N-acetylcysteine treatment ameliorates the skeletal phenotype of a mouse model of diastrophic dysplasia

Abstract

Diastrophic dysplasia (DTD) is a recessive chondrodysplasia caused by mutations in SLC26A2, a cell membrane sulfate-chloride antiporter. Sulfate uptake impairment results in low cytosolic sulfate, leading to cartilage proteoglycan (PG) undersulfation. In this work, we used the dtd mouse model to study the role of N-acetyl-l-cysteine (NAC), a well-known drug with antioxidant properties, as an intracellular sulfate source for macromolecular sulfation. Because of the important pre-natal phase of skeletal development and growth, we administered 30 g/l NAC in the drinking water to pregnant mice to explore a possible transplacental effect on the fetuses. When cartilage PG sulfation was evaluated by high-performance liquid chromatography disaccharide analysis in dtd newborn mice, a marked increase in PG sulfation was observed in newborns from NAC-treated pregnancies when compared with the placebo group. Morphometric studies of the femur, tibia and ilium after skeletal staining with alcian blue and alizarin red indicated a partial rescue of abnormal bone morphology in dtd newborns from treated females, compared with pups from untreated females. The beneficial effect of increased macromolecular sulfation was confirmed by chondrocyte proliferation studies in cryosections of the tibial epiphysis by proliferating cell nuclear antigen immunohistochemistry: the percentage of proliferating cells, significantly reduced in the placebo group, reached normal values in dtd newborns from NAC-treated females. In conclusion, NAC is a useful source of sulfate for macromolecular sulfation in vivo when extracellular sulfate supply is reduced, confirming the potential of therapeutic approaches with thiol compounds to improve skeletal deformity and short stature in human DTD and related disorders.

Skeletal Dysplasias Caused by Sulfation Defects

“…A deep understanding of the molecular mechanisms of these disorders is crucial to ultimately pave the way for innovative therapies. Recently, significant advances in the development of therapeutic approaches for some disorders have been achieved with the aim of alleviating, preventing or modifying the disease progression [70,[98][99][100][101]. This progress has been made thank to deep phenotyping of in vitro and/or in vivo models that has allowed the identification of new molecules or already existing ones through a drug repositioning approach.…”

“…Studies on the growth plate of the animal model demonstrated reduced chondrocyte proliferation due to altered Indian hedgehog (Ihh) signalling [68] and reduced phosphorylation of the pocket protein p130 that inhibits transcription factors of the E2F family leading to the block in the G1 phase of cell cycle progression [69]. Even if sulfate uptake from the extracellular environment is crucial for PG sulfation in chondrocytes, an alternative source of sulfate, namely intracellular thiol compounds, has been reported in the dtd mouse suggesting potential pharmacological approaches to DTD [70]. Recently an over activation of fibroblast growth factor receptor 3 (FGFR3) signalling has been described in a Slc26a2 knock-out mouse, an animal model of ACG1B and AO2.…”

Suppressing UPR-dependent overactivation of FGFR3 signaling ameliorates SLC26A2-deficient chondrodysplasias

“…Thus far, there has been no alternative therapy under investigation but to provide palliative care for viable ACG1B and AO2 newborns [46,47]. Drug intervention during pregnancy has been proved practical and effective to ameliorate the retarded skeletal development and rescue postnatal death in mouse models mimicking human chondrodysplasias [[77], [78], [79]]. To our best knowledge, we conducted the first preclinical study with NVP-BGJ398 in SLC26A2-related conditions.…”

Impaired proteoglycan glycosylation, elevated TGF-β signaling, and abnormal osteoblast differentiation as the basis for bone fragility in a mouse model for gerodermia osteodysplastica

“…For CS and DS disaccharide composition, GAG aliquots were digested with chondroitinase ABC (AMSBIO, Abingdon, UK) (digesting chondroitin sulfate A, B and C) or chondroitinase ACII (Seikagaku Corp., Tokyo, Japan) (digesting chondroitin sulfate A and C). Each digest was labeled with 2-aminoacridone (Thermo Fisher Scientific, MA, USA) and analyzed by reverse phase HPLC as described previously [ 61 ]. The dermatan sulfate (chondroitin sulfate B) fraction was determined as disaccharide fraction undigestable by chondroitinase ACII vs. disaccharides digested by chondroitinase ABC.…”