Influence of postwashing process on the elution of residual monomers, degree of conversion, and mechanical properties of a 3D printed crown and bridge materials.

OBJECTIVES: This study aimed to determine the effects of the postwashing method and time on the mechanical properties and biocompatibility of three-dimensional (3D) printed crown and bridge resin. METHODS: DLP (digital light processing)-printed specimens produced from Nextdent crown & bridge (C&B) resins were washed separately using an ultrasonic bath and rotary washer with TPM (tripropylene glycol monomethyl ether) for 3 min, 6 min, 10 min, 20 min, and 1 h. Postcuring was applied for 30 min to each specimen after the washing process. The flexural strength, Vickers hardness, water sorption and solubility, degree of conversion (DC), elution of residual monomers, and biocompatibility of the specimens were evaluated. RESULTS: The ultrasonic bath showed greater washing efficacy by reducing the residual HEMA (2-hydroxyethyl methacrylate) from 2.0634 ppm to 0.1456 ppm and reducing the residual TEGDMA (triethylene glycol dimethacrylate) from 1.4862 ppm to 0.1484 ppm. With prolonged washing, the flexural strength significantly decreased from 129.67 ± 6.66 MPa (mean±standard deviation) to 103.17 ± 7.20 MPa, while the Vickers hardness increased slightly for the first 6 min and then decreased thereafter significantly. The DC was 87.78 ± 1.34% after 3 min and then gradually decreased with extended washing time. The cytotoxicity significantly decreases with the increment of the washing time. SIGNIFICANCE: The washing effect on the elution of residual monomers was better for an ultrasonic bath than for a rotary washer. Extending the washing time reduces the mechanical properties and cytotoxicity of the Nextdent C&B resin.

Influence of different postcuring parameters on mechanical properties and biocompatibility of 3D printed crown and bridge resin for temporary restorations.

This study analyzed the flexural properties, Vickers hardness, degree of conversion (DC), and cell viability of 3D printed crown and bridge resin postcured using various types of postcuring equipment (PCE). 3D printed specimens were postcured for various times using different types of 3D printing PCE [for 5, 15, and 30 min using LC 3D Print Box (LC), Form Cure (FC), Cure M (CM), and Veltz 3D (VE) devices] and the VALO handheld light-curing (VA) device for 20, 40, and 60 s. Neither the flexural strength (132.27-145.79 MPa) nor the flexural modulus (1.52-1.83 GPa) differed significantly when postcuring for 30 min using the LC, FC, CM, or VE device, or for 20, 40, or 60 s of postcuring using the VA device (p > 0.05). The Vickers hardness was highest after 30 min of postcuring for all groups, and varied significantly with the postcuring time in the LC (p < 0.001) and CM (p < 0.001) groups. DC was significantly higher for the 5-min CM group (84.97 ± 4.02%) than for the GS, 30-min FC, 5-min VE, and 20-s VA groups. Cell viability of the postcured resin specimens was 56.46-92.29%, and varied significantly in the CM and VE groups according to the postcuring time (p < 0.05). Confocal laser scanning microscopy observations showed well-developed cell morphology and numerous cell-cell contacts in all groups except the GS group. This study found that the use of different types of PCE did not significantly affect the flexural properties of 3D printed crown and bridge resin, whereas there were significant variations in DC, Vickers hardness, and cell viability.

Sleeve insert scan body to predict implant placement position by using implant surgical guides: A dental technique.

In studies that assessed the accuracy of implant surgical guides, evaluations were based on the placement position of the implant by using a manufactured surgical guide. However, such assessments could involve errors that may occur during implant placement. Therefore, evaluating the 3-dimensional accuracy of the fabrication of the implant surgical guide itself is not enough. In the evaluation method described in this article, location-related information is obtained by connecting a scan body to the sleeve of the surgical guide instead of directly placing the implant. This helps to evaluate the accuracy of the surgical guide without errors in the placement of an implant.

Integration of adeno-associated virus (AAV) into the genomes of most Thai and Mongolian liver cancer patients does not induce oncogenesis.

BACKGROUND: Engineered versions of adeno-associated virus (AAV) are commonly used in gene therapy but evidence revealing a potential oncogenic role of natural AAV in hepatocellular carcinoma (HCC) has raised concerns. The frequency of potentially oncogenic integrations has been reported in only a few populations. AAV infection and host genome integration in another type of liver cancer, cholangiocarcinoma (CCA), has been studied only in one cohort. All reported oncogenic AAV integrations in HCC come from strains resembling the fully sequenced AAV2 and partly sequenced AAV13. When AAV integration occurs, only a fragment of the AAV genome is detectable in later DNA or RNA sequencing. The integrated fragment is typically from the 3' end of the AAV genome, and this positional bias has been only partly explained. Three research groups searched for evidence of AAV integration in HCC RNAseq samples in the Cancer Genome Atlas (TCGA) but reported conflicting results. RESULTS: We collected and analyzed whole transcriptome and viral capture DNA sequencing in paired tumor and non-tumor samples from two liver cancer Asian cohorts from Thailand (N = 147, 47 HCC and 100 intrahepatic cholangiocarcinoma (iCCA)) and Mongolia (N = 70, all HCC). We found only one HCC patient with a potentially oncogenic integration of AAV, in contrast to higher frequency reported in European patients. There were no oncogenic AAV integrations in iCCA patients. AAV genomic segments are present preferentially in the non-tumor samples of Thai patients. By analyzing the AAV genome positions of oncogenic and non-oncogenic integrated fragments, we found that almost all the putative oncogenic integrations overlap the X gene, which is present and functional only in the strain AAV2 among all fully sequenced strains. This gene content difference could explain why putative oncogenic integrations from other AAV strains have not been reported. We resolved the discrepancies in previous analyses of AAV presence in TCGA HCC samples and extended it to CCA. There are 12 TCGA samples with an AAV segment and none are in Asian patients. AAV segments are present in preferentially in TCGA non-tumor samples, like what we observed in the Thai patients. CONCLUSIONS: Our findings suggest a minimal AAV risk of hepatocarcinogenesis in Asian liver cancer patients. The partial genome presence and positional bias of AAV integrations into the human genome has complicated analysis of possible roles of AAV in liver cancer.

Effects of Postcuring Temperature on the Mechanical Properties and Biocompatibility of Three-Dimensional Printed Dental Resin Material.

Three-dimensional (3D) printing is an attractive technology in dentistry. Acrylic-based 3D printed resin parts have to undergo postcuring processes to enhance their mechanical and biological properties, such as UV-light and thermal polymerization. However, no previous studies have revealed how the postcuring temperature influences the biocompatibility of the produced parts. Therefore, we postprocessed 3D printed denture teeth resin under different postcuring temperatures (40, 60 and 80 °C) for different periods (15, 30, 60, 90 and 120 min), and evaluated their flexural properties, Vickers hardness, cell cytotoxicity, cell viability, and protein adsorption. In addition, confocal laser scanning was used to assess the condition of human gingival fibroblasts. It was found that increasing the postcuring temperature significantly improved the flexural strength and cell viability. The flexural strength and cell viability were 147.48 ± 5.82 MPa (mean ± standard deviation) and 89.51 ± 7.09%, respectively, in the group cured at 80 °C for 120 min, which were higher than the values in the 40 and 60 °C groups. The cell cytotoxicity increased in the 40 °C groups and for longer cultivation time. Confocal laser scanning revealed identifiable differences in the morphology of fibroblasts. This study has confirmed that the postcuring temperature influences the final mechanical and biological properties of 3D printed resin.

Effects of Groove Sealing of the Posterior Occlusal Surface and Offset of the Internal Surface on the Internal Fit and Accuracy of Implant Placements Using 3D-Printed Surgical Guides: An In Vitro Study.

This study evaluated the internal fit and the accuracy of the implant placement position in order to determine how the surface shape of the tooth and the offset influence the accuracy of the surgical guide. The acquired digital data were analyzed in three dimensions using 3D inspection software. The obtained results confirmed that the internal fit was better in the groove sealing (GS) group (164.45 ± 28.34 µm) than the original shape (OS) group (204.07 ± 44.60 µm) (p < 0.001), and for an offset of 100 µm (157.50 ± 17.26 µm) than for offsets of 30 µm (206.48 ± 39.12 µm) and 60 µm (188.82 ± 48.77 µm) (p < 0.001). The accuracy of implant placement was better in the GS than OS group in terms of the entry (OS, 0.229 ± 0.092 mm; GS, 0.169 ± 0.061 mm; p < 0.001), apex (OS, 0.324 ± 0.149 mm; GS, 0.230 ± 0.124 mm; p < 0.001), and depth (OS, 0.041 ± 0.027 mm; GS, 0.025 ± 0.022 mm; p < 0.001). In addition, the entries (30 µm, 0.215 ± 0.044 mm; 60 µm, 0.172 ± 0.049 mm; 100 µm, 0.119 ± 0.050 mm; p < 0.001) were only affected by the amount of offset. These findings indicate that the accuracy of a surgical guide can be improved by directly sealing the groove of the tooth before manufacturing the surgical guide or setting the offset during the design process.

The genomic landscape of Mongolian hepatocellular carcinoma.

Mongolia has the highest incidence of hepatocellular carcinoma (HCC) in the world, but its causative factors and underlying tumor biology remain unknown. Here, we describe molecular characteristics of HCC from 76 Mongolian patients by whole-exome and transcriptome sequencing. We present a comprehensive analysis of mutational signatures, driver genes, and molecular subtypes of Mongolian HCC compared to 373 HCC patients of different races and ethnicities and diverse etiologies. Mongolian HCC consists of prognostic molecular subtypes similar to those found in patients from other areas of Asia, Europe, and North America, as well as other unique subtypes, suggesting the presence of distinct etiologies linked to Mongolian patients. In addition to common driver mutations (TP53, CTNNB1) frequently found in pan-cancer analysis, Mongolian HCC exhibits unique drivers (most notably GTF2IRD2B, PNRC2, and SPTA1), the latter of which is associated with hepatitis D viral infection. These results suggest the existence of new molecular mechanisms at play in Mongolian hepatocarcinogenesis.

Monolithic Zirconia FPD on Modified Titanium Bonding Bases in Limited Interocclusal Distance: A Case Report.

A 62-year-old male patient sought treatment for missing maxillary teeth. A diagnostic cast demonstrated that the interocclusal distance was insufficient. A 5-unit screw-retained implant-supported fixed partial denture (FPD) was used to restore missing maxillary teeth. The restoration of multiple missing teeth using an implant-supported FPD is challenging when the interocclusal distance is limited due to lack of retention and inadequate esthetics. In this case, a hexagonal, screw-retained, and subgingivally located titanium-based zirconia implant-supported FPD with a conical abutment base was used for restoration to overcome the limited interocclusal distance. This implant-supported FPD, consisting of CAD/CAM-designed monolithic zirconia cemented to a titanium bonding base in the laboratory, is expected to facilitate predictable retention and adequate esthetics as well as provide ease of retrieval.

Microglial AGE-albumin is critical for neuronal death in Parkinson's disease: a possible implication for theranostics.

Advanced glycation end products (AGEs) are known to play an important role in the pathogenesis of neurodegenerative diseases, including Parkinson's disease (PD), by inducing protein aggregation and cross-link, formation of Lewy body, and neuronal death. In this study, we observed that AGE-albumin, the most abundant AGE product in the human PD brain, is synthesized in activated microglial cells and accumulates in the extracellular space. AGE-albumin synthesis in human-activated microglial cells is distinctly inhibited by ascorbic acid and cytochalasin treatment. Accumulated AGE-albumin upregulates the receptor to AGE, leading to apoptosis of human primary dopamine (DA) neurons. In animal experiments, we observed reduced DA neuronal cell death by treatment with soluble receptor to AGE. Our study provides evidence that activated microglial cells are one of the main contributors in AGE-albumin accumulation, deleterious to DA neurons in human and animal PD brains. Finally, activated microglial AGE-albumin could be used as a diagnostic and therapeutic biomarker with high sensitivity for neurodegenerative disorders, including PD.

Microglial AGE-albumin is critical in promoting alcohol-induced neurodegeneration in rats and humans.

Alcohol is a neurotoxic agent, since long-term heavy ingestion of alcohol can cause various neural diseases including fetal alcohol syndrome, cerebellar degeneracy and alcoholic dementia. However, the molecular mechanisms of alcohol-induced neurotoxicity are still poorly understood despite numerous studies. Thus, we hypothesized that activated microglial cells with elevated AGE-albumin levels play an important role in promoting alcohol-induced neurodegeneration. Our results revealed that microglial activation and neuronal damage were found in the hippocampus and entorhinal cortex following alcohol treatment in a rat model. Increased AGE-albumin synthesis and secretion were also observed in activated microglial cells after alcohol exposure. The expressed levels of receptor for AGE (RAGE)-positive neurons and RAGE-dependent neuronal death were markedly elevated by AGE-albumin through the mitogen activated protein kinase pathway. Treatment with soluble RAGE or AGE inhibitors significantly diminished neuronal damage in the animal model. Furthermore, the levels of activated microglial cells, AGE-albumin and neuronal loss were significantly elevated in human brains from alcoholic indivisuals compared to normal controls. Taken together, our data suggest that increased AGE-albumin from activated microglial cells induces neuronal death, and that efficient regulation of its synthesis and secretion is a therapeutic target for preventing alcohol-induced neurodegeneration.

Heat shock instructs hESCs to exit from the self-renewal program through negative regulation of OCT4 by SAPK/JNK and HSF1 pathway.

Environmental factors affect self-renewal of stem cells by modulating the components of self-renewal networks. Heat shock, an environmental factor, induces heat shock factors (HSFs), which up-regulate stress response-related genes. However, the link of heat shock to self-renewal of stem cells has not been elucidated yet. Here, we present the direct link of heat shock to a core stem cell regulator, OCT4, in the self-renewal network through SAPK/JNK and HSF1 pathway. We first showed that heat shock initiated differentiation of human embryonic stem cells (hESCs). Gene expression analysis revealed that heat shock increased the expression of many genes involved in cellular processes related to differentiation of stem cells. We then examined the effects of HSFs induced by heat shock on core self-renewal factors. Among HSFs, heat shock induced mainly HSF1 in hESCs. The HSF1 repressed the expression of OCT4, leading to the differentiation of hESCs and the above differentiation-related gene expression change. We further examined the effects of the upstream MAP (mitogen-activated protein) kinases of HSF1 on the repression of OCT4 expression by HSF1. Among the MAP kinases, SAPK/JNK controlled predominantly the repression of the OCT4 expression by HSF1. The direct link of heat shock to the core self-renewal regulator through SAPK/JNK and HSF1 provides a fundamental basis for understanding the effect of heat and other stresses involving activation of HSF1 on the self-renewal program and further controlling differentiation of hESCs in a broad spectrum of stem cell applications using these stresses.

Proteome-wide discovery of mislocated proteins in cancer.

Several studies have sought systematically to identify protein subcellular locations, but an even larger task is to map which of these proteins conditionally relocates in disease (the mislocalizome). Here, we report an integrative computational framework for mapping conditional location and mislocation of proteins on a proteome-wide scale, called a conditional location predictor (CoLP). Using CoLP, we mapped the locations of over 10,000 proteins in normal human brain and in glioma. The prediction showed 0.9 accuracy using 100 location tests of 20 randomly selected proteins. Of the 10,000 proteins, over 150 have a strong likelihood of mislocation under glioma, which is striking considering that few mislocation events have been identified in this disease previously. Using immunofluorescence and Western blotting in both primary cells and tissues, we successfully experimentally confirmed 15 mislocations. The most common type of mislocation occurs between the endoplasmic reticulum and the nucleus; for example, for RNF138, TLX3, and NFRKB. In particular, we found that the gene for the mislocating protein GFRA4 had a nonsynonymous point mutation in exon 2. Moreover, redirection of GFRA4 to its normal location, the plasma membrane, led to marked reductions in phospho-STAT3 and proliferation of glioma cells. This framework has the potential to track changes in protein location in many human diseases.

Quantitative proteomic analysis reveals that lipopolysaccharide induces mitogen-activated protein kinase-dependent activation in human microglial cells.

Microglial cells act as the first and main form of active immune defense in the central nervous system related to inflammation and neurodegenerative disease. Lipopolysaccharide (LPS) induces many genes encoding inflammatory mediators, including cytokines such as tumor necrosis factor-α, interleukin-1ß, (IL-1ß), and IL-6, chemokines, and prostaglandins in microglial cells. Quantitative proteomics methods with isobaric chemical labeling using tandem mass tags and 2D-nano LC-ESI-MS/MS were used to systematically analyze proteomic changes in microglia responding to LPS stimulation. As a result, we found that the expression level of 21 proteins in human microglial cells changed after activation. Among those, one of the strong mitogen-activated protein kinase (MAPK) regulator proteins, CMPK1 was highly upregulated after LPS stimulation in human microglial cells. We detected and validated upregulation of MAPK including ERK1/2, p38, and SAPK/JNK by immunohistochemistry and Western blotting. NFκB, strong transcription factor of CMPK1, was translocated to the nucleus from the cytosol by high contents screening after LPS stimulation. Taken together, we conclude that MAPK signaling plays an important role in LPS-induced human microglial activation related to inflammatory response.