CD20在胸腺瘤中的表达及其临床病理意义

目的 探讨CD20在胸腺瘤中的表达及其临床和病理学意义.方法 对179例手术切除的胸腺肿瘤病例,按照2004年的WHO分类标准并结合临床病理资料,重新进行病理组织学分类,并选取其中资料完整的102例肿瘤组织,运用免疫组织化学EnVision法分别标记CD20、CKpan、末端脱氧核苷酸转移酶、CD3、CD5、CD43、CD99、S-100蛋白.其中重点观察肿瘤性上皮细胞及背景淋巴细胞的CD20表达特征,其他指标用以标记细胞.同时将所有病例按是否伴有重症肌无力分为两组,对结果进行统计学分析.结果 102例胸腺瘤中,A型7例、AB型32例、B1型17例、B2型15例、B3型17例,胸腺癌14例,CD20表达于肿瘤性上皮细胞,在A、AB、B1、B2、B3型胸腺瘤和胸腺癌中的阳性率分别为3/7、84.4%(27/32)、1/17、2/15、0/17、0/14.肿瘤的背景淋巴细胞的阳性率分别为3/7、18.8%(6/32)、14/17、11/15、11/17、6/14.伴重症肌无力组(40例)肿瘤内淋巴细胞的CD20阳性表达率为67.5%(22/40),不伴重症肌无力组(62例)阳性表达率为35.5%(22/62),CD20阳性的B淋巴细胞主要分布于伴重症肌无力组,两组间差异有统计学意义(P＜0.05).结论 A型、AB型和极少数的B1、B2型胸腺瘤的上皮细胞均具有表达CD20蛋白的特征,这类上皮细胞属于肿瘤性上皮;而B3型胸腺瘤及胸腺癌的肿瘤性上皮细胞缺乏表达CD20蛋白的特征.胸腺瘤伴重症肌无力者,肿瘤内B淋巴细胞增生,数量明显多于不伴重症肌无力者.AB型胸腺瘤并非A型与B型胸腺瘤的单纯混合,可能属于不同的细胞起源而表现出不同的组织形态和免疫表型.     

Impaired expression of MHC class II molecules in response to interferon-gamma (IFN-γ) on human thymoma neoplastic epithelial cells

A human thymoma is a neoplasm derived from the thymic epithelial cell, and is well known for its association with autoimmune diseases, especially myasthenia gravis. The neoplastic epithelial cells of thymoma clearly retain thymic epithelial functions, but the development of T cells in thymoma is somewhat impaired. In this study, we quantified by flow cytometry the in vitro expression of MHC molecules on neoplastic epithelial cells precultured with IFN-γ. While MHC class I expression was comparable with that on normal thymic epithelial cells, the level of MHC class II molecules on neoplastic epithelial cells was lower than in controls, and also varied greatly from case to case. Additionally, there was a significant positive correlation between the expression level of MHC class II and the proportion of mature CD3⁺ cells in the CD4⁺CD8^? subset. Thus, accumulation of CD3^?CD4⁺CD8^? cells in thymoma may result from impaired expression of the MHC class II molecules, suggesting that the function of the neoplastic epithelial cells might determine the maturation and the positively selected repertoire of T cells in thymomas.     

Evaluation of prognostic features in thymic epithelial tumors

Based on the original proposals of Müller-Hermelink [3,8] and the study of 95 tumor specimens from the files of our institute we have established a new concept for the classification of thymic epithelial tumors. Thymomas are related to the structural components of normal thymus and divided in medullary, mixed, predominantly cortical and cortical types. In addition a well-differentiated thymic carcinoma with partial loss of organotypic differentiation is characterized and distinguished from other carcinoma types with total lack of specific thymic features. Prognostic evaluation showed, that medullary and mixed thymomas are always benign tumors, whereas predominantly cortical thymomas, cortical thymomas and well-differentiated thymic carcinomas are low-grade malignant tumors with increasing invasiveness and even metastatic capacity. Moreover the proliferation rate of neoplastic epithelial cells in vitro, which was studied in 12 cases, correlated to the different tumor types and their growth behaviour in vivo.     

T-cell development in human thymoma.

Human thymoma is derived from thymic epithelial cells and often associated with a large number of cortical thymocytes. Since thymic epithelial cells play key roles in T-cell development in the normal thymus, we hypothesized that the neoplastic epithelial cells of thymoma may support T-cell differentiation. We attempted to reconstitute the T-cell development in vitro by using neoplastic epithelial cells isolated from thymoma. CD34, a stem cell marker, was expressed on a proportion of CD4-CD8- cells in thymoma. These CD34+CD4-CD8- cells also expressed both IL-7R alpha-chain and common gamma-chain. Purified CD4-CD8- cells from thymomas were cultured with the neoplastic epithelial cells, and their differentiation into CD4+CD8+ cells via CD4 single positive intermediates was observed within 9 days' co-culture in the presence of recombinant IL-7. The CD34+CD4-CD8- cells purified from a normal thymus also differentiated to CD4+CD8+ cells in an allogeneic co-culture with the neoplastic epithelial cells of thymoma. In addition, a pleural dissemination from thymoma contained a large amount of cortical thymocytes. These results suggest that the neoplastic epithelial cells retain the function of thymic epithelium and can support T-cell development in thymomas.     

Demonstration of phenotypic abnormalities of thymic epithelium in thymoma including two cases with abundant Langerhans cells.

A panel of monoclonal antibodies that phenotypically define stages of normal human thymic epithelial (TE) cell maturation was used to compare thymic epithelium of nine thymomas with hyperplastic thymic epithelium in myasthenia gravis (MG) and thymic epithelium of normal thymuses. It has been shown previously that normal thymic epithelial cells express antigens of early TE cell maturation (A2B5, TE-4) throughout thymic ontogeny and acquire antigens 12/1-2, TE8, and TE-15 at 14 to 16 weeks of fetal gestation. Hyperplastic MG thymic epithelial cells expressed TE antigens in phenotypic patterns similar to that seen in normal postnatal thymus, ie, TE in subcapsular cortex and medulla was TE4+, A2B5+, and 12/1 - 2+ and Hassall's bodies were reactive with antibodies TE8 and TE15. In contrast, thymic epithelium in primary mediastinal thymomas was TE4+, A2B5+, TE8-, and greater than 75% of thymoma epithelium was 12/1 - 2-, a thymic epithelial phenotype similar to that seen on normal fetal thymic epithelium at 14 to 16 weeks fetal gestation. In one subject with a mature epithelial histologic pattern, thymoma epithelium was found to be strongly TE8+, a phenotype suggestive of a later stage of TE maturation. Lymphocytes in five of seven thymomas with immature thymic epithelial cells predominantly expressed immature thymocyte phenotype while two thymomas with immature epithelial phenotype showed a predominance of Langerhans cells and surrounding lymphocytes expressing a mature phenotype. Lymphocytes in the thymoma with differentiated epithelial cells expressed a mature thymocyte phenotype. Thus, in thymomas of varying histologic types, phenotypic abnormalities of thymic epithelium are present; these phenotypic abnormalities may reflect abnormal thymic epithelial maturation.     

Histopathologic changes of thymoma preoperatively treated with corticosteroids

Preoperative treatment of thymoma in advanced stages with corticosteroids may reduce the size of the tumor, but no precise histologic evaluation has been performed. We examined the histopathologic features of pretreatment biopsy and posttreatment surgical specimens of eleven cases of thymoma with such treatment to see the changes of the histologic subtypes based on Muller-Hermelink classification. All specimens were also assessed immunohistochemically for MIB-1 labeling and apoptotic cells to verify the effectiveness of this pretreatment. Seven tumors clinically diminished in size after the treatment with corticosteroids. Fungal infection occurred in three cases postoperatively. The histology of mixed thymomas (two cases) was converted to that of medullary thymoma. Predominantly cortical thymomas (four cases) and cortical thymomas (three cases) changed to show similar histologic features; both became epithelial-rich thymoma with large polygonal tumor cells having indistinct cell borders. In contrast, two well-differentiated thymic carcinomas showed at surgery more prominent squamoid appearance with distinct cell borders. The apoptotic indices of epithelial cells were increased (P = 0.001), and the MIB-1 indices tended to be decreased with corticosteroid treatment. These results suggest that there may be a histogenetic relationship between medullary and mixed thymomas and also between predominantly cortical and cortical thymomas. Corticosteroids may cause degenerative changes in the epithelial cells and lymphocytes and, in thymomas in advanced stages, corticosteroid pretreatment may be warranted, although attention should be paid to infection after surgery.     

Neuroendocrine differentiation in thymic epithelial tumors with special reference to thymic carcinoma and atypical thymoma

To determine the neuroendocrine (NE) features of thymic epithelial tumor, immunohistochemistry and electron microscopy studies were performed on eight NE tumors (thymic carcinoids) and 26 non-NE tumors (nine thymic carcinomas, five atypical thymomas, and 12 thymomas other than lymphocytic thymoma). Immunohistochemical studies were performed with antibodies against general markers for NE cells (synaptophysin, a subunit of a guanine nucleotidebinding protein, Go, and small-cell lung carcinoma cluster 1 antigen), and a broad panel of antibodies for hormonal substances. Thymic carcinoid showed synchronous diffuse immunoreactivity for the three NE markers and contained cells that were positive for a variety of hormonal products: human chorionic gonadotropin (hCG) a-subunit (eight of eight), hCG β-subunit (three of eight), adrenocorticotropic hormone (ACTH) (three of eight), calcitonin (two of eight), calcitonin gene-related peptide (two of eight), and serotonin (one of eight). Conversely, although positivity for NE markers was neither synchronous nor diffuse in non-NE tumors, seven of nine thymic carcinomas, three of five atypical thymomas (focal or dispersed distribution), and none of the five thymomas were positive for at least two of these NE markers. A small number of neoplastic cells were positive for hCGa-subunit or ACTH in three thymic carcinomas and one atypical thymoma. Ultrastructurally, dense core granules (DCG) were much more frequent in thymic carcinoid, but several DCG-like granules were identified in 12 of 13 non-NE tumors with or without immunoexpression of NE markers. The presence of focal or dispersed NE cells in thymic carcinoma and atypical thymoma may reflect multidirectional differentiation within the tumor, which, like cytological atypia, epithelial CD5 expression, and lack of immature T cell infiltration, may be another feature of this group at thymic tumors.     

Unusual thymoma subtypes

Thymomas are neoplastic proliferations of the epithelial cells of the thymus which are variably admixed with a non-neoplastic lymphoid component. Five common subtypes of thymoma are recognized in the current WHO classification. In addition to these five subtypes, three distinct unusual thymomas are currently listed as specific entities in the WHO scheme. These three subtypes, micronodular thymoma with lymphoid stroma, metaplastic thymoma and lipofibroadenoma, and their differential diagnostic consideration form the subject of this review. In addition, unusual variants of common thymomas are discussed.     

Impaired expression of MHC class II molecules in response to interferon-gamma (IFN-gamma) on human thymoma neoplastic epithelial cells.

A human thymoma is a neoplasm derived from the thymic epithelial cell, and is well known for its association with autoimmune diseases, especially myasthenia gravis. The neoplastic epithelial cells of thymoma clearly retain thymic epithelial functions, but the development of T cells in thymoma is somewhat impaired. In this study, we quantified by flow cytometry the in vitro expression of MHC molecules on neoplastic epithelial cells precultured with IFN-gamma. While MHC class I expression was comparable with that on normal thymic epithelial cells, the level of MHC class II molecules on neoplastic epithelial cells was lower than in controls, and also varied greatly from case to case. Additionally, there was a significant positive correlation between the expression level of MHC class II and the proportion of mature CD3+ cells in the CD4+CD8- subset. Thus, accumulation of CD3-CD4+CD8- cells in thymoma may result from impaired expression of the MHC class II molecules, suggesting that the function of the neoplastic epithelial cells might determine the maturation and the positively selected repertoire of T cells in thymomas.     

Tumors of the neck showing thymic or related branchial pouch differentiation: a unifying concept

A number of rare tumors occurring in the soft tissues of the neck and the thyroid gland, reported in the literature under a variety of designations, show complete to partial histologic resemblance to the fetal, mature, or involuted thymus and mediastinal thymomas. This family of tumors spans a range of histologic appearances and behaviors from completely benign lesions to metastasizing malignant tumors. After reviewing the previously reported and new cases, we have been able to delineate four reasonably well-defined clinicopathologic entities within this spectrum. On the benign end is "ectopic hamartomatous thymoma," which occurs in the soft tissues of the lower neck. It is characterized by spindle epithelial cells, solid or cystic epithelial islands, and adipose cells which intermingle haphazardly to impart a hamartomatous quality. In the middle of the spectrum are the ectopic cervical thymomas which are usually benign, but can sometimes be locally invasive and can exceptionally metastasize. They are histologically identical to mediastinal thymomas, and residual ectopic thymus is not uncommonly identifiable in the periphery of the tumor. On the malignant end are tumors we have designated as "spindle epithelial tumor with thymus-like differentiation" (SETTLE) and "carcinoma showing thymus-like differentiation" (CASTLE). Tumors of the SETTLE type occur in the thyroid gland of young patients, and are highly cellular tumors comprised of compact bundles of long spindle epithelial cells which merge with tubulopapillary structures and/or mucinous glands. Tumors of the CASTLE type are histologically similar to thymic carcinoma of the lymphoepithelioma or squamous cell variety. We postulate that this family of tumors arises either from ectopic thymus or remnants of branchial pouches which retain the potential to differentiate along the thymic line.     

Thymoma with loss of keratin expression (and giant cells): a potential diagnostic pitfall

Due to its profound therapeutic consequences, the distinction between thymoma and T-lymphoblastic lymphoma in needle biopsies is one of the most challenging in mediastinal pathology. One essential diagnostic criterion favouring thymoma is the demonstration of increased numbers of keratin-positive epithelial cells by immunohistochemistry. Loss of keratin expression in neoplastic epithelial cells could lead to detrimental misdiagnoses. We here describe a series of 14 thymic epithelial tumours (11 type B2 and B3 thymomas, 3 thymic carcinomas) with loss of expression of one or more keratins. Cases were analysed for expression of various keratins and desmosomal proteins by immunohistochemistry and immunofluorescence and compared with 45 unselected type B thymomas and 24 thymic carcinomas arranged in a multitissue histological array. All 14 cases showed highly reduced expression of at least one keratin, three cases were completely negative for all keratins studied. Of the 14 cases, 13 showed strong nuclear expression of p63. Expression of desmosomal proteins was preserved, suggesting intact cell contact structures. Loss of expression of broad-spectrum-keratins and K19 was observed in 3 and 5 % of unselected thymomas and in 30 and 60 % of thymic carcinomas. A proportion of keratin-depleted thymomas contained giant cells, reminiscent of thymic nurse cells. Loss of keratin expression in type B2 and B3 thymomas is an important diagnostic pitfall in the differential diagnosis with T-lymphoblastic lymphoma and can be expected in 5 % of cases. A panel of epithelial markers including p63 is warranted to ensure correct diagnosis of keratin-negative mediastinal tumours.     

Molecular landscape of thymic epithelial tumors

Thymic epithelial tumors (TETs) are extremely rare and represent the most frequent tumors of the anterior mediastinum originating from epithelial cells in the thymus. Thymic epithelial tumors include thymomas (TM), thymic carcinomas (TC) and thymic neuroendocrine neoplasms (TNEN). Thymomas are the most predominant and are associated with autoimmune diseases. The available data suggests that the different histological subtypes have specific molecular alterations. Thymic carcinoma shows recurrent gene mutations, but further investigations are needed to understand the role of those mutations in the pathogenetic of the TETs. Some of the new emerging identified molecular alterations have the potential to offer new targeted therapies opening new possibilities for the treatment of thymic epithelial tumors.     

Touch imprints in the intraoperative diagnosis of anterior mediastinal neoplasms

OBJECTIVE: To evaluate the accuracy of intraoperative diagnosis of mediastinal lesions using touch imprints and frozen sections. DESIGN: We studied touch imprints and frozen sections from 21 anterior mediastinal lesions retrospectively. The lesions included six cases of non-Hodgkin's lymphoma, eight thymomas, two thymic carcinomas, three Hodgkin's disease cases, and two seminomas. Slides were reviewed independently by each of the three authors, and diagnoses were recorded. RESULTS: Depending on the observer, the correct diagnosis was obtained on touch imprints alone in 76% to 81% of cases. On frozen sections alone, the correct diagnosis was made in 67% to 86% of cases. In 86% to 100% of cases, the correct diagnosis was made on either touch imprint or frozen section. CONCLUSIONS: As with frozen sections, the most common significant error in interpreting cytology preparations was in distinguishing thymic epithelial tumors (thymoma and thymic carcinoma) from lymphoma. On a modified Wright-Giemsa-stained imprint, epithelial cells in a thymoma may be inconspicuous. Clues to their presence include cells with a spindled nuclear shape or a small distinct nucleolus. The epithelial cells have scant cytoplasm with indistinct cell borders. Clumping of cells is often not prominent in lymphocytic thymomas, but may be present in epithelial or mixed lymphocytic and epithelial tumors. With practice, one can learn to recognize thymic epithelial cells on touch imprints. Familiarity with this simple inexpensive technique could improve the accuracy of intraoperative diagnosis of anterior mediastinal lesions.     

Neoplastic thymic epithelial cells of human thymoma support T cell development from CD4−CD8− cells to CD4+CD8+ cells in vitro

Human thymoma is a thymic epithelial cell tumour which often contains a large number of immature T cells and is frequently associated with autoimmune diseases. Since thymic epithelial cells play key roles in the development and selection of T cells in the normal thymus, we hypothesized that the neoplastic thymic epithelial cells of thymoma may support T cell differentiation in the tumour. We characterized CD4^?CD8^? cells in thymoma and applied an in vitro reconstitution culture system using the CD4^?CD8^? cells and the neoplastic epithelial cells isolated from thymoma. CD34, a stem cell marker, was expressed on 29.9 ± 12.2% of CD4^?CD8^? cells in thymoma. TCRγδ was expressed on 27.4 ± 15.1% of CD4^?CD8^? cells and CD19, a B cell marker, was expressed on 14.1 ± 23.1% of CD4^?CD8^? cells. CD4^?CD8^? cells expressed both IL-7R α-chain and common γ-chain. Purified CD4^?CD8^? cells from thymomas were cultured with the neoplastic epithelial cells, and their differentiation into CD4⁺CD8⁺ cells via CD4 single-positive intermediates was observed within 9 days' co-culture in the presence of recombinant IL-7. Furthermore, we examined the reconstitution culture using CD34⁺CD4^?CD8^? cells purified from normal infant thymus. The CD34⁺CD4^?CD8^? cells in normal thymus also differentiated to CD4⁺CD8⁺ cells in the allogeneic co-culture with the neoplastic epithelial cells of thymoma. These results indicate that the tumour cells of thymoma retain the function of thymic epithelial cells and can induce differentiation of T cells in thymoma.     

Immunohistological evidences of cortical and medullary differentiation in thymoma

Summary The phenotypical characteristics of human epithelial and lymphoid cells have been studied with immunohistochemical methods on frozen sections of 12 thymomas. On the basis of the cytohistological characteristics of thymoma epithelial cells (EC) the thymomas were divided in cortical, medullary and mixed types, according to recently developed light microscopical criteria. When tested with a series of monoclonal antibodies, thymoma EC were all stained by the antibody Ki-M3 (as in the thymus), but reacted with anti-HLA-DR, anti-HLA-A,B,C and with a new monoclonal antibody to cortical EC,21A6, to a lesser extent and with weaker, variable intensity in comparison with the normal thymus. Cortical type thymomas were most reactive and the medullary type almost negative. Thymomas, like normal thymus showed different immunoreactivity patterns with antibodies to prekeratins of different specificities. Cortical type thymomas and areas in mixed thymoma showed an EC staining with the antibody to non-squamous type keratin (35H11) whereas medullary type thymomas and areas showed staining with antibodies to squamous-type keratin (34E12-IV/82) in addition. Lymphoidcellswithcortical(OKT6+,Leu 1 weakly+,Leu2a+,Leu3a+) or mature medullary (OKT6-, Leu 1 strongly+, Leu 2a or Leu 3a+) phenotype were found to colonize tumours with diferent EC types. These immunohistochemical findings largely confirm our earlier cytological distinction of thymoma EC. In addition important differences have been observed in neoplastic cortical EC concerning the HLA-DR and 21A6 immunoreactivity that may be intimately related to the neoplastic process and paraneoplastic immune phenomena.This work has been supported by the Deutsche Forschungsgemeinschaft, SFB 111, project CN5     

CD5 expression in thymic carcinoma.

To determine the differences between the cellular characteristics of thymic carcinoma and thymoma, immunohistochemical analysis with lymphocyte markers (CD1a, 3, 4, 5, 8, 10, 20, 21, 25, 30, 57, and 72) was performed on 23 thymic epithelial tumors other than lymphocytic thymoma: overt thymic carcinoma (OC, n = 7), atypical thymoma (n = 5), and typical thymoma (epithelial or mixed thymoma, n = 11). Among the surface antigens examined, CD5, a type of receptor molecule that signals cell growth in T cells, was expressed in neoplastic epithelial cells of the thymus, in OC (seven of seven) and atypical thymoma (two of five), but not in typical thymoma. Double labeling immunofluorescence demonstrated expression of CD5 in cytokeratin-positive cells. The CD5 molecule extracted from an OC tumor showed the same molecular size as that in the spleen, but CD72, a ligand of CD5 on the surface of B cells, was not found in the epithelial cells of OC or atypical thymoma. Expression of CD5 was not observed in carcinomas of other organs, such as lung (n = 15), breast (n = 4), esophagus (n = 6), stomach (n = 6), colon (n = 9), and uterine cervix (n = 3). CD5 is closely related to morphological changes in thymic epithelial tumors and may play a role in the evolution of OC through receptor-ligand interaction.     

The use of antikeratin antiserum as a diagnostic tool: Thymoma versus lymphoma

Indirect immunofluorescence staining in two thymomas, one case of thymic hyperplasia, 10 malignant lymphomas and three seminomas was done with an antibody prepared against keratins from human epidermis. Staining was observed only in the epithelial cells of the thymomas and thymic hyperplasia and correlated well with electron microscopic studies. Immunofluorescence staining of thymic tumors with antikeratin antibody provides a simple, specific, and sensitive method for distinguishing thymoma from lymphoma and seminoma. The method may also prove to be useful in other instances in the distinction between epithelial and nonepithelial tumors.     

Comparative ultrastructural characteristics of epithelial cells of the parenchyma of the thymus gland and thymoma

Comparative ultrastructural investigation of the parenchyma was performed for human thymus (7 specimens) and thymoma (34 specimens). For thymic parenchyma eight types of epithelial cells were identified. These may be represented by three different histogenetic lines: 1) spindle-shaped appendiceal cortical cells (type 1-4) 2) medullary cells showing signs of squamous-epithelium differentiation (type 5-6) 3) medullary cells with signs of glandular differentiation (type 7-8). Thymomas demonstrated cells ultrastructurally similar to thymic parenchyma and exhibiting features of either cortical or medullary epithelial cells. These findings served the basis for dividing thymomas into 3 major groups: 1) with cortical cell differentiation 2) with cortical and medullary cell differentiation 3) with medullary cell differentiation.     

The Diagnosis of Thymoma: A Review

Thymoma is the most common tumor of the anterior-superior mediastinum, especially in middle-aged or older adults. Microscopically, thymomas can be differentiated from other tumors with which they can be confused by the finding of a mixed population of cells, including neoplastic thymic epithelial cells with numerous processes surrounding activated-appearing lymphocytes. Thymomas can be classified as benign or malignant, and the majority of those that are malignant appear cytologically benign and are locally invasive. Cytologically benign thymomas have been classified as being lymphocyte rich, epithelial cell rich, or spindle cell type. This classification has not been found to be prognostically useful. A new method classifies these tumors as being cortical, medullary, or mixed. This new classification appears to have prognostic significance. Malignant thymomas that are cytologically malignant are uncommon. Such tumors usually are squamous cell carcinomas. Other types include sarcomatoid carcinoma, clear cell carcinoma, basaloid carcinoma, and mucoepidermoid carcinoma. Because other tumors can occur in the anterior mediastinum, electron microscopy and/or immunocytochemistry is helpful in making the distinction. The differential features by ultrastructural and immunocytochemical analysis are reviewed.