Wnt signaling pathway inhibitors, sclerostin and DKK-1, correlate with pain and bone pathology in patients with Gaucher disease

Abstract

Patients with Gaucher disease (GD) have progressive bone involvement that clinically presents with debilitating bone pain, structural bone changes, bone marrow infiltration (BMI), Erlenmeyer (EM) flask deformity, and osteoporosis. Pain is referred by the majority of GD patients and continues to persist despite the type of therapy. The pain in GD is described as chronic deep penetrating pain; however, sometimes, patients experience severe acute pain. The source of bone pain is mainly debated as nociceptive pain secondary to bone pathology or neuropathic or inflammatory origins. Osteocytes constitute a significant source of secreted molecules that coordinate bone remodeling. Osteocyte markers, sclerostin (SOST) and Dickkopf-1 (DKK-1), inactivate the canonical Wnt signaling pathway and lead to the inhibition of bone formation. Thus, circulated sclerostin and DKK-1 are potential biomarkers of skeletal abnormalities. This study aimed to assess the circulating levels of sclerostin and DKK-1 in patients with GD and their correlation with clinical bone pathology parameters: pain, bone mineral density (BMD), and EM deformity. Thirty-nine patients with GD were classified into cohorts based on the presence and severity of bone manifestations. The serum levels of sclerostin and DKK-1 were quantified by enzyme-linked immunosorbent assays. The highest level of sclerostin was measured in GD patients with pain, BMI, and EM deformity. The multiparameter analysis demonstrated that 95% of GD patients with pain, BMI, and EM deformity had increased levels of sclerostin. The majority of patients with elevated sclerostin also have osteopenia or osteoporosis. Moreover, circulating sclerostin level increase with age, and GD patients have elevated sclerostin levels when compared with healthy control from the same age group. Pearson's linear correlation analysis showed a positive correlation between serum DKK-1 and sclerostin in healthy controls and GD patients with normal bone mineral density. However, the balance between sclerostin and DKK-1 waned in GD patients with osteopenia or osteoporosis. In conclusion, the osteocyte marker, sclerostin, when elevated, is associated with bone pain, BMI, and EM flask deformity in GD patients. The altered sclerostin/DKK-1 ratio correlates with the reduction of bone mineral density. These data confirm that the Wnt signaling pathway plays a role in GD-associated bone disease. Sclerostin and bone pain could be used as biomarkers to assess patients with a high risk of BMI and EM flask deformities.

Trial registration: ClinicalTrials.gov NCT04055831.

Keywords: DKK-1; Gaucher disease; SOST; Wnt; bone; osteoporosis; pain.

Figure 1

Plasma sclerostin concentrations. (A) Sclerostin level (pg/ml), control vs. GD. *p < 0.05 unpaired t-test and F-test. (B) Sclerostin concentrations in control subjects and GD with no bone complication (N), osteopenia (OSN), and osteoporosis (OSR). p < 0.05; ANOVA, Brown–Forsythe, and Bartlett’s multiple comparison tests. Data are means ± SEM. (C) Sclerostin level in female controls and GD patients age-related; cohort divided into two groups before and after 45 years old. *Unpaired t-test p < 0.05. (D, E) Scatterplot analysis of the correlation of sclerostin and age in healthy controls and GD female (D) and male patients (E). Pearson’s two-tailed correlation. (F–H) Sclerostin level in GD patients related to bone pain (F), bone marrow infiltration (G), and EM flask deformity (H). Biomarkers’ normal range is highlighted in green. GD, Gaucher disease; EM, Erlenmeyer.

Figure 2

(A) The Venn diagrams indicate the number of patients with bone pain, bone marrow infiltration, and EM flask deformity in the GD cohort with a normal range of sclerostin level (left) and high sclerostin level (right). Sclerostin level >250 pg/ml was assessed as an elevated level. (B) For GD patients with pain (Pain +) (n = 7, patient P29 described the pain as uncertain) and absent pain (n = 6) who had completed three or four visits, plasma sclerostin levels were measured to assess longitudinal dynamics. Three time points are represented for each participant: initial visit, 6–8 months of follow-up visit (V2), and 12–14 months of follow-up visit (V3). EM, Erlenmeyer; GD, Gaucher disease.

Figure 3

DKK-1 level. (A) DKK-1 level, control vs. GD. (B) DKK-1 concentrations in control subjects and GD with no bone complication (N), osteopenia (OSN), and osteoporosis (OSR). Data are means ± SEM. (C) For GD patients with high DKK-1 levels (n = 4) and normal DKK-1 levels (n = 5) who had completed three visits, DKK-1 was measured to assess longitudinal dynamics. Visits are represented for each participant, including an initial visit, 6–8 months of follow-up visit, and a second 12–14 months of follow-up visit, except for P20 visit 4 (18 months) and P18 visit 5 (24 months). (D) Scatterplot analysis of correlation of DKK-1 and sclerostin in healthy controls and GD patients with no bone complication, including patients without pain (blue circle) and with pain (red circle). *p < 0.05 Pearson’s and Spearman’s tests, 90%, one-tailed. (E) Scatterplot analysis of correlation of DKK-1 and sclerostin in all GD patients with OSN and OSR, including patients without pain (blue color) and with pain (red color). (G) Sclerostin/DKK-1 ratio according to the no bone pain or bone pain groups: healthy control group, all GD patients, GD group NB, GD group OSN, and GD-OSRR group. Data are means ± SEM. *p < 0.05 unpaired t-test. DKK-1 and sclerostin measurement pg/ml. GD, Gaucher disease.

Figure 4

DKK-1 correlates with RANKL in GD patients. (A) Scatterplot analysis of correlation of DKK-1 and RANKL in healthy controls. (B) Scatterplot analysis of correlation of DKK-1 and RANKL in GD patients. 95%, two-tailed. (C) Scatterplot analysis of correlation of DKK-1 and RANKL in GD patients without bone complication (GD-NB). (D, E) Scatterplot analysis of correlation of DKK-1 and RANKL in GD patients with OSN) (E) and OSR (E). (F) Correlation of sclerostin and RANKL in healthy control. (G) Sclerostin and RANKL correlation in GD. DKK-1 and sclerostin measurement pg/ml. GD, Gaucher disease; OSN, osteopenia; OSR, osteoporosis.

Figure 5

A model of inhibition of the Wnt signaling pathway in GD. The balance between bone formation and bone resorption is controlled by Wnt signaling pathway (activation of bone formation), sclerostin and DKK-1 (inhibition of bone formation), and the RANKL/OPG pathway (osteoclast activation). Elevation of secreted sclerostin or DKK-1 leads to inhibition of Wnt signaling pathway in GD. Sclerostin prevents activation of binding of Wnt 3a and LRP on the cellular membrane and, as a result, inhibits the expression of genes that stimulate bone formation, for example, RANKL inhibitor—OPG. RANKL, expressed by pre-osteoblasts and osteoblasts, promotes osteoclast maturation. Activation of osteoclasts initiates bone resorption. Elevated TRAP5b in GD plasma is the biomarker of osteoclast activity and activation of bone resorption. Activation of bone resorption with inhibition of bone formation leads to decreasing bone mineral density in GD. GD, Gaucher disease.