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Elevated levels of miR-155 in solid and liquid malignancies correlate with aggressiveness of the disease. In this manuscript, we show that miR-155 targets transcripts encoding IcosL, the ligand for Inducible T-cell costimulator (Icos), thus impairing the ability of T cells to recognize and eliminate malignant cells. We specifically found that overexpression of miR-155 in B cells of Eµ-miR-155 mice causes loss of IcosL expression as they progress toward malignancy. Similarly, in mice where miR-155 expression is controlled by a Cre-Tet-OFF system, miR-155 induction led to malignant infiltrates lacking IcosL expression. Conversely, turning miR-155 OFF led to tumor regression and emergence of infiltrates composed of IcosL-positive B cells and Icos-positive T cells forming immunological synapses. Therefore, we next engineered malignant cells to express IcosL, in order to determine whether IcosL expression would increase tumor infiltration by cytotoxic T cells and reduce tumor progression. Indeed, overexpressing an IcosL-encoding cDNA in MC38 murine colon cancer cells before injection into syngeneic C57BL6 mice reduced tumor size and increased intratumor CD8+ T cell infiltration, that formed synapses with IcosL-expressing MC38 cells. Our results underscore the fact that by targeting IcosL transcripts, miR-155 impairs the infiltration of tumors by cytotoxic T cells, as well as the importance of IcosL on enhancing the immune response against malignant cells. These findings should lead to the development of more effective anticancer treatments based on maintaining, increasing, or restoring IcosL expression by malignant cells, along with impairing miR-155 activity. [ABSTRACT FROM AUTHOR]

Background: With advances in systemic therapy translating to improved survival in metastatic malignancies, spine metastases have become an increasingly common source of morbidity. Achieving durable local control (LC) for patients with circumferential epidural disease can be particularly challenging. Circumferential stereotactic body radiotherapy (SBRT) may offer improved LC for circumferential vertebral and/or epidural metastatic spinal disease, but prospective (and retrospective) data are extremely limited. We sought to evaluate the feasibility, toxicity, and cancer control outcomes with this novel approach to circumferential spinal disease. Methods: We retrospectively identified all circumferential SBRT courses delivered between 2013 and 2019 at a tertiary care institution for postoperative or intact spine metastases. Radiotherapy was delivered to 14--27.5 Gy in one to five fractions. Feasibility was assessed by determining the proportion of plans for which ≥95% planning target volume (PTV) was coverable by ≥95% prescription dose. The primary endpoint was 1-year LC. Factors associated with increased likelihood of local failure (LF) were explored. Acute and chronic toxicity were assessed. Detailed dosimetric data were collected. Results: Fifty-eight patients receiving 64 circumferential SBRT courses were identified (median age 61, KPS ≥70, 57% men). With a median follow-up of 15 months, the 12-month local control was 85% (eight events). Five and three recurrences were in the epidural space and bone, respectively. On multivariate analysis, increased PTV and uncontrolled systemic disease were significantly associated with an increased likelihood of LF; ≥95% PTV was covered by ≥95% prescription dose in 94% of the cases. The rate of new or progressive vertebral compression fracture was 8%. There were no myelitis events or any grade 3+ acute or late toxicities. Conclusions: For patients with circumferential disease, circumferential spine SBRT is feasible and may offer excellent LC without significant toxicity. A prospective evaluation of this approach is warranted. [ABSTRACT FROM AUTHOR]

Aortic aneurism open repair surgery can cause spinal cord (SC) injury with 5–15% of patients developing paraparesis or paraplegia. Using a mouse model of transient aortic cross-clamping (ACC), we have previously found that the expression of proinflammatory microRNA miR-155 increases in motoneurons (MNs) and endothelial cells (ECs) of ischemic SCs, and that global miR-155 deletion decreases the percentage of paraplegia by 37.4% at 48-h post-ACC. Here, we investigated the cell-specific contribution of miR-155 in choline acetyltransferase-positive (ChAT+) neurons (that include all MNs of the SC) and ECs to SC injury after ACC. Mice lacking miR-155 in ChAT+ neurons (MN- miR-155 -KO mice) developed 24.6% less paraplegia than control mice at 48-h post-ACC. In contrast, mice lacking miR-155 in ECs (ECs- miR-155 -KO mice) experienced the same percentage of paraplegia as control mice, despite presenting smaller central cord edema. Unexpectedly, mice overexpressing miR-155 in ChAT+ neurons were less likely than control mice to develop early paraplegia during the first day post-ACC, however they reached the same percentage of paraplegia at 48-h. In addition, all mice overexpressing miR-155 in ECs (ECs- miR-155 -KI mice) were paraplegic at 48-h post-ACC. Altogether, our results suggest that miR-155 activity in ChAT+ neurons protects the SC against ischemic injury during the first day post-ACC before becoming deleterious during the second day, which indicates that early and late paraplegias arise from different molecular malfunctions. These results point to the need to develop specific protective therapeutics aimed at inhibiting both the early and late deleterious events after open repair surgery of aortic aneurisms. [ABSTRACT FROM AUTHOR]

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Background: Enterochromaffin cells (EC) synthesize and release 5-HT and ATP to trigger or modulate gut neural reflexes and transmit information about visceral/pain sensation. Alterations in 5-HT signaling mechanisms may contribute to the pathogenesis of IBD or IBS, but the pharmacologic or molecular mechanisms modulating Ca2+- dependent 5-HT release are not understood. Previous studies indicated that purinergic signaling via ATP and ADP is an important mechanism in modulation of 5-HT release. However, EC cells also respond to UTP and UDP suggesting uridine triphosphate receptor and signaling pathways are involved as well. We tested the hypothesis that UTP is a regulator of 5-HT release in human EC cells. Methods: UTP signaling mechanisms were studied in BON cells, a human EC model, using Fluo-4/Ca2+imaging, patch-clamp, pharmacological analysis, immunohistochemistry, western blots and qPCR. 5-HT release was monitored in BON or EC isolated from human gut surgical specimens (hEC). Results: UTP, UTPγS, UDP or ATP induced Ca2+oscillations in BON. UTP evoked a biphasic concentration-dependent Ca2+response. Cells responded in the order of UTP, ATP > UTPγS > UDP >> MRS2768, BzATP, α,β-MeATP > MRS2365, MRS2690, and NF546. Different proportions of cells activated by UTP and ATP also responded to UTPγS (P2Y4, 50% cells), UDP (P2Y6, 30%), UTPγS and UDP (14%) or MRS2768 (<3%). UTP Ca2+responses were blocked with inhibitors of PLC, IP3R, SERCA Ca2+pump, La3+sensitive Ca2+channels or chelation of intracellular free Ca2+ by BAPTA/AM. Inhibitors of L-type, TRPC, ryanodine-Ca2+pools, PI3-Kinase, PKC or SRC-Kinase had no effect. UTP stimulated voltage-sensitive Ca2+currents (ICa), Vm-depolarization and inhibited IK (not IA) currents. An IKv7.2/7.3 K+ channel blocker XE-991 mimicked UTP-induced Vm-depolarization and blocked UTP-responses. XE-991 blocked IK and UTP caused further reduction. La3+ or PLC inhibitors blocked UTP depolarization; PKC inhibitors, thapsigargin or zero Ca2+buffer did not. UTP stimulated 5-HT release in hEC expressing TPH1, 5-HT, P2Y4/P2Y6R. Zero-Ca2+buffer augmented Ca2+responses and 5-HT release. Conclusion: UTP activates a predominant P2Y4R pathway to trigger Ca2+oscillations via internal Ca2+mobilization through a PLC/IP3/IP3R/SERCA Ca2+signaling pathway to stimulate 5-HT release; Ca2+influx is inhibitory. UTP-induced Vm-depolarization depends on PLC signaling and an unidentified K channel (which appears independent of Ca2+oscillations or Ica/VOCC). UTP-gated signaling pathways triggered by activation of P2Y4R stimulate 5-HT release. [ABSTRACT FROM AUTHOR]

Recent years have seen the exploration of a puzzling number of compounds found in human diet that could be of interest for prevention or treatment of various pathologies. Although many of these natural products (NPs) have long been used as remedies, their molecular effects still remain elusive. With the advent of biotechnology revolution, NP studies turned from chemistry and biochemistry toward global analysis of gene expression. Hope is to use genetics to identify groups of patient for whom certain NPs or their derivatives may offer new preventive or therapeutic treatments. Recently, microRNAs have gained the statute of global regulators controlling cell homeostasis by regulating gene expression through genetic and epigenetic regulatory loops. Realization that certain plant polyphenols can modify microRNA expression and thus impact gene expression globally, initiated new, mainly in vitro studies, in particular to determine phytochemicals effects on inflammatory response, whose exacerbation has been linked to several disorders including cancer, auto-immune, metabolic, cardiovascular and neuro-inflammatory diseases. However, very few mechanistic insights have been provided, given the complexity of genetic regulatory networks implicated. In this review, we will concentrate on data showing the potential interest of some plant polyphenols in manipulating the expression of pro- and anti-inflammatory microRNAs in pathological conditions. [ABSTRACT FROM AUTHOR]

Lung cancer is the leading cause of cancer-related deaths worldwide. Despite advancements and improvements in surgical and medical treatments, the survival rate of lung cancer patients remains frustratingly poor. Local control for early-stage nonsmall cell lung cancer (NSCLC) has dramatically improved over the last decades for both operable and inoperable patients. However, the molecular mechanisms of NSCLC invasion leading to regional and distant disease spread remain poorly understood. Here, we identify microRNA-224 (miR-224) to be significantly up-regulated in NSCLC tissues, particularly in resected NSCLC metastasis. Increased miR-224 expression promotes cell migration, invasion, and proliferation by directly targeting the tumor suppressors TNFa-induced protein 1 (TNFAIP1) and SMAD4. In concordance with in vitro studies, mouse xenograft studies validated that miR-224 functions as a potent oncogenic miRNA in NSCLC in vivo. Moreover, we found promoter hypomethylation and activated ERK signaling to be involved in the regulation of miR-224 expression in NSCLC. Up-regulated miR-224, thus, facilitates tumor progression by shifting the equilibrium of the partially antagonist functions of SMAD4 and TNFAIP1 toward enhanced invasion and growth in NSCLC. Our findings indicate that targeting miR-224 could be effective in the treatment of certain lung cancer patients. [ABSTRACT FROM AUTHOR]

Background: The functions of long noncoding RNAs (lncRNAs) have been identified in several cancers, but the roles of lncRNAs in colorectal cancer (CRC) are less well understood. The transcription factor MYC is known to regulate lncRNAs and has been implicated in cancer cell proliferation and tumorigenesis. Methods: CRC cells and tissues were profiled to identify lncRNAs differentially expressed in CRC, from which we further selected MYC-regulated lncRNAs. We used luciferase promoter assay, ChIP, RNA pull-down assay, deletion mapping assay, LC-MS/MS and RNA immunoprecipitation to determine the mechanisms of MYC regulation of lncRNAs. Moreover, soft agar assay and in vivo xenograft experiments (four athymic nude mice per group) provided evidence of MYC-regulated lncRNAs in cancer cell transformation and tumorigenesis. The Kaplan-Meier method was used for survival analyses. All statistical tests were two-sided. Results: We identified lncRNAs differentially expressed in CRC (P < .05, greater than two-fold) and verified four lncRNAs upregulated and two downregulated in CRC cells and tissues. We further identified MYC-regulated lncRNAs, named MYCLos. The MYC-regulated MYCLos may function in cell proliferation and cell cycle by regulating MYC target genes such as CDKN1A (p21) and CDKN2B (p15), suggesting new regulatory mechanisms of MYC-repressed target genes through lncRNAs. RNA binding proteins including HuR and hnRNPK are involved in the function of MYCLos by interacting with MYCLo-1 and MYCLo-2, respectively. Knockdown experiments also showed that MYCLo-2, differentially expressed not only in CRC but also in prostate cancer, has a role in cancer transformation and tumorigenesis. Conclusions: Our results provide novel regulatory mechanisms in MYC function through lncRNAs and new potential lncRNA targets of CRC. [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]

It is now largely admitted that a pro-inflammatory environment may curtail anti-tumor immunity and favor cancer initiation and progression. The discovery that small non-coding regulatory RNAs, namely micro RNAs (mi RNAs), regulate all aspects of cell proliferation, differentiation, and function has shed a new light on regulatory mechanisms linking inflammation and cancer. Thus, mi RNAs such as miR-21, miR-125b, miR-155, miR-196, and miR-210 that are critical for the immune response or hypoxia are often overexpressed in cancers and leukemias. Given the high number of their target transcripts, their deregulation may have a number of deleterious consequences, depending on the cellular context. In this review, we focus on how the factors encoded by transcripts targeted by these five mi RNAs, be they transcription factors, tumor-suppressors, or regulators of different signaling pathways, can deregulate the immune response and favor pro-tumor immunity. Furthermore, we expose how the misdirected action of the main regulators of these mi RNAs, such as nuclear factor κB ( NF-κB), activator protein-1 ( AP-1), and signal transduction and activators of transcription ( STAT) transcription factors, or AKT and transforming growth factor β ( TGFβ) signaling pathways, can contribute to decrease anti-tumor immunity and enhance cell proliferation and oncogenesis. We conclude by briefly discussing about how these discoveries may possibly lead to the development of new mi RNA-based cancer therapies. [ABSTRACT FROM AUTHOR]

Tastant detection in the oral cavity involves selective receptors localized at the apical extremity of a subset of specialized taste bud cells called taste receptor cells (TRCs). The identification of the genes coding for the taste receptors involved in this process have greatly improved our understanding of the molecular mechanisms underlying detection. However, how these receptors signal in TRCs, and whether the components of the signaling cascades interact with each other or are organized in complexes is mostly unexplored. Here we report on the identification of three new binding partners for the mouse G protein gamma 13 subunit (Gγ13), a component of the bitter taste receptors signalling cascade. For two of these Gγ13 associated proteins, namely GOPC and MPDZ, we describe the expression in taste bud cells for the first time. Furthermore, we demonstrate by means of a yeast two-hybrid interaction assay that the C terminal PDZ binding motif of Gγ13 interacts with selected PDZ domains in these proteins. In the case of the PDZ domain-containing protein zona occludens-1 (ZO-1), a major component of the tight junction defining the boundary between the apical and baso-lateral region of TRCs, we identified the first PDZ domain as the site of strong interaction with Gγ13. This association was further confirmed by co-immunoprecipitation experiments in HEK 293 cells. In addition, we present immunohistological data supporting partial colocalization of GOPC, MPDZ or ZO-1 and Gγ13 in taste buds cells. Finally, we extend this observation to olfactory sensory neurons, another type of chemosensory cells known to express both ZO-1 and Gγ13. Taken together our results implicate these new interaction partners in the sub-cellular distribution of Gγ13 in olfactory and gustatory primary sensory cells. [ABSTRACT FROM AUTHOR]

MicroRNAs are short non-coding RNAs that posttranscriptionally modulate the expression of multiple target genes and are thus implicated in a wide array of cellular and developmental processes. miR-155 is processed from BIC, a non-coding transcript highly expressed in both activated B and T cells and in monocytes/macrophages. miR-155 levels change dynamically during both hematopoietic lineage differentiation and the course of the immune response. Different mouse models developed recently indicate that miR-155 plays a critical role during hematopoiesis and regulates lymphocyte homeostasis and tolerance. A moderate increase of miR-155 levels is observed in many types of malignancies of B cell or myeloid origin, and transgenic over-expression of miR-155 in mice results in cancer. While the high levels of miR-155 reached transiently during the course of the immune response remain unharmful for the organism, the reason why a moderate up-regulation of miR-155 can lead to cancer remains obscure. As prolonged exposure to inflammation can lead to cancer, the permanent up-regulation of miR-155 might be a link between the two. Therefore, designing miR-155 based therapies will require a better understanding of the molecular basis of its action as well as of how miR-155 levels are regulated in a cell-specific manner. [ABSTRACT FROM AUTHOR]

miRNAs are 19–24 nucleotide long noncoding RNAs found in almost all genetically dissected species, including viruses, plants, nematodes, flies, fish, mice and humans. Rapid advances have been made in understanding their physiological functions, while abnormal patterns of miRNA expression have been found in many disease states, most notably human cancer. It is now clear that miRNAs represent a class of genes with a great potential for use in diagnosis, prognosis and therapy. In this review we will focus on the discoveries that elucidate their crucial role in mammalian diseases, particularly in cancer, and propose that miRNA-based gene therapy might become the potential technology of choice in a wide range of human diseases including cancer. [ABSTRACT FROM AUTHOR]

Bax-induced lethality in yeast is accompanied by morphological changes in mitochondria, giving rise to a reduced number of swollen tubules. Although these changes are completely abolished upon coexpression of the Bax inhibitor, Bcl-2, coexpression of Bax with Bax inhibiting-glutathione S-transferase (BI-GST) leads to aggregation, but not fusion of the mitochondria. In addition, Bax affects the integrity of yeast vacuoles, resulting in the disintegration and eventual loss of the organelles, and the disruption of intracellular protein traffic. While Bcl-2 coexpression only partially corrects this phenotype, coexpression of BI-GST fully restores the organelles, indicating a different mode of protection exerted by Bcl-2 and BI-GST. [Copyright &y& Elsevier]