Name: positron emission tomography imaging for in vivo measuring of myelin content in the lysolecithin rat model of multiple sclerosis
Uploaded: 2021-02-28T14:30:00
Description: Watch this Scientific Journal Video about Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis at JoVE.com

&#946;-cube equipment (Molecubes NV, Belgium) was supported by the S&#227;o Paulo Research Foundation, FAPESP - Brazil (#2018/15167-1). LES has a PhD student scholarship from FAPESP - Brazil (#2019/15654-2).

All procedures were conducted in accordance with the guidelines of the National Council for the control of Animal Experimentation (CONCEA, Brazil) and were approved by the Ethics Committee for Animal Research of the Medical School of the University of Sao Paulo (CEUA-FMUSP, Brazil - protocol number: 25/15).
NOTE: In this protocol, we show how to induce a lysolecithin rat model of multiple sclerosis and how to acquire and analyze the myelin PET images.
1. Lysolecithin ...

<ol>
	<li>Oh, J., Vidal-Jordana, A., Montalban, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple+sclerosis:+clinical+aspects.">Multiple sclerosis: clinical aspects.</a> Current Opinion in Neurology. 31 (6), 752-759 (2018).</li><li>Sand, I. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Classification,+diagnosis,+and+differential+diagnosis+of+multiple+sclerosis.">Classification, diagnosis, and differential diagnosis of multiple sclerosis.</a> Current Opinion in Neurology. 28 (3), 193-205 (2015).</li><li>Thompson, 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=Diagnosis+of+multiple+sclerosis:+2017+revisions+of+the+McDonald+criteria.">Diagnosis of multiple sclerosis: 2017 revisions of the McDonald criteria.</a> Lancet Neurology. 17 (2), 162-173 (2018).</li><li>Veronese, 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=Quantification+of+C-11+PIB+PET+for+imaging+myelin+in+the+human+brain:+a+test-retest+reproducibility+study+in+high-resolution+research+tomography.">Quantification of C-11 PIB PET for imaging myelin in the human brain: a test-retest reproducibility study in high-resolution research tomography.</a> Journal of Cerebral Blood Flow and Metabolism. 35 (11), 1771-1782 (2015).</li><li>Carvalho, R. H. F., 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=C-11+PIB+PET+imaging+can+detect+white+and+grey+matter+demyelination+in+a+non-human+primate+model+of+progressive+multiple+sclerosis.">C-11 PIB PET imaging can detect white and grey matter demyelination in a non-human primate model of progressive multiple sclerosis.</a> Multiple Sclerosis and Related Disorders. 35, 108-115 (2019).</li><li>Stankoff, B., 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=Imaging+central+nervous+system+myelin+by+positron+emission+tomography+in+multiple+sclerosis+using+[methyl-(1)(1)C]-2-(4'-methylaminophenyl)-+6-hydroxybenzothiazole.">Imaging central nervous system myelin by positron emission tomography in multiple sclerosis using [methyl-(1)(1)C]-2-(4'-methylaminophenyl)- 6-hydroxybenzothiazole.</a> Annals of Neurology. 69 (4), 673-680 (2011).</li><li>Faria, D. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myelin+positron+emission+tomography+(PET)+imaging+in+multiple+sclerosis.">Myelin positron emission tomography (PET) imaging in multiple sclerosis.</a> Neural Regeneration Research. 15 (10), 1842-1843 (2020).</li><li>Pietroboni, A. 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=Amyloid+PET+as+a+marker+of+normal-appearing+white+matter+early+damage+in+multiple+sclerosis:+correlation+with+CSF+-amyloid+levels+and+brain+volumes.">Amyloid PET as a marker of normal-appearing white matter early damage in multiple sclerosis: correlation with CSF -amyloid levels and brain volumes.</a> European Journal of Nuclear Medicine and Molecular Imaging. 46 (2), 280-287 (2019).</li><li>Pytel, V., 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=Amyloid+PET+findings+in+multiple+sclerosis+are+associated+with+cognitive+decline+at+18+months.">Amyloid PET findings in multiple sclerosis are associated with cognitive decline at 18 months.</a> Multiple Sclerosis and Related Disorders. 39, (2020).</li><li>Faria, D. d. 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P., 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=PET+imaging+of+focal+demyelination+and+remyelination+in+a+rat+model+of+multiple+sclerosis+comparison+of+[C-11]MeDAS,+[C-11]CIC+and+[C-11]PIB.">PET imaging of focal demyelination and remyelination in a rat model of multiple sclerosis comparison of [C-11]MeDAS, [C-11]CIC and [C-11]PIB.</a> European Journal of Nuclear Medicine and Molecular Imaging. 41 (5), 995-1003 (2014).</li><li>Faria, D. d. P., 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=PET+imaging+of+focal+demyelination+and+remyelination+in+a+rat+model+of+multiple+sclerosis:+comparison+of+[11C]MeDAS,+[11C]CIC+and+[11C]PIB.">PET imaging of focal demyelination and remyelination in a rat model of multiple sclerosis: comparison of [11C]MeDAS, [11C]CIC and [11C]PIB.</a> European Journal of Nuclear Medicine and Molecular Imaging. 41 (5), 995-1003 (2014).</li><li>vander Star, B. 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=In+Vitro+and+In+Vivo+Models+of+Multiple+Sclerosis.">In Vitro and In Vivo Models of Multiple Sclerosis.</a> CNS &amp; Neurological Disorders-Drug Targets. 11 (5), 570-588 (2012).</li><li>Najm, F. 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=Drug-based+modulation+of+endogenous+stem+cells+promotes+functional+remyelination+in+vivo.">Drug-based modulation of endogenous stem cells promotes functional remyelination in vivo.</a> Nature. 522 (7555), 216 (2015).</li></ol>

The biggest advantage of using the lysolecithin model to study multiple sclerosis is the fast timeline for demyelination (about 1 week) and remyelination (about 4 weeks) to occur14. This model can also be induced in mice15, however, induction in rats is more advantageous for in vivo PET imaging due to the larger size of the rat brain compared to mice.
The first step of the induction model is to be extremely cautious. This model was valid...

An erratum was issued for:&#160;<a href="https://www.jove.com/t/62094">Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis</a>. The citation&#160;was updated.
The citation was updated from:
de Paula Faria, D., Cristiano Real, C., Estessi de Souza, L., Teles Garcez, A., Navarro Marques, F. L., Buchpiguel, C. A. Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis. J. Vis. Exp. (168), e62094, doi:10.3791/62094 (2021).
to:
de Paula Faria, D., Real, C.C., Estessi de Souza, L., Teles Garcez, A., Navarro Marques, F. L., Buchpiguel, C. A. Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis. J. Vis. Exp. (168), e62094, doi:10.3791/62094 (2021).

An erratum was issued for:&#160;<a href="https://www.jove.com/t/62094">Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis</a>. The citation&#160;was updated.

Erratum: Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis

Multiple sclerosis (MS) is a neuroinflammatory disease that affects the central nervous system, characterized by inflammation, demyelination, and axonal loss1. The prognosis of this disease is variable even with advances in treatment, and it is one of the most common causes of neurological deficits in young people1. The diagnosis of MS is based on the criteria of clinical manifestation and visualization of characteristic lesions by magnetic resonance imaging (MRI)2,3.
Positron emission tomography (PET) can be a useful tool for in vivo monitoring of MS progression and therapeutic effects. The Pittsburgh compound B radiotracer (PIB) labeled with carbon-11 (11C-PIB) is widely used to quantify &#946;-amyloid plaques; however, in the last decade, it has been studied to quantify myelin content and show dynamic demyelination and remyelination4,5,6.
Different amyloid PET tracers (11C-PIB, 18F-florbetaben,18F-florbetapir, 18F-flutemetamol) can be used to quantify myelin and provide important information about disease progression and therapeutic response, allowing identification of demyelination and remyelination processes, without the interference of neuroinflammation, which can occur with conventional magnetic resonance images (MRI)7. Amyloid PET imaging showed decreased tracer uptake in active MS patients compared to non-active patients which could be explained by early white matter damage in the active patients8. Lower amyloid tracer uptake was also associated with cognitive decline in a follow-up study, showing this technique to be a valuable tool for studying the pathophysiology of the disease and clinical outcomes9.
The lysolecithin (LPC) rat model is a chemical induced model of multiple sclerosis, where the injected toxin, LPC, induces a high response of macrophages that results in increased inflammation and, consequently, demyelination10,11. The demyelination is rapidly reversed, in approximately 4 weeks, which makes this a good model for evaluating demyelination and remyelination processes in rodents. This model has already been evaluated using PET imaging, with good results and correlation with post-mortem essays12.
Here we present the protocol for myelin PET imaging with 11C-PIB in the lysolecithin rat model, showing this imaging technique to be a useful tool for in vivo measurement of myelin content.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Analytical Balance</td>
			<td>Marte</td>
			<td>AUWZZOD</td>
			<td>max: 220 g- min: 1 mg</td>
		</tr>
		<tr>
			<td>Anestesia vaporizer</td>
			<td>Nanitech</td>
			<td>15800</td>
			<td></td>
		</tr>
		<tr>
			<td>Beta-cube</td>
			<td>Molecubes</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Bulldog clamp</td>
			<td>Stoelting</td>
			<td>5212043P</td>
			<td></td>
		</tr>
		<tr>
			<td>clorexidine</td>
			<td>Rioquimica</td>
			<td></td>
			<td>0.5%/100 mL</td>
		</tr>
		<tr>
			<td>Cotton swabs</td>
			<td>johnson e johnson</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dose calibrator</td>
			<td>Capintech</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Drill</td>
			<td>Kinzo powertools</td>
			<td>352901</td>
			<td>Model Q0M-DC3C</td>
		</tr>
		<tr>
			<td>Eppendorf tube</td>
			<td>Eppendorf</td>
			<td>30125150</td>
			<td>1.5 mL</td>
		</tr>
		<tr>
			<td>Eye lubricant</td>
			<td>ADVFARMA</td>
			<td>30049099</td>
			<td>&#160;vaseline 15 g (pharmaceutical purity)</td>
		</tr>
		<tr>
			<td>Fine forceps</td>
			<td>Stoelting</td>
			<td>52102-38P</td>
			<td></td>
		</tr>
		<tr>
			<td>Gloves</td>
			<td>Descarpack</td>
			<td>212101</td>
			<td>&#160;6.5 size</td>
		</tr>
		<tr>
			<td>Heating pad</td>
			<td>Softhear</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Injection Syringe</td>
			<td>Hamilton</td>
			<td>80314</td>
			<td>10&#181;, 32ga, model 701</td>
		</tr>
		<tr>
			<td>Insuline syringe</td>
			<td>BD</td>
			<td>328328</td>
			<td>1 mL insulin syringes with needle</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Crist&#225;lia</td>
			<td>410525</td>
			<td>100 mL , concentration 1 mL/1 mL</td>
		</tr>
		<tr>
			<td>Ketoprofen or other analgesic</td>
			<td>Sanofi</td>
			<td></td>
			<td>100 mg/2 mL</td>
		</tr>
		<tr>
			<td>lidocaine</td>
			<td>Hipolabor</td>
			<td>1.1343.0102.001-5</td>
			<td>2%/20mL</td>
		</tr>
		<tr>
			<td>L-&#945;-Lysophosphatidylcholine from egg yolk</td>
			<td>Sigma-aldrich</td>
			<td>L-4129</td>
			<td>25 mg - &#8805;99%, Type I, powder</td>
		</tr>
		<tr>
			<td>Needle holder</td>
			<td>Stoelting</td>
			<td>5212290P</td>
			<td></td>
		</tr>
		<tr>
			<td>Oxygen</td>
			<td>White Martins</td>
			<td>7782-44-7</td>
			<td>Compressed gas</td>
		</tr>
		<tr>
			<td>PMOD software</td>
			<td>PMOD technologies</td>
			<td>Version 4.1</td>
			<td>module fuse it</td>
		</tr>
		<tr>
			<td>Rat anesthesia mask</td>
			<td>KOPF</td>
			<td>Model 906</td>
			<td></td>
		</tr>
		<tr>
			<td>Saline</td>
			<td>Farmace</td>
			<td>0543325/ 14-8</td>
			<td>0.9% sodium chloride for injection, 10 mL</td>
		</tr>
		<tr>
			<td>Scapel blades</td>
			<td>Stoelting</td>
			<td>52173-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Scapel handles</td>
			<td>Stoelting</td>
			<td>52171P</td>
			<td></td>
		</tr>
		<tr>
			<td>Scissor</td>
			<td>Stoelting</td>
			<td>52136-50P</td>
			<td></td>
		</tr>
		<tr>
			<td>Semi-analytical Balance</td>
			<td>Quimis</td>
			<td>BK-3000</td>
			<td>max:3,100 g; min:0.2 g</td>
		</tr>
		<tr>
			<td>shaver</td>
			<td>Mega profissional</td>
			<td></td>
			<td>AT200 model</td>
		</tr>
		<tr>
			<td>Stereotactic Apparatus</td>
			<td>KOPF</td>
			<td>Nodel 900</td>
			<td></td>
		</tr>
		<tr>
			<td>Universal holder with needle support</td>
			<td>KOPF</td>
			<td>Model 1772-F1</td>
			<td>Hamilton support for 5 and 10 &#181;L</td>
		</tr>
	</tbody>
</table>

positron emission tomography imaging for in vivo measuring of myelin content in the lysolecithin rat model of multiple sclerosis

Multiple sclerosis (MS) is a neuroinflammatory disease with expanding axonal and neuronal degeneration and demyelination in the central nervous system, leading to motor dysfunctions, psychical disability, and cognitive impairment during MS progression. Positron emission tomography (PET) is an imaging technique able to quantify in vivo cellular and molecular alterations.
Radiotracers with affinity to intact myelin can be used for in vivo imaging of myelin content changes over time. It is possible to detect either an increase or decrease in myelin content, what means this imaging technique can detect demyelination and remyelination processes of the central nervous system. In this protocol we demonstrate how to use PET imaging to detect myelin changes in the lysolecithin rat model, which is a model of focal demyelination lesion (induced by stereotactic injection) (i.e., a model of multiple sclerosis disease). 11C-PIB PET imaging was performed at baseline, and 1 week and 4 weeks after stereotaxic injection of lysolecithin 1% in the right striatum (4 &#181;L) and corpus callosum (3 &#181;L) of the rat brain, allowing quantification of focal demyelination (injection site after 1 week) and the remyelination process (injection site at 4 weeks).
Myelin PET imaging is an interesting tool for monitoring in vivo changes in myelin content which could be useful for monitoring demyelinating disease progression and therapeutic response.

Figure 1&#160;shows illustrative 11C-PIB PET images with myelin changes over time. In the baseline scan, no differences can be seen in myelin content (i.e., no demyelination is present). In the 1-week time-point image, it is possible to see the focal demyelinated lesion (in the right hemisphere) as indicated by the white arrow. Images are presented in the 3 anatomical planes (coronal, axial, and sagittal) and it is possible to identify the demyelinated lesion in all of them. The 1...

Watch this Scientific Journal Video about Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis at JoVE.com

Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis

This protocol has the aim of monitoring in vivo myelin changes (demyelination and remyelination) by positron emission tomography (PET) imaging in an animal model of multiple sclerosis.

Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis

Multiple sclerosis (MS) is a neuroinflammatory disease with expanding axonal and neuronal degeneration and demyelination in the central nervous system, ...

This protocol has the name of monitoring in-vivo myelin chains by PET imaging with PIB labeled with carbonyl in the lysolecithin rat model of multiple sclerosis. This imaging technique has the ability of allowing in-vivo images of positions in different points. Getting point touch of the data of mining chains over time.In this video, We show how to induce the animal model by stereotactic injection and how to acquire and analyze. The mining PIP images. Take out needed material for the lysolecithin solution preparation Take the lysolecithin solution weight in an electronic balance using a glass tube take the glass tube out from the electric balance open the cell line and take the right volume for prepare 1%lysolecithin solution.Put the cell line in the plastic tube close the plastic tube and shake it using your fingers. Prepare for the surgery with the animal in the right position inject Put the eye cream the animal eyes are protecting against the hydration. use a 5%codoxygen solution for cleaning the incision area of the surgery inject lidocaine where the incision will be made.Shave the hair in the heads of their skull make an incision of about two centimeters expose the scope era with udak clamps clean area with a swab and mark the bregma position the hemato syringe on the bregma injection will be made in this three marked areas through in the right coordinate. Take the hemato needle through the drute area Go to the ventrical coordinate for start the injection, Finishing the surgery, Suture the incision. Use an appropriate place for the tracer injection.The tracer must be injected intravenously, and it can be done either, In the penile vein or in the tail vein. Image acquisition of baseline time point is performed before stereotactic surgery. And the other two time points one week and four weeks after the stereotactic surgery.Open the PET scanner software and chose the field of view to be scanned during image acquisition. Position the animal in the bed. Be sure that the animal is straight.Take the eye cream to protect the animal eyes. Cover the animal bed. Push the animal bed inside the pet scanner and start acquisition.Check the animal griefing until the end of the PET scan. When one day image acquisition is finished. Take animal out of the bed.Open remote software and choose Fuse It'model select image bio at and open with operations. Use a cylinder orientation and load the image. Go to matching and open the PET image.Adjust color scale to make the brain visible. Press crop'to crop the image and adjust the window size to cover the whole brain. It's parameters are on the bottom right side of the screen press blue shift'Be sure that the animal is positioned right in the software.Select the correct MRI template. if you need it, you can adjust the coverage restriction between PET and MRI template. Press device'button.Select the right brain template. Then you can look at different brain regions available and you can select or unselect all of them. Go to outline guides, and they will be all selected.In the top button, you can select any parameter you want to appear in this statistic data. You can touch the statistic button and then you can choose between KilobeckerCSS or SUV. Copy to clipboard and close.Your data can be pasted in an Excel sheet. For quantifying the and delusion Go to the new board, select these few and apply. You can create a new void in the contralateral side.And then go to the statistic button S2B4. It is possible to see the lesion one week after injection and the lesions covered four weeks after injection. Significant difference are seen between cell lining one week and four.Following this protocol, you'll get the three steps of taking the region of interest, being the injection site and that the principle area to be analyzed. As a large student of protocol, thus taking lesion presents the mining chains over time, where the mining nation is represented by decreased trace uptake and reimagination trace uptake. It's important to emphasize that the laser listing injection must be very slow, avoiding tissue damaging and causing only mining chains which will be detect by peep PIP imaging.

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