raw264 7 mouse macrophage cells  (ATCC)

Journal: Bioactive Materials

Article Title: A foam cell-targeted lipophagy restoration strategy stabilizes vulnerable atherosclerotic plaques

doi: 10.1016/j.bioactmat.2026.02.041

Figure Lengend Snippet: In vitro evaluation of foam cell lipid accumulation and lipophagy activation following OPN-HMCN@MLT treatment. ( A - C ) ORO and BODIPY staining images and corresponding quantification of ORO and BODIPY positive areas of RAW264.7 cells under different stimulations (n = 5, scale bar for ORO: 100 μm, scale bar for BODIPY: 20 μm). ( D ) Bio-TEM images of RAW264.7 cells post various treatments (n = 5, scale bars 1.0 μm). Green arrows indicate nanoparticles. ( E , F ) Morphometric analysis determined the mean number and area (μm 2 ) of LDs per cell section. ( G ) Confocal images depicting lipophagy flux in foam cells following different treatments (n = 5 biological replicates, scale bars: 10 μm). ( H - J ) The quantities of acidified autophagosomes (GFP-RFP+), neutral autophagosomes (GFP + RFP+), and LDs labeled with BODIPY were measured per cell for each condition. (K to N) Representative Western blot images and quantitative analysis of LC3, LAMP1, and P62 expression in foam cells. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001.

Article Snippet: RAW264.7 mouse macrophage cells (ATCC® TIB-71; RRID: CVCL_0493) and MCAECs (Procell, CP-M081) were cultured in Dulbecco's Modified Eagle Medium (DMEM) with 10% fetal bovine serum and 1% penicillin-streptomycin at 37 °C in a 5% CO 2 atmosphere.

Techniques: In Vitro, Activation Assay, Staining, Labeling, Western Blot, Expressing

Journal: Bioactive Materials

Article Title: A foam cell-targeted lipophagy restoration strategy stabilizes vulnerable atherosclerotic plaques

doi: 10.1016/j.bioactmat.2026.02.041

Figure Lengend Snippet: In vitro examination of LD degradation in foam cells through fatty acid oxidation and cholesterol efflux. (A ) Schematic diagram of the LD degradation mechanism. ( B , C ) Confocal images and quantitative analysis of LDs colocalization with fatty acids in RAW264.7 cells following different treatments (n = 5, scale bars: 5 μm). ( D , E ) Confocal images of the colocalization of mitochondria with fatty acids and quantified data of fatty acids in RAW264.7 cells under different stimulations (n = 5, scale bars: 5 μm). ( F , G ) Confocal images illustrating mitochondrial colocalization with ATP and corresponding quantification of ATP levels in RAW264.7 cells post various treatments (n = 5, scale bars: 20 μm). ( H ) Diagram illustrating the incorporation of [U- 13 C] palmitic acid into the TCA cycle and the labeling pattern of derived metabolites (n = 3). ( I ) A PCA plot illustrates the cluster separation between the two groups (n = 3). ( J ) Heatmap showing differences in metabolites between the two groups (n = 3). ( K ) Normalized total labeling of each metabolite to [U- 13 C] palmitic acid (n = 3). ( L ) Proportion of (m + 2)-labeled TCA cycle metabolites derived from [U- 13 C] palmitic acid (n = 3). ( M - R ) The study quantified NBD-cholesterol accumulation ( M , P ) and cholesterol efflux facilitated by HDL ( N , O ) and apoA-I ( Q , R ) using confocal imaging across (n = 5, Scale bar = 50 μm). ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001.

Article Snippet: RAW264.7 mouse macrophage cells (ATCC® TIB-71; RRID: CVCL_0493) and MCAECs (Procell, CP-M081) were cultured in Dulbecco's Modified Eagle Medium (DMEM) with 10% fetal bovine serum and 1% penicillin-streptomycin at 37 °C in a 5% CO 2 atmosphere.

Techniques: In Vitro, Labeling, Derivative Assay, Imaging

Journal: Bioactive Materials

Article Title: A foam cell-targeted lipophagy restoration strategy stabilizes vulnerable atherosclerotic plaques

doi: 10.1016/j.bioactmat.2026.02.041

Figure Lengend Snippet: In vivo photoacoustic imaging and analysis of the vulnerability of atherosclerotic plaque. ( A - G ) Ex vivo distribution of HMCN@Cy5.5 , Scr-HMCN@Cy5.5 , and OPN-HMCN@Cy5.5 in various organs—specifically the aorta ( B ), heart ( C ), liver ( D ), spleen ( E ), lung ( F ), and kidney ( G )—from apoE −/− mice at 0, 6, 12, and 24 h post-intravenous injection (n = 3). ( H ) Confocal images demonstrate the colocalization of OPN with CY5.5-labeled nanoparticles in aortic roots (n = 6, scale bars, 200 μm). ( I ) Quantitative analysis of the relative MFI of OPN and CY5.5 in different treatment groups. ( J , K ) Photoacoustic images and quantitative analysis of signal intensities of atherosclerotic plaque in carotid arteries of both healthy and atherosclerosis mice (n = 3). For each animal, longitudinal PA imaging was performed on the same carotid artery at predefined anatomical landmarks across different time points. Photoacoustic images were acquired with depth calibration based on acoustic time-of-flight measurements, converting ultrasound echo delay into depth using the predefined sound velocity in soft tissue. A calibrated depth scale bar is shown in each image, with an effective imaging depth of approximately 7 mm. ( L , M ) Pathological staining of atherosclerotic plaques in the carotid artery and aortic arch includes ORO and Masson staining (scale bar = 200 μm), as well as α -SMA, and CD68 fluorescent staining (scale bar = 100 μm each). ( N - Q ) The statistical analysis of ( N ) ORO staining (namely the percentage of LD area), ( O ) Masson staining (namely the percentage of collagen fiber area), ( P ) α -SMA fluorescent staining (namely the percentage of smooth muscle cell area) and ( Q ) CD68 fluorescent staining (namely the percentage of macrophage-derived foam cell area). ( R ) Vulnerability scores of aortic arch and carotid artery plaques. ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗∗ P < 0.0001.

Article Snippet: Mouse macrophage cell line (RAW264.7) was obtained from the American Type Culture Collection, USA.

Techniques: In Vivo, Imaging, Ex Vivo, Injection, Labeling, Staining, Derivative Assay

Journal: Bioactive Materials

Article Title: A foam cell-targeted lipophagy restoration strategy stabilizes vulnerable atherosclerotic plaques

doi: 10.1016/j.bioactmat.2026.02.041

Figure Lengend Snippet: In vivo atherosclerosis reversal. ( A ) Schematic illustration of the experimental timeline and treatment strategy for establishing a mature, vulnerable atherosclerosis model and evaluating therapeutic interventions. Mice were fed a high-fat diet (HFD) for 12 weeks and then divided into five groups (HFD+ 12W, Saline HFD+, OPN-HMCN@MLT HFD+, Saline HFD−, and OPN-HMCN@MLT HFD−). Except for the HFD+ 12W group, the remaining groups were further maintained for an additional 4 weeks under either HFD or non-HFD conditions with the indicated treatments. ( B , C ) Images of en face ORO-stained aortas ( B ) and quantitative analysis of ORO-positive regions ( C ) from mice subjected to different treatments and diets (n = 6, scale bar: 5 mm). ( D ) Aortic root sections stained by ORO, H&E, α-SMA antibody, Masson's trichrome, CD68 antibody, and MMP-9 antibody, respectively, following various therapeutic procedures (n = 6, scale bar: 500 μm). ( E - J ) Quantitative data of lipid accumulation ( E ), necrotic core area ( F ), collagen area ( G ), MMP-9 level ( H ), VSMC area ( I ), and macrophage-foam cell area ( J ) in aortic root sections. ( K ) Vulnerability scores of aortic root plaque. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001.

Article Snippet: Mouse macrophage cell line (RAW264.7) was obtained from the American Type Culture Collection, USA.

Techniques: In Vivo, Saline, Staining

Journal: Bioactive Materials

Article Title: A foam cell-targeted lipophagy restoration strategy stabilizes vulnerable atherosclerotic plaques

doi: 10.1016/j.bioactmat.2026.02.041

Figure Lengend Snippet: In vivo anti-atherosclerosis effects. ( A ) Diagram illustrating the treatment protocol for apoE −/− mice. ( B , C ) En face ORO staining images and quantitative analysis of the lesion area of aortic lesion areas in apoE −/− mice following various treatments (n = 6, scale bar: 5 mm). ( D ) Quantification of the reduction ratio (versus model) of ORO-positive area to the entire aorta. ( E ) Cross-sectional images of ORO-stained aortic root (scale bars, 500 μm) and brachiocephalic artery (scale bars, 200 μm). n = 6. ( F and G ) Quantitative analysis of the aortic root lesion area ( F ) and the reduction ratio (versus model) of ORO-positive area to the aortic root ( G ). ( H ) Aortic root sections stained by H&E, α-SMA antibody, Masson's trichrome, CD68 antibody, MMP-9 antibody, and OPN antibody, respectively, following various therapeutic procedures (n = 6, scale bar: 500 μm). ( I-M ) Quantitative data of necrotic core area ( I ), collagen area ( J ), VSMC area ( K ), macrophage-foam cell area ( L ), and MMP-9 level ( M ) in aortic root sections. ( N ) Representative TEM images of LDs in the aortic root and arch of apoE −/− mice following various treatments (scale bar: 1 μm). The green arrow indicates elastic fibers. ( O-R ) Quantification of lipid droplet number and average area per cell section, n = 6. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001.

Article Snippet: Mouse macrophage cell line (RAW264.7) was obtained from the American Type Culture Collection, USA.

Techniques: In Vivo, Staining

Journal: Bioactive Materials

Article Title: A foam cell-targeted lipophagy restoration strategy stabilizes vulnerable atherosclerotic plaques

doi: 10.1016/j.bioactmat.2026.02.041

Figure Lengend Snippet: Schematic of the anti-atherosclerotic mechanism of OPN-HMCN@MLT. ( A ) The study commenced with the synthesis of mesoporous carbon nanospheres (MCN) functionalized with an OPN-binding peptide and hyaluronic acid to construct the OPN-HMCN nanoplatform. The OPN-binding peptide was designed to recognize OPN enriched in the extracellular matrix and on the surface of foam cells, thereby enabling selective accumulation in OPN-rich pathological regions. Following OPN recognition, OPN-HMCN@MLT undergoes CD44-dependent endocytosis. Melatonin (MLT), a lipid autophagy–promoting agent, was subsequently encapsulated within the nanocarrier to form OPN-HMCN@MLT. Firstly, the released MLT can bind to and upregulate the expression of PPARα and PPARγ, which then promote the expression of downstream genes (ABCA1, ABCG1, ACOX-1, and CTP1A) and trigger the lipophagy. ( B ) Subsequently, its lipophagy-enhancing effects, including ABCA1/G1-mediated cholesterol efflux and CTP1A/ACOX-1-mediated mitochondrial fatty acid oxidation, were studied to confirm the reversal of foam cell formation. ( C ) These effects eventually promote foam cells to reverse into macrophages. Abbreviations: MCN, mesoporous carbon nanoparticle; OPN, osteopontin; MLT, melatonin; LDL, low-density lipoprotein; ox-LDL, oxidized low-density lipoprotein; PA, Photoacoustic.

Article Snippet: Mouse macrophage cell line (RAW264.7) was obtained from the American Type Culture Collection, USA.

Techniques: Binding Assay, Construct, Expressing

Journal: STAR Protocols

Article Title: Protocol for pro-inflammatory microRNA motif discovery using machine learning

doi: 10.1016/j.xpro.2026.104467

Figure Lengend Snippet: Microscopic images of RAW 264.7 cells in 96-well plate before starvation and transfection (related to step 10) (A) 70% confluency. (B) <50% confluency. Scale bars represent 100 μm.

Article Snippet: RAW 264.7 mouse macrophage cell line , ATCC , Cat#TIB-71; RRID: CVCL_0493.

Techniques: Transfection

Journal: iScience

Article Title: Fabrication of RADA32/Ngf_EE/MSCs composite hydrogel and its protective mechanism against radiation-induced ototoxicity

doi: 10.1016/j.isci.2026.115723

Figure Lengend Snippet: Immunomodulatory mechanism of RNM composite gel (A and B) Flow cytometric analysis of CD86 and CD206 expression in RAW264.7 macrophages after irradiation and co-culture with RN, MSCs, or RNM composite gel in a transwell system (macrophages in lower chamber). Data are represented as the mean ± SEM ( N = 3, t test). (C) Immunofluorescence staining of F4/80 (red) on cochlear sections. Scale bars, 50 μm (a, spiral ganglion; b, basilar membrane; c, stria vascularis; d, spiral ligament). (D) Apoptosis of HEI-OC1 cells analyzed by flow cytometry after radiation exposure and intervention. Data are represented as the mean ± SEM ( N = 3, t test). (E) Expression level of p-p65, a key marker of NF-κB pathway activation, in macrophages after radiation exposure and drug intervention. Data are represented as the mean ± SEM ( N = 3, t test). Significant differences between the groups are indicated by ∗ for p < 0.05, ∗∗ for p < 0.01, ∗∗∗ for p < 0.001, and ∗∗∗∗ for p < 0.0001.

Article Snippet: Mouse bone marrow-derived mesenchymal stem cells (MSCs) and the mouse monocyte/macrophage cell line RAW264.7 were purchased from Procell Life Science & Technology Co., Ltd.

Techniques: Expressing, Irradiation, Co-Culture Assay, Immunofluorescence, Staining, Membrane, Flow Cytometry, Marker, Activation Assay

Journal: Molecular Therapy Oncology

Article Title: Nicotinamide mononucleotide enhances anti-tumor effect by resetting macrophages toward the inflammatory M1-like phenotype

doi: 10.1016/j.omton.2026.201221

Figure Lengend Snippet: High-dose NMN treatment reset macrophages toward the inflammatory M1-like phenotype and enhanced the protective efficacy against mesothelioma (A) The relative changes to the initial body weight of naive mice receiving different treatments: NMN only (NMN), anti-PD1 (α-PD1), and non-treated (NC). (B) Tumor challenge schemes. Technical replicates of N = 5 were adopted for each group. (C) Tumor growth kinetics measured by tumor size. (D) Tumor growth kinetics measured by luminescence intensity. (E) Tumor luminescence intensity among different groups at the endpoint. (F) Frequencies of Gag tetramer-positive CD8 + T cells isolated from the spleen among different groups. Frequencies of PMN-MDSC, M-MDSC, NK, and Treg cells were quantified in the tumor among different groups. (G) Frequencies of M1- and M2-like macrophages among different groups. (H) Density plots representing CD206 − M1-like macrophages and CD206 + M2-like macrophages were gated in CD3 − Ly6C − F4/80 + CD11b + live cells. (I) Frequencies of Ly6C − F4/80 + CD11b + macrophages in total live CD45 + immune cells in the tumor among different groups at day 16 post tumor challenge. (J) Tumor growth kinetics measured by tumor size among NMN treatment only (NMN), NMN treatment plus macrophage depletion (NMN+M Del), and non-treated (NC). Biological replicates of N = 5 were adopted for each group in (I) and (J). two-way ANOVA was applied for (A), (C), (D), and (J). Statistics in (F), (G), and (I) were generated by one-way ANOVA with post hoc correction followed by multiple comparisons. (C and D) Data from one representative experiment of two independent experiments was shown.

Article Snippet: T cells were isolated from mouse spleen using mouse pan T cell isolation kit (Miltenyl Biotec, Cat#130-095-130), while macrophages were prepared using mouse macrophage isolation kit (Miltenyl Biotec, Cat#130-110-434), respectively.

Techniques: Isolation, Generated

Journal: Molecular Therapy Oncology

Article Title: Nicotinamide mononucleotide enhances anti-tumor effect by resetting macrophages toward the inflammatory M1-like phenotype

doi: 10.1016/j.omton.2026.201221

Figure Lengend Snippet: NMN treatment enhanced the activation and cytokine secretion of macrophages (A) Experiment schemes. Four technical replicates were set for each group. Macrophage (B) and T cell (C) purification efficacy was verified by FACS analysis using samples collected before and after isolation. (D) Fold changes in mRNA levels of TNF-α, IL-12β, and CD38 in macrophages collected from supernatant at 24 h post-co-culture. (E) Fold changes in mRNA levels of TNF-α, IL-12β, and CD38 in T cells collected from supernatant at 24 h post-co-culture. (F and G) Cells collected from tumor in mice treated with or without NMN in J at the endpoint were subjected to the FACS analysis. (F) Frequencies of TNF-α, IL-12β, CD38, IFN-γ, and granzyme B expression in macrophages. (G) Frequencies of TNF-α, IL-12β, CD38, IFN-γ, and granzyme B expression in CD8 + T cells. Statistics for (D–G) were generated by the two-tailed Student t test.

Article Snippet: T cells were isolated from mouse spleen using mouse pan T cell isolation kit (Miltenyl Biotec, Cat#130-095-130), while macrophages were prepared using mouse macrophage isolation kit (Miltenyl Biotec, Cat#130-110-434), respectively.

Techniques: Activation Assay, Purification, Isolation, Co-Culture Assay, Expressing, Generated, Two Tailed Test