Rett syndrome (RTT) is a neurodevelopmental disorder that is caused by loss-of-function mutations in the methyl-CpG binding protein 2 (MeCP2) gene. RTT patients experience a myriad of debilitating symptoms, which include respiratory phenotypes that are often associated with lethality. Our previous work established that expression of the M1 muscarinic acetylcholine receptor (mAchR) is decreased in RTT autopsy samples, and that potentiation of the M1 receptor improves apneas in a mouse model of RTT; however, the population of neurons driving this rescue is unclear. Loss of Mecp2 correlates with excessive neuronal activity in cardiorespiratory nuclei. Since M1 is found on cholinergic interneurons, we hypothesized that M1-potentiating compounds decrease apnea frequency by tempering brainstem hyperactivity. To test this, Mecp2+/- and Mecp2+/+ mice were screened for apneas before and after administration of the M1 positive allosteric modulator (PAM) VU0453595 (VU595). Brains from the same mice were then imaged for c-Fos, ChAT, and Syto16 using whole-brain light-sheet microscopy to establish genotype and drug-dependent activation patterns that could be correlated with VU595's efficacy on apneas. The vehicle-treated Mecp2+/- brain exhibited broad hyperactivity when coupled with the phenotypic prescreen, which was significantly decreased by administration of VU595, particularly in regions known to modulate the activity of respiratory nuclei (i.e. hippocampus and striatum). Further, the extent of apnea rescue in each mouse showed a significant positive correlation with c-Fos expression in non-cholinergic neurons in the striatum, thalamus, dentate gyrus, and within the cholinergic neurons of the brainstem. These results indicate that Mecp2+/- mice are prone to hyperactivity in brain regions that regulate respiration, which can be normalized through M1 potentiation.

Intellectual and developmental disabilities (IDD) are varied in their nature and presentation. Barriers to oral healthcare are reported in studies of general populations with IDD but these may not reflect the barriers experienced by individuals with rare disorders such as Rett syndrome (RTT). There are also few peer-reviewed studies in the Australian context exploring barriers to dental care access for patients living with a disability. This qualitative study explored caregivers' perceptions and experiences regarding oral health and access to dental care for those with RTT in Australia.

Parents of 31 individuals with a confirmed MECP2 mutation were sampled purposively from the Australian Rett Syndrome Database. Interview questions were based on earlier studies used in other disability populations and queried identification and management of dental pain and influence of other comorbidities in their child's oral care. Interviews were audio-recorded, transcribed and analysed using NVivo (Version 12 Plus). Directed content analysis was used to code data to a framework constructed from a literature review of factors affecting access to professional oral healthcare systems and factors affecting access to optimal at-home oral care in disability.

The most frequently cited barriers to professional dental care were dentist-related, while caregiver related financial barriers were cited by a minority of families. Dentist-related financial barriers were not present in these data. Most factors affecting access to optimal at-home oral care coded to the existing framework, with further enablers identified under training for the caregiver or parent.

The findings of this study provide a point of reference to understand factors affecting provision of at-home dental care and professional services to enable optimal oral health in RTT. Future research could explore the provision of targeted oral health information on RTT to carers and clinicians.

Arboviruses are a group of arthropod-borne viral pathogens that pose a significant threat to the public health system. The clinical manifestations associated with these viruses range from self-limiting infections to life-threatening disorders. As a group of systemic viral infections, arboviruses can affect various parts of human organ systems, such as the nervous system. In the nervous system, epigenetic mechanisms are involved in various mechanisms including adult neurogenesis, neuronal-glial differentiation, the regulation of neural behaviour and neural plasticity, as well as other brain functions such as memory, and cognition. Hence, epigenetic deregulation is a key factor in the aetiology of different neurological disorders that highlights the importance of studying the underlying mechanisms and risk factors to introduce effective therapeutic approaches. There is mounting evidence that arboviruses that affect the nervous system take advantage of various mechanisms to modulate epigenetic processes to regulate their life cycles. This phenomenon may affect the nervous system leading to neurotropic arboviral infection-associated neurological disorders. Hence, it is important to understand reciprocal interplays between neurotropic arboviral pathogens and epigenetic processes to better control these disorders. The present review provides an overview of different interactions of arboviruses with epigenetic mechanisms during neurotropic arboviral infections. It uniquely focuses on the interplay between epigenetic modifications and arboviral neurotropism, shedding light on potential therapeutic strategies that have not been comprehensively addressed before. Targeting virus-induced epigenetic alterations, such as miRNA regulation, could lead to novel antiviral therapies aimed at mitigating neuroinflammation and disease severity.

Alternative splicing is one of the processes that contributes to producing a vast protein diversity from the limited number of protein-coding genes in higher eukaryotes. The Methyl CpG Binding Protein 2 (Mecp2) gene, whose mutations cause Rett syndrome, generates two protein isoforms, MeCP2E1 and MeCP2E2, by alternative splicing. These isoforms likely possess non-redundant functions. However, the molecular mechanism for Mecp2 pre-mRNA alternative splicing remains to be understood. Here, we analyzed the alternative splicing mechanism of MeCP2 pre-mRNA and found that exon 2 is efficiently recognized through adjacent strong splice sites. In addition, exonic splicing enhancer (ESE) in exon 2 plays an important role in exon 2 inclusion, which is highly likely to be mediated by SRSF9.

Conventional methods of gene transfer lead to inconsistent transgene expression within cells. This variability can be problematic, particularly in conditions like Rett syndrome (RTT), a neurological disorder caused by mutations in the MECP2 (methyl-CpG binding protein 2) gene, because overexpression of MECP2 can also cause adverse effects. To address these challenges, we devised a gene regulation system called Expression Attenuation via Construct Tuning (EXACT), which uses a self-contained, microRNA-based feed-forward loop that not only ensures more consistent transgene expression but also protects against excessive expression. Through cell-based screening assays, we demonstrated the ability of the EXACT circuit to modulate the expression of full-length human MeCP2. Compared with a conventional construct, an EXACT-MECP2 construct exhibited a narrower range of cellular protein abundance. Furthermore, the degree of regulation by the EXACT circuit increased with higher transgene doses in vitro and in wild-type mice and mice modeling RTT. On the basis of cellular and in vivo testing, we identified an optimal configuration for the adeno-associated virus serotype 9 (AAV9) construct for self-regulated MECP2 gene therapy, designated NGN-401. Delivery of NGN-401 to neonatal male Mecp2-/y hemizygous mice via intracerebroventricular injection resulted in prolonged survival and amelioration of RTT-like phenotypes compared with vehicle-treated animals. NGN-401 was also well tolerated by female Mecp2+/- mice and healthy juvenile nonhuman primates, in contrast with a conventional construct, which caused toxicity. The results from these studies underpin a first-in-human pediatric trial of NGN-401 in RTT (ClinicalTrials.gov, NCT05898620).

Big data in drug development may not satisfactorily address the demands of precision medicine in a rare disease population, making the use of smaller clinical trials necessary. Consequently, the use of innovative design and analysis of these clinical trials using model-informed approaches have become indispensable. This requires informative exposure-outcome analysis, together with formal statistical analysis, which should include the strength of evidence for a study outcome. We demonstrate how knowledge can be gained, with supporting strength of evidence, from a small (data) clinical trial with a low dose of blarcamesine in the treatment of Rett syndrome. Based on a small data paradigm, pharmacometrics item response theory modelling and Bayes factor analysis were used to demonstrate the efficacy of blarcamesine in Rett syndrome.

Rett syndrome is a genetic neurodevelopmental disorder that predominantly affects girls. While microcephaly is a common feature, there is limited information on the detailed structural changes in the brain. This study aimed to identify regional brain volume abnormalities and explore the correlation between brain volume and clinical characteristics.

We compared the regional brain volumes of 20 female children with Rett syndrome to those of 25 healthy female children. Additionally, we assessed the correlation between regional brain volume, Clinical Severity Scores, and epilepsy status.

Significantly smaller volumes were observed in all brain regions, including the cerebral cortex, cerebral white matter, subcortical gray matter, cerebellum, and brainstem. Within the cortical regions, volume reduction was prominent in the left precentral, right lateral occipital, left precuneus, left inferior parietal, and right medial orbitofrontal cortices. After correcting for intracranial volumes, volume reduction was more prominent in the cerebral cortices than in the cerebral white matter. Small volumes were consistently observed, regardless of age. Negative correlations were observed between the volumes of multiple regions and the Clinical Severity Scores. There were no correlations among regional brain volume, seizure control, or duration of epilepsy.

The mechanism underlying the cortical-dominant volume reduction remains unclear; however, it may be caused by altered synapse development associated with methyl-CpG-binding protein 2 gene abnormalities. Characteristic impairments in visual recognition and deterioration of motor function in Rett syndrome may be associated with significant volume reduction in specific cortical regions, such as the lateral occipital cortex, precuneus, and precentral gyrus.

Objectives: Despite growing knowledge of the underlying neurobiology of autism spectrum disorder (ASD) and related neurogenetic syndromes, treatment discovery has remained elusive. In this review, we provide a blueprint for translational precision medicine in ASD and related neurogenetic syndromes. Methods: The discovery of trofinetide for Rett syndrome (RTT) is described, and the role of nonmammalian, mammalian, and stem cell model systems in the identification of molecular targets and drug screening is discussed. We then provide a framework for translating preclinical findings to human clinical trials, including the role of biomarkers in selecting molecular targets and evaluating target engagement, and discuss how to leverage these findings for future ASD drug development. Results: Multiple preclinical model systems for ASD have been developed, each with tradeoffs with regard to suitability for high-throughput small molecule screening, conservation across species, and behavioral face validity. Future clinical trials should incorporate biomarkers and intermediate phenotypes to demonstrate target engagement. Factors that contributed to the approval of trofinetide for RTT included replicated findings in mouse models, a well-studied natural history of the syndrome, development of RTT-specific outcome measures, and strong engagement of the RTT family community. Conclusions: The translation of our growing understanding of the neurobiology of ASD to human drug discovery will require a precision medicine approach, including the use of multiple model systems for molecular target selection, evaluation of target engagement, and clinical trial design strategies that address heterogeneity, power, and the placebo response.

Rett Syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked gene encoding the methyl-CpG-binding protein 2 (MECP2). Impaired function of this transcriptional regulator leads to profound neurological defects, among which respiratory distress, motor function and cognitive disorders are prominent. Despite great advances in understanding RTT neurobiology, therapies that can meaningfully improve patients' symptoms are still needed. Here, we focused on 5-HT1A receptor-mediated serotonergic signaling as a potential therapeutical route for RTT. We report the effects of a drug candidate, NLX-101, a highly selective, biased agonist of 5-HT1A post-synaptic receptors at brainstem and cortical regions, on key phenotypes of RTT. Unrestrained whole-body plethysmography studies confirmed and extended the previous observation that single i.p. administration of NLX-101 dose-dependently reduced the occurrence and length of apneic events in Mecp2tm1.1Bird heterozygous female mice and largely corrected respiratory irregularity. Although no preservation of motor function was observed, early onset chronic administration of NLX-101 entirely prevented the cognitive deficits of the Mecp2tm1.1Bird mice both in the short and the long-term memory paradigms of the Novel Object Recognition upon 10 weeks of treatment, an effect that was maintained throughout animals' age. Similar effects were observed in the Fear Conditioning paradigm, with treated Rett mice performing as well as wild-type controls, highlighting the procognitive properties of NLX-101. This work provides compelling evidence of the therapeutic potential of targeting post-synaptic 5-HT1A receptors to improve cognitive function in patients with RTT while supporting its respiratory-rescue properties.

Impaired bone health is a common morbidity in Rett syndrome (RTT). We aimed to assess lumbar bone mineral density (BMD) and trabecular bone score (TBS) in females with RTT, and to evaluate the effectiveness of bisphosphonate treatment.

This retrospective study included 40 females with RTT, aged 5-22 years, who underwent dual-energy X-ray absorptiometry (DXA) scans during 2019-2024 at a national center for RTT. Data collected included medical treatment, anthropometric measurements, and functional scores.

The median age at the first DXA scan was 10.8 years. The mean L1-4 BMD Z-score was -2.1 ± 1.4, and the mean TBS Z-score was -0.4 ± 1.3. The L1-4 BMD Z-score correlated with height (r = 0.407, p = 0.009), weight (r = 0.551, p < 0.001), BMI (r = 0.644, p < 0.001), and TBS Z-scores (r = 0.594, p = 0.009). Poor L1-4 BMD Z-scores were associated with poor mobility scores (p = 0.05) and valproate treatment (p = 0.016). Nine patients (23%) received zoledronate, for a mean 2 years. The mean age at zoledronate initiation was 9.7 ± 2.3 years. Four completed two DXA scans (pre- and post-treatment); the mean BMD Z-score improved from -2.2 ± 0.9 to -1.4 ± 0.9 after treatment.

Females with RTT have reduced lumbar BMD, which was associated with anthropometric factors, TBS, mobility, and valproate use. Zoledronate may be effective for some patients.

In a retrospective study of 40 females with Rett syndrome (RTT), low bone mineral density (BMD) correlated with lower anthropometric measurements, impaired mobility, and valproic acid use. The association between BMD and trabecular bone score (TBS) in the context of RTT is a novel finding. Our preliminary data support the effectiveness and safety of zoledronate for treating osteoporosis in patients with RTT. Our findings are important in light of the increasing life expectancy of individuals with RTT, and the consequent need to prioritize bone health in this population.

The evolving field of neuroepigenetics provides important insights into the molecular foundations of brain function. Novel sequencing technologies have identified patient-specific mutations and gene expression profiles involved in shaping the epigenetic landscape during neurodevelopment and in disease. Traditional methods to investigate the consequences of chromatin-related mutations provide valuable phenotypic insights but often lack information on the biochemical mechanisms underlying these processes. Recent studies, however, are beginning to elucidate how structural and/or functional aspects of histone, DNA, and RNA post-translational modifications affect transcriptional landscapes and neurological phenotypes. Here, we review the identification of epigenetic regulators from genomic studies of brain disease, as well as mechanistic findings that reveal the intricacies of neuronal chromatin regulation. We then discuss how these mechanistic studies serve as a guideline for future neuroepigenetics investigations. We end by proposing a roadmap to future therapies that exploit these findings by coupling them to recent advances in targeted therapeutics.

Defects in MKRN3, DLK1, KISS1, and KISS1R and some disorders, such as Temple syndrome (TS14), cause central precocious puberty (CPP). Recently, pathogenic variants (PVs) in MECP2 have been reported to be associated with CPP.

We aimed to clarify the contribution of (epi)genetic abnormalities to CPP and clinical and hormonal features in each etiology.

We conducted targeted sequencing for MKRN3, DLK1, MECP2, KISS1, and KISS1R and methylation analysis for screening of imprinting disorders such as TS14 associated with CPP in 90 patients with CPP (no history of brain injuries and negative brain magnetic resonance imaging) and collected their clinical and laboratory data. We measured serum DLK1 levels in 3 patients with TS14 and serum MKRN3 levels in 2 patients with MKRN3 genetic defects, together with some etiology-unknown patients with CPP and controls.

We detected 8 patients with TS14 (6, epimutation; 1, mosaic maternal uniparental disomy chromosome 14; 1, microdeletion) and 3 patients with MKRN3 genetic defects (1, PV; 1, 13-bp deletion in the 5'-untranslated region [5'-UTR]; 1, microdeletion) with family histories of paternal early puberty. There were no patients with PVs identified in MECP2, KISS1, or KISS1R. We confirmed low serum MKRN3 level in the patient with a deletion in 5'-UTR. The median height at initial evaluation of TS14 patients was lower than that of all patients. Six patients with TS14 were born small for gestational age (SGA).

(Epi)genetic causes were identified in 12.2% of patients with CPP at our center. For patients with CPP born SGA or together with family histories of paternal early puberty, (epi)genetic testing for TS14 and MKRN3 genetic defects should be considered.

Background/Objectives: Scoliosis is a prevalent comorbidity in Rett syndrome (RTT), often necessitating surgical intervention. This study investigated the impact of a 10-month individualized home exercise program (HEP) on scoliosis progression and gross motor function in girls aged six to 16 years with RTT. Methods: A multiple-baseline single-case design (AABA) was employed with 20 participants. A remotely supervised HEP, based on established principles focused on posture and physical activity, was implemented daily for at least one hour. The primary outcome was the rate of scoliosis progression assessed through the Cobb angle change measured via spinal radiographs at baseline, pre-intervention, and post-intervention. The secondary outcome was the gross motor function. Results: The HEP did not significantly reduce the rate of scoliosis progression. However, individual responses varied, with three participants showing scoliosis reduction. Significant improvements were observed in gross motor function, particularly in standing, walking, and stair-climbing abilities. Conclusions: The HEP did not significantly impact overall scoliosis progression, but a significant improvement was found in gross motor function. Further research into larger sample sizes is needed to confirm the effectiveness of exercise interventions in people with RTT.

Rett syndrome is a severe neurodevelopmental disorder that affects about 1 in 10,000 females. Clinical trials of disease modifying therapies are on the rise, but there are few psychometrically sound caregiver-reported outcome measures available to assess treatment benefit. We report on a new caregiver-reported outcome measure, the Rett Caregiver Assessment of Symptom Severity (RCASS). Using data from the Rett Natural History Study (n = 649), we examined the factor structure, using both exploratory and confirmatory factor analysis, and the reliability and validity of the RCASS. The four-factor model had the best overall fit, which covered movement, communication, behavior, and Rett-specific symptoms. The RCASS had moderate internal consistency. Strong face validity was found with age and mutation type, and convergent validity was established with other similar measures, including the Revised Motor-Behavior Assessment Scale, Clinical Severity Scale, Clinical Global Impression Scale, and the Child Health Questionnaire. These data provide initial evidence that the RCASS is a viable caregiver-outcome measure for use in clinical trials in Rett syndrome. Future work to assess sensitivity to change and other measures of reliability, such as test-retest and inter-rater agreement, are needed.

Epigenetics refers to heritable changes in gene expression or phenotypic changes that occur without changing the gene sequence. The main methods of epigenetics include non-coding RNA, histone modification, and DNA modification, which play an essential role in gene expression regulation and even the occurrence of diverse diseases. Naringenin, the aglycone form of naringin, is a natural flavonoid compound mainly found in fruits or plant derivatives such as citrus, tomatoes, and cherries. Naringenin and naringin exhibit a broad spectrum of biological activities and pharmacological effects, including anti-cancer, cardiovascular disease improving, anti-inflammatory, and anti-oxidant activities, all of which are advantageous for human health. Recent studies have uncovered that naringenin and naringin influence gene expression by modulating epigenetic pathways, including microRNA (miRNA) regulation. This mechanism plays a crucial role in the therapeutic potential for various diseases. This paper reviews the epigenetic researches on the physiological activities of naringenin and naringin. It highlights how these compounds can exert diverse effects through different signaling pathways, thereby ameliorating associated diseases. These findings provide valuable insights for the future applications of naringenin and naringin.

Rett syndrome (RTT) is a rare neurodevelopmental disorder that mainly affects girls and women. Trofinetide is approved for the treatment of RTT in adults and children aged ≥2 years. To gain insight into experiences with RTT and effects of trofinetide treatment at different stages of RTT, interviews with caregivers of individuals with RTT were conducted upon their exit from the open-label trofinetide trials.

Interviews were conducted with caregivers of participants in the LILAC/LILAC-2 open-label extension trials of the phase 3 LAVENDER trial in participants aged 5 to 20 years, and in DAFFODIL, an open-label trial in participants aged 2 to 4 years. Caregivers were asked about the RTT effects, experiences with trofinetide, meaningfulness of treatment effects, and satisfaction. Qualitative thematic analysis was performed.

Caregivers of 33 participants from the open-label trials were interviewed, including 26 from LILAC/LILAC-2 (mean age, 12.3 years) and 7 from DAFFODIL (mean age, 4.5 years). The most commonly reported effects of RTT in LILAC/LILAC-2 were no verbal communication (24/26 [92.3%]), unable to use hands (15/26 [57.7%]), repetitive hand movements (15/26 [57.7%]), unable to walk (15/26 [57.7%]), and seizures (14/26 [53.8%]). In DAFFODIL, the most commonly reported effects of RTT were no verbal communication (7/7 [100%]), impaired balance (4/7 [57.1%]), unable to use hands (3/7 [42.9%]), repetitive hand movements (3/7 [42.9%]), mood disturbance (3/7 [42.9%]), constipation (3/7 [42.9%]), and limited ability to use hands (3/7 [42.9%]). Caregivers most commonly reported improvements in hand use (11/26 [42.3%]), engagement with others (11/26 [42.3%]), eye gaze (8/26 [30.8%]), use of the Tobii eye tracking device (7/26 [26.9%]), and attention/focus/concentration (7/26 [26.9%]) in LILAC/LILAC-2. In DAFFODIL, caregivers reported improvements in new words (5/7 [71.4%]), hand use (4/7 [57.1%]), and eye contact (4/7 [57.1%]). Nearly all (31/32) caregivers were very satisfied or satisfied with trofinetide.

Caregivers of participants in open-label trofinetide trials reported improvements in RTT with meaningful impact in areas of motor function, communication, and engagement.

Autism spectrum disorder (ASD) is a complex neuro developmental condition characterized by significant genetic and phenotypic variability, making diagnosis and treatment challenging. The heterogeneity of ASD-associated genetic variants and the absence of clear causal factors in many cases complicate personalized care. Traditional models, such as postmortem brain tissue and animal studies, have provided valuable insights but are limited in capturing the dynamic processes and human-specific aspects of ASD pathology. Recent advances in human induced pluripotent stem cell (iPSC) technology have transformed ASD research by enabling the generation of patient-derived neural cells in both two-dimensional cultures and three-dimensional brain organoid models. These models retain the donor's genetic background, allowing researchers to investigate disease-specific cellular and molecular mechanisms while identifying potential therapeutic targets tailored to individual patients. This commentary highlights how stem cell-based approaches are advancing our understanding of ASD and paving the way for more personalized diagnostic and therapeutic strategies.

3D organization of the genome plays a critical role in regulating gene expression. How 3D-genome organization differs among different cell types and relates to cell type-dependent transcriptional regulation remains unclear. Here, we used genome-scale DNA and RNA imaging to investigate 3D-genome organization in transcriptionally distinct cell types in the mouse cerebral cortex. We uncovered a wide spectrum of differences in the nuclear architecture and 3D-genome organization among different cell types, ranging from the size of the cell nucleus to higher-order chromosome structures and radial positioning of chromatin loci within the nucleus. These cell type-dependent variations in nuclear architecture and chromatin organization exhibit strong correlations with both the total transcriptional activity of the cell and transcriptional regulation of cell type-specific marker genes. Moreover, we found that the methylated DNA binding protein MeCP2 promotes active-inactive chromatin segregation and regulates transcription in a nuclear radial position-dependent manner that is highly correlated with its function in modulating active-inactive chromatin compartmentalization.

This retrospective study compared the speech-language development of a pair of dizygotic twin girls during the first 2 years of life: one with typical development (Twin A) and one with atypical development (Twin B), who was later diagnosed with Rett syndrome (RTT). Audio snippets were extracted from home videos, with nearly equal representation from each child. The audio analysis focused on articulatory complexity, voice characteristics, and linguistic variability. Despite sharing the same social-communicative environment, the twins' speech-language development diverged. From the first to second year of life, articulatory complexity and variability increased in the typically developing Twin A. In contrast, Twin B produced a high number of vocalizations in the 7th month, including canonical sounds with substantial variability. However, her vocalization quantity, complexity, and variability subsequently decreased, resulting in only sporadically discernable canonical vocalizations during her second year. This developmental trajectory points to very early stagnation and regression in Twin B, occurring earlier than typically observed. While Twin B displayed a range of typical vocalization features, deviations in the density and distribution of inspiratory and high-pitched vocalizations during the first year further suggest early speech-language abnormalities in RTT, preceding frank developmental stagnation and regression. As the study relied on limited retrospective data, the findings should be interpreted with caution, and further investigation is needed. Nevertheless, this twin study provides a unique perspective that deepens our understanding of early speech-language developmental profiles in RTT, especially in light of the intertwinement of genetic, individual, and contextual factors.

Children with Christianson syndrome (CS), an X-linked neurodevelopmental disorder caused by loss-of-function mutations in the alkali cation/proton exchanger SLC9A6/NHE6, display severe cognitive impairments, mutism, and sensory abnormalities such as hyposensitivity to pain. However, it is unclear whether these children display other sensory abnormalities and whether their pain hyposensitivity is the result of an elevated pain threshold or a complete insensitivity to pain. To better characterize the sensory abnormalities in this disorder, we used a combination of a mouse model of CS and pain questionnaires directed at nonverbal patients with CS. We recruited 14 young male participants with CS and subjected them to a novel observational tool, the Pain Sensory and Painful Situations Questionnaire (PSQ), which takes multiple painful situations into account to broaden the description of pain expression. By analyzing social expressive behaviours of pain in these nonverbal patients, the PSQ documented that over 60% of the participants were unaffected by mechanical or inflammatory painful stimuli. This reduced pain sensitivity was also observed in the mouse CS model. Surprisingly, CS mice also displayed aversive reactions to innocuous stimuli, which prompted us to examine whether such reactions were also present in children with CS. Indeed, the results from the PSQ revealed that 30% to 50% of these patients showed an aversive response to normally innocuous stimuli like light touch and gusts of air. Our results demonstrate that children with CS have aversive reactions to innocuous stimuli and are hyposensitive to painful stimuli, the latter making them at risk for developing complications from unreported injuries.

Rett Syndrome, a neurodevelopmental disorder caused by loss-of-function mutations in the MECP2 gene, is characterized by severe motor, cognitive and emotional impairments. Some of the deficits may result from changes in cortical connections, especially downstream projections of the prefrontal cortex, which may also be targets of restoration following rearing conditions such as environmental enrichment that alleviate specific symptoms. Here, using a heterozygous Mecp2 +/- female mouse model closely analogous to human Rett Syndrome, we investigated the impact of early environmental enrichment on behavioral deficits and prefrontal cortex connectivity. Behavioral analyses revealed that enriched housing rescued fine motor deficits and reduced anxiety, with enrichment-housed Mecp2 +/- mice performing comparably to wild-type (WT) controls in rotarod and open field assays. Anatomical mapping of top-down anterior cingulate cortex (ACA) projections demonstrated altered prefrontal cortex connectivity in Mecp2 +/- mice, with increased axonal density in the somatosensory cortex and decreased density in the motor cortex compared to WT controls. ACA axons revealed shifts in hemispheric distribution, particularly in the medial network regions, with Mecp2 +/- mice exhibiting reduced ipsilateral dominance. These changes were unaffected by enriched housing, suggesting that structural abnormalities in prefrontal cortex connectivity persist despite behavioral improvements. Enriched housing rescued brain-derived neurotrophic factor (BDNF) levels in the hippocampus but failed to restore BDNF levels in the prefrontal cortex, consistent with the persistent deficits observed in prefrontal axonal projections. These findings highlight the focal nature of changes induced by reduction of MeCP2 and by exposure to environmental enrichment, and suggest that environmental enrichment starting in adolescence can alleviate behavioral deficits without reversing abnormalities in large-scale cortical connectivity.

Rett syndrome (RTT) is a severe neurodevelopmental disorder primarily caused by loss-of-function mutations in the MECP2 gene, resulting in diverse cellular dysfunctions. Here, we investigated the role of the long noncoding RNA (lncRNA) NEAT1 in the context of MeCP2 deficiency using human neural cells and RTT patient samples. Through single-cell RNA sequencing and molecular analyses, we found that NEAT1 is markedly downregulated in MECP2 knockout (KO) cells at various stages of neural differentiation. NEAT1 downregulation correlated with aberrant activation of the mTOR pathway, abnormal protein metabolism, and dysregulated autophagy, contributing to the accumulation of protein aggregates and impaired mitochondrial function. Reactivation of NEAT1 in MECP2-KO cells rescued these phenotypes, indicating its critical role downstream of MECP2. Furthermore, direct RNA-RNA interaction was revealed as the key process for NEAT1 influence on autophagy genes, leading to altered subcellular localization of specific autophagy-related messenger RNAs and impaired biogenesis of autophagic complexes. Importantly, NEAT1 restoration rescued the morphological defects observed in MECP2-KO neurons, highlighting its crucial role in neuronal maturation. Overall, our findings elucidate lncRNA NEAT1 as a key mediator of MeCP2 function, regulating essential pathways involved in protein metabolism, autophagy, and neuronal morphology.

Objective: Rett syndrome (RTT) is a severe neurodevelopmental disorder affecting predominantly females and associated with variants in the MECP2 gene. Recent success in clinical trials have resulted in an expanded use of the Rett Syndrome Behaviour Questionnaire (RSBQ) for clinical and research purposes. Implementation of the RSBQ as a global clinical severity scale has raised concerns about its construct validity considering its content, structure, and psychometric features. To further understand RSBQ data, we analyzed RSBQ scores available in the literature with a focus on variability and influencing factors. Methods: We identified publications reporting RSBQ total and/or subscale scores and summarized relevant study information, such as type of investigation, administration method, and descriptive data. We then analyzed means and standard deviations, calculating variance-to-mean ratios (VMR), as a measure of variability, when raw score descriptive statistics were available. Where appropriate, we compared means and VMRs by Welch t-tests. Results: Of the 14 publications identified, raw total scores from 5 observational studies and 4 clinical trials (baseline) were available. Raw subscale scores from four of the five observational studies were also available. We found a wide but comparable range of mean total scores for observational studies and clinical trials. However, VMRs were significantly higher in observational studies. Subscale scores showed either high (i.e., General Mood, Breathing Problems) or low (e.g., Hand Behaviours, Body Rocking and Expressionless Face) variability. Available data demonstrated greater variability in pediatric than adult groups and less variability when using interviews or electronic RSBQ administration compared with paper forms. Total score changes over time did not affect variability. Although certain studies offered insight into the relationship between the RSBQ and other measures, overall, data were insufficient for characterizing how RSBQ variability relates to other factors. Conclusions: Our findings on score variability support the need for more comprehensive reporting of RSBQ data, cohort characterization, and methodology; and the deployment of standardized RSBQ administration methods, such as advanced data capture systems. There is potential for use of subscales as outcome measures, subject to further psychometric validation studies, including prospective investigations testing the stability of RSBQ scores and influencing factors. Further examining the relationship between RSBQ scores and other instruments will aid in its interpretation as a clinical outcome measure.

Mutations in the X-linked methyl-CpG-binding protein 2 (MECP2) gene cause Rett syndrome, a severe childhood neurological disorder. MeCP2 is a well-established transcriptional repressor, yet upon its loss, hundreds of genes are dysregulated in both directions. To understand what drives such dysregulation, we deleted Mecp2 in adult mice, circumventing developmental contributions and secondary pathogenesis. We performed time series transcriptional, chromatin, and phenotypic analyses of the hippocampus to determine the immediate consequences of MeCP2 loss and the cascade of pathogenesis. We find that loss of MeCP2 causes immediate and bidirectional progressive dysregulation of the transcriptome. To understand what drives gene downregulation, we profiled genome-wide histone modifications and found that a decrease in histone H3 acetylation (ac) at downregulated genes is among the earliest molecular changes occurring well before any measurable deficiencies in electrophysiology and neurological function. These data reveal a molecular cascade that drives disease independent of any developmental contributions or secondary pathogenesis.

There is an increasing body of evidence suggesting that a single exposure to certain chemicals can have transgenerational effects, with the underlying mechanism believed to be epigenetic. However, it remains largely unknown whether psychiatric conditions like ADHD or autism, induced by environmental chemicals, can be inherited across generations. Pregnant rats were purchased from a commercial breeder. On the 7th day of gestation (E7), they were divided into two groups: one group was orally exposed to silver nanoparticles (AgNP; 4 mg/kg), while the control group received vehicle alone. The subsequent generation (F1) underwent spontaneous motor activity (SMA) measurements at 8-11 weeks of age. For breeding at 26 weeks of age, rats with higher SMA were selected from hyperactive litters, while untreated rats were randomly selected. This process was continued for four generations in both groups. The AgNP-primed rats at 4th generation displayed significantly higher SMA, 1.8 times greater than that of untreated rats. Intraperitoneal injection of valproic acid (150 mg/kg), an epigenetic modifier to 5-day-old rats causes adult hyperactivity (1.4-fold), suggesting that epigenetic modification contributes to rat hyperactivity. Global DNA methylation profiles in the mesencephalon were positively correlated in both groups of hyperactive rats. Furthermore, there were 7-8 common genes showing both hypermethylation and hypomethylation, which are involved in neuronal development, neuronal function, transcriptional activity, DNA binding activity, cell differentiation, ubiquitination processes, or histone modification, including Pax 6 and Mecp 2. Thus, it is most likely that rats retain hyperactivity through mesencephalic DNA methylation status across transgeneration.

Normal brain functioning relies on high aerobic energy production provided by mitochondria. Failure to supply a sufficient amount of energy, seen in different brain disorders, including autism spectrum disorder (ASD), may have a significant negative impact on brain development and support of different brain functions. Mitochondrial dysfunction, manifested in the abnormal activities of the electron transport chain and impaired energy metabolism, greatly contributes to ASD. The aberrant functioning of this organelle is of such high importance that ASD has been proposed as a mitochondrial disease. It should be noted that aerobic energy production is not the only function of the mitochondria. In particular, these organelles are involved in the regulation of Ca2+ homeostasis, different mechanisms of programmed cell death, autophagy, and reactive oxygen and nitrogen species (ROS and RNS) production. Several syndromes originated from mitochondria-related mutations display ASD phenotype. Abnormalities in Ca2+ handling and ATP production in the brain mitochondria affect synaptic transmission, plasticity, and synaptic development, contributing to ASD. ROS and Ca2+ regulate the activity of the mitochondrial permeability transition pore (mPTP). The prolonged opening of this pore affects the redox state of the mitochondria, impairs oxidative phosphorylation, and activates apoptosis, ultimately leading to cell death. A dysregulation between the enhanced mitochondria-related processes of apoptosis and the inhibited autophagy leads to the accumulation of toxic products in the brains of individuals with ASD. Although many mitochondria-related mechanisms still have to be investigated, and whether they are the cause or consequence of this disorder is still unknown, the accumulating data show that the breakdown of any of the mitochondrial functions may contribute to abnormal brain development leading to ASD. In this review, we discuss the multifaceted role of mitochondria in ASD from the various aspects of neuroscience.

Bone is a unique organ crucial for locomotion, mineral metabolism, and hematopoiesis. It maintains homeostasis through a balance between bone formation by osteoblasts and bone resorption by osteoclasts, which is regulated by the basic multicellular unit (BMU). Abnormal bone metabolism arises from an imbalance in the BMU. Osteoclasts, derived from the monocyte-macrophage lineage, are regulated by the RANKL-RANK-OPG system, which is a key factor in osteoclast differentiation. RANKL activates osteoclasts through its receptor RANK, while OPG acts as a decoy receptor that inhibits RANKL. In trabecular bone, high turnover involves rapid bone formation and resorption, influenced by conditions such as malignancy and inflammatory cytokines that increase RANKL expression. Cortical bone remodeling, regulated by aged osteocytes expressing RANKL, is less understood, despite ongoing research into how Rett syndrome, characterized by MeCP2 abnormalities, affects RANKL expression. Balancing trabecular and cortical bone involves mechanisms that preserve cortical bone, despite overall bone mass reduction due to aging or oxidative stress. Research into genes like sFRP4, which modulates bone mass, highlights the complex regulation by BMUs. The roles of the RANKL-RANK-OPG system extend beyond bone, affecting processes such as aortic valve formation and temperature regulation, which highlight the interconnected nature of biological research.

MECP2 variants cause X-chromosome-linked rare developmental syndromes. Typically, the mutation is sporadic, occurs in females and is fatal in men. Accurate genetic and clinical diagnostics are considered essential for the management of symptoms and the development of new treatments. These aims may be difficult to reach before more is known about factors resulting in highly variable clinical pictures among patients carrying the same MECP2 variant. We describe the clinical picture of two brothers carrying the same MECP2 variant and compare them with cases published in the literature.

Most of the MECP2 mutations are known to be de novo mutations, which is why the recurrence of the mutation in the couple's other children is unlikely. Unexpectedly, our routine genetic testing revealed a 23-year-old man (P1) and his younger brother (P2) to carry the same hemizygous pathogenic missense variant c.419C>T, p.(Ala140Val) (transcript NM_004992.3) of MECP2, which was found to be inherited from their presumably asymptomatic mother. Thus, further clinical evaluation and comparison with literature cases was considered necessary.

The P1 has a severe syndromic intellectual disorder (ID), whereas his brother has a substantially milder ID predominantly limited to problems in verbal skills. Neither P1 nor his younger brother has been diagnosed with Rett syndrome. The P1 (unlike his younger brother) has several lingual, social and motor difficulties; disruptive behavior was the most difficult symptom to treat. P1's response to several medical and non-medical treatment trials has remained inadequate, thus requiring the patient to be hospitalised for a long time. The literature review revealed that apart from our family, there are five other families with more than one male carrying the same MECP2 p.Ala140Val mutation, such as P1 and P2. The phenotypes of all 24 men from us (n = 2) and others (n = 22) carrying the same, presumably non-lethal mutation show great variability.

The p.Ala140Val mutation of MECP2 in males is associated with a rare X-chromosomal developmental disorder with highly variable phenotypes. Further studies are needed to better understand all those influencing factors that can explain phenotypic differences within the same genotype to find optimal medicinal therapies.

Rett syndrome (RTT) and MECP2 duplication syndrome (MDS) result from under- and overexpression of MECP2, respectively. Preclinical studies using genetic-based treatment showed robust phenotype recovery for both MDS and RTT. However, there is a risk of converting MDS to RTT, or vice versa, if accurate MeCP2 levels are not achieved. The aim of this study was to identify biomarkers distinguishing RTT from MDS.

We prospectively enrolled 11 MDS and 6 male RTT like (MRL) individuals for a panel of clinical and neurophysiological assessments over two visits, 8-10 months apart.

We identified numerous clinical and physiological features as promising biomarkers. MRL individuals exhibited large amplitude whole body tremor, midline stereotypies (vs. hand flapping at sides in MDS), earlier neuromotor regression, and earlier onset but less commonly refractory epilepsy. In the neurophysiological domain, we observed several marked differences in sleep physiology between MDS/MRL and typically developing (TD) individuals including reduced sleeping time, increased delta power during rapid eye movement (REM) sleep, decreased occipital alpha and increased brain-wide delta power during wakefulness, and reduced spindle density and duration. MRL individuals also had much lower delta power during NREM 2 and 3 stages than the TD group. We found differences in spindle duration in the temporal lobes and spindle amplitude in the frontal lobes between MDS and MRL.

Our study revealed distinct clinical features of MDS and MRL that can be monitored during a clinical trial and may serve as target engagement, disease progression, or safety biomarkers for interventional studies.

Weight-banded trofinetide dosing improved physician- and caregiver-rated efficacy measures and had acceptable tolerability in patients aged 2‒4 years (DAFFODIL study) and 5‒20 years (LAVENDER study) with Rett syndrome (RTT). Selection of weight-banded dosing regimens for these studies was based on population pharmacokinetic (popPK) modeling and exposure simulations. This study applied an updated popPK model to confirm steady-state trofinetide exposures achieved in DAFFODIL patients were within target range.

A popPK model was developed using data from 14 clinical studies of trofinetide in healthy volunteers and pediatric and adult patients, including the LAVENDER and DAFFODIL studies. Individual exposure measures (area under concentration-time curve over 0-12 h [AUC0-12] were generated via integration of the predicted concentration-time profile for each DAFFODIL study participant based on the popPK model and individual empiric Bayesian pharmacokinetic parameter estimates. Distributions of steady-state AUC0-12 values for each body-weight group were compared against target exposure (AUC0-12 = 800‒1200 µg·h/mL).

Distribution and box plots of simulated steady-state AUC0-12 values achieved with the weight-banded DAFFODIL dosing regimen used in younger individuals aged 2-4 years with RTT (twice daily trofinetide 5 g [≥ 9 to < 12 kg] or 6 g [≥ 12 to < 20 kg]) indicated good overlap with the target exposure range. Median steady-state AUC0-12 values for both body-weight bands fell within the target exposure range.

PopPK model-based simulations confirm that the weight-banded dosing regimen used in DAFFODIL is adequate to achieve target trofinetide exposure in 2- to 4-year-olds with RTT.

Mutations of the MECP2 gene lead to Rett syndrome (RTT), a rare developmental disease causing severe intellectual and physical disability. How the loss or defective function of MeCP2 mediates RTT is still poorly understood. MeCP2 is a global gene expression regulator, acting at transcriptional and post-transcriptional levels. Little attention has been given so far to the contribution of alternative splicing (AS) dysregulation to RTT pathophysiology. To perform a comparative analysis of publicly available RNA sequencing (RNA-seq) studies and generate novel data resources for AS, we explored 100 human datasets and 130 mouse datasets from Mecp2-mutant models, processing data for gene expression and alternative splicing. Our comparative analysis across studies indicates common species-specific differentially expressed genes (DEGs) and differentially alternatively spliced (DAS) genes. Human and mouse dysregulated genes are involved in two main functional categories: cell-extracellular matrix adhesion regulation and synaptic functions, the first category more significantly enriched in human datasets. Our extensive bioinformatics study indicates, for the first time, a significant dysregulation of AS in human RTT datasets, suggesting the crucial contribution of altered RNA processing to the pathophysiology of RTT.

Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease associated with microcephaly and poor neurodevelopmental outcomes. Here we show that the Ohia HLHS mouse model, with mutations in Sap130, a chromatin modifier, and Pcdha9, a cell adhesion protein, also exhibits microcephaly associated with mitotic block and increased apoptosis leading to impaired cortical neurogenesis. Transcriptome profiling, DNA methylation, and Sap130 ChIPseq analyses all demonstrate dysregulation of genes associated with autism and cognitive impairment. This includes perturbation of REST transcriptional regulation of neurogenesis, disruption of CREB signaling regulating synaptic plasticity, and defects in neurovascular coupling mediating cerebral blood flow. Adult mice harboring either the Pcdha9 mutation, which show normal brain anatomy, or forebrain-specific Sap130 deletion via Emx1-Cre, which show microcephaly, both demonstrate learning and memory deficits and autism-like behavior. These findings provide mechanistic insights indicating the adverse neurodevelopment in HLHS may involve cell autonomous/nonautonomous defects and epigenetic dysregulation.

Neurodevelopmental disorders (NDD) encompass a range of conditions marked by abnormal brain development in conjunction with impaired cognitive, emotional and behavioural functions. Transgenic animal models, mainly rodents, traditionally served as key tools for deciphering the molecular mechanisms driving NDD physiopathology and significantly contributed to the development of pharmacological interventions aimed at treating these disorders. However, the efficacy of these treatments in humans has proven to be limited, due in part to the intrinsic constraint of animal models to recapitulate the complex development and structure of the human brain but also to the phenotypic heterogeneity found between affected individuals. Significant advancements in the field of induced pluripotent stem cells (iPSCs) offer a promising avenue for overcoming these challenges. Indeed, the development of advanced differentiation protocols for generating iPSC-derived brain organoids gives an unprecedented opportunity to explore human neurodevelopment. This review provides an overview of how 3D brain organoids have been used to investigate various NDD (i.e. Fragile X syndrome, Rett syndrome, Angelman syndrome, microlissencephaly, Prader-Willi syndrome, Timothy syndrome, tuberous sclerosis syndrome) and elucidate their pathophysiology. We also discuss the benefits and limitations of employing such innovative 3D models compared to animal models and 2D cell culture systems in the realm of personalized medicine.

Human puberty is a dynamic biological process determined by the increase in the pulsatile secretion of GnRH triggered by distinct factors not fully understood. Current knowledge reveals fine tuning between an increase in stimulatory factors and a decrease in inhibitory factors, where genetic and epigenetic factors have been indicated as key players in the regulation of puberty onset by distinct lines of evidence. Central precocious puberty (CPP) results from the premature reactivation of pulsatile secretion of GnRH. In the past decade, the identification of genetic causes of CPP has largely expanded, revealing hypothalamic regulatory factors of pubertal timing. Among them, 3 genes associated with CPP are linked to mechanisms involving DNA methylation, reinforcing the strong role of epigenetics underlying this disorder. Loss-of-function mutations in Makorin Ring-Finger Protein 3 (MKRN3) and Delta-Like Non-Canonical Notch Ligand 1 (DLK1), 2 autosomal maternally imprinted genes, have been described as relevant monogenic causes of CPP with the phenotype exclusively associated with paternal transmission. MKRN3 has proven to be a key component of the hypothalamic inhibitory input on GnRH neurons through different mechanisms. Additionally, rare heterozygous variants in the Methyl-CpG-Binding Protein 2 (MECP2), an X-linked gene that is a key factor of DNA methylation machinery, were identified in girls with sporadic CPP with or without neurodevelopmental disorders. In this mini-review, we focus on how the identification of genetic causes of CPP has revealed epigenetic regulators of human pubertal timing, summarizing the latest knowledge on the associations of puberty with MKRN3, DLK1, and MECP2.

Rett syndrome (RS) is a rare neurodevelopmental disorder primarily caused by mutations in the X-linked methyl-CpG binding protein 2 (MECP2) gene, responsible for encoding MECP2 which plays a pivotal role in regulating gene expression. The neurological and non-neurological manifestations of RS vary widely in severity depending on the specific mutation type. Bone complications, mostly scoliosis but also osteoporosis, hip displacement, and a high rate of fractures, are among the most prevalent non-neurological comorbidities observed in girls with RS. Low bone mineral density (BMD) is primarily due to a slow rate of bone formation due to dysfunctional osteoblast activity. The use of anticonvulsants, immobilization, low physical activity, poor nutrition, and inadequate vitamin D intake all significantly hamper skeletal maturation and the accumulation of bone mass in RS girls, making them more susceptible to fragility fractures. In RS patients, the upper and lower limbs are the most common sites for fractures which are due to both a reduced BMD and a diminished bone size. This review summarizes the knowledge on risk factors for fragility fracture in patients with RS and proposes a potential diagnostic and therapeutic pathway to enhance low BMD and mitigate the risk of fragility fractures. In particular, this review focused on the importance of clinical and instrumental evaluation of bone status as a basis for adequate planning of nutritional, pharmacological, and surgical interventions to be undertaken. Additionally, the management of bone defects in individuals with RS should be customized to meet each person's specific needs, abilities, and general health.

Methyl-CpG-binding protein 2 (MeCP2) is a ubiquitously expressed nuclear protein involved in transcriptional regulation and chromatin remodeling. MeCP2 exists in two isoforms, MeCP2 E1 and E2, which share the same functional domains. Loss-of-function mutations in the MeCP2 gene are the main cause of Rett syndrome (RTT). Previous studies identified a complex formation between MeCP2 and lens epithelium derived growth factor (LEDGF), a transcriptional regulator that exists in two isoforms, LEDGF/p75 and LEDGF/p52. Here, we characterized the molecular and functional interaction between MeCP2 and LEDGF. The NCoR interaction domain (NID) domain in MeCP2 is essential for the direct binding to the PWWP-CR1 region of LEDGF. Introduction of R306C, an RTT mutation in the NID of MeCP2, reduced the interaction with LEDGF. Our data reveal mutual inhibition of MeCP2 and LEDGF multimerization due to overlapping binding sites. Aligning with this observation, LEDGF depletion resulted in larger MeCP2 and DNA foci in NIH3T3 cells, suggesting a role for the MeCP2-LEDGF complex in chromatin organization.

Ampakines are a class of compounds that are positive allosteric modulators of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and enhance glutamatergic neurotransmission. Glutamatergic synaptic transmission and AMPA receptor activation are fundamentally important to the genesis and propagation of the neural impulses driving breathing, including respiratory motoneuron depolarization. Ampakines therefore have the potential to modulate the neural control of breathing. In this paper, we describe the influence of ampakines on respiratory motor output in health and disease. We dissect the molecular mechanisms underlying ampakine action, delineate the diverse targets of ampakines along the respiratory neuraxis, survey the spectrum of respiratory disorders in which ampakines have been tested, and culminate with an examination of how ampakines modulate respiratory function after spinal cord injury. Collectively, the studies reviewed here indicate that ampakines may be a useful adjunctive strategy to pair with conventional respiratory rehabilitation approaches in conditions with impaired neural activation of the respiratory muscles.

The cases of head and neck cancer among persons with intellectual disability (PWID) are infrequently reported and therefore poorly understood. PWID often face increased barriers of access to healthcare, which can be further compounded when faced with a cancer diagnosis. This report presents the case of a 34-year-old Chinese female patient with Rett syndrome and intellectual disability, presenting with two primary cancers of the tongue and the trachea. The case highlights the importance of regular dental review in cancer surveillance, post-operative supportive dental care, and challenges in behavioral management in cancer diagnosis and treatment.