Combined chlorhexidine and PVP-I decontamination of human donor eyes prior to corneal preservation

The aim of this study was to report the efficacy of adding chlorhexidine to the protocol for decontamination of human donor globes prior to excision of corneo-scleral rims for future keratoplasty procedures. In 2005, chlorhexidine was introduced by our eye bank as an additional step in the protocol for decontaminating human donor globes. After 5?years, we prospectively evaluated the number of contaminations. Out of 2,891 globes included in our study, 2,663 globes were processed, of which 36 (1.4%) were considered contaminated. Seventeen contaminations (0.6%) were detected by culturing limbal swabs, directly after decontamination, eight (0.3%) by visible discoloration of the culture medium carrying a corneo-scleral rim, and eleven (0.4%) after inoculation of the culture medium on blood agar plates. Importantly, after 4?weeks of incubation, none of the aerobic and anaerobic cultures taken from the secondary ??transport medium?? (dextran containing medium used to transport corneal tissue to the transplantation centre) showed microbiological growth. In conclusion, the combined use of 0.02% chlorhexidine and 0.5% povidone-iodine may allow decontamination of donor globes to a level at which the risk of tissue contamination at the time of transplantation is minimized, while corneal viability is preserved.     

Preliminary investigation of variations in tooth replacement in adult Necturus maculosus

Although there are a number of studies on tooth replacement patterns in lower vertebrates, most do not indicate whether this process is continuous throughout the year or is affected by either breeding or seasonal cycles. We have surveyed the replacement patterns found in living and specifically killed Necturus maculosus (Amphibia: Proteidae) to determine the nature of their variation throughout the year prior to investigating possible controlling mechanisms of the formation and eruption of amphibian teeth. Some animals (34), kept in a large outside tank, were killed at monthly intervals and their tooth-bearing bones radiographed using a modification of the technique previously described (Miller and Radnor, '70). Other animals (9), kept at 4°C, were anesthetized with tricaine methanesulphonate (M.S. 222), and wax impressions taken with beading (carding) wax of the functioning teeth at regular intervals. Animals examined in the late spring and summer (25) showed no signs of active tooth replacement. Small replacement teeth visible beneath each functioning tooth enlarged only slightly throughout the summer. In early and late fall some functioning teeth were lost and replacement teeth grew and erupted to replace them. Replacement patterns were very irregular and classical alternate form rarely seen. In a number of animals the replacement series was formed from every third tooth. Animals kept constantly at 4°C showed no replacement phenomena. Patterns varied between the different bones of the jaws and did not support the Zahnreihe concept of Edmund ('60).     

Pattern of tooth loss in an elderly population from Ibadan, Nigeria

Objectives: To determine tooth loss, edentulousness, causes of tooth loss and pattern of tooth loss in the elderly in South East Local Government area (SELGA) in Ibadan. Background data: SELGA is one of the largest in Oyo State, Nigeria and has a population of 225 800. Design: A cross‐sectional survey. Methodology: A total of 690 elderly individuals who were 65 years and above living in various wards in SELGA were interviewed by two interviewers and examined by two trained and calibrated examiners whilst two record clerks recorded all the findings of the examination. Result: Forty‐eight per cent of the subjects in the study had not lost any teeth. The mean tooth loss was 4.5 ± 7.6. Percentage edentulousness was 1.3% and this was higher in males than in females. This difference was not statistically significant (χ² = 0.07 p = 0.7). The total number of teeth lost was 3102, 14% of the total number of teeth examined. The study showed that caries was not a major cause of tooth loss. Only 22 (0.7%) teeth were lost as a result of caries, 19 (0.6%) were lost because of trauma and periodontal disease contributed to loss of 3061 (98.7%) teeth. Mandibular teeth exhibited a higher rate of retention than maxillary teeth. The percentage of elderly individuals with tooth loss increased with age. Conclusion: The study highlights the high life expectancy of a tooth among the elderly in SELGA. Despite the different cause of tooth loss in this area, in comparison with developed countries, the pattern of tooth retention appeared similar.     

Incremental enamel development in modern human deciduous anterior teeth

This study reconstructs incremental enamel development for a sample of modern human deciduous mandibular (n = 42) and maxillary (n = 42) anterior (incisors and canines) teeth. Results are compared between anterior teeth, and with previous research for deciduous molars (Mahoney: Am J Phys Anthropol 144 (2011) 204-214) to identify developmental differences along the tooth row. Two hypotheses are tested: Retzius line periodicity will remain constant in teeth from the same jaw and range from 6 to 12 days among individuals, as in human permanent teeth; daily enamel secretion rates (DSRs) will not vary between deciduous teeth, as in some human permanent tooth types. A further aim is to search for links between deciduous incremental enamel development and the previously reported eruptionsequence. Retzius line periodicity in anterior teeth ranged between 5 and 6 days, but did not differ between an incisor and molar of one individual. Intradian line periodicity was 12 h. Mean cuspal DSRs varied slightly between equivalent regions along the tooth row. Mandibular incisors initiated enamel formation first, had the fastest mean DSRs, the greatest prenatal formation time, and based upon prior studies are the first deciduous tooth to erupt. Relatively rapid development in mandibular incisors in advance of early eruption may explain some of the variation in DSRs along the tooth row that cannot be explained by birth. Links between DSRs, enamel initiation times, and the deciduous eruption sequence are proposed. Anterior crown formation times presented here can contribute toward human infant age-at-death estimates. Regression equations for reconstructing formation time in worn incisors are given.     

Static magnetic field increases survival rate of dental pulp stem cells during DMSO-free cryopreservation

Abstract

Successful and efficient cryopreservation of living cells and organs is a key clinical application of regenerative medicine. Recently, magnetic cryopreservation has been reported for intact tooth banking and cryopreservation of dental tissue. The aim of this study was to assess the cryoprotective effects of static magnetic fields (SMFs) on human dental pulp stem cells (DPSCs) during cryopreservation. Human DPSCs isolated from extracted teeth were frozen with a 0.4-T or 0.8-T SMF and then stored at ?196?°C for 24?h. During freezing, the cells were suspended in freezing media containing with 0, 3 or 10% DMSO. After thawing, the changes in survival rate of the DPSCs were determined by flow cytometry. To understand the possible cryoprotective mechanisms of the SMF, the membrane fluidity of SMF-exposed DPSCs was tested. The results showed that when the freezing medium was DMSO-free, the survival rates of the thawed DPSCs increased 2- or 2.5-fold when the cells were exposed to 0.4-T or 0.8-T SMFs, respectively (p?<?0.01). In addition, after exposure to the 0.4-T SMF, the fluorescence anisotropy of the DPSCs increased significantly (p?<?0.01) in the hydrophilic region. These results show that SMF exposure improved DMSO-free cryopreservation. This phenomenon may be due to the improvement of membrane stability for resisting damage caused by ice crystals during the freezing procedure.     

Viability of ectomycorrhizal fungi following cryopreservation

The use of ectomycorrhizal (ECM) fungi in biotechnological processes requires their maintenance over long periods under conditions that maintain their genetic, phenotypic, and physiological stability. Cryopreservation is considered as the most reliable method for long-term storage of most filamentous fungi. However, this technique is not widespread for ECM fungi since many do not survive or exhibit poor recovery after freezing. The aim of this study was to develop an efficient cryopreservation protocol for the long-term storage of ECM fungi. Two cryopreservation protocols were compared. The first protocol was the conventional straw protocol (SP). The mycelium of the ECM isolates was grown in Petri dishes on agar and subsequently collected by punching the mycelium into a sterile straw before cryopreservation. In the second protocol, the cryovial protocol (CP), the mycelium of the ECM isolates was grown directly in cryovials filled with agar and subsequently cryopreserved. The same cryoprotectant solution, freezing, and thawing process, and re-growth conditions were used in both protocols. The survival (positive when at least 60 % of the replicates showed re-growth) was evaluated before and immediately after freezing as well as after 1 week, 1 m, and 6 m of storage at −130 °C. Greater survival rate (80 % for the CP as compared to 25 % for the SP) and faster re-growth (within 10 d for the CP compared to the 4 weeks for the SP) were observed for most isolates with the CP suggesting that the preparation of the cultures prior to freezing had a significant impact on the isolates survival. The suitability of the CP for cryopreservation of ECM fungi was further confirmed on a set of 98 ECM isolates and displayed a survival rate of 88 % of the isolates. Only some isolates belonging to Suillus luteus, Hebeloma crustuliniforme, Paxillus involutus and Thelephora terrestris failed to survive. This suggested that the CP is an adequate method for the ultra-low cryopreservation of a large set of ECM fungi and that further studies are necessary for the more recalcitrant ones.     

Viability of pulp stromal cells in cryopreserved deciduous teeth

The cryopreservation of exfoliated deciduous teeth and harvesting of stem cells from them as required would reduce the costs and efforts associated with banking stem cells from primary teeth. The aim of this study was determine whether the viability of pulp stromal cells from deciduous teeth was influenced by the cryopreservation process itself or the period of cryopreservation. In total, 126 deciduous teeth were divided into three groups: (1) fresh, (2) cryopreserved for <3 months (cryo<3), and (3) cryopreserved for 3–9 months (cryo3–9). The viability of the pulp tissues was compared among the three groups by evaluating the outgrowth from pulp tissues and cell activity within those pulp tissues. In addition, the terminal deoxynucleotidyl transferase-mediated dUTP–biotin nick end labeling (TUNEL) assay was performed to compare cell apoptosis within fresh pulp tissue and pulp tissue that had been cryopreserved for 4 months. The outgrowth from and cell activity within the pulp tissues did not differ significantly between the fresh and cryo<3 pulp tissues. However, these parameters were significantly reduced in the cryo3–9 pulp tissue. In TUNEL assay, 4-month cryopreserved pulp tissues has more apoptotic cells than fresh group. In conclusion, it is possible to acquire pulp stromal cells from cryopreserved deciduous teeth. However, as the period of cryopreservation becomes longer, it is difficult to get pulp cells due to reduced cell viability.     

Numeric anomalies of teeth in concomitant hypodontia and hyperdontia

Variations in tooth number in children, each of whom had supernumerary teeth and agenesis of teeth, is described. Among the 11, seven had cleft lip and palate, and two had clefting syndromes; two children had dental anomalies only. Only children who had both supernumerary teeth and congenitally missing teeth outside the area of the cleft alveolus were included. Concomitant hypodontia and hyperdontia were observed in the same dentition in nine subjects, in the same jaw in eight subjects, and in the same jaw quadrant in only three subjects. Supernumerary teeth and agenesis of teeth were observed simultaneously more often in the permanent dentitions than in the deciduous dentitions or in both dentitions simultaneously. The overall number of supernumeraries was 10 in the deciduous dentition and 14 in the permanent dentition of the 11 subjects. The number of congenitally absent teeth was 14 in the deciduous dentition and 40 in the permanent dentition. The etiology of concomitant hypodontia and hyperdontia is difficult to explain. It may result from disturbances in migration, proliferation, and differentiation of neural crest cells or interactions between the epithelial and mesenchymal cells during the initiation of odontogenesis.     

Cryopreservation of hybrid Cymbidium protocorm-like bodies by encapsulation–dehydration and vitrification

Cryopreservation can be a stable, long-term method of germplasm conservation, but successful application can be challenging for tropical material. To optimize survival and re-growth from cryopreserved tissues derived from protocorm-like bodies (PLBs) of hybrid Cymbidium Twilight Moon ‘Day Light’, the effects of explant type (intact PLBs, half-PLBs, or PLB longitudinal thin cell layers) and various explant treatments were studied. Encapsulation in alginate beads was essential, and intact PLBs were best for cryopreservation, based on survival and ability to form neo-PLBs and/or percentage re-growth. Osmotic hydration of intact PLBs in 2% sucrose for 24 h increased neo-PLB formation and re-growth, with the best responses seen when PLBs were excised from alginate beads prior to re-growth after cryopreservation. Both non-transgenic and transgenic PLBs were amenable to cryopreservation for up to 1 year using these methods. This optimized protocol will improve the viability of hybrid Cymbidium germplasm after long-term cryopreservation.     

Shoot regeneration and cryopreservation of shoot tips of apple (Malus) by encapsulation–dehydration

Here, we report an efficient and widely applicable method for cryopreservation of Malus shoot tips by encapsulation–dehydration using adventitious shoots. Shoots were induced from leaf segments cultured on a shoot induction medium containing 2–3 mg L^?1 thidiazuron, depending on genotype, and 0.5 mg L^?1 indole-3-butyric acid. Shoot tips (3 mm in length) containing six leaf primordia excised from 11-wk-old adventitious shoots were encapsulated and precultured with 0.5 M sucrose for 5 d, followed by air-drying for 6 h prior to direct immersion in liquid nitrogen. With our protocol, we obtained a mean organogenesis rate of 100%, a mean of 4.5 adventitious shoots per explant (leaf segment), and a mean shoot recovery of 57.0% from cryopreserved shoot tips in four Malus species. Inter-simple sequence repeat (ISSR) analysis did not reveal any polymorphic bands in regenerants recovered from either leaf segments or cryopreserved shoot tips of ‘Gala’. To the best of our knowledge, this is the first report on cryopreservation of Malus shoot tips using adventitious shoots derived from leaf segments and is the most widely applicable protocol so far reported for cryopreservation of Malus. Establishment of this protocol provides an alternative means for cryopreservation of Malus.     

Estimation of background radiation doses for the Peninsular Malaysia’s population by ESR dosimetry of tooth enamel

Background radiation dose is used in dosimetry for estimating occupational doses of radiation workers or determining radiation dose of an individual following accidental exposure. In the present study, the absorbed dose and the background radiation level are determined using the electron spin resonance (ESR) method on tooth samples. The effect of using different tooth surfaces and teeth exposed with single medical X-rays on the absorbed dose are also evaluated. A total of 48 molars of position 6–8 were collected from 13 district hospitals in Peninsular Malaysia. Thirty-six teeth had not been exposed to any excessive radiation, and 12 teeth had been directly exposed to a single X-ray dose during medical treatment prior to extraction. There was no significant effect of tooth surfaces and exposure with single X-rays on the measured absorbed dose of an individual. The mean measured absorbed dose of the population is 34 ± 6.2 mGy, with an average tooth enamel age of 39 years. From the slope of a regression line, the estimated annual background dose for Peninsular Malaysia is 0.6 ± 0.3 mGy y⁻¹. This value is slightly lower than the yearly background dose for Malaysia, and the radiation background dose is established by ESR tooth measurements on samples from India and Russia.     

On the geometry and mechanics of tooth position in the white shark,Carcharodon carcharias

The teeth of captured specimens, of prepared museum specimens, and of high-speed videotape images of the white shark, Carcharodon carcharias, were compared with respect to (1) deviation of each tooth from the animal's midline and (2) the crown angle of the functional teeth along the jaw margin. Tooth position was measured either directly using a meter stick apparatus or derived from tracings of the video footage. Tooth positions were not statistically unique in any region of the upper or lower jaw but demonstrated less variability in crown angle within 30° of the midline (71.48° ± 10°). Videotape analysis of feeding sharks indicated an 8.7° increase in crown angle of the centermost teeth during bites where the jaws were closed through an angle of 20–35° and a 15.7° reduction in this same parameter during jaw adduction through 35° or more. Such changes in tooth orientation (relative to the rear of the buccal cavity) are ascribed to flexure of the cartilaginous jaws and cranium by the cranial musculature and possibly also to sliding of the tooth bed over the jaw. Outward rotation of the teeth and jaw rami describes a plucking action during feeding or prey sampling, while larger bites rotate the frontmost teeth inward towards the gullet. Functionally, this may make the teeth more effective at grasping small prey items or gouging chunks from larger prey. However, testing of the load required to remove teeth showed no significant increase in tensile resistance with reduced crown angle. © 1995 Wiley-Liss, Inc.     

Stress analysis of irradiated human tooth enamel using finite element methods

The objectives of this project were to use finite element methods to determine how changes in the elastic modulus due to oral cancer therapeutic radiation alter the distribution of mechanical stresses in teeth and to determine if observed failures in irradiated teeth correlate with changes in mechanical stresses. A thin slice section finite element (FE) model was constructed from micro CT sections of a molar tooth using MIMICS and 3-Matic software. This model divides the tooth into three enamel regions, the dentin-enamel junction (DEJ) and dentin. The enamel elastic modulus was determined in each region using nano indentation for three experimental groups namely – control (non-radiated), in vitro irradiated (simulated radiotherapy following tooth extraction) and in vivo irradiated (extracted subsequent to oral cancer patient radiotherapy) teeth. Physiological loads were applied to the tooth models at the buccal and lingual cusp regions for all three groups (control, in vitro and in vivo). The principal tensile stress and the maximum shear stress were used to compare the results from different groups since it has been observed in previous studies that delamination of enamel from the underlying dentin was one of the major reasons for the failure of teeth following therapeutic radiation. From the FE data, we observed an increase in the principal tensile stress within the inner enamel region of in vivo irradiated teeth (9.97 ± 1.32 MPa) as compared to control/non-irradiated teeth (8.44 ± 1.57 MPa). Our model predicts that failure occurs at the inner enamel/DEJ interface due to extremely high tensile and maximum shear stresses in in vivo irradiated teeth which could be a cause of enamel delamination due to radiotherapy.     

Effects of cryopreservation with a newly-developed magnetic field programmed freezer on periodontal ligament cells and pulp tissues

The purpose of this study was to evaluate the effects of long-term cryopreservation on the isolated human periodontal ligament cells (PDL) and pulp tissues. In the first part of study, 10 freshly extracted teeth were selected and divided into two groups. In the cryopreserved group, the teeth were frozen for 5 years using a programmed freezer combined with a magnetic field, known as Cells Alive System “CAS”. As for the control group, freshly extracted teeth were used. In each group, extracted PDL tissues were cultured and gene expression and protein concentration of collagen type I, alkaline-phosphatase (ALP) and vascular endothelial growth factor (VEGF) was compared between the two groups. In the second part, pulp tissues were obtained from 10 mature and immature third molars which were freshly extracted or cryopreserved for three months. Expression of VEGF and nerve growth factor (NGF) mRNAs and the protein concentration in the supernatant were investigated. Results indicated that long-term cryopreservation with the use of CAS freezer cannot affect the growth rate and characteristics of PDL cells. There was no significant difference in VEGF expression and VEGF and NGF protein concentration of pulp cells derived from cryopreserved teeth with immature apex and control group with mature root formation. Finally, proper PDL regeneration and appropriate apexogenesis after transplanting magnetically cryopreserved immature tooth was clinically confirmed. These findings demonstrate that teeth banking with the use of magnetic field programmed freezer can be available for future autotransplantation as a treatment modality for replacing missing teeth.     

A Simple and Rapid Cryopreservation Technique for Ciliates: A Long‐Term Storage Procedure Used for Marine Scuticociliates

Pseudocohnilembus persalinus is a free‐living marine scuticociliate that, as a new model organism, has been used in a wide variety of studies. However, long‐term laboratory maintenance for this species is mainly achieved by subculture that requires rigorous culture environments and, too often, cultures of the organism die out for a variety of reasons. Successful transport of viable cultures also poses problems for researchers. This study describes a simple and rapid protocol for long‐term cryopreservation of P. persalinus. The effects of physiological states of individuals before freezing, the type and concentration of cryoprotectant, and optimal temperatures for freezing and thawing were assessed. A cryopreservation protocol, using a mixture of 30% glycerol and 70% concentrated P. persalinus cell culture, incorporating rate‐controlled freezing at ?80 °C before liquid nitrogen storage, maintained a high recovery efficiency after 8 wk of storage. These results suggest that broader application of this protocol to build a cryopreserved marine protozoa culture bank for biological studies may be possible.     

WNT10A variants are associated with non-syndromic tooth agenesis in the general population

Tooth agenesis is the most common developmental dental anomaly. Absence of one or two permanent teeth is found in the majority of affected subjects. Very few patients suffer severe tooth agenesis. Recent studies revealed that WNT10A gene mutations caused syndromic and isolated severe tooth agenesis. In this study, to determine the contribution of WNT10A variants in different severities of tooth agenesis, we investigated the association between WNT10A variants and non-syndromic tooth agenesis in a Chinese population consisting of 505 tooth agenesis patients and 451 normal controls. Twenty-three novel non-synonymous variants were identified. WNT10A variants were detected in 15.8 % (75/474) of patients with 1–3 missing teeth and 51.6 % (16/31) of patients with 4 or more missing teeth. As compared with a frequency of 3.1 % in individuals with full dentition, variant allele frequencies were significantly elevated in both groups with tooth agenesis (p values of 1.00 × 10^?6 and 3.89 × 10^?23, respectively). Our findings showed that WNT10A variants were associated with non-syndromic tooth agenesis from mild to severe tooth agenesis, and the more severe tooth agenesis, the stronger association. Biallelic genotypes of WNT10A variants may have a pathogenic effect on tooth development. Presence of a single variant allele would be predisposing for causation with low penetrance. Together with WNT10A variant, there should be other genetic or environmental factors leading to biallelic variant-related variable clinical manifestations and single allele variant-related low penetrance. The frequent missing tooth positions in the WNT10A-related cases were consistent with that in the general population, suggesting WNT10A plays a critically important role in the etiology of general tooth agenesis.     

Cryopreservation of red raspberry cultivars from the VIR <Emphasis Type="Italic">in vitro</Emphasis> collection using a modified droplet vitrification method

The goal of the present study was to analyze the post-cryogenic recovery of 12 red raspberry cultivars from the N.I. Vavilov All-Russian Institute of Plant Genetic Resources in vitro collection. The 1.1–1.8 mm shoot tips of microplants were subjected to cryopreservation using the modified droplet vitrification method. The current modifications to the droplet vitrification protocol included the elimination of the initial pretreatment stage of the microplants, and the use of modified media at the stages of initial micropropagation, explant isolation, and post-cryogenic regeneration. The optimized method reduced the duration of some cryopreservation stages compared to the initial protocol, and reduced the total procedure from 14 to 11 wk. This modified cryopreservation method also demonstrated a relatively high level of post-cryogenic regeneration. Depending on the genotype, the shoot recovery of explants after rewarming varied from 24.2–89.3% and averaged 58.8 ± 5.3%. There was a statistically significant influence of the genotype on the shoot recovery after rewarming. No differences in inter simple sequence repeats and in start codon targeted marker spectra were found between post-cryopreservation microplants and donor in vitro plants from two red raspberry cultivars.     

Optimization of a working cryopreservation protocol for Pinus patula embryogenic tissue

Summary The protocol currently used to cryopreserve Pinus patula embryogenic tissue was investigated in an attempt to improve and optimize the recovery of tissue. This investigation describes two aspects that influence tissue recovery after cryopreservation: (i) the effects of precooling tissue prior to immersion into liquid nitrogen; and (ii) whether the choice of supports onto which the recovered tissue is suspended improved the recovery rate. Results indicated that precooling tissue to −70°C prior to immersion into liquid nitrogen was superior to precooling to −30°C or direct immersion in liquid nitrogen (−196°C). Tissue recovery improved when polyester grids were used as supports, and was slowest on filter paper supports.     

Dentition and tooth replacement pattern in Chalcides (Squamata; Scincidae)

This study was undertaken as a prerequisite to investigations on tooth differentiation in a squamate, the Canarian scincid Chalcides. Our main goal was to determine whether the pattern of tooth replacement, known to be regular in lizards, could be helpful to predict accurately any stage of tooth development. A growth series of 20 laboratory-reared specimens, aged from 0.5 month after birth to about 6 years, was used. The dentition (functional and replacement teeth) was studied from radiographs of jaw quadrants. The number of tooth positions, the tooth number in relation to age and to seasons, and the size of the replacement teeth were recorded. In Chalcides, a single row of pleurodont functional teeth lies at the labial margin of the dentary, premaxillary, and maxillary. Whatever the age of the specimens, 16 tooth positions were recorded, on average, in each quadrant, suggesting that positions are maintained throughout life. Replacement teeth were numerous whatever the age and season, while the number of functional teeth was subject to variation. Symmetry of tooth development was evaluated by comparing teeth two by two from the opposite side in the four jaw quadrants of several specimens. Although the relative size of some replacement teeth fitted perfectly, the symmetry criterion was not reliable to predict the developmental stage of the opposite tooth, whether the pair of teeth compared was left-right or upper-lower. The best fit was found when comparing the size of successive replacement teeth from the front to the back of the jaw. Every replacement tooth that is 40-80% of its definitive size is followed, in the next position on the arcade, by a tooth that is, on average, 20% less developed. Considering teeth in alternate positions (even and odd series), each replacement tooth was a little more developed than the previous, more anterior, one (0.5-20% when the teeth are from 10-40% of their final size). The latter pattern showed that tooth replacement occurred in alternate positions from back to front, forming more or less regular rows (i.e., "Zahnreihen"). In Chalcides, the developmental stage of a replacement tooth in a position p can be accurately predicted provided the developmental stage of the replacement tooth in position p-1 or, to a lesser degree, in position p-2 is known. This finding will be particularly helpful when starting our structural and ultrastructural studies of tooth differentiation in this lizard.     

Analysis of Mtwo Rotary Instrument Separation During Endodontic Therapy: A Retrospective Clinical Study

To analyze the incidence of instrument separation (IS) and the factors influencing it, when associated with Mtwo rotary system (VDW, Munich, Germany) during endodontic therapy. A retrospective study involving a total of 24,108 root canals (11,036 endodontic treated teeth) was conducted at Nanjing Stomatology Hospital between January 2011 and March 2013. The information included were tooth type, root canal curvature, number of fractured instruments, length of the separated fragments, and the distance from broken tip to apex. The incidence of IS was observed to be 2.2 % according to the number of teeth and 1.0 % according to the number of root canals. Many of the separated fragments were 2–4 mm in length and the mean length was 3.07 ± 1.46 mm, and 78.4 % of fractures occurred in the apex. The mean length of separated fragments in severely curved canals was maximum, while ultra-severe curved canals was observed to be minimum. Mtwo instruments demonstrated an extremely low fracture rate during endodontic therapy. Molar teeth (especially lower molars) and the degree of canal curvature had a significant effect on the incidence of IS.