Induction of epithelial-mesenchymal transition and loss of podoplanin expression are associated with progression of lymph node metastases in human papillomavirus-related oropharyngeal carcinoma.

AIMS: To examine human papillomavirus (HPV) status, the expression of podoplanin and epithelial-mesenchymal transition (EMT) markers and lymphatic vessel counts (LVC) in oropharyngeal squamous cell carcinoma (OPSCC) tissues, and to evaluate whether these factors were associated with survival and nodal status. METHODS AND RESULTS: A total of 53 OPSCC specimens were evaluated for HPV status and expression of proteins such as podoplanin and EMT markers by immunohistochemistry. E-cadherin-negative and vimentin-positive specimens were defined as EMT-positive. Twenty-two OPSCCs were HPV-positive. There was significant progression of nodal status in patients with HPV-positive tumours (P = 0.0475). HPV-positive cases had significantly lower expression of podoplanin (P = 0.0016) and were more frequently EMT-positive (P = 0.0172). Podoplanin-negative cases and EMT-positive cases showed significantly more advanced nodal status than their respective counterparts (P = 0.0082 and P = 0.0186, respectively). LVC correlated with neither HPV nor nodal status. Multivariate analyses revealed that HPV infection was an independent marker of longer disease-specific survival (P = 0.014). CONCLUSIONS: HPV-positivity in OPSCC was associated with loss of podoplanin expression and with EMT induction, which resulted in progression of nodal status. The mechanisms leading to an improved prognosis in HPV-positive OPSCC patients requires elucidation, as this is inconsistent with the aggressive phenotype with lymph node metastases.

Clonal profiling of mixed lobular and ductal carcinoma revealed by multiplex ligation-dependent probe amplification and fluorescence in situ hybridization.

A needle biopsy of a mass in the right breast of a 36-year-old woman revealed invasive ductal carcinoma (IDC), and approximately 20% of cancer cells showed unequivocal membranous staining with the HercepTest. After systemic therapy with trastuzumab and paclitaxel followed by FEC (fluorouracil + epirubicin + cyclophosphamide), a right mastectomy was performed. By histological and immunohistochemical examinations, the resected tumor consisted mainly of E-cadherin-negative invasive lobular carcinoma (ILC), and the rest was ERBB2-positive IDC; thus, the diagnosis was mixed ductal and lobular carcinoma. Multiplex ligation-dependent probe amplification and fluorescence in situ hybridization (FISH) analyses revealed that ILC and IDC shared high-level amplification of CCND1 in homogeneously staining regions (HSR) and that IDC had an additional HSR-type amplicon of ERBB2. These findings strongly indicate that IDC and ILC had a common precursor cell with CCND1 amplification. Review of the biopsy specimen with FISH showed IDC with gene amplifications of CCND1 and ERBB2 as a minor component, IDC without amplification of CCND1 or ERBB2 as a major component, and a minute portion of ILC with CCND1 amplification. We speculate that chemotherapy and trastuzumab caused a marked reduction in IDC; however, ILC with CCND1 amplification was resistant to chemotherapy and grew.

Diagnosis and treatment of low-grade osteosarcoma: experience with nine cases.

BACKGROUND: Low-grade osteosarcoma, including low-grade central osteosarcoma and parosteal osteosarcoma, is an extremely rare variant, and the diagnosis is occasionally difficult. In this article we present cases of low-grade osteosarcomas that should be reviewed by a clinical oncologist. PATIENTS AND METHODS: Nine cases of histologically diagnosed Broder grade 1 osteosarcoma were retrospectively reviewed. The pathological diagnoses included parosteal osteosarcoma, low-grade central osteosarcoma, and low-grade chondroblastic osteosarcoma in four, four, and one cases, respectively. RESULTS: Duration from initial surgical intervention including biopsy to final diagnosis as low-grade osteosarcoma was a mean of 9.4 months. The initial benign diagnoses on biopsy specimens included fibrous dysplasia in three cases, chondroblastoma in one case, and a giant cell tumor in one case. The average number of histological examinations was 1.8. Low-grade osteosarcomas are well suited for biological reconstruction: seven cases were reconstructed by frozen autografts, distraction osteogenesis, or vascularized bone grafts. CONCLUSION: Low-grade osteosarcomas can be misdiagnosed as benign lesions, especially fibrous dysplasia. If the diagnosis of a low-grade osteosarcoma is not established on the basis of radiologic findings, care should be exercised, even when a biopsy suggests a benign lesion. Low-grade osteosarcomas should be treated with wide excision, even after an intralesional excision. Biological reconstruction might be a better option for low-grade osteosarcomas.

Lymphangiogenesis in regional lymph nodes predicts nodal recurrence in pathological N0 squamous cell carcinoma of the tongue.

AIMS: Cancer cells induce de-novo lymphatic vessel growth within draining lymph nodes before they metastasize. The aim of this study was to retrospectively evaluate lymph node lymphangiogenesis before the establishment of nodal recurrence in squamous cell carcinoma (SCC) of the tongue. METHODS AND RESULTS: Surgical specimens from 28 patients with pT2-T3N0M0 SCC of the tongue after local excision with supraomohyoid neck dissection were studied by immunohistochemistry. Intranodal lymphatic endothelium was highlighted by podoplanin staining to evaluate lymphatic vessel counts (LVCs). Primary tumour sections were examined for the expression of lymphangiogenic factors: vascular endothelial growth factor (VEGF)-C and VEGF-D. LVCs in regional lymph nodes were significantly increased in the cases with nodal recurrence (P=0.0013). Simultaneous increases in VEGF-C and VEGF-D expression were significantly associated with both an increase in LVC in regional lymph nodes (P=0.0001) and a decrease in the rate of survival without nodal recurrence (P=0.016). CONCLUSIONS: Knowing the status of lymphangiogenesis in the regional pN0 lymph nodes in tongue cancer would help in predicting which patients will develop nodal recurrence. The use of a therapeutic approach which blocks lymphangiogenic factors, such as VEGF-C and VEGF-D, may be beneficial in suppressing the lymphatic spread of tongue cancer with intense intranodal lymphangiogenesis.

Duodenal adenocarcinoma with neuroendocrine features in a patient with acromegaly and thyroid papillary adenocarcinoma: a unique combination of endocrine neoplasia.

A 67-year-old woman with familial clustering of thyroid papillary adenocarcinoma was diagnosed with acromegaly due to pituitary macroadenoma. She had multiple skin vegetations, but had no parathyroid and pancreas diseases. Before transsphenoidal surgery, she was further diagnosed as having a duodenal tumor and multiple hypervascular liver nodules. Biopsy specimens from the duodenal tumor and liver nodules were diagnosed histologically as moderately differentiated adenocarcinoma. Immunohistochemically, the tumor cells were positive for chromogranin, synaptophysin and somatostatin receptor 2a, suggestive for neuroendocrine features. After surgery, the patient was not in biochemical remission, and octreotide treatment was initiated. The duodenal cancer was treated with chemotherapy (neoadjuvant cisplatin and S-1). After 24 months, the patient's insulin-like growth factor I level had been normalized, and her liver tumors had not progressed macroscopically. This is a rare case of acromegaly associated with multiple endocrine tumors, not being categorized as conventional multiple endocrine neoplasia. Octreotide treatment might have had beneficial effects on our patient's duodenal adenocarcinoma and liver metastases, both directly via SSTR2a and indirectly via GH suppression, thereby contributing to their slow progression.

Epidermal growth factor induces cytokeratin 19 expression accompanied by increased growth abilities in human hepatocellular carcinoma.

Cytokeratin (CK) 19-positive hepatocellular carcinoma (HCC) has been reported to have a poor prognosis. The mechanism of the development of CK19-positive HCC remains to be studied. To clarify this, in vitro experiments were performed using human HCC cell lines (PLC-5, HepG2), and the phenotypic changes after stimulation with several growth factors were examined using quantitative reverse transcriptase PCR, western blotting, and immunofluorescence staining. In vivo experiments using human HCC specimens obtained from a total of 78 patients and clinicopathological analysis were also performed. Among the growth factors tested, epidermal growth factor (EGF) had prominent effects on inducing CK19 expression in PLC-5 and HepG2, which was accompanied by the reduced expression of α-fetoprotein in PLC-5. The induction of CK19 expression after EGF stimulation was accompanied by the phosphorylation of c-Jun-N-terminal kinase (JNK)/stress-activated protein kinase, which was blocked by the addition of JNK inhibitors. EGF also increased proliferative abilities and invasive properties of the HCC cell lines. In vivo, 9 (12%) of 78 HCC cases showed positive immunohistochemical staining of CK19. The extent of positive immunohistochemical signals of EGF, EGF receptor (EGFR), and JNK expression was significantly intense in CK-19-positive HCC than those of CK19-negative HCC. Clinicopathological analysis showed that CK19-positive HCC had a high incidence of portal vein invasion, extrahepatic metastasis and an early relapse, which was associated with the worsened 2-year disease free survival. These results indicate that the activation of the EGF-EGFR signaling pathway is associated with the development of CK19-positive HCC, and the EGF-induced increase in growth abilities of HCC may account for the poor prognosis of the patients.

Chondroblastoma with pulmonary metastasis in a patient presenting with spontaneous bilateral pneumothorax: Report of a case.

Chondroblastoma is a benign bone tumor with a relatively high incidence in older children and adolescents. Although it is generally regarded as a benign neoplasm, it sometimes grows aggressively or recurs but rarely metastasizes to the lung. We herein present a very rare case of a bilateral pneumothorax due to a pulmonary metastasis from a chondroblastoma. A 21-year-old man developed a bilateral pneumothorax 20 months after an operation for a chondroblastoma of the right ischium. A pertinent literature review revealed similar cases of chondroblastoma with pulmonary metastasis, but revealed no reports of a pneumothorax caused by a metastatic chondroblastoma.

Endothelial to mesenchymal transition via transforming growth factor-beta1/Smad activation is associated with portal venous stenosis in idiopathic portal hypertension.

Idiopathic portal hypertension (IPH) represents noncirrhotic portal hypertension of unknown etiology, mainly due to stenosis of peripheral portal veins. This study was performed to clarify the mechanism of portal venous stenosis in IPH from the viewpoint of the contribution of the endothelial to mesenchymal transition of the portal vein endothelium via transforming growth factor-beta1 (TGF-beta1)/Smad activation. In vitro experiments using human dermal microvascular endothelial cells demonstrated that TGF-beta1 induced myofibroblastic features in human dermal microvascular endothelial cells, including spindle cell morphology, reduction of CD34 expression, and induction of S100A4, alpha-smooth muscle actin, and COL1A1 expression, as well as the increased nuclear expression of phospho-Smad2. Bone morphogenic protein-7 preserved the endothelial phenotype of human dermal microvascular endothelial cells. Immunohistochemical analysis showed that endothelial cells of the peripheral portal veins in IPH were characterized by the decreased expression of CD34 and the enhanced nuclear expression of phospho-Smad2; these results also confirmed the expression of S100A4 and COL1A1 in the portal vein endothelium. Serum TGF-beta1 levels in patients with IPH were significantly higher than those of healthy volunteers and patients with chronic viral hepatitis/liver cirrhosis, while an elevation of serum bone morphogenic protein-7 levels was not observed. These results suggest that the endothelial to mesenchymal transition of the portal venous endothelium via TGF-beta1/Smad activation is associated with portal venous stenosis in IPH, and bone morphogenic protein-7 may therefore be a suitable therapeutic candidate for IPH.

A case of neurosarcoidosis with necrotizing granuloma expressing angiotensin-converting enzyme.

We described a case of neurosarcoidosis with necrotizing sarcoid granulomatosis in a 22-year-old man. Contrast-enhanced brain computed tomography scan and magnetic resonance imaging showed intracerebral multiple nodular lesions. Noncaseating and partial necrotizing granulomas were detected in the specimen resected by neurosurgery. In addition, immunohistochemical examination revealed the expression of angiotensin-converting enzyme in necrotizing granuloma. Thus, these findings were consistent with neurosarcoidosis. Clinical and pathological presentation, immunological features, and treatment modalities of neurosarcoidosis are discussed.

Biliary adenofibroma with ominous features of imminent malignant changes.

Biliary adenofibroma (BAF) is a rare, benign liver tumor. Herein, we report a case of BAF with histological features of imminent malignant changes. Ultrasound and CT revealed a solid 2.5-cm mass in the right liver lobe. The patient was asymptomatic and had no past medical history including liver disease. A general examination that included the abdomen and the laboratory data were normal. Because of the increase of its size, this tumor was surgically resected. Grossly, a 3.5-cm nodular mass abutted on the hepatic capsule was found, and its cut surface showed a well-circumscribed, whitish, and firm lesion that showed microcystic changes in the periphery and solid changes in the central parts. Histologically, the tumor showed a proliferation of tubulocystic structures embedded in a fibrous stroma. Microcysts were prevalent in the periphery, while tubular components with abundant fibrous stroma were in the central parts. The tubules were variably dilated and branched. This case closely resembled the previously reported cases of BAF, except that there were complicated papillary projections with fine fibrovascular cores in some of the microcysts and that the epithelial component in papillary projections showed dysplastic changes and increased cellular proliferative activities, implicating ominous features of imminent malignant changes. These dysplastic and papillary changes may be an intermediate lesion leading to malignancy, which have occasionally been reported in BAF.

Synovial sarcoma in knee joint, mimicking low-grade sarcoma confirmed by molecular detection of SYT gene split.

A 10-year-old boy underwent arthroscopic curettage for an intra-articular mass in knee joint. The tumor was diagnosed as low-grade fibrous sarcoma. Five years later, the patient presented with a recurrent tumor. The patient underwent a marginal excision with knee joint preservation and without adjuvant therapy. Two years after the last surgery, the patient is thriving with no evidence of recurrent or metastatic disease. The final diagnosis was synovial sarcoma confirmed via a SYT gene split performed with fluorescent in situ hybridization (FISH), although the tumor appeared as a low-grade fibrous type in a hematoxylin-eosin section. The first curetted specimen was also confirmed to bear a SYT gene split. Synovial sarcoma has been conventionally recognized as a high-grade sarcoma. Our patient had a tumor that exhibited the characteristics of both a histologically and clinically low-grade tumor. From the present case, we consider that low-grade variants of synovial sarcoma do exist although their existence remains controversial.

Low-grade B-cell lymphoma presenting primarily in the bone marrow.

Cases of low-grade B-cell lymphoma presenting primarily in the bone marrow are rare, and its clinicopathology remains unclear. We retrospectively examined patients with low-grade B-cell lymphoma presenting primarily in the bone marrow. Fourteen patients met the inclusion criteria, including 5 with lymphoplasmacytic lymphoma (LPL), 3 with chronic lymphocytic leukemia/small lymphocytic lymphoma, 2 with follicular lymphoma (FL), and 4 with low-grade B-cell lymphoma not otherwise specified (LGBCL-NOS). The median age was 69.5 years (range, 42-89 years), and a slight male predominance was noted (9 men and 5 women, 1.8: 1). Immunohistochemically, all cases were positive for CD20. One case was positive for CD138. Both cases of FL were positive for CD10 and B-cell lymphoma 2 (BCL-2), and immunoglobulin heavy locus (IgH)/B-cell lymphoma 2 rearrangement was observed by fluorescence in situ hybridization. The myeloid differentiation primary response gene (88) leucine to proline mutation was observed in 3 of 5 LPL, 1 of 2 FL, and 2 of 4 LGBCL-NOS patients. Paraproteinemia was observed in 10 patients; IgM and IgG paraproteinemia were observed in 6 and 3 patients, respectively. In this patient series, 3 patients had died at a median follow-up of 36.5 months; the cause of death of 1 LPL patient was malignant lymphoma itself. Thus, low-grade B-cell lymphoma presenting primarily in the bone marrow has various subtypes, and approximately one-third of the patients had LGBCL-NOS. The immunophenotypic features and myeloid differentiation primary response gene (88) leucine to proline mutation data of LGBCL-NOS suggested that some cases present with characteristics similar to those of LPL or marginal zone lymphoma.

Warthin's tumor associated with IgG4-related disease.

Immunoglobulin G4-related disease (IgG4-RD) is an inflammatory condition associated with elevated serum IgG4 levels and tissue infiltration by IgG4-expressing plasma cells. Several inflammatory conditions associated with IgG4-RD have been reported. Warthin's tumor is a benign parotid gland tumor consisting of oncocytic epithelial cells and lymphoid stroma containing lymphoid follicles with reactive germinal centers. This is the first report describing a case of Warthin's tumor with possible involvement of IgG4-RD. The patient was a 71-year-old man presenting with swollen right parotid and bilateral submandibular glands. He had a history of a Warthin's tumor of the left parotid gland, autoimmune pancreatitis, and an inflammatory abdominal aortic aneurysm. Laboratory tests revealed a high serum IgG4 level. Histological examination of the resected parotid gland showed a Warthin's tumor and a nodule showing severe lymphocytic infiltration containing many IgG4-positive plasma cells. This case shows the possible involvement of Warthin's tumor with IgG4-RD.

Utility of fluorescence in situ hybridization to detect MDM2 amplification in liposarcomas and their morphological mimics.

The atypical lipomatous tumor (ALT)/well-differentiated liposarcoma (WDLS) and the de-differentiated liposarcoma (DDLS) represent the most common category of liposarcomas. ALT/WDLSs and DDLSs are often difficult to distinguish from other tumors with similar morphological characteristics. In this study, we investigated whether the detection of amplified or overexpressed murine double-minute 2 (MDM2) can be a useful diagnostic ancillary aid. We used fluorescent in situ hybridization (FISH) and immunohistochemistry (IHC) to detect MDM2 amplification and protein overexpression, respectively, in 49 WDLSs, 5 DDLSs, 23 myxoid liposarcomas, 25 benign lipomatous tumors, and 75 spindle and pleomorphic sarcomas. MDM2 amplification was detected in 48 of 49 WDLSs, 5 of 5 DDLSs, 2 of 9 malignant peripheral nerve sheath tumors, and 2 of 10 myxofibrosarcomas. We did not detect MDM2 amplification in any of the benign lipomatous tumors. FISH-mediated detection of MDM2 amplification was the most valuable diagnostic aid for ALT/WDLS, as determined by using the Fisher exact test to compare two different diagnoses of 19 biopsies. On the contrary, unequivocal nuclear overexpression of MDM2 was found in only 10 of 50 ALT/WDLSs. The sensitivity and specificity of MDM2 amplification in distinguishing a DDLS from spindle and pleomorphic sarcomas were 100% and 95%, respectively, while those of MDM2 overexpression were 100% and 87%, respectively. In conclusion, our results indicate that FISH-mediated detection of MDM2 amplification is the most useful adjunct in the diagnosis of both ALT/WDLS and DDLS. However, IHC-mediated detection of MDM2 protein is useful only for the diagnosis of DDLS.

Evaluation of a new histologic staging and grading system for primary biliary cirrhosis in comparison with classical systems.

Recently, our research team proposed a new histologic staging and grading system for primary biliary cirrhosis (PBC) that takes into account necroinflammatory activity and histologic heterogeneity. The present study aimed to confirm the usefulness of the new evaluation system. A total of 152 liver biopsy specimens and clinical data (including outcomes in patients with PBC before treatment with ursodeoxycholic acid) were analyzed with respect to the new system. Staging was evaluated on the basis of 3 histologic components (fibrosis, bile duct loss, and deposition of orcein-positive granules), and grading was assessed on the basis of chronic cholangitis activity and hepatitis activity. Concurrently, the classical systems, that is, the Scheuer and Ludwig staging systems, were also assessed and compared with our new system. PBC cases showed different distributions in each stage of the 3 systems. The new staging and grading system reflected liver dysfunctions before specific treatment. This was on a par with the results obtained using the classical systems. Development of cirrhosis-related conditions correlated well with the new staging system compared with the 2 classical staging systems, and in particular, the amount of deposition of orcein-positive granules could reflect development of cirrhosis-related conditions (scores 0-1 versus scores 2-3 groups, P < .0001). In conclusion, the new PBC staging system was demonstrated to reflect clinicolaboratory features, and its progression was associated with the development of cirrhosis-related conditions.

Distinct morphologic, phenotypic, and clinical-course characteristics of indolent peripheral T-cell lymphoma.

Peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS) consists of a heterogeneous group of lymphomas. Patients generally show an aggressive clinical course and very poor outcome. Although the 2008 World Health Organization classification of PTCL-NOS includes 3 variants, low-grade lymphoma is not included. Of 277 PTCL-NOS cases recorded in our consultation files, we examined the clinicopathologic characteristics of 10 patients with T-cell lymphomas composed of small-sized cells with slight nuclear atypia. Eight patients showed extranodal involvement (5 patients, spleen; 3 patients, thyroid), and 5 patients were at clinical stage I or II. Histologically, all samples presented diffuse infiltrate of small lymphoid cells, with few mitotic figures. Immunohistologically, all samples were positive for CD3, and CD20 was detected in 5 samples. All samples showed a low Ki-67 labeling index (mean, 1.05%), and 7 samples were positive for central memory T-cell markers. Clonal T-cell receptor γ chain and/or α-ß chain gene rearrangements were detected in all 10 patients. Five patients received chemotherapy, whereas for 3 patients, treatment consisted only of observation following surgical resection of the spleen or thyroid. Nine patients were alive at a median follow-up time of 19.5 months, whereas 1 patient died of an unrelated disease. The present study strongly indicates that T-cell lymphoma with small-sized lymphoma cells and a low Ki-67 labeling index is a distinct variant. Recognition of this novel lymphoma subtype, which should not be defined merely as PTCL-NOS, should be seriously considered.

Induction of lymphangiogenesis through vascular endothelial growth factor-C/vascular endothelial growth factor receptor 3 axis and its correlation with lymph node metastasis in nasopharyngeal carcinoma.

The contribution of the lymphatic system to tumor metastasis is being increasingly appreciated through studies of human cancers. As the biological behavior of nasopharyngeal carcinoma (NPC) depends on its nodal status, patients with advanced nodal status show a higher tendency toward a poor outcome. Here, we examined the role of lymphangiogenesis on lymphatic spread of NPC. We also evaluated the involvement of vascular endothelial growth factor (VEGF)-C/VEGF receptor 3 (VEGFR3) signaling pathway on lymphangiogenesis in NPC. Furthermore, we tested whether Epstein-Barr virus (EBV)-latent membrane protein (LMP) 1 induces VEGF-C. Forty-one patients with NPC were evaluated for expressions of VEGF-C and its receptor, VEGFR3, and LMP1 proteins and lymphatic vessel counts (LVC) highlighted by anti-podoplanin employing immunohistochemistry. The VEGF-C induction by LMP1 was then tested with Western blotting and enzyme-linked immunosorbent assay in vitro. The LVC and VEGF-C expression were significantly higher in cases with advanced regional lymph node metastasis (N2,3) than those with no or limited lymph node involvement (N0,1) (p=0.0380 and p=0.0109, respectively). In VEGF-C/VEGFR3-positive cases, the LVC were significantly increased compared with VEGF-C/VEGFR3-negative cases (p=0.0007). However, LMP1 expression did not show significant associations with LVC and VEGF-C-expression scores (p=0.1210 and p=0.1324, respectively). Induction of VEGF-C protein by LMP1 was not detected in vitro. These results suggest the involvement of the VEGF-C/VEGFR3 axis in the induction of lymphangiogenesis which results in lymphatic spread of NPC. However, EBV-LMP1 was not associated with the mechanism.

Epithelioid glioblastoma changed to typical glioblastoma: the methylation status of MGMT promoter and 5-ALA fluorescence.

A 55-year-old man was admitted to our hospital complaining of left hemiparesis. Magnetic resonance imaging (MRI) showed a smooth ring-like enhanced cystic tumor in the right parietal lobe. He underwent gross total resection of the tumor under neuronavigation and 5-aminolevulinic acid (5-ALA) fluorescence guiding method. Histopathological examination of the tumor showed small cells formed epithelioid solid nests with some focus of duct-like structure. On the basis of the MRI and operative and histological findings, this tumor was diagnosed as a metastatic poorly differentiated carcinoma, although the primary cancer could not be detected by metastatic work-ups. Afterward, this tumor recurred repeatedly. Histopathological examination of specimen from the fourth surgery indicated that the tumor was a glioblastoma (GBM). In the review of the histology and immunohistochemistry of the first tumor, atypical fibrillary cells were seen between solid nests and positive for glial fibrillary acidic protein, therefore the tumor was retrospectively diagnosed as epithelioid GBM. We assessed whether the changes in histopathology were accompanied by changes in the methylation status of O6-methylguanine methyltransferase (MGMT) promoter and the status of 5-ALA fluorescence. The methylation status of the MGMT promoter was found to have changed from methylated to unmethylated and 5-ALA fluorescence became positive along with the histological change.