Blue Tetrazolium Research Articles

Potential Cardioprotective Role of Menaquinone-4 Against Cardiac Ischemia-reperfusion Injury.

Myocardial infarction is among the leading causes of mortality. Menaquinone-4 (MK-4), a vitamin K2 analog, might play a role in rescuing cardiac ischemia/reperfusion (I/R) injury. This work aimed to discover the potential cardioprotective role of MK-4 against myocardial I/R injury in rats. Thirty-two rats were categorized into 3 groups: (I/R) control group: subjected to I/R protocol (received vehicle), MK-4 preconditioning group: MK-4 infusion for 20 minutes before the I/R protocol, and MK-4 postconditioning group: MK-4 infusion for 20 minutes at the start of the reperfusion phase. The hearts were placed in the Langendorff apparatus, and the left ventricular developed pressure (LVDP), heart rate (HR), + (LV dP/dt) max, - (LV dP/dt) max, and Tau were calculated. The necrotic mass was determined by staining it with nitro blue tetrazolium. Creatine kinase-MB (CK-MB), lactate dehydrogenase (LDH), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and C- reactive protein (CRP), as well as cardiac superoxide dismutase (SOD), nitric oxide (NOx), malondialdehyde (MDA), and glutathione (GSH) levels were all evaluated. MK-4 postconditioning significantly reduced myocardial infarct size; increased LVDP, + (LV dp/dt) max, - (LV dp/dt) max, and HR; reduced Tau, CK-MB, LDH, CRP, IL-6, TNF-α, MDA, and NOx levels; and increased SOD activity, whereas no significant difference in the GSH level was detected. In conclusion, these data imply that MK-4 may protect the heart from the consequences of I/R.

Heptadecanoic Acid, an Odd-Chain Fatty Acid, Induces Apoptosis and Enhances Gemcitabine Chemosensitivity in Pancreatic Cancer Cells.

Odd-chain saturated fatty acids generally serve as specific biomarkers of dietary components and dairy intake, some of which have anticancer properties. This study was performed to assess the anticancer effects of heptadecanoic acid (HDNA) in human pancreatic carcinoma cells. MTT (thiazolyl blue tetrazolium bromide) assay showed that HDNA exerted stronger cytotoxic effects than pentadecanoic acid, palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1), and linoleic acid (18:2) on both Panc-1 and MIA PaCa-2 pancreatic cancer cells. In addition, HDNA reduced colony formation and induced apoptosis in these pancreatic cancer cells as indicated by Hoechst 33342 staining, Annexin V/propidium iodide staining, cell cycle analysis, and Western blotting analysis in a dose-dependent manner. Moreover, HDNA synergistically reduced cell viability and promoted apoptosis when combined with gemcitabine (GEM), a chemotherapeutic agent commonly used in the treatment of pancreatic cancer. GEM-resistant MIA PaCa-2 (GR-MIA PaCa-2) cells with a resistance indices (RI) value of 215.09 [RI = half-maximal inhibitory concentration (IC50) of GR-MIA PaCa-2 cells/IC50 of MIA PaCa-2 cells] were established, and the efficacy of HDNA on GEM chemosensitivity was confirmed. Surprisingly, HDNA exhibited even higher antiproliferative efficacy against GR-MIA PaCa-2 cells (IC50 = 71.45 ± 6.37 μM) than parental MIA PaCa-2 cells (IC50 = 77.47 ± 2.10 μM). Finally, HDNA treatment inhibited the Hippo pathway and induced apoptosis of GR-MIA PaCa-2 cells. These findings suggest the beneficial effects of a HDNA-rich diet during pancreatic cancer treatments.

Studies on Pollen Morphology, Pollen Vitality and Preservation Methods of Gleditsia sinensis Lam. (Fabaceae)

Gleditsia sinensis Lam. (Fabaceae) is an endemic species in China, which has a wide range of ecological functions and high economic value. G. sinensis belongs to androdioecy, and the stamens of perfect flowers are aborted, meaning that a perfect flower is a functional female flower. Understanding the dynamic process of flowering and the characteristics of pollen morphology effectively determine the viability of pollen vitality, and the suitable conditions for short-term storage of pollen can provide theoretical basis and technical reference for hybrid breeding and germplasm conservation of G. sinensis. In this study, the male plants of G. sinensis in Guiyang area were used as research materials. The flowering dynamic process of male flowers was recorded through field observation. The morphology of pollen was observed and analyzed with a scanning electron microscope (SEM). The germination characteristics of pollen were studied with an in vitro germination method, and the pollen vitality was also determined using four staining methods. The effects of different storage temperatures and water contents on pollen germination rate were discussed. The results showed that the male flowers of G. sinensis had a short, single flowering period, lasting 2–3 days from the opening to the shedding. The dynamic opening process of a single flower was artificially divided into five stages. Pollen grains of G. sinensis are oblate spheroidal, tricolporate with equatorial elongated endoapertures and the sporoderm surface is reticulate. The MTT (Thiazolyl Blue Tetrazolium Bromide) staining method could accurately and quickly determine the pollen vitality of G. sinensis. The highest pollen germination rate was 65.89% ± 3.41%, and the length of the pollen tube was 3.96 mm after cultured in 15% sucrose + 100 mg/L boric acid + 20 mg/L calcium chloride for 24 h. It was necessary to collect the pollen at the big bud stage, which was conducive to improving the efficiency of pollen collection because the pollen had been mature with high pollen vitality at this stage. When it came to pollen preservation, the pollen germination rate was significantly affected by storage time, storage temperature and pollen water content. The pollen still had high vitality after being stored at −80 °C for 30 days when the moisture content of the pollen decreased to 9%, and the pollen germination rate only decreased by 28.84% compared with that before storage. In conclusion, this study has comprehensively and systematically studied the morphology, vitality determination and preservation methods of the pollen of G. sinensis, providing a theoretical basis for the cross regional breeding and the conservation and utilization of germplasm resources.

MiR-340 mediates the involvement of high mobility group box 1 in the pathogenesis of liver fibrosis

Objective: To explore the pathogenic mechanism of the miR-340/high mobility group box 1 (HMGB1) axis in the formation of liver fibrosis. Methods: A rat liver fibrosis model was established by injecting CCl(4) intraperitoneally. miRNAs targeting and validating HMGB1 were selected with gene microarrays after screening the differentially expressed miRNAs in rats with normal and hepatic fibrosis. The effect of miRNA expressional changes on HMGB1 levels was detected by qPCR. Dual luciferase gene reporter assays (LUC) was used to verify the targeting relationship between miR-340 and HMGB1. The proliferative activity of the hepatic stellate cell line HSC-T6 was detected by thiazolyl blue tetrazolium bromide (MTT) assay after co-transfection of miRNA mimics and HMGB1 overexpression vector, and the expression of extracellular matrix (ECM) proteins type I collagen and α-smooth muscle actin (SMA) was detected by western blot. Statistical analysis was performed by analysis of variance and the LSD-t test. Results: Hematoxylin-eosin and Masson staining results showed that the rat model of liver fibrosis was successfully established. Gene microarray analysis and bioinformatics prediction had detected eight miRNAs possibly targeting HMGB1, and animal model validation had detected miR-340. qPCR detection results showed that miR-340 had inhibited the expression of HMGB1, and a luciferase complementation assay suggested that miR-340 had targeted HMGB1. Functional experiments results showed that HMGB1 overexpression had enhanced cell proliferation activity and the expression of type I collagen and α-SMA, while miR-340 mimics had not only inhibited cell proliferation activity and the expression of HMGB1, type I collagen, and α-SMA, but also partially reversed the promoting effect of HMGB1 on cell proliferation and ECM synthesis. Conclusion: miR-340 targets HMGB1 to inhibit the proliferation and ECM deposition in hepatic stellate cells and plays a protective role during the process of liver fibrosis.

Collagen-coated agarose-chitosan scaffold used as a skin substitute.

A single biomaterial is disadvantageous for constructing skin in vitro, so a mixed biomaterial is more conducive to skin research. In this study, agarose-chitosan scaffolds with a final concentration of 4% were constructed by freeze-drying, in which the concentration ratios of agarose to chitosan were 1:3, 2:2, and 3:1. The scaffolds were coated with a 3mg/ml collagen solution, and the mechanical properties were evaluated by studying density, porosity, swelling rate, and degradation rate. The results demonstrated that the agarose-chitosan scaffolds were porous, with porosity reaching 93%. Their densities ranged from 0.1 to 0.16g/cm3 . Analysis of Young's modulus showed that the mechanical properties of the agarose-chitosan scaffolds were significantly enhanced when the agarose content in the agarose-chitosan scaffolds was increased. Moreover, the density and Young's modulus of the agarose-chitosan scaffolds of different concentration ratios were significantly different (p < 0.01). These scaffolds can withstand a certain amount of external pressure, such as that of human skin, making them more suitable for further skin replacement research. In addition, the results of thiazolyl blue tetrazolium bromide (MTT)cell assay and immunofluorescence staining showed that the collagen-coated agarose-chitosan scaffolds were conducive to keratinocyte proliferation and differentiation. The MTT results revealed significant differences between the agarose-chitosan scaffolds coated with collagen and the agarose-chitosan scaffolds without collagen (p < 0.05). This study provides the potential for in vitro skin research and applications.

EFFECT OF MODULATION OF NITROGEN OXIDE (II) SYNTHESIS UNDER CHRONIC NORMOBARIC HYPOXIA ON THE ISOENZYME SPECTRUM OF RAT EPIDIDIMIS LACTATE DEHYDROGENASE

Lactate dehydrogenase have a key role in providing energy to cells under physiological and hypoxic conditions. Changes in the activity of the enzyme can be facilitated by nitric oxide (II), regulating the expression of individual fractions of lactate dehydrogenase. Aim. To study changes in the isoenzyme spectrum of rat epididymis lactate dehydrogenase under conditions of hypoxia and modulation of nitric oxide (II) synthesis. Material and methods. Male rats (32) were divided into 4 groups (n=8): 1) chronic normobaric hypoxia; 2) control to group 1; 3) hypoxia together with modeling of nitric oxide deficiency; 4) hypoxia against the background of induction of NO synthesis. The mitochondrial fraction and non-mitochondrial cytoplasm of the head and tail of the epididymis were isolated for laboratory research. The zymogram was obtained by electrophoresis in 7% gel followed by detection by the precipitation reaction of the reduction of nitro blue tetrazolium. The overall activity of lactate dehydrogenase was assessed and the percentage of isoenzyme fractions was calculated. Fractions 1 and X of lactate dehydrogenase were analyzed, the results were con-sidered statistically significant if p 0.05 when comparing two independent samples, and p 0.0167 when comparing three independent samples. Results. Chronic normobaric hypoxia leads to a decrease in lactate dehydrogenase activity compared to the control group. At the same time, the share of the X-fraction of the enzyme and lactate dehydrogenase 1 increases. Modulation of nitric oxide (II) deficiency against the background of hy-poxia increased the activity of the enzyme in the head of the epididymis, where a decrease in the studied fractions of lactate dehydrogenase was ob-served compared to the group of animals subjected only to hypoxia. Stimulation of the synthesis of nitric oxide (II) during hypoxia led to a decrease in the X-fraction, but an increase in the 1st fraction of lactate dehydrogenase in the head of the epididymis, in the tail of the epididymis, opposite changes were observed. Conclusion. Hypoxia causes a change in the ratio of lactate dehydrogenase isoenzymes towards an increase in lactate dehydrogenase B and lactate dehydrogenase X, while a deficiency of nitric oxide (II) contributes to a decrease in lactate dehydrogenase X synthesis.

In vitro investigation of the antibacterial and anti-inflammatory effects of LED irradiation.

This study aimed to investigate the proper wavelengths for safe levels of light-emitting diode (LED) irradiation with bactericidal and photobiomodulation effects in vitro. Cell viability tests of fibroblasts and osteoblasts after LED irradiation at 470, 525, 590, 630, and 850 nm were performed using the thiazolyl blue tetrazolium bromide assay. The bactericidal effect of 470-nm LED irradiation was analyzed with Streptococcus gordonii, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, and Tannerella forsythia. Levels of nitric oxide, a proinflammatory mediator, were measured to identify the anti-inflammatory effect of LED irradiation on lipopolysaccharide-stimulated inflammation in RAW 264.7 macrophages. LED irradiation at wavelengths of 470, 525, 590, 630, and 850 nm showed no cytotoxic effect on fibroblasts and osteoblasts. LED irradiation at 630 and 850 nm led to fibroblast proliferation compared to no LED irradiation. LED irradiation at 470 nm resulted in bactericidal effects on S. gordonii, A. actinomycetemcomitans, F. nucleatum, P. gingivalis, and T. forsythia. Lipopolysaccharide (LPS)-induced RAW 264.7 inflammation was reduced by irradiation with 525-nm LED before LPS treatment and irradiation with 630-nm LED after LPS treatment; however, the effects were limited. LED irradiation at 470 nm showed bactericidal effects, while LED irradiation at 525 and 630 nm showed preventive and treatment effects on LPS-induced RAW 264.7 inflammation. The application of LED irradiation has potential as an adjuvant in periodontal therapy, although further investigations should be performed in vivo.

Hydroxytyrosol and dopamine metabolites: Anti-aggregative effect and neuroprotective activity against α-synuclein-induced toxicity.

The abnormal aggregation of the α-synuclein (αsyn) protein is involved in the formation of Lewy bodies in the brain of patients suffering from Parkinson disease (PD). Hydroxytyrosol (HT), a polyphenolic compound present in olives, olive oil, and wine, has been shown to inhibit aggregation and destabilise the αsyn aggregates, preventing neuronal cell death. However, very limited data have been published on the study of its metabolites. Therefore, this study investigated the capacity of the metabolites 3,4-dihydroxyphenylacetaldehyde (DOPAL), 4-hydroxy-3-methoxyphenylethanol (MOPET), and 3-methoxy-4-hydroxyphenylacetaldehyde (MOPAL) to prevent the aggregation and toxic effects of αsyn fibrils. In vitro techniques, such as Thioflavin T (ThT), Transmission Electronic Microscopy (TEM), electrophoresis, thiazolyl blue tetrazolium bromide (MTT), and Real-Time PCR (RT-PCR) were used. Our results show that among these three metabolites, DOPAL exerts the greatest effect, preventing aggregation and αsyn-induced neurotoxicity. In fact, DOPAL has the ability to completely inhibit αsyn fibril formation at low doses. Moreover, this metabolite has a potent destabilising effect on the αsyn fibrils. Concerning neuroprotection, DOPAL can counteract the toxicity induced by αsyn. The vitagene expression results show a possible relationship between the neuroprotection mechanism exhibited by DOPAL and the modulation of SIRT-2 and Hsp70.

A Sensitive Spectrophotometric Determination of Silver (I) with Blue Tetrazolium Chloride

Silver(I)-assisted reduction of Blue tetrazolium chloride (3,3′-(3,3′-dimethoxy[1,1′-biphenyl]-4,4′-diyl)-bis(2,5-diphenyl-2H-tetrazolium) dichloride, BTC) by semicarbazide hydrochloride (SCH) was investigated in water-chloroform medium. The obtained colored products can be used for the visual detection and sensitive liquid-liquid extraction–spectrophotometric determination of Ag(I). Under the optimum conditions the calibration curve (which can be best approximated by a third-order polynomial; R2=0.9990) has two linear segments. For Ag(I) concentrations up to 0.011 g/ml, the linear regression equation had an intercept that was statistically indistinguishable from zero. The limit of detection, limit of quantitation, and molar absorptivity coefficient at λmax=573 nm were 0.6 ng/ml, 2 ng/ml and 1.2and#215;106 l/(molcm), respectively. The regression equation of the second linear segment (0.021–0.028 g/ml) was A = 200γ – 3.5 (R2=0.9989), where γ is the concentration in μg/ml. The relative standard deviation at the 22 ng/ml level (n = 5) was 3.8 %. The effect of concomitant ions was studied, and the analysis of real samples tested the applicability of the developed procedure.

Homo- and heterometallic complexes of Zn(II), {Zn(II)Au(I)}, and {Zn(II)Ag(I)} with pentadentate Schiff base ligands as promising anticancer agents.

Two families of homo- and heterometallic complexes, [Zn2L1(μ-OH)(H2O)2](ClO4)2, [Zn2L2(μ-OH)(H2O)2](ClO4)2, [Zn2L3(μ-OH)(H2O)2](ClO4)2, 1∞[{L1Zn2(μ-OH)}{μ-[Ag(CN)2]}](ClO4), [{L1Zn2(μ-OH)}2{μ-[Au(CN)2]}{[Au(CN)2]2}](ClO4)·H2O, 1∞[{L2Zn2(μ3-OH)}2(H2O){μ-[Ag(CN)2]}](ClO4)3·THF·0.5MeOH, 1∞[{L2Zn2(μ3-OH)}2(H2O){μ-[Au(CN)2]}](ClO4)3·THF·H2O, and 1∞[{L3Zn2(μ-OH)}{μ-[Ag(CN)2]}][Ag(CN)2]·H2O, respectively, have been synthesized and characterized. The Schiff bases used as ligands were obtained by condensation reactions of 2,6-diformyl-p-cresol with N,N-dimethyl-ethylenediamine (HL1), 2-aminomethyl-pyridine (HL2), and 2-aminoethyl-pyridine (HL3), respectively. The cytotoxic/cytostatic and genotoxic effects in cultured human MCF-7 (luminal type A breast cancer), MDA-MB-231 (triple negative breast cancer), HeLa (cervical carcinoma), and Lep-3 (non-tumor embryonal fibroblastoid cells) were studied. The investigations were performed by thiazolyl blue tetrazolium bromide test (MTT test), neutral red uptake cytotoxicity assay, crystal violet staining, hematoxylin and eosin staining, double staining with acridine orange and propidium iodide, AnnexinV/FITC, and Comet assay in short-term experiments (24-72 h, with monolayer cell cultures) as well as by 3D colony-forming method in long-term experiments (28 days, with 3D cancer cell colonies). The results obtained revealed that: (i) applied at a concentration range of 0.1-100 μg mL-1, the compounds investigated decrease in a time- and concentration-dependent manner the viability and/or proliferation of the treated cells; (ii) complexes of {Zn(II)Au(I)} show relatively higher cytotoxic/genotoxic activity and antitumor potential as compared to {Zn(II)Ag(I)}; (iii) some of the complexes demonstrate more pronounced cytotoxic potential than commercially available antitumor agents cisplatin, oxaliplatin, and epirubicin.

The effect of antibacterial agents of composite resin materials in dental caries prevention and reduction of flexural strength of the material: Systematic review and meta-analysis

Background: The present study aimed to evaluate the antibacterial agents of composite resin materials, their efficacy in caries prevention, and their impact on the mechanical properties (flexural strength) of composite resin restorative materials. Methods: In this systematic review and meta-analysis, all published articles on the effects of antibacterial properties of composite resin materials in the prevention of dental caries (between 2005 to 2020) were evaluated using valid databases, including PubMed, Google Scholar, Web of Science, ISI, Scopus, Cochrane, ProQuest, and Embase, found by searching the keywords "anti-bacterial agents," "composite resins," "dental restoration," and "dental caries" according to the MeSH system. The context of the studies was extracted and subjected to modified CONSORT. The required data were extracted and analyzed with the Comprehensive Meta-Analysis software. Results: According to the results of the random effect model, the heterogeneity of biofilm colony forming unit (CFU) average was -1.90 (CI: -1.68 to -2.13). The mean value of the control group was higher than the intervention. The heterogeneity of flexural strength (FS) average was -11.92 (CI: -8.30 to -15.55). The mean value of the control group was higher than the intervention group. The heterogeneity of thiazolyl blue tetrazolium bromide (MTT) absorbance average was -0.90 (CI: -0.65 to -1.14). The mean value of the control group was higher than the intervention group. Conclusion: The results of the present study showed that the antibacterial agents had a significant effect on the caries prevention properties of the composite resin materials; however, due to the bias related to different control groups, laboratory conditions, and mismatch between in vitro and in vivo conditions, more studies are needed in this regard.

Investigating the mechanism of interactive regulation of B-cell lymphoma-2/Beclin 1 through electroacupuncture intervention during reperfusion in myocardial ischemia-reperfusion injury in a rat model.

To observe the regulation of B-cell lymphoma-2 (Bcl-2)/Beclin 1 interaction through electroacupuncture (EA) intervention during reperfusion and to investigate the EA mechanism of apoptosis-autophagy interactive regulation against myocardial ischemia-reperfusion injury (MIRI). A total of 48 adult Sprague Dawley (SD) rats were randomly divided into the sham-operated group (group Sham), the model group (group Model), the EA group (group EA), and the JNK inhibitor (SP600125) group (group JNK), with 12 rats in each group. Biospecimens were collected randomly from six rats in each group four hours after reperfusion. Evans Blue and triphenyl tetrazolium chloride double-staining were applied to observe each group's myocardial damage area and risk area. We collected 4 ml of blood by abdominal aortic method to detect serum troponin cTnI level by enzyme-linked immunosorbent assay (ELISA). For the remaining six in each group, a part of myocardial tissue below the ligation line was stored in 4% paraformaldehyde for immunohistochemistry and TUNEL staining; the other amount of myocardial tissue was detected by Western blotting to determine the expression levels of Bcl-2, Beclin1, and the phosphorylation levels of Thr69, Ser70, and Ser87 in Bcl-2. In results: electroacupuncture (EA) intervention during reperfusion significantly reduced the myocardial infarction area, cTnI level, and myocardial apoptosis, upregulated Bcl-2 expression, downregulated Beclin 1 expression and inhibited phosphorylation levels of Thr69, Ser70, and Ser87 in Bcl-2. We concluded that EA effectively inhibited apoptosis by upregulating Bcl-2 expression and inhibiting the phosphorylation of Thr69, Ser70, and Ser87 in Bcl-2. This reduced the separation of Bcl-2 and Beclin 1, restrains excessive autophagy, alleviates MIRI, and has a protective effect on myocardial tissue.

Telangiectatic Osteosarcoma of the Tibiotarsus in a North Island Brown Kiwi (Apteryx mantelli).

A North Island brown kiwi (Apteryx mantelli) with lameness and weight loss had a telangiectatic osteosarcoma. The left proximal tibiotarsus had bony lysis with multiple blood-filled spaces separated by thick fibrous septa and neoplastic mesenchymal cells producing osteoid (5-bromo-4-chloro-3'-indolyl phosphate/nitro blue tetrazolium positive on cytology). No metastases were noted on necropsy.

MEGAMOX MEDIATES ECO-FRIENDLY SYNTHESIS OF SELENIUM NANOPARTICLES AS IMMUNOMODULATORY, ANTIOXIDANT AND ANTIMICROBIAL AGENTS

The research herein includes methodology for eco-friendly preparation of selenium nanoparticles (SeNPs) using the broad-spectrum antibiotic, Megamox (Mega.). Characterization of SeNPs was done using UV-Visible spectroscopy, FTIR, XRD, DLS and TEM imaging. Additionally, the immunomodulatory, antioxidant and antimicrobial activities of both SeNPs and Mega. were checked against different strains of bacteria and fungi. TEM and DLS images showed that SeNPs are polydisperse spheres with mean diameter of 22.4 nm. FTIR analysis indicated that the hydroxyl and nitrogen moieties in Mega. were effective for reduction plus binding manner. According to the results of the nitro blue tetrazolium reduction test, both SeNPs and Mega. presented high intracellular killing activity, which confirmed their immunostimulatory effect. Antioxidant activity of SeNPs and Mega. were 90 and 82 %, respectively. SeNPs presented great activity facing multi-drug resistant bacteria and TB. SeNPs are considered promising cost-effective and eco-friendly anti-bacterial and anti-fungal agents that can represent a new potential nano-platform in both medical and infectious diseases control.

Immune Disorders in Patients with Pulmonary Tuberculosis with Primary and Acquired Drug-Resistance of &lt;i&gt;Mycobacterium Tuberculosis&lt;/i&gt;

The objective: to evaluate main parameters of cellular, humoral immunity and natural resistance in tuberculosis patients with primary and acquired multiple drug resistance; to study the correlation of these parameters with clinical and radiological manifestations.Subjects and Methods. A prospective study was conducted which included 169 patients with pulmonary tuberculosis, 80 of them were new cases of drug susceptible pulmonary tuberculosis and they made the Comparison Group (CG), and 89 patients suffered from multiple drug resistant tuberculosis and they made the Main Group (MG). The Main Group was divided into 2 subgroups: MG-1 – 40 patients with primary multiple drug resistant tuberculosis; MG-2 ‒ 49 patients who acquired drug resistance of Mycobacterium tuberculosis during treatment of tuberculosis.The immune assays included the lymphocyte blast transformation reaction (LBTR) to phytohemagglutinin and tuberculin (PPD), immunophenotyping of CD3+ and CD19+ cells; phagocytic index and phagocytic number were calculated; nitro blue tetrazolium test was used, concentrations of immunoglobulins and antimycobacterial antibodies were determined.Results. The clinical laboratory data demonstrated a more severe course of the disease in patients with multiple drug resistant tuberculosis, especially in those with acquired multiple drug resistant tuberculosis which was confirmed by the severity of immune deficiencies of the cellular immunity and innate resistance as well as by the overactivity of humoral immunity. The levels of CD3+ and LBTR reaction to PPD had a negative correlation with clinical manifestations and radiological features demonstrating the severity of pulmonary tuberculosis in patients with multiple drug resistance, especially in those with acquired multiple drug resistance; and CD19+ and antimycobacterial antibodies positively correlated with clinical manifestations and radiological signs.Conclusions. Parameters of cellular immunity and natural resistance were the lowest in the patients with acquired multiple drug resistance of Mycobacterium tuberculosis versus primary multiple drug resistance and drug susceptible Mycobacterium tuberculosis. Parameters of humoral immunity demonstrated the increased activity to compensate for the insufficiency of cellular immunity regardless of the type of resistance of Mycobacterium tuberculosis.

Phenolic from apple blossom "Hongro" inhibits the expression of proteins related to melanogenesis in B16F10 melanoma cells.

This study aimed to assess apple blossom extracts as potential natural whitening agents due to their ability to inhibit melanogenesis. Ethanol extracts of apple blossom (ABE) were assessed for biological activity in the B16F10 mouse melanoma cell line. ABE toxicity was assessed by thiazolyl blue tetrazolium bromide (MTT) assay. Levels of melanogenic enzyme expression in response to ABE supplementation were assessed by western blotting. Also assessed purified kaempferol, one of the phenolic compounds extracted from apple blossom, was evaluated using western blot analysis. The expression levels of cellular tyrosinase, microphthalmia-associated transcription factor (MITF), tyrosinase-related protein (TRP)-1, and TRP-2 proteins related to melanogenesis decreased in a dose-dependent manner with ABE treatment of cells. Using nuclear magnetic resonance, we identified kaempferol in the ABE. Treatment of cells with purified kaempferol decreased the expression levels of tyrosinase and the MITF protein to a similar degree as that observed with ABE treatment. This suggests that the efficacy of melanogenesis-related inhibition demonstrated by ABE was due to kaempferol. ABE has an inhibitive effect on melanogenic enzymes and potentially can be applied to functional foods and cosmetics having a whitening effect as a natural material.

Interaction of Cyprinus carpio Linnaeus with the biofilm-forming Aeromonas hydrophila.

The resistance of susceptible fish populations and the adaptive potential of heterogeneous biofilms, which cause multiple antibacterial resistance and long-term persistence of microorganisms, mediate the development and outcome of the infectious process. The study of the fish immunological parameters in interaction with biofilm-forming bacteria is of practical importance for assessing the stability of the homeostasis of the fish. This study aimed to determine the immunobiological parameters of Cyprinus carpio Linnaeus when interacting with biofilm-forming bacteria Aeromonas hydrophila. Clinically healthy fish C. carpio L. (Linnaeus, 1758) of both sexes, aged 4 years, and weighing 1.0-1.5 kg (n = 10), were used in this study. The fish were taken from the pond of the VNIIR experimental base in the period of 2020-2022. The standard method was employed to determine the phagocytic activity of blood cells, the total redox activity of neutrophils, and the bactericidal activity of blood serum. After 24-48 h of cultivation in nutrient broth, the implementation of the processes of intercellular communication of bacteria had common patterns of formation of the heterogeneous structure of biofilms. Moreover, analyzing the optical density indices (density, D), it was observed that A. hydrophila was a strong producer of biofilms, as the optical density of the sample (density of sample, Ds) exceeded the optical density of the control (density of control, Dc) by more than 4 times (D = 0.464 ± 0.07). The ratio of the average number of microorganisms attached to the surface of one erythrocyte (average adhesion index) and the percentage (%) of erythrocytes having bacteria on their surface (adhesion coefficient [AC]) was 14.05 ± 0.72, and the adhesion index, AI was ≥4.00, indicating A. hydrophila to be highly adhesive. In addition, the AC of erythrocytes having bacteria on the surface was 14.05% ± 0.72%. A direct correlation was established (R2 = 0.94) between the AC (14.05% ± 0.11%-13.29% ± 0.08%) and the phagocytic index (11.3% ± 0.29%-32.0% ± 0.8%). The indicators of spontaneous nitro blue tetrazolium were 103.20 ± 11.70 when estimating the total redox activity of neutrophils. The optical density increased to 182.10 ± 21.12 with the addition of 20.0 mL of A. hydrophila bacteria (1 billion/mL) and the activity of neutrophils also increased. Among the markers of homeostasis stability, immunological indicators most fully reflect the mechanisms of initiation, development, and outcome of the infectious process mediated by the interaction of adhesive molecules of multicellular eukaryotes and adhesives of infectious disease pathogens. The research will contribute to further understanding the potential mechanism of quorum-sensing molecules and the search for new anti-adhesive drugs that reduce the formation of biofilms.

Synthesis, characterization and superoxide dismutase activity of a biomimetic Mn(III) complex covalently anchored to mesoporous silica

A mononuclear Mn(III) complex of a clickable ligand, [Mn(hbpapn)(H2O)2]ClO4·4.5H2O, where H2hbpapn = 1,3-bis[(2-hydroxybenzyl)(propargyl)amino]propane, has been prepared and fully characterized. The complex catalyzes the dismutation of superoxide employing a Mn(III)/Mn(IV) redox cycle, with catalytic rate constant of 3.9 × 106 M−1 s−1 determined through the nitro blue tetrazolium photoreduction inhibition assay, in aqueous medium of pH 7.8. The alkyne function of the ligand was used for the covalent attachment of the catalyst to azide modified mesoporous silicas with different texture and morphology, through click chemistry. In these materials the catalyst is essentially linked to the inner pore walls, isolated and protected from the external medium. The hybrid materials can be recycled, and retain or improve the superoxide dismutase activity of the free catalyst with the pore size of the solid matrix playing a role on the activity of the catalyst.

Electrochemical and in Silico Investigations of the Interaction between Nitro Blue Tetrazolium Chloride and Bovine Serum Albumin

The binding ability of the drug on its interaction with the protein will also significantly affect the apparent volume of distribution of the drugs and, in many cases the rate of elimination of the drugs. The interactions of proteins and other molecules are a fascinating topic applied to surface technologies and sensors. Therefore, it is aimed to determine the NBTCand to elucidate its interaction with Bovine Serum Albumin (BSA) by electrochemical and in silico studies in this paper. The reduction in BSA oxidation signals measured by differential pulse voltammetry upon incubation with different NBTC concentrations indicated that NBTC was bound to BSA. In addition, in silico (molecular modeling and molecular dynamics) studies have been conducted on the interactions of NBTC with proteins in plasma. As a result of the in silico studies investigated the interactions of NBTC with serum albumin, its binding affinity, and the dynamic process in the binding state. In silico studies showed that NBTC binds to BSA with high affinity (with -7.986 kcal/mol docking score), and this binding was stable (with a 3.0 average RMSD value). Eventually, the results of the electrochemical and modeling studies were perfectly matched.

Effects of 2-methoxyestradiol on hydrogen peroxide induced neuronal cell death and tau hyperphosphorylation

AimsAlzheimer's Disease (AD) is characterized by progressive cognitive impairment, and memory loss. It has been shown that depletion of estrogens renders women vulnerable to AD with menopause women presenting higher risk for AD development than men. However, women under hormone replacement therapy (HRT) with 17β-estradiol (E2) show lower risk for AD, implying that E2 may be protective. It has been shown that E2 exerts its effects through the estrogen receptor (ER) but also via its biologically active metabolites, 2-hydroxyestradiol (2OH), and 2-methoxyestradiol (2ME). We hypothesized that the neuroprotective effects of E2 are partly attributed to its metabolites. Materials and methodsSH-SY5Y neuronal cells were subjected oxidative stress (OS) cell death by hydrogen peroxide (H2O2), in the presence or absence of E2, 2ME and 2OH. Viability was assessed by trypan blue and thiazolyl blue tetrazolium bromide assays, intracellular OS with the Dichlorodihydrofluorescein Diacetate (DCFDA) assay, and Bax, p53 and PUMA quantified by RT-PCR. Tau hyperphosphorylation was studied by western blot. Key findingsE2 and its metabolites 2OH and 2ME protect from cell death as assessed by the viability assays. Their effect was partly attributed to their antioxidant properties evidenced by the reduction of intracellular OS. Treatment with 2ME resulted in a reduction of Bax, but not p53 or PUMA in cells challenged with OS. Finally, 2ME was able to inhibit tau hyperphosphorylation as well. SignificanceE2 protects neuron cells partly through its metabolites. Further studies are needed to fully delineate the mechanism for this protection.