N-acetylcysteine attenuates sodium arsenite-induced oxidative stress and apoptosis in embryonic fibroblast cells.

In recent years, the increase in environmental pollutants has been one of the most important factors threatening human and environmental health. Arsenic, a naturally occurring element found in soil, water, and air, easily enters the human body and leads to many metabolic disorders. In this study, we focused on the possible protective effects of N-acetylcysteine (NAC) against sodium arsenite (As)-induced toxic effects on embryonic fibroblast cells. The effects of As and NAC treatment on cells were evaluated, including cytotoxicity, oxidative stress, and apoptosis. Embryonic fibroblast cells were exposed to As (ranging from 0.01 µM to 10 µM) and NAC (at a concentration of 2 mM) for 24 h. The assessment of cytotoxicity markers, such as cell viability and lactate dehydrogenase (LDH), showed that As significantly reduced cell viability and increased LDH levels. Furthermore, we observed that As increased the amount of reactive oxygen species (ROS) in the cell, decreased the activity of antioxidant enzymes, and triggered apoptosis in cells. Additionally, our research revealed that the administration of NAC mitigates the detrimental effects of As. The results showed that As exerted hazardous effects on embryonic fibroblast cells through the induction of oxidative stress and apoptosis. In this context, our study provides evidence that NAC may have a protective effect against the toxicity of As in embryonic fibroblast cells.

The use of hyaluronic acid in a 3D biomimetic scaffold supports spheroid formation and the culture of cancer stem cells.

Three-dimensional (3D) bioprinting culture models capable of reproducing the pathological architecture of diseases are increasingly advancing. In this study, 3D scaffolds were created using extrusion-based bioprinting method with alginate, gelatin, and hyaluronic acid to investigate the effects of hyaluronic acid on the physical properties of the bioscaffold as well as on the formation of liver cancer spheroids. Conformational analysis, rheological characterization, and swelling-degradation tests were performed to characterize the scaffolds. After generating spheroids from hepatocellular carcinoma cells on the 3D scaffolds, cell viability and proliferation assays were performed. Flow cytometry and immunofluorescence microscopy were used into examine the expression of albumin, CD44, and E-cadherin to demonstrate functional capability and maturation levels of the spheroid-forming cells. The results show that hyaluronic acid in the scaffolds correlates with spheroid formation and provides high survival rates. It is also associated with an increase in CD44 expression and a decrease in E-cadherin, while there is no significant change in the albumin expression in the cells. Overall, the findings demonstrate that hyaluronic acid in a 3D hydrogel scaffold supports spheroid formation and may induce stemness. We present a promising 3D scaffold model for enhancing liver cancer spheroid formation and mimicking solid tumors. This model also has the potential for further studies to examine stem cell properties in 3D models.

Evaluation of small-molecule modulators of the circadian clock: promising therapeutic approach to cancer.

The circadian clock is an important regulator of human homeostasis. Circadian rhythms are closely related to cell fate because they are necessary for regulating the cell cycle, cellular proliferation, and apoptosis. Clock dysfunction can result in the development of diseases such as cancer. Although certain tumors have been shown to have a malfunctioning clock, which may affect prognosis or treatment, this has been postulated but not proven in many types of cancer. Recently, important information has emerged about the basic characteristics that underpin the overt circadian rhythm and its influence on physiological outputs. This information implies that the circadian rhythm may be managed by using particular small molecules. Small-molecule clock modulators target clock components or different physiological pathways that influence the clock. Identifying new small-molecule modulators will improve our understanding of critical regulatory nodes in the circadian network and cancer. Pharmacological manipulation of the clock may be valuable for treating cancer. The discoveries of small-molecule clock modulators and their possible application in cancer treatment are examined in this review.

Evaluation of kinetic effects of Gliotoxin in different breast cancer cell lines.

In the present research, the antiproliferative properties of Gliotoxin, which is obtained from marine fungus and thought to be a promising metabolite, on MCF-7 and MDA-MB-231 breast cancer cells, which have different molecular subtypes, were evaluated. Different cell kinetic parameters were employed for this aim. In experiments, cell viability, cell index, mitotic index, BrdU labeling index, and apoptotic index were assessed. Gliotoxin concentrations of 1.5625 µM, 3.125 µM, and 6.25 µM were used in studies for both cell lines. As a result of the values obtained from cell viability and xCELLigence Real-Time Cell Analysis (RTCA) System, 1.5625 µM concentration was determined as IC50 dose. This concentration was applied to all other parameters and anticancer activities were observed.

Anthocyanin Addition to Kefir: Metagenomic Analysis of Microbial Community Structure.

The addition of anthocyanin to kefir for the production of more functional and bio-diversified kefir beverages has the potential to increase kefir's healthful activities. In the present study, anthocyanin extracts, obtained from black carrots, were added into kefir mixture during the fermentation process in different concentrations (1% and 5%, w/v). These kefir samples were then analyzed in terms of their microbiological qualities by metagenomic analysis. The results of the analyses show that the addition of anthocyanin has significant impacts on the community structure of kefir microbiome which in turn directly affects the expected health impacts of the beverage. Kefir with no anthocyanin included predominantly probiotic bacteria such as Lactococcus lactis (34%) and Lactobacillus kefiri (34%). On the other hand, kefir with 1% anthocyanin demonstrated a more balanced distribution of probiotic species like Lb. kefiri (17%), Leuconostoc mesenteroides (9%), and Lc. lactis (5%) at similar abundance rates. 5% anthocyanin kefir demonstrated the highest polarity in the community with a strong dominance of probiotic Lb. kefiri (72%), and distinctly less abundant bacteria such as Streptococcus salivarius subsp. thermophilus (3%). These findings provide that fortification with anthocyanins can be utilized to enhance the quality, composition, and beneficial functions of kefir.

Inhibitory activity of soybean (Glycine max L. Merr.) Cell Culture Extract on tyrosinase activity and melanin formation in alpha-melanocyte stimulating Hormone-Induced B16-F10 melanoma cells.

BACKGROUND: Hyperpigmentation, which causes excessive melanin synthesis and accumulation, is an important issue in the cosmetic industry. Since compounds developed against hyperpigmentation often come with side effects such as skin irritation and contact dermatitis, new studies focus on the use of natural agents that have no side effects. METHODS AND RESULTS: In this study, it was found that the effects of soybean cell culture extract (SCE) on alpha-melanocyte-stimulating hormone (α-MSH) induced melanogenesis in B16-F10 murine melanoma cells. The cells were incubated with SCE for 48 h after treatment with α­MSH for 24 h to analysis the melanin content, cellular tyrosinase activity, and gene and protein expression. SCE at 1 mg/mL decreased melanin content and tyrosinase activity by 34% and 24%, respectively, compared to the α-MSH-treated group, which did not decrease cell viability. In addition, SCE (1 mg/mL) downregulated the expression of tyrosinase (TYR), tyrosinase-related protein (TRP)-1, tyrosinase-related protein (TRP)-2, and microphthalmia-associated transcription factor (MITF) genes 1.5-, 1.5-, 2-, and 2-fold, respectively. Furthermore, SCE inhibited the expression of TYR, TRP1, and TRP2 by decreasing the expression of MITF, as shown in a western blot assay. CONCLUSIONS: This study suggests that SCE reveals dose-dependent inhibition of melanin synthesis through the suppression of tyrosinase activity as well as molecular levels of TYR, TRP1, TRP2, and MITF in B16-F10 murine melanoma cells. Therefore, SCE has the potential to be an effective and natural skin-whitening agent for application in the cosmetic industry.

Bacteriophage cocktail as a promising bio-enhancer for methanogenic activities in anaerobic membrane bioreactors.

This study aimed to explore the effect of a bacteriophage cocktail, pyophage, on the treatment of wastewater containing antibiotics in an anaerobic membrane bioreactor (AnMBR). During the operational period, performance of the AnMBR was monitored through the changes in chemical oxygen demand (COD), antibiotic removal, transmembrane pressure, and biogas production. Microbial community structure and composition, as well as the occurrence of antibiotic resistance genes were analyzed through shotgun metagenomics analysis. When exposed to pyophage, COD removal efficiency was enhanced up to 96%, whereas membrane fouling was delayed by 25%. Average biogas production was doubled from 224.2 mL/d in control with antibiotics to 447.3 mL/d when exposed to pyophage cocktail with considerable alterations to the archaeal and bacterial community structures. Most notably, the methanogenic community shifted from dominance of Methanothermobacter to Methanoculleus, along with syntrophic bacteria. The results provide insight into the synergistic effects of phage-bacteria and methanogenic communities and illustrate the potential of bacteriophages as bio-enhancers.

Glu-370 in the large subunit influences the substrate binding, allosteric, and heat stability properties of potato ADP-glucose pyrophosphorylase.

ADP-glucose pyrophosphorylase (AGPase) is a key allosteric enzyme in plant starch biosynthesis. Plant AGPase is a heterotetrameric enzyme that consists of large (LS) and small subunits (SS), which are encoded by two different genes. In this study, we showed that the conversion of Glu to Gly at position 370 in the LS of AGPase alters the heterotetrameric stability along with the binding properties of substrate and effectors of the enzyme. Kinetic analyses revealed that the affinity of the LSE370GSSWT AGPase for glucose-1-phosphate is 3-fold less than for wild type (WT) AGPase. Additionally, the LSE370GSSWT AGPase requires 3-fold more 3-phosphogyceric acid to be activated. Finally, the LSE370GSSWTAGPase is less heat stable compared with the WT AGPase. Computational analysis of the mutant Gly-370 in the 3D modeled LS AGPase showed that this residue changes charge distribution of the surface and thus affect stability of the LS AGPase and overall heat stability of the heterotetrameric AGPase. In summary, our results show that LSE370 intricately modulate the heat stability and enzymatic activity of potato the AGPase.

Isolation, characterization and phylogenetic analysis of halophilic archaea from a salt mine in central Anatolia (Turkey).

The haloarchaeal diversity of a salt mine, a natural cave in central Anatolia, was investigated using convential microbiological and molecular biology methods. Eight halophilic archaeal isolates selected based on their colony morphology and whole cell protein profiles were taxonomically classified on the basis of their morphological, physiological, biochemical properties, polar lipid and protein profiles and 16S rDNA sequences. From the 16S rDNA sequences comparisons it was established that the isolates CH2, CH3 and CHC resembled Halorubrum saccharovorum by 98.8%, 98.9% and 99.5%, respectively. There was a 99.7% similarity between the isolate CH11 and Halobacterium noricense and 99.2% between the isolate CHA1 and Haloarcula argentinensis. The isolate CH8K and CH8B revealed a similarity rate of 99.8% and 99.3% to Halococcus dombrowskii, respectively. It was concluded that the isolates named CH2, CH3 and CHC were clustered in the genus Halorubrum and that CHA1 and CH7 in the genus Haloarcula, CH8K and CH8B in the genus Halococcus and CH11 in the genus Halobacterium.

Antimicrobial and antioxidant activities of various extracts of Verbascum antiochium Boiss. (Scrophulariaceae).

Verbascum antiochium Boiss., a member of the Scrophulariaceae family, is endemic to Turkey. The extracts obtained from V. antiochium by increased polarity and direct methanol extraction were tested by the agar well diffusion method against various Gram-positive and Gram-negative bacteria and one fungus. The methanol/water extract exhibited a larger inhibition zone against both the Gram-negative and Gram-positive bacteria than the other extracts. Haemophilus influenzae was found to be the most sensitive bacterium among the bacteria tested. The antioxidant activities of the methanolic extract of V. antiochium were examined by two complementary test systems. The 50% inhibition activity of the methanolic extract of V. antiochium against the free radical 2,2-diphenyl-1-picrylhydrazyl was determined as 4.80 mg/mL. In the case of the linoleic acid system, oxidation of linoleic acid was inhibited by the methanolic extract of V. antiochium with 79.92% inhibition, which is close to the value of the synthetic antioxidant reagent, tert-butylated hydroxytoluene. The total phenolic components of V. antiochium were determined to be 92.71 mg of gallic acid equivalents/g. Iridoid glycosides, flavonoids, and saponins were detected as the major chemical constituents in the extract.

Effective antibacterial and antioxidant properties of methanolic extract of Laurus nobilis seed oil.

This study was carried out to determine the in vitro antimicrobial and antioxidant activities of the essential oil, seed oil, and methanolic extract of seed oil obtained from Laurus nobilis L. (Lauraceae). The methanolic extract of seed oil exhibited more effective antibacterial activity comparing to essential oil and seed oil, GC-MS analyses of the essential oil resulted in the identification of 25 compounds. 1.8-Cineol (44.72%), a-Terpinyl acetate (12.95%), Sabinene (12.82%) were the main components. The fatty acid composition was characterized with the high content of linoleic acid (40.79%) and lauric acid (38.08%). The 50% (IC50) inhibition activity of the essential oil on the free radical DPPH was determined as 94.655 mg ml(-1), whereas IC50 value of methanolic extract of seed oil was found unstable. In the case of the linoleic acid system, oxidation of linoleic acid was inhibited by essential oil and methanolic extract of seed oil, which showed 64.28 and 88.76% inhibition, respectively. The inhibition value of the methanolic extract of seed oil was quite close to the synthetic antioxidant BHT, 92.46% inhibition.

Characterization of extracellular esterase and lipase activities from five halophilic archaeal strains.

A total of 118 halophilic archaeal collection of strains were screened for lipolytic activity and 18 of them were found positive on Rhodamine agar plates. The selected five isolates were further characterized to determine their optimum esterase and lipase activities at various ranges of salt, temperature and pH. The esterase and lipase activities were determined by the hydrolysis of pNPB and pNPP, respectively. The maximum hydrolytic activities were found in the supernatants of the isolates grown at complex medium with 25% NaCl and 1% gum Arabic. The highest esterase activity was obtained at pH 8-8.5, temperature 60-65 degrees C and NaCl 3-4.5 M. The same parameters for the highest lipase activities were found to be pH 8, temperature 45-65 degrees C and NaCl 3.5-4 M. These results indicate the presence of salt-dependent and temperature-tolerant lipolytic enzymes from halophilic archaeal strains. Kinetic parameters were determined according to Lineweaver-Burk plot. The KM and V (max) values were lower for pNPP hydrolysis than those for pNPB hydrolysis. The results point that the isolates have higher esterase activity comparing to lipase activity.

Comparison of chemical composition of the essential oil of Laurus nobilis L. leaves and fruits from different regions of Hatay, Turkey.

The essential oils of the leaves and fruits from bay (Laurus nobilis L.) grown in Antakya, Yayladagi and Samandagi were isolated by solvent extraction and analysed by capillary gas chromatography (GC), gas chromatography and mass spectrometry (GC/MS). In Antakya, Yayladagi and Samandagi the chemical compositions of the fruits and leaves were similar according to qualitative and quantitative analysis. Although in both fruits and leaves the major component was found to be 1.8-Cineole a concentration of about 50% compared with essential oils. The composition of the essential oil from the leaves has high content of 1.8-Cineole, Sabinene and alpha-Terpinyl acetate, but a low content of a-Pinene, alpha-Phellandrene and trans-/beta-osimen. 1.8-Cineole was found major component of the leaves essential oil collected from Samandagi (59.94%) which is sea coast of region. Interestingly alpha-Pinene, beta-Pinene, alpha-Phellandrene, 1.8-Cineole and trans-beta-osimen were found the major components of fruits of Laurus nobilis L. harvested from Antakya, Yayladagi and Samandagi Trans-beta-osimen was detected as the major component of fruits essential oil collected again from Samandagi (28.35%)

Localization of germin genes and their products in developing wheat coleoptiles.

Germination is a process which characterized with nescient synthesis of genes. Among the genes synthesized during the germination of wheat embryos, germin genes, proteins and their enzymatic activity were defined. Germin is a water soluble homopentameric glycoprotein which is remarkable resistant to degradation by a broad range of proteases including pepsin. Germin proteins found to have strong oxalate oxidase activity which produces hydrogen peroxide by degrading oxalic acid. The current study, aimed to localize the germin genes, proteins and enzymatic activities in developing coleoptiles which is a rapidly growing protective tissue of leaf primordium and shoot apex. Non-radioactively labeled germin riboprobes were employed to localize germin mRNAs in situ. FITC (Fluorescein isothiocyanate) and alkaline phosphatase linked anti-germin antibodies were used to localize germin proteins under the fluorescence and light microscopy and finally germin enzymatic activity was localized by using appropriate enzyme assay. The results revealed that in coleoptiles germin genes, proteins and their enzymatic activity were predominantly associated with the cells of epidermis and vascular bundle sheath cells.

Formation of wheat ( Triticum aestivum L.) embryogenic callus involves peroxide-generating germin-like oxalate oxidase.

In wheat ( Triticum aestivum L.), embryogenic callus formation comprises suppression of precocious germination by the zygotic embryo and the initiation of dedifferentiated cellular proliferation within it. Embryogenic calli are induced by treating immature embryos with 2,4-dichlorophenoxyacetic acid (2,4-D). Upon withdrawal from 2,4-D, somatic embryos develop from the periphery of the callus. Prior to visible callus formation, there is a striking induction of "germin-like" oxalate oxidase ("gl-OXO": EC 1.2.3.4) gene expression. Accumulation of gl-OXO mRNA is rapidly stimulated upon auxin treatment, with a consequent development of apoplastic enzyme activity producing H(2)O(2) within the cell wall. Within the dedifferentiated calli, gl-OXO enzyme activity becomes widespread over the surface of embryogenic calli. Differentiation of somatic embryos is initiated in regions of densely cytoplasmic, meristematic cells that are marked by highly localised expression of gl-OXO activity within these embryogenic cell masses. We suggest that this localised generation of H(2)O(2) by gl-OXO promotes peroxidative cross-linking of cell wall components, thereby preventing cellular expansion and maintaining these cell masses in an embryogenically competent condition.

Isolation and localization of new germination-related sequences from wheat embryos.

Subtractive library hybridization was used to isolate the cDNA clones that corresponded to the transcripts that were specifically up-regulated during wheat embryo germination. The clones with numbers 5, 6, 7, 8, 24, and 26 appeared to be more abundant in germinating wheat embryos. Among the isolated clones, we identified four new members of the wheat "germin" gene family. We also identified two novel sequences which exhibited distinct germination up-regulation, and displayed characteristic spatial patterns of expression. One of these, represented by clone pSB10, was principally expressed in the root tissue of germinating embryos. The second was represented by the pSB7 clone and was expressed in both the root and shoot primordia of the embryonic axis, as well as within the coleoptile.