Identification of root canal microbiota profiles of periapical tissue diseases using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

OBJECTIVES: The purpose of this study was to identify microorganisms isolated from various periapical tissue diseases using Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF-MS) and classify them via an unsupervised machine learning approach. METHODS: A total of 150 patients with various apical conditions and teeth in need of endodontic retreatment were divided into five groups, including Retreatment, Acute Apical Abscess, Chronic Apical Abscess, Acute Apical Periodontitis, and Chronic Apical Periodontitis. Samples were collected from root canals using paper points after agitating with a #10 K file then microorganisms were identified using MALDI-TOF-MS. Data were analyzed using a hierarchical clustering method. Quadruple clusters and dendrograms were formed according to similarities and dissimilarities. RESULTS: A total of 80 species were identified representative of six different phyla. The most similar microorganism species identified were: ''Enterococcus faecalis'' between 21 and 23-year-old female cases in Retreatment group; ''Lactobacillus rhamnosus'' between 20 and 18-year-old male cases in Symptomatic Apical Abscess cases; ''Lactobacillus paracasei'' between 26 and 40-year-old male cases in Asymptomatic Apical Abscess cases; ''Enterococcus faecalis'' between 48 and 50-year-old female cases in Symptomatic Apical Periodontitis cases; ''Lactobacillus rhamnosus'' between 48 and 60-year-old male cases in Asymptomatic Apical Periodontitis cases. CONCLUSIONS: MALDI-TOF MS can be considered a fast and high-throughput screening technique for microbial species identification in endodontics. Thus, it will provide valuable data for future research designs regarding periapical tissue diseases. As the MALDI-TOF MS database expands and comprehensive data becomes available, the relationship between microbial profiles and disease progression will become increasingly apparent.

Comparison of Four Electronic Apex Locators in Detecting Working Length: An Ex Vivo Study.

AIM: To compare the accuracy of four different electronic apex locators (EALs) in detecting a position 0.5 mm short of the major foramen. MATERIALS AND METHODS: The actual working length of thirty-five extracted human teeth was determined visually as 0.5 mm short of the apical foramen. After actual working length measurements, electronic working length was measured with four different EALs (Apex Pointer+, Raypex 5, Apex ID, and Raypex 6). Measurements were repeated three times by different operators. The data were analyzed using the intraclass correlation coefficient (ICC), the repeated measure analysis of variance (rANOVA) and Bonferroni post hoc tests. The significance level was set at p < 0.05. RESULTS: The mean differences between electronic and actual working length were-0.305 mm, 0.098 mm, 0.037, and 0.144 mm for the Apex Pointer+, the Raypex 5, the Apex ID, and the Raypex 6, respectively. Multiple paired comparisons (Bonferroni test) also showed the Apex Pointer+ is significantly different from the Raypex 5, Apex ID and Raypex 6 (p = 0.000, p = 0.001, and p = 0.001 respectively). CONCLUSION: All EALs showed an acceptable determination of the working length between the ranges of ± 0.5mm except for the Apex Pointer+ device, which had the lowest accuracy. Further studies may be beneficial especially to better evaluate the accuracy of the Apex Pointer+. CLINICAL SIGNIFICANCE: This article shows that Apex ID, which has only recently been introduced into the market, showed an acceptable determination of the working length. Its accuracy was similar to that of Raypex 5 and 6.

Solubility and apical sealing characteristics of a new calcium silicate-based root canal sealer in comparison to calcium hydroxide-, methacrylate resin- and epoxy resin-based sealers.

AIM: To assess and compare the water sorption, solubility and apical sealing ability of iRoot SP and three other widely used root canal sealers. MATERIALS AND METHODS: Solubility was assessed by immersing standardized samples of calcium silicate- (iRoot SP), calcium hydroxide- (Sealapex), methacrylate resin- (EndoREZ) and epoxy resin- (AH Plus) based sealers in distilled water and measuring weight gain and weight loss at 6 h, 24 h and daily for 14 days. Roots of extracted mandibular premolars (n = 80) were prepared with 0.04-taper nickel-titanium rotary files to a final size 40. Roots were then randomly divided into four experimental groups (n = 18) and two control groups (n = 4), root canal sealers were applied and apical leakage was assessed using the fluid filtration method. Data was analyzed using Kruskal Wallis analysis of variance and Mann-Whitney U-tests, with the level of significance set at p ≤ 0.05. RESULTS: EndoREZ exhibited the highest water sorption, followed by iRoot SP, Sealapex and AH Plus. Sealapex exhibited significantly higher solubility than the other sealers, whereas no significant differences in solubility levels were observed between the other three sealers tested. AH Plus exhibited significantly lower microleakage than Sealapex and EndoREZ, whereas no difference in microleakage was found between AH Plus and iRoot SP. CONCLUSIONS: In view of the study findings, all tested sealers except Sealapex met the ANSI/ADA's requirements for solubility and no difference was found between AH Plus and iRoot SP in terms of apical sealing ability.

Contemporary research trends on nanoparticles in endodontics: a bibliometric and scientometric analysis of the top 100 most-cited articles.

Objectives: Advancements in nanotechnology have led to the widespread usage of nanoparticles in the endodontic field. This bibliometric study aimed to determine and analyze the top 100 most-cited articles about nanoparticles in endodontics from 2000 to 2022. Materials and Methods: A detailed electronic search was conducted on the "Clarivate Analytics Web of Science, All Databases" to receive the most-cited articles related to the topic. Articles were ranked in descending order based on their citation counts, and the first 100 were selected for bibliometric analysis. Parameters such as citation density, publication year, journal, country, institution, author, study design, study field, evidence level, and keywords were analyzed. Results: The top 100 most-cited articles received 4,698 citations (16-271) with 970.21 (1.91-181) citation density in total. Among decades, citations were significantly higher in 2011-2022 (p < 0.001). Journal of Endodontics had the largest number of publications. Canada and the University of Toronto made the highest contribution as country and institution, respectively. Anil Kishen was the 1 who participated in the largest number of articles. The majority of the articles were designed in vitro. The main study field was "antibacterial effect." Among keywords, "nanoparticles" followed by "Enterococcus faecalis" were used more frequently. Conclusions: Developments in nanotechnology had an impact on the increasing number of studies in recent years. This bibliometric study provides a comprehensive view of nanoparticle advances and trends using citation analysis.

A case of a benign cementoblastoma treated by enucleation and apicoectomy.

Cementoblastoma is a rare, benign, odontogenic neoplasm of ectomesenchymal origin, representing less than 6% of all odontogenic tumors. Despite its well-known typical features, there are still controversies regarding the management of the condition. This article presents the case of a benign cementoblastoma in a 17-year-old girl. The lesion was typical and associated with the mandibular right first molar. Endodontic treatment of the involved tooth, enucleation of the cementoblastoma, and apicoectomy of the affected roots were performed. Removal of the tumor while preserving the associated tooth resulted in normal osseous healing and no evidence of recurrence after one year. Based on the findings of the current report, it can be suggested that, in properly selected cases, it is possible to remove cementoblastomas that affect molars without extracting the involved teeth.

Functional characterization of GNAS mutations found in patients with pseudohypoparathyroidism type Ic defines a new subgroup of pseudohypoparathyroidism affecting selectively Gsα-receptor interaction.

Pseudohypoparathyroidism type Ia (PHPIa) is caused by GNAS mutations leading to deficiency of the α-subunit of stimulatory G proteins (Gsα) that mediate signal transduction of G protein-coupled receptors via cAMP. PHP type Ic (PHPIc) and PHPIa share clinical features of Albright hereditary osteodystrophy (AHO); however, in vitro activity of solubilized Gsα protein is normal in PHPIc but reduced in PHPIa. We screened 32 patients classified as PHPIc for GNAS mutations and identified three mutations (p.E392K, p.E392X, p.L388R) in four unrelated families. These and one novel mutation associated with PHPIa (p.L388P) were introduced into a pcDNA3.1(-) expression vector encoding Gsα wild-type and expressed in a Gsα-null cell line (Gnas(E2-/E2-) ). To investigate receptor-mediated cAMP accumulation, we stimulated the endogenous expressed ß(2) -adrenergic receptor, or the coexpressed PTH or TSH receptors, and measured the synthesized cAMP by RIA. The results were compared to receptor-independent cholera toxin-induced cAMP accumulation. Each of the mutants associated with PHPIc significantly reduced or completely disrupted receptor-mediated activation, but displayed normal receptor-independent activation. In contrast, PHPIa associated p.L388P disrupted both receptor-mediated activation and receptor-independent activation. We present a new subgroup of PHP that is caused by Gsα deficiency and selectively affects receptor coupling functions of Gsα.

Is endodontic treatment necessary during coronectomy procedure?

PURPOSE: Close proximity of the inferior alveolar nerve (IAN) to the third molar roots can result in nerve injuries during extraction of third molars. Consequently, it is necessary to determine the relationship of the nerve and roots to avoid damage to the IAN. Computed tomography scans are widely used to determine the correct relationship between the IAN and lower third molars. PATIENTS AND METHODS: The study consisted of 10 patients with 16 lower third molars in close relationship with the IAN who were divided into a study group and a control group. The patients in the study group were treated via coronectomies performed with endodontic treatments. The patients in the control group underwent coronectomies without endodontic treatment. The patients were followed up for at least 1 year. RESULTS: We had to extract 7 of the roots because of the infection in 8 patients belonging to the study group, which were treated endodontically. Moreover, there were 3 cases of IAN damage because of the extraction in the study group. However, in the control group, no infection was determined and IAN damage was absent. CONCLUSION: Coronectomy appears to be a reliable technique to protect the IAN from damage. This procedure has a low incidence of complications. Endodontic treatment does not affect the success of this method according to our results.

In vitro evaluation of matched-taper single-cone obturation with a fluid filtration method.

OBJECTIVE: Widespread use of rotary nickel-titanium systems has caused manufacturers to produce gutta-percha cones that match the taper of the canals prepared with these systems. Obturations with these cones may provide 3-dimensional obturation of the root canal in less time. The aim of this study was to use a fluid filtration method to compare the apical sealing ability of matched-taper single-cone, Thermafil and lateral condensation obturations. MATERIALS AND METHODS: Sixty-six extracted human lower premolar teeth were instrumented to size F3 with ProTaper instruments. The teeth were divided into 3 groups and obturated with matched-taper single-cone, Thermafil and lateral condensation techniques. AH Plus was used as a sealer. Six teeth were used as positive and negative controls. A fluid filtration system was used to evaluate the apical seal. Measurements were made at 2-minute intervals for 8 minutes and the average of these values was recorded. One-way analysis of variance test was used for statistical analysis. RESULTS: Mean leakage values for the lateral condensation, single-cone and Thermafil groups were 0.49 +/- 0.37 microL min(-1), 0.39 +/- 0.24 microL min(-1) and 0.44 +/- 0.25 microL min(-1), respectively. The difference between the groups was not statistically significant (p > 0.05). CONCLUSIONS: The apical sealing ability of matched-taper single-cone obturation was comparable with that of lateral condensation and Thermafil techniques.

A G protein-coupled, IP3/protein kinase C pathway controlling the synthesis of phosphaturic hormone FGF23.

Dysregulated actions of bone-derived phosphaturic hormone fibroblast growth factor 23 (FGF23) result in several inherited diseases, such as X-linked hypophosphatemia (XLH), and contribute substantially to the mortality in kidney failure. Mechanisms governing FGF23 production are poorly defined. We herein found that ablation of the Gq/11α-like, extralarge Gα subunit (XLαs), a product of GNAS, exhibits FGF23 deficiency and hyperphosphatemia in early postnatal mice (XLKO). FGF23 elevation in response to parathyroid hormone, a stimulator of FGF23 production via cAMP, was intact in XLKO mice, while skeletal levels of protein kinase C isoforms α and δ (PKCα and PKCδ) were diminished. XLαs ablation in osteocyte-like Ocy454 cells suppressed the levels of FGF23 mRNA, inositol 1,4,5-trisphosphate (IP3), and PKCα/PKCδ proteins. PKC activation in vivo via injecting phorbol myristate acetate (PMA) or by constitutively active Gqα-Q209L in osteocytes and osteoblasts promoted FGF23 production. Molecular studies showed that the PKC activation-induced FGF23 elevation was dependent on MAPK signaling. The baseline PKC activity was elevated in bones of Hyp mice, a model of XLH. XLαs ablation significantly, but modestly, reduced serum FGF23 and elevated serum phosphate in Hyp mice. These findings reveal a potentially hitherto-unknown mechanism of FGF23 synthesis involving a G protein-coupled IP3/PKC pathway, which may be targeted to fine-tune FGF23 levels.

Re-eruption of an intruded immature permanent incisor with necrotic pulp: a case report.

Intrusive luxation is a serious dental injury that causes damage to the pulp and supporting structures of a tooth because of its dislocation into the alveolar process. This paper presents the case of the re-eruption of a severely intruded immature permanent incisor with a crown-root fracture. A 9-year-old boy was referred to the clinic 1 day after a fall. Clinical and radiographic examinations revealed intrusive luxation of the immature left permanent incisor and a crown-root fracture without pulp exposure. Palatal gingivectomy was done 2 weeks later to facilitate re-eruption. Root canal therapy with intracanal calcium hydroxide paste was initiated during the first month owing to severe spontaneous pain. Six months later, the tooth re-erupted to a normal position, after which root canal obturation and a final esthetic restoration were done. The present case demonstrates the possibility of obtaining re-eruption of intruded immature permanent teeth with interim medication (calcium hydroxide) in the root canal.

Evaluation of the surface characteristics of used and new ProTaper Instruments: an atomic force microscopy study.

The purpose of this study was to evaluate the topography of new and used ProTaper rotary nickel-titanium (NiTi) instruments by using atomic force microscope. Four new and four used size S1, S2, F1, and F2 instruments were selected for this study. New and used instruments were analyzed on 11 points along a 3-mm section at the tip of the instrument. Quantitative measurements according to the topographic deviations (root mean square) were recorded. Data were analyzed by paired samples t test. Mean root mean square values for used ProTaper instruments were higher than the new ones, and the difference between them was statistically significant. The results of this study showed that used ProTaper instruments demonstrated more surface deformation and wear.

The effect of mineral trioxide aggregate on the contractility of the rat thoracic aorta.

Mineral trioxide aggregate (MTA) is being advocated for vital pulp therapy. In the case of direct pulp capping, hemorrhage control is an important step on success, but little is known about the effect of MTA on pulpal bleeding. In addition, there has been a lack of information on the effect of MTA on smooth muscle contraction, such as can occur in the blood vessels of dental pulp. The present study assessed the vascular effects of MTA, using the rat aortic ring preparations as a tissue model. MTA (100-500 mg) induced dose-dependent contraction in rat thoracic aorta. The contractile effect of MTA was blocked by calcium channel blocker nifedipine (1 muM). These data suggest that the vasoconstrictor property of MTA is related to calcium influx and it may allow proper control of hemorrhage which is critical for the success of any pulp-capping treatment.

Pedestrian self-reports of factors influencing the use of pedestrian bridges.

The study was designed to find out factors that influence use/non-use of pedestrian bridges. The use rate of five pedestrian bridges was observed in the central business district (CBD) of Ankara. After the observations, a survey was conducted among pedestrians using those bridges and crossing contrary to safe practice under them at street level (n=408). In the present data, the use rate of pedestrian bridges varied from 6 to 63%. The frequent use of the bridge when crossing the road concerned, and seeing bridge use as time saving and safe in general were positively related to respondents' bridge use. Frequent visits to CBD decreased the likelihood of using the bridge. Other factors accounted only for a small proportion of variance in bridge use. The study suggests that bridge use or non-use is a habit and not coincidental behaviour. For increasing the pedestrians' bridge use, escalators seem to be a good solution, but traffic signals under a bridge may deteriorate the use rate. In addition, increasing the number of legs leading to the bridge may not increase the use rate. The use rate is likely to improve, if the safety benefits and convenience of using the bridge without considerable time loss are clearly visible to pedestrians.

Ablation of the Stimulatory G Protein α-Subunit in Renal Proximal Tubules Leads to Parathyroid Hormone-Resistance With Increased Renal Cyp24a1 mRNA Abundance and Reduced Serum 1,25-Dihydroxyvitamin D.

PTH regulates serum calcium, phosphate, and 1,25-dihydroxyvitamin D (1,25(OH)2D) levels by acting on bone and kidney. In renal proximal tubules (PTs), PTH inhibits reabsorption of phosphate and stimulates the synthesis of 1,25(OH)2D. The PTH receptor couples to multiple G proteins. We here ablated the α-subunit of the stimulatory G protein (Gsα) in mouse PTs by using Cre recombinase driven by the promoter of type-2 sodium-glucose cotransporter (Gsα(Sglt2KO) mice). Gsα(Sglt2KO) mice were normophosphatemic but displayed, relative to controls, hypocalcemia (1.19 ±0.01 vs 1.23 ±0.01 mmol/L; P < .05), reduced serum 1,25(OH)2D (59.3 ±7.0 vs 102.5 ±12.2 pmol/L; P < .05), and elevated serum PTH (834 ±133 vs 438 ±59 pg/mL; P < .05). PTH-induced elevation in urinary cAMP excretion was blunted in Gsα(Sglt2KO) mice (2- vs 4-fold over baseline in controls; P < .05). Relative to baseline in controls, PTH-induced reduction in serum phosphate tended to be blunted in Gsα(Sglt2KO) mice (-0.39 ±0.33 vs -1.34 ±0.36 mg/dL; P = .07). Gsα(Sglt2KO) mice showed elevated renal vitamin D 24-hydroxylase and bone fibroblast growth factor-23 (FGF23) mRNA abundance (∼3.4- and ∼11-fold over controls, respectively; P < .05) and tended to have elevated serum FGF23 (829 ±76 vs 632 ±60 pg/mL in controls; P = .07). Heterozygous mice having constitutive ablation of the maternal Gsα allele (E1(m-/+)) (model of pseudohypoparathyroidism type-Ia), in which Gsα levels in PT are reduced, also exhibited elevated serum FGF23 (474 ±20 vs 374 ±27 pg/mL in controls; P < .05). Our findings indicate that Gsα is required in PTs for suppressing renal vitamin D 24-hydroxylase mRNA levels and for maintaining normal serum 1,25(OH)2D.

The accuracy of three different electronic root canal measuring devices: an in vitro evaluation.

The main objective of this in vitro study was to evaluate the effects of the operator's experience level and pre-flaring on the accuracy of the results of three different brands of a new generation of root canal measuring devices, as well as the comparison among them. Extracted human teeth were prepared and then actual length and electronic length measurements were made by three different operators according to a double-blind technique. Three different operators performed electronic measurements on each specimen separately with three different electronic root canal measuring devices using in vitro models. Measurements were repeated by all operators after the pre-flaring. Taking the clinical tolerance of +/- 0.5 mm into account, there was no statistically significant difference between the accuracy of the instruments (P > 0.01). However, the results obtained from the Bingo electronic apex locator in pre-flared canals by the beginner operator were statistically significant (P < 0.01). All of the instruments had a clinically acceptable result at the tolerance of +/- 0.5 mm. If the instruments are used in accordance with the manufacturer's instructions, experience with electronic root canal measurement is not essential. However, the operator has to be more careful when working on pre-flared canals.

Postnatal establishment of allelic Gαs silencing as a plausible explanation for delayed onset of parathyroid hormone resistance owing to heterozygous Gαs disruption.

Pseudohypoparathyroidism type-Ia (PHP-Ia), characterized by renal proximal tubular resistance to parathyroid hormone (PTH), results from maternal mutations of GNAS that lead to loss of α-subunit of the stimulatory G protein (Gαs) activity. Gαs expression is paternally silenced in the renal proximal tubule, and this genomic event is critical for the development of PTH resistance, as patients display impaired hormone action only if the mutation is inherited maternally. The primary clinical finding of PHP-Ia is hypocalcemia, which can lead to various neuromuscular defects including seizures. PHP-Ia patients frequently do not present with hypocalcemia until after infancy, but it has remained uncertain whether PTH resistance occurs in a delayed fashion. Analyzing reported cases of PHP-Ia with documented GNAS mutations and mice heterozygous for disruption of Gnas, we herein determined that the manifestation of PTH resistance caused by the maternal loss of Gαs, ie, hypocalcemia and elevated serum PTH, occurs after early postnatal life. To investigate whether this delay could reflect gradual development of paternal Gαs silencing, we then analyzed renal proximal tubules isolated by laser capture microdissection from mice with either maternal or paternal disruption of Gnas. Our results revealed that, whereas expression of Gαs mRNA in this tissue is predominantly from the maternal Gnas allele at weaning (3 weeks postnatal) and in adulthood, the contributions of the maternal and paternal Gnas alleles to Gαs mRNA expression are equal at postnatal day 3. In contrast, we found that paternal Gαs expression is already markedly repressed in brown adipose tissue at birth. Thus, the mechanisms silencing the paternal Gαs allele in renal proximal tubules are not operational during early postnatal development, and this finding correlates well with the latency of PTH resistance in patients with PHP-Ia.

The evaluation of two rotary instrumentation techniques under "operator-related variables" standardized conditions.

OBJECTIVE: We aimed here to evaluate the enlargement characteristics of two nickel-titanium rotary instrument systems that use two different preparation techniques, in simulated root canals under "operator-related variables" standardized conditions. MATERIALS AND METHODS: A total of 40 simulated canals in resin blocks were divided into two groups as the Mtwo group for use with a single-length technique and the ProTaper instrument system group for use with crown down technique. To standardize the operator-related variables, all preparations were carried out with a computer controlled device that was developed and used previously as described in a published study. The pre- and post-operative digital images of the canals were superimposed and measured at 11 levels from the apical tip. The distances between the pre- and post-operative outer lines of the root canals were measured at both the inner and outer sides of the curve. The amount of removed material and the symmetry of the preparations were evaluated. Statistical analyses were performed with Mann-Whitney test. RESULTS: Significantly more material was removed by Mtwo at 9 levels and by ProTaper at 2 levels (P < 0.05) while no significant difference was determined at 11 levels. The preparations made with ProTaper were more symmetrical at 4 levels while no significant difference was determined at 7 levels. CONCLUSION: Mtwo removed significantly more material than ProTaper at different levels of the curved root canals. Mtwo and ProTaper made symmetrical preparations and maintained the original shape of the root canal at different levels under controlled operator-related variables.

Comparison of cyclic fatigue resistance of three different rotary nickel-titanium instruments designed for retreatment.

INTRODUCTION: A number of rotary nickel-titanium (NiTi) systems have been developed to provide better, faster, and easier cleaning and shaping of the root canal system, and recently, rotary NiTi systems designed for root canal retreatment have been introduced. Because the main problem with the rotary NiTi files is fracture, the aim of this study was to compare the cyclic fatigue resistance of 3 different rotary NiTi systems designed for root canal retreatment. METHODS: Total of 60 instruments of 3 different rotary NiTi systems designed for root canal retreatment were used in this study. Twenty R-Endo R3, 20 ProTaper D3, and 20 Mtwo R (Retreatment) 25.05 instruments were tested. Cyclic fatigue testing of instruments was performed by using a device that allowed the instruments to rotate freely inside an artificial canal. Each instrument was rotated until fracture occurred, and the number of cycles to fracture for each instrument was calculated. Representative samples were also evaluated under a scanning electron microscope to confirm the fracture was flexural. Data were analyzed by using 1-way analysis of variance test. RESULTS: R-Endo R3 instruments showed better cyclic fatigue resistance than ProTaper D3 and Mtwo R 25.05 instruments, and the difference was statistically significant (P < .05). There was no significant difference between ProTaper D3 and Mtwo R 25.05 groups. CONCLUSIONS: The R-Endo R3 instruments were more resistant to fatigue failure than ProTaper D3 and Mtwo R 25.05.

Cyclic fatigue resistance of new and used Mtwo rotary nickel-titanium instruments in two different radii of curvature.

The fracture of nickel-titanium rotary instruments is a serious problem that can jeopardise the outcome of root canal treatment. The aim of this study was to compare the cyclic fatigue resistance of new and used Mtwo rotary nickel-titanium instruments in artificial canals with two different radii of curvature. Four hundred and twenty Mtwo instruments were used in the present study. Thirty new and 30 used instruments of each size were tested for cyclic fatigue in artificial canals with radii of curvature of 5 or 10 mm. Each instrument was rotated until fracture occurred and the number of cycles to fracture was calculated by multiplying time to fracture and rotational speed. Data were analysed using a t-test. According to the results of the study, radius of curvature was an important factor on cyclic fatigue resistance and clinical use significantly reduced the instrument lifespan.

Potent constitutive cyclic AMP-generating activity of XLαs implicates this imprinted GNAS product in the pathogenesis of McCune-Albright syndrome and fibrous dysplasia of bone.

Patients with McCune-Albright syndrome (MAS), characterized primarily by hyperpigmented skin lesions, precocious puberty, and fibrous dyslasia of bone, carry postzygotic heterozygous mutations of GNAS causing constitutive cAMP signaling. GNAS encodes the α-subunit of the stimulatory G protein (Gsα), as well as a large variant (XLαs) derived from the paternal allele. The mutations causing MAS affect both GNAS products, but whether XLαs, like Gsα, can be involved in the pathogenesis remains unknown. Here, we investigated biopsy samples from four previously reported and eight new patients with MAS. Activating mutations of GNAS (Arg201 with respect to the amino acid sequence of Gsα) were present in all the previously reported and five of the new cases. The mutation was detected within the paternally expressed XLαs transcript in five and the maternally expressed NESP55 transcript in four cases. Tissues carrying paternal mutations appeared to have higher XLαs mRNA levels than maternal mutations. The human XLαs mutant analogous to Gsα-R201H (XLαs-R543H) showed markedly higher basal cAMP accumulation than wild-type XLαs in transfected cells. Wild-type XLαs demonstrated higher basal and isoproterenol-induced cAMP signaling than Gsα and co-purified with Gß1γ2 in transduced cells. XLαs mRNA was measurable in mouse calvarial cells, with its level being significantly higher in undifferentiated cells than those expressing preosteoblastic markers osterix and alkaline phosphatase. XLαs mRNA was also expressed in murine bone marrow stromal cells and preosteoblastic MC3T3-E1 cells. Our findings are consistent with the possibility that constitutive XLαs activity adds to the molecular pathogenesis of MAS and fibrous dysplasia of bone.