MicroRNAs (miRNAs) are small regulatory RNAs that act by blocking the translation and increasing the degradation of target transcripts. MiRNAs play a critical role in many biological processes ...including development and differentiation and many studies have shown that major changes in miRNA levels occur in cancer. Since miRNAs degrade target messages, we used this property to develop a novel computational method aimed at determining the actual biological activity of miRNAs using variations in gene expression. Using the method described here, we quantified miRNA activity in papillary thyroid carcinoma and breast cancer, and found a strong and distinctive signal of increased global miRNA activity, embedded in the pertaining gene expression measurements. Interestingly, we found that in these two cancers, miRNA activity is globally increased, and is associated with a global downregulation of miRNA target genes. This downregulation of miRNA regulated genes is particularly noticeable for genes carrying multiple target sites for miRNAs. Among the miRNA-repressed genes, we found a significant enrichment of known tumor suppressors, thereby suggesting that the increased miRNA activity was indeed tumorigenic.
microRNAs (miRNAs) are small noncoding RNAs that regulate cognate mRNAs post-transcriptionally. Human embryonic stem cells (hESC), which exhibit the characteristics of pluripotency and self-renewal, ...may serve as a model to study the role of miRNAs in early human development. We aimed to determine whether endodermally-differentiated hESC demonstrate a unique miRNA expression pattern, and whether overexpression of endoderm-specific miRNA may affect hESC differentiation.
miRNA expression was profiled in undifferentiated and NaButyrate-induced differentiated hESC of two lines, using microarray and quantitative RT-PCR. Then, the effect of lentiviral-based overexpression of liver-specific miR-122 on hESC differentiation was analyzed, using genomewide gene microarrays.
The miRNA profiling revealed expression of three novel miRNAs in undifferentiated and differentiated hESC. Upon NaButyrate induction, two of the most upregulated miRNAs common to both cell lines were miR-24 and miR-10a, whose target genes have been shown to inhibit endodermal differentiation. Furthermore, induction of several liver-enriched miRNAs, including miR-122 and miR-192, was observed in parallel to induction of endodermal gene expression. Stable overexpression of miR-122 in hESC was unable to direct spontaneous differentiation towards a clear endodermal fate, but rather, delayed general differentiation of these cells.
Our results demonstrate that expression of specific miRNAs correlates with that of specific genes upon differentiation, and highlight the potential role of miRNAs in endodermal differentiation of hESC.
The human cytomegalovirus (HCMV) is extremely prevalent in the human population. Infection by HCMV is life threatening in immune compromised individuals and in immune competent individuals it can ...cause severe birth defects, developmental retardation and is even associated with tumor development. While numerous mechanisms were developed by HCMV to interfere with immune cell activity, much less is known about cellular mechanisms that operate in response to HCMV infection. Here we demonstrate that in response to HCMV infection, the expression of the short form of the RNA editing enzyme ADAR1 (ADAR1-p110) is induced. We identified the specific promoter region responsible for this induction and we show that ADAR1-p110 can edit miR-376a. Accordingly, we demonstrate that the levels of the edited-miR-376a (miR-376a(e)) increase during HCMV infection. Importantly, we show that miR-376a(e) downregulates the immune modulating molecule HLA-E and that this consequently renders HCMV infected cells susceptible to elimination by NK cells.
The toll-like receptor-9 (TLR9) agonist cytosine phosphate guanine (CpG), activates hepatic stellate cells (HSCs) and mediates fibrosis. We investigated the TLR9 effects on lymphocyte/HSCs ...interactions. Liver fibrosis was induced in wild-type (WT) mice by intra-peritoneal carbon-tetrachloride (CCl4) induction for 6 weeks. Fibrotic groups were intravenously treated by a vehicle versus CpG along last 2 weeks. Compared to vehicle-treated fibrotic WT, the in-vivo CpG-treatment significantly attenuated hepatic fibrosis and inflammation, associated with decreased CD8 and increased NK liver cells. In-vitro, co-cultures with vehicle-treated fibrotic NK cells increased HSCs proliferation (P<0.001) while their CpG-treated counterparts achieved a significant decrease. To investigate the role of lymphocytes, TLR9(-/-) mice induced-hepatic fibrosis were used. Although TLR9(-/-) mice manifested lower fibrotic profile as compared to their wild-type (WT) counterparts, senescence (SA-β-Gal activity) in the liver and ALT serum levels were significantly greater. In an adoptive transfer model; irradiated WT and TLR9(-/-) recipients were reconstituted with naïve WT or TLR9(-/-) lymphocytes. The adoptive transfer of TLR9(-/-) versus WT lymphocytes led to increased fibrosis of WT recipients. TLR9(-/-) fibrotic recipients reconstituted with TLR9(-/-) or WT lymphocytes showed no changes in hepatic fibrosis severity or ALT serum levels. TLR9 activation had inconsistent effects on lymphocytes and HSCs. The net balance of TLR9 activation in WT, displayed significant anti-fibrotic activity, accompanied by CD8 suppression and increased NK-cells, activity and adherence to HSCs. The pro-fibrotic and pro-inflammatory properties of TLR9(-/-) lymphocytes fail to activate HSCs with an early senescence in TLR9(-/-) mice.
Aging is associated with progressive decline in cell function and with increased damage to macromolecular components. DNA damage, in the form of double-strand breaks (DSBs), increases with age and in ...turn, contributes to the aging process and age-related diseases. DNA strand breaks triggers a set of highly orchestrated signaling events known as the DNA damage response (DDR), which coordinates DNA repair. However, whether the accumulation of DNA damage with age is a result of decreased repair capacity, remains to be determined. In our study we showed that with age there is a decline in the resolution of foci containing γH2AX and pKAP-1 in diethylnitrosamine (DEN)-treated mouse livers, already evident at a remarkably early age of 6-months. Considerable age-dependent differences in global gene expression profiles in mice livers after exposure to DEN, further affirmed these age related differences in the response to DNA damage. Functional analysis identified p53 as the most overrepresented pathway that is specifically enhanced and prolonged in 6-month-old mice. Collectively, our results demonstrated an early decline in DNA damage repair that precedes 'old age', suggesting this may be a driving force contributing to the aging process rather than a phenotypic consequence of old age.
The long-term prognosis after resection of hepatocellular carcinoma (HCC), which is one of the treatment options for early-stage HCC, remains unsatisfactory as a result of a high incidence of disease ...recurrence. Recent studies performed in murine models revealed a link between liver regeneration under chronic inflammation and hepatic tumorigenesis. Sorafenib is a potent drug for advanced HCC with multikinase inhibition activity. We propose that inhibition of signal transduction pathways which are activated during hepatectomy, using Sorafenib, will reduce accelerated tumorigenesis. To test this hypothesis, we studied the Mdr2-knockout (KO) mouse strain, a model of inflammation-associated cancer, which underwent partial hepatectomy (PHx) at three months of age, with or without Sorafenib.Here we show that Sorafenib treatment during PHx inhibited different signal transduction pathways at the multikinase levels, but did not result in increased morbidity or mortality. At the early stages after PHx, Sorafenib treatment had no effect on the course of proliferation, apoptosis and DNA repair in the regenerating liver, but resulted in decreased stellate cells activation and inflammatory response. Finally, we show that Sorafenib treatment during PHx at three months of age resulted in decreased fibrosis and tumor formation at 8.5 months.In conclusion our study indicates that short-term Sorafenib treatment during PHx is safe and effective in inhibiting inflammation-associated cancer, and is therefore a potential strategy for recurrence prevention in patients with early-stage HCC treated with PHx.
An increase in the number of stem cells in blood following mobilization is required to enhance engraftment after high‐dose chemotherapy and improve transplantation outcome. Therefore, an approach ...that improves stem cell mobilization is essential. The interaction between CXCL12 and its receptor, CXCR4, is involved in the retention of stem cells in the bone marrow. Therefore, blocking CXCR4 may result in mobilization of hematopoietic progenitor and stem cells. We have found that the CXCR4 antagonist known as 4F‐benzoyl‐TN14003 (T‐140) can induce mobilization of hematopoietic stem cells and progenitors within a few hours post‐treatment in a dose‐dependent manner. Furthermore, although T‐140 can also increase the number of white blood cells (WBC) in blood, including monocytes, B cells, and T cells, it had no effect on mobilizing natural killer cells. T‐140 was found to efficiently synergize with granulocyte colony‐stimulating factor (G‐CSF) in its ability to mobilize WBC and progenitors, as well as to induce a 660‐fold increase in the number of erythroblasts in peripheral blood. Comparison between the CXCR4 antagonists T‐140 and AMD3100 showed that T‐140 with or without G‐CSF was significantly more potent in its ability to mobilize hematopoietic stem cells and progenitors into blood. These results demonstrate that different CXCR4 antagonists may have different therapeutic potentials.
Disclosure of potential conflicts of interest is found at the end of this article.
Mouse models of hepatocellular carcinoma (HCC) simulate specific subgroups of human HCC. We investigated hepatocarcinogenesis
in Mdr2-knockout (Mdr2-KO) mice, a model of inflammation-associated HCC, ...using gene expression profiling and immunohistochemical
analyses. Gene expression profiling showed that although Mdr2-KO mice differ from other published murine HCC models, they
share several important deregulated pathways and many coordinately differentially expressed genes with human HCC data sets.
Analysis of genome positions of differentially expressed genes in liver tumors revealed a prolonged region of down-regulated
genes on murine chromosome 8 in three of the six analyzed tumor samples. This region is syntenic to human chromosomal regions
that are frequently deleted in human HCC and harbor multiple tumor suppressor genes. Real-time reverse transcription-PCR analysis
of 16 tumor samples confirmed down-regulation of several tumor suppressors in most tumors. We show that in the aged Mdr2-KO
mice, cyclin D1 nuclear level is increased in dysplastic hepatocytes that do not form nodules; however, it is decreased in
most dysplastic nodules and in liver tumors. We found that this decrease is mostly at the protein, rather than the mRNA, level.
These findings raise the question on the role of cyclin D1 at early stages of hepatocarcinogenesis in the Mdr2-KO HCC model.
Furthermore, we show that most liver tumors in Mdr2-KO mice were characterized by the absence of β-catenin activation. In
conclusion, the Mdr2-KO mouse may serve as a model for β-catenin–negative subgroup of human HCCs characterized by low nuclear
cyclin D1 levels in tumor cells and by down-regulation of multiple tumor suppressor genes. (Mol Cancer Res 2007;5(11):1159–70)
The function of imprinted H19 long non-coding RNA is still controversial. It is highly expressed in early embryogenesis and decreases after birth and re-expressed in cancer. To study the role of H19 ...in oncogenesis and pluripotency, we down-regulated H19 expression in vitro and in vivo in pluripotent human embryonic carcinoma (hEC) and embryonic stem (hES) cells. H19 knockdown resulted in a decrease in the expression of the pluripotency markers Oct4, Nanog, TRA-1-60 and TRA-1-81, and in the up-regulation of SSEA1; it further attenuated cell proliferation, decreased cell-matrix attachment, and up-regulated E-Cadherin expression. SCID-Beige mice transplanted with H19 down-regulated hEC cells exhibited slower kinetics of tumor formation, resulting in an increased animal survival. Tumors derived from H19 down-regulated cells showed a decrease in the expression of pluripotency markers and up-regulation of SSEA-1 and E-cadherin. Our results suggest that H19 oncogenicity in hEC cells is mediated through the regulation of the pluripotency state.
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