Fukumoto Masayuki, Miyamoto Daisuke, Soyama Akihiko, Hara Takanobu, Maruya Yasuhiro, Li Peilin, Matsushima Hajime, Migita Kazushige, Enjoji Takahiro, Tetsuo Hanako, Fujita Takuro, Yamashita Mampei, Imamura Hajime, Adachi Tomohiko, Kanetaka Kengo, Ochiya Takahiro, Eguchi Susumu
Department of Surgery, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan.
Department of Molecular and Cellular Medicine, Tokyo Medical University, Tokyo, Japan.
PLoS One. 2024 Dec 5;19(12):e0313312. doi: 10.1371/journal.pone.0313312. eCollection 2024.
We previously reported the efficacy of chemically induced liver progenitors (CLiP) as a source of cells for transplantation in patients with liver disease. This study aimed to characterize CLiP derived from steatotic livers using a pig model for future clinical applications. Livers were removed from miniature pigs with diet-induced steatosis and normal livers by laparoscopic hepatectomy. Mature hepatocytes (MH) isolated from the livers of each group were cultured in differentiation medium composed of Y-27632, A-83-01, and CHIR99021 (YAC medium). The characteristics of CLiP, including liver-specific function, proliferative capacity in vivo, and extracellular vesicles (EVs) production, were evaluated. Although CLiP in both groups expressed hepatic progenitor cell markers (Epithelial cell adhesion molecule and Trophoblast cell surface antigen 2), the proliferative potential was higher for the disease group than the healthy group. In contrast, markers of functional MH after re-differentiation were only detected in the healthy group. Both groups showed high cell viability and the ability to differentiate into albumin-positive cells in vivo. EVs counts were lower in disease-derived CLiP than in the normal group; however, there were no differences in microRNA expression within EVs. Using a pig model, CLiP was successfully produced from a liver that reproduced steatotic liver disease. Although there were slightly fewer EVs from CLiP in the disease group than in the normal liver group, the in vivo proliferative capacity of CLiP was high. Therefore, CLiP induced in the steatotic liver are a promising source for cell therapy in patients with liver disease.
我们之前报道了化学诱导肝祖细胞（CLiP）作为肝病患者移植细胞来源的疗效。本研究旨在利用猪模型对源自脂肪变性肝脏的CLiP进行表征，以供未来临床应用。通过腹腔镜肝切除术从饮食诱导脂肪变性的小型猪和正常肝脏中取出肝脏。从每组肝脏中分离出的成熟肝细胞（MH）在由Y-27632、A-83-01和CHIR99021（YAC培养基）组成的分化培养基中培养。评估了CLiP的特性，包括肝脏特异性功能、体内增殖能力和细胞外囊泡（EVs）产生。尽管两组中的CLiP均表达肝祖细胞标志物（上皮细胞粘附分子和滋养层细胞表面抗原2），但疾病组的增殖潜力高于健康组。相比之下，再分化后功能性MH的标志物仅在健康组中检测到。两组均显示出高细胞活力以及在体内分化为白蛋白阳性细胞的能力。疾病来源的CLiP中的EVs计数低于正常组；然而，EVs内的微小RNA表达没有差异。利用猪模型，成功地从再现脂肪变性肝病的肝脏中产生了CLiP。尽管疾病组CLiP产生的EVs略少于正常肝脏组，但CLiP的体内增殖能力很高。因此，脂肪变性肝脏中诱导产生的CLiP是肝病患者细胞治疗的一个有前景的来源。
