Journal: Frontiers in Microbiology

Article Title: Impact of age and sex on neuroinflammation following SARS-CoV-2 infection in a murine model

doi: 10.3389/fmicb.2024.1404312

Figure Lengend Snippet: SARS-CoV-2 lineage B.1.135 induced weight loss is higher in older male mice. Male and female C57BL/6J mice at 4, 10, and 16 months of age were infected with 1 × 10 5 FFU of SARS-CoV-2 lineage B.1.351 via intranasal route. Change in body weight was monitored for 7 days. Error bars indicate standard error of mean (SEM) ( n = 3 to 4 in each group).

Article Snippet: The following reaction conditions were used: 95°C for 3 min followed by 40 cycles of 95°C for 10 s, 65.5°C for 30 s, and 72°C for 30 s. A standard curve was generated to determine genome copy numbers in the tissue sample by performing a RT-qPCR using synthetic SARS-CoV-2 RNA control (Twist Bioscience, Cat #104043).

Techniques: Infection

Journal: Frontiers in Microbiology

Article Title: Impact of age and sex on neuroinflammation following SARS-CoV-2 infection in a murine model

doi: 10.3389/fmicb.2024.1404312

Figure Lengend Snippet: Older male mice are more susceptible to SARS-CoV-2 lung infection. Viral load in the lung homogenate was quantified at 3 dpi and 7 dpi by panel (A,B) RT-qPCR for SARS-CoV-2 N gene and expressed as copies of N gene per μg of total RNA, and (C,D) by focus forming assay on Vero E6 cells. Error bars indicate standard error of mean (SEM). The dotted line indicates the limit of detection of the assay ( n = 3 to 4 per group).

Article Snippet: The following reaction conditions were used: 95°C for 3 min followed by 40 cycles of 95°C for 10 s, 65.5°C for 30 s, and 72°C for 30 s. A standard curve was generated to determine genome copy numbers in the tissue sample by performing a RT-qPCR using synthetic SARS-CoV-2 RNA control (Twist Bioscience, Cat #104043).

Techniques: Infection, Quantitative RT-PCR, Focus Forming Assay

Journal: Frontiers in Microbiology

Article Title: Impact of age and sex on neuroinflammation following SARS-CoV-2 infection in a murine model

doi: 10.3389/fmicb.2024.1404312

Figure Lengend Snippet: Pulmonary inflammation in SARS-CoV-2 infected mice. Representative images of H&E stained sections. All infected male (A–C) and female (E–G) mice demonstrated perivascular and interstitial inflammation that was not observed in their uninfected counterparts (D,H , respectively). The inflammatory foci were multifocal (black arrows), composed predominantly of lymphocytes and plasma cells and surrounding small and medium-sized vessels (black star, insets) ( n = 3 males and 3 females in SARS-CoV-2 infected and uninfected group). (A–H) = 40X and insets = 400X.

Article Snippet: The following reaction conditions were used: 95°C for 3 min followed by 40 cycles of 95°C for 10 s, 65.5°C for 30 s, and 72°C for 30 s. A standard curve was generated to determine genome copy numbers in the tissue sample by performing a RT-qPCR using synthetic SARS-CoV-2 RNA control (Twist Bioscience, Cat #104043).

Techniques: Infection, Staining

Journal: Frontiers in Microbiology

Article Title: Impact of age and sex on neuroinflammation following SARS-CoV-2 infection in a murine model

doi: 10.3389/fmicb.2024.1404312

Figure Lengend Snippet: Viral RNA is undetectable in the brain of the majority of C57BL/6J mice. (A) Viral RNA in the brain homogenate of C57BL/6J mice infected with 1 × 10 5 FFU of SARS-CoV-2 was quantified at 3 dpi and 7dpi by RT-qPCR for SARS-CoV-2 N gene and expressed as copies of N gene per μg of total RNA. The dotted line indicates the limit of detection of the assay ( n = 3 to 4 per group). (B) Viral RNA in the lung and brain homogenate of K18-hACE2 mice ( n = 3) infected with 1 × 10 4 FFU of SARS-CoV-2 was determined at 7dpi by RT-qPCR for SARS-CoV-2 N gene. (C) Representative immunofluorescence image of sagittal brain section of K18-hACE2 mice infected with SARS-CoV-2. Brain sections ( n = 3 infected and 2 uninfected mice) were immunostained with rabbit polyclonal antibody to SARS-CoV-2 nucleocapsid (N) protein followed by Alexa Fluor 594 conjugated goat anti-rabbit IgG (red). Nucleus were stained with DAPI (blue).

Article Snippet: The following reaction conditions were used: 95°C for 3 min followed by 40 cycles of 95°C for 10 s, 65.5°C for 30 s, and 72°C for 30 s. A standard curve was generated to determine genome copy numbers in the tissue sample by performing a RT-qPCR using synthetic SARS-CoV-2 RNA control (Twist Bioscience, Cat #104043).

Techniques: Infection, Quantitative RT-PCR, Immunofluorescence, Staining

Journal: Frontiers in Microbiology

Article Title: Impact of age and sex on neuroinflammation following SARS-CoV-2 infection in a murine model

doi: 10.3389/fmicb.2024.1404312

Figure Lengend Snippet: SARS-CoV-2 lineage B.1.135 infection induces an inflammatory response in the brain of C57BL/6J mice despite the absence of virus in the brain at 7 dpi. Relative mRNA expression levels of IL-6, TNF-α, and CCL2 in the (A) lung and (B) brain of C57BL/6J mice infected with SARS-CoV-2 were analyzed by RT-qPCR. The ΔCT values were normalized to Rpl27 gene expression and represented as fold change (2 ΔΔCT ) over uninfected control. Error bars indicate standard error of mean (SEM). The dotted line represents the mean expression level in uninfected control ( n = 3 to 4 per group).

Article Snippet: The following reaction conditions were used: 95°C for 3 min followed by 40 cycles of 95°C for 10 s, 65.5°C for 30 s, and 72°C for 30 s. A standard curve was generated to determine genome copy numbers in the tissue sample by performing a RT-qPCR using synthetic SARS-CoV-2 RNA control (Twist Bioscience, Cat #104043).

Techniques: Infection, Virus, Expressing, Quantitative RT-PCR, Control

Journal: Frontiers in Microbiology

Article Title: Impact of age and sex on neuroinflammation following SARS-CoV-2 infection in a murine model

doi: 10.3389/fmicb.2024.1404312

Figure Lengend Snippet: Brain transcriptional response to SARS-CoV-2 infection. (A) Volcano plot indicating differentially expressed genes (DEGs) (adjusted p < 0.05) in the brains of mice infected with SARS-CoV-2 vs. control. Upregulated DEGs in SARS-CoV-2 infected mice are indicated in red, and downregulated DEGs in SARS-CoV-2 infected mice are indicated in blue. The dotted line corresponds to adjusted p = 0.05. (B) Hierarchical clustering heatmap view of DEGs. DEGs are presented in the dendrogram along the Y -axis. Each column contains expression values for an individual mouse, with groups indicated by the color bars along the X -axis. The expression levels from low to high are represented as a color gradient from blue to red, respectively. (C) Barplot visualization of the top 8 (by adjusted p -value) enriched GO biological process pathways. The significance level is indicated as -Log10 (adjusted p -value). (D) Cnetplot visualization of the genes in the top 8 (by adjusted p -value) enriched GO biological process pathways. Connections between genes and pathway nodes are color coded. The size of each node represents the number of overlapped genes in each GO term and the color represents log-fold change of each gene between groups.

Article Snippet: The following reaction conditions were used: 95°C for 3 min followed by 40 cycles of 95°C for 10 s, 65.5°C for 30 s, and 72°C for 30 s. A standard curve was generated to determine genome copy numbers in the tissue sample by performing a RT-qPCR using synthetic SARS-CoV-2 RNA control (Twist Bioscience, Cat #104043).

Techniques: Infection, Control, Expressing