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GBM, epithelioid type. a: H & E demonstrates a vascular lumen. b: ERG exclusively highlights the nuclei of endothelial cells. c: CD34 highlights not only endothelial cells, but also tumor cells. d: α-SMA highlights smooth muscle cells within the wall of a vascular channel (a-d: 200× magnification).
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ETS-related gene (ERG) is a transcription factor that has been linked to angiogenesis. Very little research has been done to assess ERG expression in central nervous system (CNS) tumors. We evaluated 57 CNS tumors, including glioblastomas (GBMs) and hemangioblastomas (HBs), as well as two arteriovenous malformations and four samples of normal brain...
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... all 15 GBMs, all 4 AAs, and the 2 OGs, we observed strong nuclear immunoreactiv- ity for ERG exclusively in endothelial cells lining vascular lumens (Figure 2b, 3b, 4b). For example, in the glomeruloid microvas- cular proliferation composed of hyperplastic vascular complexes adjacent to pseudopali- sading cells surrounding areas of necrosis, ERG was only detected in endothelial cells (Figure 3b). In contrast, α-SMA immunore- activity was detected within the abluminal cells of hyperplastic vessels in GBMs (Fig- ure 2e, 3e, 4d). In the 1 GBM case where microvascular proliferation was absent, en- dothelial cells were also highlighted by the ERG immunoreactivity. In the post-irradiat- ed GBM, secondary microvascular changes were present and with endothelial cells that were strongly reactive for the ERG immu- nostain. In GBMs, AAs, and OGs immuno- reactivity for CD31 and α-SMA was variable and sometimes weak or even absent within non-hyperplastic vascular channels ( Figure 2d, e, 3d, e, 4d), while immunoreactivity for CD34 was moderate (Figure 2c, 3c, 4c). In partially sclerosed vessels α-SMA immuno- reactivity was reduced, whereas ERG im- munoreactivity was present. In addition, in 1 GBM where ERG only stained endothelial cells (Figure 4b), CD34 stained both endo- thelial and tumor cells (Figure 4c). ERG- positive endothelial cells were seen at the invasive edge of all GBMs as ...
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... all 15 GBMs, all 4 AAs, and the 2 OGs, we observed strong nuclear immunoreactiv- ity for ERG exclusively in endothelial cells lining vascular lumens (Figure 2b, 3b, 4b). For example, in the glomeruloid microvas- cular proliferation composed of hyperplastic vascular complexes adjacent to pseudopali- sading cells surrounding areas of necrosis, ERG was only detected in endothelial cells (Figure 3b). In contrast, α-SMA immunore- activity was detected within the abluminal cells of hyperplastic vessels in GBMs (Fig- ure 2e, 3e, 4d). In the 1 GBM case where microvascular proliferation was absent, en- dothelial cells were also highlighted by the ERG immunoreactivity. In the post-irradiat- ed GBM, secondary microvascular changes were present and with endothelial cells that were strongly reactive for the ERG immu- nostain. In GBMs, AAs, and OGs immuno- reactivity for CD31 and α-SMA was variable and sometimes weak or even absent within non-hyperplastic vascular channels ( Figure 2d, e, 3d, e, 4d), while immunoreactivity for CD34 was moderate (Figure 2c, 3c, 4c). In partially sclerosed vessels α-SMA immuno- reactivity was reduced, whereas ERG im- munoreactivity was present. In addition, in 1 GBM where ERG only stained endothelial cells (Figure 4b), CD34 stained both endo- thelial and tumor cells (Figure 4c). ERG- positive endothelial cells were seen at the invasive edge of all GBMs as ...
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... like CD31, CD34 is affected by several drawbacks which should allow us to question the prevalence of its use as an endo- thelial cell marker. For instance, CD34-pos- itive immunostaining has also been reported in non-vascular cells within CNS tumors, including solitary fibrous tumor and ganglio- glioma [29,30,31], thus limiting the use of CD34 as a specific marker for endothelial cells. In addition, in 1 case of a GBM in our study, CD34 highlighted not only endothelial cells, but also tumor cells (Figure ...
Citations
... Measuring MVD is considered a standard method for the evaluation of tumor angiogenesis, moreover, the level of tumor angiogenesis based on MVD could potentially be valuable as a prognostic biomarker of tumors [12][13][14]. Multiple endothelial markers of immunohistochemistry (IHC) can be used to recognize the recently developed small blood vessels and differentiate them from the existing blood vessels [12][13][14][15]. Among the commonly used IHC markers are CD31, CD34, alpha smooth muscle actin (α-SMA), and erythroblastosis virus E26 oncogene homolog (ERG). ...
... ERG is an avian-vets member of the ETS family of transcription factors that plays a role in tumor angiogenesis and is effectively expressed in vascular endothelial cells, not stromal or tumor cells. Conversely, CD31 and CD34 exhibited weak to moderate reactivity [15]. Moreover, it has been demonstrated that α-SMA has effective expression in vessels and pericytes [12]. ...
The harmony between malignant cells and the adjacent microenvironment is a sophisticated subject; however, it seems to play an important role in cancer evolution. This study aimed to assess the microvascular density (MVD) and the mean pericyte number in the tumor and adjacent tissue, and to correlate the results with special histopathological prognostic variables of the tumor. The study included 48 colorectal cancer (CRC) cases diagnosed in the central lab of Duhok. The immunohistochemical (IHC) expressions of the mesenchymal and vascular dissemination markers, erythroblastosis virus E26 oncogene homolog (ERG, a member of the ETS family of transcription factors) and alpha smooth muscle actin (α-SMA) for microvascular density and pericytes, were assessed in tumor cells and in adjacent tissue around the tumor and then correlated to clinicopathological variables with a special concentration on inflammatory reaction, tumor budding, tumor deposition, and lymphovascular invasion. The results showed that the MVD was significantly higher outside the tumor in T1 and T2 compared with T3 and T4. Moreover, it was significantly higher in grade I when compared to grades II and III within the tumor. However, no correlation was found between the MVD and the special histopathological variables that had been studied. On the other hand, the low mean pericyte showed multiple significant associations outside tumor areas, with special histopathological features including a severe inflammatory reaction, a positive tumor deposit, and a negative lymphovascular invasion. These findings may indicate that defective or transformed pericytes around the tumor can participate in the development of the tumor and, subsequently, the outcome and prognosis.
... ERG is specifically expressed in endothelial cells and abundant in the lung and brain vasculature (56,57). Inflammatory cytokines TNF-a, LPS, and IL-1b decrease ERG expression in endothelial cells (5860). ...
Oxidative stress induces a prothrombotic state through enhancement of adhesion properties of the endothelium. E-selectin, an endothelial cell adhesion molecule, becomes a therapeutic target for venous thrombosis, whereas the regulatory mechanisms of its expression have not been fully understood. In the present study, we report that H2O2 treatment increases expression of E-selectin but decreases expression of the endothelial transcription factor ETS-related gene (ERG) in HUVECs in a dose- and time-dependent manner. In BALB/c mice treated with hypochlorous acid, E-selectin expression is increased and ERG expression is decreased in endothelial cells of the brain and lung. RNA interference of ERG upregulates E-selectin expression, whereas transfection of ERG-expressing plasmid downregulates E-selectin expression in HUVECs. Knockdown or overexpression of ERG comprises H2O2-induced E-selectin expression in HUVECs. Deletion of the Erg gene in mice results in embryonic lethality at embryonic days 10.5–12.5, and E-selectin expression is increased in the Erg−/− embryos. No chromatin loop was found on the E-selectin gene or its promoter region by capture high-throughput chromosome conformation capture. Chromatin immunoprecipitation and luciferase reporter assay determined that the −127 ERG binding motif mediates ERG-repressed E-selectin promoter activity. In addition, ERG decreases H2O2-induced monocyte adhesion. Together, ERG represses the E-selectin gene transcription and inhibits oxidative stress–induced endothelial cell adhesion.
... ETS proteins are nuclear DNA-binding phosphoproteins that act as activators or repressors of transcription [2]. ERG is expressed in endothelial cells (ECs) [3], particularly in the blood vessels surrounding the neural tube [4], heart vasculature, and the pre-cartilage [5]. It plays a key role in the regulation of tissue-specific processes such as haematopoiesis, vascular inflammation, and angiogenesis [6], by regulating the WNT/β-catenin signaling pathway and the transcriptional control of EC-specific genes including angiopoietin 2, a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 endoglin, von Willebrand factor (vWF), VEGF-A [7] and VE-cadherin [8]. ...
Ets-related gene (ERG) is overexpressed as a fusion protein in prostate cancer. During metastasis, the pathological role of ERG is associated with cell proliferation, invasion, and angiogenesis. Here, we hypothesized that miRNAs regulate ERG expression through its 3'UTR. Several bioinformatics tools were used to identify miRNAs and their binding sites on 3'UTR of ERG. The selected miRNAs expression was analyzed in prostate cancer samples by qPCR. The miRNAs overexpression was induced in prostate cancer cells (VCaP) to analyze ERG expression. Reporter gene assay was performed to evaluate the ERG activity in response to selected miRNAs. The expression of ERG downstream target genes was also investigated through qPCR after miRNAs overexpression. To observe the effects of selected miRNAs on cell proliferation and migration, scratch assay was performed to calculate the cell migration rate. miR-4482 and miR-3912 were selected from bioinformatics databases. miR-4482 and -3912 expression were decreased in prostate cancer samples, as compared to controls (p<0.05 and p<0.001), respectively. Overexpression of miR-4482 and miR-3912 significantly reduced ERG mRNA (p<0.001 and p<0.01), respectively) and protein (p<0.01) in prostate cancer cells. The transcriptional activity of ERG was significantly reduced (p<0.01) in response to miR-4482 and-3912. ERG angiogenic targets and cell migration rate was also reduced significantly (p<0.001) after miR-4482 and -3912 over-expression. This study indicates that miR-4482 and -3912 can suppress the ERG expression and its target genes, thereby, halt prostate cancer progression. These miRNAs may be employed as a potential therapeutic target for the miRNA-based therapy against prostate cancer.
... Interference of ERG expression reduces vWF expression and this effect is abolished by mutations of the -56 ETS-motif of the vWF promoter 92 . Although ERG is known for its role in endothelial homeostasis, aberrant expression of endothelial ERG has been observed in pathological conditions including vascular malignancies [93][94][95][96][97] , arterial calcification 98 , and tumour neovascularization 99,100 . In addition, overexpression of ERG in the Xenopus embryo results in developmental defects and ectopic endothelial differentiation 101 . ...
Vascular aging is a major contributing factor to cardiovascular disease. The aged blood vessels, characterized by vascular wall thickening and stiffening, are instigated by endothelial cell dysfunction induced by oxidative stress and inflammation. von Willebrand Factor (vWF) is a glycoprotein known for its role in coagulation, and plasma levels of vWF are increased with age. Elevated vWF promotes thrombosis, atherosclerotic plaque formation, inflammation and proliferation of vascular smooth muscle cells. Cadmium (Cd) is an environmental pollutant associated with increased morbidity and mortality of cardiovascular disease. At low concentrations, Cd activates pro-survival signaling in endothelial cells, however enhances intima-media thickness and atherogenesis. A non-cytotoxic dose of Cd also increases endothelial vWF expression and secretion in vivo and in vitro. In this review, we summarize the molecular mechanisms underlying vWF-promoted vascular aging-associated pathologies and Cd-induced vWF expression. In addition, we propose that exposure to low-dose Cd is a risk factor for vascular aging, through elevation of plasma vWF.
... Intriguingly, NOVA2 expression was restricted to the nuclei of ECs ( Figure 4B). Linear regression analysis showed a statistically significant correlation between the expression levels of NOVA2 and ETS-related gene (ERG) (R 2 = 0.5845, p < 0.001), a specific EC marker [84]. ...
... Intriguingly, NOVA2 expression was restricted to the nuclei of ECs ( Figure 4B). Linear regression analysis showed a statistically significant correlation between the expression levels of NOVA2 and ETS-related gene (ERG) (R² = 0.5845, p < 0.001), a specific EC marker [84]. The vessel-restricted expression of NOVA2 was confirmed in an independent cohort of GC patients ( Figure S6A-D). ...
Angiogenesis is crucial for cancer progression. While several anti-angiogenic drugs are in use for cancer treatment, their clinical benefits are unsatisfactory. Thus, a deeper understanding of the mechanisms sustaining cancer vessel growth is fundamental to identify novel biomarkers and therapeutic targets. Alternative splicing (AS) is an essential modifier of human proteome diversity. Nevertheless, AS contribution to tumor vasculature development is poorly known. The Neuro-Oncological Ventral Antigen 2 (NOVA2) is a critical AS regulator of angiogenesis and vascular development. NOVA2 is upregulated in tumor endothelial cells (ECs) of different cancers, thus representing a potential driver of tumor blood vessel aberrancies. Here, we identified novel AS transcripts generated upon NOVA2 upregulation in ECs, suggesting a pervasive role of NOVA2 in vascular biology. In addition, we report that NOVA2 is also upregulated in ECs of gastric cancer (GC), and its expression correlates with poor overall survival of GC patients. Finally, we found that the AS of the Rap Guanine Nucleotide Exchange Factor 6 (RapGEF6), a newly identified NOVA2 target, is altered in GC patients and associated with NOVA2 expression, tumor angiogenesis, and poor patient outcome. Our findings provide a better understanding of GC biology and suggest that AS might be exploited to identify novel biomarkers and therapeutics for anti-angiogenic GC treatments.
... Analysis of both mouse and human brain tissues has shown that high levels of Erg are nearly exclusively expressed in endothelial cells (Figs. 2 and 3). Antibody staining of both mouse and human tissues has confirmed the specificity of this transcription factor to endothelial cells within brain tissue, and shown that nearly all endothelial cells are positively stained [21][22][23] . While endothelial-to-mesenchymal-like transitions in endothelial cells in tumors have been linked to reduced Erg expression 24 , our own in vivo data does not suggest a loss of Erg transcript following activation in vivo or in vitro 25 . ...
Endothelial cells are important contributors to brain development, physiology, and disease. Although RNA sequencing has contributed to the understanding of brain endothelial cell diversity, bulk analysis and single-cell approaches have relied on fresh tissue digestion protocols for the isolation of single endothelial cells and flow cytometry-based sorting on surface markers or transgene expression. These approaches are limited in the analysis of the endothelium in human brain tissues, where fresh samples are difficult to obtain. Here, we developed an approach to examine endothelial RNA expression by using an endothelial-specific marker to isolate nuclei from abundant archived frozen brain tissues. We show that this approach rapidly and reliably extracts endothelial nuclei from frozen mouse brain samples, and importantly, from archived frozen human brain tissues. Furthermore, isolated RNA transcript levels are closely correlated with expression in whole cells from tissue digestion protocols and are enriched in endothelial markers and depleted of markers of other brain cell types. As high-quality RNA transcripts could be obtained from as few as 100 nuclei in archived frozen human brain tissues, we predict that this approach should be useful for both bulk analysis of endothelial RNA transcripts in human brain tissues as well as single-cell analysis of endothelial sub-populations.
Understanding of endothelial functions would be accelerated by methods for the specific isolation of these cells from archived human specimens. Here, the authors use the endothelial transcription factor Erg to isolate nuclei from mouse and human tissues, paving the way for high-throughput characterization of the function of endothelium in homeostasis and disease.
... ERG expression was randomly imputed using by drawing from a normal distribution with mean and standard deviation 0.199 ± 0.114 % positive area. [88]. Virtual biopsies were assumed to be IDH wild-type. ...
The overall aggressiveness of a glioma is measured by histologic and molecular analysis of tissue samples. However, the well-known spatial heterogeneity in gliomas limits the ability for clinicians to use that information to make spatially specific treatment decisions. Magnetic resonance imaging (MRI) visualizes and assesses the tumor. But, the exact degree to which MRI correlates with the actual underlying tissue characteristics is not known. In this work, we derive quantitative relationships between imaging and underlying pathology. These relations increase the value of MRI by allowing it to be a better surrogate for underlying pathology and they allow evaluation of the underlying biological heterogeneity via imaging. This provides an approach to answer questions about how tissue heterogeneity can affect prognosis. We estimated the local pathology within tumors using imaging data and stereotactically precise biopsy samples from an ongoing clinical imaging trial. From this data, we trained a random forest model to reliably predict tumor grade, proliferation, cellularity, and vascularity, representing tumor aggressiveness. We then made voxel-wise predictions to map the tumor heterogeneity and identify high-grade malignancy disease. Next, we used the previously trained models on a cohort of 1,850 glioma patients who previously underwent surgical resection. High contrast enhancement, proliferation, vascularity, and cellularity were associated with worse prognosis even after controlling for clinical factors. Patients that had substantial reduction in cellularity between preoperative and postoperative imaging (i.e. due to resection) also showed improved survival. We developed a clinically implementable model for predicting pathology and prognosis after surgery based on imaging. Results from imaging pathology correlations enhance our understanding of disease extent within glioma patients and the relationship between residual estimated pathology and outcome helps refine our knowledge of the interaction of tumor heterogeneity and prognosis.
... ETV2, an early regulator of vascular development, was found to be overexpressed in high-grade gliomas, and was reported to play a critical role in endothelial transdifferentiation of CD133+ GBM stem cells, which is thought to render them resistant to anti-angiogenic therapy [23]. Another ETS-related gene, ERG, was found to be a novel and highly specific marker for endothelial cells within CNS tumors, a feature that can be used in studying the vascularization of gliomas [24]. A transposon-based study of gliomagenesis identified friend leukemia integration 1 transcription factor (Fli1), among other genes, to be expressed in gliomas, although Fli1 expression is limited to a subset of glioma cells [25], and ETS protein PU.1, known for its critical role in hematopoietic development, was also reported to be highly expressed in glioma patients, indicating its role in the progression of glioma [26]. ...
The ETS domain family of transcription factors is involved in a number of biological processes, and is commonly misregulated in various forms of cancer. Using microarray datasets from patients with different grades of glioma, we have analyzed the expression profiles of various ETS genes, and have identified ETV1, ELK3, ETV4, ELF4, and ETV6 as novel biomarkers for the identification of different glioma grades. We have further analyzed the gene regulatory networks of ETS transcription factors and compared them to previous microarray studies, where Elk-1-VP16 or PEA3-VP16 were overexpressed in neuroblastoma cell lines, and we identify unique and common regulatory networks for these ETS proteins.
... ERG has been described in several tumors, such as prostatic carcinoma [18], acute myeloid leuke-mia [19], and Ewing sarcoma [20]. Haber [21] suggested that ERG could be a novel, reliable, and specific marker for endothelial cells in central nervous system (CNS) tumors. However, ERG expression in RCCs has not been studied to date and its value compared with other vascular endothelial (VE) markers has not been reported. ...
... As mentioned above, ERG is essential for postnatal vascular development and is involved in tumor angiogenesis and growth [17]. Haber [21] compared immunostaining for CD31, CD34, α-SMA, and ERG in CNS tumors and found that ERG was exclusively and strongly expressed in VE cells but not in stromal or tumor cells. Conversely, α-SMA was expressed in abluminal cells of hyperplastic vessels and CD31 and CD34 showed weak or moderate [11][12][13][14][15][16][17][18]. ...
... Conversely, α-SMA was expressed in abluminal cells of hyperplastic vessels and CD31 and CD34 showed weak or moderate [11][12][13][14][15][16][17][18]. The IRB waived the need for patient consent in this study because the folloreactivity [21]. The use of CD31 and CD34 as endothelial cell markers is limited because they are not only expressed in endothelial cells. ...
Objectives:
Renal cell carcinoma (RCC) accounts for approximately 90% of all renal malignancy. Because a rich vasculature is an outstanding feature of RCC, information on the blood vessels of RCC might explain its tumor characteristics. Several researchers have noted the effects of tumor vessels on the clinicopathologic characteristics and prognosis of tumors; however, a clear association has not been established. We hypothesized that the immaturity of the neovasculature may be an important clinicopathologic characteristic forprognosis of RCC patients. ERG and nestin are new vascular markers that regulate vascular homeostasis and angiogenesis. Therefore, in the present study, we investigated how ERG and nestin were expressed with respect to tumor characteristics.
Materials and methods:
IHC staining for ERG, nestin, CD31, and CD34 was performed for 217 renal tumors, including clear-cell RCC (ccRCC; n = 184), papillary RCC (pRCC; n = 14), chromophobe RCC (chRCC; n = 14), and oncocytoma (n = 5).
Results:
Vascular endothelial cells from normal kidney consistently showed strong nuclear expression of ERG and nestin. Conversely, a loss of ERG and nestin expression was observed in endothelial cells of some tumor blood vessels, which was associated with tumor progression. In particular, the loss of ERG expression was significantly associated with progression-free survival and overall survival (univariate analyses: P = 0.027 and P = 0.004, respectively; multivariate analyses: P = 0.030 and P = 0.046, respectively).
Conclusion:
A loss of ERG and nestin expression is associated with tumor progression, and loss of ERG is a powerful prognostic marker for ccRCC.
... For instance, it could be due to the fact that endothelial cells in different vessel types may express endothelial cell antigens heterogeneously (Pusztaszeri et al., 2006). Haber et al. (2015) reported that endothelial cells were only variably immunoreactive for CD31, while immunoreactivity for CD34 was more intense. Moreover, CD34 is expressed not only by endothelial cells, but also hematopoietic stem cells (Pusztaszeri et al., 2006). ...
To support survival and growth of follicles, the transplantable artificial ovary should mimic the original organ, offering a physical (3D matrix) and biological support (cells). In order to replicate the ovarian cell populations, the aim of this study is to assess the proportions of stromal and endothelial cells in the ovarian cortex. To this end, ovarian biopsies were obtained from six women (mean age: 49 years). The epithelial layer and medulla were carefully removed. The cortex was finely minced and enzymatically digested and the isolated cells were fixed. For cell characterization, immunostaining for CD31 (for endothelial cells) and inhibin-α (for granulosa cells) was performed. Positive cells in each staining were counted and the proportion of the different cell populations was estimated from the total number of isolated cells. Since there is no specific marker for ovarian stromal cells, we estimated the proportion of these cells by performing a vimentin immunostaining and subtracting the proportions of CD31- and inhibin-α-positive cells. Immunostaining showed that 84% of isolated cells were vimentin-positive. From this pool, 3% were endothelial cells and 1% granulosa cells. Consequently, the population of ovarian stromal cells was 80%. In conclusion, our findings show that stromal cells represent the larger population of cells in the human ovarian cortex. While this ensures follicle survival and development in a normal ovary, we believe that the low proportion of endothelial cells could have a negative impact on the angiogenesis in the artificial ovary after the first days of transplantation.
