Different immunoreactivity of endothelial markers in well and poorly differentiated areas of angiosarcomas

This study evaluated the immunohistochemical staining of four endothelial cell markers in well differentiated and poorly differentiated areas of angiosarcomas. Formaldehyde-fixed, paraffin-embedded sections from eight angiosarcomas were studied using the antibodies anti-factor VIII-related antigen (FVIII-RA), Ulex europaeus I agglutinin, anti-CD34 (QBEND/10) and anti-CD31 (JC70). The immunostaining of the angiomatous (well differentiated) and solid (poorly differentiated) areas was separately analysed and specificity was evaluated in 20 non-vascular tumours. The antibody anti-CD31 and Ulex europaeus were the most sensitive markers staining well differentiated vasoformative structures and poorly differentiated solid areas. Anti-FVIII-RA and anti-CD34 did not stain undifferentiated malignant endothelial cells from solid areas. Ulex europaeus and anti-CD34 showed very low specificity; in contrast, none of the non-vascular tumours expressed CD31 or FVIII-RA. JC70 (anti-CD31) appears to be the most useful marker in elucidating the vascular nature of angiosarcomas. Is important to emphasize the lack of specificity of Ulex europaeus and the low sensitivity of anti-CD34 and anti-FVIII-RA for poorly differentiated lesions.     

Staining for factor VIII related antigen and Ulex europaeus agglutinin I (UEA-I) in 230 tumours. An assessment of their specificity for angiosarcoma and Kaposi''s sarcoma

In this study we examined the staining reactivity of commercially available antisera to factor VIII related antigen (F VIII RAg) and Ulex europaeus agglutinin I (UEA-I) on sections from 230 formalin fixed paraffin embedded tumours. These included 196 sarcomas, 20 carcinomas and 14 angiomas. All angiomas showed positive staining for F VIII RAg; all carcinomas showed negative staining; the vasoformative areas of all angiosarcomas stained positively but only four of six angiosarcomas showed positive staining of their solid areas; of seven Kaposi's sarcomas, all showed positive staining of vessels and six showed positive staining of the spindle cell component. In the remaining 181 non-vascular sarcomas there was a false positive result in four tumours (2.2%), three of which had a history of irradiation. Pre-radiotherapy biopsies of these three tumours stained negatively with anti-F VIII RAg. UEA-I was demonstrated in all the angiomas studied, in all angiosarcomas (including the solid components) and in well-formed vessels of all Kaposi's sarcomas, but only in the spindle cell component of 3/6. However, there was an unacceptably high rate of false positive staining amongst the carcinomas and non-vascular sarcomas. In conclusion, F VIII RAg is a specific but not a sensitive marker of angiosarcomas; UEA-I is a sensitive but not a specific marker of angiosarcomas.     

Cushing's disease arising from a clinically nonfunctioning pituitary adenoma

A 49-yr-old woman with a large pituitary tumor leading to visual loss and galactorrhea-amenorrhea was submitted to transcranial pituitary surgery, when a clinically nonfunctioning pituitary adenoma was partially removed. Histopathology and immunohistochemistry confirmed the diagnosis of “non-secreting atypical adenoma”. At that time, serum and urinary free cortisol were normal, with low T4 levels and hyperprolactinemia. The patient was discharged on thyroxine and bromocriptine and treated with conventional radiotherapy. Two years later, she presented high free urinary cortisol levels and a positive ACTH response to desmopressin testing on dexametasone 2 mg overnight. A pituitary biopsy confirmed aggressive growth as well as positive immunoreactivity for ACTH, p53, Ki-67, and c-erb-B2. The patient was then treated with radiosurgery on ketoconazole therapy. The overall clinical, laboratory, and pathological data suggest a transition from a clinically nonfunctioning to a hypersecreting ACTH-producing tumor. Putative mechanisms of tumor transformation and the possibility of a silent corticotropinoma evolving into clinical Cushing's syndrome are discussed.     

人喉鳞状细胞癌p53蛋白和PCNA的表达

目的：探讨Ｐ５３蛋白和增殖细胞核抗原（ＰＣＮＡ）在人喉鳞状细胞癌的表达。方法：应用免疫组织化学Ｓ－Ｐ法对８０例喉鳞状细胞癌、１０例声带息肉、１０例喉乳头状瘤进行检测。结果：（１）Ｐ５３蛋白在喉鳞状细胞癌阳性率为４１％，阳性细胞指数为５４．５９±２２．８２，与临床Ｔ分期，分化程度以及淋巴结转移无关；（２）ＰＣＮＡ指数在声带息肉，喉乳头状瘤，喉鳞的逐渐提高，在有淋巴结转移的比无淋巴结转移的指数明显增高     

星形细胞肿瘤p53蛋白和PCNA免疫组化及其临床病理意义

应用免疫组化和图象分析技术对人脑星形细胞肿瘤中抑癌基因ｐ５３蛋白（４７例）和增殖细胞核抗原（ＰＣＮＡ）（９７例）的定位、分布及反应强度与分级和预后的关系进行了研究。结果显示；３种ｐ５３蛋白抗体的阳性率为２３．４％～６６．７％，其中ＣＭ－１抗体阳性率高于ｐＡｂ１８０１和ｐＡｂ２４０；ｐ５３表达水平在Ⅱ～Ⅳ级高于１级，并与ＰＣＮＡ标记指数之间相关。ＰＣＮＡ反应强度既与分级相关，又与预后相关，对于分析星形细胞瘤增殖活性和恶性程度有重要意义。     

免疫磁珠分选系统在分离大鼠骨髓干细胞群中的应用

采用免疫磁珠分选系统(MACS)分离大鼠骨髓Thy-1.1 干细胞群。收集大鼠胫骨和股骨的骨髓细胞,Percoll密度梯度离心分离单个核细胞,Thy1.1单抗标记,间接MACS分离纯化Thy-1.1 干细胞群,流式细胞仪检测其纯度和分选前后CD34 百分比,台盼兰拒染法检测细胞活力,计算回收率。结果表明分选后的Thy-1.1 细胞纯度达94.2%,回收率64.7%;分选后细胞活力为99.6%,与分选前99.8%无明显差异;分选前CD34 百分比为1.2%,与分选后1.3%无差异。MACS能有效分选大鼠骨髓Thy-1.1 干细胞群,所得细胞纯度高,细胞活力保持好。大鼠Thy-1.1抗原与CD34分子无明显相关性。     

Expression of bcl-2 oncoprotein in renal cell tumours

Expression of bcl-2 is associated with inhibition of apoptosis and extension of cell survival. The importance of apoptosis in relation to the development and progression of renal cell neoplasia remains undefined so far. In order to determine the expression of bcl-2 oncoprotein in normal and neoplastic renal cells, 37 renal tumours were investigated by immunolabelling, including 13 clear cell carcinomas, ten tubulopapillary carcinomas, four chromophobic renal cell carcinomas, and ten oncocytomas. Twenty-six samples of adjacent normal renal tissue served as controls. bcl-2 expression was correlated with cell proliferation activity as estimated by Ki67 antigen expression, and p53 protein expression in the tumour samples. The results demonstrate that in the normal kidney, positive bcl-2 immunostaining was present in glomerular parietal epithelial cells, in distal tubular cells, and in sparse proximal tubule cells. Renal cell tumours showed heterogeneous bcl-2 expression according to the tumour cell type. While the majority of carcinomas of clear cell type were usually negative or contained sparsely distributed positive cells, all tubulopapillary carcinomas were consistently positive for bcl-2. In oncocytomas and chromophobic carcinomas, there was a low percentage of bcl-2 immunoreactive tumour cells; some nuclear bcl-2 positivity was detected in one chromophobic tumour. These findings indicate variable bcl-2 oncoprotein expression in different types of renal cell tumours, with the highest level of expression in tubulopapillary carcinomas. No clear relationship was found between nuclear grade, cell proliferation activity, and level of bcl-2 expression. p53 protein was detected in only one tubulopapillary carcinoma.     

小鼠p27Kip1基因的克隆及与外周B细胞凋亡和生存的关系

目的：克隆小鼠的p27^kip1基因，并以此为探针分析p27^kip1在外周血B细胞中的表达，以及其在抗原受体和CD40介导的抑制和促进增殖信号中的作用。方法：从培养的B细胞中提取总RNA，用RT-PCR扩增并克隆p27^kip1基因，用流式细胞仪检测B细胞凋亡，用Northern blot观察p27^kip1mRNA的表达水平。结果：成功地克隆了小鼠的p27^kip1基因，发现抗原受体交联在引起外周血B细胞凋亡之前，出现p27^kip1表达上调，若同时激活CD40则使p27^kip1表达降低，B细胞也免于凋亡用针对p27^kip1的反义寡核苷酸处理，可阻断抗原受体交联引起的外周血B细胞凋亡。结论：p27^kip1可能在抗原受体信号介导的B细胞凋亡和CD40信号介导的B细胞生存中发挥一定的作用。     

SV40大T抗原在人脑肿瘤中与p53形成特异性复合物

目的 探讨SV40早期区域基因编码产物大T抗原（Tag) 表达及与抑癌蛋白p53的相互作用在人脑肿瘤发生发展中的意义。方法 采用免疫共沉淀及Western印迹法检测43例人脑肿瘤组织及5例正常人脑组织中Tag的表达,并对18例Tag阳性瘤组织检测Tag-p53复合物的存在。结果 Tag在5例室管膜瘤及2例脉络丛乳头状瘤中全部表达,垂体腺瘤（5／6）、星形胶质细胞瘤（7／10）、脑膜瘤（4／6）、多形性胶质母细胞瘤（3／5）及髓母细胞瘤（2／5）均有Tag的表达,3例少枝胶质细胞瘤、1例松果体瘤及5例正常人脑组织无Tag表达;检测18例Tag阳性瘤组织均发现Tag与p53形成特异性复合物。结论 在人脑肿瘤组织中Tag广泛表达,Tag可与p53形成特异性复合物,Tag-p53特异性复合物的形成导致p53失活,可能是SV40致人脑肿瘤发生的一个重要机理。     

Expression of aquaporin 5 increases proliferation and metastasis potential of lung cancer

Water channel aquaporin 5 (AQP5) is highly expressed at the apical membrane of alveolar type I epithelial cells and confers high osmotic water permeability. AQP5 is also expressed in lung cancer tissue. Previous studies showed there was an up‐regulation of AQP5 expression in cancer tissue compared to surrounding normal tissue. In addition, expression of AQP5 in lung cancer tissue was associated with poor prognosis. Herein, we tested the role of AQP5 in lung cancer oncogenesis and development. Lung cancer cells with different expression of AQP5 were used to study cell proliferation and migration, two important parameters for tumour cell biology. We found enhanced proliferation and migration potential in cancer cells with high AQP5 expression, while reduced proliferation and metastasis potential in cancer cells with low AQP5 expression. Oncogene analysis showed significantly increased PCNA and c‐myc expression in AQP5 transfected cells. AQP5 transfected cells also showed significant increased MUC5AC mucin expression, which might contribute to the enhanced metastasis potential of lung cancer. AQP5 overexpression resulted in enhanced activation of the epidermal growth factor receptor (EGFR), extracellular receptor kinase (ERK1/2), and p38 mitogen‐activated protein kinase (p38 MAPK) pathway in cancer cells. Moreover, deletion of AQP5 demonstrated decreased activation of the EGFR/ERK/p38 MAPK pathway in AQP5 knockout mice lungs, while deletion of AQP1 or AQP3 did not exhibit significant changes on activation of the EGFR/ERK/p38 MAPK pathway in lung tissue. In conclusion, our results provide evidence for AQP5‐facilitated lung cancer cell proliferation and migration, possibly through activation of the EGFR/ERK/p38 MAPK signalling pathway, but why AQP5 but not other aquaporin expression affects the EGFR/ERK/p38 MAPK pathway still needs further exploration. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Extrapleural solitary fibrous tumor: clinicopathologic study of 17 cases and molecular analysis of the p53 pathway

Solitary fibrous tumor (SFT) occurring at various extrapleural sites is sometimes difficult to diagnose because of its histologic variability. Although a solitary fibrous tumor is usually a slow-growing tumor with favorable prognosis, a small number of malignant cases have been reported. In the present study, we examined the clinical behavior, histologic, immunohistochemical and molecular features of 17 cases of extrapleural SFT. Four tumors were located in the pelvic cavity, two in the nasal cavity, two were confined to the pulmonary parenchyma, and there was one each in the meninges, kidney, mediastinum, retroperitoneum, temporal region, neck, groin, buttock and thigh. Histologically, all the tumors were characterized by the presence of areas consisting of a proliferation of bland spindle cells with variable amounts of thick, often hyalinized or keloid-like intercellular collagen bundles. Highly cellular areas were observed in three tumors, frequent mitoses in two, and cellular pleomorphism and tumor necrosis in one each. All 17 tumors showed immunoreactivity to CD34 and 15 (88%) to bcl-2 protein. The labeling indices of p53, mdm2 protein and Ki-67 were generally low. PCR-SSCP and a subsequent sequence analysis of the p53 gene disclosed point mutation at codon 161 in exon 5 in one of the 13 cases analyzed. According to follow-up information, none of the patients had developed local recurrence or distant metastasis. Our results suggest that most extrapleural SFTs behave in a benign fashion even in a higher histologic grade group, and it is difficult to predict their clinical outcome. Complete surgical excision in order to obtain clear margins and long-term follow-up is advisable for patients with an extrapleural SFT.     

Expression of Complement Regulatory Proteins on Human Eggs and Preimplantation Embryos

PROBLEM : To investigate the relation between the complement system and reproduction, expression of complement regulatory proteins (C3b receptors and inhibitor of the membrane attack complex) were screened on unfixed human eggs and preimplantation embryos. METHODS : Unfixed unfertilized oocytes and preimplantation embryos obtained from an in vitro fertilization program were stained by indirect immunofluorescence using monoclonal antibodies raised against membrane cofactor protein, (MCP or CD46), decay accelerating factor (DAF or CD55), protectin (CD59), human C3b/C4b receptor (CR1 or CD35), and major histocompatibility complex class I antigen (MHC class I). RESULTS : CD55 and CD59 were both expressed by the plasma membrane of unfertilized oocytes and pre-implantation embryos. CD46 was not expressed by unfertilized oocytes but appeared at the 6-to-8 cell stage embryo when human gene expression first occurs. CD35 and MHC class I antigens were not expressed at all on oocytes and preimplantation embryos. CONCLUSIONS : Selective expression of complement regulatory proteins (DAF and protectin) associated with the lack of MHC class I antigens may represent an immune protective mechanism by which human oocytes and preimplantation embryos escape complement-mediated damage during their travel through the female genital tract. Furthermore, participation of these complement regulatory proteins including MCP in cell to cell interaction during fertilization and/or implantation cannot be excluded.     

P21,p53蛋白和增殖细胞核抗原在膀胱癌中的表达及意义

目的研究同一膀胱癌中癌基因蛋白产物和抗癌基因蛋白产物的表达。方法应用免疫组织化学ＡＢＣ方法检测５例正常膀胱组织和５８例膀胱癌组织中增殖细胞核抗原（ＰＣＮＡ）,Ｐ２１和ｐ５３蛋白的表达水平。结果５例正常膀胱组织中未发现Ｐ２１,ｐ５３和ＰＣＮＡ的表达,５８例膀胱癌组织中Ｐ２１,ｐ５３和ＰＣＮＡ阳性检出率分别为６２．１％、５５．２％和５８．６％。表达阳性的ＰＣＮＡ和ｐ５３均定位于肿瘤细胞核内,Ｐ２１蛋白定位于细胞膜上。Ｐ２１,ｐ５３和ＰＣＮＡ阳性表达率与膀胱癌的病理分级,临床分期和患者术后生存率关系密切。结论检测ｐ２１,ｐ５３和ＰＣＮＡ有助于评估膀胱癌的预后。     

p53 overexpression and mutation in metaplastic carcinoma of the breast: genetic evidence for a monoclonal origin of both the carcinomatous and the heterogeneous sarcomatous components

Metaplastic carcinoma of the breast (MCB) is characterized by the biphasic presence of both a carcinomatous component (CC) and heterogeneous sarcomatous components (HSCs). Although an epithelial or myoepithelial origin of the HSCs has been suggested, molecular evidence for a common origin for the CC and HSCs is limited and the mechanism underlying the sarcomatous or metaplastic change is unknown. The present study investigated the frequency and nature of p53 expression and mutation in 11 biphasic and three monophasic MCBs by immunohistochemistry and either needle-assisted or laser-capture microdissection, followed by PCR and direct sequencing. In all 11 biphasic MBCs, staining for p53 was concordant in the CC and HSCs (8/11 positive and 3/11 negative), consistent with a monoclonal origin of both components. Significantly, whenever a component of carcinoma in situ was present (5/11), the p53 staining was always concordant with that in the CC and HSCs. Screening of the 14 cases for p53 mutation identified four mutants, each in a single case of biphasic MCB with concordant p53 overexpression. Both the CC and each of the HSCs revealed identical p53 mutation in all four cases; in addition, one of the four cases also had an in situ component and the same mutant was found simultaneously in the in situ, invasive, and sarcomatous components. The concordant pattern of p53 alteration (overexpression or mutation) implies that early p53 mutation, occurring prior to invasion, was maintained throughout tumour progression and metaplastic change. The findings therefore support a monoclonal histogenesis of the various components, but are neutral regarding the role of p53 alteration in the development of metaplastic change in MCBs.     

Immunohistochemical assessment of Ki67 and p53 expression assists the diagnosis and grading of ulcerative colitis-related dysplasia

AIMS: To assess whether Ki67 and p53 immunostaining may assist the diagnosis and grading of ulcerative colitis-related dysplasia. METHODS AND RESULTS : Location of Ki67 staining and location and intensity of p53 staining were assessed in ulcerative colitis (UC) cases showing the features of high-grade dysplasia (HGD, n = 14), low-grade dysplasia (LGD, n = 22), 'indefinite for dysplasia' (n = 12), or regenerative atypia (RA, n = 22). Good intra- and inter-observer reproducibilities were demonstrated in the performance of these assessments. All the dysplasia cases showed extension of Ki67 staining above the basal third of the crypt. Moderate intensity p53 staining was seen in 10/22 RA cases, but strong intensity p53 staining was seen only in cases of dysplasia. All the cases of HGD showed extension of Ki67 and p53 staining above the basal two thirds of the crypt. CONCLUSIONS: Restriction of Ki67 staining to the basal third of the crypt appears to exclude a diagnosis of dysplasia whereas strong intensity p53 staining suggests a diagnosis of dysplasia. Restriction of Ki67 or p53 staining to the basal two-thirds of the crypt appears to exclude a diagnosis of HGD.     

Correlation of Nrf2, NQO1, MRP1, cmyc and p53 in colorectal cancer and their relationships to clinicopathologic features and survival

Due to emergence of resistant tumor populations, prognosis for metastatic colorectal cancer (CRC) patients remains poor and five-year survival rate is still very low. To guide clinicians in selecting treatment option for CRC patients, reliable markers predictive of poor clinical outcome are desirable. This study analyzed the correlation of NF-E2-related factor 2 (Nrf2), NAD(P)H quinine oxidoreductase 1 (NQO1), multidrug resistant protein 1 (MRP1), cmyc and p53 in CRC and their relationships to Duke’s stage and clinical prognosis. 76 specimens of CRC tissues were immunohistochemically investigated using Nrf2, NQO1, MRP1, cmyc and p53 antibodies. IHC stain showed that Nrf2, NQO1, MRP1, cmyc and p53 were highly expressed in CRC tissues compared with adjacent non-tumor tissues. Significant positive correlations were found between the expression of Nrf2 and that of NQO1, MRP1, cmyc and p53. Moreover, there was significant correlation between the high level of Nrf2, NQO1, MRP1, p53 expression and Duke’s stage, as well as poor clinical prognosis. We confirmed that Nrf2, NQO1, MRP1, and p53 expression exhibits considerable heterogeneity according to CRC clinical stage and prognosis. Nrf2 is the most promising biomarker in identifying a poor prognostic group of CRC.     

Creatine kinase subunits M and B as markers in the diagnosis of poorly differentiated rhabdomyosarcomas in children

Antibodies against the M and B subunits of creatine kinase were assessed for their usefulness in the diagnosis of poorly differentiated rhabdomyosarcoma. Routinely processed formaldehyde-fixed tissue and the avidin-biotin-peroxidase complex technique were used. The majority of the poorly differentiated and all of the moderately and well-differentiated rhabdomyosarcomas studied showed immunostaining for the M subunit. The rhabdomyoblastic component of malignant "triton" tumors was also positive. Staining, although weak compared with that of the rhabdomyosarcomas, was also observed in a few leiomyosarcomas, hemangioendotheliosarcomas, malignant fibrous histiocytomas, and ganglioneuroblastomas. On the other hand, staining for the B subunit was seen in many types of soft tissue tumors, including rhabdomyosarcoma, Ewing's sarcoma, and (ganglio)neuroblastoma. The results indicate that creatine kinase subunit M is a useful marker for distinguishing poorly differentiated rhabdomyosarcoma from other types of small round cell tumors in children, such as neuroblastoma, Ewing's sarcoma, and malignant lymphoma.