Journal: Immunology

Article Title: Human Alveolar Macrophages Detect SARS‐CoV‐2 Envelope Protein Through TLR2 and TLR4 and Secrete Cytokines in Response

doi: 10.1111/imm.13922

Figure Lengend Snippet: SARS‐CoV‐2 envelope protein, but not spike protein, induces the secretion of cytokines from human alveolar macrophages. AMs were isolated from BAL fluid of patients undergoing routine bronchoscopy by adherence purification. AMs were pre‐treated with an isotype control antibody (control IgG) (10 μg/mL) or untreated for 1 h and then stimulated with SARS‐CoV‐2 spike protein or envelope protein (both 1 μg/mL). Untreated AMs that were stimulated with LPS (100 ng/mL) or unstimulated were used as positive and negative controls, respectively. Supernatant was collected at 24 h poststimulation. The concentration of cytokines was measured using an MSD multiplex assay ( n = 9). Statistical comparisons were made using the Friedman test with Dunn's multiple comparisons test.

Article Snippet: Supernatant was collected at 24‐h poststimulation, and the concentration of cytokines was measured using a Meso Scale Discovery (MSD) multiplex assay (Human Proinflammatory 7 spot plate, N75008B‐1 or Human Proinflammatory‐ 4 spot plate, N45009B‐1) according to the manufacturer's instructions (Meso Scale Diagnostics, Rockville, Maryland, USA).

Techniques: Isolation, Purification, Control, Concentration Assay, Multiplex Assay

Journal: Immunology

Article Title: Human Alveolar Macrophages Detect SARS‐CoV‐2 Envelope Protein Through TLR2 and TLR4 and Secrete Cytokines in Response

doi: 10.1111/imm.13922

Figure Lengend Snippet: Blocking antibodies against TLR2 and TLR4 alter the cytokine responses of human alveolar macrophages to SARS‐CoV‐2 envelope protein. AMs were isolated and treated with blocking antibodies against TLR2, TLR4, or an isotype control antibody (control IgG) (all 10 μg/mL) and stimulated as previously described. Supernatant was collected at 24‐h post stimulation. The concentration of cytokines was measured using an MSD multiplex assay ( n = 9). Statistical comparisons were made using the Friedman test with Dunn's multiple comparisons test.

Article Snippet: Supernatant was collected at 24‐h poststimulation, and the concentration of cytokines was measured using a Meso Scale Discovery (MSD) multiplex assay (Human Proinflammatory 7 spot plate, N75008B‐1 or Human Proinflammatory‐ 4 spot plate, N45009B‐1) according to the manufacturer's instructions (Meso Scale Diagnostics, Rockville, Maryland, USA).

Techniques: Blocking Assay, Isolation, Control, Concentration Assay, Multiplex Assay

Journal: Immunology

Article Title: Human Alveolar Macrophages Detect SARS‐CoV‐2 Envelope Protein Through TLR2 and TLR4 and Secrete Cytokines in Response

doi: 10.1111/imm.13922

Figure Lengend Snippet: Alveolar macrophages from donors aged over 70 years had higher cytokine responses to SARS‐CoV‐2 envelope protein than younger donors. AMs were isolated, treated,and stimulated as previously described. Supernatant was collected at 24 h poststimulation. The concentration of cytokines was measured using an MSD multiplex assay. The statistical comparison presented here is younger ( n = 5) and older ( n = 4) donor AMs treated with control IgG, then stimulated with envelope protein, or unstimulated. Planned statistical comparisons were made between stimulated cells from donors aged < 70 years versus those from donors aged > 70 years using two‐way ANOVA with Šídák's multiple comparison test.

Article Snippet: Supernatant was collected at 24‐h poststimulation, and the concentration of cytokines was measured using a Meso Scale Discovery (MSD) multiplex assay (Human Proinflammatory 7 spot plate, N75008B‐1 or Human Proinflammatory‐ 4 spot plate, N45009B‐1) according to the manufacturer's instructions (Meso Scale Diagnostics, Rockville, Maryland, USA).

Techniques: Isolation, Concentration Assay, Multiplex Assay, Comparison, Control

Journal: Immunology

Article Title: Human Alveolar Macrophages Detect SARS‐CoV‐2 Envelope Protein Through TLR2 and TLR4 and Secrete Cytokines in Response

doi: 10.1111/imm.13922

Figure Lengend Snippet: Alveolar macrophages from donors aged over 70 years did not have higher cytokine responses to LPS than younger donors. AMs were isolated, treated, and stimulated as previously described. Supernatant was collected at 24 h poststimulation. The concentration of cytokines was measured using an MSD multiplex assay. The statistical comparison presented here is younger ( n = 5) and older ( n = 4) donor untreated AMs stimulated with LPS or unstimulated. Planned statistical comparisons were made between stimulated cells from donors aged < 70 years versus those from donors aged > 70 years using two‐way ANOVA with Šídák's multiple comparison test.

Article Snippet: Supernatant was collected at 24‐h poststimulation, and the concentration of cytokines was measured using a Meso Scale Discovery (MSD) multiplex assay (Human Proinflammatory 7 spot plate, N75008B‐1 or Human Proinflammatory‐ 4 spot plate, N45009B‐1) according to the manufacturer's instructions (Meso Scale Diagnostics, Rockville, Maryland, USA).

Techniques: Isolation, Concentration Assay, Multiplex Assay, Comparison

Journal: Immunology

Article Title: Human Alveolar Macrophages Detect SARS‐CoV‐2 Envelope Protein Through TLR2 and TLR4 and Secrete Cytokines in Response

doi: 10.1111/imm.13922

Figure Lengend Snippet: Alveolar macrophages from smokers had lower cytokine responses to SARS‐CoV‐2 envelope protein than non‐smokers. AMs were isolated, treated, and stimulated as previously described. Supernatant was collected at 24‐h poststimulation. The concentration of cytokines was measured using an MSD multiplex assay. The statistical comparison presented here is smokers' ( n = 5) and non‐smokers' ( n = 4) AMs treated with control IgG, then stimulated with envelope protein, or unstimulated. Planned statistical comparisons were made between stimulated cells from never/ex‐smokers versus those from current smokers using two‐way ANOVA with Šídák's multiple comparison test.

Article Snippet: Supernatant was collected at 24‐h poststimulation, and the concentration of cytokines was measured using a Meso Scale Discovery (MSD) multiplex assay (Human Proinflammatory 7 spot plate, N75008B‐1 or Human Proinflammatory‐ 4 spot plate, N45009B‐1) according to the manufacturer's instructions (Meso Scale Diagnostics, Rockville, Maryland, USA).

Techniques: Isolation, Concentration Assay, Multiplex Assay, Comparison, Control

Journal: Immunology

Article Title: Human Alveolar Macrophages Detect SARS‐CoV‐2 Envelope Protein Through TLR2 and TLR4 and Secrete Cytokines in Response

doi: 10.1111/imm.13922

Figure Lengend Snippet: Alveolar macrophages from smokers did not have lower cytokine responses to LPS than non‐smokers. AMs were isolated, treated, and stimulated as previously described. Supernatant was collected at 24 h poststimulation. The concentration of cytokines was measured using an MSD multiplex assay. The statistical comparison presented here is smokers' ( n = 5) and nonsmokers' ( n = 4) untreated AMs stimulated with LPS, or unstimulated. Planned statistical comparisons were made between stimulated cells from never/ex‐smokers versus those from current smokers using two‐way ANOVA with Šídák's multiple comparison test.

Article Snippet: Supernatant was collected at 24‐h poststimulation, and the concentration of cytokines was measured using a Meso Scale Discovery (MSD) multiplex assay (Human Proinflammatory 7 spot plate, N75008B‐1 or Human Proinflammatory‐ 4 spot plate, N45009B‐1) according to the manufacturer's instructions (Meso Scale Diagnostics, Rockville, Maryland, USA).

Techniques: Isolation, Concentration Assay, Multiplex Assay, Comparison