Radiographic technical quality of root canal treatment performed ex vivo by dental students at Valencia University Medical and Dental School, Spain

performed on extracted teeth by undergraduate dental students. Study Design: A total of 561 premolars and molars extracted teeth were prepared using nickel-titanium rotary files or manual instrumentation and filled with gutta-percha using a cold lateral condensation technique, by 4th grade undergraduate students. Periapical radiographs were used to assess the technical quality of the root canal filling, evaluating three variables: length, density and taper. These data were recorded, scored and used to study the "technical success rate" and the "overall score". The length of each root canal filling was classified as acceptable, short and overfilled, based on their relationship with the radiographic apex. Density and taper of filling were evaluated based on the presence of voids and the uniform tapering of the filling, respectively. Statistical analysis was used to evaluate the quality of root canal treatment, considering p < 0.05 as a statistical significant level. Results: The percentage of technical success was 44% and the overall score was 7.8 out of 10. Technical success and overall score were greater with rotary instruments (52% against 28% with a manual one, p < 0.001; 8.3 against 6.7 respectively, p < 0.001). Conclusions: It appears that inexperienced operators perform better root canal treatment (RCT) with the use of rotary instrumentation

Dentin growth after direct pulp capping with the different fractions of plasma rich in growth factors (PRGF) vs MTA: experimental study in animal model

Background: The study aimed to evaluate the area of dentin growth in rabbit incisors after pulp capping with plasma rich in growth factors (PRGF) compared with mineral trioxide aggregate (MTA) by fluorescence. Methods: 27 upper and lower incisors of rabbits were divided into 4 groups: poor PRGF (F1) (n = 9 teeth), rich PRGF (F2) (n = 8 teeth), ProRoot MTA (positive control, n = 5 teeth) and untreated (NC) (negative control, n=5). Fluorochrome markers were injected 24 hours before surgery and the day before euthanasia, 28 days after the vital pulp therapy (VPT). Two transverse cuts were performed to every tooth: the first cut (A), 1 mm incisal to the gingival margin, and the second cut (B), 5 mm apical to the first cut. The sections were assessed with histomorphometric evaluation by fluorescence microscopy, comparing the dentin area between fluorescence marks and the total mineralized area. Results: The higher percentage of dentin growth was observed in the F2 group (B=63.25%, A=36.52%), followed by F1 (B=57.63%, A=30,12%) and MTA (B=38.64%, A=15,74%) The group with lowest percentage of dentin growth was the NC group (B=29.22%, A=7.82%). Significant difference (p <0.05) was found between F2 group and MTA, also statistically significant difference has been observed comparing dentin growth areas of NC group with F1 and F2 groups. Conclusions: The application of PRGF rich and poor fraction as a pulp capping material stimulated dentin formation significantly more intensively than MTA and NC

Spontaneous mouse lymphoma in patient-derived tumor xenografts: The importance of systematic analysis of xenografted human tumor tissues in preclinical efficacy trials

Patient-derived tumor xenograft (PDX) is now largely recognized as a key preclinical model for cancer research, mimicking patient tumor phenotype and genotype. Immunodeficient mice, well-known to develop spontaneous lymphoma, are required for PDX growth. As for all animal models used for further clinical translation, a robust experimental design is strongly required to lead to conclusive results. Here we briefly report unintentional co-engraftment of mouse lymphoma during expansion of well-established PDXs to illustrate the importance of systematic check of the PDX identity to avoid misinterpretation. Besides, this quality control based on complementary approaches deserves a more detailed description in materials and methods section to ensure experimental validity and reproducibility

In situ protein expression in tumour spheres: development of an immunostaining protocol for confocal microscopy.

International audienceBACKGROUND: Multicellular tumour sphere models have been shown to closely mimic phenotype characteristics of in vivo solid tumours, or to allow in vitro propagation of cancer stem cells (CSCs). CSCs are usually characterized by the expression of specific membrane markers using flow cytometry (FC) after enzymatic dissociation. Consequently, the spatial location of positive cells within spheres is not documented. Confocal microscopy is the best technique for the imaging of thick biological specimens after multi-labelling but suffers from poor antibody penetration. Thus, we describe here a new protocol for in situ confocal imaging of protein expression in intact spheroids. METHODS: Protein expression in whole spheroids (150 mum in diameter) from two human colon cancer cell lines, HT29 and CT320X6, has been investigated with confocal immunostaining, then compared with profiles obtained through paraffin immunohistochemistry (pIHC) and FC. Target antigens, relevant for colon cancer and with different expression patterns, have been studied. RESULTS: We first demonstrate that our procedure overcomes the well-known problem of antibody penetration in compact structures by performing immunostaining of EpCAM, a membrane protein expressed by all cells within our spheroids. EpCAM expression is detected in all cells, even the deepest ones. Likewise, antibody access is confirmed with CK20 and CD44 immunostaining. Confocal imaging shows that 100% of cells express beta-catenin, mainly present in the plasma membrane with also cytoplasmic and nuclear staining, in agreement with FC and pIHC data. pIHC and confocal imaging show similar CA 19-9 cytoplasmic and membranar expression profile in a cell subpopulation. CA 19-9+ cell count confirms confocal imaging as a highly sensitive method (75%, 62% and 51%, for FC, confocal imaging and pIHC, respectively). Finally, confocal imaging reveals that the weak expression of CD133, a putative colon CSC marker, is restricted to the luminal cell surface of colorectal cancer acini, with CD133+ cellular debris into glandular lumina. CONCLUSION: The present protocol enables in situ visualization of protein expression in compact three-dimensional models by whole mount confocal imaging, allowing the accurate localization and quantification of cells expressing specific markers. It should prove useful to study rare events like CSCs within tumour spheres

Cancer cells in the tumor core exhibit spatially coordinated migration patterns

International audienceIn early stages of metastasis, cancer cells exit the primary tumor and enter the vasculature. Although most studies have focused on the tumor invasive front, cancer cells from the tumor core can also potentially metastasize. To address cell motility in the tumor core, we imaged tumor explants from spontaneously-forming tumors in real time using long-term two-photon microscopy. Cancer cells in the tumor core are remarkably dynamic and exhibit correlated migration patterns, giving rise to local "currents" and large-scale tissue dynamics. Although cells exhibit stop-and-start migration with intermittent pauses, pausing does not appear to be required during division. Use of pharmacological inhibitors indicates that migration patterns in tumors are actively driven by the actin cytoskeleton. Under these conditions, we also observed a relationship between migration speed and correlation length, suggesting that cells in tumors are near a jamming transition. Our study provides new insight into the dynamics of cancer cells in the tumor core, opening new avenues of research in understanding the migratory properties of cancer cells and later metastasis

Active cell migration is critical for steady-state epithelial turnover in the gut

International audienceSteady-state turnover is a hallmark of epithelial tissues throughout adult life. Intestinal epithelial turnover is marked by continuous cell migration, which is assumed to be driven by mitotic pressure from the crypts. However, the balance of forces in renewal remains ill-defined. Combining biophysical modeling and quantitative three-dimensional tissue imaging with genetic and physical manipulations, we revealed the existence of an actin-related protein 2/3 complex-dependent active migratory force, which explains quantitatively the profiles of cell speed, density, and tissue tension along the villi. Cells migrate collectively with minimal rearrangements while displaying dual-apicobasal and front-back-polarity characterized by actin-rich basal protrusions oriented in the direction of migration. We propose that active migration is a critical component of gut epithelial turnover

Active cell migration is critical for steady-state epithelial turnover in the gut

International audienceSteady-state turnover is a hallmark of epithelial tissues throughout adult life. Intestinal epithelial turnover is marked by continuous cell migration, which is assumed to be driven by mitotic pressure from the crypts. However, the balance of forces in renewal remains ill-defined. Combining biophysical modeling and quantitative three-dimensional tissue imaging with genetic and physical manipulations, we revealed the existence of an actin-related protein 2/3 complex-dependent active migratory force, which explains quantitatively the profiles of cell speed, density, and tissue tension along the villi. Cells migrate collectively with minimal rearrangements while displaying dual-apicobasal and front-back-polarity characterized by actin-rich basal protrusions oriented in the direction of migration. We propose that active migration is a critical component of gut epithelial turnover

Active cell migration is critical for steady-state epithelial turnover in the gut

International audienceSteady-state turnover is a hallmark of epithelial tissues throughout adult life. Intestinal epithelial turnover is marked by continuous cell migration, which is assumed to be driven by mitotic pressure from the crypts. However, the balance of forces in renewal remains ill-defined. Combining biophysical modeling and quantitative three-dimensional tissue imaging with genetic and physical manipulations, we revealed the existence of an actin-related protein 2/3 complex-dependent active migratory force, which explains quantitatively the profiles of cell speed, density, and tissue tension along the villi. Cells migrate collectively with minimal rearrangements while displaying dual-apicobasal and front-back-polarity characterized by actin-rich basal protrusions oriented in the direction of migration. We propose that active migration is a critical component of gut epithelial turnover