This work was supported by grants from the Ministry of Education Innovation Team Development Program of China (No. IRT1279).

The protocol was carried out in accordance with the Guidelines for the Care and Use of Laboratory Animals and was approved by the Committee on the Ethics of Animal Experiments of Xi&#39;an Jiaotong University, Xi&#39;an, Shaanxi Province, China.
1. Preparation prior to the training
NOTE: The vascular reconstruction training machine is shown in Figure 1. It consists of a control panel and an operating floor.
<ol>
	<li>Click the Clean button on the control panel to clean and drain the residual liquid from the operating floor.</li>
	<li>Click the Add Liquid button on the control panel and add 0.9% saline into the machine from the operating floor until the prompt &#34;The testing liquid is adequate&#34; appears on the control panel.</li>
	<li>Prepare the magnetic tractor, which consists of a circular permanent magnet with a diameter of 20 mm and a thickness of 1 mm, an acrylonitrile butadiene styrene (ABS) plastic casing, a spiral spring, a 30 cm nylon traction wire, and a stainless-steel clamp with plastic sleeves or a vascular clamp.
	<ol>
		<li>Glue the circular magnet and the plastic casing together using an acrylate adhesive. The traction force will increase with the lengthening of the traction wire. Use a universal testing machine to test the association between the length of the traction wire and the traction force (Figure 2).</li>
		<li>Fix the clamp and the plastic casing on the upper holder and the lower holder of the universal testing machine, respectively. Gradually elevate the upper holder to stretch the traction wire between the two holders. Test the strength of the traction wire while it is being stretched. 
		NOTE: The magnetic suture tractor and magnetic vascular tractor are shown in Figure 3.</li>
	</ol>
	</li>
	<li>Prepare a magnetic suture puller.
	<ol>
		<li>Use a quasi-oval polylactic acid board with a thickness of 2 mm, a major axis diameter of 10 cm, a minor axis diameter of 2 cm, three magnetic balls with a diameter of 5 mm, and three magnetic cylinders with a diameter of 5 mm and a height of 5 mm.</li>
		<li>Punch three holes with a diameter of 3 mm and a depth of 0.5 mm on the polylactic acid board, so that the magnetic balls can cling to the board by magnetic attraction force from the magnetic cylinders under the board. 
		NOTE: The magnetic suture puller is shown in Figure 4.</li>
	</ol>
	</li>
	<li>Fix the suture under the magnetic ball after one stitch. This plays the role of a suture puller, preventing the previous suture from loosening. Extract the end with the suture needle with a force of about 0.3 N, closely in parallel to the polylactic acid board, and continue the next stitch.</li>
	<li>Ligate all the vein branches using 3-0 silk sutures to avoid leakage after anastomosis. Use tissue scissors to trim the ends of the veins and clear the excess tissue on the wall of the veins to make the veins smooth. 
	NOTE: The vasculature used in this study included the right and left iliac veins (diameter ~10 mm) harvested from Bama pigs weighing 50&#8211;60 kg. To simplify the reconstruction, only a few branches of the iliac veins were picked, and the two veins were similar in size. The vasculature was kept at -20 &#176;C. Before training, it was immersed in 0.9% saline at room temperature.</li>
</ol>
2. Fix the veins on the operating floor
<ol>
	<li>Tie the two veins on the water inlet and water outlet of the training machine with 2-0 silk sutures. 
	NOTE: This study uses the two-point vascular anastomosis5.</li>
	<li>Adjust the length of the water outlet of the training machine and ensure that the ends of the two veins are tension-free in a parallel direction.</li>
	<li>Straighten the veins and lay two 4-0 polypropylene traction sutures at the 6 o&#8217;clock and 12 o&#8217;clock positions.</li>
	<li>Insert the needle of the traction sutures from the outside of the vein and then insert from the inside of the other vein.</li>
	<li>Wet the surgical glove and sutures to avoid damaging the sutures. Gently tie at least five knots to avoid tearing the walls of the veins.</li>
	<li>Use the two stainless-steel clamps of the magnetic suture tractor to grasp the traction sutures and attract the circular magnets of the magnetic suture tractors to the ferromagnetic stainless-steel operating floor. Adjust the position of magnetic attraction and ensure that the ends of the two veins are stretched in a vertical direction.</li>
	<li>Use the two vascular clamps of the magnetic vascular tractor to clamp the anterior wall of the veins and attract the circular magnets of the magnetic vascular tractors on the operating floor. Adjust the position of attraction and ensure that the anterior walls of the veins are retracted, and the posterior walls of the veins are clearly exposed.</li>
</ol>
3. Anastomosis of posterior walls
<ol>
	<li>Use the two stainless-steel clamps of the magnetic suture tractor to grasp the traction sutures and attract the circular magnets of the magnetic suture tractors on the ferromagnetic stainless-steel operating floor. Leave the tail segment of the polypropylene suture at the 12 o&#8217;clock position for the traction suture and use the segment with the needle for continuous suture.</li>
	<li>Ensure intima-to-intima contact between the two veins.</li>
	<li>Insert the first suture from the outside of the vein to the inside.</li>
	<li>In subsequent sutures, insert the needle from the inside of the vein and then insert from the outside of the other vein.</li>
	<li>Check that the sutures are not loose.</li>
	<li>After one suture, ensure that the polypropylene suture is hung on the magnetic suture puller and pull the polypropylene gently until the magnetic ball presses the polypropylene.</li>
	<li>Extract the end with the needle of the suture with a force of about 0.3 N, closely in parallel to the polylactic acid board, and continue the next stitch. 
	NOTE: By using this technique, the tail of the polypropylene suture will be sufficiently tight. As the sutures continue, the polypropylene suture will become shorter. According to the length of the suture, select the most suitable of the three magnetic balls, and then manually press the suture under it.</li>
	<li>Insert the last suture from the inside of the vein to the outside to ensure intima-to-intima contact between the two veins.</li>
	<li>Avoid stenosis after anastomosis by two means: Keep same, proper margin and needle spacing when stitching, and keep the &#34;growth factor&#34;15 when knotting. 
	NOTE: The &#34;growth factor&#34; is the reserved space away from the vessel wall when tying the first knot after the anastomosis so that vessels can remain flexible rather than stenose.
	<ol>
		<li>Maintain the same suture margin and needle spacing. 
		NOTE: In this study, iliac veins with a diameter of approximately 10 mm were used, so the suture margin and needle spacing was about 1 mm.</li>
		<li>Keep the &#34;growth factor&#34;15 when tying the knots. After anastomosis of the posterior walls, tie the end of the suture and the tail segment of the suture at the 6 o&#8217;clock position together away from the vein wall in order to prevent suture stenosis. Use the standard method for tying knots.</li>
	</ol>
	</li>
</ol>
4. Anastomosis of anterior walls
<ol>
	<li>After anastomosis of the posterior walls, remove the magnetic vascular tractor, leave the tail as a traction suture, and use the segment with the needle at the 6 o&#8217;clock position for the anastomosis of the anterior walls.</li>
	<li>Insert the needle from the outside of the vein and then insert from the inside of the other vein. 
	NOTE: The methods used in the anastomosis of the posterior walls to ensure the intima-to-intima contact between the two veins (the suture not being loose and avoiding stenosis after anastomosis) were followed in the anastomosis of the anterior walls5,15.</li>
	<li>After anastomosis of the anterior walls, cut off two traction sutures using suture scissors.</li>
</ol>
5. Test the effect of anastomosis
<ol>
	<li>Set the test parameters.
	<ol>
		<li>Set the perfusion pressure as 2.0 kPa on the control panel. 
		NOTE: The normal vein pressure will not exceed 2.0 kPa.</li>
		<li>Set the duration of peak pressure as 5 s on the control panel.</li>
		<li>Set the temperature as 25 &#176;C on the control panel.</li>
		<li>Set the pressure deviation as 0.1 kPa on the control panel.</li>
	</ol>
	</li>
	<li>Click the Test button and observe the time and pressure on the control panel and whether the reconstructed vein leaks. 
	NOTE: If the vein does not leak during the peak pressure, the anastomosis is successful. If leaks are found, the leakage position should be located and sutured, and then the test should be performed again. The results of the test in this video are shown in Figure 5.</li>
</ol>
6. Checking the safety of the magnetic suture puller
NOTE: To test whether the magnetic suture puller damaged the polypropylene suture, perform the breaking strength and light microscopy tests. In this experiment, three groups with six segments of 4-0 polypropylene suture in each were tested: a control group with no intervention on the polypropylene suture, a manual group in which the polypropylene suture was manually pulled with sterile gloves 20x, and a magnetic puller group in which the magnetic puller pulled the polypropylene suture 20x.
<ol>
	<li>Test the breaking strength of the polypropylene suture on the universal testing machine. Fix the two ends of the polypropylene suture on the upper holder and the lower holder of the universal testing machine. Gradually elevate the upper holder. Test the strength of the polypropylene suture while it is being stretched. Set the breaking strength as the tension force when the suture snaps. Compare the breaking strength among the three groups and perform pairwise comparisons.</li>
	<li>Observe the damage of the polypropylene suture under a light microscope. Define the number of damage points as the number of fibrous or coarse fracture points visible at 200x magnification. Compare the number of damage points among the three groups and perform pairwise comparisons.</li>
</ol>

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	<li>Obora, Y., Tamaki, N., Matsumoto, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nonsuture+microvascular+anastomosis+using+magnet+rings:+preliminary+report.">Nonsuture microvascular anastomosis using magnet rings: preliminary report.</a> Surgical Neurology. 9 (2), 117-120 (1978).</li><li>Holt, G. P., Lewis, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+technique+for+end-to-end+anastomosis+of+small+arteries.">A new technique for end-to-end anastomosis of small arteries.</a> Surgical Forum. 11, 242-243 (1960).</li><li>Enzmann, F. 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=Trans-Iliac+Bypass+Grafting+for+Vascular+Groin+Complications.">Trans-Iliac Bypass Grafting for Vascular Groin Complications.</a> European Journal of Vascular and Endovascular. , 1-6 (2019).</li><li>Bala, M., 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=Acute+mesenteric+ischemia:+Guidelines+of+the+World+Society+of+Emergency+Surgery.">Acute mesenteric ischemia: Guidelines of the World Society of Emergency Surgery.</a> World Journal of Emergency Surgery. 12 (1), 1-11 (2017).</li><li>Makowka, 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=Surgical+Technique+of+Orthotopic+Liver+Transplantation.">Surgical Technique of Orthotopic Liver Transplantation.</a> Gastroenterology Clinics of North America. 17 (1), 33-51 (1998).</li><li>Tang, A. 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=The+elimination+of+anastomosis+in+open+trauma+vascular+reconstruction:+A+novel+technique+using+an+animal+model.">The elimination of anastomosis in open trauma vascular reconstruction: A novel technique using an animal model.</a> Journal of Trauma and Acute Care Surgery. 79 (6), 937-942 (2015).</li><li>Rivas, 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=Magnetic+Surgery:+Results+from+First+Prospective+Clinical+Trial+in+50+Patients.">Magnetic Surgery: Results from First Prospective Clinical Trial in 50 Patients.</a> Annals of Surgery. 267 (1), 88-93 (2018).</li><li>Arain, N. 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The authors have nothing to disclose.

With the help of magnetic tractors and a magnetic suture puller, a trainee can complete vein anastomosis individually and precisely. Magnetic tractors pull the tissue that blocks the anastomosis field and provide suitable strength for stretching the veins in a vertical direction, thus achieving clear exposure for vein anastomosis. In traditional manual anastomosis, at least one assistant is required for surgical exposure. The use of magnetic tractors could achieve the required exposure and substitute for assistants. In a...

Vascular reconstruction is a basic skill required for surgeons. Although Obora1 and Holt2 invented several mechanical reconstruction methods to simplify the reconstruction of small vessels (diameters &#60;10 mm), these methods are not commonly applied in macrovascular anastomosis. Manual vascular anastomosis is still performed in many operations, including vascular surgery3, emergency surgery4, and solid organ transplantation5. Thus, it is essential for surgeons to practice manual vascular anastomosis. However, an optimal training system for vascular reconstruction in vitro is uncommon, and inexperienced surgeons must undergo considerable training in vivo on large animals6 before they can master the technique. Because failure is inevitable during initial training, many animals are likely to die of vascular complications, which is concerning regarding animal welfare. Further, during the procedure of end-to-end vascular reconstruction, to avoid mistakes in stitch positions or loose sutures, the surgeon needs at least one assistant to expose the posterior vascular wall and pull the suture. Thus, vascular reconstruction usually cannot be performed by the surgeon individually, and the efficiency of preparation is usually limited by the proficiency of the assistant.
Magnetic anchoring surgery has become a topic of interest in recent years7,8,9,10,11. The clinical trial by Rivas et al.7 showed that with his magnetic surgical instrument and following the principle of magnetic anchoring, surgeons can perform reduced-port laparoscopic cholecystectomy. The use of this instrument also allows for a reduced role for the assistant during open surgery. Through the magnetic field, the magnetic device is adsorbed onto an anchoring point. This magnetic anchoring device can act as a mechanical arm, grasping and retracting the tissue or organ, exposing the surgical field, and simplifying the operation. Based on this rationale, we invented magnetic tractors to retract the vascular wall and suture, and a magnetic suture puller to pull the polypropylene sutures.
The use of a vascular reconstruction training machine was another milestone in this study. It consists of an operating floor and a control panel: the vasculature is fixed on the operating floor, and the trainee can practice on it. After anastomosis, the trainee can set the perfusion parameters on the control panel in order to test the quality of anastomosis. Compared to previous vascular anastomosis training systems6,12,13,14, the use of this system provides two main advantages: First, magnetic devices can be used to expose the surgical field, so that the trainees can practice on it individually. Second, the trainee can check the effect of anastomosis using a perfusion test.
In the present study, we introduce a training and testing system where the trainee can complete manual vascular reconstruction in vitro individually using a magnetic anchoring technique and the quality of reconstruction can also be tested. Limited by the design and size of the water inlet and water outlet on the operating floor, the training system can only perform end-to-end reconstruction on vessels with a &#62;5 mm diameter.

<table border="0" cellpadding="0" cellspacing="0" style="width:1140px;" width="1140">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Circular permanent magnet</td>
			<td style="width:244px;">Hangzhou Permanent Magnet Group Co.LTD</td>
			<td style="width:161px;">20*1mm</td>
			<td style="width:519px;">Magnetic tractor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Magnetic balls</td>
			<td style="width:244px;">Hangzhou Permanent Magnet Group Co.LTD</td>
			<td style="width:161px;">5mm</td>
			<td style="width:519px;">Magnetic suture puller</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Magnetic cylinders</td>
			<td style="width:244px;">Hangzhou Permanent Magnet Group Co.LTD</td>
			<td style="width:161px;">5*5mm</td>
			<td style="width:519px;">Magnetic suture puller</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Polypropylene suture</td>
			<td style="width:244px;">Johnson and Johnson</td>
			<td style="width:161px;">PROLENE 4-0</td>
			<td style="width:519px;">Used for anastomosis</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Silk suture</td>
			<td style="width:244px;">SILK</td>
			<td style="width:161px;">2-0&#65292;3-0</td>
			<td style="width:519px;">Used for fixing vascular and ligation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Surgical insturments</td>
			<td style="width:244px;">Jinzhong Shanghai</td>
			<td style="width:161px;">JZ-2018</td>
			<td style="width:519px;">Suture scissors, tissue scissors&#65292; forceps, needle and needle holder</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Universal testing machine</td>
			<td style="width:244px;">Zwick GmbH&#38;Co</td>
			<td style="width:161px;">Z010</td>
			<td style="width:519px;">Used for testing the association between the length of traction wire and the traction force</td>
		</tr>
	</tbody>
</table>

a training and testing system for performing vascular reconstruction in vitro

Manual vascular reconstruction training is essential for a beginner surgeon. However, an optimal training system for vascular reconstruction in vitro has yet to be developed. In this study, we introduce an in vitro training and testing system using a magnetic anchoring technique with which a trainee can practice manual vascular reconstruction individually. Additionally, this system can also be used to test the quality of the reconstruction. The described system includes a vascular reconstruction training machine, magnetic tractors, and a magnetic suture puller. In this manuscript, we detail an end-to-end vein anastomosis using porcine right and left iliac veins. To identify the potential damage caused by a magnetic suture puller on the suture, we created three groups with six segments of 4-0 polypropylene sutures each: a control group with no intervention on the polypropylene suture, a group in which the polypropylene suture is manually pulled with sterile gloves 20x, and a magnetic puller group in which the magnetic puller pulled the polypropylene suture 20x. These groups were tested by light microscopy and breaking strength tests, and the effect of reconstruction was assessed. In the light microscopy test, the control group was less likely to be damaged (p &#60; 0.05) and the number of damaged points of the manual group and magnetic puller group were similar (p &#62; 0.05). The results of the breaking strength test were compared across groups and no significant difference was observed (p &#62; 0.05). The end-to-end anastomosis of the porcine iliac veins was successfully performed using this training system, and the reconstructed veins could undergo 2.0 kPa perfusion pressure. Using this training and testing system the trainee can practice manual vascular reconstruction in vitro individually with the aid of magnetic tractors and a magnetic suture puller, and the quality of the reconstruction can be tested.

The vascular reconstruction training machine is shown in Figure 1 and includes two main parts: the operating floor and the control panel. The operating floor consists of a water inlet, a water outlet, and a water storage basin. The two ends of the vasculature are tied to the water inlet and water outlet to test the effect of anastomosis. The length of the water outlet is adjustable, and we set the parameters (e.g., the perfusion pressure, duration of peak pressure, temperature, and pressure ...

Watch this Scientific Journal Video about A Training and Testing System for Performing Vascular Reconstruction In Vitro at JoVE.com

A Training and Testing System for Performing Vascular Reconstruction In Vitro

Here we present a training and testing system where a trainee can complete manual vascular reconstruction in vitro individually using a magnetic anchoring technique. The system can also be used to test the quality of reconstruction.

Manual vascular reconstruction training is essential for a beginner surgeon. However, an optimal training system for vascular reconstruction in vitro has ...

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7.9K Views.  First Affiliated Hospital of Xi'an Jiaotong University. Here we present a training and testing system where a trainee can complete manual vascular reconstruction in vitro individually using a magnetic anchoring technique. The system can also be used to test the quality of reconstruction.

Video: A Training and Testing System for Performing Vascular Reconstruction In Vitro

Vascular Occlusion Training for Inclusion Body Myositis: A Novel Therapeutic Approach

Inclusion body myositis (IBM) is a rare idiopathic inflammatory myopathy. It is known to produces remarkable muscle weakness and to greatly compromise function and quality of life. Moreover, clinical practice suggests that, unlike other inflammatory myopathies, the majority of IBM patients are not responsive to treatment with immunosuppressive or immunomodulatory drugs to counteract disease progression1. Additionally, conventional resistance training programs have been proven ineffective in restoring muscle function and muscle mass in these patients2,3. Nevertheless, we have recently observed that restricting muscle blood flow using tourniquet cuffs in association with moderate intensity resistance training in an IBM patient produced a significant gain in muscle mass and function, along with substantial benefits in quality of life4. Thus, a new non-pharmacological approach for IBM patients has been proposed. Herein, we describe the details of a proposed protocol for vascular occlusion associated with a resistance training program for this population.

The present article presents the details pertaining to the application of resistance training associated to vascular occlusion in IBM patients.

Inclusion body myositis (IBM) is a rare idiopathic inflammatory myopathy. It is known to produces remarkable muscle weakness and to greatly compromise ...

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 ...

Evaluation of Tumor-infiltrating Leukocyte Subsets in a Subcutaneous Tumor Model

Specialized immune cells that infiltrate the tumor microenvironment regulate the growth and survival of neoplasia.&#160; Malignant cells must elude or subvert anti-tumor immune responses in order to survive and flourish. Tumors take advantage of a number of different mechanisms of immune &#8220;escape,&#8221; including the recruitment of tolerogenic DC, immunosuppressive regulatory T cells (Tregs), and myeloid-derived suppressor cells (MDSC) that inhibit cytotoxic anti-tumor responses. Conversely, anti-tumor effector immune cells can slow the growth and expansion of malignancies: immunostimulatory dendritic cells, natural killer cells which harbor innate anti-tumor immunity, and cytotoxic T cells all can participate in tumor suppression. The balance between pro- and anti-tumor leukocytes ultimately determines the behavior and fate of transformed cells; a multitude of human clinical studies have borne this out. Thus, detailed analysis of leukocyte subsets within the tumor microenvironment has become increasingly important. Here, we describe a method for analyzing infiltrating leukocyte subsets present in the tumor microenvironment in a mouse tumor model. Mouse B16 melanoma tumor cells were inoculated subcutaneously in C57BL/6 mice. At a specified time, tumors and surrounding skin were resected en bloc and processed into single cell suspensions, which were then stained for multi-color flow cytometry. Using a variety of leukocyte subset markers, we were able to compare the relative percentages of infiltrating leukocyte subsets between control and chemerin-expressing tumors. Investigators may use such a tool to study the immune presence in the tumor microenvironment and when combined with traditional caliper size measurements of tumor growth, will potentially allow them to elucidate the impact of changes in immune composition on tumor growth. Such a technique can be applied to any tumor model in which the tumor and its microenvironment can be resected and processed.

This protocol describes a method for the detailed evaluation of leukocyte subsets within the tumor microenvironment in a mouse tumor model. Chemerin-expressing B16 melanoma cells were implanted subcutaneously into syngeneic mice. Cells from the tumor microenvironment were then stained and analyzed by flow cytometry, allowing for detailed leukocyte subset analyses.

Specialized immune cells that infiltrate the tumor microenvironment regulate the growth and survival of neoplasia.&#160; Malignant cells must elude or ...

Comparing the Effects of Electronic Cigarette Vapor and Cigarette Smoke in a Novel In Vivo Exposure System

Electronic cigarettes (E-cigarettes) are being widely used, and growing in popularity. It is estimated that more than 9 million adults use them regularly. The potential adverse health effects of electronic cigarette vapor (E-vapor) exposure are poorly defined. While several animal models of E-vapor exposure have been developed, few models expose rodents to clinically relevant quantities of nicotine and make direct comparisons to cigarette smoke within the same exposure system. Here, we present a method for constructing and operating an E-vapor chamber and cigarette smoke chamber. The chambers are constructed by outfitting anesthesia chambers with a computer controlled pumping system that delivers consistent amounts of E-vapor or cigarette smoke to rodents. Nicotine exposure is measured indirectly by quantifying pre and post-exposure serum cotinine levels. This exposure system can be modified to accommodate various types of E-cigarettes&#160;and tobacco cigarettes, and can be used to compare the effects of E-vapor&#160;and cigarette smoke in vivo.

Comparing the Effects of Electronic Cigarette Vapor and Cigarette Smoke in a Novel In Vivo Exposure System

This protocol describes a method for exposing rodents to electronic cigarette vapor (E-vapor) and cigarette smoke. Exposure chambers are constructed by modifying anesthesia chambers with an automated pumping system that delivers E-vapor or cigarette smoke to rodents. This system can easily be modified to accommodate many experimental endpoints.

Electronic cigarettes (E-cigarettes) are being widely used, and growing in popularity. It is estimated that more than 9 million adults use them regularly. ...

Muscle Imbalances: Testing and Training Functional Eccentric Hamstring Strength in Athletic Populations

Many hamstring injuries that occur during physical activity occur while the muscles are lengthening, during eccentric hamstring muscle actions. Opposite of these eccentric hamstring actions are concentric quadriceps actions, where the larger and likely stronger quadriceps straighten the knee. Therefore, to stabilize the lower limbs during movement, the hamstrings must eccentrically combat against the strong knee-straightening torque of the quadriceps. As such, eccentric hamstring strength expressed relative to concentric quadricep strength is commonly referred to as the &#34;functional ratio&#34; as most movements in sports require simultaneous concentric knee extension and eccentric knee flexion. To increase the strength, resiliency, and functional performance of the hamstrings, it is necessary to test and train the hamstrings at different eccentric speeds. The main purpose of this work is to provide instructions for measuring and interpreting eccentric hamstring strength. Techniques for measuring the functional ratio using isokinetic dynamometry are provided and sample data will be compared. Additionally, we briefly describe how to address hamstring strength deficiencies or unilateral strength differences using exercises that specifically focus on increasing eccentric hamstring strength.

The hamstrings are a group of muscles that are sometimes problematic for athletes, resulting in soft tissue injury in the lower limbs. To prevent such injuries, functional training of the hamstrings requires intensive eccentric contractions. Additionally, hamstring function should be tested in relation to quadricep function at different contraction speeds.

Many hamstring injuries that occur during physical activity occur while the muscles are lengthening, during eccentric hamstring muscle actions. Opposite ...

Porcine As a Training Module for Head and Neck Microvascular Reconstruction

Live models that resemble surgical conditions of humans are needed for training free-flap harvesting and anastomosis. Animal models for training purposes have been available for years in many surgical fields. We used the female (because they are easy to handle for the procedure) Yorkshire pigs for the head and neck reconstruction by harvesting the deep inferior epigastric artery perforator or the superior epigastric artery perforator flap. The anastomosis site (neck skin defect or tracheal wall defect) was prepared via the dissection of the common carotid artery and the internal jugular vein, in which 3.5&#215; loupe magnification was used for anastomosis as we use on human cases in real life. This procedure demonstrates a new training method using a reliable learning model and provides a detailed anatomy in a live scenario. We focused on the ischemia time, harvesting, vessel anastomosis, and designing the flap to fit the defect site. This model improves tissue handling and with the use of proper instruments can be repeated many times so that the surgeon is fully confident before starting the surgery on humans.

Here we present a protocol for the use of the pig superior epigastric artery perforator flap as a learning module for head and neck microvascular reconstruction.

Live models that resemble surgical conditions of humans are needed for training free-flap harvesting and anastomosis. Animal models for training purposes ...

Laser Doppler Perfusion Imaging in the Mouse Hindlimb

Blood flow recovery is a critical outcome measure after experimental hindlimb ischemia or ischemia-reperfusion. Laser Doppler perfusion imaging (LDPI) is a common, noninvasive, repeatable method for assessing blood flow recovery. The technique calculates overall blood flow in the sampled tissue from the Doppler shift in frequency caused when a laser hits moving red blood cells. Measurements are expressed in arbitrary perfusion units, so the contralateral non-intervened upon leg is usually used to help control measurements. Measurement depth is in the range of 0.3-1 mm; for hindlimb ischemia, this means that dermal perfusion is assessed. Dermal perfusion is dependent on several factors&#8212;most importantly skin temperature and anesthetic agent, which must be carefully controlled to result in reliable readings. Furthermore, hair and skin pigmentation can alter the ability of the laser to either reach or penetrate to the dermis. This article demonstrates the technique of LDPI in the mouse hindlimb.

Here, we present a protocol that demonstrates the technique and necessary controls for Laser Doppler perfusion imaging to measure blood flow in the mouse hindlimb.

Blood flow recovery is a critical outcome measure after experimental hindlimb ischemia or ischemia-reperfusion. Laser Doppler perfusion imaging (LDPI) is ...

A Real-World High-Intensity Interval Training Protocol for Cardiorespiratory Fitness Improvement

High-Intensity Interval Training (HIIT) has emerged as an interesting time-efficient approach to increase exercise adherence and improve health. However, few studies have tested the efficiency of HIIT protocols in a &#34;real world&#34; setting, e.g., HIIT protocols designed for outdoor spaces without specialized equipment. This study presents a &#34;real world&#34; training protocol, named &#34;beep training&#34;, and compares the efficiency of a HIIT regiment versus a traditional long-duration Moderate-Intensity Continuous Training (MICT) regiment using this beep training protocol on VO2 max of overweight untrained men. Twenty-two subjects performed outdoor running with MICT (n = 11) or HIIT (n = 11). Cardiorespiratory fitness was assessed before and after training protocols using a metabolic analyzer. Both training protocols were performed 3 days a week for 8 weeks using the Beep Test results. The MICT group performed the exercise program at 60%-75% of the maximum speed of the 20 m shuttle test (Vmax) and with a progression of the distance of 3,500-5,000 m. The HIIT group performed the interval exercise with 7-10 bouts of 200 m at 85%-100% of the maximum speed of the 20 m shuttle test (Vmax), interspersed with 1 min of passive recovery. Although the HIIT group presented a significantly lower training volume than the MICT group (p &#60; 0.05) after 8 weeks of beep training, HIIT was superior to MICT in improving VO2 max (MICT: ~4.1%; HIIT: ~7.3%; p &#60; 0.05). The &#34;real world&#34; HIIT regiment based on beep training protocol is a time-efficient, low-cost, and easy-to-implement protocol for overweight untrained men.

This study presents a low-cost and easy-to-implement "real world" high-intensity interval training (HIIT) protocol for scientific research and discusses its efficiency for cardiorespiratory fitness.

High-Intensity Interval Training (HIIT) has emerged as an interesting time-efficient approach to increase exercise adherence and improve health. However, ...

Strategies for Efficient Vascular Anastomosis in Mouse Liver Transplantation

Vascular Reconstruction with the Cuff Technique in Mouse Orthotopic Liver Transplantation

Mouse orthotopic liver transplantation is an effective methodology for investigating the underlying mechanisms of liver ischemia and reperfusion injury. However, the technical challenges pose a barrier to utilizing this valuable experimental model and passing on these skills to the next generation. The most challenging aspect of this procedure is vascular reconstruction, including the portal vein (PV), infrahepatic inferior vena cava (IHIVC), and suprahepatic inferior vena cava. The use of plastic cuffs, rather than sutures, allows for smoother PV and IHIVC reconstruction. Vessels are reconstructed by attaching a cuff made from an intravenous catheter to the tip of the graft vessel and interposing the cuff into the recipient vessel. The two most crucial aspects are properly visualizing the inner lumen of the vessel and avoiding the use of excessive force. Our aim is to provide a technical overview of vascular reconstructions using the cuff technique in recipient surgery. These technical tips for the cuff technique are expected to help microsurgeons facilitate vascular reconstruction and advance their research.

Author Spotlight: Exploring Cold Ischemia and Warm Reperfusion Injury in Fatty Liver Transplantation

This protocol provides technical information for vessel reconstruction using the cuff technique in mouse orthotopic liver transplantation.

Mouse orthotopic liver transplantation is an effective methodology for investigating the underlying mechanisms of liver ischemia and reperfusion injury. ...

Enhancing Stroke Rehabilitation with Virtual Reality-Based Occupational Training

Cognitive Function and Upper Limb Rehabilitation Training Post-Stroke Using a Digital Occupational Training System

Stroke rehabilitation often requires frequent and intensive therapy to improve functional recovery. Virtual reality (VR) technology has shown the potential to meet these demands by providing engaging and motivating therapy options. The digital occupational training system is a VR application that utilizes cutting-edge technologies, including multi-touch screens, virtual reality, and human-computer interaction, to offer diverse training techniques for advanced cognitive capacity and hand-eye coordination abilities. The objective of this study was to assess the effectiveness of this program in enhancing cognitive function and upper extremity rehabilitation in stroke patients. The training and assessment consist of five cognitive modules covering perception, attention, memory, logical reasoning, and calculation, along with hand-eye coordination training. This research indicates that after eight weeks of training, the digital occupational training system can significantly improve cognitive function, daily living skills, attention, and self-care abilities in stroke patients. This software can be employed as a time-saving and clinically effective rehabilitation aid to complement traditional one-on-one occupational therapy sessions. In summary, the digital occupational training system shows promise and offers potential financial benefits as a tool to support the functional recovery of stroke patients.

Author Spotlight: Rehabilitation of Stroke Patients With a Digital Occupational Training System

The current protocol outlines how the VR-based digital occupational training system enhances the rehabilitation of patients with cognitive impairment and upper limb dysfunction following a stroke.