Search

Introduction Acute myeloid leukemia (AML) is one of the most common types of leukemia diagnosed in adults. While many advancements have been made in the understanding of the genetic components [...], Natural killer (NK) cells can have potent antileukemic activity following haplo-mismatched, T cell-depleted stem cell transplantations for the treatment of acute myeloid leukemia (AML), but they are not successful in eradicating de novo AML. Here, we have used a mouse model of de novo AML to elucidate the mechanisms by which AML evades NK cell surveillance. NK cells in leukemic mice displayed a marked reduction in the cytolytic granules perforin and granzyme B. Further, as AML progressed, we noted the selective loss of an immature subset of NK cells in leukemic mice and in AML patients. This absence was not due to elimination by cell death or selective reduction in proliferation, but rather to the result of a block in NK cell differentiation. Indeed, NK cells from leukemic mice and humans with AML showed lower levels of TBET and EOMES, transcription factors that are critical for terminal NK cell differentiation. Further, the microRNA miR-29b, a regulator of T-bet and EOMES, was elevated in leukemic NK cells. Finally, deletion of miR-29b in NK cells reversed the depletion of this NK cell subset in leukemic mice. These results indicate that leukemic evasion of NK cell surveillance occurs through miR-mediated dysregulation of lymphocyte development, representing an additional mechanism of immune escape in cancer.

miR-122, an abundant liver-specific microRNA (miRNA), regulates cholesterol metabolism and promotes hepatitis C virus (HCV) replication. Reduced miR-122 expression in hepatocellular carcinoma (HCC) correlates with metastasis and poor prognosis. Nevertheless, [...]

1. Introduction Fragile sites can be defined as heritable-specific loci on human chromosomes that exhibit nonrandom gaps or breaks when chromosomes are exposed to specific cell culture conditions [1]. Classification [...], Chromosomal common fragile sites (CFSs) are specific mammalian genomic regions that show an increased frequency of gaps and breaks when cells are exposed to replication stress in vitro. CFSs are also consistently involved in chromosomal abnormalities in vivo related to cancer. Interestingly, several CFSs contain one or more tumor suppressor genes whose structure and function are often affected by chromosomal fragility. The two most active fragile sites in the human genome are FRA3B and FRA16D where the tumor suppressor genes FHIT and WWOX are located, respectively. The best approach to study tumorigenic effects of altered tumor suppressors located at CFSs in vivo is to generate mouse models in which these genes are inactivated. This paper summarizes our present knowledge on mouse models of cancer generated by knocking out tumor suppressors of CFS.

Background: The self-renewing ability of HSCs is fundamental for the maintenance of a pool of bone marrow precursors throughout the life of an individual. The genetic mechanisms underlying such a complex process are still poorly understood. Results and Significance: Here, we show that constitutive in vivo deletion of miR29ab1 leads to reduced number of HSCs and that miR29ab1 deficient bone marrow cannot repopulate the bone marrow of irradiated mice. An Affymetrix analysis of the miR29ab1 knockout mice identifies key proteins that could be responsible for this phenotype, as DNMT3a and b. Moreover, our findings reveal that whereas miR29b2c knockout mice do not exhibit any spontaneous abnormality, the double knock out – miR29ab1b2c – has marked generalized atrophy, raising the possibility that the two bi-cistrons might cooperate in order to maintain the stem cell number in general, not only limited to the bone marrow. [ABSTRACT FROM AUTHOR]

Differentiation of lung vascular smooth muscle cells (vSMCs) is tightly regulated during development or in response to challenges in a vessel specific manner. Aberrant vSMCs specifically associated with distal pulmonary arteries have been implicated in the pathogenesis of respiratory diseases, such as pulmonary arterial hypertension (PAH), a progressive and fatal disease, with no effective treatment. Therefore, it is highly relevant to understand the underlying mechanisms of lung vSMC differentiation. miRNAs are known to play critical roles in vSMC maturation and function of systemic vessels; however, little is known regarding the role of miRNAs in lung vSMCs. Here, we report that miR-29 family members are the most abundant miRNAs in adult mouse lungs. Moreover, high levels of miR-29 expression are selectively associated with vSMCs of distal vessels in both mouse and human lungs. Furthermore, we have shown that disruption of miR-29 in vivo leads to immature/synthetic vSMC phenotype specifically associated with distal lung vasculature, at least partially due to the derepression of KLF4, components of the PDGF pathway and ECM-related genes associated with synthetic phenotype. Moreover, we found that expression of FBXO32 in vSMCs is significantly upregulated in the distal vasculature of miR-29 null lungs. This indicates a potential important role of miR-29 in smooth muscle cell function by regulating FBXO32 and SMC protein degradation. These results are strongly supported by findings of a cell autonomous role of endogenous miR-29 in promoting SMC differentiation in vitro. Together, our findings suggested a vessel specific role of miR-29 in vSMC differentiation and function by targeting several key negative regulators. [ABSTRACT FROM AUTHOR]

Invariant natural killer T cells (iNKT cells) are CD1d-restricted, lipid antigen-reactive T lymphocytes with immunoregulatory functions. iNKT cell development in the thymus proceeds through subsequent stages, defined by the expression of CD44 and NK1.1, and is dictated by a unique gene expression program, including microRNAs. Here, we investigated whether miR-155, a microRNA involved in differentiation of most hematopoietic cells, played any role in iNKT cell development.To this end, we assessed the expression of miR- 155 along iNKT cell maturation in the thymus, and studied the effects of miR-155 on iNKT cell development using Lck-miR-155 transgenic mice, which over express miR-155 in T cell lineage under the lymphocyte-specific protein tyrosine kinase (Lck) promoter. We show that miR-155 is expressed by newly selected immature wild-type iNKT cells and turned off along iNKT cells differentiation. In transgenic mice, miR-155 over-expression resulted in a substantial block of iNKT cell maturation at Stage 2, in the thymus toward an overall reduction of peripheral iNKT cells, unlike mainstream T cells. Furthermore, the effects of miR-155 over-expression on iNKT cell differentiation were cell autonomous. Finally, we identified Ets1 and ITK transcripts as relevant targets of miR-155 in iNKT cell differentiation. Altogether, these results demonstrate that a tight control of miR-155 expression is required for the development of iNKT cells. [ABSTRACT FROM AUTHOR]

MicroRNAs are being exploited for diagnosis, prognosis and monitoring of cancer and other diseases. Their high tissue specificity and critical role in oncogenesis provide new biomarkers for the diagnosis and classification of cancer as well as predicting patients' outcomes. MicroRNAs signatures have been identified for many human tumors, including colorectal cancer (CRC). In most cases, metastatic disease is difficult to predict and to prevent with adequate therapies. The aim of our study was to identify a microRNA signature for metastatic CRC that could predict and differentiate metastatic target organ localization. Normal and cancer tissues of three different groups of CRC patients were analyzed. RNA microarray and TaqMan Array analysis were performed on 66 Italian patients with or without lymph nodes and/or liver recurrences. Data obtained with the two assays were analyzed separately and then intersected to identify a primary CRC metastatic signature. Five differentially expressed microRNAs (hsa-miR-21, -103, -93, -31 and -566) were validated by qRT-PCR on a second group of 16 American metastatic patients. In situ hybridization was performed on the 16 American patients as well as on three distinct commercial tissues microarray (TMA) containing normal adjacent colon, the primary adenocarcinoma, normal and metastatic lymph nodes and liver. Hsa-miRNA-21, -93, and -103 upregulation together with hsa-miR-566 downregulation defined the CRC metastatic signature, while in situ hybridization data identified a lymphonodal invasion profile. We provided the first microRNAs signature that could discriminate between colorectal recurrences to lymph nodes and liver and between colorectal liver metastasis and primary hepatic tumor. [ABSTRACT FROM AUTHOR]

Micro RNAs (mi RNAs) are small, regulatory non-coding RNAs that have potent effects on gene expression. Several mi RNA are deregulated in cellular processes involved in human liver diseases and regulation of cellular processes. Recent studies have identified the involvement of mi R-29 in hepatic fibrosis and carcinogenesis. Although several targets of mi R-29 have been identified, there is limited information regarding the cell-type specific roles of mi R-29 in the liver, and we sought to evaluate the role of this mi RNA in hepatic pathobiology. We report the generation of a tissue-specific knockout mouse to evaluate the role of mi R-29 in hepatic fibrosis and carcinogenesis in response to injury. We hypothesized that mi R-29 contributes to the hepatocyte driven response to chronic cellular injury that results in fibrosis. In support of this hypothesis, fibrosis and mortality were enhanced in mi R29 knockout mice in response to carbon tetrachloride. Genome-wide gene expression analysis identified an over-representation of genes associated with fibrosis. The oncofetal RNA H19 was modulated in a mi R-29 dependent manner following exposure to carbon tetrachloride in vivo. The impact of a hepatocyte specific mi R-29 knockout on survival following chronic hepatic injury in vivo implicates this mi RNA as a potential target for intervention. These results provide evidence of the involvement of mi R-29 in chronic hepatic injury, and suggest a role for deregulated hepatocyte expression of mi R-29 in the response to hepatic injury, fibrosis and carcinogenesis. [ABSTRACT FROM AUTHOR]

Ptprj is a ubiquitously expressed murine gene encoding a receptor-type protein tyrosine phosphatase, which has recently been proposed as a candidate gene on the locus Scc1 for colon cancer susceptibility. It has been demonstrated that PTPRJ, the human homologue of Ptprj, is involved in the control of cell growth and adhesion, being furthermore altered in several types of cancer including mammary, thyroid, lung, colon, and pancreatic cancers. To investigate the biological functions of Ptprj, we have generated mice deficient in this receptor protein tyrosine phosphatase. Ptprj-deficient mice are viable, fertile, and show no gross anatomical alterations. Furthermore, neither changes in life span nor spontaneous tumor appearance were observed in Ptprj-null mice. Our results indicate that Ptprj is dispensable for normal growth and development in mice. [ABSTRACT FROM AUTHOR]

Th cell programming and function is tightly regulated by complex biological networks to prevent excessive inflammatory responses and autoimmune disease. The importance of microRNAs (miRNAs) in this process is highlighted by the preferential Th1 polarization of Dicer-deficient T cells that lack miRNAs. Using genetic knockouts, we demonstrate that loss of endogenous miR-29, derived from the miR-29ab1 genomic cluster, results in unrestrained T-bet expression and IFN-γ production. miR-29b regulates T-bet and IFN-γ via a direct interaction with the 3' untranslated regions, and IFN-γ itself enhances miR-29b expression, establishing a novel regulatory feedback loop. miR-29b is increased in memory CD4(+) T cells from multiple sclerosis (MS) patients, which may reflect chronic Th1 inflammation. However, miR-29b levels decrease significantly upon T cell activation in MS patients, suggesting that this feedback loop is dysregulated in MS patients and may contribute to chronic inflammation. miR-29 thus serves as a novel regulator of Th1 differentiation, adding to the understanding of T cell-intrinsic regulatory mechanisms that maintain a balance between protective immunity and autoimmunity.

Metallothioneins (MT) are potent scavengers of free radicals that are silenced in primary hepatocellular carcinomas (HCC) of human and rodent origin. To examine whether loss of MT promotes hepatocarcinogenesis, male Mt-1 and Mt-2 double knockout (MTKO) and wild-type (WT) mice were exposed to diethylnitrosamine (DEN) and induction of HCC was monitored at 23 and 33 weeks. The size and number of liver tumors, the ratio between liver and body weight, and liver damage were markedly elevated in the MTKO mice at both time points compared with the WT mice. At 23 weeks, MTKO mice developed HCC whereas WT mice developed only preneoplastic nodules suggesting that loss of MT accelerates hepatocarcinogenesis. MTKO tumors also exhibited higher superoxide anion levels. Although NF-κB activity increased in the liver nuclear extracts of both genotypes after DEN exposure, the complex formed in MTKO mice was predominantly p50/65 heterodimer (transcriptional activator) as opposed to p50 homodimer (transcriptional repressor) in WT mice. Phosphorylation of p65 at Ser276 causing its activation was also significantly augmented in DEN-exposed MTKO livers. NF-κB targets that include early growth response genes and proinflammatory cytokines were significantly upregulated in MTKO mice. Concurrently, there was a remarkable increase (∼100-fold) in Pai-1 expression; significant increase in c-Jun, c-Fos, c-Myc, Ets2, and ATF3 expressions; and growth factor signaling that probably contributed to the increased tumor growth in MTKO mice. Taken together, these results demonstrate that MTs protect mice from hepatocarcinogen-induced liver damage and carcinogenesis, underscoring their potential therapeutic application against hepatocellular cancer., (©2010 AACR.)

We report here that miR-155 and miR-125b play a role in innate immune response. LPS stimulation of mouse Raw 264.7 macrophages resulted in the up-regulation of miR-155 and down-regulation of miR-125b levels. The same changes also occurred when C57BL/6 mice were i.p. injected with LPS. Furthermore, the levels of miR-155 and miR-125b in Raw 264.7 cells displayed oscillatory changes in response to TNF-alpha. These changes were impaired by pretreating the cells with the proteasome inhibitor MG-132, suggesting that these two microRNAs (miRNAs) may be at least transiently under the direct control of NF-kappaB transcriptional activity. We show that miR-155 most probably directly targets transcript coding for several proteins involved in LPS signaling such as the Fas-associated death domain protein (FADD), IkappaB kinase epsilon (IKKepsilon), and the receptor (TNFR superfamily)-interacting serine-threonine kinase 1 (Ripk1) while enhancing TNF-alpha translation. In contrast, miR-125b targets the 3'-untranslated region of TNF-alpha transcripts; therefore, its down-regulation in response to LPS may be required for proper TNF-alpha production. Finally, Emu-miR-155 transgenic mice produced higher levels of TNF-alpha when exposed to LPS and were hypersensitive to LPS/d-galactosamine-induced septic shock. Altogether, our data suggest that the LPS/TNF-alpha-dependent regulation of miR-155 and miR-125b may be implicated in the response to endotoxin shock, thus offering new targets for drug design.

The expression of the WWOX tumor suppressor gene is lost or reduced in a large fraction of various cancers, including prostate cancer. We previously reported that Wwox overexpression induced apoptosis and suppressed prostate cancer growth in vitro and in vivo. In this study, pathways through which Wwox contributes to control of prostate cancer cell growth have been investigated. We found that Wwox interacts with Ap2gamma and prevents it from entering the nucleus to bind the ERBB2 promoter region to activate transcription of ERBB2, a mediator of androgen receptor activity and prostate cancer cell growth at limiting androgen concentration. Ectopic expression of Wwox reduced ErbB2 protein expression in vitro and expression of Wwox protein inversely correlated with expression of ErbB2 protein in prostate cancer tissues. Furthermore, Wwox suppressed Ap2gamma/ErbB2-induced prostate cancer cell growth and suppressed prostate-specific antigen secretion through interaction with Ap2gamma and down-modulation of ErbB2, an effect that required functional androgen receptor.

Chronic lymphocytic leukemia (CLL) is the most common leukemia in the world. The TCL1 gene, responsible for prolymphocytic T cell leukemia, is also overexpressed in human B cell malignancies and overexpression of the Tcl1 protein occurs frequently in CLL. Aging transgenic mice that overexpress TCL1 under control of the mu immunoglobulin gene enhancer, develop a CD5+ B cell lymphoproliferative disorder mimicking human CLL and implicating TCL1 in the pathogenesis of CLL. In the current study, we exploited this transgenic mouse to investigate two different CLL-related issues: potential treatment of CLL and characterization of neoplasms that accompany CLL. We successfully transplanted CLL cells into syngeneic mice that led to CLL development in the recipient mice. This approach allowed us to verify the involvement of the Tcl1/Akt/mTOR biochemical pathway in the disease by testing the ability of a specific pharmacologic agent, rapamycin, to slow CLL. We also showed that 36% of these transgenic mice were affected by solid malignancies, in which the expression of the Tcl1 protein was absent. These findings indicate that other oncogenic mechanism(s) may be involved in the development of solid tumors in Emu-TCL1 transgenic mice.