Targeted Gene Correction in Osteopetrotic-Induced Pluripotent Stem Cells for the Generation of Functional Osteoclasts.
Autosomal recessive osteopetrosis is a human bone disease mainly caused by TCIRG1 gene mutations that prevent osteoclasts resorbing activity, recapitulated by the oc/oc mouse model. Bone marrow transplantation is the only available treatment, limited by the need for a matched donor. The use of induced pluripotent stem cells (iPSCs) as an unlimited source of autologous cells to generate gene corrected osteoclasts might represent a powerful alternative. We generated iPSCs from oc/oc mice, corrected the mutation using a BAC carrying the entire Tcirg1 gene locus as a template for homologous recombination, and induced hematopoietic differentiation. Similarly to physiologic fetal hematopoiesis, iPSC-derived CD41(+) cells gradually gave rise to CD45(+) cells, which comprised both mature myeloid cells and high proliferative potential colony-forming cells. Finally, we differentiated the gene corrected iPSC-derived myeloid cells into osteoclasts with rescued bone resorbing activity. These results are promising for a future translation into the human clinical setting.
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Tui
T
Neri
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Sharon
S
Muggeo
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Marianna
M
Paulis
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Maria Elena
ME
Caldana
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Laura
L
Crisafulli
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Dario
D
Strina
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Maria Luisa
ML
Focarelli
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Francesca
F
Faggioli
Mouse & Animal Pathology Laboratory, Fondazione Filarete, 20139 Milano, Italy.
Camilla
C
Recordati
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Samantha
S
Scaramuzza
Mouse & Animal Pathology Laboratory, Fondazione Filarete, 20139 Milano, Italy; Department of Veterinary Sciences and Public Health, University of Milan, Via Celoria 10, 20133 Milan, Italy.
Eugenio
E
Scanziani
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Stefano
S
Mantero
Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Chiara
C
Buracchi
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Cristina
C
Sobacchi
San Raffaele Telethon Institute for Gene Therapy (TIGET), San Raffaele Scientific Institute and Vita Salute San Raffaele University, 20132 Milan, Italy.
Angelo
A
Lombardo
San Raffaele Telethon Institute for Gene Therapy (TIGET), San Raffaele Scientific Institute and Vita Salute San Raffaele University, 20132 Milan, Italy.
Luigi
L
Naldini
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Paolo
P
Vezzoni
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy. Electronic address: anna.villa@humanitasresearch.it.
Anna
A
Villa
Milan Unit, Istituto di Ricerca Genetica e Biomedica, CNR, 20138 Milan, Italy; Humanitas Clinical and Research Center, via Manzoni 56, 20089 Rozzano (Mi), Italy.
Francesca
F
Ficara
D000818:Animals; D002454:Cell Differentiation; D002460:Cell Line; D006410:Hematopoiesis; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D022423:Myeloid Cells / Q000166:cytology / Q000378:metabolism; D010010:Osteoclasts / Q000166:cytology* / Q000378:metabolism; D010022:Osteopetrosis / Q000235:genetics / Q000628:therapy*; D052416:Targeted Gene Repair / Q000379:methods*; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics*
C484226:Atp6ap1 protein, mouse; D025262:Vacuolar Proton-Translocating ATPases