These results suggest that targeted delivery of MYCN siRNA by folate receptor-targeted lipid vesicles into LA-N-5 cells is efficacious and capable of suppressing MYCN mRNA expression both in vitro and in vivo.

BackgroundThe cyclic AMP (cAMP) signaling pathway has been reported to either promote or suppress cell death, in a cell context-dependent manner. Our previous study has shown that the induction of dynein light chain (DLC) by cAMP response element-binding protein (CREB) is required for cAMP-mediated inhibition of mitogen-activated protein kinase (MAPK) p38 activation in fibroblasts, which leads to suppression of NF-κB activity and promotion of tumor necrosis factor-α (TNF-α)-induced cell death. However, it remains unknown whether this regulation is also applicable to fibroblastoma cells.MethodsIntracellular cAMP was determined in L929 fibroblastoma cells after treatment of the cells with various cAMP elevation agents. Effects of cAMP in the presence or absence of the RNA synthesis inhibitor actinomycin D or small interfering RNAs (siRNAs) against CREB on TNF-α-induced cell death in L929 cells were measured by propidium iodide (PI) staining and subsequent flow cytomety. The activation of p38 and c-Jun N-terminal protein kinase (JNK), another member of MAPK superfamily, was analyzed by immunoblotting. JNK selective inhibitor D-JNKi1 and p38 selective inhibitor SB203580 were included to examine the roles of JNK and p38 in this process. The expression of DLC or other mediators of cAMP was analyzed by immunoblotting. After ectopic expression of DLC with a transfection marker GFP, effects of cAMP on TNF-α-induced cell death in GFP+ cells were measured by PI staining and subsequent flow cytomety.ResultsElevation of cAMP suppressed TNF-α-induced necrotic cell death in L929 fibroblastoma cells via CREB-mediated transcription. The pro-survival role of cAMP was associated with selective unresponsiveness of L929 cells to the inhibition of p38 activation by cAMP, even though cAMP significantly inhibited the activation of JNK under the same conditions. Further exploration revealed that the induction of DLC, the major mediator of p38 inhibition by cAMP, was impaired in L929 cells. Enforced inhibition of p38 activation by using p38 specific inhibitor or ectopic expression of DLC reversed the protection of L929 cells by cAMP from TNF-α-induced cell death.ConclusionThese data suggest that the lack of a pro-apoptotic pathway in tumor cells leads to a net survival effect of cAMP.

MKK7 works as a cytoplasmic anchoring protein for JNK1 in various cell lines but exhibits aberrant nuclear entry in Jurkat cells, which leads to resistance to Fas-mediated apoptosis.

The high expression of folate receptor (FR) on cancer cells might be a potential target for cancer therapy. In this study, the FR-β expression and the modulation effect of all-trans retinoic acid (ATRA) in a number of cancer cell lines were analyzed. The gateway of ATRA activity on FR-β expression was further studied by a panel of retinoid activators and inhibitors. The results revealed that ATRA was capable of upregulating the expression of FR-β protein in KG-1 cells in a dosage-dependent manner, not in KG-1a, NB4, HL60, 293, L1210, JAR, and Hela cells. FR-β mRNA expression in KG-1 cells was higher when ATRA was present in culture medium at 10⁻⁶ mol/L for 5 days, and it went down to baseline when ATRA was removed from the medium, vice versa. The upregulation of FR-β expression in KG-1 cells by ATRA was not associated with cell proliferation and differentiation. In addition, activators of retinoid acid receptor (RAR)α and RARγ, CD336, and CD2781 also induced FR-β expression. The induction of FR-β expression by CD336 could be inhibited by RARγ antagonist CD2665; RARβ agonist CD-417 and CD-2314 as well as retinoid X receptor (RXR) agonist LG100364 could not induce FR-β expression. These results indicate that ATRA within a certain range of concentration could reversibly induce the expression of FR-β in a dosage- and cell type-dependent manner, and its action in KG-1 cells might be associated with the signal transduction of retinoid receptor RARα and RARγ, rather than RARβ and RXRs.

microRNA‑34a (miRNA‑34a) plays an important role in the pathogenesis of leukemia. This study aimed to explore its role in the proliferation of HL‑60 cells and the correlation with some of its predicted target genes: the cyclin‑dependent kinase 4 (CDK4), the oncogene MYB and the silent information regulator 1 (SIRT1). We first analyzed the expression of miR‑34a, CDK4, MYB and SIRT1 in peripheral blood samples from acute leukemia (AL) patients and healthy controls, and conducted a correlation analysis. HL‑60 cells were then transfected with miR‑34a and control 'scramble̓ miRNA, and quantitative RT‑PCR and western blotting were used to analyze the effects of the interfering sequence in HL‑60 cells. The expression of miR‑34a was significantly reduced in AL patients compared to healthy controls (P<0.01), and negatively correlated with the expression of CDK4 and MYB. Sub‑group analysis revealed that the expression of MYB was significantly lower in AL children <3 years old compared to those >3 years. Following the transfection of HL‑60 cells with miR‑34a, the mRNA level of CDK4, MYB and SIRT1 decreased by 53.2, 43.3 and 33.5%, respectively, compared to the control, similarly to the respective changes in protein levels. This study showed that the expression of miR‑34a negatively correlates with the expression of CDK4 and MYB in pediatric patients with acute leukemia. miRNA‑34a downregulates the expression of the CDK4, MYB and SIRT1 genes in vitro; it may thus represent a novel therapeutic target for acute leukemia.