MicroRNAs (miRNAs) are a class of recently discovered noncoding RNA genes that post-transcriptionally regulate gene expression. It is becoming clear that miRNAs play an important role in the regulation of gene translation during development. However, in mammals, expression data are principally based on whole tissue analysis and are still very incomplete. We isolated CD34 þ CD38À hematopoietic stem cells (HSCs) from human umbilical cord blood, on the basis of cell-surface markers using fluorescence-activated cell sorting (FACS). Also, CD34þ subpopulation was FACS isolated as the control. Next, using microarray containing oligonucleotides corresponding to 517 miRNAs from human, mouse, and rat genomes, we obtained miRNA gene expression profiles of both subpopulations. We focused on the HSCs correlative miRNAs with comparison to the control. The miRNAs of particular interest were confirmed by real-time RT-PCR. HSCs-overexpressed hsa-miR-520h and underexpressed hsa-miR-129 were selected for target prediction and analysis. The result showed that EIF2C3 and CAMTA1, genes related to miRNAs processing or transcription regulation, were proved to be real targets for hsa-miR-129. And ABCG2, involved in stemness maintaining, a real target for hsa-miR520h. Finally, we chose hsa-miR-520h, enriched in HSCs but low in CD34 þ cells, for functional characterization, because of its possible role in differentiation of HSCs by regulating ABCG2. As a result, hsa-miR-520h transduction into CD34 þ cells greatly increased number of different progenitor colonies in Colony-Forming-Cell assays, suggesting that hsa-miR520h may promote differentiation of HSCs into progenitor cells by inhibiting ABCG2 expression. This study paves the way for identifying HSC-specific miRNAs and their roles in HSC development.
FXR is a member of the nuclear receptor superfamily and is the primary bile acid receptor. We previously showed that FXR was required for the promotion of liver regeneration/repair after physical resection or liver injury. However, the mechanism by which FXR promotes liver regeneration/repair is still unclear. Here we showed that both hepatic-FXR and intestine-FXR contributed to promoting liver regeneration/repair after either 70% partial hepatectomy or CCl4-induced liver injury. Hepatic FXR, but not intestine FXR, is required for the induction of Foxm1b gene expression in liver during liver regeneration/repair. In contrast, intestine FXR is activated to induce FGF15 expression in intestine after liver damage. Ectopic expression of FGF15 was able to rescue the defective liver regeneration/repair in intestine-specific FXR null mice. Conclusion These results demonstrate that, in addition to the cell-autonomous effect of hepatic FXR, the endocrine FGF15 pathway activated by FXR in intestine also participates in the promotion of liver regeneration/repair.
Farnesoid X receptor (FXR) (nuclear receptor subfamily 1, group H, member 4) is a member of nuclear hormone receptor superfamily, which plays essential roles in metabolism of bile acids, lipid, and glucose. We previously showed spontaneously hepatocarcinogenesis in aged FXR(-/-) mice, but its relevance to human hepatocellular carcinoma (HCC) is unclear. Here, we report a systematical analysis of hepatocarcinogenesis in FXR(-/-) mice and FXR expression in human liver cancer. In this study, liver tissues obtained from FXR(-/-) and wild-type mice at different ages were compared by microarray gene profiling, histological staining, chemical analysis, and quantitative real-time PCR. Primary hepatic stellate cells and primary hepatocytes isolated from FXR(-/-) and wild-type mice were also analyzed and compared. The results showed that the altered genes in FXR(-/-) livers were mainly related to metabolism, inflammation, and fibrosis, which suggest that hepatocarcinogenesis in FXR(-/-) mice recapitulated the progression of human liver cancer. Indeed, FXR expression in human HCC was down-regulated compared with normal liver tissues. Furthermore, the proinflammatory cytokines, which were up-regulated in human HCC microenvironment, decreased FXR expression by inhibiting the transactivity of hepatic nuclear factor 1α on FXR gene promoter. Our study thereby demonstrates that the down-regulation of FXR has an important role in human hepatocarcinogenesis and FXR(-/-) mice provide a unique animal model for HCC study.
Bcr-Abl tyrosine kinase inhibitors (TKIs) have been a remarkable success for the treatment of Ph ؉ chronic myeloid leukemia (CML). However, a significant proportion of patients treated with TKIs develop resistance because of leukemia stem cells (LSCs) and T315I mutant Bcr-Abl. Here we describe the unknown activity of the natural product berbamine that efficiently eradicates LSCs and T315I mutant BcrAbl clones. Unexpectedly, we identify CaMKII ␥ as a specific and critical target of berbamine for its antileukemia activity. Berbamine specifically binds to the ATPbinding pocket of CaMKII ␥, inhibits its phosphorylation and triggers apoptosis of leukemia cells. More importantly, CaMKII ␥ is highly activated in LSCs but not in normal hematopoietic stem cells and coactivates LSC-related -catenin and Stat3 signaling networks. The identification of CaMKII ␥ as a specific target of berbamine and as a critical molecular switch regulating multiple LSC-related signaling pathways can explain the unique antileukemia activity of berbamine. These findings also suggest that berbamine may be the first ATP-competitive inhibitor of CaMKII ␥, and potentially, can serve as a new type of molecular targeted agent through inhibition of the CaMKII ␥ activity for treatment of leukemia. (Blood. 2012; 120(24):4829-4839) IntroductionChronic myeloid leukemia (CML), which accounts for approximately 20% of all adult leukemias, 1 is characterized by the presence of the Philadelphia chromosome (Ph ϩ ), which results from a chromosomal translocation between the Bcr gene on chromosome 22 and the Abl gene on chromosome 9. 2 This translocation produces the fusion protein Bcr-Abl that has constitutive kinase activity 3 and is essential for the growth of CML cells and has become an attractive target for treatment of Ph ϩ CML cases, and the Abl tyrosine kinase inhibitors (TKIs) are now first-line therapeutic agents. [4][5][6] Inhibition of Bcr-Abl with Abl tyrosine kinase inhibitors (TKIs), such as imatinib (IM), is highly effective in controlling CML at chronic phase but not curing the disease. This is largely because of the inability of these kinase inhibitors to kill leukemia stem cells (LSCs) responsible for initiation, drug resistance, and relapse of CML 4-6 and Bcr-Abl gene mutation, particularly T315I mutant Bcr-Abl clones. 7-9 Thus, drug resistance associated with TKIs has created a need for more potent and safer therapies against other targets apart from the Bcr-Abl oncogenic kinase.Increasing evidence shows that traditional Chinese medicine (TCM) products not only play important roles in the discovery and development of drugs, but can also be used as molecular probes for identifying therapeutic targets. Homoharringtonine, arsenic trioxide, and triptolide are 3 famous examples. 9-11 Berbamine (BBM) is a structurally unique bisbenzylisoquinoline isolated from TCM Berberis amurensis, and has been used in traditional Chinese medicine for treating a variety of diseases from inflammation to tumors for many years. 12,13 It possesses a unique profile ...
ERRα is an orphan nuclear receptor emerging as a novel biomarker of breast cancer. Over-expression of ERRα in breast tumor is considered as a prognostic factor of poor clinical outcome. The mechanisms underlying the dysexpression of this nuclear receptor, however, are poorly understood. MicroRNAs (miRNAs) regulate gene expression at the post-transcriptional level and play important roles in tumor initiation and progression. In the present study, we have identified that the expression of ERRα is regulated by miR-137, a potential tumor suppressor microRNA. The bioinformatics search revealed two putative and highly conserved target-sites for miR-137 located within the ERRα 3′UTR at nt 480–486 and nt 596–602 respectively. Luciferase-reporter assay demonstrated that the two predicted target sites were authentically functional. They mediated the repression of reporter gene expression induced by miR-137 in an additive manner. Moreover, ectopic expression of miR-137 down-regulated ERRα expression at both protein level and mRNA level, and the miR-137 induced ERRα-knockdown contributed to the impaired proliferative and migratory capacity of breast cancer cells. Furthermore, transfection with miR-137mimics suppressed at least two downstream target genes of ERRα–CCNE1 and WNT11, which are important effectors of ERRα implicated in tumor proliferation and migration. Taken together, our results establish a role of miR-137 in negatively regulating ERRα expression and breast cancer cell proliferation and migration. They suggest that manipulating the expression level of ERRα by microRNAs has the potential to influence breast cancer progression.
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