Differential Nanoparticle Accumulation Kinetics in a Mouse Model of Atherosclerotic Plaque Phenotypes
Thomas Ripperda
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04/05/2021
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UCARE research presentation video for the academic year 2020-2021. The research consisted of using nanoparticles as a diagnostic method to determine the advancement of atherosclerotic plaque and the plaque phenotype.
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- [00:00:01.160]Hello, my name is Thomas Ripperda,
- [00:00:03.270]and my UCARE project was titled
- [00:00:05.657]"Differential Nanoparticle Accumulation Kinetics
- [00:00:07.917]"in a Mouse Model of Atherosclerotic Plaque Phenotypes."
- [00:00:11.290]I worked on this project with Ian McCue,
- [00:00:15.070]a graduate student, Morgan Schake,
- [00:00:16.640]and under Dr. Ryan Pedrigi,
- [00:00:18.570]in the Mechanical and Materials Engineering Department.
- [00:00:21.810]Atherosclerosis is a leading cause
- [00:00:23.350]of death in the Western world.
- [00:00:25.000]A major reason that it's a leading cause of death
- [00:00:26.760]is that advanced atherosclerosis
- [00:00:29.230]can cause symptoms such as myocardial infarctions
- [00:00:32.130]and ischemic strokes.
- [00:00:35.210]Previous research has shown
- [00:00:36.566]that atherosclerosis plaque tends to form
- [00:00:41.470]at disturbed blood flow regions in the artery.
- [00:00:44.020]This can be seen in the picture shown on the right,
- [00:00:46.510]on the left, my bad.
- [00:00:49.470]Disturbed blood flow regions are shown in red
- [00:00:51.620]and the laminar flow region is shown in blue.
- [00:00:57.160]The research question that we focused on
- [00:00:58.700]was to test the hypothesis
- [00:00:59.980]that nanoparticles will accumulate
- [00:01:01.410]in advance plaque phenotypes.
- [00:01:03.830]We will also focus on
- [00:01:04.780]whether smooth muscle cells accumulate more
- [00:01:06.690]in one phenotype than the other.
- [00:01:10.080]For this study, we utilized ApoE-negative knockout mice.
- [00:01:14.400]They were immediately fed a high-fat diet
- [00:01:16.070]that lasted the remainder of the experiment upon arrival.
- [00:01:19.230]At two weeks after arrival,
- [00:01:21.100]a blood modifying cuff, shown in section C,
- [00:01:24.170]was placed on the left carotid artery.
- [00:01:25.870]The cuff has a 500 micrometer inlet
- [00:01:28.270]and a 250 micrometer outlet diameter.
- [00:01:31.620]The geometry of the,
- [00:01:34.510]the blood modifying cuff
- [00:01:35.560]creates a different flow environment,
- [00:01:37.190]and thus different plaque phenotypes
- [00:01:40.410]in the low wall shear stress region,
- [00:01:42.150]as seen in section B, which is upstream of the cuff,
- [00:01:44.950]and multidirectional wall shear stress region,
- [00:01:47.480]seen in section D,
- [00:01:48.540]which is downstream with the cuff.
- [00:01:50.790]Representative histology sections are shown below,
- [00:01:53.090]along with Ktrans map and DCE-MRI images,
- [00:01:56.270]which will be explained next.
- [00:01:57.220]The low wall shear stress region
- [00:02:00.700]is close to the arch
- [00:02:01.533]and promotes a vulnerable plaque phenotype
- [00:02:03.670]that has a high risk of rupturing,
- [00:02:05.100]a common cause of ischemic strokes.
- [00:02:07.280]The multidirectional wall shear stress region
- [00:02:09.750]is close to the bifurcation
- [00:02:12.060]and creates a stable plaque phenotype,
- [00:02:16.437]of the carotid bifurcation.
- [00:02:20.450]At five and nine weeks after cuff placement,
- [00:02:23.300]the mice were injected
- [00:02:24.140]with a hundred microliters of folic acid-coated gadolinium,
- [00:02:28.450]a custom diagnostic nanoparticles
- [00:02:31.730]into the lateral tail vein.
- [00:02:33.290]Previous research has shown nanoparticle accumulation
- [00:02:35.480]in plaque bodies.
- [00:02:36.770]The FA-GD nanoparticles used were designed to bond
- [00:02:41.080]to the type M1 and M2 macrophages.
- [00:02:44.520]This provides a diagnostic approach
- [00:02:46.240]to determine the advancement of the plaque.
- [00:02:49.570]After nanoparticle injection,
- [00:02:51.040]individual mice or image using DCE-MRI relaxation rate
- [00:02:57.850]and generation of transfer constants
- [00:02:59.880]using MATLAB.
- [00:03:01.310]This helps us to determine the nanoparticle distribution.
- [00:03:03.980]An example of these images is shown on the right.
- [00:03:08.150]The pixels that are darker red
- [00:03:09.720]represent higher nanoparticle accumulation,
- [00:03:11.907]and this is under the column labeled Ktrans,
- [00:03:15.570]and a representative histology image sustained
- [00:03:17.800]with the most smooth muscle alpha-actin
- [00:03:20.905]is under the,
- [00:03:22.430]is on the farthest right column.
- [00:03:23.660]The top row is low wall shear stress,
- [00:03:26.720]and the bottom row is multidirectional wall shear stress.
- [00:03:32.500]The mice were sacrificed at nine weeks after cuff placement.
- [00:03:35.160]Histology was performed on the carotid arteries,
- [00:03:37.210]and the stains that were used focused on classifying lipids,
- [00:03:41.730]collagen, and the smooth muscle alpha-actin
- [00:03:44.480]in sections of the plaque.
- [00:03:48.134]In overview of the methods,
- [00:03:50.160]as seen in the bottom image,
- [00:03:52.210]and it has the time point that each process was done at.
- [00:04:01.350]Results.
- [00:04:04.180]Image on the left
- [00:04:05.013]demonstrates a statistically significant correlation
- [00:04:08.250]between sections with the highest plaque burden
- [00:04:10.540]and sections with the highest Ktrans value,
- [00:04:13.120]but only in the upstream region,
- [00:04:16.610]which is a low wall shear stress region.
- [00:04:18.650]The multidirectional wall shear stress region
- [00:04:20.760]didn't have statistically significant correlation,
- [00:04:24.500]but we believe that's because it was close
- [00:04:26.260]to the carotid bifurcation.
- [00:04:31.560]The image on the right shows
- [00:04:33.530]the atherogenic flow environments
- [00:04:35.130]at five weeks and nine weeks
- [00:04:36.570]contain a statistically significant higher amount
- [00:04:40.070]of nanoparticle accumulation than the control regions.
- [00:04:43.850]The question that I focused on the most
- [00:04:45.320]was why the nine-week low wall sheer stress region
- [00:04:47.740]had statistically significant,
- [00:04:50.300]lower nanoparticle accumulation
- [00:04:52.610]than the five-week low wall shear stress?
- [00:04:55.020]Furthermore,
- [00:04:56.190]why it also had lower nanoparticle accumulation
- [00:04:59.993]than the nine-week sections
- [00:05:01.970]in the multidirectional wall shear stress region?
- [00:05:05.817]The difference wasn't statistically significant,
- [00:05:09.120]but it was trending in that direction.
- [00:05:11.050]This warranted the group to address
- [00:05:12.470]why this trend was occurring.
- [00:05:17.160]Let's start with the low wall shear stress region.
- [00:05:19.760]The analysis performed correlated the intensity
- [00:05:21.900]of the smooth muscle alpha-actin stain
- [00:05:24.300]done on a confocal microscope
- [00:05:27.350]to the value of the Ktrans.
- [00:05:29.410]This was performed in the plaque region,
- [00:05:32.440]the plaque cap, which is the first 13 micrometers
- [00:05:36.220]of plaque thickness on the border of the lumen, right there,
- [00:05:42.400]and the plaque body.
- [00:05:45.440]Plaque as a whole demonstrated a strong negative correlation
- [00:05:47.950]between the intensity of the smooth muscle alpha-actin stain
- [00:05:51.460]and the Ktrans value.
- [00:05:54.580]Furthermore, this,
- [00:05:58.290]on the plaque cap region
- [00:05:59.700]demonstrated a stronger negative correlation
- [00:06:01.880]and a higher p-value.
- [00:06:04.670]This was not demonstrated by the plaque body,
- [00:06:07.240]which had a weaker correlation
- [00:06:08.670]and a nonspecific, non-significant p-value.
- [00:06:13.150]This means, this demonstrates the importance
- [00:06:16.550]of that plaque cap region.
- [00:06:22.550]Also, the plaque body, the plaque as a whole,
- [00:06:25.720]did not have a significant correlation or p-value
- [00:06:29.250]at the five week time point.
- [00:06:34.590]We believe the cap region is to be,
- [00:06:38.090]is like a fibrous cap that is a common feature
- [00:06:40.710]in vulnerable advanced plaque phenotypes in humans.
- [00:06:45.610]Next image,
- [00:06:46.550]the next image is the same analysis performed
- [00:06:48.450]in the multidirectional wall flow region.
- [00:06:51.250]In this multidirectional flow region,
- [00:06:52.900]the analysis yielded different results.
- [00:06:55.040]The analysis returned a weak correlation
- [00:06:57.140]between smooth muscle alpha-actin stain intensity
- [00:06:59.910]and the Ktrans value in the plaque as a whole,
- [00:07:03.410]the plaque cap region, and the plaque body,
- [00:07:05.340]even though the plaque region
- [00:07:09.220]had more smooth muscle alpha-actin,
- [00:07:13.300]demonstrated by higher intensity.
- [00:07:16.810]With this in mind and the evidence,
- [00:07:19.480]as a reminder the flow region showed it
- [00:07:21.440]with a higher Ktrans value
- [00:07:23.610]in the nine-week time point
- [00:07:26.130]than the low wall shear stress region.
- [00:07:27.820]With this in mind and the evidence provided
- [00:07:29.890]from the smooth muscle alpha-actin stain,
- [00:07:32.900]it is within reason to conclude that
- [00:07:34.320]the organization of the smooth muscle alpha-actin
- [00:07:36.930]in the plaque region
- [00:07:38.210]of the low wall shear stress
- [00:07:39.360]inhibits proliferation of FA-GD nanoparticles.
- [00:07:43.960]The future work is,
- [00:07:46.520]that will be conducted for this study
- [00:07:49.400]is using the nanoparticles in a de-cuffed mouse model
- [00:07:52.530]and to stain macrophage markers
- [00:07:54.370]to determine healing and inflammatory responses.
- [00:07:56.860]Thank you.
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