30. Validation of a Novel Transgenic Mouse Model Expressing GFP in Motoneurons for the Investigation of Peripheral Nerve Injury
Notice bibliographique
Résumé
PURPOSE: Peripheral nerve injury (PNI) is a significant clinical problem, and novel therapies are evidently necessary to optimize functional outcomes. However, the regenerative environment of motor fibers must first be thoroughly elucidated, an effort which is currently limited by inherent drawbacks of conventional visualization techniques. Mnx1 is a homeobox transcription factor that has been established as a spinal motoneuron-specific marker. Here, we propose a transgenic mouse model (Mnx-GFP) in which expression of a ZsGreen1 GFP is driven by the Mnx1 promoter for the specific investigation of motoneuron regeneration. This study aims to confirm an absence of off-target effects of recombination on nerve regeneration, fiber characteristics, and grip strength in male Mnx-GFP mice after median nerve injury. METHODS: 4-week old founder mice (males: heterozygous Mnx1-cre, females: homozygous Rosa expressing ZsGeen1 GFP) were obtained from the Jackson Laboratory and cross-bred in-house, followed by genotyping to confirm recombination. Male transgenic Mnx-GFP (n=2) and wildtype C57BL/6 (n=10) mice underwent surgical transection and immediate repair of the right median nerve. Every 3d following the intervention, unilateral volitional forelimb grip strength was assessed in all mice. On day 27, the median nerve was re-exposed and transected 4mm distal to the original repair site, at which point a 2mm distal segment was collected for histomorphometry. Fluorogold (FG) neurotracer was applied to the nerve stump to label regenerated neurons, followed by collection of DRG and spinal cord tissue 1 week later for immunofluorescent imaging and immunostaining for the neuronal marker Isl1. RESULTS: No significant differences in grip strength (g) were observed between Mnx-GFP and wildtype at baseline (122.7±11.74 vs. 122.8±6.72; p=0.99) or 4-weeks post-injury (31.83±4.41 vs. 39.29±1.45; p=0.071). Retrograde-labelling revealed that motor (274±36 vs. 169±20; p=0.057) and sensory (1160±104 vs. 1072±118; p=0.76) neuron counts were comparable between cohorts. Furthermore, the total pool of Mnx+ motoneurons in the cervical enlargement was observed to include neurons that successfully regenerated to the point of neurotracer application (FG+) as well as those that did not. Interestingly, a considerable number of non-brachial motoneurons were both GFP+ and Isl1+. Histomorphometry of the median nerve revealed no significant difference in myelin thickness (0.56±0.020μm vs. 0.51±0.014μm; p=0.19), axon diameter (1.89±0.040μm vs. 1.81±0.090μm; p=0.75), fiber diameter (3.025±0.025μm vs. 2.84±0.10μm; p=0.44), or g-ratio (0.64±0.025 vs. 0.64±0.012; p>0.99) at the 4-week timepoint. All values are presented as Mnx-GFP vs. C57BL/6. CONCLUSION: Currently, no transgenic models exist to specifically visualize motoneuron regeneration. This data demonstrates that the Mnx-GFP transgene had no measurable impact on regenerative capacity, fiber characteristics, or motor function, supporting this model as a valid tool to study PNI. The use of Mnx-GFP mice alongside more traditional analytical metrics thus has the potential to improve the efficiency and reproducibility of data collected in this field. Further, the Mnx+ motoneuron population was not limited to neurons that successfully regenerated to the site of retrograde-labelling, suggesting a potential utility of this model for assessing regenerative efficacy. Future investigations may aim to examine spinal motor column-specific markers to aid in identifying the cell population of interest within the larger motoneuron pool.
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Comment cette classification a été obtenuedéplier
Prédiction distillée sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Apprise à partir de 10 348 étiquettes directes de Codex et de 10 348 étiquettes directes de Gemma. Le mode candidate est l'union des têtes enseignantes seuillées; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont ni des étiquettes humaines ni des étiquettes directes de modèles de pointe.
Scores Codex et Gemma par catégorie
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,001 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,001 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,001 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,000 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule tête enseignante, pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».