We thank Miss Isabelle Lu for the English narration.

All procedures for animal use were approved by the National Yang-Ming University Institutional Animal Care and Use Committee and complied with the &#8220;Guide for the Care and Use of Laboratory Animals. 8th edition, 2011&#8221;. The anesthetic procedure was performed according to the guideline for the pain control, anesthesia, pre-operative, and postoperative care for the experimental animals from National Yang Ming University, ICAUC-016, 2015, 1st edition.
1. Animal preparation
<ol>
	<li>Obtain 20 8-week-old wild type C57BL/6 male mice from the National Laboratory Animal Center (NLAC) that weigh around 16-18 grams. Randomly assign these mice into the SG or the sham operation group with 10 mice each.</li>
	<li>Start the mice on chow or a high-fat diet (see <a href="https://www.jove.com/files/ftp_upload/60719/High fat diet.pdf" target="_blank">Supplemental File</a>) prior to the bariatric or sham surgeries9. Produce diet-induced obesity with at least 2 preoperative weeks of the HFD.</li>
	<li>House the mice in the National Yang-Ming University Laboratory Animal Center under a light-dark cycle of 12:12 with light onset at 07:00 A.M. and allow free feeding to standard rodent chow and water.</li>
	<li>Restrict all animals from food but allow free water access on the night before surgery.</li>
</ol>
2. General preoperative preparation
<ol>
	<li>Fast the mice for at least 6 hours before the surgery. Induce anesthesia in an induction chamber with 5% isoflurane and oxygen (both 3-4 L/min).
	<ol>
		<li>Check the depth of anesthesia ensuring that pinch stimulation (blunt curved serrated smooth micro-forceps) of the hind paw, the forepaw, and the ear does not evoke any motor reflex. Administer antibiotics (25 mg/kg cefazolin) via subcutaneous injections before the operation.</li>
	</ol>
	</li>
	<li>Assign a particular work zone for surgical procedures. Clean the surgical area and spray the surgical table with 75% alcohol solution before an operation.</li>
	<li>Place the mouse in the nose cone and maintain anesthesia with 2% isoflurane (2 L/min) and oxygen (4 L/min).</li>
	<li>For sleeve gastrectomy and sham operation, after confirming the appropriate depth of anesthesia by toe stimulation, apply the 0.2% Carbomer eye gel on the mouse&#8217;s eyes to prevent dry eyes.</li>
	<li>Use an electric razor to shave hair from the abdomen to the sternum. Remove and clean the remaining hair with a depilatory cream. Place and fix the mouse on the surgical table in a supine position.</li>
	<li>Disinfect the abdominal wall of the mouse with povidone-iodine solutions alternating with 75% alcohol for 3 applications.</li>
</ol>
3. Sleeve gastrectomy and sham procedures
<ol>
	<li>Median laparotomy
	<ol>
		<li>Use a micro-scissor to make a 1-1.5 cm length midline incision at the upper abdomen.</li>
	</ol>
	</li>
	<li>Stomach and intestine externalization
	<ol>
		<li>Perform the SG procedure with the aid of a magnifying dissecting microscope or magnifier as necessary to prevent the unpredictable bleeding and hypovolemic shock.</li>
		<li>Use two curved smooth serrated micro-forceps to gently move the stomach and complete externalize it.</li>
		<li>Carefully divide the gastrosplenic ligament connecting the left stomach to the spleen using a cotton-tipped probe and electrocautery as needed, thereby dissecting the gastric fundus from the surrounding spleen and other internal organs.</li>
	</ol>
	</li>
</ol>
4. Stomach isolation and clip
NOTE: The SG for mice was performed using the new clip applier technique as previously published10.
<ol>
	<li>Stretch the fundus and pylorus of the stomach gently and laterally with forceps, identify the midline (Figure 1 and Figure 2), and carefully apply surgical clips to half of the medial side of the stomach midline from the gastroesophageal junction inferiorly and the lower pole superiorly. Clamp and exclude around 75-80% of stomach, thereby creating the entire lateral sleeve of the stomach (Figure 1 and Figure 2).</li>
	<li>Move the intestine to the bare skin on the side and cover it with a warm-saline moistened gauze and perform intraperitoneal hydration as needed to prevent dehydration and hypothermia.</li>
	<li>Resect the lateral clipped stomach, remove the excluded sleeve of the stomach with microscissors and then sterilize the cut edge of the stomach with povidone-iodine solution.</li>
	<li>Oversew the clip line with a 5-0 monofilament nonabsorbable suture to ensure no leakage. Knot the ends of the suture and anchor to the clips on either end.</li>
	<li>Return the stomach and intestine to the proper position in the abdominal cavity and close the abdomen with a running 5-0 monofilament nonabsorbable continuous and discontinuous suture to the fascia and abdominal wall.</li>
	<li>Administer analgesics with ketoprofen (2-5 mg/kg) and antibiotics with cefazolin (25 mg/kg) intraperitoneally after the whole procedure.</li>
</ol>
5. Sham procedure of SG
<ol>
	<li>Perform a similar procedure as described previously with a midline laparotomy, and externalize the intestine and stomach using 37 &#176;C wet warm saline gauze coverage for 5 min.</li>
	<li>Return the stomach and intestine to the proper sites of these internal organs.</li>
	<li>Close the abdominal wall carefully as previously described with 2 layers of continuous and discontinuous closure to the fascia and skin using slow or non-absorbable monofilament sutures. Administer analgesics with ketoprofen (2-5 mg/kg) and antibiotics with cefazolin (25 mg/kg) intraperitoneally after the whole procedure.</li>
</ol>
6. General postoperative care
<ol>
	<li>Stop isoflurane and continue with a room air flow of 3-5 L/min until the mouse is fully awake.</li>
	<li>Keep watching the mouse while it regains mobility and begins to walk around the cage.</li>
	<li>Place the mouse in a 30 &#176;C independent incubator for 5 days. Make sure there is only one mouse per cage to prevent the mice from injuring each other.</li>
	<li>Return free access to a gel diet food (high-fat gel diet: 10% lard, 10% liquid sugar, 57% water) for 3 days after surgery and reintroduce the previous assigned diet 3 days after surgery.</li>
	<li>Subcutaneously or intraperitoneally inject ketoprofen (2-5 mg/kg), and cefazolin (25 mg/kg) for 1 day after the operation.</li>
	<li>Assess the mouse body weight weekly through the whole study period. Provide ketoprofen (2-5 mg/kg) by intraperitoneal injection for pain control once daily as needed if the animal is in distress after the operation. 
	NOTE: Here, the survival rate of SG was 90% after the learning period.</li>
</ol>
7. Metabolic parameter assessment
<ol>
	<li>Fast mice for 6 h, and take baseline blood samples (0 min). Obtain all blood samples from the tip of the tail vein of freely moving mice11,12.</li>
	<li>Deliver 1 mg/g of 50% dextrose by intraperitoneal injection for the glucose tolerance test.</li>
	<li>Measure the blood glucose from the tips of the tail veins of freely moving mice at 0, 5, 15, 30, 60, and 120 min after glucose administration on duplicate samples using glucometers and test strips.</li>
	<li>Keep the whole blood sample (n = 10 for each group) at room temperature with clotting for 30 minutes.</li>
	<li>Centrifuge the blood sample at 3,000 x g for 10 minutes at 4 &#176;C.</li>
	<li>Transfer the plasma into separate tubes without disturbing blood clots and store at -80 &#176;C.</li>
	<li>Study the plasma samples at the end of the study using commercial mice ELISA kits for hemoglobin A1c (HBA1c), glucose, cholesterol, and insulin levels according to the manufacturer&#8217;s guidelines.</li>
</ol>

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The authors declare no conflict of interest.

Most of the prospective cohort studies have confirmed that increasing the body mass index is related to increasing the mortality. Obesity is associated with a higher risk of diabetes, cardiovascular diseases (CVD), and death in both sexes. The specific pathologic processes have established a link between diabetes, obesity, and mortality14. The systematic reviews indicate the benefits of bariatric surgery in reducing risk factors for CVD with evidence for ventricular hypertrophy regression and impr...

The prevalence of obesity has increased nearly threefold worldwide since 1975. In 2016, more than 1.9 billion adults older than 18 years were overweight and over 650 million adults were obese. The prevalence of T2D in the adult population has also doubled from 4.7% to 8.5% with the number rising from 108 million to 422 million adults between 1980 and 2014 globally1. Most bariatric surgeries result in significant weight loss, a reduction of body fat, and improvement in glucose tolerance, insulin resistance and diabetes control2. In addition to weight loss and remission of T2D, bariatric surgery further produces additional health advantages such as hypertension control and a lower incidence of certain types of obesity related cancer development and progression3. Bariatric surgery also induces durable complete, partial remission of T2D, and decreases the risk of ten-year major coronary heart disease (CHD) and some degree of cerebrovascular risk. However, the underlying mechanisms are not completely understood4.
There are several procedures for bariatric/metabolic surgery. Both restrictive and gastrointestinal bypass surgeries provide positive effects on metabolism5. Of the bariatric surgical models, SG and modified Roux-en-Y gastric bypass have higher success and lower mortality rates and demonstrate reliable restrictive and gastrointestinal bypass surgery models in mice6. Although the gastrointestinal bypass surgery provides more weight loss and a significant improvement of glucose tolerance and liver steatosis than the restrictive procedure in the long term, the SG still produces good control of body weight and glucose levels and is easy to perform with a low incidence of complications6. The proportion of SG increased from 30% to 54% and Roux-en-Y gastric bypass surgery decreased from 52% to 32% from 2008 to 20147. Currently, laparoscopic SG is the most commonly performed bariatric procedure at the national level within academic centers in the United States8. Although there have been many published reports regarding the pathophysiological processes between bariatric surgery, diabetes and obesity, we need more animal experiments to further explore unknown mechanisms.
This protocol aims to produce an animal method to investigate the mechanisms underlying the outcomes seen after bariatric surgery in humans. The current translational study may provide insights on the mechanism of obesity and T2D treatment from the SG.

<table>
	<tbody>
		<tr>
			<td>0.9% normal saline</td>
			<td>China Chemical &#38; Pharmaceutical Co., Ltd.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>5-0 monofilament suture</td>
			<td>Shineteh Inc. Taiwan</td>
			<td></td>
			<td>Ethicon</td>
		</tr>
		<tr>
			<td>Alm</td>
			<td>Shineteh Inc. Taiwan</td>
			<td>ST-A072PK</td>
			<td>UNIK</td>
		</tr>
		<tr>
			<td>Animal Anesthesia</td>
			<td>Step Technology</td>
			<td>0712VAP11076</td>
			<td>Matrx</td>
		</tr>
		<tr>
			<td>Barraqer-Troutman forceps</td>
			<td>Shineteh Inc. Taiwan</td>
			<td>ST-N309(H-4410)</td>
			<td>UNIK</td>
		</tr>
		<tr>
			<td>cefazolin</td>
			<td>Taiwan veteran pharmacy company</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Chow diet</td>
			<td>Research Diets</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Glucose strips and glucometer</td>
			<td>BeneCheck</td>
			<td>BKM13-1</td>
			<td>BeneCheck</td>
		</tr>
		<tr>
			<td>HbA1c kit</td>
			<td>Level</td>
			<td>OKEH00661</td>
			<td></td>
		</tr>
		<tr>
			<td>Hematology system</td>
			<td>Fuji</td>
			<td>FUJI DRI-CHEM 4000i</td>
			<td>Europe</td>
		</tr>
		<tr>
			<td>High Fat diet</td>
			<td>Research Diets</td>
			<td>1810724</td>
			<td>LabDiet</td>
		</tr>
		<tr>
			<td>Iris Forceps</td>
			<td>Shineteh Inc. Taiwan</td>
			<td>ST-1210</td>
			<td>UNIK</td>
		</tr>
		<tr>
			<td>Iris Scissors</td>
			<td>Shineteh Inc. Taiwan</td>
			<td>ST-S011</td>
			<td>UNIK</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Panion &#38; BF biotech INC</td>
			<td>8547</td>
			<td>Panion &#38; BF</td>
		</tr>
		<tr>
			<td>Ketoprofen</td>
			<td>Sigma</td>
			<td>22071154</td>
			<td>Sigma-Aldrich</td>
		</tr>
		<tr>
			<td>Stereo microscope</td>
			<td>MicroTech</td>
			<td>SZ-5T</td>
			<td>MicroTech</td>
		</tr>
		<tr>
			<td>Surgical clip (M)</td>
			<td>Echicon Inc., Somerville, NJ</td>
			<td></td>
			<td>Size M, 5mm</td>
		</tr>
		<tr>
			<td>Vidisic gel</td>
			<td>Dr. Gerhard Mann Chem.-pharm. Fabrik GmbH</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

sleeve gastrectomy in mice using surgical clips

The number of people who are overweight and obese is continually increasing both in the adult and adolescent populations. This coincides with the increased universal phenomenon of type 2 diabetes (T2D) and other metabolic problems. Bariatric surgery, such as SG, is currently one of the most effective and commonly used long-term treatment for obesity and T2D, but the association between them is not completely explored yet. The mechanisms underlying the outcomes seen after bariatric surgery in humans can be investigated based on preclinical animal studies. The SG reduces body weight, glucose levels and many metabolic parameters, and is easy to perform with a low incidence of complications. The goal of this work is to provide a simple method and an uncomplicated preclinical model of bariatric surgery in animals for researchers.

Results of the operation are shown in Figure 3 and Figure 4. The survival rate of the study was 90%. One mouse died in the sham group because of weakness and another mouse died in the SG group on the third day after operation for unknown reasons. Results obtained on weight loss (Figure 3A) after operation (at week 3) on HFD fed mice were quite similar to those observed in humans, with a final body weight loss of about 15-20%<sup cla...

Watch this Scientific Journal Video about Sleeve Gastrectomy in Mice using Surgical Clips at JoVE.com

Sleeve Gastrectomy in Mice using Surgical Clips

The prevalence of diabetes and obesity is continuously increasing worldwide. The mechanisms between diabetes, obesity and their associated mortality and co-morbidities need to be further investigated. Here, we present a protocol for sleeve gastrectomy (SG) in animals as an uncomplicated preclinical model of bariatric surgery.

The number of people who are overweight and obese is continually increasing both in the adult and adolescent populations. This coincides with the ...

sleeve-gastrectomy-in-mice-using-surgical-clips

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6.4K Views.  Min-Sheng General Hospital. The prevalence of diabetes and obesity is continuously increasing worldwide. The mechanisms between diabetes, obesity and their associated mortality and co-morbidities need to be further investigated. Here, we present a protocol for sleeve gastrectomy (SG) in animals as an uncomplicated preclinical model of bariatric surgery.

Video: Sleeve Gastrectomy in Mice using Surgical Clips

Implantation of Radiotelemetry Transmitters Yielding Data on ECG, Heart Rate, Core Body Temperature and Activity in Free-moving Laboratory Mice

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a manageable size and relatively easy to house. These factors, together with the availability of a wealth of spontaneous and experimentally induced mutants, make laboratory mice ideally suited to a wide variety of research areas.
In cardiovascular, pharmacological and toxicological research, accurate measurement of parameters relating to the circulatory system of laboratory animals is often required. Determination of heart rate, heart rate variability, and duration of PQ and QT intervals are based on electrocardiogram (ECG) recordings. However, obtaining reliable ECG curves as well as physiological data such as core body temperature in mice can be difficult using conventional measurement techniques, which require connecting sensors and lead wires to a restrained, tethered, or even anaesthetized animal. Data obtained in this fashion must be interpreted with caution, as it is well known that restraining and anesthesia can have a major artifactual influence on physiological parameters1, 2.
Radiotelemetry enables data to be collected from conscious and untethered animals. Measurements can be conducted even in freely moving animals, and without requiring the investigator to be in the proximity of the animal. Thus, known sources of artifacts are avoided, and accurate and reliable measurements are assured. This methodology also reduces interanimal variability, thus reducing the number of animals used, rendering this technology the most humane method of monitoring physiological parameters in laboratory animals3, 4. Constant advancements in data acquisition technology and implant miniaturization mean that it is now possible to record physiological parameters and locomotor activity continuously and in realtime over longer periods such as hours, days or even weeks3, 5.
Here, we describe a surgical technique for implantation of a commercially available telemetry transmitter used for continuous measurements of core body temperature, locomotor activity and biopotential (i.e. onelead ECG), from which heart rate, heart rate variability, and PQ and QT intervals can be established in freeroaming, untethered mice. We also present pre-operative procedures and protocols for post-operative intensive care and pain treatment that improve recovery, well-being and survival rates in implanted mice5, 6.

A surgical technique for implantation of commercially available telemetry transmitters used for continuous measurement of biopotential (one-lead ECG), heart rate, core body temperature and locomotor activity in freely moving mice is shown. Recommendations and protocols for post-operative care and pain relief, improving recovery, well being and survival rate are also presented.

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a ...

Minimal Invasive Surgical Procedure of Inducing Myocardial Infarction in Mice

Myocardial infarction still remains the main cause of death in western countries, despite considerable progress in the stent development area in the last decades. For clarification of the underlying mechanisms and the development of new therapeutic strategies, the availability of valid animal models are mandatory. Since we need new insights into pathomechanisms of cardiovascular diseases under in vivo conditions to combat myocardial infarction, the validity of the animal model is a crucial aspect. However, protection of animals are highly relevant in this context. Therefore, we establish a minimally invasive and simple model of myocardial infarction in mice, which assures a high reproducibility and survival rate of animals. Thus, this models fulfils the requirements of the 3R principle (Replacement, Refinement and Reduction) for animal experiments and assure the scientific information needed for further developing of therapeutical strategies for cardiovascular diseases.

A highly reproducible model for myocardial infarction in mice with minimal invasive manipulations is described. The model can be easily performed, resulting in a high reproducibility and survival rate. Thus, the described model will reduce the number of required animals as requested by the 3R principle (Replacement, Refinement and Reduction).

Myocardial infarction still remains the main cause of death in western countries, despite considerable progress in the stent development area in the last ...

Surgical Models of Gastroesophageal Reflux with Mice

Multiple surgical procedures have been reported to induce gastroesophageal reflux in animals. Herein, we report three surgical models with mice aiming to induce reflux of gastric contents, duodenal contents or mixed contents. Surgical procedures and general principles have been described in detail. A researcher with surgical experience should be able to grasp the technique after a short period of practice. After surgery, most mice can survive and develop reflux esophagitis similar to that in humans. However, it should be noted that histological differences between mouse and human esophagus are the inherent limitations of these surgical models. If used for research on Barrett&#8217;s esophagus and adenocarcinoma, these procedures may need to be combined with genetic modifications.

This article demonstrates surgical procedures of gastroesophageal reflux with mice. These models are useful tools for research on mechanisms and treatment of gastroesophageal reflux disease and potentially Barrett&#8217;s esophagus and esophageal adenocarcinoma.

Multiple surgical procedures have been reported to induce gastroesophageal reflux in animals. Herein, we report three surgical models with mice aiming to ...

Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ischemic in origin and occur in the territory of the middle cerebral artery (MCA). Currently the only Food and Drug Administration (FDA) approved drug for the treatment of this devastating disease is tissue plasminogen activator (tPA). However, tPA has a small therapeutic window for administration (3 - 6 hr), and is only effective in 4% of the patients who actually receive it. Current research focuses on understanding the pathophysiology of stroke in order to find potential therapeutic targets. Thus, reliable models are crucial, and the MCA occlusion (MCAo) model (also termed the intraluminal filament or suture model) is deemed to be the most clinically relevant surgical model of ischemic stroke, and is fairly non-invasive and easily reproducible. Typically the MCAo model is used with rodents, especially with mice due to all the genetic variations available for this species. Here we describe (and present in the video) how to successfully perform the MCAo model (with reperfusion) in mice to generate reliable and reproducible data.

In order to understand the pathophysiology of stroke, it is important to use reliable models. This paper will describe one of the most frequently used stroke models in mice, termed the middle cerebral artery occlusion (MCAo) model (also termed the intraluminal filament or suture model) with reperfusion.

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ...

Techniques of Sleeve Gastrectomy and Modified Roux-en-Y Gastric Bypass in Mice

Obesity is a major public health issue, with a prevalence of 4 to 28% for men and 6.2 to 36.5% for women in Europe (from 2003 to 2008). Morbid obesity is frequently associated with metabolic complications, such as type 2 diabetes, hypertension, and dyslipidemia, reducing life expectancy and quality. In the absence of any effective noninvasive treatments, bariatric surgery is a valuable therapeutic option for patients with morbid obesity (body mass index (BMI) &#62;40 kg/m2), leading to long-term, sustained weight loss and improvements in metabolic complications. However, the underlying cellular and molecular mechanisms sustaining the beneficial effects of bariatric surgery are not yet fully understood. Due to the numerous genetically-modified strains available, the mouse model is the most convenient animal model to explore the molecular mechanisms behind the pleiotropic beneficial effects of bariatric surgeries. Here, we detailed the optimized healthcare methods and surgical protocols in mice for the two most widely-used bariatric surgeries: the sleeve gastrectomy and the modified Roux-en-Y gastric bypass. Deciphering the molecular mechanisms underlying the therapeutic effects of bariatric surgeries offers the promise of identifying new therapeutics targets.

Bariatric surgery is the most efficient way to reduce body weight and the deadly metabolic complications (diabetes, obesity, and dyslipidemia) frequently associated with morbid obesity. Mouse models of bariatric surgery represent a unique asset for deciphering molecular mechanisms behind the beneficial effects of these surgeries on diabetes, hypertension, and dyslipidemia.

Obesity is a major public health issue, with a prevalence of 4 to 28% for men and 6.2 to 36.5% for women in Europe (from 2003 to 2008). Morbid obesity is ...

Using the Sleeve Technique in a Mouse Model of Aortic Transplantation - An Instructional Video

Orthotopic aortic transplantation using the sleeve technique reduces injury to the aorta with failure rate of only 10-20%. The time to anastomose the aorta in mice using the sleeve method was short and easy averaging 20 min, permitting studies of iso/allo grafts. The following article describes the aortic transplantation procedure used in our laboratory. The mice were anesthetized with a mixture of 1.5% volume isoflurane and 100% oxygen through a face mask. At this point, the segment of the aorta between the renal arteries and its bifurcation was separated from the vena cava, freely prepared and clampedat the proximal and distal segments with a single silk suture. Prior to the removal of the aorta, a saline solution containing heparin was injected into the inferior vena cava. Then the aorta was cut between the clamps and a saline heparin solution was used to flush the lumen. The sleeve technique with monofilament sutures was used in order to transplant the abdominal aorta in the orthotopic position.

We present an orthotopic aortic transplantation model using the sleeve technique in mice. It is a very rapid anastomosis method, which can be employed in studies of vascular disease.

Orthotopic aortic transplantation using the sleeve technique reduces injury to the aorta with failure rate of only 10-20%. The time to anastomose the ...

Intravital Microscopy of Tumor-associated Vasculature Using Advanced Dorsal Skinfold Window Chambers on Transgenic Fluorescent Mice

Tumor and tumor vessel development, as well as tumor response to therapeutics, are highly dynamic biological processes. Histology provides static information and is often not sufficient for a correct interpretation. Intravital evaluation, in which a process is followed in time, provides extra and often unexpected information. With the creation of transgenic animals expressing cell-specific markers and live cell tracers, improvements to imaging equipment, and the development of several imaging chambers, intravital microscopy has become an important tool to better understand biological processes. This paper describes an experimental design for the investigation of tumor vessel development and of therapeutic effects in a spatial and temporal manner. Using this setup, the stage of vessel development, tip cell and lumen formation, blood flow, extravasation, an established vascular bed, and vascular destruction can be visualized and followed. Furthermore, therapeutic effects, intratumoral fate, and the localization of chemotherapeutic compounds can also be followed.

This protocol describes the intravital imaging of transgenic mice expressing cell-specific fluorescent markers. Intravital imaging provides a non-invasive method for high-resolution observations in living animals using implantable windows through which the microscopic visualization of processes is possible. This method is especially useful to study longitudinal processes.

Tumor and tumor vessel development, as well as tumor response to therapeutics, are highly dynamic biological processes. Histology provides static ...

Using In Vitro Live-cell Imaging to Explore Chemotherapeutics Delivered by Lipid-based Nanoparticles

Conventional imaging techniques can provide detailed information about cellular processes. However, this information is based on static images in an otherwise dynamic system, and successive phases are easily overlooked or misinterpreted. Live-cell imaging and time-lapse microscopy, in which living cells can be followed for hours or even days in a more or less continuous fashion, are therefore very informative. The protocol described here allows for the investigation of the fate of chemotherapeutic nanoparticles after the delivery of doxorubicin (dox) in living cells. Dox is an intercalating agent that must be released from its nanocarrier to become biologically active. In spite of its clinical registration for more than two decades, its uptake, breakdown, and drug release are still not fully understood. This article explores the hypothesis that lipid-based nanoparticles are taken up by the tumor cells and are slowly degraded. Released dox is then translocated to the nucleus. To prevent fixation artifacts, live-cell imaging and time-lapse microscopy, described in this experimental procedure, can be applied.

Using In Vitro Live-cell Imaging to Explore Chemotherapeutics Delivered by Lipid-based Nanoparticles

Live-cell imaging is a powerful tool to visualize dynamic processes. The examination of fixed cells provides only static pictures, which can lead to misinterpretation and confusion about the process. This work presents a method to study uptake, drug release, and intracellular localization of liposomal nanoparticles in living cells.

Conventional imaging techniques can provide detailed information about cellular processes. However, this information is based on static images in an ...

Designing CAD/CAM Surgical Guides for Maxillary Reconstruction Using an In-house Approach

Computer-aided design/computer-assisted manufacturing (CAD/CAM) is now being evaluated as a preparative technique for maxillofacial surgery. Because this technique is expensive and available in only limited areas of the world, we developed a novel CAD/CAM surgical guide using an in-house approach. By using the CAD software, the maxillary resection area and cutting planes and the fibular cutting planes and angles are determined. Once the resection area is decided, the necessary faces are extracted using a Boolean modifier. These superficial faces are united to fit the surface of the bones and thickened to stabilize the solids. Not only the cutting guides for the fibula and maxilla but also the location arrangement of the transferred bone segments is defined by thickening the superficial faces. The CAD design is recorded as .stl files and three-dimensionally (3-D) printed as actual surgical guides. To check the accuracy of the guides, model surgery using 3-D-printed facial and fibular models is performed. These methods may be used to assist surgeons where commercial guides are not available.

Methods for designing a computer-aided design/computer-aided manufacturing (CAD/CAM) surgical guide are shown. Cutting planes are separated, united, and thickened to easily visualize the necessary bone transfer. These designs can be three-dimensional printed and checked for accuracy.

Computer-aided design/computer-assisted manufacturing (CAD/CAM) is now being evaluated as a preparative technique for maxillofacial surgery. Because this ...

Real-Time Electrocardiogram Monitoring During Treadmill Training in Mice

Regular physical exercise is a major contributor to cardiovascular health, influencing various metabolic as well as electrophysiological processes. However, in certain cardiac diseases such as inherited arrhythmia syndromes, e.g., arrhythmogenic cardiomyopathy (ACM) or myocarditis, physical exercise may have negative effects on the heart leading to a proarrhythmogenic substrate production. Currently, the underlying molecular mechanisms of exercise-related proarrhythmogenic remodeling are largely unknown, thus it remains unclear which frequency, duration, and intensity of exercise can be considered safe in the context of disease(s).
The proposed method allows to study proarrhythmic/antiarrhythmic effects of physical exercise by combining treadmill training with real-time monitoring of the ECG. Implantable telemetry devices are used to continuously record the ECG of freely moving mice over a period of up to 3 months both at rest and during treadmill training. Data acquisition software with its analysis modules is used to analyze basic ECG parameters such as heart rate, P wave duration, PR interval, QRS interval, or QT duration at rest, during and after training. Furthermore, heart rate variability (HRV) parameters and occurrence of arrhythmias are evaluated. In brief, this manuscript describes a step-by-step approach to experimentally explore exercise induced effects on cardiac electrophysiology, including potential proarrhythmogenic remodeling in mouse models.

Electrocardiogram (ECG) is the key variable to understanding cardiac electrophysiology. Physical exercise has beneficial effects but may also be harmful in the context of cardiovascular diseases. This manuscript provides a method of recording real-time ECG during exercise, which can serve to investigate its effects on cardiac electrophysiology in mice.

Regular physical exercise is a major contributor to cardiovascular health, influencing various metabolic as well as electrophysiological processes. ...

Methods Article

Sleeve Gastrectomy in Mice using Surgical Clips