Journal: Molecular Therapy. Nucleic Acids

Article Title: ACE-2 blockade and TMPRSS2 inhibition mitigate SARS-CoV-2 severity following cigarette smoke exposure in airway epithelial cells in vitro

doi: 10.1016/j.omtn.2025.102580

Figure Lengend Snippet: SARS-CoV-2 genomic replication is increased in CSE-exposed normal HBE cells and HBE cells derived from COPD patients (A) Transepithelial electrical resistance (TEER) measurements in vehicle-treated, cigarette smoke extract (CSE)-treated, and COPD human bronchial epithelial (HBE) monolayer cultures infected with SARS-CoV-2 or mock infection. The data were obtained by averaging three independent Transwell reads, each of which represented the mean of three separate readings. ( N = 3–5) HBE. Each line corresponds to a distinct donor (3–5 donors in total), and each donor is represented by 3–5 replicates. (B) Quantification of viral load by real-time qPCR of SARS-CoV-2 mRNA at 72 h post-infection with SARS-CoV-2 in CSE- or vehicle-exposed HBE cells. N = 3 different donors, with each line representing a separate donor. (C) Quantification of viral load by real-time qPCR of SARS-CoV-2 mRNA at 72 h post-infection with SARS-CoV-2 in healthy non-smoker or COPD HBE cells. N = monolayers/condition derived from 3 to 5 different donors. (D and E) Representative images using RNAscope in situ hybridization for comparable detection of genomic RNA with only the SARS-CoV-2-specific S probe. Images show CSE- or vehicle-exposed HBE cells, and healthy non-smoker or COPD HBE cells, at 72 h post-inoculation with (D) mock infection or (E) SARS-CoV-2. SARS-CoV-2 (red), TMPRSS2 (green), ACE-2 (white), nuclei (blue). ∗∗ p < 0.01.

Article Snippet: Primary HBE cells were isolated from lung explants at the time of organ transplantation from COPD patients, healthy donors without lung disease, and healthy donors, expanded in submerged culture for one or two passages in bronchial epithelial growth medium (BEGM, Lonza, Walksville, MD), and then seeded on Transwell membranes (Corning, New York, NY) as described previously., , Cells were grown at ALI in PneumaCult-ALI media (Stem Cell Technology) for 3–4 weeks until terminally differentiated.

Techniques: Derivative Assay, Infection, RNAscope, In Situ Hybridization

Journal: bioRxiv

Article Title: Dynamic Culture Improves the Predictive Power of Bronchial and Alveolar Airway Models of SARS-CoV-2 Infection

doi: 10.1101/2025.07.21.665885

Figure Lengend Snippet: (A) Schematic of the PhysioMimix® lung MPS co-culture system. Epithelial cells were seeded on the apical side of the Transwell® insert and human pulmonary microvascular endothelial cells (HPMVECs) on the basolateral side, under air–liquid interface (ALI) and dynamic flow conditions. (B) Representative H&E and Alcian blue-stained histological section of bronchial MPS co-culture after 14 days. Mucus is stained blue; endothelial cells are indicated by white arrows. Scale bar, 50 µm. (C) TEER measurements comparing epithelial monoculture, endothelial monoculture, and epithelial–endothelial co-culture over 14 days under ALI conditions. (D) TEER comparison of bronchial and alveolar co-cultures grown under static or dynamic flow MPS conditions over the 14-day ALI differentiation period. (E) Gene expression of Club cell (SCGB1A1) and Goblet cell (MUC5AC) markers in NHBE cells before culture (NHBE pellet) and after 14 days of differentiation in MPS co- culture. (F) Gene expression of alveolar markers - AT1 (AQP5) and AT2 (SFTPB) - in SAEC cells before culture (SAEC pellet) and after 14 days of MPS co-culture. (G) Immunofluorescence staining of bronchial MPS co-culture tissue for acetylated α-tubulin (yellow), mucus (MUC5AC, green), actin (phalloidin, red), and nuclei (Hoechst 33342, blue). Top row shows epithelial layer; bottom row shows endothelial layer. Scale bar, 100 µm. (H) Immunofluorescence staining of alveolar MPS co-culture tissue after 14 days of differentiation, showing surfactant (SFTPB, green), actin (phalloidin, magenta), and nuclei (Hoechst 33342, blue). Endothelial cells are marked with white arrows. Scale bar, 20 µm.

Article Snippet: All airway models were produced in 24-well, 6mm, 0.4μM pore, PET Transwell® membranes (Corning; 3470) coated with 10 μg/mL type I rat tail collagen (Corning; 354236) in PBS at 37°C for 1 hour.

Techniques: Co-Culture Assay, Staining, Comparison, Gene Expression, Immunofluorescence

Journal: Communications Biology

Article Title: HBV DNA integration gene CCDC91 is oncogenic and a potential therapeutic target for hepatocellular carcinoma

doi: 10.1038/s42003-025-08369-1

Figure Lengend Snippet: A The mRNA level of CCDC91 in CCDC91-overexpressing HCC cells and control cells revealed by qPCR (n = 3). B The protein level of CCDC91 in CCDC91-overexpressing HCC cells and control cells detected by western blot. C Cell proliferation of CCDC91-overexpressing HCC cells and control cells detected by CCK8 assay (n = 3). D , E The migration abilities of CCDC91-overexpressing HCC cells and control cells measured by wound healing assay (n = 3). The migration ( F ) and invasion ( G ) abilities of CCDC91-overexpressing HCC cells and control cells measured by transwell assay (n = 3). H Cell proliferation of CCDC91 knockdown HCC cells and control cells detected by CCK8 assay (n = 3). I , J The migration abilities of CCDC91 knockdown HCC cells and control cells measured by wound healing assay (n = 3). K , L The migration and invasion abilities of CCDC91 knockdown HCC cells and control cells measured by transwell assay (n = 3). Statistical analysis was performed using Student’s t test in ( A , C , E , F , G , H , J , and L ). Data represents mean ± SEM. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Cell migration or invasion was measured using Transwell inserts with an 8 μm pore size culture membrane (Corning, Ithaca, NY, USA) covered without or with Matrigel (BD Biosciences, Franklin Lakes, NJ, USA) before use.

Techniques: Control, Western Blot, CCK-8 Assay, Migration, Wound Healing Assay, Transwell Assay, Knockdown

Journal: Communications Biology

Article Title: HBV DNA integration gene CCDC91 is oncogenic and a potential therapeutic target for hepatocellular carcinoma

doi: 10.1038/s42003-025-08369-1

Figure Lengend Snippet: A Co-IP with antibody against CCDC91 followed by Western blot in HepG2 and PLC/PRF/5 cells (n = 3). B Immunofluorescence staining of CCDC91 and LDHA in CCDC91-overexpressing HCC cells and control cells. C The mRNA level of LDHA in CCDC91-overexpressing HCC cells with or without LDHA knockdown revealed by qPCR (n = 3). D The protein level of LDHA in CCDC91-overexpressing HCC cells with or without LDHA knockdown detected by western blot (n = 3). E Cell proliferation of CCDC91-overexpressing HCC cells with or without LDHA knockdown detected by CCK8 assay (n = 3). ( F – I ). F , G The migration abilities of CCDC91-overexpressing HCC cells with or without LDHA knockdown measured by transwell assay (n = 3). The migration ( H , I ) and invasion ( J , K ) abilities of CCDC91-overexpressing HCC cells with or without LDHA knockdown measured by wound healing and transwell assay, respectively (n = 3). L Glucose levels were detected in CCDC91-overexpressing HCC cells with or without LDHA knockdown (n = 3). M Lactate levels were detected in CCDC91-overexpressing HCC cells with or without LDHA knockdown (n = 3). Statistical analysis was performed using Student’s t test in ( C , E , G , I , K , L , and M ). Data represents mean ± SEM. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Cell migration or invasion was measured using Transwell inserts with an 8 μm pore size culture membrane (Corning, Ithaca, NY, USA) covered without or with Matrigel (BD Biosciences, Franklin Lakes, NJ, USA) before use.

Techniques: Co-Immunoprecipitation Assay, Western Blot, Immunofluorescence, Staining, Control, Knockdown, CCK-8 Assay, Migration, Transwell Assay

Journal: Communications Biology

Article Title: HBV DNA integration gene CCDC91 is oncogenic and a potential therapeutic target for hepatocellular carcinoma

doi: 10.1038/s42003-025-08369-1

Figure Lengend Snippet: A The Kaplan–Meier analysis revealed the association of CCDC91 with the progression free survival of HCC patients that treated with sorafenib from TMA cohort (n = 77). B The Kaplan–Meier analysis revealed the association of CCDC91 with the overall survival of HCC patients that treated with sorafenib from TMA cohort (n = 80). C Cell proliferation of CCDC91-overexpressing HCC cells and control cells with or without sorafenib detected by CCK8 assay for 24 h, 48 h and 72 h separately (n = 3). The migration ( D ) and invasion ( E ) abilities of CCDC91-overexpressing HCC cells with or without sorafenib measured by transwell assay for 18 h (n = 3). F – H Huh-7 cells stably expressing SC or shCCDC91 were injected subcutaneously into the flanks of nude mice administrated without or with sorafenib (n = 5). Representative images of dissected tumors at the end of the experiment were shown. G Tumor growth curves of mice during sorafenib treatment were analyzed (n = 5). And xenografts were weighted (n = 5, H ). All control cells were treated with an equivalent volume of DMSO to account for potential solvent effects, while the experimental groups were treated with sorafenib dissolved in DMSO. Statistical analysis was performed using Student’s t test in ( C , D , E , and H ). Two-way ANOVA was used for tumor volume in G Data represents mean ± SEM. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Cell migration or invasion was measured using Transwell inserts with an 8 μm pore size culture membrane (Corning, Ithaca, NY, USA) covered without or with Matrigel (BD Biosciences, Franklin Lakes, NJ, USA) before use.

Techniques: Control, CCK-8 Assay, Migration, Transwell Assay, Stable Transfection, Expressing, Injection, Solvent

Journal: Communications Biology

Article Title: HBV DNA integration gene CCDC91 is oncogenic and a potential therapeutic target for hepatocellular carcinoma

doi: 10.1038/s42003-025-08369-1

Figure Lengend Snippet: A The protein level of HBx in HepG2 and PLC/PRF/5 cells with or without Ct-HBx transfection detected by western blot (n = 3). B The mRNA level of CCDC91 in HepG2 cells with or without Ct-HBx transfection revealed by qPCR (n = 3). C Co-IP with antibody against HBx followed by Western blot in HepG2 and PLC/PRF/5 cells (n = 3). D Immunofluorescence staining of CCDC91 and HBx in HepG2 and PLC/PRF/5 cells with or without Ct-HBx transfection. E Cell proliferation of HepG2 and PLC/PRF/5 cells with or without Ct-HBx transfection detected by CCK8 assay (n = 3). F , G ) The migration and invasion abilities of HepG2 and PLC/PRF/5 cells with or without Ct-HBx transfection measured by transwell assay (n = 3). H The protein level of HBx, CCDC91, and LDHA in HepG2 and PLC/PRF/5 cells with or without Ct-HBx transfection revealed by western blot (n = 3). I Schematic diagram illustrating the proposed mechanisms of the effects of the HBV-integrated gene CCDC91 on HCC. Statistical analysis was performed using Student’s t test in ( B , E , F , and G ). Data represents mean ± SEM.*, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Cell migration or invasion was measured using Transwell inserts with an 8 μm pore size culture membrane (Corning, Ithaca, NY, USA) covered without or with Matrigel (BD Biosciences, Franklin Lakes, NJ, USA) before use.

Techniques: Transfection, Western Blot, Co-Immunoprecipitation Assay, Immunofluorescence, Staining, CCK-8 Assay, Migration, Transwell Assay