Impaired astrocyte-to-neuron cholesterol trafficking drives synaptic dysfunction in Rett syndrome

Selective reactivation of Mecp2 expression in astrocytes of Mecp2 knock-out mice results in significant rescue of behavioral and neuronal phenotype.

“selective reactivation of Mecp2 expression in astrocytes of Mecp2 KO mice resulted in a significant rescue of both behavioral and neuronal phenotype”

• cites Lioy et al., 2011 sourcePartly supported

  The passages on behavior and neuronal anatomy report improved behavioral measures and restored dendritic branching and other neuronal measures after astrocytic MeCP2 re-expression. However, the experiments used Mecp2Stop mice, not complete Mecp2 knockouts: the paper notes residual MeCP2 expression from the stop allele and distinguishes these mice from Mecp2-/y mice. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Lipidomic analyses revealed reduced cholesterol levels in cerebrospinal fluid, but not plasma, of Rett syndrome patients compared with age-matched healthy controls.

“Lipidomic analyses performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) on CSF and plasma of RTT patients revealed reduced cholesterol levels in the CSF, but not in plasma, compared with agematched healthy controls (Zandl-Lang et al., 2022).”

• cites Zandl-Lang et al., 2022 sourceSupported

MECP2 mutant astrocytes contribute to neuronal alterations through non-cell-autonomous mechanisms.

“and MECP2 mutant astrocytes contribute to neuronal alterations through non-cell-autonomous mechanisms (Albizzati et al., 2024;Ballas et al., 2009;Sun et al., 2023;Williams et al., 2014).”

• cites Albizzati et al., 2024 sourceSupported
• cites Ballas et al., 2009 sourceSupported
• cites Sun et al., 2023 sourceSupported

Mecp2 KO astrocytes influence both synaptogenesis and synapse maintenance through release of synaptotoxic molecules and/or reduced secretion of synaptogenic factors.

“Indeed, through the release of synaptotoxic molecules and/or the reduced secretion of synaptogenic factors, Mecp2 KO astrocytes influence both synaptogenesis and synapse maintenance (Albizzati et al., 2024;Sun et al., 2023).”

• cites Albizzati et al., 2024 sourcePartly supported

  The Results section reports that molecules secreted by Mecp2 KO cortical astrocytes impair synaptogenesis in WT neurons, and the Discussion proposes toxic molecules and/or reduced synaptogenic cues as possible mechanisms. The paper does not establish an effect on the maintenance of existing synapses, so that part of the claim is unsupported. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Sun et al., 2023 sourcePartly supported

  The Results section on astrocyte-dependent neuronal structural plasticity (page unavailable) reports that MECP2-mutant astrocytes enhanced synapse formation less efficiently than wild-type astrocytes. It does not establish an effect on synapse maintenance or show that synaptotoxic release or reduced secretion of synaptogenic factors caused the difference. The study describes MECP2-mutant human astrocytes, not specifically Mecp2-knockout astrocytes. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Lcat is significantly down-regulated in the Mecp2 KO cortex.

“In addition, our previous study reported a significant down-regulation of Lcat in the Mecp2 KO cortex (Albizzati et al., 2022).”

• cites Albizzati et al., 2022 sourceSupported

Intracellular cholesterol accumulation provides a potential explanation for the reduced nuclear localization of Srebp2 in Mecp2 KO astrocytes, given negative feedback.

“Notably, the accumulation of intracellular cholesterol provides a potential explanation for the reduced nuclear localization of Srebp2 observed in KO astrocytes, given the negative feedback exerted by intracellular cholesterol on Srebp2 activation (Duan et al., 2022;N.-Q. Wang et al., 2025).”

• cites Duan et al., 2022 sourceSupported
• cites N.-Q. Wang et al., 2025 sourceNot checked — no full text of the source

Cholesterol treatment recovers pre- and post-synaptic defects in Mecp2 HET neurons.

“By measuring the density of synaptic puncta, we proved that cholesterol treatment recovers pre-and post-synaptic defects, typically shown by HET neurons (Frasca et al., 2024) (Figure 5A-D).”

• cites Frasca et al., 2024 sourcePartly supported

  In the Results section on NPC-secreted factors (Fig. 1J–K; page unavailable), Frasca et al. report reduced presynaptic puncta in Mecp2 Het neurons. The cited paper documents postsynaptic puncta defects in Mecp2 KO neurons, but does not establish the corresponding puncta defect in Het neurons. The cholesterol-treatment result is presented as the citing authors’ own finding, not as a finding attributed to Frasca et al. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Nsdhl protein levels are reduced by approximately 50% in the brain of KI Mecp2 Y120D/y mouse model.

“Importantly, Nsdhl protein levels were similarly reduced by approximately 50% in the brain of the KI Mecp2 Y120D/y mouse model, which carries a phospho-mimetic missense mutation and phenotypically recapitulates the null mouse (Supplementary Figure 6A) (D'Annessa et al., 2018;Gandaglia et al., 2019).”

• cites D'Annessa et al., 2018 sourceCould not verify

  Section 3.3 (pp. 9–10) tests DNA binding by MeCP2 Y120D protein fragments, not Nsdhl levels in mouse brain. Searches for Nsdhl and the Y120D knock-in mouse model found no passage reporting an approximately 50% reduction. (LLM, conf=0.78)

  Experimental — accuracy not yet measured
• cites Gandaglia et al., 2019Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Intracellular cholesterol inhibits Srebp2 activation through a negative feedback mechanism.

“However, intracellular cholesterol is known to inhibit Srebp2 activation through a negative feedback mechanism (Duan et al., 2022;Wang et al., 2025).”

• cites Duan et al., 2022 sourceSupported
• cites Wang et al., 2025 sourceNot checked — no full text of the source

There is accumulating evidence of altered brain cholesterol metabolism in RTT.

“as well as by accumulating evidence of altered brain cholesterol metabolism in RTT (Ben-Shachar et al., 2009;Buchovecky et al., 2013b;Lopez et al., 2017;Lütjohann et al., 2018;Urdinguio et al., 2008;Villani et al., 2016;Zandl-Lang et al., 2022).”

• cites Ben-Shachar et al., 2009 sourceCould not verify

  The Results describe altered gene expression in the cerebellum and hypothalamus of MeCP2-disorder mouse models, including a change involving a low-density-lipoprotein-related receptor. They do not say that brain cholesterol metabolism is altered in RTT. Searches for cholesterol, lipid metabolism, and sterol biosynthesis found no passage addressing that point. (LLM, conf=0.75)

  Experimental — accuracy not yet measured
• cites Buchovecky et al., 2013bNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Lopez et al., 2017 sourceSupported

Restoring Cyp46a1 expression in Mecp2 KO mice significantly ameliorated disease manifestations.

“reporting that restoring Cyp46a1 expression in Mecp2 KO mice significantly ameliorated disease manifestations (Audouard et al., 2024), providing independent evidence that improving cholesterol homeostasis leads to behavioral benefits.”

• cites Audouard et al., 2024 sourcePartly supported

  Results §3.2 reports significantly reduced hindlimb clasping and improved survival in treated Mecp2 KO male mice. But the treatment overexpressed human CYP46A1 rather than restoring deficient Cyp46a1 expression: the Discussion states that endogenous mouse Cyp46A1 expression was not altered in Mecp2 mice. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Several signaling pathways disrupted in RTT, including BDNF/TrkB signaling, glutamatergic neurotransmission and activity-dependent synaptic plasticity, depend on the integrity of lipid rafts.

“and several signaling pathways that are disrupted in RTT, including BDNF/TrkB signaling, glutamatergic neurotransmission and activity-dependent synaptic plasticity, are known to depend on the integrity of lipid rafts for their proper organization and function (Sebastião et al., 2013;Zonta & Minichiello, 2013).”

• cites Sebastião et al., 2013Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Zonta & Minichiello, 2013 sourcePartly supported

  The section “Lipid raft regulation of BDNF signaling at synapses” reports that disrupting rafts abolished BDNF’s effects on synaptic transmission and plasticity, and discusses raft-associated glutamate receptors. It does not establish that these pathways are disrupted in RTT, and it explicitly says lipid rafts are not required for BDNF/TrkB signaling involved in neuronal survival. Thus the claim’s unqualified dependence on raft integrity is broader than the review supports. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Cholesterol treatment induces a significant distal shift of the AIS in HET neurons.

“whereas cholesterol treatment induced a significant distal shift of the AIS in HET neurons, suggesting an activity-dependent remodeling of the AIS associated with the restoration of synaptic function (Figure 5H).”

Nsdhl protein expression is markedly reduced in Mecp2 KO astrocytes compared with WT cells.

“Consistent with the mRNA data, Nsdhl protein expression was markedly reduced in KO astrocytes compared with WT cells (Figure 1F).”

Nsdhl downregulation in symptomatic Mecp2 HET female mice (P200) is restricted to the cortex.

“In contrast, Nsdhl downregulation in symptomatic Mecp2 HET female mice (P200) was restricted to the cortex (Supplementary Figure 6B).”

Mecp2 knock-out astrocytes exhibit reduced nuclear localization of Srebp2.

“Mecp2 KO astrocytes exhibited reduced nuclear localization of the transcriptional regulator Srebp2”

• Also stated in: § Introduction · plain

  “We found that Mecp2 deficient astrocytes exhibit reduced nuclear translocation of the master regulator Srebp2”

Mecp2 knock-out astrocytes accumulate desmosterol.

“These molecular alterations were associated with intracellular cholesterol and desmosterol accumulation”

• Also stated in: § Introduction · plain

  “Mecp2 deficient astrocytes accumulated intracellular cholesterol and desmosterol”

ApoE and Lcat protein levels are not affected in the cortex of Mecp2 KO animals.

“Obtained results indicated that, whereas ApoE and Lcat protein levels were not affected, a significant reduction of Abca1 was evident in the cortex of KO animals (Figure 3F-I).”

Cholesterol treatment restores axon initial segment length in Mecp2 heterozygous neurons.

“cholesterol treatment restored pre-and post-synaptic density, as well as axon initial segment length in Mecp2 heterozygous (HET) neurons”

Lcat protein shows only a trend toward reduced expression in Mecp2 KO astrocytes.

“whereas Lcat (lecithin-cholesterol acyltransferase), which contributes to the maturation of ApoEcontaining lipoprotein particles, showed only a trend toward reduced expression (Figure 1H).”

Nuclear Srebp2 staining is significantly reduced in Mecp2 KO astrocytes compared with WT cells.

“Immunofluorescence (IF) analysis revealed a significant reduction in nuclear Srebp2 staining in KO astrocytes compared with WT cells (Figure 1A,C)”

Similar alterations are detected in acutely isolated astrocytes and in the cerebral cortex of Mecp2 deficient mice.

“similar alterations were detected in acutely isolated astrocytes and in the cerebral cortex of Mecp2 deficient mice”

In primary cortical astrocytes from P1-P3 mice, genes involved in cholesterol biosynthesis and transport are broadly downregulated in Mecp2 KO cells compared with WT.

“In primary cortical astrocytes isolated from P1-P3 mice, qRT-PCR analysis revealed a broad downregulation of genes involved in both cholesterol biosynthesis and transport in KO cells compared with WT.”

GC-MS analysis revealed a significant accumulation of desmosterol in Mecp2 KO astrocytes.

“Gas chromatography-mass spectrometry (GC-MS) analysis revealed a trend toward increased cholesterol levels in KO astrocytes, together with a significant accumulation of its precursor desmosterol (Figure 2A).”

Approximately 40% of HET neurons display an AIS shorter than 12 μm, compared to about 20% of the treated experimental groups.

“Specifically, we estimated that approximately 40% of HET neurons displayed an AIS shorter than 12 µm, compared to about 20% of the treated experimental groups (Figure 5G).”

Abca1 protein is significantly reduced in the cortex of Mecp2 KO animals.

“Obtained results indicated that, whereas ApoE and Lcat protein levels were not affected, a significant reduction of Abca1 was evident in the cortex of KO animals (Figure 3F-I).”

Mecp2 knock-out astrocytes have defective ApoE lipidation.

“defective ApoE lipidation”

Abca1 protein is significantly decreased in Mecp2 KO astrocytes.

“Similarly, Abca1 (ATP-binding cassette transporter A1), a key mediator of ApoE lipidation, was significantly decreased in Mecp2 KO astrocytes (Figure 1G)”

Mecp2 knock-out astrocytes accumulate intracellular cholesterol.

“These molecular alterations were associated with intracellular cholesterol and desmosterol accumulation”

• Also stated in: § Introduction · plain

  “Mecp2 deficient astrocytes accumulated intracellular cholesterol and desmosterol”

Transcripts encoding Nsdhl, Hmgcr, Mvk and Sqle are significantly reduced in Mecp2 KO astrocytes.

“Specifically, transcripts encoding key enzymes of the cholesterol biosynthetic pathway, including Nsdhl, Hmgcr, Mvk and Sqle, were significantly reduced.”

Mecp2 deficient astrocytes release reduced levels of lipidated ApoE.

“released reduced levels of lapidated ApoE”

Cholesterol concentration in KO-ACM is not altered compared with WT secretome, as measured by GC-MS and Amplex Red assay.

“GC-MS reported no alteration in cholesterol concentration in the KO-ACM, with respect to WT secretome, and similar values resulted from the enzymatic fluorometric Amplex Red Cholesterol Assay (Figure 2D).”

GSEA of cortical transcriptomic dataset revealed a significant impairment of cholesterol homeostasis pathway in Mecp2 KO cortex.

“Gene set enrichment analysis (GSEA) of our previously published cortical transcriptomic dataset (Pozzer et al., 2025) revealed a significant impairment of cholesterol homeostasis pathway in Mecp2 KO cortex (Figure 3B).”

• cites Pozzer et al., 2025Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

There are no significant differences in AIS positioning relative to the soma between untreated HET and WT neurons.

“By analyzing AIS positioning relative to the soma, no significant differences were detected between untreated HET and WT neurons”

Addition of cholesterol fully reverses the reduced density of pre- and postsynaptic compartments and colocalized puncta in WT neurons treated with KO-ACM.

“In particular, the reduced density of both pre-and postsynaptic compartments, as well as that of colocalized puncta, was fully reversed by cholesterol treatment (Figure 4A-D).”

Mecp2 knock-out astrocytes have preserved cholesterol secretion.

“despite preserved ApoE expression and cholesterol secretion”

• Also stated in: § Introduction · plain

  “Although cholesterol secretion remained unchanged”

Expression of Abca1, Abcg1 and Lcat is markedly decreased in Mecp2 KO astrocytes.

“Similarly, the expression of Abca1, Abcg1 and Lcat, which participate in extracellular cholesterol transport, was markedly decreased.”

ApoE expression is unaffected in the ACM collected from Mecp2 KO astrocytes.

“Through WB, we proved that ApoE expression remained unaffected in the ACM collected from KO astrocytes (Figure 2E).”

Cholesterol supplementation improves synaptic morphology in Rett syndrome neurons.

“we demonstrate for the first time that cholesterol supplementation improves synaptic morphology in RTT neurons”

Nsdhl protein expression is markedly reduced in the cerebral cortex, hippocampus and cerebellum of Mecp2 KO mice.

“By WB, we reported a marked reduction in the expression of Nsdhl, not only in the cerebral cortex but also in the hippocampus and cerebellum.”

In acutely sorted cortical astrocytes from P7 WT and KO mice, most cholesterol-related genes are downregulated in KO astrocytes compared with WT.

“Consistent with the findings obtained in pups-derived astrocyte cultures, qRT-PCR analysis confirmed the downregulation of most cholesterol-related genes in KO astrocytes compared with WT cells (Figure 1E).”

GC-MS analysis revealed a trend toward increased cholesterol levels in Mecp2 KO astrocytes.

“Gas chromatography-mass spectrometry (GC-MS) analysis revealed a trend toward increased cholesterol levels in KO astrocytes, together with a significant accumulation of its precursor desmosterol (Figure 2A).”

Filipin staining quantification confirmed a significant accumulation of free intracellular cholesterol in Mecp2 KO astrocytes compared with WT.

“Quantification of the integrated fluorescence density per cell confirmed a significant accumulation of free intracellular cholesterol compared with WT astrocytes (Figure 2B,C).”

The amount of cholesterol secreted by KO astrocytes does not change in acutely isolated astrocytes.

“The amount of cholesterol secreted by KO astrocytes did not change even when the analysis was extended to the ACM of acutely isolated astrocytes, thus excluding the possibility that a lack of difference was attributed to the protocol used (Supplementary Figure 3).”

Mecp2 knock-out astrocytes have reduced Abca1 expression.

“reduced Abca1 expression”

• Also stated in: § Introduction · plain

  “exhibited reduced Abca1 expression”

Mecp2 knock-out astrocytes have preserved ApoE expression.

“despite preserved ApoE expression and cholesterol secretion”

Mecp2 knock-out astrocytes exhibit downregulation of genes involved in cholesterol biosynthesis and transport.

“together with the downregulation of genes involved in cholesterol biosynthesis and transport”

• Also stated in: § Introduction · plain

  “leading to a general downregulation of genes involved in cholesterol biosynthesis and trafficking”

Levels of lipidated ApoE complexes are strongly reduced in KO-ACM compared with WT-ACM.

“However, analysis of its lipidic state, qualitatively assessed by an electrophoresis in non-denaturating native gel, indicated a strong impairment, as reported by an evident reduction of the levels of lipidated ApoE complexes in KO-ACM with respect to WT-ACM (Figure 2F).”

Cholesterol treatment restores pre- and post-synaptic density in Mecp2 heterozygous neurons.

“cholesterol treatment restored pre-and post-synaptic density, as well as axon initial segment length in Mecp2 heterozygous (HET) neurons”

Cholesterol supplementation of astrocyte-conditioned medium rescues synaptic defects induced in wild-type neurons by Mecp2 knock-out astrocytes.

“cholesterol supplementation of astrocyte-conditioned medium rescued the synaptic defects induced in wild-type neurons by Mecp2 KO astrocytes”

Cortical cholesterol levels are significantly increased in symptomatic (P40) Mecp2 KO mice compared with WT controls.

“High Performance Thin Layer Chromatography (HPTLC) analysis revealed a significant increase in cortical cholesterol levels in KO mice compared with WT controls (Figure 3A, Supplementary Figure 4).”

ApoE mRNA levels are not altered in Mecp2 KO astrocytes.

“In contrast, ApoE mRNA levels were not altered (Figure 1D).”

Lcat protein expression level in the cortex of WT and KO mice at P60 differs (p<0.05).

“Histograms report the protein expression level of Abca1(F), Lcat (G) and ApoE (H) in the cortex of WT and KO mice at P60.Data are expressed as percentages of corresponding WT animals and shown as mean±SEM.n=8-13/exp.group.*p<0.05 by Student's t-test or Mann-Whitney test according to data distribution,”

Mecp2 deficiency does not affect astrocytic cholesterol secretion.

“Mecp2 deficiency does not affect astrocytic cholesterol secretion but impairs ApoE lipidation”

Nsdhl protein expression levels in cortex, hippocampus, and cerebellum differ between Mecp2 KO and WT animals at pre-symptomatic (P20) and symptomatic (P40) stages (p<0.05 etc).

“Histograms show the protein expression levels of Nsdhl measured in the cortex (CTX), hippocampus (HP) and cerebellum (CB) of pre-symptomatic (at P20) (D) and symptomatic (P40) (E) Mecp2 KO animals.Data are expressed as percentages of corresponding WT animals and shown as mean±SEM.n=5-10/exp.groups.*p<0.05,**p<0.01 by Student's t test.”

Nsdhl protein levels differ between Mecp2 KO and WT astrocytes.

“Histograms depict the protein levels of Nsdhl (F), Abca1 (G) and Lcat (H) in Mecp2 KO astrocytes, compared to WT astrocytes.”

Abca1 protein expression level in the cortex of WT and KO mice at P60 differs (p<0.05).

“Histograms report the protein expression level of Abca1(F), Lcat (G) and ApoE (H) in the cortex of WT and KO mice at P60.Data are expressed as percentages of corresponding WT animals and shown as mean±SEM.n=8-13/exp.group.*p<0.05 by Student's t-test or Mann-Whitney test according to data distribution,”

Cholesterol homeostasis gene set shows a negative enrichment (NES = -1.57, FDR q = 0.044) in the cortex of Mecp2 KO P60 mice compared with WT.

“Gene set enrichment analysis (GSEA) of cholesterol homeostasis using RNAsequencing data from the cortex of WT and Mecp2 KO P60 mice.NES = -1.57and FDR q=0.044.”

ApoE protein expression level in the cortex of WT and KO mice at P60 differs (p<0.05).

“Histograms report the protein expression level of Abca1(F), Lcat (G) and ApoE (H) in the cortex of WT and KO mice at P60.Data are expressed as percentages of corresponding WT animals and shown as mean±SEM.n=8-13/exp.group.*p<0.05 by Student's t-test or Mann-Whitney test according to data distribution,”

Srebp2 integrated density measured in whole astrocytes and in the nucleus differs between Mecp2 KO and WT cortical astrocytes (p<0.01).

“Histograms report the integrated density measured in the whole astrocytes (B) and selectively in the nucleus (C).Data are shown as mean±SEM.n=25/30 fields deriving from n=6/7 biological replicates.**p<0.01 by Student's t-test.”

Desmosterol content in Mecp2 KO cortical astrocytes differs from WT (p<0.01).

“Histograms show the content of cholesterol and its precursor desmosterol in WT and Mecp2 KO cortical astrocytes analysed through GC-MS and normalized to protein concentration.Data are expressed as mean±SEM, n=6-8/exp.group.**p<0.01,by Mann-Whitney test.”

Mecp2 deficiency impairs ApoE lipidation.

“Mecp2 deficiency does not affect astrocytic cholesterol secretion but impairs ApoE lipidation”

• Also stated in: § Discussion · plain

  “Native gel electrophoresis probed for ApoE, showing differences in the lipidation of ApoE in the medium of Mecp2 KO astrocytes compared to WT.”

Cholesterol treatment rescues synaptic alterations in Mecp2 HET neurons.

“Figure 5. Cholesterol treatment rescues synaptic alterations in Mecp2 HET neurons.”

Mecp2 KO astrocytes show molecular deregulation of the expression of cholesterol-related genes.

“Figure 1.Molecular deregulation of the expression of cholesterol-related genes in Mecp2 KO astrocytes.”

Cholesterol treatment rescues synaptic alterations induced by Mecp2 KO-ACM.

“Figure 4. Cholesterol treatment rescues synaptic alterations induced by Mecp2 KO-ACM.”

Lcat protein levels differ between Mecp2 KO and WT astrocytes.

“Histograms depict the protein levels of Nsdhl (F), Abca1 (G) and Lcat (H) in Mecp2 KO astrocytes, compared to WT astrocytes.”

Cholesterol content in the cortex of Mecp2 KO (P40) mice differs from age-matched WT littermates (p<0.05).

“Graph shows the content of cholesterol in the cortex of Mecp2 KO (P40) mice and age-matched WT littermates analysed through HPTLC.Data are normalized on protein content.n=8/exp.groups*p<0.05 by Student's t test.”

Cholesterol metabolism is deregulated in the cerebral cortex of Mecp2 deficient animals.

“Figure 3. Molecular evidence of cholesterol metabolism deregulation in the cerebral cortex of Mecp2 deficient animals.”

Cholesterol content in Mecp2 KO cortical astrocytes differs from WT (p<0.01).

“Histograms show the content of cholesterol and its precursor desmosterol in WT and Mecp2 KO cortical astrocytes analysed through GC-MS and normalized to protein concentration.Data are expressed as mean±SEM, n=6-8/exp.group.**p<0.01,by Mann-Whitney test.”

Filipin signal integrated density within the area of astrocytes differs between Mecp2 KO and WT cortical astrocytes (p<0.0001).

“The graph shows the integrated density of Filipin signal within the area of the astrocytes.n=80/100.Data are shown as mean±SEM.****p<0.0001by Student's t-test.”

mRNA levels of genes involved in the synthesis and transport of cholesterol are altered in the cortex of symptomatic Mecp2 KO (P40) mice compared with WT littermates (p<0.05 etc).

“Graph shows the mRNA levels of genes involved in the synthesis and transport of cholesterol in the cortex of symptomatic Mecp2 KO (P40).Data are expressed as percentages with respect to male WT littermates, set at 100% and represented by the dotted line.Data are shown as mean±SEM.n=8/exp.groups.;*p<0.05,**p<0.01,***p<0.001by Student's t test.”

Astrocytes are the main source of cholesterol in the brain.

“Astrocytes are the main source of cholesterol in the brain, and cholesterol secreted by astrocytes has a role in synaptic formation and maintenance (Goritz et al., 2005;J.-P. Liu et al., 2010;Pfrieger, 2003).”

• cites Goritz et al., 2005 sourceNot checked — no full text of the source
• cites J.-P. Liu et al., 2010Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Pfrieger, 2003 sourceNot checked — no full text of the source

Impaired cholesterol homeostasis extends beyond in vitro models.

“demonstrating that impaired cholesterol homeostasis extends beyond in vitro models”

Mecp2 deficiency affects Srebp2 activation rather than its expression.

“suggesting that Mecp2 deficiency affects Srebp2 activation rather than its expression.”

Reduced nuclear translocation of Srebp2 leads to downregulation of genes involved in cholesterol biosynthesis and trafficking in Mecp2 deficient astrocytes.

“We found that Mecp2 deficient astrocytes exhibit reduced nuclear translocation of the master regulator Srebp2, leading to a general downregulation of genes involved in cholesterol biosynthesis and trafficking.”

Cholesterol treatment suggests activity-dependent remodeling of the AIS associated with restoration of synaptic function in HET neurons.

“whereas cholesterol treatment induced a significant distal shift of the AIS in HET neurons, suggesting an activity-dependent remodeling of the AIS associated with the restoration of synaptic function (Figure 5H).”

Mecp2 deficiency impairs Srebp2 translocation to the nucleus.

“indicating impaired translocation to nucleus, the compartment where Srebp2 is transcriptionally active.”

Cholesterol secreted by astrocytes has a role in synaptic formation and maintenance.

“Astrocytes are the main source of cholesterol in the brain, and cholesterol secreted by astrocytes has a role in synaptic formation and maintenance (Goritz et al., 2005;J.-P. Liu et al., 2010;Pfrieger, 2003).”

• cites Goritz et al., 2005 sourceNot checked — no full text of the source
• cites J.-P. Liu et al., 2010Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Pfrieger, 2003 sourceNot checked — no full text of the source

Conditioned medium from Mecp2 deficient astrocytes is sufficient to impair neuronal development.

“the observation that conditioned medium from Mecp2 deficient astrocytes is sufficient to impair neuronal development suggests that soluble factors are major mediators of astrocyte-neuron communication in RTT (Albizzati et al., 2024;Ballas et al., 2009).”

• cites Albizzati et al., 2024;Ballas et al., 2009Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Astrocytic cholesterol metabolism is a potential therapeutic target.

“supporting astrocytic cholesterol metabolism as a potential therapeutic target.”

Alterations in brain cholesterol metabolism have been reported in Rett syndrome.

“Multiple lines of evidence suggest that cholesterol metabolism is altered in RTT.”

• Also stated in: § Abstract · plain

  “Although alterations in brain cholesterol metabolism have been reported in RTT”

• cites Buchovecky et al., 2013a;Lopez et al., 2017;Lütjohann et al., 2018;Zandl-Lang et al., 2022;Segatto et al., 2014Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Astrocyte dysfunction contributes to Rett syndrome pathogenesis through non-cell-autonomous mechanisms.

“Increasing evidence indicates that astrocyte dysfunction contributes to RTT pathogenesis through non-cell-autonomous mechanisms”

• Also stated in: § Introduction · boosted

  “recent studies have also demonstrated a role for non-cell-autonomous mechanisms largely mediated by astrocytes, which strongly influence neuronal maturation and function”

• cites Allen & Eroglu, 2017;Postogna et al., 2025Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

The reduction in ApoE lipidation in KO samples might be ascribed to the reduction of Abca1 transporter.

“Of relevance, the reduction in ApoE lipidation in KO samples might be ascribed to the reduction of Abca1 transporter, considering its key role in free cholesterol transfer to ApoE (C.Liu et al., 2025) (Figure 1).”

• cites C.Liu et al., 2025Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Defective astrocyte-to-neuron cholesterol trafficking is a key mechanism contributing to neuronal dysfunction in Rett syndrome.

“Together, these findings identify defective astrocyte-toneuron cholesterol trafficking as a key mechanism contributing to neuronal dysfunction in RTT”

• AssumptionRests on an assumption: Mecp2 knock-out mouse astrocytes are a valid model of human Rett syndrome astrocyte cholesterol metabolism.

Impaired cholesterol trafficking from astrocytes to neurons is a novel mechanism contributing to synaptic dysfunction in RTT.

“In conclusion, our study identifies impaired cholesterol trafficking from astrocytes to neurons as a novel mechanism contributing to synaptic dysfunction in RTT”

• AssumptionRests on an assumption: The relevance of cholesterol metabolism in astrocyte-neuron communication has not been previously addressed, particularly with respect to the metabolically active pool of cholesterol involved in intercellular trafficking.

The fraction of cholesterol released in a functional lipoprotein-associated form and available for delivery to neurons is significantly reduced in Mecp2 KO astrocytes, despite preserved cholesterol secretion.

“One of the most relevant findings of this study is the marked reduction in ApoE lipidation in Mecp2 KO astrocytes, indicating that, despite preserved cholesterol secretion, the fraction of cholesterol that is released in a functional lipoprotein-associated form and available for delivery to neurons is significantly reduced.”

Restoration of membrane lipid rafts might contribute to the beneficial effects of cholesterol supplementation.

“One mechanism that might contribute to these effects is the restoration of membrane lipid rafts, cholesterol-enriched microdomains that organize signaling molecules and synaptic proteins and are required for efficient synaptic transmission (Korinek et al., 2020;Linetti et al., 2010).”

• cites Korinek et al., 2020Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Linetti et al., 2010 sourceNot checked — no full text of the source

• AssumptionRests on an assumption: Direct evidence for lipid raft alterations in RTT is currently lacking.

Total cholesterol levels do not necessarily reflect the fraction of cholesterol that is effectively available for astrocyte-to-neuron transport.

“Indeed, total cholesterol levels do not necessarily reflect the fraction of cholesterol that is effectively available for astrocyte-to-neuron transport”

Reduced Abca1 expression is expected to limit cholesterol and other lipids loading onto ApoE particles, thereby impairing the formation of mature lipoproteins.

“Reduced Abca1 expression is expected to limit cholesterol and other lipids loading onto ApoE particles, thereby impairing the formation of mature lipoproteins.”

The primary defect in RTT might not reside in the total amount of cholesterol present in the brain, but rather in its functional availability and trafficking to neurons.

“Collectively our findings suggest that the primary defect in RTT might not reside in the total amount of cholesterol present in the brain, but rather in its functional availability and trafficking to neurons.”

Cholesterol accumulation might account for the reduced nuclear translocation of Srebp2 and the widespread transcriptional inactivation of its target genes in Mecp2 KO astrocytes.

“Thus, cholesterol accumulation itself might account for the reduced nuclear translocation of Srebp2 and, consequently, for the widespread transcriptional inactivation of its target genes.”

Mecp2 deficiency primarily impairs ApoE maturation rather than its synthesis.

“Notably, this defect occurred despite unchanged ApoE expression, suggesting that Mecp2 deficiency primarily impairs ApoE maturation rather than its synthesis.”