Bregalda Alessandro, Carducci Claudia, Viscomi Maria Teresa, Pierigè Francesca, Biagiotti Sara, Menotta Michele, Biancucci Federica, Pascucci Tiziana, Leuzzi Vincenzo, Magnani Mauro, Rossi Luigia
Department of Biomolecular Sciences, University of Urbino "Carlo Bo", via Saffi 2, 61029 Urbino, PU, Italy.
Department of Experimental Medicine, Sapienza University, viale del Policlinico 155, 00161 Rome, Italy.
Neurobiol Dis. 2023 May;180:106093. doi: 10.1016/j.nbd.2023.106093. Epub 2023 Mar 21.
Untreated phenylketonuria (PKU) patients and PKU animal models show hypomyelination in the central nervous system and white matter damages, which are accompanied by myelin basic protein (MBP) impairment. Despite many assumptions, the primary explanation of the mentioned cerebral outcomes remains elusive. In this study, MBP protein and mRNA expression on brains of wild type (WT) and phenylketonuric (ENU2) mice were analyzed throughout mice lifespan (14-60-180-270-360-540 post-natal days, PND). The results confirmed the low MBP expression at first PND times, while revealed an unprecedented progressive MBP protein expression recovery in aged ENU2 mice. Unexpectedly, unaltered MBP mRNA expression between WT and ENU2 was always observed. Additionally, for the same time intervals, a significant decrease of the phenylalanine concentration in the peripheral blood and brain of ENU2 mice was detected, to date, for the first time. In this scenario, a translational hindrance of MBP during initial and late cerebral development in ENU2 mice was hypothesized, leading to the execution of a microRNA microarray analysis on 60 PND brains, which was followed by a proteomic assay on 60 and 360 PND brains in order to validate in silico miRNA-target predictions. Taken together, miR-218-1-3p, miR-1231-3p and miR-217-5p were considered as the most impactful microRNAs, since a downregulation of their potential targets (MAG, CNTNAP2 and ANLN, respectively) can indirectly lead to a low MBP protein expression. These miRNAs, in addition, follow an opposite expression trend compared to MBP during adulthood, and their target proteins revealed a complete normalization in aged ENU2 mice. In conclusion, these results provide a new perspective on the PKU pathophysiology understanding and on a possible treatment, emphasizing the potential modulating role of differentially expressed microRNAs in MBP expression on PKU brains during PKU mouse lifespan.
未经治疗的苯丙酮尿症（PKU）患者和PKU动物模型在中枢神经系统中表现出髓鞘形成不足和白质损伤，同时伴有髓鞘碱性蛋白（MBP）受损。尽管有许多假设，但上述脑部病变的主要原因仍不清楚。在本研究中，分析了野生型（WT）和苯丙酮尿症（ENU2）小鼠在整个生命周期（出生后14 - 60 - 180 - 270 - 360 - 540天，PND）大脑中MBP蛋白和mRNA的表达。结果证实，在出生后早期，MBP表达较低，而在老年ENU2小鼠中发现了前所未有的MBP蛋白表达逐渐恢复的现象。出乎意料的是，WT和ENU2之间的MBP mRNA表达始终未改变。此外，在相同的时间间隔内，首次检测到ENU2小鼠外周血和大脑中苯丙氨酸浓度显著降低。在这种情况下，推测ENU2小鼠在大脑发育早期和晚期存在MBP的翻译障碍，因此对60天龄的大脑进行了微小RNA微阵列分析，随后对60天龄和360天龄的大脑进行了蛋白质组学分析，以验证计算机模拟的微小RNA - 靶标预测。综合来看，miR - 218 - 1 - 3p、miR - 1231 - 3p和miR - 217 - 5p被认为是最具影响力的微小RNA，因为它们潜在靶标（分别为MAG、CNTNAP2和ANLN）的下调可间接导致MBP蛋白表达降低。此外，在成年期，这些微小RNA与MBP呈现相反的表达趋势，并且它们的靶蛋白在老年ENU2小鼠中显示完全正常化。总之，这些结果为理解PKU的病理生理学和可能的治疗提供了新的视角，强调了差异表达的微小RNA在PKU小鼠生命周期中对PKU大脑中MBP表达的潜在调节作用。
