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. Author manuscript; available in PMC: 2024 May 1.
Published in final edited form as: Annu Rev Pathol. 2023 Oct 23;19:507–540. doi: 10.1146/annurev-pathmechdis-051222-121126

Figure 6.

Figure 6

The T238A mouse. (a) Plasma PPi was absent in both Enpp1T238A and Enpp1asj mice, demonstrating abrogation of ENPP1 catalytic activity in both genotypes. (b) Western blot of primary osteocytes derived from calvaria cells confirms comparable ENPP1 protein expression in T238A and WT mice and reduced expression in ASJ mice (normalized to GAPDH). Analysis of skeletal mineralization demonstrated preservation of trabecular (c), and cortical (d) mineralization parameters as well as biomechanical performance (e) in the tibias and femurs of Enpp1T238A mice, as compared with Enpp1asj mice. The data in panels c–e are displayed as change (Δ) relative to WT siblings using box plots to display median values and interquartile range. Superimposed on the box plots in green is the relative mean (bar) and standard error of the mean (whiskers) for each measurement. (f) Calvaria cells derived from WT, Enpp1asj, and Enpp1T238A mice and differentiated into osteocytes in a phosphate source for 21 days exhibit dramatic reductions in calcium phosphate in the Enpp1asj mice (stain, Alizarin Red). (g) Quantitation of mRNA expression of Sfrp1 in calvaria osteoblasts derived from WT mice (black symbols) and Enpp1asj mice (gray symbols) reveals elevations of Sfrp1 at days 7 and 21. Expression normalized to Hprt1. (h) Western blot of Sfrp1 knockout in Enpp1asj calvaria cells demonstrates increased nuclear β-catenin, demonstrating that suppression of β-catenin signaling via SFRP1 accounts for the mineralization defect in ENPP1 deficiency. Values of p are explicitly stated as 0.05 ≥ p ≥ 0.001; *** indicates p < 0.001 and **** indicates p < 0.0001, Student’s unpaired t test (to respective WT sibling pairs). Abbreviations: ASJ, associated with stiff joints; BV/TV, bone volume to total volume fraction; Ct.Th, cortical thickness; ENPP1, ectonucleotide pyrophosphatase/phosphodiesterase 1; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; PPi, pyrophosphate; Tb.N, trabecular number; Tb.Th, trabecular thickness; WT, wild type. Figure adapted from Reference 102.