Giant benign lymphangioendothelioma with positive expression of Wilms tumor 1: A case report.

Benign lymphangioendothelioma (BL, acquired progressive lymphangioma) is a rare, slow-growing lymphatic tumor, first described 40 years ago, with fewer than 50 published cases. Clinically, it presents as a skin-colored or erythematous patch. Definitive diagnosis requires histopathological examination. The immunohistochemical staining profile is still controversial regarding Wilms tumor 1 (WT1) expression, a marker of proliferative and neoplastic, rather than malformative nature. Here, we report a case of a 60-cm-long BL on the breast of an adult female. Biopsy revealed irregular vascular spaces dissecting the collagen bundles lined by swollen endothelial cells but without cellular atypia. Positivity for podoplanin (D2-40), CD31, and WT1 was observed, supporting the neoplastic nature of this lesion. Dermatologists and pathologists must be aware of this entity for early diagnosis and treatment.

Signaling pathways and inhibitors of cells from patients with kaposiform lymphangiomatosis.

BACKGROUND: Kaposiform lymphangiomatosis (KLA) is a rare lymphatic anomaly with significant morbidity and mortality. KLA is characterized by diffuse multifocal lesions comprised of focal areas of "kaposiform" spindled cells accompanying malformed lymphatic channels. The goal of this study was to identify activated signaling pathways in cells isolated from three KLA patients for the purpose of testing new therapies. PROCEDURE: Cells were obtained from the lungs of one patient isolated at autopsy and the spleen of two patients removed in surgery due to disease complications. A protein kinase array was performed on the KLA cell lysates and normal lymphatic endothelial cells. RESULTS: Higher activation of key signaling pathways in the KLA cells, including PRAS40, AKT1/2/3, and ERK-1/2, was identified by protein kinase array and confirmed by Western blot analysis. This indicated a role for highly activated PI3K-AKT and MAPK-ERK-1/2 signaling pathways in KLA cells. Cell proliferation studies assessed PI3K inhibitors (LY294002; BYL719), AKT inhibitor ARQ092, mTOR inhibitor rapamycin, and MAPK inhibitor U0126. These studies demonstrated that PI3K-AKT-mTOR and MAPK signaling are important mediators of KLA cell proliferation. BYL719 and rapamycin were more effective at inhibiting KLA cell proliferation than U0126. CONCLUSIONS: Our studies using cells from KLA patient lesions demonstrate that these cells are highly proliferative and the PI3K-AKT-mTOR and MAPK pathways are promising therapeutic targets. Development and clinical trials of PI3K, AKT, and MAPK inhibitors for cancer treatment and the data in this study lend support for early clinical trials assessing the efficacy of these inhibitors in KLA patients.

Benign lymphangioendothelioma presenting as a giant flank mass.

Benign lymphangioendothelioma is a rare lesion of controversial etiology and a histopathologic mimic of Kaposi sarcoma and so-called 'well-differentiated' angiosarcoma. Its most typical clinical presentation is as a slowly expanding, erythematous patch or plaque; it rarely presents as a large mass. We report the second case of a giant benign lymphangioendothelioma, which arose as a serpiginous mass involving most of the flank of an elderly male with no prior radiation exposure and with a remote history of herpes zoster infection. A biopsy revealed numerous anastomosing vascular channels extending from the superficial dermis to the subcutis that were dilated to progressively slit-like in architecture. The endothelial cells lacked cytologic atypia, hobnailing, or significant mitotic activity, and human herpesvirus-8 expression was absent. Positivity for podoplanin (D2-40) was observed in the endothelial cells, supporting a lymphatic phenotype. Furthermore, the lesional cells lacked immunohistochemical expression of Wilms tumor 1, providing further support of a malformative - rather than neoplastic - pathogenesis.

Maldevelopment of dermal lymphatics in Wnt5a-knockout-mice.

Maintenance of tissue homeostasis and immune surveillance are important functions of the lymphatic vascular system. Lymphatic vessels are lined by lymphatic endothelial cells (LECs). By gene micro-array expression studies we recently compared human lymphangioma-derived LECs with umbilical vein endothelial cells (HUVECs). Here, we followed up on these studies. Besides well-known LEC markers, we observed regulation of molecules involved in immune regulation, acetylcholine degradation and platelet regulation. Moreover we identified differentially expressed WNT pathway components, which play important roles in the morphogenesis of various organs, including the blood vascular system. WNT signaling has not yet been addressed in lymphangiogenesis. We found high expression of FZD3, FZD5 and DKK2 mRNA in HUVECs, and WNT5A in LECs. The latter was verified in normal skin-derived LECs. With immunohistological methods we detected WNT5A in LECs, as well as ROR1, ROR2 and RYK in both LECs and HUVECs. In the human, mutations of WNT5A or its receptor ROR2 cause the Robinow syndrome. These patients show multiple developmental defects including the cardio-vascular system. We studied Wnt5a-knockout (ko) mouse embryos at day 18.5. We show that the number of dermal lymphatic capillaries is significantly lower in Wnt5a-null-mice. However, the mean size of individual lymphatics and the LEC number per vessel are greater. In sum, the total area covered by lymphatics and the total number of LECs are not significantly altered. The reduced number of lymphatic capillaries indicates a sprouting defect rather than a proliferation defect in the dermis of Wnt5a-ko-mice, and identifies Wnt5a as a regulator of lymphangiogenesis.

Novel characterization of bEnd.3 cells that express lymphatic vessel endothelial hyaluronan receptor-1.

Murine bEnd.3 endothelioma cell line has been widely used in vascular research and here we report the novel finding that bEnd.3 cells express lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) and vascular endothelial growth factor receptor-3 (VEGFR-3). Moreover, these cells express progenitor cell markers of Sca-1 and CD133. Upon stimulation with tumor necrosis factor-alpha (TNF-alpha), the bEnd.3 cells demonstrate enhanced formation of capillary-type tubes, which express LYVE-1. As the bEnd.3 cell line is derived from murine endothelioma, we further examined human tissues of endothelioma and identified lymphatic vessels in the tumor samples which express both LYVE-1 and podoplanin. Moreover, a significantly higher number of lymphatic vessels were detected in the endothelioma samples compared with normal control. Taken together, this study not only redefines bEnd.3 cells for vascular research, but also indicates a broader category of human diseases that are associated with lymphatics, such as endothelioma.

Benign lymphangioendothelioma: a clinical, histopathologic and immunohistochemical analysis of four cases.

BACKGROUND: Benign lymphangioendothelioma represents a rare lymphatic vascular proliferation characterized by proliferation of irregular and thin-walled vessels dissecting amongst dermal collagen. Immunohistochemical analysis has been lacking in most previously reported cases. METHODS: Herein, we report the clinical and histopathologic characteristics of four cases of benign lymphangioendothelioma. Immunohistochemical study was completed for all lesions. RESULTS: All lesions presented as large, red to brown patches or plaques. Three lesions were located on the thigh and one lesion was located on the neck. Histopathologically, all lesions showed proliferation of anastomotic or retiform thin-walled vessels with a single layer of endothelial cells that dissect the dermis. D2-40 and Prox1 immunostains were positive and Wilms tumor 1 (WT-1) immunostain was negative in all cases. CONCLUSION: Benign lymphangioendothelioma represents a lymphatic vascular proliferation. A lack of expression of WT-1 suggests it represents a lymphatic vascular malformation.

Multiple expressions of lymphatic markers and morphological evolution of newly formed lymphatics in lymphangioma and lymph node lymphangiogenesis.

The rapid evolution of reliable technology combined with increasing number of specific markers for lymphatic endothelial cells (LECs) facilitates the investigation of lymphangiogenesis in developing and diseased tissues. Here, we injected incomplete Freund's adjuvant (IFA) peritoneally into BALB/c and nonobese diabetic (NOD) mice to induce lymphangioma and found atypical lymphatic accumulations with intervening fibrous tissue and lymphoid aggregates. Lymphatic markers, LYVE-1 and podoplanin, were used to specifically define the morphological features of the neoplastic lymphatics. The NOD mice (affected by an autoimmune disorder) had fewer and smaller lymphangiomas than the BALB/c mice. Injection of IFA in the footpad skin of the mice also disturbed draining regional lymph node lymphangiogenesis and caused enlargement of popliteal lymph nodes. Molecular analyses of the LECs indicated potential interventions for lymphangioma through vascular endothelial growth factor (VEGF)-A/-C/-D and their receptors, VEGF receptors-2/-3, and Prox-1 signaling pathways. These findings represent an important link between multiple factors and lymphatic disorders.

VEGF-D deficiency in mice does not affect embryonic or postnatal lymphangiogenesis but reduces lymphatic metastasis.

Vascular endothelial growth factor-D (VEGF-D) is one of the two ligands of the VEGFR-3 receptor on lymphatic endothelial cells. Gene-silencing studies in mice and Xenopus tadpoles recently showed that the role of endogenous VEGF-D in lymphatic development is moderate. By contrast, exogenous VEGF-D is capable of stimulating lymphangiogenesis. Nonetheless, its endogenous role in pathological conditions remains largely unknown. Hence, we reassessed its role in disease, using Vegf-d(null) mice. Vegf-d(null) mice were generated that, under physiological conditions, displayed normal embryonic and postnatal lymphangiogenesis and lymphatic remodelling, efficient lymphatic functioning and normal health. Vegf-d(null) mice also reponded normally in models of skin wound healing and healing of infarcted myocardium, despite enhanced expression of VEGF-D in these models in wild-type mice. In contrast, Vegf-d(null) mice displayed reduced peritumoral lymphangiogenesis and lymph node metastasis in an orthotopic pancreatic tumour model. Together, our data indicate that endogenous VEGF-D in mice is dispensible for lymphangiogenesis during development, in postnatal and adult physiology and in several pathological conditions, but significantly contributes to lymphatic metastasis.

Acquired vulvar lymphangioma circumscriptum: a comparison of 12 cases with Crohn's associated lesions or radiation therapy induced tumors.

BACKGROUND: Lymphangioma circumscriptum (LC) is a benign lesion of lymphatic origin. Vulvar involvement occurs in various clinical settings. METHODS: We present 12 cases, and compare lesions in patients with Crohn's disease and those associated with pelvic radiation. RESULTS: The average age at presentation was 49 years. Thirty-three percent of the patients had Crohn's disease, 58% had radiation therapy and 9% had no significant medical history. Sixty-seven percent of the patients had multifocal lesions in anatomically distinct regions. Patients presented on average 16 years after onset of predisposing factors. Presenting complaints were pruritus, wetness and vulvar edema. Lesions were clinically heterogeneous, often found on the labia majora. Lesions consisted of dilated lymphatic channels at the junction of the reticular and papillary dermis. The cells lining these spaces lacked cytologic atypicality or mitotic activity. All lesions so examined were immunoreactive for D240. Patients were most often treated with surgical excision followed by laser ablation. Four of twelve patients, all with radiation-associated lesions, experienced disease progression necessitating additional surgery. CONCLUSIONS: Patients with LC secondary to radiation, when compared to those with Crohn's disease, were 10 years younger, more likely to have associated co-morbidities, and frequently experienced disease progression needing additional surgeries. Acquired vulvar LC has multiple causes with differing prognosis.

The Roles of Neuropilin 2/VEGF-C Axis in a Series of Recurrent Lymphangioma.

INTRODUCTION: Vascular endothelial growth factor (VEGF) and its receptor act as a major contributor to lymphangioma, but their role on nonrecurrent and recurrent lymphangiomas remain unclear. We aim to investigate those factors in the generation of recurrent lymphangioma. MATERIALS AND METHODS: Patients diagnosed with lymphangioma from January 2005 to December 2012 in our hospital were collected and divided into nonrecurrent and recurrent lymphangiomas. The clinical characteristics including age, sex, symptoms, location, and size of lymphangioma were collected. Surgical resection samples were collected for histology, protein and mRNA detection of VEGF-C, VEGF receptor-3 (VEGFR-3), and neuropilin 2 (Nrp2). Follow-ups including lymphangioma recurrent and the local symptoms such as ulcer were reviewed. RESULTS: A total of 80 patients aged from 5 months to 12 years were enrolled in this study, 51 patients had no recurrence and other 29 patients suffered from recurrent lymphangioma. There was no significant difference in demographic data and clinical characters between the two groups (p > 0.05). Immunohistochemistry staining showed that VEGFR-3 remained unchanged between nonrecurrent and recurrent lymphangiomas (p > 0.05), and VEGF-C and Nrp2 were significantly increased in recurrent lymphangioma compared with nonrecurrent lymphangioma (p < 0.05). The same expression trend was proved as detected by protein and mRNA levels. CONCLUSION: The VEGF-C/Nrp2 axis was significantly increased in the recurrent lymphangioma, indicating that VEGF-C/Nrp2 targeted therapy may serve as a potential therapeutic strategy for recurrent lymphangioma.

Intussusceptive Lymphangiogenesis in Lymphatic Malformations/Lymphangiomas.

Intussusception in lymphatic vessels has received less attention than in blood vessels. In tumors and pseudotumors of blood vessels with intravascular papillary structures, including sinusoidal hemangioma and intravascular papillary endothelial hyperplasia, we observed exuberant intussusceptive angiogenesis, as well as the similarity between papillae (term used by pathologists) and pillars/folds (hallmarks of intussusceptive angiogenesis). A similar response could be expected in lymphangiomas (lymphatic malformations and reactive processes rather than tumors) with papillae. The aim of this work is to assess whether papillae/pillars/folds and associated structures (vessel loops and septa) are present in lymphangiomas, and to establish the characteristics and formation of these structures. For this purpose, we selected lymphangiomas with intraluminal papillae (n = 18), including cystic, cavernous, circumscriptum, and progressive types, of which two cases of each type with a greater number of papillae were used for serial histologic sections and immunohistochemistry. The studies showed a) dilated lymphatic spaces giving rise to lymphatic-lymphatic vascular loops, which dissected and encircled perilymphatic structures (interstitial tissue structures/ITSs and pillars/posts), b) ITSs and pillars, surrounded by anti-podoplanin-positive endothelial cells, protruding into the lymphatic spaces (papillary aspect), and c) splitting, remodeling, linear arrangement, and fusion of papillae/pillars/folds, forming papillary networks and septa. In conclusion, as occurs in blood vessel diseases, the development of lymphatic vessel loops, papillae/pillars/folds, and septa (segmentation) supports intussusceptive lymphangiogenesis and suggests a piecemeal form of intussusception. This intussusceptive lymphangiogenesis in lymphatic diseases can provide a basis for further studies of lymphatic intussusception in other conditions, with clinical and therapeutic implications. Anat Rec, 302:2003-2013, 2019. © 2019 American Association for Anatomy.

Elevated expression of VEGFR-3 in lymphatic endothelial cells from lymphangiomas.

BACKGROUND: Lymphangiomas are neoplasias of childhood. Their etiology is unknown and a causal therapy does not exist. The recent discovery of highly specific markers for lymphatic endothelial cells (LECs) has permitted their isolation and characterization, but expression levels and stability of molecular markers on LECs from healthy and lymphangioma tissues have not been studied yet. We addressed this problem by profiling LECs from normal dermis and two children suffering from lymphangioma, and also compared them with blood endothelial cells (BECs) from umbilical vein, aorta and myometrial microvessels. METHODS: Lymphangioma tissue samples were obtained from two young patients suffering from lymphangioma in the axillary and upper arm region. Initially isolated with anti-CD31 (PECAM-1) antibodies, the cells were separated by FACS sorting and magnetic beads using anti-podoplanin and/or LYVE-1 antibodies. Characterization was performed by FACS analysis, immunofluorescence staining, ELISA and micro-array gene analysis. RESULTS: LECs from foreskin and lymphangioma had an almost identical pattern of lymphendothelial markers such as podoplanin, Prox1, reelin, cMaf and integrin-alpha1 and -alpha9. However, LYVE-1 was down-regulated and VEGFR-2 and R-3 were up-regulated in lymphangiomas. Prox1 was constantly expressed in LECs but not in any of the BECs. CONCLUSION: LECs from different sources express slightly variable molecular markers, but can always be distinguished from BECs by their Prox1 expression. High levels of VEGFR-3 and -2 seem to contribute to the etiology of lymphangiomas.

Benign lymphangiomatous papules of the skin associated with ovarian fibroma.

Benign lymphangiomatous papules of the skin are considered reactive lymphatic proliferations either caused by disruption of the lymphatic flow or tissue damage produced by operation or radiation therapy. We report a 72-year-old woman with umbilical papules and vesicle-like lesions that led to the diagnosis of a large ovarian fibroma. Histologic study revealed dilated lymphatic spaces manifesting an anastomosing and branched pattern in the papillary and reticular dermis dissecting collagen bundles. The vessels were lined by plump endothelial cells with foci of intravascular papillary endothelial cell hyperplasia. After the ovarian fibroma was removed by laparotomy, umbilical lesions almost disappeared, leaving small flesh-colored papules. A periumbilical dermatosis may herald certain intra-abdominal diseases including those of neoplastic derivation. A heightened awareness of this association may lead to an early diagnosis with a potential for improved patient outcome. Benign lymphangiomatous papules have not been previously described in association with an untreated tumor, without previous operation or radiotherapy. This case advocates for disruption of the lymphatic drainage as the probable pathogenetic mechanism.

[Splenic lymphangioma: clinicopathologic and immunohistochemical study of 18 cases and review of literature].

OBJECTIVE: To study the clinicopathologic features and differential diagnosis of splenic lymphangioma. METHODS: Eighteen cases of splenic lymphangioma were retrieved from the pathology archives during the period between January 1990 to December 2005. The clinicopathologic features were analyzed. Immunohistochemical study was performed on the paraffin sections of 16 cases. RESULTS: The age of the patients ranged from 9 to 72 years (median = 40 years). Thirteen patients were males and 5 were females. Clinically, the tumor could be asymptomatic or present with abdominal symptoms and hypersplenism. Follow-up information was available in 13 patients (72.2%) and the duration varied from 5 months to 15 years. All 13 patients had an uneventful clinical course, with no evidence of residual disease, local recurrence or metastasis. Gross examination showed splenic enlargement. The tumor appeared as cystic (8/18), solid (5/18) or honeycomb mass (5/18), either solitary (5/18) or multifocal (13/18). Histologically, splenic lymphangioma could be subclassified as cavernous (9/18), cystic (5/18) or mixed (4/18). Immunohistochemical study showed that the positivity rates for CD9 and D2-40 were 100% and 43.8% respectively. CONCLUSIONS: Splenic lymphangioma is a rarely encountered entity that can be misdiagnosed as a splenic hemangioma. A definite diagnosis depends on pathologic examination.

Altered pigment epithelium-derived factor and vascular endothelial growth factor levels in lymphangioma pathogenesis and clinical recurrence.

OBJECTIVE: To determine the role of angiogenesis in the clinical behavior and pathogenesis of lymphangioma tumors. DESIGN: A retrospective study. Median follow-up period was 44.5 months. SETTING: Children's Memorial Hospital, Chicago, Ill. PATIENTS: Tumor specimens from 12 pediatric patients who underwent surgical excision of cervicofacial lymphangioma were examined for expression of angiogenic inducer vascular endothelial growth factor (VEGF) and angiogenic inhibitor pigment epithelium-derived factor (PEDF) using immunohistochemical analysis. Specimens were divided into recurrent and nonrecurrent tumors based on clinical information. MAIN OUTCOME MEASURES: Staining patterns of VEGF and PEDF were evaluated in lymphangioma specimens. Staining patterns were then compared in both recurrent and nonrecurrent groups and graded in a blinded fashion. Histological evidence of increased angiogenesis including microvascular density, stromal fibrosis, and inflammation were graded in each group and correlated with recurrence. RESULTS: Lymphangioma specimens demonstrated histological evidence of increased angiogenic activity including multiple areas of increased VEGF staining combined with little PEDF staining. Sex, age at onset, or tumor location did not correlate with recurrence. Furthermore, recurrent specimens had increased histological evidence of angiogenesis as well as increased VEGF and decreased PEDF activity compared with nonrecurrent lesions. CONCLUSIONS: Lymphangiomas exhibit tumorlike pathogenesis owing to the high expression of angiogenic inducers compared with the low expression of inhibitors. Recurrence may be influenced by this imbalance of angiogenic mediators. Further research with antiangiogenic therapy using agents such as PEDF analogues or anti-VEGF receptor antibodies is indicated because they may stabilize or suppress the growth of these neoplasms.

Differential Prox-1 and CD 31 expression in mucousae, cutaneous and soft tissue vascular lesions and tumors.

The study of lymphatic vessels and lymphatic tumors has been hampered with difficulty due to the overlapping morphological features between blood and lymphatic endothelial cells, as well as to the lack of specific lymphatic endothelial markers. Over the last few years, lymphatic vessels and lymphangiogenesis have received great attention owing to their putative implications in terms of metastatic dissemination and the promise of targets for lymphangiogenic therapy. Prox-1 is a nuclear transcription factor that plays a major role during embryonic lymphangiogenesis and is deemed to be a useful marker for differentiating lymphatic endothelial cells from the other blood vessels endothelial cells. Here, we describe a double-immunostaining strategy for formalin-fixed, paraffin-embedded tissues that aims at evaluating the distribution of Prox-1 and CD 31 - a cytoplasmic pan-endothelial marker - in a series of 28 mucousae, cutaneous and soft tissue vascular lesions and tumors, including hemangiomas, lymphangiomas, lymphangiectasia, and Kaposi's sarcomas. Our results showed that in non-lesional mucousae and skin, Prox-1 decorated exclusively the nuclei of endothelial cells in lymphatic vessels. Prox-1 stained almost all the benign lymphatic vascular lesions/tumors (91%) and was absent or only focally positive in 75% of blood vascular tumors. CD 31 stained endothelial cells of blood vessels of superficial and deep dermal plexuses, lymphatics, and all blood vascular lesions/tumors. Kaposi's sarcomas were all positive for both CD 31 and Prox-1 markers. In conclusion, although Prox-1 expression in vascular lesions/tumors was not entirely restricted to tumors with known lymphatic differentiation, CD 31/Prox-1 double-immunolabeling can be used as an adjunct marker to identify lymphatic vessels in routinely processed formalin-fixed, paraffin-embedded samples.

Lymphangiomatosis. Histologic and immunohistochemical analysis of four cases.

Lymphangiomatosis is a rare condition that involves bone, soft tissue, or viscera in a diffuse fashion. We report four examples affecting male infants and boys (aged 9 months to 11 years: mean, 5.3 years). All four cases presented with respiratory symptoms. Investigations showed chylothorax in all patients and chylopericardium in one patient. Three patients showed multiple lytic lesions in several bones on the x-rays. Imaging studies of lungs and spleen suggested lymphangiomatosis. Biopsy specimens of the parietal pleura, lung, skin, or bone revealed an increase in the size and number of thin-walled channels lined by attenuated endothelial lining. All had histological evidence of parietal pleural involvement. In addition, lung involvement was seen in three cases and skin involvement in two cases. Factor VIII-related antigen and CD31 were the most reliable immunocytochemical markers in highlighting the endothelia. All the patients had repeated thoracocentesis and pleurodesis to control chylothorax; three died within 6 to 33 months of presentation (mean, 15 months). Autopsy performed in two cases revealed additional involvement of spleen, bone, and mediastinum. Further, lymphangiectasia was seen in the liver, kidney, tests, lymph node, intestines, and adrenals.

Distribution of keratins in normal endothelial cells and a spectrum of vascular tumors: implications in tumor diagnosis.

Vascular endothelial cells are specialized mesenchyme-derived epithelial-like lining cells which are the essential participants in benign and malignant vascular tumors. Although endothelia in lower animals often express keratins (K), human endothelia are generally K negative and vimentin-positive. However, K expression has been noted in some endothelia and in some epithelioid vascular tumors. In this study, we systematically examined normal human vascular endothelia and a spectrum of human vascular tumors (n = minimum of 137 tumors with each marker) for simple epithelial keratin polypeptides of the Moll catalogue (K7, K8, K18, and K19). Selected vascular tumors were also evaluated with antibodies to K14 and the monoclonal antibody 34betaE12 that recognizes several keratins of stratified epithelia. Endothelia of normal veins, venules, and lymphatics commonly exhibited focal positivity for K7 and K18, whereas K8, K14, and K19 were not seen in non-neoplastic endothelia with the antibodies used. Lymphangiomas (6 of 7) and venous hemangiomas (6 of 13) often showed K7-positive endothelial cells; K18 was detected less commonly, whereas K8 and K19 were not detected. Epithelioid hemangioendotheliomas (EHEs) showed K7 and K18 expression in the majority of cases (50% and 100%, respectively), while K8 was seen in 10% cases and K14 and K19 in none. In contrast, epithelioid angiosarcomas (EAs) were often positive for K8 and K18 (approximately 50%), whereas they less commonly showed K7 and only occasionally K19; all tumors were negative for K14 and with the antibody 34betaE12. Nonepithelioid angiosarcomas (AS) less commonly showed keratin expression with K7, K8, and K18 being positive in 20% of cases, and K14 and K19 in none of the cases. Epithelial membrane antigen (EMA) was occasionally detectable in EHE (2/19) but was present in 4 of 16 (25%) EAs and 17 of 48 (35%) nonepithelioid AS. These findings document the common presence of focal reactivity for K7 and K18 in subsets of normal endothelia and also the frequent presence of simple epithelial keratins in malignant vascular tumors, while such expression is uncommon in nonepithelioid angiosarcomas. K- and EMA-positivity in neoplastic endothelia needs to be considered in the evaluation of human tumors. K antibodies such as those specific to K19 or AE1 that do not react with K8 and K18 should be used in the differential diagnosis of epithelioid vascular tumors and carcinomas.

Effect of hepatoma H22 on lymphatic endothelium in vitro.

AIM: To determine the effect of metastatic hepatoma cells on lymphangioma-derived endothelium, and to establish in vitro model systems for assessing metastasis-related response of lymphatic endothelium. METHODS: Benign lymphangioma, induced by intraperitoneal injection of the incomplete Freund's adjuvant in BALB/c mice, was embedded in fibrin gel or digested and then cultured in the conditioned medium derived from hepatoma H22. Light and electron microscopy, and the transwell migration assay were used to determine the effect of H22 on tissue or cell culture. Expressions of Flt-4, c-Fos, proliferating cell nuclear antigen (PCNA), and inducible nitric oxide synthase (iNOS) in cultured cells, and content of nitric oxide in culture medium were also examined. RESULTS: The embedded lymphangioma pieces gave rise to array of capillaries, while separated cells from lymphangioma grew to a cobblestone-like monolayer. H22 activated growth and migration of the capillaries and cells, induced expressions of Flt-4, c-Fos, PCNA and iNOS in cultured cells, and significantly increased the content of NO in the culture medium. CONCLUSION: Lymphangioma-derived cells keep the differentiated phenotypes of lymphatic endothelium, and the models established in this study are feasible for in vitro study of metastasis-related response of lymphatic endothelium.