组织工程心脏瓣膜体外构建的初步研究

目的:初步体外构建组织工程人工心脏瓣膜。方法:种子细胞选用胸骨骨髓,抽取后分离出人骨髓间充质干细胞,体外培养,扩增待用。脱细胞瓣膜支架的构建应用破膜剂曲拉通-脱氧胆酸钠对新鲜猪主动脉瓣膜36片(实验组)进行处理48h后,与对照组未经处理的猪主动脉瓣膜(36片)进行对比,在光镜和电镜下观察脱细胞效果。将人骨髓间充质干细胞接种于脱细胞瓣膜支架上静态培养7d,观察细胞-支架复合体生长状况。结果:曲拉通-脱氧胆酸钠法脱细胞彻底,保持原有纤维结构。种植的人骨髓间充质干细胞7d后可生长于瓣膜支架的表面。结论:曲拉通-脱氧胆酸钠法是一种较为理想的猪瓣膜脱细胞方法;人骨髓间充质干细胞能在脱细胞猪瓣膜支架上较好地黏附和生长,从而初步构建组织工程人工心脏瓣膜。     

人脐静脉血管内皮细胞构建组织工程心脏瓣膜及其生理功能

目的 探讨人脐静脉血管内皮细胞(HUVECs)构建组织工程心脏瓣膜(TEHV)及其生理功能.方法 以脱细胞猪主动脉瓣作支架;将扩增的HUVECs种植在瓣膜上,体外静态构建TEHV,观察内皮细胞的形态和生长状况.收集瓣膜培养液,检测瓣膜内皮细胞分泌组织型纤溶酶原激活物(t-PA)、组织型纤溶酶原激活物抑制物(PAI-1)、前列环素(PGI2)、一氧化氮(NO)、内皮素(ET-1)的功能.结果 猪主动脉瓣膜中的细胞成分能完全去除,脱细胞瓣叶的生物力学特性同新鲜瓣叶相比无明显变化以HUVECs做种子细胞,成功构建TEHV;瓣膜表面的内皮细胞生长状态良好,长成连续的细胞层.细胞能够分泌t-PA、PAI-1、PGI2、NO、ET-1等.结论 以脱细胞猪主动脉瓣膜为支架,种植HUVECs成功构建TEHV,其表面HUVECs具有正常内皮细胞的生理功能.     

猪主动脉瓣去细胞的初步研究

目的　对猪主动脉瓣进行了酶水化法去细胞的研究,探讨应用猪去细胞瓣膜支架体外构建组织工程心脏瓣膜的可行性。方法　应用胰蛋白酶/乙二胺四乙酸去除猪主动脉瓣组织中的细胞,对处理后的组织进行组织学与免疫组化,扫描电子显微镜观察,并进行理化性能测定。结果　酶水化法去细胞效果良好,且能较好地保持胶原纤维和弹性纤维原有的排列和分布。与新鲜猪主动脉瓣比较,热皱缩温度、抗张强度差异无统计学意义(P>0.05)。结论　酶水化法去细胞效果良好,初步制造了猪主动脉瓣脱细胞基质材料,为进一步制作组织工程瓣膜提供了较合适的支架材料。     

人主动脉壁脱细胞血管基质的制备

目的 探讨应用同种去细胞主动脉血管壁支架体外构建组织工程管道的可行性.方法 取液氮保存的同种主动脉带瓣管道.首先置于Tris低渗缓冲液室温下溶解血管壁细胞,血管壁用D-Hanks液充分清洗后,再用含十二烷基硫酸钠(SDS)的Tris低渗缓冲液室温下再孵育24h,洗涤脱去细胞;最后用Tris等渗缓冲液充分冲洗残留细胞,完成脱细胞支架制备.并对脱细胞后的管壁进行光镜及电镜形态学观察以及进行理化性能测定.结果 人主动脉血管壁经SDS法去细胞后,经光镜和电镜观察,脱细胞后血管壁无细胞残留,胶原纤维和弹性纤维保留完整,形态学无明显变化;生物学性能也没有明显改变.结论 SDS法去细胞效果良好,初步制造了同种主动脉血管壁脱细胞基质材料,为构建同种带瓣管道提供了较合适的材料.     

脱除人主动脉瓣膜细胞的研究

目的探讨人主动脉瓣叶去细胞后作为组织工程心脏瓣膜支架的可行性。方法经胰酶-EDTA、表面活性剂和核酸酶处理,去除人主动脉瓣叶的细胞成分,测定瓣叶去细胞前、后的生物力学特性。结果人主动脉瓣去细胞后,经光镜和电镜观察,脱细胞后血管壁无细胞残留,胶原纤维和弹性纤维保留完整,形态学无明显变化;生物学性能也没有明显改变。结论去细胞效果良好,初步制造了同种主动脉血管壁脱细胞基质材料,为构建同种带瓣管道提供了较合适的材料。     

新一代人工心脏瓣膜材料血液相容性的实验研究

目的：探讨表面改性后的人工心脏瓣膜材料的血液相容性。方法：氧化钛（Ti-O)试样薄膜与目前的心脏瓣膜材料热解碳（LTIC）薄膜成对置入动物犬体内。1．腹主动脉内埋置60天。2．右心房内埋置17天,60天,100天。3．改性后Ti-O瓣膜支架与未改性瓣膜支架成对置入右心房内17天,60天,100天。结果：电镜扫描观察17天,60天Ti-O的薄膜及支架表面无血细胞粘附,100天Ti-O的薄膜支架仅有个别形态完整的血细胞,LTIC薄膜及未改性的瓣膜支架17天即可见到血球沉着,100天时血细胞和纤维状物大量粘附,支架被沉着物密布,血细胞变形有伪足,电 见血小板内有空泡,脱颗粒,微管扩张。结论：Ti-O有极好的血液相容性,有希望成为 新一代人工心脏瓣膜材料。     

聚羟基丁酸酯-羟基戊酸酯作为软骨细胞支架的实验研究

目的探讨聚羟基丁酸酯-羟基戊酸酯（PHBV）作为软骨细胞支架的可行性。方法采用“压片-热处理-粒子析出”技术制备PHBV多孔支架并观测其结构。体外分离培养软骨细胞后接种到PHBV支架进行培养，第3、7、14天后取材行扫描电镜观察；培养至2、6、10周后行组织学观察。结果PHBV支架孔径在200-300μm之间孔隙率80％；软骨细胞-PHBV复合物在体外培养期间支架无塌陷及形变，软骨细胞在支架上黏附、增殖良好，并能分泌细胞外基质。随着培养时间延长，新生软骨的组织学特征日益明显。结论PHBV支架具有适宜的孔结构和生物相容性，可以作为软骨组织工程支架材料。     

海藻酸钠在组织工程中的应用

摘要：组织工程是以细胞生物学和材料科学为基础,在体内或体外构建组织或器官的新兴学科。组织工程主要研究种子细胞和支架材料,支架材料为种子细胞提供三维立体空间结构,并且可以为细胞的气体交换、营养物质获取等提供通路,本文将针对一种具有良好安全性、可生物降解性等的天然材料—海藻酸钠(SA)的性质及在组织工程中的应用进行综述。     

循环应力对构建组织工程软骨的影响

目的 探讨循环应力对体外构建组织工程软骨的影响。方法 分离兔关节软骨细胞,体外扩增培养后种植于PGA无纺网支架上。实验组在循环应力下培养（0-200kPa,0．1Hz）,对照组静态培养。实验组和对照组均培养4周。4周后取出组织工程软骨行大体标本观察。组织化学和免疫组织化学染色观察组织工程软骨结构以及Ⅱ型胶原分泌情况。结果 体外培养4周时,实验组的组织工程软骨形态和色泽更加接近正常透明软骨,厚度和组织弹性优于对照组,实验组的软骨细胞数量和细胞外基质合成明显增加,分泌Ⅱ型胶原能力更强。结论 循环应力刺激软骨细胞合成分泌Ⅱ型胶原增加,有利于在体外构建组织工程软骨。     

经导管瓣膜镍钛合金支架镍离子释放体外模拟及细胞毒性评价

目的：研究经导管瓣膜镍钛合金支架动态疲劳中镍离子释放及其潜在体外细胞毒性,为该类创新医疗器械产品的临床前安全性评价提供参考,为推动医疗器械创新发展提供技术支撑。方法：以经导管瓣膜镍钛合金支架为研究对象,利用脉动疲劳试验模拟支架临床植入10年的情况(4亿次疲劳测试),研究其动态疲劳过程中镍离子的释放情况,并利用细胞模型评估所释放镍离子的潜在毒性效应。结果：随着镍钛合金支架疲劳试验时间的增加,测试溶液中镍离子浓度增大,测试液中镍离子最大浓度为0.075 mg·L^-1。 利用不同浓度梯度的镍离子溶液对L929细胞进行细胞毒性试验,当离子浓度为小于等于6.25 mg·L^-1时, 无体外细胞毒性,当镍离子浓度大于等于12.5 mg·L^-11时,有潜在的细胞毒性。结论：经4亿次动态疲劳测试后(模拟临床使用10年)经导管瓣膜支架中镍离子的释放量低于安全限值,无体外细胞毒性。     

Efficacy of gut lavage,hemodialysis, and hemoperfusion in the therapy of paraquat or diquat intoxication

Clinical and in vitro investigations were carried out to test the efficacy of gut lavage, hemodialysis, and hemoperfusion in the treatment of poisoning with paraquat or diquat. In a patient suffering from diquat intoxication 130 times more diquat was removed by gut lavage 30 h after ingestion than was removed by complete aspiration of the gastric contents.Determination of in vitro clearances for paraquat and diquat by hemodialysis showed that, at serum concentrations of 1–2 ppm, such as are frequently encountered in poisoning in man, toxicologically relevant quantities of herbicide cannot be removed from the body. At a concentration of 20 ppm, on the other hand, hemodialysis proved to be effective, the clearance being 70 ml/min at a blood flow rate of 100 ml/min. The efficacy of hemoperfusion with coated activated charcoal was on the whole better. Especially at concentrations around 1–2 ppm, the clearance values for hemoperfusion were some 5–7 times higher than those for hemodialysis.In a patient suffering from paraquat poisoning, both hemodialysis as well as hemoperfusion were carried out. The in vitro results could be confirmed: At serum concentrations of paraquat less than 1 ppm no clearance could be obtained by hemodialysis while by hemoperfusion with activated charcoal quite high clearance values were measured and the serum level dropped down to zero.

---

Zusammenfassung Klinische Untersuchungen und Laboratoriumsversuche wurden durchgeführt, um die Wirksamkeit von Darmspülung, Hämodialyse und Hämoperfusion bei Paraquat- und Deiquat-Vergiftungen zu prüfen.Bei einem Patienten wurde 30 Std nach Deiquat-Aufnahme durch Darmspülung 130mal mehr Deiquat entfernt als durch vollständige Aspiration des Mageninhaltes. In vitro-Versuche ergaben, daß bei Blutserumkonzentrationen von 1–2 ppm, die bei Vergiftungen oft gemessen werden, durch Hämodialyse keine toxikologisch relevanten Paraquat- oder Deiquat-Mengen entfernt werden können. Dagegen erwies sich die Hämodialyse bei 20 ppm und einer Blutumlaufgeschwindigkeit von 100 ml/min mit einer Clearance von 70 ml/min als wirksam. Die Hämoperfusion mit beschicheter Aktivkohle war in diesen Versuchen aber eindeutig überlegen, denn insbesondere bei Konzentrationen um 1–2 ppm waren die Clearance-Werte 5–7mal höher als bei der Hämodialyse.Die in vitro-Ergebnisse wurden bei einem Patienten mit einer Paraquat-Vergiftung bestätigt: Bei Konzentrationen unter 1 ppm war die Hämodialyse wirkungslos, während durch Hämoperfusion relativ hohe Clearance-Werte erreicht wurden, so daß der Serumspiegel rasch unter die Nachweisgrenze abfiel.

     

In vitro studies on sterically stabilized liposomes (SL) as enzyme carriers in organophosphorus (OP) antagonism

This study describes a new approach for organophosphorous (OP) antidotal treatment by encapsulating an OP hydrolyzing enzyme, OPA anhydrolase (OPAA), within sterically stabilized liposomes. The recombinant OPAA enzyme was derived from Alteromonas strain JD6. It has broad substrate specificity to a wide range of OP compounds: DFP and the nerve agents, soman and sarin. Liposomes encapsulating OPAA (SL)* were made by mechanical dispersion method. Hydrolysis of DFP by (SL)* was measured by following an increase of fluoride ion concentration using a fluoride ion selective electrode. OPAA entrapped in the carrier liposomes rapidly hydrolyze DFP, with the rate of DFP hydrolysis directly proportional to the amount of (SL)* added to the solution. Liposomal carriers containing no enzyme did not hydrolyze DFP. The reaction was linear and the rate of hydrolysis was first order in the substrate. This enzyme carrier system serves as a biodegradable protective environment for the recombinant OP-metabolizing enzyme, OPAA, resulting in prolongation of enzymatic concentration in the body. These studies suggest that the protection of OP intoxication can be strikingly enhanced by adding OPAA encapsulated within (SL)* to pralidoxime and atropine.     

Aquatic Insects and Trace Metals: Bioavailability,Bioaccumulation, and Toxicity

Abstract

The uptake of metals from food and water sources by insects is thought to be additive. For a given metal, the proportions taken up from water and food will depend both on the bioavailable concentration of the metal associated with each source and the mechanism and rate by which the metal enters the insect. Attempts to correlate insect trace metal concentrations with the trophic level of insects should be made with a knowledge of the feeding relationships of the individual taxa concerned. Pathways for the uptake of essential metals, such as copper and zinc, exist at the cellular level, and other nonessential metals, such as cadmium, also appear to enter via these routes. Within cells, trace metals can be bound to proteins or stored in granules. The internal distribution of metals among body tissues is very heterogeneous, and distribution patterns tend to be both metal and taxon specific. Trace metals associated with insects can be both bound on the surface of their chitinous exoskeleton and incorporated into body tissues. The quantities of trace meals accumulated by an individual reflect the net balance between the rate of metal influx from both dissolved and particulate sources and the rate of metal efflux from the organism. The toxicity of metals has been demonstrated at all levels of biological organization: cell, tissue, individual, population, and community. Much of the literature pertaining to the toxic effects of metals on aquatic insects is based on laboratory observations and, as such, it is difficult to extrapolate the data to insects in nature. The few experimental studies in nature suggest that trace metal contaminants can affect both the distribution and the abundance of aquatic insects. Insects have a largely unexploited potential as biomonitors of metal contamination in nature. A better understanding of the physico-chemical and biological mechanisms mediating trace metal bioavailability and exchange will facilitate the development of general predictive models relating trace metal concentrations in insects to those in their environment. Such models will facilitate the use of insects as contaminant biomonitors.     

Pharmacological treatment of COVID-19: an opinion paper

The precocity and efficacy of the vaccines developed so far against COVID-19 has been the most significant and saving advance against the pandemic. The development of vaccines has not prevented, during the whole period of the pandemic, the constant search for therapeutic medicines, both among existing drugs with different indications and in the development of new drugs. The Scientific Committee of the COVID-19 of the Illustrious College of Physicians of Madrid wanted to offer an early, simplified and critical approach to these new drugs, to new developments in immunotherapy and to what has been learned from the immune response modulators already known and which have proven effective against the virus, in order to help understand the current situation.     

Alzheimer’s disease – current trends in basic and clinical research. Highlights from the 16th ECNP

Advances in the molecular biological knowledge of neuronal nicotinic acetylcholine receptors (nAChRs) have led to a growing interest by the pharmaceutical industry in the development of novel compounds that selectively modulate nAChR function. The ability of (-)-nicotine, an activator of nAChRs, to enhance attentional aspects of cognition in animals and humans, to exert neuroprotective and anxiolytic-like effects, and presumably to mediate the negative correlation between smoking and Alzheimer's (and Parkinson's) Disease, has focused interest on the potential therapeutic utility of modulators of nAChR function for treatment of some of the deficits associated with these progressive, neurodegenerative conditions. Numerous compounds are known which activate nAChRs and which might serve as lead compounds toward the development of such agents. The pharmacologic diversity of neuronal nAChR subtypes suggests the possibility of developing selective compounds which would have more favourable side-effect profiles than existing agents. This broader class of agents, collectively called cholinergic channel modulators (ChCMs), is anticipated to encompass compounds which would have more favourable side-effect profiles than existing agents, which generally exhibit low selectivity. This selectivity may be achieved by preferentially activating some subtypes of nAChRs (i.e., Cholinergic Channel Activators, ChCAs) or inhibiting the function of other subtypes (Cholinergic Channel Inhibitors, ChCIs). An overview of the biology of nAChRs and the rationale for the use of ChCMs for the treatment of dementia related to neurodegenerative diseases are presented, followed by a discussion of lead compounds and compounds under consideration for clinical evaluation.     

Steroidal sapogenin contents in some domestic plants

In order to find out the values of the steroid resources for the future use. the compositions and contents of steroidal sapogenins from 13 domestic plants have been investigated. As a result,Dioscorea nipponica, D. quinqueloba andSmilax china were found to have large amount of diosgenin. And pennogenin inTrillium kamtschaticum andParis verticillata, yuccagenin inAllium fistulosum, hecogenin inAgave americana and neochlorogenin inSolanum nigum were appeared to be major steroidal sapogenins.