Enhanced bioprocess control to advance the manufacture of mesenchymal stromal cell-derived extracellular vesicles in stirred-tank bioreactors.

Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) act as signaling mediators of cellular responses. However, despite representing a promising alternative to cell-based therapies, clinical translation of EVs is currently limited by their lack of scalability and standardized bioprocessing. Herein, we integrated scalable downstream processing protocols with standardized expansion of large numbers of viable cells in stirred-tank bioreactors to improve EV production. Higher EV yields were linked to EV isolation by tangential flow filtration followed by size exclusion chromatography, rendering 5 times higher number of EVs comparatively to density gradient ultracentrifugation protocols. Additionally, when compared to static culture, EV manufacture in bioreactors resulted in 2.2 higher yields. Highlighting the role of operating under optimal cell culture conditions to maximize the number of EVs secreted per cell, MSCs cultured at lower glucose concentration favored EV secretion. While offline measurements of metabolites concentration can be performed, in this work, Raman spectroscopy was also applied to continuously track glucose levels in stirred-tank bioreactors, contributing to streamline the selection of optimal EV collection timepoints. Importantly, MSC-derived EVs retained their quality attributes and were able to stimulate angiogenesis in vitro, therefore highlighting their promising therapeutic potential.

Retreatability of two endodontic sealers, EndoSequence BC Sealer and AH Plus: a micro-computed tomographic comparison.

OBJECTIVES: Recently, bioceramic sealers like EndoSequence BC Sealer (BC Sealer) have been introduced and are being used in endodontic practice. However, this sealer has limited research related to its retreatability. Hence, the aim of this study was to evaluate the retreatability of two sealers, BC Sealer as compared with AH Plus using micro-computed tomographic (micro-CT) analysis. MATERIALS AND METHODS: Fifty-six extracted human maxillary incisors were instrumented and randomly divided into 4 groups of 14 teeth: 1A, gutta-percha, AH Plus retreated with chloroform; 1B, gutta-percha, AH Plus retreated without chloroform; 2A, gutta-percha, EndoSequence BC Sealer retreated with chloroform; 2B, gutta-percha, EndoSequence BC Sealer retreated without chloroform. Micro-CT scans were taken before and after obturation and retreatment and analyzed for the volume of residual material. The specimens were longitudinally sectioned and digitized images were taken with the dental operating microscope. Data was analyzed using an ANOVA and a post-hoc Tukey test. Fisher exact tests were performed to analyze the ability to regain patency. RESULTS: There was significantly less residual root canal filling material in the AH Plus groups retreated with chloroform as compared to the others. The BC Sealer samples retreated with chloroform had better results than those retreated without chloroform. Furthermore, patency could be re-established in only 14% of teeth in the BC Sealer without chloroform group. CONCLUSION: The results of this study demonstrate that the BC Sealer group had significantly more residual filling material than the AH Plus group regardless of whether or not both sealers were retreated with chloroform.

Effect of apical size and taper on volume of irrigant delivered at working length with apical negative pressure at different root curvatures.

INTRODUCTION: The purpose of this investigation was to determine the effect that apical preparation size and preparation taper have on the volume of irrigant delivered at the working length for different canal curvatures using apical negative pressure irrigation. METHODS: One hundred fifty-five human teeth (55 maxillary canines and 100 mandibular molars) were used in this study. Root canals were prepared with rotary instruments to a size 35.04 and separated into 3 experimental groups according to their degree of curvature: group A (n = 50) included canal curvatures ranging from 0° to 10°, group B (n = 50) from 11° to 30°, and group C (n = 50) from 31° to 65°. Samples of each curvature group were further randomized to experimental subgroups according to the apical size and taper as follow: 35.06, 40.04, 40.06, 45.04, and 45.06. The apical third was irrigated using a microcannula and the volume of NaOCl suctioned at the working length under negative pressure was measured over a period of 30 seconds using a fluid recovery trap. Positive controls consisted of measuring the maximum volume of 5.25% NaOCl capable of being suctioned by the microcannula from an open glass vial over 30 seconds. Negative control was the volume of irrigant aspirated by the microcannula with a preparation size of 25.04 over 30 seconds. RESULTS: The volume of irrigant was significantly greater when the apical preparation size increased from 35.06 to 40.04. As the apical preparation taper increased further from 40.04 to 40.06, the volume of irrigant significantly improved in group B, but it was not significant in group A. Apical preparation sizes greater than 40.06 did not show an increase of the volume of irrigant aspirated. CONCLUSIONS: The degree of root canal curvature decreased the volume of irrigant at the working length for a given apical size and taper. An apical preparation of 40.06 significantly increased the volume and exchange of irrigant at the working length regardless of curvature.