Journal: Carcinogenesis

Article Title: Microvesicles derived from normal and multiple myeloma bone marrow mesenchymal stem cells differentially modulate myeloma cells' phenotype and translation initiation.

doi: 10.1093/carcin/bgx045

Figure Lengend Snippet: Figure 4. TI and autophagy are essential for MM cell lines’ phenotype. U266 and RPMI 8226 MM cell lines were incubated with MM-MVs (50 μg/ml) and with eIF4E/eIF4G complex inhibitor for 3 days. Then, the MM cells were assayed for cell count and migration (transwell assay) and immunoblotted for PCNA, and Cyclin D1. Analysis of our observations (A) and representative immunoblots (B) are presented. Tubulin served as loading control. Results are expressed as percent (mean ± SE, n ≥ 4) of respec- tive protein expression in control cells not treated with MVs (dotted line). Next, U266 and MM1S cell lines were co-treated with MM-MSCs MVs and autophagy inhibitor (3MA) for 3 days then assayed for dead cell count (trypan blue, C), and apoptosis (Annexin/7AAD analysis and representative dot plots, C). Representative immunoblot of LC3-II in 3MA treated U266 is presented (D). Asterisks depict statistical significance (*P < 0.05, **P < 0.01).

Article Snippet: The following proteins were detected using rabbit/mouse anti-human: peIF4E (Ser209)/total eIF4E, peIF4GI(Ser1108)/total eIF4GI, p4EBP(Ser65)/total 4EBP, pmTOR(Ser2448)/ total mTOR, pMNK(Thr197/Thr202)/total MNK/pERK1/2 and pJNK (Cell Signaling Technology, Danvers, MA, USA); SMAD5 (Epitomics, Burlingame, CA); c-Myc, HIF1α, NFkB, PCNA (Santa-Cruz, CA); tubulin, LC3-II (Sigma).

Techniques: Incubation, Cell Counting, Migration, Transwell Assay, Western Blot, Control, Expressing

Journal: Molecular Cancer Therapeutics

Article Title: Dual targeting of AKT and mammalian target of rapamycin: A potential therapeutic approach for malignant peripheral nerve sheath tumor

doi: 10.1158/1535-7163.mct-08-1008

Figure Lengend Snippet: Figure 1. MPNST exhibit dereg- ulation of the mTOR and AKT path- ways. A, expression of activated mTOR downstream effectors 4EBP1 (top) and S6RP (bottom) in MPNST was assessed via immuno- histochemistry. A tissue microarray consisting of human NF1-associated plexiform neurofibromas and NF1 associated as well as sporadic MPNST was used for immuno- histochemistry. A significantly higher proportion of MPNST cells expressed p4EBP1 and pS6RP compared with neurofibromas (rep- resentative tissue microarray spots are shown; original pictures were captured at ×200). Similarly, stain- ing intensity for both markers was significantly higher in the malignant tumors compared with the benign lesions. Results of Wilcoxon rank- sum tests: blue line, median of the data; bars, 25% and 75% quar- tiles; dots, mean outliers. The two lines are whisker lines, depicting the largest or smallest values that are not outliers. Statistical P values are shown. B, pAKT was evaluated as above; a significantly higher pro- portion of MPNST cells were found to exhibit a stronger pAKT expres- sion level. Furthermore, metastatic lesions expressed higher levels of pAKT compared with localized (pri- mary and recurrent) tumors.

Article Snippet: AKT, pAKT (Ser473), mTOR, pmTOR (Ser2448), S6K1, pS6K1 (Thr398), S6 ribosomal protein (S6RP), pS6RP (Ser235/ Ser236), 4EBP1, p4EBP1 (Thr70), eIF4E, peIF4E (Ser209), pGSK-3α/β (Ser21/Ser9), GSK-3α/β, pMDM2 (Ser166), LC3B, PTEN, and cyclin D1 were all purchased from Cell Signaling; c-Met, MDM2, BCN1, EGFR, β-actin, and antimouse and anti-rabbit second antibody were acquired from Santa Cruz Biotechnology.

Techniques: Expressing, Immunohistochemistry, Microarray, Staining, Whisker Assay

Journal: Molecular Cancer Therapeutics

Article Title: Dual targeting of AKT and mammalian target of rapamycin: A potential therapeutic approach for malignant peripheral nerve sheath tumor

doi: 10.1158/1535-7163.mct-08-1008

Figure Lengend Snippet: Figure 3. Rapamycin abrogates MPNST cell growth but induces the activation of AKT and eIF4E. A, MTS assays showing a rapamycin (48 h) dose- dependent decrease in MPNST cell growth. B, rapamycin (4 h) blocks the activation of the mTOR downstream targets S6K1 and 4EBP1 (Western blot). C, rapamycin at doses indicated induces activation of AKT in both cell lines tested (Western blot). D, rapamycin (4 h) induces the phosphorylation of eIF4E in MPNST cells. This activation is blocked when combining rapamycin with a PI3K inhibitor (LY294002; Western blot).

Article Snippet: AKT, pAKT (Ser473), mTOR, pmTOR (Ser2448), S6K1, pS6K1 (Thr398), S6 ribosomal protein (S6RP), pS6RP (Ser235/ Ser236), 4EBP1, p4EBP1 (Thr70), eIF4E, peIF4E (Ser209), pGSK-3α/β (Ser21/Ser9), GSK-3α/β, pMDM2 (Ser166), LC3B, PTEN, and cyclin D1 were all purchased from Cell Signaling; c-Met, MDM2, BCN1, EGFR, β-actin, and antimouse and anti-rabbit second antibody were acquired from Santa Cruz Biotechnology.

Techniques: Activation Assay, Western Blot, Phospho-proteomics

Journal: Oncotarget

Article Title: Simultaneous targeting of androgen receptor (AR) and MAPK-interacting kinases (MNKs) by novel retinamides inhibits growth of human prostate cancer cell lines

doi:

Figure Lengend Snippet: (A) Effect of NRs treatment (5 μmol/L) on PC-3 cell migration was assessed by wound healing assay after 24 h of wound formation. Representative photomicrographs of initial and final wounds are shown at 100x magnification. (B) Effect of NRs on PC-3 cell invasion was evaluated by transwell migration assay. PC-3 cells were seeded on matrigel coated boyden chamber and treated with NRs (5 μmol/L, 24 h). Photographs represent the extent of cell invasion in each of the treated cells. (C) Western blot analysis of the effect of NRs on the expression of cadherins in PC-3 cells. Cells were treated with indicated compound 20 μmol/L for 24 h. Total cell lysates were separated by SDS-PAGE and probed with E- and N-cadherin antibodies. Vehicle treated cells were included as a control and all blots were reprobed for β-actin for equal protein loading and transfer.

Article Snippet: Unless otherwise described, 30 μg protein was resolved by SDS–polyacrylamide gel electrophoresis, transferred, and immunoblotted with using the following antibodies: AR, Bax, caspase-3, CHIP, cyclin B, cyclin D1, E-cadherin, eIF4E, MNK1, MDM2, N-cadherin, cleaved PARP, peIF4E ser209 , PSA procured from Cell Signaling Technology, Danvers, MA, USA; anti-MNK2 was purchased from Sigma-Aldrich, St. Louis, MO, USA; and normal rabbit IgG, cyclin D1, E-cadherin, and Ub was from Santa Cruz Biotechnology, CA, USA (19).

Techniques: Migration, Wound Healing Assay, Transwell Migration Assay, Western Blot, Expressing, SDS Page, Control