Extracellular ATP induces cytokine expression and apoptosis through P2X7 receptor in murine mast cells

Extracellular ATP and other nucleotides act through specific cell surface receptors and regulate a wide variety of cellular responses in many cell types and tissues. In this study, we demonstrate that murine mast cells express several P2Y and P2X receptor subtypes including P2X(7), and describe functional responses of these cells to extracellular ATP. Stimulation of bone marrow-derived mast cells (BMMC), as well as MC/9 and P815 mast cell lines with millimolar concentrations of ATP, resulted in Ca(2+) influx across the cellular membrane and cell permeabilization. Moreover, brief exposures to ATP were sufficient to induce apoptosis in BMMCs, MC/9, and P815 cells which involved activation of caspase-3 and -8. However, in the time period between commitment to apoptosis and actual cell death, ATP triggered rapid but transient phosphorylation of multiple signaling molecules in BMMCs and MC/9 cells, including ERK, Jak2, and STAT6. In addition, ATP stimulation enhanced the expression of several proinflammatory cytokines, such as IL-4, IL-6, IL-13, and TNF-alpha. The effects of ATP were mimicked by submillimolar concentrations of 3-O-(4'-benzoyl)-benzoyl-benzoyl-ATP, and were inhibited by pretreatment of mast cells with a selective blocker of human and mouse P2X(7) receptor, 1[N,O-bis(5-isoquinolinesulphonyl)-N-methyl-l-tyrosyl]-4-phenylpiperazine, as well as oxidized ATP. The nucleotide selectivity and pharmacological profile data support the role for P2X(7) receptor as the mediator of the ATP-induced responses. Given the importance of mast cells in diverse pathological conditions, the ability of extracellular ATP to induce the P2X(7)-mediated apoptosis in these cells may facilitate the development of new strategies to modulate mast cell activities.     

Assessment of thermal load on transported goats administered with ascorbic acid during the hot-dry conditions

The major factor in the induction of physiological stress during road transportation of livestock is the complex fluctuations of the thermal transport microenvironment, encountered when animals are transported across different ecological zones. Recommended guidelines on optimum “on-board” conditions in which goats should be transported are lacking, and there are no acceptable ranges and limits for the thermal loads to which goats may be subjected during long-distance road transportation in hot-dry conditions. Panting score (PS), rectal temperature (RT), heart rate (HR) and respiratory rate (RR) were employed as reliable stress indices to assess the effects of different thermal loads, measured as temperature humidity index (THI), encountered in the vehicle during 12 h of road transportation of 40 goats, and to suggest the administration of 100 mg/kg body weight of ascorbic acid (AA) as an ameliorating agent. The results obtained showed that the PS, RT, HR and RR rose above normal reference values with increase in the THI and journey duration. The rise in PS value, which is a visual indicator of the severity of thermal load, was the most pronounced. The results suggest that values of THI in the vehicle up to 94.6 constitute no risk, while at of 100 it presents a moderate risk and above 100 may result in severe stress. The relationships between the thermal load and the physiological variables were positive and significant (P < 0.05). They reflect the degree of stress imposed by each THI value during the transportation, and may be used as recommended ranges and limit thermal load values in transported goats. The results demonstrated that administration of 100 mg/kg body weight of AA before road transportation mitigated the risk of adverse effects of high THI values and other stress factors due to road transportation in goats.     

How a cytokine is chaperoned through the secretory pathway by complexing with its own receptor: lessons from interleukin-15 (IL-15)/IL-15 receptor alpha

While it is well appreciated that receptors for secreted cytokines transmit ligand-induced signals, little is known about additional roles for cytokine receptor components in the control of ligand transport and secretion. Here, we show that interleukin-15 (IL-15) translocation into the endoplasmic reticulum occurs independently of the presence of IL-15 receptor α (IL-15Rα). Subsequently, however, IL-15 is transported through the Golgi apparatus only in association with IL-15Rα and then is secreted. This intracellular IL-15/IL-15Rα complex already is formed in the endoplasmic reticulum and, thus, enables the further trafficking of complexed IL-15 through the secretory pathway. Just transfecting IL-15Rα in cells, which transcribe but normally do not secrete IL-15, suffices to induce IL-15 secretion. Thus, we provide the first evidence of how a cytokine is chaperoned through the secretory pathway by complexing with its own high-affinity receptor and show that IL-15/IL-15Rα offers an excellent model system for the further exploration of this novel mechanism for the control of cytokine secretion.     

Effect of a pseudovolume image in multifocus radiography

The effect of a pseudovolume image was theoretically substantiated on the basis of calculations. This was experimentally studied using with microfocus x-ray diagnostic apparatuses. Digital microfocus and standard x-ray study were compared using an experimental material. Microfocus x-ray study is a highly informative technique that shows a greater possibility to provide volume images of an object than does standard x-ray study.     

Capabilities of digital microfocal x-ray study in the evaluation of reparative regeneration of bone tissue in an experiment

Digital microfocal x-ray study was experimentally studied in animals to examine the time course of changes in their bone regeneration. Sixteen Chinchila rabbits whose bone defect in the angle of the mandibular ramus had been closed with the osteoplastic material Gapcol with the applied allogeneic, autologous stem cells isolated from rabbit adipose tissue and human plasma enriched with thrombocytic growth factors were examined. The capabilities of digital microfocal x-ray study versus x-ray computed tomography were compared in the evaluation of reparative regeneration of bone tissue. The results of radiation studies were verified with the data of scanning electron microscopy.     

Novel aryl and heteroaryl substituted N-[3-(4-phenylpiperazin-1-yl)propyl]-1,2,4-oxadiazole-5-carboxamides as selective GSK-3 inhibitors

Synthesis, biological evaluation, and SAR dependencies for a series of novel aryl and heteroaryl substituted N-[3-(4-phenylpiperazin-1-yl)propyl]-1,2,4-oxadiazole-5-carboxamide inhibitors of GSK-3beta kinase are described. The inhibitory activity of the synthesized compounds is highly dependent on the character of substituents in the phenyl ring and the nature of terminal heterocyclic fragment of the core molecular scaffold. The most potent compounds from this series contain 3,4-di-methyl or 2-methoxy substituents within the phenyl ring and 3-pyridine fragment connected to the 1,2,4-oxadiazole heterocycle. These compounds selectively inhibit GSK-3beta kinase with IC(50) value of 0.35 and 0.41 microM, respectively.     

Strongyloides ratti infection induces expansion of Foxp3+ regulatory T cells that interfere with immune response and parasite clearance in BALB/c mice

To escape expulsion by their host's immune system, pathogenic nematodes exploit regulatory pathways that are intrinsic parts of the mammalian immune system, such as regulatory T cells (Tregs). Using depletion of Treg mice, we showed that Foxp3(+) Treg numbers increased rapidly during infection with the nematode Strongyloides ratti. Transient depletion of Tregs during the first days of infection led to dramatically reduced worm burden and larval output, without aggravation of immune pathology. The transient absence of Tregs during primary infection did not interfere with the generation of protective memory. Depletion of Tregs at later time points of infection (i.e., day 4) did not improve resistance, suggesting that Tregs exert their counterregulatory function during the priming of S. ratti-specific immune responses. Improved resistance upon early Treg depletion was accompanied by accelerated and prolonged mast cell activation and increased production of types 1 and 2 cytokines. In contrast, the blockade of the regulatory receptor CTLA-4 specifically increased nematode-specific type 2 cytokine production. Despite this improved immune response, resistance to the infection was only marginally improved. Taken together, we provide evidence that Treg expansion during S. ratti infection suppresses the protective immune response to this pathogenic nematode and, thus, represents a mechanism of immune evasion.