Name: establishment of a simple and effective rat model for intraoperative parathyroid gland imaging
Uploaded: 2022-08-17T14:05:00
Description: Watch this Scientific Journal Video about Establishment of a Simple and Effective Rat Model for Intraoperative Parathyroid Gland Imaging at JoVE.com

This study was supported by the National Natural Science Foundation of China (NSFC) (82172598), the Natural Science Foundation of Zhejiang Province, China (LZ22H310001), the 551 Health Talent Training Project of Health Commission of Zhejiang Province, China, and the Medical and Health Science and Technology Project of Zhejiang Province, China (2021KY110).

All animal studies have been approved by the Institutional Animal Care and Use Committee (IACUC) of the Institute of Basic Medicine and Cancer, Chinese Academy of Sciences. This is a non-survival surgery.
1. Animal
<ol>
	<li>Use a 6-8-week-old female SD rat, weighing 250 g, for intraoperative PG imaging.</li>
</ol>
2. Anesthesia
<ol>
	<li>Turn on the anesthesia machine.</li>
	<li>Before beginning, ensure the isoflur...

<ol>
	<li>Cope, O., Donaldson, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relation+of+thyroid+and+parathyroid+glands+to+calcium+and+phosphorus+metabolism.+Study+of+a+case+with+coexistent+hypoparathyroidism+and+hyperthyroidism.">Relation of thyroid and parathyroid glands to calcium and phosphorus metabolism. Study of a case with coexistent hypoparathyroidism and hyperthyroidism.</a> The Journal of Clinical Investigation. 16 (3), 329-341 (1937).</li><li>Mansberger, A. R., Wei, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surgical+embryology+and+anatomy+of+the+thyroid+and+parathyroid+glands.">Surgical embryology and anatomy of the thyroid and parathyroid glands.</a> Surgical Clinics of North America. 73 (4), 727-746 (1993).</li><li>Koch, A., Hofbeck, M., Dorr, H. G., Singer, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hypocalcemia-induced+heart+failure+as+the+initial+symptom+of+hypoparathyroidism.">Hypocalcemia-induced heart failure as the initial symptom of hypoparathyroidism.</a> Zeitschrift f&#252;r Kardiologie. 88 (1), 10-13 (1999).</li><li>Shoback, D. 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=Presentation+of+hypoparathyroidism:+etiologies+and+clinical+features.">Presentation of hypoparathyroidism: etiologies and clinical features.</a> The Journal of Clinical Endocrinology &amp; Metabolism. 101 (6), 2300-2312 (2016).</li><li>Arneiro, A. J., 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=Self-report+of+psychological+symptoms+in+hypoparathyroidism+patients+on+conventional+therapy.">Self-report of psychological symptoms in hypoparathyroidism patients on conventional therapy.</a> Archives of Endocrinology Metabolism. 62 (3), 319-324 (2018).</li><li>Olson, E., Wintheiser, G., Wolfe, K. M., Droessler, J., Silberstein, 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=Epidemiology+of+thyroid+cancer:+a+review+of+the+national+cancer+database,+2000-2013.">Epidemiology of thyroid cancer: a review of the national cancer database, 2000-2013.</a> Cureus. 11 (2), 4127 (2019).</li><li>Du, 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=Epidemiology+of+thyroid+cancer:+incidence+and+mortality+in+China,+2015.">Epidemiology of thyroid cancer: incidence and mortality in China, 2015.</a> Frontiers in Oncology. 10, 1702 (2020).</li><li>Barrios, 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=Incidental+parathyroidectomy+in+thyroidectomy+and+central+neck+dissection.">Incidental parathyroidectomy in thyroidectomy and central neck dissection.</a> Surgery. 169 (5), 1145-1151 (2021).</li><li>Sitges-Serra, A., 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=Inadvertent+parathyroidectomy+during+total+thyroidectomy+and+central+neck+dissection+for+papillary+thyroid+carcinoma.">Inadvertent parathyroidectomy during total thyroidectomy and central neck dissection for papillary thyroid carcinoma.</a> Surgery. 161 (3), 712-719 (2017).</li><li>Sakorafas, G. 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=Incidental+parathyroidectomy+during+thyroid+surgery:+an+underappreciated+complication+of+thyroidectomy.">Incidental parathyroidectomy during thyroid surgery: an underappreciated complication of thyroidectomy.</a> World Journal of Surgery. 29 (12), 1539-1543 (2005).</li><li>Sahyouni, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rate+of+incidental+parathyroidectomy+in+a+pediatric+population.">Rate of incidental parathyroidectomy in a pediatric population.</a> OTO Open. 5 (4), (2021).</li><li>Erickson, A. 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=Incidence,+survival+time,+and+surgical+treatment+of+parathyroid+carcinomas+in+dogs:+100+cases+(2010-2019).">Incidence, survival time, and surgical treatment of parathyroid carcinomas in dogs: 100 cases (2010-2019).</a> Journal of the American Veterinary Medical Association. 259 (11), 1309-1317 (2021).</li><li>Bi, R., Fan, Y., Luo, E., Yuan, Q., Mannstadt, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+techniques+to+create+hypoparathyroid+mice:+parathyroidectomy+using+GFP+glands+and+diphtheria-toxin-mediated+parathyroid+ablation.">Two techniques to create hypoparathyroid mice: parathyroidectomy using GFP glands and diphtheria-toxin-mediated parathyroid ablation.</a> Journal of Visualized Experiments. (121), e55010 (2017).</li><li>Soulsby, S. N., Holland, M., Hudson, J. A., Behrend, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasonographic+evaluation+of+adrenal+gland+size+compared+to+body+weight+in+normal+dogs.">Ultrasonographic evaluation of adrenal gland size compared to body weight in normal dogs.</a> Veterinary Radiology &amp; Ultrasound. 56 (3), 317-326 (2015).</li><li>Zheng, W. 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=Biodegradable+iron+oxide+nanoparticles+for+intraoperative+parathyroid+gland+imaging+in+thyroidectomy.">Biodegradable iron oxide nanoparticles for intraoperative parathyroid gland imaging in thyroidectomy.</a> PNAS Nexus. 1 (3), 087 (2022).</li></ol>

We demonstrate an IONPs-guided negative imaging technique of rat PG&#160;using black IONPs, which were injected locally in the center of the thyroid gland and diffused within the thyroid gland but not the PG. It enables clear identification of the PG&#160;with naked eyes without the aid of any microscope. Although transgenic mice with green fluorescent protein selectively expressed in the PG&#160;have been reported13, the model described in this paper is more straightforward to perform. It only ta...

Parathyroid gland&#160;(PG) is small, oval-shaped endocrine glands located in the neck of humans and other vertebrates, which produce and secrete parathyroid hormones to regulate and balance calcium and phosphorus levels in the blood and in bones1. Humans usually have two pairs of PG&#160;located behind the thyroid gland lobes in variable locations; the size of human PG&#160;typically measures 6 mm x 4 mm x 2 mm, with a weight of approximately 35-40 mg2. Removal or damage of the PG&#160;causes hypoparathyroidism (HP), an endocrine disorder characterized by hypocalcemia and low or undetectable levels of parathyroid hormones, which cause a wide range of symptoms from cramp-like spasms to malformed teeth to chronic kidney diseases. Some of these complications are fatal (e.g., heart failure and seizure)3,4,5; thus, PG&#160;is essential in regulating the body&#39;s metabolism and sustaining life.
HP is one of the most common complications after anterior neck surgery, especially in thyroidectomy, a well-established curative treatment for thyroid cancer, which is the most common endocrine cancer worldwide6,7. Post-thyroidectomy HP is predominantly caused by direct trauma, ischemia, or removal of the PG in surgery because of a severe lack of ability to reliably discriminate the PG&#160;from thyroid gland lobes and other surrounding tissues (e.g., lymph nodes and peripheral fat particles) in real-time in the operation room. In 2021, Barrios et al. reported an average PG mis-section rate of 22.4% within 1,114 thyroidectomy cases, and even experienced surgeons who had a minimum error rate of 7.7%8. Such high PG mis-section rates are consistent with other similar reports9,10,11. Thus, incorrect parathyroidectomy is an independent risk factor for transient and permanent postoperative HP.
Developing effective intraoperative PG identification methods holds the key to addressing this critical unmet medical need; however, it has been severely limited by the lack of animal models in preclinical research. To date, most intraoperative PG identification investigations have been performed on human patients and large animals (e.g., dogs)12, which are expensive and difficult to receive ethical approval, expand subject numbers, and repeat tests. Meanwhile, the mouse, the most commonly used vertebrate model in biological research, has extremely small PG, with a size of less than 1 mm13. Due to this limitation, mouse PG models have seldom been used in intraoperative PG identification research.
This paper reports the establishment of a simple, straightforward, and effective rat model for intraoperative PG identification studies. We investigated the usage of native Sprague-Dawley (SD) rats without any surgical modifications or genetic engineering as a reliable animal model for testing a PG imaging contrast agent, IONPs, in a thyroidectomy surgery. This rat model demonstrates a highly similar physiological structure of PG&#160;and the surrounding microenvironment to that of humans, and the size of rat PG&#160;is large enough to be visually detected in comparison with those of mice. Most rats have one PG on each side of the thyroid gland. The simplicity and effectiveness of this rat model have been demonstrated by performing intraoperative IONP-enhanced PG imaging in thyroidectomy surgery.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>alcohol</td>
			<td>Li feng</td>
			<td>9400820067</td>
			<td></td>
		</tr>
		<tr>
			<td>anesthesia machine</td>
			<td>RWD Company</td>
			<td>R520IE</td>
			<td>Machine number</td>
		</tr>
		<tr>
			<td>blade</td>
			<td>Daopian</td>
			<td>TB-JZ-10#</td>
			<td></td>
		</tr>
		<tr>
			<td>cylindrical pillow</td>
			<td></td>
			<td></td>
			<td>made by ourselves</td>
		</tr>
		<tr>
			<td>depilatory cream</td>
			<td>Nair</td>
			<td>TMG-001</td>
			<td></td>
		</tr>
		<tr>
			<td>electronic scale</td>
			<td>Hong xingda</td>
			<td>CN-HXD2</td>
			<td></td>
		</tr>
		<tr>
			<td>eosin</td>
			<td>Thermo Fisher (Waltham, USA).</td>
			<td>C0105S-2</td>
			<td></td>
		</tr>
		<tr>
			<td>erythromycin</td>
			<td>Shuang ji (Beijing, China)</td>
			<td>200409</td>
			<td></td>
		</tr>
		<tr>
			<td>gauze</td>
			<td>Fulanns</td>
			<td>YY0331-2006</td>
			<td></td>
		</tr>
		<tr>
			<td>heating pad</td>
			<td>Johon (ShenZhen,China)</td>
			<td>JH-36-2006</td>
			<td></td>
		</tr>
		<tr>
			<td>hematoxylin</td>
			<td>Thermo Fisher (Waltham, USA).</td>
			<td>C0105S-1</td>
			<td></td>
		</tr>
		<tr>
			<td>insulin injection needle</td>
			<td>Jiangyin NanquanMacromolecule</td>
			<td>20170702</td>
			<td></td>
		</tr>
		<tr>
			<td>iodophor cotton ball</td>
			<td>HOYON</td>
			<td>19-6007</td>
			<td></td>
		</tr>
		<tr>
			<td>iron oxide nanoparticle solution</td>
			<td>Zhongke Leiming Technology (Beijing, China)</td>
			<td>Mag9110-05</td>
			<td></td>
		</tr>
		<tr>
			<td>isoflurane</td>
			<td>Sigma Aldrich (St Louis USA).</td>
			<td>21112801</td>
			<td></td>
		</tr>
		<tr>
			<td>needle holder</td>
			<td>Meijun</td>
			<td>MH0587</td>
			<td></td>
		</tr>
		<tr>
			<td>operation table</td>
			<td>BioJane</td>
			<td>BJ-P-M</td>
			<td></td>
		</tr>
		<tr>
			<td>paraformaldehyde solution</td>
			<td>Biosharp</td>
			<td>21269333</td>
			<td></td>
		</tr>
		<tr>
			<td>rubber</td>
			<td>G-CLONE</td>
			<td> 
			XT41050</td>
			<td></td>
		</tr>
		<tr>
			<td>scanning machine</td>
			<td>Olympus</td>
			<td>Slideview VS200</td>
			<td></td>
		</tr>
		<tr>
			<td>surgical forceps</td>
			<td>Suping</td>
			<td>SPHC-0676</td>
			<td></td>
		</tr>
		<tr>
			<td>surgical knife handle</td>
			<td>Aladdin</td>
			<td>S3052-06-1EA</td>
			<td></td>
		</tr>
		<tr>
			<td>surgical retractor</td>
			<td>TOCYTO</td>
			<td>18-4010</td>
			<td></td>
		</tr>
		<tr>
			<td>surgical scissors</td>
			<td>Suping</td>
			<td>SPHC-0795</td>
			<td></td>
		</tr>
		<tr>
			<td>surgical towel</td>
			<td>Along technology</td>
			<td>YCKJ-RJ-036205</td>
			<td></td>
		</tr>
		<tr>
			<td>suture</td>
			<td>Ethicon</td>
			<td>SA84G</td>
			<td></td>
		</tr>
		<tr>
			<td>suture with needle</td>
			<td>Jinhuan (Shanghai,China)</td>
			<td>F301</td>
			<td></td>
		</tr>
		<tr>
			<td>vascular forceps</td>
			<td>Along technology</td>
			<td>YCKJ-RJ-016218</td>
			<td></td>
		</tr>
		<tr>
			<td>Water Bath-Slide Drier</td>
			<td>Hua su (Jinhua, China)</td>
			<td>HS-1145</td>
			<td></td>
		</tr>
	</tbody>
</table>

establishment of a simple and effective rat model for intraoperative parathyroid gland imaging

Parathyroid gland (PG) identification is a critical unmet need in thyroidectomy. The identification of the PG is challenging in thyroid surgery as it is similar in color to the thyroid gland. The lack of effective animal models in preclinical research is a severe limitation for the development of PG identification techniques. This protocol allows for the establishment of a simple and effective rat model for PG identification. In this model, black iron oxide nanoparticles (IONPs) are injected locally in the thyroid gland and rapidly diffuse within the thyroid gland but not the PG. A negatively stained PG and a positively stained thyroid gland can be easily identified by the naked eye without requiring external microscopes. The position of the PG can be identified by increasing the color contrast between the thyroid gland and the PG, based on the color of the black IONPs. This rat model is low-cost and convenient for PG identification, and the IONPs are a novel PG contrast agent.

In this animal model, we surgically incised the neck of a SD rat to expose the trachea, larynx, and surrounding tissues. Then, the thyroid gland was visually located on both sides of the trachea; it is butterfly-shaped and approximately 3 mm x 5 mm in size. A pair of PG&#160;are usually located in the upper part of the thyroid gland, and their color is very similar to that of the thyroid gland lobes, making it extremely difficult to distinguish them with the naked eye (Figure 1).&#160;
<...

Watch this Scientific Journal Video about Establishment of a Simple and Effective Rat Model for Intraoperative Parathyroid Gland Imaging at JoVE.com

Establishment of a Simple and Effective Rat Model for Intraoperative Parathyroid Gland Imaging

To date, the development of parathyroid gland (PG) identification methods is limited by the lack of animal models in preclinical research. Here, we establish a simple and effective rat model for intraoperative PG imaging and evaluate its effectiveness by using iron oxide nanoparticles as a novel PG contrast agent.

Parathyroid gland (PG) identification is a critical unmet need in thyroidectomy. The identification of the PG is challenging in thyroid surgery as it is ...

Parathyroid gland is a small organ with very important functions. Our protocol can be used for developing new PG identification technologies which can help surgeons to protect it during the thyroidectomy for thyroid cancer. The main advantage of this model is that it can observe a clear difference between the thyroid and PGs through the negative imaging of IONP.In addition, the cost and operational difficulty are considerably lower for this rat model than for large animal models. To begin, place a prewarmed heating pad on the surgical table in order to sustain the animal's body temperature during the surgery, and then transfer the anesthetized rat down to a surgical drape placed on the operation table. Using rubber bands, fix the rat's limbs to the operation table and place a cylindrical pillow made of drape under the rat's shoulder to lean its head back, completely exposing the neck area, then apply depilatory cream to the neck area, up to the submandibular space, down to the xiphoid process, and on both sides to the outside of the sternocleidomastoid muscle.After three minutes, gently wipe the hair and depilatory cream with a tissue. Next, using an iodophor cotton ball, sterilize the middle of the neck and surrounding area. from which hair is removed.Cover the animal with a surgical drape, keeping the hole aligned with disinfected area. Using the scalpel, make approximately a five-centimeter longitudinal incision in the anterior midline of the rat's neck. Lift the skin along both sides of the incision and using scissors, longitudinally cut along the linea alba cervicalis.Use forceps to separate the sternohyoid and sternothyroid muscle. Clamp the separated muscles in front of the neck using vascular forceps and pull the clamped tissue outside. Using the needle, pass the suture through the clamped tissue.Make a knot and fix the suture to the surgical drape of the operation table. Locate the thyroid cartilage based on its shield shape and cricoid cartilage based on its ring shape as the upper boundary in the operation area, then locate the trachea based on its tubular cartilage ring shape in the front and middle of the neck as the lower boundary in the operation area. Locate the thyroid gland, a red butterfly-shaped gland on the opposite side of the trachea between the upper and lower boundaries.Next, locate two parathyroid glands in a fusiform shape that are reddish, but lighter than the surrounding thyroid gland with a certain boundary on the upper and outer sides of the thyroid gland. Take a frontal photograph of the parathyroid glands with the trachea, thyroid, and larynx. Dissect the back of the esophagus and using a retractor, expose the right side of the parathyroid glands.Take a right-side photograph of the parathyroid glands with the thyroid gland and trachea, and similarly expose and capture the left side of the glands. Use an insulin syringe to locally inject 10 microliters of black iron oxide nanoparticles suspension into the center of the thyroid gland, and gently press the injection site with gauze for five seconds, then observe the rapid diffusion of the black iron oxide nanoparticles within the thyroid glands, but not the parathyroid glands, as it negatively stains the parathyroid glands and differentiates them from the surrounding thyroid. Capture the front, left, and right-side photographs of the negatively stained parathyroid glands together with the trachea, thyroid gland, and larynx.After injection, the contrast agent, black iron oxide nanoparticles, readily diffuse within the thyroid gland and stain it black, but cannot infiltrate the parathyroid gland due to their high tissue density. Histological analysis of black iron oxide nanoparticles injected-thyroid gland revealed that the parathyroid glands are enriched with closely aligned chief cells, whereas the thyroid gland features many loose lumens indicating much lower tissue density. Two important things should be remembered in this procedure.First, the posture of the rat. We recommended to lean ears and head back, completely exposing the neck area. Second, when we inject the IONPs, we should be use fine-ensuring needle because the thyroid surface is full of blood vessels.This procedure can also be performed by autofluorescence imaging of the PG, which is non-invasive, provides real-time information, and does not use dye as a contrast agent, but the autofluorescence signal is weak and the imaging efficiency is also poor. Other researchers can use this simple and effective rat model to develop and evaluate new PG identification technologies and products instead of using more complicated and expensive large animals such as dogs and pigs.

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MRI-guided Disruption of the Blood-brain Barrier using Transcranial Focused Ultrasound in a Rat Model

Focused ultrasound (FUS) disruption of the blood-brain barrier (BBB) is an increasingly investigated technique for circumventing the BBB1-5. The BBB is a significant obstacle to pharmaceutical treatments of brain disorders as it limits the passage of molecules from the vasculature into the brain tissue to molecules less than approximately 500 Da in size6. FUS induced BBB disruption (BBBD) is temporary and reversible4 and has an advantage over chemical means of inducing BBBD by being highly localized. FUS induced BBBD provides a means for investigating the effects of a wide range of therapeutic agents on the brain, which would not otherwise be deliverable to the tissue in sufficient concentration. While a wide range of ultrasound parameters have proven successful at disrupting the BBB2,5,7, there are several critical steps in the experimental procedure to ensure successful disruption with accurate targeting. This protocol outlines how to achieve MRI-guided FUS induced BBBD in a rat model, with a focus on the critical animal preparation and microbubble handling steps of the experiment.

Microbubble-mediated focused ultrasound disruption of the blood-brain barrier (BBB) is a promising technique for non-invasive targeted drug delivery in the brain1-3. This protocol outlines the experimental procedure for MRI-guided transcranial BBB disruption in a rat model.

Focused ultrasound (FUS) disruption of the blood-brain barrier (BBB) is an increasingly investigated technique for circumventing the BBB1-5. The BBB is a ...

A Simple Critical-sized Femoral Defect Model in Mice

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all limb bone fractures fail to heal completely due to the extent of the trauma, disease, or age of the patient. Our ability to improve bone regenerative strategies is critically dependent on the ability to mimic serious bone trauma in test animals, but the generation and stabilization of large bone lesions is technically challenging. In most cases, serious long bone trauma is mimicked experimentally by establishing a defect that will not naturally heal. This is achieved by complete removal of a bone segment that is larger than 1.5 times the diameter of the bone cross-section. The bone is then stabilized with a metal implant to maintain proper orientation of the fracture edges and allow for mobility. 
Due to their small size and the fragility of their long bones, establishment of such lesions in mice are beyond the capabilities of most research groups. As such, long bone defect models are confined to rats and larger animals. Nevertheless, mice afford significant research advantages in that they can be genetically modified and bred as immune-compromised strains that do not reject human cells and tissue.
Herein, we demonstrate a technique that facilitates the generation of a segmental defect in mouse femora using standard laboratory and veterinary equipment. With practice, fabrication of the fixation device and surgical implantation is feasible for the majority of trained veterinarians and animal research personnel. Using example data, we also provide methodologies for the quantitative analysis of bone healing for the model.

Animal models are frequently employed to mimic serious bone injury in biomedical research. Due to their small size, establishment of stabilized bone lesions in mice are beyond the capabilities of most research groups. Herein, we describe a simple method for establishing and analyzing experimental femoral defects in mice.

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all ...

Establishment and Characterization of UTI and CAUTI in a Mouse Model

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

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

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

Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction

Normal tissues that lie within the portals of radiation are inadvertently damaged. Salivary glands are often injured during head and neck radiotherapy. Irreparable cell damage results in a chronic loss of salivary function that impairs basic oral activities, and increases the risk of oral infections and dental caries. Salivary hypofunction and its complications gravely impact a patient's comfort. Current symptomatic management of the condition is ineffective, and newer therapies to assuage the condition are needed. 
Salivary glands are exocrine glands, which expel their secretions into the mouth via excretory ducts. Cannulation of these ducts provides direct access to the glands. Retroductal delivery of a contrast agent to major salivary glands is a routine out-patient procedure for diagnostic imaging. Using a similar procedure, localized treatment of the glands is feasible. However, performing this technique in preclinical studies with small animals poses unique challenges. In this study we describe the technique of retroductal administration in rat submandibular glands, a procedure that was refined in Dr. Bruce Baum's laboratory (NIH)1, and lay out a procedure for local gland irradiation.

Salivary gland hypofunction, a major adverse effect of head and neck radiotherapy diminishes a patient's quality of life. The demonstration of efficacy of new therapies in animal models is a prerequisite before clinical transition. This protocol describes retroductal administration and local irradiation of rat submandibular glands.

Normal tissues that lie within the portals of radiation are inadvertently damaged. Salivary glands are often injured during head and neck radiotherapy. ...

3D Ultrasound Imaging: Fast and Cost-effective Morphometry of Musculoskeletal Tissue

The developmental goal of 3D ultrasound imaging (3DUS) is to engineer a modality to perform 3D morphological ultrasound analysis of human muscles. 3DUS images are constructed from calibrated freehand 2D B-mode ultrasound images, which are positioned into a voxel array. Ultrasound (US) imaging allows quantification of muscle size, fascicle length, and angle of pennation. These morphological variables are important determinants of muscle force and length range of force exertion. The presented protocol describes an approach to determine volume and fascicle length of m.&#160;vastus lateralis and m.&#160;gastrocnemius medialis. 3DUS facilitates standardization using 3D anatomical references. This approach provides a fast and cost-effective approach for quantifying 3D morphology in skeletal muscles. In healthcare and sports, information on the morphometry of muscles is very valuable in diagnostics and/or follow-up evaluations after treatment or training.

3D ultrasound imaging (3DUS) allows fast and cost-effective morphometry of musculoskeletal tissues. We present a protocol to measure muscle volume and fascicle length using 3DUS.

The developmental goal of 3D ultrasound imaging (3DUS) is to engineer a modality to perform 3D morphological ultrasound analysis of human muscles. 3DUS ...

Investigation of the Electrophysiological and Thermographic Safety Parameters of Surgical Energy Devices During Thyroid and Parathyroid Surgery in a Porcine Model

In thyroid and parathyroid surgery, surgical energy devices (SEDs) provide more efficient hemostasis than conventional clamp-and-tie hemostasis in areas with rich blood supply. However, when a SED is activated near the recurrent laryngeal nerve (RLN), the heat generated by the SED may injure the nerve irreversibly. To safely apply SEDs in thyroid/parathyroid surgery, this article introduces experimental porcine model studies to investigate the activation and cooling safety parameters of SEDs in standardized electrophysiological (EP) and thermographic (TG) procedures, respectively. In the EP safety parameter experiments, continuous intraoperative neuromonitoring (C-IONM) is applied to demonstrate the RLN function in real-time. The EP activation study evaluates the safe activation distance of SEDs; the EP cooling study evaluates the safe cooling time of SEDs. In the TG safety parameter experiment, a thermal imaging camera is used to record the temperature change after activating the SED. The TG activation study evaluates the lateral thermal spread distance after SED activation in a dry or humid environment and whether smoke and splashing are generated; the TG cooling study evaluates the cooling time. This will help establish the safety parameters of newly developed SEDs used in thyroid/parathyroid surgery and provide safety guidelines to avoid RLN injury and related complications.

The safe application of newly developed surgical energy devices in thyroid/parathyroid surgery attracts the attention of surgeons. Animal experimental models can avoid unnecessary trials and errors in human surgery. This report aims to demonstrate electrophysiological and thermographic methods to evaluate the safety parameters of SEDs in thyroid/parathyroid surgery.

In thyroid and parathyroid surgery, surgical energy devices (SEDs) provide more efficient hemostasis than conventional clamp-and-tie hemostasis in areas ...

A Simple and Effective Method to Consistently Isolate Mouse Cardiomyocytes

The need for reproducible yet technically simple methods yielding high-quality cardiomyocytes is essential for research in cardiac biology. Cellular and molecular functional experiments (e.g., contraction, electrophysiology, calcium cycling, etc.) on cardiomyocytes are the gold standard for establishing mechanism(s) of disease. The mouse is the species of choice for functional experiments and the described technique is specifically for the isolation of mouse cardiomyocytes. Previous methods requiring a Langendorff apparatus require high levels of training and precision for aortic cannulation, often resulting in ischemia. The field is shifting toward Langendorff-free isolation methods that are simple, are reproducible, and yield viable myocytes for physiological data acquisition and culture. These methods greatly diminish ischemia time compared to aortic cannulation and result in reliably obtained cardiomyocytes. Our adaptation to the Langendorff-free method includes an initial perfusion with ice-cold clearing solution, use of a stabilizing platform that ensures a steady needle during perfusion, and additional digestion steps to ensure reliably obtained cardiomyocytes for use in functional measurements and culture. This method is simple and quick to perform and requires little technical skill.

The gold standard in cardiology for cellular and molecular functional experiments are cardiomyocytes. This article describes adaptations to the non-Langendorff technique to isolate mouse cardiomyocytes.

The need for reproducible yet technically simple methods yielding high-quality cardiomyocytes is essential for research in cardiac biology. Cellular and ...

Orthopedic Robot-Assisted Femoral Neck System in the Treatment of Femoral Neck Fracture

Cannulated screw fixation is the main therapy for femoral neck fractures, especially in young patients. The traditional surgical procedure uses C-arm fluoroscopy to place the screw freehand and requires several guide wire adjustments, which increases the operation time and radiation exposure. Repeated drilling can also cause damage to the blood supply and bone quality of the femoral neck, which can be followed by complications such as screw loosening, nonunion, and femoral head necrosis. In order to make fixation more precise and reduce the incidence of complications, our team applied robot-assisted orthopedic surgery for screw placement using the femoral neck system to modify the traditional procedure. This protocol introduces how to import a patient's X-ray information into the system, how to perform screw path planning in software, and how the robotic arm assists in screw placement. Using this method, the surgeons can place the screw successfully the first time, improve the accuracy of the procedure, and avoid radiation exposure. The whole protocol includes the diagnosis of femoral neck fracture; the collection of intraoperative X-ray images; screw path planning in the software; precise placement of the screw under the assistance of the robotic arm by the surgeon; and verification of the implant placement.

This article introduces a method of robot-assisted orthopedic surgery for screw placement during the treatment of femoral neck fracture using the femoral neck system, which allows for more accurate screw placement, improved surgical efficiency, and fewer complications.

Cannulated screw fixation is the main therapy for femoral neck fractures, especially in young patients. The traditional surgical procedure uses C-arm ...

Development of an Effective Mice Model for Studying Oronasal Fistulas

Establishment of an Oronasal Fistula Mice Model

This study presents a method utilizing heated ophthalmologic cautery to develop a viable model for investigating oronasal fistulas. C57BL/6 mice were used to establish the oronasal fistula (ONF) model. To create the ONF, the mice were anesthetized, immobilized, and their hard palates were exposed. During the surgical procedure, a 2.0 x 1.5 mm full-thickness mucosal injury was induced in the midline of the hard palate using ophthalmologic cautery. It was crucial to control the size of the ONF and minimize bleeding in order to ensure the success of the experiment. Verification of the ONF model's effectiveness was conducted on the 7th-day post-operation, encompassing both anatomical and functional assessments. The presence of the nasal septum within the oral cavity and the outflow of sterile water from the nostrils upon injection into the oral cavity confirmed the successful establishment of the ONF model. The model demonstrated a practical and successful oronasal fistula, characterized by a low mortality rate, significant weight changes, and minimal variation in ONF size. Future studies may consider adopting this methodology to elucidate the mechanisms of palate wound healing and explore novel treatments for oronasal fistulas.

Author Spotlight: Investigating Wound Healing in Mice Models of Oronasal Fistulas 

This article outlines a step-by-step procedure for establishing a mice model with an oronasal fistula. The oronasal fistula was created by employing heated ophthalmologic cautery to damage the midline portion of the hard palate, resulting in the formation of an opening between the oral and nasal cavities.

This study presents a method utilizing heated ophthalmologic cautery to develop a viable model for investigating oronasal fistulas. C57BL/6 mice were used ...

A Murine Maxillary Orthodontic Model Guide to Bone Remodeling Analysis

The Establishment of a Murine Maxillary Orthodontic Model

Due to the lack of reproducible protocols for establishing a murine maxillary orthodontic model, we present a reliable and reproducible protocol to provide researchers with a feasible tool to analyze mechanical loading-associated bone remodeling. This study presents a detailed flowchart in addition to different types of schematic diagrams, operation photos, and videos. We performed this protocol on 11 adult wide-type C57/B6J mice and harvested samples on postoperative days 3, 8, and 14. The micro-CT and histopathological data have proven the success of tooth movements coupled with bone remodeling using this protocol. Furthermore, according to the micro-CT results on days 3, 8, and 14, we have divided bone modeling into three stages: preparation stage, bone resorption stage, and bone formation stage. These stages are expected to help researchers concerned with different stages to set sample collection time reasonably. This protocol can equip researchers with a tool to carry out regenerative analysis of bone remodeling.

Author Spotlight: Insights into an Efficient Murine Maxillary Orthodontic Model Protocol

Here we demonstrate step-by-step a manageable, orthodontic tooth movement protocol operated on a murine maxillary model. With the explicit explanation of each step and visual demonstration, researchers can master this model and apply it to their experimental needs with a few modifications.

Due to the lack of reproducible protocols for establishing a murine maxillary orthodontic model, we present a reliable and reproducible protocol to ...