Investigation of the impact of the Caspr3 receptor in brain microvascular endothelial bEnd.3 cells on invasion function of Escherichia coli K1
Year: 2024
Type: preprint
Abstract: Objective To investigate the role of the Caspr3 receptor in bEnd.3 cells on the invasion function of Escherichia coli K1 strain. Methods 1. Cell Culture and Bacterial Infection:bEnd.3 cells and E. coli K1 (E44) were cultured in vitro. Confocal laser microscopy was used to observe the changes in F-actin and Caspr3 in bEnd.3 cells following bacterial addition. 2. Caspr3 Knockdown: bEnd.3 cells were transfected with lentivirus to establish stable Caspr3 knockdown (Lenti-sh-Caspr3) and null-loaded (Lenti-sh-NC). Transfection efficiency was assessed via Western blot and confocal laser microscopy. 3. Bacterial Adhesion and Invasion Assays: Lenti-sh-NC and Lenti-sh-Caspr3 cells were infected with E. coli E44 to evaluate bacterial adhesion and invasion. 4. Changes in F-actin: Confocal laser microscopy was used to observe the changes in F-actin after E. coli E44 invaded Lenti-sh-Caspr3. Results 1. bEnd.3 cells expressed Caspr3. Compared with the control group, there was no significant change in the fluorescence intensity of Caspr3, F-actin aggregation occurred, and the co-localization with Caspr3 at the aggregation was more significant after E44 infection. 2. Western blot and confocal microscopy confirmed a significant reduction in Caspr3 protein expression in Lenti-sh-Caspr3 compared to bEnd.3 and Lenti-sh-NC. 3. No significant difference in E44 adhesion rate was observed between Lenti-sh-Caspr3 and the control group. However, the invasion rate of E44 in Lenti-sh-Caspr3 was significantly lower than in the control group. 4. F-actin aggregation was significantly reduced in Lenti-sh-Caspr3 after E44 infection. Conclusions bEnd.3 cells expresses Caspr3, which co-localizes with F-actin during E. coli meningitis invasion. Knockdown of Caspr3 did not affect the adhesion of E. coli E44 to host cells, but the number of bacterial invasions was significantly decreased, and F-actin aggregation of the host cells was reduced. Caspr3 may influence the invasion of host cells by E. coli meningitis by regulating F-actin rearrangement.
Source: Research Square
Authors: Zhi-Dong Qiao, Hong Zhao, Lei Xu, Jun-Xian Fu, Tian-Xia Li, Yue-Xin Jia, Wei-Dong Zhao, Guang-Lu Yang
Language: English
Subfield: Neurology
Field: Neuroscience
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W4404840416