Aside from MMP28, we also measured MMP13 expres sion, whose levels have been described in the literature to be elevated with degeneration. In our samples, we also found a significant and relatively high increase of MMP13 expression in the grade V degeneration group, compared to all lower grades of degeneration, thus con further information firming previously published data. However, when testing whether inflammation regulates MMP28 expression, we could not find any changes in MMP28 mRNA levels after treatment with LPS, IL 1b or TNF a, although inflammatory mediators regulate many other MMPs, as shown in the literature. Indeed, when measuring changes in MMP13 expression in our samples, we were able to detect a significant increase after stimulation with all three agents.

This clearly indi cates that the absence of MMP28 regulation observed in this study is not due to lack of sensitivity of our model system. As effects on gene expression after stimulation can depend strongly on the used concentrations Inhibitors,Modulators,Libraries Inhibitors,Modulators,Libraries as well as on the chosen time point for analysis, variations in dose and sampling points were considered in this study, yet no effects were observed under any condition. In human keratinocytes, TNF a induced MMP28 at least to a minor degree, while multiple other growth factors and cytokines did not influence its expression levels at all. All this data indicates that compared to other MMPs, MMP28 seems to be rather unresponsive to external inflammatory sti muli in disc cells, although being expressed in degenera tive diseases that are characterized by inflammation.

It should however be noted that, in this part of the study, no distinction was made between annulus fibrosus and nucleus pulposus cells as a clear separation of the two zones is not Inhibitors,Modulators,Libraries possible in later stage Inhibitors,Modulators,Libraries degenerated disc tissue. Considering the fact that no effect was observed in this mixed cell population, it is however unli kely that a significant alteration would have been observed if distinct cell types had been used. As TNF a was not able to induce MMP28 in human IVD cells, we investigated the potential of trichostatin A, a HDAC inhibitor, which was previously shown to strongly regulate MMP28 in HeLa cells. It is assumed that HDAC inhibitors induce MMP28 promoter by acetylation of Inhibitors,Modulators,Libraries spe cificity protein 1, which can alter protein protein interactions and can modify the SP1 containing protein complexes that act at the GC GT boxes.

However, in our experiments, trichostatin A did not have any effect on the expression levels of MMP28 in disc cells, but the sti mulatory www.selleckchem.com/products/azd9291.html effect in HeLa cells could be confirmed in our experimental setting. So far, no other studies have been performed concerning the responsiveness of MMP28 to HDAC inhibitors. Therefore, it is unknown whether most other cell types would show a behavior similar to HeLa cells or to IVD cells.