Rett Syndrome (RTT) is a genetic neurodevelopmental disorder characterized by progressive regression of mental and psychomotor development during the symptomatic phase, following a period of apparent normal growth in the first months of life (presymptomatic phase). Evidence suggests that metabolic abnormalities, including mitochondrial dysfunction, aberrant redox homeostasis, and subclinical inflammation, significantly contribute to RTT. The unique pathogenic development of RTT involves a relatively slow transition to the symptomatic stage, providing a critical time window for early diagnosis and intervention. Our research has focused on identifying early alterations in mitochondrial function and structure, specifically related to the molecular events that drive the switch from the presymptomatic to symptomatic stage of RTT.Given the challenges in studying human brain diseases, we utilized in vitro and ex vivo models, with a particular emphasis on molecular processes involved in mitochondrial dynamics in MeCP2 knockout primary neurons and brains from MeCP2 knockout mice. To validate the role of these processes in disease manifestation, we also compared our findings with human serum samples, establishing correlations between cellular and systemic pathophysiological features of RTT. Here we present data highlighting the alterations in mitochondrial molecular composition, morphology, dynamics and oxidative state in the pathological model.These data enhance the understanding of RTT pathophysiology and facilitate the refinement of extensive preclinical studies. Ultimately, this knowledge may accelerate the development of innovative therapies aimed at delaying the onset of clinical symptoms and improving the quality of life for RTT patients.