Delie Roselyn Cerutis

This study builds on our previous findings on the role of salivary lysophosphatidic acid (LPA) species in humans to investigate their presence, together with salivary gland LPA receptor (LPAR) expression in a Porphyromonas gingivalis -infected murine (C57BL/6J) model of periodontal disease (PD). Utilizing LC-MS/MS for LPA analysis alongside confocal LPAR imaging and second harmonic (SHG) imaging for collagen visualization, we compared mouse salivary LPA levels and gland LPAR expression to previously established human and mouse data. The findings reveal that while healthy mouse saliva maintains low homeostatic LPA levels, PD triggers an ∼ 10-fold increase, mirroring the elevation we observed in PD patients. Furthermore, the study confirmed the presence of LPA1, LPA3, and LPA4 within submandibular gland (SMG) tissue. Notably, LPA3 was identified as the most widely distributed subtype, while providing the first evidence of LPA4 expression in adult mouse salivary glands. The presence of multiple LPARs suggests that LPA signaling is a critical factor in salivary gland biology. The documented existence of multiple LPARs within salivary glands indicates that they must be taken into consideration in future research concerning autoimmune conditions, and in pharmacological studies involving drugs that impact salivary gland biology and secretory function.

Oral health is crucial to overall health, and periodontal disease (PDD) is a chronic inflammatory disease. Over the past decade, PDD has been recognized as a significant contributor to systemic inflammation. Here, we relate our seminal work defining the role of lysophosphatidic acid (LPA) and its receptors (LPARs) in the oral system with findings and parallels relevant to cancer. We discuss the largely unexplored fine-tuning potential of LPA species for biological control of complex immune responses and suggest approaches for the areas where we believe more research should be undertaken to advance our understanding of signaling at the level of the cellular microenvironment in biological processes where LPA is a key player so we can better treat diseases such as PDD, cancer, and emerging diseases.

ID 25340 Poster Board 79 Our laboratory has pioneered studying the biology of lysophosphatidic acid (LPA) in oral homeostasis and periodontal disease. LPA is the simplest lipid mediator and plays key homeostatic and inflammatory roles. Saliva is essential for maintaining oral health, and salivary glands are thus pivotal, as they participate in regulating immunity and inflammation, especially in oral disease prevention, oral infection control (Papagerakis et al., 2014), and autoimmunity. Salivary gland secretion and function is also affected by many pharmacological agents. We (Bathena et al. 2011) and others have reported the presence of multiple LPA species in human healthy normal and in periodontal disease saliva. Therefore, we hypothesized that the same LPA species would also be present in mouse saliva, and that mouse salivary glands would also express multiple LPARs. Methods: the simple, sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method we developed in 2011 was used to quantify LPA species (LPA 18:0, LPA 16:0, and LPA 18:1) in C57BL/6 mouse saliva. To further our work, we previously also developed whole-mount in situ LPA receptor (LPAR) subtype localization for various oral soft and hard tissues (Cerutis et al. 2016). Polyclonal antibodies against LPA1, LPA3, and LPA4 were used in indirect immunofluorescence for labeling normal mouse salivary glands using this whole-mount in situ technique. Results: As we reported for humans, LPA species are also present in low, homeostatic concentrations in mouse saliva. As periodontal disease develops, paralleling what we reported in Bathena et al. (2011) for human saliva, some of the LPA species increase approximately 10-fold as well. Confocal microscopy confirmed LPA1, LPA3, and substantial LPA4 labeling; similar but different patterns of distribution were seen for each GPCR. In conclusion, this confirms LPA1 and LPA3 expression (like in NOD mice, Park et al., (2017) but is the first report of the presence of LPA4 in mouse salivary gland tissue. The presence of multiple confirmed LPARs in mouse salivary glands suggests that these receptors need to be factored into not only studies of autoimmune conditions but also for pharmacological studies of drugs affecting salivary gland biology and secretion. Support: NIDCR/NIGMS 1R15DE028687-01(D.R.C).

Mucosal seal formation around dental abutments is critical to the successful integration of dental implants into the human oral cavity. No information exists for how clinically relevant polishing procedures for computer-aided design and computer-aided manufactured (CAD/CAM) zirconia abutments affects cellular responses important to mucosal seal formation. CAD/CAM zirconia was divided into four groups for clinically relevant polishing utilizing commercial polishing heads: control, coarse, coarse plus medium, and coarse plus medium plus fine. Surfaces were analyzed with scanning electron microscopy (SEM), atomic force microscopy (AFM), and optical profilometry (OP). Subsequently, human gingival fibroblasts (HGFs) were seeded onto the zirconia surfaces. Proliferation was measured via a quantitative SEM technique and focal adhesion kinase (FAK) phosphorylation status was measured by an enzyme-linked immunosorbent assay (ELISA). Results showed an increase in proliferation on all polished surfaces as compared to the control. Phosphorylation of FAK at tyrosine 397 (Y397) was up-modulated on the control surfaces. The associated cell adaptation is discussed. In all cases, FAK phosphorylation was greater at 24 h than 48 h. These results suggest that clinicians should be mindful of the effects of abutment polishing methodology, as this may have an impact on early mucosal seal formation.

Melissa S. Lang, DDS, MS/D. Roselyn Cerutis, BS, PhD/Takanari Miyamoto, DDS, PhD, CAGS, MS, MBA/Martha E. Nunn, DDS, MS, PhD: Purpose: The aim of this study was to evaluate the surface characteristics and gingival fibroblast adhesion of disks composed of implant and abutment materials following brief and repeated instrumentation with instruments commonly used in procedures for implant maintenance, stage-two implant surgery, and periimplantitis treatment. Materials and Methods: One hundred twenty disks (40 titanium, 40 titaniumzirconium, 40 zirconia) were grouped into treatment categories of instrumentation by plastic curette, titanium curette, diode microlaser, rotary titanium brush, and no treatment. Twenty strokes were applied to half of the disks in the plastic and titanium curette treatment categories, while half of the disks received 100 strokes each to simulate implant maintenance occurring on a repetitive basis. Following analysis of the disks by optical laser profilometry, disks were cultured with human gingival fibroblasts. Cell counts were conducted from scanning electron microscopy (SEM) images. Results: Differences in surface roughness across all instruments tested for zirconia disks were negligible, while both titanium disks and titaniumzirconium disks showed large differences in surface roughness across the spectrum of instruments tested. The rotary titanium brush and the titanium curette yielded the greatest overall mean surface roughness, while the plastic curette yielded the lowest mean surface roughness. The greatest mean cell counts for each disk type were as follows: titanium disks with plastic curettes, titanium-zirconium disks with titanium curettes, and zirconia disks with the diode microlaser. Conclusion: Repeated instrumentation did not result in cumulative changes in surface roughness of implant materials made of titanium, titanium-zirconium, or zirconia. Instrumentation with plastic implant curettes on titanium and zirconia surfaces appeared to be more favorable than titanium implant curettes in terms of gingival fibroblast attachment on these surfaces.

The small bioactive lipid lysophosphatidic acid (LPA) plays critical roles in both normal physiology and inflammation in many systems. However, its actions are just beginning to be defined in oral biology and pathophysiology.