Faculty Bibliography

Head and neck squamous cell carcinoma (HNSCC) presents a major public health concern because of delayed diagnosis and poor prognosis. Malignant cells often reprogram their metabolism in order to promote their survival and proliferation. Aberrant glutaminase 1 (GLS1) expression enables malignant cells to undergo increased glutaminolysis and utilization of glutamine as an alternative nutrient. In this study, we found a significantly elevated GLS1 expression in HNSCC, and patients with high expression levels of GLS1 experienced shorter disease-free periods after therapy. We hypothesized that the GLS1 selective inhibitor, bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide (BPTES), which curtails cells' glutamine consumption, may inhibit HNSCC cell growth. Our results support the idea that BPTES inhibits HNSCC growth by inducing apoptosis and cell cycle arrest. Considering that metformin can reduce glucose consumption, we speculated that metformin would enhance the anti-neoplasia effect of BPTES by suppressing malignant cells' glucose utilization. The combination of both compounds exhibited an additive inhibitory effect on cancer cell survival and proliferation. All of our data suggest that GLS1 is a promising therapeutic target for HNSCC treatment. Combining BPTES with metformin might achieve improved anti-cancer effects in HNSSC, which sheds light on using novel therapeutic strategies by dually targeting cellular metabolism.

Our understanding of the microbiome and its role in immunity, cancer initiation, and cancer progression has evolved significantly over the past century. The "germ theory of cancer" was first proposed in the early 20^th century, and shortly thereafter the bacterium Helicobacter pylori and later, Fusobacterium nucleatum were implicated in the development of gastric and colorectal cancers respectively. However, with the development of reliable mouse models and affordable sequencing technologies, the most fascinating aspect of the microbiome-cancer relationship, where microbes undermine cancer immune surveillance and indirectly promote oncogenesis, has only recently been described. In this review, we highlight the essential role of the microbiome in immune system development and maturation. We review how microbe-induced immune activation promotes oncogenesis, focusing particularly on pancreatic carcinogenesis, and show that modulation of the microbiome augments the anti-cancer immune response and enables successful immunotherapy against pancreatic cancer.

Knowledge Transfer Statement: This article discusses the proceedings of the conference organized by the Task Force on Design and Analysis in Oral Health Research on the understanding of the translational evidence on the etiology and pathogenesis of medication-related osteonecrosis of the jaw as well as the clinical protocols on patient management.

Background: Ageing occurs as a complex interaction of cell autonomous and non-cell autonomous mechanisms. Functional decline is a hallmark of aging in multiple tissues, including the intestine, and this process is thought to be driven in part by deterioration in resident stem cell function. The intestine has a rapid turnover of every 3-5 days, owing this to the presence of Lgr5+ intestinal stem cells (ISCs), that resides at the bottom of the crypt along with their niche providing Paneth cells. Therefore, we investigated how this compartment is affected with aging.
Method(s): Four and 22 months old C57BL/6 male mice were used for aging phenotype studies. Young and old mice received rapamycin (4 mg/kg) or salicylate (2 mg/mL) for 28 days. Neutralizing antibodies to IL-1beta, TNF-alpha, IFN-gamma or IgG1 were i.p. injected at a 300 mug dose for 3 weeks. Isolated crypts from Lgr5-EGFP mice were re-suspended in TrypLE Express with Rock inhibitor and DNAse I, filtered and centrifuged. Cells were then re-suspended in FACS buffer containing PE-conjugated anti-CD24 antibody and APC-conjugated anti-Epcam antibody for 15 min at 4degreeC, and analyzed by MoFlo. ISC proliferation was assessed using an ex vivo organoid assay. Briefly, isolated crypts were re-suspended in matrigel, transferred to a 48-well plate to solidify at 37degreeC, and 250 muL crypt culture medium (ADF, Pen/Strep, HEPES, Glutamax, N2, B27, N-acetyl-L-cysteine, Noggin, EGF and R-Spondin) was added to each well and maintained at 37degreeC. The organoid formation was quantified on day 9. Parabiosis surgery was carried out by the Einstein Chronobiosis core.
Result(s): As compared to young mice, old mice harbor declines in ISC proliferation, impaired mucosal barrier integrity, and shifts in the gut immune cell composition, including increased CD8⁺ T cells and dendritic cells (P<0.05). We next generated isochronic (Y-Y, O-O) and heterochronic (Y-O) C57BL/6 male parabionts and observed impaired ISC function in young parabionts exposed to old blood (P<0.05). Rapamycin or salicylate treatment restored ISC function in old mice (P<0.05), without further suppressing mTOR signaling. However, rapamycin reduced plasma cytokines to more youthful levels (P<0.05), suggesting a potential role for inflammation in mediating the stem cell dysfunction with aging. Ex vivo screening assays confirmed that TNFalpha and IFNgamma potently disrupt ISC proliferation (P<0.05), while in vivo treatment with a TNFalpha-or IFNgamma-neutralizing-antibody restored ISC function in old mice (P<0.05). Upon single cell ablation in the crypts, the motion of neighboring cells was dramatically impaired, and cell debris 'lingered' in the crypts of old animals (P<0.05).
Conclusion(s): We propose that intestinal aging happens as a complex interplay of cell autonomous and non-cell autonomous pathways. Moreover, IFN-gamma and TNF-alpha can act as lead progeronic factors to drive a decline in intestinal stem cell function with aging

Singlet oxygen is a potent agent for the selective killing of a wide range of harmful cells, however current delivery methods pose significant obstacles to its widespread use as a treatment method. Limitations include: the need for photosensitizer proximity to tissue because of the short (3.5 µs) lifetime of singlet oxygen in contact with water; the strong optical absorption of the photosensitizer, which limits penetration depth; and hypoxic environments that restrict the concentration of available oxygen. In this article, we describe a novel superhydrophobic singlet oxygen delivery device for the selective inactivation of bacterial biofilms. The device addresses the current limitations by: immobilizing photosensitizer molecules onto inert silica particles; embedding the photosensitizer containing particles into the plastron (i.e. the fluid-free space within a superhydrophobic surface between the solid substrate and fluid layer); distributing the particles along an optically transparent substrate such that they can be uniformly illuminated; enabling the penetration of oxygen via the contiguous vapor space defined by the plastron; and stabilizing the superhydrophobic state while avoiding direct contact of the sensitizer to biomaterials. In this way, singlet oxygen generated on the sensitizer-containing particles can diffuse across the plastron and kill bacteria even deep within hypoxic periodontal pockets. For the first time, we demonstrate complete biofilm inactivation (>5 log killing) of Porphyromonas gingivalis, a bacterium implicated in periodontal disease. The biofilms were cultured on hydroxyapatite discs and exposed to active and control surfaces to assess killing efficiency as monitored by colony counting and confocal microscopy. Two sensitizer particle types: a silicon phthalocyanine sol-gel (Si-Pc), and a chlorin e6 derivative covalently bound to fluorinated silica, were evaluated; the biofilm killing efficiency was found to correlate with the amount of singlet oxygen detected in separate trapping studies. Finally, we discuss applications for such devices in the treatment of periodontitis.

We found that the cancerous pancreas harbors a markedly more abundant microbiome compared with normal pancreas in both mice and humans, and select bacteria are differentially increased in the tumorous pancreas compared with gut. Ablation of the microbiome protects against preinvasive and invasive pancreatic ductal adenocarcinoma (PDA), whereas transfer of bacteria from PDA-bearing hosts, but not controls, reverses tumor protection. Bacterial ablation was associated with immunogenic reprogramming of the PDA tumor microenvironment, including a reduction in myeloid-derived suppressor cells and an increase in M1 macrophage differentiation, promoting TH1 differentiation of CD4⁺T cells and CD8⁺T-cell activation. Bacterial ablation also enabled efficacy for checkpoint-targeted immunotherapy by upregulating PD-1 expression. Mechanistically, the PDA microbiome generated a tolerogenic immune program by differentially activating select Toll-like receptors in monocytic cells. These data suggest that endogenous microbiota promote the crippling immune-suppression characteristic of PDA and that the microbiome has potential as a therapeutic target in the modulation of disease progression.SIGNIFICANCE:We found that a distinct and abundant microbiome drives suppressive monocytic cellular differentiation in pancreatic cancer via selective Toll-like receptor ligation leading to T-cell anergy. Targeting the microbiome protects against oncogenesis, reverses intratumoral immune tolerance, and enables efficacy for checkpoint-based immunotherapy. These data have implications for understanding immune suppression in pancreatic cancer and its reversal in the clinic.Cancer Discov; 8(4);1-14. ©2018 AACR.

The mechanism underlying bone impairment in patients with diabetes mellitus, a metabolic disorder characterized by chronic hyperglycaemia and dysregulation in metabolism, is unclear. Here we show the difference in the metabolomics of bone marrow stromal cells (BMSCs) derived from hyperglycaemic (type 2 diabetes mellitus, T2D) and normoglycaemic mice. One hundred and forty-two metabolites are substantially regulated in BMSCs from T2D mice, with the tricarboxylic acid (TCA) cycle being one of the primary metabolic pathways impaired by hyperglycaemia. Importantly, succinate, an intermediate metabolite in the TCA cycle, is increased by 24-fold in BMSCs from T2D mice. Succinate functions as an extracellular ligand through binding to its specific receptor on osteoclastic lineage cells and stimulates osteoclastogenesis in vitro and in vivo. Strategies targeting the receptor activation inhibit osteoclastogenesis. This study reveals a metabolite-mediated mechanism of osteoclastogenesis modulation that contributes to bone dysregulation in metabolic disorders.

Various surface modifications have been tried for enhancing osseointegration of the dental implants like mechanical and/or chemical treatments and deposition of calcium phosphate coatings. The objective of this research was to develop calcium-phosphate based thin coatings with antibacterial and bioactive properties for potential application in dental implants. Titanium (Ti) discs were immersed in different calcifying solutions: CaP (positive control), F-CaP, Zn-CaP and FZn-CaP and incubated for 24 h. Negative control was uncoated Ti discs. Coated surfaces were characterized using X-ray diffraction, scanning electron microscopy and energy dispersive spectroscopy. Antibacterial properties were tested using Porphyromonas gingivalis because of its strong association with periodontal and peri-implant infections. Bacterial adhesion and colonization were studied at different timepoints. The coated surfaces had compositional characteristics similar to that of bone mineral and they inhibited the growth, colonization and adherence of P. gingivalis, resulted in reduced thickness of biofilms and bacterial inhibition in the culture medium as compared to the positive and negative controls (p < 0.05). There was no significant difference between the experimental groups (p > 0.05). All CaP coatings demonstrated in vitro bioactivity (formed carbonate hydroxyapatite when immersed in SBF). Such coatings can enhance osseointegration and prevent infection in implants, thereby improving the success rates of implants

People with Down syndrome (DS) are at an increased risk for Alzheimer's disease (AD). After 60 years of age, >50% of DS subjects acquire dementia. Nevertheless, the age of onset is highly variable possibly because of both genetic and environmental factors. Genetics cannot be modified, but environmental risk factors present a potentially relevant intervention for DS persons at risk for AD. Among them, inflammation, important in AD of DS type, is potential target. Consistent with this hypothesis, chronic peripheral inflammation and infections may contribute to AD pathogenesis in DS. People with DS have an aggressive form of periodontitis characterized by rapid progression, significant bacterial and inflammatory burden, and an onset as early as 6 years of age. This review offers a hypothetical mechanistic link between periodontitis and AD in the DS population. Because periodontitis is a treatable condition, it may be a readily modifiable risk factor for AD.