Comparison of the Effects of Mycobacterium tuberculosis H37Rv and Brucella ovis Infiltration on Tumor Necrosis Factor-Alpha and Interleukin-1 Beta Expression in Osteoblasts.

Abstract

Objective: This study aims to compare the expression levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β) in osteoblasts infiltrated with Mycobacterium tuberculosis H37Rv (H37Rv) and Brucella ovis to understand the differential bone destruction in spinal tuberculosis (STB) versus Brucella spondylitis (BS). Methods: Primary osteoblasts were isolated and cultured from the cranial bones of 2-5 days old mice and characterized by alkaline phosphatase (ALP) staining and alizarin red staining (ARS). H37Rv and B. ovis were cultured to the logarithmic phase, and transfection solutions were prepared. Osteoblasts were infiltrated with these bacteria at various multiplicities of infection (MOI) and time points. Cell survival post-infiltration was assessed using CCK-8 to determine optimal infection conditions. Osteoblasts were divided into three groups: the H37Rv group (infiltrated with optimal MOI H37Rv), the B. ovis group (infiltrated with optimal MOI B. ovis), and a negative control group. TNF-α and IL-1β expression in the cytoplasm was observed using immunohistochemical staining, whereas their levels in cell supernatants were measured using enzyme-linked immunosorbent assay. Protein expression was analyzed by Western blot. Differences between groups were compared with using one-way analysis of variance and t-tests, with p < 0.05 indicating statistical significance. Results: Both H37Rv and B. ovis infiltrated osteoblasts, substantially increasing TNF-α and IL-1β expression. The H37Rv group showed substantially higher levels of TNF-α and IL-1β compared with the B. ovis group (p < 0.05). Conclusion: Infiltration of osteoblasts with H37Rv and B. ovis substantially increases TNF-α and IL-1β expression, with higher levels observed in H37Rv-infected osteoblasts. This overexpression may contribute to the more severe vertebral bone destruction seen in STB compared with BS.