Ali A M, Rossouw H C, Silove M, Walt J G
Department of Physiology, Faculty of Veterinary Science, University of Pretoria, Onderstepoort 0110, South Africa.
Exp Physiol. 2000 Sep;85(5):469-78.
The study was designed to develop an improved technique for perfusing the isolated caudal lobe of sheep liver. Twenty caudal lobes were perfused for 3-4 h, in a non-recirculating mode, with Krebs-Henseleit bicarbonate buffer. The perfusion system was designed to give a constant flow. The hepatic viability and functional normality of the perfused lobe were assessed by measuring the perfusion flow rate, pH, K+ efflux, O2 uptake, substrate uptake, gluconeogenesis from propionate and amino acids, and ureagenesis from ammonia and amino acids. Liver tissue was sampled for histological examination, as well as for the determination of liver glycogen and wet : dry weight ratio. The perfusion flow rate and pH were both stable throughout the perfusion. The potassium concentration in the effluent perfusate did not increase during the perfusion, suggesting that there was no loss of viability or hypoxia. The perfused lobe extracted more than 50% of the O2 supply. The rate of oxygen consumption was comparable to the rate reported in vivo. The initial glycogen content was reduced by about 40% after 4 h perfusion. The wet : dry weight ratio was 3.6, consistent with the absence of tissue oedema. Urea production was stimulated when NH4Cl (0.3 mM) was added to the medium but there was no significant increase in urea release when alanine (0.15 mM), glutamine (0.2 mM) or lysine (0.2 mM) was added. Urea production, however, increased by about 171% when a physiological mixture of amino acids was added. Propionate (0.5 mM), alanine and glutamine stimulated glucose production but not lysine or the complete amino acid mixture. Glutamine release was lower than that reported in the rat liver. Changing the direction of flow also revealed an apparent difference between livers from sheep and rats in their metabolism of ammonia. The improved technique offers a simple practical and inexpensive approach to many problems in ruminant physiology and nutritional biochemistry.
本研究旨在开发一种改进的技术，用于灌注绵羊肝脏的离体尾叶。20个尾叶以非循环模式用Krebs-Henseleit碳酸氢盐缓冲液灌注3 - 4小时。灌注系统设计为提供恒定流量。通过测量灌注流速、pH值、钾离子外流、氧气摄取、底物摄取、丙酸盐和氨基酸的糖异生以及氨和氨基酸的尿素生成，评估灌注叶的肝脏活力和功能正常性。采集肝脏组织进行组织学检查，以及测定肝糖原和湿重：干重比。在整个灌注过程中，灌注流速和pH值均保持稳定。灌注流出液中的钾浓度在灌注过程中没有增加，表明没有活力丧失或缺氧。灌注叶摄取了超过50%的氧气供应。氧气消耗速率与体内报道的速率相当。灌注4小时后，初始糖原含量降低了约40%。湿重：干重比为3.6，与无组织水肿一致。当向培养基中添加NH4Cl（0.3 mM）时，尿素生成受到刺激，但当添加丙氨酸（0.15 mM）、谷氨酰胺（0.2 mM）或赖氨酸（0.2 mM）时，尿素释放没有显著增加。然而，当添加氨基酸的生理混合物时，尿素生成增加了约171%。丙酸盐（0.5 mM）、丙氨酸和谷氨酰胺刺激葡萄糖生成，但赖氨酸或完整氨基酸混合物则无此作用。谷氨酰胺释放低于大鼠肝脏中的报道。改变血流方向还揭示了绵羊和大鼠肝脏在氨代谢方面的明显差异。这种改进的技术为反刍动物生理学和营养生物化学中的许多问题提供了一种简单、实用且廉价的方法。
