title: Cysteamine treatment restores the in vitro ability to differentiate along the osteoblastic lineage of mesenchymal stromal cells isolated from bone marrow of a cystinotic patient.
abstract: BACKGROUND
  Cystinosis is a rare autosomal recessive disease caused by mutations of the CTNS gene, which encodes for a lysosomal cystine/H(+) symporter. In mice, inactivation of the CTNS gene causes intralysosomal cystine accumulation and progressive organ damage that can be reversed, at least in part, by infusion of mesenchymal stromal cells (MSCs). Little is known on the mesenchymal compartment of cystinotic patients. The aim of the study was to test the phenotypical and functional properties of cystinotic MSCs (Cys-MSCs) isolated from bone marrow (BM) aspirate of a patient with nephropathic cystinosis.
  
  
  METHODS
  Morphology, proliferative capacity (measured as population doublings), immunophenotype (by flow-cytometry) and immunomodulatory properties (as phytohemagglutinin-induced peripheral blood mononuclear cell proliferation) were analyzed. The osteogenic differentiation potential of Cys-MSCs was evaluated by histological staining (alkaline phosphatase activity, Alzarin Red and von Kossa staining) spectrophotometry and Quantitative Reverse Transcriptase Polymerase Chain Reaction for osteigenic markers in the presence and in the absence of cysteamine. Cys-MSCs were compared with those isolated and expanded ex vivo from three healthy donors (HD-MSCs).
  
  
  RESULTS
  Despite a slightly lower proliferative capacity, Cys-MSCs displayed a characteristic spindle-shaped morphology and similar immunephenotype as HD-MSCs. Cys-MSCs and HD-MSCs prevented proliferation of PHA-stimulated allogeneic peripheral blood mononuclear cells to the same extent. After in vitro induction into osteoblasts, Cys-MSCs showed reduced alkaline phosphatase (ALP) activity, calcium depositions and expression of ALP and collagen type 1. When Cys-MSCs were treated in vitro with increasing doses of cysteamine (50-100-200 μM/L) during the differentiation assay, recovery of Cys-MSCs differentiation capacity into osteoblasts was observed. No difference in adipogenic differentiation was found between Cys-MSCs and HD-MSCs.
  
  
  CONCLUSIONS
  Our results indicate that, as compared to HD-MSCs, Cys-MSCs show reduced ability to differentiate into osteoblasts, which can be reverted after cysteamine treatment.
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. antonella.conforti@opbg.net.
authors.forename: Antonella
authors.initials: A
authors.lastname: Conforti
authors.affiliation: Department of Nephrology and Urology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. anna.taranta@opbg.net.
authors.forename: Anna
authors.initials: A
authors.lastname: Taranta
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. simone.biagini@opbg.net.
authors.forename: Simone
authors.initials: S
authors.lastname: Biagini
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. nadia.starc85@gmail.com.|University of Rome, Tor Vergata, Rome. nadia.starc85@gmail.com.
authors.forename: Nadia
authors.initials: N
authors.lastname: Starc
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. angela.pitisci@opbg.net.
authors.forename: Angela
authors.initials: A
authors.lastname: Pitisci
authors.affiliation: Department of Nephrology and Urology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. francesco.bellomo@opbg.net.
authors.forename: Francesco
authors.initials: F
authors.lastname: Bellomo
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. valentina.cirillo@opbg.net.
authors.forename: Valentina
authors.initials: V
authors.lastname: Cirillo
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. franco.locatelli@opbg.net.|University of Pavia, Pavia, Italy. franco.locatelli@opbg.net.
authors.forename: Franco
authors.initials: F
authors.lastname: Locatelli
authors.affiliation: Department of Pediatric Hematology/Oncology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. mebernardo@gmail.com.
authors.forename: Maria Ester
authors.initials: ME
authors.lastname: Bernardo
authors.affiliation: Department of Nephrology and Urology, IRCCS Bambino Gesù Children's Hospital, P.le S. Onofrio, 00165, Rome, Italy. francesco.emma@opbg.net.
authors.forename: Francesco
authors.initials: F
authors.lastname: Emma
mesh_terms: D000293:Adolescent; D001853:Bone Marrow / Q000473:pathology*; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D019070:Cell Lineage; D049109:Cell Proliferation; D002648:Child; D003543:Cysteamine / Q000737:chemistry*; D003554:Cystinosis / Q000235:genetics* / Q000473:pathology*; D006801:Humans; D016130:Immunophenotyping; D007963:Leukocytes, Mononuclear / Q000166:cytology; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D055815:Young Adult
chemical_list: D003543:Cysteamine