Distribution of cell surface glycoconjugates during secondary neurulation in the chick embryo

Lectin histochemistry was used to examine the expression of cell surface glycoconjugates during secondary neurulation in chick embryos. Fourteen lectins were applied to serial sections of the caudal region of embryos at the various stages of tail bud development. The lectins Bandeiraea simplicifolia, Dolichos biflorus agglutinin, Phaseolus vulgaris leukoagglutinin, soybean agglutinin, Sophora japonica agglutinin, Ulex europaeus agglutinin and succinylated wheat germ agglutinin (sWGA) showed very light or no binding to the developing medullary cord of the tail bud. With the other lectins, staining occurred throughout the early tail bud and solid medullary cord. During cavitation, however, differential expression of cell surface glycoconjugates by different cell populations was observed. The lectins concanavalin A, Lens culinaris agglutinin, Pisum sativum agglutinin, Phaseolus vulgaris erythroagglutinin, Ricinus communis agglutinin and WGA showed basic similarities in the distribution of lectin binding. Of these, the binding pattern of WGA was the most striking. As the medullary cord cells were separating into central mesenchymal and peripheral epithelial populations, WGA bound preferentially to the epithelial cells and the notochord. The lectin PNA, however, became preferentially bound to the mesenchymal cells. Heavy staining by WGA (specific for N-acetylglucosamine and sialic acid) where sWGA staining (specific for N-acetylglucosamine only) was faint suggested that WGA binding was due to the presence of sialic acid containing glycoconjugates.     

D2-40 immunohistochemistry in the differential diagnosis of seminoma and embryonal carcinoma: a comparative immunohistochemical study with KIT (CD117) and CD30.

The distinction between seminoma and embryonal carcinoma based on morphology alone can sometimes be problematic, requiring the use of immunohistochemistry to facilitate diagnosis. D2-40 is a monoclonal antibody that reacts with an oncofetal antigen expressed by fetal germ cells and testicular germ cell tumors. The diagnostic value of D2-40 immunohistochemistry in distinguishing seminoma from embryonal carcinoma has not been determined. D2-40 immunoreactivity was evaluated in a series of testicular germ cell tumors and compared with that of KIT (CD117) and CD30, to assess the relative utility of this marker in discriminating between seminoma and embryonal carcinoma. Forty testicular germ cell neoplasms were examined, which included 19 seminomas, three embryonal carcinomas, three teratomas, one yolk sac tumor, and 14 mixed germ cell tumors. The 14 cases of mixed germ cell tumors contained components of seminoma (n=7), embryonal carcinoma (n=11), teratoma (n=10), yolk sac tumor (n=2), and choriocarcinoma (n=1). All cases of pure seminoma and the seminomatous components of mixed germ cell tumors exhibited positive immunoreactivity for D2-40. Focal positivity for D2-40 was also observed in 29% of the embryonal carcinoma samples. D2-40 immunoreactivity in seminomas was characterized by diffuse membrane staining, whereas for embryonal carcinomas, staining was focal and distributed along the apical surfaces of the neoplastic cells. Immunohistochemical staining for KIT was observed in 92% of the seminoma samples and in none of the embryonal carcinomas. Conversely, CD30 expression was identified in 93% of the embryonal carcinoma samples and in none of the seminomas. Other germ cell components showed no immunoreactivity for D2-40, KIT, or CD30. KIT and CD30 are effective immunohistochemical markers in separating seminoma from embryonal carcinoma. Although a highly sensitive marker for seminomas, D2-40 positivity was also observed in a subset of embryonal carcinomas, thus limiting the utility of this antibody for discriminating between these two malignancies.     

外阴营养不良、外阴鳞状上皮细胞癌凝集素标记免疫组化分析

目的：探讨外阴营养不良增生型,硬化苔藓型表皮细胞和外阴鳞状上皮细胞癌等细胞膜结构与凝集素受体结合表达特征及它们三者之间的关系。方法：对慢性外阴营养不良表皮增生型,苔藓硬化型及外阴鳞状上皮细胞癌3种状态下的细胞进行了8种凝集素免疫组织化学标记,分析,比较。结果：外阴营养不良增生型表皮各层细胞细胞膜与刀豆凝集素（ConA),扁豆凝集素（LCA）,花生凝集素（PNA）,蓖麻凝集素（RCA－1）和大豆凝集素（SBA）结合强度弱,其表皮细胞细胞核膜与ConA,PNA和麦胚凝集素（WGA）有中等强度结合,外阴营养不良硬化苔藓型表皮各层细胞细胞膜与ConA,RCA-1 SBA和WGA结合强度弱,而与LCA不结合,其12例中的8例与荆豆凝集素（UEA－1）发生较弱的结合,外阴鳞状上皮细胞癌癌细胞与8种凝集素均结合,其12例中的7例其癌细胞膜与双花扁豆凝集素（DBA）弱结合,癌细胞膜与UEA－1强结合,但癌巢中央细胞团不与UEA－1结合,癌细胞细胞核膜与ConA,LCA,RCA-1呈强结合。结论：凝集素标记免疫组织化学染色可作为诊断,鉴别诊断外阴营养不良增生,硬化苔藓型和外阴鳞状上皮细胞癌新的指标,具有恶变潜能的增生型外阴营养不良与外阴鳞状上皮细胞癌,两者细胞核膜凝集素标记有相似之处,提示在外阴组织的癌变过程中,细胞核膜凝集素标记的异常表达是一个早期标识,糖基和糖蛋白代谢的异常可能是细胞发生癌变最早期的表现之一。     

Immunohistochemical staining of germ cell tumors and intratubular malignant germ cells of the testis using antibody to placental alkaline phosphatase and a monoclonal anti-seminoma antibody.

Antibody to placental alkaline phosphatase (PLAP) has been previously used in immunoperoxidase (IMP) staining studies of germ cell tumors and intratubular malignant germ cells (ITMGC) of the testis, the latter believed to be the precursor of these tumors. In this study, we compared staining by IMP using monoclonal antibody (mAb) and polyclonal antibody to PLAP with that seen using a mAb, M2A, which was previously shown to react with testicular seminomas and ITMGC. Antibody to PLAP and M2A reacted with different cellular components, as assessed by IMP staining of placenta and prepubertal testis and by Western blotting of seminoma lysates. Antibody to PLAP stained pure seminomas (seven of seven), pure embryonal carcinomas (four of four), and the seminoma (three of three) and embryonal carcinoma (six of six) components of mixed testicular germ cell tumors. M2A stained pure seminomas (26 of 26) and the seminoma component (three of three) of the mixed tumors, but failed to stain pure embryonal carcinomas (zero of four) or the embryonal carcinoma component (zero of five) of the mixed tumors. Both antibody to PLAP and M2A stained ITMGC of the testis. Since M2A stained seminomas and ITMGC but not embryonal carcinomas, seminomas would appear to be more closely related to ITMGC than embryonal carcinomas. This result has led us to speculate on the histogenesis of testicular germ cell tumors.     

Placental alkaline phosphatase immunohistochemistry of intratubular malignant germ cells and associated testicular germ cell tumors

Two hundred three testicular germ cell tumors were studied immunohistochemically for the presence of placental alkaline phosphatase (PLAP). Special emphasis was placed on the pattern and incidence of positive staining of intratubular malignant germ cells (ITMGCs) adjacent to tumors. 99% of cases with adjacent ITMGCs showed a positive staining reaction in some or all IT-MGCs present. Other germ cell elements showed at least a focal positive staining reaction in the following proportions: seminomas, 96%; embryonal carcinomas, 96%; yolk sac tumors, 25%; mature teratomas, 5%; immature teratomas, 4%; choriocarcinomas, 45%; and syncytiotrophoblasts, 43%. The staining pattern for seminomas tended to be diffuse, whereas for embryonal carcinomas the staining pattern was more focal. Yolk sac tumors stained inconsistently for PLAP and a positive reaction was limited to a small percentage of cells. Syncytiotrophoblasts, singly or in choriocarcinomas, also showed variable positivity. These results corroborate the fact that PLAP is a sensitive marker for ITMGC, seminoma, and embryonal carcinoma.     

Lectin Binding Patterns in Normal Liver, Chronic Active Hepatitis, Liver Cirrhosis, and Hepatocellular Carcinoma An Immunohistochemical and Immunoelectron Microscopic Study

We studied the binding patterns of 14 lectins in human normal and cirrhotic liver (LC) tissues and hepatocellular carcinomas (HCC) using the ABC method. Lectins were divided into 4 groups according to their binding patterns in normal tissues: (A) PHA, MPA, LcH, RCA I, and WGA, which bound to hepatocytes and all three types of sinusoidal cells; (B) BPA, GS-I, PNA, and SBA, which bound to Kupffer cells and endothelia of interlobular arteries and veins and bile duct epithelia in the portal tract, but not to hepatocytes; (C) UEA-I, which bound only to endothelia of interlobular arteries and veins and bile duct epithelia in the portal tract; (D) LBA, Lotus, LPA, and SJA, which showed no binding. Thus group B lectins may be useful markers of Kupffer cells. Only electron microscopic examination revealed the precise binding sites of lectins in sinusoidal cells and hepatocytes. Hepatocyte cell surface polarities demonstrated by lectin binding in LC and HCC were different from those in the normal liver. The binding pattern of PHA to LC hepatocytes changed from a membranous to both a membranous and a cytoplasmic pattern, and that of LcH to HCC cells changed to dot-like staining in the cytoplasm. These changes of polarities in LC and HCC might be caused by changes in the distribution of lectin binding carbohydrates or by the altered glycosylation of glycoconjugates. Acta Pathol Jpn 42: 566–572, 1992.     

Evaluation of cyclin expression in testicular germ cell tumors: cyclin E correlates with tumor type, advanced clinical stage, and pulmonary metastasis.

The measurement of proliferative index has yielded promising yet conflicting results in the evaluation of testicular tumors. We have examined the role of Ki-67, along with the cyclins A and E in testicular tumorigenesis. We compared the immunoreactivity of 20 pure seminomas with 20 mixed germ cell tumors composed predominantly of embryonal carcinoma with a variety of proliferation markers, including Ki-67, cyclin A, and cyclin E. All 40 tumors stained for Ki-67, and 19 of 20 (95%) seminomas and 18 of 20 (90%) embryonal carcinomas stained positively for cyclin A. Cyclin E stained 14 of 19 (74%) of the embryonal carcinomas and only 4 of 20 (20%) of the seminomas (Fisher's exact two-tailed test, P = .0012). There was a trend toward larger tumor size for cyclin E-positive seminomas (median, 5.92 cm versus 3.96 cm; P = .08), although the same correlation was not significant in embryonal carcinomas. For both seminomas and embryonal carcinomas, staining with cyclin E did not correlate with the presence of lymphovascular invasion or capsular invasion. However, patients who had cyclin E-positive tumors presented with higher clinical stage (P = .0015). In addition, pulmonary spread in embryonal carcinomas (four patients) and seminomas (one patient) occurred only in patients whose tumors were cyclin E positive (P = .014). Although Ki-67 and cyclin A offer little prognostic information in testicular germ cell tumors, cyclin E immunoreactivity correlates with tumor type and is strongly predictive of distant tumor spread.     

Lectin cytochemistry of carbohydrates on cell membranes of rat cerebellum

Summary Wheat germ agglutinin (WGA),Ricinis communis agglutinin (RCA) andLens culinaris (LC) lectins have been used to characterize carbohydrates of neuronal membrane systems in rat and chick cerebellum. WGA and RCA both label Golgi membrane cisternae on the side of the membrane facing the cisternal space but they do not label other elements of the granular and agranular endoplasmic reticulum (ER). WGA labels the plasma membrane generally and WGA binding sites are concentrated within the synaptic cleft and at specialized sites along the axolemma at the node of Ranvier. RCA labelling of plasma membranes is sparse. Neuraminidase treatment of tissue slices prior to lectin cytochemistry does not alter the Golgi membrane distribution of WGA and RCA binding sites and other ER membranes remain unlabelled. The plasma membrane staining with WGA is decreased and that with RCA is increased after neuraminidase pretreatment. The pattern of lectin labelling with LC resembles that seen with concanavalin A (con A) in that all elements of the ER within cell bodies label on the side of the membrane facing the cisternal space. A major difference when compared to con A labelling is that the hypolemmal cisternae lying adjacent to the plasma membrane of Purkinje cell axons and dendrites do not label with LC. Collectively, these results suggest that elements of the ER of Purkinje cells are heterogeneous with respect to their lectin binding properties in a manner which is consistent with the formation of more complex oligosaccharides on membranes of the cell periphery.     

Human chorionic gonadotrophin and alpha-fetoprotein in testicular germ cell tumours: a retrospective immunohistochemical study

A series of testicular germ cell tumours (46 seminomas and 27 non-seminomas) was studied immunohistochemically with regard to the presence of alpha FP and HCG. In three seminomas, HCG reactive syncitiotrophoblast-like giant cells (STLG) were found. Immunoreactive alpha FP did not occur in seminomas. In differentiated mature teratomas HCG or alpha FP could not be demonstrated. In embryonal carcinomas with or without teratoma (MTI/MTU/MTT) HCG immunoreactivity was found in 83%, usually localized in STLG. In 75% of these tumours alpha FP could be demonstrated. This protein was localized in foci of endodermal sinus or yolk sac differentiation, but also in single cells and cell clusters in areas of embryonal carcinoma. In some cases syncitial cells were present which contained both HCG and alpha FP. Immunostaining of tumour markers appeared not to provide important additional criteria for classification of these tumours in the currently available classifications. The significance of HCG containing STLG in seminomas deserves further investigation. Prospective studies of embryonal carcinoma with or without teratoma (MTI/MTU/MTT) will be necessary to evaluate the possible prognostic importance of the presence of alpha FP or HCG or both.     

Detection of high-mobility group proteins A1 and A2 represents a valid diagnostic marker in post-pubertal testicular germ cell tumours

The high-mobility group A (HMGA) non-histone chromosomal proteins HMGA1 and HMGA2 are architectural factors. They are abundantly expressed during embryogenesis and in most malignant neoplasias, whereas their expression is low or absent in normal adult tissues. Their over-expression is known to have a causal role in cellular neoplastic transformation. Previous studies from our group have shown that their expression is restricted to specific germinal cells. In this study we have evaluated, by immunohistochemistry, the expression of HMGA1 and HMGA2 in a series of post-pubertal testicular tumours of different histological types, including 30 seminomas, 15 teratomas, 15 embryonal carcinomas and 10 mixed germinal tumours with a prominent yolk sac tumour component. HMGA1 protein expression was detected in all seminomas and embryonal carcinomas analysed, but not in teratomas or yolk sac carcinomas. Conversely, HMGA2 was present only in embryonal carcinomas and yolk sac carcinomas, but not in seminomas or teratomas. The immunohistochemical data were further confirmed by Western blot and, at the mRNA level, by RT-PCR analyses. These findings indicate that HMGA1 and HMGA2 are differently expressed with respect to the state of differentiation of testicular germ cell tumours (TGCTs), with over-expression of both proteins in pluripotential embryonal carcinoma cells and loss of expression of HMGA1 in yolk sac tumours and of both proteins in the mature adult tissue of teratoma areas. Therefore, the different profiles of HMGA1 and HMGA2 protein expression could represent a valuable diagnostic tool in some cases in which the histological differential diagnosis is problematic.     

Use of lectin histochemistry in pancreatic cancer.

Lectin peroxidase histochemical analysis was carried out on pancreatic tissue from patients with pancreatic carcinoma and chronic pancreatitis and from subjects with normal pancreas to find a tumour specific pattern of lectin binding that would aid histological and cytological diagnosis. There were striking differences between the lectin binding characteristics of the different cell types in the normal pancreas. Acinar cells were uniformly positive for binding with wheat germ agglutinin and soy bean agglutinin while islet cells were usually negative for these lectins. Ulex europaeus I lectin however, was found not to be specific for endothelium, showing positivity also for acinar and ductal tissue. Griffonia simplicifolia II lectin was found to be highly specific for ductal epithelium, and because of this was tested in a hamster pancreatic cancer model where it was not specific for ductal epithelium, reflecting differing carbohydrate expression in the hamster pancreas. Pancreatic carcinomas and chronic pancreatitis bound all five lectins without any qualitative distinction from each other or from normal pancreatic tissue, but there was increased intensity of peanut agglutinin binding to secreted mucins in pancreatic carcinoma, which may be of potential use in radiolabelled lectin scanning.     

Lectin binding patterns in amphibian skin epithelium.

Seven lectins (PNA, DBA, WGA, UEA-I, RCA, SBA, Con A) were used to localize glycoconjugates in the skin of 10 species of Amphibia, 7 anurans (Bufo marinus, Bufo bufo, Rana ridibunda, Rana pipiens, Hyla arborea, Pelobates syriacus and Xenopus laevis) and 3 urodeles (Salamandra salamandra, Triturus vulgaris and Ambystoma mexicanum). It was found that every lectin has a specific binding pattern in the skin of each species. No common pattern could be established, either among frogs or toads, nor for a particular lectin. Each lectin bound specifically and selectively to a particular epithelial component, which differed from one species to the other. A number of lectins showed selective binding to mitochondria-rich cells, but, again, a pattern in positivity could not be found. It is concluded that lectin histochemistry does correlate with cellular function. Our data can be applied in studies of epithelium and skin development, and of changes that occur during adaptation to the environment by amphibian species.     

NUCLEAR LAMIN EXPRESSION IN NORMAL TESTIS AND TESTICULAR GERM CELL TUMOURS OF ADOLESCENTS AND ADULTS

Nuclear A- and B-type lamins are differentially expressed in tissues, depending on the degree of cellular differentiation and proliferative status. By studying lamin expression in testis parenchyma and testicular germ cell tumours, further insight may be gained into the degree of cellular differentiation in normal testis and into the whole spectrum of differentiation lineages found in testicular germ cell tumours. Frozen tissue sections of normal testis and the different types of testicular germ cell tumours were immunostained with monoclonal antibodies to distinct lamin subtypes. Lamin reactivity was evaluated in relation to the lineage and degree of cellular differentiation and the reactivity patterns were compared with each other and with those in normal testis. In normal testis, both A- and B-type lamins were expressed in Sertoli, Leydig, and peritubular cells, while in spermatogonia only B-type lamins were found and spermatocytes showed weak reactivity with the A-type lamin antibodies. Carcinoma in situ was most often positive for both of the B-type lamins and negative for the A-type lamins (lamins A and C). In testicular germ cell tumours, B-type lamins were always expressed, while A-type lamins were differentially expressed. Differentiated non-seminomas were positive for both of the A-type lamins, whereas embryonal carcinomas were positive for lamin C and negative for lamin A. Seminomas were negative for both of the A-type lamins, with the exception of seminomas containing a Ras mutation. Spermatogonia and seminoma cells, which follow a differentiation pathway along the spermatogenic lineage and show characteristics of germ cells, do not express A-type lamins. Non-seminomas, showing embryonal or extraembryonal differentiation, express A-type lamins to varying degrees, distinguishing embryonal carcinoma cells from other non-seminomatous components. This may aid in the evaluation of the percentage of embryonal carcinoma in non-seminomatous testicular germ cell tumours as a prognostic parameter. © 1997 John Wiley & Sons, Ltd.     

Laminin in testicular germ cell tumours. An immunohistochemical study

Thirty-five testicular germ cell tumours comprising 16 yolk sac tumours, 15 embryonal carcinomas and 13 seminomas were examined for the presence and distribution of laminin using an indirect immunoperoxidase technique. In addition, nine normal yolk sacs and 23 carcinomas of the lung were studied. All the yolk sac tumours were positively stained for laminin. Both extra- and intracellular staining were found. Hyaline, eosinophilic material present within the tumours was positively stained, although with varying intensity. In 12 out of 15 embryonal carcinomas, laminin was found as a membrane staining but cytoplasmic staining also occurred. In 10 out of 13 classical seminomas, a membrane staining of many tumour cells was found, while cytoplasmic staining occurred in only a few seminomas. In all but one of the yolk sacs, laminin was present in the membrane beneath both the mesoblastic outer cell layer and the visceral endoderm. Intracellular staining was seen in some of the cells in both cell layers. In nine out of 23 carcinomas of the lung, laminin occurred extra- as well as intracellularly. Thus, this study showed that in normal yolk sacs the presence of laminin was not found to be particularly associated with any of the cell layers. Likewise, demonstration of laminin within yolk sac tumours did not define different patterns or subtypes of the yolk sac tumour. In addition, demonstration of laminin was not found to be useful in differentiating either between yolk sac tumours and embryonal carcinomas or between seminomas and non-seminomatous germ cell tumours. The findings add, however, interesting knowledge to histogenesis and embryogenesis.     

Lectin binding pattern in human retinal pigment epithelium.

A comparative lectin histochemical study of human retinal pigment epithelium (RPE) was performed to investigate the lectin binding pattern of normal, reactive and proliferating RPE. Normal RPE with attached sensory retina was found to bind the lectins Con A, WGA, PNA and RCA I. Reactive and proliferating RPE in retinal detachment and in photocoagulation scars revealed the same lectin binding pattern although its cellular topography changed. RPE-macrophages showed an additional reaction with SBA. In periretinal membranes of human PVR the typical lectin binding pattern of Con A, WGA, PNA and RCA I was found in pigmented and in a subpopulation of non-pigmented cells, suggesting that these lectin-positive elements were of RPE-origin. Additionally, single pigmented cells positive for SBA were found indicating macrophage differentiation. Thus lectin histochemistry provides a tool for cytochemical identification of RPE and its morphologic variants by revealing a specific combination of sugar-binding sites.     

Light and electron microscopic investigation of the lectin-binding pattern in the oxyntic gland region of bovine abomasum.

For the first time the expression of glycoconjugate residues in the oxyntic gland region of bovine abomasum has been investigated by means of lectin histochemistry. For light microscopic investigations, a battery of ten lectins, Con A, PSA, UEA I, WGA, LEA, SNA, RCA120, MPA, DBA and SBA was used. For electron microscopic examinations, WGA and RCA120 were utilized. The staining pattern of the lectins in all exocrine cell types of the oxyntic gland region is described. Compared to the results of monogastric species our study reveals some similarities, but just as many differences in the composition of glycoconjugate residues in bovine exocrine cell types. Typical for surface mucous cells is the amount of L-fucose, N-acetyl glucosamine residues and Galbeta1, 4GlcNAc sequences in the secretory granules. SNA could serve as a marker for surface mucous cells, because this lectin exclusively stains the plasma membrane and the secretory granules of surface mucous cells and the extracellular mucus. L-fucose and N-acetyl glucosamine are typical for the secretory granules of mucous neck cells. In addition, the secretory granules show the highest amount of N-acetyl galactosamine residues of all exocrine cells, so that DBA and SBA are recommended as marker lectins for mucous neck cells. Most lectins strongly stain the intracellular membrane system of oxyntic cells. The cocktail of glycoconjugates in the vicinity of the HCI production site provide protection against chemical injury. In chief cells only the apical plasma membrane is more or less labeled with all lectins apart from SNA. Specific marker lectins for oxyntic cells or chief cells of the bovine have not been characterized.     

Histochemically demonstrable changes in cell surface carbohydrates of human germ cell tumors

Cell surface carbohydrate chains of human germ cell tumors were investigated histochemically using peanut agglutinin (PNA), Dolichos biflorus agglutinin (DBA), Ulex europaeus agglutinin I (UEA-1), and anti-I-(Ma) antibody. Of 16 cases of embryonal carcinoma in adult testis, peanut agglutinin, a lectin specific for terminal beta-galactosyl residues, bound to the surface of tumor cells in 11 cases, D. biflorus agglutinin, a lectin specific for terminal alpha-N-acetyl galactosamine residues, in 10, and U. europaeus agglutinin, a lectin specific for terminal alpha-L-fucosyl residues in 7. I-(Ma) antigen, a branched carbohydrate chain composed of three N-acetyl lactosamine units, was found in 12 cases as well. The cell surface of tubular or organoid architecture tended to be positive for some of these four reagents, whereas that of a papillary pattern tended to be negative. All yolk sac tumors (three in adult testis, 10 in infantile testis, and four in ovary) were positive for peanut and U. europaeus agglutinins and anti-I-(Ma) antibody but were negative for D. biflorus agglutinin. Seventeen of 18 seminomas (in adult testis) and one dysgerminoma (in ovary) did not react with these four reagents. Four choriocarcinomas (three in testis and one in ovary) were not positive for any of them either. In 10 cases of immature teratoma, the appearance of the binding sites to these lectins and antibody depended on the direction of differentiation and the degree of maturity. These findings suggest that carbohydrate chains of human germ cell tumors change greatly in the process of differentiation from embryonal carcinoma cells to somatic cell (teratoid) component or yolk sac carcinoma component, and that histochemical detection of the binding sites described may assist in the classification of germ cell tumors, in spite of the lack of precise knowledge about the biologic role of cell surface carbohydrates.     

Frequent Fas gene mutations in testicular germ cell tumors

The Fas (Apo-1/CD95)/Fas ligand (L) system is involved in cell death signaling, and has been suggested to be important for the regulation of germ cell apoptosis in the testis. Mutations of the Fas gene may result in accumulation of germ cells and thus might contribute to testicular carcinogenesis. The open reading frame of Fas cDNA was examined in 24 cases of testicular germ cell tumors (TGCTs), comprised of 19 pure histological type (15 seminomas, 3 embryonal carcinomas, 1 immature teratoma) and 5 mixed-type tumors. Mutations of the Fas gene were found in nine (37.5%) of these cases. Each lesion with a homogeneous histological picture was selectively microdissected using a laser capture microdissection method: samples consisted of 18 lesions from seminomas, 7 embryonal carcinomas, 4 immature teratomas, 2 choriocarcinomas, and 1 from a yolk sac tumor. Microdissected genomic DNA was examined to determine which mutations were derived from which kind of histological lesion. Eleven mutations were detected in 10 TGCT lesions from nine cases, but none were found in benign lesions. All were point mutations, and eight missense mutations occurred in exon 9 encoding the core protein of the death domain essential for apoptotic signal transduction. Three were silent mutations. Mutations were found in the seminoma (27.8%) and embryonal carcinoma lesions (62.5%), but none were found in the one yolk sac tumor, two choriocarcinomas, or four immature teratoma lesions. Each seminoma and embryonal carcinoma lesion found in the same case had a different type of Fas mutation from the others. Mouse T-cell lymphoma cells transfected with missense mutated genes were resistant to apoptosis induced by anti-Fas antibody, indicating these to be loss-of-function mutations. These findings suggested a role of Fas gene mutations in the pathogenesis of TGCTs.     

Monoclonal antibody 43-9F: an immunohistochemical marker of embryonal carcinoma of the testis.

Discrimination between different types of germ cell tumours may be difficult in routine histological preparations. Additionally, none of the established immunohistochemical markers is completely reliable in diagnosis of embryonal carcinoma. A pilot study indicated that monoclonal antibody 43-9F--a marker of carcinoma-in-situ germ cells--may also react with embryonal carcinoma of the testis. In order to elucidate the applicability of 43-9F in diagnosis of embryonal carcinoma, 42 consecutive testicular germ cell tumours were tested immunohistochemically. Among the 42 tumours, 23 were seminomas and 19 were non-seminomas with seminomatous components in seven of them. Embryonal carcinomas were found in 15 tumours, two being of pure type and the remaining 13 a part of mixed tumours. Additionally, the material included 11 teratomas, nine yolk sac tumours and one choriocarcinoma. Immunohistochemical stainings were performed with 43-9F and additionally with antibodies against placental-like alkaline phosphatase, cytokeratins, alpha-foetoprotein and human chorionic gonadotropin. Using 43-9F a strong colour reaction was found in 13 of the embryonal carcinomas, whereas the reaction was moderate in the remaining two cases. A weak positive reaction was found in six seminomas and the remaining 24 did not react at all. 43-9F exhibited a positive reaction in four of 11 teratomas. The reactivity was generally weak with some focal areas with strong staining. In five cases the yolk sac tumour elements did not stain with this monoclonal antibody. The reaction was weak in three cases and in only one case was the staining intensity scored as moderate. Finally, no reaction was found in the choriocarcinoma element. Compared to the other antibodies tested, including the antibody against cytokeratins, in embryonal carcinoma immunohistochemical staining with 43-9F was more specific, stronger and more constantly expressed. The monoclonal antibody 43-9F may be of value in histological diagnosis of germ cell tumours. Additionally, the study confirmed the pathogenetical link between pre-invasive carcinoma in situ and embryonal carcinoma.     

Ultrastructural characteristics of seminoma and embryonal cancer of the testis

Twenty-three germ cell tumors of the testis were investigated. Electron microscopic examinations of four seminomas and four embryonal carcinomas revealed cells of different degrees of differentiation. Tumor cells of seminoma acquired the ultrastructural features of the cells of developing seminal epithelium whereas in embryonal carcinoma there was a pronounced tendency of formation of mature tissues of teratoma. These data confirmed the cytogenetic resemblance of seminoma and embryonal carcinoma originating from the seminal epithelium which could undergo malignization at different stages of development. The latter predetermined diverse direction of differentiation in germ cell tumors of the testis.