Bone Abstracts

Objectives: Osteogenesis Imperfecta (OI) is a heritable brittle bone disease mainly caused by mutation of COL1A1 or COL1A2. Treatment with bisphosphonate is not effective enough in patients with severe OI. 4-phenylbutyric acid (4-PBA) may become a new medicine, which was reported to ameliorate the phenotype of an OI zebrafish model. In the present study, we aimed to analyze the pathology of OI and the effects of 4-PBA on patient-derived fibroblasts and induced pluripotent stem cells (iPSCs).

Methods: Dermal fibroblasts were obtained from 6 patients with OI: 1 had a nonsense mutation, 2 had a glycine substitution mutation and 3 had an exon skipping mutation in COL1A1 or COL1A2. Endoplasmic reticulum (ER) retention of type I procollagen was observed by immunofluorescent staining. The expression of ER stress markers was quantified by real-time PCR. Protein level and mRNA level of type I collagen were measured by ELISA and real-time PCR, respectively. The molecular weight of secreted type I collagen was analyzed by SDS-PAGE. Post-translational modifications of type I collagen were analyzed by LC-MS. Normal and patient-specific iPSCs were established from fibroblasts, and induced to osteoblasts. Mineralization was assessed by Alizarin Red S staining. We evaluated the effects of 4-PBA on above experiments.

Results: In OI fibroblasts, immunofluorescent staining showed excessive amount of procollagen in ER. 4-PBA addition decreased the retention in a dose-dependent manner. The expression of BIP and CHOP was not changed by 4-PBA. In OI with a glycine substitution, the protein and mRNA levels of type I collagen were increased and canceled by 4-PBA. SDS-PAGE showed the retarded band and LC-MS analysis showed overglycosylation of type I collagen with a glycine substitution. 4-PBA did not change the retarded band, but partially improved overglycosylation. Osteoblasts differentiated from patient-specific iPSCs exhibited less calcification than normal, and 4-PBA improved the capability of calcification to the normal level.

Conclusion: 4-PBA decreased the excessive production and accumulation of type I collagen in OI fibroblasts, and partially improved overglycosylation. In addition, 4-PBA improved mineralization of osteoblasts differentiated from patient-derived iPSCs. Since 4-PBA has already been approved for other disease, we may reposition 4-PBA for OI.

Disclosure: The authors declared no competing interests.