Nitroblue Tetrazolium Test Research Articles

Chronic exposure to TiO2 micro- and nano particles: A biochemical and histopathological experimental study.

The aim of this work was to analyze the effects of long-term exposure to titanium dioxide (TiO2) micro- (MPs) and nanoparticles (NPs) (six and 12 months) on the biochemical and histopathological response of target organs using a murine model. Male Wistar rats were intraperitoneally injected with a suspension of TiO2 NPs (5 nm; TiO2-NP5 group) or MPs (45 μm; TiO2-NP5 group); the control group was injected with saline solution. Six and 12 months post-injection, titanium (Ti) concentration in plasma and target organs was determined spectrometrically (ICP-MS). Blood smears and organ tissue samples were evaluated by light microscopy. Liver and kidney function was evaluated using serum biochemical parameters. Oxidative metabolism was assessed 6 months post-injection (determination of superoxide anion by nitroblue tetrazolium (NBT) test, superoxide dismutase (SOD) and catalase (CAT), lipid peroxidation, and paraoxonase 1). Titanium (Ti) concentration in target organs and plasma was significantly higher in the TiO2-exposed groups than in the control group. Histological evaluation showed the presence of titanium-based particles in the target organs, which displayed no structural alterations, and in blood monocytes. Oxidative metabolism analysis showed that TiO2 NPs were more reactive over time than MPs (p < .05) and mobilization of antioxidant enzymes and membrane damage varied among the studied organs. Clearance of TiO2 micro and nanoparticles differed among the target organs, and lung clearance was more rapid than clearance from the lungs and kidneys (p < .05). Conversely, Ti concentration in plasma increased with time (p < .05). In conclusion, neither serum biochemical parameters nor oxidative metabolism markers appear to be useful as biomarkers of tissue damage in response to TiO2 micro- and nanoparticle deposits at chronic time points.

BENEFICIAL EFFECTS OF ANGIOSTATIN K1-3 AND LACTOFERRIN IN ALKALI-BURNED RABBIT CORNEA: A COMPARATIVE STUDY

Corneal injury is associated with hypoxia-induced neovascularization, which interferes optical transparency of the cornea, resulting in vision loss and blindness. The treatment of corneal damage remains a ignificant unmet medical need. Lactoferrin (Lf) and angiostatins (AS) are naturally occurring antiinflammatory and antiangiogenic proteins, which play important roles in eye physiology and can be used as protectors against corneal diseases. The aim of the study was to evaluate and compare the effects of plasminogen fragment AS K1-3 and Lf (as a referent preparation) on the levels of the principal markers of angiogenesis, apoptosis, and autophagy in the burn-injured cornea. Additionally, effects of AS on the metabolic activity of macrophages have been studied. Materials and methods. Experimental model of corneal burn was induced by NaOH application to rabbit’s eye surface. One of the group of animals with injured cornea topically received AS, which were applied as eye drops (1 μM solution) daily for 14 days after injury, another one was treated by the equimolar solution of Lf. The levels of protein markers of angiogenesis (VEGF), apoptosis (caspase-3), and autophagy (beclin-1) were evaluated in corneal lysates by western blot. Histological analysis was performed by hematoxylin/eosin staining of corneal slices followed by light microscopy. Effects of AS in the range of concentrations 10-200 nM on the activity of phagocytic cells were assessed in murine macrophage cell line J744.2 by the test with nitroblue tetrazolium (NBT). The quantitative results were analysed with the use of Mann-Whitney U-test. The difference between group mean values was considered significant at P &lt; 0.05. Results. Dramatically increased levels of protein markers related to angiogenesis, apoptosis, and utophagy were found in the corneas with burn-induced injury. AS or Lf-based treatment reduced burninduced overexpression of VEGF, caspase-3, and beclin-1. It is important to note that AS, along with anti-VEGF activity, appeared to more effectively suppress apoptosis and autophagy by decreasing tested markers near the control level, as compared with Lf. Histochemical examination revealed typical signs of fibrotic and necrotic changes in the injured corneas, which were associated with excessive inflammatory infiltration and intense neovascularization. Treatment of burn-induced injury with AS alleviated histopathological changes in cornea, which is evidenced by improving epithelial regeneration, reducing neovascularization, nd moderating leucocyte infiltration. These observations are in agreement with the ata of NBT test suggesting that AS in the concentrations &gt;50 nM is able to moderately inhibit metabolic activity of macrophages up to 30% as compared with intact cells (P&lt;0.05). Conclusions. Obtained results indicate that AS may serve as an effective treatment option to relief alkali-induced corneal injury with the efficacy comparable or even higher than that of firmly established eye protective protein Lf.

SOURCES OF SUPEROXIDE ANION RADICAL IN SMALL INTESTINE TISSUES IN RATS SUBJECTED TO SURGICAL TRAUMA SIMULATED UNDER EXPERIMENTAL MODEL OF POSTTRAUMATIC STRESS DISORDER

The sources of superoxide anion radical (.О ) in the tissues of the small intestine in rats subjected to surgical trauma simulated under an experimental model of posttraumatic stress disorder (PTSD) were investigated. The study involved 42 white Wistar rats weighing 210-230 g, divided into 6 groups: group 1 comprised intact animals, group 2 included animals with the PTSD induced through single-prolonged stress (SPS), group 3 consisted of rats subjected to a sham surgical operation, group 4 comprised animals undergoing laparotomy, group 5 involved rats undergoing a sham surgical operation following SPS, and group 6 included animals undergoing laparotomy under modeled SPS. The rate of .О generation in the small intestine homogenate was measured spectrophotometrically using the nitroblue tetrazolium test. The .О production by NADPH-dependent (microsomal and NO synthase) electron transport chains (ETC), NADH-dependent (mitochondrial) ETC and leukocyte NADPH oxidase was assessed. The findings obtained have demonstrated that experimental SPS modeling leads to an increase in oxidative stress in the small intestine tissues of rats. This is manifested by an increase in the rate of .О formation with the participation of microsomes, mitochondria and leukocyte NADPH oxidase. On the 7th day after laparotomy under the experimental PTSD model, the .О production in the tissues of the small intestine by different sources (microsomes, mitochondria and leukocyte NADPH oxidase) exceeds their values obtained after a single laparotomy or after performing a sham operation under single long-term stress.

The Relationship between Serum Zonulin and Innate Immunity in Patients with Inflammatory Bowel Disease

The aim of the study was to investigate the relationship between the serum zonulin and the cellular immunity in patients with ulcerative colitis (UC) and Crohn’s disease (CD). The study included 97 patients, 13 (13.4%) patients with CD and 84 (86.6%) patients with UC. The concentration of zonulin in the serum was studied using the ELISA kits. The investigation of the circulated leukocyte subpopulation was carried out through flow cytometry. The functional activity of the circulating neutrophils was studied using the absorption capacity of the monodisperse polystyrene latex particles and indicators of oxygen-dependent metabolism in the nitroblue tetrazolium test. The serum zonulin concentration in CD patients was significantly higher compared with UC patients (p = 0.003). The zonulin concentration directly correlated with the functional activity of the circulating neutrophils. Patients with a zonulin concentration &gt; 472.4 pg/mL had a significantly higher number of band neutrophils (p = 0.0104), CD3+CD8+ cells (p = 0.0212), NK cells (p = 0.0161), and lower–CD19+ cells (p = 0.0034). Among the IBD patients, zonulin was associated with IBD severity. An increase in the serum concentration of zonulin was associated with an increase in the functional activity of circulating neutrophils and an increase in the number of CD3+CD8+ cells, NK cells, and a decrease in the number of CD19+ cells.

Estimation of Phagocytic activity by normal human peripheral blood mononuclear cells on various oral isolates of Candida species – An in-vitro study

Polymorphonuclear neutrophils (PMN) and mononuclear phagocytes represent an important first line and effector function in control of Candida infections. The aim of the study is to determine the in-vitro phagocytic activity of human peripheral blood mononuclear cells against oral isolates of Candida species and its antifungal susceptibility. The study also evaluates the degree of respiratory burst activity of PBMCs. Phagocytic and lytic indices by PBMCs were determined for Candida spp. The respiratory burst activity was evaluated by nitroblue tetrazolium test. Antifungal disc diffusion susceptibility testing was performed. A total of 100 Candida were isolated belonging to the species C.albicans, C. tropicalis, C.krusei and C.auris. Phagocytic and lytic indices of C.albicans was significant when compared to standard strain of C.albicans. For C.tropicalis and C.krusei phagocytic index was significant while lytic index was not significant when compared to standard strain. The inter species comparison of both the indices was not significant for the clinical isolates of Candida. A significant reduction in phagocytic activity was observed for clinical isolates of Candida spp. but lytic activity was variable when compared to the standard strain of C.albicans.

Effects of exposure to alternating low-intensity, intermediate-frequency electric fields on the differentiation of human leukemic cell line U937.

Studying the bioeffects of electric fields have been the subject of ongoing research which led to promising therapeutic effect, particularly in cancer treatment. Here, we investigated the impact of low-intensity, intermediate-frequency alternating electric fields on the differentiation of human myeloid leukemia cell line U937. The results showed a near twofold increase in differentiation of U937 cells treated for 24 h by alternating 600 kHz, 150 V/m electric fields, in comparison to their control groups. This measure was evaluated by latex bead phagocytosis assay, nitro blue tetrazolium test, and cell cycle analysis which revealed a significant shift in the number of cells from G2 +M to G0 +G1 phases. The simulation result for the intracellular field intensity showed around 50% attenuation with respect to the applied external field for our setup which ruled out masking of the applied field by the internal electric noise of the cell. Based on previous studies we postulate a possible calcium-related effect for the observed differentiation, yet the exact underlying mechanism requires further investigation. Finally, our results may offer a potential therapeutic method for leukemia in the future.

Exploring the Relationship between Neutrophil Activation and Different States of Canine L. infantum Infection: Nitroblue Tetrazolium Test and IFN-γ.

This study aimed to investigate the role of neutrophils in canine leishmaniosis by assessing neutrophil activation and its relationship with different states of L. infantum infection and antibody and IFN-γ production. Dogs were categorized into five groups: healthy-seronegative (n = 25), healthy-seropositive (n = 21), LeishVet-stage I (n = 25), Leishvet-stage II (n = 41), and LeishVet-stage III-IV (n = 16). Results of the nitroblue tetrazolium reduction test (NBT) showed significantly higher neutrophil activation in stage I (median:17.17, range: [7.33-31.50]%) compared to in healthy-seronegative (4.10 [1.20-18.00]%), healthy-seropositive (7.65 [3.98-21.74]%), stage II (6.50 [1.50-28.70]%), and stage III-IV (7.50 [3.00-16.75]%) groups (p < 0.0001). Healthy-seropositive dogs also displayed higher values than all groups except stage I. Stages II and III-IV did not show significant differences compared to healthy-seronegative. Regarding IFN-γ, stage I dogs had higher concentrations (median:127.90, range: [0-3998.00] pg/mL) than healthy-seronegative (0 [0-109.50] pg/mL) (p = 0.0002), stage II (9.00 [0-5086.00] pg/mL) (p = 0.045), and stage III-IV (3.50 [80.00-548.80] pg/mL) (p = 0.02) dogs. Stage II dogs showed increased IFN-γ compared to healthy-seronegative dogs (p = 0.015), while stage III-IV dogs had no significant differences compared to healthy-seronegative dogs (p = 0.12). Healthy-seropositive dogs had elevated IFN-γ concentrations compared to healthy-seronegative dogs (p = 0.001) and dogs in stage III-IV (p = 0.03). In conclusion, neutrophil activation was higher in dogs with mild disease and healthy-seropositive dogs, and a relationship between neutrophil activation and the production of IFN-γ was found.

Omega-3 Fatty Acids as Supplements and Cryoprotectants and Positive Outcomes for Human Seminal Cryopreservation: A Study in the Iraqi Context

Background: With the increasing deterioration in male fertility rates over the past few decades, assisted reproductive therapy via sperm banking has gained considerable attention from different health authorities worldwide. The process of sperm cryopreservation exerts certain harmful effects on sperm quality parameters. The aim of the current study was to examine the effect of omega-3 on human sperm cryopreservation as a dietary supplement and as a cryoprotectant stimulant.&#x0D; Methods: From healthy men, 120 seminal fluid samples were randomly collected for cryopreservation (for 30 days). All samples were divided into three groups (40 in each). The supplement group (SG) (first group) had been given dietary supplements of omega-3 (30% EPA: 20% DHA) for eight weeks before sample collection. The samples of the cryoprotectant group (CG) (second group) were treated with omega-3 additive in the diluent at the time of being collected and before we cryopreserved them. The samples from the control group (third group) were cryopreserved without any dietary or cryoprotectant supplementation. Initial seminal analysis was recorded and post-thawing assessment of sperm motility, vitality (using eosin test) and oxidative stress (using a nitro blue tetrazolium (NBT) test) were compared.&#x0D; Results and Conclusion: SG samples had greater initial seminal fluid volumes, as well as better sperm motility and vitality, but the oxidative stress assessment did not differ significantly pre-cryo. Post-thawing assessment revealed that the CG group had the best parameters of motility, vitality and oxidative stress. These results may be related to the positive effects of omega-3 fatty acids on reproductive glands, hormonal milieu, sperm function and structure, making it a suitable biostimulant for improving the outcome of human sperm banking. Results were comparable to multiple previous animal studies.

Rosmarinic acid alone or in combination with Lactobacillus rhamnosus ameliorated resistance to ammonia stress in the rainbow trout, Oncorhynchus mykiss: growth, immunity, antioxidant defense and liver functions

Abstract Rosmarinic acid (RS) and Lactobacillus rhamnosus (LR) were added singularly or in combination to rainbow trout (Oncorhynchus mykiss) diets to test their efficacy in the protection against ammonia stress. Fish (31.4±0.6 g) were randomly allocated to six groups in three replicates, as follows: T1: basic food as control, T2: LR with a concentration of 1.5 × 108 CFU /g, T3: LR with a concentration of 3 × 108 CFU/g, T4: 1 g RS/kg, T5: 3 g RS/kg, and T6: 1.5 × 108 CFU/g LR + 1 g RS/kg and T7: 3 × 108 CFU/g LR + 3 g RS/kg. After 60 days feeding, fish were exposed to ammonia stress. After the feeding period, the supplemented fish had the highest final body weight (FW), weight gain (WG), and specific growth rate (SGR), and the lowest feed conversion ratio (FCR) as compared with the control group (P&lt;0.05). Amylase, protease and lipase activities were noticed markedly higher in fish supplemented with 1.5 × 108 CFU/g LR + 1 g RS/kg and 1.5 × 108 CFU/g LR diets compared to the control (P&lt;0.05). Generally, fish in supplemented diets, particularly T2 and T6 groups, had the highest lysozyme, alternative complement activity (ACH50), total Ig, nitroblue tetrazolium test (NBT), myeloperoxidase (MPO), complement component 3 (C3), complement component 4 (C4), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx). On the other hand, T2 and T6 groups had the lowest malondialdehyde (MDA), glucose, and cortisol concentrations as well as alanine aminotransferase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) enzyme levels when compared with the control (P&lt;0.05). After ammonia stress, fish in the supplemented groups, particularly T2 and T6, generally showed significantly higher values of lysozyme, ACH50, total Ig, NBT, MPO, C3, C4, SOD, CAT, GPx and lower levels of MDA, glucose, cortisol, ALT, ALP, LDH when compared with the control (P&lt;0.05). In conclusion, a combined administration of RS and L. rhamnosus effectively improved growth performance and health status as well as enhanced the resistance of rainbow trout against ammonia toxicity.

Evaluation of NADPH Oxidase (NOX) Activity by nitro Blue Tetrazolium (NBT) Test in SLE Patients.

Evaluation of NADPH Oxidase (NOX) Activity by nitro Blue Tetrazolium (NBT) Test in SLE Patients.

Pot Marigold (Calendula officinalis) Powder in Rainbow Trout (Oncorhynchus mykiss) Feed: Effects on Growth, Immunity, and Yersinia ruckeri Resistance

The current research investigated the effects of pot marigold (Calendula officinalis) powder on growth, biochemical parameters, digestive enzymes, serum and mucus immune responses, antioxidant defense, and resistance of rainbow trout (Oncorhynchus mykiss) against Yersinia ruckeri. Fish (No. 450, 15.06 ± 0.11 g; mean ± SE ) were randomly distributed to five groups fed with a diet containing 0 (control group), 0.5%, 1%, 1.5%, and 2% of pot marigold powder (MP) for 60 days. Then, fish were challenged with Y. ruckeri infection. Specific growth rate (SGR), weight gain (WG), final weight (FW), feed conversion ratio (FCR), mucus lysozyme (LYZ), mucus protease, serum nitroblue tetrazolium test (NBT), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) values in the 1%, 1.5%, and 2% MP groups significantly increased ( P &lt; 0.05 ) compared to the other groups. Regression analysis exhibited that rainbow trout growth indices were polynomially linked to dietary MP concentrations. In this regard, the optimal levels of MP, according to growth parameters (SGR and FCR), were 1.31 and 1.4% diet, respectively. The intestinal protease, serum complement component 3 (C3), complement component 4 (C4), and LYZ activities in fish fed with the 1% and 1.5% MP-supplemented regime were higher ( P &lt; 0.05 ) compared to the other groups. Also, fish fed with diets containing 1.5% MP had significantly higher intestinal lipase and mucus ALP activities than the other groups ( P &lt; 0.05 ). Aspartate aminotransferase (AST) activity in all MP groups remarkably increased ( P &lt; 0.05 ) compared to the control. Moreover, fish fed with a diet containing 1.5% MP had remarkably higher ( P &lt; 0.05 ) lactate dehydrogenase (LDH) and total protein (TP) values than the other groups. Serum myeloperoxidase (MPO), total immunoglobulin (Ig), and mucus peroxidase values in the 1% MP group remarkably increased ( P &lt; 0.05 ) compared to the other groups. In addition, catalase (CAT) and superoxide dismutase (SOD) activities in the 1.5% and 2% MP groups were significantly higher ( P &lt; 0.05 ) compared to the others. However, malondialdehyde (MDA) levels in fish fed with the 1%, 1.5%, and 2% MP-supplemented diet remarkably decreased than in the other groups ( P &lt; 0.05 ). The highest survival rate after a challenge with Y. ruckeri was recorded in the 1.5% group compared to the control group. Our findings revealed that using pot marigold powder in diets, especially at the 1.5% level, has positive effects on growth, digestive enzyme activities, antioxidant capacity, immune response, and disease resistance in O. mykiss.

S-Doped rGO-Enwrapped Magnetically Porous NiFe2O4/CuS Heterojunction: An Efficient Z-Scheme Nano-Photocatalyst for Visible-Light-Driven Tetracycline Degradation and 4-Nitrophenol Reduction

The photocatalytic degradation of economically hazardous compounds has been thoroughly researched with the aid of numerous photocatalysts and techniques. The efficient degradation and removal of lingering contaminants from the aquatic environment are still difficult to accomplish. Hereby, we present a sulfur-doped reduced graphene oxide (SrGO) enwrapped magnetic porous nickel ferrite (NiFe2O4)/copper sulfide (CuS) nanocatalyst (SrGO/NiFe2O4/CuS:GNFC) constructed through a facile solvothermal-reflux route. Under exposure to visible light, the fabricated GNFC photocatalyst exhibits outstanding catalytic activity for the photodegradation of tetracycline hydrochloride (TCH) and photoreduction of 4-nitrophenol (4-NP). Diverse analytical techniques were used to characterize the fabricated nanomaterials. In terms of photocatalytic activity, GNFC-13 outperforms all other nanocatalysts in eliminating model pollutants. According to our findings, 94.39% of TCH (80 ppm) can be degraded within 90 min, and 90.62% of 4-NP (30 ppm) can be reduced within 120 min by the GNFC-13 nanohybrid, which is significantly better than that of pristine and doublet nanomaterials, respectively. The explanations for the improved photocatalytic efficiency of GNFC nanocatalysts are due to the porous structures of the magnetic NiFe2O4 and the SrGO surface, which can offer a lot of adsorption sites and, therefore, advantageous for the adsorption enrichment of harmful pollutants. Additionally, in situ photocatalytic degradation and adsorption enrichment working together synergistically may lead to improved pollutant removal efficacy. The Raman and XPS analytical techniques verified the formation of SrGO in the GNFC nanocomposites. The free-radical trapping studies, terephthalic acid test, nitroblue tetrazolium test, and electron spin resonance test disclosed that holes (h+), hydroxy radicals (·OH), electrons (e–), and superoxide radicals (·O2–) are cardinal reactive species in the photocatalytic system. A Z-scheme charge transfer channelization mechanism of the as-produced photocatalyst is explained in light of the various experimental findings. The developed Z-scheme system has led to the increase of catalytic activity due to the effective photoinduced carrier separation, wider photoabsorption range, high hole oxidation capacity, and high electron reduction power. In addition, the GNFC photocatalyst ability to catalyze TCH and 4-NP shows no discernible decline even after five recycles. Moreover, the GNFC nanocatalysts are magnetically detachable for recycling. Thus, this study reveals that the as-obtained GNFC nanocatalysts have an excellent prospect for environmental remediation of toxic pollutants.

Degradation of Fusobacterium nucleatum biofilm and quantity of reactive oxygen species due to a combination of photodynamic therapy and 2.5% sodium hypochlorite

Background: The persistence of microorganisms in the root canal system is one of the leading causes of root canal treatment failure. Biofilms of putative pathogens hidden inside dentin tubules and other root canal ramifications may limit current disinfection protocols. Photodynamic therapy (PDT) with a wavelength of 628 nm can be used as an antimicrobial strategy that uses low-power laser energy to activate a non-toxic photosensitizer to produce singlet oxygen with the ability to kill microorganisms in root canals. Fusobacterium nucleatum was used because this bacterium is one of the bacteria involved in root canal infection. Purpose: The aim of this study was to compare the bactericidal efficacy of sodium hypochlorite (NaOCl) 2.5%, PDT, and a combination of PDT and NaOCl 2.5% against Fusobacterium nucleatum. Methods: Mature biofilm Fusobacterium nucleatum was divided into four groups according to the protocol of decontamination: K1 (negative control – biofilm), K2 (NaOCl 2.5%), K3 (PDT), and K4 (NaOCl 2.5% + PDT). Biofilm degradation was observed using optical density (OD) at 570 nm using a microplate reader. A reactive oxygen species quantity check was carried out using a nitroblue tetrazolium test, and OD observation was done with a microplate reader at 540 nm. Results: Group 4 (NaOCl 2.5% + PDT) showed more biofilm bacteria elimination than the other groups. Conclusion: A combination of PDT and NaOCl 2.5% can be considered an effective protocol for the elimination of Fusobacterium nucleatum. There is a potentiation relationship between NaOCl 2.5% and PDT FotoSan. Biofilm degradation occurs because of the effect of antibacterial NaOCl 2.5% and the irradiation effect of the Toluidine blue O photosensitizer.

Single or combined consumption of resveratrol and the probiotic, Lactobacillus acidophilus attenuate the effects of crowding stress on growth, immune characteristics, and antioxidant defense in the common carp, (Cyprinus carpio)

In the present study, dietary resveratrol (RE) and Lactobacillus acidophilus (LAB) were individually or combined added to the diet of common carp (Cyprinus carpio) to protect against crowding stress. Fish (30.16 ± 0.7; Mean ± SE) were randomly allocated to seven groups in three replicates, as follows: T1: basic food as control, T2: LAB with a concentration of 1.5 × 107 CFU/g, T3: LAB with a concentration of 3 × 107 CFU/g, T4: 300 mg resveratrol/kg, T5: 600 mg resveratrol/kg, and T6: 1.5 × 107 CFU/g + 300 mg resveratrol/kg and T7: 3 × 107 CFU/g + 600 mg resveratrol/kg. After 60 days feeding, the supplemented fish had the highest final body weight (FBW), weight gain (WG), and specific growth rate (SGR), and the lowest feed conversion ratio (FCR) as compared with the control group (P < 0.05). The activities of amylase, protease and lipase were noticed markedly higher in fish supplemented with 1.5 × 107 CFU/g + 300 mg resveratrol/kg and 1.5 × 107 CFU/g diets compared to the control (P < 0.05). Generally, fish in supplemented diets, particularly T2 and T6 groups, had the highest lysozyme, alternative complement activity (ACH50), total immunoglobulin (Ig), nitroblue tetrazolium test (NBT), myeloperoxidase (MPO), complement component 3 (C3), complement component 4 (C4), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and lower levels of malondialdehyde (MDA), glucose, cortisol, alanine aminotransferase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) when were compared with the control before crowding stress (P < 0.05). After crowding stress challenge, fish in the supplemented groups, particularly T2 and T6, generally showed significantly higher values of lysozyme, ACH50, total Ig, NBT, MPO, C3, C4, SOD, CAT, GPx and lower levels of MDA, glucose, cortisol, ALT, ALP, LDH when compared with the control (P < 0.05). Also, recovered fish in the control group demonstrated significantly declined levels of lysozyme, ACH50, total Ig, NBT, MPO, C3, C4, SOD, CAT, GPx and higher levels of MDA, glucose, cortisol, ALT, ALP, LDH as compared to other group (P < 0.05). In conclusion, a combined administration of RE and LAB effectively improved growth performance and health status as well as protected common carp against crowding stress.

Эффекты пептида GHК и его структурных аналогов D-Ala-GHK и GHК-D-Ala на состояние врожденного иммунитета и перекисного окисления липидов в условиях кожной раны

Objective: to study the effects of the GHK peptide and its structure analogues on the mechanisms of innate immunity and lipid peroxidation in wound. Materials and methods. The experiments were performed on 70 male Wistar rats. The skin wound was modeled by applying a full-layer wound with an area of 250 mm2 on the animal's back. The peptides Gly-His-Lys (GHK), D-Ala-Gly-His-Lys (D-Ala-GHK), and Gly-His-Lys-D-Ala (GHK-D-Ala) were used in doses of 0.5 μg/kg and 1.5 μg/kg, which it was administered intradermally in doses of 0.5 μg/kg and 1.5 μg/kg in a volume of 0.1 ml for 3, 7 or 10 days. The activity of lipid peroxidation (LPO) processes in blood serum was assessed by the content of malonic dialdehyde (MDA) and acylhydroperoxides (AHP). The antioxidant mechanisms were evaluated by determining the activity of catalase and the total antioxidant activity (OAA) of blood serum. Phagocytic activity of blood neutrophils was assessed by phagocytic index (PI) and phagocytic number (PN). The activity of oxygen-dependent mechanisms in phagocytes was evaluated in a spontaneous nitroblue tetrazolium test (NBT). Results. After the administration of GHK, the tendency to PI, PN and completeness of phagocytosis prevailed, which mainly persisted with the use of peptides D-Ala-GHK and GHK-D-Ala. At the same time, the GHK-D-Ala peptide at a dose of 1.5 μg/kg had the most stable effect on phagocytic activity. The data obtained in the NBT are largely consistent with the PN indicators. At the same time, the effects of structural analogues, unlike GHK, were manifested throughout the experiment. Significantly significant changes in the activity of LPO and antioxidant mechanisms were observed with the use of all peptides. However, their dynamics, orientation and severity throughout the experiment were quite complex. Conclusion. GHK and its structural analogues, D-Ala-GHK and GHK-D-Ala, had an effect on the indicators of innate immunity and LPO in a skin wound, the severity and direction of which depends on the healing period. At the same time, the most pronounced and stable effects were observed when using GHK-D-Ala. That demonstrates the importance of protecting the tripeptide molecule from the action of carboxypeptidases.

Effect of reactive inflammation in osteoarthritis on extracellular traps formation by circulating neutrophils

The results of recent studies demonstrating the presence of pro-inflammatory mediators in osteoarthritis (OA), as well as known similarity of histological signs of inflammation in rheumatoid arthritis (RA) and OA, suggest an important role of NETosis in immune inflammation in OA patients. Our objective was to assess ability of blood neutrophils from ОA patients to generate NETosis spontaneously and after in vitro induction, and impact of reactive synovitis on the dynamics of NETosis indexes. Thirty-one patients with verified OA and 30 healthy volunteers were included into the study. Circulating neutrophils were isolated with one-step density gradient centrifugation using double layers of Iohexol gradient. Subpopulational profile of isolated neutrophil fractions, their viability, and nonspecific activation were evaluated microscopically using Trypan Blue exclusion test, as well as nitro-blue tetrazolium test. NETs were induced by phorbol-12-myristate13-acetate (PMA). Spontaneous and induced formation of NETs was assessed using fluorescence microscopy. The ОA patients were in clinical remission at the time of inclusion in the study. In 23 OA patients, an exacerbation was diagnosed during the study. The neutrophil fractions showed high purity and a high content of viable nonactivated cells. These parameters were comparable in the study groups. Mean percentage of spontaneous NETs in OA patients in remission was significantly increased comparing to healthy controls. Usage of PMA, as inducing agent was accompanied by a significant increase in ability of neutrophils to form NETs. Transition of OA to exacerbation was characterized by further significant increase in spontaneous and PMA-induced NETs. Spontaneous and induced NETs in OA patients at acute stage of the disease are significantly higher than in OA patients in remission state. The growth rate of spontaneous NET formation is 3.74 times higher than the induced NET formation in OA patients upon exacerbation. Statistically significant increase in the ability of peripheral neutrophils to spontaneous and induced formation of extracellular traps was found, depending on the stage of osteoarthritis. The data obtained suggest an opportunity for participation of circulating neutrophils via NETosis in pathogenesis of immune inflammation in OA.

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.

Use of Nitroblue Tetrazolium Test: Revisited in Context of COVID-19.

Use of Nitroblue Tetrazolium Test: Revisited in Context of COVID-19.

Age-specific features of the absorptive and cytokine-inducing function of phagocytosis in children with recurrent acute respiratory infections

Background. The study of the absorbing and cytokine-inducing function of phagocytosis is a mandatory starting point in assessing the state of anti-infective protection in children who suffer from frequent respiratory infections. It is important to understand and compare the features of phagocytosis in these children depending on their age. The purpose: to improve the diagnosis of immune disorders in children aged 2–5 and 6–10 years, who often suffer from acute respiratory diseases, by means of a comparative analysis of phagocytosis parameters and serum levels of some cytokines. Materials and methods. Under observation, there were 60 children aged 2 to 10 years who often suffer from acute infectious diseases of the respiratory tract. Two groups were formed: 1) children aged 2–5 years (n = 30); 2) children aged 6–10 years (n = 30). Results. Among patients of the first group compared to those of group 2, an increase in the frequency of low indicators of the phagocytic number (by 36.7 %, p &lt; 0.05), phagocytic index (by 46.7 %, p &lt; 0.05), the index of completion of phagocytosis (by 66.6 %, p &lt; 0.05) and the staphylococcus-stimulated nitroblue tetrazolium test (by 46.4 %, p &lt; 0.05), an increase in the frequency of high serum levels of interleukin-1β (by 33.3 %, p &lt; 0.05), interleukin-10 (by 27.7 %, p &lt; 0.05), a decrease in the frequency of high levels of interleukin-6 (by 16.7 %, p &lt; 0.05). In the children of group 2 compared to those in the first one, high index of completion of phagocytosis was more often registered (by 70.0 %, p &lt; 0.05), as well as more than a twofold increase in the ratio of interleukin-6/interleukin-10. Conclusions. In children aged 6–10 years, more frequent morbidity is due to a cytokine imbalance that was reflected in a more than a twofold increase in interleukin-6/interleukin-10 ratio, which is a negative factor indicating an enhancement of pro-inflammatory processes and a longer damage to the cells of the immune microenvironment.

Стан неспецифічної резистентності ротової порожнини у працівників промислових виробництв із шкідливими чинниками

The purpose of the study was to investigate local resistance of the oral cavity in industrial workers with harmful factors, patients with generalized periodontitis. Materials and methods. For this study, 2 examination groups were formed. The main group (61 people) includes employees of three industrial productions of the Ivano-Frankivsk region: employees of chemical production, glass production and agro-industrial complex. The control group consisted of 30 conditionally healthy individuals with intact periodontium. The age of the examinees was 25–55 years. Work experience in production is from 0.5 to 15 years. In order to study the state of local resistance in generalized periodontitis under the influence of harmful factors of the production environment, individual indicators of the immunity of the capillary blood of the gums and oral fluid were studied: the total number of leukocytes and lymphocytes; to evaluate the T-system of immunity, calculation of subpopulations of lymphocytes was performed using monoclonal antibodies CD-3, CD-4, CD-8; the level of immunoglobulins IgA, IgG and sIA; circulating immune complexes; phagocytic index, spontaneous nitroblue tetrazolium test. Results and discussion. A comparative evaluation of the results of immunological research in 61 patients with generalized chronic periodontitis of the main group and 30 healthy individuals of the control group indicated certain changes in the quantitative data of cellular and humoral factors and deviations in the functional activity of individual parameters of non-specific protection in the capillary blood of the gums of patients compared to the indicators of individuals with intact periodontium. The total number of leukocytes and lymphocytes in workers with generalized periodontitis remained within the generally accepted norm and differed insignificantly from the similar indicators of the healthy group. At the same time, a difference in the subpopulation composition of T-lymphocytes was revealed. According to absolute indicators, the number of CD4+ lymphocytes in patients with generalized periodontitis was 0.57 ± 0.04109/l, which is by 1.2 times lower compared to 0.68 ± 0.03109/l in healthy individuals. The obtained indicators of CD4+ and CD8+ lymphocytes made it possible to state a decrease in the value of the immunoregulatory index in workers with generalized periodontitis by 19.31% from the value of healthy people (1.63 ± 0.04 versus 2.02 ± 0.02), which is characteristic of immunodepressive conditions. In patients of the main group, an increase in IgA production was recorded on average to 2.05 ± 0.06 g/l, which is by 11.41% higher compared to the indicator (1.84 ± 0.02 g/l) of the control group. In workers of industries with harmful factors, with a dystrophic-inflammatory process in the periodontium, the level of average globular circulating immune complexes exceeded the value of healthy people (0.10 ± 0.03 mg/ml) by 2.5 times. This may indicate exhaustion of the clearance function of the phagocytic system and the development of hyperimmune complex syndrome. Conclusion. Having studied individual indicators of the capillary blood of the gums and oral fluid, we can conclude that in chronic exogenous intoxication with industrial xenobiotics, phenomena of acquired combined immune deficiency were observed, which were manifested by a decrease in the immuno-regulatory index CD4+/CD8+; a violation in the humoral link of immunity – activation of IgA, IgG production and a decrease in sIgA secretion, disorders of the phagocytic system, manifested by a decrease in the phagocytic activity of neutrophils and activation of oxygen-dependent metabolism (nitroblue tetrazolium test), which indicated, on the one hand, the strengthening of redox processes in these cells and a certain antibacterial potential of them, but on the other hand – the deepening of the inflammatory process of autogenous origin. Exhaustion of the body's compensatory capabilities confirmed the increase in the level of medium-globular circulating immune complexes. Therefore, significant changes in the non-specific protection of the oral cavity were observed in industrial workers suffering from generalized periodontitis