Long-Term Effect of Heat Shock Protein 60 from Actinobacillus actinomycetemcomitans on Epithelial Cell Viability and Mitogen-Activated Protein Kinases

Abstract

Our previous studies showed that bacterial heat shock protein 60 (hsp60) induces cultured epithelial cell proliferation within 24 h. Here we investigated the long-term effects of heat shock protein 60 isolated from Actinobacillus actinomycetemcomitans on skin keratinocyte (HaCaT cell line) viability and the cell signaling involved. Prolonged incubation in the presence of hsp60 increased the rate of epithelial cell death. The number of viable cells in hsp60-treated culture was 37% higher than the number in the control at 24 h but 27% lower at 144 h. A kinetics study of the effect of hsp60 on the phosphorylation of mitogen-activated protein kinases (MAPKs) involving Western blotting with phospho-specific antibodies showed that in addition to a transient early increase in p38 levels, a second peak appeared in keratinocytes 24 h after the addition of hsp60. In contrast, prolonged incubation with hsp60 caused a decrease in the level of phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2) compared with that in the controls, possibly as a result of protein phosphatase activity. We found that hsp60 increased the levels of several phosphatases, including MAP-2, which strongly dephosphorylates ERK1/2. Moreover, hsp60 increased the level of tumor necrosis factor alpha (TNF-␣) in culture medium in a dose-dependent manner. TNF-␣ added to culture showed a cytotoxic effect on epithelial cells, particularly with longer incubation periods. TNF-␣ also induced the phosphorylation of p38. Finally, our results show that bacterial hsp60 inhibited stress-induced synthesis of cellular hsp60. Therefore, several cell behavior changes caused by long-term exposure to bacterial hsp60 may lead to impaired epithelial cell viability.

The Purified and Recombinant Legionella pneumophila Chaperonin Alters Mitochondrial Trafficking and Microfilament Organization

“…Alternatively, HtpB could interact with plasma membrane surface signaling molecules (e.g., Ras) that also signal from endomembranes while in transit to the plasma cell membrane (10,11). Evidence to support an HtpB-mediated signaling mechanism includes the following: (i) the striking morphological resemblance between our altered actin phenotype and that induced in porcine aortic endothelial cells by the constitutive activation of the signaling molecule Rnd1 (2); (ii) bacterial chaperonins, in general, modulate cell signaling pathways (reviewed in reference 51) and, in particular, can activate extracellular signalregulated kinase 1/2-mitogen-activated protein kinase pathways when added to the cell culture medium (88,89); (iii) HtpB modulates cytokine mRNA levels and interleukin-1 secretion in macrophages via a protein kinase C-dependent signaling pathway (64); and (iv) HtpB has the ability to deliver an intracellular signal in Saccharomyces cerevisiae that activates two signaling pathways controlled by Ras2p and results in pseudohyphal growth (66). Moreover, the fact that Lp02 and HtpB-coated beads did not colocalize in the same phagosome (Fig.…”

Fusobacterium nucleatum Increases Collagenase 3 Production and Migration of Epithelial Cells

“…Bacterial infection may modulate the rate of cell proliferation and migration. Bacteria may also modify the effects of cytokines in infected tissue (76). We studied the effect of F. nucleatum on the rate of closure of the epithelial wound.…”

Bartonella bacilliformis GroEL: Effect on Growth of Human Vascular Endothelial Cells in Infected Cocultures

“…12 Actinobacillus actinomycetemcomitans GroEL is mitogenic for cultured epithelial cells at low concentration, but is cytotoxic at higher concentrations or upon prolonged exposure. [13][14][15] The purpose of this study was to examine the effect of B. bacilliformis on HUVEC cocultures and to study the consequences of excess bacterial GroEL during an active infection in vitro.…”