human microglia cell line hmc3  (ATCC)

Journal: Scientific Reports

Article Title: Interferon-β and FTY720 ameliorate progressive CNS inflammation via SOCS1-associated astrocyte signaling

doi: 10.1038/s41598-026-45303-9

Figure Lengend Snippet: Glial cells respond to FTY720 and IFN-β treatment with cell type–specific modulation of inflammatory and regenerative programs. Primary cultures of murine glial cells were treated with vehicle, FTY720, or the combination of FTY720 and IFN-β overnight and then stimulated with IL-1β and TNF-ɑ for 4 h. qPCR of the indicated genes was performed in ( A-D ) microglia and ( n = 3 biological replicates) and ( E–F ) astrocytes (n = 6 biological replicates). ( G ) Astrocyte-conditioned medium (ACM) was generated as described in Fig. H. In brief, primary astrocytes were activated with IL-1β (10 ng/mL) and TNF-α (5 ng/mL) for 24 h, followed by treatment with FTY720/IFN-β and either the control peptide or pJAK2(1001–1013) (20 µM) for 6 h in FCS-free medium. The medium was replaced the next day and collected after an additional 24 h. The harvested ACM was added to activated astrocytes under the respective conditions for subsequent qPCR analysis. qPCR was performed of indicated genes (n = 6 biological replicates). ( H ) Migration assay was performed as described in Fig. H, and the number of migrated monocytes was measured after 3 h ( n = 5 biological replicates). ( I ) Activated human microglia (HMC3 cells; n = 6) and ( J ) human astrocytes (Human astrocytes Catalog #1800, ScienCell; n = 3) were treated with vehicle, FTY720 and vehicle, or the combination of FTY720 and IFN-β overnight and then stimulated with IL-1β and TNF-ɑ for 4 h. qPCR of the indicated genes was performed. All qPCR results are displayed by fold change in mRNA expression. P-values of p < 0.05 were considered significant and determined by one-way ANOVA followed by Tukey´s multiple-comparisons test.

Article Snippet: For further in vitro experiments we used the human microglia cell line HMC3 (American Type Culture Collection [ATCC], Manassas, VA, USA; CRL-3304) and primary human astrocytes (ScienCell Research Laboratories, Carlsbad, CA, USA; Human Astrocytes, Catalog #1800).

Techniques: Generated, Control, Migration, Expressing

Journal: bioRxiv

Article Title: Circulating extracellular vesicles drive microglial senescence and neurodegeneration in Parkinson’s disease

doi: 10.64898/2026.03.10.709299

Figure Lengend Snippet: (a) Immunostaining of HMC3 cells with DAPI (blue) and IBA1 (red) after 24 hours of EV treatment. Scale bar: 100 μm; zoom scale bar: 50 μm (b-c). Percentage and surface area of activated HMC3 cells after 24 hours of EV treatment. Data are represented as mean ± SEM and median with interquartile range (n=8 images per group). P-values are represented in the figures. (d) Heatmap showing normalized signal intensity of cytokine expression in HMC3 cells treated with PD-EV- and HC-EV . (e) Western blot quantification of p16 INK4a and p21 WAF /CIP in HMC3 cells after 48 and 96 hours of EV treatment (n=3). (f-g) Semi-quantification of p16 INK 4a and p21 WAF /CIP expressed as percentage change: (Normalized signal (EV-treated))/ (Normalized signal (PBS control)-1) *100. Data are represented as mean ± SEM. P-value is represented in the figures.

Article Snippet: Human microglia HMC3 cells (ATCC, Cat. No. CRL-3304) were maintained at 37 °C, 5% CO 2 in a DMEM/F-12 (Thermo Fisher Scientific) supplemented with GlutaMAX, 10% v/v FBS, 1% NEAA, and 1% P/S.

Techniques: Immunostaining, Expressing, Western Blot, Control

Journal: bioRxiv

Article Title: Circulating extracellular vesicles drive microglial senescence and neurodegeneration in Parkinson’s disease

doi: 10.64898/2026.03.10.709299

Figure Lengend Snippet: (a-c) p16 INK4a, p21 WAF /CIP , and SERPINE1 mRNA relative expression in HMC3 cells after 48 and 96 hours of treatment with HC-EV, PD-EV, or PBS.

Article Snippet: Human microglia HMC3 cells (ATCC, Cat. No. CRL-3304) were maintained at 37 °C, 5% CO 2 in a DMEM/F-12 (Thermo Fisher Scientific) supplemented with GlutaMAX, 10% v/v FBS, 1% NEAA, and 1% P/S.

Techniques: Expressing

Journal: bioRxiv

Article Title: Circulating extracellular vesicles drive microglial senescence and neurodegeneration in Parkinson’s disease

doi: 10.64898/2026.03.10.709299

Figure Lengend Snippet: (a) Schematic experimental setup of EV treatment on HMC3 cells followed by CM isolation and SH-SY5Y neurons treatment. (b,f) Immunostaining with DAPI (blue) and TUBB3 (red) of SH-SY5Y neurons after treatment with CM and EV - CM. Scale bar: 200 µm. (c,g) Neurite length quantification of SH-SY5Y neurons after 24 hours treatment with CM and EV - CM (n= 550-580 neurons per group). (d,h) Flow cytometry analysis of SH-SY5Y neurons exposed to CM and EV - CM for 24 hours. Annexin V FITC (x-axis) and propidium iodide (y-axis). Q1: dead cells (PI positive), Q2: late-stage apoptotic cells (Annexin V and PI positive), Q3: early-stage apoptotic cells (Annexin V positive), Q4: live cells (Annexin V and PI negative). (e,i) Bar graph showing percentage of cells in Q1 and Q2 stages after 24 hours treatment with CM and EV - CM. Results correspond to 3 replicates from at least three independent differentiations. (j) Neurite length and (k) apoptosis comparison of SH-SY5Y neurons treated with CM and EV - CM, respectively. Scatter plots show all data points, with median as a straight line and the IQR as an error bar. Bar plots show mean ± SEM. P-values are represented in the figures.

Article Snippet: Human microglia HMC3 cells (ATCC, Cat. No. CRL-3304) were maintained at 37 °C, 5% CO 2 in a DMEM/F-12 (Thermo Fisher Scientific) supplemented with GlutaMAX, 10% v/v FBS, 1% NEAA, and 1% P/S.

Techniques: Isolation, Immunostaining, Flow Cytometry, Comparison

Journal: Nature Communications

Article Title: A common 19 bp APOE enhancer deletion is protective against Alzheimer’s disease in African Americans

doi: 10.1038/s41467-026-68808-3

Figure Lengend Snippet: a Schematic of the luciferase experimental design and constructs. The APOE 3′UTR ( APOE 3′UTR only) plus ~400 bp of downstream DNA containing the 19 bp deletion region and SPI1 binding site were cloned into a psiCheck2.2 dual luciferase reporter construct. b Renilla:Firefly luciferase expression data in HMC3 cells after delivery of APOE SPI1 WT sequence ( p = 0.0016 relative to the APOE 3′UTR only) or delivery of the 19 bp deletion [ p = 0.8252; one-way ANOVA, n = 12 samples/group (3 biological replicates, 4 technical replicates), ± SD]. c –e HMC3 gene expression following SPI1 overexpression. c Effect of SPI1 overexpression on c APOE expression ( p = 0.3831, F = 1.789, unpaired two-tailed t-test, n = 12 control and 12 SPI1 biological replicates derived from the average of 4 technical replicates/sample, ± SEM), d APOC1 expression ( p = <0.0001, F = 2.987, unpaired two-tailed t-test, n = 11 control and 12 SPI1 biological replicates derived from the average of 4 technical replicates/sample, ± SEM), and e lncRNA ENSG00000280087 expression ( p = 0.0170, F = 5.414, unpaired two-tailed t-test, n = 12 control and 10 SPI1 biological replicates derived from the average of 4 technical replicates/sample, ± SEM). f – h HMC3 gene expression following Aβ 1:42 treatment. f Effect of Aβ 1:42 treatment on APOE expression ( p = 0.2098, F = 1.243, unpaired two-tailed t-test, n = 12 control and 12 Aβ biological replicates derived from the average of 4 technical replicates/sample, ± SEM), g APOC1 expression ( p = 0.0015, F = 4.657, unpaired two-tailed t-test, n = 11 control and 12 Aβ biological replicates derived from the average of 4 technical replicates/sample, ± SEM) and h, lncRNA ENSG00000280087 expression ( p = 0.0255, F = 2.502, unpaired two-tailed t-test, n = 12 control and 12 Aβ biological replicates derived from the average of 4 technical replicates/sample, ± SEM) in HMC3 cells.

Article Snippet: HMC3 human microglia cell lines (ATCC CRL-3304) were grown in EMEM media and seeded at 3.34 × 10 4 cells plated in 12-well plates for luciferase assays.

Techniques: Luciferase, Construct, Binding Assay, Clone Assay, Expressing, Sequencing, Gene Expression, Over Expression, Two Tailed Test, Control, Derivative Assay

Journal: Neural Regeneration Research

Article Title: Mesenchymal stem cell–derived small extracellular vesicles enhance the therapeutic effect of retinal progenitor cells in retinal degenerative disease rats

doi: 10.4103/NRR.NRR-D-23-02108

Figure Lengend Snippet: Microglial suppression of RCS rats mediated by co-transplantation of RPCs with M-sEVs. (A) Representative confocal images of Iba1 staining in the RPCs and RPCs + M-sEVs groups at 2 and 4 weeks post transplantation. Enlarged orthogonal view of M-sEVs at the injected area (right panel) shows co-localization of the M-sEVs with microglia in the subretinal space. (B) Statistical analysis of the number of Iba1-positive cells in each group in A. (C, D) Representative confocal images showing the localization of MSC-sEVs (PKH26 staining) with F-actin-labeled BV2 (C) and HMC3 (D) microglial cells stimulated with LPS. Scale bars: 20 μm (A), 10 μm (A, enlarged orthogonal view), 50 μm (C, D). (E) RT-PCR analysis showing relative mRNA expression levels of IL-1β, IL-6, and TNF-α in the mouse (BV2) and human (HMC3) microglial cell lines. Data are expressed as means ± SD ( n ≥ 3 for Iba1 staining; n = 3 for real-time PCR); * P < 0.05, ** P < 0.01, *** P < 0.001 (B: Welch’s t -test; D: Welch’s one-way analysis of variance followed by Dunnett’s T3 multiple-comparison test). DAPI: 4′,6-Diamidino-2-phenylindole; EGFP: enhanced green fluorescent protein; Iba1: ionized calcium-binding adapter molecule 1; IL-1β; interleukin 1 beta; IL-6: interleukin 6; LPS: lipopolysaccharide; M-sEVs: mesenchymal stem cell–small extracellular vesicles; RT-PCR: real-time polymerase chain reaction; RPCs: retinal progenitor cells; RCS: Royal College of Surgeons; TNF-α: tumor necrosis factor α; w: weeks.

Article Snippet: Human microglia clone 3 (HMC3) cells were purchased from Procell Life Science & Technology Co., Ltd. (Wuhan, China, Cat# CL-0620, RRID: CVCL_II76) and were cultured in alpha minimum essential medium (Cat# SH30265.01, Hyclone) containing 10% FBS.

Techniques: Transplantation Assay, Staining, Injection, Labeling, Reverse Transcription Polymerase Chain Reaction, Expressing, Real-time Polymerase Chain Reaction, Comparison, Binding Assay