Journal: bioRxiv

Article Title: Single-cell analysis of sterol-induced Ca 2+ signaling in human astrocytes by dynamic mode decomposition

doi: 10.64898/2026.02.09.704834

Figure Lengend Snippet: Simulation of calcium oscillations in astrocytes under varying SERCA pump activity. (A) Schematic representation of intracellular calcium handling components, illustrating selected proteins involved in astrocytic calcium signaling. The ordinary differential equation (ODE)–based framework incorporates cytosolic calcium influx pathways, inositol 1,4,5-trisphosphate receptor (IP 3 R)-mediated endoplasmic reticulum (ER) calcium release (Green), and SERCA-driven reuptake (Red), while other signaling modules are greyscale for simplicity. Components shown include G-protein-coupled receptors (GPCRs), phospholipase C (PLC), store-operated calcium entry (SOCE), purinergic P2X receptors (P2X), Piezo1 channels, transient receptor potential (TRP) channels, voltage-gated calcium channels (VGCCs), plasma membrane Ca 2+ -ATPase (PMCA), Na + /Ca 2+ exchanger (NCX), mitochondrial calcium uniporter (MCU), and secretory pathway Ca 2+ -ATPase (SPCA). (B) Simulated cytosolic Ca 2+ dynamics for three levels of SERCA activity ( v m 2 = 5, 10, and 15 µ M/s). Higher SERCA rates lead to more frequent and higher-amplitude oscillations. (C) Corresponding ER Ca 2+ dynamics show complementary depletion and refilling behavior across the same SERCA conditions. (D) Phase-plane trajectories (ER Ca 2+ vs. cytosolic Ca 2+ ) illustrate distinct dynamical regimes that emerge as a function of SERCA strength. (E) Distribution of 100 sampled v m 2 values drawn from three Gaussian distributions centered at 5 µ M/s (green), 10 µ M/s (orange), and 15 µ M/s (blue), used to generate the simulated variability in Ca 2+ dynamics. Together, the simulations demonstrate how altering SERCA activity modulates the frequency, amplitude, and qualitative form of astrocytic calcium oscillations.

Article Snippet: Normal human astrocytes immortalized with SV40 large T antigen (P10251-IM, Innoprot) were maintained in T25 culture flasks (Nunc EasYFlask, 156367, Thermo Fisher Scientific) at 37 °C in a humidified incubator with 5% CO 2 .

Techniques: Activity Assay, Clinical Proteomics, Membrane

Journal: bioRxiv

Article Title: Single-cell analysis of sterol-induced Ca 2+ signaling in human astrocytes by dynamic mode decomposition

doi: 10.64898/2026.02.09.704834

Figure Lengend Snippet: DMD reconstruction and classification of simulated single-cell Ca 2+ dynamics. (A) Cytosolic Ca 2+ traces generated by numerically integrating the astrocyte ODE model for three values of the SERCA rate parameter v m 2 , producing 100 simulated cells per condition. (B) Amplitudes of the DMD modes and (C) corresponding eigenvalues computed from a rank-400 decomposition with a delay dimension of 300. (D) Kernel PCA applied to the DMD mode amplitudes followed by k -means clustering separated the simulated cells into four dynamical clusters. Silhouette score = 0.64 (E–F) Example Ca 2+ traces (dots) and their DMD reconstructions (lines) for cells representative of clusters 1–2 (E) and clusters 3–4 (F). (G) Cluster composition across experimental conditions indicating the distribution of cells in each cluster.

Article Snippet: Normal human astrocytes immortalized with SV40 large T antigen (P10251-IM, Innoprot) were maintained in T25 culture flasks (Nunc EasYFlask, 156367, Thermo Fisher Scientific) at 37 °C in a humidified incubator with 5% CO 2 .

Techniques: Single Cell, Generated

Journal: bioRxiv

Article Title: Single-cell analysis of sterol-induced Ca 2+ signaling in human astrocytes by dynamic mode decomposition

doi: 10.64898/2026.02.09.704834

Figure Lengend Snippet: Classification of cholesterol-dependent single-cell Ca 2+ dynamics using DMD-TDE and kernel-PCA. (A) Pooled and normalized cytosolic Ca 2+ traces from all experimental conditions. Astrocytes were loaded with Cal-520 and imaged at 1 Hz; at t = 300 s, either control medium (M1), methyl-beta-cyclodextrin (MCD), or cholesterol–cyclodextrin complexes (20, 100, or 250 µM cholesterol) were added. Traces from all cells across all conditions were concatenated into a single cell-time matrix. (B) Amplitudes of the delay-embedded DMD modes computed from the full dataset (delay embedding: 400 steps; truncation rank: 400). (C) Kernel-PCA followed by k -means clustering identified three distinct dynamical clusters of Ca 2+ activity. Silhouette score = 0.708 (D) Cluster representation across conditions, showing strong enrichment of cluster 1 in control and cholesterol-depleted cells, and a shift toward clusters 2 and 3 with increasing cholesterol concentration. (E–G) Ca 2+ heatmaps (identical intensity scaling) for the three clusters, illustrating that cluster 1 exhibits low activity, cluster 2 displays frequent high-amplitude spikes, and cluster 3 shows dense, moderate-amplitude oscillations. (H) Spatial maps of classified cells for each experimental condition. Cells are color-coded according to their assigned cluster (blue: cluster 1; green: cluster 2; red: cluster 3), and cell outlines are derived from CellPose segmentation (see Materials and Methods).

Article Snippet: Normal human astrocytes immortalized with SV40 large T antigen (P10251-IM, Innoprot) were maintained in T25 culture flasks (Nunc EasYFlask, 156367, Thermo Fisher Scientific) at 37 °C in a humidified incubator with 5% CO 2 .

Techniques: Single Cell, Control, Activity Assay, Concentration Assay, Derivative Assay

Journal: bioRxiv

Article Title: Single-cell analysis of sterol-induced Ca 2+ signaling in human astrocytes by dynamic mode decomposition

doi: 10.64898/2026.02.09.704834

Figure Lengend Snippet: Temporal correlation structure and synchronization in the three astrocyte clusters. (A-C) Temporal correlation matrices for clusters 1-3, computed from the Ca 2+ activity time series within each cluster. Clusters 2 and 3 show pronounced off-diagonal structure, indicating recurrent and coordinated activity patterns over time, whereas cluster 1 exhibits minimal off-diagonal correlations, consistent with weak temporal coupling. (D) Kuramoto order parameter r ( t ) for each cluster, quantifying instantaneous phase synchrony. The dashed vertical line marks the addition of cholesterol-cyclodextrin complexes at 300 s.

Article Snippet: Normal human astrocytes immortalized with SV40 large T antigen (P10251-IM, Innoprot) were maintained in T25 culture flasks (Nunc EasYFlask, 156367, Thermo Fisher Scientific) at 37 °C in a humidified incubator with 5% CO 2 .

Techniques: Activity Assay

Journal: bioRxiv

Article Title: Single-cell analysis of sterol-induced Ca 2+ signaling in human astrocytes by dynamic mode decomposition

doi: 10.64898/2026.02.09.704834

Figure Lengend Snippet: DMD classification of astrocytic Ca 2+ dynamics after oxysterols treatment and cholesterol loading. (A) Pooled and normalized Ca 2+ activity traces from all recorded astrocytes across conditions (Control, 24-HC, 25-HC, 27-HC). Each row represents a single cell.(B) Time-delay Dynamic Mode Decomposition amplitudes for all cells, showing the contribution of each DMD mode to individual cellular activity. (C) Three-dimensional embedding of the DMD amplitude features using kernel PCA followed by k -means clustering. Four distinct dynamical clusters emerge, separating cells based on shared temporal motifs in their Ca 2+ activity. Silhouette score = 0.718. (D) Cluster composition across experimental conditions. Oxysterol-treated astrocytes are predominantly assigned to Cluster 1, which is characterized by weak or quiescent Ca 2+ activity, whereas clusters exhibiting oscillatory or spike-like behavior (Clusters 2 and 3) are enriched in control cells. (E-H) Ca 2+ activity heatmaps for cells belonging to each cluster. Cluster 1 (E) displays recurrent, temporally coherent activity. Cluster 2 (F) shows sparse or unstructured signals with minimal coordination. Cluster 3 (G) exhibits pronounced bursting patterns, while Cluster 4 (H) contains a small subset of cells with irregular or atypical Ca 2+ dynamics. Together, these results demonstrate that DMD-TDE combined with unsupervised clustering reveals distinct dynamical phenotypes of astrocytic Ca 2+ signaling and highlights condition-dependent shifts in astrocyte activity patterns.

Article Snippet: Normal human astrocytes immortalized with SV40 large T antigen (P10251-IM, Innoprot) were maintained in T25 culture flasks (Nunc EasYFlask, 156367, Thermo Fisher Scientific) at 37 °C in a humidified incubator with 5% CO 2 .

Techniques: Activity Assay, Control, Single Cell

Journal: Cancer Research and Treatment : Official Journal of Korean Cancer Association

Article Title: CYBC1 Drives Glioblastoma Progression via Reactive Oxygen Species and NF-κB Pathways

doi: 10.4143/crt.2024.827

Figure Lengend Snippet: Analysis of cytochrome b-245 chaperone 1 (CYBC1) expression. (A) CYBC1 mRNA expression in different glioma types. Non-tumor, n=4; oligodendroglioma, n=191; oligoastrocytoma, n=130; astrocytoma, n=194; glioblastoma (GBM), n=152. One-way ANOVA with Tukey’s multiple comparison test was conducted for statistical analysis. (B) CYBC1 expression in non-tumors and gliomas. Non-tumor, n=4; GBM, n=156. This data was derived from the GlioVis database ( http://gliovis.bioinfo.cnio.es/ ). TCGA, The Cancer Genome Atlas.(C) Differential mRNA expression of the CYBC1 gene in Gene Expression Omnibus (GEO) datasets (accession number: GSE15824 ). Non-tumor, n=5; GBM, n=12. (D) Kaplan-Meier survival curves derived from GEPIA ( http://gepia.cancer-pku.cn/index.html ). (E) CYBC1 immunostaining in normal cortex, low-grade glioma (LGG), and high-grade glioma (HGG). Data are derived from the Human Protein Atlas. (F) CYBC1 immunostaining in normal (n=10), astrocytoma (n=22), and GBM (n=23) tissue microarrays (GL807a). Scale bars=500 μm. (G) Western blot analysis of CYBC1 expression in normal human astrocytes (NHA) and GBM cell lines. Immunofluorescence (H) and immunoblot (I) assay demonstrating CYBC1 localization in GBM cells. Scale bars=200 μm. n.s, not significant.

Article Snippet: U373 and U87 cells were purchased from the Korean Cell line bank, and normal human astrocytes (NHA) were purchased from iXCells Biotechnologies.

Techniques: Expressing, Comparison, Derivative Assay, Gene Expression, Immunostaining, Western Blot, Immunofluorescence

Journal: Biomolecules

Article Title: Expression of Reelin, Aβ1-42, Tau and FTH1 in Idiopathic Epiretinal Membranes: Exploring the Link Between Reelin and Neurodegenerative Biomarkers

doi: 10.3390/biom15081187

Figure Lengend Snippet: Cultures of normal human astrocytes respond to vitreous exposure. RELN, Aβ1-42, FTH1, TAU and GFAP transcript expressions by ERM vitreal fluid exposed astrocytes. The lines represent the exclusivity of expression for: ( A ) red line for stage 3 and green line for stage 2 and ( B ) red line for stage 4 and green line for stage 2; p < 0.001, REST–ANOVA and multiparametric analysis with Bonferroni correction. Legend: RELN—Reelin; Aβ1-42—amyloid β 1-42; FTH1—ferritin heavy chain; TAU—TAU protein; GFAP—glial fibrillary acidic protein.

Article Snippet: We developed in vitro primary cultures of human normal retinal astrocytes (Innoprot; Bizkaia, Spain).

Techniques: Expressing

Journal: Biomolecules

Article Title: Expression of Reelin, Aβ1-42, Tau and FTH1 in Idiopathic Epiretinal Membranes: Exploring the Link Between Reelin and Neurodegenerative Biomarkers

doi: 10.3390/biom15081187

Figure Lengend Snippet: Graphical illustration of inflammation loop during Alzheimer’s disease. A representative image showing how brain and eye (retina) are involved in Alzheimer’s disease (a neurodegenerative disease) and some ocular conditions (vitreoretinal traction and AMD). As stated, β-amyloid accumulation (mainly Aβ1-42 inside soluble and insoluble plaques) and neurofibrillary tangles (chiefly the hyperphosphorylated form) result in a substantial synaptic and neuronal loss . In AD brains, activated microglia and reactive astrocytes are associated with plaques and tangles, so their activation in retinal tissues might be of great relevance for understanding the neurodegenerative microenvironment . The concept behind “burning person” is that the mechanisms of inflammation and oxidative stress that occur in the brain may have similarities or effects in the retina. Legend: Aβ—beta-amyloid; TAU—microtubule-associated protein TAU (tangles); ILs—interleukins; Fe—iron; ROS—reactive oxygen species indicative of oxidative stress; AMD—age-related macular degeneration; NU—neutrophil; M—microglia; AS—astrocyte; NU—neuron; AD—Alzheimer’s disease.

Article Snippet: We developed in vitro primary cultures of human normal retinal astrocytes (Innoprot; Bizkaia, Spain).

Techniques: Activation Assay