Assessing Antioxidant Nanoparticle Efficacy Following Traumatic Brain Injury in Mice

Aria Tarudji Author

04/01/2021 Added

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Assessing antioxidant nanoparticle efficacy following traumatic brain injury in mice. Utilizing controlled cortical impact to induce brain injury, and look for the reduction in oxidative stress with nanoparticle treatment.

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[00:00:00.870]Hi everyone. My name is Aria Tarudji.
[00:00:03.030]I am a PhD student in the Kievit lab at the biological system engineering
[00:00:06.660]department. And today I want to talk about my research in assessing antioxidant
[00:00:10.740]nanoparticle efficacy following traumatic brain injury in mice.
[00:00:15.880]A little bit of background of the traumatic brain injury.
[00:00:18.090]Traumatic brain injury is the main cause of death and disability of people below
[00:00:21.750]45 years old.
[00:00:23.280]And has 69 million new cases worldwide annually. It was estimated to costs over
[00:00:28.130]60 billion dollar a year in the U.S. alone.
[00:00:31.590]And the problem is that after primary injury,
[00:00:33.900]from the damage to the head or the brain, it doesn't stop there. However,
[00:00:38.380]there is the progression that induce more degeneration
[00:00:43.260]of the brain, it's called the secondary injury.
[00:00:47.820]So after the primary injury that causing cell death,
[00:00:52.440]releasing oxidative stress, which induces more oxidative stress.
[00:00:56.640]And this oxidative stress inducing cell death.
[00:01:00.030]Therefore the cycles continue, and this is called the secondary injury of TBI.
[00:01:05.940]And this cascade can continue more than 18 years after the injury.
[00:01:09.840]And it was hypothesized,
[00:01:11.340]It is hypothesized to increase the chance of Alzheimer's disease.
[00:01:16.210]However, currently, there is no treatment that can reduce the secondary injury in TBI.
[00:01:22.110]We think the failure is partly because it's using the small molecule drugs.
[00:01:26.640]Small molecule drugs,
[00:01:28.800]have small accumulation and small retention time in the brain injury.
[00:01:32.910]Moreover, the small molecule drugs has fairly small therapeutic window in
[00:01:37.830]which the drugs can be administered to the patient of TBI.
[00:01:42.540]And with these limitations we think that the smaller molecule drugs
[00:01:47.580]cannot be used in the traumatic brain injury to reduce the effect of
[00:01:52.500]secondary injury.
[00:01:55.980]And therefore I would like to propose the use of antioxidant nanoparticle.
[00:02:00.180]Nanoparticle is a very small molecule, according to us,
[00:02:03.120]but it's quite high in molecular weight compared to small molecule drug,
[00:02:07.470]and usually the shape is sphere.
[00:02:09.360]However, there is multiple shapes of the nanoparticles that people use.
[00:02:15.600]We found that nanoparticles,
[00:02:17.400]regardless of the size and the type of the nanoparticle's able to accumulate
[00:02:22.350]more than the gadolinium DTPA, which is a small molecule resembling
[00:02:27.150]the small molecule drugs.
[00:02:29.850]And, we also found that the small molecule have
[00:02:34.740]lower retention rate in the brain injury,
[00:02:38.160]as well as lower blood circulation
[00:02:41.640]half-life compared to the nanoparticles.
[00:02:44.370]And the evidence is not only found by our group, but also by other groups.
[00:02:51.450]And with this higher accumulation and higher retention in the brain injury,
[00:02:54.930]we hope the nanoparticle is able to have more time to react with
[00:02:59.830]the oxidative stress.
[00:03:02.050]Therefore, reducing the secondary injury cascade.
[00:03:06.880]And by that we used the antioxidant nanoparticle,
[00:03:10.300]the nanoparticle has a thioether bond and free thiol group,
[00:03:14.740]both of them is able to react with the oxidative stress.
[00:03:17.440]Therefore, inhibiting the oxidative stress to damage the cells nearby.
[00:03:22.870]And because it's reducing the oxidative stress level,
[00:03:27.130]that's why it's called the antioxidant nanoparticle.
[00:03:31.180]And what we want is that the nanoparticle will be able to react with the oxidative
[00:03:35.500]stress, therefore inhibiting the propagation of the oxidative stress,
[00:03:40.390]therefore inhibiting the cell death, and inhibiting the
[00:03:44.920]secondary injury of TBI. The test, this hypothesis,
[00:03:48.940]we use the controlled cortical impact as the primary injury in the
[00:03:53.740]mice model of TBI.
[00:03:56.560]We use controlled cortical impact or CCI because it's very repeatable
[00:04:01.900]and it can induce severe injury in the mice.
[00:04:07.120]So we can know how efficacy,
[00:04:11.080]how effective is the nanoparticle in reducing the oxidative stress,
[00:04:15.220]even in the very severe TBI.
[00:04:18.670]And we induce it on the left side of the brain
[00:04:23.830]and also the good thing with the CCI, even though it's very severe,
[00:04:27.580]it's very high survivability rate.
[00:04:31.060]So almost no mice died from this surgery.
[00:04:35.710]So, we put the mice in the stereotaxic to keep the head level and hit the
[00:04:39.760]brain with the
[00:04:41.450]piston.
[00:04:44.610]And to test the oxidative stress level.
[00:04:48.060]We use the DHE assay. DHE is a chemical that readily oxidized
[00:04:53.070]with the oxidative stress and become fluorescence.
[00:04:56.700]And the fluorescence level is able to be correlated with the
[00:05:02.070]level of the oxidative stress in the brain injury.
[00:05:06.600]Therefore we did the CCI and the nanoparticle or MnTMPyP
[00:05:11.040]injection right after the injury.
[00:05:13.680]MnTMPyP is a small molecule that also act as the antioxidant and
[00:05:18.300]currently in the clinical trials.
[00:05:20.520]So we use them as the positive control against the nanoparticle. And then we,
[00:05:25.410]at three hours time point, we inject the DHE and at four hours,
[00:05:30.210]we collect the brain for a confocal microscope. In the confocal
[00:05:34.650]microscope, we can
[00:05:37.500]quantify the florescence in the injured area compared to the uninjured
[00:05:42.360]area.
[00:05:43.410]And we found that there's huge reduction in the ROS
[00:05:48.420]with the MnTMPyP and nanoparticle treatment.
[00:05:51.420]And there are more reduction in nanoparticle compared to the MnTMPyP.
[00:05:57.920]We also test the alpha spectrin degradation product.
[00:06:02.900]So alpha spectrin is the cytoskeleton of the cells. And when the cells died,
[00:06:07.730]either by the necrosis or apoptosis,
[00:06:10.160]it degraded differently. With the necrosis,
[00:06:12.710]it'll be degraded by calpain pathway and therefore producing 145
[00:06:17.210]kilodalton, while on the apoptosis,
[00:06:20.180]it will be dominated by caspase-3 pathway,
[00:06:22.640]therefore it will be inducing the 120 kilodalton.
[00:06:27.470]And because this is more downstream compared to the oxidative stress,
[00:06:30.770]we compare it at the three days after the injury, collect the brains for the
[00:06:35.210]Western blot. And with the Western blot,
[00:06:37.640]we found the significant reduction in the 150
[00:06:42.050]kilodalton in that area of the brain,
[00:06:45.950]the left and the right brains. And,
[00:06:50.180]we also found significant reduction in the 145 kilodalton
[00:06:54.950]of the injured side of the brain, compared to the untreated CCI mice.
[00:06:59.960]As a conclusion,
[00:07:00.920]antioxidant nanoparticle's able to reduce the ROS in the acute phase of TBI,
[00:07:04.880]and also reducing cell death in the subacute phase of TBI. And the next,
[00:07:09.920]we want to look for nanoparticle therapeutic window,
[00:07:13.520]which is the nanoparticle concentration that give the
[00:07:18.470]highest efficacy in reducing the oxidative stress, and also the treatment
[00:07:23.330]window, which is the
[00:07:25.460]how long after the injury that we can still administer nanoparticle,
[00:07:29.030]and still give accumulation as well as reducing the secondary injury.
[00:07:33.650]And by that, I would like to acknowledge Dr. Forrest Kievit, my primary advisor,
[00:07:38.030]Dr. Greg Bashford, Dr. Rebecca Wachs, and Dr. Aaron Mohs as my committee members,
[00:07:42.970]Connor Gee and Brandon McDonald as helping me in handling the mice.
[00:07:48.470]And you to listening to my presentation. Thank you very much.

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Assessing Antioxidant Nanoparticle Efficacy Following Traumatic Brain Injury in Mice

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