Immunohistochemical Examination of Novel Rat Monoclonal Antibodies against Mouse and Human Podoplanin

This study aims to develop new monoclonal antibodies (mAbs) against mouse and human podoplanin. Rats were immunized with synthetic peptides, corresponding to amino acids 38–51 of mouse podoplanin or human podoplanin which is 100% homologous to the same site of monkey podoplanin; anti-mouse podoplanin mAb PMab-1 (IgG_2a) and anti-human mAb NZ-1.2 (IgG_2a) were established. In immunocytochemistry, the mouse melanoma B16-F10 and mouse podoplanin (mPDPN)-expressed CHO transfectant were stained by PMab-1; human lymphatic endothelial cells (LEC) and human podoplanin (hPDPN)-expressed squamous cell carcinoma HSC3 transfectant, were stained by NZ-1.2. Western-blot analysis detected an about 40-kDa protein in CHO-mPDPN and B16-F10 by PMab-1, and in HSC3-hPDPN and LEC by NZ-1.2. In frozen sections, PMab-1 reacted with mouse kidney, pulmonary alveoli, pulmonary pleura, and salivary gland myoepithelial cells while NZ-1.2 reacted to the human salivary gland myoepithelial cells. The immunostaining of paraffin-embedded sections also showed the reaction of PMab-1 or NZ-1.2 to the mouse or monkey kidney glomerulus, pulmonary alveoli, and lung lymphatic vessels. These results indicate that the two novel rat mAbs to the mouse and human/monkey podoplanin are useful for Western-blot and immunostaining of somatic tissues on paraffin-embedded sections as well as frozen sections.     

Diagnostic utility of D2-40 and podoplanin in effusion cell blocks

The distinction between malignant mesothelioma and adenocarcinoma is a diagnostic challenge in cytologic specimens of effusion fluids. As for today, no single antibody has demonstrated absolute sensitivity or specificity for Mesothelioma. D2-40 and podoplanin have recently been recognized to stain mesothelial cells. Our aim for this study was to evaluate the utility of these two markers as indicators of mesothelial cells using cell blocks by comparison with two other established mesothelial markers. A total of 40 cell blocks of effusion fluids including cases of epithelioid mesotheliomas, metastatic carcinomas and benign cases with reactive mesothelial cells were selected. A panel of immunostains including D2-40, podoplanin, CK5, and calretinin was performed. D2-40 and podoplanin were positive in 100% of mesothelioma cases in comparison to metastatic adenocarcinoma cases where the positivity was 0%. It is concluded that D2-40 and podoplanin are very useful markers for mesotheliomas. Since these markers are extremely helpful in differentiating epithelioid mesothelioma from metastatic adenocarcinoma, they shall be a valuable addition to the battery of markers used to differentiate the two entities.     

Tissue factor expression and tumor-infiltrating T lymphocytes in ovarian carcinomas and their association with venous thromboembolism

Ovarian cancer is a known risk factor of venous thromboembolism (VTE). Thrombogenic factor expression and lymphocytic infiltrate have been reported in endometriosis and ovarian cancers. We reviewed 30 cases of ovarian carcinomas (high grade serous carcinoma, 10; endometrioid carcinoma, 10; clear cell carcinoma (CCC), 10) and 16 endometriotic lesions. We immunohistochemically investigated the expressions of tissue factor (TF), podoplanin, P-selectin, and number of CD4 and CD8 positive lymphocytes in cancer tissue and endometriotic lesions, along with their relationship with VTE. The expression of TF was higher in CCC. The TF expression and the number of CD8 positive cells were higher in cancer tissues with VTE than in those without VTE. The podoplanin or P-selectin expression did not differ among histological types or between cases with and without VTE. Our results demonstrated a high TF expression and intraepithelial CD8 cells in CCC, which were associated with VTE. The results suggest that infiltrating lymphocytes may affect TF expression that, in turn, influences VTE.     

podoplanin在非小细胞肺癌组织中的表达及其与肿瘤淋巴转移的关系

目的研究podoplanin在非小细胞肺癌(NSCLC)组织中的表达及其与肺癌淋巴结转移的关系。方法应用免疫组化方法检测46例NSCLC组织标本和8例炎性假瘤组织标本中podoplanin、CD31和血管内皮生长因子受体-3(VEGFR-3)的表达,并记数淋巴管密度(LVD);分析podoplanin表达和肺癌淋巴转移的关系。结果Podoplanin显示的LVD与CD31显示的血管密度值(MVD)、VEGFR-3显示的微管密度值不相关(Spearman相关系数0.23,P=0.61;0.09,P=0.84);VEGFR-3阳性管数与CD31阳性管数相关(系数0.80,P=0.031,P<0.05)。在NSCLC组织LVD明显高于肺炎性假瘤组织LVD(P<0.01),与淋巴结转移阴性组比较,淋巴结转移阳性组LVD明显增高(P<0.05)。结论在NSCLC组织中,podoplanin是合适的淋巴管标记物,而VEGFR-3标记的微管多数是血管而不是淋巴管;podoplanin可作为判断NSCLC淋巴转移的指标,进而评价肺癌患者的预后。     

Significance of podoplanin expression in keratocystic odontogenic tumor

J Oral Pathol Med (2010) 39 : 110–114
Background:  The most important clinical features of the keratocystic odontogenic tumor (KCOT) are its potential for locally destructive behavior, a tendency to recur, and its origin in the odontogenic epithelium. The clinical features of KCOT are similar to those of ameloblastoma (AM). Histologically, KCOT is distinguished from jaw cyst with keratinization (orthokeratinized odontogenic cyst; OOC). However, current scientifically based clinical parameters cannot predict any potential for neoplastic behavior, or aggressive and localized invasiveness, in patients with KCOT. We have shown that podoplanin, a lymphatic endothelial marker, is highly expressed in AM. The purpose of this study was to determine the usefulness of podoplanin for reclassification of the odontogenic keratocyst (OKC) from cyst to tumor status.
Methods:  Paraffin-embedded tissue specimens of 57 OKCs (46 KCOTs and 11 OOCs) and 15 dentigerous cysts (DCs) were immunohistochemically examined using antibody against podoplanin.
Results:  Immunohistochemical reactivity for podoplanin was detected in the cell membrane and cytoplasm of most of the basal and suprabasal layer, areas of budding basal cell proliferation, epithelial nests and peripheral cells of daughter cysts in the stromal connective tissue in KCOTs. In the case of OOC and DC, only cases associated with inflammation were positive for podoplanin.
Conclusion:  Podoplanin is strongly expressed in KCOTs in comparison with OOCs. The pattern of staining for podoplanin in KCOT could be related to its neoplastic nature, and suggests a role of the protein in tumor invasiveness.     

Lymphatic Neoangiogenesis in Human Renal Allografts: Results from Sequential Protocol Biopsies

Neoangiogenesis of lymphatic vessels may be important for the cellular immune response in renal transplants. To determine the prevalence and chronology of lymph vessel proliferation and its relation to cellular infiltrates and allograft function, we analyzed sequential protocol biopsies (n = 162), taken at 6, 12 and 26 weeks after transplantation. Biopsies were stained with an antibody against podoplanin and lymphatic vessel density was quantified per square millimeter. The prevalence of lymph vessel-positive biopsies and the lymph vessel density were similar at 6, 12 and 26 weeks after transplantation. Biopsies with acute cellular rejection showed no significantly different lymph vessel density compared to those below the threshold for acute rejection or chronic allograft nephropathy. While lymphatic neoangiogenesis was equally prevalent in biopsies with and without infiltrates, the lymph vessel density was significantly higher in areas with cellular infiltrates than in areas without. Graft function at 1 year after transplantation was better in cases with lymph vessels in their infiltrates compared to cases with lymph vessel-free infiltrates. In conclusion, lymphangiogenesis not only shows a clear association with cellular infiltrates but might also have an impact on the pathogenicity of these cellular infiltrates.     

肝细胞癌中podoplanin的表达及其与淋巴道转移关系的研究

目的 观察肝细胞癌组织淋巴管内皮细胞podoplanin的表达,为研究肝细胞癌淋巴道转移的分子机制提供形态学依据.方法 用SP免疫组化检测法对肝细胞癌组织、淋巴结及正常肝组织中podoplanin的表达情况进行观察;用podoplanin来标记淋巴管,检测癌组织中微淋巴管密度.结果 肝细胞癌组织中血管内皮细胞podophnin不染色,淋巴管内皮细胞podoplanin阳性表达.经计数微淋巴管数量,癌组织和癌旁肝组织微淋巴管密度明显高于正常肝组织(P<0.01).结论 podophnin可选择性的在肝细胞癌组织淋巴管内皮表达,能明确的区分血管及淋巴管成分,可作为一个淋巴管内皮细胞较特异的标记物.由于微淋巴管密度在癌组织中的表达明显增高,说明淋巴管数量的增加可能与肝细胞癌的淋巴转移有关.     

First international consensus on the methodology of lymphangiogenesis quantification in solid human tumours

The lymphatic system is the primary pathway of metastasis for most human cancers. Recent research efforts in studying lymphangiogenesis have suggested the existence of a relationship between lymphatic vessel density and patient survival. However, current methodology of lymphangiogenesis quantification is still characterised by high intra- and interobserver variability. For the amount of lymphatic vessels in a tumour to be a clinically useful parameter, a reliable quantification technique needs to be developed. With this consensus report, we therefore would like to initiate discussion on the standardisation of the immunohistochemical method for lymphangiogenesis assessment.