脑室给予不同剂量的Capsazepine对LPS致大鼠发热过程的影响

目的在大鼠侧脑室内给予不同剂量的瞬时受体电位香草酸受体1(transient receptor potential vanilloid 1,TRPV1)拮抗剂Capsazepine,观察对LPS致热过程的影响。方法在侧脑室内预先给与3种剂量Capsazepine,随之腹腔给与LPS致热。在腹腔内埋植发射子遥测体温变化过程,同时应用荧光测定法检测下丘脑细胞内钙离子浓度的变化,Western blot方法检测TRPV1表达水平。结果随着Capsazepine剂量增大,LPS致大鼠发热水平升高,下丘脑细胞内钙离子浓度减少,TRPV1表达水平降低。结论 Capsazepine作用于大鼠下丘脑,可使LPS诱导的发热水平提高,此效应可能与TRPV1的作用有关。     

地佐环平对短暂电击所致创伤后应激障碍模型小鼠行为的影响

目的研究N-甲基-D-天冬氨酸(NMDA)受体非竞争性拮抗剂地佐环平(MK-801)对短暂电击所致创伤后应激障碍(PTSD)模型小鼠行为学的影响,探讨NMDA受体拮抗剂在PTSD治疗中应用的可能性。方法 1 ICR小鼠ip给予MK-801 0.0125,0.025,0.05和0.1 mg·kg-1,5和10 d后进行自发活动测试,观察小鼠自发活动。2连续2 d给予ICR小鼠不可逃避的足底电击(每天15次,0.8 m A,间隔10 s,持续10 s)制备PTSD模型。第1天电击后5 min,ip给予MK-801 0.0125,0.025和0.05 mg·kg-1或ig给予盐酸舍曲林15 mg·kg-1,每天1次,连续16 d。其中第3,8和15天测定僵住时间;第9天进行自发活动实验,观察水平跨越次数和垂直站立次数;第13天进行高架十字迷宫实验,观察进入开臂的次数和在开臂的滞留时间;第16天进行爬梯实验,观察爬梯数和站立次数。结果 1 MK-801 0.1 mg·kg-1连续给药5或10 d可显著抑制小鼠自发活动(P<0.01),其他各给药组对小鼠自发活动无显著影响。2与正常对照组比较,电击组、MK-801各剂量组及舍曲林给药组小鼠的水平跨越格数和垂直站立次数均无显著性差异;与正常对照组小鼠相比,电击组小鼠的僵住时间显著增加(P<0.01);小鼠进入开臂的次数和在开臂滞留时间的百分比显著减少(P<0.01);爬梯实验中小鼠的站立次数显著增加(P<0.01),提示模型建立成功。电击后小鼠的PTSD样行为学变化可以被伴随给予舍曲林15 mg·kg-1逆转,但MK-801无法逆转电击后小鼠的PTSD样行为。结论在不影响小鼠自发活动的剂量范围内,MK-801在小鼠短暂电击模型上无抗PTSD的行为学效应,单纯拮抗NMDA受体未必能发挥抗PTSD作用,这可能与PTSD发病机制的复杂性和NMDA受体结构与功能的复杂性有关。     

苦味受体抑制剂研究进展

自然界存在多种苦味物质,绝大多数苦味物质通过激活苦味受体而产生苦味.苦味受体抑制剂通过与苦味物质竞争结合位点而起到抑制苦味的作用,在食品和药物掩味方面发挥着重要作用.本文综述了苦味受体抑制剂的生理活性及其在药物中的应用,并根据其化学结构进行了分类.     

Surface‐expressed insulin receptors as well as IGF‐I receptors both contribute to the mitogenic effects of human insulin and its analogues

There is a medical need for new insulin analogues. Yet, molecular alterations to the insulin molecule can theoretically result in analogues with carcinogenic effects. Preclinical carcinogenicity risk assessment for insulin analogues rests to a large extent on mitogenicity assays in cell lines. We therefore optimized mitogenicity assay conditions for a panel of five cell lines. All cell lines expressed insulin receptors (IR), IGF‐I receptors (IGF‐IR) and hybrid receptors, and in all cell lines, insulin as well as the comparator compounds X10 and IGF‐I caused phosphorylation of the IR as well as IGF‐IR. Insulin exhibited mitogenicity EC₅₀ values in the single‐digit nanomolar to picomolar range. We observed correlations across cell types between (i) mitogenic potency of insulin and IGF‐IR/IR ratio, (ii) Akt phosphorylation and mitogenic potency and (iii) Akt phosphorylation and IR phosphorylation. Using siRNA‐mediated knockdown of IR and IGF‐IR, we observed that in HCT 116 cells the IR appeared dominant in driving the mitogenic response to insulin, whereas in MCF7 cells the IGF‐IR appeared dominant in driving the mitogenic response to insulin. Together, our results show that the IR as well as IGF‐IR may contribute to the mitogenic potency of insulin. While insulin was a more potent mitogen than IGF‐I in cells expressing more IR than IGF‐IR, the hyper‐mitogenic insulin analogue X10 was a more potent mitogen than insulin across all cell types, supporting that the hyper‐mitogenic effect of X10 involves the IR as well as the IGF‐IR. These results are relevant for preclinical safety assessment of developmental insulin analogues. Copyright © 2014 John Wiley & Sons, Ltd.     

Bisphenol A promotes X‐linked inhibitor of apoptosis protein‐dependent angiogenesis via G protein‐coupled estrogen receptor pathway

Bisphenol A (BPA), one of the high‐volume chemicals worldwide, has a core structure resembling that of natural estradiol. Recent evidence has demonstrated that exposure to BPA has a relationship with the risk of cancer. The objective of our study is to investigate the mechanisms underlying the pro‐angiogenic effects of BPA. We demonstrated that BPA markedly induces endothelial cell proliferation, migration and tube formation by activating endothelial nitric oxide synthase. BPA‐induced nitric oxide generation appeared to be associated with the X‐linked inhibitor of apoptosis protein (XIAP), which competes with endothelial nitric oxide synthase for caveolin‐1. BPA was shown to exert its pro‐angiogenic effect by upregulating XIAP expression via G protein‐coupled estrogen receptor (ER) activation but not via ERα or ERβ. Our data suggest that 100 nM BPA promote angiogenesis in a G protein‐coupled ER‐dependent genomic pathway, and provide a novel insight into the potential role of XIAP in mediating the pro‐angiogenic effects of BPA in endothelial cells. Copyright © 2015 John Wiley & Sons, Ltd.     

Mechanism of adrenocortical toxicity induced by quinocetone and its bidesoxy-quinocetone metabolite in porcine adrenocortical cells in vitro

Quinocetone (QCT) is a new feeding antibacterial agent in the QdNOs family. The mechanism of its adrenal toxicity is far from clear. This study was conducted to estimate the adrenal cell damage induced by QCT and its bidesoxy-quinocetone (B-QCT) metabolite and to further investigate their mechanisms. Following doses of QCT increasing from 5 to 50 μM, cell apoptosis and necrosis, mitochondrial dysfunction and redox imbalance were observed in porcine adrenocortical cells. The mRNA levels of the six components of intermediary enzymes and the adrenal renin-angiotensin-aldosterone system (RAAS) displayed a dysregulation induced by QCT, indicating that QCT might influence aldosterone secretion not only through the upstream of the production but also through the downstream of the adrenal RAAS pathway. In contrast, B-QCT had few toxic effects on the cell apoptosis, mitochondrial dysfunction and redox imbalance. Moreover, LCMS-IT-TOF analysis showed that no desoxy metabolites of QCT were found in either cell lysate or supernatant samples. In conclusion, we reported on the cytotoxicity in porcine adrenocortical cells exposed to QCT via oxidative stress, which raised awareness that its toxic effects resulted from N→O groups, and its toxic mechanism might involve the interference of the steroid hormone biosynthesis pathway.     

CGRP receptor antagonists and antibodies against CGRP and its receptor in migraine treatment

Recently developed calcitonin gene-related peptide (CGRP) receptor antagonistic molecules have shown promising results in clinical trials for acute treatment of migraine attacks. Drugs from the gepant class of CGRP receptor antagonists are effective and do not cause vasoconstriction, one of the major limitations in the use of triptans. However their use had to be discontinued because of risk of liver toxicity after continuous exposure. As an alternative approach to block CGRP transmission, fully humanized monoclonal antibodies towards CGRP and the CGRP receptor have been developed for treatment of chronic migraine (attacks >15 days/month). Initial results from phase I and II clinical trials have revealed promising results with minimal side effects and significant relief from chronic migraine as compared with placebo.The effectiveness of these various molecules raises the question of where is the target site(s) for antimigraine action. The gepants are small molecules that can partially pass the blood–brain barrier (BBB) and therefore, might have effects in the CNS. However, antibodies are large molecules and have limited possibility to pass the BBB, thus effectively excluding them from having a major site of action within the CNS. It is suggested that the antimigraine site should reside in areas not limited by the BBB such as intra- and extracranial vessels, dural mast cells and the trigeminal system. In order to clarify this topic and surrounding questions, it is important to understand the localization of CGRP and the CGRP receptor components in these possible sites of migraine-related regions and their relation to the BBB.     

67‐kDa Laminin receptor contributes to hypoxia‐induced migration and invasion of trophoblast‐like cells by mediating matrix metalloproteinase‐9

Insufficient trophoblast invasion often occurs in patients experiencing preeclampsia. The 67‐kDa laminin receptor (LR1) is a multifunctional protein that binds to laminin and interacts with the extracellular matrix. We recently demonstrated that LR1 is implicated in trophoblast migration and invasion. However, whether LR1 is involved in hypoxia‐mediated trophoblastic invasion remains unclear and requires further investigation. This study demonstrates that two trophoblast‐like cell lines (JEG3 and BeWo cells) cultured at 3% oxygen exerted enhanced migratory and invasive capabilities as compared with their counterparts exposed to 20% oxygen. LR1 expression was increased in hypoxic JEG3 cells but decreased after transfection with hypoxia‐inducible factor 1 alpha (HIF‐1α) specific siRNA. Moreover, shRNA targeting LR1 mRNA significantly inhibited hypoxia‐induced increase in matrix metalloproteinase (MMP)‐9 activity in JEG3 cells. Forced overexpression of LR1 augmented JEG3 cell migration and invasion, and enhanced MMP‐9 expression and activity. Additionally, the blockade of the MMP‐9 effect with its neutralizing antibody reduced LR1 elevation‐promoted trophoblastic invasion. In summary, this study demonstrates that LR1 contributes to hypoxia‐induced migration and invasion of trophoblast cells at least partly by mediating MMP‐9 in vitro.     

Canavanine increases glucose uptake in C2C12 cells through the activation of imidazoline I‐2B receptors

Canavanine is a guanidinium derivative that contains the basic structure of the ligand(s) of imidazoline receptor (I‐R). Canavanine has been reported to activate the imidazoline I‐3 receptor (I‐3R) both in vivo and in vitro. Additionally, the activation of the imidazoline I‐2B receptor (I‐2BR) by guanidinium derivatives may increase glucose uptake. Therefore, the effect of canavanine on the I‐2BR was investigated in the present study. Glucose uptake into cultured C₂C₁₂ cells was determined using the radio‐ligated tracer 2‐[¹⁴C]‐deoxy‐glucose. The changes in 5′ AMP‐activated protein kinase (AMPK) expression were also identified using Western blotting analysis. The canavanine‐induced glucose uptake was inhibited in a dose‐dependent manner by BU224 (0.01–1 μmol/L), which is a specific I‐2BR antagonist, in the C₂C₁₂ cells. Additionally, the canavanine‐stimulated AMPK phosphorylation and glucose transporter (GLUT4) expression were also sensitive to BU224 inhibition in the C₂C₁₂ cells. Moreover, both canavanine‐stimulated glucose uptake and AMPK phosphorylation were attenuated by high concentrations of amiloride (1–2 μmol/L), which is another established I‐2BR inhibitor, in a dose‐dependent manner in C₂C₁₂ cells. Additionally, compound C abolished the canavanine‐induced glucose uptake and AMPK phosphorylation at a concentration (0.1 μmol/L) sufficient to inhibit AMPK. In conclusion, these data demonstrated that canavanine has an ability to activate I‐2BR through the AMPK pathway to increase glucose uptake, which indicates I‐2BR as a new target for diabetic therapy.