| Mechanical unloading (reduced mechanical load) leads to significant bone loss, as evidenced by disuse osteoporosis which is a critical issue for bedridden patients and astronauts. Osteocytes have recently been shown to be the major cell population responsible for mechanotransduction; however, the cellular and molecular mechanisms underlying the response of bone to unloading are just starting to emerge. Wnt/β-catenin signaling plays a key role in controlling bone mass, but a role of this signaling in the bone response to mechanical unloading remains elusive. Sclerostin emerged as a potent inhibitor of bone growth, but the cellular and molecular mechanism of its function remains unclear. Importantly, its expression pattern suggested the involvement of Sclerostin in bone mechanotransduction. In this dissertation, we addressed the mechanism underlying Sclerostin function and the role of Sclerostin and Wnt/β-catenin signaling in the response of bone to mechanical unloading. First we subjected wildtype mice to tail-suspension which simulated the condition of mechanical unloading in the hindlimb. The activity Wnt/β-catenin signaling was decreased in association with upregulation of Sost in response to mechanical unloading. This inspired us to disrupt Sost in vivo through genetic approach. We made a conventional knockout by replacing the Sost coding sequence with a LacZ gene and a PGK-Neo cassette. The Sost null mice were viable and fertile, but radiography and Micro-CT analysis revealed strikingly higher bone mass in Sost null mice compared to their wildtype littermates. Moreover, they exhibited high Bone mineral density (BMD) throughout the life. However, bone development in Sost-/- embryo was normal.In order to explore the molecular mechanism of Sclerostin function, we performed Western blot using total protein and nuclear fraction of total protein extracted from femora of 7-week-old Sost-/- mice and WT littermates. In Sost-/- bones, nuclear accumulation ofβ-catenin was increased, phosphorylatedβ-catenin level (the form targeted for degradation) was decreased, and the level of a Wnt/β-catenin signaling target Connexin 43 was increased. Besides, immunohistochemitry on the section of tibiae consistently showed the nuclear localization ofβ-catenin in bone cells were increased in Sost-/-. Finally, Wnt/β-catenin signaling target Lef-1, was upregulated in Sost-/- bones. Together these results demonstrated the activity of Wnt/β-catenin signaling was enhanced in Sost-/- bones. Meanwhile, the level of phosphorylated SMAD1/5/8 and expression of a BMP signaling target Id1 was not altered in Sost-/- bones. Taken together, we present first in vivo evidence that Sclerostin was an endogenous antagonist to Wnt/β-catenin signaling, rather than to BMP signaling.We next explore the mechanism of Sclerostin function in cellular level. Ratio of Bcl-2/BAX and level of Bcl-xL was elevated Sost-/- long bone, implying decreased apoptosis in bone cells. TUNEL assay showed the apoptosis of osteocytes and osteoblasts were impaired in Sost-/-. In addition, analysis of histological and molecular markers of osteoblasts activity both indicated the activity of Sost-/- osteoblasts were enhanced. But primary osteoblast isolated from Sost-/- calvarium displayed similar function with that from WT calvarium, implying the enhancement in Sost-/- osteoblasts was non-cell-autonomous. Taken together, physiologically Sclerostin secreted from osteocytes suppressed the activity and viability of osteoblasts as well as oteocytes viability.To directly investigate the role of Sclerostin in response of bone to mechanical loading, 17-week-old and 7-week-old female Sost-/- and WT mice were subjected to hindlimb unloading for 2 weeks. Remarkably, while unloading significantly reduced the BMD and bone volume of wildtype mice, the BMD and bone volume were not altered in Sost-/- mice. And other bone microstructure parameters displayed similar results. Thus, Sost-/- mice were resistant to unloading-induced bone loss. In cellular level, while bone formation rate was markedly decreased in WT by unloading, it was not decreased by unloading in Sost-/- mice. In molecular level, Wnt/β-catenin signaling activity, which was decreased in WT, was unchanged in Sost-/- mice. Therefore, Sclerostin was the physiological link between osteocytes sensing mechanical stress and cell signaling in effector cells, and Wnt/β-catenin signaling was a key signaling mediating the effect of unloading.This study presented first in vivo evidence that Sclerostin specifically antagonized Wnt/β-catenin signaling. Moreover, we revealed the essential role of Sclerostin and Wnt/β-catenin signaling in bone response to mechanical unloading. Our findings clearly indicated Sclerostin was a promising target toward disuse osteoporosis.Besides, we identified Sostdc1 gene as a target under Ihh induction. Thus we made Sostdc1 conditional knockout. But mice deficient in Sostdc1 showed no defect in bone, but supernumerary in teeth. We also conducted a case-control study on the association of 5-HTT and schizophrenia in the Chinese Han population. We showed two haplotype of 5-HTT was significantly associated with schizophrenia.
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