Reverse the progression of Alzheimer’s disease through Nano-ERASER-based adult neuroregeneration

Subsequent lineage-tracing studies yielded conflicting results, suggesting that PTBP1 depletion alone may be insufficient to drive genuine astrocyte-to-neuron conversion in the adult mammalian brain.

“However, subsequent lineage-tracing studies yielded conflicting results, suggesting that PTBP1 depletion alone may be insufficient to drive genuine astrocyte-to-neuron conversion in the adult mammalian brain, raising concerns that some reported conversions reflected viral promoter leakage into pre-existing neurons.”

• cites [16] Wang, 2021, Revisiting astrocyte to neuron conversion with lineage tracing in vivo, Cell sourceSupported
• cites [17] Chen, 2022, Repressing PTBP1 fails to convert reactive astrocytes to dopaminergic neurons in a 6-hydroxydopamine mouse model of Parkinson's disease, eLife sourceSupported
• cites [18] Hoang, 2023, Ptbp1 deletion does not induce astrocyte-to-neuron conversion, Nature sourceSupported

PTBP1 is an RNA-binding protein that maintains non-neuronal splicing programs.

“PTBP1 is an RNA-binding protein that maintains non-neuronal splicing programs; its downregulation is a hallmark of neuronal differentiation during development.”

• cites [12] Xue, 2013, Genetic programs in human and mouse early embryos revealed by single-cell RNA sequencing, Nature sourceCould not verify

  The supplied passages discuss gene-expression networks in early embryos, not PTBP1 or its role in splicing. Searches for PTBP1 and for polypyrimidine-tract binding protein in connection with neuronal splicing returned no passage addressing the claim; the copy therefore neither supports nor contradicts it. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

PTBP1 downregulation is a hallmark of neuronal differentiation during development.

“PTBP1 is an RNA-binding protein that maintains non-neuronal splicing programs; its downregulation is a hallmark of neuronal differentiation during development.”

• cites [12] Xue, 2013, Genetic programs in human and mouse early embryos revealed by single-cell RNA sequencing, Nature sourceCould not verify

  The paper’s Abstract (page unavailable) describes gene-expression changes from oocyte to morula in human and mouse pre-implantation embryos, not neuronal differentiation. Searches for PTBP1, non-neuronal splicing, and neuronal differentiation found no passage in our copy addressing the claim. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Astrocyte-to-neuron reprogramming has recently emerged as an appealing regenerative strategy because astrocytes represent an endogenous cellular reservoir from which new neurons might be generated in situ.

“Astrocyte-to-neuron reprogramming has recently emerged as an appealing regenerative strategy because astrocytes, which are abundant, regionally distributed, and proliferative, represent an endogenous cellular reservoir from which new neurons might be generated in situ.”

• cites [10] Mattugini, 2019, Inducing Different Neuronal Subtypes from Astrocytes in the Injured Mouse Cerebral Cortex, Neuron sourceSupported
• cites [11] Heinrich, 2014, Sox2-mediated conversion of NG2 glia into induced neurons in the injured adult cerebral cortex, Stem Cell Rep. sourceSupported

Prior mechanistic studies have shown that PTBP1 normally suppresses productive expression of PTBP2 through alternative splicing coupled to nonsense-mediated decay.

“Prior mechanistic studies have shown that PTBP1 normally suppresses productive expression of PTBP2 through alternative splicing coupled to nonsense-mediated decay.”

• cites [22] Dawicki-McKenna, 2023, Mapping PTBP2 binding in human brain identifies SYNGAP1 as a target for therapeutic splice switching, Nat. Commun. sourceCould not verify

  The Results section on PTBP2-dependent synaptic genes reports that PTBP1 levels fall while PTBP2 levels rise during neuronal differentiation, but it does not say that PTBP1 suppresses PTBP2 expression. Searches for PTBP1, PTBP2, alternative splicing, and nonsense-mediated decay found NMD mechanisms involving SYNGAP1, not PTBP2. No page numbers are available in our copy. (LLM, conf=0.75)

  Experimental — accuracy not yet measured
• cites [23] Ge, 2016, Polypyrimidine tract binding protein 1 protects mRNAs from recognition by the nonsense-mediated mRNA decay pathway, eLife sourceSupported

RAGE receptors are overexpressed in the BBB and astrocytes, especially in brain tissue from AD animal models and AD patients.

“Our previous research and that of others found that RAGE receptors are overexpressed in the BBB and astrocytes, especially in brain tissue from AD animal models and AD patients.33–35”

• cites [33] Wang, 2009, Receptor for advanced glycation end products is upregulated in optic neuropathy of Alzheimer's disease, Acta Neuropathol. sourceSupported
• cites [34] Sasaki, 2001, Immunohistochemical distribution of the receptor for advanced glycation end products in neurons and astrocytes in Alzheimer's disease, Brain Res. sourceNot checked — no full text of the source
• cites [35] Choi, 2014, Increased expression of the receptor for advanced glycation end products in neurons and astrocytes in a triple transgenic mouse model of Alzheimer's disease, Exp. Mol. Med. sourcePartly supported

  The Results (Distribution and cellular localization of RAGE-positive cells; page unavailable) report more RAGE-expressing astrocytes in the hippocampal CA1 region of aged AD-model mice than in controls. The Introduction reports increased RAGE in AD patient brains. The Discussion mentions RAGE-mediated transport across the blood–brain barrier, but does not establish RAGE overexpression at the barrier. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Serum exposure to hiPSC-derived brain organoids recapitulates AD-like pathologies.

“To mimic BBB leakage-induced sporadic AD, human induced pluripotent stem cell (hiPSC)-derived brain organoids (BOs, composed of neurons and astrocytes) were exposed to serum to recapitulate AD-like pathologies (Figure 3I).31”

• cites [31] Chen, 2021, Modeling Sporadic Alzheimer's Disease in Human Brain Organoids under Serum Exposure, Adv. Sci. (Weinh) sourceSupported

TN-PTBP1 penetrates the blood-brain barrier.

“TN-PTBP1 penetrates the blood–brain barrier, delivers anti-PTBP1 antibodies into astrocytes, and triggers rapid TRIM21-mediated PTBP1 degradation.”

TN-PTBP1 delivers anti-PTBP1 antibodies into astrocytes.

“TN-PTBP1 penetrates the blood–brain barrier, delivers anti-PTBP1 antibodies into astrocytes, and triggers rapid TRIM21-mediated PTBP1 degradation.”

TN-PTBP1 triggers rapid TRIM21-mediated PTBP1 degradation.

“TN-PTBP1 penetrates the blood–brain barrier, delivers anti-PTBP1 antibodies into astrocytes, and triggers rapid TRIM21-mediated PTBP1 degradation.”

Free aPTBP1 and TN-IgG failed to generate functional and interconnected neurons.

“In contrast, free aPTBP1 and TN-IgG failed to generate functional and interconnected neurons.”

TN-PTBP1 treatment reduces TNF-α and IL-6 in brains of AD animals.

“To our surprise, the conversion of astrocytes to neurons did not exacerbate glial dysfunction but was instead associated with an overall improvement in the inflammatory milieu of the AD brain, as evidenced by reduced TNF-α and IL-6 in the brains of animals receiving TN-PTBP1 treatment (Figure 5L–M).”

A significantly higher fraction of DCX+, Myelin+, and MAP2+ cells was observed in TN-PTBP1 treatment group.

“In contrast, a significantly higher fraction of DCX+, Myelin+, and MAP2+ cells was observed in the TN-PTBP1 treatment group, validating TN-PTBP1’s capacity to convert astrocytes into neurons.”

TN-PTBP1 improves nesting skills of AD mice.

“The gradually improved nesting score in the TN-PTBP1 treatment group suggests its potent and progressive therapeutic effect (Figure 4E&G).”

TN-PTBP1 effectively downregulates PTBP1 and TRIM21 expressions in HA after penetrating the BBB.

“As expected, the TN-PTBP1 effectively downregulated PTBP1 and TRIM21 expressions in HA after penetrating the BBB (Figure 2Q–S), suggesting that TN-PTBP1 might deplete PTBP1 in HA through the TRIM21-mediated proteasome degradation process when applied in vivo.”

TN-PTBP1 has a surface charge of about −10.0 mV.

“The surface charge of TN-PTBP1 was about −10.0 mV (Figure 1F), which ensures their stability during circulation in the bloodstream.”

TN-PTBP1 treatment group showed a reduction in Aβ burden.

“Benefiting from the improved pathological brain microenvironment, the TN-PTBP1 treatment group showed a reduction in Aβ burden (Figure 5N, Supplementary Figure 12).”

TN-PTBP1 effectively downregulated PTBP1 to a level comparable to that in normal BOs.

“As expected, TN-PTBP1 effectively downregulated PTBP1 to a level comparable to that in normal BOs.”

TN-PTBP1 has a hydrodynamic diameter of about 128 nm.

“The hydrodynamic diameters of TN-PTBP1 were about 128 nm (Figure 1E).”

N-IgG barely enters human cerebral microvascular endothelial cells (hCMEC/D3) and primary human astrocytes (HA).

“As expected, N-IgG could barely enter human cerebral microvascular endothelial cells (hCMEC/D3) (Figure 2A–C) and primary human astrocytes (HA) (Figure 2D–F).”

aPTBP1 and TN-IgG treatments did not improve mice's nesting skills.

“In contrast, aPTBP1 and TN-IgG treatments did not improve mice’s nesting skills.”

TN-IgG treatment does not reduce PTBP1 levels.

“In contrast, TN-IgG treatment did not reduce PTBP1 levels, suggesting that the depletion of PTBP1 shown in Figure 2J–K was due to the specific protein-degradation effect of TN-PTBP1, rather than to the carrier system itself.”

Most astrocytes in TN-PTBP1 treatment group changed their morphology from a star shape to a cell body with a long axon.

“Interestingly, most astrocytes in the TN-PTBP1 treatment group changed their morphology from a star shape to a cell body with a long axon (Figure 3A).”

Encapsulated aPTBP1 can be released under reducing conditions that mimic the intracellular environment.

“Furthermore, Figure 1D validated that the encapsulated aPTBP1 could be released under reducing conditions that mimic the intracellular environment.”

TN-IgG can effectively escape from endo-lysosomes in HA.

“The standing-alone red signals in TN-IgG-treated HA cells revealed that TN-IgG could effectively escape from the endo-lysosomes (Figure 2G–H), which is a required step for Nano-ERASER based Trim-Away.”

5XFAD mice brain exhibits significantly higher levels of RAGE compared to normal mice.

“Compared to normal mice, the brain of 5XFAD mice exhibited significantly higher levels of RAGE (Supplementary Figure 7).”

TN-PTBP1 achieves high brain-targeting efficacy, with most signals penetrating the BBB and entering the brain parenchyma.

“In contrast to the non-targeted nanogel, the boosted, stand-alone red signals within the hippocampus in the TN-Cy5 treatment group confirmed that TN-PTBP1 achieved high brain-targeting efficacy, with most signals penetrating the BBB and entering the brain parenchyma (Figure 4C).”

TN-PTBP1 treatment causes a significant increase of DCX in HA 1-week post-treatment.

“Remarkably, a significant increase of doublecortin (DCX), a key neuronal marker for immature neurons, was observed in TN-PTBP1 treated cells 1-week post-treatment (Figure 2J–L).”

TN-PTBP1 improves expression of DCX and NeuN in AD brain.

“As expected, TN-PTBP1 improved the expression of DCX and NeuN (Figure 5C–D).”

At three weeks post-treatment, fractions of DCX+, Myelin+, and MAP2+ cells in aPTBP1-treated group were similar to control group.

“However, at three weeks post-treatment, the fractions of DCX+, Myelin+, and MAP2+ cells in the aPTBP1-treated group were similar to those in the control group, suggesting that aPTBP1 could initiate the astrocyte-to-neuron conversion process but failed to complete it.”

TN-BSA penetrates the in vitro BBB significantly faster than BSA under typical culture conditions.

“Results shown in Figure 2O revealed that TN-BSA penetrated the BBB significantly faster than BSA under typical culture conditions.”

TN-PTBP1-treated BOs exhibited widespread high-amplitude spikes and strong, synchronized responses upon electrical stimulation.

“Compared with control BOs, TN-PTBP1-treated BOs exhibited widespread high-amplitude spikes and strong, synchronized responses upon electrical stimulation (Figure 3R–S), confirming that the regenerated neurons within the BOs were functional and had integrated into the existing neural circuits.”

TN-PTBP1 treatment induces upregulation of PTBP2.

“As expected, TN-PTBP1 treatment induced upregulation of PTBP2 (Figure 2J and 2N).”

PTBP1 level in AD brain is higher than in normal ones.

“It was revealed that the PTBP1 level in the AD brain was higher than that in the normal ones.”

Most TN-Cy3-Cy5 nanogels were intact after transcytosis through the BBB.

“The little change in the FRET signal of the transcytosized nanogels and significantly lower than that of fully dissociated ones (TN-Cy3/TN-Cy5) suggests that most nanogels were intact (Figure 2P) after transcytosis through the BBB.”

TN-Cy5 effectively targets the brains of AD animals.

“Figure 4B also confirmed that TN-Cy5 could effectively target the brains of AD animals.”

TN-PTBP1 reduces PTBP1 level in HA to less than 40% compared with control cells two weeks post-treatment.

“Figure 2J–K revealed that the PTBP1 level in TN-PTBP1 treated HA was less than 40% compared with that in control cells two weeks post-treatment.”

Most antibodies in HA had been released within 24 h post-treatment.

“It was revealed that most of the antibodies in the HA had been released within 24 h post-treatment, evidenced by the disappearance of red FRET signals and the recovery of green Cy3 signals (Figure 2I).”

Transcytosis of TN-BSA is substantially enhanced in LPS-stimulated BV2 conditioned medium compared to BV2 conditioned medium.

“Interestingly, when conditioned medium from BV2 stimulated with LPS (CMBV-2 + LPS) was presented, the transcytosis of TN-BSA was substantially enhanced compared to that in BV2 conditioned medium (CMBV-2) (Figure 2O), confirming that targeted nanogels may utilize the receptors overexpressed on the BBB during an inflammatory condition, such as AD.”

aPTBP1 and TN-PTBP1 yielded similar fractions of DCX+, Myelin+, and SOX2+ cells at 1 week post-treatment.

“It was revealed that aPTBP1 and TN-PTBP1 yielded similar fractions of DCX+, Myelin+, and SOX2+ cells at 1 week post-treatment.”

TN-PTBP1-treated AD mice showed increased expression of tight junction proteins ZO-1 and Occludin relative to control AD mice.

“Notably, TN-PTBP1-treated AD mice showed increased expression of the tight junction proteins ZO-1 and Occludin relative to control AD mice, suggesting that TN-PTBP1 does not compromise BBB integrity (Supplementary Figure 13); rather, by reducing reactive astrocytosis, neuroinflammation, and amyloid burden, it may improve the neurovascular microenvironment despite inducing astrocyte-to-neuron”

NeuN level in TN-PTBP1 BOs recovered to that in normal BOs.

“Furthermore, the NeuN level in the TN-PTBP1 BOs recovered to that in the normal BOs (Figure 3O and 3Q).”

MEA recordings confirm that only TN-PTBP1 treatment induces widespread, synchronized neural firing.

“MEA recordings confirm that only TN-PTBP1 treatment induces widespread, synchronized neural firing (Figure 3H).”

Ten days post-treatment, PTBP1 expression in BOs was upregulated upon serum stimulation.

“Ten days post-treatment, PTBP1 expression in BOs was upregulated upon serum stimulation (Figure 3O–P).”

TN-IgG shows enhanced cellular uptake in hCMEC/D3 and HA due to functionalization of MPC and RAGE-targeting ligands.

“In contrast, abundant red signals were observed in TN-IgG-Cy5-treated cells, confirming the enhanced cellular uptake resulting from the functionalization of MPC and RAGE-targeting ligands (Figure 2A–F).”

Cells linked with a long axon were positive for both MAP2 and NeuN.

“As expected, the cells linked with a long axon were positive for both MAP2 and NeuN (Figure 3A), which validated that TN-PTBP1 had successfully converted astrocytes to neurons.”

In serum-injured organoids modeling sporadic AD conditions, neuronal markers are diminished.

“and neuronal markers are diminished (Figure 3J–K).”

TN-PTBP1 releases antibody cargo intracellularly.

“release antibody cargo intracellularly (Figure 2I)”

TN-PTBP1 leverages dual-targeting ligands to penetrate the BBB.

“The platform leverages dual-targeting ligands to penetrate the BBB (Figures 2O)”

TN-PTBP1 significantly improves learning performance during the Morris water maze in AD mice.

“learning performance during the Morris water maze (Figures 4H)”

TN-PTBP1 restores NeuN levels in serum-injured organoids.

“restores NeuN (Figure 3J–K, 3O, and 3Q)”

TN-PTBP1 treatment coincides with increased hippocampal neurogenesis.

“These improvements coincide with increased hippocampal neurogenesis”

Nanogel transport across the hCMEC/D3 monolayer was enhanced under inflammatory conditions.

“Their transport was further enhanced under inflammatory conditions.”

In vivo, TN-PTBP1 penetrates the BBB.

“penetrates the BBB”

Free aPTBP1 fails to achieve comparable outcomes to TN-PTBP1.

“Free aPTBP1 fails to achieve comparable outcomes”

TN-PTBP1 escapes endolysosomal compartments.

“escape endolysosomal compartments (Figure 2G–H)”

Live-cell imaging reveals a dynamic, multistep astrocyte-to-neuron transition.

“Live-cell imaging further reveals a dynamic, multistep astrocyte-to-neuron transition (Supplementary Videos 2)”

In serum-injured organoids modeling sporadic AD conditions, PTBP1 is upregulated.

“In serum-injured organoids modeling sporadic AD conditions, PTBP1 is upregulated (Figure 3O–P)”

In vivo, TN-PTBP1 accumulates selectively in AD brain regions.

“TN-PTBP1 accumulates selectively in AD brain regions (Figure 4A–B)”

TN-PTBP1 significantly improves memory retention in AD mice.

“and memory retention (Figures 4F and 4I, Supplementary Figure 8).”

TN-PTBP1 significantly improves nesting ability in AD mice.

“Behavioral tests confirm that TN-PTBP1 significantly improves nesting ability (Figures 4E and 4G)”

TN-PTBP1 enables rapid TRIM21-mediated degradation of PTBP1 at the protein level.

“These design features enable rapid TRIM21-mediated degradation of PTBP1 at the protein level (Figure 2J–K and 2Q–R)”

In vitro, TN-PTBP1 robustly downregulates PTBP1.

“In vitro, TN-PTBP1 robustly downregulates PTBP1”

In vivo, TN-PTBP1 localizes within the hippocampal parenchyma.

“and localizes within the hippocampal parenchyma (Figure 4C).”

TN-PTBP1 treatment results in the presence of DCX+ immature neurons and NeuN+ mature neurons.

“as evidenced by the presence of DCX+ immature neurons and NeuN+ mature neurons (Figures 5I–K, Supplementary Figure 10–11).”

Free aPTBP1 compromises BBB integrity.

“and even compromises BBB integrity”

TN-PTBP1 treatment coincides with restoration of neuronal populations.

“and restoration of neuronal populations”

TN-PTBP1 treatment is accompanied by decreased GFAP expression.

“accompanied by decreased GFAP expression (Figure 5E–F)”

TN-PTBP1 efficiently enters astrocytes.

“efficiently enter astrocytes (Figures 2A–F)”

TN-PTBP1 treatment attenuates neuroinflammation.

“the attenuated neuroinflammation (Figure 5L–M)”

In vitro, TN-PTBP1 induces expression of neuronal markers such as DCX, MAP2, and NeuN.

“induces expression of neuronal markers such as DCX, MAP2, and NeuN (Figures 2J, 2M, and 3A–E).”

TN-PTBP1 restores MAP2 levels in serum-injured organoids.

“and MAP2 levels (Figure 3J and 3M)”

TN-PTBP1 normalizes PTBP1 expression in serum-injured organoids.

“TN-PTBP1 normalizes PTBP1 expression (Figure 3O–P)”

Targeted nanogels penetrated the hCMEC/D3 monolayer more efficiently than non-targeted controls.

“Importantly, the targeted nanogels penetrated the monolayer more efficiently than the non-targeted controls.”

PTBP1 is selectively downregulated in astrocytes after TN-PTBP1 treatment.

“Furthermore, PTBP1 is selectively downregulated in astrocytes (Figures 5E and 5H)”

TN-PTBP1 is a nanoparticle-based Trim-Away platform that enables efficient, astrocyte-targeted depletion of PTBP1 through intracellular delivery of anti-PTBP1 antibodies and TRIM21-mediated proteasomal degradation.

“we developed and validated a nanoparticle-based Trim-Away platform (TN-PTBP1) that enables efficient, astrocyte-targeted depletion of PTBP1 through intracellular delivery of anti-PTBP1 antibodies and TRIM21-mediated proteasomal degradation.”

MEA analyses in vitro and in human brain organoids support the capacity of TN-PTBP1-generated neurons to participate in active neuronal networks.

“MEA analyses in vitro and in human brain organoids support the capacity of TN-PTBP1-generated neurons to participate in active neuronal networks”

TN-PTBP1 achieves robust and selective PTBP1 depletion in human astrocytes, brain organoids, and AD mouse brains.

“Our results demonstrate that TN-PTBP1 achieves robust and selective PTBP1 depletion in human astrocytes, brain organoids, and AD mouse brains.”

TN-PTBP1 treatment decreases amyloid burden.

“and decreased amyloid burden”

In vitro human astrocytes and human brain organoid data provide direct support for TN-PTBP1-induced astrocyte-to-neuron conversion.

“while our in vitro human astrocytes and human brain organoid data provide direct support for TN-PTBP1-induced astrocyte-to-neuron conversion”

TN-PTBP1 treatment improves BBB integrity.

“improved BBB integrity (Supplementary Figure 13)”

TN-PTBP1 treatment reduced measured inflammatory cytokines, including TNF-α and IL-6.

“Although TN-PTBP1 treatment reduced measured inflammatory cytokines, including TNF-α and IL-6”

Increased DCX+ immature neurons and NeuN+ mature neurons in the hippocampus and other brain regions support in vivo neurogenesis/neuroregeneration.

“although increased DCX+ immature neurons and NeuN+ mature neurons in the hippocampus and other brain regions support in vivo neurogenesis/neuroregeneration”

This work establishes a translational biomaterials-based platform for in situ neuronal regeneration.

“By resolving the long-standing controversy surrounding PTBP1 reprogramming and establishing a translational biomaterials-based platform for in situ neuronal regeneration, this work introduces a transformative therapeutic paradigm aimed not only at slowing AD but at rebuilding the neuronal circuitry it destroys.”

TN-PTBP1 induces astrocyte-to-neuron conversion.

“This non-genomic protein depletion induces astrocyte-to-neuron conversion, generates functional neurons in vitro and in human brain organoids, and restores hippocampal neurogenesis in AD mice.”

Timely PTBP1 downregulation may be required to unlock neuronal reprogramming competence.

“These discrepancies suggest that timely PTBP1 downregulation may be required to unlock neuronal reprogramming competence.”

TN-PTBP1 treatment demonstrates true functional reversal of AD-associated deficits.

“Importantly, TN-PTBP1 treatment significantly improves cognition, learning, and memory, demonstrating true functional reversal of AD-associated deficits.”

TN-PTBP1 restores hippocampal neurogenesis in AD mice.

“This non-genomic protein depletion induces astrocyte-to-neuron conversion, generates functional neurons in vitro and in human brain organoids, and restores hippocampal neurogenesis in AD mice.”

TN-PTBP1 treatment has potent and progressive therapeutic effect.

“The gradually improved nesting score in the TN-PTBP1 treatment group suggests its potent and progressive therapeutic effect (Figure 4E&G).”

TN-PTBP1 does not compromise BBB integrity.

“Notably, TN-PTBP1-treated AD mice showed increased expression of the tight junction proteins ZO-1 and Occludin relative to control AD mice, suggesting that TN-PTBP1 does not compromise BBB integrity (Supplementary Figure 13); rather, by reducing reactive astrocytosis, neuroinflammation, and amyloid burden, it may improve the neurovascular microenvironment despite inducing astrocyte-to-neuron”

TN-PTBP1 treatment suggests neuronal regeneration and subsequent maturation from astrocytes in BOs.

“Immunohistostaining further confirmed the increased density of NeuN+ and MAP+ cells (Figure 3J–K, 3M) and the reduced density of GFAP+ and DCX+ cells (Figure 3K, 3L, and 3N) in BOs after TN-PTBP1 treatment, suggesting neuronal regeneration and subsequent maturation from astrocytes.”

TN-PTBP1 might deplete PTBP1 in HA through TRIM21-mediated proteasome degradation when applied in vivo.

“As expected, the TN-PTBP1 effectively downregulated PTBP1 and TRIM21 expressions in HA after penetrating the BBB (Figure 2Q–S), suggesting that TN-PTBP1 might deplete PTBP1 in HA through the TRIM21-mediated proteasome degradation process when applied in vivo.”

The depletion of PTBP1 by TN-PTBP1 is due to specific protein-degradation effect of TN-PTBP1, rather than carrier system itself.

“In contrast, TN-IgG treatment did not reduce PTBP1 levels, suggesting that the depletion of PTBP1 shown in Figure 2J–K was due to the specific protein-degradation effect of TN-PTBP1, rather than to the carrier system itself.”

The surface charge of TN-PTBP1 ensures their stability during circulation in the bloodstream.

“The surface charge of TN-PTBP1 was about −10.0 mV (Figure 1F), which ensures their stability during circulation in the bloodstream.”

Presence of DCX-positive cells in hippocampus and whole brains of TN-PTBP1-treated animals demonstrates neurogenic effect of TN-PTBP1.

“The presence of DCX-positive cells in the hippocampus and whole brains of TN-PTBP1-treated animals demonstrated the neurogenic effect of TN-PTBP1 (Supplementary Figure 10, Figure 5I).”

Downregulation of PTBP1 in HA was due to intracellularly released aPTBP1-induced PTBP1 degradation via Trim-Away.

“Taken together, we can conclude that the downregulation of PTBP1 in HA was due to intracellularly released aPTBP1-induced PTBP1 degradation via Trim-Away.”

TN-PTBP1 has capacity to convert astrocytes into neurons.

“In contrast, a significantly higher fraction of DCX+, Myelin+, and MAP2+ cells was observed in the TN-PTBP1 treatment group, validating TN-PTBP1’s capacity to convert astrocytes into neurons.”

By enabling non-genomic, reversible, and cell-type-directed reprogramming, nanoparticle-mediated Trim-Away offers a biomaterials-driven strategy for regenerative therapy that complements current disease-modifying approaches.

“Moreover, by enabling non-genomic, reversible, and cell-type-directed reprogramming, nanoparticle-mediated Trim-Away offers a biomaterials-driven strategy for regenerative therapy that complements current disease-modifying approaches.”

TN-PTBP1 improves learning capacity of AD mice.

“The TN-PTBP1 treatment decreased the latency time of AD mice during training and yielded a latency time and average distance to the platform similar to those of normal mice (Figure 4H, Supplementary Figure 8), indicating improved learning capacity.”

Reduced GFAP expression and decreased correlation between GFAP and PTBP1 expression in AD brain treated with TN-PTBP1 suggests TN-PTBP1 effectively downregulated PTBP1 in astrocytes.

“Furthermore, reduced GFAP expression and decreased correlation between GFAP and PTBP1 expression in the AD brain treated with TN-PTBP1 suggested that TN-PTBP1 effectively downregulated PTBP1 in astrocytes (Figure 5E–H).”

Higher MAP2 density in TN-PTBP1-treated cells than in aPTBP1-treated cells suggests better conversion efficiency.

“The higher MAP2 density in TN-PTBP1-treated cells than in aPTBP1-treated cells suggests better conversion efficiency.”

TN-PTBP1 successfully converted astrocytes to neurons.

“As expected, the cells linked with a long axon were positive for both MAP2 and NeuN (Figure 3A), which validated that TN-PTBP1 had successfully converted astrocytes to neurons.”

The astrocyte-converted, functionally active neurons formed connected networks capable of electrically evoked activity.

“In addition, the high degree and widespread synchrony of spikes imply that the astrocyte-converted, functionally active neurons formed connected networks capable of electrically evoked activity.”

TN-PTBP1 can induce astrocyte-to-neuron reprogramming.

“All these results proved that TN-PTBP1 can induce astrocyte-to-neuron reprogramming.”

TN-PTBP1-induced adult neuroregeneration successfully reversed AD progression in the 5XFAD mouse model.

“Combining the results above, we conclude that TN-PTBP1-induced adult neuroregeneration successfully reversed AD progression in the 5XFAD mouse model.”

Targeted nanogels may utilize receptors overexpressed on the BBB during an inflammatory condition, such as AD.

“Interestingly, when conditioned medium from BV2 stimulated with LPS (CMBV-2 + LPS) was presented, the transcytosis of TN-BSA was substantially enhanced compared to that in BV2 conditioned medium (CMBV-2) (Figure 2O), confirming that targeted nanogels may utilize the receptors overexpressed on the BBB during an inflammatory condition, such as AD.”

These findings support the physiological relevance of the targeting strategy.

“Together, these findings support the physiological relevance of the targeting strategy.”

The Nano-ERASER system (TN-PTBP1) was engineered to overcome the barriers of PTBP1 depletion and astrocyte targeting.

“By contrast, our nanoparticle-based Nano-ERASER system (TN-PTBP1) was engineered explicitly to overcome both barriers.”

Intracellular delivery, not mere antibody presence, is essential for inducing reprogramming and generating an interconnected neural network.

“reinforcing that intracellular delivery, not mere antibody presence, is essential for inducing reprogramming and generating an interconnected neural network (Figure 3H).”

Efficient PTBP1 depletion is sufficient to activate endogenous neuronal programs.

“These results indicate that efficient PTBP1 depletion is sufficient to activate endogenous neuronal programs, resolving the central mechanistic uncertainty that has divided the field.”

When PTBP1 is efficiently and selectively downregulated, astrocytes adopt neuronal transcriptional and morphological identities.

“Crucially, our results demonstrate that when PTBP1 is efficiently and selectively downregulated, astrocytes adopt neuronal transcriptional and morphological identities.”

TN-PTBP1 treatment indicates effective neuronal regeneration in serum-injured organoids.

“indicating effective neuronal regeneration.”

The newly generated neurons demonstrate truly functional integration.

“demonstrating truly functional integration.”

The astrocyte-to-neuron transition is consistent with lineage progression from immature to mature neuronal states.

“consistent with lineage progression from immature to mature neuronal states.”

TN-PTBP1 achieves a recoverable suppressive potency not attainable with conventional gene-silencing methods.

“achieving a recoverable suppressive potency not attainable with conventional gene-silencing methods.”

PTBP1 may not have been reduced to the threshold required to unlock astrocytic plasticity in some prior studies.

“PTBP1 may simply not have been reduced to the threshold required to unlock astrocytic plasticity.”

TN-PTBP1 treatment has broader effects on AD pathology.

“suggest broader effects of TN-PTBP1 treatment on AD pathology (Figure 5N, Supplementary Figure 12).”

TN-PTBP1 was designed to enhance receptor-mediated uptake at the BBB and to enrich delivery to astrocytes after transcytosis, rather than to achieve absolute cell-type specificity.

“TN-PTBP1 was designed to enhance receptor-mediated uptake at the BBB and to enrich delivery to astrocytes after transcytosis, rather than to achieve absolute cell-type specificity.”

TN-PTBP1 overcomes key barriers that have limited both PTBP1-targeting strategies and the in vivo application of Trim-Away.

“TN-PTBP1 overcomes key barriers that have limited both PTBP1-targeting strategies and the in vivo application of Trim-Away.”

Efficient PTBP1 downregulation can subsequently reverse the progression of AD.

“and subsequently reverse the progression of AD.”

• AssumptionRests on an assumption: The adult mammalian brain has very limited regenerative capacity, and progressive neuronal loss is a central driver of irreversible functional decline in disorders such as AD, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, traumatic brain injury, and stroke.
• AssumptionRests on an assumption: Endogenous neurogenesis in the adult human brain is highly restricted and insufficient to compensate for neuronal loss.

The study was not powered for definitive sex-stratified analysis.

“However, the study was not powered for definitive sex-stratified analysis.”

PTBP1 depletion is not inherently sufficient to drive reprogramming.

“Our results suggest that PTBP1 depletion is not inherently sufficient to drive reprogramming.”

Traditional genetic approaches may fail to meet the threshold for successful conversion because they often produce incomplete knockdown, limited recoverability, or confounding artifacts.

“Traditional genetic approaches may fail to meet this threshold because they often produce incomplete knockdown, limited recoverability, or confounding artifacts.”

Successful conversion appears to depend on whether PTBP1 can be depleted with sufficient potency, efficiency, and time dependency.

“Rather, successful conversion appears to depend on whether PTBP1 can be depleted with sufficient potency, efficiency, and time dependency.”

In the context of AD, effective PTBP1 downregulation can contribute to the reversal of disease-associated pathology and behavioral deficits.

“and, in the context of AD, can contribute to the reversal of disease-associated pathology and behavioral deficits.”

• AssumptionRests on an assumption: The adult mammalian brain has very limited regenerative capacity, and progressive neuronal loss is a central driver of irreversible functional decline in disorders such as AD, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, traumatic brain injury, and stroke.
• AssumptionRests on an assumption: Endogenous neurogenesis in the adult human brain is highly restricted and insufficient to compensate for neuronal loss.

TN-PTBP1 is a powerful biomaterials-enabled platform for non-genomic, reversible, and cell-type-specific reprogramming of astrocytes.

“our findings establish TN-PTBP1 as a powerful biomaterials-enabled platform for non-genomic, reversible, and cell-type–specific reprogramming of astrocytes.”

These findings help reconcile the conflicting reports surrounding PTBP1-mediated astrocyte-to-neuron conversion.

“Together, these findings help reconcile the conflicting reports surrounding PTBP1-mediated astrocyte-to-neuron (AtN) conversion.”

The platform is astrocyte-enriched rather than astrocyte-exclusive.

“We therefore interpret this platform as astrocyte-enriched rather than astrocyte-exclusive.”

The present study did not directly analyze microglial activation, microglial phenotypes, or microglia-specific inflammatory signaling.

“the present study did not directly analyze microglial activation, microglial phenotypes, or microglia-specific inflammatory signaling.”

TN-PTBP1 offers compelling evidence that efficient PTBP1 downregulation can drive astrocyte conversion under AD-relevant conditions.

“offers compelling evidence that efficient PTBP1 downregulation can drive astrocyte conversion under AD-relevant conditions”

Marker-based data alone do not definitively distinguish direct astrocyte-to-neuron conversion from stimulation of endogenous neural progenitor-mediated neurogenesis.

“these marker-based data alone do not definitively distinguish direct astrocyte-to-neuron conversion from stimulation of endogenous neural progenitor-mediated neurogenesis.”

Effective PTBP1 downregulation can induce AtN conversion in both in vitro and in vivo settings.

“our data support the conclusion that effective PTBP1 downregulation can induce AtN conversion in both in vitro and in vivo settings”