Consequences of 1,4-Butanediol Misuse: A Review.

Gamma-hydroxybutyrate (GHB), along with its precursors, 1,4-butanediol (1,4-BD) and gamma-butyrolactone (GBL), are potent central depressant agents widely illicitly used for their euphoric and relaxant effects. The article presents a review of the literature on the 1,4-BD misuse, the clinical picture of intoxication, development of addiction and delirium. The available evidence shows that 1,4-BD is a substance with its own psychoactive effects, a high addiction potential and potentially severe withdrawal symptoms.

[The Thymic Hormone Thymosin-1α Reduces the Pro-Inflammatory Response of Raw 264.7 Cells Induced by Endotoxin].

The aim of this work was to study the effects of thymosin-1 alpha (Tα1) on the anti-inflammatory response of RAW 264.7 macrophages cultured in the presence of lipopolysaccharide (LPS) from the walls of gram-negative bacteria. As well, we evaluated production of pro-inflammatory cytokines and the activity of the NF-κB and SAPK/JNK signaling pathways. In addition, the level of expression of a number of genes that regulate cell apoptosis, as well as the activity of receptors involved in the pro-inflammatory response, was determined. First, the addition of Tα1 normalized the level of cytokine production to varying degrees, with a particularly noticeable effect on IL-1ß and IL-6. Second, the addition of Tα1 normalized the activity of the NF-κB and SAPK/JNK signaling cascades and the expression of the Tlr4 gene. Third, Tα1 significantly reduced p53 and the activity of the P53 gene, which is a marker of cell apoptosis. Fourth, it was shown that the increase in Ar-1 gene expression under the influence of LPS was significantly reduced using Tα1. Thus, it was found that the presence of Tα1 in the RAW 264.7 cell culture medium significantly reduced the level of the pro-inflammatory response of cells.

Protective effect of exogenous peroxiredoxin 6 and thymic peptide thymulin on BBB conditions in an experimental model of multiple sclerosis.

This study aimed to assess the effects of the immunomodulator thymulin, a thymic peptide with anti-inflammatory effects, and peroxiredoxin 6 (Prdx6), an antioxidant enzyme with dual peroxidase and phospholipase A2 activities, on the blood‒brain barrier (BBB) condition and general health status of animals with relapsing-remitting experimental autoimmune encephalomyelitis (EAE), which is a model of multiple sclerosis in humans. Both thymulin and Prdx6 significantly improved the condition of the BBB, which was impaired by EAE induction, as measured by Evans blue dye accumulation, tight-junction protein loss in brain tissue, and lymphocyte infiltration through the BBB. The effect was associated with significant amelioration of EAE symptoms. Thymulin treatment was accompanied by a decrease in immune cell activation as judged by interleukin-6, -17, and interferon-gamma cytokine levels in serum and NF-kappaB cascade activation in splenocytes of mice with EAE. Prdx6 did not induce significant immunomodulatory effects but abruptly decreased EAE-induced NOX1 and NOX4 gene expression in brain tissue, which may be one of the possible mechanisms of its beneficial effects on BBB conditions and health status. The simultaneous administration of thymulin and Prdx6 resulted in complete symptomatic restoration of mice with EAE. The results demonstrate prospective strategies for multiple sclerosis treatment.

Geldanamycin Enhances the Radioprotective Effect of Peroxyredoxin 6 in Irradiated 3T3 Fibroblasts.

The aim of the study was to evaluate the possibility of increasing the radioprotective potential of peroxiredoxin 6 (Prdx6) and its mutant form S32A by their combined use with geldanamycin (GA) for 3T3 fibroblasts irradiated with X-rays at a dose of 6 Gy. The mutant enzyme S32A, which does not have phospholipase activity, exhibits a more pronounced radioprotective activity when combined with GA. The use of this combination of radioprotective drugs completely abolishes the peak of NF-κB activity in irradiated 3T3 cells. Another transcription factor, p53, which is an indicator of the level of cell apoptosis and increases upon irradiation, is also reduced by S32A in combination with GA. The low-molecular-weight protein p21, which is a marker of cell senescence and whose production increases upon irradiation, is also normalized when S32A is used in combination with GA. In addition, the use of this combination of radioprotective drugs significantly reduces the stress response of 3T3 cells to X-ray irradiation.

[Antitumor Activity of the Combination of Topotecan and Tyrosyl-DNA-Phosphodiesterase 1 Inhibitor on Model Krebs-2 Mouse Ascite Carcinoma].

Topotecan is a cytostatic drug from the camptothecin group, it acts by inhibiting topoisomerase 1 (TOP1). Tyrosyl-DNA phosphodiesterase 1 (TDP1) is capable of interfering with the action of TOP1 inhibitors, reducing their therapeutic efficacy. Suppression of TDP1 activity may enhance the effects of topotecan. In this work, we investigated the effect of the antitumor drug topotecan alone and in combination with a TDP1 inhibitor, a hydrazinothiazole derivative of usnic acid, on Krebs-2 mouse ascites tumors. We have previously shown that this derivative efficiently inhibits TDP1. In the present work, we show that both topotecan and the TDP1 inhibitor have an antitumor effect when evaluated separately. The combination of topotecan and the TDP1 inhibitor additively reduces both the weight of the ascites tumor and the number of cells in ascites. In mice, the TDP1 inhibitor alone or in combination with topotecan eliminated the tumor cells. After the combined intraperitoneal administration of these two compounds, we observed cells in which lipid droplets occupied almost the entire cytoplasm and the accumulation of cell detritus, which was absent in the samples collected from mice treated with each compound separately.

Participation of Hsp70 and Hsp90α Heat Shock Proteins in Stress Response in the Course of Type 1 Diabetes Mellitus.

The role of two heat shock proteins, Hsp70 and Hsp90α, on stress response in mice with severe diabetes mellitus induced by a high dose of alloxan (500 mg/kg body weight), as well as in RIN-m5F ß cells cultured in the presence of cytokines (IL-1 and TNF-α) was studied. Our results showed that severe type 1 diabetes mellitus (T1D) caused a higher expression of Hsp90α, but not Hsp70. Moreover, injections of the peroxiredoxin 6 antioxidant enzyme (PRDX6) did not affect the expression of these chaperones. Conversely, pro-inflammatory cytokines added to ß-cells caused a significant increase in the expression of Hsp90α and, substantially, Hsp70. Moreover, cells cultivated in the presence of PRDX6 were more susceptible to the cytokine effect. Thus, in the course of severe alloxan-induced T1D, no protective role of the heat shock proteins, was revealed, and their expression level was not increased by PRDX6. At the same time the protective potential of these proteins was shown in vitro with the use of RIN-m5F ß cells. Thus, the system of heat shock proteins was unable to prevent the devastating effects of severe T1D accompanied by high animal mortality.

Precursors of thymic peptides as stress sensors.

INTRODUCTION: A large volume of data indicates that the known thymic hormones, thymulin, thymopoietin, thymosin-α, thymosin-ß, and thymic humoral factor-y2, exhibit different spectra of activities. Although large in volume, available data are rather fragmented, resulting in a lack of understanding of the role played by thymic hormones in immune homeostasis. AREA COVERED: Existing data compartmentalizes the effect of thymic peptides into 2 categories: influence on immune cells and interconnection with neuroendocrine systems. The current study draws attention to a third aspect of the thymic peptide effect that has not been clarified yet, wherein ubiquitous and highly abundant intranuclear precursors of so called 'thymic peptides' play a fundamental role in all somatic cells. EXPERT OPINION: Our analysis indicated that, under certain stress-related conditions, these precursors are cleaved to form immunologically active peptides that rapidly leave the nucleus and intracellular spaces, to send 'distress signals' to the immune system, thereby acting as stress sensors. We propose that these peptides may form a link between somatic cells and immune as well as neuroendocrine systems. This model may provide a better understanding of the mechanisms underlying immune homeostasis, leading thereby to the development of new therapeutic regimes utilizing the characteristics of thymic peptides.

Protective Effect of Peroxiredoxin 6 Against Toxic Effects of Glucose and Cytokines in Pancreatic RIN-m5F ß-Cells.

Taking into account a special role of pancreatic ß-cells in the development of diabetes mellitus, the effects of peroxiredoxin 6 (Prx6) on the viability and functional activity of rat insulinoma RIN-m5F ß-cells were studied under diabetes-simulating conditions. For this purpose, the cells were cultured at elevated glucose concentrations or in the presence of pro-inflammatory cytokines (TNF-α and IL-1) known for their special role in the cytotoxic autoimmune response in diabetes. It was found that the increased glucose concentration of 23-43 mM caused death of 20-60% ß-cells. Prx6 added to cells significantly reduced the level of reactive oxygen species and protected the RIN-m5F ß-cells from hyperglycemia, reducing the death of these cells by several fold. A measurement of insulin secretion by the RIN-m5F ß-cells showed a significant stimulatory effect of Prx6 on the insulin-producing activity of pancreatic ß-cells. It should be noted that the stimulatory activity of Prx6 was detected during culturing the cells under both normal and unfavorable conditions. The regulation of the NF-κB signaling cascade could be one of the mechanisms of Prx6 action on ß-cells, in particular, through activation of RelA/p65 phosphorylation at Ser536.

Immune response in the relapsing-remitting experimental autoimmune encephalomyelitis in mice: The role of the NF-κB signaling pathway.

Characteristics of the mouse model of relapsing-remitting experimental autoimmune encephalomyelitis (rEAE) closely resemble manifestations of multiple sclerosis in humans. In the present study, we investigated the mechanisms of inflammatory response, focusing on NF-κB pathway activation. Cytokine response in rEAE mice was multiphasic: the early phase was characterized by the increase in interferon-γ level in plasma. In the later stage, the level of interleukin-17, but not of interferon-γ, was increased. The early phase of rEAE was also accompanied by increased RelA/p65 phosphorylation at Ser276 in spleen cells, whereas the rEAE maintenance phase was characterized by RelA/p65 phosphorylation at Ser536 and IKK phosphorylation. The IKKα/ß inhibitor reduced interleukin-17 and interferon-γ levels in plasma and alleviated rEAE symptoms. The IKKα/ß inhibitor decreased IKK and p65(Ser536) phosphorylation, but doubled p65(Ser276) phosphorylation in rEAE mice. The increased RelA/p65(Ser276) phosphorylation coincided in time with the production of interferon-γ, Hsp72, and the early phase of IL-17 generation, whereas increased RelA/p65(Ser536) phosphorylation coincided with the activation of IKK, SAPK/JNK, and p53, as well as the late phase of IL-17 production, indicating the role of the RelA/p65 phosphorylation events in the induction and maintenance of rEAE.

The Role of p38 and CK2 Protein Kinases in the Response of RAW 264.7 Macrophages to Lipopolysaccharide.

The role of protein kinases p38 and CK2 (casein kinase II) in the response of RAW 264.7 macrophages to the lipopolysaccharide (LPS) from gram-negative bacteria was studied. Using specific p38 and CK2 inhibitors (p38 MAP kinase Inhibitor XI and casein kinase II Inhibitor III, respectively), we investigated the effects of these protein kinases on (i) LPS-induced activation of signaling pathways involving nuclear factor κB (NF-κB), stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), p38, and interferon regulatory factor 3 (IRF3); (ii) expression of Toll-like receptor 4 (TLR4) and inducible heat-shock proteins HSP72 and HSP90; and (iii) production of interleukins IL-1α, IL-1ß, IL-6, tumor necrosis factor α, and IL-10. Activation of the proapoptotic signaling in the macrophages was evaluated from the ratio between the active and inactive caspase-3 forms and p53 phosphorylation. Six hours after LPS addition (2.5 µg/ml) to RAW 264.7 cells, activation of the TLR4 signaling pathways was observed that was accompanied by a significant increase in phosphorylation of IκB kinase α/ß, NF-κB (at both Ser536 and Ser276), p38, JNK, and IRF3. Other effects of macrophage incubation with LPS were an increase in the contents of TLR4, inducible heat-shock proteins (HSPs), and pro- and anti-inflammatory cytokines, as well as slight activation of the pro-apoptotic signaling in the cells. Using inhibitor analysis, we found that during the early response of macrophages to the LPS, both CK2 and p38 modulate activation of MAP kinase and NF-κB signaling pathways and p65 phosphorylation at Ser276/Ser536 and cause accumulation of HSP72, HSP90 and the LPS-recognizing receptor TLR4. Suppression of the p38 MAP kinase and CK2 activities by specific inhibitors (Inhibitor XI and Inhibitor III, respectively) resulted in the impairment of the macrophage effector function manifested as a decrease in the production of the early-response proinflammatory cytokines and disbalance between the pro- and anti-apoptotic signaling pathways leading presumably to apoptosis development. Taken together, our data indicate the inefficiency of therapeutic application of p38 and CK2 inhibitors during the early stages of inflammatory response.

Involvement of the p38 MAPK signaling cascade in stress response of RAW 264.7 macrophages.

The role of the p38 MAPK signaling cascade was studied in stress response of RAW 264.7 macrophages to extremely low-intensity centimeter microwaves. Irradiation stimulated production of a number of cytokines (IL-1, IL-6, TNF-α, INF-γ and IL-10), as well as induced activation of the signaling cascades NF- κB and p38 MAPK, and enhanced expression of Hsp72 heat shock protein. In the presence of the cascade p38 MAPK inhibitor (p38 MAP kinase inhibitor XI), the stimulating effects of electromagnetic waves were abrogated either completely (for NF-κB and Hsp72) or partially (for p38 MAPK and cytokines). The results obtained are indicative of a high sensitivity of the signaling cascade p38 MAPK to the effect of low-intensity physical fields.

Antiretroviral Activity Of a Novel Pyrimidyl-Di(Diazaspiroalkane) Derivative.

A novel compound, 3,3'-(5-nitropyrimidine-4,6-diyl)bis-3,12-diaza-6,9-diazoniadispiro[5.2.5.2]hexadecane tetrachloride dihydrochloride, was synthesized. The compound was found to inhibit the replication of various viral families by blocking specific heparan sulfate receptors on the host cell's surface. In experiments, the compound was found to be highly effective against several strains of HIV retroviruses.

The role of CK2 protein kinase in stress response of RAW 264.7 macrophages.

The role of casein kinase 2 (CK2) in the activation of the key NF-κB signaling cascade and other signaling and stress proteins during stress induced by exposure to nonthermal low-intensity electromagnetic radiation has been investigated. The ability of CK2 to regulate the activation of the NF-κB signaling cascade and upregulate the proinflammatory cytokine production and TLR4 receptor expression has been demonstrated. The existence of an alternative signaling pathway for NF-κB cascade activation directly involving protein kinase CK2 has been demonstrated using inhibition analysis in cells stressed by microwave irradiation.

Modulation of inflammatory response in mice with severe autoimmune disease by thymic peptide thymulin and an inhibitor of NF-kappaB signalling.

To investigate some cellular and molecular aspects of the autoimmune response and anti-inflammatory efficiency of potential therapeutic agents in a severe form of experimental autoimmune encephalomyelitis (sEAE), an inhibitor of NF-kappaB signalling, IKK Inhibitor XII, and/or thymic peptide thymulin, were injected intraperitoneally at 1.8 and 0.15mg/kg e.o.d, respectively, to C57BL/6 mice immunized with myelin oligodendrocyte glycoprotein and several adjuvants. The immunization induced high lethality in three weeks. The biphasic cytokine response observed in earlier and delayed phases was attributed to the activity of Th1 and Th17 cells, respectively. Phosphorylation of RelA protein from the NF-kappaB family increased during the earlier phase and decreased in the delayed phase. SAPK/JNK signalling protein and heat shock protein Hsp72 significantly increased in lymphocytes. Both the IKK Inhibitor XII and thymulin reduced disease severity, attenuated immune imbalance, and increased mouse life-span. Co-administration of the agents produced no additive effect. Both the inhibitor and thymulin reduced the Th1 response but not the Th17 response. Therefore, RelA-associated Th1 activation and RelA-independent Th17 activation occurred in sEAE. Thymulin and the inhibitor demonstrate similar patterns of activity, potentially through the RelA pathway inhibition, resulting in a partial therapeutic effect on the animals' health status.

Changes in immune cell signalling, apoptosis and stress response functions in mice returned from the BION-M1 mission in space.

To explore the effect of the spaceflight environment on immunity in animals, C57/BL6 mice flown on a 30-day space high-orbit satellite mission (BION-M1) were analyzed. Cytokine response in mice was measured in tandem with the following parameters: the synthesis of inducible forms of the heat shock proteins HSP72 and HSP90α; activity of the NF-κB, IFR3, and SAPK/JNK signalling pathways; and TLR4 expression. In addition, apoptosis in the thymus was measured by caspase-3 and ph-p53/p53 ratio testing. In response to flight environment exposure, mice had a reduction in spleen and thymus masses and decreased splenic and thymic lymphocyte counts. Plasma concentration of IL-6 and IFN-γ but not TNF-α was decreased in C57BL6 mice. The NF-κB activity in splenic lymphocytes through the canonical pathway involving IκB degradation was significantly increased at 12h after landing. One week after landing, however, the activity of NF-κB was markedly decreased below even the control values. Non-canonical NF-κB activity increased during the whole observation period. The activities of SAPK/JNK and IRF-3 were invariable at 12h but significantly increased 7 days after landing. The expression of Hsp72 and Hsp90α was somewhat increased 12h (Hsp72) and 7 days (Hsp90α). TLR4 expression in splenic cells was significantly increased only at 12h, returning to normal 7 days after landing. To assess the apoptosis in thymus lymphocytes, caspase-3 and levels of p53 protein along with its phosphorylated form were measured in thymic lymphocytes. The results indicated that the high-orbit spaceflight environment caused an increase in the level of p53 but more notably in the activated, phosphorylated form of the p53 protein. The calculated ratio of the active to inactive forms of the protein (ph-53/p53) 12h after landing increased by more than twofold, indicating the apparent induction of apoptosis in thymus cells. Interestingly, 7 days after the landing, this ratio was not restored, but rather increased: the specified ratio was four times higher compared to the ground-based control. Measurements of caspase-3 in thymic cells indicated more expressive increase in apoptosis. Taken together, the results of the present study indicate that spaceflight induces an imbalance in the immunity of mice, showing variation in signalling, apoptosis and stress response that are not restored by 7 days after landing. These changes are distinguished from classic stress-related alterations usually caused by conventional stressors.

Anti-inflammatory effects of IKK inhibitor XII, thymulin, and fat-soluble antioxidants in LPS-treated mice.

The present study was designed to compare the anti-inflammatory effects of several agents applied in vivo, namely, a synthetic inhibitor of the NF-κB cascade, fat-soluble antioxidants, and the thymic peptide thymulin. Cytokine response in LPS-treated mice was analysed in tandem with the following parameters: the synthesis of inducible forms of the heat shock proteins HSP72 and HSP90α; activity of the NF-κB and SAPK/JNK signalling pathways; and TLR4 expression. Inflammation-bearing Balb/c male mice were pretreated with an inhibitor of IKK-α/ß kinases (IKK Inhibitor XII); with thymulin; with dietary coenzyme Q9, α-tocopherol, and ß-carotene; or with combinations of the inhibitor and peptide or antioxidants. Comparable anti-inflammatory effects were observed in inflammation-bearing mice treated separately with thymulin or with dietary antioxidants administered daily for two weeks before LPS treatment. When LPS-injected mice were treated with the inhibitor and antioxidants together, neither plasma cytokines, signal proteins, nor heat shock proteins recovered more efficiently than when mice were treated with these agents separately. In contrast to antioxidant diet, the thymulin was shown to increase the effect of IKK Inhibitor XII in preventing IKK activation in LPS-treated mice.

Thymic peptides restrain the inflammatory response in mice with experimental autoimmune encephalomyelitis.

Modulation of autoimmune inflammation by the thymic peptides thymulin and thymopentin was studied in mice with acute experimental autoimmune encephalomyelitis (EAE), which resembles multiple sclerosis in humans. EAE was induced in NZW mice by a single immunisation with myelin basic protein coupled with adjuvants. Visible signs of pathology appeared on days 12-14 after the immunisation, peaked on days 20-25, were retained up to day 45, and then reverted. A biphasic cytokine response was also detected. In the "early" phase, which started at day 35, increased levels of interferon-gamma and interleukin-6 in the blood were observed; during the "delayed" phase, which started at day 48, the levels of plasma interleukin-17 and tumour necrosis factor-alpha were also raised. In addition, the phosphorylation of NF-kappaB signalling proteins and the production of heat shock protein Hsp72 were significantly increased in splenic lymphocytes from EAE-bearing mice. When applied intraperitoneally every other day for 30 days, either thymulin or thymopentin (15 µg per 100g of body weight) significantly reduced the disease severity compared to untreated EAE mice. The effect of thymulin but not thymopentin remained after its withdrawal. Thymulin reduced the cytokine response in both the early and the delayed phases, whereas thymopentin only reduced the "early phase cytokines" (IL-6 and interferon-gamma). Both peptides significantly reduced the level of phosphorylation of the NF-kappaB signalling protein IKK and the production of Hsp72 protein. The data presented here indicate the presence of time-dependent immune responses in EAE-bearing mice, which may be associated with the Th1 and Th17 subpopulations of T-cells. Thymulin and thymopentin demonstrated different patterns of activity, most likely via mechanisms involved in NF-kappa B signalling and Hsp72 expression.

[Significance of static pressure-volume loop and lung computed tomography for differential diagnostics of parenchymal respiratory insufficiency].

UNLABELLED: Purpose of the study was to determine a significance of static pressure-volume loop and lung computed tomography for differential diagnostics of parenchymal lung failure developing during mechanical ventilation. MATERIALS AND METHODS: 75 patients (42 males and 33 females) with acute lung failure due to parenchymal lung injury during mechanical ventilation were included in to the research. Criteria of including into the research were age over 15, ARDS symptoms absence before respiratory support beginning and modified American-European Consensus Conference ARDS criteria presence during mechanical ventilation (AECC ARDS criteria, 1994--PaO2/FiO2 < 250 mmHg). Lung computed tomography (CT), static compliance and plateau measurement were performed in all patients. Static pressure-volume loop was plotted in 23 patients. RESULTS: diffuse alveolar damage was diagnosed by CT in 24.3% of patients and "wet sponge" symptom in 10.7% of patients. Dorsal atelectasis (77.3%) and ventilator-associated pneumonia (VAP) (82.7%) were diagnosed in most of patients with AECC ARDS criteria. Sensitivity and specificity of PaO2/FiO2 ratio were too low for diagnostics of ARDS (AUROC 0.67) Patients with diffuse alveolar damage had plateau pressure 25 mbar (95% CI 22-32), while patients with local lung injury (VAP or atelectasis) had significantly lower plateau pressure--20 mbar (95% CI 18-22) (p = 0.014). Elevation of plateau pressure over 30 mbar predicted diffuse alveolar damage with specificity of 100%. Lower inflection point values on the static pressure-volume loop was higher in patients with diffuse alveolar damage than in patients with local lung injury--12 mbar (95% CI 7-17) vs. 6 mbar (95% CI 5-10), (p = 0.042, n = 23). Effective (linear) compliance had poor prognostic value for differential diagnostics of acute respiratory failure due to parenchimal lung injury (p = 0.023). CONCLUSION: Lung CT plays leading role in differential diagnostics of parenchymal lung failure developing during mechanical ventilation. In the luck of CT scan elevation of plateau pressure over 30 mbar and values of lower inflection point on the static pressure-volume loop over 12 mbar can predict ARDS.