Intercellular junctions in Ewing sarcoma/primitive neuroectodermal tumor: additional evidence of epithelial differentiation.

Ewing sarcoma/primitive neuroectodermal tumor (ES/PNET) has recently been shown to frequently express cytokeratins, suggesting partial epithelial differentiation. Older ultrastructural studies have documented primitive cell-cell junctions in ES/PNET, reportedly resembling poorly formed desmosomes. Recently, paraffin-reactive antibodies have become available to proteins found in a variety of intercellular junctions indicative of epithelial differentiation, including tight junctions, desmosomes and adherens junctions. We examined intercellular junction protein expression in a large number of genetically confirmed ES/PNET. Formalin-fixed, paraffin-embedded sections from 23 primary and seven recurrent or metastatic cases of genetically confirmed ES/PNET were immunostained for claudin-1 and occludin (tight junction structural proteins), zonula occludens-1 (ZO-1, tight junction linker protein), desmoglein 1/2 (desmosomal adherens protein), desmoplakin (desmosomal structural protein) and E-cadherin (epithelial adherens junction protein), using steam heat-induced epitope retrieval and the Dako Envision system. Cases with >5% positive cells were scored as 'positive'. Normal colonic epithelium and skin served as external positive controls. Claudin-1 was expressed by 19 of 30 specimens (63%), ZO-1 was expressed by 15 of 29 specimens (51%), and occludin was expressed by three of 28 specimens (11%). In 28 specimens all three tight junction markers were evaluable. In all, 15 samples (54%) expressed only one tight junction marker, and 10 samples (36%) expressed two tight junction markers. No case expressed all three tight junction markers. Desmoglein was expressed in one of 30 (3%) samples. Desmoplakin was expressed in two of 28 (7%) samples. E-cadherin was negative in all cases. Our data suggest that many of the previously described cell-cell junctions in ES/PNET are poorly formed tight junctions, given the high frequency of claudin-1 and ZO-1 expression. This may underestimate the true frequency of tight junction protein expression in ES/PNET, as there are at least 20 different claudins and other ZO proteins. These tight junctions are almost certainly abnormal, given the absence of occludin expression in most cases. Desmosomal and adherens junction protein expression was rare to absent. Our findings provide additional evidence that ES/PNET frequently show partial epithelial differentiation.     

滑膜肉瘤的融合基因检测分析

目的：基于存在染色体易位所致特异的SYT－SSX融合基因,证实可在滑膜肉瘤组织石蜡切片上检测出并探讨其在诊断中的价值。方法：采用逆转录－聚合酶链反应,检测并分析20例滑膜肉瘤（组织学亚型15例单相,5例双相）的SYT－SSX转录物,并对照相应病理学所见,结果：所检20例滑膜肉瘤中,有19例（95％）出现特异SYT－SSX逆转录聚酶链反应产物,其中13例具有SYT－SSX2融合基因的肿瘤有10例呈组织学单相分化。结论：SYT－SSX融合基因转录物可在石蜡切片和组织块中获得满意结果。具有较好的灵敏性,是滑膜肉瘤所特有的诊断标志物,它的亚类分型（SYT－SSX1和SYT－SSX2）,可能成为预后推测指征。     

Alterations in tight junction molecules of uterine epithelial cells during early pregnancy in the rat

Distribution patterns of the tight junction associated proteins ZO-1, claudin-1 and occludin were investigated in rat uterine epithelial cells during early pregnancy. Light microscopy and immunohistochemical labelling were used to detect these proteins on days 1, 3, 6 and 7 of pregnancy. Intense staining of claudin-1 at the apical region of the lateral plasma membrane accompanied diffuse staining throughout the cytoplasm. ZO-1 was also localised in the apical region, but ZO-1 was not present in the lower two thirds of the lateral plasma membrane or in the cytoplasm. Occludin was present only on days 6 and 7 of pregnancy. Labelling was also localised in the apical region of the lateral plasma membrane where tight junctions are known to be present. Our results show that ZO-1, claudin-1 and occludin are present in the apical region of uterine epithelial cells, and appear to play a role in the very dynamic tight-junctional network of uterine epithelial cells during early pregnancy. In particular, occludin appears only during uterine receptivity for implantation.     

滑膜肉瘤石蜡包埋组织SYT-SSX融合基因检测的临床病理意义

目的：探讨在石蜡包埋组织中检测SYT－SSX融合基因的可行性及其对滑膜肉瘤的诊断,分型和鉴别诊断的价值。方法：收集滑膜肉瘤标本38例,以恶性周围神经鞘膜瘤,纤维肉瘤,平滑肌肉瘤,尤文肉瘤,血管外皮肉瘤和转移性腺癌作为对照,共40例,均为甲醛固定,石蜡包埋组织,用逆转录－聚合酶链反应（RT－PCR）方法检测SYT－SSX融合基因mRNA表达,以看家基因PBGD作为内对照检测mRNA质量。结果：78例标本中64例（占82．1％）可检出PBGD mRNA表达,38例滑膜肉瘤中33例中可检出SYT－SSX融合基因mRNA表达,对照组无一例检出SYT－SSX基因,去除PBGD及SYT－SSX均阴性病例1例,滑膜肉瘤SYT－SSX融合基因检出率为89．2％（33／37）,33例SYT－SSX阳性滑膜肉中,SYT－SSX1型22例,SYT－SSX2型6例,5例无法区分。融合基因类型与滑膜肉瘤组织学类型有关。10例双相型滑膜肉瘤均为SYT－SSX1型,而18例单相型滑膜肉瘤中SYT－SSX1型12例,SYT－SSX2型6例,二者差异有统计学意义（P＜0．05）,结论：（1）从石蜡包埋组织中检测SYT－SSX融合基因对滑膜肉瘤有较高的敏感性和特异性,可用于滑膜肉瘤的诊断和鉴别诊断;（2）SYT－SSX融合基因类型与滑膜肉瘤组织学类型相关,SYT－SSX2型仅见于单相型。     

34例滑膜肉瘤分子遗传学改变的诊断学意义

目的 探讨石蜡包埋滑膜肉瘤组织中t(x；18)(p11．2；q11．2)染色体易位融合基因SYT-SSX mRNA表达的诊断学意义和应用价值。方法 收集滑膜肉瘤标本34例，以14例梭形细胞肉瘤和小圆细胞肉瘤做对照(包括2例纤维肉瘤、2例平滑肌肉瘤、1例恶性神经鞘膜瘤、4例Ewing肉瘤、2例腺泡型横纹肌肉瘤、2例恶性黑色素瘤、1例血管外皮瘤)。在进行免疫组织化学指标检测的基础上，用一步法逆转录-聚合酶链反应(RT-PCR)技术检测34例石蜡包埋滑膜肉瘤组织中SYT-SSX的表达。结果 34例滑膜肉瘤中30例获得有效RNA，28例(93．3％)检出SYT-SSX融合基因表达。其中14例表达SYT-SSXl型者中10例为双相型，9例表达SYT-SSX2型者中5例为单相分化型，5例SYT-SSXl／2均未检出。对照组均未检出SYT-SSX基因的表达。结论 SYT-SSX融合基因表达可作为诊断滑膜肉瘤新的分子诊断指标。一步法RT-PCR是一种理想而可行的用于石蜡包埋滑膜肉瘤组织SYT-SSX融合基因检测的分子诊断技术。     

Claudin-1 is a strong prognostic indicator in stage II colonic cancer: a tissue microarray study.

Tight junction associated proteins are key molecular components governing cellular adhesion, polarity and glandular differentiation. Tight junction proteins also play critical roles in cellular proliferation and neoplastic pathways via their functions as couplers of the extracellular milieu to intracellular signaling pathways and the cytoskeleton. Neoplastic cells frequently exhibit structural and functional deficiencies in the tight junction. The purpose of this study was to determine the pattern of expression and prognostic value of four tight junction associated proteins, claudin-1, claudin-4, occludin and ZO-1 in a cohort of TNM stage II colon cancer using tissue microarray technology. In this study, we retrospectively analyzed, resected and otherwise untreated paraffin embedded specimens from 129 consecutive patients with TNM stage II colonic carcinomas for claudin-1, claudin-4, occludin and ZO-1 protein expression by immunohistochemistry. Seventy-five, 58, 56 and 44% of the tumors exhibited normal to elevated expression levels (+2 and +3 immunopositivity) of claudin-1, claudin-4, occludin and ZO-1 respectively. Low expression levels of claudin-1 and ZO-1 were directly associated with higher tumor grade (P=0.05 and 0.03 respectively). Multivariate analysis indicated that lymphovascular invasion (P=0.01) and low levels of claudin-1 (P=0.0001) expression were independent predictors of recurrence and that reduced claudin-1 expression (P=0.0001) was associated with poor survival. This study is the first to comprehensively examine the expression of several tight junction associated proteins in colonic neoplasms and to correlate their expression with disease progression. Loss of claudin-1 expression proved to be a strong predictor of disease recurrence and poor patient survival in stage II colon cancer.     

IL-8对血管内皮细胞紧密连接的影响

目的 研究白细胞介素-8(IL-8)对血管内皮细胞紧密连接的影响.方法 采用免疫荧光染色检测经不同浓度和时间IL-8处理后EA.hy926细胞的3种紧密连接蛋白occludin、claudin-5和ZO-1的形态和分布;逆转录PCR(RT-PCR)检测3种蛋白mRNA的表达水平.结果 IL-8可改变内皮细胞紧密连接蛋白...     

Synovial sarcoma. Inter-relationship of the biphasic and monophasic subtypes.

In order to assess minimum diagnostic criteria for synovial sarcoma, particularly the monophasic variety, and the inter-relationship between the monophasic and biphasic types, 32 examples were studied histologically, immunohistochemically (26 cases), and ultrastructurally (13 cases). Of the six biphasic synovial sarcomas examined by electron microscopy, the spindle cell component did not show evidence of epithelial differentiation or resemble the epithelial phase, but did appear fibroblastic; no tumor cells transitional between the spindle and epithelial component were evident. In contrast, all of the seven monophasic lesions had ultrastructural growth patterns and some cellular features approximating the epithelial cells of the biphasic variant. In 11 biphasic synovial sarcomas, epithelial membrane antigen was detected in the glandular epithelium of all cases and cytokeratins in eight cases; in no case were these antigens detected in the spindle cell regions of biphasic lesions. Of the 15 monophasic synovial sarcomas, two were positive for cytokeratins and four for epithelial membrane antigen. Thus, the detection of epithelial markers either immunohistochemically or by electron microscopy (or both) should be the minimal diagnostic criteria for monophasic synovial sarcomas. Based on these findings, it is suggested that monophasic synovial sarcomas do not represent the spindle cell or "stromal" phase of biphasic synovial sarcomas, but are a poorly differentiated variant of the latter. As others have suggested, these tumors are, in fact, carcinosarcomas and carcinomas of the soft tissues and the designation synovial sarcoma is inappropriate for this tumor class.     

Tight junctions in thyroid carcinogenesis: diverse expression of claudin-1, claudin-4, claudin-7 and occludin in thyroid neoplasms.

Claudins and occludin are integral constituents of tight junctions and are deregulated in a variety of malignancies. Their role in thyroid carcinogenesis has not yet been elucidated. This study investigates the expression of occludin and claudin-1, -4 and -7 in thyroid neoplasms. Ninety-one thyroid neoplasms (15 follicular adenomas, 15 follicular carcinomas, 26 papillary carcinomas, 16 papillary microcarcinomas, 8 medullary carcinomas, 3 poorly differentiated carcinomas, 8 undifferentiated carcinomas) were immunostained with antibodies against occludin and claudin-1, -4 and -7. Occludin was mainly expressed in the form of intracytoplasmic vesicles, whereas all claudins tested exhibited membranous immunostaining. Thirteen out of 15 follicular adenomas, 10/15 follicular carcinomas, 24/26 papillary carcinomas, 15/16 papillary microcarcinomas, 1/8 medullary carcinomas, 2/3 poorly differentiated carcinomas and 2/8 undifferentiated carcinomas exhibited claudin-1 expression, whereas claudin-4 was expressed in 13/15, 12/15, 23/26, 13/16, 7/8, 2/3 and 2/8 of the tumors, respectively, and claudin-7 expression was found in 67, 33, 73, 69, 25, 0 and 13% of the cases, respectively. Occludin was expressed in 100% follicular adenomas, 80% follicular carcinomas, 96% papillary carcinomas, 50% papillary microcarcinomas, 50% medullary carcinomas, 33% poorly differentiated carcinomas and 88% undifferentiated carcinomas. Occludin expression was reduced in papillary microcarcinomas, medullary carcinomas and poorly differentiated carcinomas. All claudins exhibited reduced expression in undifferentiated carcinomas. Claudin-1 was additionally reduced in medullary carcinomas and claudin-7 in follicular, medullary and poorly differentiated carcinomas. A correlation between loss of claudin-1 expression and worse disease-free survival was noted on univariate analysis. Dedifferentiation of the thyroid carcinomas is accompanied by reduction in claudin-1, -4 and -7 expression. A differential expression of tight junction proteins in the different histologic types of thyroid gland is noted. Additionally, claudin-1 expression may be an important prognostic indicator of recurrence in thyroid carcinomas.     

Calponin and h-caldesmon expression in synovial sarcoma; the use of calponin in diagnosis

AIMS: To assess the frequency of expression and potential diagnostic utility of calponin and caldesmon in synovial sarcomas. METHODS AND RESULTS: Immunohistochemistry with antibodies to calponin and h-caldesmon was undertaken in paraffin sections from 50 synovial sarcomas (19 biphasic and 31 monophasic, of which one biphasic and eight monophasic tumours had poorly differentiated areas), 10 each of malignant peripheral nerve sheath tumour (MPNST), solitary fibrous tumour (SFT), dermatofibrosarcoma (DFSP), Ewing's sarcoma (ES/PNET), and neuroblastoma, eight alveolar rhabdomyosarcomas, five adult fibrosarcomas and five carcinosarcomas. Nine of 19 biphasic synovial sarcomas were positive for calponin in spindle and glandular areas, nine in spindle cells only, and one in glands only. All monophasic synovial sarcomas and poorly differentiated areas expressed calponin; in monophasic tumours this was focally (29% of cases), moderately (39%), or diffusely (32%) positive and the poorly differentiated areas were usually moderately or diffusely positive. Four synovial sarcomas showed focal reactivity for h-caldesmon. Calponin was found in 4/10 MPNST, 7/10 SFT, 4/10 DFSP, 3/5 fibrosarcomas and the spindle component of the carcinosarcomas. H-caldesmon was weakly positive in 1/10 MPNST, 4/10 SFT, 0/10 DFSP, 0/5 fibrosarcomas and 1/5 carcinosarcomas (glands only). Both markers were negative in the other small round cell tumours. CONCLUSIONS: Calponin can be used as an additional marker for synovial sarcoma. Its absence argues against the diagnosis. The presence of calponin might be useful in distinguishing poorly differentiated synovial sarcoma from other small round cell tumours. H-caldesmon is not helpful in diagnosis of synovial sarcoma.     

Amyloid beta-peptide1-42 alters tight junction protein distribution and expression in brain microvessel endothelial cells

Alzheimer's disease is characterised by neuronal loss, numerous intraneuronal deposits of neurofibrillary tangles, senile plaques, and cerebrovascular amyloid deposits. The major component of senile plaques and cerebrovascular deposits is the 39-43 amino acid beta-amyloid peptide (Abeta). The effects of Abeta on cerebral endothelium and thus the blood-brain barrier remain unclear. Utilising endothelial cells isolated from rat cerebral cortex microvessels, we have examined effects of Abeta peptides on tight junction protein behaviour. The transmembrane tight junction proteins occludin, claudin-1 and claudin-5, as well as the cytoplasmic accessory proteins ZO-1 and ZO-2 displayed a continuous distribution at cell boundaries. Endothelial cells exposed to Abeta1-42 (20 microM) for 3 days showed a disrupted plasma membrane pattern of claudin-5 and ZO-2 with relocation to the cytoplasm. These effects were not seen with Abeta25-35 or Abeta1-40[Gln22] (Dutch type). Abeta1-42 treatment altered also protein expression: occludin was lower at 1st day, claudin-1 increased at all times, and ZO-2 increased after 1 day and then decreased. These data suggest that Abeta1-42 effects on tight junction protein complexes may alter blood-brain barrier integrity and contribute to the neuropathological sequelae of Alzheimer's disease.     

Patterns of keratin polypeptides in 110 biphasic, monophasic, and poorly differentiated synovial sarcomas

Synovial sarcoma is a mesenchymal neoplasm of unknown histogenesis that shows various degrees of epithelial differentiation. It is known to contain simple epithelial keratins, and the possibility of complex epithelial keratin expression has been suggested. In this study, we immunohistochemically examined 110 well-documented synovial sarcomas including 44 biphasic, 48 monophasic, and 18 poorly differentiated (undifferentiated, highly mitotically active) tumors for 11 different keratin (K) polypeptides of the Moll catalogue. The epithelia of biphasic synovial sarcomas showed consistent, extensive reactivity for K7, K8, K14, K18, and K19. Other keratins seen in the epithelia of biphasic tumors included K17 (variable, in 77%), K13 (25%), K16 (23%), and K6 (24%) in the minority of biphasic tumors, predominantly in stratified-appearing epithelia. K10 was detected only focally in one case that showed keratinizing squamous differentiation. Focal expression of K20 was seen in 27% of cases. Monophasic synovial sarcomas had a more limited keratin repertory. Simple epithelial keratin positivity was detected, usually focally for K7 (79%), K19 (60%), K8 (45%), and K18 (46%). Two cases showed more extensive keratin positivity in the spindle cells. The monophasic tumors showed limited positivity for complex epithelial keratins: K14 (28%) and K17 (10%). K20 was detected focally in 6% of the monophasic tumors; other keratins were not detected. The poorly differentiated synovial sarcomas showed limited simple epithelial keratin reactivity, usually limited to scattered cells: K19 (61%), K7 (50%), K18 (47%), K8 (33%), but five cases showed more extensive positivity. Complex epithelial keratins were scant: K14 in one case and K17 in two cases. The immunoreactivity of capillary endothelia seen for K7 and K18 (but not for K8 and K19 with the antibodies used) is a potential diagnostic pitfall, and may cause overdiagnosis of synovial sarcoma if not properly recognized. In summary, we show complex patterns of keratins in synovial sarcoma, especially in the biphasic tumors. Such patterns establish a baseline in differential diagnostic considerations, and give an insight into the complex epithelial differentiation of this enigmatic mesenchymal tumor. Received: 17 February 2000 / Accepted: 3 April 2000     

帕金森病模型大鼠十二指肠黏膜紧密连接蛋白表达下调

目的研究紧密连接蛋白在6-羟多巴(6-OHDA)制备的帕金森病(PD)大鼠模型十二指肠黏膜的表达变化。方法用6-OHDA损毁双侧中枢黑质多巴胺能神经元建立大鼠模型。用免疫荧光组织化学和蛋白免疫印迹检测紧密连接蛋白claudin-1、occludin和ZO-1肠黏膜的定位和表达。结果紧密连接蛋白claudin-1、occludin和ZO-1在PD大鼠模型十二指肠黏膜上均有表达,但仅ZO-1(P0.001)和occludin(P0.01)表达明显下调,而claudin-1无显著变化。结论 PD大鼠模型十二指肠黏膜紧密连接蛋白ZO-1、occludin的表达显著下调,可能与帕金森病十二指肠溃疡的发生发展相关。     

Synovial sarcoma of the upper digestive tract: a report of two cases with demonstration of the X;18 translocation by fluorescence in situ hybridization.

Two cases of synovial sarcoma that arose in the upper digestive tract are reported. One case was a polypoid mass that arose at the gastroesophageal junction; the other was a large intramural mass that arose in the wall of the stomach. Both cases had a classic biphasic pattern. In the stomach tumor, the biphasic morphology was focal and there was an abrupt transition to poorly differentiated synovial sarcoma. The tumors had immunohistochemical features that were consistent with synovial sarcoma. Ultrastructural evaluation of the gastroesophageal tumor supported the diagnosis. The diagnostic X;18 translocation was demonstrated by fluorescence in situ hybridization on sections from paraffin-embedded tissue in 86% and 50% of interphase nuclei from the gastroesophageal and gastric tumor, respectively. The translocation was present in equal frequency in the epithelial and spindle cells in the biphasic areas and the poorly differentiated areas of the gastric tumor, indicating that the development of the more aggressive subclone was probably due to genetic mutations not encompassing the SYT-SSX gene fusion product. We are aware of only five reported cases of synovial sarcoma arising in the digestive tract, all in the proximal esophagus. These cases are the first reported arising in the gastroesophageal junction and stomach and the only cases of synovial sarcoma of the digestive tract in which the diagnostic translocation was demonstrated. Sarcomatoid carcinoma (carcinosarcoma) and gastrointestinal stromal tumor are the main differential diagnoses for synovial sarcoma in this site. Synovial sarcoma of the digestive tract may be underdiagnosed, and its recognition may have important clinical implications. Fluorescence in situ hybridization is helpful in making this distinction.