Journal: Cell Research

Article Title: Coinfection with influenza A virus enhances SARS-CoV-2 infectivity

doi: 10.1038/s41422-021-00473-1

Figure Lengend Snippet: a, b A549 cells were mock-infected or infected with WSN at an MOI of 0.1. At 12 h.p.i., total RNA was extracted from cells, and ACE2, TMPRSS2, Furin, and CatL mRNA levels ( a ) or NP, Mx1, and ISG54 mRNA levels ( b ) were evaluated via qRT-PCR using the SYBR green method. The data are expressed as fold changes relative to the mock infections. c A549 cells were infected with WSN at an MOI of 0.1. IAV NP protein (red) and ACE2 (green) were detected with an immunofluorescence assay at 12 h.p.i. Scale bars are shown. A549 ( d ), Calu-3 ( e ), and NHBE ( f ) cells were preinfected with WSN at an MOI of 0.1 for 12 h. Cells were then infected with live SARS-CoV-2 at an MOI of 0.01 for another 48 h. Total RNA was extracted from cells, and ACE2 mRNA was evaluated via qRT-PCR using the SYBR green method. The protein expression levels of ACE2, SARS-CoV-2 N gene, IAV NP, and β-actin were measured via western blotting assay. * means increased exposure to visualize ACE2. ( g ) The relative mRNA levels of ACE2 were measured in lung homogenates from the indicated groups, and the protein expression of IAV NP and ACE2 was detected via western blotting. ( d – g ) The data are expressed as fold changes relative to the non-IAV infection control. ( h – i ) To establish ACE2 knockdown cells, A549 cells were transduced with lentivirus encoding the CRISPR-Cas9 system with two guide RNAs targeting ACE2 (sgRNA1 and sgRNA2) or control guide RNA. Cells were infected with live SARS-CoV-2 at an MOI of 0.01 with or without IAV infection using the same procedure described above. The ACE2 (qRT-PCR) ( h ) and SARS-CoV-2 E gene (Taqman-qRT-PCR) ( i ) mRNA levels were detected. The data are expressed as the fold change relative to the non-IAV infection control. Values represent means ± SD of three independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: The primary antibodies used in this study were rabbit polyclonal antibody against ACE2 for immunofluorescence (Sino Biological, 10108-T26) and anti-influenza virus-NP antibody (kindly provided by Professor Ningshao Xia).

Techniques: Infection, Quantitative RT-PCR, SYBR Green Assay, Immunofluorescence, Expressing, Western Blot, Transduction, CRISPR

Journal: PLoS Pathogens

Article Title: Infection of human Nasal Epithelial Cells with SARS-CoV-2 and a 382-nt deletion isolate lacking ORF8 reveals similar viral kinetics and host transcriptional profiles

doi: 10.1371/journal.ppat.1009130

Figure Lengend Snippet: Immunofluorescence staining for expression of ACE2 and TMPRSS2 on (A) primary nasal tissue, and (B) cross-section of NECs. ACE2 or TMPRSS2 staining in the relevant panels are represented in red, βIV-tubulin in green, and nuclear staining with DAPI in blue. Representative images from at least two independent stains are shown, at a magnification of 400x.

Article Snippet: Rabbit polyclonal antibodies against ACE2 [21115-1-AP] (Proteintech), rabbit polyclonal antibody against TMPRSS2 [14437-1-AP] (Proteintech) and mouse monoclonal antibody against βIV-Tubulin [ab11315] (Abcam) were used at 1:500, 1:100, and 1:800, respectively.

Techniques: Immunofluorescence, Staining, Expressing