Fig 2: Oral Glucose Tolerance Test. Glucose tolerance decreased with Sparc KO and increased with exercise. The ageing also decreased the glucose tolerance only in WT mice (not in Sparc KO mice). The 4-way ANOVA analysis also revealed an interaction (age × genotype × exercise) in which ageing decreased the tolerance only in WT-Ex mice (not WT-Sed mice) and increased the glucose tolerance in KO-Ex mice (not KO-Sed mice). All data are mean ± SEM. The number of mice: 11–12 mice per experimental condition. Abbreviations: AUC, area under the curve; Ex, exercise; KO, knockout; O, old; Sed, sedentary; Sparc, secreted protein acidic and rich in cysteine; WT, wild-type; Y, young.

Fig 3: Total muscle and inguinal adipose tissue weights: (A) Sparc overexpression tended to increase the total skeletal muscles (gastrocnemius, soleus, tibialis anterior, and extensor digitorum longus) weight. Male mice had a heavier muscle weight than female mice. For the inguinal adipose tissue (B), the Sparc overexpression decreased its weight in male mice by 23%. Male mice had more inguinal adipose tissue than female mice. All data are mean ± SEM. The number of mice: 5–11 mice per experimental condition. *: Trend (0.05 ≤ p ≤ 0.1). Abbreviations: g, gram; Sparc, secreted protein acidic and rich in cysteine; Tg, transgenic (Sparc overexpression); WT, wild-type.

Fig 4: Altered expression of ECM proteins in Cyp1b1-/- LSEC.Western blot analysis of ECM proteins in the conditioned medium (CM) and cell lysates from LSEC was performed. (A) The levels of SPARC, Tenascin-C, TSP2, and Osteopontin were below the level of detection. Cyp1b1 LSEC produced TSP1, periostin, and fibronectin. (B) The quantitative assessment of the data. TSP1 level was significantly decreased in lysates from Cyp1b1-/- LSEC (***P< 0.001; n = 3), while periostin level was increased in lysates from Cyp1b1-/- LSEC. The level of fibronectin secreted into the conditioned medium was decreased in Cyp1b1-/- LSEC. These experiments were repeated with two isolation of LSEC with similar results.

Fig 5: Expression of Shh target genes is upregulated in Ptch1cKO astrocytes(A and B) Kir4.1 fluorescence intensity in WT and Ptch1cKO cortex. n = 4 mice for each condition. Scale bar, 100 μm.(C, G, K, and O) RNA-seq fold change of Shh target genes in Ptch1cKO astrocytes compared with WT. n = 4 mice for each condition.(D, H, L, and P) Real-time PCR measurement of Shh target genes in FACS-sorted tdTomato+ astrocytes of WT and Ptch1cKO. n = 4 mice for each condition.(E and F) Density of Lrig1+ cells in WT and Ptch1cKO cortex. WT: n = 5 mice, KO: n = 6 mice. Scale bar, 100 μm.(I and J) Density of Il33+ cells in WT and Ptch1cKO cortex. n = 6 mice for each condition. Scale bar, 100 μm.(M and N) Density of Sparc+ cells in WT and Ptch1cKO layer V cortex. n = 5 mice for each condition. Scale bar, 30 μm.Data represent mean ± SEM; statistical analyses were t test (D, H, L, P) or multiple t test (B, F, J, N). See also Figures S3 and S1’4.