Electrophysiological actions of N-[1-[4-(4-fluorophenoxy)butyl]-4-piperidinyl]-N-methyl-2-benzothiazolamine (R56865) on CA1 neurons of the rat hippocampal slice during hypoxia

The electrophysiological effects of N-[1-[4-(4-fluorophenoxy)butyl]-4-piperidinyl]-N-methyl-2-benzothiazolamine (R56865), a drug which protects heart cells from ischemia-induced arrhythmias, was studied on intracellularly-recorded CA1 neurons of the rat hippocampal slice under normal or hypoxic conditions. On normoxic cells R56865 (1 μM) reduced firing accommodation without changing passive membrane properties, spike characteristics or synaptic transmission. On hypoxic cells R56865 selectively reduced the amplitude of hypoxia-induced membrane depolarization and partly counteracted the depression of synaptic transmission evoked by Schaffers collateral stimulation. Despite its influence on repetitive firing properties, R56865 might be useful to limit the extent of cellular depolarizing responses to hypoxia.     

Baclofen antagonism by 4-amino-3-(5-methoxybenzo[b]furan-2-yl) butanoic acid in the cat spinal cord

When administered microelectrophoretically, 4-amino-3-(5-methoxybenzo[b]furan-2-yl)butanoic acid (MBFG) reversibly reduced the presynaptic depression by (−)-baclofen of the monosynaptic excitation of spinal interneurones by impulses in muscle low-threshold afferent fibres of the cat as well as the postsynaptic depression by (−)-baclofen of the firing of these neurones. MBFG, as an antagonist of (−)-baclofen, may be useful in investigating the structure-activity relationships of central and peripheral baclofen receptors.     

TCR-independent CD137 (4-1BB) signaling promotes CD8+-exhausted T cell proliferation and terminal differentiation

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Expression and function of B7-1 (CD80) and B7-2 (CD86) on human epidermal Langerhans cells

In addition to T cell receptor triggering, activation of T cells requires co-stimulatory signals that have been shown to be mainly initiated through CD28. We analyzed the expression and function of the two ligands for CD28, B7-1 (CD80) and B7-2 (CD86), on human Langerhans cells (LC), the antigen-presenting cells from epidermis. Human LC freshly isolated from epidermis (fLC) expressed significant level of B7-2, which was increased upon a short culture in vitro. In contrast, B7-1 was undetectable on fLC but appeared at the cell surface after a 3-day culture in vitro. Pre-incubation of 18-h cultured LC with anti-B7-2 monoclonal antibodies (mAb) was sufficient to abrogate the binding of CTLA4-Ig fusion protein, while a combination of both mAb against B7-1 and B7-2 was necessary to obtain a complete inhibition of CTLA4-Ig binding on 3-day cultured LC, showing the absence of a third CTLA4 ligand. The function of B7-1 and B7-2 on human LC has been analyzed by adding mAb at the beginning of mixed epidermal cell lymphocyte reactions. Anti-B7-2 mAb and CTLA4-Ig, but not anti-B7-1 mAb, strongly inhibited allogeneic, as well as recall antigen-induced T cell proliferation supported by fLC or 3-day cultured LC. Collectively, these results demonstrate that B7-2 is the major ligand for CD28/CTLA4 at the LC surface and that it plays a crucial role in human LC co-stimulatory function with little, if any, dependence on B7-1 expression.     

Specificity of base substitution mutations induced by the dietary carcinogens 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) in Salmonella

The base pair substitution mutational profiles induced by the heterocyclic amine cooked food mutagens PhIP and IQ in Salmonella typhimurium strains TA100 and TA1535 were determined by colony hybridization analysis. Both PhIP and IQ induced predominantly G→TA transversions in strain TA100 (rfa,ΔuvrB/pKM101) with a pronounced preference for the second codon position (CC→CAC; 72% of total). PhIP also reverted strain TA1535 (rfa, ΔuvrB) efficiently at concentrations similar to those required for strain TA100. In contrast to the PhIP-induced mutational profile observed in strain TA100, in strain TA1535 PhIP induced exclusively G→AT transitions at the second codon position (CC→CTC; 96–99% of total). Base substitution mutagenesis induced by heterocyclic amines related to PhIP is generally SOS-dependent, requiring the presence of plasmid pKM101 in Salmonella hisG46 strains. Thus, the SOS dependent reversion of S. typhimurium strain TA100 probably reflects error-prone lesion bypass at the major PhIP- guanosine adduct at the C-8 position. The G→AT transition mutations induced by PhIP in strain TA1535 appear to be SOS-independent, however, suggesting that these mutations may arise from the formation of PhIP-DNA adducts other than the replication-blocking C8-dG lesion. Environ. Mol. Mutagen. 31:327–332, 1998 © 1998 Wiley-Liss, Inc.     

不同剂量(+)-2-(1-羟基-4-环己酮)乙基咖啡酸酯对大鼠关节炎模型和全血炎症模型的药效学研究

目的:对(+)-2-(1-羟基-4-环己酮)乙基咖啡酸酯(HOEC)在大鼠关节炎模型上进行药效学评价,并利用大鼠全血花生四烯酸(AA)代谢模型,探究引起HOEC对关节炎的非剂量依赖性治疗作用的可能机理。方法:(1)采用大鼠胶原诱导性关节炎(CIA)模型,研究3个不同剂量的HOEC对CIA的治疗情况,并用免疫组化方法检测关节组织中胞质磷脂酶A_2(cPLA_2)、5-脂氧合酶(5-LOX)和环氧合酶2 (COX-2)的表达水平;(2) ELISA检测HOEC和其体内代谢物咖啡酸对大鼠全血AA代谢模型中代谢产物的作用。结果:(1) HOEC对大鼠CIA有治疗作用,但高剂量(10 mg/kg)组的治疗效果不如低(1 mg/kg)和中剂量(3 kg/kg)组;(2) HOEC对大鼠关节组织中cPLA2、5-LOX和COX-2的表达水平均有抑制作用;(3) HOEC对大鼠全血AA代谢模型中LOX和COX通路的代谢产物均有抑制作用,而咖啡酸对这些代谢产物的抑制作用弱于HOEC。结论:HOEC对大鼠CIA模型的抗炎作用可能与抑制关节组织中cPLA_2、5-LOX和COX-2的表达有关; HOEC对治疗大鼠CIA的非剂量依赖现象可能与其代谢产物咖啡酸对AA代谢产物生成的抑制作用弱于HOEC有关。     

Human recombinant IL-1 receptor antagonist (IL-1Ra) inhibits leukotriene B4 generation from human monocyte suspensions stimulated by lipopolysaccharide (LPS).

The effect of human recombinant IL-1 receptor antagonist (hrIL-1Ra) on leukotriene B4 (LTB4) release was investigated in activated human monocyte cultures. To stimulate LTB4 generation, LPS was used as an agonist. Detection was performed with the highly sensitive radioimmunoassay method. The cells were treated with scalar concentrations using LPS at 1-1000 ng/ml for different periods of time. The greater LTB4 stimulation was found at LPS 100 ng/ml for 18 h incubation time. Preincubation of monocytes with cytochalasin B (CB) (5 micrograms/ml) for 15 min augmented the release of LTB4 when LPS was used. A dose-dependent inhibition was found when human monocytes were pretreated for 10 min with hrIL-1Ra at different concentrations (0.25-250 ng/ml) and then treated with LPS 100 ng/ml for 18 h. Maximum inhibition was observed at the highest concentration of hrIL-1Ra (250 ng/ml). Macrophages treated with a non-selective 5-lipoxygenase inhibitor, nordihydroguaiaretic acid (NDGA), used at 10 microM, added 15 min before LPS 100 ng/ml, produce a dose-dependent inhibition of LTB4. Cells pretreated with arachidonic acid, at various concentrations (10(-9)-10(-5) M) for 10 min and then treated with LPS 100 ng/ml for 18 h, were also inhibited in a dose-dependent manner by hrIL-1Ra in their production of LTB4. The inhibition of LTB4 release by hrIL-1Ra, in LPS-stimulated human monocytes, may suggest an important modulatory role for this new cytokine (monokine) in inflammation and immunity and may hold future therapeutic implications for diseases involving LTB4 as a mediator.     

Swelling Response of Radiation Synthesized 2-Hydroxyethylmethacrylate-co-[2-(methacryloyloxy)ethyl] Trimethylammonium Chloride Hydrogels Under Various In Vitro Conditions

High-energy ⁶⁰Co gamma radiation has been used to synthesize 2-hydroxyethylmethacrylate-co-[2-(methacryloyloxy)ethyl]trimethylammonium chloride (HEMA-co-MAETC) polyelectrolyte hydrogels. HEMA-co-MAETC co-polymer gels were characterized and investigated for swelling behaviour in different swelling conditions. Fourier transformed infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM) techniques were used to characterize the co-polymer gels. Swelling extent of the gels was found to be a linear function of MAETC content in the gels. The effect of ionic strength, temperature, pH, some solutes of biological importance like glucose, urea, and surfactants such as Triton-X and deoxycholic acid on swelling behavior have been reported. The swelling of gels at higher temperature enhanced the swelling rates but not the swelling extent. HEMA-co-MAETC hydrogel exhibited an excellent responsive characteristic to the ionic strength of the swelling medium. It was found that the swelling of the co-polymer gel at 60°C reduced the swelling–deswelling cycle time by approx. 30% without altering the swelling extent. The gels were also investigated for their swelling in aqueous solutions of anionic dyes, acid blue 25 (AB25), acid blue (AB74) and acid yellow 99 (AY99), and were found to be suitable for dye uptake applications.     

Interaction of 2-(9H-purin-6-ylamino)-4-(methylthio) Butanoic Acid with Human Serum Albumin: Fluorescence and Modeling Studies

This study was designed to investigate the interaction between (S)-2-(9H-purin-6-ylamino)-4-(methylthio) butanoic acid (PYMBA) and human serum albumin (HSA) using fluorescence spectroscopy. The combination of UV absorption and molecular docking under simulative physiological conditions was also applied to comprehensively understand the binding mechanism of PYMBA to HSA. Fluorescence data indicated that PYMBA has a strong ability to quench the intrinsic fluorescence of HSA. The binding constants (K) at different temperatures, thermodynamic parameters including enthalpy change (ΔH) and entropy change (ΔS) of PYMBA–HSA were correlated to the relevant fluorescence data, which suggested that the hydrophobic force played a very important role for the PYMBA binding to the HSA. The experimental results were in agreement with the results obtained via a molecular docking study. The effects of other ions on the binding constants were also studied.     

Immunochemical characterization of mouse brain-associated theta (Thy-1) antigen.

The Thy-1 antigens or rat brain and thymus have been isolated and chemically characterized, but those of mice have not been identified. Moreover, it is uncertain whether the antigens are glycolipids or glycoproteins. This study with highly purified preparations of gangliosides GM₁, ¹GD_1a, GD_1b and GT_1b from bovine brain and several ganglioside fractions from mouse brain showed that Thy-1 activity does not reside in gangliosides, but rather in the chloroform-methanol-insoluble residue of brain remaining after extraction of gangliosides. The antigen could be solubilized from this residue with a non-ionic detergent. The antigenic activity of the solubilized preparation was heat-labile but resistant to periodate. The chemical properties of the Thy-1 antigen of mouse brain are discussed.     

Elucidating the role of 5-HT(1A) and 5-HT(7) receptors on 8-OH-DPAT-induced behavioral recovery after experimental traumatic brain injury

8-OH-DPAT is a 5-HT(1A/7) receptor agonist that enhances behavioral recovery after traumatic brain injury (TBI). This study is a first attempt to decipher whether the benefits induced by 8-OH-DPAT after TBI are mediated by 5-HT(1A) or 5-HT(7) receptors. A single i.p. injection of 8-OH-DPAT (0.5 mg/kg) alone or co-administered with either the 5-HT(1A) or 5-HT(7) receptor antagonists WAY 100635 (0.5 mg/kg) or SB 269970 HCl (2.0 mg/kg), respectively, or vehicle control (1.0 mL/kg) was given 15 min after cortical impact or sham injury. Function was assessed by established motor and cognitive tests. No difference in motor performance was observed among the TBI groups. Spatial acquisition was enhanced, relative to vehicle controls, by 8-OH-DPAT alone and when co-administered with WAY 100635, but not when combined with SB 269970 HCl. These data imply that 5-HT(1A) receptor antagonism does not abate the 8-OH-DPAT-induced cognitive benefits, but 5-HT(7) receptor antagonism does, which suggests that the 8-OH-DPAT-induced benefits in this single administration paradigm may be mediated more by 5-HT(7) versus 5-HT(1A) receptors. Evaluation of a specific 5-HT(7) receptor agonist will further elucidate the contribution of 5-HT(1A) and 5-HT(7) receptors on behavioral recovery conferred by acute 8-OH-DPAT treatment after TBI.     

血管紧张素-(1-7)通过抑制TLR4激活和坏死性凋亡的相互作用对抗高糖引起的H9c2心肌细胞损伤

目的:研究血管紧张素-(1-7)[Ang-(1-7)]能否通过抑制Toll样受体4(TLR4)激活和坏死性凋亡的相互作用对抗高糖(HG)引起的H9c2心肌细胞损伤。方法:应用Western blot检测心肌细胞受体相互作用蛋白3(RIP3;反映坏死性凋亡的指标)和TLR4的表达水平;CCK-8法测定心肌细胞存活率;用试剂盒检测细胞培养液中乳酸脱氢酶(LDH)的活性;ELISA检测细胞培养液中白细胞介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)的分泌水平;双氯荧光素染色法测定细胞内活性氧簇(ROS)水平;罗丹明123染色法测定线粒体膜电位(MMP)。结果:HG(35 mmol/L葡萄糖)作用H9c2心肌细胞24 h可使RIP3的表达水平明显升高,应用30μmol/L TAK-242(TLR4抑制剂)和HG共处理心肌细胞24 h可抑制HG对RIP3的上调;另一方面,HG可上调TLR4的表达,100μmol/L坏死性凋亡的特异性抑制剂necrostatin-1(Nec-1)和HG共处理心肌细胞24 h可抑制HG对TLR4的上调;而1μmol/L Ang-(1-7)和HG共处理心肌细胞24 h能同时抑制HG对RIP3和TLR4的上调。此外,1μmol/L Ang-(1-7)、30μmol/L TAK-242或100μmol/L Nec-1与HG共处理心肌细胞均能对抗HG引起的心肌细胞损伤,细胞存活率升高,LDH活性降低,ROS生成和MMP丢失减少,同时IL-1β和TNF-α的分泌减少。结论:Ang-(1-7)通过抑制TLR4激活和坏死性凋亡的相互作用对抗高糖引起的H9c2心肌细胞损伤。     

Five-aminoimidazole-4-carboxamide-1-β-4-ribofuranoside attenuates poly (I:C)-induced airway inflammation in a murine model of asthma

BACKGROUND: Asthma can frequently be induced or exacerbated by respiratory viral infections. Oxidative stress might also play an essential role in the pathogenesis of allergic airway diseases, indicating that antioxidant therapy may have a potential effect in controlling allergic airway diseases. Recent studies showed that 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR) has the potential ability to modulate NADPH oxidase activity, indicating the antioxidant activity of AICAR. This study investigated the inhibitory effects of AICAR as an anti-inflammatory modulator on allergic airway inflammation in murine animal models. METHODS: The anti-inflammatory effects of AICAR were evaluated in two experimental asthma models: (1) an ovalbumin (OVA)-induced experimental asthma model and (2) an OVA plus polyinosinic-polycytidylic acid [poly (I:C)]-induced experimental asthma model to mimic respiratory viral infections. The inhibitory effects of AICAR in poly (I:C)-mediated signalling for NF-kappaB activation and production of TNF-alpha were analysed in vitro. RESULTS: AICAR was shown to have a marginal inhibitory effect in an OVA-induced asthma model. Interestingly, AICAR significantly attenuated poly (I:C)-induced airway hyperresponsiveness and airway inflammation, as shown by the attenuation of the influx of total inflammatory cells and soluble products into bronchoalveolar lavage fluid, such as macrophages, eosinophils, IL-5, IL-13, TNF-alpha and IFN-gamma. AICAR also significantly reduced the serum levels of OVA-specific IgE and IgG2a antibodies. Histologic and flow cytometric studies showed that AICAR inhibited poly (I:C)-induced lung inflammation and the infiltration of CD11b+CD11c+ dendritic cells into the lung. Moreover, AICAR effectively inhibited poly (I:C)-mediated activation of NF-kappaB and the production of TNF-alpha. CONCLUSION: These findings suggest that AICAR may be a novel immunomodulator with promising beneficial effects for the treatment of respiratory viral infection in airway allergic diseases.     

A Novel Synthetic Compound 3-Amino-3-(4-Fluoro-Phenyl)-1H-Quinoline-2,4-Dione (KR22332) Exerts a Radioprotective Effect via the Inhibition of Mitochondrial Dysfunction and Generation of Reactive Oxygen Species

Purpose

Acute side effects of radiation such as oral mucositis are observed in most patients. Although several potential radioprotective agents have been proposed, no effective agent has yet been identified. In this study, we investigated the effectiveness of synthetic compound 3-amino-3-(4-fluoro-phenyl)-1H-quinoline-2,4-dione (KR22332) as a radioprotective agent.

Materials and Methods

Cell viability, apoptosis, the generation of reactive oxygen species (ROS), mitochondrial membrane potential changes, and changes in apoptosis-related signaling were examined in human keratinocyte (HaCaT).

Results

KR22332 inhibited irradiation-induced apoptosis and intracellular ROS generation, and it markedly attenuated the changes in mitochondrial membrane potential in primary human keratinocytes. Moreover, KR22332 significantly reduced the protein expression levels of ataxia telangiectasia mutated protein, p53, and tumor necrosis factor (TNF)-α compared to significant increases observed after radiation treatment.

Conclusion

KR22332 significantly inhibited radiation-induced apoptosis in human keratinocytes in vitro, indicating that it might be a safe and effective treatment for the prevention of radiation-induced mucositis.     

超声微泡介导转染FKBP12.6基因对小鼠H9c2(2-1)心肌细胞中Ca2+浓度的影响

目的:探讨超声微泡介导转染FKBP12.6基因后,对小鼠H9c2(2-1)心肌细胞中Ca2 浓度的影响。方法:将pcDNA3.1-FKBP12.6质粒与白蛋白包裹微泡造影剂混合,经超声转染H9c2(2-1)细胞后,通过倒置显微镜观察心肌细胞生长状况的变化;激光共聚焦显微镜检测细胞内Ca2 浓度的变化;免疫组织化学方法检测FKBP12.6蛋白的表达。结果:超声触发微泡破裂转染的FKBP12.6基因可在心肌细胞高效表达,细胞生长良好。高表达FKBP12.6的心肌细胞中,总的钙离子浓度增加。结论:超声微泡介导FKBP12.6基因转染心肌细胞,可以明显增加心肌细胞中的Ca2 浓度,心肌细胞的收缩能力增强。     

ACE2-Ang(1-7)-Mas轴的生物学功能及其下游信号通路

ACE2-Ang(1-7)-Mas轴具有拮抗ACE-AngⅡ-AT1R经典轴的作用,参与体内众多的生理反应.在循环、泌尿和消化等系统及炎性反应中发挥着重要的保护作用.ACE2-Ang(1-7)-Mas轴功能的发挥与其信号通路密切相关.探讨此调节轴的生物学功能及其信号传导通路将为相关疾病的治疗提供新的思路.     

Effect of Betulonic Acid and Its Derivative [3-Oxo-20(29)-Lupene-28-Oyl]-3-Aminopropionic Acid on Liver Structure in Mice with RLS Lymphoma

Morphological study of the effects of semisynthetic derivatives of betulin (betulonic acid and [3-oxo-20(29)-lupene-28-oyl]-3-aminopropionic acid) on the liver of CBA/Lac mice with transplanted RLS lymphoma was studied in the control and after cytostatic polychemotherapy. The number of small focal necroses decreased, while the counts of hepatocytes in a state of slight hyaline droplet degeneration increased. Morphometry of the main elements of liver parenchyma showed that alanine amide derivative of betulonic acid decreases the severity of necrotic and degenerative changes in the liver parenchyma, induced by cytostatic polychemotherapy. Betulonic acid exhibited no appreciable hepatoprotective effect under these conditions. __________ Translated from Byulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 140, No. 9, pp. 348–351, September, 2005