OBJECTIVE: The study aimed to assess the microleakage of nanoparticle-based (NPB) cyanoacrylate sealer and epoxy resin-based (ERB) sealer using radioisotope method and confocal laser scanning microscopy (CLSM).
METHODS: A total of 100 single-rooted teeth were collected; specimens were accessed, instrumented, and irrigated, and randomly distributed into 4 groups of 25 samples each: Group I: Positive control, group II: Negative control, group III: Obturated with NPB sealer, and group IV: Obturated with ERB sealer. All samples were immersed in 99mTc pertechnetate solution, for 3 hours after which radioactivity was estimated under a Gamma camera. The radioactivity released by specimens before and after nail varnish removal was statistically analyzed. After 2 weeks, the same samples were used for CLSM analysis. The sealer tubular penetration depth was measured at the deepest level for each group using ZEN lite 2012. Data collected was statistically evaluated.
RESULTS: The amount of radioactivity observed at first in group III and group IV was 194.76 and 599.12 units, respectively, with p-value < 0.001, indicating significant interaction, and after nail varnish removal, it was 89.68 and 468.44 units, respectively, with a p-value < 0.001; again, indicating statistical significance. Hence, the radioactivity of NPB sealer was found to be lower than ERB sealer in both cases, indicating better sealing of the former. The photomicrographs show that mean value of dye penetration in NPB sealer in first, second, and third segment from apex was 85.06, 75.73, and 66.09, respectively; while in the case of ERB sealer, those were 597.28, 461.17, and 195.68, respectively; with p-value < 0.001; signifying that NPB sealer exhibited higher resistance to microleakage than ERB sealer.
CONCLUSIONS: The NPB sealer can become a potential root canal sealer in future endodontics due to superior physiochemical properties attributed to the cyanoacrylate and incorporated nanoparticles.
CONCLUSIONS: The study clinically signifies that we can equally use the radioisotopic method along with confocal method while conducting the microleakage studies. In addition, NPB sealer can be an emerging replacement with better properties than gold standard root canal sealers for clinical use. How to cite this article: Shetty C, Qaiser S, Shetty A, et al. Evaluation of Microleakage of Nanoparticle-incorporated Cyanoacrylate Root Canal Sealer Using the Radioisotopic Method: An In Vitro Study. J Contemp Dent Pract 2024;25(4):335-341.

Interleukin-17A therapeutic inhibitors are among the most effective treatment methods for moderate-to-severe plaque psoriasis (PP). Reflectance confocal microscopy is a non-invasive imaging technique already documented to be beneficial in evaluating the follow-up of PP under treatment with topical actives and phototherapy. This study aimed to assess the epidermal and dermal changes associated with psoriasis and its treatment with RCM during systemic secukinumab treatment in patients with moderate-to-severe PP. A pilot study was conducted to evaluate RCM as a non-invasive tool for monitoring secukinumab treatment in patients with PP. For patients receiving secukinumab treatment, lesional skin was selected for RCM imaging, which were recorded at all scheduled times. The RCM evaluation criteria were established based on the histopathological diagnostic criteria for psoriasis. The clinical severity of psoriasis was assessed utilizing the psoriasis area severity index. A total of 23 patients with PP were included in the study. Each patient received 300 mg of subcutaneous secukinumab as induction therapy at baseline and weeks 1-4, followed by maintenance therapy every four weeks. Microscopic confocal changes were observed during the treatment. The results identified early microscopic evidence of the anti-inflammatory activity of secukinumab, which was not detected during the clinical examination. RCM findings correlating with the PASI were used to observe the patient\'s response to treatment and were identified as follows: acanthosis and parakeratosis, presence of epidermal and dermal inflammatory cells, presence of non-edge dermal papillae, and vascularization in the papillary dermis. This study is the first to demonstrate the use of RCM as an effective tool for non-invasive monitoring of secukinumab therapeutic response at a cellular level in a clinical or research setting. Early detection of RCM parameters associated with secukinumab activity may facilitate the identification of an early treatment response. RCM appears to be capable of providing practical and helpful information regarding follow-up in patients with PP undergoing secukinumab treatment. RCM may also provide novel perspectives on the subclinical evaluation of PP\'s response to biological therapy.

Assessing programmed death ligand 1 (PD-L1) expression through immunohistochemistry (IHC) is the golden standard in predicting immunotherapy response of non-small cell lung cancer (NSCLC). However, observation of heterogeneous PD-L1 distribution in tumor space is a challenge using IHC only. Meanwhile, immunofluorescence (IF) could support both planar and three-dimensional (3D) histological analyses by combining tissue optical clearing with confocal microscopy. We optimized clinical tissue preparation for the IF assay focusing on staining, imaging, and post-processing to achieve quality identical to traditional IHC assay. To overcome limited dynamic range of the fluorescence microscope\'s detection system, we incorporated a high dynamic range (HDR) algorithm to restore the post imaging IF expression pattern and further 3D IF images. Following HDR processing, a noticeable improvement in the accuracy of diagnosis (85.7%) was achieved using IF images by pathologists. Moreover, 3D IF images revealed a 25% change in tumor proportion score for PD-L1 expression at various depths within tumors. We have established an optimal and reproducible process for PD-L1 IF images in NSCLC, yielding high quality data comparable to traditional IHC assays. The ability to discern accurate spatial PD-L1 distribution through 3D pathology analysis could provide more precise evaluation and prediction for immunotherapy targeting advanced NSCLC.

Activation of G protein-coupled receptors upon chemoattractant stimulation induces activation of multiple signaling pathways. To fully understand how these signaling pathway coordinates to achieve directional migration of neutrophils, it is essential to determine the dynamics of the spatiotemporal activation profile of signaling components at the level of single living cells. Here, we describe a detailed methodology for monitoring and quantitatively analyzing the spatiotemporal dynamics of 1,4,5-inositol trisphosphate (IP3) in neutrophil-like HL60 cells in response to various chemoattractant fields by applying Förster resonance energy transfer (FRET) fluorescence microscopy.

Autophagy is an intracellular clearance and recycling pathway that delivers different types of cargos to lysosomes for degradation. In recent years, autophagy has attracted considerable medical interest, and many different techniques are being developed to study this process in experimental models such as Dictyostelium. Here we describe the use of different autophagic markers in confocal microscopy, in vivo and also in fixed cells. In particular, we describe the use of the GFP-Atg8-RFP-Atg8ΔG marker and the optimization of the GFP-PgkA cleavage assay to detect small differences in autophagy flux.

BACKGROUND: Merkel cell carcinoma (MCC) is a rare, aggressive, cutaneous tumour with high mortality and frequently delayed diagnosis. Clinically, it often manifests as a rapidly growing erythematous to purple nodule usually located on the lower extremities or face and scalp of elderly patients. There is limited available data on the dermoscopic findings of MCC, and there are no specific features that can be used to definitively diagnose MCC.
OBJECTIVE: Here, we aimed to summarize existing published literature on dermatoscopic and reflectance confocal microscopy (RCM) features of MCC.
METHODS: To find relevant studies, we searched the PubMed and Scopus databases from inception to April 12, 2023. Our goal was to identify all pertinent research that had been written in English. The following search strategy was employed: (\" dermoscopy\" OR \" dermatoscopy\" OR \" videodermoscopy\" OR \" videodermatoscopy\" OR \" reflectance confocal microscopy\") AND \" Merkel cell carcinoma\". Two dermatologists, DK and GE, evaluated the titles and abstracts separately for eligibility. For inclusion, only works written in English were taken into account.
RESULTS: In total 16 articles were retrieved (68 cases). The main dermoscopic findings of MCC are a polymorphous vascular pattern including linear irregular, arborizing, glomerular, and dotted vessels on a milky red background, with shiny or non-shiny white areas. Pigmentation was lacking in all cases. The RCM images showed a thin and disarranged epidermis, and small hypo-reflective cells that resembled lymphocytes arranged in solid aggregates outlined by fibrous tissue in the dermis. Additionally, there were larger polymorphic hyper-reflective cells that likely represented highly proliferative cells.
CONCLUSIONS: Dermoscopic findings of MCC may play a valuable role in evaluating MCC, aiding in the early detection and differentiation from other skin lesions. Further prospective case-control studies are needed to validate these results.

In cereal products, the use of flour containing clusters of intact cells has been indicated as a potential strategy to decrease starch digestion. Rye possesses more uniform and thicker cell walls than wheat but its protective effect against starch digestion has not been elucidated. In this study, rye flours with three different particle sizes, large (LF) (∼1700 μm), medium (MF) (∼1200 μm), and small (SF) (∼350 μm), were used to produce model bread. The textural properties of these breads were analysed using Textural Profile Analysis (TPA). The starch digestibility of both the flour and the bread was measured using Englyst\'s method, while the presence of intact cell clusters was examined using Confocal Laser Scanning Microscopy (CLSM). Additionally, the disintegration of bread digesta during simulated digestion was assessed through image analysis. CLSM micrographs revealed that bread made with MF and LF retained clusters of intact cells after processing, whereas bread made with SF showed damaged cell walls. Starch digestibility in LF and MF was lower (p ≤ 0.05) than that in SF. Bread produced with MF and LF exhibited the least (p ≤ 0.05) cohesive and resilient texture, disintegrated more during digestion, and exhibited higher starch digestibility (p ≤ 0.05) than bread made with SF. These results highlight the central role of bread texture on in vitro starch digestibility.

Corneal nerves and dendritic cells are increasingly being visualised to serve as clinical parameters in the diagnosis of ocular surface diseases using intravital confocal microscopy. In this review, different methods of image analysis are presented. The use of deep learning algorithms, which enable automated pattern recognition, is explained in detail using our own developments and compared with other established methods.
Hornhautnerven und dendritische Zellen werden zunehmend bei der Diagnostik von Erkrankungen der Augenoberfläche als klinische Parameter mittels intravitaler Konfokalmikroskopie dargestellt. In dieser Übersichtsarbeit werden unterschiedliche Verfahren der Bildauswertung dargestellt. Die Verwendung von Deep-Learning-Algorithmen, die eine automatisierte Mustererkennung ermöglichen, wird anhand eigener Entwicklungen detailliert erläutert und mit anderen etablierten Verfahren verglichen.

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This study aimed to evaluate the antimicrobial activity of calcium hypochlorite (Ca (OCl)2) and sodium hypochlorite (NaOCl) using confocal laser scanning microscopy (CLSM) and dentin organic matrix alteration by picrosirius staining and light microscopy (LM). Samples of human extracted teeth were infected with Enterococcus faecalis by centrifugation of the bacterial suspension and were treated with Ca(OCl)2 or NaOCl at 0.5%, 2.5%, and 6% for 15, 30, and 60 seconds. CLSM and viability staining were used to quantitatively analyze the proportions of dead/live bacteria in the canal lumen and border of the root canal. The data were analyzed by ANOVA and Fisher test. For LM analysis, one hundred bovine teeth were randomly divided into 10 test groups (n=10): G1- Without treatment; G2- 17% EDTA; G3- 6% NaOCl; G4- 6% NaOCl + EDTA; G5- 0.5% Ca(OCl)2; G6- 0.5% Ca(OCl)2 + EDTA; G7- 2.5% Ca(OCl)2; G8- 2.5% Ca(OCl)2 + EDTA; G9- 6% Ca(OCl)2; G10- 6% Ca(OCl)2 + EDTA. The samples were fragmented and stained with Picrosirius. Data were analyzed by Kruskal-Wallis and Dunn (P<0.05). There was a strong correlation between the results of the canal lumen and the border of the root canal (r=0.962). Both hypochlorites at a concentration of 0.5% showed less microbial reduction compared to 2.5% and 6% (P<0.05). There was less antimicrobial activity at 15 seconds compared to 30 and 60 seconds (P<0.05). Ca(OCl)2 and NaOCl showed similar results at the same concentrations (P>0.05). In conclusion, Ca(OCl)2 caused fewer alterations to the dentin organic matrix at concentrations of 0.5% and 2.5%. Ca(OCl)2 presents antimicrobial activity similar to NaOCl, and collagen damage is concentration-dependent.