Expression and protein localisation of IGF2 in the marsupial placenta

<p>Abstract</p> <p>Background</p> <p>In eutherian mammals, genomic imprinting is critical for normal placentation and embryo survival. <it>Insulin-like growth factor 2 </it>(<it>IGF2</it>) is imprinted in the placenta of both eutherians and marsupials, but its function, or that of any imprinted gene, has not been investigated in any marsupial. This study examines the role of <it>IGF2 </it>in the yolk sac placenta of the tammar wallaby, <it>Macropus eugenii</it>.</p> <p>Results</p> <p><it>IGF2 </it>mRNA and protein were produced in the marsupial placenta. Both IGF2 receptors were present in the placenta, and presumably mediate IGF2 mitogenic actions. <it>IGF2 </it>mRNA levels were highest in the vascular region of the yolk sac placenta. IGF2 increased <it>vascular endothelial growth factor </it>expression in placental explant cultures, suggesting that IGF2 promotes vascularisation of the yolk sac.</p> <p>Conclusion</p> <p>This is the first demonstration of a physiological role for any imprinted gene in marsupial placentation. The conserved imprinting of <it>IGF2</it> in this marsupial and in all eutherian species so far investigated, but not in monotremes, suggests that imprinting of this gene may have originated in the placenta of the therian ancestor.</p

Targeting the angiotensin II type 2 receptor (AT2R) in colorectal liver metastases

BACKGROUND: Blockade of the angiotensin (ANG) II type 1 receptor (AT1R) inhibits tumour growth in several cancers, including colorectal cancer (CRC) liver metastases. While AT1R blockade has been extensively studied, the potential of targeting the antagonistically acting AT2R in cancer has not been investigated. This study examined the effect of AT2R activation with the agonist CGP42112A in a mouse model of CRC liver metastases. RESULTS: In vitro, mouse CRC cell (MoCR) proliferation was inhibited by treatment with CGP42112A in a dose dependent manner while apoptosis was increased. Immunofluorescent staining for key signalling and secondary messengers, PLA2 and iNOS, were also increased by CGP42112A treatment in vitro. Immunohistochemical staining for proliferation (PCNA) and the apoptosis (active caspase 3) markers confirmed a CGP42112A-associated inhibition of proliferation and induction of apoptosis of mouse CRC cells (MoCR) in vivo. However, angiogenesis and vascular endothelial growth factor (VEGF) appeared to be increased by CGP42112A treatment in vivo. This increase in VEGF secretion by MoCRs was confirmed in vitro. Despite this apparent pro-angiogenic effect, a syngenic orthotopic mouse model of CRC liver metastases showed a reduction in liver to body weight ratio, an indication of tumour burden, following CGP42112A treatment compared to untreated controls. CONCLUSIONS: These results suggest that AT2R activation might provide a novel target to inhibit tumour growth. Its potential to stimulate angiogenesis could be compensated by combination with anti-angiogenic agents

Evolution of the CDKN1C-KCNQ1 imprinted domain

<p>Abstract</p> <p>Background</p> <p>Genomic imprinting occurs in both marsupial and eutherian mammals. The <it>CDKN1C </it>and <it>IGF2 </it>genes are both imprinted and syntenic in the mouse and human, but in marsupials only <it>IGF2 </it>is imprinted. This study examines the evolution of features that, in eutherians, regulate <it>CDKN1C </it>imprinting.</p> <p>Results</p> <p>Despite the absence of imprinting, CDKN1C protein was present in the tammar wallaby placenta. Genomic analysis of the tammar region confirmed that <it>CDKN1C </it>is syntenic with <it>IGF2</it>. However, there are fewer LTR and DNA elements in the region and in intron 9 of <it>KCNQ1</it>. In addition there are fewer LINEs in the tammar compared with human and mouse. While the CpG island in intron 10 of <it>KCNQ1 </it>and promoter elements could not be detected, the antisense transcript <it>KCNQ1OT1 </it>that regulates <it>CDKN1C </it>imprinting in human and mouse is still expressed.</p> <p>Conclusion</p> <p>CDKN1C has a conserved function, likely antagonistic to IGF2, in the mammalian placenta that preceded its acquisition of imprinting. CDKN1C resides in synteny with IGF2, demonstrating that imprinting of the two genes did not occur concurrently to balance maternal and paternal influences on the growth of the placenta. The expression of <it>KCNQ1OT1 </it>in the absence of CDKN1C imprinting suggests that antisense transcription at this locus preceded imprinting of this domain. These findings demonstrate the stepwise accumulation of control mechanisms within imprinted domains and show that <it>CDKN1C </it>imprinting cannot be due to its synteny with <it>IGF2 </it>or with its placental expression in mammals.</p

Analysis of the platypus genome suggests a transposon origin for mammalian imprinting

Comparisons between the platypus and eutherian mammalian genomes provides new insights into how epigenetic imprinting may have evolved in mammalian genomes

Recent Assembly of an Imprinted Domain from Non-Imprinted Components

Genomic imprinting, representing parent-specific expression of alleles at a locus, raises many questions about how—and especially why—epigenetic silencing of mammalian genes evolved. We present the first in-depth study of how a human imprinted domain evolved, analyzing a domain containing several imprinted genes that are involved in human disease. Using comparisons of orthologous genes in humans, marsupials, and the platypus, we discovered that the Prader-Willi/Angelman syndrome region on human Chromosome 15q was assembled only recently (105–180 million years ago). This imprinted domain arose after a region bearing UBE3A (Angelman syndrome) fused with an unlinked region bearing SNRPN (Prader-Willi syndrome), which had duplicated from the non-imprinted SNRPB/B′. This region independently acquired several retroposed gene copies and arrays of small nucleolar RNAs from different parts of the genome. In their original configurations, SNRPN and UBE3A are expressed from both alleles, implying that acquisition of imprinting occurred after their rearrangement and required the evolution of a control locus. Thus, the evolution of imprinting in viviparous mammals is ongoing

Blockade of MMP14 Activity in Murine Breast Carcinomas: Implications for Macrophages, Vessels, and Radiotherapy

Background: Matrix metalloproteinase (MMP) 14 may mediate tumor progression through vascular and immune-modulatory effects. Methods: Orthotopic murine breast tumors (4T1 and E0771 with high and low MMP14 expression, respectively; n = 5-10 per group) were treated with an anti-MMP14 inhibitory antibody (DX-2400), IgG control, fractionated radiation therapy, or their combination. We assessed primary tumor growth, transforming growth factor β (TGFβ) and inducible nitric oxide synthase (iNOS) expression, macrophage phenotype, and vascular parameters. A linear mixed model with repeated observations, with Mann-Whitney or analysis of variance with Bonferroni post hoc adjustment, was used to determine statistical significance. All statistical tests were two-sided. Results: DX-2400 inhibited tumor growth compared with IgG control treatment, increased macrophage numbers, and shifted the macrophage phenotype towards antitumor M1-like. These effects were associated with a reduction in active TGFβ and SMAD2/3 signaling. DX-2400 also transiently increased iNOS expression and tumor perfusion, reduced tissue hypoxia (median % area: control, 20.2%, interquartile range (IQR) = 6.4%-38.9%; DX-2400: 1.2%, IQR = 0.2%-3.2%, P = .044), and synergistically enhanced radiation therapy (days to grow to 800mm3: control, 12 days, IQR = 9-13 days; DX-2400 plus radiation, 29 days, IQR = 26-30 days, P < .001) in the 4T1 model. The selective iNOS inhibitor, 1400W, abolished the effects of DX-2400 on vessel perfusion and radiotherapy. On the other hand, DX-2400 was not capable of inducing iNOS expression or synergizing with radiation in E0771 tumors. Conclusion: MMP14 blockade decreased immunosuppressive TGFβ, polarized macrophages to an antitumor phenotype, increased iNOS, and improved tumor perfusion, resulting in reduced primary tumor growth and enhanced response to radiation therapy, especially in high MMP14-expressing tumor

Tetraspanin CD53 promotes lymphocyte recirculation by stablising L-selectin surface expression

Tetraspanins regulate key processes in immune cells; however, the function of the leukocyterestricted tetraspanin, CD53 has remained unknown. Here we show that CD53 is essential for lymphocyte recirculation. Lymph nodes of Cd53-/- mice were smaller than wild-type mice due to a marked reduction in B cells and a 50% decrease in T cells. This reduced cellularity reflected an inability of Cd53-/- B and T cells to efficiently home to lymph nodes, due to the near absence of L-selectin from Cd53-/- B cells and reduced stability of L-selectin on Cd53-/- T cells. Further analyses, including on human lymphocytes, showed that CD53 inhibits L-selectin shedding via both ADAM17-dependent and -independent mechanisms. The disruption in lymphocyte recirculation in Cd53-/- mice led to impaired immune responses dependent on antigen delivery to lymph nodes. Together these findings demonstrate a previously unrecognized essential role for CD53 in lymphocyte trafficking and immune responses

The IDENTIFY study: the investigation and detection of urological neoplasia in patients referred with suspected urinary tract cancer - a multicentre observational study

Objective To evaluate the contemporary prevalence of urinary tract cancer (bladder cancer, upper tract urothelial cancer [UTUC] and renal cancer) in patients referred to secondary care with haematuria, adjusted for established patient risk markers and geographical variation. Patients and Methods This was an international multicentre prospective observational study. We included patients aged ≥16 years, referred to secondary care with suspected urinary tract cancer. Patients with a known or previous urological malignancy were excluded. We estimated the prevalence of bladder cancer, UTUC, renal cancer and prostate cancer; stratified by age, type of haematuria, sex, and smoking. We used a multivariable mixed-effects logistic regression to adjust cancer prevalence for age, type of haematuria, sex, smoking, hospitals, and countries. Results Of the 11 059 patients assessed for eligibility, 10 896 were included from 110 hospitals across 26 countries. The overall adjusted cancer prevalence (n = 2257) was 28.2% (95% confidence interval [CI] 22.3–34.1), bladder cancer (n = 1951) 24.7% (95% CI 19.1–30.2), UTUC (n = 128) 1.14% (95% CI 0.77–1.52), renal cancer (n = 107) 1.05% (95% CI 0.80–1.29), and prostate cancer (n = 124) 1.75% (95% CI 1.32–2.18). The odds ratios for patient risk markers in the model for all cancers were: age 1.04 (95% CI 1.03–1.05; P < 0.001), visible haematuria 3.47 (95% CI 2.90–4.15; P < 0.001), male sex 1.30 (95% CI 1.14–1.50; P < 0.001), and smoking 2.70 (95% CI 2.30–3.18; P < 0.001). Conclusions A better understanding of cancer prevalence across an international population is required to inform clinical guidelines. We are the first to report urinary tract cancer prevalence across an international population in patients referred to secondary care, adjusted for patient risk markers and geographical variation. Bladder cancer was the most prevalent disease. Visible haematuria was the strongest predictor for urinary tract cancer