Human Hepatocyte Isolation: Does Portal Vein Embolization Affect the Outcome?
Human Hepatocyte Isolation: Does Portal Vein Embolization Affect the Outcome?
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DOI：
10.1089/ten.tec.2015.0190
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发表时间：
2016-01-01
影响因子：
3
通讯作者：
Raschzok, Nathanael
中科院分区：
文献类型：
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作者：
Kluge, Martin;Reutzel-Selke, Anja;Raschzok, Nathanael
原代人肝细胞广泛用于再生医学的基础研究、药物测试和治疗概念。人肝细胞可以从切除的肝组织中分离。术前门静脉栓塞（PVE）越来越多地用于通过诱导未来残余肝组织的选择性肥大来降低术后肝再生延迟的风险。本研究的目的是研究PVE对肝细胞分离结果的影响。采用两步胶原酶灌注技术，然后进行Percoll纯化，从部分肝切除术（n=190）获得的肝组织中分离原代人肝细胞。在这些肝切除术中，27例（14.2%）使用术前接受PVE的患者的肝组织进行分离。使用文献中描述的与肝细胞分离结果相关的参数对所有分离物进行表征。比较PVE组和非PVE组Percoll纯化前后的分离效果。在培养6天以上的肝细胞上清液中测量代谢参数（转氨酶、尿素、白蛋白和血管内皮生长因子分泌）（PVE：n=4和非PVE：n=3）。PVE和非PVE组在供体参数（性别、年龄和手术适应症）、隔离参数（肝脏重量和冷缺血时间）和肝脏组织质量方面相似。PVE组和非PVE组的平均初始活细胞产量无差异（10.16 +/- 2.03x10（6）个细胞/g vs. 9.70 +/- 0.73x10（6）个细胞/g，p=0.499）。PVE组的初始存活率略好（77.8%+/- 2.03% vs. 74.4%+/- 1.06%）。Percoll纯化后的平均活细胞产量（p=0.819）和平均活力（p=0.141）在组间无差异。PVE对培养肝细胞的酶渗漏和代谢活性无影响。尽管PVE导致肝血流的剧烈代谢改变和变化，但栓塞的肝组织是分离原代人肝细胞的合适来源，并且在细胞产量和活力方面与未处理的肝组织等同。
Primary human hepatocytes are widely used for basic research, pharmaceutical testing, and therapeutic concepts in regenerative medicine. Human hepatocytes can be isolated from resected liver tissue. Preoperative portal vein embolization (PVE) is increasingly used to decrease the risk of delayed postoperative liver regeneration by induction of selective hypertrophy of the future remnant liver tissue. The aim of this study was to investigate the effect of PVE on the outcome of hepatocyte isolation. Primary human hepatocytes were isolated from liver tissue obtained from partial hepatectomies (n=190) using the two-step collagenase perfusion technique followed by Percoll purification. Of these hepatectomies, 27 isolations (14.2%) were performed using liver tissue obtained from patients undergoing PVE before surgery. All isolations were characterized using parameters that had been described in the literature as relevant for the outcome of hepatocyte isolation. The isolation outcomes of the PVE and the non-PVE groups were then compared before and after Percoll purification. Metabolic parameters (transaminases, urea, albumin, and vascular endothelial growth factor secretion) were measured in the supernatant of cultured hepatocytes for more than 6 days (PVE: n=4 and non-PVE: n=3). The PVE and non-PVE groups were similar in regard to donor parameters (sex, age, and indication for surgery), isolation parameters (liver weight and cold ischemia time), and the quality of the liver tissue. The mean initial viable cell yield did not differ between the PVE and non-PVE groups (10.16 +/- 2.03x10(6) cells/g vs. 9.70 +/- 0.73x10(6) cells/g, p=0.499). The initial viability was slightly better in the PVE group (77.8%+/- 2.03% vs. 74.4%+/- 1.06%). The mean viable cell yield (p=0.819) and the mean viability (p=0.141) after Percoll purification did not differ between the groups. PVE had no effect on enzyme leakage and metabolic activity of cultured hepatocytes. Although PVE leads to drastic metabolic alterations and changes in hepatic blood flow, embolized liver tissue is a suitable source for the isolation of primary human hepatocytes and is equivalent to untreated liver tissue in regard to cell yield and viability.