Examination of cellular and molecular changes associated with neuronal thioredoxin-1 deficiency

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Shcholok, Tetiana
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Introduction: Oxidative distress is a potent inducer and modulator of pro-neurodegenerative processes, although the exact mechanisms that underlie these changes remain controversial. Based on the pathomorphological features, all neurodegenerative disorders commonly express a set of so-called hallmarks of neurodegeneration, i.e., accumulation of protein aggregates, cytoskeleton impairments, decreased mitochondrial metabolism, etc. Numerous attempts to tackle these traditional targets did not yield any clinically efficient treatment of neurodegenerative disorders. Therefore, there is an urgent need for a profound investigation into the pathophysiology of neurodegeneration that might lead to the development of novel strategies in diagnostics, prevention, and treatment. The involvement of oxidative distress in neurodegenerative disorders is well-documented by increased levels of oxidized proteins, lipids, and nucleic acids in neural tissues. Thioredoxin-1 (Trx-1) is a thiol antioxidant protein responsible for the reduction of disulfide bonds in oxidized proteins. Altered Trx-1 activity is reported in various neurodegenerative disorders. However, the knowledge about the role of Trx-1 system in the prevention of neurodegenerative processes remains limited. In vitro reports from our lab and others have demonstrated that downregulation of Trx-1 disrupts several vital cellular processes in neurons, including alteration of nuclear integrity. Trx-1 is involved in the prevention of nuclear lamina invagination, a newly identified mechanism typical for several neurodegenerative diseases known as laminopathy. The events consequent to nuclear lamina damage are currently the area of active investigation in neurodegeneration research. Functional, morphological, аand molecular changes associated with Trx-1 downregulations in neurons remain to be studied in animal models. Hypothesis: Trx-1 neuronal depletion is sufficient for induction of neurodegenerative-associated pathophysiology. Methods: Trx-1 neuron-specific knockout mice (Trx-nKO) were generated using Cre-Lox recombination. Synapsin-1 Cre-driver was used to achieve Trx-1 depletion in all mature neurons. I then performed phenotypical (mean survival, body weight, hindlimb circumference measurements), behavioural (hindlimb clasping test, rotarod challenge, sensory-motor, novel object recognition, and Y-maze tests), morphological (MRI), histological (immunofluorescence and H&E staining), and molecular (western blotting and enzymatic activity assay) assessments on 4- and 8-week-old animals. Results: Neuron-specific knockout of Trx-1 in mice results in severe phenotypical abnormalities as evidenced by significantly shortened lifespan, decreased body weight and muscular volume. These mice have dramatic motor deficits with no significant change in cognitive functions. Morphologically, the knockouts have significantly reduced brain and spinal cord size as well as prominent deterioration in white matter organization. Thus, these animals have significantly reduced corpus callosum thickness and lower number and diameter of axons in spinal cord and sciatic nerve. These findings coincided with a significant reduction in myelin content and perinuclear accumulation of axon-specific proteins. Molecular and histological characterization revealed significantly increased incidence of nuclear lamina damage in cortices and lumbar spinal cords of the knockouts. These mice also demonstrated a significant increase in c-Jun kinase (JNK)-medicated Tau phosphorylation and TAR DNA-binding protein (TDP-43) mislocatization and phosphorylation, pathological hallmarks typical for several neurodegenerative disorders. Altered neuronal homeostasis triggered glial responses in cortex and spinal cord of the mutants. There was a significant increase in the density of astroglia however, no significant signs of pro-inflammatory activation in the microglia population. Conclusion: This study pioneered the characterization of novel neuron-specific Trx-1 knockout in rodents. It demonstrates the significance of Trx-1 antioxidant system for neuronal homeostasis as evidenced by severe phenotypical, behavioural, morphological, and molecular abnormalities identified in Trx-nKO mice. Detailed examination of molecular players involved in activation of pro-neurodegenerative changes is beneficial for designing novel tools for diagnostics, prevention, and treatment of neurodegeneration.
Thioredoxin-1, Antioxidants, Oxidative stress, Neurodegeneration, Neuron-specific knockout, ALS, Alzheimer's disease, Nuclear lamina damage, Animal model