蓖麻毒素对原代培养小鼠肺泡上皮细胞的影响

目的探讨蓖麻毒素气源性中毒中的肺损伤机制。方法取BALB/c雌性小鼠,采用心脏灌注排血、胶原酶和胰蛋白酶原位消化法及组织块细胞迁出培养法,分离培养小鼠肺泡上皮细胞,经刮除法及差相消化贴壁法纯化小鼠肺泡上皮细胞,通过兔抗鼠角蛋白单克隆抗体细胞爬片组化染色法对细胞进行纯度鉴定;经不同终浓度的蓖麻毒素(0、0.000 1、0.001、0.01、0.1、1、10、100μg/ml)处理细胞进行毒性分析;倒置显微镜下观察0.05μg/ml蓖麻毒素对细胞的毒性作用。结果获得的小鼠肺泡上皮细胞纯度达90%以上;随着蓖麻毒素浓度的增加,细胞活力降低,蓖麻毒素浓度与肺泡上皮细胞增殖抑制呈正相关;0.05μg/ml的蓖麻毒素即可引起细胞皱缩、空泡化变性。结论蓖麻毒素可引起原代培养肺泡上皮细胞变性损伤,损伤程度随蓖麻毒素浓度增高而加重,为蓖麻毒素气源性中毒中的肺损伤机制提供了理论依据,同时为进一步探讨蓖麻毒素的毒理作用提供了新的思路。     

The phospholipids of rabbit type II alveolar epithelial cells: Comparison with lung lavage,lung tissue,alveolar macrophages,and a human alveolar tumor cell line

The phospholipid composition of type II alveolar epithelial cells from the rabbit was compared with that of alveolar macrophages, lung lavage and lung tissue. In addition, the phospholipid composition of a human alveolar tumor cell line, which is morphologically similar to type II cells, was examined. Phosphatidylcholine accounted for 48% of the total phospholipid in the type II cells, 41% in the tumor cells, and 30% in the macrophages. Phosphatidylcholine was 51% disaturated in the type II cells, 54% in lung lavage, 39% in whole lung, 29% in lavaged lung and macrophages, and 16% in the tumor cells. Palmitic acid was the major fatty acid in phosphatidylcholine from all samples with the exception of the tumor cells in which almost half of the fatty acids were accounted for by oleic acid. The phospholipids of the type II cells were more similar to those of lung lavage, and thus surfactant, than to lung tissue and macrophages. This is consistent with their supposed role in surfactant production. The tumor cells, although morphologically similar to type II cells, were quite different with respect to phospholipid composition.     

Comparative toxicity of 24 manufactured nanoparticles in human alveolar epithelial and macrophage cell lines

Background  

A critical issue with nanomaterials is the clear understanding of their potential toxicity. We evaluated the toxic effect of 24 nanoparticles of similar equivalent spherical diameter and various elemental compositions on 2 human pulmonary cell lines: A549 and THP-1. A secondary aim was to elaborate a generic experimental set-up that would allow the rapid screening of cytotoxic effect of nanoparticles. We therefore compared 2 cytotoxicity assays (MTT and Neutral Red) and analyzed 2 time points (3 and 24 hours) for each cell type and nanoparticle. When possible, TC50 (Toxic Concentration 50 i.e. nanoparticle concentration inducing 50% cell mortality) was calculated.     

Antiproliferative and apoptotic effects of tocopherols and tocotrienols on normal mouse mammary epithelial cells

Studies were conducted to determine the comparative effects of tocopherols and tocotrienols on normal mammary epithelial cell growth and viability. Cells isolated from midpregnant BALB/c mice were grown within collagen gels and maintained on serum-free media. Treatment with 0–120 μM α-and γ-tocopherol had no effect, whereas 12.5–100 m μM tocotrienol-rich fraction of palm oil (TRF), 100–120 μM δ-tocopherol, 50–60 μM α-tocotrienol, and 8–14 μM γ- or δ-tocotrienol significantly inhibited cell growth in a dose-responsive manner. In acute studies, 24-h exposure to 0–250 μM α-, γ-, and δ-tocopherol had no effect, whereas similar treatment with 100–250 μM TRF, 140–250 μM α-, 25–100 μM γ- or δ-tocotrienol significantly reduced cell viability. Growth-inhibitory doses of TRF, δ-tocopherol, and a-, γ-, and δ-tocotrienol were shown to induce apoptosis in these cells, as indicated by DNA fragmentation. Results also showed that mammary epithelial cells more easily or preferentially took up tocotrienols as compared to tocopherols, suggesting that at least part of the reason tocotrienols display greater biopotency than tocopherols is because of greater cellular accumulation. In summary, these findings suggest that the highly biopotent γ- and δ-tocotrienol isoforms may play a physiological role in modulating normal mammary gland growth, function, and remodeling.     

Oxidative stress mediated cytotoxicity of biologically synthesized silver nanoparticles in human lung epithelial adenocarcinoma cell line

The goal of the present study was to investigate the toxicity of biologically prepared small size of silver nanoparticles in human lung epithelial adenocarcinoma cells A549. Herein, we describe a facile method for the synthesis of silver nanoparticles by treating the supernatant from a culture of Escherichia coli with silver nitrate. The formation of silver nanoparticles was characterized using various analytical techniques. The results from UV-visible (UV-vis) spectroscopy and X-ray diffraction analysis show a characteristic strong resonance centered at 420 nm and a single crystalline nature, respectively. Fourier transform infrared spectroscopy confirmed the possible bio-molecules responsible for the reduction of silver from silver nitrate into nanoparticles. The particle size analyzer and transmission electron microscopy results suggest that silver nanoparticles are spherical in shape with an average diameter of 15 nm. The results derived from in vitro studies showed a concentration-dependent decrease in cell viability when A549 cells were exposed to silver nanoparticles. This decrease in cell viability corresponded to increased leakage of lactate dehydrogenase (LDH), increased intracellular reactive oxygen species generation (ROS), and decreased mitochondrial transmembrane potential (MTP). Furthermore, uptake and intracellular localization of silver nanoparticles were observed and were accompanied by accumulation of autophagosomes and autolysosomes in A549 cells. The results indicate that silver nanoparticles play a significant role in apoptosis. Interestingly, biologically synthesized silver nanoparticles showed more potent cytotoxicity at the concentrations tested compared to that shown by chemically synthesized silver nanoparticles. Therefore, our results demonstrated that human lung epithelial A549 cells could provide a valuable model to assess the cytotoxicity of silver nanoparticles.     

Toxic effects of brake wear particles on epithelial lung cells in vitro

Background  

Fine particulate matter originating from traffic correlates with increased morbidity and mortality. An important source of traffic particles is brake wear of cars which contributes up to 20% of the total traffic emissions. The aim of this study was to evaluate potential toxicological effects of human epithelial lung cells exposed to freshly generated brake wear particles.     

Investigation on the role of the molecular weight of polyvinyl pyrrolidone in the shape control of high-yield silver nanospheres and nanowires

Serving as shape control agent, polyvinyl pyrrolidone (PVP) has been widely used in chemical synthesis of metal nanoparticles. However, the role of molecular weight (MW) of PVP has been rarely concerned. In this study, we show a facile method to control the shapes of silver nanocrystals using PVP with different MWs. PVP_MW=8,000, PVP_MW=29,000, PVP_MW=40,000, and PVP_MW=1,300,000 are compared in the present study. Surprisingly, high-yield silver rodlike nanostructures, nanospheres, and nanowires can be obtained under the same growth environment and reactant concentrations by simply changing the MW of PVP. The mechanism studies of the role of PVP with different MWs in the growth process were carried out systemically using the morphology and spectroscopic measurement, FT-IR spectrum analysis, and seed crystallization monitoring. The results indicate that the MW of PVP plays a determinant role in the morphology and optical property control of the silver nanocrystals. Meantime, the concentration of PVP was found to be an assistant factor to further improve the shape and the yield of the synthesized nanocrystals.     

Functional and ultrastructural effects of essential fatty acid deficiency in kidney epithelial cells

Madin-Darby canine kidney (MDCK) epithelial cells were grown in culture medium supplemented with 1% fetal bovine serum (FBS) to provide a cell culture model of essential fatty acid deficiency (EFAD). 5,8,11-Eicosatrienoic acid (20∶3n−9) accumulated in cellular phospholipids, and arachidonic acid (20∶4) decreased. A large increase in cellular cholesterol/phospholipid ratio was observed. Hemicyst formation was greatly reduced from normal levels in the EFAD-MDCK cells. Scanning and transmission electron microscopy revealed that EFAD-MDCK were much flatter than their normal counterparts. They had much less dense surface microvilli, mitochondria and other organelles were very sparse, except in the perinuclear area, and much of the peripheral cytoplasm was amorphous. The EFAD was rapidly reversed by the addition of as little as 10 μM linoleic or arachidonic acid to the medium. Cells supplemented with 10% FBS, the usual culture condition, displayed borderline EFAD, with intermediate levels of 20∶3n−9 and 20∶4 and hemicyst formation. These studies suggest that EFAD reduces water and electrolyte transport in renal tubular epithelium.     

Parameters influencing the effects of quartz on alveolar macrophages

Studies on the interactions of quartz particles with macrophages have contributed considerably to a better understanding of the mechanisms eventually leading to lung fibrosis. The aim of this study was to compare the effects of a number of test or standard quartz materials on alveolar macrophages from different species. Also the geometry of exposure was varied.     

Shape and size effects on the compressive strength of high-strength concrete

In this paper we investigate the influence of the shape and of the size of the specimens on the compressive strength of high-strength concrete. We use cylinders and cubes of different sizes for performing stable stress-strain tests. The tests were performed at a single axial strain rate, 10^{− 6} s^{− 1}. This value was kept constant throughout the experimental program. Our results show that the post-peak behavior of the cubes is milder than that of the cylinders, which results in a strong energy consumption after the peak. This is consistent with the observation of the crack pattern: The extent of cracking throughout the specimen is denser in the cubes than in the cylinders. Indeed, a main inclined fracture surface is nucleated in cylinders, whereas in cubes we find that lateral sides get spalled leading to the so-called hour-glass failure mode. The remaining cube core gets fragmented due to crushing, in some cases exhibiting a dense columnar cracking in the bulk of the specimen. Finally, we investigate the relationship between the compressive strength given by both types of specimen for several specimen sizes.     

人气管上皮细胞的原代分离及气液界面培养

目的分离人气管上皮细胞,并进行气液界面(air-liquid interface,ALI)培养,为呼吸道病毒研究提供良好的细胞模型。方法采用低温消化法分离人气管上皮细胞,用预包被胶原的0.4μm Transwell培养皿对传代后的气管上皮细胞进行ALI培养。利用倒置显微镜观察细胞生长状态,免疫细胞化学染色和免疫组化鉴定培养细胞的生长和分化情况,同时测定细胞分化过程中的跨上皮电阻(trans epithelial electrical resistance,TEER)。结果分离培养的气管上皮细胞光镜下细胞形态及免疫荧光角蛋白染色阳性,证实培养细胞为气管上皮细胞。ALI培养后的人气管上皮细胞的形态学、MUC5AC蛋白、Ⅳ型β-微管蛋白(β-tubulinⅣ)的表达及紧密连接和假复层的形成情况均与人气管组织结构类似。结论低温消化法可成功分离到活性高的上皮细胞。ALI培养的上皮细胞形态和功能与体内接近,且能较长时间维持其正常形态及功能,为呼吸道相关疾病的研究提供了理想的平台。     

Regulation of cholesterol synthesis in isolated epithelial cells of human small intestine

We have investigated the regulation of cholesterol synthesis in isolated human small intestine epithelial cells (enterocytes). It was established that the amount of cholesterol synthesized increased linearly with the incubation time and the number of cells in the incubation mixture; the synthesis was suppressed by 7-ketocholesterol. Cholic, dehydrocholic, chenodeoxycholic, glycocholic, taurocholic, taurochenodeoxycholic and taurodeoxycholic acids inhibited cholesterol synthesis in enterocytes to different degrees in a dose-dependent manner. Lithocholic acid enhanced the rate of cholesterol synthesis. Deoxycholic acid, methyl ester of cholic acid and cholesterol did not affect the process. No bile acids tested, with the exception of taurodeoxycholic acid, affected fatty acid synthesis in enterocytes. Most bile acids also decreased cholesterol synthesis in cultured human skin fibroblasts. The results obtained make it possible to postulate that cholesterol synthesis in human enterocytes may be subject to a complex regulation by bile acids.     

The effects of quercetin on SW480 human colon carcinoma cells: a proteomic study

Background  

High fruit and vegetable intake is known to reduce the risk of colon cancer. To improve understanding of this phenomenon the action of different phytochemicals on colon cells has been examined. One such compound is quercetin that belongs to the group known as flavonoids. The purpose of this study was to determine the influence of quercetin on the proteome of the SW480 human colon adenocarcinoma cell line, specifically to identify proteins that could be the molecular targets of quercetin in its amelioration of the progression of colon cancer. To this end, two-dimensional gel electrophoresis and mass spectrometry were used to identify proteins that underwent a change in expression following treatment of the cells with 20 μM quercetin. This could elucidate how quercetin may reduce the progression of colon cancer.     

Novel silicon-containing analogues of the retinoid agonist bexarotene: syntheses and biological effects on human pluripotent stem cells

Twofold sila‐substitution (C/Si exchange) of the clinically used RXR‐selective retinoid agonist bexarotene leads to disila‐bexarotene, which displays pharmacological potency similar to that of the parent carbon compound, as shown in a HeLa‐cell‐based RXR assay. Formal exchange of the SiCH₂CH₂Si group in disila‐bexarotene with a SiCH₂Si or SiOSi moiety leads to the disila‐bexarotene analogues 8 and 9 . The silicon compounds 8 and 9 were synthesized in multistep syntheses, starting from HC?C(CH₃)₂SiCH₂Si(CH₃)₂C?CH and HC?C(CH₃)₂SiOSi(CH₃)₂C?CH, respectively. The key step in the syntheses of 8 and 9 is a cobalt‐catalyzed [2+2+2] cycloaddition reaction that affords the 1,3‐disilaindane and 2‐oxa‐1,3‐disilaindane skeletons. Disila‐bexarotene and its analogues 8 and 9 were studied for their biological effects relative to all‐trans retinoic acid in cultured human pluripotent stem cells. The parent carbon compound bexarotene was included in some of these biological studies. Although the silicon‐containing bexarotene analogues disila‐bexarotene, 8 , and 9 appear not to regulate the differentiation of TERA2.cl.SP12 stem cells, preliminary evidence indicates that these compounds may possess enhanced functions over the parent compound bexarotene, such as induction and regulation of cell death and cell numbers. The biological data obtained indicate that bexarotene, contrary to the silicon‐containing analogues disila‐bexarotene, 8 , and 9 , may partially act to induce cell differentiation.     

Multi-walled carbon nanotube length as a critical determinant of bioreactivity with primary human pulmonary alveolar cells

Multiwalled carbon nanotube (MWCNT) length is suggested to critically determine their pulmonary toxicity. This stems from in vitro and in vivo rodent studies and in vitro human studies using cell lines (typically cancerous). There is little data using primary human lung cells. We addressed this knowledge gap, using highly relevant, primary human alveolar cell models exposed to precisely synthesised and thoroughly characterised MWCNTs. In this work, transformed human alveolar type-I-like epithelial cells (TT1), primary human alveolar type-II epithelial cells (ATII) and alveolar macrophages (AM) were treated with increasing concentrations of MWCNTs before measuring cytotoxicity, inflammatory mediator release and MAP kinase signalling. Strikingly, we observed that short MWCNTs (∼0.6 μm in length) induced significantly greater responses from the epithelial cells, whilst AM were particularly susceptible to long MWCNTs (∼20 μm). These differences in the pattern of mediator release were associated with alternative profiles of JNK, p38 and ERK1/2 MAP kinase signal transduction within each cell type. This study, using highly relevant target human alveolar cells and well defined and characterised MWCNTs, shows marked cellular responses to the MWCNTs that vary according to the target cell type, as well as the aspect ratio of the MWCNT.     

乳酸杆菌对子宫内膜上皮细胞增殖的影响

目的研究乳酸杆菌对子宫内膜上皮细胞增殖的影响。方法以内蒙古地区68名健康生育期女性人群作为研究对象,取阴道分泌物,经条件培养基分离纯化乳酸杆菌,通过染色、酶触反应、产H2O2等试验确定其生物学性质,通过16sRNA基因扩增及测序对乳酸杆菌进行分类。将不同浓度(10、102、103和104个/mL)优势乳酸杆菌与子宫内膜上皮细胞共培养12、24、36和48 h,采用CCK-8法检测乳酸杆菌对子宫内膜上皮细胞增殖作用的影响,以确定乳酸杆菌对子宫内膜上皮细胞作用的最适浓度和时间。结果经过分离纯化得到141株乳酸杆菌,草酸铵结晶紫染色呈紫色,为革兰阳性菌,所有菌落均能产生H2O2,但产H2O2能力有差别。经16sRNA基因扩增及测序共分为6种乳酸杆菌,其中卷曲乳杆菌(L. crispatus)、格氏乳杆菌(L. asseri)和阴道乳杆菌(L. vaginalis)为内蒙古女性人群阴道中优势乳酸杆菌菌属,分别占54. 61%、24. 82%和12. 06%;不同浓度L. crispatus均能促进子宫内膜上皮细胞增殖,且乳酸杆菌浓度103个/mL、作用36 h时,子宫内膜上皮细胞增殖最显著,增殖率可达93. 10%。结论乳酸杆菌可促进子宫内膜上皮细胞的增殖,为乳酸杆菌促进子宫内膜局部的损伤修复提供了理论依据。     

Shape and “gap” effects on the behavior of variably confined concrete

Factors affecting the behavior of variably confined concrete are presented. The effect of debonding the fiber-reinforced polymer (FRP) jacket to the concrete substrate and providing a gap between the concrete and confining jacket is investigated. A second parameter—the shape of the cross section—is also investigated. An experimental program involving the compression testing of standard cylinders and similarly sized square specimens having external FRP jackets providing passive confinement is presented. Factors affecting jacket efficiency and the appropriateness of factors accounting for specimen shape are determined experimentally and discussed.The provision of a gap affected the axial stress at which the confining jacket was engaged, resulting in a reduced maximum attainable concrete strength. The jacket efficiency was not affected by the provision of the gap. The shape of the specimens was observed to affect the level of confinement generated. Square specimens exhibit lower confinement levels than circular specimens having the same jacket.     

All that is silver is not toxic: silver ion and particle kinetics reveals the role of silver ion aging and dosimetry on the toxicity of silver nanoparticles

Background

When suspended in cell culture medium, nano-objects composed of soluble metals such as silver can dissolve resulting in ion formation, altered particle properties (e.g. mass, morphology, etc.), and modulated cellular dose. Cultured cells are exposed not just to nanoparticles but to a complex, dynamic mixture of altered nanoparticles, unbound ions, and ion-ligand complexes. Here, three different cell types (RAW 264.7 macrophages and bone marrow derived macrophages from wild-type C57BL/6?J mice and Scavenger Receptor A deficient (SR-A^(?/?)) mice) were exposed to 20 and 110?nm silver nanoparticles, and RAW 264.7 cells were exposed to freshly mixed silver ions, aged silver ions (ions incubated in cell culture medium), and ions formed from nanoparticle dissolution. The In Vitro Sedimentation, Diffusion, Dissolution, and Dosimetry Model (ISD3) was used to predict dose metrics for each exposure scenario.

Results

Silver nanoparticles, freshly mixed ions, and ions from nanoparticle dissolution were toxic, while aged ions were not toxic. Macrophages from SR-A^(?/?) mice did not take up 20?nm silver nanoparticles as well as wild-types but demonstrated no differences in silver levels after exposure to 110?nm nanoparticles. Dose response modeling with ISD3 predicted dose metrics suggest that amount of ions in cells and area under the curve (AUC) of ion amount in cells are the most predictive of cell viability after nanoparticle and combined nanoparticle/dissolution-formed-ions exposures, respectively.

Conclusions

Results of this study suggest that the unbound silver cation is the ultimate toxicant, and ions formed extracellularly drive toxicity after exposure to nanoparticles. Applying computational modeling (ISD3) to better understand dose metrics for soluble nanoparticles allows for better interpretation of in vitro hazard assessments.

     

Processing and shape effects on silver paste electrically conductive adhesives (ECAs)

Various amounts of silver flakes and dendrites were used as conductive fillers in an electrically conductive adhesive (ECA) resin with DPM, BCA and xylene as diluent to help uniform distribution of filler particles in the matrix. Due to the fact that the higher the temperature, the higher the shrinkage rate of the polymer resin and, consequently, the larger the connecting area in-between fillers, a better curing condition for processing silver filled ECA was found to be a relatively higher curing temperature. The mechanism of conductivity achievement in conductive adhesives was analyzed by comparing processing conditions, resistivity and microstructures. In addition, the influence of adding nano-sized silver particles on the resistivity of the conductive adhesives was also investigated and the addition of nano-sized silver particles resulted in a lower percolation threshold for ECAs.