title: Useful properties of undifferentiated mesenchymal stromal cells and adipose tissue as the source in liver-regenerative therapy studied in an animal model of severe acute fulminant hepatitis.
abstract: OBJECTIVE
  End-stage liver diseases frequently require liver transplantation. Cell therapy could be an alternative. This study aimed to analyze whether undifferentiated mesenchymal stromal cells (U-MSCs) or MSC-derived hepatocyte-like cells (DHLCs) from adipose tissue (AT), umbilical cord blood (UCB) and bone marrow (BM) would better restore damaged liver.
  
  
  METHODS
  AT was obtained from lipo-aspiration, UCB from an Umbilical Cord Blood Bank and BM from a BM Transplantation Unit. AT (collagenase digestion), UCB and BM (Ficoll gradient) were cultured (Dulbecco's modified Eagle's medium, low glucose, FBS) for 3 days. Detached adherent cells, at passage 4, were characterized as MSCs. Genetic stability was investigated by means of telomerase enzyme activity and karyotype. Hepatocyte differentiation protocol was performed with the use of Dulbecco's modified Eagle's medium, hepatocyte growth factor, basic fibroblast growth factor and nicotinamide (7 days); maturation medium (oncostatin, dexamethasone, insulin, transferrin and selenium) was added at 36 days. Hepatogenesis analyses were performed by use of morphology and albumin, AF, tyrosine-aminotransferase and glutamine synthetase gene expression and quantitative reverse transcription-polymerase chain reaction on days 9, 18, 25 and 36. Functionality was assessed through glycogen storage detection, indocyanine green absorption and transplantation procedure. U-MSCs and DHLCs were injected 48 h after induced fulminant hepatitis (intraperitoneal injection of carbon tetrachloride) in SCID/BALB-c mice. Histopathologic analyses were performed on days 7 and 15. Human origin included albumin and CK19 human markers.
  
  
  RESULTS
  All MSCs differentiated into functional hepatocyte-like cells, stored glycogen and absorbed indocyanine green. AT-MSC DHLC gene expression was more consistent with a normal hepatogenic-differentiation profile. UCB-MSCs expanded weakly, impairing their use for the transplantation procedure. AT and BM U-MSCs and DHLCs regenerated liver injury equally. Regenerated hepatocytes exhibited human origin.
  
  
  CONCLUSIONS
  AT might be the source and U-MSCS the stem cells useful for liver-regenerative therapy.
authors.affiliation: Umbilical Cord Blood Bank, Hematology Hemotherapy Centre/INCT do Sangue, University of Campinas, São Paulo, Brazil.
authors.forename: Bruna Maria
authors.initials: BM
authors.lastname: Manzini
authors.affiliation: Umbilical Cord Blood Bank, Hematology Hemotherapy Centre/INCT do Sangue, University of Campinas, São Paulo, Brazil.
authors.forename: Adriana
authors.initials: A
authors.lastname: da Silva Santos Duarte
authors.affiliation: Hematology Hemotherapy Centre/INCT do Sangue, University of Campinas, São Paulo, Brazil.
authors.forename: Sundararaj
authors.initials: S
authors.lastname: Sankaramanivel
authors.affiliation: Hematology Hemotherapy Centre/INCT do Sangue, University of Campinas, São Paulo, Brazil.
authors.forename: Aline Lisie
authors.initials: AL
authors.lastname: Ramos
authors.affiliation: Research Center in Pediatrics, Faculty of Medical Sciences, University of Campinas, São Paulo, Brazil.
authors.forename: Paulo
authors.initials: P
authors.lastname: Latuf-Filho
authors.affiliation: Pathology Department, Faculty of Medical Sciences, University of Campinas, São Paulo, Brazil.
authors.forename: Cecilia
authors.initials: C
authors.lastname: Escanhoela
authors.affiliation: Plastic Surgery Department, Faculty of Medical Sciences, University of Campinas, São Paulo, Brazil.
authors.forename: Paulo
authors.initials: P
authors.lastname: Kharmandayan
authors.affiliation: Internal Medicine Department, Faculty of Medical Sciences, Haematology Hemotherapy Centre/INCT do Sangue, University of Campinas, São Paulo, Brazil.
authors.forename: Sara Teresinha
authors.initials: ST
authors.lastname: Olalla Saad
authors.affiliation: Liver Transplantation Unit-Gastroenterology Department, Faculty of Medical Sciences, University of Campinas São Paulo, Brazil.
authors.forename: Ilka
authors.initials: I
authors.lastname: Boin
authors.affiliation: Umbilical Cord Blood Bank, Hematology Hemotherapy Centre/INCT do Sangue, University of Campinas, São Paulo, Brazil; Liver Transplantation Unit-Gastroenterology Department, Faculty of Medical Sciences, University of Campinas São Paulo, Brazil. Electronic address: luzo@unicamp.br.
authors.forename: Ângela Cristina
authors.initials: ÂC
authors.lastname: Malheiros Luzo
mesh_terms: D000273:Adipose Tissue / Q000166:cytology; D000818:Animals; D015415:Biomarkers; D001854:Bone Marrow Cells / Q000166:cytology; D002251:Carbon Tetrachloride; D002454:Cell Differentiation / Q000502:physiology; D064987:Cell- and Tissue-Based Therapy / Q000379:methods*; D004195:Disease Models, Animal; D005312:Fetal Blood / Q000166:cytology; D015870:Gene Expression; D006003:Glycogen / Q000378:metabolism; D006505:Hepatitis / Q000628:therapy*; D017228:Hepatocyte Growth Factor; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism / Q000637:transplantation; D006801:Humans; D017114:Liver Failure, Acute / Q000628:therapy*; D008115:Liver Regeneration / Q000502:physiology*; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology; D051379:Mice; D008807:Mice, Inbred BALB C; D016513:Mice, SCID
chemical_list: D015415:Biomarkers; C066855:HGF protein, human; D017228:Hepatocyte Growth Factor; D006003:Glycogen; D002251:Carbon Tetrachloride
keywords: adipose tissue and umbilical cord blood sources; bone marrow; differentiation; functional hepatocyte; mesenchymal stromal cells