Abnormal Microvessels Research Articles

Vascularization ability of glioma stem cells in different three-dimensional microenvironments.

Glioblastoma (GBM) is among the most common and aggressive adult central nervous system tumors. One prominent characteristic of GBM is the presence of abnormal microvessels. A significant correlation between angiogenesis and prognosis has been observed. Accurately reconstructing this neovascularization and tumor microenvironment through personalized in vitro disease models presents a significant challenge. However, it is crucial to develop new anti-angiogenic therapies for GBM. In this study, 3D bioprinted glioma stem cell (GSC)-laden hydrogel scaffolds, hybrid GSC hydrogels and cell-free hydrogel scaffolds were manufactured to investigate the vascularization ability of GSCs in varying 3D microenvironments. Our results demonstrated that the bioactivity of GSCs in the 3D bioprinted GSC-laden hydrogel scaffold was preferable and stable, and the amounts of vascular endothelial growth factor A and basic fibroblast growth factor were the highest in the microenvironment. When the three different models were co-cultured with human umbilical vein endothelial cells, the expression of angiogenesis-related markers was the most abundant in the bioprinted GSC-laden hydrogel scaffold. Additionally, xenograft tumors formed by bioprinted GSC-laden hydrogel scaffolds more closely resembled human gliomas regarding color, texture and vascularization. Notably, in xenograft tumors derived from 3D bioprinted GSC-laden hydrogel scaffolds, the number of human CD105+ cells was significantly higher, and human endothelial vascular lumen-like structures were observed. This indicates that the 3D bioprinted GSC-laden hydrogel scaffold is a suitable model for mimicking the glioma microenvironment and studying tumor angiogenesis.

Diagnostic performance of endoscopy for subsquamous extension of superficial adenocarcinoma of the esophagogastric junction

Objectives Extension of adenocarcinoma of the esophagogastric junction under the squamous epithelium may lead to errors when determining lateral margins. However, the characteristics of subsquamous extension are unclear. Herein, we evaluated the prevalence and characteristics of subsquamous extension of adenocarcinoma of the esophagogastric junction and the diagnostic performance of endoscopy for this condition. Methods Eighty-nine consecutive patients with superficial adenocarcinoma of the esophagogastric junction who underwent endoscopic or surgical resection at a tertiary cancer center between January 2010 and December 2017 were retrospectively evaluated. Endoscopic subsquamous extension was defined as a submucosal tumor-like elevation covered by squamous epithelium and/or a brownish area with abnormal microvessels on the squamous epithelium observed using narrow-band imaging. The diagnostic performance of endoscopy for subsquamous extension was evaluated using histological subsquamous extension as gold standard. Results Thirty-nine patients (44%) had histological subsquamous extension. Proton pump inhibitor use was significantly associated with histological subsquamous extension [odds ratio: 4.65; 95% confidence interval (CI): 1.77–12.2]. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy of endoscopic subsquamous extension were 56% (95% CI: 40–72%), 96% (86–99%), 92% (73–99%), 74% (62–84%) and 79% (69–87%), respectively. The median length difference between histological and endoscopic subsquamous extension was 2 mm (range: −6 to 9 mm). Conclusions The sensitivity of endoscopic diagnosis of subsquamous extension was unsatisfactory. The endoscopic length of subsquamous extension tended to be underestimated. An oral safety margin of one centimeter is reasonable during endoscopic resection of adenocarcinoma of the esophagogastric junction. IMPACT STATEMENT This study will contribute significantly to the literature because this is the first study to determine the difference between the lengths of subsquamous extension detected endoscopically and histologically. This study determines the prevalence of subsquamous extension and identifies characteristics associated with subsquamous extension. An understanding of the risk of subsquamous extension is important when choosing a treatment strategy and planning the resection margins in patients with adenocarcinoma of the esophagogastric junction. This study provides patients with subsquamous extension characteristics and suggests a method for accurately diagnosing this condition.

Treatment of neovascularization in the eye by photo-mediated ultrasound therapy

Neovascularization in the eye is a common disorder. Abnormal microvessels can develop in the cornea, retina, or choroid of the eye, leading to severe vision loss if left untreated. Current treatment methods carry disadvantages or side effects, such as drug-resistance or damage to the surrounding healthy tissues. By applying synchronized laser pulses and ultrasound bursts, we developed a technique, termed photo-mediated ultrasound therapy (PUT), as a localized antivascular method based on the enhanced cavitation mechanism. PUT takes advantage of the high native optical contrast between blood and surrounding tissues, and has the unique capability to self-target microvessels without damaging surrounding tissues. The technique utilizes nanosecond laser pulses, which is spatially and temporally synchronized with low duty-cycle ultrasound bursts of sub-MHz frequencies at the targeted microvessels. We have tested the feasibility of PUT on the removal of neovascularization in the cornea, retina and choroid in a rabbit eye model. We found that neovascularization in the eye could be greatly reduced after PUT, and the occlusion of microvessels could last up to 4 weeks. In conclusion, PUT holds significant promises as a non-invasive method to precisely remove microvessels in neurovascular eye diseases by more selectively treating vasculature with minimized side-effects.

CALD1 Modulates Gliomas Progression via Facilitating Tumor Angiogenesis.

Simple SummaryCaldesmon has recently attracted attention in cancer due to its roles in cell migration, invasion and proliferation. l-CALD1 was also considered a potential serum marker for glioma. However, little is known about mechanisms underlying the effect of CALD1 on the microvascular facilitation and architecture in glioma. The purpose of this study was to explore the role of CALD1 for prediction glioma patient prognosis and in glioma angiogenesis. The findings of this study suggested that l-CALD1 could imply abnormal microvessels in anaplastic astrocytoma and GBM. In addition, high CI (calmodulin index) predicted worse prognosis in glioma, and furthermore, CALD1 may serve as a key marker for monitoring the progress of glioma and a novel target for therapy.Angiogenesis is more prominent in anaplastic gliomas and glioblastoma (GBM) than that in pilocytic and diffuse gliomas. Caldesmon (CALD1) plays roles in cell adhesion, cytoskeletal organization, and vascularization. However, limited information is available on mechanisms underlying the effect of CALD1 on the microvascular facilitation and architecture in glioma. In this study, we explored the role of CALD1 in gliomas by integrating bulk RNA-seq analysis and single cell RNA-seq analysis. A positive correlation between CALD1 expression and the gliomas’ pathological grade was noticed, according to the samples from the TCGA and CGGA database. Moreover, higher CALD1 expression samples showed worse clinical outcomes than lower CALD1 expression samples. Biofunction prediction suggested that CALD1 may affect glioma progression through modulating tumor angiogenesis. The map of the tumor microenvironment also depicted that more stromal cells, such as endothelial cells and pericytes, infiltrated in high CALD1 expression samples. CALD1 was found to be remarkably upregulated in neoplastic cells and was involved in tumorigenic processes of gliomas in single cell sequencing analysis. Histology and immunofluorescence analysis also indicated that CALD1 associates with vessel architecture, resulting in glioma grade progression. In conclusion, the present study implies that CALD1 may serve as putative marker monitoring the progress of glioma.

Real time SVD-based clutter filtering using randomized singular value decomposition and spatial downsampling for micro-vessel imaging on a Verasonics ultrasound system

Singular value decomposition (SVD)-based clutter filters can robustly reject the tissue clutter as compared with the conventional high pass filter-based clutter filters. However, the computational burden of SVD makes real time SVD-based clutter filtering challenging (e.g. frame rate at least 10–15 Hz with region of interest of about 4 × 4 cm2). Recently, we proposed an acceleration method based on randomized SVD (rSVD) clutter filtering and randomized spatial downsampling, which can significantly reduce the computational complexity without compromising the clutter rejection capability. However, this method has not been implemented on an ultrasound scanner and tested for its performance. In this study, we implement this acceleration method on a Verasonics scanner using a multi-core CPU architecture, and evaluate the selections of the imaging and processing parameters to enable real time micro-vessel imaging. The Blood-to-Clutter Ratio (BCR) performance was evaluated on a Verasonics machine with different settings of parameters such as block size and ensemble size. The demonstration of real time process was implemented on a 12-core CPU (downsampling factor of 12, 12-threads in this study) host computer. The processing time of the rSVD-based clutter filter was less than 30 ms and BCRs were higher than 20 dB as the block size, ensemble size and the rank of tissue clutter subspace were set as 30 × 30, 45 and 26 respectively. We also demonstrate that the micro-vessel imaging frame rate of the proposed architecture can reach approximately 22 Hz when the block size, ensemble size and the rank of tissue clutter subspace were set as 20 × 20 pixels, 45 and 26 respectively (using both images and supplementary videos). The proposed method may be important for real time 2D scanning of tumor microvessels in 3D to select and store the most representative 2D view with most abnormal micro-vessels for better diagnosis.

Hypoxia accelerates intraplaque neovascularization derived from endothelial progenitor cells in carotid stenosis.

The relationship between intraplaque hypoxia and intraplaque hemorrhage (IPH) has been reported, but the details remain obscure. In this study, the authors aimed to clarify the relationship among intraplaque hypoxia, endothelial progenitor cells (EPCs), and neovascularization, which causes IPH. The histological findings of specimens obtained from carotid endarterectomy were assessed. This study included 49 patients who underwent carotid endarterectomy. Magnetic resonance plaque imaging was performed to analyze the components of the carotid plaques, and surgical specimens were subjected to immunohistochemical analysis. The numbers of hypoxia-inducible factor-1 alpha (HIF-1α)-, CD34-, CD133-, and vascular endothelial growth factor receptor-2 (VEGFR-2)-positive cells in the carotid plaques were precisely quantified, as were the number and maximum diameter of CD31-positive microvessels. Plaque components were judged as fibrous in 7 samples, lipid-rich in 22, and IPH in 20. The number of CD34-, VEGFR-2-, and CD133-positive cells as an EPC-specific marker was significantly correlated with the number of HIF-1α-positive cells (r = 0.9, r = 0.82, and r = 0.81, respectively). These numbers varied among the 3 plaque components (IPH > lipid-rich > fibrous). The number and maximum luminal diameter of CD31-positive microvessels were also significantly correlated with the number of HIF-1α-positive cells (r = 0.85 and r = 0.89, respectively) and varied among the 3 plaque components (IPH > lipid-rich > fibrous). The present findings suggest that intraplaque hypoxia may accelerate abnormal microvessel formation derived from EPCs, which in turn promotes IPH. The results also suggest that microvessel enlargement is a pivotal characteristic of IPH and these enlarged microvessels are immature endothelial tubes with disorganized branching and are fragile and prone to rupture.

The correlation between chorioamnionitis-induced abnormal placental microvessel and platelet metabolism of preterm infants

Objective To observe the effect of chorioamnionitis on placental microvessel and platelet metabolism in premature and the correlation between them. Methods With clinical randomized controlled trial (RCT), the cases were matched by 11 according to gestational ages and divided into 2 groups according to the placental pathology results: chorioamnionitis group and control group, 32 cases in each group.Dates were obtained for preterm infants (gestational age<37 weeks) admitted to the Department of Neonatology at Guangdong Women and Children Hospital, born between June and December 2016.The platelet parameter [platelet count (PLT), mean platelet volume (MPV), platelet distribution width (PDW), plateletcrit (PCT)], birth weight, thrombocytopenia, hemorrhage complication, miscrovascular density (MVD) in placenta, platelet activating factors (CD62p, CD63) and thrombopoietin (TPO) in preterm infants were recorded and compared. Results In chorioamnionitis group, the infant birth weight[(1.90±0.41) kg vs.(2.31±0.62) kg] and the PLT in 72 hours [<24 h (197.97±63.43)×109/L vs.(266.34±69.92)×109/L; 24-72 h (202.28±29.70)×109/L vs.(256.38±69.96)×109/L] were significantly lower compared with the control group, and the differences were statistically significant(all P<0.05). The incidence of early thrombocytopenia(37.50% vs.9.38%), intracranial hemorrhage(40.62% vs.15.63%), MPV [(8.73±0.89) fL vs.(8.27±0.64) fL] and PDW[(59.46±5.90)% vs.(55.20±5.37)%] in 24 hours were significantly higher in chorioamnionitis group, and the differences were statistically significant (all P<0.05). The placental MVD in chorioamnionitis group significantly decreased[(9.08±1.35)% vs.(12.89±1.36)%, P<0.05 ]. The level of CD62p, CD63 and TPO in umbilical cord blood were significantly higher in chorioamnionitis group[(25.37±5.20)% vs.(10.35±2.94)%, (9.49±1.58)% vs.(4.04±1.21)%, (271.08±197.22) μg/L vs.(141.87±78.10) μg/L, all P<0.05]. The placental MVD was positively associated with PLT (r=0.74, P<0.05) and negatively associated with CD62p, CD63 and TPO among infants with chorioamnionitis (r=-0.64, -0.44, -0.44, all P<0.05). Conclusions The chorioamnionitis may decrease the MVD in placenta and activate platelet in fetal circulation, damaged placental microvessel may activate platelet further. Key words: Chorioamnionitis; Platelet; Platelet activation; Placenta; Infant, premature

Sa1256 ARTIFICIAL INTELLIGENCE USING CONVOLUTIONAL NEURAL NETWORK SHOWS HIGH DIAGNOSTIC PERFORMANCE OF MICROVESSELS ON SUPERFICIAL ESOPHAGEAL SQUAMOUS CELL CARCINOMA SIMILAR TO EXPERTS

The morphological diagnosis of microvessels on the surface of superficial esophageal squamous cell carcinoma (SESCC) with magnifying endoscopy using narrowband imaging (NBI-ME) is widely used in clinical practice. In particular, the presence of abnormal microvessels without a loop-like formation is very important, because it represents that the tumor invades muscularis mucosae (MM) or deeper, which is associated with lymph node metastasis. We constructed a convolutional neural network (CNN) system which diagnosed microvessels of SESCCs, and we evaluated the diagnostic performance of the system.

Fibulin-7 is overexpressed in glioblastomas and modulates glioblastoma neovascularization through interaction with angiopoietin-1.

Glioblastoma (GBM) is pathologically characterized by highly malignant neoplastic cells, focal necrosis and aberrant blood vessels composed of disorganized endothelial cells and pericytes. The recent cancer microarray database revealed upregulation of fibulin-7 (Fbln7), a member of the fibulin family, but provided no information on the tissue localization or biological function. In the present study, we demonstrated that Fbln7 is markedly overexpressed by the GBM tissue among astrocytic tumors, and immunolocalized mainly to endothelial cells and pericytes of the glomeruloid and hypertrophied microvessels. The production of Fbln7 by endothelial cells and pericytes was confirmed in cultured human umbilical vein endothelial cells (HUVEC) and human brain vascular pericytes (HBVP) and vascular endothelial growth factor (VEGF) stimulated the Fbln7 expression in HUVEC. Fbln7 bound to angiopoietin-1, but not angiopoietin-2 or Tie2 receptor, through interaction between the N-terminal portions of Fbln7 and angiopoietin-1, and it blocked phosphorylation of Tie2 receptor in HUVEC. In a coculture assay using HUVEC and HBVP, multilayered and irregular-shaped tube-like structures of HUVEC were induced by treatment with a high concentration of VEGF. This was accompanied by Fbln7 overproduction by HUVEC and angiopoietin-1 expression by HBVP. The production of aberrant VEGF-induced tube-like structures was attenuated by treatment with antibody or synthetic peptides specific to the Fbln7 N-terminal domain or knockdown of Fbln7. These data demonstrate that Fbln7 is overexpressed by endothelial cells and pericytes of the abnormal microvessels in GBM, and suggest that Fbln7 may contribute to the aberrant vessel formation by modulation of the angiopoietin-1/angiopoietin-2-Tie2 axis.

DCE-MRI and Quantitative Histology Reveal Enhanced Vessel Maturation but Impaired Perfusion and Increased Hypoxia in Bevacizumab-Treated Cervical Carcinoma

This study had a dual purpose: to investigate (1) whether bevacizumab can change the microvasculature and oxygenation of cervical carcinomas and (2) whether any changes can be detected with dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). Two patient-derived xenograft models of cervical cancer (BK-12 and HL-16) were included in the study. Immunostained histologic preparations from untreated and bevacizumab-treated tumors were analyzed with respect to microvascular density, vessel pericyte coverage, and tumor hypoxia using CD31, α-SMA, and pimonidazole as markers, respectively. DCE-MRI was performed at 7.05T, and parametric images of Ktrans and ve were derived from the data using the Tofts pharmacokinetic model. The tumors of both models showed decreased microvascular density, increased vessel pericyte coverage, and increased vessel maturation after bevacizumab treatment. Bevacizumab-treated tumors were more hypoxic and had lower Ktrans values than untreated tumors in the BK-12 model, whereas bevacizumab-treated and untreated HL-16 tumors had similar hypoxic fractions and similar Ktrans values. Significant correlations were found between median Ktrans and hypoxic fraction, and the data for untreated and bevacizumab-treated tumors were well fitted by the same curve in both tumor models. Bevacizumab-treated tumors show less abnormal microvessels than untreated tumors do, but because of treatment-induced vessel pruning, the overall function of the microvasculature might be impaired after bevacizumab treatment, resulting in increased tumor hypoxia. DCE-MRI has great potential for monitoring bevacizumab-induced changes in tumor hypoxia in cervical carcinoma.

Usefulness of narrow band imaging with magnifying endoscopy for the differential diagnosis of cancerous and noncancerous laryngeal lesions.

We evaluated the accuracy of endoscopic findings observed by narrow band imaging (NBI) combined with magnifying gastrointestinal endoscopy (GIE) for the differential diagnosis of cancerous and noncancerous laryngeal lesions. A total of 166 vocal cord lesions for which good images were obtained on NBI with magnifying GIE were evaluated with respect to the following 6 variables: macroscopic type, tumor location, color, white coat, keratinization, and abnormal microvessels. Multivariate analysis showed that white coat (odds ratio [OR], 2.95, P = 0.05), keratosis (OR, 3.14, P = 0.02) and abnormal microvessels (OR, 31.1, P < 0.0001) were significantly related to laryngeal cancer. In the diagnosis of laryngeal cancer, the sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of abnormal microvessels were 84.4%, 88.6%, 91%, 80.5%, and 86.1%, respectively. The abnormal microvessels on NBI combined with magnifying GIE are useful for the differential diagnosis of laryngeal lesions.

Inhibitory Effects On Retinal Neovascularization by Ranibizumab and sTie2-Fc in An Oxygen-Induced Retinopathy Mouse Model

ABSTRACTPurpose: To study retinal neovascularization (RNV) inhibition by intravitreal injections (IVs) of ranibizumab, sTie2 fusion protein (sTie2-Fc), and a combined therapy in an oxygen-induced retinopathy (OIR) model.Materials and methods: An OIR mouse model was used to simulate RNV in retinopathy of prematurity (ROP); and the effect of blocking the angiopoietin (Ang) and its receptor (Tie2) and the vascular endothelial growth factor (VEGF) and its receptor (VEGFR) signaling pathways was compared using an IV of sTie2-Fc (Ang inhibitor) and/or ranibizumab (aVEGF antagonist). The effects were assessed using fluorescein isothiocyanate (FITC)-dextran cardiac perfusion, isolectin B4 (IB4) staining with whole retinal mounting, and hematoxylin and eosin (HE) staining to count the endothelial cells (ECs) that broke through the internal limiting membrane (ILM). The mRNA and protein levels of VEGF-A, VEGFR-2, Ang1, Ang2, and Tie2 were also determined by reverse transcriptase polymerase chain reaction (RT-PCR) and western blot analysis.Results: Compared with the control group injected with phosphate-buffered saline (PBS), all three experimental groups, ranibizumab, sTie2-Fc, and ranibizumab + sTie2-Fc, had a significant decrease in micro-vessel densities and neovascular clusters, and fewer ECs broke through the ILM (all p < 0.05). The non-perfusion areas decreased in both mono-treated groups, although the combined therapy had larger non-perfusion areas. All three treatments decreased the mRNA and protein levels of VEGFA, Ang1, and Tie2.Conclusion: In this study, it was confirmed that blocking the Ang/Tie2 and/or VEGF/VEGFR pathways could inhibit RNV and decrease abnormal micro-vessel density; and the mono-blockage of Ang/Tie2 might cause a smaller non-perfusion area.

Challenges and opportunities of using stereotactic body radiotherapy with anti-angiogenesis agents in tumor therapy.

Microvessels promote proliferation of tumor cells by delivering oxygen and nutrients, but rapid growth of tumors results in unmet demands for oxygen and nutrients, thereby creating a hypoxia microenvironment. Under hypoxic conditions, vascular endothelial cells (ECs) initiate the formation of immature and abnormal microvasculature. This results in leakage and tortuosity that facilitates tumor cell invasion, metastasis and resistance to cytotoxic treatment. Radiotherapy (RT) is a vital tumor treatment modality. Currently, more than 60% of patients with malignant tumors receive RT at certain points during their treatment. Hypoxia induced by abnormal microvessels can hamper the cytotoxic effect of ionizing radiation, particularly, stereotactic body radiotherapy (SBRT). Anti-angiogenesis (AA) agents are known to reduce and renormalize microvessels in tumors, and hence alleviate hypoxia. The combination of AA agents with SBRT may have a synergistic role in inhibiting the growth of tumors. On the contrary, large doses of irradiation may affect tumor microvessels itself. In this review, we aim to clarify the relationship between SBRT and microvessel formation in tumors. In addition, we provide a retrospective analysis of the combination therapy involving SBRT and AA agents in preclinical and clinical practice to define its role in anti-tumor treatment.

A high-yield isolation and enrichment strategy for human lung microvascular endothelial cells.

Vasculopathies, characterized by the formation of fragile and abnormal microvessels, are associated with the severity of many chronic lung diseases, including pulmonary fibrosis, emphysema/chronic obstructive pulmonary disease, systemic sclerosis, and hypertension. However, the study of human lung vasculature has been limited by the ability to isolate generous quantities of microvascular endothelial cells (MVEC) free from mesenchymal contamination. Expansion and passaging of primary human MVEC in vitro typically results in loss of a traditional phenotype in favor of an intermediate mesenchymal one, as early as passage five. Here we provide a detailed protocol for the selection of large quantities of enriched primary human lung MVEC based upon differential adherence from mesenchyme and simple magnetic separation, which decreases the need for excessive passaging, in order to obtain sufficient cell numbers to successfully freeze stock cultures. Additional protocols are provided for Ac-di-LDL selection, characterization, and a sandwich angiogenesis method of functional tube formation. The complete protocol including cell isolation and characterization takes approximately six weeks to complete.

New Antiangiogenetic Therapy Beyond Bevacizumab in the Treatment of Advanced Non Small Cell Lung Cancer.

Lung cancer is the leading cause of death from cancer worldwide with limited available treatment options in absence of specific molecular alteration. New therapeutic approaches for addressing non small cell lung cancer (NSCLC) are urgently needed. Angiogenesis plays a central role in the tumor growth and metastatic dissemination which stimulates multiple cells to build new abnormal microvessels and leads to tumor microenvironment alterations. This process involves many factors, such as, the vascular endothelial growth factor (VEGF) that has a dominant role, the fibroblast growth factors (FGFs) and the plateled-derived growth factor (PDGF) that together contribute to resistance to VEGF/VEGFR- directed therapy. To date, bevacizumab is currently the only angiogenesis inhibitor that has been approved for the treatment of patients with advanced NSCLC in first-line setting. Moreover, in the last year, two new antiangiogenic agents have been approved for the treatment of patients with advanced NSCLC in second line setting. This review describes the new antiangiogenic agents in the treatment of advanced NSCLC.

Dual-modal ablation treatment for refractory radiation proctitis

Dear Editor: A 77-year-old male with recent history of radiation therapy for prostate cancer was referred with symptoms of fresh bleeding per rectum. Radiation proctitis (RP) was previously diagnosed on endoscopy at an outside facility. He had undergone three endoscopic ablation treatments with argon plasma coagulation (APC) but lower GI bleeding persisted, requiring multiple blood transfusions. On flexible sigmoidoscopy, multiple bleeding telangiectatic lesions with erythema and friability extending to 10 cm from the anal verge were noted, consistent with moderate grade radiation proctitis. Radiofrequency ablation (RFA) was offered as a rescue treatment. RFA was performed using the HALO 90 system (Covidien, Sunnyvale, CA), and the scope was rotated circumferentially to target all areas. Energy was delivered at settings of 12–15 J/cm and power density of 40 W/cm. At least two ablation sets were applied to each area. Post ablation, the coagulum was not scraped to promote hemostasis. The patient underwent a total of three RFA sessions at 4to 6-week intervals. His symptoms markedly improved with treatment sessions. On repeat endoscopy, the patient was found to have few spotty telangiectasias that were ablated with APC, 30 W, pulsed at 2 s (Erbe, Tubingen, Germany). On close follow-up, he was asymptomatic and hemoglobin levels were stable from prior. Rectal bleeding can occur from chronic radiation proctitis in up to 20 % of patients as a late complication following radiation therapy for pelvic malignancy. Ablation of the abnormal microvessels with subsequent re-epithelization is beneficial, and APC is currently offered as treatment of choice. However, in refractory cases, endoscopic RFA has recently been proposed as an alternative treatment for RP. RFA confers additional advantages over other modalities such as APC, as illustrated as follows. Firstly, RFA allows simultaneous treatment of broader areas of targeted tissue compared to APC. Also, RFA delivers a consistent amount of energy to the surface causing uniform coagulation of vessels achieving adequate hemostasis. Finally, by the tightly spaced bipolar array, RFA has limited energy penetration, which avoids energy dissipation and further injury to deeper tissue. The technical challenge was to maintain an optimal apposition to the targeted tissue applying torque and rotation of the scope with the electrode attached over the tip in a clockwise fashion, such that circumferential ablation to all areas was feasible. Targeting lesions close to the dentate line may be better using APC. This unique case exhibits a Bdual^ approach to achieve complete resolution of symptoms and bleeding related to chronic RP. In this proposed dual approach, RFA serves to deliver an initial ablative Bhit^ by significantly decreasing the burden of lesions, and APC is used to subsequently target the remaining point lesions. We believe that this combined approach also will prevent ulceration and long-term structuring compared to a single-mode RFA treatment. In conclusion, dual-modal ablation offers an ideal approach to management of RP, especially in refractory cases. * Kevin P. Cowley kcowley@uabmc.edu

Quantitative study on appearance of microvessels in spectral endoscopic imaging.

Increase in abnormal microvessels in the superficial mucosa is often relevant to diagnostic findings of neoplasia in digestive endoscopy; hence, observation of superficial vasculature is crucial for cancer diagnosis. To enhance the appearance of such vessels, several spectral endoscopic imaging techniques have been developed, such as narrow-band imaging and blue laser imaging. Both techniques exploit narrow-band blue light for the enhancement. The emergence of such spectral imaging techniques has increased the importance of understanding the relation of the light wavelength to the appearance of superficial vasculature, and thus a new method is desired for quantitative analysis of vessel visibility in relation to the actual structure in the tissue. Here, we developed microvessel-simulating phantoms that allowed quantitative evaluation of the appearance of 15-μm-thick vessels. We investigated the relation between the vascular contrast and light wavelength by the phantom measurements and also verified it in experiments with swine, where the endoscopically observed vascular contrast was investigated together with its real vascular depth and diameter obtained by microscopic observation of fluorescence-labeled vessels. Our study indicates that changing the spectral property even in the wavelength range of blue light may allow selective enhancement of the vascular depth for clinical use.

Gastrointestinal: Gastric mucosa‐associated lymphoid tissue (MALT) lymphoma observed by magnifying endoscopy with narrow band imaging

Mucosa-associated lymphoid tissue (MALT) lymphoma derived from the B-lymphocytes, rarely occurs in the gastrointestinal (GI) tract. The commonest site of occurrence is the stomach. Narrow band imaging (NBI) with magnifying endoscopy can identify MALT lymphoma and there have been several reports and case series on this. We presently report a 79-year-old man who underwent GI endoscopy as part of a health checkup. Conventional endoscopy showed a depressed reddish lesion in the posterior wall of the mid-gastric body (Figure 1A). Chromoendoscopy with indigo carmine identified this to be a depressed lesion. (Figure 1B) Magnifying endoscopy with NBI showed a cleare demarcation line of this depressed lesion (Figure 1C, arrows), and revealed an loss of the normal epithelium and abnormal micro-vessels that did not have the typical tree-branching calibre changes around white round lesions (Figure 1D). Endoscopic biopsy specimens taken from the lesion showed a diffuse proliferation of abnormal lymphoid cells within the mucosa (low-power histology—not shown). High-power histology showed a diffuse proliferation of small centrocyte-like cells and lymphoepithelial lesions. Immunohistochemical analysis was positive for CD20 but negative for CD3. He was diagnosed as having gastric MALT lymphoma. The positron-emission tomography/computed tomography showed only gastric uptake and no other extra-nodal disease. Based on the histopathological findings, a diagnosis of gastric MALT lymphoma (high-grade) was made, and combination-chemotherapy with pirarubicin hydrochloride, cyclophosphamide, vincristine sulfate and rituximab was started.

BENEFICIAL USE OF MAGNIFYING ENDOSCOPY WITH NARROW‐BAND IMAGING FOR DIAGNOSING A PATIENT WITH SQUAMOUS CELL CARCINOMA OF THE ANAL CANAL

The patient was a 74-year-old woman. She visited a dermatology clinic with a complaint of discomfort in the anal region. Erosion was observed in the anal region, and biopsies were performed. She was diagnosed with Bowen's disease and was referred to the dermatology department of our hospital for treatment. At our department, an endoscopic examination was performed for assessing the extent of Bowen's disease in the rectum. A retroflexed view of the anal canal revealed a slightly raised lesion with a faded color and an irregular surface. Narrow-band imaging (NBI) revealed a whitish lesion with a relatively clear margin and brown dots on the inside. Magnifying endoscopy with NBI revealed abnormal microvessels with dilatation, tortuosity, caliber change and various shapes, which were quite similar to the intrapapillary capillary loop patterns of superficial esophageal cancer. A complete transanal resection of the anal mucosa was performed thereafter. The patient was diagnosed with a well-differentiated squamous cell carcinoma on the basis of a pathological examination. An early diagnosis is critical for successful treatment of anal canal cancer. In this regard, magnifying endoscopy with NBI may be useful for determining the presence and extent of anal canal cancer.

Characterization of Targets of Plitidepsin In JAK2V617F-Mutated Cells From Myeloproliferative Neoplasms

AbstractAbstract 4093 Background:In a previous study (Verrucci M et al, J. Cell Physiol, 2010, Epub May10) we reported that the cyclic depsipeptide Aplidin®, a potent cytotoxic agent currently in phase II/III clinical trials for solid and hematologic neoplasia, displayed activity in a murine model of myelofibrosis, the GATA1-low mice. In fact, Aplidin improved the proliferation of Gata1-low hematopoietic cells, corrected abnormal microvessel density and reduced bone marrow fibrosis. These effects were largely attributed to improved maturation of megakaryocytes, as suggested by the increased platelet count in thrombocytopenic Gata1-low mice. It has been previously shown that a low expression of p27 in tumor cells might correlate with their response to Aplidin; of interest, GATA1-low megakaryocytes expressed reduced levels of p27. Overall, these results suggested that Aplidin could have the potential to alter the course of myelofibrosis-like disease in Gata1-low mice and could be useful for the treatment of myelofibrosis. Aims:to evaluate activity and targets of Aplidin in cellular models of myeloproliferative neoplasms (MPN). Methods:We measured the effect of Aplidin on the proliferation of JAK2V617F-mutated cell lines (UKE-1, HEL, SET2) and of primary cells from MPN patients in liquid cultures and semisolid medium, the rate of apoptosis using Annexin V flow cytometry analysis, and the cell cycle by propidium iodide staining. Expression of mRNA was quantified by real-time PCR, while protein level was measured by western blotting. Gene expression profiling was accomplished with Agilent Whole Human Genome Oligo Microarrays (44K). Results:Aplidin reduced the proliferation of all human V617F-mutated cell lines in the low nanomolar range, with IC50 from 0.5+/−0.03nM for UKE-1 to 1.5+/−0.05nM for HEL cells. Aplidin increased the proportion of cells in the G1/G0 phase of cell cycle, up to a mean of 80+/−5% from 60+/−3% in control cells (P<0.01). Also the proportion of apoptotic cells dose-dependently increased from 20+/−3% in control cultures to 50+/−6% at 10 nM in UKE-1 cells. After 24 h of incubation with Aplidin, the level of p27 mRNA was dose-dependently increased in all cell lines evaluated, while the BCR/ABL mutated K562 cells, whose proliferative rate was inhibited at higher Aplidin concentration, did not show significant changes of p27 mRNA. By western blotting analysis, level of p27 was overtly increased in UKE-1 cells treated with 5 and 10 nM Aplidin for 24h, and at the highest concentration we also observed reduction of phosphorylated STAT5. On the other hand, there was no change in JAK2, pJAK2, STAT5, STAT3, pSTAT3, Akt, pAkt. We then measured the level of p27 mRNA in peripheral blood cells obtained from patients with polycythemia vera (PV) or primary myelofibrosis (PMF). We found that PMF patients had significantly reduced p27 mRNA and protein level compared to controls and PV patients (P<.001 for both). After incubation with 1nM Aplidin there was a 2.5+/−2.0-fold increase in the p27 mRNA level and protein in cells of PMF patients while it was substantially unaffected in controls or PV patients. Gene expression analysis was performed in SET2 and UKE-1 cells that had been treated with 3nM of Aplidin for 24 hr. Aplidin treatment of SET2 cell line resulted in the up-regulation (UR) of 268 probsets and down-regulation (DR) of 247 (in total, about 1.5% of 40,961 genes called as “present”); corresponding figures in UKE-1 cells were 261 for UR (0.6%) and 364 (0.9%) for DR genes. MAPK signalling, TGF-beta signaling pathway, Cytokine-cytokine receptor interaction, cell adhesion molecules (CAMs), neuroactive ligand-receptor interaction, T cell receptor signaling, calcium signaling pathway, and regulation of actin cytoskeleton were the most involved pathways. Conclusions:Current results suggest that Aplidin has growth inhibitory activity and induces apoptosis in MPN cells. Response to Aplidin involves increasing p27 level, similar to findings in myelofibrotic GATA1-low mice; furthermore, we have identified low p27 expression as a characteristic of PMF cells compared to normal and PV cells. Finally, gene expression analysis allowed to identify a set of genes and involved pathways undergoing differential regulation in response to Aplidin. These information could be of value also for analyzing the response to Aplidin of PMF patients enrolled in an ongoing Phase II clinical trial. Disclosures:Aracil:PharmaMar: Employment. Vannucchi:PharmaMar: Research Funding.