The Characteristics of Human iPS Cells and siRNA Transfection Under Hypoxia.

The characteristics of pluripotent cells have great potential for basic and clinical research and application. We describe the effect of normoxia or hypoxia regarding the proliferation and pluripotency of human iPS cells using colony number counting and real-time polymerase chain reaction (PCR). In addition, the function of hypoxia-inducible factors (HIFs) in human iPS cells under hypoxic conditions is evaluated in relation to the expression of pluripotency markers by siRNA and real-time PCR. Furthermore, we introduce the change of HIF-2α expression when signal transducer and activator of transcription 3 (STAT3) is suppressed by its inhibitor, Stattic or S31 201, using RT-PCR.

Perceived general health in relation to oral health status in a rural Kenyan elderly population.

BACKGROUND: This study aimed to determine the present oral health status of the rural Kenyan elderly population and to investigate whether oral health status is associated with the perceived general health. METHODS: A total of 131 individuals aged 65 years and over in Mbita Constituency, Homa Bay County in Kenya were randomly selected and visited at home. The home visit study, which was conducted from 2014 to 2016, included oral examination by a dentist and administration of a self-reporting questionnaire. The number of teeth present and functional tooth unit (FTU) points were calculated using the dental chart. Perceived general health, subjective masticatory ability, and self-reported periodontal symptoms were collected using a questionnaire sheet. Fisher's exact test and nonparametric test were used to determine the difference in percentage and means. The odds ratio of good general health based on the participants' masticatory satisfaction was calculated by logistic analysis. RESULTS: Satisfactory mastication was dependent on the number of teeth present, FTU points, and self-reported "gum bleeding" and "tooth mobility". Furthermore, satisfactory mastication was associated with perceived general health status independent of sex and age. The adjusted odds ratio of good perceived general health was 2.29 (95% confidence interval 1.05-4.99) for participants who had the subjective masticatory ability. CONCLUSION: Among the Kenyan elderly population, satisfactory mastication was related to the number of teeth present, FTU points, and self-reported periodontal symptoms. Furthermore, satisfactory mastication was associated with perceived general health status independently.

Laparoscopic Hysterectomy for Benign Pathology Does Not Yield More Perioperative Complications than Abdominal or Vaginal Hysterectomies: Our Experience in Introducing Laparoscopic Hysterectomy.

OBJECTIVES: Total laparoscopic hysterectomy (TLH) is increasing as a substitute for total abdominal hysterectomy (TAH) and total vaginal hysterectomy (TVH) with the growing prevalence of laparoscopic surgery. The aim of this study is to assess perioperative complications of the chosen hysterectomy techniques performed for benign indications when started performing TLH. This was retrospective cohort study. This study was conducted at Nagahama City Hospital. MATERIALS AND METHODS: There were 176 patients who underwent hysterectomy for benign indications from 2013 to 2016. Perioperative and postoperative outcomes were compared for the three different hysterectomy approaches laparoscopic; abdominal; and vaginal. Data were analyzed using the t-test or Chi-square and Fisher's exact test. RESULTS: TAH, TLH, and TVH were performed on 118 patients (67.0%), 32 (18.2%), and 26 (14.8%), respectively. Operation time was significantly longer for the TLH group than for the TAH and TVH groups. Blood loss was lower for the TVH and TLH groups than for the TAH group. Three days after surgery, C-reactive protein was lower in the TVH group than in the TAH group. The average uterus size in the TAH group was larger than in the TVH and TLH groups. Patients undergoing TLH experienced fewer perioperative complications than patients in the TAH and TVH groups; however, this difference was not statistically significant. CONCLUSION: TLH for benign pathology does not yield more perioperative complications than TAH or TVH. However, vaginal hysterectomy is the least invasive approach. The final choice for the route of hysterectomy depends on many factors, including body mass index, uterus size, and experience of the gynecologist.

Small-cell lung carcinoma with ovarian metastasis 4 years after the first-line treatment.

Small-cell lung carcinoma rarely metastasizes to the ovary. Only few cases of this condition have been reported to date. A 42-year-old female nonsmoker was an outpatient after receiving treatment for small-cell lung carcinoma. Approximately 45 months after the first-line treatment, the pro-gastrin-releasing peptide level exhibited a gradual increase. Positron emission tomography-computed tomography revealed abnormal accumulation in the left ovary. Accordingly, we performed laparoscopic salpingo-oophorectomy. Both pathological and immunohistochemical examinations (thyroid transcription factor-1, synaptophysin, and chromogranin A staining) led to the diagnosis of ovarian metastasis of small-cell lung carcinoma. The pro-gastrin-releasing peptide level declined postoperatively, and no recurrence has been reported thus far. Here we reported an extremely rare case of small-cell lung carcinoma metastatic to the ovary after several years of receiving the initial treatment for small-cell lung carcinoma, which, however, exhibited an excellent course postoperatively.

Anaplastic lymphoma kinase-negative uterine inflammatory myofibroblastic tumor containing the ETV6-NTRK3 fusion gene: a case report.

Inflammatory myofibroblastic tumors (IMTs) are neoplasms with low malignant potential, and the most common tumor in the lung and orbit. Their occurrence in the uterus is rare. Approximately 50% of IMT patients have anaplastic lymphoma kinase gene ( ALK) rearrangements. Recent studies described novel fusions involving ROS1, platelet-derived growth factor receptor beta ( PDGFR-ß), and ETS translocation variant ( ETV6) genes in a subset of ALK-negative patients. We report a 44-year-old woman with anemia and uterine IMT. Ultrasonography and magnetic resonance imaging revealed a myxoid degenerative myoma-like mass, 7.4 cm in maximum diameter, on the left uterine side wall. Hysterectomy was performed as a definitive treatment. Microscopic examination revealed spindle cell proliferation with numerous lymphocytes and plasma cells. Immunohistochemically, the spindle cells were negative for ALK-1, desmin, and smooth muscle actin. The pathological diagnosis was IMT arising from the uterus. Fluorescence in situ hybridization demonstrated an ETV6-neurotrophic tyrosine kinase, receptor, type 3 gene ( NTRK3) translocation but no ALK, ROS1, or PDGFR-ß translocations. Lung and abdomen computed tomography at 31 months postoperatively revealed no disease recurrence. This association of an ETV6-NTRK3 fusion oncogene with an ALK-negative uterine IMT increases our understanding of this neoplasm, which may help the development of specific therapies.

Application of astatine-210: Evaluation of astatine distribution and effect of pre-injected iodide in whole body of normal rats.

We proposed use of astatine-210 in preclinical study. Astatine-210 has higher yield of production and is easier to quantify than astatine-211. We produced astatine-210 with Bi target and 40 MeV alpha beam accelerated by cyclotron, free astatine-210 was separated and injected to normal rats. Three male rats (blocking group) were injected non-radioactive iodide before injection of astatine-210. Compared with the control group, the astatine-210 accumulations in the blocking group decreased to 24% in the thyroid.

Cell migration capability of vascular endothelial growth factor into the root apex of a root canal model in vivo.

Once a tooth develops deep caries and the dental pulp tissue is irreversibly infected, the infected dental pulp tissue should be removed, and filling material should be placed in the root canal. Endodontically treated teeth are prone to root fracture or periapical periodontitis; however, dental pulp tissue has the potential to prevent root fracture or periapical periodontitis. Therefore, dental pulp regeneration after pulpectomy may help prolong tooth life. In this study, a new method of dental pulp regeneration was developed. Vascular endothelial growth factor-adsorbed collagen gel was injected into the root canal of a prepared root canal model, placed into the dorsum of a rat, and cultured for 3 weeks. After retrieving the implant, histological analysis was performed. It was found that rat somatic cells were recruited into the root apex of the transplanted root canal model. These findings suggest a new potential technique for engineering dental pulp tissue.

Effects of hypoxia inducible factors on pluripotency in human iPS cells.

A hypoxic condition is known to contribute to pluripotency. In the present article, the effects of transcription factors were first assessed regarding the proliferation and differentiation of human induced pluripotent stem (iPS) cells under hypoxic conditions using cell morphology and real-time polymerase chain reaction (RT-PCR). Morphology evaluations and RT-PCR revealed that the colony formation was promoted and the expression of pluripotent markers was increased under hypoxic conditions. In addition, the function of hypoxia inducible factors (HIFs) in human iPS cells under hypoxic conditions was evaluated in relation to the morphology and the expression of pluripotency markers by siRNA and RT-PCR. The HIF-2α silencing group showed a reduction in the colony size of human iPS cells and a statistically significant reduction in the expression of undifferentiation markers compared to the control group. Furthermore, the expression of HIF-2α was decreased when signal transducer and activator of transcription 3 (STAT3) was suppressed by its inhibitor, Stattic or S31 201. The inhibition using Stattic did not produce colony formation. The expression of pluripotent markers was also decreased using Stattic or S31 201. This study indicates that the HIF-2α expression in human iPS cells was activated under hypoxic conditions, similarly to that in murine iPS cells, and that HIF-2α among HIFs is the most effective compound for maintaining the pluripotency of human iPS cells. Furthermore, the STAT3 signal pathway regulates the expression of HIF-2α.

The clinical findings and managements in 44 cases of cracked vital molars.

BACKGROUND/PURPOSE: The aim of this study was to evaluate the associations between the clinical findings and managements in cracked vital molars that were caused by various factors including restoration and occlusion. MATERIALS AND METHODS: The subjects' gender, age, chief complaint, type of tooth, percussion test results, pulp vitality, restoration material and cavity classification, clinical depth of the crack, evaluation of occlusion, depth of periodontal probing, and final management were recorded. RESULTS: A total of 44 vital cracked teeth (molars) were diagnosed in 40 patients. Regarding the type of tooth, a greater number of mandibular molars were affected than maxillary molars. Nonworking-side interference (NWI) was recognized in 38 cases (86.4%). Eight teeth (18.2%) had not been restored. Thirty-six teeth (81.8%) had been restored; 26 teeth (72.2%) with a metal inlay, 6 (16.7%) with an amalgam, and 4 (11.1%) with a composite resin. Regarding the final treatment in the endodontically-treated group, all 17 teeth were covered with a metal full crown. Regarding the final treatment in the pulp-reserved group, 19 teeth (70.4%) were covered with a metal full crown, and the other managements were as follows: occlusal adjustment (n = 4, 14.8%), composite resin (n = 2, 7.4%), and only follow-up without treatment (n = 2, 7.4%). All of the cases showed a good clinical prognosis. CONCLUSION: The NWI group restored with 58% of metal inlay accounted for more than 86% of the cracked teeth. Thus, in order to achieve a good outcome, cracked teeth, particularly those originating due to occlusal interference should be protected with coverage-type restorations.

Boron Accelerates Cultured Osteoblastic Cell Activity through Calcium Flux.

A low concentration of boron (B) accelerates the proliferation and differentiation of mammalian osteoblasts. The aim of this study was to investigate the effects of 0.1 mM of B on the membrane function of osteoblastic cells in vitro. Genes involved in cell activity were investigated using gene expression microarray analyses. The Ca2+ influx and efflux were evaluated to demonstrate the activation of L-type Ca2+ channel for the Ca2+ influx, and that of Na+/K+-ATPase for the Ca2+ efflux. A real-time PCR analysis revealed that the messenger RNA (mRNA) expression of four mineralization-related genes was clearly increased after 3 days of culture with a B-supplemented culture medium. Using microarray analyses, five genes involved in cell proliferation and differentiation were upregulated compared to the control group. Regarding the Ca2+ influx, in the nifedipine-pretreated group, the relative fluorescence intensity for 1 min after adding B solution did not increase compared with that for 1 min before addition. In the control group, the relative fluorescence intensity was significantly increased compared with the experimental group (P < 0.05). Regarding the Ca2+ efflux, in the experimental group cultured in 0.1 mM of B-supplemented medium, the relative fluorescence intensity for 10 min after ouabain treatment revealed a significantly lower slope value compared with the control group (P < 0.01). This is the first study to demonstrate the acceleration of Ca2+ flux by B supplementation in osteoblastic cells. Cell membrane stability is related to the mechanism by which a very low concentration of B promotes the proliferation and differentiation of mammalian osteoblastic cells in vitro.

Scanning electron microscopy reveals severe external root resorption in the large periapical lesion.

The present study was designed to investigate the relationships between clinicopathological findings and the resorptive conditions of root apices of teeth with periodontitis. The samples included 21 root apices with large periapical radiolucent lesions. The preoperative computed tomography (CT) and intraoperative findings were correlated with the presence, extension, and the progression pattern of periapical resorption using a scanning electron microscope. The subjects' age, gender, chief complaint, type of tooth, percussion test results, size of periapical lesion using CT, and intraoperative findings were recorded. All apicoectomies were performed under an operative microscope for endodontic microsurgery. A significant large size was observed in cystic lesions compared with granulomatous lesions. The cementum surface at the periphery of the lesion was covered with globular structures (2-3 µm in diameter). Cementum resorption started as small defect formations at the surface. As the defect formation progressed, a lamellar structure appeared at the resorption area, and the size of globular structures became smaller than that of globules at the surface. Further resorption produced typical lacuna formation, which was particularly observed in fracture cases. The most morphologically severe destructive pattern of dentin resorption was observed in large cystic lesions. This study is the first report to elucidate the relationships between three clinical types of undesirable periapical lesions: (1) undertreatment, (2) periapical fracture, (3) macro-level resorption, and the microstructure of external root resorption including from small defects at the cementum surface to a significant destructive pattern inside the dentin. Microsc. Res. Tech. 79:495-500, 2016. © 2016 Wiley Periodicals, Inc.

The Characteristics of Murine iPS Cells and siRNA Transfection Under Hypoxia.

iPS cells are attractive for the regenerative medicine. The creation of pluripotent cells from somatic cells has great potential for basic and clinical research and application. Retroviral transduction of four or three transfection factors has been shown to initiate a reprogramming process. Here, we describe the effect of transcription factors regarding the growth and differentiation of mouse iPS cells in normoxia or hypoxia. Furthermore, we introduce the function of hypoxia-inducible factors (HIFs) in mouse iPS cells in hypoxia using RT-PCR and western blotting together with HIFs knockdown techniques.

Feasibility studies towards future self-sufficient supply of the (99)Mo-(99m)Tc isotopes with Japanese accelerators.

In order to establish a self-sufficient supply of (99m)Tc, we studied feasibilities to produce its parent nucleus, (99)Mo, using Japanese accelerators. The daughter nucleus, (99m)Tc, is indispensable for medical diagnosis. (99)Mo has so far been imported from abroad, which is separated from fission products generated in nuclear reactors using enriched (235)U fuel. We investigated (99m)Tc production possibilities based on the following three scenarios: (1) (99)Mo production by the (n, 2n) reaction by spallation neutrons at the J-PARC injector, LINAC; (2) (99)Mo production by the (p, pn) reaction at Ep = 50-80 MeV proton at the RCNP cyclotron; (3) (99m)Tc direct production with a 20 MeV proton beam from the PET cyclotron. Among these three scenarios, scenario (1) is for a scheme on a global scale, scenario (2) works in a local area, and both cases take a long time for negotiations. Scenario (3) is attractive because we can use nearly 50 PET cyclotrons in Japan for (99m)Tc production. We here consider both the advantages and disadvantages among the three scenarios by taking account of the Japanese accelerator situation.

Comparison of conventional and new-generation nickel-titanium files in regard to their physical properties.

INTRODUCTION: This study investigated the surface, fractured structure, and physicochemical properties related to cyclic fatigue in various nickel-titanium (NiTi) files. METHODS: Among a total of 10 groups of NiTi files, conventional NiTi files (ProFile [Dentsply Maillefer, Ballaigues, Switzerland] and K3 [SybronEndo, Orange, CA]) and new-generation NiTi files (ProFile Vortex [PV; Tulsa Dental Specialties, Tulsa, OK], Vortex Blue [VB; Tulsa Dental Specialties], and K3 XF [XF; SybronEndo, Orange, CA]) with the same tip diameter (ISO size 25) and 2 types of taper (0.04 and 0.06) were used in this study. Scanning electron microscopy of the file surface structure, differential scanning calorimetry, and cyclic fatigue resistance tests were conducted. RESULTS: Many mechanical grooves were recognized on the file surface. The surface in the ProFile group was extremely smooth compared with that observed for the other files. Many shallow hollows besides mechanical grooves were noted on the surface in the XF group. A smooth curve was observed in the ProFile, K3, and PV groups. Defined peaks in differential scanning calorimetry were observed in the VB and XF groups. The 0.04 taper files exhibited a statistically higher number of cycles to fracture than the 0.06 taper files in all groups (P < .05). Cracks along the mechanical grooves were observed in the NiTi files, with the exception of the XF group. The start of cracking was detected at U-shape sites in the ProFile group, the cutting edge in the PV and VB groups, and radial islands in the K3 and XF groups. CONCLUSIONS: The present findings suggest that new-generation NiTi files are not necessarily improved compared with conventional files.

Potency of fish collagen as a scaffold for regenerative medicine.

Cells, growth factors, and scaffold are the crucial factors for tissue engineering. Recently, scaffolds consisting of natural polymers, such as collagen and gelatin, bioabsorbable synthetic polymers, such as polylactic acid and polyglycolic acid, and inorganic materials, such as hydroxyapatite, as well as composite materials have been rapidly developed. In particular, collagen is the most promising material for tissue engineering due to its biocompatibility and biodegradability. Collagen contains specific cell adhesion domains, including the arginine-glycine-aspartic acid (RGD) motif. After the integrin receptor on the cell surface binds to the RGD motif on the collagen molecule, cell adhesion is actively induced. This interaction contributes to the promotion of cell growth and differentiation and the regulation of various cell functions. However, it is difficult to use a pure collagen scaffold as a tissue engineering material due to its low mechanical strength. In order to make up for this disadvantage, collagen scaffolds are often modified using a cross-linker, such as gamma irradiation and carbodiimide. Taking into account the possibility of zoonosis, a variety of recent reports have been documented using fish collagen scaffolds. We herein review the potency of fish collagen scaffolds as well as associated problems to be addressed for use in regenerative medicine.

Biological safety of fish (tilapia) collagen.

Marine collagen derived from fish scales, skin, and bone has been widely investigated for application as a scaffold and carrier due to its bioactive properties, including excellent biocompatibility, low antigenicity, and high biodegradability and cell growth potential. Fish type I collagen is an effective material as a biodegradable scaffold or spacer replicating the natural extracellular matrix, which serves to spatially organize cells, providing them with environmental signals and directing site-specific cellular regulation. This study was conducted to confirm the safety of fish (tilapia) atelocollagen for use in clinical application. We performed in vitro and in vivo biological studies of medical materials to investigate the safety of fish collagen. The extract of fish collagen gel was examined to clarify its sterility. All present sterility tests concerning bacteria and viruses (including endotoxin) yielded negative results, and all evaluations of cell toxicity, sensitization, chromosomal aberrations, intracutaneous reactions, acute systemic toxicity, pyrogenic reactions, and hemolysis were negative according to the criteria of the ISO and the Ministry of Health, Labour and Welfare of Japan. The present study demonstrated that atelocollagen prepared from tilapia is a promising biomaterial for use as a scaffold in regenerative medicine.

Fabrication and characteristics of chitosan sponge as a tissue engineering scaffold.

Cells, growth factors, and scaffolds are the three main factors required to create a tissue-engineered construct. After the appearance of bovine spongiform encephalopathy (BSE), considerable attention has therefore been focused on nonbovine materials. In this study, we examined the properties of a chitosan porous scaffold. A porous chitosan sponge was prepared by the controlled freezing and lyophilization of different concentrations of chitosan solutions. The materials were examined by scanning electron microscopy, and the porosity, tensile strength, and basic fibroblast growth factor (bFGF) release profiles from chitosan sponge were examined in vitro. The morphology of the chitosan scaffolds presented a typical microporous structure, with the pore size ranging from 50 to 200 µm. The porosity of chitosan scaffolds with different concentrations was approximately 75-85%. A decreasing tendency for porosity was observed as the concentration of the chitosan increased. The relationship between the tensile properties and chitosan concentration indicated that the ultimate tensile strength for the sponge increased with a higher concentration. The in vitro bFGF release study showed that the higher the concentration of chitosan solution became, the longer the releasing time of the bFGF from the chitosan sponge was.

Suppressive effects of D-glucosamine on the 5-HT sensitive nociceptive units in the rat tooth pulpal nerve.

It is well known that D-glucosamine hydrochloride (DGL) has a variety of biological activities and is regarded as a nutritional supplement effective in improving various disorders, including osteoarthritis and atherosclerosis. Although it has been reported that DGL has a significant pain relief effect in treating osteoarthritis, little is known about the characteristics of the effects of this compound on dental pain. The present study was undertaken to evaluate the applicability of DGL as a medicament to control pulpalgia. Using an in vitro rat mandible-inferior alveolar nerve preparation (jaw-nerve preparation), we evaluated the effects of DGL on 5-hydroxytryptamine (5-HT) sensitive nociceptive responses in the tooth pulpal nerve. 5-HT-induced nociceptive responses were fairly suppressed by direct application of DGL, suggesting that DGL have a pain relief effect on patients with dental pain.

D-glucosamine conjugation accelerates the labeling efficiency of quantum dots in osteoblastic cells.

Quantum dots (QDs) are useful imaging tools in the medical and biological fields due to their optical properties, such as a high fluorescence intensity, remarkable resistance to photobleaching, broad absorption spectra, and narrow emission spectra. This is the first study to investigate the uptake of carboxylated QDs conjugated with D-glucosamine (core size: approximately 3 nm, final modified size: 20-30 nm) into cultured osteoblastic cells. The QDs attached to the cell surface and were transported into the cytoplasm within approximately three hours of culture, whose process was clearly demonstrated using specific fluorescent staining of the cell membrane. Although the intranuclear distribution was not observed, a dramatic decrease in the transfer of quantum dots into the cytoplasm was recognized after approximately seven days of culture. Other interesting phenomena include the escape of the quantum dots from lysosomes in the cytoplasm, as confirmed by the merging of both QD fluorescence and specific fluorescent staining of lysosomes in the cytoplasm. These findings suggest that D-glucosamine conjugation enhances proton absorption in acid organelles and promotes the lysosomal escape of QDs.