Auswirkungen einer PACAP-Gendefizienz auf Purkinjezell- Degeneration und Atrophie des Kleinhirns im Superoxiddismutase-1 Mausmodell der Amyotrophen Lateralsklerose

Die Amyotrophe Lateralsklerose (ALS) ist eine neurodegenerative Erkrankung. Hauptsymptom ist ein Verlust der Willkürmotorik, bedingt durch ein kontinuierliches Absterben motorischer Nervenzellen in Cortex, Hirnstamm und Rückenmark. Aktuell wird davon ausgegangen, dass sich die Pathologie der ALS nic...

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Bibliographic Details
Main Author: Burrichter, Jannik
Contributors: Schütz, Burkhard (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2022
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Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease. The main symptom is a loss of voluntary motor skills, caused by the continuous death of motor nerve cells in the cortex, brain stem and spinal cord. It is assumed that the pathology of ALS is not only limited to motor neurons, but is a multisystem network disease with characteristic impaired connectivity. Based on a genetic mutation that occurs in 20% of all hereditary ALS cases, the establishment of the superoxide dismutase 1 (SOD1) transgenic mouse model by Gurney et al. in 1994 contributed significantly to the elucidation of possible underlying pathomechanisms. The neuropeptide Pituitary-adenylate-cyclase-activating-polypeptide (PACAP) was initially isolated from the hypothalamus and belongs to a gene family that also includes the vasoactive intestinal polypeptide (VIP), secretin, the growth hormone (GH), as well as glucagon. PACAP has various neuroprotective, but also neuroinflammatory functions and effects on the CNS and is involved in acute and chronic neurodegenerative diseases such as ALS. This dissertation thesis dealt with the effects of a PACAP deficiency in the SOD1 (G93A) mouse model of ALS with a focus on microscopic and macroscopic changes of the cerebellum. For this purpose, the examined mouse groups either harbored the genetic SOD1 mutation, or a PACAP knockout, or both factors, and effects at the two measurement times of the presymptomatic stage (p60/70) and end stage (p120) were evaluated. Histologically, the Purkinje cells were counted and a possible macroscopic atrophy of the cerebellum (based on the transverse diameter and length) assessed with MRI. Finally, histological microtome sections were compared with the MRT diagnosis based on an area calculation in the sagittal plane. In the Purkinje cell analysis, neither a monofactorial SOD1 mutation, nor the sole PACAP knockout resulted in Purkinje cell death at either point in time. A simultaneous occurrence of the SOD1 mutation with an existing PACAP knockout (WT/KO vs. SOD1/KO) in the early stage led to a reduced Purkinje cell count. For the group SOD1/WT vs. SOD1/KO, this effect could not be proven, both in the early stage and endstage. The transverse diameter of the cerebellum calculated in the MRI showed significant macroscopic atrophy for the pure SOD1 mutation (WT / WT vs. SOD1 / WT) at the p60/70 and endstage time. For the sole PACAP knockout, this effect was not observed. In the calculated length of the cerebellum, however, only the group with the sole PACAP knockout (WT/WT p60/70 vs. WT/KO p60/70) was found to be significantly atrophic. This atrophy could not be replicated in the cerebellar length for the SOD1 mutation at p60/70 or endstage (WT/WT p60/70 vs. SOD1/WT p60/70 or SOD1/WT p60/70 vs. SOD1/WT endstage). These MRI results indicated a possible tissue destruction in the case of deficiency or a neuroprotective effect in the case of normal function, possibly also to the cell genesis and cell death of the cerebellum. Translated into humans, this diagnostic method could also be useful in ALS cerebellar diagnostics in vivo and may be used more forcefully in future studies. In the last step of this thesis it was compared to what extent a macroscopic atrophy can be assessed equally by histology and radiology by calculating the area of the SOD1-mutated and PACAP-deficient cerebellum. Histologically, the usability of an area calculation could only be proven in the presymptomatic ALS mice (p60/70 time point). Here, PACAP knockout mice showed significant cerebellar atrophy. In the PACAP knockout end-stage group, but also for the monofactor of the SOD1 mutation, however, this significance could not be confirmed at either time point. In the MRT area calculation, a significant atrophy of the cerebellum was detectable for both the PACAP knockout and SOD1 mutated animals at the p60/70 time point. Only the comparison between SOD1/WT p60/70 vs. SOD1/WT End showed no progression in surface atrophy. In summary, with both diagnostic methods, histology and MRI, there is the possibility of a meaningful analysis of the cerebellar surface in the early stages of ALS. In contrast to histology, an effect of the SOD1 mutation could also be clearly demonstrated with MRI. In addition, the undisputed advantage of MRI lies in the possibility of in vivo diagnostics, which can also be used for humans. Thus, in comparison to histological sectioning, the use of a radiological evaluation of the cerebellum in the SOD1 mouse model of ALS seems beneficial and could play a greater role in future research.