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1.
Electrophoretic patterns of seminoma- and normal-testis-derived alkaline phosphatase isozymes, the placental alkaline phosphatase (PLAP)-like enzyme and the tissue-nonspecific (liver) alkaline phosphatase (LAP), were studied on starch gel and isoelectric focusing (IEF). Different migration patterns of the PLAP-like enzyme were observed with respect to both seminomas and normal testes on starch gel electrophoresis. On IEF, seminomas showed different staining patterns among different tumors; however, a common main activity was focused at pIs of 4.3-4.6, corresponding to pIs of PLAP. Normal testes showed two enzyme-staining regions, at pIs of 4.1 and 5.0-5.2, which were discriminated from pIs of PLAP and the PLAP-like enzyme in seminoma. The PLAP-like enzyme in seminoma was differentiated from PLAP by digestion with neuraminidase. Neuraminidase treatment simplified the distribution patterns of the PLAP-like enzyme in normal testis, but did not alter the pattern of microheterogeneity in seminoma. Two factors other than sialylation, namely structural modification of the carbohydrate moiety and variation of hydrophobicity, were shown to contribute to the microheterogeneity of the PLAP-like enzyme in seminoma. LAP in seminoma and in normal testis also showed marked electrophoretic heterogeneity and differences in pI distributions from LAP of liver. However, the migration patterns after desialylation were very similar to each other. The findings imply that electrophoretic heterogeneity demonstrated in LAP in seminoma and in normal testis is caused by a difference in sialic acid content in the molecule, and the heterogeneity of the PLAP-like enzyme in seminoma is considerable.  相似文献   

2.
The three human isozymes of alkaline phosphatases were quantitatively determined in normal testis and seminoma tissues. The highly selective assays were based on isozyme specific monoclonal antibodies. In the normal testis approximately 90% of the catalytic activity originates from the tissue unspecific alkaline phosphatase, and the remaining activity was due to trace expression of both intestinal (approximately 5%) and placental alkaline phosphatase (PLAP) or PLAP-like isozyme (approximately 5%). In homogenates of seminoma tissues, highly increased levels of all three isozymes were identified. Both the tissue unspecific alkaline phosphatase and PLAP-like enzymes displayed relative increases of 10- to 100-fold and intestinal alkaline phosphatase 2- to 10-fold compared with normal testis. This finding indicates that the entire genome coding for alkaline phosphatases may be activated in seminomas. The PLAP-like enzyme from seminoma cells comprises a heterogenous population of molecules demonstrating partial heat sensitivity and microheterogeneity upon starch gel electrophoresis in contrast to the pregnancy related PLAP. These findings have implications for the different PLAP assays used in the clinical monitoring of seminoma patients.  相似文献   

3.
Seminomas and control tissues were analyzed for several tumor markers. Very high levels of placental alkaline phosphatase (PLAP)-like enzyme levels were found in all 18 seminomas studied. The majority of the seminomas were of phenotype I, thus differing from palcental PLAP. The mean amount of enzyme protein as measured by monoclonal antibodies, was 100 times higher than in non-malignant tissues and 10 times lower than in placental tissue. The specific enzymatic activity in seminomas was about half of that observed in placenta. Similarly, the specific activity of PLAP-like enzymes in sera of patients with seminoma was only about half of that found in pregnancy sera. HCG was strongly elevated in 3 seminomas, but not obviously related to PLAP. Thirteen of the 17 pure seminomas had HCG over 100 IU/g, which was not seen in normal testes. Liver alkaline phosphatase (LAP) and intestinal alkaline phosphatase (IAP) were high in seminomatous tissues, the mean increases being 60-fold and 20-fold, respectively. The highest IAP levels were found in 2 yolk-sac tumors. Ferritin was moderately elevated in seminomas, but high in several control tissues. Carcinoembryonic antigen (CEA) was not elevated and alpha-fetoprotein (AFP) was not detected at all in pure seminomas. A decrease in carbohydrate antigen 50 (CA-50) content was noted in seminomas as compared to normal testes, yolk-sac tumors and choriocarcinomas. Defects in tumor-related enzymes may account for increase of PLAP and decrease of CA-50.  相似文献   

4.
Very high levels of placental-like alkaline phosphatases (PLAP-like enzymes) were observed in tissues from 13 typical seminomas. Four tumors with seminoma components contained these enzymes to varying degrees, and other testicular tumors had them in smaller or non-detectable amounts. Analysis using monoclonal antibodies produced against the common placental alkaline phosphatase (PLAP) phenotypes and enzyme inhibition studies with amino acids and peptides showed the PLAP-like enzymes present in seminoma to be similar to those PLAP-like enzymes which are expressed in lower amounts in two embryonal carcinomas and in trace amounts in normal testicular tissue. These similarities suggest that the increased expression of PLAP-like enzymes in seminomas results from enhanced eutopic expression of enzymes found in normal testis.  相似文献   

5.
The four known isozymes of the human alkaline phosphatase (ALP) were detected by isoelectric focusing in extracts of various types of germ cell tumors, three related cell lines, and their precancerous elements (atypical germ cells). In seminoma, placental alkaline phosphatase (PLAP) and germ cell alkaline phosphatase (PLAP-like) could be separated by isoelectric focusing following isolation by immunoaffinity. The occurrence of both isozymes in seminoma could explain partial heat sensitivity and variation in electrophoretic patterns of the seminoma isozyme frequently observed upon starch gels, in comparison to the normal placental phenotype. The four ALP isozymes are produced not only in germ cell tumors, but already in precancerous tissues. Quantitative analysis showed that the amount of the four isozymes varies in parallel in the tumors tested. Maximal expression was found in seminoma. The relation between ALP gene overexpression and gene amplification by polyploidy of chromosomes 1 and 2 in these lesions is discussed. On the other hand, the ectopic expression of intestinal alkaline phosphatase and PLAP associated with overexpression of PLAP-like in tumor cells as well as in their precancerous stage indicates gene activation by some unknown mechanisms, probably a regulatory process affecting the three tissue-specific ALP genes simultaneously.  相似文献   

6.
A monoclonal antibody (H17E2) recognising both placental alkaline phosphatase (PLAP) and testicular PLAP-like alkaline phosphatase was incorporated in a solid phase immunoassay. This was used to measure levels of PLAP in 257 sera from 148 patients with germ cell neoplasms of the testis. High levels of PLAP were found in all patients with active seminomas (mean 0.85 O.D.) compared to those in clinical remission (mean 0.20 O.D.) (P less than 0.0001). More importantly, changing levels of PLAP correlated with the course of disease in 79 samples from 33 patients with seminoma (P less than 0.0001). Elevated PLAP levels were also noted in patients in remission who were smokers (mean 0.32 O.D.) compared to non-smokers (mean 0.15 O.D.) (P less than 0.001). These data demonstrate that determination of PLAP levels using this sensitive immunoassay is an important new adjunct in the monitoring of the response to treatment in patients with seminoma.  相似文献   

7.
PLAP-like enzymes could be detected in serum of patients with primary testicular tumors, in particular seminomas. The use of a panel of monoclonal antibodies (MAbs) permitted typing into 6 different testicular serum phenotypes, of which one appeared similar to a placental type (II) and 2 have not been previously described. Most tumor sera belonged to type I, as described for seminoma tissues. With a more advanced tumor the mean serum PLAP-like levels increased. After operation, after radiotherapy or with no evidence of disease lower or non-detectable enzyme levels were found. In typing the tissue PLAP-like antigen in serum of the same patient or sera from recurrences of a seminoma in the same patient, the same phenotypes of PLAP-like antigen were usually but not always found. None of the 6 phenotypes appeared to confer a poorer prognosis. We conclude that the expression of PLAP-like antigen is eutopic and is enhanced by testicular malignancy, especially in seminoma cells.  相似文献   

8.
A431 human epidermoid carcinoma cells monophenotypically express the placental alkaline phosphatase (PLAP)-like enzyme shown by its catalytic and antigenic characteristics, properties which are shared by the Nagao isozyme. More specifically, it is L-leucine sensitive just as is the rare placental D-variant of PLAP and the testicular heat-stable enzyme. Collectively, these are all referred to as PLAP-like enzymes. The enzyme was localized to the surface of the plasma membrane since it was released in an active form by bromelain treatment of cells. The number of molecules per A431 cell was estimated by radioimmunoassay at 7.5 X 10(5), a value significantly higher than that observed for HeLa TCRC-1 cells (5 X 10(4) which express the S-variant of PLAP, also referred to as the Regan isozyme. The quantity of the enzyme was increased significantly (10-fold) by treating the cells with modulating agents including sodium butyrate, prednisolone, and hyperosmolar sodium chloride. The identification of a cell line such as A431 with enhanced expression in the amount of the PLAP-like enzyme and which can be further enhanced by modulating agents will facilitate studies of the differences and the similarities between this protein and other variants of PLAP. The A431 cell line now takes its place with other cell lines which are phenotypically restricted in their expression of alkaline phosphatase. Finally, the A431 cell line is also shown here to be a suitable model system for in vivo tumor studies such as immunolocalization.  相似文献   

9.
Indirect immunofluorescence and radioimmunoassay with specific rabbit antisera demonstrated the occurrence of alphafetoprotein (AFP), carcinoembryonic antigen (CEA) and placental alkaline phosphatase (PLAP) in primary testicular tumor cells. Embryonal carcinomas had AFP- and CEA-containing cells, sometimes PLAP. PLAP and sometimes CEA were found in seminoma cells. Sera from patients with advanced non-seminomatous tumors could contain any of these antigens or any combination of them. Sera from patients with seminomas had raised PLAP or CEA. PLAP appears to be a new marker for seminoma.  相似文献   

10.
Immunohistochemical analysis was done on 7 testicular tumors classified as spermatocytic seminoma (SS) and 25 classic seminomas. Except for a few scattered cells, the spermatocytic seminomas were negative for placental-like alkaline phosphatase (PLAP); the classic seminomas were all positive for this enzyme. The SS also were negative for alpha-fetoprotein (AFP), beta-human chorionic gonadotropin (hCG), and leukocyte common antigen (LCA). The ploidy of the seven tumors of SS was as follows: two, diploid; two, near-diploid; one, tetraploid; one, aneuploid; and one, uninterpretable. The essentially negative staining of SS for PLAP was strikingly different from the pattern in classic seminoma. Thus, staining for this enzyme is useful for making the differential diagnosis between classic seminoma and SS. To differentiate between malignant lymphoma and SS, staining for leukocyte common antigen is helpful.  相似文献   

11.
The clinical course of colorectal carcinoma may be monitored by tumor markers such as carcinoembryonic antigen (CEA), carcinoma antigen (CA) 19-9 and CA-50. Alkaline phosphatase isozymes were previously used to study the clinical course of testicular and gynecologic tumors. In this study we investigated 8 patients with advanced colorectal carcinoma. Their sera were analyzed for the tumor markers CEA, CA 19-9, CA-50 and three alkaline phosphatase isozymes: the nonspecific liver isozyme LAP, the intestinal isozyme IAP and the placental isozyme PLAP. Rising levels of CEA, CA 19-9 and CA-50 were seen as expected, and PLAP also showed rising levels during tumor progression. LAP remained elevated. This indicates an association between progression of colorectal carcinoma and a raised serum content of alkaline phosphatase isozymes.  相似文献   

12.
Using alkaline phosphatase isozyme-specific immunocatalytical assays, the content of isozymes was determined in normal mucosas and adenocarcinomas from human colon or rectum. Tumor levels of both the tissue (liver)-unspecific and the placental-like alkaline phosphatase (PLAP-like) were elevated compared to normal mucosas of the same patients. Such elevations have been reported previously, particularly in seminomas and ovarian tumors. In several tumors, moreover, the intestinal isozyme was expressed in lesser amounts than in the adjacent mucosa. The present results indicate that the activation of two of the phosphatase isozymes, including expression of the typical germ cell line phosphatase (the PLAP-like isozyme), may occur even in nongonadal tumors. This may reflect an induction pattern of phosphatase isozymes, with implications for malignant transformation also in other tumors.  相似文献   

13.
Placental and placental-like alkaline phosphatase (PLAP) levels in the culture media of 87 cell lines of neoplastic and ''normal'' origin were measured by a conventional immunosorbent enzymatic assay (IAEA) and by a new immunoradiometric assay (IRMA). The IRMA detected immunoreactive PLAP in 37 of 80 (46%) human epithelial and germ cell cultures, while the IAEA detected PLAP in only 25 (33%). Of the 52 non-germ cell tumour cultures, the IRMA detected expression in 24 (46%) and the IAEA in only 16 (31%). In 17 cases (21%) the IRMA recorded levels double that of the IAEA, while in five cultures (6%) the reverse was true. The IRMA was much more robust than the IAEA and had considerably lower inter- and intra-assay coefficients of variation (3.75-8.5% vs 5.2-46%). Detection of PLAP(-like) expression by IAEA is dependent on neoplastic expression of enzymatically functional molecules and quantification assumes constant enzyme kinetics. PLAP-like material has a higher catalytic rate constant than PLAP and thus will give higher values on a stoichiometric basis in an IAEA. The higher detection rate and levels of PLAP-like material in neoplastic cultures when measured by the IRMA clearly demonstrate ectopic expression of non-enzymatic PLAP and PLAP-like genes. The incidence of PLAP(-like) expression by non-germ cell and possible germ cell tumours has been underestimated and its utility as a tumour marker should be re-examined using assays which measure antigen mass rather than phosphatase activity.  相似文献   

14.
M E Lowe  A W Strauss 《Cancer research》1990,50(13):3956-3962
The alkaline phosphatase (AP) synthesized by human tumor cells closely resembles human placental AP (PLAP). Little is known about the molecular events that lead to the expression of a placenta-like AP in tumor cells. The complementary DNA encoding the AP expressed by a choriocarcinoma cell line, BeWo, was isolated and characterized. The complementary DNA is the product of the germ cell AP (Nagao isozyme) gene and not of the term PLAP gene. Like placental AP, the tumor AP can be released from the cell membrane by a phosphaditylinositol-specific phospholipase C and has a phosphaditylinositol-glycan (PI-glycan) moiety at the COOH terminus. Immunoprecipitation of phosphaditylinositol-specific phospholipase C-treated AP and analysis by polyacrylamide gel electrophoresis or isoelectric focusing demonstrates that at least 95% of the AP contains PI-glycan. Two-dimensional gel electrophoresis reveals two precursors of the mature AP. One of these does not bind an antibody against the Trypanosoma variable surface glycoprotein cross-reacting determinant and probably does not contain PI-glycan. This precursor had a shorter half-life than the more prominent PI-glycan-containing precursor in pulse-chase experiments, suggesting a precursor-product relationship between the two proteins. These data demonstrate that BeWo AP is the product of a gene normally expressed in testis, thymus, and germ cells, but not in placenta. Thus, the expression of BeWo AP results from the repression of the PLAP gene and derepression of the germ cell AP gene and, as such, the expression is ectopic. The BeWo AP (Nagao isozyme) is modified with PI-glycan that is added soon after translation, not cotranslationally.  相似文献   

15.
Serum samples from 62 patients with seminoma were assayed for placental alkaline phosphatase-like activity using the monoclonal antibody H17 E2, in order to evaluate its utility as a serum tumour marker. Fifteen of 16 patients (94%) with active seminoma had elevated serum PLAP levels. Sixteen of 46 (35%) of patients considered to be in remission had elevated PLAP levels (false positive rate 35%). Fifteen false positive results were considered attributable to concomitant smoking, and if these patients are excluded, only one false positive case was detected. In 7 out of 7 patients sequential PLAP assays reflected clinical response to treatment.  相似文献   

16.
The purpose of this study was a complementary method to the diagnosis and prognosis of complete hydatidiform mole (CHM) and differentiate it from the other cases of gestational trophoblatic diseases. This was done by examining the quality and quantity of the total and the placental alkaline phosphatase activity. The ALP in the tissues and sera from 12 patients were compared with 13 control normal non-pregnant and 30 control pregnant females. The enzyme activities were determined by biochemical and histochemical examination.The placental tissues were obtained from uterine curettage, or after delivery which then were frozen in a liquid nitrogen and processed for biochemical study. Cryosections were histochemically stained for ALP and PLAP by the azo coupling method. Isoenzyme specificity was evaluated by heating the tissue at 65 degrees C for 15 min while the including L-phenylalanine (50 mM), D-phenylalanine (50 mM) and L-homoarginine (50 mM) were used for chemical inhibition study. The activity of ALP and PLAP of patients were reduced in comparison with pregnant control group (P<0.05). There was no significant difference between the patients and non-pregnant control (P<0.05) group. The localization of enzyme activities in cryosections of all groups were in the basal, apical, and the cytoplasm of syncytiotrophoblast cells. The ALP in all the groups was thermostable (65 degrees C for 15 min) and was inhibited by L-phenylalanine, but no inhibition was seen with L homoarginine in patients group only. These findings suggest that the PLAP is a useful marker in the diagnosis and prognosis of hydatidiform mole.  相似文献   

17.
O Koide  S Iwai  K Baba  H Iri 《Cancer》1987,60(6):1325-1330
The identification of atypical testicular germ cells is often difficult by by routine histologic examination. By immunohistochemical detection of placental alkaline phosphatase (PLAP) and by periodic acid Schiff staining of glycogen, atypical germ cells were easily identified in testicular samples. Forty-one fetal and adult testes were used for a preliminary study, and 121 testes from infants and adults with either cryptorchidism or germ cell tumors were studied for the presence of atypical germ cells. Two types of clear germ cells were differentiated histochemically, and one with PLAP-positive cell surfaces and glycogen-rich cytoplasm was considered to be atypical. The alkaline phosphatase of atypical germ cells appeared to be similar to that found in a few germ cells of early fetal testes. The atypical germ cells seemed to be multi-potential malignant cells capable of developing not only into seminoma but also into other germ cell tumors. Only in yolk sac tumor of infants were the atypical germ cells absent from tumor-adjacent seminiferous tubules.  相似文献   

18.
A monoclonal antibody (H17E2) was used in a solid-phase localisation of enzyme activity (ILEA) assay to evaluate placental-like alkaline phosphatase (PLAP) as a serum marker of testicular germ cell tumours. Single or repeated assays were performed on 213 normal blood donor and a smaller number of term pregnancy and testicular cancer sera. The detection limit of PLAP by this system was 0.14 O.D. units equivalent to 0.04iul-1. Of 50 patients with established metastatic disease tested before treatment, 88% of 16 with seminoma, 54% of 13 with mixed seminoma and malignant teratoma and 33% of 21 with malignant teratoma had serum PLAP greater than 0.2 O.D. units. This compared to an incidence of 2% in non-smokers and of 29% in smokers who had been free of disease for more than 12 months. In 15 of 22 successfully treated patients, pre-treatment serum PLAP exceeded 0.2 O.D. units (mean 0.69 O.D.) and varying (53-97%) reductions in the initial levels occurred with treatment. These results with monoclonal antibody ILEA assay suggest that measurement of PLAP levels will be useful in the management of patients with germ cell tumours, particularly seminoma.  相似文献   

19.
In 11 adult testes studied, about 0.3 to 4.6% of the total alkaline phosphatase activity was heat stable and L-phenylalanine sensitive but L-homoarginine insensitive. The testicular heat-stable enzyme was more susceptible to inhibition by L-leucine and ethylenediaminetetraacetate than were the normal placental and intestinal enzymes. By antibody-directed enzyme inhibition test, the testicular heat-stable enzyme cross-reacted completely with normal placental enzyme but clearly distinguished itself from a heat-stable component of normal intestinal enzyme. Thus, placental alkaline phosphatase D-variant is synthesized in testis, indicating that the gene for elaborating this placental protein is probably already active in the testicular cells. The high incidence of this protein in cancers of testis and ovary is probably due to its increased production by gonadal genes present in the genome of these particular tumors.  相似文献   

20.
Non-specific testicular accumulation of radiolabeled intact anti-CEA monoclonal antibody (MAb), (A431/26, Behringwerke AG) was observed in 11 out of 12 patients with the testes and prostate included in the examination field at radioimmunoscintigraphy (RIS). Previous studies have shown that placental alkaline phosphatase (PLAP) serves as an Fc-receptor, mediating IgG transport through the placenta. A closely related protein, the germ cell alkaline phosphatase (GCAP), is expressed in the testes. The testicular uptake of IgG is observed only when intact but not fragmented MAbs are used, indicating involvement of Fc-receptors. MDCK cells (dog kidney cell line) transfected with the plasmid pSVT7 containing the GCAP gene were shown to acquire the capacity to both express membrane bound GCAP and to bind IgG on the cell surface. This might indicate that GCAP is responsible for the non-specific accumulation of intact MAb in the testes and prostate often observed when intact murine MAbs are used for radioimmunolocalization (RIL)  相似文献   

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