The Efficacy of Commercial Tooth Storage Media for Maintaining the Viability of Human Periodontal Ligament Fibroblasts

Aim To evaluate Save‐A‐Tooth (SAT), EMT Toothsaver (EMT) and Hank\u27s Balanced Salt Solution (HBSS) for their influence on the viability and proliferative capacity of human periodontal ligament fibroblasts (HPDLFs). Methodology Primary HPDLFs were seeded into 96‐well cell culture plates and exposed to SAT, EMT, HBSS and water (negative control) for 0.5, 1, 3, 6, 12 and 24 h at room temperature (22 °C). After each exposure time, cell viability was measured through quantifying adenosine triphosphate (ATP) using a luminescent dye. The proliferative capacity was also quantified using the PrestoBlue assay after 12 or 24 h storage in each medium. The data were analysed statistically by two‐way anova and post hoc Least Significant Difference (LSD) test (P \u3c 0.05). The morphology of the cells after 12 h storage was also investigated through live/dead viability/cytotoxicity kit together with fluorescence microscopy. Results There was no significant difference in cell viability amongst HBSS, SAT and EMT groups up to 6 h. SAT was effective in maintaining cell viability only up to 12 h and then became detrimental to HPDLF; after 24 h, the effectiveness of SAT in maintaining cell viability was similar to that of water (P \u3e 0.05). Amongst all the media, only EMT could maintain the proliferative capacity of HPDLFs significantly higher than the negative control, that is water (P \u3c 0.05) after 24 h storage. Conclusion EMT maintained the proliferative capacity of HPDLFs after 24 h storage

Multistep, sequential control of the trafficking and function of the multiple sulfatase deficiency gene product, SUMF1 by PDI, ERGIC-53 and ERp44.

Sulfatase modifying factor 1 (SUMF1) encodes for the formylglicine generating enzyme, which activates sulfatases by modifying a key cysteine residue within their catalytic domains. SUMF1 is mutated in patients affected by multiple sulfatase deficiency, a rare recessive disorder in which all sulfatase activities are impaired. Despite the absence of canonical retention/retrieval signals, SUMF1 is largely retained in the endoplasmic reticulum (ER), where it exerts its enzymatic activity on nascent sulfatases. Part of SUMF1 is secreted and paracrinally taken up by distant cells. Here we show that SUMF1 interacts with protein disulfide isomerase (PDI) and ERp44, two thioredoxin family members residing in the early secretory pathway, and with ERGIC-53, a lectin that shuttles between the ER and the Golgi. Functional assays reveal that these interactions are crucial for controlling SUMF1 traffic and function. PDI couples SUMF1 retention and activation in the ER. ERGIC-53 and ERp44 act downstream, favoring SUMF1 export from and retrieval to the ER, respectively. Silencing ERGIC-53 causes proteasomal degradation of SUMF1, while down-regulating ERp44 promotes its secretion. When over-expressed, each of three interactors favors intracellular accumulation. Our results reveal a multistep control of SUMF1 trafficking, with sequential interactions dynamically determining ER localization, activity and secretion

"Wink to grasp" – comparing eye, voice & EMG gesture control of grasp with soft-robotic gloves

The ability of robotic rehabilitation devices to support paralysed end-users is ultimately limited by the degree to which human-machine-interaction is designed to be effective and efficient in translating user intention into robotic action. Specifically, we evaluate the novel possibility of binocular eye-tracking technology to detect voluntary winks from involuntary blink commands, to establish winks as a novel low-latency control signal to trigger robotic action. By wearing binocular eye-tracking glasses we enable users to directly observe their environment or the actuator and trigger movement actions, without having to interact with a visual display unit or user interface. We compare our novel approach to two conventional approaches for controlling robotic devices based on electromyo-graphy (EMG) and speech-based human-computer interaction technology. We present an integrated software framework based on ROS that allows transparent integration of these multiple modalities with a robotic system. We use a soft-robotic SEM glove (Bioservo Technologies AB, Sweden) to evaluate how the 3 modalities support the performance and subjective experience of the end-user when movement assisted. All 3 modalities are evaluated in streaming, closed-loop control operation for grasping physical objects. We find that wink control shows the lowest error rate mean with lowest standard deviation of (0.23 ± 0.07, mean ± SEM) followed by speech control (0.35 ± 0. 13) and EMG gesture control (using the Myo armband by Thalamic Labs), with the highest mean and standard deviation (0.46 ± 0.16). We conclude that with our novel own developed eye-tracking based approach to control assistive technologies is a well suited alternative to conventional approaches, especially when combined with 3D eye-tracking based robotic end-point control

MULTIPLE PLURIPOTENT STEM CELL MARKERS IN HUMAN ANAPLASTIC THYROID CANCER: THE PUTATIVE UPSTREAM ROLE OF SOX-2

Background: Anaplastic Thyroid Carcinoma (ATC) is a rare and aggressive endocrine tumor, with highly undifferentiated morphology. It has been suggested that cancer stem cells (CSCs) might play a central role in ATC. The objectives of this study were the following: 1) to characterize CSCs from ex vivo ATC specimens by investigating the expression of several pluripotent stem cell markers; 2) to evaluate in vitro drug resistance modifications after specific CSC transcription factor switch off. Methods: Ex vivo: eight formalin-fixed, paraffin-embedded ATC specimens were analyzed by RT and qRT-PCR and immunohistochemistry. In vitro: in ATC SW1736 cells the expression levels of OCT-4, NANOG and ABCG2 and the sensitivity to either cisplatin or doxorubicin were evaluated after silencing. Results: OCT-4, KLF4 and SOX2 transcription factors and C-KIT and THY-1 stem surface antigens showed variable up-regulation in all ATC cases. The SW1736 cell line was characterized by a high percentage of stem population (10.4 \ub1 2.1 % of cells were aldehyde dehydrogenase positive) and a high expression of several CSC markers (SOX2, OCT4, NANOG, C-MYC, SSEA4). SOX2 silencing down-regulated OCT-4, NANOG and ABCG2. SOX2 silencing sensitized SW1736 cells, causing a significant cell death increase (1.8 fold) in comparison to control cells with 10 \ub5M cisplatin (93.9\ub13.4% vs. 52.6\ub19.4%, p<0.01) and 2.7 fold with 0.5\ub5M doxorubicin (45.8\ub19.9% vs. 17.1\ub13.4% p<0.01). ABCG2 silencing caused increased cell death with both cisplatin (74.9\ub11.4%) and doxorubicin treatment (74.1\ub10.1%) vs. no-target-treated cells (respectively, 45.8\ub11.0% and 48.6\ub11.0%, p<0.001). Conclusions: The characterization of CSCs in ATC through the analysis of multiple pluripotent stem cell markers might be useful in identifying cells with a stem-like phenotype capable of resisting conventional chemotherapy. In addition, our data demonstrate that SOX2 switch-off through ABCG2 transporter down-regulation has a major role in overcoming CSC chemotherapy resistance

Cellular models and assays to study NLRP3 inflammasome biology

The NLRP3 inflammasome is a multi-protein complex that initiates innate immunity responses when exposed to a wide range of stimuli, including pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs). Inflammasome activation leads to the release of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 and to pyroptotic cell death. Over-activation of NLRP3 inflammasome has been associated with several chronic inflammatory diseases. A deep knowledge of NLRP3 inflammasome biology is required to better exploit its potential as therapeutic target and for the development of new selective drugs. To this purpose, in the past few years, several tools have been developed for the biological characterization of the multimeric inflammasome complex, the identification of the upstream signaling cascade leading to inflammasome activation, and the downstream effects triggered by NLRP3 activation. In this review, we will report cellular models and cellular, biochemical, and biophysical assays that are currently available for studying inflammasome biology. A special focus will be on those models/assays that have been used to identify NLRP3 inhibitors and their mechanism of action

BRSGO 7460RR: cultivar de soja transgênica.

bitstream/item/57492/1/BRSGO7460RR.pdf1 folder