The authors have no funding to declare.

NOTE: The University of Stirling NHS, Invasive, or Clinical Research Committee approved the biopsy procedure described below. All research studies using this procedure must be approved by the appropriate independent ethics committee. The biopsy taker must have completed formal training in the described technique in accordance with their institution&#39;s requirements. Typically, this involves observing a demonstration of the described adipose tissue biopsy technique by a board-certified physician, followed by supervised practice. Once the trainee has performed 10 practice adipose tissue biopsies on volunteer subjects under supervision, they will be examined by a board-certified physician to ensure good knowledge and practice of the procedure. The board-certified physician then provides the individual with a signed examination form.
1. Laboratory room preparation
<ol>
	<li>Ensure that the laboratory has an appropriately private room with clean, wipeable non-porous surfaces and a clean, comfortable (preferably non-porous) bed on which the participant may lie supine. Clean all required surfaces for the biopsy procedure using 70% ethanol spray and clean paper towels. Provide clean pillows or cushions to support the participant if required.</li>
	<li>Keep appropriate sharps disposal bins and biohazard waste bags within easy reach of the area where the biopsy is being performed and within easy reach of the person taking the biopsy.</li>
	<li>Prepare the equipment required for the procedure and set up on a freshly cleaned general medical trolley prior to the participant arriving to the laboratory (Figure 1). For a complete list of consumables required, see the Table of Materials.</li>
</ol>
2. Participant preparation
<ol>
	<li>Ensure that all participants provide written informed consent prior to undergoing the procedure in accordance with protocols required by the institution&#39;s independent ethics committee. Additionally, ask the participants to complete a written questionnaire to ensure they are not allergic to any materials used in the procedure (namely, nickel, chromium, local anesthetic, iodine, shellfish, and plasters).</li>
	<li>Confirm the identity of the participant. Ensure the participant understands the procedure to be carried out and potential secondary effects, including bruising, pain, and infection (Table 1). Gather verbal consent in addition to previously obtained written informed consent.</li>
	<li>Describe to the participant how the procedure will be carried out, with emphasis on how the administration of the anesthetic and biopsy itself will feel. Ensure that the participant is comfortable with proceeding. 
	NOTE: Local subcutaneous anesthetic will produce a stinging sensation, similar to a bee sting of short duration. Many participants report the anesthetic administration as the most uncomfortable part of the technique. Once the anesthetic has taken effect, the participant should feel no more than a slight tugging sensation during the biopsy.</li>
	<li>Ensure that the participant has no allergies to the local anesthetic (specifically from the amino-amide type, if using lidocaine or similar), certain metals (nickel and chromium), and shellfish (if using iodine-based solutions). Additionally, ensure that the participants are not taking any form of anticoagulant medication.</li>
	<li>Provide the participant with an opportunity to go and empty their bladder if required, to ensure they do not have to interrupt the procedure or experience undue discomfort in step 4.1.</li>
</ol>
3. Biopsy procedure - instructions for the biopsy taker
<ol>
	<li>Once the participant is lying in a supine position, identify the biopsy site approximately 5-10 cm lateral to the umbilicus. 
	NOTE: If the participant is to undergo multiple biopsies on the same day, identify biopsy sites on opposing sides of the umbilicus for each biopsy. This will ensure maximal distance between each biopsy site.</li>
	<li>Wash hands with soap and warm water according to standard medical guidelines4.</li>
	<li>Place the sterile sheet on the cleaned trolley or work area, taking care to only touch the outer edges of the sheet.</li>
	<li>Put on sterile surgical gloves using proper aseptic technique. Have the assistant open the rest of the equipment in such a way that it drops onto the prepared sterile sheet without touching/contaminating the equipment. Ensure that the assistant takes care not to touch items when removing tools from their sterile wrappings.</li>
	<li>Instruct the assistant to dispense a small amount of iodine-based solution on some sterile gauze (without oversaturating the gauze) on the work surface.</li>
	<li>Sterilize approximately 5-10 cm2 around the chosen biopsy site using the sterile gauze and iodine-based solution. Ensure the skin is cleaned in a spiraling motion moving outward from the proposed biopsy site. Repeat the skin cleaning procedure twice. Remove excess liquid (e.g., running off sterile area) by wiping with fresh sterile gauze.</li>
	<li>Along with the assistant, verbally confirm the content of the local anesthetic vial (2% lidocaine in this protocol) and that this is within its expiry date. Instruct the assistant to hold the opened vial upside down and draw 5 mL of local anesthetic into a syringe, using a 21 G needle. Dispose of the needle into the sharps bin, and ensure the syringe is free of air bubbles.</li>
	<li>Apply a 26 G needle to the syringe and expel any air bubbles. Gently pinch the abdominal skin and adipose tissue, moving it away from the abdominal wall. Then, insert the needle horizontally into the subcutaneous tissue at an angle no greater than 10&#176; relative to the surface of the skin.
	<ol>
		<li>Withdraw the syringe&#39;s plunger an additional 0.5 mL (to ensure the needle is not in a blood vessel). If blood appears in the syringe, withdraw and reinsert the needle at a different angle.</li>
		<li>Raise a bleb of 2-4 mm diameter to anesthetize the insertion area.</li>
		<li>Advance the needle into the subcutaneous tissue and administer ~1 mL of lidocaine in a fan-shaped pattern (Figure 2), taking care to withdraw the plunger each time before injecting the anesthetic.</li>
		<li>Remove and dispose of the 26 G needle, apply a 21 G needle to the syringe, expel any air bubbles, and administer the remaining ~4 mL of lidocaine in a fan-shaped pattern (Figure 2), taking care to withdraw the plunger each time before injecting the anesthetic.</li>
	</ol>
	</li>
	<li>Wait approximately 5 min for the local anesthetic to take effect. Use a sterile scalpel to gently prod the biopsy area to i) ensure the local anesthetic has taken effect and ii) identify the boundaries of the anesthetized area. If necessary, wait an additional minute or two and reassess.</li>
	<li>Once satisfied that the local anesthetic is working, gently pinch the skin and adipose tissue (as in step 3.8) and use a sterile scalpel to make a small 1-2 mm puncture in the skin. 
	NOTE: This only needs to be large enough to ease the entry of the 14 G needle and must be small enough that no suture is required to close it. It is common for some bleeding to occur from this point onwards, which can be controlled with a piece of sterile gauze.</li>
	<li>First, apply a 14 G needle to a 5 or 10 mL syringe. Then, while gently pinching the skin and adipose tissue, gradually insert the needle through the puncture into the adipose tissue approximately centrally in the anesthetized area and at an angle no greater than 10&#176; relative to the surface of the skin. 
	NOTE: For all cases of needle advancement in step 3.11, a syringe angle of no greater than 10&#176; must be maintained.
	<ol>
		<li>Apply suction by withdrawing the plunger to approximately the 2.5 mL mark. Take the biopsy by moving the needle in a quick backwards and forwards motion to slice fragments of adipose tissue. After approximately 30 s, twist the needle and syringe through 90&#176; and repeat this procedure to break up the fragments of adipose tissue, which are then aspirated into the syringe by the suction. 
		&#8203;NOTE: Other syringe sizes can be used. It is essential that the researcher selects a syringe size that permits both a good grip on the syringe and to comfortably maintain plunger retraction for maintenance of the vacuum. Locking syringes are available that maintain the vacuum, which can improve needle control and reduce perceived difficulty for the biopsy taker 5.</li>
		<li>After approximately 45-60 s of step 3.11.1, remove the needle and empty the syringe content onto a layer of gauze covering a weighing boat. Ensure that the lumen of the needle is facing down to avoid potential blood spatter.</li>
		<li>Repeat steps 3.11.1 and 3.11.2 for a maximum of 3 times. Check that the participant is content to proceed before each repeat of the above procedure.</li>
		<li>Whilst performing steps 3.11.1 and 3.11.2, instruct the assistant to process and prepare the samples for analysis/storage (see section 5).</li>
	</ol>
	</li>
</ol>
4. Post-biopsy procedure
<ol>
	<li>Once a satisfactory sample (i.e., ~200 mg) of adipose tissue has been obtained, place 1-2 layers of sterile gauze over the puncture wound, then place an ice pack over these, and apply firm pressure for approximately 10 min to induce hemostasis.</li>
	<li>When hemostasis has occurred, wipe away any iodine-based solution/dried blood with sterile gauze, and apply an adhesive wound dressing with absorbent pad to the site. Check that the participant feels well and provide verbal and written instructions on biopsy site aftercare.
	<ol>
		<li>Emphasize that the participants will likely exhibit some bruising for the next few days. Inform them that this may be substantial, although it is minimized by the ice pack in step 4.1 and will resolve without lasting effects.</li>
		<li>Recommend that should the participants feel any discomfort/pain once the anesthetic has worn off, they should take analgesics such as paracetamol following the instructions on the packet but refrain from taking analgesics that have anticoagulant activities (e.g., ibuprofen or aspirin).</li>
		<li>Explain that swelling, redness, or discharge from the biopsy site are indications of infection. In the unlikely event that these signs or symptoms occur, instruct the participant to urgently seek medical advice from a doctor or local Accident &#38; Emergency unit. Inform the participant that if they seek medical advice, they must also notify the research team. 
		NOTE: As research staff, neither the biopsy taker nor the assistant can provide medical advice or treatment; however, it is important that the research team are aware of and record all instances of complications resulting from the biopsy procedure.</li>
		<li>Recommend that participants should avoid swimming or overly strenuous activity for 48 h until the site of incision has closed.</li>
	</ol>
	</li>
	<li>Clear away any used sharps and contaminated materials into designated sharps and/or clinical waste containers.</li>
	<li>Clean all surfaces used in the biopsy procedure using 70% ethanol spray and clean paper towels. Place disposable and non-disposable items of bedding in appropriate clinical bags for disposal or cleaning, respectively.</li>
</ol>
5. Sample processing - instructions for the assistant
<ol>
	<li>Use sterile tweezers and 0.9% saline to rinse the adipose tissue sample to remove visible contaminants (i.e., blood, vasculature). Then, weigh the adipose tissue samples using digital scales. Split the tissue into appropriately sized pieces for downstream analysis and place them into appropriate storing tubes using sterile tweezers. Immerse the tubes containing the adipose tissue biopsies in liquid nitrogen at -190 &#176;C to flash-freeze until the samples are stored at -80 &#176;C. 
	NOTE: The assistant must complete sample processing as quickly as possible, typically within 3 min of sample aspiration, to minimize potential sample degradation.</li>
</ol>

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M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aseptic+technique:+evidence-based+approach+for+patient+safety.">Aseptic technique: evidence-based approach for patient safety.</a> British Journal of Nursing. 14 (10), 540-546 (2005).</li><li>Handiyani, H., Meily Kurniawidjaja, L., Irawaty, D., Damayanti, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effective+needle+stick+injury+prevention+strategies+for+nursing+students+in+the+clinical+settings:+a+literature+review.">The effective needle stick injury prevention strategies for nursing students in the clinical settings: a literature review.</a> Enfermeria Clinica. 28, 167-171 (2018).</li><li>Raggio, B. S., Barton, B. M., Kandil, E., Friedlander, P. 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(127), e55593 (2017).</li><li>Civelek, 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=Genetic+regulation+of+human+adipose+microRNA+expression+and+its+consequences+for+metabolic+traits.">Genetic regulation of human adipose microRNA expression and its consequences for metabolic traits.</a> Human Molecular Genetics. 22 (15), 3023-3037 (2013).</li><li>Chen, Y. C., 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=Feeding+influences+adipose+tissue+responses+to+exercise+in+overweight+men.">Feeding influences adipose tissue responses to exercise in overweight men.</a> American Journal of Physiology Endocrinology and Metabolism. 313 (1), 84-93 (2017).</li><li>Mendham, A. 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The authors have no conflicts of interest to declare.

The described protocol and associated video provide a step-by-step overview of a mini-liposuction technique to obtain subcutaneous adipose tissue samples from the abdominal area. This research group has performed a total of 124 biopsies over the course of 19 months with no adverse effects in participants. The procedure is safe and associated with minimum risk to participants or the biopsy team, provided that the described safety measures are followed. Aseptic technique (including opening and dispensing of sterile equipme...

Adipose tissue can provide useful information on the metabolic function of humans. Human subcutaneous adipose tissue is readily accessible. A technique for subcutaneous adipose tissue extraction was first described in the mid-80s1; since then, the initial protocol has been improved to increase the yield and improve study participant tolerability. Subcutaneous adipose tissue can be obtained from numerous sites, most commonly from the glutei1 and abdominal area2. Samples from the latter may be more desirable as they provide more valuable information in metabolic disease-related contexts3.
Subcutaneous adipose tissue biopsy using the mini-liposuction method can be safely and efficiently performed in a non-clinical setting. Following appropriate training by a board-certified physician and using strict aseptic technique, researchers can routinely perform these biopsies with minimal risk to both participant and investigators. The biopsy team must consist of at least 2 individuals: the person who will perform the biopsy and an assistant.
The person responsible for the biopsy is tasked with confirming the participant&#39;s identity, checking the participant can safely undergo the procedure (see protocol steps 2.1-2.4 below), ensuring the participant is comfortable throughout the procedure, ensuring sterile technique is maintained throughout the procedure, carrying out the procedure, and providing the participant with verbal and written after-care procedures. The assistant&#39;s role is to handle and rapidly process the adipose tissue obtained for later analysis and/or storage. The assistant also helps by being the &#34;non-sterile hands&#34; and ensuring the participant is at ease throughout the procedure. The purpose of this video and paper is to describe the step-by-step biopsy procedure to safely obtain subcutaneous adipose tissue from the abdominal area.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Item: 14 G needle 
			14 G x 3 1/8&#34; 210 mm x 80 mm</td>
			<td>B Braun</td>
			<td>4665473</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>Item: 21 G needle 
			21 G x 1 1/2&#34; 0.8 mm x 38 mm</td>
			<td>Terumo</td>
			<td>AN*2138R1</td>
			<td>Per biopsy: 2</td>
		</tr>
		<tr>
			<td>Item: 26 G&#160; needle 
			26 G x 1/2&#34; microlance needle 0.45 mm x 13 mm</td>
			<td>BD</td>
			<td>303800</td>
			<td>Per biopsy: 2</td>
		</tr>
		<tr>
			<td>Item: 5 mL syringe 
			5 mL luer</td>
			<td>DB plastipak</td>
			<td>302187</td>
			<td>Per biopsy: 2</td>
		</tr>
		<tr>
			<td>Item: Adhesive wound dressing 
			Opsite Post-Op Dressing 9.5 x 8.5</td>
			<td>Smith &#38; Nephew</td>
			<td>6600709</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>Item: Disposable sterile scalpel 
			Disposable Scalpel Sterile Blade no. 10</td>
			<td>Swann Morton</td>
			<td>/0501</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>Item: Icepack 
			BlueDot Reusable Hot/Cold Pack 26.5 cm x 13.0 cm</td>
			<td>NuCare</td>
			<td>F711</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>Item: Iodine based antiseptic 
			Videne antiseptic solution</td>
			<td>Ecolab Videne</td>
			<td>3030440</td>
			<td>Per biopsy: q.s</td>
		</tr>
		<tr>
			<td>Item: Lidocaine 2% w/o epinephrine 
			Lidocaine 2% injection 5 mL</td>
			<td>B Braun</td>
			<td>3558553</td>
			<td>Per biopsy: 5 mL</td>
		</tr>
		<tr>
			<td>Item: Non-sterile gloves 
			Starguard sensitive powder free nitrile gloves</td>
			<td>Starguard</td>
			<td>SG-N-S</td>
			<td>Per biopsy: pair</td>
		</tr>
		<tr>
			<td>Item: Sodium chloride 0.9% 
			Sodium chlride 0.9% w/v intravenous infusion BP</td>
			<td>BBraun</td>
			<td>S8004-5384</td>
			<td>Per biopsy: q.s.</td>
		</tr>
		<tr>
			<td>Item: Stabilization solution* 
			RNAlater Stabilization Solution</td>
			<td>ThermoFisher Scientific</td>
			<td>AM7020</td>
			<td>Per biopsy: q.s</td>
		</tr>
		<tr>
			<td>Item: Sterile forceps 
			Sterile forceps</td>
			<td>Rocialle</td>
			<td>RML109-006</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>Item: Sterile gauze swabs 
			Non woven swabs sterile 7.5 x 7.5 cm</td>
			<td>Prestige</td>
			<td>1860</td>
			<td>Per biopsy: 5</td>
		</tr>
		<tr>
			<td>Item: Sterile gloves 
			Prestige soft vinyl sterile powder free medical gloves</td>
			<td>Prestige</td>
			<td>S: P4301 
			M:P3302 
			L:P3301</td>
			<td>Per biopsy: pair</td>
		</tr>
		<tr>
			<td>Item: Sterile Microcentrifuge tubes 
			1.5 mL Sterile Microcentrifuge Tubes</td>
			<td>StarLab</td>
			<td>I1415-5510</td>
			<td>Per biopsy: q.s</td>
		</tr>
		<tr>
			<td>Item: Sterile sheet 
			Paper plain white 90 x 90 cm</td>
			<td>Rocialle</td>
			<td>RML 126-216</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>Item: Weighing boat 
			Diamond shape weigh boats</td>
			<td>Heathrow Scientific</td>
			<td>HS1427C</td>
			<td>Per biopsy: 1</td>
		</tr>
		<tr>
			<td>* denotes optional materials</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

human subcutaneous adipose tissue sampling using a mini-liposuction technique

Studies on adipose tissue are useful in understanding metabolic and other conditions. Human subcutaneous adipose tissue is accessible. With appropriate training and strict adherence to aseptic technique, subcutaneous adipose samples can be safely and efficiently obtained in a non-clinical setting by researchers. Following the administration of local anesthetic lateral to the umbilicus, a 14 G needle attached to a 5 or 10 mL syringe is inserted through the skin into the subcutaneous tissue. Under suction, the syringe is moved in a reciprocating, slicing motion to isolate fragments of adipose tissue. Withdrawing the plunger is enough to ensure that adipose tissue fragments are aspirated through the needle into the syringe. A single biopsy can collect about 200 mg of tissue. This biopsy technique is very safe for both participants and research staff. Following the biopsy, participants can resume most everyday activities, although they should avoid swimming and overly strenuous activities for 48 h to avoid excessive bleeding. Participants can safely undergo 2 biopsies within a single day, meaning that the technique can be applied in before-after acute intervention studies.

The described adipose tissue biopsy procedure is an efficient and low-risk technique for researchers to obtain subcutaneous adipose tissue samples from human volunteers. We performed 39 subcutaneous adipose tissue biopsies using the described procedure in 11 healthy, normal weight females (age, 27.4 &#177; 3.3 years; body mass index (BMI), 22.6 &#177; 1.5 kg.m2). All participants attended the laboratory between 07:00 and 10:00 following an 8-12 h fasting period. Sample yield using this adipose tissue biopsy pr...

Watch this Scientific Journal Video about Human Subcutaneous Adipose Tissue Sampling Using a Mini-Liposuction Technique at JoVE.com

Human Subcutaneous Adipose Tissue Sampling Using a Mini-Liposuction Technique

The manuscript and associated video demonstrate a percutaneous biopsy technique to obtain samples of subcutaneous adipose tissue from areas surrounding the umbilicus. This method is a low-risk and efficient way to investigate a range of parameters (e.g., gene or protein expression, enzyme activity, lipid content) within adipose tissue.

Studies on adipose tissue are useful in understanding metabolic and other conditions. Human subcutaneous adipose tissue is accessible. With appropriate ...

human-subcutaneous-adipose-tissue-sampling-using-mini-liposuction

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JoVE Journal

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2.8K Views.  University of Stirling. The manuscript and associated video demonstrate a percutaneous biopsy technique to obtain samples of subcutaneous adipose tissue from areas surrounding the umbilicus. This method is a low-risk and efficient way to investigate a range of parameters (e.g., gene or protein expression, enzyme activity, lipid content) within adipose tissue.

Video: Human Subcutaneous Adipose Tissue Sampling Using a Mini-Liposuction Technique

Cerenkov Luminescence Imaging of Interscapular Brown Adipose Tissue

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely related to metabolism and energy expenditure, and its dysfunction is one important contributor for obesity and diabetes. Contrary to previous belief, recent PET/CT imaging studies indicated the BAT depots are still present in human adults. PET imaging clearly shows that BAT has considerably high uptake of 18F-FDG under certain conditions. In this video report, we demonstrate that Cerenkov luminescence imaging (CLI) with 18F-FDG can be used to optically image BAT in small animals. BAT activation is observed after intraperitoneal injection of norepinephrine (NE) and cold treatment, and depression of BAT is induced by long anesthesia. Using multiple-filter Cerenkov luminescence imaging, spectral unmixing and 3D imaging reconstruction are demonstrated. Our results suggest that CLI with 18F-FDG is a practical technique for imaging BAT in small animals, and this technique can be used as a cheap, fast, and alternative imaging tool for BAT research.

In this video report, we show the application of Cerenkov Luminescence Imaging (CLI) for interscapular brown adipose tissue in mice under activated and depressed conditions.

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely ...

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

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

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

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

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular weight metabolites in tissues of the treated host to achieve long-term survival. The semipermeable membrane allows engrafted encapsulated cells to avoid rejection by the immune system. The encapsulation procedure was designed to enable a controlled release of bioactive compounds, such as insulin, other hormones, and cytokines. Here we describe a method for encapsulation of catabolic cells, which consume lipids for heat production and energy dissipation (thermogenesis) in the intra-abdominal adipose tissue of obese mice. Encapsulation of thermogenic catabolic cells may be potentially applicable to the prevention and treatment of obesity and type 2 diabetes. Another potential application of catabolic cells may include detoxification from alcohols or other toxic metabolites and environmental pollutants.

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular ...

Working with Human Tissues for Translational Cancer Research

Medical research for human benefit is greatly impeded by the necessity for human tissues and subjects. However, upon obtaining consent for human specimens, precious samples must be handled with the greatest care in order to ensure integrity of organs, tissues, and cells to the highest degree. Unfortunately, tissue processing by definition requires extraction of tissues from the host, a change which can cause great cellular stress and have major repercussions on subsequent analyses. These stresses could result in the specimen being no longer representative of the site from which it was retrieved. Therefore, a strict protocol must be adhered to while processing these specimens to ensure representativeness. The desired assay(s) must also be taken into consideration in order to ensure that an optimal technique is used for sample processing. Outlined here is a protocol for tissue retrieval, processing and various analyses which may be performed on processed tissue in order to maximize downstream production from limited tissue samples.

Translational cancer research is dependent on extraction of human tissues. Much work has gone into optimizing extraction methods for ex vivo analysis. Here, we describe tissue processing methods allowing for maximal data output from limited samples.

Medical research for human benefit is greatly impeded by the necessity for human tissues and subjects. However, upon obtaining consent for human ...

Subcutaneous Angiotensin II Infusion using Osmotic Pumps Induces Aortic Aneurysms in Mice

Osmotic pumps continuously deliver compounds at a constant rate into small animals. This article introduces a standard protocol used to induce aortic aneurysms via subcutaneous infusion of angiotensin II (AngII) from implanted osmotic pumps. This protocol includes calculation of AngII amount and dissolution, osmotic pump filling, implantation of osmotic pumps subcutaneously, observation after pump implantation, and harvest of aortas to visualize aortic aneurysms in mice. Subcutaneous infusion of AngII through osmotic pumps following this protocol is a reliable and reproducible technique to induce both abdominal and thoracic aortic aneurysms in mice. Infusion durations range from a few days to several months based on the purpose of the study. AngII 1,000 ng/kg/min is sufficient to provide maximal effects on abdominal aortic aneurysmal formation in male hypercholesterolemic mouse models such as apolipoprotein E deficient or low-density lipoprotein receptor deficient mice. Incidence of abdominal aortic aneurysms induced by AngII infusion via osmotic pumps is 5 - 10 times lower in female hypercholesterolemic mice and also lower in both genders of normocholesterolemic mice. In contrast, AngII-induced thoracic aortic aneurysms in mice are not hypercholesterolemia or gender-dependent. Importantly, multiple features of this mouse model recapitulate those of human aortic aneurysms.

Subcutaneous implantation of osmotic pumps provides a convenient approach for prolonged and consistent delivery of compounds. This approach has been used extensively to study both abdominal and thoracic aortic aneurysms in mice.

Osmotic pumps continuously deliver compounds at a constant rate into small animals.  This article introduces a standard protocol used to induce aortic ...

Using a Laminating Technique to Perform Confocal Microscopy of the Human Sclera

The sclera is a dense connective tissue that covers and protects the eye. It mainly consists of dense collagen bundles (types I, III, IV, V, VI, and VII). Due to its autofluorescence, opaqueness, and thickness, it has not been found suitable for confocal microscopy. An alternative approach to the one presented here, which uses formalin-fixed sclera embedded in paraffin for immunohistochemistry, has technical challenges, especially when preheating the tissue for antigen retrieval. Since the sclera is relatively poor in both cells and vessels, the use of larger tissue samples was explored to help prevent overlooking cells and to understand their localization in relation to vessels and other anatomical sites. To allow for the analysis of larger tissue samples under the confocal microscope, a laminating technique was performed to create thin layers from the sclera. Following the analysis of results of CD31 blood vessels and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1) positive cells, for which approval for scientific examination was obtained, the advantages and limitations of this method are discussed.

Human sclera tissue is mainly collagen; therefore, it is not easily usable for immunohistochemistry. To achieve the goal of performing immunohistochemistry for confocal microscopy of scleral tissue, a laminating technique was used.


The sclera is a dense connective tissue that covers and protects the eye. It mainly consists of dense collagen bundles (types I, III, IV, V, VI, and ...

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.

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 ...

Assessment of Human Adipose Tissue Microvascular Function Using Videomicroscopy

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. It is hypothesized that pro-atherogenic mediators released from fat tissues particularly in association with central/visceral adiposity may promote pathogenic vascular changes locally and systemically, and the notion that cardiovascular disease may be the consequence of adipose tissue dysfunction continues to evolve. Here, we describe a unique method of videomicroscopy that involves analysis of vasodilator and vasoconstrictor responses of intact small human arterioles removed from the adipose depot of living human subjects. Videomicroscopy is used to examine functional properties of isolated microvessels in response to pharmacological or physiological stimuli using a pressured system that mimics in vivo conditions. The technique is a useful approach to gain understanding of the pathophysiology and molecular mechanisms that contribute to vascular dysfunction locally within the adipose tissue milieu. Moreover, abnormalities in the adipose tissue microvasculature have also been linked with systemic diseases. We applied this technique to examine depot-specific vascular responses in obese subjects. We assessed endothelium-dependent vasodilation to both increased flow and acetylcholine in adipose arterioles (50 - 350 &#181;m internal diameter, 2 - 3 mm in length) isolated from two different adipose depots during bariatric surgery from the same individual. We demonstrated that arterioles from visceral fat exhibit impaired endothelium-dependent vasodilation compared to vessels isolated from the subcutaneous depot. The findings suggest that the visceral microenvironment is associated with vascular endothelial dysfunction which may be relevant to clinical observation linking increased visceral adiposity to systemic disease mechanisms. The videomicroscopy technique can be used to examine vascular phenotypes from different fat depots as well as compare findings across individuals with different degrees of obesity and metabolic dysfunction. The method can also be used to examine vascular responses longitudinally in response to clinical interventions.

Videomicroscopy systems are used to examine functional properties of isolated adipose tissue arterioles in response to physiological and pharmacological stimuli. This technique can be used to examine microvascular phenotypes in different adipose tissue domains in obese humans.

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. ...

Characterization of Immune Cells in Human Adipose Tissue by Using Flow Cytometry

Infiltration of immune cells in the subcutaneous and visceral adipose tissue (AT) deposits leads to a low-grade inflammation contributing to the development of obesity-associated complications such as type 2 diabetes. To quantitatively and qualitatively investigate the immune cell subsets in human AT deposits, we have developed a flow cytometry approach. The stromal vascular fraction (SVF), containing the immune cells, is isolated from subcutaneous and visceral AT biopsies by collagenase digestion. Adipocytes are removed after centrifugation. The SVF cells are stained for multiple membrane-bound markers selected to differentiate between immune cell subsets and analyzed using flow cytometry. As a result of this approach, pro- and anti-inflammatory macrophage subsets, dendritic cells (DCs), B-cells, CD4+ and CD8+ T-cells, and NK cells can be detected and quantified. This method gives detailed information about immune cells in AT and the amount of each specific subset. Since there are numerous fluorescent antibodies available, our flow cytometry approach can be adjusted to measure various other cellular and intracellular markers of interest.

This article describes a method to analyze immune cell content of adipose tissue by isolation of immune cells from adipose tissue and subsequent analysis using flow cytometry.

Infiltration of immune cells in the subcutaneous and visceral adipose tissue (AT) deposits leads to a low-grade inflammation contributing to the ...

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to expend energy has made it a potential target for novel therapies to ameliorate obesity and associated metabolic disorders in humans. Though this tissue has been well studied in small animals, BAT&#39;s thermogenic capacity in humans remains largely unknown due to the difficulties of measuring its volume, activity, and distribution. Identifying and quantifying active human BAT is commonly performed using 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography and computed tomography (PET/CT) scans following cold-exposure or pharmacological activation. Here we describe a detailed image-analysis approach to quantify total-body human BAT from 18F-FDG PET/CT scans using an open-source software. We demonstrate the drawing of user-specified regions of interest to identify metabolically active adipose tissue while avoiding common non-BAT tissues, to measure BAT volume and activity, and to further characterize its anatomical distribution. Although this rigorous approach is time-consuming, we believe it will ultimately provide a foundation to develop future automated BAT quantification algorithms.

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

Using free, open-source software, we have developed an analytical approach to quantify total and regional brown adipose tissue (BAT) volume and metabolic activity of BAT using 18F-FDG PET/CT.

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to ...