mouse brain endothelial cells bend3  (ATCC)

Journal: Frontiers in Immunology

Article Title: Serum EMMPRIN/CD147 promotes the lung pre-metastatic niche in a D2A1 mammary carcinoma mouse model

doi: 10.3389/fimmu.2025.1568578

Figure Lengend Snippet: EMMPRIN promotes angiogenesis in the lung PMN. Mice were injected with the different tumor cells and treatments as described in the legend of Figure 1 . (A) Representative images of the endothelial cell marker CD31 staining in the lungs, and (C) their quantitation (n=9 for the D2A1-WT group, n=8 for the D2A1-KD group, n=7 for the healthy group, n=7 for the D2A1-WT + m161-pAb group, n=9 for the Healthy + rec. EMMRPIN group). Bar size is 100 μm. (B) Representative images of bEND3 endothelial cell migration length after their incubation for 18 h with serum-starvation media containing 5µg of lung lysates from the different experimental groups. (D) Quantitation of the migration length (n=14 for the D2A1-WT group, n=10 for the D2A1-KD group, n=8 for the healthy group, n=9 for the D2A1-WT + m161-pAb group, n=7 for the Healthy + rec. EMMRPIN group). Bar size is 150 μm. Data are presented as mean ± SEM. Three groups were analyzed using one-way ANOVA followed by Bonferroni’s post-hoc test, and two groups were compared using the non-parametric two-tailed Mann-Whitney t test. The increased CD31 staining of endothelial cells in the mice implanted with the D2A1-WT cells and their longer migration distance suggests that they proliferate and migrate, two properties necessary for angiogenesis, more than the endothelial cells in healthy mice or mice implanted with the D2A1-KD cells. The importance of EMMPRIN to this process is exemplified by the results from the addition of h161-pAb or the injection of recombinant EMMPRIN.

Article Snippet: The mouse brain endothelial cell line bEND3 (ATCC CRL-2299) was cultured in the same full medium that contained 1% glutamine, whereas D2A1 full medium contained 2% glutamine.

Techniques: Injection, Marker, Staining, Quantitation Assay, Migration, Incubation, Two Tailed Test, MANN-WHITNEY, Recombinant

Journal: CNS Neuroscience & Therapeutics

Article Title: Inhibition of HDAC6 promotes microvascular endothelial cells to phagocytize myelin debris and reduces inflammatory response to accelerate the repair of spinal cord injury

doi: 10.1111/cns.14439

Figure Lengend Snippet: Phagocytosis of myelin debris by microvascular endothelial cells (MECs) increases the secretion of inflammatory factors. (A, B) MECs were inoculated on the matrix glue, and the x‐y and x‐z views show the formation of the tubular structure, indicating the successful generation of a vascular‐like structure (CD31, red). (C) After 72 h of coculture, we examined the dynamic process of myelin debris entering vascular‐like structures using x‐y and x‐z views (myelin debris, green): 1. The myelin debris was close to the vascular‐like structure. 2. The myelin debris had just come into contact with the vascular‐like structure. 3. The myelin debris had completely entered the vascular‐like structure. (D) The microstructure of MECs was observed using a transmission electron microscope. The red arrow indicates endocytosis of myelin debris. (E‐H) Levels of IL‐1β, IL‐6, MCP‐1, and TNF‐α in the supernatant were examined via ELISA (myelin debris was cocultured with MECs in the model group). (I) A transwell experiment was performed to examine the effect of phagocytosis of myelin debris by MECs on macrophage recruitment (macrophages through micropores were stained purple). All data are expressed as the mean ± standard deviation ( n ≥ 3 replicates per group). ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse brain microvascular endothelial cells (BEND3) and mouse monocytic macrophage leukemia cells (RAW264.7) were purchased from Procell Life Science & Technology Co., Ltd. (CL‐0598, CL‐0190) and were cultured in DMEM under high‐glucose conditions (containing 10% FBS and 1% P/S).

Techniques: Transmission Assay, Microscopy, Enzyme-linked Immunosorbent Assay, Staining, Standard Deviation

Journal: CNS Neuroscience & Therapeutics

Article Title: Inhibition of HDAC6 promotes microvascular endothelial cells to phagocytize myelin debris and reduces inflammatory response to accelerate the repair of spinal cord injury

doi: 10.1111/cns.14439

Figure Lengend Snippet: Opsonization of IgM or IgG affects the efficiency of phagocytosis. (A–C) The efficiency with which microvascular endothelial cells (MECs) phagocytize myelin debris under different conditions was examined via ELISA, flow cytometry, and immunofluorescence (myelin debris, green). In the absence of serum or in case of inactivation of IgM and IgG, supplementation with IgM, IgG, or myelin debris opsonized by IgM or IgG increased the efficiency of phagocytosis. (D, E) IgM and IgG were added at concentrations of 0 μg/mL, 10 μg/mL, 100 μg/mL, 200 μg/mL, and 400 μg/mL to interfere with the coculture system (myelin debris, red). Immunofluorescence and ELISA experiments revealed a positive correlation between the concentration of IgM or IgG and the efficiency with which MECs phagocytized myelin debris. (F) After collecting the cell supernatant, ELISA experiments were performed without serum and with inactivation of IgG; supplementation of IgG or myelin debris opsonized by IgG had no effect on MCP‐1 levels. However, when experiments were performed without serum, supplementation of IgM or myelin debris opsonized by IgM upregulated MCP‐1 levels. All data are expressed as the mean ± standard deviation ( n ≥ 3 replicates per group). ns p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse brain microvascular endothelial cells (BEND3) and mouse monocytic macrophage leukemia cells (RAW264.7) were purchased from Procell Life Science & Technology Co., Ltd. (CL‐0598, CL‐0190) and were cultured in DMEM under high‐glucose conditions (containing 10% FBS and 1% P/S).

Techniques: Enzyme-linked Immunosorbent Assay, Flow Cytometry, Immunofluorescence, Concentration Assay, Standard Deviation

Journal: CNS Neuroscience & Therapeutics

Article Title: Inhibition of HDAC6 promotes microvascular endothelial cells to phagocytize myelin debris and reduces inflammatory response to accelerate the repair of spinal cord injury

doi: 10.1111/cns.14439

Figure Lengend Snippet: Tubastatin‐A increases the efficiency with which microvascular endothelial cells (MECs) phagocytize myelin debris and decrease inflammatory secretion. (A, B) Tubastatin‐A was used to interfere with the coculture system (myelin debris, green). Immunofluorescence and ELISA experiments revealed that Tubastatin‐A increased the efficiency with which MECs phagocytize myelin debris and upregulated intracellular MBP levels. (C–G) Tubastatin‐A was used to interfere with the coculture system. Western blot and ELISA experiments revealed that Tubastatin‐A reduced IL‐1 β, IL‐6, MCP‐1, and TNF‐α content in the cell supernatant. (H) The effect of phagocytosis of myelin debris by MECs on macrophage recruitment (macrophages passed through micropores were stained purple) was examined in a transwell experiment. All data are expressed as the mean ± standard deviation ( n ≥ 3 replicates per group). ns p > 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse brain microvascular endothelial cells (BEND3) and mouse monocytic macrophage leukemia cells (RAW264.7) were purchased from Procell Life Science & Technology Co., Ltd. (CL‐0598, CL‐0190) and were cultured in DMEM under high‐glucose conditions (containing 10% FBS and 1% P/S).

Techniques: Immunofluorescence, Enzyme-linked Immunosorbent Assay, Western Blot, Staining, Standard Deviation

Journal: CNS Neuroscience & Therapeutics

Article Title: Inhibition of HDAC6 promotes microvascular endothelial cells to phagocytize myelin debris and reduces inflammatory response to accelerate the repair of spinal cord injury

doi: 10.1111/cns.14439

Figure Lengend Snippet: Tubastatin‐A promotes microvascular endothelial cells (MECs) to phagocytize myelin debris by regulating the HDAC6‐mediated autophagy‐lysosome pathway. (A) Tubastatin‐A was used to interfere with the coculture system. HDAC6 expression and the endocytosis of myelin debris were examined using immunofluorescence (myelin debris in green and HDAC6 in pink). (B) Tubastatin‐A and Baf‐A1 were used to interfere with the coculture system. LC3B expression and the endocytosis of myelin debris were examined using immunofluorescence (myelin debris in green and LC3B in pink). (C) The microstructure of MECs was observed using a transmission electron microscope. The red arrow indicates autophagy. (D, E) Expression levels of microtubule‐related and autophagy‐related proteins were examined via Western blotting. (F) The binding levels of Syntaxin17 and VAMP8 were examined using Co‐IP experiments.

Article Snippet: Mouse brain microvascular endothelial cells (BEND3) and mouse monocytic macrophage leukemia cells (RAW264.7) were purchased from Procell Life Science & Technology Co., Ltd. (CL‐0598, CL‐0190) and were cultured in DMEM under high‐glucose conditions (containing 10% FBS and 1% P/S).

Techniques: Expressing, Immunofluorescence, Transmission Assay, Microscopy, Western Blot, Binding Assay, Co-Immunoprecipitation Assay

Journal: CNS Neuroscience & Therapeutics

Article Title: Inhibition of HDAC6 promotes microvascular endothelial cells to phagocytize myelin debris and reduces inflammatory response to accelerate the repair of spinal cord injury

doi: 10.1111/cns.14439

Figure Lengend Snippet: Phagocytosis of myelin debris by microvascular endothelial cells (MECs) may play a role in the repair of SCI. (A, B) Following the generation of the spinal cord injury (SCI) model, the injured spinal cord was removed for HE (overall structure and inflammatory infiltration) and Nissl staining (neuronal morphology and intracellular condition) on days 1, 3, and 7 postinjury. (C–G) On the first, third, and seventh days after SCI, levels of MCP‐1, IL‐6, IL‐1β, TNF‐α, and IL‐10 in the serum were examined via ELISA. (H) On the third day after SCI, immunofluorescence experiments were used to examine the distributions of microglia (CD11b, red), astrocytes (GFAP, green), and MECs (CD31, pink). (I) On the third day after SCI, phagocytosis of myelin debris by MECs was examined using 3D confocal microscopy. The process of myelin debris entering vascular‐like structures was observed using x‐y, y‐z, and x‐z views (MBP in green, CD31 in red). All data are expressed as the mean ± standard deviation ( n ≥ 3 replicates per group). ns p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse brain microvascular endothelial cells (BEND3) and mouse monocytic macrophage leukemia cells (RAW264.7) were purchased from Procell Life Science & Technology Co., Ltd. (CL‐0598, CL‐0190) and were cultured in DMEM under high‐glucose conditions (containing 10% FBS and 1% P/S).

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Confocal Microscopy, Standard Deviation

Journal: CNS Neuroscience & Therapeutics

Article Title: Inhibition of HDAC6 promotes microvascular endothelial cells to phagocytize myelin debris and reduces inflammatory response to accelerate the repair of spinal cord injury

doi: 10.1111/cns.14439

Figure Lengend Snippet: Tubastatin‐A promotes SCI repair by promoting microvascular endothelial cells (MECs) to phagocytize myelin debris and inhibit inflammatory secretion. (A, B) Tubastatin‐A was used to intervene in mice with spinal cord injury (SCI). On the third day after SCI, HE staining was performed to examine changes in global structure and inflammatory infiltration, while Nissl staining was performed to examine changes in neuronal morphology and intracellular conditions. (C–J) On the third day after SCI, ELISA was performed to examine changes in MCP‐1, IL‐6, IL‐1β, TNF‐α, IL‐10, C3, IgM, and IgG levels in the serum. (K) On the third day after SCI, flow cytometry was performed to examine the proliferation and differentiation of B cells in the mouse spleen. (L) On the third day after SCI, the phagocytosis of myelin debris by MECs was detected using 3D confocal microscopy. The process of myelin debris entering vascular‐like structures was observed using x‐y, y‐z, and x‐z views (MBP in green, CD31 in red). (M) On the third day after SCI, levels of HDAC6 (rose red), LC3B (green), NLRP3 (red), and CD31 (pink) expression were examined using immunofluorescence. (N) The co‐localization of HDAC6 (rose red) and NLRP3 (red) was detected based on the results of immunofluorescence experiments. All data are expressed as the mean ± standard deviation ( n ≥ 3 replicates per group). ns p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse brain microvascular endothelial cells (BEND3) and mouse monocytic macrophage leukemia cells (RAW264.7) were purchased from Procell Life Science & Technology Co., Ltd. (CL‐0598, CL‐0190) and were cultured in DMEM under high‐glucose conditions (containing 10% FBS and 1% P/S).

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Confocal Microscopy, Expressing, Immunofluorescence, Standard Deviation