Antibodies to sclerostin or G-CSF receptor partially eliminate bone or marrow adipocyte loss, respectively, following vertical sleeve gastrectomy

AUTHORS

Ziru Li, Kevin Qiu, Jingtong Zhao, Katrina Granger, Hui Yu, Alfor G. Lewis, Andriy Myronovych, Mouhamadoul H. Toure, Sarah J. Hatsell, Aris N. Economides, Randy J. Seeley, Ormond A. MacDougald

ABSTRACT

Vertical sleeve gastrectomy (VSG), the most utilized bariatric procedure in clinical practice, greatly reduces body weight and improves a variety of metabolic disorders. However, one of its long-term complications is bone loss and increased risk of fracture. Elevated circulating sclerostin (SOST) and granulocyte-colony stimulating factor (G-CSF) concentrations have been considered as potential contributors to VSG-associated bone loss. To test these possibilities, we administrated antibodies to SOST or G-CSF receptor and investigated alterations to bone and marrow niche following VSG. Neutralizing either SOST or G-CSF receptor did not alter beneficial effects of VSG on adiposity and hepatic steatosis, and anti-SOST treatment provided a further improvement to glucose tolerance. SOST antibodies partially reduced trabecular and cortical bone loss following VSG by increasing bone formation, whereas G-CSF receptor antibodies had no effects on bone mass. The expansion in myeloid cellularity and reductions in bone marrow adiposity seen with VSG were partially eliminated by treatment with Anti-G-CSF receptor. Taken together, these experiments demonstrate that antibodies to SOST or G-CSF receptor may act through independent mechanisms to partially block effects of VSG on bone loss or marrow niche cells, respectively.

Targeted postnatal knockout of Sclerostin using a bone-targeted adeno-associated viral vector increases bone anabolism and decreases canalicular density

AUTHORS

Alexandra K. O'Donohue, Ya Xiao, Lucinda R. Lee, Timothy Schofield, Tegan L. Cheng, Craig Munns, Paul A. Baldock, Aaron Schindeler

ABSTRACT

Purpose

The creation of murine gene knockout models to study bone gene functions often requires the resource intensive crossbreeding of Cre transgenic and gene-floxed strains. The developmental versus postnatal roles of genes can be difficult to discern in such models. For example, embryonic deletion of the Sclerostin (Sost) gene establishes a high-bone mass phenotype in neonatal mice that may impact on future bone growth. To generate a postnatal skeletal knockout of Sost in adult mice using a single injection of a bone-targeted recombinant adeno-associated virus (rAAV) vector.

Methods

8-week-old Sostflox/flox mice were injected with saline (control) or a single injection containing 5 × 1011 vg AAV8-Sp7-Cre vector. Ai9 fluorescent Cre reporter mice were dosed in parallel to confirm targeting efficiency. After 6 weeks, detailed bone analysis performed via microCT, biomechanical testing, and bone histology on vertebral and long bone specimens.

Results

The AAV8-Sp7-Cre vector induced widespread persistent recombination in the bone compartment. Regional microCT analyses revealed significant increases in bone with vector treatment. In the L3 vertebrae, Sostflox/flox:AAV-Cre showed a 22 % increase in bone volume and 21 % in trabecular bone fraction compared to controls; this translated to a 17 % increase in compressive strength. In the tibiae, Sostflox/flox:AAV-Cre led to small but statistically significant increases in cortical bone volume and thickness. These were consistent with a 25 % increase in mineral apposition rate, but this did not translate into increased four-point bending strength. Ploton silver nitrate stain on histological sections revealed an unexpected increase in canalicular density associated with Sost ablation.

Conclusion

This report demonstrates a proof-of-concept that the AAV8-Sp7-Cre vector can efficiently produce postnatal skeletal knockout mice using gene-floxed strains. This technology has the potential for broad utility in the bone field with existing conditional lines. These data also confirm an important postnatal role for Sost in regulating bone homeostasis, consistent with prior studies using neutralizing Sclerostin antibodies, and highlights a novel role of Sost in canalicular remodeling.

Reactivation of Bone Lining Cells are Attenuated Over Repeated Anti-sclerostin Antibody Administration

AUTHORS

A Ram Hong, Jae-Yeon Yang, Ji Yeon Lee, Joonho Suh, Yun-Sil Lee, Jung-Eun Kim & Sang Wan Kim

ABSTRACT

Reactivation of bone lining cells (BLCs) is a crucial mechanism governing the anabolic action of anti-sclerostin antibody (Scl-Ab) via modeling-based bone formation; however, it remains unclear whether this reactivation can be attenuated after persistent administration of Scl-Ab. Here, we aimed to investigate the reproducibility of persistent Scl-Ab administration for the reactivation of BLCs, and to elucidate the relationship between the activity of BLCs and serum levels of N-terminal procollagen type I (P1NP) during chronic Scl-Ab administration. We conducted an osteoblast lineage tracing study. Briefly, Dmp1-CreERt2(+):Rosa26R mice were injected with 1 mg of 4-hydroxy-tamoxifen weekly from postnatal weeks four to eight. Mice were treated twice with either vehicle or Scl-Ab (25 mg/kg) at weeks 12, 16, and 20, and were euthanized at weeks 8, 12, 13, 16, 17, 20, and 21 (4–6 mice in each group). After euthanization, the number and thickness of X-gal (+) cells on the periosteum of the femoral bones and the serum levels of P1NP were quantified at each time point. Scl-Ab induced a significant increase in the thickness of X-gal (+) cells on periosteal bone surfaces at postnatal weeks 13 (after 1st dose), 17 (after 2nd dose), and 21 (after 3rd dose) compared to that in vehicle-treated mice (all P < 0.001). In the Scl-Ab group, significant increases in the thickness of labeled cells were observed between weeks 16 and 17 and weeks 20 and 21 (both P < 0.001). The percentage increase in X-gal (+) cell thickness was 108.9% from week 12 to week 13, 54.6% from week 16 to week 17, and 49.2% from week 20 to week 21 in the Scl-Ab group. Although Scl-Ab treatment increased the serum levels of P1NP at postnatal weeks 13 and 17 compared with those at week 12 (P = 0.017 and P = 0.038, respectively), the same was not observed at week 21 (P = 0.296). A significant increase in P1NP levels was observed between weeks 16 and 17 and weeks 20 and 21 in the Scl-Ab group (P = 0.005 and P = 0.007, respectively). The percentage increase in P1NP levels was 141.7% from weeks 12 to 13, 114.8% from weeks 16 to 17, and 99.4% from weeks 20 to 21. Serum P1NP levels were positively correlated with X-gal (+) cell thickness (R2 = 0.732, P < 0.001). Reactivation of BLCs is modestly attenuated, but reproducible, during persistent Scl-Ab administration. Serum P1NP levels appear to be an indicator of the impact of Scl-Ab on the conversion of BLCs into mature osteoblasts on periosteal bone surfaces, thus contributing to modeling-based bone formation.