Three-dimensional perfused cell culture

Compelling evidence suggests the limitation and shortcomings of the current and well established cell culture method using multi-well plates, flasks and Petri dishes. These are particularly important when cell functions are sensitive to the local microenvironment, cell–cell and cell–extracellular matrix interactions. There is a clear need for advanced cell culture systems which mimic in vivo and more physiological conditions. This review summarises and analyses recent progress in three dimensional (3D) cell culture with perfusion as the next generation cell culture tools, while excluding engineered tissue culture where three dimensional scaffold has to be used for structural support and perfusion for overcoming mass transfer control. Apart from research activities in academic community, product development in industry is also included in this review.     

Hepatic metabolism of vasoactive intestinal polypeptide (VIP) in the rat

Vasoactive intestinal polypeptide (VIP) is released into the portal circulation by a meal stimulus, but is rapidly cleared from plasma. Although it is known to bind to receptors on liver cells, the role of the liver in the clearance of VIP is not clearly defined. We therefore studied the disappearance of VIP in recirculating and in single pass isolated perfused rat liver (IPRL) preparations. Disappearance of added VIP was rapid in recirculating IPRL experiments with a half life of ca. 30 min. In single-pass steady-state studies in which livers were perfused at 16 ml/min for 30 min, clearance of VIP was complete (16 ml/min) at concentrations of 500 fmol/ml, but clearance fell to 3 and 1 ml/min at perfusate concentrations of 8 and 40 pmol/ml respectively. Further experiments to evaluate whether VIP was disappearing in perfusate itself demonstrated substantial metabolism of VIP in perfusate which had previously been circulated through a liver for 90 min. The products of metabolism were identical to those found in the IPRL. We conclude that VIP is rapidly cleared as it passes through the isolated perfused rat liver model with a significant proportion of clearance attributable to release of a peptidase from the liver into the perfusate.     

Stereoselective Nicotine-Induced Release of Dopamine from Striatal Synaptosomes: Concentration Dependence and Repetitive Stimulation

Using a sensitive perfusion system we have studied the nicotine-induced release of [3H]dopamine ([( 3H]DA) from striatal synaptosomes. Nicotine-evoked release was concentration dependent with an EC50 of 3.8 microM. The response to 1 microM nicotine was comparable to that to 16 mM K+; 10 microM veratridine evoked a larger response. All three stimuli were Ca2+ dependent but only the response to veratridine was blocked by tetrodotoxin. Repetitive stimulations by 1 microM (-)-nicotine (100 microliters) at 30-min intervals resulted in similar levels of [3H]DA release; higher concentrations of (-)-nicotine resulted in an attenuation of the response particularly following the third stimulation. This may reflect desensitisation or tachyphylaxis of the presynaptic nicotinic receptor. The action of nicotine was markedly stereoselective: a 100-fold higher concentration of (+)-nicotine was necessary to evoke the same level of response as 1 microM (-)-nicotine. It is proposed that these presynaptic nicotinic receptors on striatal terminals are equivalent to high-affinity nicotine binding sites described in mammalian brain.     

Effect of brief vascular perfusion on the ultrastructure and activity of mitochondria isolated from the rat heart

Summary Mitochondria isolated from heart tissue after a 1-min perfusion with Hanks medium were found to have significantly lower rates of State-3 respiration and respiratory control ratios compared to mitochondria isolated from non-perfused hearts. Examination of the mitochondrial preparations by electron microscopy revealed that a large proportion of the mitochondria isolated from perfused heart tissue were swollen and broken compared to mitochondria from non-perfused hearts.     

Measurement of motive force for cytoplasmic streaming in characean internodes

Summary With an attempt to measure the motive force responsible for cytoplasmic streaming in characean internodal cells, the difference between densities of cytoplasm and vacuolar sap was heightened by about 10 times (density of vacuolar sap was made larger than that of cytoplasm) by replacing the natural vacuolar sap ofChara corallina with an artificial one of higher density. Endoplasmic flow contiguous to the peripheral actin cables (peripheral flow of endoplasm) in the centrifugal direction was not influenced at all by the application of centrifugal acceleration up to 1400 g. We thus concluded that the motive force for the peripheral flow should be much larger than 12dyn/cm², a figure more than 10 times larger than that for bulk endop lasmic flow so far reported.Dedicated to Emeritus Professor Noburo Kamiya on the occasion of his 80th birthday     

原发性肾病综合征合并血栓栓塞的临床研究

摘要 目的：肺栓塞及下肢深静脉血栓是影响肾病综合征(Nephrotic Syndrome,NS)预后的重要因素,但在其发生率以及治疗方面仍存在争议。本文将通过回顾性的病例研究对NS患者中血栓栓塞事件的发生率、危险因素以及是否需要早期干预等问题进行探讨。方法：收集上海交通大学医学院附属新华医院肾内科2014年1-12月诊断为原发性NS患者,并同时收集患者基本临床信息,血栓检查结果以及NS相关血液及尿液检查结果等在内的临床资料,采用多种统计方法分析血栓发生率及其危险因素,并进一步讨论NS患者的预防性抗凝措施。结果：NS患者下肢静脉血栓或肺栓塞的发生率均为15.4%,两种血栓同时发生的概率为9.6%。NS患者血栓事件的发生率与D-二聚体、血浆白蛋白、病程、病理类型呈明显相关,且具有统计学意义。结论：NS患者血栓栓塞的发生率与D-二聚体、血浆白蛋白、病程、病理类型呈显著相关,膜性肾病患者血栓栓塞发生率最高。如NS患者合并高危因素,建议定期筛查,综合评估后预防性抗凝。     

脑内微透析采样技术及其在神经科学中的应用

作为一种新的在体化学采样技术,脑内微透析引起了神经科学家的关注。它与迅速发展起来的高灵敏度的微量分析技术相结合,实现了对体内细胞外环境中化学物质变化的动态监测,从而在神经科学领域获得应用。本系统地介绍了这一新技术的原理和方法,并扼要地介绍了一这一技术在神经科学中的应用及其取得的新进展,并结合本实验室的工作经验,对该技术存在的一些问题进行了讨论。     

A comparison of the effects of different perfusion regimes on the structure of the isolated human placental lobule

Summary Isolated lobules of normal term human placentas were perfused using two different procedures. In the first more conventional system, open-circuit perfusion of both the maternal and the fetal circulations with Earle's solution containing dextran was established and maintained for either 30 min or 1 h. In the second series of experiments both circulations were perfused in separate closed circuits with a mixture of fresh autologous fetal blood and Earle's solution for 0, 1, 2 or 3 h. In both series the lobule was then fixed by perfusion through the fetal circulation.Light and electron-microscopic examination of a set of tissue samples from each perfused lobule showed substantial differences between the effects of these two types of perfusion procedure. Tissue from lobules perfused by the open-circuit blood-free procedure showed patchy but severe cell swelling and vacuolation of the trophoblast after only one hour's perfusion. Particularly striking was swelling and disruption of a large proportion of the mitochondria in all placental cell types. By contrast, placental tissue from the closed-circuit perfusion with blood-containing medium showed little change over a period of two hours, while after three hours it showed oedema and microvillous damage, but no sign of cell swelling and little mitochondrial damage.It is concluded that the viability of the perfused human placental lobule depends on the type of perfusate used, and that the use of a fetal blood-enriched perfusate is of considerable value in maintenance of the preparation as assessed by structural criteria.     

The isolated perfused rabbit ovary — A model for studies of ovarian function

Summary In the present investigation the ultrastructure of isolated rabbit ovaries, perfused with different media for various time periods, was studied. The steroid hormone production by the perfused ovary was also determined. Perfusion with Medium 199 results in prominent interstitial ovarian oedema which increases with perfusion time. Even after the addition of 6–10 % Dextran T40, oedema appears in the interstitial tissue of the ovary. Perfusion solutions with osmotically active colloid particles of large molecular size (Dextran T70; average molecular weight 70,000 and bovine serum albumin), cause less distortion in the ovarian structure, and ultrastructurally the ovarian tissues appear essentially the same as in the control ovaries.The results indicate that the perfused rabbit ovary, under strictly controlled conditions, can be used as an experimental model for studies of various aspects of ovarian function, including follicular rupture.