Ulex europaeus I lectin as a marker for tumors derived from endothelial cells

Some skin and soft tumors, which generally are assumed to be derived from endothelial cells or blood vessels, were characterized with fluorochrome-labeled Ulex europaeus I agglutinin (UEA I), recently shown to bind specifically to endothelial cells in various normal human tissues. The staining pattern was compared with that obtained with immunostaining using antibodies against factor-VIII-related antigen (FVIII-RAG), a known marker for endothelial cells. The results showed that UEA-I is a specific and a more sensitive marker for the endothelial cells in benign vascular lesions as compared with anti-FVIII-RAG. UEA-I also stained many neoplastic cells of endothelial sarcomas, which generally were negative for FVIII-RAG. Melanomas, anaplastic carcinomas, and other types of sarcomas were negative for both UEA-I and FVIII-RAG. The results suggest that UEA-I lectin is a specific and sensitive adjunct tool in demonstrating endothelial cells and endothelial derivation of human tumors.     

Thrombomodulin as a marker for vascular tumors. Comparative study with factor VIII and Ulex europaeus I lectin

Thrombomodulin (TM) is a newly described endothelial cell-associated protein that functions as a potent natural anticoagulant by converting thrombin from a procoagulant protease to an anticoagulant. Various vascular tumors were characterized with immunoperoxidase staining with the use of a polyclonal anti-TM serum. The staining patterns of TM were compared with those of Factor VIII-related antigen (FVIII-RAG) and Ulex europaeus agglutinin-I (UEA-I), which have been used as markers for endothelial cells. The results showed that TM is a specific and a highly sensitive marker for angiosarcomas in comparison with FVIII-RAG or UEA-I. In contrast, UEA-I is more sensitive for benign vascular tumors than TM or FVIII-RAG. The other mesenchymal tumors of nonvascular origin showed negative staining for three endothelial markers. These results indicate that TM is a new specific and sensitive tool for the diagnosis of angiosarcomas.     

Demonstration of vascular endothelium in thyroid carcinomas using Ulex europaeus I agglutinin

The usefulness of using peroxidase-labelled Ulex europaeus agglutinin I for the staining of small vessels and capillaries in the capsule of thyroid tumours is demonstrated. With this procedure the scanning for small tumour deposits in those vessels and, consequently, the diagnosis of follicular carcinoma of the thyroid is facilitated.     

Comparison of Ulex europaeus I lectin and factor VIII-related antigen in vascular lesions

Factor VIII-related antigen (FVIIIR:Ag) has been widely used as a marker for endothelial cells. Recent studies have shown that Ulex europaeus agglutinin I lectin (UEAI) also binds specific endothelial cells. To determine the relative utility of the two endothelial cell markers, a group of vascular and nonvascular tumors were studied with both markers, using an immunoperoxidase method. Our results indicate that UEAI is a more sensitive marker for endothelial cells than FVIIIR:Ag, and that it should be used as an additional diagnostic marker for endothelial cell-derivated tumors.     

Comparison of Ulex europaeus I lectin binding and factor VIII-related antigen as markers of vascular endothelium in follicular carcinoma of the thyroid

Factor VIII-related antigen (F VIII-RAg), a well established marker for endothelial cells, and the lectin Ulex europaeus agglutinin I (UEA-1), a newly described marker, were used in immunoperoxidase techniques to demonstrate endothelial cells in 30 follicular neoplasms of the thyroid and so to assist in detection of vascular invasion. Both endothelial cell markers made visible a greater number of instances of vascular invasion than were detected on routine elastic stains, but UEA-1 lectin gave more reliable staining of endothelium lining large capsular veins and of blood vessels within the tumour than did F VIII-RAg. In vessels completely occluded by tumour, examples of endothelial staining by UEA-1 in the absence of F VIII-RAg staining were found, but some such vessels appeared to lack surviving endothelial cells and in these no staining occurred. It is concluded that UEA-1 lectin is a more reliable endothelial marker in this setting than F VIII-RAg and may assist the demonstration of vascular invasion in these tumours.     

Ulex europeus I--peroxidase as a marker of vascular endothelium: its application in routine histopathology

Peroxidase-labelled Ulex europeus I lectin has been applied to a series of routinely fixed and processed vascular lesions to assess its value in the identification of endothelium. A group of breast carcinomas previously examined for binding of the lectin to the tumour have been re-assessed for evaluation of the use of Ulex europeus I in the determination of vascular invasion. There was consistent binding to normal vascular endothelium but this could be variable, there being stronger reactivity in small vessels of some tissues. All benign lesions stained showed clear definition of vascular channels but angiosarcomas were much less consistent in their staining reaction and the peroxidase-labelled lectin was considered to be a less satisfactory marker. Lymphatic endothelium was always negative; this proved to be a useful feature in differentiating between blood vessel and lymphatic invasion by breast carcinoma. Ulex europeus I-peroxidase could certainly have a role in routine histopathology in this area.     

Hepatic sinusoidal endothelium: Ulex lectin binding

The sinusoidal endothelial cells of human liver can be identified by light and electron microscopy, but there appear to be no specific immunocytochemical markers of these cells. Among specific markers available for vascular endothelial cells in general, Ulex europaeus I lectin (UEA I) is the most sensitive. In the present study, 37 liver biopsies were examined for UEA I binding and for Factor VIII related antigen (F VIII RAg) to determine if sinusoidal endothelial cells were positive. The material included normal liver, biopsies from patients with cirrhosis and biopsies in a variety of other liver diseases. Three embryonal human livers were also included in the immunocytochemical analysis. Eleven oesophageal rings obtained at mechanical transection for variceal bleeding in cirrhotic patients were used as control tissue. Sinusoidal endothelial cells of normal liver did not stain with UEA I, but six of seven with alcoholic cirrhosis and only one of 25 non-cirrhotic liver specimens (a case of acute hepatitis with bridging necrosis) were positive. In two of the six cirrhoses the sinusoidal endothelial cells were stained for F VIII RAg as well. Embryonal sinusoidal endothelial cells were stained with UEA I but were negative for F VIII RAg. The results of the study confirm that sinusoidal endothelial cells of normal adult human liver are phenotypically different from those lining blood vessels in other sites. In cirrhosis, positive staining may be related to the transformation of hepatic sinusoids into true capillaries and thus be a marker of the severity of physiological disturbance in the liver.     

Nestin as a marker for proliferative endothelium in gliomas

Nestin is one of the intermediate filaments abundantly produced in the developing central nervous system and somites in the embryonic stage. Nestin is also reportedly detected in gliomas/glioblastomas. We retested nestin expression in brain tumors having a range of malignancy grades using immunostaining. The intensity of nestin immunostaining roughly paralleled the malignancy grade of the gliomas. However, many tumors were negative for nestin immunostaining, while nestin immunostaining was invariably detected in tumor endothelium regardless of glioma malignancy grades or brain tumor types. We suspected that angiogenic epithelial cells may express nestin, and we found that nestin was highly positive in bovine aortic endothelial cells in static culture. However, nestin expression decreased when the endothelial cells underwent laminar shear stress flow, under which endothelial cells exhibit differentiated features and a decreased rate of growth. Because nestin is highly expressed in growing endothelial cells, we examined its expression in hemangioblastomas because hemangioblasts are thought to be a precursor for angiogenic epithelial cells. As expected, nestin immunostained strongly in all four samples of hemangioblastomas. We suggest that nestin is not only a marker for neuroepithelial stem cells and glioma cells but also for tumor endothelial cells during rapid growth.     

An immunocytochemical study of the germinal layer vasculature in the developing fetal brain using Ulex europaeus 1 lectin

The characteristics of the germinal matrix vasculature were studied in the developing fetal brain using immunocytochemical methods. A preliminary comparative immunocytochemical study was made on six fetal brains to compare endothelial staining by Ulex europaeus I lectin with that of antibody to Factor VIII related antigen. Ulex was found to stain germinal layer vessels better than Factor VIII related antigen. Subsequently, the germinal layers of a further 15 fetal and preterm infant brains ranging from 13 to 35 weeks' gestation were stained with Ulex europaeus I to demonstrate the vasculature. With increasing gestation, there was a gradual increase in vessel density, particularly of capillaries. This was not a uniform process. A plexus of capillaries was prominent immediately beneath the ependyma while the more central parts of the germinal matrix contained fewer, but often larger diameter, vessels. The variation in vessel density which was a feature of the later gestation brains may have implications for local blood flow and may be a factor in haemorrhage at this site.     

Lectin I of ulex europaeus as a marker for a subset of histiocytic tumours of the lymph node

Summary We describe four lymph node based tumours in which numerous neoplastic cells and some mitotic figures were characterized by staining affinity for Lectin I of Ulex europaeus (UEA-I). The patients had no vascular or epithelial tumours and presented symptoms suggestive of a systemic lymphoproliferative disease. Histologically, the tumours were composed of large, cohesive, cells which were mainly located in the paracortex. UEA-I reactivity was more evident in the Golgi area and was present in large mononucleated cells often arranged to delimit vascular-like spaces. The neoplastic cells were weakly muramidase-positive in one case, and were ANAE+/AP + in two other cases. Large dots of UEA-I reactivity were detected in S-100+/muramidase-negative Langerhans-like cells present in one case of Letterer-Siwe disease. UEA-I staining was consistently negative in 20 cases of B cell- or T cell lymphoma and in 9 other cases of histiocytic lymphoma. It is suggested that UEA-I+ tumours of the lymph nodes are part of a distinct subset of histiocytic malignancies whose neoplastic cells present some morphological and phenotypic properties normally associated with endothelial cells.Supported by CNR contract N. 84.00440.44 Progetto Finalizzato Oncologia     

Helix pomatia and Ulex europeus lectin binding in human breast carcinoma

Formalin fixed, paraffin embedded tissue from 100 consecutive cases of breast carcinoma were studied for binding with Helix pomatia (HPA) and Ulex Europeus (UEA1) lectins. Serial sections were pretreated with trypsin or neuraminidase to determine the effect of these enzymes on lectin binding. The lectins were visualized by the peroxidase antiperoxidase technique and the cell staining proportion assessed in a semi-quantitative manner under the light microscope. Correlating staining with prognostic factors and patient follow-up details showed that UEA1 related to disease-free interval and survival, and HPA to lymph node stage, time to loco regional recurrence and to survival. Relationships with both lectins were abolished by pretreatment with neuraminidase. The study demonstrates that a simple assessment of lectin binding can provide prognostic information in breast cancer. This may be useful particularly when conservational surgical practice restricts the amount of nodal tissue for staging.     

Specific carbohydrate expression by small-diameter subclasses of rabbit trigeminal, glossopharyngeal, and vagal afferent fibers studied with the lectin Ulex europaeus agglutinin I

Lectin Ulex europaeus agglutinin I (UEAI), which binds to some alpha-L-fucose containing carbohydrates, specifically labeled a subset of trigeminal, glossopharyngeal, and vagal ganglion cells and their axons in the rabbit. The UEAI-positive trigeminal afferent fibers traveled through the spinal tract of the trigeminal nerve and terminated predominantly at the superficial laminae of the caudal region of the nucleus of the spinal trigeminal tract. The trajectory and termination regions of the UEAI-positive trigeminal afferent fibers as well as their small diameters suggest that they are C-fibers which mediate nociceptive and/or thermoceptive inputs. The UEAI-positive vagal and glossopharyngeal fibers were also of small diameter and traveled through the solitary tract. They projected to the nucleus of the solitary tract and the area postrema. No central neurons in the brainstem were labeled with UEAI. These results indicate that subsets of trigeminal, glossopharyngeal, and vagal primary sensory neurons express specific carbohydrates on their cell surfaces.     

Immunobiology of human vascular endothelium

The author’s laboratory studies interactions between human T lymphocytes and vascular endothelial cells (EC). Our work is organized around three hypotheses. First, we propose that vascular EC can initiate secondary (i.e., recall) immune reactions by presenting antigenic peptide-major histocompatibility complex (MHC) complexes to those circulating memory T cells whose cognate antigen is locally present within a peripheral tissue, e.g., as a consequence of infection or allogeneic transplantation. In this way, EC can increase the efficiency of immune surveillance. Second, we propose that T cell signals, both secreted (e.g., cytokines) and contact-dependent (e.g., CD40 ligand), activate new gene expression in EC that induce the capacity to perform new effector functions, such as leukocyte recruitment and activation or initiation of intravascular coagulation. In this way, EC can particpate as effector cells for cell-mediated immune reactions. Third, we propose that EC are major targets of immune-mediated injury. Consequently, increasing resistance of endothelium to immune effector mechanisms may protect tissues from damage, e.g., in allograft rejection. These three hypotheses are explored through in vitro experiments, through analyses of human tissue specimens, and through in vivo studies employing novel human-mouse chimeric animals.     

Evaluation of claspin as a proliferation marker in human cancer and normal tissues

Claspin is a nuclear protein involved in DNA replication and the DNA damage response. Its structural and functional properties suggest that it may represent a potentially useful proliferation marker. To this end, a monoclonal antibody was generated and the expression of claspin was investigated in normal fibroblasts and various cancer cell lines, as well as in tumour and normal tissues from patients with primary epithelial carcinomas. Immunoblotting analysis confirmed the specificity of the antibody, while immunohistochemistry demonstrated its applicability in archival material. In normal cells and tissues, claspin expression was weak, whereas increased levels were observed in cancer cell lines and tumour specimens. Claspin staining correlated strongly with Ki67 staining in both normal (p < 0.001) and tumour tissues (p < 0.001). However, the labelling index (LI) of claspin was consistently lower than that of Ki67, suggesting that claspin expression may be limited to a narrower part of the cell cycle. Co-localization assays with cyclin A and cell synchronization experiments indicated that claspin expression coincides with the S phase. Interestingly, the relative increase of the claspin LI in tumour samples compared with normal tissues was significantly higher (14-fold) than that of the Ki67 LI (five-fold), suggesting that claspin may be a more sensitive marker of aberrant proliferation.     

Endothelial cell markers in vascular neoplasms: an immunohistochemical study comparing factor VIII-related antigen, blood group specific antigens, 6-keto-PGF1 alpha, and Ulex europaeus 1 lectin

Markers for endothelial cells including Ulex europaeus 1 lectin, blood group A, B, and H, and the prostaglandin metabolite 6-keto-PGF1 alpha were evaluated in paraffin secretions from formalin-fixed benign and malignant vascular neoplasms using a variety of immunohistochemical techniques, and results compared with staining for factor VIII-related antigen. Staining for Ulex appeared more sensitive than factor VIII-related antigen in identifying poorly differentiated neoplasms including haemangiosarcomas and spindle cell proliferations in Kaposi's sarcoma. Staining for blood group related antigens correlated with blood group in all cases. Ulex europaeus 1 lectin was the only marker for endothelial cells in lymphangiomas.     

Use of Ulex europaeus agglutinin I (UEAI) to distinguish vascular and "pseudovascular" invasion in transitional cell carcinoma of bladder with lamina propria invasion

We used Ulex europaeus agglutinin I (UEAI)-immunoperoxidase staining of endothelium to study the accuracy of hematoxylin and eosin (H&E) diagnosis, occurrence, and significance of lymphvascular invasion in transitional cell carcinoma (TCC) of the bladder invading the lamina propria (Stage T1). Original histologic slides from cases (1967 to 1985) with and without vascular invasion were destained and restained with UEAI-immunoperoxidase. Only 5 of 36 biopsies originally diagnosed with lymphvascular invasion had tumor nests within endothelium-lined spaces. The 31 negative biopsies had extensive retraction artifacts lined by connective tissue and fibroblasts around tumor nests. Thirty-five control biopsies remained negative for lymphvascular invasion. Clinical follow-up of the five patients with proven lymphvascular invasion found three without progression of disease 3 to 10 yr postbiopsy, one dead of a local recurrence of TCC 1.67 yr postbiopsy, and one lost to follow-up. Based on this study, we feel that lymphvascular invasion by TCC in Stage T1 tumors is unusual, is frequently misdiagnosed on H&E stain, and does not necessarily portend a poor prognosis.