Figure 4.

Role of VEGFR1 in Cancer Pain following Osteolytic Sarcoma Cell Implantation in the Calcaneus Bone

(A) Western blot analyses for VEGFR1 or VEGFR2 expression in ipsilateral L3–L4 DRG of tumor-bearing mice or sham-treated mice and their quantitative densitometric analysis (n = 3 independent experiments; ^∗p = 0.02; Student’s t test).

(B) Typical examples and quantitative summary of VEGFR1 expression in PGP9.5-positive peripheral nerves overlying bone metastases in tumor-affected or sham hind paw (n = 6 mice/group). Negative controls for immunostaining and H&E-stained sections from same animals (not adjacent sections) are included to judge morphology. epi, epidermis; der, dermis.

(C and D) Whole-mount images (C) or cryosection (D) of a DRG 3 weeks after injection of lentivirions expressing EGFP and shRNA.

(E) Western blot analysis of VEGFR1 expression in L3–L4 DRGs injected ipsilaterally (ipsi) with lentivirions, using contralateral DRGs as control.

(F–H) Tumor-induced mechanical hypersensitivity (F) and tumor-induced hypertrophy and sprouting of epidermal sensory nerves expressing the marker protein PGP9.5 (G and H) in mice injected with lenti-VEGFR1-shRNA as compared with lenti-non targeting shRNA in the DRG (n = 5 mice/group).

^∗p < 0.05 as compared with sham in (B), (H) and compared with basal in (F); ^†p < 0.05 as compared with lenti-non-targeting control; ANOVA followed by post hoc Fisher’s test. Scale bars represent 50 μm in (B), (D), and (G) and 250 μm in (C). Data are presented as mean ± SEM. See also Figure S4.