Ab69858

The mice were anesthetized by intraperitoneal injection of 10% uratan and 2% chloral hydrate (0.1 mL/10 g). Cardiac perfusion was then performed with 0.9% normal saline and 4% paraformaldehyde solution respectively. The whole brain was excised, paraffin-embedded and sectioned. Four brain sections were taken from each group and stained. The SNc tissue sections of PD mice were baked at 60°C for 1 h, deparaffinized using routine xylene, and hydrated with gradient ethanol, followed by antigen retrieval under high pressure for 2 min. After that, the sections were incubated with the primary rabbit polyclonal antibody to TH (ab112, dilution ratio of 1 : 200, Abcam Inc., Cambridge, UK) or rabbit anti-mouse NPTX2 (ab69858, dilution ratio of 1 : 1000, Abcam Inc., Cambridge, UK) at 37°C for 2 h. The detailed steps were performed according to previously published protocols [44 (link)]. The number of positive cells was counted in 5 high-power randomly selected visual fields from each section.

Total protein was extracted and protein concentration was detected by BCA assay. Western blot analysis was performed as previously described [39 (link)]. EFNB2 (ab69858, Abcam, Cambridge, UK), EPHB4 (ab150545, ab98933, Abcam), LDLR (ab52818, Abcam), β-catenin (ab32572, Abcam), VLDLR (ab203271, Abcam), SCARB1(ab52629, Abcam), STAT3(ab68153, Abcam), p-STAT3(ab267373, Abcam), JAK2(ab108596, Abcam), p-JAK2(ab108596, Abcam), SREBP2(ab30682, Abcam), proliferating cell nuclear antigen (PCNA; Proteintech Group, Inc., Sankt Leon-Rot, Germany) primary antibodies were used. Horseradish peroxidase (HRP)-conjugated Affinipure Goat Anti-Rabbit IgG (H + L) (SA00001-2) and HRP-conjugated Affinipure Goat Anti-Mouse IgG (H + L) (SA00001-1) were obtained from Proteintech Group, Inc (Chicago, US).

Total protein was extracted using radioimmunoprecipitation assay (RIPA) buffer supplemented with 1% protease inhibitors and phosphatase inhibitors. Protein concentration was measured with the bicinchoninic acid (BCA) assay. Western blot analysis was performed as previously described^{12 (link)}. NPTX2 (ab69858, Abcam), FZD6 (ab150545, ab98933, Abcam), β-catenin (ab32572, Abcam), MYC (ab39688, Abcam), cyclin D1 (CCND1; ab16663, Abcam), snail-1 (ab53519, Abcam), N-cadherin (ab76011, Abcam), E-cadherin (ab1416, Abcam), Ki-67, and proliferating cell nuclear antigen (PCNA; Proteintech Group, Inc.) primary antibodies were used. Horseradish peroxidase (HRP)-conjugated Affinipure goat anti-rabbit IgG (H + L) and HRP-conjugated Affinipure goat anti-mouse IgG (H + L) were obtained from Proteintech Group, Inc (Jackson).