Yovchev Mladen I, Locker Joseph, Oertel Michael
Dept. of Pathology (Division of Experimental Pathology), University of Pittsburgh, Pittsburgh, PA, United States.
Dept. of Pathology (Division of Experimental Pathology), University of Pittsburgh, Pittsburgh, PA, United States; McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, PA, United States.
J Hepatol. 2016 Jun;64(6):1348-57. doi: 10.1016/j.jhep.2016.01.036. Epub 2016 Feb 5.
BACKGROUND & AIMS: Current research focuses on developing alternative strategies to restore decreased liver mass prior to the onset of end-stage liver disease. Cell engraftment/repopulation requires regeneration in normal liver, but we have shown that severe liver injury stimulates repopulation without partial hepatectomy (PH). We have now investigated whether a less severe injury, secondary biliary fibrosis, would drive engraftment/repopulation of ectopically transplanted mature hepatocytes.
Ductular proliferation and progressive fibrosis in dipeptidyl-peptidase IV (DPPIV)(-) F344 rats was induced by common bile duct ligation (BDL). Purified DPPIV(+)/green fluorescent protein (GFP)(+) hepatocytes were infused without PH into the spleen of BDL rats and compared to rats without BDL.
Within one week, transplanted hepatocytes were detected in hepatic portal areas and at the periphery of expanding portal regions. DPPIV(+)/GFP(+) repopulating cell clusters of different sizes were observed in BDL rats but not untreated normal recipients. Surprisingly, some engrafted hepatocytes formed CK-19/claudin-7 expressing epithelial cells resembling cholangiocytes within repopulating clusters. In addition, substantial numbers of hepatocytes engrafted at the intrasplenic injection site assembled into multicellular groups. These also showed biliary "transdifferentiation" in the majority of intrasplenic injection sites of rats that received BDL but not in untreated recipients. PCR array analysis showed upregulation of osteopontin (SPP1). Cell culture studies demonstrated increased Itgβ4, HNF1β, HNF6, Sox-9, and CK-19 mRNA expression in hepatocytes incubated with osteopontin, suggesting that this secreted protein promotes dedifferentiation of hepatocytes.
Our studies show that biliary fibrosis stimulates liver repopulation by ectopically transplanted hepatocytes and also stimulates hepatocyte transition towards a biliary epithelial phenotype.
目前的研究聚焦于开发替代策略，以在终末期肝病发作前恢复减少的肝脏质量。细胞植入/再增殖在正常肝脏中需要再生，但我们已表明，严重肝损伤可在不进行部分肝切除术（PH）的情况下刺激再增殖。我们现在研究了一种不太严重的损伤，即继发性胆汁性纤维化，是否会驱动异位移植的成熟肝细胞的植入/再增殖。
通过胆总管结扎（BDL）诱导二肽基肽酶IV（DPPIV）（-）F344大鼠的小胆管增殖和进行性纤维化。将纯化后的DPPIV（+）/绿色荧光蛋白（GFP）（+）肝细胞在不进行PH的情况下注入BDL大鼠的脾脏，并与未进行BDL的大鼠进行比较。
在一周内，在肝门区域和扩张的门脉区域周边检测到移植的肝细胞。在BDL大鼠中观察到不同大小的DPPIV（+）/GFP（+）再增殖细胞簇，而未处理的正常受体中未观察到。令人惊讶的是，一些植入的肝细胞在再增殖簇中形成了表达细胞角蛋白19（CK-19）/闭合蛋白7的上皮细胞，类似于胆管细胞。此外，大量在脾内注射部位植入的肝细胞聚集形成多细胞团。在接受BDL的大鼠的大多数脾内注射部位，这些细胞团也显示出胆管“转分化”，而未处理受体中则未出现。PCR阵列分析显示骨桥蛋白（SPP1）上调。细胞培养研究表明，与骨桥蛋白一起孵育的肝细胞中整合素β4（Itgβ4）、肝细胞核因子1β（HNF1β）、肝细胞核因子6（HNF6）、性别决定区Y盒9（Sox-9）和CK-19的mRNA表达增加，表明这种分泌蛋白促进肝细胞去分化。
我们的研究表明，胆汁性纤维化可刺激异位移植的肝细胞进行肝脏再增殖，并刺激肝细胞向胆管上皮表型转变。
