摘要: Testicular ageing is accompanied by a series of morphological changes, while the features of mitochondrial dysfunction remain largely unknown. Herein, we observed a range of age-related modifications in testicular morphology and spermatogenic cells, and conducted single-cell RNA sequencing on young and old testes in Drosophila. Pseudotime trajectory revealed significant changes in germline subpopulations during ageing. Our examination unveiled that genes showing bias in spermatids exhibited higher dN/dS than those in GSCs_Spermatogonia. Genes biased towards young GSCs_Spermatogonia displayed higher dN/dS than those in old GSCs_Spermatogonia. Interestingly, genes biased towards young spermatids demonstrated lower dN/dS in contrast to those in old spermatids, revealing the complexity of evolutionary adaptations during ageing. Furthermore, mitochondria associated events, including oxidative phosphorylation, TCA cycle and pyruvate metabolism, were significantly enriched in germline subpopulations. Specifically, mitochondrial function was significantly impaired during the process of testicular ageing, concurrently emphasising the role of several key nuclear genome-encoded mitochondrial regulatory genes, such as Hsp60B, fzo, Tim17b1 and mRpL12. Our data offer insights into testicular homeostasis regulated by mitochondrial function during the ageing process.

摘要: Human midbrain organoids with functional dopaminergic (DA) neurons are invaluable for the therapeutic development of Parkinson's disease (PD). However, current methods face significant limitations, including challenges in generating pint-sized organoids enriched with DA neurons and the lack of robust functional assays for efficiently evaluating neural networks over extended periods. Here we present an innovative approach that combines developmental patterning with mechanical cutting to produce small midbrain organoids, with diameters less than 300 mu m, suitable for long-term evaluation, along with a comprehensive functional assay system consisting of calcium transient assay, neurite extension assay, and multielectrode array (MEA) assay. Radial cutting of organoids into four to eight portions according to their sizes at the appropriate developmental stage significantly increases the yield of viable organoids while reducing necrotic cell regions. Using the functional assay system, we demonstrate that DA neurons within the organoids extend long projections, respond to dopamine stimulation, and form neural networks characterised by giant depolarising potential-like events. Our approach supports the generation of midbrain organoids and PD models that can be used for long-term functional testing.