Name: a technique for subcutaneous abdominal adipose tissue biopsy via a non-diathermy method
Uploaded: 2017-09-30T13:00:00
Description: Watch this Scientific Journal Video about A Technique for Subcutaneous Abdominal Adipose Tissue Biopsy via a Non-diathermy Method at JoVE.com

This work received funding from the European Union&#39;s Horizon 2020 (grant agreement no. 645710) and 7th Framework (grant agreements no. 612547 and 319010) Programmes. The&#160;authors wish to thank Dr. Fotini Sourli-Chasioti for the assistance with the biopsy procedure. The authors also thank Mr. Yiorgos Paterakis for his valuable help in the preparation of the video.

The study conformed to the standards set by the Declaration of Helsinki and was approved by the University of Thessaly, Department of Exercise Science, &#914;i&#959;ethics Committee.
1. Preparation of the Surgery Equipment and Consumables
<ol>
	<li>Position a disinfected surgical field on a 4-wheel surgical Mayo stand.</li>
	<li>Position the following consumables and surgical equipment on a 4-wheel surgical Mayo stand: 
	One pair of straight operating scissors (15 cm) 
	One scalpel...

<ol>
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C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ligasure+versus+diathermy+scissors+tonsillectomy:+A+controlled+randomized+study.">Ligasure versus diathermy scissors tonsillectomy: A controlled randomized study.</a> Acta Otolaryngol. 130, 1180-1184 (2010).</li><li>Ecker, T., Carvalho, A. L., Choe, J. H., Walosek, G., Preuss, K. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hemostasis+in+thyroid+surgery:+harmonic+scalpel+versus+other+techniques--a+meta-analysis.">Hemostasis in thyroid surgery: harmonic scalpel versus other techniques--a meta-analysis.</a> Otolaryngol Head Neck Surg. 143, 17-25 (2010).</li><li>Campbell, K. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+pilot+study+of+sampling+subcutaneous+adipose+tissue+to+examine+biomarkers+of+cancer+risk.">A pilot study of sampling subcutaneous adipose tissue to examine biomarkers of cancer risk.</a> Cancer Prev Res. 2, 37-42 (2009).</li><li>Jamurtas, A. Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+a+single+bout+of+exercise+on+resting+energy+expenditure+and+respiratory+exchange+ratio.">The effects of a single bout of exercise on resting energy expenditure and respiratory exchange ratio.</a> Eur J Appl Physiol. 92, 393-398 (2004).</li><li>Adriaens, P. E. M., Schoffelen, F. M. P., Westerterp, K. 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N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Infiltrative+anesthesia+in+office+practice.">Infiltrative anesthesia in office practice.</a> Am Fam Phys. 89, 956-962 (2014).</li><li>Jurevic, R., 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=Plasma+levels+of+2%+lidocaine+with+1:100,000+epinephrine+with+young+children+undergoing+dental+procedures.">Plasma levels of 2% lidocaine with 1:100,000 epinephrine with young children undergoing dental procedures.</a> Anesth Prog. 45, 87-90 (1998).</li><li>Kale, S. 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The aim of this study was to standardize a surgical technique for subcutaneous abdominal adipose tissue biopsy, performed under local anesthesia with a non-diathermy method in healthy men. Diathermy is commonly used in the operating room; however, possible side effects (i.e., flash fires and environmental toxicity)9 and delays in healing10 may occur. These unwanted effects of diathermy can generate health risks for both the operating staff and the patients/particip...

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide an insightful overview of mechanisms relating to metabolism and disease. For instance, they are used for breast cancer tumor identification1, fatty acid composition examination2, and research on important metabolic and hormonal disease pathways3. To obtain subcutaneous adipose tissue biopsies in the abdominal area, researchers and physicians use either a surgical or a needle-based technique. However, surgical subcutaneous fat biopsies can offer tissue samples that may provide a more comprehensive overview of complex biological mechanisms [e.g., messenger ribonucleic acid (mRNA) expression, histological analysis, etc.]4. They have previously been used in healthy adults4 and can be safely performed at the bedside5.
The needle-based biopsy technique requires intense liposuction that may destroy the integrity of the tissue, and it usually retrieves a small amount of tissue (100-500 mg)6. These tissue samples may not be appropriate for complex research studies. Furthermore, compared to the needle-based technique, surgical subcutaneous fat biopsies can offer tissue samples that may draw a more complete picture of several inflammatory indices (e.g., B- and T-cell signaling); cytoskeleton regulation; cellular pathways; metabolic pathways of lipids, carbohydrates and amino acids; and oxidative phosphorylation pathways4. Also, the needle-based technique includes a higher risk for blood cell contamination than the surgical one4, while providing little information about the fibrotic regions of the tissue4,7,8.
Usually, a surgical adipose tissue biopsy includes a diathermy treatment for cauterizing blood vessels to prevent excessive bleeding9. A diathermy is also used to burn and destroy tissues affected by neoplasms and warts or tissues that are infected by disease9. This technique is particularly applicable in neurosurgery and eye surgery9.
The excessive use of diathermy may lead to the formation of necrotic tissue, causing infections and delaying the healing process10. Indeed, side effects such as flash fires and skin lesions in the tissue have been reported following diathermy9. Additionally, concerns have been raised about the toxicity of surgical smoke released by diathermy into the surgery environment, which may be inhaled by the participant/patient and the operating staff9. Indeed, the use of diathermy may cause air toxicity in the operating room equal to that produced by smoking six cigarettes11 and, although operating rooms typically have good ventilation systems, the contamination can last for at least 20 min12. However, in patients undertaking a tonsillectomy surgery, the diathermy method has shown less, but non-significant, post-operative pain13 and bleeding14 than the classic ligation hemostasis method. Overall, evidence regarding differences in the post-operative pain and bleeding rates after adipose tissue biopsies in healthy individuals barely exists. Given the lack of evidence and the side effects of diathermy, standardization of a non-diathermy method for adipose tissue biopsy is warranted. Accordingly, the aim of this study was to standardize a surgical technique for subcutaneous abdominal adipose tissue biopsy, performed under local anesthesia using a non-diathermy method in healthy men. Moreover, we outline the procedures for tissue collection to determine mRNA expression and protein concentration and to perform histological analyses.

<table border="0" cellpadding="0" cellspacing="0" width="981">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:327px;">&#160;Disinfected surgical field&#160;</td>
			<td style="width:183px;">Hartmann</td>
			<td style="width:175px;">23516-00</td>
			<td style="width:296px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Stainless Steel Instrument Stands (surgical Mayo)</td>
			<td>Medical Device Depot Inc.</td>
			<td>32000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Operating Scissors straight 15 cm</td>
			<td>Karl Storz</td>
			<td>791903</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Scalpel No 11&#160;</td>
			<td>Swann-Morton</td>
			<td>203</td>
			<td>Bipolar, catalog number 429011</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Scarpel handle No 3</td>
			<td>Karl Storz</td>
			<td>488090</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Scissors curved 14 cm</td>
			<td>Karl Storz</td>
			<td>511514, 752918</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mosquito forceps</td>
			<td>Karl Storz</td>
			<td>535012</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tweezers Kocher&#160;</td>
			<td>Karl Storz</td>
			<td>530416</td>
			<td>Bipolar, catalog number 830316</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Surgical tweezers</td>
			<td>Karl Storz</td>
			<td>793216</td>
			<td>Bipolar, catalog number 831016</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Povidone-iodine&#160;</td>
			<td>Various</td>
			<td>Not applicable</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Suture 4.0 Vicryl</td>
			<td>Johnsons</td>
			<td>V4970H&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Scissors straight 11 cm</td>
			<td>Karl Storz</td>
			<td>512511 DS</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Needle holder 15 cm</td>
			<td>Karl Storz</td>
			<td>515515</td>
			<td>Bipolar, catalog number 213015</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">&#160;Sterile gauzes</td>
			<td>Hartmann</td>
			<td>3021</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">&#913;dhesive sterile gauze&#160;</td>
			<td>Digas medical equipment</td>
			<td>2889</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">10 mL syringe with disinfected needle</td>
			<td>Alpha medical solutions</td>
			<td>10310-55-0010</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">2%-xylocaine (no adrenaline)</td>
			<td>Various</td>
			<td>Not applicable</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sterile surgical gloves&#160;</td>
			<td>Sempermed</td>
			<td>10001</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Eppendorf tubes</td>
			<td>Kartell</td>
			<td>16283</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">10% formalin&#160;</td>
			<td>Alpha medical solutions</td>
			<td>11016-30-2909</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tubes 120 mL</td>
			<td>Digas medical equipment</td>
			<td>9025</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Liquid nitrogen at -190&#176;C</td>
			<td>Revival</td>
			<td>Not applicable</td>
			<td></td>
		</tr>
	</tbody>
</table>

a technique for subcutaneous abdominal adipose tissue biopsy via a non-diathermy method

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide insightful overviews of mechanisms relating to metabolism and disease. To obtain subcutaneous adipose tissue biopsies in the abdominal area, researchers and physicians use either a surgical or a needle-based technique. However, surgical subcutaneous fat biopsies can offer tissue samples that may provide a more comprehensive overview of the complexities of biological indices in white adipose tissue. Usually, a surgical adipose tissue biopsy includes a diathermy treatment for cauterizing blood vessels to prevent excessive bleeding. Nevertheless, side effects, such as flash fires and skin lesions in the tissue, have been reported after diathermy. Therefore, we aimed to standardize a surgical abdominal adipose tissue biopsy performed under local anesthesia using a non-diathermy method. We conducted 115 subcutaneous adipose tissue biopsies in healthy men using a non-diathermy abdominal surgical biopsy method. Our results showed three cases of excessive post-operation bleeding out of 115 operations (2.61%).In conclusion, our standardized subcutaneous abdominal adipose tissue surgical biopsy using a non-diathermy method can be safely applied to healthy men at the bedside, with minimal side effects.

The characteristics of the participants are provided in Table 1. The stitches in the area of the incision fell off within the next 8-12 days. We completed a total of 115 subcutaneous adipose tissue biopsies using the non-diathermy abdominal surgical biopsy method in healthy adult men (age: 35.2 &#177; 6.9 years; body mass index: 27.3 &#177; 4.4; and body fat percentage: 27.0 &#177; 9.0%). Post-biopsy excessive bleeding was observed in three cases (prevalence: 2.61%) two d...

Watch this Scientific Journal Video about A Technique for Subcutaneous Abdominal Adipose Tissue Biopsy via a Non-diathermy Method at JoVE.com

A Technique for Subcutaneous Abdominal Adipose Tissue Biopsy via a Non-diathermy Method

We standardized an abdominal adipose tissue biopsy using a non-diathermy method performed under local anesthesia. Three cases of excessive post-operation bleeding out of 115 operations (2.61%) occurred.We conclude that an abdominal adipose tissue surgical biopsy using a non-diathermy method can be safely applied to healthy men.

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide insightful overviews of mechanisms relating to metabolism and disease. To ...

Subcutaneous adipose tissue biopsies are very important for research and health because they can reveal mechanisms of adipose tissue related to the disease. In this paper we describe a technique for obtaining subcutaneous adipose tissue biopsies in the abdominal area. To obtain such samples, researchers and physicians use either a needle-based technique or a surgical-based technique.However, it is important to note that the surgical technique provides a more complete overview of the biological indices in white adipose tissue compared to the needle-based technique, which may lead to misdiagnosis. This is true, for example, for histological analysis, which is vital for both research and health purposes. Overall, it is important to note that the surgical technique has been performed in both men and women and can be safely performed in the bedside.The participant undertaking a surgical fat biopsy must undergo an eight hour fast and refrain from exercise, excessive stressors, alcohol, as well as active and passive smoking in the 72 hours prior to the biopsy procedure. These precautions are meant to minimize the risk for misleading results when the white adipose tissue sample is analyzed. We first prepare the materials that are needed for the adipose tissue biopsy procedure.On a wheeled surgical Mayo we position a sterile surgical sheet. The materials are placed on Mayo and are the following. One pair of operating scissors, straight, 15 centimeters.One scalpel, number 11. One scalpel handle. One pair of scissors, curved, 14 centimeters.One Mosquito forceps. One pair of tweezers Kocher. One surgical tweezers.Povidone iodine on a sterile gauze. One suture, 4.0. One pair of scissors, straight, 11 centimeters.One needle holder, 15 centimeters. Five sterile gauzes. One adhesive sterile gauze.One 10 milliliter syringe with a disinfected needle. 10 mL Xylocaine, 2%no adrenaline. And one pair of disinfected surgical gloves.On a separate bench we prepare the following materials that are needed for the tissue collection. Two Eppendorf tubes. Using a sterile needle, we open a small hole on the cap of each Eppendorf.This is because we want to avoid the cap failure when we immerge the Eppendorf in liquid nitrogen in minus 190 degrees Celsius. One tube in which we put five mL of formalin, 10%One small container with liquid nitrogen, in minus 190 degrees Celsius. We need this to immediately immerse the Eppendorf tubes containing the tissue to ensure tissue's integrity until final deposition.Following preparation of all materials and consumables, we position the patient on a surgical bed in a supine position. The surgeon's position is on the one side of the surgical bed. The surgeon first performs disinfection of the abdominal region using povidone iodine, 15 centimeters around the point of the incision.10 mL of Xylocaine, 2%no adrenaline, is then injected in the point of the incision for local anesthesia. A sterile surgical field is then positioned in the area. Once this is completed, we confirm that local anesthesia has been achieved.This requires approximately three to five minutes. Thereafter, the surgeon connects the scalpel number 11 with the scalpel handle and performs an incision of the skin and the subcutaneous tissue until the underlying adipose tissue is revealed. The incision is approximately two to two and a half centimeters, and it is performed three to five centimeters away from the navel.Thereafter, with a pair of operating scissors, straight, 15 centimeters, the surgeon removes the subcutaneous tissue. Once the adipose tissue is revealed, it is captured with a pair of tweezers and a pair of curved scissors, 14 centimeters. Then the adipose tissue is prepared and cut.The total amount of adipose tissue removed depends on the analysis that will be performed. Based on our experience, approximately one gram of adipose tissue will be enough for the purposes of most histological and gene expression analyses. Once the adipose tissue is removed, the surgeon places sterile gauzes on the incision area for approximately 30 seconds to ensure hemostasis.Once hemostasis is confirmed, the surgeon captures a 4-0 suture with a 14 centimeters needle and the skin with the surgical tweezers until subcutaneous tissue is visible. The suture needle enters the skin towards the subcutaneous tissue while the skin and subcutaneous are then brought together. The suture needle then follows and continues zig-zag technique into the subcutaneous tissue until being driven externally off the skin.This continues until the skin incision is closed. Finally, the suture tying is done externally. Usually the stitches fall off within eight to 12 days, and this whole technique reduces the possibility of a post-operation scar on the skin.The trauma is then cleaned with saline, and it is covered with adhesive sterile gauze. You may have noticed that we did not use the tweezers Mosquito. This instrument would be used in the case that the vessel is bleeding.If so, the vessel is captured with a tweezers Mosquito to ligate and close it until achieving hemostasis. Remember that for our purposes the sample that we collected weighed approximately one gram. The collected tissue is cut into three pieces and weighed 350 milligrams, 150 milligrams, and 500 milligrams.The 350 milligram sample is placed into an Eppendorf tube and will be used for protein levels analysis. The 150 milligram sample is placed in another Eppendorf tube and will be used for mRNA expression analysis. These two Eppendorf tubes are immediately immersed in a small container with liquid nitrogen at minus 190 degrees Celsius to ensure the integrity of the samples until final deposition in a minus 80 degrees freezer.Finally, the 500 milligram sample is immersed in a tube containing 5 mL of formalin, 10%and will be used for histological analysis. Our technique is based on a non-diathermy method. We perform hemostasis by using sterile gauzes in order to eliminate the risk for a skin lesion, such as a scar after the surgery.Overall, our non-diathermy method has insignificant side effects. Indeed, we performed in this study a total of 115 biopsies and we occurred only three excessive bleedings. These occurred in the second day following the biopsy procedure.We treated these individuals by reopening the trauma, draining, applying hemostasis, and placing surgical stapling. No further complications were observed and normal healing occurred in the following days. Finally, it's important to note that three months after each biopsy procedure for all subjects no side effects were observed, including skin scars.Our non-diathermy surgical biopsy technique showed minimal side effects, and it's proposed to be used when a complete overview of the biological indices of white adipose tissue is required.

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