Investigating the Potential Neuroprotective Effects of ACMSD Across Sex in P301S Mice

Description
The kynurenine pathway (KP) is a biosynthetic pathway for the catabolism of tryptophan, the amino acid precursor to serotonin. The KP has been linked to neuroinflammation, as inflammatory agents upregulate an early enzyme in the pathway (Davis & Liu, 2015;

The kynurenine pathway (KP) is a biosynthetic pathway for the catabolism of tryptophan, the amino acid precursor to serotonin. The KP has been linked to neuroinflammation, as inflammatory agents upregulate an early enzyme in the pathway (Davis & Liu, 2015; Wang et al., 2009). In addition, the metabolite quinolinic acid (QA) has been found to induce tau phosphorylation and excess glutamate release, inducing further neuroinflammation (Guillemin, 2012; Rahman et al., 2009). However, in the presence of 2-amino-3-carboxymuconate-6-semialdehyde decarboxylase (ACMSD), the KP shifts away from QA production towards that of picolinic acid (PA), a metabolite with antimicrobial and antiviral properties. Our lab has previously shown that overexpressing ACMSD via adeno-associated virus (AAV) delivery in the alpha-synuclein (α -syn) model of Parkinson's disease (PD) animal model exhibits neuroprotective effects by preventing the loss of dopaminergic neurons in the substantia nigra and limiting motor impairments caused by nigrostriatal denervation. Based on these findings, we predicted that ACMSD would provide neuroprotective effects in the P301S tauopathy model of neurodegenerative disease, a mouse model of frontotemporal dementia (FTD). Specifically, we hypothesized that ACMSD would ameliorate behavioral deficits, including those related to cognitive and emotional processing. We also predicted that ACMSD overexpression would prevent histological indices of pathology, including the expression of hyperphosphorylated tau, gliosis, and neurodegeneration. As previous findings in the literature have denoted sex differences in pathological outcomes of the P301S mouse model (Sun et al., 2020) with males showing more pronounced behavioral deficits and increased hyperphosphorylated tau than females, we hypothesized that ACMSD would show a higher degree of neuroprotection in male P301S mice. P301S and litter/age-matched wild-type controls underwent stereotaxic surgery at two months of age prior to pathological onset to deliver either an AAV-dHS-ACMSD or AAV-dHS-GFP (control) to the dorsal hippocampus. Prior to euthanasia, the mice underwent the Barnes maze test to assess cognitive function focusing on learning and memory recall. Analysis of this assay revealed that male P301S mice treated with ACMSD displayed a trend toward shorter latency in locating the escape hatch during the Barnes Maze test than untreated males, albeit not significant, suggesting a potential enhancement in spatial learning. At eight and a half months, the mice were killed, and their brains harvested. The tissue underwent immunohistochemistry staining for a marker of hyperphosphorylated tau (AT8), markers of gliosis (Iba1 and GFAP), and the pan-neuronal marker (HuC/D) to quantify pathological indices. Preliminary histological analyses show decreased immunoreactivity of AT8 in the hippocampus of P301S mice injected with ACMSD, compared to those injected with GFP control, indicating potential neuroprotective effects by limiting the amount of hyperphosphorylated tau.
Date Created
2024-05
Agent

Examining the degree of neuroprotective effects of ACMSD in the AAV-α-synuclein Parkinson’s Disease animal model

Description
Parkinson's disease (PD) is the second most common neurodegenerative disease and is characterized by impaired motor function including tremor, rigidity, akinesia and postural instability as well as non-motor symptoms including cognitive impairment, depression, and anxiety. Pathological indices of PD consist

Parkinson's disease (PD) is the second most common neurodegenerative disease and is characterized by impaired motor function including tremor, rigidity, akinesia and postural instability as well as non-motor symptoms including cognitive impairment, depression, and anxiety. Pathological indices of PD consist of protein aggregation of α-synuclein (ɑ-syn), striatal dopaminergic denervation and the loss of dopaminergic neurons in the substantia nigra. While the exact causes of PD remain unknown, substantial evidence suggests that the kynurenine pathway (KP), a pathway upregulated in response to inflammation, is implicated in its pathology. Under normal physiological conditions, the KP catabolizes the amino acid tryptophan to the metabolite 2-amino-3-carboxymuconate-6-semialdehyde (ACMS). ACMS is then spontaneously converted to quinolinic acid (QA), which is an intermediate in the formation of nicotinamide adenine dinucleotide (NAD+), a cofactor in mitochondrial respiration, thus serving as an essential molecule in cellular energy metabolism. QA itself may serve as a contributing factor in PD pathology as this metabolite promotes oxidative stress, mitochondrial dysfunction, excitotoxicity, and positively correlates with increased scores in the Unified Parkinson’s Disease Rating Scale (UPDRS) in motor experiences of daily living (UPDRS II) and motor complications (UPDRS III). ACMS is enzymatically converted to picolinic acid (PA), an iron chelator and anti-viral agent, by 2-amino-3-carboxymuconate-6-semialdehyde decarboxylase (ACMSD), thus limiting the formation of QA and potentially conferring neuroprotective effects. To model Parkinson’s disease, we overexpressed α-synuclein unilaterally in the substantia nigra with an adeno-associated virus (AAV), which leads to dose-dependent neurodegeneration, aggregation of α-syn and motor impairments. This model may therefore serve to examine perturbations of the KP in the context of PD including increased QA/PA expression. We hypothesize that the overexpression of the enzyme ACMSD in the AAV-α-syn model of PD will reduce the degree of neurodegeneration and inflammation and prevent motor decline. Neurodegeneration and neuroinflammation will be quantified histologically, through stereology and densitometry respectively. Observable PD-like symptoms will be quantified using behavioral assessments, particularly the cylinder test and amphetamine induced rotations to assess the degree of motor decline.
Date Created
2024-05
Agent

Chitinases as a Novel Therapeutic Target in Parkinson's Disease

Description
Parkinson’s disease (PD) is a debilitating neurodegenerative disease characterized primarily by physical impairments such as tremors, poor balance, and bradykinesia; however, some individuals with PD will additionally experience numerous nonmotor symptoms such as dementia, depression, and sleep disturbances amongst various

Parkinson’s disease (PD) is a debilitating neurodegenerative disease characterized primarily by physical impairments such as tremors, poor balance, and bradykinesia; however, some individuals with PD will additionally experience numerous nonmotor symptoms such as dementia, depression, and sleep disturbances amongst various other life-altering ailments. Two of the key pathological hallmarks of PD include the death of melanated dopaminergic neurons in the nigrostriatal pathway and the accumulation of Lewy bodies, which are primarily composed of aggregates of the protein α-synuclein (α-syn). Interestingly, members of the chitinase protein family, namely chitinase-3-like protein-1 (L1), have heightened concentrations in a number of neurodegenerative diseases other than PD. To investigate the specific role L1 plays in PD etiology, we evaluated if astrocytic L1 expression was elevated in postmortem brain tissue of PD patients as well as in an α-syn overexpression rat model, and further tested if manipulating astrocytic-specific L1 expression correlated with neuroinflammation and nigral neuronal degeneration in the model. Preliminary histological analysis has shown increased levels of L1 expression in the α-syn model before neuronal loss occurs, and in human tissue, L1 was found to be significantly increased in the postmortem tissue of individuals with PD versus non-diseased controls. Investigations in identifying an astrocytic-specific virus capsid and manipulating L1 expression in the α-syn model are ongoing. This preliminary data thus far supports that increased astrocytic expression of L1 is associated with PD pathology.
Date Created
2023-12
Agent

Herpes Simplex Virus 1 Amplicon Vectors

161592-Thumbnail Image.png
Description
Globally, about two-thirds of the population is latently infected with herpes simplex virus type 1 (HSV-1). HSV-1 is a large double stranded DNA virus with a genome size of ~150kbp. Small defective genomes, which minimally contain an HSV-1 origin of

Globally, about two-thirds of the population is latently infected with herpes simplex virus type 1 (HSV-1). HSV-1 is a large double stranded DNA virus with a genome size of ~150kbp. Small defective genomes, which minimally contain an HSV-1 origin of replication and packaging signal, arise naturally via recombination during viral DNA replication. These small defective genomes can be mimicked by constructing a bacterial plasmid containing the HSV-1 origin of replication and packaging signal, transfecting these recombinant plasmids into mammalian cells, and infecting with a replicating helper virus. The absence of most viral genes in the amplicon vector allows large pieces of foreign DNA (up to 150kbp) to be incorporated. The HSV-1 amplicon is replicated and packaged by the helper virus to form HSV-1 particles containing the amplicon DNA. We constructed a novel HSV-1 amplicon vector system containing lambda phage-derived attR sites to facilitate insertion of transgenes by Invitrogen Gateway recombination. To demonstrate that the amplicon vectors work as expected, we packaged the vector constructs expressing Emerald GFP using the replication-competent helper viruses OK-14 or HSV-mScartlet-I-UL25 in Vero cells and demonstrate that the vector stock can subsequently transduce and express Emerald GFP. In further work, we will insert transgenes into the amplicon vector using Invitrogen Gateway recombination to study their functionality.
Date Created
2021
Agent