Confocal Laser Endomicroscopy for the Morphometric Evaluation of Microvessels in Human Colorectal Cancer Using Targeted Anti-CD31 Antibodies

Cârţână, Tatiana; Săftoiu, Adrian; Gruionu, Lucian Gheorghe; Gheonea, Dan Ionuţ; Pirici, Daniel; Georgescu, Claudia Valentina; Ciocalteu, Adriana; Gruionu, Gabriel

doi:10.1371/journal.pone.0052815

Cited by 15 publications

(17 citation statements)

References 22 publications

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“…CD31 is a pan-endothelial marker for small and large vessels (29). In order to assess the MVD, CD31 and associated antibodies are used to mark tumor vascular endothelial cells and the capillary number per unit area is counted (30). In the present study, immunohistochemical staining for CD31 revealed that the average MVD in the pLXSN-Tum-5 group was significantly lower compared with that in the pLXSN and wild-type H22 HCC groups.…”

Section: Discussionsupporting

confidence: 47%

Vector-mediated Tum-5 expression in neovascular endothelial cells for treating hepatocellular carcinoma

Guan

Sun

et al. 2017

Experimental and Therapeutic Medicine

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Abstract. Hypervascular hepatocellular carcinoma (HCC) is one of the leading causes of cancer-associated mortality. Angiogenesis is an important contributor to HCC progression and metastasis; therefore, inhibiting angiogenesis may be an effective method of treating HCC. Tumstatin is a novel type of efficient endogenous vascular endothelial cell growth inhibiting factor. The anti-angiogenic activity of tumstatin is localized to the 54-132 amino acid region (Tum-5). In a previous study performed by our group, the gene fragment encoding Tum-5 was cloned and inserted into a pLXSN retroviral vector. In the present study, the anti-angiogenic effects of Tum-5 and the antitumor effects exerted by the pLXSN-Tum-5 vector in vivo were investigated. The results demonstrated that pLXSN-Tum-5 significantly inhibited the growth of human umbilical vein endothelial cells compared with pLXSN, but had no obvious effect on HepG2 cell growth. Moreover, the antitumor and anti-angiogenic activity of Tum-5 was examined in vivo using a xenograft of H22 HCC cells. The results indicated that pLXSN-Tum-5 significantly inhibited tumor growth following 5 injections over 10 days. The size and weight of tumors in the pLXSN-Tum-5 group were lower than those in the saline and pLXSN groups. Furthermore, immunohistochemical analysis with CD31 antibodies indicated that the average microvessel density in the pLXSN-Tum-5 group were significantly lower than that in the saline and pLXSN groups. These results suggested that Tum-5 exerts its antitumor activity by suppressing vascular endothelial cells. The gene fragment of Tum-5 may be developed as an effective inhibitor of angiogenesis and used to treat patients with HCC.

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Section: Discussionsupporting

confidence: 47%

Vector-mediated Tum-5 expression in neovascular endothelial cells for treating hepatocellular carcinoma

Guan

Sun

et al. 2017

Experimental and Therapeutic Medicine

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“…Afterwards, the excess antibodies were washed away in saline and the samples were immediately visualized in CLE imaging to assess the microvascularization ex vivo up to a maximum depth of 250 μm. CLE images were acquired using Pentax EC-3870 CIFK, Tokyo, Japan, a dedicated endomicroscopy system with an excitation wavelength of 488 nm and with a maximum laser power output of ≤ 1 mW at the surface of the tissue [16,17] . To assess both endothelial markers more accurately, we used the color overlay function in the ImageJ image processing software (National Institutes of Health, United States).…”

Section: Targeted Anti-cd105 Antibodies For Cle Imaging Of Normal Colmentioning

confidence: 99%

“…To confirm the role of CD105 vs CD31 in tumor neoangiogenesis, adjacent samples from the same patient were processed for immunohistochemistry, for normal and tumor samples as described previously [16,17] . Briefly, after formaldehyde fixation and paraffin embedding, 4 μm tissue sections were sliced from these blocks, deparaffinized, re-hydrated and processed for antigen retrieval by microwaving for 20 min in citrate buffer pH 6.…”

Section: Immunohistochemistrymentioning

confidence: 99%

“…Recent meta-analyses performed to determine the diagnostic accuracy of CLE in the detection of colorectal neoplasia showed high sensitivity and specificity of the method [14,15] . Recently, we have used CLE to assess tumor vasculature by fluorescence labelled antibodies targeted against endothelial markers [16,17] . In the present feasibility study, we used CLE to compare the selective expression of fluorescently labeled anti-CD105 antibodies in newly-formed vessels to fluorescently labeled anti-CD31 total vessel staining, and the gold standard of histopathology.…”

Section: Introductionmentioning

confidence: 99%

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Tumor neoangiogenesis detection by confocal laser endomicroscopy and anti-CD105 antibody: Pilot study

Ciocalteu¹,

Săftoiu²,

Pirici³

et al. 2015

WJGO

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AIM:To evaluate neoangiogenesis in patients with colon cancer by two fluorescently labeled antibodies on fresh biopsy samples imaged with confocal laser endomicroscopy (CLE). . When using the anti-CD31 antibody, the average diameter of vessels in the normal colon tissue was 7.67 ± 0.5 μm and the vessel density was 3191.60 ± 387.8 vessels/mm 3 , while in the tumors we obtained an average diameter of 10.88 ± 0.8 μm and a vessel density of 4707.30 ± 448.85 vessels/mm 3 . Thus, there were more vessels stained with CD31 than CD105 (P < 0.05). The average vessel diameter was similar for both CD31 and CD105 staining. A qualitative comparison between CLE vs immunohistochemistry lead to similar results. CONCLUSION:Specific imaging and quantification of tumor microvessels are feasible in human rectal cancer using CLE examination and CD105 immunostaining of fresh tissue samples. Core tip: We evaluated CD105 expression from fresh tissue samples of human rectal adenocarcinoma, using confocal laser endomicroscopy (CLE). While vessels marked with fluorescent CD31 were visible in both normal and malignant tissue, CD105 was predominantly expressed in tumor lesions, having reduced affinity for normal rectal mucosa. Our data showed that CLE using CD105 antibody for tumor vascular network imaging is feasible and that CD105 represents a more specific marker for rectal cancer neoangiogenesis than panendothelial markers. To our knowledge, this is the first study to report the use of fluorescently-labeled CD105 antibody in conjunction with CLE in patients with rectal tumor.

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“…In a previous study, we showed that the CLE method is feasible for imaging different aspects of the tumor vasculature from fresh biopsy samples similar to the currently accepted histopathology techniques [9]. In this study, we evaluated new vascular network parameters difficult to assess with conventional immunohistochemistry in patients with clinically staged T3 primary colorectal carcinoma, without metastatic spread, by using CLE combined with fluorescently labeled anti-CD31 antibodies for labeling of both normal and tumor blood vessels.…”

Section: Introductionmentioning

confidence: 98%

Evaluation of New Morphometric Parameters of Neoangiogenesis in Human Colorectal Cancer Using Confocal Laser Endomicroscopy (CLE) and Targeted Panendothelial Markers

et al. 2014

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The tumor microcirculation is characterized by an abnormal vascular network with dilated, tortuous and saccular vessels. Therefore, imaging the tumor vasculature and determining its morphometric characteristics represent a critical goal for optimizing the cancer treatment that targets the blood vessels (i.e. antiangiogenesis therapy). The aim of this study was to evaluate new vascular morphometric parameters in colorectal cancer, difficult to achieve through conventional immunohistochemistry, by using the confocal laser endomicroscopy method. Fresh biopsies from tumor and normal tissue were collected during colonoscopy from five patients with T3 colorectal carcinoma without metastasis and were marked with fluorescently labeled anti-CD31 antibodies. A series of optical slices spanning 250 µm inside the tissue were immediately collected for each sample using a confocal laser endomicroscope. All measurements were expressed as the mean ± standard error. The mean diameter of tumor vessels was significantly larger than the normal vessels (9.46±0.4 µm vs. 7.60±0.3 µm, p = 0.0166). The vessel density was also significantly higher in the cancer vs. normal tissue samples (5541.05±262.81 vs. 3755.79±194.96 vessels/mm3, p = 0.0006). These results were confirmed by immunohistochemistry. In addition, the tortuosity index and vessel lengths were not significantly different (1.05±0.016 and 28.30±3.27 µm in normal tissue, vs. 1.07±0.008 and 26.49±3.18 µm in tumor tissue respectively, p = 0.5357 and p = 0.7033). The daughter/mother ratio (ratio of the sum of the squares of daughter vessel radii over the square of the mother vessel radius) was 1.15±0.09 in normal tissue, and 1.21±0.08 in tumor tissue (p = 0.6531). The confocal laser endomicroscopy is feasible for measuring more vascular parameters from fresh tumor biopsies than conventional immunohistochemistry alone. Provided new contrast agents will be clinically available, future in vivo use of CLE could lead to identification of novel biomarkers based on the morphometric characteristics of tumor vasculature.

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Confocal Laser Endomicroscopy for the Morphometric Evaluation of Microvessels in Human Colorectal Cancer Using Targeted Anti-CD31 Antibodies

Cited by 15 publications

References 22 publications

Vector-mediated Tum-5 expression in neovascular endothelial cells for treating hepatocellular carcinoma

Vector-mediated Tum-5 expression in neovascular endothelial cells for treating hepatocellular carcinoma

Tumor neoangiogenesis detection by confocal laser endomicroscopy and anti-CD105 antibody: Pilot study

Evaluation of New Morphometric Parameters of Neoangiogenesis in Human Colorectal Cancer Using Confocal Laser Endomicroscopy (CLE) and Targeted Panendothelial Markers

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