{Reference Type}: English Abstract
{Title}: [Characterization of a 3-dimensional tuberculosis spheroid model constructed using human monocytic THP-1 cells and Bacillus Calmette-Guerin].
{Author}: Guo J;Qiu Y;Hu C;Li D;DU Y;
{Journal}: Nan Fang Yi Ke Da Xue Xue Bao
{Volume}: 43
{Issue}: 12
{Year}: 2023 Dec 20
暂无{DOI}: 10.12122/j.issn.1673-4254.2023.12.14
{Abstract}: OBJECTIVE: To establish a 3-dimensional tuberculosis spheroid model for studying the formation and characteristics of tuberculous granuloma in vivo.
METHODS: Human myeloid leukemia mononuclear THP-1 cells and Bacillus Calmette-Guerin (BCG) were mixed in a 3D cell culture plate and co-cultured in the presence of PMA for 3 days. The growth of the spheroid was examined every 24 h, and the distribution of bacteria, cell survival rate, transformation of the monocytes into macrophages, and penetration of fluorescently labeled nanoparticles into the cell spheroids and tuberculosis spheroids were observed using confocal laser scanning microscopy. The BCG and cell architecture within the 3D tuberculosis spheroid was observed using transmission electron microscopy. Image-iTTM red hypoxia probe, H2O2 test kit, and a waterproof pen PH meter were used to detect the differences in the microenvironment between BCG-infected and non-infected 3D tuberculous spheroids. The utility of this 3D tuberculous spheroids for assessing antibiotic effects of rifampicin and levofloxacin was evaluated by plate colony counting.
RESULTS: In the cell-bacterial suspensions, stable 3-D tuberculous spheroids (50-200 μm) occurred slowly, in which the cells adhered tightly with numerous bacteria in the center, and necrotic cells and monocytederived macrophages were seen within the spheroids. Drug penetration was difficult in the 3D tuberculous spheroids as compared with the non-infected cell spheroids. Transmission electron microscopy revealed the presence of cell necrosis and a large number of BCG in the macrophages in the tuberculous spheroids. The tuberculosis spheroid had a more hypoxic microenvironment than the non-infected cell spheroids with higher H2O2 content and nearly a neutral PH. The tuberculous spheroid model was capable of evaluating the efficacy of anti-tuberculosis drugs, and among them rifampicin showed a stronger antibacterial effect.
CONCLUSIONS: The 3-D tuberculous spheroid model established in this study provides a useful platform for studies of tuberculous granuloma.