Characterization of mesenchymal progenitor cell populations from non-epithelial oral mucosa.

OBJECTIVES: The characteristics of cell populations extracted from oral mucosal non-epithelial tissues and their ability to differentiate were evaluated in vitro as a potential source of cells for mandibular and corneal regeneration. MATERIALS AND METHODS: Oral mucosal non-epithelial cells (OMNECs) were extracted from tissue samples and were studied by flow cytometry and RT-PCR. Cells differentiating into osteoblasts, adipocytes, chondrocytes, neurocytes, or keratocytes were characterized by RT-PCR and cell staining. RESULTS: OMNECs expressed CD44, CD90, CD105, CD166, and STRO-1 antigens, which are markers for mesenchymal stem cells. In addition, Oct3/4, c-Myc, Nanog, KLF4, and Rex, which are expressed by embryonic or pluripotent stem cells, were detected by RT-PCR. Expression of CD49d, CD56, and PDGFRα, proteins closely associated with the neural crest, was observed in OMNECs, as was expression of Twist1, Sox9, Snail1 and Snail2, which are early neural crest and neural markers. Specific differentiation markers were expressed in OMNECs after differentiation into osteoblasts, adipocytes, chondrocytes, or keratocytes. CONCLUSIONS: Populations of OMNECs may contain both mesenchymal stem cells and neural crest origin cells and are a potential cell source for autologous regeneration of mandibular or corneal stroma.

IGF-1 Mediates EphrinB1 Activation in Regulating Tertiary Dentin Formation.

Eph receptors belong to a subfamily of receptor tyrosine kinases that are activated by membrane-spanning ligands called ephrins. Previously, we demonstrated that the ephrinB1-EphB2 interaction regulates odontogenic/osteogenic differentiation from dental pulp cells (DPCs) in vitro. The goal of this study was to identify the molecular mechanisms regulated by the EphB2/ephrinB1 system that govern tertiary dentin formation in vitro and in vivo. During tooth development, ephrinB1, and EphB2 were expressed in preodontoblast and odontoblasts at postnatal day 4. EphrinB1 was continuously expressed in odontoblasts and odontoblastic processes until the completion of tooth eruption. In addition, ephrinB1 was expressed in odontoblastic processes 2 wk following tooth injury without pulp exposure, whereas EphB2 was expressed in the center of pulp niches but not odontoblasts. In a model of tooth injury with pulp exposure, ephrinB1 was strongly expressed in odontoblasts 4 wk postinjury. In vitro studies with human and mouse DPCs treated with calcium hydroxide (CH) or mineral trioxide aggregate (MTA) showed an increased expression of insulin-like growth factor 1 (IGF-1). Experiments using several inhibitors of IGF-1 receptor signaling revealed that inhibiting the Ras/Raf-1/MAPK pathway inhibited EphB2 expression, and inhibiting the PI3K/Akt/mTOR pathway specifically inhibited ephrinB1 gene expression. Tooth injury in mice with odontoblast-specific IGF-1 receptor ablation exhibited a reduced tertiary dentin volume, mineral density, and ephrinB1 expression 4 wk following injury. We conclude that the IGF-1/ephrinB1 axis plays significant roles in the early stages of tooth injury. Further research is needed to fully understand the potential of targeting ephrinB1 as a regenerative pulp therapy.

The Piccolo Intronic Single Nucleotide Polymorphism rs13438494 Regulates Dopamine and Serotonin Uptake and Shows Associations with Dependence-Like Behavior in Genomic Association Study.

Piccolo (PCLO) inhibits methamphetamine-induced neuropharmacological effects via modulation of dopamine (DA) uptake and regulation of the transport of synaptic vesicles in neuronal cells. Clinical studies have recently suggested that the single nucleotide polymorphism (SNP) rs13438494 in the intron 24 of the PCLO gene is associated with psychiatric disorder, in the meta-analysis of GWAS. Therefore, in this study, we attempted to evaluate the possible role of the PCLO SNP in the mechanisms of uptake of monoamines. To characterize rs13438494 in the PCLO gene, we constructed plasmids carrying either the C or A allele of the SNP and transiently transfected them into SH-SY5Y cells to analyze genetic effects on the splicing of PCLO mRNA. The C and A allele constructs produced different composition of the transcripts, indicating that the intronic SNP does affect the splicing pattern. We also transfected DA and serotonin (5-hydroxytryptamine; 5- HT) transporters into cells and analyzed their uptakes to elucidate the association to psychiatric disorders. In the cells transfected with the C allele, both the DA and 5-HT uptake were enhanced compared to the A allele. We also conducted a clinical study, in order to clarify the genetic associations. PCLO rs13438494 exhibits a relationship with the symptoms of drug dependence or related parameters, such as the age of first exposure to methamphetamine, eating disorders, tobacco dependence and fentanyl requirement. Our findings suggest that rs13438494 is associated with drug abuse and contributes to the pathogenesis of psychiatric disorders via modulation of neurotransmitter turnover.

Organization of the functional domains in membrane cytoskeletal protein talin.

Talin, a putative homodimer of 230-kDa polypeptides, was cleaved into the N-terminal 47-kDa and C-terminal 190-kDa fragments with calpain II. The 190-kDa fragment, but not the 47-kDa fragment, was found to bind to actin. The 190-kDa fragment possessed similar levels of activities to stimulate both polymerization of G-actin and alpha-actinin-dependent gelation of F-actin as did intact talin. Limited digestions of the 190-kDa fragment with chymotrypsin and papain resulted in partial and complete reductions, respectively, of both activities, although these digests contained 95- and 46-kDa major polypeptides, respectively, which were able to bind to actin. Whereas the 190-kDa fragment generated fully cross-linked oligomeric polypeptides on treatment with 1-ethyl-3[3-(dimethylamino)-propyl]carbodiimide, the 95-kDa chymotryptic polypeptide generated heterologous polypeptides cross-linked partially with smaller polypeptides. The papain digest did not contain any cross-linkable polypeptide. Intact talin and the 47-kDa calpain fragment, but not the 190-kDa calpain fragment, were found to bind to phospholipid vesicles containing phosphatidylserine. These results indicate that the N-terminal and C-terminal domains play distinct roles in interacting with the membrane and cytoskeletal elements, respectively, and that the dimeric structure is also required for the latter interactions.

Novel lipase-catalyzed ring-opening copolymerization of lactide and trimethylene carbonate forming poly(ester carbonate)s.

Poly(lactide-co-trimethylene carbonate)s were prepared by the lipase-catalyzed ring-opening copolymerization of lactide and trimethylene carbonate having carbonate content from 0 to 100%. Their thermal properties and enzymatic degradability were measured. The L,L-, D,D- and D,L-lactides were copolymerized with trimethylene carbonate by porcine pancreatic lipase to produce random copolymers having molecular weights of up to 21000. The glass transition temperature (Tg of the copolymer was dependent on the carbonate content and the Tg values linearly decreased from 35 degrees C (polylactide) to -8 degrees C [poly(trimethylene carbonate)]. Among the lipases tested, the porcine pancreatic lipase and proteinase K showed biodegradability towards poly(ester-carbonate)s.

Results of a dental fear survey in Japan: implications for dental public health in Asia.

The purpose of this investigation was to initiate the study of dental fear in Japan. 415 college students, aged 18-22 yr were surveyed. A standardized questionnaire which has been used in the United States was translated into Japanese and was administered to the students. More than 80% of those surveyed reported some dental fear. Six to 14% of the students reported extreme fear of the dentist. The majority of the subjects admitted that they delayed making dental appointments due to fear. Muscle tension was the most common physiological symptom reported. The dental drill and needle were the most fear-provoking stimuli.

Histopathologic and radiographic findings of the simple bone cyst.

OBJECTIVE: The purpose of this study is to examine the correlation between histopathologic and radiographic findings and to discuss the cause of the simple bone cyst. STUDY DESIGN: Histopathologically, we classified 53 simple bone cysts into two types. Type A has a connective tissue membrane and type B has a partially thickened wall with dysplastic bone formation. Radiographically, we evaluated the following: margin, radiolucency, or radiopacity, relationship with tooth apices, bucco-lingual bone expansion, and displacement of the mandibular canal. RESULTS: Bone expansion and radiopacity were closely related to histopathologic findings although there was no correlation between the histopathologic findings and radiographic margin, relationship with tooth apices, and displacement of mandibular canal. Local recurrence was more likely to be observed in patients diagnosed as having type B than type A lesions. CONCLUSIONS: Type A and type B bone cysts may have different causes. Cysts determined radiographically to be radiopaque, those diagnosed as type B histopathologically, and cysts that have been treated surgically should all be followed by radiographic examinations.

Dental fear in Japan: Okayama Prefecture school study of adolescents and adults.

A total of 3,041 students and staff in middle school in Okayama Prefecture, Japan, were surveyed regarding dental fear. Over 88% reported fear, with 42.1% classified as having high fear. Almost 70% reported acquiring dental fear prior to junior high school. A majority reported being hurt at the last appointment. Delay of dental work was also reported for over 50% of the sample. Coping, pattern of physiological upset, nondental fears, and sex and age differences were also reported. Results suggest intervention is needed to address the major dental public health problems associated with dental fear.

Lipase-catalyzed transformation of poly(epsilon-caprolactone) into cyclic dicaprolactone.

The enzymatic degradation and polymerization using an enzyme were carried out with respect to the establishment of a sustainable chemical recycling system for poly(epsilon-caprolactone) (PCL) which is a typical biodegradable synthetic plastic. The enzymatic transformation of PCL having an Mn of 110,000 using Candida antarctica lipase (lipase CA) in water-containing toluene at 40 degrees C afforded the corresponding cyclic dicaprolactone (DCL, 1,8-dioxacyclotetradecane-2,9-dione) in a yield of up to 97%. Thus the obtained DCL readily polymerized again using both the fresh and recovered lipase CAs.

Assessment of the caries activity test (Cariostat) based on the infection levels of mutans streptococci and lactobacilli in 2- to 13-year-old children's dental plaque.

It is generally agreed that mutans streptococci and lactobacilli are associated etiologically with dental caries. The caries activity test, Cariostat, was designed to measure the pH decrease caused by microorganisms in the plaque sample obtained from the buccal surfaces. Researchers found the test to be a reliable, diagnostic, and predictive device. Incubation was done on MS and MSB plates in an atmosphere of 95 percent N and 5 percent CO at 37 degrees C and for 48 hours. The relationship of the Cariostat scores and the pH values are shown in a table. The test scores are shown for two age-groups: Ages two-to-six years with primary dentitions, and ages five-to-thirteen years with mixed dentitions. The advantages of the Cariostat test are: the sampling method is simple and the time of analysis is short; the test can be used for the very young and for patients difficult to manage; and it requires no specialized knowledge or equipment.

Large scale production and characterization of lyophilized pyridoxalated hemoglobin-polyoxyethylene (PHP).

Lyophilized PHP as an oxygen carrier is prepared from outdated red cell and dicarboxymethylated polyoxyethylene. In order to apply PHP for a clinical use, a large scale production of high quality PHP has been studied. We have set up a 20 L scale production flow of PHP88. The product was tested to confirm the quality and lot-to-lot consistency. The blood group specific materials were weakly positive in stroma-free hemoglobin (SFH), however, were found negative in the PHP of this scale. The amount of phosphatidylethanolamine (PE) in purified SFH and PHP88 reconstituted solution was 0.19 +/- 0.04 and 0.03 +/- 0.01 ppm, respectively. Contamination of viruses such as HBV and Non A non B hepatitis virus could not be observed in the final product. Elimination and inactivation of HIV was validated through a spike test. The characterizations of the final products in 20 L scale were done through MW, P50, Hill coefficient, viscosity, and molecular weight distribution by SDS-PAGE and batch to batch consistency was also confirmed. The results show that production process is appropriate to eliminate the blood group materials, PE and virus, and produce PHP of high quality. Lyophilized PHP88 can be produced by addition of maltose and can be stored over 1 year.

Involvement of catalytic amino acid residues in enzyme-catalyzed polymerization for the synthesis of polyesters.

Recently, a variety of aliphatic polyesters have been synthesized using hydrolases such as lipases and PHB depolymerases, and the reaction mechanism for these enzyme-catalyzed polymerization has been discussed. In this paper, we have studied the involvement of the catalytic amino acid residues of the hydrolase in enzyme-catalyzed polymerization with an extracellular PHB depolymerase from Alcaligenes faecalis T1. A wild-type PHB depolymerase and three kinds of site-specific mutants (catalytic amino acids were substituted) were prepared and their polymerization activities for the ring-opening polymerization of (R)-beta-butyrolactone (BL) were compared. BL was polymerized at 80 degrees C in bulk by the wild-type enzyme to yield polymers consisting of cyclic and linear structures in a high monomer conversion. In contrast, none of the mutant enzymes showed obvious polymerization activity. These results have clearly demonstrated that the catalytic triad is indeed responsible for the enzyme-catalyzed polymerization of BL.

[A behavioral dental science research concerning prevention of dental fear. A report on the surveys of dental fear in four countries Japan, Indonesia, Brazil and Argentina].

We conducted a behavioral scientific research from the point of view of dental fear prevention. This time, the writers carried out the surveys with the goal of understanding and making clear the realities concerning dental anxiety in the four countries: Japan, Indonesia, Brazil and Argentina. The subjects were 728 dental students from the four countries, and the surveys consisted of 25 multiple choice questions which were divided into three major areas. The first area consisted of preliminary questions such as chief complaints and treatment experiences, the second area dealt with the reactions and physical responses on receiving dental treatment and the third area was on the different levels of anxiety shown during treatment expressed through feelings and reactions. We concluded as follows: 1. In Japan, 77% of the subjects reported having at least a little fear and this percentage was higher than those of other countries (45-65%). But in all countries, 4-7% of subjects reported having extreme dental fear. 2. In Japan and Brazil, women reported the higher fear than men. In Indonesia and Argentina, these results were the opposite. 3. According to canonical correlation analysis, a significant correlation between the level of physical reactions and the degree of dental anxiety was found in all countries. 4. In the high fear group, dental anxiety increased by psychological stress before treatment and the expressions of physical reactions were stronger than in the low fear group.