SHP1 expression and down-regulation of the JAK3/STAT3 pathway. (A) SHP1 expression induced down-regulation of the JAK3/STAT3 signaling pathway in SU-DHL-1 and Karpas 299 cells. SU-DHL1 cells transfected with pIRES2-EGFP or pIRES2-EGFP-SHP1 were sorted using flow cytometry and subjected to Western blot analysis 24 hours after gene transfection. The results showed substantial decreases in the protein expression of pSTAT3, pJAK3, and JAK3. In contrast, there were relatively few changes in the protein level of STAT3 between the 2 samples. Karpas 299 cells were transfected with pCI (empty vector) and pCI-SHP1, and subjected to Western blot analysis 24 hours after gene transfection. Similar to the SU-DHL-1 cells, Karpas 299 cells showed down-regulation of the JAK3/STAT3 signaling. (B) Modulation of STAT3 downstream targets as well as NPM-ALK in Karpas 299 and SU-DHL-1 cells after SHP1 gene transfection. SHP1 expression in Karpas 299 and SU-DHL-1 cells induced similar changes in the STAT3 downstream targets including bcl-2, mcl-1, and cyclin D3. Survivin was only slightly down-regulated. The protein level of NPM-ALK was also decreased. Cell lysates were prepared 24 hours after gene transfection. Cells transfected with pCI empty vector and pIRES2-EGFP served as negative controls for Karpas 299 and SU-DHL-1 cells, respectively. (C) Blockade of SHP1 expression using siRNA in SUP-M2. Inhibition of SHP1 in SU-DHL-1 cells using siRNA induced down-regulation of the expression of SHP1, with 200 pM more effective than 100 pM. There were increases in the expression of pSTAT3, pJAK3, JAK3, and NPM-ALK. One of the STAT3 downstream targets, cyclin D3, was also up-regulated. SUP-M2 cells transfected with the sense SHP1 siRNA served as negative controls.