Figure 5. SHP1 inhibition enhances the antigen cross-presentation of DCs.

A, Growth curves of orthotopic B16-GMCSF-OVA (B16GM-OVA) melanomas in C57BL/6 mice treated with vehicle or 50mg/kg of VitE (n=5).

B, Flow cytometry analysis and quantification of CD103⁺CD11c⁺ cross-presenting DCs in B16GM-OVA tumor tissues collected at 16 days post-implantation from vehicle- or VitE-treated C57BL/6 mice (n=5).

C, Flow cytometry analysis and quantification of OVA antigen-specific OVA-tetramer⁺CD8⁺ T (SIINFEKL⁺CD8⁺) cells in tumor tissues as in B above (n=5).

D, Flow cytometric analysis and quantification of H-2K^b-SIINFEKL MFI in OVA-loaded sgCtrl or sgPtpn6-DC2.4 cells cultured in RPMI1640+10%FBS without (medium) or with TCM (n=3).

E, FCM analysis and quantification of H-2K^b-SIINFEKL (%) in BMDCs cultured in medium without or with TCM and treated with the vehicle- or VitE (n=3).

F, Quantification of H-2K^b-SIINFEKL MFI in the vehicle- or VitE-treated sgCtrl-DC2.4 or sgPtpn6-DC2.4 (n=3).

G, Confocal microscopy projections of staining of LysoTracker (red) and LAMP-1 (green) of scrambled siRNA (control) or Ptpn6 siRNA transfected BMDCs.

H, Immunoblot of indicated Rab GTPase proteins in Ptpn6-transfected versus control HEK-293 cells.

I, Immunoblot of Rab34 protein in HEK-293 cells with Ptpn6-overexpression (OE) or Ptpn6-OE-HEK-293 cells with Ptpn6 knockdown (KD).

J, Cytosol and membrane Rab34 levels in siCtrl- and siPtpn6-BMDCs.

K, Immunoblot of Rab34 proteins in BMDCs treated with vehicle (Veh) or VitE for 48h.

L, Cytosol and membrane Rab34 levels in BMDCs pre-treated with vehicle (Veh) or VitE for 48h.

M, CFSE-dilution analysis of OT-1 T cells co-cultured with OVA-pulsed BMDCs with knockdown of Rab34 by siRab34 versus control BMDCs transfected with siCtrl (n=3).

Error bars, s.e.m. Two-sided Student’s t-test (B, C, M). One-way ANOVA with Tukey’s multiple comparison post-test (D, E, F). The statistical significance is defined by P-value: ns, no significant. *P < 0.05, **P < 0.01.