05a  (Cell Applications Inc)

Journal: Viruses

Article Title: Dissecting Phenotype from Genotype with Clinical Isolates of SARS-CoV-2 First Wave Variants.

doi: 10.3390/v15030611

Figure Lengend Snippet: Figure 2. Evaluation of SARS-CoV-2 variants in primary epithelial cells and K18-hACE2 mouse model. (A) Normal human primary tracheal epithelial cells (NHTE) and normal human primary nasal epithelial cells (NHNE) were inoculated with a MOI = 1 and virus titer were assessed at 1 to 3 dpi with a plaque assay. Data are represented as mean ± standard error of the mean. Virus isolates within the A lineage, WA1/2020 (red), and A.1, UT21 (green), are represented by dashed lines. Virus isolates within the B.1 lineage, UT12 (black), UT23 (navy blue), UT27 (orange), and UT29 (violet), and B.1.577, UT5 (brown), are represented by solid lines. In B-D, female K18-hACE2 mice were intranasally challenged at 104 PFU with either SARS-CoV-2 WA1/2020 (n = 2) or one of six clinical isolates UT5, UT12, UT21, UT23, UT27, or UT29 (n = 3). Graphs of lung viral titers (B), weight and body temperature (C), or selected immune responses in lung (D) are presented.

Article Snippet: Normal human tracheal epithelial cells (NHTE) donated from a 42-year-old Caucasian male were purchased through Cell Applications Cat# 504-05a, Lot# 2559.

Techniques: Virus, Plaque Assay

Journal: Journal of Clinical Investigation

Article Title: SPDEF regulates goblet cell hyperplasia in the airway epithelium

doi: 10.1172/jci29176

Figure Lengend Snippet: Figure 1 Spdef mRNA in mouse respiratory epithelial cells, trachea, and tracheal glands. (A) SPDEF and GAPDH expression was identified by RT-PCR using RNA extracts from human lung adenocarcinoma H441 cells, cervical adenocarcinoma HeLa cells, normal human tracheal epithelial HTEpC cells, SV40 large T immortalized mouse lung epithe- lial MLE-12 cells, mouse lung (mLu), and mouse trachea (mTra). Spdef mRNA was detected in H441 cells, HTEpC cells, and mouse lung and trachea, but not in HeLa or MLE-12 cells. PCR without RT (–) showed no product. (B–E) In situ hybridization for Spdef mRNA was performed on sections of trachea and tracheal glands (B and D) and lung (C and E) in adult mice. Spdef mRNA was detected in the epithelium lining trachea, bronchi (B), and tracheal glands (arrows), but not in bronchioles (Br) or blood vessels (V). Inset shows phase microscopy of the hybridized tracheal glands; original magnifica- tion, ×4. Scale bars: 200 μm. C, cartilage.

Article Snippet: Total RNA was prepared from a mouse lung epithelial cell line (MLE-12), a human pulmonary adenocarcinoma cell line (H441), and human tracheal epithelial cells (HTEpc; Cell Applications Inc.) using TRIzol according to the manufacturer’s protocol (Invitrogen).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, In Situ Hybridization, Microscopy

Journal: Journal of Clinical Investigation

Article Title: SPDEF regulates goblet cell hyperplasia in the airway epithelium

doi: 10.1172/jci29176

Figure Lengend Snippet: Figure 2 SPDEF immunohistochemistry in mouse trachea and tracheal glands. Immunohistochemistry was performed on sections of adult lungs. (A and B) SPDEF staining was detected in nuclei in epithelial cells lining the trachea (A) and tracheal glands (B). (C–F) Immunohistochemistry for SPDEF (A and B), TTF‑1 (C), SOX17 (D), FOXJ1 (E), and SCGB1a1 (F) in tracheal epithelium. Scale bars: 50 μm.

Article Snippet: Total RNA was prepared from a mouse lung epithelial cell line (MLE-12), a human pulmonary adenocarcinoma cell line (H441), and human tracheal epithelial cells (HTEpc; Cell Applications Inc.) using TRIzol according to the manufacturer’s protocol (Invitrogen).

Techniques: Immunohistochemistry, Staining

Journal: Journal of Clinical Investigation

Article Title: SPDEF regulates goblet cell hyperplasia in the airway epithelium

doi: 10.1172/jci29176

Figure Lengend Snippet: Figure 8 Morphometric analysis of goblet cell hyperplasia. (A) Alcian blue stain- ing (μm2/mm) was increased significantly in CCSP-rtTA/TRE2-Spdef mice exposed to doxycycline compared with unexposed mice (P = 0.004; Kruskal-Wallis 1-way ANOVA on ranks). Staining was increased in all 4 categories of airways: cartilagenous, proximal (non- cartilagenous), central, and distal. Pairwise comparisons of the data for the 4 different airway categories demonstrated that the increase in Alcian blue staining was most significant in the proximal airways of the doxycycline-exposed double-transgenic mice. *P = 0.026; Mann-Whit- ney rank-sum test. (B) Alcian blue staining was observed only in air- ways associated with cartilage, not in the epithelia of noncartilagenous airways, in double-transgenic mice without doxycycline. (C) Alcian blue staining was observed in epithelial cells lining these regions conducting airways in the presence of doxycycline. (D) Minimal SPDEF staining was observed in control mice at this antibody dilution. (E) Increased staining for SPDEF was observed in conducting airways of doxycycline- exposed double-transgenic mice. Scale bars: 500 μm.

Article Snippet: Total RNA was prepared from a mouse lung epithelial cell line (MLE-12), a human pulmonary adenocarcinoma cell line (H441), and human tracheal epithelial cells (HTEpc; Cell Applications Inc.) using TRIzol according to the manufacturer’s protocol (Invitrogen).

Techniques: Staining, Transgenic Assay, Control