Objectives: Parkinson's disease (PD) is a common neurodegenerative disorder characterized by the progressive and selective degeneration of dopaminergic neurons. Microglial activation and neuroinflammation are associated with the pathogenesis of PD. However, the relationship between microglial activation and PD pathology remains to be explored.Materials and Methods: An acute regimen of MPTP was administered to adult C57BL/6J mice with normal, much reduced or repopulated microglial population. Damages of the dopaminergic system were comprehensively assessed. Inflammation‐related factors were assessed by quantitative PCR and Multiplex immunoassay. Behavioural tests were carried out to evaluate the motor deficits in MPTP‐challenged mice.Results: The receptor for colony‐stimulating factor 1 inhibitor PLX3397 could effectively deplete microglia in the nigrostriatal pathway of mice via feeding a PLX3397‐formulated diet for 21 days. Microglial depletion downregulated both pro‐inflammatory and anti‐inflammatory molecule expression at baseline and after MPTP administration. At 1d post‐MPTP injection, dopaminergic neurons showed a significant reduction in PLX3397‐fed mice, but not in control diet (CD)‐fed mice. However, partial microglial depletion in mice exerted little effect on MPTP‐induced dopaminergic injuries compared with CD mice at later time points. Interestingly, microglial repopulation brought about apparent resistance to MPTP intoxication.Conclusions: Microglia can inhibit PD development at a very early stage; partial microglial depletion has little effect in terms of the whole process of the disease; and microglial replenishment elicits neuroprotection in PD mice.

Prime editing was used to insert and correct various pathogenic mutations except for beta-thalassemia variants, which disrupt functional beta-globin and prevent hemoglobin assembly in erythrocytes. This study investigated the effect of gene correction using prime editor version 3 (PE3) in a mouse model with the human beta-thalassemia IVS-II-654 mutation (C > T). The T conversion generates a 5′ donor site at intron 2 of the beta-globin gene resulting in aberrant splicing of pre-mRNA, which affects beta-globin expression. We microinjected PE3 components (pegRNA, nick sgRNA, and PE2 mRNA) into the zygotes from IVS-II-654 mice to generate mutation-edited mice. Genome sequencing of the IVS-II-654 site showed that PE3 installed the correction (T > C), with an editing efficiency of 14.29%. Reverse transcription-PCR analysis showed that the PE3-induced conversion restored normal splicing of beta-globin mRNA. Subsequent comprehensive phenotypic analysis of thalassemia symptoms, including anemic hematological parameters, anisocytosis, splenomegaly, cardiac hypertrophy, extramedullary hematopoiesis, and iron overload, showed that the corrected IVS-II-654 mice had a normal phenotype identical to the wild type mice. Off-target analysis of pegRNA and nick sgRNA additionally showed the genomic safety of PE3. These results suggest that correction of beta-thalassemia mutation by PE3 may be a straightforward therapeutic strategy for this disease.

Studies reveal a linkage of miR-29s in aging and Parkinson’s disease (PD). Here we show that the serum levels of miR-29s in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice exhibited dynamic changes. The role of miR-29b2/c in aging and PD was studied utilizing miR-29b2/c gene knockout mice (miR-29b2/c KO). miR-29b2/c KO mice were characterized by a markedly lighter weight, kyphosis, muscle weakness and abnormal gait, when compared with wild-type (WT) mice. The WT also developed apparent dermis thickening and adipose tissue reduction. However, deficiency of miR-29b2/c alleviated MPTP-induced damages of the dopaminergic system and glial activation in the nigrostriatal pathway and consequently improved the motor function of MPTP-treated KO mice. Knockout of miR-29b2/c inhibited the expression of inflammatory factors in 1-methyl-4-phenylpyridinium (MPP⁺)-treated primary cultures of mixed glia, primary astrocytes, or LPS-treated primary microglia. Moreover, miR-29b2/c deficiency enhanced the activity of AMPK but repressed the NF-κB p65 signaling in glial cells. Our results show that miR-29b2/c KO mice display the progeria-like phenotype. Less activated glial cells and repressed neuroinflammation might bring forth dopaminergic neuroprotection in miR-29b2/c KO mice. Conclusively, miR-29b2/c is involved in the regulation of aging and plays a detrimental role in Parkinson’s disease.

Introduction: The new targeted gene editing technologies, such as the CRISPR/Cas system, enable researchers to insert or delete genes at targeted loci efficiently. The Cre-loxp recombination system is widely used to activate or inactivate genes with high spatial and temporal specificity.Methods: Using the CRISPR/Cas9 system, we inserted the CreERT2 transgene expression cassette into the Cd2 gene locus to generate conditional Cre-driver line Cd2-CreERT2 knock-in mice, which drove the expression of CreERT2 by the endogenous Cd2 promoter. By mating the Cd2-CreERT2 strain with a Rosa26-LSL-tdTomato reporter mouse strain which contains a tdTomato expression fragment blocked with a loxP-flanked STOP cassette (LSL) driven by a CAG promoter, a Cd2-CreERT2;Rosa26-LSL-tdTomato reporter strain was obtained to evaluate the expression pattern of CD2 in different cell types.Results: After treatment with tamoxifen, the Cd2-CreERT2 knock-in mice were induced to perform efficient recombination at the loxP site following CreERT2 activation and cause the expression of tdTomato fluorescence. The tdTomato and CD2 were expressed in the T cells of peripheral blood, spleen and mesenteric lymph nodes, whereas detected in a low proportion in the B cells. While about 20% of cells labeled with tamoxifen-induced tdTomato were CD2⁺ monocytes in peripheral blood, 10% of dendritic cells were tdTomato⁺/CD2⁺ cells. Tamoxifen-independent expression of tdTomato occurred in approximately 3% of CD2⁺ macrophages, but in negligible (~0.5%) in CD2⁺ granulocytes.Discussion: This work supplied a new transgenic mouse as a valuable tool for lineage tracing in CD2-expressing cells, for conditional mutant studies of immune modulatory effects in a time-dependent manner, and analysis of the potential therapeutic effect of CD2-targeting biologics.