糖链加工酶抑制剂地衣霉素和Swainsonine对运铁蛋白受体再循环的影响

作者应用两种糖链加工酶抑制剂,即抑制糖蛋白N-糖链合成的地衣霉素及使糖蛋白N-糖链变为高甘露糖型和杂合型的Swainsonine,来研究SMMC-7721人肝癌细胞表面运铁蛋白受体(TR)N-糖链结构的改变对受体再循环过程的影响。实验结果表明,1μg/ml的地衣霉素处理细胞后,细胞生长、蛋白质合成、DNA合成及细胞膜糖蛋白N-糖链的合成均受到不同程度的抑制,但对TR的再循环并无明显影响。2μg/ml的Swainsonine处理细胞后,对细胞生长、蛋白质合成、DNA合成均无明显影响,它使细胞膜糖蛋白N-糖链中甘露糖的掺入增加,并可加快TR再循环过程。     

Clathrin assembly protein AP-2 is detected in both neurons and glia, and its reduction is prominent in layer II of frontal cortex in Alzheimer's disease

There are several adaptor proteins associated with clathrin coated vesicles. Among them are AP180 and AP-2. We and others have previously described synaptic localization of AP180. AP180 immunoreactivity is altered in both the superior frontal gyrus and hippocampus in Alzheimer’s disease (AD). We here investigate the location and alteration of another adaptor protein, AP-2. In contrast to AP180, we have found that AP-2 is expressed by both neurons and glia. Furthermore, the only noticeable change of AP-2 in AD is a loss of its immunoreactivity in layer II of the superior frontal gyrus.     

Targeting cell surface trafficking of pain-facilitating receptors to treat chronic pain conditions

Introduction: Chronic pain conditions are serious clinical concerns. Its genesis is closely associated with sensitization of nociceptive primary sensory neurons (nociceptors) and dorsal horn neurons by various pain mediators produced during inflammation and tissue injury. Growing evidence showed that increasing cell surface trafficking of pain-facilitating receptors is an important mechanism underlying the peripheral and central sensitization.

Areas covered: We summarized the progress of this area over the past decade by showing that inflammation, tissue damage or pain mediators facilitate cell surface trafficking of pain-facilitating receptors such as transient receptor potential vanilloid-1, transient receptor potential ankyrin-1, voltage-gated sodium channel 1.8, P2X3 and EP4 in primary sensory neurons, GluR1 and GluR2 of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, NR1 and NR2 of N-methyl-d-aspartate receptors and acid-sensing ion channels 1 in dorsal horn neurons and P2X4 in spinal microglia. The anti-allodynic effects of gabapentin was mediated by blocking surface trafficking of α2δ1 and α2δ2 subunits of voltage-gated calcium channels in primary sensory and dorsal horn neurons.

Expert opinion: Pain mediators stimulate forward surface trafficking of their own and/or other pain-facilitating receptors to amplify pain intensity and duration. Enhancing surface abundance of pain-facilitating receptors in nociceptors and dorsal horn neurons is an important mechanism underpinning chronic pain states. Targeting specific trafficking events of pain-facilitating receptors may open a novel therapeutic avenue to more efficiently treat chronic pain conditions.     

妇科再造胶囊与克罗米芬联合治疗无排卵性不孕的临床研究

目的：探讨妇科再造胶囊与克罗米芬联合治疗无排卵性不孕的临床效果。方法收集2012年4月～2013年10月来我院治疗的无排卵性不孕患者58例,将其随机分为两组,观察组29例,采用克罗米芬联合妇科再造胶囊治疗;对照组29例,采用克罗米芬治疗。两组都以3个月经周期作为1个治疗疗程。结果卵巢优势卵泡直径比较中,观察组C12、C14均大于对照组,差异有统计学意义（P＜0.05）。观察组的排卵率为84.8%,妊娠率为51.7%,对照组为75.0%,31.0%,观察组排卵率及妊娠率均高于对照组,差异有统计学意义（P＜0.05）。观察组子宫内膜厚度≥8～13mm的患者比例高于对照组,差异有统计学意义（P＜0.05）。结论无排卵性不孕症患者采用妇科再造胶囊与克罗米芬联合治疗效果与单独应用克罗米芬治疗效果显著,可以在临床推广应用。     

Highly specific detection of muscarinic M3 receptor,G protein interaction and intracellular trafficking in human detrusor using Proximity Ligation Assay (PLA)

Purpose

Muscarinic acetylcholine receptors (mAChRs) regulate a number of important physiological functions. Alteration of mAChR expression or function has been associated in the etiology of several pathologies including functional bladder disorders (e.g bladder pain syndrome/interstitial cystitis – BPS/IC). In a previous study we found specific mAChR expression patterns associated with BPS/IC, while correlation between protein and gene expression was lacking. Posttranslational regulatory mechanisms, e.g. altered intracellular receptor trafficking, could explain those differences. In addition, alternative G protein (GP) coupling could add to the pathophysiology via modulation of muscarinic signaling. In our proof-of-principle study, we addressed these questions in situ. We established PLA in combination with confocal laserscanning microscopy (CLSM) and 3D object reconstruction for highly specific detection and analysis of muscarinic 3 receptors (M3), G protein (GP) coupling and intracellular trafficking in human detrusor samples.

Compartmentalization of signaling by vesicular trafficking: a shared building design for the immune synapse and the primary cilium

Accumulating evidence underscores the immune synapse (IS) of naive T cells as a site of intense vesicular trafficking. At variance with helper and cytolytic effectors, which use the IS as a secretory platform to deliver cytokines and/or lytic granules to their cellular targets, this process is exploited by naive T cells as a means to regulate the assembly and maintenance of the IS, on which productive signaling and cell activation crucially depend. We have recently identified a role of the intraflagellar transport (IFT) system, which is responsible for the assembly of the primary cilium, in the non-ciliated T-cell, where it controls IS assembly by promoting polarized T-cell receptor recycling. This unexpected finding not only provides new insight into the mechanisms of IS assembly but also strongly supports the notion that the IS and the primary cilium, which are both characterized by a specialized membrane domain highly enriched in receptors and signaling mediators, share architectural similarities and are homologous structures. Here, we review our current understanding of vesicular trafficking in the regulation of the assembly and maintenance of the naive T-cell IS and the primary cilium, with a focus on the IFT system.     

Methadone Recycling Sustains Drug Reservoir in Tissue

We hypothesize that there is a tissue store of methadone content in humans that is not directly accessible, but is quantifiable. Further, we hypothesize the mechanism by which methadone content is sustained in tissue stores involves methadone uptake, storage, and release from tissue depots in the body (recycling). Accordingly, we hypothesize that such tissue stores, in part, determine plasma methadone levels. We studied a random sample of six opioid-naïve healthy subjects. We performed a clinical trial simulation in silico using pharmacokinetic modeling. We found a large tissue store of methadone content whose size was much larger than methadone's size in plasma in response to a single oral dose of methadone 10 mg. The tissue store measured 13–17 mg. This finding could only be explained by the contemporaneous storage of methadone in tissue with dose recycling. We found that methadone recycles 2–5 times through an inaccessible extravascular compartment (IAC), from an accessible plasma-containing compartment (AC), before exiting irreversibly. We estimate the rate of accumulation (or storage) of methadone in tissue was 0.029–7.29 mg/h. We predict 39 ± 13% to 83 ± 6% of methadone's tissue stores “spillover” into the circulation. Our results indicate that there exists a large quantifiable tissue store of methadone in humans. Our results support the notion that methadone in humans undergoes tissue uptake, storage, release into the circulation, reuptake from the circulation, and re-release into the circulation, and that spillover of methadone from tissue stores, in part, maintain plasma methadone levels in humans.     

Synaptic Vesicle Recycling: Genetic and Cell Biological Studies

Abstract

We review mainly the work from our research group here. Our focus has been on the use of genetic methods to delineate the mechanisms of synaptic vesicle recycling and cellular trafficking. Acute temperature-sensitive paralytic mutants have been of particular value in this approach. We have primarily used screens for suppressor and enhancer mutations to identify genetic loci coding for proteins that interact with Dynamin in Drosophila. In addition, we have used reverse genetic approaches to investigate few other candidate molecules that may play a role in synaptic vesicle endocytosis. We have in particular discussed at some length the role of endocytic accessory proteins Stoned and Eps15 in vesicle recycling.     

Clathrin-mediated endocytosis is inhibited during mitosis

A long-standing paradigm in cell biology is the shutdown of endocytosis during mitosis. There is consensus that transferrin uptake is inhibited after entry into prophase and that it resumes in telophase. A recent study proposed that endocytosis is continuous throughout the cell cycle and that the observed inhibition of transferrin uptake is due to a decrease in available transferrin receptor at the cell surface, and not to a shutdown of endocytosis. This challenge to the established view is gradually becoming accepted. Because of this controversy, we revisited the question of endocytic activity during mitosis. Using an antibody uptake assay and controlling for potential changes in surface receptor density, we demonstrate the strong inhibition of endocytosis in mitosis of CD8 chimeras containing any of the three major internalization motifs for clathrin-mediated endocytosis (YXXΦ, [DE]XXXL[LI], or FXNPXY) or a CD8 protein with the cytoplasmic tail of the cation-independent mannose 6-phosphate receptor. The shutdown is not gradual: We describe a binary switch from endocytosis being "on" in interphase to "off" in mitosis as cells traverse the G(2)/M checkpoint. In addition, we show that the inhibition of transferrin uptake in mitosis occurs despite abundant transferrin receptor at the surface of HeLa cells. Our study finds no support for the recent idea that endocytosis continues during mitosis, and we conclude that endocytosis is temporarily shutdown during early mitosis.     

The Impact of Ground Tire Rubber Oxidation with H2O2 and KMnO4 on the Structure and Performance of Flexible Polyurethane/Ground Tire Rubber Composite Foams

The use of waste tires is a very critical issue, considering their environmental and economic implications. One of the simplest and the least harmful methods is conversion of tires into ground tire rubber (GTR), which can be introduced into different polymer matrices as a filler. However, these applications often require proper modifications to provide compatibility with the polymer matrix. In this study, we examined the impact of GTR oxidation with hydrogen peroxide and potassium permanganate on the processing and properties of flexible polyurethane/GTR composite foams. Applied treatments caused oxidation and introduction of hydroxyl groups onto the surface of rubber particles, expressed by the broad range of their hydroxyl numbers. It resulted in noticeable differences in the processing of the polyurethane system and affected the structure of flexible composite foams. Treatment with H₂O₂ resulted in a 31% rise of apparent density, while the catalytic activity of potassium ions enhanced foaming of system decreased density by 25% and increased the open cell content. Better mechanical performance was noted for H₂O₂ modifications (even by 100% higher normalized compressive strength), because of the voids in cell walls and incompletely developed structure during polymerization, accelerated by KMnO₄ treatment. This paper shows that modification of ground tire rubber is a very promising approach, and when properly performed may be applied to engineer the structure and performance of polyurethane composite foams.