Intermittent fasting along with hydroalcoholic extract of Centella-asiatica ameliorates sub-acute hypoxia-induced ischemic stroke in adult zebrafish.

Reduced blood flow (hypoxia) to the brain is thought to be the main cause of strokes because it deprives the brain of oxygen and nutrients. An increasing amount of evidence indicates that the Centella-Asiatica (HA-CA) hydroalcoholic extract has a variety of pharmacological benefits, such as antioxidant activity, neuroprotection, anti-inflammatory qualities, and angiogenesis promotion. Intermittent fasting (IF) has neurological benefits such as anti-inflammatory properties, neuroprotective effects, and the ability to enhance neuroplasticity. The current study evaluates the combined effect of IF (for 1, 6, and 12 days) along with HA-CA (daily up to 12 days) in adult zebrafish subjected to hypoxia every 5 min for 12 days followed by behavioral (novel tank and open-field tank test), biochemical (SOD, GSH-Px, and LPO), inflammatory (IL-10, IL-1ß, and TNF-α), mitochondrial enzyme activities (Complex-I, II, and IV), signaling molecules (AMPK, MAPK, GSK-3ß, Nrf2), and imaging/staining (H&E, TTC, and TEM) analysis. Results show that sub-acute hypoxia promotes the behavioral alterations, and production of radical species and alters the oxidative stress status in brain tissues of zebrafish, along with mitochondrial dysfunction, neuroinflammation, and alteration of signaling molecules. Nevertheless, HA-CA along with IF significantly ameliorates these defects in adult zebrafish as compared to their effects alone. Further, imaging analysis significantly provided evidence of infarct damage along with neuronal and mitochondrial damage which was significantly ameliorated by IF and HA-CA. The use of IF and HA-CA has been proven to enhance the physiological effects of hypoxia in all dimensions.

Investigating temperature variability on antioxidative behavior of synthesized cerium oxide nanoparticle for potential biomedical application.

Cerium oxide nanoparticles (CNP) have garnered significant attention due to their versatile redox properties and wound-healing applications. The antioxidative nature of CNP is due to its ability to be oxidized and reduced, followed by the capture or release of oxygen which is used for scavenging reactive oxygen species (ROS). Herein, CNP is produced through a wet chemistry approach and its tunable redox property is tested across a range of temperatures. The synthesized CNP was observed to reveal the signature peak at 245 nm indicating a high Ce+3/Ce+4 ratio. Towards evaluating the redox antioxidative behavior, CNPs were subjected to a comprehensive analysis for superoxide dismutase mimetic analysis with riboflavin-mediated nitroblue tetrazolium scavenging assay. The results demonstrated that the redox activity of cerium oxide nanoparticles was strongly influenced by the different temperature ranges. Superoxide dismutase mimetic activity was observed to be reduced with a decrease in temperature as we moved from 4°C (80% activity) to -80°C (47% activity) at 1 mM conc of CNP. Similarly, the SOD mimetic activity increased with an increase in temperature from 40°C (72% activity) to 70°C (94% activity). Further, CNP was found to inhibit E. coli (gram+ve) and Enterobacter (gram-ve) beyond 70% simultaneously at 1 mM conc, indicating its potential application as a remarkable antimicrobial agent. CNP also inhibited the alpha-amylase activity up to the 60% at 1 mM conc suggesting its potential application in antidiabetic wound healing therapy. Overall, the CNP finds its application in mitigating the oxidative stress-related disorder exhibited by its high antioxidative, antimicrobial, and antidiabetic behavior.

Monomer release, cell adhesion, and cell viability of indirect restorative materials manufactured with additive, subtractive, and conventional methods.

PURPOSE: The aim of this study was to measure residual monomer, cell adhesion, and cell viability of 3-dimensional printable permanent resin (PR), hybrid ceramic block (HCB), and indirect composite (IC) produced with additive, subtractive, and conventional techniques. METHODS: Five 8 × 8 × 2 mm3 samples of each material were prepared for each experiment. In a 24-h period, monomer release was analyzed with high-performance liquid chromatography, and cell viability and adhesion were evaluated with the water-soluble tetrazolium salt test. Data were analyzed with IBM SPSS Statistics 26.0 statistical software, and results were regarded as significant at α = 0.05. RESULTS: Monomer release (triethylene glycol dimethacrylate, urethane dimethacrylate, and Bisphenol A glycerolate dimethacrylate) was significantly higher in the IC group. Mean cell viability was significantly lower in the HCB group than in the IC group. CONCLUSION: All monomers in the tested materials were released at rates that were below clinical significance. Cell adhesion rates in the groups were similar. Cytotoxic response was classified as minor in the HCB and PR groups and non-cytotoxic in the IC group.

Immunomodulatory effect of Myrtenol on benzo (a) pyrene-induced lung cancer in Swiss albino mice via modulation of tumor markers, cytokines and inhibition of PCNA expression.

Lung cancer is one of the most common cancers in men. Although many diagnostic and treatment regimens have been followed in the treatment for lung cancer, increasing mortality rate due to lung cancer is depressing and hence requires alternative plant based therapeutics with with less side-effects. Myrtenol exhibits anti-inflammatory and antioxidant properties. Hence we intended to study the effect of Myrtenol on B(a)P-induced lung cancer. Our study showed that B(a)P lowered hematological count, decreased phagocyte and avidity indices, nitroblue tetrazolium (NBT) reduction, levels of immunoglubulins, antioxidant levels, whereas Myrtenol treatment restored them back to normal levels. On the other hand, xenobiotic and liver dysfunction marker enzymes and pro-inflammatory cytokines were elevated on B(a)P exposure, which retuned back to normal by Myrtenol. This study thus describes the immunomodulatory and antioxidant effects of Myrtenol on B[a]P-induced immune destruction.

2,3,5-triphenyl tetrazolium chloride as colorimetric indicator for drug susceptibility testing against nontuberculous mycobacteria.

This study aimed to propose and evaluate a drug susceptibility testing (DST) using the 2,3,5-triphenyl tetrazolium chloride (TTC) as a colorimetric indicator against Mycobacterium abscessus complex (MABC), M. avium complex (MAC), and M. kansasii strains, main nontuberculous mycobacteria (NTM) of clinical relevance. Our results demonstrated that the assay using TTC and the broth microdilution method recommended by the Clinical and Laboratory Standards Institute had essential agreement above 91%, 92%, and 100%, for drugs tested against MABC, MAC, and M. kansasii strains, respectively. Categorical agreement above 91% was obtained for most drugs tested against MABC, except to cefoxitin (76.5%). For drugs tested against MAC and M. kansasii, categorical agreement above 92% and 100% was observed, respectively. TTC showed to be a promising colorimetric indicator of growth to be used in DST for NTM, allowing an easier reading of results.

Formation of morpholine-acetamide derivatives as potent anti-tumor drug candidates: Pharmacological evaluation and molecular docking studies.

Heterocyclic amines and acetamide derivatives are known for their chemotherapeutic potential. Hence, in the present study, morpholine was taken as a principal product and novel morpholine derivatives were designed, formulated, characterized, and screened for the mechanism of inhibition of carbonic anhydrase and their anticancer potential. In addition, in vitro inhibition of hypoxia-inducible factor-1 (HIF-1) protein was also investigated. Results revealed that compounds 1c, 1d, and 1h possessed significant inhibitory activities against carbonic anhydrase with IC50 of 8.80, 11.13, and 8.12 µM, respectively. Interestingly, the carbonic anhydrase inhibitory activity of compound 1h was comparable with that of standard acetazolamide (IC50 7.51 µM). The compounds 1h and 1i significantly inhibited the proliferation of ovarian cancer cell line ID8 with IC50 of 9.40, and 11.2 µM, respectively while the standard cisplatin exhibited an IC50 8.50 µM. In addition, compounds 1c, 1b, 1h and 1i also exhibited significant inhibitory effects on HIF-1α. In conclusion, we report first time the biological potential of morpholine based compounds against ovarian cancer and HIF-1α that may serve as lead molecules for drug discovery.

Comparative Evaluation of Nano Inorganic Metal Oxides as Intracanal Medicaments for Cytotoxicity and Antimicrobial Activity in the Root Canal System.

Aim and objective: To evaluate and compare the cytotoxicity and antimicrobial activity of various inorganic metal oxide nanoparticles along with vehicles when used as intracanal medicaments in the root canal system. Materials and methods: The study included triplicates (n = 36 times) that were subjected to n calcium oxide (CaO), n zinc oxide (ZnO), n magnesium oxide (MgO), and metapaste as intracanal medicaments. The efficacy of novel intracanal medicaments was evaluated for biocompatibility assay using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reagent following antimicrobial efficacy against Enterococcus faecalis (E. faecalis) was evaluated using zone of inhibition (ZOI) and minimum inhibitory concentration (MIC). The statistical analysis Kruskal-Wallis test, student t-test, and analysis of variance (ANOVA) using Statistical Package for the Social Sciences (SPSS) software (v.20.0). Results: The order of proliferative activity of experimental groups on L929 mouse fibroblast cells using MTT assay was: metapaste > nCaO > nMgO > nZnO). After evaluation of antimicrobial efficacy, group I: nCaO showed maximum ZOI and MIC against E. faecalis, which showed high statistically significant differences between all four groups after ANOVA (p < 0.0001*). Conclusion: n calcium oxide (CaO) mixed with propylene glycol (PPG) 400 has a potential role as an intracanal medicament with minimum cytotoxic effect and maximum antimicrobial activity against endodontic pathogens. Clinical significance: Nanoparticles-based intracanal medicament can provide a promising future in reducing endodontic flareups when used as intracanal medicament. How to cite this article: Barge P, Gugawad S, Devendrappa SN, et al. Comparative Evaluation of Nano Inorganic Metal Oxides as Intracanal Medicaments for Cytotoxicity and Antimicrobial Activity in the Root Canal System. Int J Clin Pediatr Dent 2023;16(S-2):S168-S175.

Trimethylamine increases intestinal fatty acid absorption: in vitro studies in a Caco-2 cell culture system.

Although elevated blood levels of trimethylamine N-oxide (TMAO) have been associated with atherosclerosis development in humans, the role of its gut microbiota-derived precursor, TMA, in this process has not been yet deciphered. Taking this into account, and the fact that increased intestinal fatty acid absorption contributes to atherosclerosis onset and progression, this study aimed to evaluate the effect of TMA on fatty acid absorption in a cell line that mimics human enterocytes. Caco-2 cells were treated with TMA 250 µM for 24 h. Fatty acid absorption was assessed by measuring the apical-to-basolateral transport and the intracellular levels of BODIPY-C12, a fluorescently labelled fatty acid analogue. Gene expression of the main intestinal fatty acid transporters was evaluated by real-time quantitative reverse transcription PCR. Compared to control conditions, TMA increased, in a time-dependent manner and by 20-50 %, the apical-to-basolateral transport and intracellular levels of BODIPY-C12 fatty acid in Caco-2 cells. Fatty acid transport protein 4 (FATP4) and fatty acid translocase (FAT)/CD36 gene expression were not stimulated by TMA, suggesting that TMA-induced increase in fatty acid transport may be mediated by an increase in FAT/CD36 and/or FATP4 activity and/or fatty acid passive transport. This study demonstrated that TMA increases the intestinal absorption of fatty acids. Future studies are necessary to confirm if this may constitute a novel mechanism that partially explains the existing positive association between the consumption of a diet rich in TMA sources (e.g. red meat) and the increased risk of atherosclerotic diseases.

Development and validation of a simple method for the determination of triamcinolone acetonide in nasal spray.

A rapid, convenient, and sensitive analytical technique for quantitative analysis of triamcinolone acetonide (TAC) in pharmaceutical nasal spray dosage form using the blue tetrazolium colorimetric reaction and UV spectrophotometric method was developed and validated. Beer's law of the developed method was proven in the concentration range of 10-40 µg/mL and showed a specific linear relationship with coefficient value R2 = 0.998. The LOQ level was 9.99 µg/mL, with (RSD = 0.26%). From precision assay, RSD values have been obtained for the repeatability and intermediate precision, which were found to be (RSD = 1.65%) and (RSD = 2.01%), respectively, indicating that the method is reproducible. Recovery studies showed mean recoveries in the range of (100.08-103.65 %), meeting the acceptance criteria for accuracy. In addition, we compared the results of the developed method UV-Vis spectrophotometric procedure with those of a well-established official USP analytical procedure (HPLC), and the results showed good agreement. The proposed UV method represents a potential alternative to the official USP analytical assay procedure (HPLC) for estimating TAC in nasal spray forms. Furthermore, it has the potential to be implemented in routine use for rapid qualitative and quantitative determinations for TAC.

A Rapid and Quantitative Method for Determining Seed Viability Using 2,3,5-Triphenyl Tetrazolium Chloride (TTC): With the Example of Wheat Seed.

Current colorimetric methods for quantitative determination of seed viability (SV) with 2,3,5-triphenyl tetrazolium chloride (TTC) have been plagued by issues of being cumbersome and time-consuming during the experimental process, slow in extraction and staining, and exhibiting inconsistent results. In this work, we introduced a new approach that combines TTC-staining with high-temperature extraction using dimethyl sulfoxide (DMSO). The optimization of the germination stage, TTC-staining method, and 1,3,5-triphenylformazan (TTF) extraction method were meticulously carried out as follows: When the majority of wheat seeds had grown the radicle, and the length of radicles was approximately equal to the seed length (24 h-germination), 2 g germinating seeds were placed into a beaker (20 mL) containing 5 mL 10 g·L-1 TTC solution. The seeds were stained with TTC in the dark at 25 °C for 1 h. Following the staining, 1 mL 1 mol·L-1 H2SO4 was added to stop the reaction for 5 min. The H2SO4 solution was then removed, and the seeds were gently rinsed with deionized water. Subsequently, the TTF produced in the seeds was extracted directly with 5 mL DMSO solution at 55 °C for 1 h. The absorbance of the extract was measured at 483 nm, and the index of SV was calculated according to a predetermined TTC calibration curve and expressed by mg TTC·g-1 (seed)·h-1. The new method has been demonstrated to be rapid, stable, and highly sensitive, as evidenced by the accurate measurement of seed viability with different aging degrees.

Extractive Spectrophotometric Determination and Theoretical Investigations of Two New Vanadium(V) Complexes.

Two new vanadium (V) complexes involving 6-hexyl-4-(2-thiazolylazo)resorcinol (HTAR) and tetrazolium cation were studied. The following commercially available tetrazolium salts were used as the cation source: tetrazolium red (2,3,5-triphenyltetrazol-2-ium;chloride, TTC) and neotetrazolium chloride (2-[4-[4-(3,5-diphenyltetrazol-2-ium-2-yl)phenyl]phenyl]-3,5-diphenyltetrazol-2-ium;dichloride, NTC). The cations (abbreviated as TT+ and NTC+) impart high hydrophobicity to the ternary complexes, allowing vanadium to be easily extracted and preconcentrated in one step. The complexes have different stoichiometry. The V(V)-HTAR-TTC complex dimerizes in the organic phase (chloroform) and can be represented by the formula [(TT+)[VO2(HTAR)]]2. The other complex is monomeric (NTC+)[VO2(HTAR)]. The cation has a +1 charge because one of the two chloride ions remains undissociated: NTC+ = (NT2+Cl-)+. The ground-state equilibrium geometries of the constituent cations and final complexes were optimized at the B3LYP and HF levels of theory. The dimer [(TT+)[VO2(HTAR)]]2 is more suitable for practical applications due to its better extraction characteristics and wider pH interval of formation and extraction. It was used for cheap and reliable extraction-spectrophotometric determination of V(V) traces in real samples. The absorption maximum, molar absorptivity coefficient, limit of detection, and linear working range were 549 nm, 5.2 × 104 L mol-1 cm-1, 4.6 ng mL-1, and 0.015-2.0 µg mL-1, respectively.

Tetrazolium-based colorimetric assays underestimat the direct antitumor effects of anti-VEGF agent bevacizumab.

The direct antitumor effect of bevacizumab (BEV) has long been debated. Assessment of the direct cytotoxic activities of drugs is usually conducted via in vitro experiments, of which tetrazolium-based colorimetric assays are widely employed to measure the direct antitumor activity of BEV. This study aimed to investigate whether tetrazolium-based colorimetric assays are applicable when evaluating the cytotoxicity of BEV against tumor cells. Our results showed that BEV significantly augmented tumor-cell mitochondrial metabolism. Enhanced mitochondrial metabolism caused changes in cellular oxidation-and-reduction environment and upregulated succinate dehydrogenase, which in turn promoted the reduction of tetrazolium to produce formazan. Increased formazan formation resulted in underestimation of the in vitro direct antitumor effect of BEV. Furthermore, inhibition of mitochondrial hypermetabolism partially corrected the underestimation of colorimetric assays in evaluating the direct antitumor activity of BEV. Our findings suggest that tetrazolium-based colorimetric assays are unsuitable for accurately assessing the in vitro cytotoxicity of anti-VEGF drugs and may be the methodological reason for the controversial direct antitumor effect of BEV.

Gold Nanoparticles Functionalized with 2-Thiouracil for Antiproliferative and Photothermal Therapies in Breast Cancer Cells.

Nanoparticles have been used to transport drugs to various body parts to treat cancer. Our interest is in gold nanoparticles (AuNPs) since they have the capacity to absorb light and convert it to heat, inducing cellular damage. This property is known as photothermal therapy (PTT) and has been studied in cancer treatment. In the present study, biocompatible citrate-reduced AuNPs were functionalized with a biologically active compound, 2-thiouracil (2-TU), of potential anticancer activity. Both the unfunctionalized (AuNPs) and functionalized (2-TU-AuNPs) were purified and characterized by UV-Vis absorption spectrophotometry, Zeta potential, and Transmission Electron Microscopy. Results showed monodispersed, spherical AuNPs with a mean core diameter of 20 ± 2 nm, a surface charge of -38 ± 5 mV, and a localized surface plasmon resonance peak at 520 nm. As a result of functionalization, the mean core diameter of 2-TU-AuNPs increased to 24 ± 4 nm, and the surface charge increased to -14 ± 1 mV. The functionalization of AuNPs and the load efficiency were further established through Raman spectroscopy and UV-Vis absorption spectrophotometry. The antiproliferative activities of AuNPs, 2-TU and 2-TU-AuNPs were examined by a 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay in the MDA-MB-231 breast cancer cell line. It was established that AuNPs significantly enhanced the antiproliferative activity of 2-TU. Furthermore, the irradiation of the samples with visible light at 520 nm decreased the half-maximal inhibitory concentration by a factor of 2. Thus, the 2-TU drug concentration and its side effect during treatments could be significantly reduced by synergistically exploiting the antiproliferative activity of 2-TU loaded onto AuNPs and the PTT effect of AuNPs.

Optimization of the Dose Rate Effect in Tetrazolium Gellan Gel Dosimeters.

Tetrazolium salts provide an appealing candidate for 3D gel dosimeters as they exhibit a low intrinsic color, no signal diffusion and excellent chemical stability. However, a previously developed commercial product (the ClearView 3D Dosimeter) based on a tetrazolium salt dispersed within a gellan gum matrix presented a noticeable dose rate effect. The goal of this study was to find out whether ClearView could be reformulated in order to minimize the dose rate effect by optimizing of the tetrazolium salt and gellan gum concentrations and by the addition a thickening agent, ionic crosslinkers, and radical scavengers. To that goal, a multifactorial design of experiments (DOE) was conducted in small-volume samples (4-mL cuvettes). It showed that the dose rate could be effectively minimized without sacrificing the integrity, chemical stability, or dose sensitivity of the dosimeter. The results from the DOE were used to prepare candidate formulations for larger-scale testing in 1-L samples to allow for fine-tuning the dosimeter formulation and conducting more detailed studies. Finally, an optimized formulation was scaled-up to a clinically relevant volume of 2.7 L and tested against a simulated arc treatment delivery with three spherical targets (diameter 3.0 cm), requiring different doses and dose rates. The results showed excellent geometric and dosimetric registration, with a gamma passing rate (at 10% minimum dose threshold) of 99.3% for dose difference and distance to agreement criteria of 3%/2 mm, compared to 95.7% in the previous formulation. This difference may be of clinical importance, as the new formulation may allow the quality assurance of complex treatment plans, relying on a variety of doses and dose rates; thus, expanding the potential practical application of the dosimeter.

Short-chain fatty acids reprogram metabolic profiles with the induction of reactive oxygen species production in human colorectal adenocarcinoma cells.

Short-chain fatty acids (SCFAs) exhibit anticancer activity in cellular and animal models of colon cancer. Acetate, propionate, and butyrate are the three major SCFAs produced from dietary fiber by gut microbiota fermentation and have beneficial effects on human health. Most previous studies on the antitumor mechanisms of SCFAs have focused on specific metabolites or genes involved in antitumor pathways, such as reactive oxygen species (ROS) biosynthesis. In this study, we performed a systematic and unbiased analysis of the effects of acetate, propionate, and butyrate on ROS levels and metabolic and transcriptomic signatures at physiological concentrations in human colorectal adenocarcinoma cells. We observed significantly elevated levels of ROS in the treated cells. Furthermore, significantly regulated signatures were involved in overlapping pathways at metabolic and transcriptomic levels, including ROS response and metabolism, fatty acid transport and metabolism, glucose response and metabolism, mitochondrial transport and respiratory chain complex, one-carbon metabolism, amino acid transport and metabolism, and glutaminolysis, which are directly or indirectly linked to ROS production. Additionally, metabolic and transcriptomic regulation occurred in a SCFAs types-dependent manner, with an increasing degree from acetate to propionate and then to butyrate. This study provides a comprehensive analysis of how SCFAs induce ROS production and modulate metabolic and transcriptomic levels in colon cancer cells, which is vital for understanding the mechanisms of the effects of SCFAs on antitumor activity in colon cancer.

Dried human-cultured epidermis accelerates wound healing in a porcine partial-thickness skin defect model.

Introduction: Autologous cultured epidermis (CE) is an effective approach for overcoming the deficiency of donor sites to treat extensive burns. However, the production of autologous CE takes 3-4 weeks, which prevents its use during the life-threatening period of severe burns. In contrast, allogeneic CE can be prepared in advance and used as a wound dressing, releasing several growth factors stimulating the activity of recipient cells at the application site. Dried CE is prepared by drying CEs under controlled temperature and humidity conditions until all the water is completely removed and no viable cells are present. Dried CE accelerates wound healing in a murine skin defect model and is potentially a new therapeutic strategy. However, the dried CE safety and efficacy have not yet been studied in large animal models. Therefore, we studied the safety and efficacy of human-dried CE in wound healing using a miniature swine model. Methods: Human CE was manufactured using Green's method from donor keratinocytes. Three types of CEs (Fresh, Cryopreserved, and Dried) were prepared, and the ability of each CE to promote keratinocyte proliferation was confirmed in vitro. Extracts of the three CEs were added to keratinocytes seeded in 12-well plates, and cell proliferation was evaluated using the WST-8 assay for 7 days. Next, we prepared a partial-thickness skin defect on the back of a miniature swine and applied three types of human CE to evaluate wound healing promotion. On days 4 and 7, the specimens were harvested for hematoxylin-eosin, AZAN, and anti-CD31 staining to assess epithelialization, granulation tissue, and capillary formation. Results: The conditioned medium containing dried CE extract significantly enhanced keratinocyte proliferation compared to the control group (P < 0.05). In vivo experiments revealed that human-dried CE significantly accelerated epithelialization at day 7 to the same extent as fresh CE, compared to the control group (P < 0.05). The three CE groups similarly affected granulation formation and neovascularization. Conclusions: Dried CE accelerated epithelialization in a porcine partial-thickness skin defect model, suggesting that it may be an effective burn treatment alternative. A clinical study with a long-term follow-up is needed to assess the applicability of CEs in clinics.

Potential Application of the WST-8-mPMS Assay for Rapid Viable Microorganism Detection.

To ensure clean drinking water, viable pathogens in water must be rapidly and efficiently screened. The traditional culture or spread-plate process-the conventional standard for bacterial detection-is laborious, time-consuming, and unsuitable for rapid detection. Therefore, we developed a colorimetric assay for rapid microorganism detection using a metabolism-based approach. The reaction between a viable microorganism and the combination of 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium sodium salt (WST-8) and 1-methoxy-5-methylphenazinium methyl sulfate (mPMS) results in a color change. In combination with a microplate reader, WST-8-mPMS reactivity was leveraged to develop a colorimetric assay for the rapid detection of various bacteria. The detection limit of the WST-8-mPMS assay for both gram-negative and gram-positive bacteria was evaluated. This WST-8-mPMS assay can be used to perform colorimetrical semi-quantitative detection of various bacterial strains in buffers or culture media within 1 h without incubation before the reaction. The easy-to-use, robust, rapid, and sensitive nature of this novel assay demonstrates its potential for practical and medical use for microorganism detection.

Detection of microscopic filamentous fungal biofilms - Choosing the suitable methodology.

Microscopic filamentous fungi are ubiquitous microorganisms that adapt very easily to a variety of environmental conditions. Due to this adaptability, they can colonize a number of various surfaces where they are able to start forming biofilms. Life in the form of biofilms provides them with many benefits (increased resistance to desiccation, UV radiation, antimicrobial compounds, and host immune response). The aim of this study is to find a reliable and reproducible methodology to determine biofilm growth of selected microscopic filamentous fungi strains. Several methods (crystal violet staining, MTT assay, XTT assay, resazurin assay) for the determination of total biofilm biomass and its metabolic activity were tested on four fungi - Alternaria alternata, Aspergillus niger, Fusarium culmorum and Fusarium graminearum, and their biofilm was also imaged by spinning disc confocal microscopy using fluorescent dyes. A reproducible biofilm quantification method is essential for the subsequent testing of the biofilm growth suppression using antifungal agents or physical methods. Crystal violet staining was found to be a suitable method for the determination of total biofilm biomass of selected strains, and the MTT assay for the determination of metabolic activity of the biofilms. Calcofluor white and Nile red fluorescent stains successfully dyed the hyphae of microscopic fungi.

Targeting miR-5088-5p attenuates radioresistance by suppressing Slug.

Radiotherapy is widely used for cancer treatment, but paradoxically, it has been reported that surviving cancer cells can acquire resistance, leading to recurrence or metastasis. Efforts to reduce radioresistance are required to increase the effectiveness of radiotherapy. miRNAs are advantageous as therapeutic agents because it can simultaneously inhibit the expression of several target mRNAs. Therefore, this study discovered miRNA that regulated radioresistance and elucidated its signaling mechanism. Our previous study confirmed that miR-5088-5p was associated with malignancy and metastasis in breast cancer. As a study to clarify the relationship between radiation and miR-5088-5p identified as onco-miRNA, it was confirmed that radiation induced hypomethylation of the promoter of miR-5088-5p and its expression increased. On the other hand, miR-5088-5p inhibitors were confirmed to reduce radiation-induced epithelial-mesenchymal transition, stemness, and metastasis by reducing Slug. Therefore, this study showed the potential of miR-5088-5p inhibitors as therapeutic agents to suppress radioresistance.