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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
Codon Optimized Pljm1 Sars Cov 2 N Flag, supplied by Sino Biological, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
Pcmv3 Sars Cov 1 Spike Flag, supplied by Sino Biological, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
Codon Optimized R2 Orfs And Other Dna Modules, supplied by GenScript corporation, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min <t>before</t> <t>Codon-optimized</t> <t>pLJM1-SARS-CoV-2</t> N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001
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SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min before Codon-optimized pLJM1-SARS-CoV-2 N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001

Journal: Molecular Medicine

Article Title: The SARS-CoV-2 nucleocapsid protein induces microglia senescence-mediated cognitive impairment via Glycolysis

doi: 10.1186/s10020-025-01410-3

Figure Lengend Snippet: SARS-CoV-2 N protein drives the senescence of BV2 microglial cells by triggering mitochondrial dysfunction. BV2 microglial cells were treated with Mdivi-1 (100 nM) 30 min before Codon-optimized pLJM1-SARS-CoV-2 N-FLAG transfection. A Representative image of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). B Mitochondrial morphology stained with TOM20 in BV2 cells (bar = 2 μm). C - D The expression levels of p-DRP1 and DRP1 proteins in BV2 microglial cells detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to DRP1) in the experimental group relative to that in the control group. E Representative images of BV2 cells loaded with the mitochondrial membrane potential indicator JC-1 (bar = 20 μm). F The expression levels of p19 , p21 , p53 , Il-1β , and Tnf-α mRNA in BV2 microglial cells detected by real-time PCR ( n = 3). G SA-β-gal staining was performed after transfection of Codon-optimized pLJM1-SARS-CoV-2 N-FLAG and treatment with Mdivi-1 for 36 h (bar = 10 μm). H The fluorescence intensity of Ki67 (green) was detected by immunofluorescence (bar = 50 μm). I - J The expression levels of p-DRP1, p53, p21, p16, and γ-H2AX proteins in BV2 microglial cells were detected by Western blot ( n = 3). Bar graph data represent the mean ± SD of the target protein intensity (normalized to α-tubulin) in the experimental group relative to that in the control group. Comparisons between the two groups were made with an unpaired t -test. Differences among multiple groups were performed using ANOVA. * P < 0.05, ** P < 0.01, and *** P < 0.001

Article Snippet: Codon-optimized pLJM1-SARS-CoV-2 N-FLAG was cloned from SinoBiological (cat# VG40588-NF) using BamHI and EcoRI.

Techniques: Transfection, Membrane, Staining, Expressing, Western Blot, Control, Real-time Polymerase Chain Reaction, Fluorescence, Immunofluorescence