Increased expression of histone demethylase JHDM1D under nutrient starvation suppresses tumor growth via down-regulating angiogenesis.
ABSTRACT: Histone demethylase JHDM1D (also known as KDM7A) modifies the level of methylation in histone and participates in epigenetic gene regulation; however, the role of JHDM1D in tumor progression is unknown. Here, we show that JHDM1D plays a tumor-suppressive role by regulating angiogenesis. Expression of JHDM1D was increased in mouse and human cancer cells under long-term nutrient starvation in vitro. Expression of JHDM1D mRNA was increased within avascular tumor tissue at the preangiogenic switch, along with increased expression of angiogenesis-regulating genes such as Vegf-A. Stable expression of JHDM1D cDNA or siRNA silencing of JHDM1D in cancer cells did not affect cell proliferation, anchorage-independent cell growth, or cell cycle progression in vitro. Notably, JHDM1D-expressing mouse melanoma (B16) and human cervical carcinoma (HeLa) cells exhibited significantly slower tumor growth in vivo compared with the original cells. This reduction in tumor growth was associated with decreased formation of CD31(+) blood vessels and reduced infiltration of CD11b(+) macrophage linage cells into tumor tissues. Expression of multiple angiogenic factors such as VEGF-B and angiopoietins was decreased in tumor xenografts of JHDM1D-expressing B16 and HeLa cells. Our results provide evidence that increased JHDM1D expression suppressed tumor growth by down-regulating angiogenesis under nutrient starvation.
Project description:Long noncoding RNAs play a pivotal role in tumor progression, but their role in cancer cells in the nutrient-starved tumor microenvironment remains unknown. Here, we show that a nutrient starvation-responsive long noncoding RNA, JHDM1D antisense 1 (JHDM1D-AS1), promotes tumorigenesis by regulating angiogenesis in response to nutrient starvation. Expression of JHDM1D-AS1 was increased in cancer cells. In addition, expression of JHDM1D-AS1 was increased in clinical tumor samples compared to that in normal tissue. Stable expression of JHDM1D-AS1 in human pancreatic cancer (PANC-1 and AsPC-1) cells promoted cell growth in vitro Remarkably, these JHDM1D-AS1-expressing cells showed a significant increase in tumor growth in vivo that was associated with increased formation of CD31+ blood vessels and elevated infiltration of CD11b+ macrophage lineage cells into tumor tissues. Genome-wide analysis of tumor xenografts revealed that expression of genes for tumor-derived angiogenic factors such as hHGF and hFGF1 concomitant with host-derived inflammation-responsive genes such as mMmp3, mMmp9, mS100a8, and mS100a9 was increased in tumor xenografts of JHDM1D-AS1-expressing pancreatic cancer cells, leading to a poor prognosis. Our results provide evidence that increased JHDM1D-AS1 expression under nutrient starvation accelerates tumor growth by upregulating angiogenesis, thus laying the foundation for improved therapeutic strategies.
Project description:JHDM1D antisense 1 (JHDM1D-AS1), a long non-coding RNA (lncRNA), has been shown to promote pancreatic cancer growth by inducing an angiogenic response. However, its biological and clinical significance in non-small-cell lung cancer (NSCLC) is still unclear. In this study, we examined the expression and prognostic significance of JHDM1D-AS1 in NSCLC. The effects of JHDM1D-AS1 knockdown or overexpression on NSCLC growth and metastasis were investigated. We show that JHDM1D-AS1 is upregulated in NSCLC relative to adjacent normal lung tissues. High JHDM1D-AS1 expression is significantly correlated with advanced tumor, node, and metastasis (TNM) stage and lymph node metastasis. JHDM1D-AS1 expression serves as an independent prognostic factor for overall survival of patients with NSCLC. Functionally, JHDM1D-AS1 knockdown inhibits NSCLC cell aggressiveness both in vitro and in vivo, which is rescued by ectopic expression of JHDM1D-AS1. JHDM1D-AS1 binding stabilizes DHX15 protein in NSCLC cells. DHX15 overexpression enhances NSCLC cell proliferation and invasion, whereas knockdown of DHX15 exerts opposite effects. JHDM1D-AS1-mediated aggressive phenotype is impaired when DHX15 is silenced. Ectopic expression of DHX15 restores the defects in proliferation and invasion of JHDM1D-AS1-depleted NSCLC cells. Collectively, the interaction between JHDM1D-AS1 and DHX15 accounts for NSCLC growth and metastasis. This work provides potential additional therapeutic targets for treatment of NSCLC.
Project description:Monocytes are key players in innate immunity, with their ability to regulate inflammatory responses and combat invading pathogens. There is a growing body of evidence indicating that long non-coding RNA (lncRNA) participate in various cellular biological processes, including the innate immune response. The immunoregulatory properties of numerous lncRNAs discovered in monocytes remain largely unexplored. Here, by RNA sequencing, we identified a lncRNA JHDM1D-AS1, which was upregulated in blood monocytes obtained from patients with sepsis relative to healthy controls. JHDM1D-AS1 expression was induced in primary human monocytes exposed to Toll-like receptor ligands, such as lipopolysaccharide (LPS), or bacteria. The inducibility of JHDM1D-AS1 expression in monocytes depended, at least in part, on nuclear factor–κB activation. JHDM1D-AS1 knockdown experiments in human monocyte-derived macrophages revealed significantly enhanced expression of inflammatory mediators, before and after exposure to LPS, relative to control cells. Specifically, genes involved in inflammatory responses were upregulated (e.g., CXCL2, CXCL8, IL1RN, TREM1, TNF, and IL6), whereas genes involved in anti-inflammatory pathways were downregulated (e.g., SOCS1 and IL10RA). JHDM1D-AS1 overexpression in a pro-monocytic cell line revealed diminished pro-inflammatory responses subsequent to LPS challenge. Collectively, these findings identify JHDM1D-AS1 as a potential anti-inflammatory mediator induced in response to inflammatory stimuli.
Project description:Although death receptor 6 (DR6) is aberrantly expressed in certain cancer cell lines, its function, signaling pathway and potential clinical significance in tumor progression are not well characterized. We report here that knocking down DR6 in the mouse B16 cell line has no effect on B16 cell death in vitro but suppresses xenograft B16 tumor growth by preventing tumor blood vessel formation in vivo. Deficiency of DR6 changes cytokine expression and secretion; in particular, it inhibits the proinflammatory cytokine interleukin-6 (IL-6), which is able to induce the expression of the angiogenesis-related factors: vascular endothelial growth factor-A, platelet-derived growth factor-?, vascular endothelial growth factor-D and platelet-derived growth factor receptor-?. Further experiments demonstrate that DR6-dependent angiogenesis is involved in the IL-6/P38 MAPK and IL-6/STAT3 pathways. Our novel findings demonstrate for the first time that DR6 expression in B16 cells facilitates tumor growth by accelerating tumor angiogenesis. Moreover, these results suggest that DR6 is involved in three important intracellular pathways that lead to homeostatic angiogenesis in tumor growth.
Project description:The Forkhead/Fox transcription factor Foxc2 is a critical regulator of vascular development. However, the role of Foxc2 in pathological angiogenesis in cancer remains unknown. Here we show that FoxC2 is highly expressed in human breast and colonic tumors and in the tumor endothelium in human and mouse melanomas. Using the B16 melanoma tumor model, we investigated the function of Foxc2 in tumor angiogenesis. After subcutaneous injection of B16 melanoma cells, primary tumor growth as well as neovascularization was markedly reduced in mice lacking one copy of the Foxc2 gene (Foxc2+/-). Consistently, expression levels of several angiogenic factors, including vascular endothelial growth factor (Vegf), matrix metallopeptidase 2 (Mmp2), and platelet-derived growth factor-B (Pdgfb), were significantly decreased in B16 tumors grown in Foxc2+/- mice, and tumor blood vessels formed in Foxc2+/- mice showed reduced coverage of mural cells and endothelial cell apoptosis. In addition, the tumor tissue in Foxc2+/- mice had an accumulation of necrotic cells. Taken together, these findings demonstrate that haplodeficiency of Foxc2 results in impaired formation of tumor blood vessels as well as reduced tumor growth and thereby provide evidence that Foxc2 is critical for tumor development and angiogenesis.
Project description:Tumor necrosis factor-alpha (TNF-?) binds to two distinct receptors, TNFR1/p55 and TNFR2/p75. TNF-? is implicated in the processes of tumor growth, survival, differentiation, invasion, metastases, secretion of cytokines and pro-angiogenic factors. We have shown that TNFR2/p75 signaling promotes ischemia-induced angiogenesis via modulation of several angiogenic growth factors. We hypothesized that TNFR2/p75 may promote tumor growth and angiogenesis. Growth of mouse Lewis lung carcinoma (LLC1) and/or mouse melanoma B16 cell was evaluated in wild type (WT), p75 knockout (KO) and double p55KO/p75KO mouse tumor xenograft models. Compared with WT and p55KO/p75KO mice, growth of tumors in p75KO mice was significantly decreased (twofold) in both LLC and B16 tumors. Tumor growth inhibition was correlated with decreases in vascular endothelial growth factor (VEGF) expression and capillary density, as well as bone marrow-derived endothelial progenitor cells incorporation into the functional capillary network, and an increase in apoptotic cells in LLC xenografts. Gene array analysis of tumor tissues showed a decrease in gene expression in pathways that promote tumor angiogenesis and cell survival. Blocking p75 by short-hairpin RNA in cultured LLCs led to increases in TNF-mediated apoptosis, as well as decreases in the constitutive and TNF-mediated expression of angiogenic growth factors (VEGF, HGF, PLGF), and SDF-1? receptor CXCR4. In summary, p75 is essential for tumor angiogenesis and survival in highly vascularized murine lung tumor xenografts. Blocking p75 expression may lead to tumor regression. This may represent new and effective therapy against lung neoplasms and potentially tumors of other origin.
Project description:The Ets transcription factors regulate a wide variety of biologic processes. Several members have been shown to play a role in regulating angiogenesis and vascular development. For example, the Ets factor ELF-1 is enriched in the developing vasculature of the embryo, where it regulates the expression of the Tie2 gene. We have determined that ELF-1 and Tie2 expression is also enriched in tumor blood vessels, and have identified a short peptide, 34 amino acids in length, corresponding to the terminal portion of the highly conserved ETS domain that potently blocks the function of ELF-1. A tailored ELF-1 blocking peptide, containing a 12-amino acid HIV-1 TAT protein, readily crosses the cell membrane and enters into the nucleus of endothelial cells, leading to a marked reduction in the expression of ELF-1 gene targets including Tie2 and endothelial nitric oxide synthase. Furthermore, the ELF-1 blocking peptide potently inhibits angiopoietin-1-mediated endothelial cell migration. Systemic administration of this peptide markedly attenuates B16 melanoma tumor growth and tumor-associated angiogenesis in nude mice. These results support the function of ELF-1 in the regulation of Tie2 gene expression during the development of tumor angiogenesis.
Project description:In established tumors, angiogenic endothelial cells (ECs) coexist next to "quiescent" EC in matured vessels. We hypothesized that angio-gene expression of B16.F10 melanoma would differ depending on the growth stage. Unraveling the spatiotemporal nature thereof is essential for drug regimen design aimed to affect multiple neovascularization stages. We determined the angiogenic phenotype-represented by 52 angio-genes-and vascular morphology of small, intermediate, and large s.c. growing mouse B16.F10 tumors and demonstrated that expression of these genes did not differ between the different growth stages. Yet vascular morphology changed dramatically from small vessels without lumen in small to larger vessels with increased lumen size in intermediate/large tumors. Separate analysis of these vascular morphologies revealed a significant difference in ?SMA expression in relation to vessel morphology, while no relation with VEGF, HIF-1?, nor Dll4 expression levels was observed. We conclude that the tumor vasculature remains actively engaged in angiogenesis during B16.F10 melanoma outgrowth and that the major change in tumor vascular morphology does not follow molecular concepts generated in other angiogenesis models.
Project description:<h4>Background</h4>Recent studies have suggested that adenosine generated by ecto-5'-nucleotidase (CD73) in the tumor microenvironment plays a major role in promoting tumor growth by suppressing the immune response and stimulating angiogenesis via A2A and A2B receptors. However, adenosine has also been reported to inhibit tumor growth acting via A1 and A3 receptors. Therefore the aim of this study was to clarify the role of host CD73, which catalyzes the extracellular hydrolysis of AMP to adenosine, on tumor growth and metastasis of B16-F10 melanoma cells.<h4>Methods</h4>CD73 and alkaline phosphatase (AP) activity of B16-F10 melanoma cells were measured by HPLC. Tumor cells were injected either subcutaneously or intradermally in WT and CD73-/- mice and tumor growth was monitored by MRI at 9.4 T. Immune cell subpopulations within tumors were assessed by FACS after enzymatic digestion. An endothelium specific CD73-/- was created using Tie2-Cre+ mice and CD73flox/flox (loxP) mice. Chimeric mice lacking CD73-/- on hematopoietic cells was generated by bone marrow transplantation. Lung metastatic spread was measured after intravenous B16-F10 application.<h4>Results</h4>B16-F10 cells showed very little CD73 and negligible AP activity. Neither complete loss of host CD73 nor specific knockout of CD73 on endothelial cells or hematopoietic cells affected tumor growth after subcutaneous or intradermal tumor cell application. Only peritumoral edema formation was significantly attenuated in global CD73-/- mice in the intradermal model. Immune cell composition revealed no differences in the different transgenic mice models. Also lung metastasis after intravenous B16-F10 injection was not altered in CD73-/- mice.<h4>Conclusions</h4>CD73 expression on host cells, particularly on endothelial and hematopoietic cells, does not modulate tumor growth and metastatic spread of B16-F10 melanoma cells most likely because of insufficient adenosine formation by the tumor itself.
Project description:The role of B cells in cancer and the underlying mechanisms remain to be further explored. Here, we show that tumor-associated B cells with activated STAT3 contribute to tumor development by promoting tumor angiogenesis. B cells with or without Stat3 have opposite effects on tumor growth and tumor angiogenesis in both B16 melanoma and Lewis Lung Cancer mouse models. Ex vivo angiogenesis assays show that B cell-mediated tumor angiogenesis is mainly dependent on the induction of pro-angiogenic gene expression, which requires Stat3 signaling in B cells. Furthermore, B cells with activated STAT3 are mainly found in or near tumor vasculature and correlate significantly with overall STAT3 activity in human tumors. Moreover, the density of B cells in human tumor tissues correlates significantly with expression levels of several STAT3-downstream pro-angiogenic genes, as well as the degree of tumor angiogenesis. Together, these findings define a novel role of B cells in promoting tumor progression through angiogenesis and identify STAT3 in B cells as potential therapeutic target for anti-angiogenesis therapy.