Tordjmann T, Berthon B, Lardeux B, Moreau A, Jacquemin E, Combettes L, Feldmann G, Claret M
Unité de Recherche U442, Institut National de la Santé et de la Recherche Médicale, Université Paris-Sud, Orsay, France.
Hepatology. 1997 Dec;26(6):1592-9. doi: 10.1053/jhep.1997.v26.pm0009398003.
Morphological and functional heterogeneity of hepatocytes according to their position in the liver lobule has been known for many years. The digitonin-collagenase perfusion technique is widely used to study hepatocyte heterogeneity and has yielded reliable data. However, with this procedure, periportal (PP) or perivenous (PV) hepatocytes are isolated from different livers, allowing only comparison between cell populations issued from two separate animals. To overcome this drawback, we have modified this technique by perfusing the two main rat liver lobes of a single animal in succession. The procedure involved alternate clamping of the median and the left lateral lobes, restricting digitonin infusion to one lobe via the portal vein, and to the other via the caudal vena cava. Lobe exclusion during digitonin perfusion, and zonal restriction of digitonin-induced damage, were monitored using macroscopic and histological controls. We compared our results with previous data on PP and PV hepatocytes issued from two different livers using the conventional digitonin-collagenase perfusion technique. First, we found that the cellular sensitivity to angiotensin II, a calcium-mobilizing agonist, was 60% to 80% higher in PV than in PP hepatocytes, whereas, previously, no difference had been recorded. Second, we found that albumin messenger RNAs (mRNAs) were 35% more abundant in PP than in PV hepatocytes, whereas, previously, larger differences had been reported. Our results show that PP and PV hepatocytes may be isolated from a single liver using an improved digitonin-collagenase perfusion technique. Furthermore, we suggest that zonal differences can be artificially masked or amplified when comparing PP and PV cell populations from two different livers, indicating that it is preferable to use a single liver for accurate zonal comparisons between hepatocytes.
多年来，人们已经知道肝细胞根据其在肝小叶中的位置存在形态和功能异质性。洋地黄皂苷 - 胶原酶灌注技术被广泛用于研究肝细胞异质性，并产生了可靠的数据。然而，通过这种方法，门周（PP）或中央静脉周围（PV）肝细胞是从不同的肝脏中分离出来的，这仅允许对来自两只不同动物的细胞群体进行比较。为了克服这一缺点，我们对该技术进行了改进，通过依次灌注同一只动物的两个主要大鼠肝叶。该过程包括交替夹闭中叶和左外叶，通过门静脉将洋地黄皂苷注入一个叶，通过尾静脉将其注入另一个叶。在洋地黄皂苷灌注期间监测叶的排除情况，以及洋地黄皂苷诱导损伤的区域限制，通过宏观和组织学对照进行监测。我们将我们的结果与先前使用传统洋地黄皂苷 - 胶原酶灌注技术从两个不同肝脏获得的关于PP和PV肝细胞的数据进行了比较。首先，我们发现，作为一种钙动员激动剂，血管紧张素II对PV肝细胞的细胞敏感性比对PP肝细胞高60%至80%，而之前未记录到差异。其次，我们发现PP肝细胞中的白蛋白信使核糖核酸（mRNA）比PV肝细胞中的丰富35%，而之前报道的差异更大。我们的结果表明，使用改进的洋地黄皂苷 - 胶原酶灌注技术可以从单个肝脏中分离出PP和PV肝细胞。此外，我们认为，在比较来自两个不同肝脏的PP和PV细胞群体时，区域差异可能会被人为掩盖或放大，这表明最好使用单个肝脏来进行肝细胞之间准确的区域比较。
