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PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
TIM-3 receptor-ligand engagement has been proposed to inhibit the Extracellular signal-regulated kinase (ERK) and NF-κB pathways in NK cells, yet Nuclear Factor of Activated T-cells (NFAT) signaling remains unaffected, thereby enabling NK cells to sustain cytokine production.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Nevertheless, further investigation is warranted to fully elucidate the underlying mechanism behind the effect of TIM-3 on cytokine production in NK cells.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
We validated our observations in a haploidentical allo-HCT murine model of NBL relapse to establish preclinically the combination of adoptive transfer of NK cells and TIM-3 blockade treatment.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
We observed that the combination of NK cell donor lymphocyte infusions and TIM-3 blockade treatment is a viable therapeutic platform.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Mice treated with multiple infusions of 15–4P stimulated NK cells and anti-TIM-3 antibody showed significantly smaller NBL tumors and prolonged survival.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
However, tumors began to grow shortly after treatment ceased.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Prior studies show that following IL-15 expansion, allogeneic ex vivo expanded NK cells remain detectable for up to 16 days after infusion in a murine model.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
The limited persistence of adoptive NK cell therapies may require frequent infusions to effectively impact clinical outcomes, highlighting a limitation of this therapeutic approach.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
However, clinical studies have demonstrated favorable outcomes with the use of NK cell infusions combined with immunotherapy as an effective bridge therapy before consolidation.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Future studies may aim to generate a sustained NK cell response by designing an in vivo tumor vaccine with costimulatory molecules and/or cytokines specific to activating NK cells.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
TIM-3 also plays a crucial role in modulating immune responses during acute GVHD following allo-HCT.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Donor T cells in the allogeneic setting rapidly upregulate TIM-3 expression.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Inhibition of the TIM-3/Gal-9 interaction has been shown to lead to increased proliferation of T cells and heightened GVHD lethality.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
However, TIM-3 inhibition after regulatory T-cell depletion has been shown to increase IFN-γ production and reduce GVHD severity.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
In our experimental model, mice received TCD BMCs with 1×10 T cells added back to the graft.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Notably, GVHD development was absent despite the use of TIM-3 blockade in the presence of residual donor T cells in the bone marrow graft.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
We then investigated whether the anti-NBL response was driven by NK cells or T cells in the graft.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Only NK cell depletion reversed the clinical benefit of TIM-3 blockade, leading to larger tumors and worse survival compared with T-cell depletion, which had no effect.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
It is possible that the 15–4P NK cell infusions increased IFN-γ production while mediating GVT responses, which indirectly facilitated CD8 T cell and macrophage antitumor responses as well.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
However, it is important to acknowledge the divergence between human and mouse TIM-3 function.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
In mice, TIM-3 engagement inhibits NK cell activation and cytokine production, primarily by promoting apoptosis of activated NK cells.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
In contrast, human NK cells exhibit a gradient response where TIM-3 can elicit co-stimulatory or inhibitory signals based on cellular conditions.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
Nevertheless, these results support the conclusion that the anti-NBL response is primarily driven by NK cells, although the potential contribution of blocking TIM-3 macrophages and dendritic cells on NBL clearance needs to be explored.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
In summary, this study explored the combination of ex vivo activated NK cells with TCD allo-HCT and revealed the enhanced NK cell potency against relapsed NBL, particularly when paired with TIM-3 blockade.
PMC12699621
TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant
TCD allo-HCT has emerged as a safe and effective treatment for multiple hematologic malignancies, and in combination with cell-based immunotherapy may become a platform for treating solid tumors to maximize the GVT effect without exacerbating lethal GVHD.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
Sirtuin-2 (SIRT2), a member of the NAD+-dependent deacetylase family, is a key regulator of cellular processes such as metabolism, stress response and aging.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
We hypothesize that SIRT2 isoforms (v1, v2, v3) exert isoform-specific regulatory effects on mitochondrial-related genes, influencing mitochondrial dynamics, energy metabolism, and cellular homeostasis.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
To investigate this, SH-SY5Y cells, from a neuroblastoma cell line were transfected with empty vector (EV) or SIRT2 isoforms (v1, v2, v3), and the expression of mitochondrial genes, including Complex I subunits (NDUFAB1, NDUFV1, NDUFV2, NDUFS1), mitochondrial biogenesis and energy metabolism regulators (PGC-1α, PGC-1β), and mitochondrial dynamics and morphology markers (MFN-1, MFN-2, FiS1), were analyzed using quantitative RT-PCR (QuantStudio 3 Real-Time PCR System).
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
Results revealed that SIRT2-v1 significantly upregulated mRNA levels of NDUFAB1 (p < 0.001), NDUFV1 (p < 0.01), and NDUFS1 (p < 0.05) compared to v2 and v3.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
SIRT2-v1 also increased PGC-1α expression (p < 0.05), numerically increased PGC-1β expression too and MFN-2 (p < 0.01), which are vital for mitochondrial biogenesis and mitochondrial fusion.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
These findings highlight the isoform-specific roles of SIRT-2 in modulating mitochondrial gene expression, with SIRT2-v1 predominantly influencing Complex I and PGC-1α.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
This study provides insights into the distinct regulatory effects of SIRT-2 v1 isoforms on mitochondrial function and metabolic pathways, underscoring their potential therapeutic relevance in mitochondrial-associated disorders.
PMC12761066
Isoform-specific roles of SIRT2 in regulating mitochondrial gene expression
Additionally, the differential age-related increase in SIRT2 isoforms warrants further investigation.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
Alzheimer's disease (AD) is the progressive neurological disorder, which causes degeneration of neuronal cells and memory loss.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
The neuropathological presentation of AD constitute amyloid beta (Aβ) plaques and hyperphosphorylated tau tangles.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
Hyperactivation of GSK‐3b and p53 results in the formation of these neuropathological hallmarks.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
In this study, we have explored the potential of GSK‐3b and p53 as blood‐based biomarkers for early detection of AD.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
The circulatory level of GSK‐3b, phospho‐GSK‐3b, p53 and phospho‐p53 in serum from AD, mild cognitive impairment (MCI), and geriatric‐control (GC) subjects were measured using surface plasmon resonance and further validated by western blot.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
The rescue of neurotoxicity by an antioxidant plant extract, Emblica Officinalis (EO) was also checked by MTT assay on SH‐SY5Y cells (neuroblastoma cell line).
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
The level of all the proteins were found to be significantly upregulated (p > 0.001) in AD and MCI compared to GC and can easily differentiate AD and MCI from GC.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
The expression of these proteins was found to be decreased in a dose dependent manner when treated with EO in SH‐SY5Y cells.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
These proteins can serve as potential blood based biomarkers for the diagnosis of AD and EO can suppress their level.
PMC12782009
Circulatory GSK‐3b; A blood based biomarker and a therapeutic target for Alzheimer's Disease
This work has translational value and clinical utility in the future.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Effective treatments for early cognitive impairment and Alzheimer’s disease (AD) remain limited.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Given the multifactorial nature of AD, therapeutic strategies must focus on targeting disease‐specific biochemical pathways.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Metabolic disruptions play a critical role in cognitive decline, prompting research into alternative disease modifiers like phytochemicals.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
This study investigated the therapeutic potential of resveratrol, a biologically active terpenoid derived from Ginkgo biloba, to alleviate amyloid‐induced cellular dysfunction.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Known for its antioxidant and anti‐inflammatory properties, resveratrol may modulate oxidative stress and inflammation, key contributors to neurodegeneration.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
The study utilized the BE(2)‐M17 neuroblastoma cell line to model AD.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Cells were differentiated into mature neurons using 10 µM retinoic acid and subsequently treated with 20 µM amyloid beta (Aβ) for 24 hours.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Prior to Aβ exposure, cells were pre‐treated with varying concentrations of resveratrol (1 µM, 5 µM, 10 µM, 15 µM, and 20 µM) for 24 hours.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Three assays—CellTiter‐Glo, ROS, and MTT—were performed to assess resveratrol’s effects on cell viability, oxidative stress, and mitochondrial function.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
The CellTiter‐Glo assay revealed that pretreatment with 10 µM resveratrol resulted in the highest ATP production and cell viability (p = 0.0386), indicating improved cellular energy status and survival under Aβ exposure.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
The ROS assay demonstrated that 10 µM resveratrol significantly reduced oxidative stress induced by Aβ (p = 0.049), highlighting its ability to counteract a major driver of neurodegeneration.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Lastly, the MTT assay showed that 10 µM resveratrol effectively mitigated Aβ‐induced mitochondrial dysfunction (p = 0.013), emphasizing its protective impact on mitochondrial integrity.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
Resveratrol at 10 µM emerged as the most effective concentration across all assays, demonstrating its ability to enhance cell viability, reduce oxidative stress, and restore mitochondrial function.
PMC12740619
Protective Effects of Resveratrol Against Amyloid Beta‐Induced Toxicity in Neuroblastoma Cells: Insights from Cell Viability and cellular energetics.
These findings suggest that resveratrol’s modulation of energy metabolism and cellular homeostasis holds promise as a therapeutic approach for Alzheimer’s disease.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
The SH‐SY5Y neuroblastoma cell line is a valuable in vitro model for studying neuronal differentiation and neurodegenerative diseases like Alzheimer's disease (AD).
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Traditional differentiation protocols mainly use retinoic acid (RA); however, they lack extracellular matrix (ECM) components that are critical for mechanotransduction and cellular adhesion, which limits their physiological relevance.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Laminins, a key ECM glycoprotein, play an essential role in neurite outgrowth and synaptic formation, indicating their potential to enhance neuronal differentiation.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
SH‐SY5Y cells were cultured in DMEM/F12 supplemented with fetal bovine serum (FBS) and essential additives.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Differentiation was induced using RA (10 µM and 25 µM) and a laminin‐rich ECM (LrECM).
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Plates were pre‐coated with Matrigel® (a laminin‐rich ECM) before seeding the cells.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Differentiation efficiency was monitored over 10 days through light microscopy, immunofluorescence for neuronal markers (NeuN and β3‐tubulin), and acetylcholinesterase (AChE) activity assays.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Western blotting assessed β3‐tubulin expression, and neurite lengths were quantified using FIJI software.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
The combined RA and LrECM treatment significantly enhanced SH‐SY5Y differentiation when compared to RA alone.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Neuronal morphology, marked by extensive neurite outgrowth, became evident as early as day 4 and was sustained for up to 10 days.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Immunofluorescence confirmed increased NeuN expression, showing a shift from cytoplasmic to perinuclear localization over time.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
β3‐tubulin levels remained consistently high in LrECM‐treated cells, unlike those treated with RA alone, which demonstrated a decline after day 7.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
Enhanced cholinergic differentiation was indicated by elevated AChE activity, particularly at 25 µM RA, although higher RA concentrations were unable to sustain neuronal characteristics and raised concerns about cytotoxicity.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
The incorporation of LrECM into SH‐SY5Y differentiation protocols significantly enhances neuronal differentiation and maintains neuron‐like characteristics, providing a more physiologically relevant in vitro model for studying AD and other neurodegenerative diseases.
PMC12779348
An Optimized Approach for Neuron‐Like Differentiation of SH‐SY5Y Neuroblastoma Cells Using Retinoic Acid and a Laminin‐Rich Extracellular Matrix
This approach enables cost‐effective, rapid differentiation and more accurately mimics the brain microenvironment, establishing a strong platform for neurobiological research and therapeutic screening.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
The cholinergic system is one of the most ancient and widespread signaling systems in the body, implicated in a range of pathological conditions—from neurodegenerative disorders to cancer.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Given its broad relevance, there is growing interest in characterizing this system across diverse cellular models to enable drug screening, mechanistic studies, and exploration of new therapeutic avenues.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
In this study, we investigated four cancer cell lines: one of neuroblastoma origin previously used in cholinergic signaling studies (SH-SY5Y), one non-small cell lung adenocarcinoma line (A549), and two small cell lung carcinoma lines (H69 and H82).
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
We assessed the expression and localization of key components of the cholinergic system, along with the cellular capacity for acetylcholine (ACh) synthesis and release.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Whole-cell flow cytometry following membrane permeabilization revealed that all cell lines expressed the ACh-synthesizing enzyme choline acetyltransferase (ChAT).
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
HPLC-MS analysis confirmed that ChAT was functionally active, as all cell lines synthesized and released ACh into the conditioned media, suggesting the presence of autocrine and/or paracrine ACh signaling circuits, consistent with previous reports.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
The cell lines also demonstrated choline uptake, indicative of functional choline and/or organic cation transporters.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Additionally, all lines expressed the ACh-degrading enzymes acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), as well as the alfa seven (α7) nicotinic and M1 muscarinic ACh receptor subtypes.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Notably, flow cytometry of intact SH-SY5Y cells revealed two novel findings: (1) ChAT was localized to the extracellular membrane, a feature not observed in the lung cancer cell lines, and (2) BChE, rather than AChE, was the predominant membrane-bound ACh-degrading enzyme.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
These results were corroborated by both whole-cell and surface-confocal microscopy.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
In conclusion, our findings suggest that a functional cholinergic phenotype is a shared feature of several carcinoma cell lines, potentially serving as a survival checkpoint that could be therapeutically explored.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
The discovery of extracellular membrane-bound ChAT uniquely in neuroblastoma SH-SY5Y cells points to a novel form of in situ ACh signaling that warrants further investigation.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
The cholinergic signaling system, which relies on acetylcholine (ACh), is one of the most widespread and evolutionarily conserved communication systems in the body.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Cholinergic cells express the enzyme choline acetyltransferase (ChAT), which synthesizes ACh by transferring the acetyl group from acetyl-coenzyme A (A-CoA) to choline.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
The canonical view is that ChAT is a cytosolic enzyme, with ACh biosynthesis occurring exclusively in the cytoplasm.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
However, recent studies challenge this view, demonstrating that ChAT can be released by lymphocytes and astrocytes into extracellular fluids, where it contributes to maintaining extracellular ACh equilibrium even in the presence of active ACh-degrading enzymes .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
In addition to ChAT, the cholinergic system comprises two major receptor families: nicotinic acetylcholine receptors (nAChRs), which are ligand-gated ion channels, and muscarinic acetylcholine receptors (mAChRs), which are G protein-coupled receptors .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Two key transporter proteins also support cholinergic function: the vesicular ACh transporter (VAChT), responsible for packaging ACh into synaptic vesicles, and the high-affinity choline transporter (hChT), which facilitates choline reuptake following ACh hydrolysis at synapses.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
ACh signaling is terminated by enzymatic hydrolysis into choline and acetate .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Acetylcholinesterase (AChE) has been considered to be the primary enzyme responsible for ACh hydrolysis while butyrylcholinesterase (BChE) may act as a decoy enzyme, protecting AChE from dietary or environmental inhibitors .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Cells that express ChAT and synthesize ACh are classified as cholinergic, while those expressing ACh receptors are termed cholinoceptive.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Importantly, cholinergic cells can also be cholinoceptive, as ACh may exert autocrine effects via autoreceptors .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Cholinergic cells are broadly categorized into neuronal and non-neuronal types.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Neuronal cholinergic systems include the central cholinergic network originating from the basal forebrain, comprising four major nuclei (Ch1–Ch4) that project throughout the brain .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
This system plays a critical role in cognition, attention, and memory, and is notably affected in dementia disorders such as Alzheimer’s disease (AD), dementia with Lewy bodies, and Parkinson’s disease dementia .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Cholinergic interneurons within the nigrostriatal system, particularly in the putamen, are compromised in Parkinson’s disease and related disorders such as corticobasal degeneration and progressive supranuclear palsy .
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Another neuronal cholinergic system comprises the parasympathetic system, consisting of the 12 cranial nerves which connect directly to organs, muscles, and glands, controlling various bodily functions.
PMC12608971
Comparative Analysis of Cholinergic Machinery in Carcinomas: Discovery of Membrane-Tethered ChAT as Evidence for Surface-Based ACh Synthesis in Neuroblastoma Cells
Most of these together with cholinergic neuromotor neurons in the brainstem and spinal cord are progressively lost in neuromotor disorders like amyotrophic lateral sclerosis (ALS) .