Establishment of an Animal Model of Bisphosphonate-Related Osteonecrosis of the Jaws in Spontaneously Diabetic Torii Rats.

BACKGROUND: We evaluated the side effects of bisphosphonate (BP) on tooth extraction socket healing in spontaneously diabetic Torii (SDT) rats, an established model of non-obese type 2 diabetes mellitus, to develop an animal model of BP-related osteonecrosis of the jaws (BRONJ). MATERIALS AND METHODS: Male Sprague-Dawley (SD) rats and SDT rats were randomly assigned to the zoledronic acid (ZOL)-treated groups (SD/ZOL or SDT/ZOL) or to the control groups (SD/control or SDT/control). Rats in the SD/ZOL or SDT/ZOL groups received an intravenous bolus injection of ZOL (35 µg/kg) every 2 weeks. Each group consisted of 6 rats each. Twenty-one weeks after ZOL treatment began, the left maxillary molars were extracted. The rats were euthanized at 2, 4, or 8 weeks after tooth extraction, and the total maxillae were harvested for histological and histochemical studies. RESULTS: In the oral cavity, bone exposure persisted at the tooth extraction site in all rats of the SDT/ZOL group until 8 weeks after tooth extraction. In contrast, there was no bone exposure in SD/control or SDT/control groups, and only 1 of 6 rats in the SD/ZOL group showed bone exposure. Histologically, necrotic bone areas with empty lacunae, microbial colonies, and less invasion by inflammatory cells were observed. The number of tartrate-resistant acid phosphatase-positive osteoclasts was lower in the SDT/ZOL group than in the SD/control group. The mineral apposition rate was significantly lower in the SDT/ZOL group compared with the SD/control group. CONCLUSIONS: This study demonstrated the development of BRONJ-like lesions in rats and suggested that low bone turnover with less inflammatory cell infiltration plays an important role in the development of BRONJ.

Cytoreductive surgery under aminolevulinic acid-mediated photodynamic diagnosis plus hyperthermic intraperitoneal chemotherapy in patients with peritoneal carcinomatosis from ovarian cancer and primary peritoneal carcinoma: results of a phase I trial.

BACKGROUND: We conducted a phase I clinical trial to evaluate the sensitivity, specificity, and safety of cytoreductive surgery (CRS) under aminolevulinic acid-mediated photodynamic diagnosis (ALA-PDD) plus hyperthermic intraperitoneal chemotherapy (HIPEC) on 20 patients with peritoneal carcinomatosis (PC) from ovarian cancer and primary peritoneal carcinoma (PPC). PATIENTS AND METHODS: Patients took 5-aminolevulinic acid (5-ALA) at a dose of 20 mg/kg orally with 50 mL of water 2 h before surgery. During surgery, the abdominal cavity was observed under blue light (wavelength of 440 nm) before and after CRS plus HIPEC. Specimens were excised and submitted for pathological examination to evaluate the specificity of ALA-PDD. Postoperative course was closely monitored and detailed information was recorded. RESULTS: CRS under ALA-PDD plus HIPEC was performed 21 times in 20 patients with PC (16 ovarian cancer, 4 PPC) between June 2011 and October 2013. With the exception of 1 (5 %) patient, strong red fluorescence was detected in 19 patients with ovarian cancer, with a sensitivity of 95 %. All specimens from red fluorescent lesions were invaded by cancer cells, with a specificity of 100 %. No severe adverse events occurred during the perioperative period, with the exception of some abnormal laboratory results and mild complications. All patients were alive until the last follow-up. CONCLUSION: ALA-PDD provided a high sensitivity and specificity in detecting peritoneal metastasis in patients with PC from ovarian serous carcinoma and PPC. CRS under ALA-PDD plus HIPEC was a feasible and safe treatment option for patients with PC from ovarian cancer and PPC.

Functionally cloned pdrM from Streptococcus pneumoniae encodes a Na(+) coupled multidrug efflux pump.

Multidrug efflux pumps play an important role as a self-defense system in bacteria. Bacterial multidrug efflux pumps are classified into five families based on structure and coupling energy: resistance-nodulation-cell division (RND), small multidrug resistance (SMR), major facilitator (MF), ATP binding cassette (ABC), and multidrug and toxic compounds extrusion (MATE). We cloned a gene encoding a MATE-type multidrug efflux pump from Streptococcus pneumoniae R6, and designated it pdrM. PdrM showed sequence similarity with NorM from Vibrio parahaemolyticus, YdhE from Escherichia coli, and other bacterial MATE-type multidrug efflux pumps. Heterologous expression of PdrM let to elevated resistance to several antibacterial agents, norfloxacin, acriflavine, and 4',6-diamidino-2-phenylindole (DAPI) in E. coli KAM32 cells. PdrM effluxes acriflavine and DAPI in a Na(+)- or Li(+)-dependent manner. Moreover, Na(+) efflux via PdrM was observed when acriflavine was added to Na(+)-loaded cells expressing pdrM. Therefore, we conclude that PdrM is a Na(+)/drug antiporter in S. pneumoniae. In addition to pdrM, we found another two genes, spr1756 and spr1877,that met the criteria of MATE-type by searching the S. pneumoniae genome database. However, cloned spr1756 and spr1877 did not elevate the MIC of any of the investigated drugs. mRNA expression of spr1756, spr1877, and pdrM was detected in S. pneumoniae R6 under laboratory growth conditions. Therefore, spr1756 and spr1877 are supposed to play physiological roles in this growth condition, but they may be unrelated to drug resistance.

The promotional effect of IL-22 on mineralization activity of periodontal ligament cells.

OBJECTIVES: Interleukin (IL)-22 acts on non-immune cells to induce anti-microbial responses, protection from tissue damage, and enhance cell regeneration. However, little is known about the involvement of IL-22 in periodontal biology. This study investigated the biological effects of IL-22 on periodontal ligament (PDL) cells as part of studies to assess the involvement of IL-22 in periodontal disease. MATERIALS AND METHODS: Gene expression levels of IL-22 and its receptors in PDL cells and gingival tissue samples were evaluated by real-time PCR. Proliferative responses and mineralized-matrix forming activities of PDL cells were examined in the presence and absence of IL-22. RESULTS: In contrast to the expression of IL-22 receptors detected in PDL tissues and their cell lines, gingival tissues showed modest or no gene expressions of IL-22. The production of several cytokines including IL-11, IL-8 and CCL2 was upregulated by IL-22 treatment of PDL cells in a dose-dependent manner. IL-22 treatment had no effect on the proliferative response in PDL cells. Meanwhile, IL-22 precipitated mineralized nodule formation and induced gene expressions of RUNX2, MSX2 and osteocalcin in PDL cells, suggesting that IL-22 enhances the mineralized matrix-forming activities of PDL cells. CONCLUSION: IL-22 has the potential to promote mineralizing activity in PDL cells and to develop appropriate regenerative therapy.

Sodium valproate, a histone deacetylase inhibitor, augments the expression of cell-surface NKG2D ligands, MICA/B, without increasing their soluble forms to enhance susceptibility of human osteosarcoma cells to NK cell-mediated cytotoxicity.

MHC class I-related chain molecules A and B (MICA and B) expressed on the cell-surface of tumor cells are ligands for an activating receptor, NKG2D, expressed on natural killer (NK) cells and stimulate the NK cell-mediated cytotoxicity. On the other hand, the soluble form of MICA and B produced by proteolytic cleavage of cell-surface MIC interferes with NK cell-mediated cytotoxicity. We investigated effect of sodium valproate (VPA), a histone deacetylase inhibitor, on the production of cell-surface and soluble MICA and B and NK cell-mediated cytotoxicity in four human osteosarcoma cells. VPA at 0.5 and 1.0 mM induced acetylation of histones bound to MICA and B gene promoters, increased cell-surface but not soluble MICA and B, and augmented the susceptibility of osteosarcoma cells to NK cell-mediated cytotoxicity. The present results indicate that VPA sensitizes human osteosarcoma cells to cytotoxicity of NK cells.

Gene cloning and characterization of EfmA, a multidrug efflux pump, from Enterococcus faecium.

A DNA fragment responsible for resistance to antimicrobial agents was cloned from chromosomal DNA of Enterococcus faecium FN-1, a clinically isolated strain. Escherichia coli KAM32, a drug-hypersusceptible mutant, was used as a host for gene cloning. Cells of E. coli KAM32 harboring a recombinant plasmid (pTFM8) carrying the DNA fragment became resistant to fluoroquinolones, macrolides, ethidium bromide, 4',6-diamidino-2-phenylindole (DAPI) and tetraphenylphosphonium chloride (TPPCl). Three complete open reading frames (ORFs) were found in the DNA insert of pTFM8, and the deduced amino acid sequences of one of the ORFs showed high similarity to Mdt(A) from Lactococcus lactis. Mdt(A) is a multidrug efflux pump belonging to a major facilitator superfamily. We designated the ORF efmA. E. coli KAM32 cells harboring the efmA showed energy-dependent efflux of DAPI and TPP(+). We also observed norfloxacin/H(+) antiport due to EfmA. The mRNA expression of efmA was observed in E. faecium FN-1 grown without any exogenously added antimicrobial agents. Thus, we conclude that efmA is constitutively expressed under laboratory growth conditions and would contribute to intrinsic resistance against multiple antimicrobial agents in E. faecium FN-1.