AXL-Initiated Paracrine Activation of pSTAT3 Enhances Mesenchymal and Vasculogenic Supportive Features of Tumor-Associated Macrophages

Tumor-associated macrophages (TAMs) are integral to the development of complex tumor microenvironments (TMEs) and can execute disparate cellular programs in response to extracellular cues. However, upstream signaling processes underpinning this phenotypic plasticity remain to be elucidated. Here, we report that concordant AXL-STAT3 signaling in TAMs is triggered by lung cancer cells or cancer-associated fibroblasts in the cytokine milieu. This paracrine action drives TAM differentiation toward a tumor-promoting M2-like phenotype with upregulation of CD163 and putative mesenchymal markers, contributing to TAM heterogeneity and diverse cellular functions. One of the upregulated markers, CD44, mediated by AXL-IL-11-pSTAT3 signaling cascade, enhances macrophage ability to interact with endothelial cells and facilitate formation of primitive vascular networks. We also found that AXL-STAT3 inhibition can impede the recruitment of TAMs in a xenograft mouse model, thereby suppressing tumor growth. These findings suggest the potential application of AXL-STAT3-related markers to quantitatively assess metastatic potential and inform therapeutic strategies in lung cancer

Tandem Repeat Hypothesis in Imprinting: Deletion of a Conserved Direct Repeat Element Upstream of H19 Has No Effect on Imprinting in the Igf2-H19 Region

Igf2 and H19 are reciprocally imprinted genes on mouse distal chromosome 7. They share several regulatory elements, including a differentially methylated region (DMR) upstream of H19 that is paternally methylated throughout development. The cis-acting sequence requirements for targeting DNA methylation to the DMR remain unknown; however, it has been suggested that direct tandem repeats near DMRs could be involved. Previous studies of the imprinted Rasgrf1 locus demonstrate indeed that a direct repeat element adjacent to a DMR is responsible for establishing paternal allele-specific methylation at the DMR and therefore allelic expression of the Rasgrf1 transcript. We identified a prominent and conserved direct tandem repeat 1 kb upstream of the H19 DMR and proposed that it played a similar role in imprinted regulation of H19. To test our hypothesis, we generated mice harboring a 1.7-kb targeted deletion of the direct repeat element and analyzed fetal growth, allelic expression, and methylation within the Igf2-H19 region. Surprisingly the deletion had no effect on imprinting. These results together with deletions of other repeats close to imprinted genes suggest that direct repeats may not be important for the targeting of methylation at the majority of imprinted loci and that the Rasgrf1 locus may be an exception to this rule

Epigenetic asymmetry in the mammalian zygote and early embryo: relationship to lineage commitment?

Epigenetic asymmetry between parental genomes and embryonic lineages exists at the earliest stages of mammalian development. The maternal genome in the zygote is highly methylated in both its DNA and its histones and most imprinted genes have maternal germline methylation imprints. The paternal genome is rapidly remodelled with protamine removal, addition of acetylated histones, and rapid demethylation of DNA before replication. A minority of imprinted genes have paternal germline methylation imprints. Methylation and chromatin reprogramming continues during cleavage divisions, but at the blastocyst stage lineage commitment to inner cell mass (ICM) or trophectoderm (TE) fate is accompanied by a dramatic increase in DNA and histone methylation, predominantly in the ICM. This may set up major epigenetic differences between embryonic and extraembryonic tissues, including in X-chromosome inactivation and perhaps imprinting. Maintaining epigenetic asymmetry appears important for development as asymmetry is lost in cloned embryos, most of which have developmental defects, and in particular an imbalance between extraembryonic and embryonic tissue development

Alterations in the placental methylome with maternal obesity and evidence for metabolic regulation

<div><p>The inflammatory and metabolic derangements of obesity in pregnant women generate an adverse intrauterine environment, increase pregnancy complications and adverse fetal outcomes and program the fetus for obesity and metabolic syndrome in later life. We hypothesized that epigenetic modifications in placenta including altered DNA methylation/hydroxymethylation may mediate these effects. Term placental villous tissue was collected following cesarean section from lean (prepregnancy BMI<25) or obese (BMI>30) women. Genomic DNA was isolated, methylated and hydroxymethylated DNA immunoprecipitated and hybridized to the NimbleGen 2.1M human DNA methylation array. Intermediate metabolites in placental tissues were measured by HPLC-ESI-MS, ascorbate levels by reverse phase HPLC and gene expression by RT-PCR. Differentially methylated and hydroxymethylated regions occurred across the genome, with a 21% increase in methylated but a 31% decrease in hydroxymethylated regions in obese vs lean groups. Whereas increased methylation and decreased methylation was evident around transcription start sites of multiple genes in the <i>GH/CSH</i> and <i>PSG</i> gene clusters on chromosomes 17 and 19 in other areas there was no relationship. Increased methylation was associated with decreased expression only for some genes in these clusters. Biological pathway analysis revealed the 262 genes which showed reciprocal differential methylation/ hydroxymethylation were enriched for pregnancy, immune response and cell adhesion-linked processes. We found a negative relationship for maternal BMI but a positive relationship for ascorbate with α-ketoglutarate a metabolite that regulates ten eleven translocase (TET) which mediates DNA methylation. We provide evidence for the obese maternal metabolic milieu being linked to an altered DNA methylome that may affect placental gene expression in relation to adverse outcomes.</p></div

Relationship between maternal adiposity, intermediary metabolites and placenta-specific gene expression.

<p>(A-C) Correlations between placental levels of αKG and maternal BMI (A) or ascorbate (B), and between αKG and <i>CSHL1</i> mRNA levels (C). The relations between continuous variables were evaluated by Spearman correlations (*<i>P</i> < 0.05). Pre-pregnancy or first trimester BMI was used as maternal early BMI.</p