Validation of a high-throughput method for simultaneous determination of areca nut and tobacco biomarkers in hair using microwave-assisted extraction and isotope dilution liquid chromatography tandem mass spectrometry.

For people with habits of chewing betel nuts and smoking, the probability of suffering from oral cancer is ten to a hundred times higher than others. Due to the serious health consequences of areca nut and tobacco, a reliable cessation program is needed. Hair is the best option to document long-term exposure. Unfortunately, the research on betel nut in hair did not attract much attention. In this study, a high-throughput method based on microwave-assisted extraction (MAE) and isotope dilution liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed to measure the four biomarkers of betel nuts and cigarettes, including areca alkaloids (arecoline), tobacco alkaloids (nicotine), and their metabolites (arecaidine and cotinine). The hair sample was washed, cut, weighed, and incubated for 3 min MAE with methanol/trifluoroacetic acid, then evaporated and reconstituted for LC-MS/MS analysis. The total experiment time was 50 min. The lower limits of quantification (LOQ) were 5-10 pg/mg. The intra-day and inter-day precision were 2.2-7.6%. Intra-day and inter-day accuracy were - 6.1-8.2%. The method showed good linearity (r2 > 0.995) over LOQ - 1000 pg/mg concentration ranges. It was successfully applied to analyze 11 subjects of regular areca nut chewers, also smokers. Eight samples were black hair; three samples were naturally black hair with partially gray hair. Measured concentrations in black hair were in the range 56.9 pg/mg to 3.2 ng/mg for arecoline, 12.8 pg/mg to 222.2 pg/mg for arecaidine, 3.8 ng/mg to 33.4 ng/mg for nicotine and 1.1 ng/mg to 6.1 ng/mg for cotinine. The results showed lower levels in gray hair. This method was utilized successfully to analyze pg/mg levels of arecoline, arecaidine, nicotine, and cotinine, and good recoveries were obtained. The mean concentration of arecaidine and cotinine in hair was 15% and 20% of arecoline and nicotine, respectively. A good positive correlation was found between the concentrations of these compounds and self-report. This method improved extraction speed, concentration, and analysis of samples and is useful for monitoring betel nut and smoking cessation programs.

Impact of Zr-Doped Bi2O3 Radiopacifier by Spray Pyrolysis on Mineral Trioxide Aggregate.

Mineral trioxide aggregates (MTA) have been developed as a dental root repair material for a range of endodontics procedures. They contain a small amount of bismuth oxide (Bi2O3) as a radiopacifier to differentiate adjacent bone tissue on radiographs for endodontic surgery. However, the addition of Bi2O3 to MTA will increase porosity and lead to the deterioration of MTA's mechanical properties. Besides, Bi2O3 can also increase the setting time of MTA. To improve upon the undesirable effects caused by Bi2O3 additives, we used zirconium ions (Zr) to substitute the bismuth ions (Bi) in the Bi2O3 compound. Here we demonstrate a new composition of Zr-doped Bi2O3 using spray pyrolysis, a technique for producing fine solid particles. The results showed that Zr ions were doped into the Bi2O3 compound, resulting in the phase of Bi7.38Zr0.62O12.31. The results of materials analysis showed Bi2O3 with 15 mol % of Zr doping increased its radiopacity (5.16 ± 0.2 mm Al) and mechanical strength, compared to Bi2O3 and other ratios of Zr-doped Bi2O3. To our knowledge, this is the first study of fabrication and analysis of Zr-doped Bi2O3 radiopacifiers through the spray pyrolysis procedure. The study reveals that spray pyrolysis can be a new technique for preparing Zr-doped Bi2O3 radiopacifiers for future dental applications.

Construction of polycistronic baculovirus surface display vectors to express the PCV2 Cap(d41) protein and analysis of its immunogenicity in mice and swine.

To increase expression levels of the PCV2 Cap(d41) protein, novel baculovirus surface display vectors with multiple expression cassettes were constructed to create recombinant baculoviruses BacSC-Cap(d41), BacDD-2Cap(d41), BacDD-3Cap(d41), and BacDD-4Cap(d41). Our results reveal that the recombinant baculovirus BacDD-4Cap(d41) was able to express the highest levels of Cap(d41) protein. Optimum conditions for expressing the PCV2 Cap(d41) protein were determined, and our results show that 107 of Sf-9 infected with the recombinant baculovirus BacDD-4Cap(d41) at an MOI of 5 for 3 days showed the highest level of protein expression. Mice immunized with the 4Cap(d41) vaccine which was prepared from the recombinant baculovirus-infected cells (107) elicited higher ELISA titers compared to the Cap (d41) vaccine. The 4Cap(d41) vaccine could elicit anti-PCV2 neutralizing antibodies and IFN-γ in mice, as confirmed by virus neutralization test and IFN-γ ELISA. Moreover, the swine lymphocyte proliferative responses indicated that the 4Cap(d41) vaccine was able to induce a clear cellular immune response. Flow cytometry analysis showed that the percentage of CD4+ T cells and CD4+/CD8+ ratio was increased significantly in SPF pigs immunized with the 4Cap(d41) vaccine. Importantly, the 4Cap(d41) vaccine induced an IFN-γ response, further confirming that its effect is through cellular immunity in SPF pigs. An in vivo challenge study revealed that the 4Cap(d41) and the commercial vaccine groups significantly reduce the viral load of vaccinated pigs as compared with the CE negative control group. Taken together, we have successfully developed a 4Cap(d41) vaccine that may be a potential subunit vaccine for preventing the disease associated with PCV2 infections.

The association between Parkinson's disease and temporomandibular disorder.

The prevalence of temporomandibular disorder (TMD) among elderly people with Parkinson's disease (PD) is relatively high, but a population-based study of the relationship between PD and TMD is still lacking. This study, therefore, sought to investigate the association between TMD and PD by using data for one million randomly sampled beneficiaries of Taiwan's National Health Insurance program, including 6,185 PD patients who were matched through propensity score matching with 18,555 non-PD patients. Both the PD and non-PD cohorts were followed until death, any diagnosis of TMD, or December 31, 2013, whichever occurred first. Each diagnosis of TMD was made by a qualified physician according to the International Classification of Diseases, 9th Revision, Clinical Modification (ICD-9-CM), using the diagnosis codes 524.60, 524.62, 524.63, and 524.69 while excluding tooth abscess, wisdom tooth eruption, herpes zoster and postherpetic neuralgia, mastoiditis, otitis externa, otitis media, parotitis, sialadenitis, and trigeminal neuralgia. We used Cox proportional hazard regression models to calculate the relative risk of TMD and found a 2.11-fold (95% CI: 1.35-3.30) increased risk of TMD overall in the PD group compared with the non-PD group. Stratified by follow-up period, there was a 4.25-fold (95% CI: 1.51-11.93) increased risk in the PD group in the first year after the initial PD diagnosis and a 3.88-fold (95% CI: 1.33-11.28) increased risk in the second year. Over the long-term (>5 years), PD was significantly associated with an increased risk of TMD. These findings suggest that it is important to closely monitor the temporomandibular joint health of PD patients.

Vastus lateralis muscle flap used for reconstruction of the maxilla after radical resection of recurrent ameloblastoma.

Maxillary ameloblastoma is a benign odontogenic neoplasm. Excision should involve wide margins because of its high recurrence rate and, ideally, be followed by reconstruction. A 23 year-old female patient presented with recurrent ameloblastoma. Two years previously, she had undergone a curettage excision. The recurrent lesion was managed by a total maxillectomy including inferior orbital rim. Simultaneous reconstruction was performed with an iliac crest bone graft for the orbital floor, and a vastus lateralis muscle flap for obliteration of the maxillary sinus and repair the oral and nasal cavities. The transferred vastus lateralis muscle already had spontaneous mucosalization over its surface two weeks after surgery. Postoperative follow up after fourteen months revealed no recurrence and no diplopia. The patient was satisfied with her appearance. The use of a single vastus lateralis muscle flap to repair both oral and nasal cavities is advantageous. It helps avoid hyper-nasal speech and maintain the stability of removable partial dentures. The conventional iliac bone graft supported by a well vascularized muscle eventually survived and provided a good functional and cosmetic result.