Phase Contrast Microscopic Analysis Research Articles

Brucine Entrapped Titanium Oxide Nanoparticle for Anticancer Treatment: An In Vitro Study.

Backgroundand Objective. The public's health has been seriously threatened by cervical cancer during recent times. In terms of newly diagnosed cases worldwide, it ranks as the ninth most prevalent malignancy. Multiple investigations have proven that nanoparticles can effectively combat cancer due to their small dimensions and extensive surface area. In the meantime, bioactive compounds which are biocompatible are being loaded onto nanoparticles to promote cancer therapy. The current study investigates the anticancerous potential of Brucine-entrapped titanium oxide nanoparticles (TiO2 NPs) in cervical cancer cell line (HeLa). Materials and Methods. The physiochemical, structural, and morphological aspects of Brucine-entrapped TiO2 NPs were evaluated by UV-visible spectrophotometer, Fourier transform-infrared spectroscopy (FT-IR), dynamic light scattering (DLS), scanning electron microscopy (SEM), and energy dispersive X-ray (EDAX). The cytotoxic effect against the HeLa cell line was assessed using a tetrazolium-based colorimetric assay (MTT), a trypan blue exclusion (TBE) assay, phase contrast microscopic analysis, and a fluorescence assay including ROS and DAPI staining. Furthermore, estimation of antioxidant markers includes catalase (CAT), glutathione (GSH), and superoxide dismutase (SOD). Results. The UV spectrum at 266 nm revealed the formation of TiO NPs. The FT-IR peaks confirmed the effective entrapment of brucine with TiO2 NPs. The average size (100.0 nm) of Brucine-entrapped TiO2 NPs was revealed in DLS analysis. The micrograph of the SEM revealed the formation of ellipsoidal to tetragonal-shaped NPs. The Ti, O, and C signals were observed in EDAX. In MTT assay, Brucine-entrapped TiO2 NPs showed inhibition of cell proliferation in a dose-wise manner and IC50 was noticed at the concentration of 30 µg/mL. The percentage of viable cells gradually reduced in the trypan blue exclusion assay. The phase contrast microscopic analysis of Brucine-entrapped TiO2 NP-treated cells showed cell shrinkage, cell wall deterioration, and cell blebbing. The intracellular ROS level was increased in a dose-wise manner when compared to control cells in ROS staining. The condensed nuclei and apoptotic cells were increased in treated cells, as noted in DAPI staining. In treated cells, the antioxidant markers such as CAT, GSH, and SOD levels were substantially lower compared to the control cells. Conclusion. The synthesized Brucine entrapped TiO2 NPs exhibited remarkable anticancer activity against the HeLa cell line.

Workers exposure to synthetic textiles microfibers

The textile industry is one of the most polluting sectors, its contribution to the growing contamination with microfibers being of recent interest. Microfibers are an important part of microplastics, a real problem in our society. Textiles are dominated by synthetic fibres, generating microfibres as environmental contaminants. Exposure may also occur in the workplace, but fewer studies address this topic. Although synthetic microfibres toxicity is not fully understood, is accepted that their dimensions can influence health effects. This study analysed the length and diameter of 201 airborne polyester fibres sampled in the working environment of a textile factory. Phase contrast microscopic analysis was performed using NIOSH method 7400. Three size categories were considered for the length: small (≤ 10 μm), medium (10 - 100 μm) and large (≥ 100 μm). Four diameter categories: thin (≤ 1 μm), medium (≤ 2 μm) and thick (≤ 3 μm) were defined. Diameters over 3 microns were considered elongated microplastics. Results show that most of the fibres (93%) were less than 100 μ long (20.1% being less than 10 μ) and the fibres with 1μ diameter were predominant (42.8%) Such fibres may reach into the bronchial tree and even further, where they may cause health problems.

Periodontal Maintenance Therapy: Efficacy of Oral Irrigator in the Home Oral Hygiene Protocol Associated with Microbiological Analysis with Phase Contrast Microscope

Objective: The aim of this research was to assess how the application of an oral irrigator modifies the subgingival bacterial flora in patients undergoing periodontal maintenance therapy. We used a qualitative microbiological analysis with a phase contrast microscope that can differentiate a non-pathogenic (immobile) bacterial flora from a pathogenic (mobile). Methods: In this study, 60 patients with a diagnosis of moderate periodontitis were enrolled. They were treated with non-surgical laser-assisted causal therapy and after one month a re-evaluation was performed that established the stabilization of the periodontal pathology. They were instructed in the home oral hygiene protocol with sonic toothbrush, interdental brushes and oral irrigator, and were included in the periodontal maintenance therapy program with a phase contrast microscope control every 6 months. At T0 phase contrast–phase microscopic examinations, the existence of non-pathogenic bacterial flora was displayed in all patients. They were then randomly divided into two groups, A and B. After professional oral hygiene treatment, group B suspended the usage of oral irrigator. Patients of group A, after the professional oral hygiene treatment, were motivated to continue their oral hygiene protocol at home. After three months, the patients underwent microscopic analysis of the bacterial plaque. Results: At T1 in Group B, 90% of patients had undergone a pathogenic bacterial flora change. In group A, in which patients kept using the oral irrigator for the three months, 100% of them showed immobile plaque on phase contrast microscopic analysis. Conclusions: This research showed that oral irrigator in the practice of home hygiene protocols plays a role in the long-term maintenance of a non-pathogenic bacterial flora in periodontal patients.

Coumarin–gold nanoparticle bioconjugates: preparation, antioxidant, and cytotoxic effects against MCF-7 breast cancer cells

In recent, the conjugation of gold nanoparticles (AuNPs) with biomolecules has shown great potential especially in disease diagnostics and treatment. Taking this in account, we report the methodology involved in the conjugation of coumarin onto the surface of citrate-capped AuNPs by a simple in situ method. Herein, we systematically performed UV–Vis spectroscopy, transmission electron microscopy, dynamic light scattering, and zeta potential measurements to characterize citrate-capped AuNPs and bioconjugates. Our results demonstrate in-depth surface chemistry of bioconjugates with improved surface plasmon resonance (529 nm), morphology (near spherical shape), hydrodynamic diameter (25.3 nm) as well as surface charge (− 35 mV). Furthermore, the bioconjugates displayed dose-dependent response in scavenging free radicals and exhibited cytotoxicity against MCF-7 breast cancer cell lines. In addition, phase-contrast microscopic analysis revealed that bioconjugates promote apoptosis in cancer cells in a time-dependent manner. Overall, we ascertain the fact that this kind of bioconjugation of AuNPs with coumarin further enhances the efficacy of inorganic nanomaterials and thus make them a better bio-therapeutic candidate.

In vitro evaluation of anticancer potentials of lupeol isolated from Elephantopus scaber L. on MCF-7 cell line.

Lupeol is a triterpenoid, present in most of the medicinally effective plants and possess a wide range of biological activity against human diseases. The present study aims at evaluating the anticancer potentials of lupeol, isolated from the leaves of Elephantopus scaber L. and thereby explores its action on key cancer marker, Bcl-2. The effect of lupeol on the cell viability of MCF-7 was determined by MTT and lactate dehydrogenase assays at different concentrations. The efficacy of the compound to induce cell death was analyzed using AO/EtBr staining. Phase contrast microscopic analysis provided the changes in cell morphology of the compound treated normal breast cells (MCF-10A) and MCF-7 cells. The expression of Bcl-2 and Bcl-xL proteins in the normal, cancer and lupeol treated cancer cell was analyzed by western blotting. Lupeol induced an effective change in the cell viability of MCF-7 cells with IC50 concentration as 80 μM. Induction of cell death, change in cell morphology and population of the cancer cells was observed in the lupeol treated cells, but the normal cells were not affected. The compound effectively downregulated Bcl-2 and Bcl-xL protein expressions, which directly contribute for the induction of MCF-7 cell apoptosis. Conclusion: Thus, lupeol acts as an anticancer agent against MCF-7 cells and is a potent phytodrug to be explored further for its cytotoxic mechanism.

Antibacterial activity of Aquilaria crassna leaf extract against Staphylococcus epidermidis by disruption of cell wall.

BackgroundAquilaria crassna Pierre ex Lecomte has been traditionally used in Thailand for treatment of infectious diseases such as diarrhoea and skin diseases for a long time. The main objectives of this study were to examine antibacterial activity of the Aquilaria crassna leaf extract against Staphylococcus epidermidis and its underlying mechanism. The antioxidant activity and acute toxicity were studied as well.MethodsAntioxidant activities were examined by FRAP, ABTS and DPPH scavenging methods. Antibacterial activity was conducted using disc diffusion assay and the minimum inhibitory concentration (MIC) was determined by dilution method. The minimum bactericidal concentration (MBC) was reported as the lowest concentration producing no growth of microbes in the subcultures. Morphological changes of the microbe were observed by scanning electron microscopy, while an inhibitory effect on biofilm formation was evaluated by phase contrast microscopic analysis. Bacterial cell wall integrity was assessed by transmission electron microscopy. Acute toxicity was conducted in accordance with the OECD for Testing of Chemicals (2001) guidelines.ResultsThe extract exhibited considerable antioxidant activity. Staphylococcus epidermidis was susceptible to the extract with the MIC and MBC of 6 and 12 mg/ml, respectively. The extract caused swelling and distortion of bacterial cells and inhibited bacterial biofilm formation. Rupture of bacterial cell wall occurred after treated with the extract for 24 h. Acute toxicity test in mice showed no sign of toxicity or death at the doses of 2,000 and 15,000 mg/kg body weight.ConclusionThe aqueous extract of Aquilaria crassna leaves possesses an in vitro antibacterial activity against Staphylococcus epidermidis, with no sign of acute oral toxicity in mice, probably by interfering with bacterial cell wall synthesis and inhibiting biofilm formation.

Involvement of Selective Reactive Oxygen Species Upstream of Proapoptotic Branches of Unfolded Protein Response

Cadmium triggers apoptosis of LLC-PK1 cells through induction of endoplasmic reticulum (ER) stress. We found that cadmium caused generation of reactive oxygen species (ROS) and that cadmium-induced ER stress was inhibited by antioxidants. In contrast, suppression of ER stress did not attenuate cadmium-triggered oxidative stress, suggesting that ER stress occurs downstream of oxidative stress. Exposure of the cells to either O(2)(*), H(2)O(2), or ONOO(-) caused apoptosis, whereas ER stress was induced only by O(2)(*) or ONOO(-). Transfection with manganese superoxide dismutase significantly attenuated cadmium-induced ER stress and apoptosis, whereas pharmacological inhibition of ONOO(-) was ineffective. Interestingly, transfection with catalase attenuated cadmium-induced apoptosis without affecting the level of ER stress. O(2)(*) caused activation of the activating transcription factor 6-CCAAT/enhancer-binding protein-homologous protein (CHOP) and the inositol-requiring ER-to-nucleus signal kinase 1-X-box-binding protein 1 (XBP1) proapoptotic cascades, and overexpression of manganese superoxide dismutase attenuated cadmium-triggered induction of both pathways. Furthermore, phosphorylation of proapoptotic c-Jun N-terminal kinase by O(2)(*) or cadmium was suppressed by dominant-negative inhibition of XBP1. These data elucidated 1) cadmium caused ER stress via generation of ROS, 2) O(2)(*) was selectively involved in cadmium-triggered, ER stress-mediated apoptosis through activation of the activating transcription factor 6-CHOP and inositol-requiring ER-to-nucleus signal kinase 1-XBP1 pathways, and 3) phosphorylation of JNK was caused by O(2)(*)-triggered activation of XBP1.

Detoxification of industrial asbestos waste by low-temperature heating in a vacuum

Industrial asbestos waste such as asbestos cement board, sprayed crocidolite and sprayed amosite was heated in a vacuum. The detoxification conditions were then established in reference to JIS A 1481:2006 and JIS K 3850-1:2006. When asbestos cement board was heated at 700°C for 1 h, detoxification of the cement board was completed. On the other hand, in the case of amphibolic asbestos waste, such as sprayed crocidolite and sprayed amosite, was heated, a phase-contrast microscope analysis was not applicable for identification of asbestos heated at more than 500°C. According to XRD analysis, crocidolite was eliminated by heating at 800°C for 3 h. Furthermore, amosite was not detected after heating at 700°C for 3 h. Finally, the grindability of heated amphibolic asbestos was investigated. When sprayed crocidolite was heated at 900°C for 3 h and then ground with a mortar for 300 s, the crushed grains had a length of less than 5 μm and an aspect ratio of 3. In addition, when sprayed amosite was heated at 1000°C for 3 h and then ground, the crushed grains had a length of less than 5 μm and an aspect ratio of 3.

Detoxification of Asbestos-Containing Building Material Waste and Its Application to Cement Product

According to phase-contrast microscope analysis based on the JIS A 1481: 2006, chrysotile in the asbestos-containing building material waste was not detected after heating at 800°C for over 2 h. The asbestos-cement which was ground to below 180 μm after heating asbestos-containing building material waste at 800°C for 3 h was added into OPC to fabricate secondary cement product. When 50 mass% of the asbestos-cement was added to OPC, the compressive strength of the secondary cement product after 7 days was 32 MPa. Further chrysotile was not detected in the secondary cement product using phase contrast microscope analysis.

Detoxification of Sprayed Amosite

According to XRD analysis, when sprayed amosite was heated at over 900°C for 3 h, amosite crystal was not detected in the sample. In contrast, a part of the needlelike amosite grains were not detected in the sprayed amosite using a phase-contrast microscope analysis after heating at 500°C for 3 h. Then the color of the needlelike amosite grains changed to red, because Fe2O3 (hematite) was precipitated at the surface of the amosite grains. The grindability of the needlelike amosite grains was improved with an increase of the heating temperature. When the sprayed amosite was heated at 1100°C for 3 h, a fine powder with under aspect ratio of 3 was obtained after grinding.

Morphology of renal parenchymal culture in collagen gels

Explants of rabbit renal parenchyma have been grown in primary tissue culture suspended within hydrated collagen gels. Light and phase contrast microscopic analysis of the first 17 days in culture is described. Pieces of NZW rabbit renal parenchyma were suspended in collagen gels and bathed in supplemented RPMI 1640 medium and incubated at 37 degrees C in 5% CO2 in air. Tubules demonstrated a fine granularity by phase contrast microscopy and glomeruli appeared as red spheres. Blebs formed at the sides and ends of the explant and a monolayer outgrowth of tightly packed polygonal cells occurred from day 4. Histologically an immediate phase of necrosis was followed by regeneration whereby tubules became lined with a confluent epithelium composed of a single layer of flat to cuboidal-shaped cells sitting on an intact tubular basement membrane (TBM). Intraluminal casts of organized cellular debris as well as material presumed to be Tamm Horsfall protein were present. Glomeruli demonstrated collapsed capillary loops. The interstitium became widened by eosinophilic material. The tissue surface contained epithelial cells arranged in places into sac-like structures enclosing a space.

A predictive model for determining asbestos concentrations for fibers less than five micrometers in length

The controversy of whether small asbestos fibers are biologically significant has not been resolved. The present standard method for evaluating asbestos fiber concentrations in workroom air excludes fibers less than 5 micron long even though it has been shown that small fiber concentrations dominate in a dust cloud. This research project was conducted to develop a mathematical model whereby one could predict small (less than 5 micron length) asbestos fiber concentration based on the fiber count concentration determined by phase contrast microscope analysis. Dry chrysotile asbestos was aerosolized into a chamber and sampled by membrane filtration. Segments from each filter were analyzed by both the NIOSH technique using phase contrast microscopy (PCM) and scanning electron microscopy (SEM) at 2000 X for fiber concentrations. A linear relationship was found to exist between the natural logarithm of the SEM-determined concentration and the natural logarithm of the PCM-determined concentration (r = 0.852). Using these data, a mathematical model was developed to predict SEM concentrations based on PCM counts. This model may have application in retrospective epidemiological studies for estimating small fiber exposure levels to determine if small fibers play a role in disease production. The greatest utility would be in those retrospective studies where the only exposure information available is based on PCM counts.

Orthodontic therapy in patients with juvenile periodontitis: Clinical and microbiologic effects

The correction of malocclusions in juvenile periodontitis (JP) patients completing periodontal therapy is a problem of increasing clinical concern to orthodontists, since many teeth with severe alveolar bone loss in these patients can now be successfully treated without extraction. In this report, fixed edgewise orthodontic therapy was carried out after the completion of periodontal therapy on four JP patients. The orthodontic therapy included extensive intrusion of teeth severely affected by JP. Phase-contrast microscopic analysis of subgingival plaque from orthodontically treated teeth was used to monitor longitudinally the effects of fixed orthodontic bands on the subgingival flora and also to monitor the efficacy of topical and systemic antimicrobial therapy aimed at suppression of suspected periodontopathic bacteria. Orthodontic movement was completed on most periodontally compromised teeth without significant evidence of additional deterioration in periodontal status. However, within the first 6 months of orthodontic band placement, all patients had significant increases in the number of spirochetes and motile rods in their subgingival flora. Three of the patients also developed high levels of crevicular polymorphonuclear leukocytes around orthodontically treated teeth, indicating significant subgingival inflammation. Intensive antimicrobial measures, including topical inorganic salt applications and systemic tetracycline, were helpful in limiting clinical inflammation and subgingival colonization by periodontopathogens during orthodontic therapy. The results demonstrate that successful orthodontic repositioning can be carried out in treated JP patients. In addition, bacteriologic monitoring and chemotherapeutic suppression of periodontal pathogens may be valuable in the prevention of further destructive periodontal disease activity in periodontitis patients undergoing orthodontic therapy.