@misc{rssler_efficient_generation_2021, 
author={Rössler, U.,Hennig, A.,Stelzer, N.,Bose, S.,Kopp, J.,Søe, K.,Cyganek, L.,Zifarelli, G.,Ali, S.,von der Hagen, M.,Strässler, E.,Hahn, G.,Pusch, M.,Stauber, T.,Izsvák, Z.,Gossen, M.,Stachelscheid, H.,Kornak, U.},
title={Efficient generation of osteoclasts from human induced pluripotent stem cells and functional investigations of lethal CLCN7-related osteopetrosis},
year={2021},
howpublished = {journal article},
doi = {https://doi.org/10.1002/jbmr.4322},
abstract = {Human induced pluripotent stem cells (hiPSCs) hold great potential for modeling human diseases and the development of innovative therapeutic approaches. Here, we report on a novel, simplified differentiation method for forming functional osteoclasts from hiPSCs. The three-step protocol starts with embryoid body formation, followed by hematopoietic specification, and finally osteoclast differentiation. We observed continuous production of monocyte-like cells over a period of up to 9 weeks, generating sufficient material for several osteoclast differentiations. The analysis of stage-specific gene and surface marker expression proved mesodermal priming, the presence of monocyte-like cells, and of terminally differentiated multinucleated osteoclasts, able to form resorption pits and trenches on bone and dentine in vitro. In comparison to peripheral blood mononuclear cell (PBMC)-derived osteoclasts hiPSC-derived osteoclasts were larger and contained a higher number of nuclei. Detailed functional studies on the resorption behavior of hiPSC-osteoclasts indicated a trend towards forming more trenches than pits and an increase in pseudoresorption. We used hiPSCs from an autosomal recessive osteopetrosis (ARO) patient (BIHi002-A, ARO hiPSCs) with compound heterozygous missense mutations p.(G292E) and p.(R403Q) in CLCN7, coding for the Cl−/H+-exchanger ClC-7, for functional investigations. The patient's leading clinical feature was a brain malformation due to defective neuronal migration. Mutant ClC-7 displayed residual expression and retained lysosomal co-localization with OSTM1, the gene coding for the osteopetrosis-associated transmembrane protein 1, but only ClC-7 harboring the mutation p.(R403Q) gave strongly reduced ion currents. An increased autophagic flux in spite of unchanged lysosomal pH was evident in undifferentiated ARO hiPSCs. ARO hiPSC-derived osteoclasts showed an increased size compared to hiPSCs of healthy donors. They were not able to resorb bone, underlining a loss-of-function effect of the mutations. In summary, we developed a highly reproducible, straightforward hiPSC-osteoclast differentiation protocol. We demonstrated that osteoclasts differentiated from ARO hiPSCs can be used as a disease model for ARO and potentially also other osteoclast-related diseases. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).},
note = {Online available at: \url{https://doi.org/10.1002/jbmr.4322} (DOI). Rössler, U.; Hennig, A.; Stelzer, N.; Bose, S.; Kopp, J.; Søe, K.; Cyganek, L.; Zifarelli, G.; Ali, S.; von der Hagen, M.; Strässler, E.; Hahn, G.; Pusch, M.; Stauber, T.; Izsvák, Z.; Gossen, M.; Stachelscheid, H.; Kornak, U.: Efficient generation of osteoclasts from human induced pluripotent stem cells and functional investigations of lethal CLCN7-related osteopetrosis. Journal of Bone and Mineral Research. 2021. vol. 36, no. 8, 1621-1635. DOI: 10.1002/jbmr.4322}}