Sales K M, Kingston S T, Doyle K M, Purcell W M
Department of Surgery, Royal Free and University College Medical School, Rowland Hill Street, Hampstead NW3 2PF, UK.
Toxicology. 2004 Feb 15;195(2-3):187-202. doi: 10.1016/j.tox.2003.10.002.
Organophosphate induced delayed neuropathy (OPIDN) has been studied extensively but the mechanisms of toxicity remain unclear. It is generally accepted that the inhibition and ageing (dealkylation) of the B-esterase neuropathy target esterase (NTE) is integral to axonal loss. At present, the only way of detecting compounds that induce OPIDN is the hen test, an animal model. In this study, we preliminary validated hen embryo brain spheroids (HEBS) for the study of organophosphate (OP) toxicity. Hen brain spheroids have been characterised previously, although they have never been fully optimised for OP testing. We optimised the levels of acetylcholine esterase (AChE) and neuropathy target esterase by adapting the culture technique and using chemically defined media. Spheroid cultures were maintained for 35 days and viability and enzyme levels were monitored over this time. Levels of AChE and NTE in this system remained stable over the 35 day period. Using transmission electron microscopy, we have shown synaptogenesis within HEBS earlier than previously suggested in spheroid culture. These studies indicate that HEBS may be useful for the study of OP-induced toxicity and that the long-term stability of the cultures makes it an ideal candidate for studying OPIDN.
有机磷诱导的迟发性神经病（OPIDN）已得到广泛研究，但毒性机制仍不清楚。人们普遍认为，B酯酶神经病变靶酯酶（NTE）的抑制和老化（脱烷基化）是轴突丧失的必要条件。目前，检测诱导OPIDN的化合物的唯一方法是母鸡试验，一种动物模型。在本研究中，我们初步验证了母鸡胚胎脑球体（HEBS）用于有机磷（OP）毒性研究的可行性。母鸡脑球体此前已有特征描述，尽管它们从未针对OP测试进行过全面优化。我们通过调整培养技术和使用化学成分明确的培养基，优化了乙酰胆碱酯酶（AChE）和神经病变靶酯酶的水平。球体培养维持了35天，并在此期间监测了活力和酶水平。该系统中AChE和NTE的水平在35天内保持稳定。使用透射电子显微镜，我们已证明HEBS内的突触形成比此前球体培养中所表明的时间更早。这些研究表明，HEBS可能对OP诱导的毒性研究有用，并且培养物的长期稳定性使其成为研究OPIDN的理想候选对象。
