|
1
|
Zhou X, Liu WM, Sun HY, Peng Y, Huang RJ, Chen CY, Zhang HD, Zhou SA, Wu HP, Tang D, Huang WJ, Wu H, Li QG, Zhai B, Xia Q, Yu WF, Yan HX. Hepatocyte-derived liver progenitor-like cells attenuate liver cirrhosis via induction of apoptosis in hepatic stellate cells. Hepatol Commun 2025; 9:e0614. [PMID: 39878682 PMCID: PMC11781762 DOI: 10.1097/hc9.0000000000000614] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/28/2024] [Accepted: 10/12/2024] [Indexed: 01/31/2025] Open
Abstract
BACKGROUND Cell therapy demonstrates promising potential as a substitute therapeutic approach for liver cirrhosis. We have developed a strategy to effectively expand murine and human hepatocyte-derived liver progenitor-like cells (HepLPCs) in vitro. The primary objective of the present study was to apply HepLPCs to the treatment of liver cirrhosis and to elucidate the underlying mechanisms responsible for their therapeutic efficacy. METHODS The effects of allogeneic or xenogeneic HepLPC transplantation were investigated in rat model of liver cirrhosis. Liver tissues were collected and subjected to immunostaining to assess changes in histology. In vitro experiments used HSCs to explore the antifibrotic properties of HepLPC-secretomes and their underlying molecular mechanisms. Additionally, proteomic analysis was conducted to characterize the protein composition of HepLPC-secretomes. RESULTS Transplantation of HepLPCs resulted in decreased active fibrogenesis and net fibrosis in cirrhosis models. Apoptosis of HSCs was observed in vivo after HepLPC treatment. HepLPC-secretomes exhibited potent inhibition of TGF-β1-induced HSC activation and promoted apoptosis through signal transducer and activator of transcription (STAT)1-mediated pathways in vitro. Furthermore, synergistic effects between amphiregulin and FGF19 within HepLPC-secretomes were identified, contributing to HSC apoptosis and exerting antifibrotic effects via activation of the janus kinase-STAT1 pathway. CONCLUSIONS HepLPCs have the potential to ameliorate liver cirrhosis by inducing STAT1-dependent apoptosis in HSCs. Amphiregulin and FGF19 are key factors responsible for STAT1 activation, representing promising novel therapeutic targets for the treatment of liver cirrhosis.
Collapse
Affiliation(s)
- Xu Zhou
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
- Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China
- Key Laboratory of Anesthesiology (Shanghai Jiao Tong University), Ministry of Education, Shanghai, China
| | - Wen-Ming Liu
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
- Key Laboratory of Anesthesiology (Shanghai Jiao Tong University), Ministry of Education, Shanghai, China
- Institute for Regenerative Medicine, Shanghai East Hospital, School of Life Sciences and Technology, Tongji University, Shanghai, China
| | - Han-Yong Sun
- Department of Hepatic Surgery, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Yuan Peng
- Department of Interventional Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Ren-Jie Huang
- Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China
| | - Cai-Yang Chen
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Hong-Dan Zhang
- Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China
| | - Shen-Ao Zhou
- Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China
| | - Hong-Ping Wu
- Molecular Epidemiology Laboratory, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, China
| | - Dan Tang
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
- Key Laboratory of Anesthesiology (Shanghai Jiao Tong University), Ministry of Education, Shanghai, China
| | - Wei-Jian Huang
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Han Wu
- Hubei Key Laboratory of Tumour Biological Behaviors, Department of Radiation and Medical Oncology, Zhongnan Hospital of Wuhan University, Wuhan, China
| | - Qi-Gen Li
- Department of Hepatic Surgery, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Bo Zhai
- Department of Interventional Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Qiang Xia
- Department of Hepatic Surgery, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| | - Wei-Feng Yu
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
- Key Laboratory of Anesthesiology (Shanghai Jiao Tong University), Ministry of Education, Shanghai, China
| | - He-Xin Yan
- Department of Anesthesiology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
- Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China
- Key Laboratory of Anesthesiology (Shanghai Jiao Tong University), Ministry of Education, Shanghai, China
- Department of Interventional Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
- Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
| |
Collapse
|
|
2
|
Nobakht Lahrood F, Saheli M, Farzaneh Z, Taheri P, Dorraj M, Baharvand H, Vosough M, Piryaei A. Generation of Transplantable Three-Dimensional Hepatic-Patch to Improve the Functionality of Hepatic Cells In Vitro and In Vivo. Stem Cells Dev 2020; 29:301-313. [PMID: 31856676 DOI: 10.1089/scd.2019.0130] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022] Open
Abstract
Cell therapy and tissue engineering (TE) are considered alternative therapeutic approaches to organ transplantation. Since cell therapy approaches achieved little success for liver failure treatment, liver TE is considered a more promising alternative. In this study, we produced a liver tissue equivalent (called "liver-derived extracellular matrix scaffold [LEMS]-Patch") by co-culture of human bone marrow stromal cells, human umbilical vein endothelial cells, and a hepatoma cell line, Huh7, within an artificial three-dimensional liver-extracellular matrix scaffold. The results showed significant increase in the liver-specific gene expression and hepatic functions, in terms of albumin (ALB) and fibrinogen secretion, urea production, and cytochrome inducibility in the LEMS-Patch compared to controls. In addition, transplanted LEMS-Patch was successfully incorporated into the recipient liver of acute liver failure mice and produced human ALB. Consequently, our data demonstrated that the generated LEMS-Patch could be used as a good platform for functional improvement of hepatic cells in vitro and in vivo.
Collapse
Affiliation(s)
- Fatemeh Nobakht Lahrood
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Mona Saheli
- Department of Anatomy, Afzalipour School of Medicine, Kerman University of Medical Sciences, Kerman, Iran
| | - Zahra Farzaneh
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Payam Taheri
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Mahshad Dorraj
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Hossein Baharvand
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
- Department of Developmental Biology, University of Science and Culture, Tehran, Iran
| | - Massoud Vosough
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
- Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Abbas Piryaei
- Department of Biology and Anatomical Sciences, School of Medicine, and School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
- Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| |
Collapse
|
|
3
|
Park JY, Han J, Jung HS, Lee G, Kim HJ, Cho GS, Park HJ, Han C, Kim JS, Kim JH. Synthetic probes for in vitro purification and in vivo tracking of hepatocytes derived from human pluripotent stem cells. Biomaterials 2019; 222:119431. [DOI: 10.1016/j.biomaterials.2019.119431] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2019] [Revised: 07/23/2019] [Accepted: 08/13/2019] [Indexed: 02/06/2023]
|
|
4
|
de Vries RJ, Banik PD, Nagpal S, Weng L, Ozer S, van Gulik TM, Toner M, Tessier SN, Uygun K. Bulk Droplet Vitrification: An Approach to Improve Large-Scale Hepatocyte Cryopreservation Outcome. LANGMUIR : THE ACS JOURNAL OF SURFACES AND COLLOIDS 2019; 35:7354-7363. [PMID: 30514081 PMCID: PMC6548701 DOI: 10.1021/acs.langmuir.8b02831] [Citation(s) in RCA: 30] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/27/2023]
Abstract
Loss of hepatocyte viability and metabolic function after cryopreservation is still a major issue. Although vitrification is a promising alternative, it has generally been proven to be unsuitable for vitrification of large cell volumes which is required for clinical applications. Here, we propose a novel bulk droplet (3-5 mm diameter) vitrification method which allows high throughput volumes (4 mL/min), while using a low preincubated CPA concentration (15% v/v) to minimize toxicity and loss of cell viability and function. We used rapid (1.25 s) osmotic dehydration to concentrate a low preincubated intracellular CPA concentration ahead of vitrification, without the need of fully equilibrating toxic CPA concentrations. We compared direct postpreservation viability, long-term viability, and metabolic function of bulk droplet vitrified, cryopreserved, and fresh hepatocytes. Simulations and cooling rate measurements confirmed an adequate concentration of the intracellular CPA concentration (up to 8.53 M) after dehydration in combination with high cooling rates (960-1320 °C/min) for successful vitrification. In comparison to cryopreserved hepatocytes, bulk droplet vitrified hepatocytes had a significantly higher viability, directly after preservation and after 1 day in culture. Moreover, bulk droplet vitrified hepatocytes had evidently better morphology and showed significantly higher metabolic activity than cryopreserved hepatocytes in long-term collagen sandwich cultures. In conclusion, we developed a novel bulk droplet vitrification method of which we validated the theoretical background and demonstrated the feasibility to use this method to vitrify large cell volumes. Moreover, we showed that this method results in improved hepatocyte viability and metabolic function as compared to cryopreservation.
Collapse
Affiliation(s)
- Reinier J. de Vries
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
- Department of Surgery, University of Amsterdam, Amsterdam, the Netherlands
| | - Peony D. Banik
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| | - Sonal Nagpal
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| | - Lindong Weng
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| | - Sinan Ozer
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| | | | - Mehmet Toner
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| | - Shannon N. Tessier
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| | - Korkut Uygun
- Department of Surgery, Massachusetts General Hospital, Boston, MA, USA
- Center for Engineering in Medicine, Harvard Medical School, Boston MA, USA
| |
Collapse
|
|
5
|
Raju R, Chau D, Notelaers T, Myers CL, Verfaillie CM, Hu WS. In Vitro Pluripotent Stem Cell Differentiation to Hepatocyte Ceases Further Maturation at an Equivalent Stage of E15 in Mouse Embryonic Liver Development. Stem Cells Dev 2018; 27:910-921. [PMID: 29851366 DOI: 10.1089/scd.2017.0270] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Hepatocyte-like cells (HLCs) can be derived from pluripotent stem cells (PSCs) by sequential treatment of chemical cues to mimic the microenvironment of embryonic liver development. However, these HLCs do not reach the full maturity level of primary hepatocytes. In this study, we carried out a meta-analysis of cross-species transcriptome data of in vitro differentiation of human PSCs to HLCs and in vivo mouse embryonic liver development to identify the developmental stage at which HLC maturation was blocked at. Systematic variations were found associated with the data source and removed by batch correction. Using principal component analysis, HLCs from different stages of differentiation were aligned with mouse embryonic liver development chronologically. A "unified developmental time" (DT) scale was developed after aligning in vitro HLC differentiation and in vivo embryonic liver development. HLCs were found to cease further maturation at an equivalent stage of mouse embryonic day (E)13-15. Genes with discordant time dynamics were identified by aligning in vivo and in vitro data set onto a common DT scale. These genes may be targets of genetic intervention for enhancing the maturity of PSC-derived HLCs.
Collapse
Affiliation(s)
- Ravali Raju
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota
| | - David Chau
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota.,3 Department of Biomedical Engineering, University of Minnesota , Minneapolis, Minnesota
| | - Tineke Notelaers
- 4 Department of Development and Regeneration, KU Leuven , Leuven, Belgium .,5 Stem Cell Institute Leuven , KU Leuven, Leuven, Belgium
| | - Chad L Myers
- 6 Department of Computer Science and Engineering, University of Minnesota , Minneapolis, Minnesota
| | - Catherine M Verfaillie
- 4 Department of Development and Regeneration, KU Leuven , Leuven, Belgium .,5 Stem Cell Institute Leuven , KU Leuven, Leuven, Belgium
| | - Wei-Shou Hu
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota
| |
Collapse
|
|
6
|
Hegab MH, Abd-Allah SH, Badawey MS, Saleh AA, Metwally AS, Fathy GM, Nada SM, Abdel-Rahman SA, Saleh AA, Fawzy A, El-Magd MA. Therapeutic potential effect of bone marrow-derived mesenchymal stem cells on chronic liver disease in murine Schistosomiasis Mansoni. J Parasit Dis 2018; 42:277-286. [PMID: 29844633 DOI: 10.1007/s12639-018-0997-8] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2018] [Accepted: 04/11/2018] [Indexed: 12/11/2022] Open
Abstract
Some reports have shown that mesenchymal stem cells (MSCs) therapy could ameliorate chemically-induced hepatic fibrosis. This research assesses the therapeutic action of bone marrow mesenchymal stem cells (BM-MSCs) on chronic diseased liver in Schistosoma mansoni infected mice. All infected female mice divided into three groups, one group (15 mice) treated with oral praziquantel (PZQ), second group (15 mice) received intravenous injection of BM-MSCs and third group (15 mice) treated with both MSCs + PZQ. Two control groups (15 mice each) subdivided into one infected and second healthy one. BM-MSCs were obtained from bones of both femur and tibia of male mice (30 mice), then cultured and characterized morphologically by detection of CD105 by flow cytometer. Liver tissues for all groups were examined histopathologically. Measuring of the collagen 1 gene expression was done by real-time PCR and immunohistochemical study to detect stem cells differentiation for detection of MSCs engraftments in liver tissue. MSCs treatment caused marked improvement and regression of fibrosis, and prevents deposition of collagen and reduced the expression of collagen 1 gene in infected mice on their liver tissues, especially when used with PZQ in mice treatment. It can be concluded that, MSCs is a good therapeutic method for liver fibrosis caused by S. mansoni infection.
Collapse
Affiliation(s)
- Mohamed H Hegab
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Somia H Abd-Allah
- 2Department of Biochemistry, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Maha S Badawey
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Ayman A Saleh
- 3Department of Animal Wealth Development, Genetics & Genetic Engineering, of Veterinary Medicine, Zagazig University, Zagazig, Egypt
| | - Ashraf S Metwally
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Ghada M Fathy
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Soad M Nada
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Sara A Abdel-Rahman
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Amira A Saleh
- 1Department of Parasitology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Amal Fawzy
- 2Department of Biochemistry, Faculty of Medicine, Zagazig University, Zagazig, Egypt
| | - Mohammed Abu El-Magd
- 4Department of Anatomy & Embryology, Faculty of Veterinary Medicine, Kafrelsheikh University, Kafrelsheikh, Egypt
| |
Collapse
|
|
7
|
Saheli M, Sepantafar M, Pournasr B, Farzaneh Z, Vosough M, Piryaei A, Baharvand H. Three-dimensional liver-derived extracellular matrix hydrogel promotes liver organoids function. J Cell Biochem 2018; 119:4320-4333. [PMID: 29247536 DOI: 10.1002/jcb.26622] [Citation(s) in RCA: 96] [Impact Index Per Article: 13.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2017] [Accepted: 12/12/2017] [Indexed: 12/25/2022]
Abstract
An important advantage of employing extracellular matrix (ECM)-derived biomaterials in tissue engineering is the ability to tailor the biochemical and biophysical microenvironment of the cells. This study aims to assess whether three-dimensional (3D) liver-derived ECM hydrogel (LEMgel) promotes physiological function of liver organoids generated by self-organization of human hepatocarcinoma cells together with human mesenchymal and endothelial cells. We have optimized the decellularization method to fabricate liver ECM derived from sheep to preserve the greatest content of glycosaminoglycans, collagen, laminin, and fibronectin in produced LEMgel. During gelation, complex viscoelasticity modulus of the LEMgel (3 mg/mL) increased from 186.7 to 1570.5 Pa and Tan Delta decreased from 0.27 to 0.18. Scanning electron microscopy (SEM) determined that the LEMgel had a pore size of 382 ± 71 µm. Hepatocarcinoma cells in the self-organized liver organoids in 3D LEMgel (LEMgel organoids) showed an epithelial phenotype and expressed ALB, CYP3A4, E-cadherin, and ASGPR. The LEMgel organoid had significant upregulation of transcripts of ALB, CYP3A4, CYP3A7, and TAT as well as downregulation of AFP compared to collagen type I- and hydrogel-free-organoids or organoids in solubilized LEM and 2D culture of hepatocarcinoma cells. Generated 3D LEMgel organoids had significantly more ALB and AAT secretion, urea production, CYP3A4 enzyme activity, and inducibility. In conclusion, 3D LEMgel enhanced the functional activity of self-organized liver organoids compared to traditional 2D, 3D, and collagen gel cultures. Our novel 3D LEMgel organoid could potentially be used in liver tissue engineering, drug discovery, toxicology studies, or bio-artificial liver fabrication.
Collapse
Affiliation(s)
- Mona Saheli
- Department of Biology and Anatomical Sciences, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Mohammadmajid Sepantafar
- Department of Cell Engineering, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.,Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Behshad Pournasr
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Zahra Farzaneh
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Massoud Vosough
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Abbas Piryaei
- Department of Biology and Anatomical Sciences, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.,Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technology in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Hossein Baharvand
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.,Department of Developmental Biology, University of Science and Culture, Tehran, Iran
| |
Collapse
|
|
8
|
Hui H, Ma W, Cui J, Gong M, Wang Y, Zhang Y, He T, Bi Y, He Y. Periodic acid‑Schiff staining method for function detection of liver cells is affected by 2% horse serum in induction medium. Mol Med Rep 2017; 16:8062-8068. [PMID: 28944920 PMCID: PMC5779889 DOI: 10.3892/mmr.2017.7587] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2016] [Accepted: 03/08/2017] [Indexed: 02/06/2023] Open
Abstract
Developing a thorough understanding of experimental methods of hepatic differentiation in hepatic progenitor cells (HPCs) should expand the knowledge of hepatocyte induction in vitro and may help to develop cell transplantation therapies for the clinical usage of HPCs in liver diseases. A previous induction method effectively induced differentiation and metabolic abilities in HPCs. Periodic acid-Schiff (PAS) staining is used to identify glycogen synthesis and hepatocyte function; however, this method failed to detect induced hepatocytes. The present study aimed to investigate the possible factors affecting the previous confusing results of PAS staining. Removal of single induction factors, including dexamethasone, hepatic growth factor and fibroblast growth factor 4 from the induction media did not restore PAS staining, whereas replacement of 2% horse serum (HS) with 10% fetal bovine serum (FBS) significantly increased the number of PAS positive cells. Following 12 days of basal induction, replacing the induction medium with media containing 10% FBS for 12–72 h significantly improved PAS staining, but did not influence indocyanine green uptake. Furthermore, incubation in induction medium with 10% FBS following 12 days of normal induction did not affect the expression of hepatic markers and mature function of HPCs. Therefore, the present study suggested that 2% HS in the induction medium did not affect the hepatic function of induced cells, but did affect glycogen storage, whereas replacement of medium with 10% FBS in advance of PAS staining may restore the failure of PAS staining in low serum concentrations of induced hepatocytes.
Collapse
Affiliation(s)
- Hui Hui
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Wenjun Ma
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Jiejie Cui
- Stem Cell Biology and Therapy Laboratory, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Mengjia Gong
- Stem Cell Biology and Therapy Laboratory, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Yi Wang
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Yuanyuan Zhang
- Stem Cell Biology and Therapy Laboratory, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Tongchuan He
- Stem Cell Biology and Therapy Laboratory, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Yang Bi
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Yun He
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| |
Collapse
|
|
9
|
Asai A, Aihara E, Watson C, Mourya R, Mizuochi T, Shivakumar P, Phelan K, Mayhew C, Helmrath M, Takebe T, Wells J, Bezerra JA. Paracrine signals regulate human liver organoid maturation from induced pluripotent stem cells. Development 2017; 144:1056-1064. [PMID: 28275009 DOI: 10.1242/dev.142794] [Citation(s) in RCA: 98] [Impact Index Per Article: 12.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2016] [Accepted: 02/01/2017] [Indexed: 12/17/2022]
Abstract
A self-organizing organoid model provides a new approach to study the mechanism of human liver organogenesis. Previous animal models documented that simultaneous paracrine signaling and cell-to-cell surface contact regulate hepatocyte differentiation. To dissect the relative contributions of the paracrine effects, we first established a liver organoid using human induced pluripotent stem cells (iPSCs), mesenchymal stem cells (MSCs) and human umbilical vein endothelial cells (HUVECs) as previously reported. Time-lapse imaging showed that hepatic-specified endoderm iPSCs (HE-iPSCs) self-assembled into three-dimensional organoids, resulting in hepatic gene induction. Progressive differentiation was demonstrated by hepatic protein production after in vivo organoid transplantation. To assess the paracrine contributions, we employed a Transwell system in which HE-iPSCs were separately co-cultured with MSCs and/or HUVECs. Although the three-dimensional structure did not form, their soluble factors induced a hepatocyte-like phenotype in HE-iPSCs, resulting in the expression of bile salt export pump. In conclusion, the mesoderm-derived paracrine signals promote hepatocyte maturation in liver organoids, but organoid self-organization requires cell-to-cell surface contact. Our in vitro model demonstrates a novel approach to identify developmental paracrine signals regulating the differentiation of human hepatocytes.
Collapse
Affiliation(s)
- Akihiro Asai
- Pediatric Liver Care Center, Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Eitaro Aihara
- Department of Molecular and Cellular Physiology, University of Cincinnati, Cincinnati, OH 45229, USA
| | - Carey Watson
- Division of Pediatric Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Reena Mourya
- Pediatric Liver Care Center, Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Tatsuki Mizuochi
- Pediatric Liver Care Center, Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Pranavkumar Shivakumar
- Pediatric Liver Care Center, Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Kieran Phelan
- Pediatric Liver Care Center, Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Christopher Mayhew
- Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Michael Helmrath
- Division of Pediatric Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Takanori Takebe
- Department of Regenerative Medicine, Yokohama City University, Yokohama, Kanagawa 236-0004, Japan
| | - James Wells
- Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| | - Jorge A Bezerra
- Pediatric Liver Care Center, Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
| |
Collapse
|
|
10
|
Kholodenko IV, Kholodenko RV, Manukyan GV, Yarygin KN. [The hepatic differentiation of adult and fetal liver stromal cells in vitro]. BIOMEDIT︠S︡INSKAI︠A︡ KHIMII︠A︡ 2017; 62:674-682. [PMID: 28026812 DOI: 10.18097/pbmc20166206674] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
The liver has a marked capacity for regeneration. In most cases the liver regeneration is determined by hepatocytes. The regenerative capacity of hepatocytes is significantly reduced in acute or chronic damage. In particular, repair mechanisms are not activated in patients with alcoholic cirrhosis. Organ transplantation or advanced methods of regenerative medicine can help such patients. The promising results were obtained in clinical trials involving patients with various forms of liver disease who received transplantation of autologous bone marrow stem cells. However, to improve the effectiveness of such treatment it is necessary to search for more optimal sources of progenitor cells, as well as to evaluate the possibility of using descendants of these cells differentiated in vitro. In this study we isolated stromal cells from the liver biopsies of three patients with alcoholic cirrhosis, conducted their morphological and phenotypic analysis, and evaluated the hepatic potential of these cells in vitro. The stromal cells isolated from fetal liver were used for comparison. The results of this can serve as a basis for the development of a new method for the treatment of end-stage liver disease. The stromal cells isolated from the liver biopsies for a long time proliferate in a culture and this which makes it possible to expand them to large amounts for subsequent differentiation into hepatocyte-like cells and autologous transplantation.
Collapse
Affiliation(s)
| | - R V Kholodenko
- Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia
| | - G V Manukyan
- Russian National Research Center of Surgery, Moscow, Russia
| | - K N Yarygin
- Institute of Biomedical Chemistry, Moscow, Russia
| |
Collapse
|
|
11
|
Xiang Y, Pang BY, Zhang Y, Xie QL, Zhu Y, Leng AJ, Lu LQ, Chen HL. Effect of Yi Guan Jian decoction on differentiation of bone marrow mesenchymalstem cells into hepatocyte-like cells in dimethylnitrosamine-induced liver cirrhosis in mice. Mol Med Rep 2016; 15:613-626. [PMID: 28035356 PMCID: PMC5364852 DOI: 10.3892/mmr.2016.6083] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2015] [Accepted: 11/08/2016] [Indexed: 12/14/2022] Open
Abstract
Yi Guan Jian decoction (YGD) may induce the differentiation of bone marrow mesenchymal stem cells (BMSCs) into hepatocyte-like cells (HLCs); however, the underlying mechanisms remain to be elucidated. The present study aimed to investigate this process. To do this, a dimethylnitrosamine (DMN)-induced liver cirrhosis mouse model was established. The mice from the model group were randomly divided into three subgroups: i) Negative control, ii) hepatocyte growth factor and iii) YGD. The overall health, liver function and histological alterations were monitored. The expression of α‑smooth muscle actin (α‑SMA), C‑X‑C chemokine receptor type 4 (CXCR4), extracellular signal‑regulated kinase (ERK1/2), nuclear factor κB p65 subunit (NF‑κB p65) and β‑catenin were measured by immunohistochemistry, western blotting and reverse transcription‑quantitative polymerase chain reaction. Following administration of DMN, the overall health of the mice significantly decreased, with an increase in pathological developments and liver damage resulting in a decrease in liver function. Immunohistochemistry revealed that the expression of α‑SMA, CXCR4, ERK1/2, NF‑κB p65 and β‑catenin was upregulated. Following treatment with YGD, the overall health, liver function and pathology improved. The mRNA and protein expression levels of CXCR4 and ERK1/2 were upregulated, where as α‑SMA, NF‑κB p65 and β‑catenin levels were downregulated. The results demonstrated that YGD may induce the differentiation of BMSCs into HLCs to reverse DMN‑induced liver cirrhosis; this may be achieved via an upregulation of the SDF‑1/CXCR4 axis to activate the mitogen activated protein kinase/ERK1/2 signaling pathway.
Collapse
Affiliation(s)
- Yan Xiang
- Department of Infectious Diseases, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Bing-Yao Pang
- Department of Infectious Diseases, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Yuan Zhang
- Department of Infectious Diseases, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Qiao-Ling Xie
- Department of Infectious Diseases, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Ying Zhu
- Department of Infectious Diseases, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Ai-Jing Leng
- Department of Pharmacy, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Long-Qing Lu
- Department of Infectious Diseases, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| | - Hai-Long Chen
- Department of General Surgery, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China
| |
Collapse
|
|
12
|
Alternative Cell Sources to Adult Hepatocytes for Hepatic Cell Therapy. Methods Mol Biol 2016; 1506:17-42. [PMID: 27830543 DOI: 10.1007/978-1-4939-6506-9_2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/17/2023]
Abstract
Adult hepatocyte transplantation is limited by scarce availability of suitable donor liver tissue for hepatocyte isolation. New cell-based therapies are being developed to supplement whole-organ liver transplantation, to reduce the waiting-list mortality rate, and to obtain more sustained and significant metabolic correction. Fetal livers and unsuitable neonatal livers for organ transplantation have been proposed as potential useful sources of hepatic cells for cell therapy. However, the major challenge is to use alternative cell sources for transplantation that can be derived from reproducible methods. Different types of stem cells with hepatic differentiation potential are eligible for generating large numbers of functional hepatocytes for liver cell therapy to treat degenerative disorders, inborn hepatic metabolic diseases, and organ failure. Clinical trials are designed to fully establish the safety profile of such therapies and to define target patient groups and standardized protocols.
Collapse
|
|
13
|
Raju R, Chau D, Cho DS, Park Y, Verfaillie CM, Hu WS. Cell Expansion During Directed Differentiation of Stem Cells Toward the Hepatic Lineage. Stem Cells Dev 2016; 26:274-284. [PMID: 27806669 DOI: 10.1089/scd.2016.0119] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
The differentiation of human pluripotent stem cells toward the hepatocyte lineage can potentially provide an unlimited source of functional hepatocytes for transplantation and extracorporeal bioartificial liver applications. It is anticipated that the quantities of cells needed for these applications will be in the order of 109-1010 cells, because of the size of the liver. An ideal differentiation protocol would be to enable directed differentiation to the hepatocyte lineage with simultaneous cell expansion. We introduced a cell expansion stage after the commitment of human embryonic stem cells to the endodermal lineage, to allow for at least an eightfold increase in cell number, with continuation of cell maturation toward the hepatocyte lineage. The progressive changes in the transcriptome were measured by expression array, and the expression dynamics of certain lineage markers was measured by mass cytometry during the differentiation and expansion process. The findings revealed that while cells were expanding they were also capable of progressing in their differentiation toward the hepatocyte lineage. In addition, our transcriptome, protein and functional studies, including albumin secretion, drug-induced CYP450 expression and urea production, all indicated that the hepatocyte-like cells obtained with or without cell expansion are very similar. This method of simultaneous cell expansion and hepatocyte differentiation should facilitate obtaining large quantities of cells for liver cell applications.
Collapse
Affiliation(s)
- Ravali Raju
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota
| | - David Chau
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota.,3 Department of Biomedical Engineering, University of Minnesota , Minneapolis, Minnesota
| | - Dong Seong Cho
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota
| | - Yonsil Park
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota
| | - Catherine M Verfaillie
- 4 Department of Development and Regeneration, Stem Cell Institute Leuven , KU Leuven, Leuven, Belgium
| | - Wei-Shou Hu
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota.,2 Stem Cell Institute, University of Minnesota , Minneapolis, Minnesota
| |
Collapse
|
|
14
|
Alizadeh E, Eslaminejad MB, Akbarzadeh A, Sadeghi Z, Abasi M, Herizchi R, Zarghami N. Upregulation of MiR-122 via Trichostatin A Treatments in Hepatocyte-like Cells Derived from Mesenchymal Stem Cells. Chem Biol Drug Des 2015; 87:296-305. [PMID: 26360933 DOI: 10.1111/cbdd.12664] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/20/2015] [Revised: 08/29/2015] [Accepted: 08/31/2015] [Indexed: 12/13/2022]
Abstract
The miR-122 is a tissue-specific miRNA; its expression is abundant in liver. MiR-122 upregulation is crucial for differentiation, functionality, and maintenance of differentiated phenotype in hepatocytes. The improving effects of trichostatin A (TSA) on hepatic differentiation have been reported previously. The aim of this study was to determine whether TSA can affect the expression of miR-122 in hepatocyte-like cells (HLCs) generated from human adipose tissue-derived mesenchymal stem cells (hAT-MSCs). The hepatic differentiation of hAT-MSCs induced by a mixture of growth factors and cytokines either with or without TSA treatments. The functionality of HLCs generated with or without TSA and the expression levels of miR-122 were studied. The expression levels of miR-122 in TSA-treated HLCs was significantly (p < 0.05) higher than those generated by growth factors and cytokines, only. The downregulation of a-fetoprotein (AFP) levels but enhanced albumin synthesis (p < 0.05) and upregulation of liver-enriched transcription factors (LETFs) HNF4α (hepatocyte nuclear factor 4α) and HNF6 (hepatocyte nuclear factor 6) were observed in TSA-treated HLCs (p < 0.05). In conclusion, administration of TSA in hepatogenic differentiation of hAT-MSCs resulted in higher expression levels of miR-122, facilitation of differentiation, and subsequently attenuation of AFP levels.
Collapse
Affiliation(s)
- Effat Alizadeh
- Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran
| | - MohamadReza Baghaban Eslaminejad
- Department of Stem Cells and Developmental Biology at Cell Sciences Research Center, Royan Institute for Stem Cell Biology and Technology, ACER, Royan Institute, Tehran, I.R. Iran
| | - Abolfazl Akbarzadeh
- Department of Medical Nanotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran
| | - Zohre Sadeghi
- Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran
| | - Mozghan Abasi
- Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran
| | - Roya Herizchi
- Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran
| | - Nosratollah Zarghami
- Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran.,The Umbilical Cord Stem Cell Research Center (UCSRC), Tabriz University of Medical Sciences, Golgasht Ave., Tabriz 3137851656, I.R. Iran
| |
Collapse
|
|
15
|
He Y, Cui J, He T, Bi Y. 5-azacytidine promotes terminal differentiation of hepatic progenitor cells. Mol Med Rep 2015; 12:2872-8. [PMID: 25975647 DOI: 10.3892/mmr.2015.3772] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2014] [Accepted: 03/24/2015] [Indexed: 11/06/2022] Open
Abstract
5-azacytidine (5-azaC) is known to induce cardiomyocyte differentiation. However, its function in hepatocyte differentiation is unclear. The present study investigated the in vitro capability of 5-azaC to promote maturation and differentiation of mouse embryonic hepatic progenitor cells, with the aim of developing an approach for improving hepatic differentiation. Mouse embryonic hepatic progenitor cells (HP14.5 cells) were treated with 5-azaC at concentrations from 0 to 20 μmol/l, in addition to hepatocyte induction culture medium. Hepatocyte induction medium induces HP14.5 cell differentiation. 5-azaC may enhance the albumin promotor-driven Gaussia luciferase (ALB-GLuc) activity in induced HP14.5 cells. In the present study 2 μmol/l was found to be the optimum concentration with which to achieve this. The expression of hepatocyte-associated factors was not significantly different between the group treated with 5-azaC alone and the control group. The mRNA levels of ALB; cytokeratin 18 (CK18); tyrosine aminotransferase (TAT); and cytochrome p450, family 1, member A1 (CYP1A1); in addition to the protein levels of ALB, CK18 and uridine diphosphate glucuronyltransferase 1A (UGT1A) in the induced group with 5-azaC, were higher than those in the induced group without 5-azaC, although no significant differences were detected in expression of the hepatic stem cell markers, DLK and α-fetoprotein, between the two groups. Treatment with 5-azaC alone did not affect glycogen synthesis or indocyanine green (ICG) metabolic function in HP14.5 cells, although it significantly increased ICG uptake and periodic acid-Schiff-positive cell numbers amongst HP14.5 cells. Therefore, the present study demonstrated that treatment with 5-azaC alone exerted no effects on the maturation and differentiation of HP14.5 cells. However, 5-azaC exhibited a synergistic effect on the terminal differentiation of induced hepatic progenitor cells in association with a hepatic induction medium.
Collapse
Affiliation(s)
- Yun He
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Jiejie Cui
- Stem Cell Biology and Therapy Laboratory, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Tongchuan He
- Stem Cell Biology and Therapy Laboratory, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Yang Bi
- Department of Pediatric Surgery, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| |
Collapse
|
|
16
|
Tonsil-derived mesenchymal stem cells ameliorate CCl4-induced liver fibrosis in mice via autophagy activation. Sci Rep 2015; 5:8616. [PMID: 25722117 PMCID: PMC4342568 DOI: 10.1038/srep08616] [Citation(s) in RCA: 91] [Impact Index Per Article: 9.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2014] [Accepted: 01/28/2015] [Indexed: 12/24/2022] Open
Abstract
Liver transplantation is the treatment of choice for chronic liver failure, although it is complicated by donor shortage, surgery-related complications, and immunological rejection. Cell transplantation is an alternative, minimally invasive treatment option with potentially fewer complications. We used human palatine tonsil as a novel source of mesenchymal stem cells (T-MSCs) and examined their ability to differentiate into hepatocyte-like cells in vivo and in vitro. Carbon tetrachloride (CCl4) mouse model was used to investigate the ability of T-MSCs to home to the site of liver injury. T-MSCs were only detected in the damaged liver, suggesting that they are disease-responsive. Differentiation of T-MSCs into hepatocyte-like cells was confirmed in vitro as determined by expression of hepatocyte markers. Next, we showed resolution of liver fibrosis by T-MSCs via reduction of TGF-β expression and collagen deposition in the liver. We hypothesized that autophagy activation was a possible mechanism for T-MSC-mediated liver recovery. In this report, we demonstrate for the first time that T-MSCs can differentiate into hepatocyte-like cells and ameliorate liver fibrosis via autophagy activation and down-regulation of TGF-β. These findings suggest that T-MSCs could be used as a novel source for stem cell therapy targeting liver diseases.
Collapse
|
|
17
|
Zhao S, Wang Y, Gao C, Zhang J, Bao H, Wang Z, Gong P. Superparamagnetic iron oxide magnetic nanomaterial-labeled bone marrow mesenchymal stem cells for rat liver repair after hepatectomy. J Surg Res 2014; 191:290-301. [DOI: 10.1016/j.jss.2014.03.064] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2013] [Revised: 03/18/2014] [Accepted: 03/21/2014] [Indexed: 12/14/2022]
|
|
18
|
Li J, Xin J, Zhang L, Wu J, Jiang L, Zhou Q, Li J, Guo J, Cao H, Li L. Human hepatic progenitor cells express hematopoietic cell markers CD45 and CD109. Int J Med Sci 2013; 11:65-79. [PMID: 24396288 PMCID: PMC3880993 DOI: 10.7150/ijms.7426] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/16/2013] [Accepted: 12/11/2013] [Indexed: 01/11/2023] Open
Abstract
OBJECTIVE To clarify the precise characteristics of human hepatic progenitor cells (HPCs) for future cytotherapy in liver diseases. METHODS Hepatic progenitor-like cells were isolated and cultured from the livers of patients who had undergone partial hepatectomy for various pathologies but displayed no sign of hepatic dysfunction. These cells were characterized by transcriptomic profiling, quantitative real-time PCR and immunocyto/histochemistry. RESULTS Cultured HPCs contained polygonal, high nucleus/cytoplasm ratio and exhibited a global gene expression profile similar (67.8%) to that of primary hepatocytes. Among the genes with more than 20-fold higher expression in HPCs were a progenitor marker (CD90), a pentraxin-related gene (PTX3), collagen proteins (COL5A2, COL1A1 and COL4A2), cytokines (EGF and PDGFD), metabolic enzymes (CYBRD1, BCAT1, TIMP2 and PAM), a secreted protein (SPARC) and an endothelial protein C receptor (PROCR). Moreover, eight markers (ALB, AFP, CK8, CK18, CK19, CD90, CD117 and Oval-6) previously described as HPC markers were validated by qRT-PCR and/or immunocyto/histochemistry. Interestingly, human HPCs were also positive for the hematopoietic cell markers CD45 and CD109. Finally, we characterized the localization of HPCs in the canals of Hering and periportal areas with six previously described markers (Oval-6, CK8, CK18, CK19, CD90 and CD117) and two potential markers (CD45 and CD109). CONCLUSION The human HPCs are highly similar to primary hepatocytes in their transcriptional profiles. The CD45 and CD109 markers could potentially be utilized to identify and isolate HPCs for further cytotherapy of liver diseases.
Collapse
Affiliation(s)
- Jun Li
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Jiaojiao Xin
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Liyuan Zhang
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Jian Wu
- 2. Department of Surgery, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Longyan Jiang
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Qian Zhou
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Jun Li
- 3. Department of Pathology, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, China. 310003
| | - Jing Guo
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Hongcui Cao
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| | - Lanjuan Li
- 1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University. 79 Qingchun Rd., Hangzhou, 310003. China
| |
Collapse
|
|
19
|
Subramanian K, Owens DJ, Raju R, Firpo M, O'Brien TD, Verfaillie CM, Hu WS. Spheroid culture for enhanced differentiation of human embryonic stem cells to hepatocyte-like cells. Stem Cells Dev 2013; 23:124-31. [PMID: 24020366 DOI: 10.1089/scd.2013.0097] [Citation(s) in RCA: 64] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Stem cell-derived hepatocyte-like cells hold great potential for the treatment of liver disease and for drug toxicity screening. The success of these applications hinges on the generation of differentiated cells with high liver specific activities. Many protocols have been developed to guide human embryonic stem cells (hESCs) to differentiate to the hepatic lineage. Here we report cultivation of hESCs as three-dimensional aggregates that enhances their differentiation to hepatocyte-like cells. Differentiation was first carried out in monolayer culture for 20 days. Subsequently cells were allowed to self-aggregate into spheroids. Significantly higher expression of liver-specific transcripts and proteins, including Albumin, phosphoenolpyruvate carboxykinase, and asialoglycoprotein receptor 1 was observed. The differentiated phenotype was sustained for more than 2 weeks in the three-dimensional spheroid culture system, significantly longer than in monolayer culture. Cells in spheroids exhibit morphological and ultrastructural characteristics of primary hepatocytes by scanning and transmission electron microscopy in addition to mature functions, such as biliary excretion of metabolic products and cytochrome P450 activities. This three-dimensional spheroid culture system may be appropriate for generating high quality, functional hepatocyte-like cells from ESCs.
Collapse
Affiliation(s)
- Kartik Subramanian
- 1 Department of Chemical Engineering and Materials Science, University of Minnesota , Minneapolis, Minnesota
| | | | | | | | | | | | | |
Collapse
|
|
20
|
The generation of hepatocytes from mesenchymal stem cells and engraftment into the liver. Curr Opin Organ Transplant 2013; 16:69-75. [PMID: 21150616 DOI: 10.1097/mot.0b013e3283424f5b] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
PURPOSE OF REVIEW Liver transplantation is the ultimate therapeutic option for the treatment of end-stage liver diseases, which, however, is restricted by the shortage of donor organs. Instead hepatocyte transplantation seemed to be a way out, but again marginal donor livers for the isolation of primary human hepatocytes are scarce. The hepatocyte differentiation capacity of mesenchymal stem cells might open a new cell resource to generate hepatocyte-like cells for therapeutical use. RECENT FINDINGS Apart from their potency of hepatocyte differentiation mesenchymal stem cells display pleiotropic biological features including modulation of immunogenicity, anti-inflammatory and anti-apoptotic as well as pro-proliferative impact at the site of tissue or organ lesions. They are mobilized from the bone marrow and migrate to the liver along chemoattractive gradients thus contributing to the humoral and cellular response in tissue repair. The cause of different liver diseases is varying depending on, for example, viral, toxic, nutritional, neoplastic challenges. As known from animal studies mesenchymal stem cells seem to have a beneficial impact on liver regeneration and tissue repair under a variety of liver disease conditions. SUMMARY Their versatile biological features render mesenchymal stem cells an alternate cell resource for the treatment of liver diseases. It is important to know the mechanisms of integration of transplanted cells into the recipient tissue and to understand the communication between donor cells and the host tissue on the molecular level in order to support efficacy of cell transplantation and thus optimize the therapeutical outcome.
Collapse
|
|
21
|
He Y, Zhou JW, Xu L, Gong MJ, He TC, Bi Y. Comparison of proliferation and differentiation potential between mouse primary hepatocytes and embryonic hepatic progenitor cells in vitro. Int J Mol Med 2013; 32:476-84. [PMID: 23756629 DOI: 10.3892/ijmm.2013.1413] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2013] [Accepted: 05/22/2013] [Indexed: 11/05/2022] Open
Abstract
Cell therapy may be a novel and effective treatment strategy for liver diseases, replacing liver transplantation. The potential of two alternative cell types (hepatic progenitor/stem cells and mature hepatocytes) has not yet been fully assessed; the issues of low amplification efficiency and recovery function remain to be resolved. In this study, we investigated the proliferation, differentiation and function of primary mouse mature hepatocytes and embryonic hepatic progenitor cells. Primary cells were obtained from the livers of mouse embryos at 14.5 days post coitus [hepatic progenitor 14.5d (HP14.5d) cells], as well as from the livers of 3-month-old mice [liver cells 3m (LC3m)]. Using trypan blue staining and crystal violet staining to detect cell viability, we found that compared with the limited growth capability of primary LC3m cells, primary HP14.5d cells exhibited an active cell proliferation; however, proliferative ability of passaged HP14.5d cells significantly decreased. After the HP14.5d cells were treated in hepatic induction medium, the expression of progenitor cell markers decreased and that of mature hepatic markers increased, to levels similar to those of LC3m cells. On day 12 of induction, the HP14.5d cells showed comparable indocyanine green (ICG) uptake and glycogen storage to that of the LC3m cells. Therefore, our study demonstrates that primary hepatic progenitor cells have a stronger proliferation capacity and differentiation potential, supporting their clinical application in liver cell transplantation.
Collapse
Affiliation(s)
- Yun He
- Department of Pediatric Surgery, Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, Chongqing Stem Cell Therapy Engineering Technical Center, The Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | | | | | | | | | | |
Collapse
|
|
22
|
Ghaedi M, Duan Y, Zern MA, Revzin A. Hepatic differentiation of human embryonic stem cells on growth factor-containing surfaces. J Tissue Eng Regen Med 2012; 8:886-95. [DOI: 10.1002/term.1595] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2011] [Revised: 06/13/2012] [Accepted: 07/19/2012] [Indexed: 12/14/2022]
Affiliation(s)
- Mahboobe Ghaedi
- Department of Biomedical Engineering; University of California at Davis; CA USA
- National Institute for Genetic Engineering and Biotechnology (NIGEB); Tehran Iran
| | - Yuyou Duan
- Department of Medicine, Transplant Research Institute; University of California at Sacramento; CA USA
| | - Mark A. Zern
- Department of Medicine, Transplant Research Institute; University of California at Sacramento; CA USA
| | - Alexander Revzin
- Department of Biomedical Engineering; University of California at Davis; CA USA
| |
Collapse
|
|
23
|
Jorns C, Ellis EC, Nowak G, Fischler B, Nemeth A, Strom SC, Ericzon BG. Hepatocyte transplantation for inherited metabolic diseases of the liver. J Intern Med 2012; 272:201-23. [PMID: 22789058 DOI: 10.1111/j.1365-2796.2012.02574.x] [Citation(s) in RCA: 89] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Inherited metabolic diseases of the liver are characterized by deficiency of a hepatic enzyme or protein often resulting in life-threatening disease. The remaining liver function is usually normal. For most patients, treatment consists of supportive therapy, and the only curative option is liver transplantation. Hepatocyte transplantation is a promising therapy for patients with inherited metabolic liver diseases, which offers a less invasive and fully reversible approach. Procedure-related complications are rare. Here, we review the experience of hepatocyte transplantation for metabolic liver diseases and discuss the major obstacles that need to be overcome to establish hepatocyte transplantation as a reliable treatment option in the clinic.
Collapse
Affiliation(s)
- C Jorns
- Division of Transplantation Surgery, Department of Clinical Science, Intervention and Technology (CLINTEC), Karolinska University Hospital Huddinge, Stockholm, Sweden.
| | | | | | | | | | | | | |
Collapse
|
|
24
|
Zhou WL, Medine CN, Zhu L, Hay DC. Stem cell differentiation and human liver disease. World J Gastroenterol 2012; 18:2018-25. [PMID: 22563188 PMCID: PMC3342599 DOI: 10.3748/wjg.v18.i17.2018] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/02/2011] [Revised: 02/08/2012] [Accepted: 02/26/2012] [Indexed: 02/06/2023] Open
Abstract
Human stem cells are scalable cell populations capable of cellular differentiation. This makes them a very attractive in vitro cellular resource and in theory provides unlimited amounts of primary cells. Such an approach has the potential to improve our understanding of human biology and treating disease. In the future it may be possible to deploy novel stem cell-based approaches to treat human liver diseases. In recent years, efficient hepatic differentiation from human stem cells has been achieved by several research groups including our own. In this review we provide an overview of the field and discuss the future potential and limitations of stem cell technology.
Collapse
|
|
25
|
Christ B, Brückner S. Rodent animal models for surrogate analysis of cell therapy in acute liver failure. Front Physiol 2012; 3:78. [PMID: 22485094 PMCID: PMC3317270 DOI: 10.3389/fphys.2012.00078] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2012] [Accepted: 03/16/2012] [Indexed: 12/27/2022] Open
Abstract
Without therapeutic intervention acute liver failure (ALF) is the consequence of a progredient destruction of the liver parenchyma due to metabolic exhaustion of the hepatocytes. Perivenous hepatocytes are responsible for the detoxification of noxious compounds via the cytochrome P450 enzyme system. Liver transplantation is the only remaining therapeutic option in the end-stage of the disease. Assuming that metabolic capacity could be provided by healthy hepatocytes and thus substitute for the genuine parenchymal cells hepatocyte transplantation since quite some time is considered to be an alternative to whole liver transplantation. While this hypothesis achieved proof-of-concept in animal trials clinical breakthrough is still awaiting success, the reasons of which are ongoing matter of debate. In recent times mesenchymal stem cells (MSC) came into focus as a transplantable cell source to treat ALF. Interestingly, as demonstrated in various rodent animal models their mode of action is rather based on trophic support of hepatocytes remaining in the damaged host parenchyma rather than substitution of tissue loss. Mechanistically, either direct or indirect paracrine effects from the transplanted cells acting pro-proliferative, anti-apoptotic, and anti-inflammatory seem to trigger the regenerative response of the residual healthy hepatocytes in the otherwise lethally injured liver parenchyma. Thus, allogeneic MSC may be the best choice for the treatment of ALF taking advantage of their short-term benefit to sustain the critical phase of the acute insult avoiding long-term immunosuppression.
Collapse
Affiliation(s)
- Bruno Christ
- Applied Molecular Hepatology Laboratory, Department of Visceral, Transplantation, Thoracic and Vascular Surgery, University Hospital Leipzig Leipzig, Germany
| | | |
Collapse
|
|
26
|
Zhang W, Li W, Liu B, Wang P, Li W, Zhang H. Efficient generation of functional hepatocyte-like cells from human fetal hepatic progenitor cells in vitro. J Cell Physiol 2012; 227:2051-8. [PMID: 21751216 DOI: 10.1002/jcp.22934] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Differentiation of human hepatic progenitor cells to functional hepatocytes holds great potential to develop new therapeutic strategies for liver disease and to provide a platform for drug toxicity screens and identification of novel pharmaceuticals. We report here that human fetal hepatic progenitor cells (hFHPCs) efficiently differentiate to hepatocyte-like cells by continuous exposure to a combination of soluble factors for 7 days in vitro. We compared the effect of hepatocyte growth factor (HGF), oncostatin M (OSM), dexamethasone (DEX), or a combination on the expression of a liver-specific marker, albumin (ALB). Real-time RT-PCR analysis showed that, upon exposure to a combination of OSM, DEX, and HGF, the expression of ALB gradually increased in a time-dependent manner. In contrast, the level of the hepatic progenitor cell marker alpha-fetoprotein (AFP) decreased as differentiation progressed. Moreover, cells exposed to the combination of OSM, DEX, and HGF gradually featured highly differentiated hepatic functions, including ALB secretion, glycogen storage, urea production, and cytochrome P450 (CYP) activity. The effect of these factors on the differentiation of hFHPCs may be blocked by U0126, an inhibitor of the ERK1/2 signaling pathway. In conclusion, we demonstrate that a combination of soluble factors facilitates the efficient generation of highly differentiated hepatocyte-like cells from hFHPCs and ERK1/2 signaling pathway involved in this process. Results suggest that this system will be useful for generating functional hepatocytes and, hence, may serve as a cell source suitable for preclinical pharmacological research and testing.
Collapse
Affiliation(s)
- Weitao Zhang
- Department of Cell Biology, Municipal Laboratory for Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China
| | | | | | | | | | | |
Collapse
|
|
27
|
Use of indocyanine green for functional assessment of human hepatocytes for transplantation. Asian J Surg 2012; 35:9-15. [DOI: 10.1016/j.asjsur.2012.04.017] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2011] [Revised: 12/15/2011] [Accepted: 02/24/2012] [Indexed: 12/13/2022] Open
|
|
28
|
Resaz R, Emionite L, Vanni C, Astigiano S, Puppo M, Lavieri R, Segalerba D, Pezzolo A, Bosco MC, Oberto A, Eva C, Chou JY, Varesio L, Barbieri O, Eva A. Treatment of newborn G6pc(-/-) mice with bone marrow-derived myelomonocytes induces liver repair. J Hepatol 2011; 55:1263-71. [PMID: 21703205 PMCID: PMC6541203 DOI: 10.1016/j.jhep.2011.02.033] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/23/2010] [Revised: 02/03/2011] [Accepted: 02/28/2011] [Indexed: 12/16/2022]
Abstract
BACKGROUND & AIMS Several studies have shown that bone marrow-derived committed myelomonocytic cells can repopulate diseased livers by fusing with host hepatocytes and can restore normal liver function. These data suggest that myelomonocyte transplantation could be a promising approach for targeted and well-tolerated cell therapy aimed at liver regeneration. We sought to determine whether bone marrow-derived myelomonocytic cells could be effective for liver reconstitution in newborn mice knock-out for glucose-6-phosphatase-α. METHODS Bone marrow-derived myelomonocytic cells obtained from adult wild type mice were transplanted in newborn knock-out mice. Tissues of control and treated mice were frozen for histochemical analysis, or paraffin-embedded and stained with hematoxylin and eosin for histological examination or analyzed by immunohistochemistry or fluorescent in situ hybridization. RESULTS Histological sections of livers of treated knock-out mice revealed areas of regenerating tissue consisting of hepatocytes of normal appearance and partial recovery of normal architecture as early as 1 week after myelomonocytic cells transplant. FISH analysis with X and Y chromosome paints indicated fusion between infused cells and host hepatocytes. Glucose-6-phosphatase activity was detected in treated mice with improved profiles of liver functional parameters. CONCLUSIONS Our data indicate that bone marrow-derived myelomonocytic cell transplant may represent an effective way to achieve liver reconstitution of highly degenerated livers in newborn animals.
Collapse
Affiliation(s)
- Roberta Resaz
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | - Laura Emionite
- Department of Translational Oncology, National Institute for Research on Cancer, Genova, Italy
| | - Cristina Vanni
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | - Simonetta Astigiano
- Department of Translational Oncology, National Institute for Research on Cancer, Genova, Italy
| | - Maura Puppo
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | - Rosa Lavieri
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | - Daniela Segalerba
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | | | - Maria Carla Bosco
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | - Alessandra Oberto
- Neuroscience Institute of the Cavalieri-Ottolenghi Foundation, Orbassano (Torino), Italy
- Neuroscience Institute of Torino (NIT), Torino, Italy
- Department of Anatomy, Pharmacology and Forensic Medicine, Pharmacology Section, University of Torino, Torino, Italy
| | - Carola Eva
- Neuroscience Institute of the Cavalieri-Ottolenghi Foundation, Orbassano (Torino), Italy
- Neuroscience Institute of Torino (NIT), Torino, Italy
- Department of Anatomy, Pharmacology and Forensic Medicine, Pharmacology Section, University of Torino, Torino, Italy
| | - Janice Y. Chou
- Section on Cellular Differentiation, Program on Developmental Endocrinology and Genetics, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-1830, USA
| | - Luigi Varesio
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| | - Ottavia Barbieri
- Department of Translational Oncology, National Institute for Research on Cancer, Genova, Italy
- Department of Experimental Medicine (DIMES), University of Genova, Genova, Italy
| | - Alessandra Eva
- Laboratory of Molecular Biology, G. Gaslini Institute, Genova, Italy
| |
Collapse
|
|
29
|
Tolosa L, Bonora-Centelles A, Teresa Donato M, Pareja E, Negro A, López S, Castell JV, José Gómez-Lechón M. Steatotic liver: a suitable source for the isolation of hepatic progenitor cells. Liver Int 2011; 31:1231-8. [PMID: 22093411 DOI: 10.1111/j.1478-3231.2011.02609.x] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
BACKGROUND Alternative and/or complementary sources of cells such as hepatic progenitor cells (HPC) are under investigation for hepatic cell therapy purposes. Steatotic livers are those most commonly rejected for clinical transplantation and are also unsuitable for good quality hepatocyte isolation. AIM Taken together these two facts, our aim was to investigate whether they could represent a suitable source for the isolation of progenitor cells. METHODS Rats fed for 7 weeks with methionine-choline deficient diets showing proved steatotic signs (i.e. increase in hepatic lipids; macrovesicular steatosis) and steatotic and normal human liver samples were used to study the expression of HPC markers and to isolate these cells. RESULTS In the liver of the steatotic rats there was a significant increase in HPC (known as oval cells in rodents) markers such as Thy-1, epithelial cell adhesion molecule (EpCAM) and OV-6 (2-, 3- and 5-fold increase respectively). Additionally, there was an increase in the yield of isolated oval cells compared to control rats. Similarly, studies using human livers clearly confirmed an increase in the expression of HPC markers in the steatotic tissue and a significant rise in the number of isolated progenitor cells (EpCAM+, Thy-1+, OV-6+) (10, 12 and 11.6 × 10(4) cells/g of tissue respectively). CONCLUSIONS These data suggest that steatotic livers, discarded for orthotopic liver transplantation and hepatocyte isolation, could be a suitable source for large scale isolation of HPC which might be potential candidates in liver cell therapy.
Collapse
Affiliation(s)
- Laia Tolosa
- Unidad de Hepatología Experimental, Centro de Investigación, Hospital La Fe, Valencia, Spain
| | | | | | | | | | | | | | | |
Collapse
|
|
30
|
Xu C, Chen X, Chang C, Wang G, Wang W, Zhang L, Zhu Q, Wang L, Zhang F. Genome-wide expression profiling of hepatic oval cells after partial hepatectomy in rats. Tissue Cell 2011; 43:291-303. [PMID: 21764095 DOI: 10.1016/j.tice.2011.06.001] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2011] [Revised: 06/03/2011] [Accepted: 06/06/2011] [Indexed: 11/15/2022]
Abstract
To examine the changes of biological activities in hepatic oval cells (HOCs) elicited by 70% partial hepatectomy (PH) and understand the relationship between this cell and liver regeneration (LR), this study isolated and obtained the high purity HOCs (≥ 95%) from rat regenerating livers, and then monitored gene expression profiling of rat hepatic oval cells following surgical operation. Results showed that there were LR-related 1059 genes. These genes were grossly classified into three groups using a fold change cut-off threshold of three-fold: up-regulation, down-regulation and up/down regulation. Analyses of gene expression patterns combined with gene functional categorization suggested that genes in the categories "nucleic acid metabolism" and "cell cycle" were dominated by up-regulated expression. Genes in the functional groups "cell metabolism" and "oxidation reduction" were significantly enriched in expression pattern characterized by down-regulation. According to above mentioned results, the synchronized induction of DNA replication and cell proliferation-involved genes suggested that the peak of oval cell proliferation might occur between 30 and 36 h post-PH. The amino acid transformation-involved genes were down-regulated at the early phase of LR, which perhaps trigger the storage of those amino acids essential for protein synthesis. Reduced oxidative-reduction activity at early phase might be related to negative influence of surgical operation on its detoxification capacity. Conclusively, the genome-wide transcriptional analysis of oval cells would contribute to our understanding of the nature of LR at cell level.
Collapse
Affiliation(s)
- Cunshuan Xu
- College of Life Science, Henan Normal University, Xinxiang 453007, China.
| | | | | | | | | | | | | | | | | |
Collapse
|
|
31
|
Trebol Lopez J, Georgiev Hristov T, García-Arranz M, García-Olmo D. Stem Cell Therapy for Digestive Tract Diseases: Current State and Future Perspectives. Stem Cells Dev 2011; 20:1113-29. [DOI: 10.1089/scd.2010.0277] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Affiliation(s)
- Jacobo Trebol Lopez
- General and Digestive Tract Surgery Department, University Hospital “La Paz”, Madrid, Spain
- Cell Therapy Laboratory, Investigation Institute IdiPAZ, University Hospital “La Paz”, Madrid, Spain
| | - Tihomir Georgiev Hristov
- General and Digestive Tract Surgery Department, University Hospital “La Paz”, Madrid, Spain
- Cell Therapy Laboratory, Investigation Institute IdiPAZ, University Hospital “La Paz”, Madrid, Spain
| | - Mariano García-Arranz
- Cell Therapy Laboratory, Investigation Institute IdiPAZ, University Hospital “La Paz”, Madrid, Spain
| | - Damián García-Olmo
- General and Digestive Tract Surgery Department, University Hospital “La Paz”, Madrid, Spain
- Cell Therapy Laboratory, Investigation Institute IdiPAZ, University Hospital “La Paz”, Madrid, Spain
- Surgery Department, Autonomous University of Madrid, Madrid, Spain
| |
Collapse
|
|
32
|
Nussler AK, Zeilinger K, Schyschka L, Ehnert S, Gerlach JC, Yan X, Lee SML, Ilowski M, Thasler WE, Weiss TS. Cell therapeutic options in liver diseases: cell types, medical devices and regulatory issues. JOURNAL OF MATERIALS SCIENCE. MATERIALS IN MEDICINE 2011; 22:1087-1099. [PMID: 21461918 DOI: 10.1007/s10856-011-4306-7] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/15/2011] [Accepted: 03/24/2011] [Indexed: 05/30/2023]
Abstract
Although significant progress has been made in the field of orthotopic liver transplantation, cell-based therapies seem to be a promising alternative to whole-organ transplantation. The reasons are manifold but organ shortage is the main cause for this approach. However, many problems such as the question which cell type should be used or which application site is best for transplantation have been raised. In addition, some clinicians have had success by cultivating liver cells in bioreactors for temporary life support. Besides answering the question which cell type, which injection site or even which culture form should be used for liver support recent international harmonization of legal requirements is needed to be addressed by clinicians, scientists and companies dealing with cellular therapies. We here briefly summarize the possible cell types used to partially or temporarily correct liver diseases, the most recent development of bioreactor technology and important regulatory issues.
Collapse
Affiliation(s)
- Andreas K Nussler
- Department of Traumatology, MRI, Klinikum rechts der Isar, Technische Universität München, Ismaningerstr. 22, 81675, Munich, Germany.
| | | | | | | | | | | | | | | | | | | |
Collapse
|
|
33
|
Abstract
INTRODUCTION Due to a lack of adequate liver donors and post-surgical complications, researchers propose that cell therapy should be an alternative treatment for patients with end-stage liver diseases. DATA SOURCES We performed a literature review on cell-based therapy for liver disorders. AREAS OF AGREEMENT Due to growing numbers of patients on waiting lists for liver transplantation, a substitute treatment strategy is needed for our patients. Cell therapy can save patients who are in life-threatening situations, enabling them to have more time and increase their chances of survival. Pluripotent stem cells can be a good resource for cell-based therapy after the establishment of efficient differentiation protocols in addition to the settlement of ethical and immunological issues. Cell-based therapy will be applicable after the approval of its efficiency via animal model studies. AREAS OF CONTROVERSY Transplanted cells cannot integrate into the recipient liver and lose their functionality after a limited time. The rate of homing and transdifferentiation of transplanted cells into hepatocytes is scant. GROWING POINTS Application of autologous bone marrow mononuclear cells (MNCs), hematopoietic and mesenchymal stem cells (HSCs and MSCs) has improved the general conditions of certain patients. Although this improvement is temporary, new studies have focused on increasing their performance. TIMELY AREAS FOR DEVELOPING RESEARCH: The safety, feasibility and efficacy of applying MNCs, HSCs and MSCs in liver disorders have been proven in clinical trials. Patient-specific cell therapy after the production of induced pluripotent stem cells and new discoveries in somatic cell conversion during transdifferentiation are promising insights.
Collapse
Affiliation(s)
- Massoud Vosough
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | | | | | | |
Collapse
|
|
34
|
Raschzok N, Teichgräber U, Billecke N, Zielinski A, Steinz K, Kammer NN, Morgul MH, Schmeisser S, Adonopoulou MK, Morawietz L, Hiebl B, Schwartlander R, Rüdinger W, Hamm B, Neuhaus P, Sauer IM. Monitoring of Liver Cell Transplantation in a Preclinical Swine Model Using Magnetic Resonance Imaging. CELL MEDICINE 2010; 1:123-35. [PMID: 27004132 DOI: 10.3727/215517910x551053] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Liver cell transplantation (LCT) is a promising treatment approach for certain liver diseases, but clinical implementation requires methods for noninvasive follow-up. Labeling with superparamagnetic iron oxide particles can enable the detection of cells with magnetic resonance imaging (MRI). We investigated the feasibility of monitoring transplanted liver cells by MRI in a preclinical swine model and used this approach to evaluate different routes for cell application. Liver cells were isolated from landrace piglets and labeled with micron-sized iron oxide particles (MPIO) in adhesion. Labeled cells (n = 10), native cells (n = 3), or pure particles (n = 4) were transplanted to minipigs via intraportal infusion into the liver, direct injection into the splenic parenchyma, or intra-arterial infusion to the spleen. Recipients were investigated by repeated 3.0 Tesla MRI and computed tomography angiography up to 8 weeks after transplantation. Labeling with MPIO, which are known to have a strong effect on the magnetic field, enabled noninvasive detection of cell aggregates by MRI. Following intraportal application, which is commonly applied for clinical LCT, MRI was able to visualize the microembolization of transplanted cells in the liver that were not detected by conventional imaging modalities. Cells directly injected into the spleen were retained, whereas cell infusions intra-arterially into the spleen led to translocation and engraftment of transplanted cells in the liver, with significantly fewer microembolisms compared to intraportal application. These findings demonstrate that MRI can be a valuable tool for noninvasive elucidation of cellular processes of LCT and-if clinically applicable MPIO are available-for monitoring of LCT under clinical conditions. Moreover, the results clarify mechanisms relevant for clinical practice of LCT, suggesting that the intra-arterial route to the spleen deserves further evaluation.
Collapse
Affiliation(s)
- Nathanael Raschzok
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Ulf Teichgräber
- † Radiology, Charité-Campus Mitte, Universitätsmedizin Berlin , Berlin , Germany
| | - Nils Billecke
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Anja Zielinski
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Kirsten Steinz
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Nora N Kammer
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Mehmet H Morgul
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin, Berlin, Germany; ‡Visceral, Transplantation, Thorax, and Vascular Surgery, Universitätsklinikum Leipzig, Leipzig, Germany
| | - Sarah Schmeisser
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Michaela K Adonopoulou
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Lars Morawietz
- § Institute of Pathology, Charité-Campus Mitte, Universitätsmedizin Berlin , Berlin , Germany
| | - Bernhard Hiebl
- ¶ Centre for Biomaterial Development and Berlin-Brandenburg Centre for Regenerative Therapies (BCRT), Institute for Polymer Research, GKSS Research Centre Geesthacht GmbH , Teltow , Germany
| | | | | | - Bernd Hamm
- † Radiology, Charité-Campus Mitte, Universitätsmedizin Berlin , Berlin , Germany
| | - Peter Neuhaus
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| | - Igor M Sauer
- General, Visceral, and Transplantation Surgery, Experimental Surgery and Regenerative Medicine, Charité-Campus Virchow, Universitätsmedizin Berlin , Berlin , Germany
| |
Collapse
|
|
35
|
Chun YS, Chaudhari P, Jang YY. Applications of patient-specific induced pluripotent stem cells; focused on disease modeling, drug screening and therapeutic potentials for liver disease. Int J Biol Sci 2010; 6:796-805. [PMID: 21179587 PMCID: PMC3005346 DOI: 10.7150/ijbs.6.796] [Citation(s) in RCA: 73] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2010] [Accepted: 12/13/2010] [Indexed: 01/04/2023] Open
Abstract
The recent advances in the induced pluripotent stem cell (iPSC) research have significantly changed our perspectives on regenerative medicine by providing researchers with a unique tool to derive disease-specific stem cells for study. In this review, we describe the human iPSC generation from developmentally diverse origins (i.e. endoderm-, mesoderm-, and ectoderm- tissue derived human iPSCs) and multistage hepatic differentiation protocols, and discuss both basic and clinical applications of these cells including disease modeling, drug toxicity screening/drug discovery, gene therapy and cell replacement therapy.
Collapse
Affiliation(s)
| | | | - Yoon-Young Jang
- Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| |
Collapse
|
|
36
|
Farzaneh Z, Pournasr B, Ebrahimi M, Aghdami N, Baharvand H. Enhanced functions of human embryonic stem cell-derived hepatocyte-like cells on three-dimensional nanofibrillar surfaces. Stem Cell Rev Rep 2010; 6:601-610. [PMID: 20694582 DOI: 10.1007/s12015-010-9179-5] [Citation(s) in RCA: 46] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
Human embryonic stem cell (hESC)-derived hepatocytes provide a promising unlimited resource for the treatment of liver disease. However, current protocols for the generation of mature and functional hepatocytes are inefficient. Therefore, in order to better differentiate and maintain the function of differentiating hESCs, we have hypothesized that hESCs undergo better differentiation into hepatocyte-like cells (HLCs) when induced on three-dimensional nanofibrillar surfaces. We have demonstrated that, during stepwise differentiation of induction, the markers of hepatic lineage expressed and finally lead to the generation of functional mature cells. In the presence of an ultraweb nanofiber, HLCs produced lower AFP, greater urea, glycogen storage, metabolic PROD activity, uptake of LDL and organic anion ICG, all of which are indicative of the differentiation of HLCs. These results show that topographically treated hESCs at the nano level have a distinct hepatic functionality profile which has implications for cell therapies.
Collapse
Affiliation(s)
- Zahra Farzaneh
- Department of Stem Cells and Developmental Biology, Royan Institute for Stem Cell Biology and Technology, ACECR, P.O. Box 19395-4644, Tehran, Iran
| | | | | | | | | |
Collapse
|
|
37
|
Gennero L, Roos MA, Sperber K, Denysenko T, Bernabei P, Calisti GF, Papotti M, Cappia S, Pagni R, Aimo G, Mengozzi G, Cavallo G, Reguzzi S, Pescarmona GP, Ponzetto A. Pluripotent plasticity of stem cells and liver repopulation. Cell Biochem Funct 2010; 28:178-89. [PMID: 20232487 DOI: 10.1002/cbf.1630] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Different types of stem cells have a role in liver regeneration or fibrous repair during and after several liver diseases. Otherwise, the origin of hepatic and/or extra-hepatic stem cells in reactive liver repopulation is under controversy. The ability of the human body to self-repair and replace the cells and tissues of some organs is often evident. It has been estimated that complete renewal of liver tissue takes place in about a year. Replacement of lost liver tissues is accomplished by proliferation of mature hepatocytes, hepatic oval stem cells differentiation, and sinusoidal cells as support. Hepatic oval cells display a distinct phenotype and have been shown to be a bipotential progenitor of two types of epithelial cells found in the liver, hepatocytes, and bile ductular cells. In gastroenterology and hepatology, the first attempts to translate stem cell basic research into novel therapeutic strategies have been made for the treatment of several disorders, such as inflammatory bowel diseases, diabetes mellitus, celiachy, and acute or chronic hepatopaties. In the future, pluripotent plasticity of stem cells will open a variety of clinical application strategies for the treatment of tissue injuries, degenerated organs. The promise of liver stem cells lie in their potential to provide a continuous and readily available source of liver cells that can be used for gene therapy, cell transplant, bio-artificial liver-assisted devices, drug toxicology testing, and use as an in vitro model to understand the developmental biology of the liver.
Collapse
Affiliation(s)
- Luisa Gennero
- Department of Internal Medicine, University of Turin, Turin, Italy.
| | | | | | | | | | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
38
|
Song Z, Cai J, Liu Y, Zhao D, Yong J, Duo S, Song X, Guo Y, Zhao Y, Qin H, Yin X, Wu C, Che J, Lu S, Ding M, Deng H. Efficient generation of hepatocyte-like cells from human induced pluripotent stem cells. Cell Res 2009; 19:1233-42. [PMID: 19736565 DOI: 10.1038/cr.2009.107] [Citation(s) in RCA: 363] [Impact Index Per Article: 22.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Human induced pluripotent stem (iPS) cells are similar to embryonic stem (ES) cells, and can proliferate intensively and differentiate into a variety of cell types. However, the hepatic differentiation of human iPS cells has not yet been reported. In this report, human iPS cells were induced to differentiate into hepatic cells by a stepwise protocol. The expression of liver cell markers and liver-related functions of the human iPS cell-derived cells were monitored and compared with that of differentiated human ES cells and primary human hepatocytes. Approximately 60% of the differentiated human iPS cells at day 7 expressed hepatic markers alpha fetoprotein and Alb. The differentiated cells at day 21 exhibited liver cell functions including albumin Asecretion, glycogen synthesis, urea production and inducible cytochrome P450 activity. The expression of hepatic markers and liver-related functions of the iPS cell-derived hepatic cells were comparable to that of the human ES cell-derived hepatic cells. These results show that human iPS cells, which are similar to human ES cells, can be efficiently induced to differentiate into hepatocyte-like cells.
Collapse
Affiliation(s)
- Zhihua Song
- The MOE Key Laboratory of Cell Proliferation and Differentiation, Department of cell biology, College of Life Sciences, Box 38, Peking University, Beijing 100871, China
| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |
Collapse
|