolerance induction, thus providing the foundation to propagate superior allograft survival strategies for larger animal models and future clinical application. Herein, we developed a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique. The experience in syngeneic and allogeneic transplant settings is described for future broader immunological investigations via an instructional manuscript and video supplement.

To study combined solid organ and vascularized composite allotransplantation, we describe a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique.

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression ...

Vessel-sparing Excision and Primary Anastomosis

Urethroplasty is considered to be the standard treatment for urethral strictures since it provides excellent long-term success rates. For isolated short bulbar or posterior urethral strictures, urethroplasty by excision and primary anastomosis (EPA) is recommended. As EPA only requires the excision of the narrowed segment and the surrounding spongiofibrosis, a full-thickness transection of the corpus spongiosum, as performed in the traditional transecting EPA (tEPA), is usually unnecessary. Jordan et al. introduced the idea of a vessel-sparing approach in 2007, aiming to reduce surgical trauma, especially to the dual arterial blood supply of the urethra, and, thus, potentially reducing the risk of postoperative erectile dysfunction or glans ischemia. This approach could also be beneficial for subsequent urethral interventions such as redo urethroplasty using a free graft, in which a well-vascularized graft bed is imperative. Nevertheless, these potential benefits are only assumptions as prospective studies comparing the functional outcome of both techniques with validated questionnaires are currently lacking. Moreover, vessel-sparing EPA (vsEPA) should at least be able to provide similar surgical outcomes as tEPA. The aim of this paper is to give an elaborate, step-by-step overview of how to manage patients with isolated short bulbar or posterior urethral strictures with vsEPA. The main objective of this manuscript is to outline the surgical technique and to report the representative surgical outcome. A total of 117 patients were managed according to the described protocol. The analysis was performed on the entire patient cohort and on the bulbar (n = 91) and posterior (n = 26) vsEPA group separately. Success rates were 93.4% and 88.5% for the bulbar and posterior vsEPA, respectively. To conclude, vsEPA, as outlined in the protocol, provides excellent success rates with low complication rates for isolated short bulbar and posterior urethral strictures.

Here, we present an elaborate and efficient protocol to treat isolated short bulbar or posterior urethral strictures with vessel-sparing excision and primary anastomosis.

Urethroplasty is considered to be the standard treatment for urethral strictures since it provides excellent long-term success rates. For isolated short ...

Contusion Spinal Cord Injury via a Microsurgical Laminectomy in the Regenerative Axolotl

The purpose of this study is to establish a standardized and reproducible regenerative blunt spinal cord injury model in the axolotl (Ambystoma mexicanum). Most clinical spinal cord injuries occur as high energy blunt traumas, inducing contusion injuries. However, most studies in the axolotl spinal cord have been conducted with sharp traumas. Hence, this study aims to produce a more clinically relevant regenerative model. Due to their impressive ability to regenerate almost any tissue, axolotls are widely used as models in regenerative studies and have been used extensively in spinal cord injury (SCI) studies. In this protocol, the axolotls are anesthetized by submersion in a benzocaine solution. Under the microscope, an angular incision is made bilaterally at a level just caudal to the hind limbs. From this incision, it is possible to dissect and expose the spinous processes. Using forceps and scissors, a two-level laminectomy is performed, exposing the spinal cord. A custom trauma device consisting of a falling rod in a cylinder is constructed, and this device is used to induce a contusion injury to the spinal cord. The incisions are then sutured, and the animal recovers from anesthesia. The surgical approach is successful in exposing the spinal cord. The trauma mechanism can produce contusion injuries to the spinal cord, as confirmed by histology, MRI, and neurological examination. Finally, the spinal cord regenerates from the injury. The critical step of the protocol is removing the spinous processes without inflicting damage to the spinal cord. This step requires training to ensure a safe procedure. Furthermore, wound closure is highly dependent on not inflicting unnecessary damage to the skin during incision. The protocol was performed in a randomized study of 12 animals.

This manuscript presents protocols for surgically inflicting controlled blunt and sharp spinal cord injuries to a regenerative axolotl (Ambystoma mexicanum).