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Secretion of interleukin-8 by human-derived cell lines infected with Mycobacterium bovis
Patricia Méndez-Samperio1, Janet Palma-Barrios, Abraham Vázquez-Hernández
1Departmento de Inmunología, Escuela Nacional de Ciencias Biológicas, México, D F, México. pmendezs@bios.encb.ipn.mx
Insights
Bacillus Calmette-Guérin (BCG) infection triggers interleukin-8 (IL-8) secretion in human cells. Nuclear factor-kappaB (NF-kappaB) activation and calcium influx are key mechanisms driving this response.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- The variable efficacy of Bacillus Calmette-Guérin (Mycobacterium bovis BCG) necessitates understanding its immune-modulating mechanisms.
- Chemokines, such as interleukin-8 (IL-8), play a crucial role in host defense against tuberculosis.
- Investigating the signaling pathways involved in BCG-induced chemokine production is vital for developing improved tuberculosis vaccines and therapies.
Purpose of the Study:
- To elucidate the roles of nuclear factor-kappaB (NF-kappaB) and intracellular calcium in M. bovis BCG-induced IL-8 production.
- To determine the contribution of NF-kappaB signaling and calcium influx to IL-8 secretion in monocytic (U937) and epithelial (HEp-2) cell lines.
Main Methods:
- Enzyme-linked immunosorbent assay (ELISA) was used to quantify IL-8 concentrations in cell culture supernatants.
- Specific inhibitors of NF-kappaB (sulfasalazine, curcumin) were employed to assess its role in IL-8 secretion.
- Calcium influx was manipulated using ethylenediamine tetraacetic acid (EDTA) and 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl) ester (BAPTA-AM) to evaluate calcium's effect on IL-8 production.
Main Results:
- M. bovis BCG infection significantly elevated IL-8 protein secretion in both U937 and HEp-2 cell lines (p < 0.01).
- Inhibition of calcium influx markedly reduced M. bovis BCG-induced IL-8 secretion.
- Specific NF-kappaB inhibitors, sulfasalazine and curcumin, effectively suppressed M. bovis BCG-induced IL-8 secretion.
Conclusions:
- Activation of the NF-kappaB signaling pathway is a critical mechanism mediating M. bovis BCG-induced IL-8 secretion in monocytic and epithelial cells.
- Calcium influx directly influences IL-8 secretion in response to M. bovis BCG infection in these cell types.
- These findings provide insights into the molecular mechanisms underlying BCG's immunomodulatory effects and potential therapeutic targets for tuberculosis.
Background:
The variable efficacy of bacillus Calmette-Guérin (Mycobacterium bovis BCG) in protecting humans against tuberculosis has prompted a search for the mechanisms through which BCG induces chemokines. In this study, our experiments were designed to determine the role of the transcription factor nuclear factor-kappaB (NF-kappaB) and intracellular calcium in the production of interleukin (IL)-8, a main chemotactic factor, by human-derived monocytic cell line U937 and by a human epithelial HEp-2 cell line infected with M. bovis BCG.
Methods:
The concentrations of IL-8 in culture supernatants of U937 cells or HEp-2 cells infected with M. bovis BCG were determined by enzyme-linked immunosorbent assay. We used sulfasalazine and curcumin, which are well-described inhibitors of NF-kappaB activity, and we used ethylenediamine tetraacetic acid to deplete extracellular Ca2+ or used the cell-permeable agent 1,2-bis (2-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid tetra (acetoxymethyl) ester to chelate releasable intracellular stores of Ca2+ in order to investigate the mechanisms through which M. bovis BCG induces IL-8 secretion in our system.
Results:
The enzyme-linked immunosorbent assay showed that IL-8 protein secretion was elevated in M. bovis-infected cell lines. This effect was statistically significant (p < 0.01). When calcium influx was suppressed in M. bovis-infected cell lines, IL-8 secretion was inhibited. Notably, specific inhibitors of NF-kappaB (sulfasalazine and curcumin) inhibited M. bovis-induced IL-8 secretion from U937 cells or HEp-2 cells.
Conclusions:
Collectively, these results indicate that activation of NF-kappaB is an important signal transduction pathway in M. bovis-induced IL-8 secretion in monocytic or epithelial cells. Furthermore, the results showed that calcium influx had a direct effect on IL-8 secretion in U937 cells or HEp-2 cells infected with M. bovis.
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