Ceramides in the cytoplasm of host cells are attracting interests as second messengers in a variety of cell events, especially pro-inflammatory response. However, membrane structures of most bacteria, as distinct from that of vertebrate, do not contain ceramide, indicating that production of ceramides by bacteria is rare. Nonetheless, phosphoglycerol dihydroceramide (PGDHC), but not LPS, produced by Porphyromonas gingivalis is found abundantly in the human periodontitis lesion. Contrast to the membrane impermeable ceramides derived from vertebrates, we reported that PGDHC penetrates the cellular membrane of host osteoclast precursor (OCP) and promotes cell fusion required for osteoclastogenesis by acting on a non-muscle myosin IIA (Myh9). Myh9 is an intracellular down-regulatory factor for cell fusion event in osteoclastogenesis. In contrast, it was also demonstrated that host ceramides directly act on cathepsin B to activate its catalytic activity. Furthermore, cathepsin B promotes RANKL-mediated osteoclastogenesis via temporally controlled proteolysis of Myh9 for initiation of cell fusion between OCPs. Our preliminary data demonstrated that PGDHC-mediated upregulation of osteoclastogenesis is attenuated by the chemical inhibitor for cathepsin B, indicating that cathepsin B is also engaged in the PGDHC-mediated osteoclastogenesis. Importantly, the intracellular levels of ceramides, either derived from host or bacteria, are regulated by ceramidases in the physiological context. Dysregulated function of acid ceramidase is implicated in the pathogenesis of Farber Disease and other medical complications. We discovered and published that expression of acid ceramidase in patients with periodontitis is diminished. Nonetheless, our knowledges about the ability of acid ceramidase to regulate the level of PGDHC as well as molecular mechanisms underlying PGDHC-mediated osteoclastogenesis are not yet understood well. Thus, we hypothesized that diminished aCDase activity in periodontal lesions retains the pathogenic property of tissue- penetrating PGDHC which, in turn, elevates the osteoclastogenesis via novel Cathepsin B/Myh9 axis in the bone resorption lesion of periodontitis.
The Aim 1 is designed to test the protective role of acid ceramidase against PGDHC-mediated osteoclastognesis in a mouse model of periodontitis. We will evaluate the effect of endogenous acid ceramidase as well as exogenously administered recombinant acid ceramidase on neutralizing the PGDHC's pathogenic activity.
In Aim 2, using loss- and gain-of-function approaches, we will evaluate the engagement of cathepsin B in the PGDHC-mediated OCP fusion via Myh9 proteolysis. The proposed research project will, for the first time, elucidate the molecular mechanism underlying the pathogenically elevated periodontal bone loss targeting the unique P. gingivalis ceramides that promote osteoclastogenesis by modulating the cell fusion regulatory factor, Myh9, and investigate the effects of host-derived acid ceramidase that can potentially counteract such a pathogenic action of PGDHC.
We recently discovered new class of inflammatory ceramides, phosphoglycerol dihydroceramide (PGDHC) produced by Porphyromonas gingivalis, that is not only abundantly present in human periodontitis lesions, but also engaged in promotion of osteoclastogenesis. The proposed studies will investigate molecular mechanism underlying PGDHC-mediated pathologic bone resorption in periodontitis, in relation to the host-derived acid ceramidase that can potentially degrade PGDHC. Once our hypothesis is proven to be true, we will be well placed to develop a novel therapeutic approach for suppression of bone loss in P. gingivalis-associated periodontitis. .