Experimental study of CD40 as a target molecule for targeted treatment of spinal tuberculosis

English Keywords:Osteoblasts  CD40  Target molecule  Mycobacterium tuberculosis

English Abstract:

  【Abstract】 Objectives: To explore the possibility of CD40 as a target molecule for targeted therapy of spinal tuberculosis, and to investigate the differential expression of CD40 molecules on the surface of normal human osteoblasts infected by different multiplicity of mycobacterium tuberculosis. Methods: Normal human osteoblasts from American Type Culture Collection(ATCC) were used in this study and logarithmic phase culture of mycobacterium tuberculosis(H37Ra-GFP) was used for infection experiment. The human osteoblasts were randomly divided into four groups: A, B, C and D. Group A was the control group, which was not infected by H37Ra-GFP. The other groups were all treatment groups, these groups were infected by Mycobacterium tuberculosis with the different multiplicity of infection(MOI) 1∶1000, 1∶100, 1∶10. The expressions of CD40 mRNA in the human osteoblasts induced by Mycobacterium tuberculosis with different infections were detected by real-time quantitative PCR. Western blot was used to analyze the changes of CD40 protein levels on the surface of osteoblasts. Results: The analysis of mycobacterium tuberculosis uptake by osteoblast showed different MOI of bacterial infection caused dose-dependently increasing the uptake. Real-time quantitative PCR results showed that the up-regulated expression levels of CD40 mRNA in human osteoblasts had been observed. There were significant differences(F=74.005, P<0.05) in the expressions of CD40 mRNA in group A(1.0337±0.0715), B(1.4083±0.1145), C(1.7172±0.1294) and D(2.0378±0.1573). Compared with group A, the expression of CD40 mRNA in mycobacterium tuberculosis infection group B, C, D was higher than that in non-infection group A, and there were significant differences(P<0.05). There were differences in group B, C and D when comparing with each other(P<0.05), and the expression of CD40 mRNA from group B to group D was up-regulated. The Western Blot results showed that the expression of CD40 on osteoblast surface was correlated well with the CD40 mRNA expression. Conclusions: The CD40 mRNA levels were dose-dependently up-regulated by different dose of mycobacterium tuberculosis infection. And the CD40 protein levels were correlated well with the mRNA levels, that showed the higher the multiplicity of infection, the more expression of CD40 molecules. Considering CD40 molecules can specifically bind to their corresponding aptamers, this study provides a theoretical basis for targeted treatment of spinal tuberculosis using CD40 molecules as targeting molecules.