Cell Plate Research Articles

Mobility of the syntaxin PEN1 in Arabidopsis reflects functional specialization of the conserved SYP12 clade

ABSTRACT Plant innate immunity toward cell-wall penetrating filamentous pathogens relies on the conserved SYP12 clade of secretory syntaxins. In Arabidopsis, the two closely related SYP12 clade members, PEN1 and SYP122, play an overlapping role in this general immunity, which can be complemented by two SYP12 clade members from Marchantia (MpSYP12A and MpSYP12B). However, in addition to the conserved SYP12 clade function, PEN1 alone mediates pre-invasive immunity toward powdery mildew fungi, which likely reflects a specialization of its functionality. Here, we show that the PEN1-specific specialization in immunity correlates with a continuous BFA-sensitive recycling and the ability to accumulate strongly at the growing cell plate. This contrasts with the behavior of SYP122, MpSYP12A, and MpSYP12B, all being more stable at the plasma membrane. We suggest that the highly mobile SYP12 specialization observed for PEN1 is required for a fast pre-invasive immune response to resist attack from powdery mildew fungi.

Dihydrotanshinone I Enhances Cell Adhesion and Inhibits Cell Migration in Osteosarcoma U-2 OS Cells through CD44 and Chemokine Signaling.

In the screening of novel natural products against cancer using an in vitro cancer cell model, we recently found that tanshinones from a traditional Chinese medicine, the rhizome of Salvia miltiorrhiza Bunge (Danshen), had potent effects on cell proliferation and migration. Especially for human osteosarcoma U−2 OS cells, tanshinones significantly enhanced the cell adherence, implying a possible role in cell adhesion and cell migration inhibition. In this work, therefore, we aimed to provide a new insight into the possible molecule mechanisms of dihydrotanshinone I, which had the strongest effects on cell adhesion among several candidate tanshinones. RNA−sequencing-based transcriptome analysis and several biochemical experiments indicated that there were comprehensive signals involved in dihydrotanshinone I-treated U−2 OS cells, such as cell cycle, DNA replication, thermogenesis, tight junction, oxidative phosphorylation, adherens junction, and focal adhesion. First, dihydrotanshinone I could potently inhibit cell proliferation and induce cell cycle arrest in the G0/G1 phase by downregulating the expression of CDK4, CDK2, cyclin D1, and cyclin E1 and upregulating the expression of p21. Second, it could significantly enhance cell adhesion on cell plates and inhibit cell migration, involving the hyaluronan CD44−mediated CXCL8–PI3K/AKT–FOXO1, IL6–STAT3–P53, and EMT signaling pathways. Thus, the increased expression of CD44 and lengthened protrusions around the cell yielded a significant increase in cell adhesion. In summary, these results suggest that dihydrotanshinone I might be an interesting molecular therapy for enhancing human osteosarcoma U−2 OS cell adhesion and inhibiting cell migration and proliferation.

Effect of berberine on cancer cell motility in glioma, lung cancer, and prostate cancer cell cultures.

e15079 Background: Initially, berberine was used as an antimicrobial agent in traditional medicine, but its anticancer properties were later discovered. In addition to its cytotoxic effect, berberine also has the ability to inhibit cell motility, which has been demonstrated in some permanent cancer cell lines. The objective of this study was to assess berberine anti-migratory activity in permanent cell cultures compared to primary cell cultures which are generally thought to better reflect tumor characteristics. Methods: H1299 lung cancer, PC3 prostate cancer, and T98G glioma cells, as well as primary cell cultures of the corresponding cancer obtained in our laboratory, were planted in an amount of 15*104 cells per well of a 24-well plate (Biofil, China) in DMEM medium (Gibco, USA) supplemented with 10% FBS (HyClone, USA). After cell adhesion berberine was added in concentration 5 µM, and a wound healing assay was performed according to the standard procedure. Cell plates were continuously incubated and photographed in Lionheart FX imager (BioTek, USA) at 37°C and 5.0% CO2. The extent of cell migration was measured as the percentage reduction in wound area after 48 hours of incubation relative to baseline. Data are presented as Mean ± 95% confidence interval (n = 12). Results: The use of berberine at a concentration of 5 µM led to a significant decrease in cell motility in permanent cultures of lung cancer, glioma and prostate cancer. Namely, the reduction in the wound area after 48 hours of incubation with bereberine was 74.52±12.3% (compared with 94.56±6.2% in the control) in H1299 cell culture, 38.22±10.6% (compared with 83.89±15.5% in the control) in T98G cell culture, and 48.6±7.5% (compared with 69.56±8.1% in the control) in PC3 cell culture. The resulting difference between the control and experimental groups in permanent cell cultures was statistically significant at a significance level of 5% (df = 22). At the same time, the values of wound area reduction for primary cultures of the same cancers did not differ significantly in the control and under the influence of 5 μM berberine at the accepted level of significance. Namely, the reduction in the wound area after 48 hours of incubation with bereberine was 84.79±11.2% (compared with 81.47±15.3% in the control) in lung cancer primary cell culture, 94.64±5.1% (compared with 91.73±6.8% in the control) in glioma primary culture, and 62.63±5.8% (compared with 61.1±8.9% in the control) in prostate cancer primary cell culture. Conclusions: Permanent cell lines are more sensitive to berberine anti-migratory activity than primary cancer cell lines of the same localization.

Fuzheng Kang' ai decoction inhibits cell proliferation, migration and invasion by modulating mir-21-5p/human phosphatase and tensin homology deleted on chromosome ten in lung cancer cells.

To elucidate the potential molecular mechanism by which Fuzheng Kang'ai decoction (, FZKA) inhibits proliferation, migration, and invasion of lung cancer cells. Varying FZKA concentrations were used to manage lung cancer cells (A549 and PC9). We employed: cell counting kit-8 (CCK-8) and plate clone for-mation assays to examine the cell viability; flow cytometry (FCM) to analyze the cycle arrest; transwell and wound-healing assays to assess the cell invasion and migration, respectively. Further, a quantitative real-time polymerase chain reaction (qRT-PCR) assay was adopted to evaluate the miR-21-5p expression. For protein expression analysis, we employed the Western blot technique. Recombinant miR-21-5p overexpression adenovirus vector harboring GFP was constructed and transfected into A549 and PC9, after which we explored the effect of FZKA on miR-21-5p overexpression. Notably, treatment with FZKA inhibited viability, clone-formation ability, invasion, and migration of lung cancer cells. Mechanistically, FZKA markedly suppressed miR-21-5p expression but elevated the human phosphatase and tensin homology deleted on chromosome ten (PTEN) protein level in both A549 and PC9 cells. Over-expression of miR-21-5p lowered PTEN protein expression. Besides, overexpressed miR-21-5p levels with adenovirus antagonized FZKA-upregulated PTEN protein expression. The present study demonstrates how FZKA modulates cell biological behaviors, for instance, it impedes the proliferation by upregulating PTEN expression with miR-21-5p as the target. These findings unveil the potential novel molecular mechanisms from the microRNA aspect by which FZKA suppresses the growth of human lung cancer cells.

Connective tissue growth factor expression maintains the epithelial phenotype of ovarian cancer in early epithelial to mesenchymal transition.

e17544 Background: Epithelial to Mesenchymal Transition (EMT) is essential to the process of metastasis in ovarian cancer (OC). Early expressions of proteins and pathways during EMT essential for understanding mechanism and potential therapeutic targets in OC. The aim of our study is to evaluate the role and mechanism of Connective Tissue Growth Factor (CTGF) in OC. Methods: R182 and R2615 are well-characterized epithelial OC cell lines. CTGF expression +/- Transforming Growth Factor -b (TGF-b) was determined via Western blot. R182 and R2615 CTGF knock out (KO) were derived utilizing a Cas9/CRISPR-Cas9 lentivirus plasmid vector and confirmed via PCR. Anoikis resistance and invasion assays were performed to characterize phenotypes of R182 and R2615 wild type (WT) and KO cells. For anoikis resistance, cells were plated in triplicate in an ultra-low adhesive (ULA) cell plate Promega CellTiter assay which measured cell viability quantified by absorbance at 450 nm at 0, 24, 48, and 72 hrs. For invasion assay, 3000 cells were suspended in 50% reduced growth factor Basement Membrane Extract. Human recombinant CTGF was added at 50 and 100 ng/mL concentrations. Cells were plated in tissue culture plate and placed in Cytation 5/Biospa and imaged at 4-hour interval for up to 6 days. Western blot evaluated expression of mesenchymal markers. Chemo-sensitivity was measured by half maximal inhibitory concentration (IC50) for Cisplatin between the WT and KO cells. RNA sequence analysis was performed for both R182 WT and KO cells and analyzed utilizing iPathway Guide. Results: CTGF is constitutively expressed in R182 and R2615 OC cells line as early as 6 hrs of culture and after 30 min of TGF-b treatment. Loss of CTGF promoted anoikis resistance. At 72 hr, R182 CTGF WT cells displayed 75% viability while R182 KO only have 10% viability. Further, we demonstrated that loss of CTGF increases invasion. Administration of exogeneous CTGF in KO cells suppresses invasion in a dose dependent manner demonstrated. Along with epithelial markers CK-18 and beta-catenin, the KO cells expressed the mesenchymal marker SNAIL. No difference in chemo-sensitivity to Cisplatin was seen between WT and KO in either cell line. RNA sequence analysis identified 1106 out of 14054 differentially expressed genes between the WT and KO cells. PI3KAkt, cell adhesion molecules and ECM receptor interaction were pathway that were most significantly regulated. Conclusions: CTGF expression maintains the epithelial phenotype of OC cells during EMT. Loss of CTGF allows anoikis resistance and invasion which are vital characteristics in the metastatic nature of OC. We suggest that loss of CTGF expression in OC cells could be a biomarker for more aggressive OC.

Inhibitory Effects and Mechanism of the Natural Compound Diaporthein B Extracted from Marine-Derived Fungi on Colon Cancer Cells.

This study aimed to investigate the inhibitory effects and mechanism of diaporthein B (DTB), a natural compound extracted from the fungus Penicillium sclerotiorum GZU-XW03-2, on human colon cancer cells. The inhibitory effect of DTB at different concentrations on the proliferation of colon cancer cells HCT116 and LOVO was detected at 24 and 48 h. The effect of cell migration and clone formation ability were detected by cell scratch and plate cloning experiments. Morphological changes were observed by Hoechst 33342 and Annexin-V/PI staining, and flow cytometry was used to detect the proportion of apoptotic cells. DTB significantly inhibited colon cancer cell proliferation, migration, and apoptosis in a dose-dependent manner without significant effects on normal colonic epithelial cells NCM460. The IC50 inhibition effect can be achieved after treatment with 3 μmol/L DTB for 24 h. Compared with the blank group, the migration and clonal-forming ability of colon cancer cells in the DTB group was significantly decreased (p < 0.01), while the apoptotic cells were significantly increased (p < 0.01) in a concentration-dependent manner. DTB can inhibit the proliferation and migration of human colon cancer cells HCT116 and LOVO and promote the apoptosis of human colon cancer cells.

Automated Analysis of Acetaminophen Toxicity on 3D HepaRG Cell Culture in Microbioreactor.

Real-time monitoring of bioanalytes in organotypic cell cultivation devices is a major research challenge in establishing stand-alone diagnostic systems. Presently, no general technical facility is available that offers a plug-in system for bioanalytics in diversely available organotypic culture models. Therefore, each analytical device has to be tuned according to the microfluidic and interface environment of the 3D in vitro system. Herein, we report the design and function of a 3D automated culture and analysis device (3D-ACAD) which actively perfuses a custom-made 3D microbioreactor, samples the culture medium and simultaneously performs capillary-based flow ELISA. A microstructured MatriGrid® has been explored as a 3D scaffold for culturing HepaRG cells, with albumin investigated as a bioanalytical marker using flow ELISA. We investigated the effect of acetaminophen (APAP) on the albumin secretion of HepaRG cells over 96 h and compared this with the albumin secretion of 2D monolayer HepaRG cultures. Automated on-line monitoring of albumin secretion in the 3D in vitro mode revealed that the application of hepatotoxic drug-like APAP results in decreased albumin secretion. Furthermore, a higher sensitivity of the HepaRG cell culture in the automated 3D-ACAD system to APAP was observed compared to HepaRG cells cultivated as a monolayer. The results support the use of the 3D-ACAD model as a stand-alone device, working in real time and capable of analyzing the condition of the cell culture by measuring a functional analyte. Information obtained from our system is compared with conventional cell culture and plate ELISA, the results of which are presented herein.

A spatial localization model of mobile robot based on entorhinal-hippocampal cognitive mechanism in rat brain

Physiological studies reveal that rats rely on multiple spatial cells for spatial navigation and memory. In this paper, we investigated the firing mechanism of spatial cells within the entorhinal-hippocampal structure of the rat brain and proposed a spatial localization model for mobile robot. Its characteristics were as follows: on the basis of the information transmission model from grid cells to place cells, the neural network model of place cells interaction was introduced to obtain the place cell plate with a single-peaked excitatory activity package. Then the solution to the robot's position was achieved by establishing a transformation relationship between the position of the excitatory activity package on the place cell plate and the robot's position in the physical environment. In this paper, simulation experiments and physical experiments were designed to verify the model. The experimental results showed that compared with RatSLAM and the model of grid cells to place cells, the positioning performance of the model in this paper was more accurate, and the cumulative error in the long-time path integration process of the robot was also smaller. The research results of this paper lay a foundation for the robot navigation method that mimics the cognitive mechanism of rat brain.

Differentiation of Human Induced Pluripotent Stem Cells into Keratinocytes.

Investigating basic biological mechanisms underlying human diseases relies on the availability of sufficient quantities of patient cells. As most primary somatic cells have a limited lifespan, obtaining sufficient material for biological studies has been a challenge. The development of induced pluripotent stem cell (iPSC) technology has been a game changer, especially in the field of rare genetic disorders. iPSC are essentially immortal, can be stored indefinitely, and can thus be used to generate defined somatic cells in unlimited quantities. Further, the availability of genome editing technologies, such as CRISPR/CAS, has provided us with the opportunity to create “designer” iPSC lines with defined genetic characteristics. A major advancement in biological research stems from the development of methods to direct iPSC differentiation into defined cell types. In this article, we provide the basic protocol for the generation of human iPSC‐derived keratinocytes (iPSC‐K). These cells have the characteristics of basal epidermal keratinocytes and represent a tool for the investigation of normal epidermal biology, as well as genetic and acquired skin disorders. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Directed differentiation of human iPSC into keratinocytes Support Protocol 1: Coating cell culture dishes or plates with Vitronectin XF™ Support Protocol 2: Freezing iPSC Support Protocol 3: Preparing AggreWell™400 6‐well plates for EB formation Support Protocol 4: Coating cell culture dishes or plates with Collagen IV Support Protocol 5: Immunofluorescence staining of cells

Circular RNA-UBE2D2 accelerates the proliferation and metastasis of non-small cell lung cancer cells via modulating microRNA-376a-3p/Eukaryotic Translation Initiation Factor 4γ2 axis

ABSTRACT Non-small cell lung cancer (NSCLC) ranks first in the morbidity and mortality of malignant tumors in China. As reported, circular RNAs (circRNAs) are emerged in the progress of NSCLC. The study was to figure out the potential mechanism of circ-UBE2D2 in the progression of NSCLC. First, plasmid vectors intervening circ-UBE2D2, microRNA (miR)-376a-3p or Eukaryotic Translation Initiation Factor 4γ2 (EIF4G2) expression were transfected into NSCLC cells, and the expression of circ-UBE2D2, miR-376a-3p and EIF4G2 was detected by reverse transcription quantitative polymerase chain reaction or Western blot. Then, cell proliferation was detected by Cell counting kit-8 assay and plate cloning. Cell apoptosis was tested by flow cytometry. Plate scratches and Transwell were used to detect cell migration and invasion. Finally, the binding sites of circRNA UBE2D2, EIF4G2 and miR-376a-3p were verified by bioinformatics website starBase analysis and dual luciferase reporter gene assay. The results manifested the up-regulation of circ-UBE2D2 expression in NSCLC tissues and cells. Circ-UBE2D2 promoted the proliferation, migration and invasion, but repressed apoptosis of NSCLC cells. Interestingly, circ-UBE2D2 directly targeted miR-376a-3p and up-regulated miR-376a-3p restrained proliferation, migration and invasion, but accelerated apoptosis of NSCLC cells. More importantly, EIF4G2 was the target of miR-376a-3p, and overexpression of EIF4G2 reversed the effects of circ-UBE2D2 downregulation on proliferation, migration, invasion and apoptosis of NSCLC cells. These results suggest that circ-UBE2D2 promotes the proliferation, migration and invasion but restrains apoptosis of lung cancer cells by regulating miR-376a-3p/EIF4G2 axis.

In vitro degradation, photo-dynamic and thermal antibacterial activities of Cu-bearing chlorophyllin-induced Ca-P coating on magnesium alloy AZ31.

Surgical failures, caused by postoperative infections of bone implants, are commonly met, which cannot be treated precisely with intravenous antibiotics. Photothermal therapy (PTT) and photodynamic therapy (PDT) have attracted widespread attention due to their non-invasive antibacterial effects on tissues and no bacterial resistance, which may be an excellent approach to solve infections related to bone implants for biodegradable magnesium alloys. Herein, a sodium copper chlorophyllin (SCC) with a porphyrin ring induced Ca–P coating was prepared on AZ31 magnesium alloy via layer-by-layer (LbL) assembly. The morphology and composition of the samples were characterized through field emission scanning electron microscope (FE-SEM) with affiliated energy dispersive spectrometer (EDS), X-ray diffractometer (XRD), and Fourier infrared spectrometer (FTIR) and X-ray photoelectron spectrometer (XPS) as well. Potentiodynamic polarization, electrochemical impedance spectroscopy (EIS) and hydrogen evolution experiments were employed to evaluate the corrosion behavior of the samples. Atomic absorption spectrophotometer was used to measure Cu elemental content of different immersion periods. Cytocompatibility and antibacterial performance of the coatings were probed using in vitro cytotoxicity tests (MTT assay), live/dead cell staining and plate counting method. The results showed that the obtained (Ca–P/SCC)10 coating exhibited good corrosion resistance, antimicrobial activity (especially under 808 nm irradiation) and biocompatibility. The antibacterial rates for E. coli and S. aureus were 99.9% and 99.8%, respectively; and the photothermal conversion efficiency was as high as 42.1%. Triple antibacterial mechanisms including photodynamic, photothermal reactions and copper-ions release were proposed. This coating exhibited a promising application for biodegradable magnesium alloys.

Enhancement of photoluminescence intensity of epoxy thin-film resin by nano graphitic carbon nitride

In this paper, epoxy resin was prepared with graphitic carbon nitride (GCN) via the casting method. Different concentrations of GCN (0.5%, 1%, and 1.5%) were added in the epoxy matrix and studied the effect of filler on the optical properties of the composite. A lower optical bandgap of 2.53 eV harnessing UV and visible light was obtained for 1.5% GCN in an epoxy matrix. Higher optical conductivity was obtained for a higher concentration of GCN in epoxy resin. The structural property and thermal property have been discussed in detail. The room temperature photoluminescence studies indicated enhanced luminescence for a higher concentration of GCN in epoxy resin. The high concentration of GCN creates more traps for electrons hence delayed radiative emission is observed. As this composite possesses excellent optical properties, it could be employed in flat panel displays, LED, optical fibers, and base solar cell plates.

Transcript Profiling Analysis and ncRNAs' Identification of Male-Sterile Systems of Brassica campestris Reveal New Insights Into the Mechanism Underlying Anther and Pollen Development.

Male-sterile mutants are useful materials to study the anther and pollen development. Here, whole transcriptome sequencing was performed for inflorescences in three sterile lines of Chinese cabbage (Brassica campestris L. ssp. chinensis Makino, syn. B. rapa ssp. chinensis), the genic male-sterile line (A line), the Polima cytoplasmic male-sterile (CMS) line (P line), and the Ogura CMS line (O line) along with their maintainer line (B line). In total, 7,136 differentially expressed genes (DEGs), 361 differentially expressed long non-coding RNAs (lncRNAs) (DELs), 56 differentially expressed microRNAs (miRNAs) (DEMs) were selected out. Specific regulatory networks related to anther cell differentiation, meiosis cytokinesis, pollen wall formation, and tapetum development were constructed based on the abortion characteristics of male-sterile lines. Candidate genes and lncRNAs related to cell differentiation were identified in sporocyteless P line, sixteen of which were common to the DEGs in Arabidopsis spl/nzz mutant. Genes and lncRNAs concerning cell plate formation were selected in A line that is defected in meiosis cytokinesis. Also, the orthologs of pollen wall formation and tapetum development genes in Arabidopsis showed distinct expression patterns in the three different sterile lines. Among 361 DELs, 35 were predicted to interact with miRNAs, including 28 targets, 47 endogenous target mimics, and five precursors for miRNAs. Two lncRNAs were further proved to be functional precursors for bra-miR156 and bra-miR5718, respectively. Overexpression of bra-miR5718HG in B. campestris slowed down the growth of pollen tubes, caused shorter pollen tubes, and ultimately affected the seed set. Our study provides new insights into molecular regulation especially the ncRNA interaction during pollen development in Brassica crops.

MicroRNA-29c-3p participates in insulin function to modulate polycystic ovary syndrome via targeting Forkhead box O 3

ABSTRACT MicroRNAs (miRNAs) are gene expression regulators and changes in miRNA levels are associated with diabetes, insulin resistance, and inflammation, the latter two of which are characteristic of polycystic ovary syndrome (PCOS). The purpose of this study was to explore the specific mechanism in which miR-29 c-3p participated in insulin function to regulate PCOS by targeting Forkhead box O 3 (Foxo3). Peripheral blood from PCOS patients and healthy volunteers were first collected, and the expression levels of miR-29 c-3p and Foxo3 were detected by reverse transcription quantitative polymerase chain reaction or Western blot. Then human granular tumor cell line (KGN) was treated with insulin, and transfected with plasmid vectors interfering with miR-29 c-3p or Foxo3 expression. Cell proliferation was detected by Cell counting kit-8 and plate cloning, and cell apoptosis was tested by flow cytometry. In addition, PCOS rat model was established. PCOS rats were injected with plasmids vectors interfering with miR-29 c-3p or Foxo3 expression, respectively. Pathological changes in ovarian tissues of rats in each group were observed by hematoxylin-eosin staining, and serum sex hormones and glucose metabolism-related indicators were detected. Finally, via bioinformatics website, luciferase digestion report assay was detected the targeting relationship between miR-29 c-3p and Foxo3. The experimental results showed that miR-29 c-3p was down-regulated in PCOS, but Foxo3 was up-regulated. Up-regulated miR-29 c-3p or down-regulated Foxo3 promoted KGN cell proliferation, inhibited apoptosis in vitro, restored PCOS rat sex hormone levels and improved glucose metabolism in vivo. These results suggest that miR-29 c-3p is involved in insulin function to improve PCOS by targeting Foxo3.

Arabidopsis exocyst subunit SEC6 is involved in cell plate formation during Microgametogenesis

Cytokinesis during pollen mitosis I is critical for cell division and differentiation in the male gametophyte development, but the vesicle trafficking mechanisms in this process are largely unknown. Exocyst is an octameric tethering complex which plays multiple important roles in plant cell vesicle trafficking. Here we report the characterization of exocyst subunit SEC6 in the cytokinesis during pollen mitosis I. We found that significantly amount of pollen from two sec6/+ mutant alleles arrested at the transition from unicelluar stage microspore to bicellular stage. Further analysis showed that sec6 mutation impaired cell plate formation and led to vesicles accumulation in cytoplasm. The localization of KNOLLE on the cell plate was compromised. Consistently, SEC6 gene was expressed start from early pollen development stage and SEC6-GFP localized to the cell plate. These results indicated that SEC6 participated in the cell plate formation during pollen mitosis I, where it might help to tether the vesicles before fusion.

Evaluation of the In-Vitro Cytotoxicity of Heat Cure Denture Base Resin Modified with Recycled PMMA-Based Denture Base Resin.

Aim:The aim of the present study was to evaluate the in-vitro cytotoxicity of heat-cure denture base resin (PMMA) modified with recycled denture base resin at 10%, 20%, 30%, 40%, and 50% (w/w) concentration.Materials and Methods:A total of 30 disk-shaped specimens were prepared and divided into six groups (n = 5). The Control group (R0) consisted of unmodified processed denture base resin, the experimental group consisted of denture base resin processed with substitution of 10% 20%, 30%, 40%, and 50% (w/w) of recycled denture base resin (R10, R20, R30, R40, and R50). Eluates were prepared using five sterile specimens of each group. The mouse fibroblast cell line (L929) was seeded in a 96-well cell plate system at a concentration of 1 × 104 cells/well in the DMEM medium with 1× antibiotic and antimycotic solution and 10% fetal bovine serum at 37°C with 5% CO2 and incubated in a CO2 incubator for 48 h. MTT assay was applied and the absorbance was measured at 570 nm using a microplate reader to assess the in-vitro cytotoxicity. One-way analysis of variance (ANOVA) along with post hoc Scheffe test was used to statistically compare the mean optical density (OD) values and cell survival/viability % amongst the groups.Results:No statistically significant difference was observed in the mean and standard deviation of the optical density and cell viability % of the test groups that were compared.Conclusion:Modification of denture base resin using recycled PMMA does not have a cytotoxic effect on the mouse fibroblast cell line L929.

Analysis of fluid flow and heat transfer in the novel liquid volumetric plated cavity receiver

The article mainly focuses on the study of thermal performance of liquid volumetric plated cavity receiver which includes the flow characteristics of heat transfer fluid and estimation of heat losses. Due to higher heat capacity, liquids are considered more suitable heat transfer fluid than air. However, it is difficult to use liquid as heat transfer fluid (HTF) in the conventional air volumetric receivers, therefore a plated volumetric receiver is proposed, as it can heat the fluid within the volume of the receiver. The plated receiver consists of a parallel arrangement of rectangular cells suspended in an enclosure. Each cell is enclosed by two large rectangular metallic plates joined together, creating a hollow space between them with inlet and outlet nozzles. The incoming solar radiation is absorbed by outer surface of the cell and transferred to the HTF. Large absorbing surface area and large contact area between the plates and HTF is the key feature for avoiding hot spots on the cell’s surfaces. Numerical analysis is carried out for molten salt flow inside the cells and air flow in the cavity. The results show a uniform distribution of flow (with Re = 212) resulting into no hotspots on the receiver surface. Local heat transfer coefficient is estimated to be maximum and minimum in the high and low flux regions, respectively. The thermal efficiency of the receiver is calculated as 90.76%. The cell thickness and plate size of the receiver is 6 mm and 100 mm × 50 mm, respectively.

An in vitro study on the effects of serum proteins on Enterococcus faecalis adhesion to three types of root sealers and gutta-percha

BackgroundThe extrusion of overfilled materials that extend beyond the apical foramina into the periradicular tissue may serve as a reservoir for bacterial adhesion and further affect recovery from periapical diseases. The aim of this study was to evaluate the effects of serum proteins on Enterococcus faecalis adhesion and survival on the surface of a calcium hydroxide-based root canal sealer (Apexit Plus), an epoxy resin sealer (AH-Plus) and a bioceramic sealer (iRoot SP).MethodsApexit Plus, AH-Plus and iRoot SP were evenly coated on gutta-percha, using gutta-percha alone as the control. After root canal sealer setting, the number of E. faecalis adhering to the root canal sealers and gutta-percha was counted in fetal bovine serum (FBS) or tryptic soy broth supplemented with 1% glucose (TSBG) by viable cell plate counts. The morphology of 7-day-old E. faecalis biofilms in FSB and TSBG was observed by scanning electron microscopy (SEM). Furthermore, E. faecalis biofilms on the three root canal sealers were labeled with a LIVE/DEAD BacLight™ Bacterial Viability Kit, and the ratios of viable to dead cells were analyzed using laser scanning microscopy operative software (Zen software).ResultsIn the assays, after 1 and 7 days, the number of E. faecalis adhering to the root canal sealers or gutta-percha in FBS were significantly lower than those in TSBG (P < 0.05). In FBS, E. faecalis adhesion to iRoot SP and gutta-percha was reduced to a greater extent than that adhered to Apexit Plus and AH-Plus. Few E. faecalis accumulated on iRoot SP in FBS, whereas many bacteria assembled on iRoot SP and formed biofilms in TSBG. The ratio of viable cells in the E. faecalis biofilm on iRoot SP was the lowest.ConclusionsCalcium hydroxide-based root canal sealers, epoxy resin sealers and bioceramic sealers may provide a substrate for E. faecalis adhesion, and the bioceramic sealer in this study showed the least E. faecalis adhesion in the presence of serum proteins compared to the other two sealers.

Comparative transcriptome and DNA methylation analysis in temperature-sensitive genic male sterile wheat BS366

BackgroundKnown as the prerequisite component for the heterosis breeding system, the male sterile line determines the hybrid yield and seed purity. Therefore, a deep understanding of the mechanism and gene network that leads to male sterility is crucial. BS366, a temperature-sensitive genic male sterile (TGMS) line, is male sterile under cold conditions (12 °C with 12 h of daylight) but fertile under normal temperature (20 °C with 12 h of daylight).ResultsDuring meiosis, BS366 was defective in forming tetrads and dyads due to the abnormal cell plate. During pollen development, unusual vacuolated pollen that could not accumulate starch grains at the binucleate stage was also observed. Transcriptome analysis revealed that genes involved in the meiotic process, such as sister chromatid segregation and microtubule-based movement, were repressed, while genes involved in DNA and histone methylation were induced in BS366 under cold conditions. MethylRAD was used for reduced DNA methylation sequencing of BS366 spikes under both cold and control conditions. The differentially methylated sites (DMSs) located in the gene region were mainly involved in carbohydrate and fatty acid metabolism, lipid metabolism, and transport. Differentially expressed and methylated genes were mainly involved in cell division.ConclusionsThese results indicated that the methylation of genes involved in carbon metabolism or fatty acid metabolism might contribute to male sterility in BS366 spikes, providing novel insight into the molecular mechanism of wheat male sterility.

A biophysical model for plant cell plate maturation based on the contribution of a spreading force.

Plant cytokinesis, a fundamental process of plant life, involves de novo formation of a “cell plate” partitioning the cytoplasm of dividing cells. Cell plate formation is directed by orchestrated delivery, fusion of cytokinetic vesicles, and membrane maturation to form a nascent cell wall by timely deposition of polysaccharides. During cell plate maturation, the fragile membrane network transitions to a fenestrated sheet and finally a young cell wall. Here, we approximated cell plate sub-structures with testable shapes and adopted the Helfrich-free energy model for membranes, including a stabilizing and spreading force, to understand the transition from a vesicular network to a fenestrated sheet and mature cell plate. Regular cell plate development in the model was possible, with suitable bending modulus, for a two-dimensional late stage spreading force of 2–6 pN/nm, an osmotic pressure difference of 2–10 kPa, and spontaneous curvature between 0 and 0.04 nm−1. With these conditions, stable membrane conformation sizes and morphologies emerged in concordance with stages of cell plate development. To reach a mature cell plate, our model required the late-stage onset of a spreading/stabilizing force coupled with a concurrent loss of spontaneous curvature. Absence of a spreading/stabilizing force predicts failure of maturation. The proposed model provides a framework to interrogate different players in late cytokinesis and potentially other membrane networks that undergo such transitions. Callose, is a polysaccharide that accumulates transiently during cell plate maturation. Callose-related observations were consistent with the proposed model’s concept, suggesting that it is one of the factors involved in establishing the spreading force.