Myopericytoma arising from myopericytosis-a hitherto unrecognized entity within the lung.

Two cases of myopericytosis combined with pericytoma originating within the lung are reported. These are rare pulmonary tumors. The differential diagnosis for hemangiopericytoma and pericytic tumors with glomus elements is discussed. Both myopericytic lesions mimic other lesions, which are more commonly seen in the lung. Based on the expression of vascular growth factor receptors 2 and 3, an antiangiogenic therapy was suggested for the patient with the myopericytoma. A treatment with an angiogenesis inhibitor resulted in a regression of the tumor, but not the precursor lesion. Probably a more specific therapy using tyrosine kinase inhibitors for VEGFR2/3 might better control these myopericytic proliferations.

Autotaxin and vascular endothelial growth factor receptor-2 and -3 are related to vascular development during the progression of chronic viral hepatitis C.

Vascular endothelial growth factor (VEGF) and autotaxin (ATX) play important roles in embryonic vasculogenesis and cancer progression. This study examines whether these two angiogenic factors cooperate in the mechanism that regulates vascular development during the progression of chronic viral hepatitis C (CVH-C) (Inuyama classification, F1-F4). First, surgical wedge biopsy specimens and needle biopsy specimens were obtained. Immunohistochemical staining for ATX and vascular endothelial growth factor receptor was assessed in serial sections. Immunoelectron microscopy was conducted with a perfusion-fixation method. In normal control liver tissue specimens, ATX was expressed at low levels within the branches of the hepatic artery and hepatic sinusoids. In F1 CVH-C liver tissue specimens, ATX was expressed within the branches of the hepatic artery. Additionally, VEGFR-2 was expressed within the branches of the hepatic artery and capillaries. In F3-F4 CVH-C liver tissue specimens, positive staining for ATX and VEGFR-2 or VEGFR-3 was detected in the branches of the hepatic artery or microlymphatic vessels. ATX-1 reaction products were specifically expressed on the plasma membrane of some microvascular endothelial cells (ECs) in the proliferative capillary artery. VEGFR-2 was expressed on caveolae in ECs and vascular smooth muscle cells. VEGFR-3 immunogold particles were also observed in lymphatic ECs. These results suggest functional interactions among ATX, VEGFR-2, and VEGFR-3 in the modulation of hemovascular and lymphovascular cell activation during vascular development.

Co-expression of PDGF-B and VEGFR-3 strongly correlates with poor prognosis in hepatocellular carcinoma patients after hepatectomy.

BACKGROUND: The ability to evaluate the prognosis of hepatocellular carcinoma (HCC) patients following hepatectomy with biological markers is of great importance. METHODS: In this study, we collected samples from 90 patients with HCC after hepatectomy. Immunohistochemistry was used to detect the expression of PDGF-B and VEGFR-3 in these HCC samples. RESULTS: According to the immunohistochemical results, PDGF-B and VEGFR-3 staining were significantly associated with clinical features. Additionally, a significant association between high PDGF-B and VEGFR-3 levels and shorter overall survival was noted, when PDGF-B and VEGFR-3 co-expression been analyzed. CONCLUSION: These results suggest that the correlative expression level of PDGF-B and VEGFR-3 has strong value in the prognosis of HCC patients following hepatectomy.

Ascending Vasa Recta Are Angiopoietin/Tie2-Dependent Lymphatic-Like Vessels.

Urinary concentrating ability is central to mammalian water balance and depends on a medullary osmotic gradient generated by a countercurrent multiplication mechanism. Medullary hyperosmolarity is protected from washout by countercurrent exchange and efficient removal of interstitial fluid resorbed from the loop of Henle and collecting ducts. In most tissues, lymphatic vessels drain excess interstitial fluid back to the venous circulation. However, the renal medulla is devoid of classic lymphatics. Studies have suggested that the fenestrated ascending vasa recta (AVRs) drain the interstitial fluid in this location, but this function has not been conclusively shown. We report that late gestational deletion of the angiopoietin receptor endothelial tyrosine kinase 2 (Tie2) or both angiopoietin-1 and angiopoietin-2 prevents AVR formation in mice. The absence of AVR associated with rapid accumulation of fluid and cysts in the medullary interstitium, loss of medullary vascular bundles, and decreased urine concentrating ability. In transgenic reporter mice with normal angiopoietin-Tie2 signaling, medullary AVR exhibited an unusual hybrid endothelial phenotype, expressing lymphatic markers (prospero homeobox protein 1 and vascular endothelial growth factor receptor 3) as well as blood endothelial markers (CD34, endomucin, platelet endothelial cell adhesion molecule 1, and plasmalemmal vesicle-associated protein). Taken together, our data redefine the AVRs as Tie2 signaling-dependent specialized hybrid vessels and provide genetic evidence of the critical role of AVR in the countercurrent exchange mechanism and the structural integrity of the renal medulla.

Whole-body imaging of lymphovascular niches identifies pre-metastatic roles of midkine.

Cutaneous melanoma is a type of cancer with an inherent potential for lymph node colonization, which is generally preceded by neolymphangiogenesis. However, sentinel lymph node removal does not necessarily extend the overall survival of patients with melanoma. Moreover, lymphatic vessels collapse and become dysfunctional as melanomas progress. Therefore, it is unclear whether (and how) lymphangiogenesis contributes to visceral metastasis. Soluble and vesicle-associated proteins secreted by tumours and/or their stroma have been proposed to condition pre-metastatic sites in patients with melanoma. Still, the identities and prognostic value of lymphangiogenic mediators remain unclear. Moreover, our understanding of lymphangiogenesis (in melanomas and other tumour types) is limited by the paucity of mouse models for live imaging of distal pre-metastatic niches. Injectable lymphatic tracers have been developed, but their limited diffusion precludes whole-body imaging at visceral sites. Vascular endothelial growth factor receptor 3 (VEGFR3) is an attractive 'lymphoreporter' because its expression is strongly downregulated in normal adult lymphatic endothelial cells, but is activated in pathological situations such as inflammation and cancer. Here, we exploit this inducibility of VEGFR3 to engineer mouse melanoma models for whole-body imaging of metastasis generated by human cells, clinical biopsies or endogenously deregulated oncogenic pathways. This strategy revealed early induction of distal pre-metastatic niches uncoupled from lymphangiogenesis at primary lesions. Analyses of the melanoma secretome and validation in clinical specimens showed that the heparin-binding factor midkine is a systemic inducer of neo-lymphangiogenesis that defines patient prognosis. This role of midkine was linked to a paracrine activation of the mTOR pathway in lymphatic endothelial cells. These data support the use of VEGFR3 reporter mice as a 'MetAlert' discovery platform for drivers and inhibitors of metastasis.

[Epithelioid sarcoma with mesothelial and lymphatic endothelial differentiation: a clinicopathologic analysis of 10 cases].

Objective: To investigate the multidirectional differentiation potential in epithelioid sarcoma (ES), with special emphasis on its mesothelial and lymphatic endothelial markers expression. Methods: Ten cases of distal-type ES were included. The clinical, histological, and immunohistochemical(including mesothelial and lymphatic endothelial markers expression)features and follow-up data were evaluated. Results: The patients aged between 8 to 66 years. Five cases were male and five were female. The tumors were located at the palm (2 cases), wrist (3 cases), upper arm (2 cases), poplitealfossa (1 case), lower leg (1 case) and thigh (1 case), respectively. Clinically, most cases presented as painful, firm subcutaneous nodules. Histologically, the tumors were mainly composed of epithelioid, rhabdoid, spindle, or transitional cells, with abundant eosinophilic cytoplasm, oval and vesicular nucleus, and one or more prominent nucleoli. They were arranged in nodular, diffuse nodular or sheet like growth patterns, frequently with necrosis at the center with vague granulomatous configuration. Immunohistochemically, all tumors expressed cytokeratins, epithelial membrane antigen, CD34, desmin, mesothelial markers such as Calretinin, WT1, D2-40, M2A, vascular and lymphatic endothelial markers FLI-1, and VEGFR-3. The tumor cells did not express CD31, FⅧRAg, HHF35, HMB45, Melan A, MyoD1, myogenin, S-100 protein and SMA. All 10 patients underwent radical resection or extended excision, with additional radiotherapy or chemotherapy. During the follow-up from October 2012 to August 2016, seven cases showed recurrences and metastases within 2 months to 2 years. Five patients died of the disease due to widespread metastases. Conclusions: ES may show a wide spectrum of morphology, and display a multidirectional differentiating capabilities including towards mesothelial and lymphatic endothelial cells. As such, its diagnosis and differential diagnosis are particularly important as it is easily confused with other tumors with similar morphology or immune phenotype.

Establishment of an Acquired Lymphedema Model in the Mouse Hindlimb: Technical Refinement and Molecular Characteristics.

BACKGROUND: The pathophysiology of secondary lymphedema remains poorly understood. This study aimed to establish a consistent mouse hindlimb lymphedema model for further investigation of the mechanism and treatment of lymphedema. METHODS: The authors developed a novel postsurgical lymphedema model in the mouse hindlimb by modifying previously described methods. Lymphedema in the hindlimb was created by removing both the inguinal and popliteal lymph nodes together with the surrounding fat pads, followed by silicone splint placement in the inguinal region. Using this modified mouse model, the authors analyzed lymphatic function, histologic changes, and the expression of lymphangiogenic factors including vascular endothelial growth factor C at various time points. RESULTS: The splinted lymphedema model showed a significant increase of edema formation in the hindlimb compared with the sham surgery control animals. Indocyanine green lymphography revealed lymphatic drainage impairment shown by dermal backflow and rerouting of lymph flow in the lymphedema model. Histopathologic and immunohistochemical examinations showed a significant increase of skin thickness and abnormally dilated lymphatics in the lymphedema model. The expression of lymphangiogenic factors in lymphedematous tissues of the splinted lymphedema model was significantly increased compared with controls, depending on the degree of lymphedema. CONCLUSIONS: This splinted lymphedema model closely simulates the volume response, histopathology, and lymphography characteristics of human acquired lymphedema. Given these similarities to human lymphedema, this refinement of a mouse hindlimb model of acquired lymphedema represents a promising platform for the study of lymphatic vascular insufficiency and for the evaluation of new therapeutic modalities.

VEGF receptor subtypes may serve as novel prognostic factors and putative indicators for anti VEGF receptor treatment response in renal cell carcinoma cases.

PURPOSE: Targeted therapies are novel treatment options for renal cell carcinoma (RCC). Of the target molecules investigated, vascular endothelial growth factor receptors (VEFGRs) were seldom evaluated. The current study investigated the prognostic significance of VEGFRs and IMP-3 as a potential prognostic markers. METHODS: Pathological material and clinical files of 100 patients with RCC were retrospectively evaluated. For each case, the clinical outcome and disease stage were assessed and resected materials were histologically reevaluated. VEGFR-2, VEGFR-3 and IMP-3 expression of tumor samples were analyzed with immunohistochemistry. These expressions were compared with prognosis and clinicopathological variables. RESULTS: Five-year overall survival (OS) was 80% in the whole cohort. Mean survival was 20.3±1.9 months in metastatic disease (95%CI:16.4-24.2). Two-year OS was 20% and 5-year OS was zero in the metastatic group. Survival was significantly longer in VEGFR-2 expressing group than in the nonexpressing group (78.7±2.6 vs 63.9±6; 95%CI:73.7-84 and 52.1-75.7, respectively; p=0.031). VEGFR-3 and IMP-3 expressions were not significantly correlated with survival. In the non-metastatic group mean OS was 82.6±2.1 months and 2- and 5-year OS were 96 and 88%, respectively. CONCLUSIONS: Since VEGFRs were expressed on all histological subtypes and significantly correlated with survival, assessment of VEGFR-2 and VEGFR-3 on tumor samples might serve as a putative prognostic factor in RCC cases. These expressions might identify a subset of patients that may benefit from antiangiogenic treatments targeting VEGFR receptors.

Interleukin 12 shows a better curative effect on lung cancer than paclitaxel and cisplatin doublet chemotherapy.

BACKGROUND: Interleukin 12 (IL-12) is a cytokine that has been reported to exhibit potent tumoricidal effects in animal tumor models. A combined approach using Paclitaxel and platinum-based doublet chemotherapy is the most commonly used backbone regimen for treating lung cancer. Despite numerous studies regarding the anti-tumor effects of IL-12 and the widespread use of conventional chemotherapy, few direct comparisons of IL-12 and conventional chemotherapy in the treatment of lung cancer have been performed. METHODS: We compared IL-12 to paclitaxel and cisplatin doublet chemotherapy in terms of efficacy against lung cancer in mouse models. The antitumor effect was measured by survival assays, histological analyses and imaging analyses. The cytokine levels were assessed using enzyme linked immunosorbent assay (ELISA) and flow cytometry (FACS). The spleen sizes were measured. CD31, CD105 and Vascular endothelial growth factor receptor 3 (VEGFR3) were analyzed using immunofluorescence. Matrix metalloprotein-9 (MMP-9) and cadherin 1 (CDH1) transcript levels were measured by quantitative PCR. Tumor cells apoptosis were examined by Tunel assay. RESULTS: The results showed that IL-12 treatment inhibited lung tumor growth, resulting in the long-term survival of lung cancer-bearing mice. Further examination revealed that IL-12 rapidly activated NK cells to secrete IFN-γ, resulting in the inhibition of tumor angiogenesis. In contrast, paclitaxel and cisplatin doublet chemotherapy did not show the expected efficacy in orthotopic lung cancer models; the IFN-γ levels were not increased after this treatment, and the number of peripheral lymphocytes was reduced. CONCLUSION: Together, these animal model data indicate that IL-12 shows a better curative effect than PTX + CDDP doublet chemotherapy.

Heterogeneity of Vascular Endothelial Growth Factor Receptors 1, 2, 3 in Primary Human Colorectal Carcinoma.

BACKGROUND: The vascular endothelial growth factor (VEGF) pathway plays an important role in growth and progression of human cancer, including colorectal carcinomas (CRC). The key mediators of VEGF signaling are VEGFR1, VEGFR2, and VEGFR3, part of a family of related receptor tyrosine kinases. The relative expression, activity, or interplay among these receptors may determine the response of CRC patients to anti-angiogenic therapies. MATERIALS AND METHODS: We developed technically sound immunohistochemical (IHC) assays to quantify VEGFR1, 2 and 3, and using a well-annotated CRC tissue microarray (TMA), we carried out comprehensive comparative evaluation of the three VEGFRs in archival primary CRC tissues (n=84). For each TMA core, tumor cell VEGFR1 expression was reported as H-score (range=0-300); vascular VEGFR2/VEGFR3 expression was manually scored as the number of receptor-positive tumor stromal vessels. Each case was defined as VEGFR1/ VEGFR2/VEGFR3-negative, low, medium or high. RESULTS: Based on the differential expression of the three VEGFRs, eight VEGFR staining profiles were observed: Triple VEGFR positive (n=12, 14%), VEGFR1 predominant (n=17, 20%), VEGFR2 predominant (n=7, 8%), VEGFR3 predominant (n=1, 1%), VEGFR1/2 predominant (n=39, 46%), VEGFR1/3 predominant (n=2, 2%), VEGFR2/3 predominant (n=3, 4%), and triple-VEGFR-negative (n=3, 4%). CONCLUSION: Herein we demonstrated heterogeneity of expression of VEGFRs in human CRC stromal vessels and tumor cells. The observed VEGFR expression-based subsets of human CRCs may reflect differences in biology of pathologic angiogenesis in primary CRC tissues. Furthermore, the heterogeneity of expression of VEGFRs unraveled in this analysis merits independent validation in larger cohorts of primary and metastatic human CRC tissues and in pertinent experimental models treated with various anti-angiogenic therapies.

Selective Stimulation of Cardiac Lymphangiogenesis Reduces Myocardial Edema and Fibrosis Leading to Improved Cardiac Function Following Myocardial Infarction.

BACKGROUND: The lymphatic system regulates interstitial tissue fluid balance, and lymphatic malfunction causes edema. The heart has an extensive lymphatic network displaying a dynamic range of lymph flow in physiology. Myocardial edema occurs in many cardiovascular diseases, eg, myocardial infarction (MI) and chronic heart failure, suggesting that cardiac lymphatic transport may be insufficient in pathology. Here, we investigate in rats the impact of MI and subsequent chronic heart failure on the cardiac lymphatic network. Further, we evaluate for the first time the functional effects of selective therapeutic stimulation of cardiac lymphangiogenesis post-MI. METHODS AND RESULTS: We investigated cardiac lymphatic structure and function in rats with MI induced by either temporary occlusion (n=160) or permanent ligation (n=100) of the left coronary artery. Although MI induced robust, intramyocardial capillary lymphangiogenesis, adverse remodeling of epicardial precollector and collector lymphatics occurred, leading to reduced cardiac lymphatic transport capacity. Consequently, myocardial edema persisted for several months post-MI, extending from the infarct to noninfarcted myocardium. Intramyocardial-targeted delivery of the vascular endothelial growth factor receptor 3-selective designer protein VEGF-CC152S, using albumin-alginate microparticles, accelerated cardiac lymphangiogenesis in a dose-dependent manner and limited precollector remodeling post-MI. As a result, myocardial fluid balance was improved, and cardiac inflammation, fibrosis, and dysfunction were attenuated. CONCLUSIONS: We show that, despite the endogenous cardiac lymphangiogenic response post-MI, the remodeling and dysfunction of collecting ducts contribute to the development of chronic myocardial edema and inflammation-aggravating cardiac fibrosis and dysfunction. Moreover, our data reveal that therapeutic lymphangiogenesis may be a promising new approach for the treatment of cardiovascular diseases.

Expression of vascular endothelial growth factor-C (VEGF-C) and its receptor (VEGFR-3) in the glial reaction elicited by human mesenchymal stem cell engraftment in the normal rat brain.

To determine whether vascular endothelial growth factor-C (VEGF-C) and its receptor (VEGFR-3) are involved in the glial reaction elicited by transplanted mesenchymal stem cells (MSCs), we examined the cellular localization of VEGF-C and VEGFR-3 proteins in the striatum of adult normal rats that received bone marrow-derived human MSCs. The MSC grafts were infiltrated with activated microglia/macrophages and astrocytes over a 2-week period post-transplantation, which appeared to parallel the loss of transplanted MSCs. VEGF-C/VEGFR-3 was expressed in activated microglia/macrophages recruited to the graft site, where the induction of VEGF-C protein was rather late compared with that of its receptor. VEGF-C protein was absent or very weak on day 3, whereas VEGFR-3 immunoreactivity was evident within the first three days. Furthermore, within three days, VEGF-C could be detected in the brain macrophages localized immediately adjacent to the needle track. At the same time, almost all the brain macrophages in both regions expressed VEGFR-3. Reactive astrocytes at the graft site expressed VEGFR-3, but not VEGF-C. These data demonstrated the characteristic time- and cell-dependent expression patterns for VEGF-C and VEGFR-3 within the engrafted brain tissue, suggesting that they may contribute to neuroinflammation in MSC transplantation, possibly through the recruitment and/or activation of microglia/macrophages and astrogliosis.

Roles of serum and biliary CEA, CA19-9, VEGFR3, and TAC in differentiating between malignant and benign biliary obstructions.

BACKGROUND/AIMS: Despite the presence of many diagnostic methods, the differential diagnosis between benign and malignant biliary obstructions is still not easy. We aimed to evaluate the role of serum/biliary carcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (CA 19-9), vascular endothelial growth factor receptor-3(VEGFR-3), and total antioxidant capacity (TAC) tests in this differential diagnosis. MATERIALS AND METHODS: Patients (n:225; 110♂, 115♀) with diagnosis of malignant (n:96) or benign (n:129) biliary obstruction were included in this cross-sectional study. Serum and biliary CEA, CA 19-9, VEGFR-3, and TAC tests were analyzed, statistics were obtained, and significance was defined as p<0.05. RESULTS: Mean age was 54.9±16.4 for the benign and 54.2±19.6 for the malignant group (p=0.89). Head of pancreas cancer (18.2%), cholangiocarcinoma (11.4%) and choledochal stone (48%) were the most common etiologies. The area under the curve (AUC)s by ROC analysis of serum/biliary CA 19-9, VEGFR-3, and TAC and serum CEA were 0.701/0.616, 0.622/0.663, 0.602/0.581, and 0713, respectively. Serum TAC had higher sensitivity (61.1%) and CEA had lower sensitivity (42.7%), whereas CEA had higher specificity (89.9%) and TAC had lower specificity (60.5%). In biliary tumor markers, CA 19-9 had higher sensitivity (74%) and VEGFR-3 had lower sensitivity (56.2%); however, VEGFR-3 had higher specificity (79.1%) and CA 19-9 had lower specificity (34.1%). Additionally, combination of serum CEA (p<0.001), CA 19-9 (p<0.001), VEGFR-3 (p<0.001), and biliary CA 19-9 (p=0.028) markers achieved 95% estimation probability, and the sensitivity, specificity, and accuracy were 88.5%, 45.7%, and 64%, respectively. CONCLUSION: Serum and biliary CEA, CA 19-9, VEGFR-3, and TAC tests would not be useful in the differentiation between malignant and benign biliary obstructions.

Localization and signaling patterns of vascular endothelial growth factors and receptors in human periapical lesions.

INTRODUCTION: Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are key players in vasculogenesis and are also involved in pathologic conditions with bone destruction. Vasculogenesis is critical for disease progression, and bone resorption is a hallmark of apical periodontitis. However, the localization of VEGFs and VEGFRs and their gene signaling pathways in human apical periodontitis have not been thoroughly investigated. The aim of this study was to localize VEGFs and VEGFRs and analyze their gene expression as well as signaling pathways in human periapical lesions. METHODS: Tissue was collected after endodontic surgery from patients diagnosed with chronic apical periodontitis. Periodontal ligament samples from extracted healthy wisdom teeth was also collected and used as control tissue. In lesion cryosections, VEGFs/VEGFRs were identified by immunohistochemistry/double immunofluorescence by using specific antibodies. A human VEGF signaling polymerase chain reaction array system was used for gene expression analysis comparing lesions with periodontal ligament samples. RESULTS: The histologic evaluation revealed heterogeneous morphology of the periapical lesions with various degrees of inflammatory infiltrates. In the lesions, all investigated factors and receptors were identified in blood vessels and various immune cells. No lymphatic vessels were detected. Gene expression analysis revealed up-regulation of VEGF-A and VEGFR-3, although not significant. Phosphatidylinositol-3-kinases, protein kinase C, mitogen-activated protein kinases, and phospholipases, all known to be involved in VEGF-mediated angiogenic activity, were significantly up-regulated. CONCLUSIONS: The cellular and vascular expressions of VEGFs and VEGFRs in chronic apical periodontitis, along with significant alterations of genes mediating VEGF-induced angiogenic responses, suggest ongoing vascular remodeling in established chronic periapical lesions.

Expression of vascular endothelial growth factor C in human pterygium.

Vascular endothelial growth factor C (VEGF-C) and its receptor VEGFR-3 mediate lymphangiogenesis. In this study, we analyzed the expression of VEGF-C and VEGFR-3 as well as lymphatic vessels in the pterygium and normal conjunctiva of humans. Fifteen primary nasal pterygia and three normal bulbar conjunctivas, surgically removed, were examined in this study. The lymphatic vessel density (LVD) and blood vessel density were determined by the immunolabeling of D2-40 and CD31, markers for lymphatic and blood vessels, respectively. VEGF-C and VEGFR-3 expression in pterygial and conjunctival tissue proteins was detected by Western blotting and were evaluated using immunohistochemistry. The LVD was significantly higher in the pterygium than normal conjunctiva (p < 0.05). Western blot demonstrated high-level expression of VEGF-C and VEGFR-3 in the pterygium compared with normal conjunctiva. VEGF-C immunoreactivity was detected in the cytoplasm of pterygial and normal conjunctival epithelial cells. The number of VEGF-C-immunopositive cells in pterygial epithelial cells was significantly higher than in normal conjunctival cells (p < 0.05). VEGFR-3 immunoreactivity was localized in the D2-40-positive lymphatic endothelial cells. The present findings suggest the potential role of VEGF-C in the pathogenesis and development of a pterygium through lymphangiogenesis and the VEGF-C/VEGFR-3 pathway as a novel therapeutic target for the human pterygium.

Papillary thyroid carcinoma involving cervical neck lymph nodes: correlations with lymphangiogenesis and ultrasound features.

Stratification of risk factors for cervical lymph node metastasis (LNM) in thyroid papillary carcinoma is important for providing standards for post-operative adjuvant radio-iodine therapy and for patient prognosis. We investigated pathological factors based on the lymphatic vessel system and radiological features associated with tumor with cervical neck LNM. Among patients who had undergone thyroidectomy confirmed to be papillary thyroid carcinoma, we selected 126 age-sex matched paired patients without cervical LNM (group 1) and with LNM (group 2) to evaluate risk factors. Pathological factors evaluated were size, multiplicity, and extra thyroid extension state, based on the pathological reports using stored data. The lymphatic vessel density (LVD) of each tumor was evaluated by staining for VEGFR-3 and D2-40 and correlated with cervical LNM state. Malignant ultrasound features were evaluated to compare the differences between these two groups. Larger tumor size, multiplicity, extrathyroid extension were more common in group 2 (p<0.05). The median percentage of VEGFR-3 for group 1 was 20 (range 0-30) and D2-40 was 13 (range 7-23) while for group 2, VEGFR-3 was 80 (70-90) and D2-40 was 78 (54-114). LVD measured by intratumoral D2-40 staining was 20.6% and 79.4% for group 1 and group 2, respectively. Intra-tumoral lymphatics measured by D2-40 stain had a strong correlation with cervical LNM (Odds 1.230, CI 1.01.-1.499 p value 0.040). Ultrasound (US) features had no significant differences between the two groups although calcifications tended to be higher in group 2 (84% vs. 76% p=0.264). Lymphatic vessel density and nodule echogenicity were not associated with LNM. Intratumoral lymphangiogenesis was most strongly associated with LNM and thus, could be a useful predictive marker for cervical LNM.

Vascular endothelial growth factors and their receptors and regulators in gestational trophoblastic diseases and normal placenta.

OBJECTIVE: To study the expression of vascular endothelial growth factors (VEGFs), placental growth factor (PLGF) and their receptors (VEGFR-1, -2, -3) and their regulators (IL-6, CD147) in normal placenta and gestational trophoblastic disease (GTD) in order to evaluate their potential role in the biology of GTD. STUDY DESIGN: Paraffin sections of 10 normal, first-trimester placentas, 10 partial moles, 10 complete moles, 5 choriocarcinomas and 5 placental site trophoblastic tumors (PSTTs) were studied immunohistochemically for expression of VEGFR-1, VEGFR-2, VEGFR-3, IL-6, PLGF and CD147. Immunolocalization of VEGF, Angiopoietin-1 and Angiopoietin-2 was performed on 5 choriocarcinomas and 5 PSTTs. The levels of VEGF and VEGFR-2 were determined in supernatants and lysates of normal trophoblast, JEG-3 and JAR choriocarcinoma cells with electrochemiluminescence assays. RESULTS: The normal placenta had significantly stronger expression of VEGFR-2 than did those of partial and complete mole (p = 0.001, p = 0.003). VEGF, Angiopoietin-1 and Angiopoietin-2 expression in PSTT were significantly higher than those in choriocarcinoma (p = 0.002, p= 0.01, p = 0.038). Choriocarcinoma showed stronger intensity of staining for VEGFR-3 than did normal placenta, partial and complete mole (p = 0.036, p = 0.038, p = 0.05). Choriocarcinoma had significantly stronger staining of CD147 than did partial and complete mole (p<0.01, p<0.01). PSTT exhibited significantly stronger staining for IL-6 than did choriocarcinoma (p = 0.03). CONCLUSION: PSTTs exhibited strong staining for VEGF, and choriocarcinoma showed strong staining for VEGFR-3. Agents that inhibit the activity of VEGF and VEGF receptors may prove to be useful in the therapy of gestational trophoblastic neoplasia.

Vascular endothelial growth factors and receptors are up-regulated during development of apical periodontitis.

INTRODUCTION: Apical periodontitis is a common inflammatory disease caused by persistent root canal infection and is characterized by bone resorption. Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) have been described in many pathologic and inflammatory conditions, but their involvement in the development of apical periodontitis has not been thoroughly investigated. The aim of this study was to quantify gene expression and localize VEGF-A, VEGF-C, and VEGF-D and VEGFR-2 and VEGFR-3 in a rat model of apical periodontitis. METHODS: Molar pulps were unilaterally exposed to the oral cavity for 10 or 21 days. Jaw sections were used for localization of VEGFs and VEGFRs with immunohistochemistry and identification of cells with double immunofluorescence. Gene expression analysis for VEGF-A, VEGF-C, and VEGFR-3 of periapical tissues was performed with quantitative real-time polymerase chain reaction. RESULTS: All investigated factors and receptors were expressed immunohistochemically in blood vessels at the periodontal ligament of control teeth and were up-regulated during lesion development. In apical lesions, macrophages and neutrophils expressed all studied factors and receptors, with macrophages being an important source of VEGF-C and VEGF-D. Osteoclasts expressed VEGFR-2 and VEGFR-3, and the latter was also identified in fibroblast-like cells in the lesions. VEGF-A and VEGFR-3 gene expression was up-regulated at days 10 and 21 (P < .05). CONCLUSIONS: The current findings indicate that the VEGF family and receptors are involved in vascular remodeling and immune functions during disease development. The presence of VEGFR-2 and VEGFR-3 on osteoclasts indicates that bone resorbing activity is influenced by VEGFs.