Biomedical subjects

T F Orntoft

Publications and source records attributed to T F Orntoft.

At least 55 records · Page 3Linked to original sources

PubMed1996

Bladder squamous cell carcinomas express psoriasin and externalize it to the urine.

PURPOSE: To report a single biomarker, psoriasin (Mr 11.0 kd, pI 6.2), a calcium binding protein which is expressed largely by stratified squamous epithelia and is externalized to the urine of bladder squamous cell carcinoma (SCC) bearing patients. MATERIALS AND METHODS: Protein expression profiles of SCCs obtained immediately after surgery were analyzed by two-dimensional gel electrophoresis and Coomassie blue staining. Protein identity was determined by microsequencing and immunoblotting. Protein expression in cryosections was studied by immunofluorescence. RESULTS: Four patients with SCC were identified from 100 samples of patients with suspected transitional cell carcinoma (TCC). The protein profiles of the 4 SCCs (56-1, grade III, T4; 181-1, grade I, T3; 219-1, grade III, T3 and 239-1, grade not determined, T2-4) resembled that of keratinocytes, suggesting that these cells express an early developmental pattern of gene expression. Besides expressing markers characteristic of keratinizing stratified squamous epithelia, the SCCs exhibited psoriasin, a protein externalized to the medium by keratinocytes. Immunohistochemistry of 3 of the SCCs with psoriasin antibodies showed that the positive cells were confined chiefly to the "squamous pearls." The presence of psoriasin in the urine of the 4 SCC patients was demonstrated by two-dimensional gel immunoblotting. Similar analysis of 43 urines from patients with bladder tumors other than SCC revealed 7 positives, some of which may reflect squamous differentiation. Analysis of the urine of 13 control individuals (12 males matched by age and a 42-year-old female) revealed 2 positives. Immunoblotting of the SCC patients' serum proteins with psoriasin antibodies failed to reveal the protein. CONCLUSION: The results point towards psoriasin, alone or as part of a biomarker profile, as a potential marker for the noninvasive follow-up of patients with SCC.

Adult↗

PubMed1996

Towards a comprehensive database of proteins from the urine of patients with bladder cancer.

PURPOSE: To establish a comprehensive two-dimensional database of proteins from the urine of patients with bladder cancer. MATERIALS AND METHODS: Urines dialized against distilled H2O were freeze-dried and subjected to isoelectrofocusing two-dimensional gel electrophoresis. Coomassie brilliant blue stained dry gels were scanned and analyzed with the PDQUEST software. Proteins were identified by one or more of the following techniques: microsequencing (44 proteins), mass spectrometry, comigration and immunoblotting. RESULTS: The urine protein database, which includes all the polypeptides detected in the urines of 50 patients, lists 339 proteins (including variants) of which 124 have been identified. Of these, psoriasin, the psoriasis-associated fatty acid binding protein 5, the gelsolin fragments and prostaglandin D2 synthetase have not been previously described. CONCLUSIONS: The database provides a solid basis for further studies aiming at identifying tumor markers in the urine that may serve as prognostic factors in bladder cancer.

Amino Acid Sequence↗

PubMed1995

Loss of ABH antigen expression in bladder cancer is not caused by loss of heterozygosity of the ABO locus.

The significance of loss of heterozygosity (LOH) of chromosome 9 as an early event in human bladder tumors has been demonstrated by several studies. We have studied LOH of the ABO gene locus at 9q34.2 by means of polymerase chain reaction (PCR) genotyping of tumor preparations and leukocytes. Twenty-two tumors, all of which were immunohistochemically negative for ABH antigens, were examined. Eleven tumors were from blood-group O individuals, and were examined by denaturing gradient gel electrophoresis (DGGE). They showed normal band patterns with no sign of single base mutations or LOH. All II A and B tumors were sorted on a fluorescence-activated cell sorter (FACS) according to DNA content, thereby making it possible to study differences in genotype between subpopulations of cells in the same tumor. In 2 genotypically AO cases, we found 2 aneuploid subpopulations. In both cases, the most abnormal, with the highest DNA content, showed complete loss of the O allele, leaving the A allele intact. As all tumors were ABH antigen-negative, this study demonstrates that LOH of the ABO locus on chromosome 9q34 is not the cause of loss of blood-group ABH expression in human bladder cancer.

ABO Blood-Group System↗

PubMed1995

Circulating blood group related carbohydrate antigens as tumour markers.

The various blood group related carbohydrate structures which are in clinical use as circulating tumour makers are reviewed. Their location on carbohydrate chains and their structural characteristics are shown, and their clinical performance in various malignant diseases is reviewed. The available data on their sensitivity, specificity and predictive value are shown; and carcinomas of the pancreas, ventricle, colon-rectum and ovary are identified as diseases in which these markers can be of good benefit for follow-up. Future research should be devoted to studies of the function of these structures, and to studies of their gene-transcription.

Antigens, Tumor-Associated, Carbohydrate↗

PubMed1995

Chronic treatment with epidermal growth factor causes esophageal epithelial hyperplasia in pigs and rats.

Epidermal growth factor (EGF) is an important factor for maintaining the esophageal functional integrity. Goettingen minipigs were treated with either placebo or subcutaneous EGF (30 micrograms/kg/day) for four weeks. Wistar rats were treated with either placebo or subcutaneous EGF (150 micrograms/kg/day) for four weeks. At sacrifice, esophageal samples were obtained for histology, immunochemistry, and lectin characterization. In pigs, the thickness of the esophageal epithelium was almost doubled in the EGF-treated animals. Characterization with lectins revealed a normal pattern of differentiation. Subcutaneously administered EGF was visualized on cells located basally in the esophageal epithelium. In rats, EGF-treatment increased the esophageal volume of the epithelium, the lamina propria of the mucosa, and the submucosa. In conclusion, systemic EGF challenge induces growth of the esophageal epithelium with an unaltered pattern of differentiation. This supports previous studies demonstrating a beneficial effects of systemic EGF-treatment on sclerotherapy-induced esophageal damage.

Animals↗

PubMed1995

Chronic systemic treatment with epidermal growth factor in pigs causes pronounced urothelial growth with accumulation of glycoconjugates.

Epidermal growth factor (EGF) is present in large amounts in the urine, but the effects of systemically administered EGF on the urinary tract have not been described previously. In the present paper, we describe a potent growth induction of EGF on the urinary tract. Goettingen minipigs were treated with solvent (n = 5), EGF 30 micrograms/kg/day (n = 6) for 4 weeks, or EGF 30 micrograms/kg/day for 5 weeks followed by 3 weeks of recovery (n = 5). The ureters and bladders were examined by routine histology and electron microscopy and were immunostained for proliferating cell nuclear antigen. Four weeks of EGF treatment increased the median cross sectional area of the ureter fourfold with growth of all wall layers. The urothelium was widened from 5 cell layers in the controls to 10 in the EGF-treated animals. Proliferating cell nuclear antigen immunostaining revealed an increased mitotic activity in the basal zone of the urothelium. In the luminal zone, glycoconjugates accumulated in goblet cells, in cells with intracytoplasmic lumina, and beneath the luminal cell membrane in the umbrella cells. Our studies present a new experimental approach to growth induction of the urinary tract. The findings implicate the EGF system in regulating urothelial growth and glycoconjugate biosynthesis.

Animals↗

PubMed1995

A comparative study of peanut agglutinin and amaranthin binding to human urinary bladder tumor glycoproteins.

The reactivity of two T-antigen specific lectins, Amaranthin (ACA) and Peanut Agglutinin (PNA), was compared by immunohistochemical staining of serial sections of human bladder tumors and by Western Blot analysis of glycoproteins extracted from human bladder tumors prior to and after sialic acid removal. In low grade tumors ACA reacted prior to neuraminidase treatment. PNA on the other hand only showed positive reaction in these tumors after neuraminidase treatment. The two lectins showed identical staining patterns after neuraminidase treatment in tumors. In Western blottings both lectins detected a 28 kD glycoprotein. PNA also reacted with several other bands after neuraminidase treatment. The reactivity of ACA was not enhanced after neuraminidase treatment. In monosaccharide inhibition tests ACA- and PNA-binding to different T-antigens was most efficiently inhibited by GalNAc and Gal respectively. PNA binding was also inhibited by Glc, GlcNAc and GalNAc at higher concentrations, while the binding of ACA was only scarcely affected by Glc, GlcNAc and Gal.

Antigens, Tumor-Associated, Carbohydrate↗

PubMed1994

Blood group ABO-related glycosylation of urothelial cell lines: immunocytological, enzymatic, and genetic characterization.

Three immortalized, human urothelial cell lines were characterized with respect to their ABO-related carbohydrate phenotypes using a panel of monoclonal antibodies directed to a series of carbohydrate epitopes (Lac, sialylated Lac, Le(a), sialylated Le(a), Le(x), sialylated Le(x), H types I and II, Ley, Leb, A monofucosylated types I and II, ALey, Aleb, and A type III). The glycosyltransferases forming some of these epitopes (beta 1-3/4 galactosyltransferase, alpha 1-2 fucosyltransferase, alpha 1-3 galactosyltransferase, and alpha 1-3-N-acetyl-galactosaminyltransferase) were determined by enzyme assays. The ABO gene complex was analyzed by Southern blotting, Northern blotting, and polymerase chain reaction across the O deletion and across base differences between the A and B alleles. The immunocytochemical stainings showed marked differences between the three cell lines; the high grade (tumorigenic, metastatic) cell line showed difucosylated types I and II structures, and the low grade (nontumorigenic, nonmetastatic) cell lines showed monofucosylated types I and II structures. Polymerase chain reaction genotyping of the cell lines indicated that one was OO, one was AA, and one was A plus a mutated allele. Northern blotting showed RNA encoding the A transferase. However, even though both of the A cell lines seemed to have an intact gene, which could produce A transferase and transcribed RNA, none of them showed any activity of the A gene encoded enzyme or any A-structures at the cell surface. In contrast, the three other examined glycosyltransferases were active. The three urothelial cell lines reflect in vivo findings in humans. They represent a competent system for in vitro studies of the different carbohydrate transferase genes responsible for the carbohydrate structures expressed on the cell surface in bladder tumors.

ABO Blood-Group System↗

PubMed1994

Hereditary spherocytosis: diagnostic and anaemia-associated aberrations of ghost proteins.

Various disorders of the red cell skeleton and membrane have been described in hereditary spherocytosis. To elucidate which aberrations could be used for identification of HS patients in a Danish population, we examined ghosts from 17 HS patients and 20 normals by use of SDS-gel scanning, native spectrin extraction, and limited tryptic digestion. Compared to normals, HS patients had significantly lowered alpha-spectrin (p < 0.004), protein 4.2 (p < 0.025), and actin (p < 0.05), and significantly increased anion-transporter (p < 3 x 10(-6)) and glyceraldehyde-3-phosphate dehydrogenase (G3PD, p < 0.04). Sixteen out of 17 HS patients could be identified by aberrations of the anion-transporter or protein 4.2 outside a 95% confidence interval for normals. Extraction of native spectrin and limited tryptic digest showed no difference between normals and HS patients. RBC separated into young and old fractions were used to examine the occurrence of protein aberrations associated with RBC age. Young RBC contained more G3PD (35%) and less protein 4.1 (6.5%) and actin (8.7%) than old. In male HS patients an increased G3PD content showed a linear correlation (p < 0.001) with a low concentration of blood haemoglobin. We conclude that aberrations of G3PD, and possibly protein 4.1, and actin, are associated with anaemia in HS. Increased anion-transporter or lowered protein 4.2 may be useful for diagnosis of HS, and were inherited in five out of six families where two generations were available.

Actins↗

PubMed1994

ELISA-type titertray assay of IgM anti-GM1 autoantibodies.

We report an ELISA-type titertray assay for autoantibodies against the ganglioside GM1. Trays were coated with ganglioside GM1 and reacted with patients' sera; bound IgM was detected with rabbit antibody to human IgM. High-titer serum from a patient was used as calibrator, another patient's serum as the positive control, and the GM1-specific cholera toxin as the control for GM1 coating. Regression curves of serum titers obtained from different patients were linear and parallel. Intra- and interassay CVs were 4.0-7.8% and 5.5-16%, respectively. We detected antibodies at a titer of 1:250 in normal subjects. Analytical specificity of the calibrator serum against GM1 was demonstrated by immune thin-layer chromatography. Anti-GM1 antibodies were increased in patients with chronic inflammatory demyelinating polyradiculoneuropathy (P < 0.002) or multiple sclerosis (P < 0.01). In Guillain-Barré syndrome, preliminary longitudinal studies showed a decrease in anti-GM1 titer that was related to clinical recovery.

Autoantibodies↗

PubMed1992

Human urinary bladder carcinoma glycoconjugates expressing T-(Gal beta(1-3)GalNAc alpha 1-O-R) and T-like antigens: a comparative study using peanut agglutinin and poly- and monoclonal antibodies.

T- and T-like antigens on glycoproteins and glycolipids were examined in extracts of human urinary bladder tumors and normal tissue by Western blot analysis and reagent binding to thin layer chromatograms. Three different anti-T-reagents were used: peanut (Arachis hypogaea) lectin (PNA) and mono- and polyclonal antibodies specific for T-antigen (Gal beta(1-3)GalNAc alpha 1-O-R). Immunodetection with the T-specific reagents in nitrocellulose replicas of bladder tumor glycoproteins, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, demonstrated tumor-specific T-antigen-bearing glycoproteins compared to normal urothelial glycoproteins. In addition, a remarkable difference in binding was found between the immunological reagents and PNA lectin. PNA showed major reactivity to a 28-kD glycoprotein extracted from tumors. Monoclonal anti-T-antibody (clone HH8) showed major reactivity with an M(r) 34,000 glycoprotein, and polyclonal anti-T-antibody showed major reactivity with an M(r) 36,000 glycoprotein. PNA agarose column affinity-purified tumor glycoproteins did not bind the antibodies. Glycoproteins, M(r) 28,000 and 34,000, were shown to be O-linked by stepwise deglycosylation. In solid phase monosaccharide inhibition tests, galactose followed by N-acetyl-galactosamine were the most potent monosaccharides inhibiting binding to immobilized bladder tumor glycoproteins. None of the anti-T-reagents reacted with glycolipids extracted from tumor tissue. It is concluded that PNA lectin, in addition to the T-disaccharide, reacts with other protein-anchored carbohydrate structures in carcinomas.

Antibodies, Monoclonal↗

PubMed1992

Expression of histo-blood-group-A/B-gene-defined glycosyltransferases in normal and malignant epithelia: correlation with A/B-carbohydrate expression.

Malignant transformation of oral and bladder epithelia is often associated with loss of histo-blood-group-A- and -B-carbohydrate antigens, whereas these antigens, which are absent in normal adult distal colon (but present in fetal colon) reappear in malignant distal colon. In order to gain insight into the genetic basis of the biosynthetic regulation for these changes, we have correlated the expression of the A- and B-carbohydrate antigens with that of the A/B-gene-defined glycosyltransferases in colon, bladder and oral carcinomas by immunohistology. A newly developed anti-A/B-transferase monoclonal antibody (MAb) was used to demonstrate the in situ localization of transferase expression at the individual cell level with correlation to carbohydrate antigen expression, and gave the essential information that the transferase is derived from the ABO gene complex. The reappearance of A- and B-carbohydrate antigens in carcinomas of the distal colon was found to be unrelated to the expression of the A/B-transferase proteins, which were expressed throughout normal adult colon in accordance with previous enzymatic studies. In contrast, the loss of A- and B-carbohydrate antigens in malignant bladder and oral epithelia was accompanied by concordant loss of enzymes.

ABO Blood-Group System↗

PubMed1992

Specificity and immunobiological properties of monoclonal antibody IMH2, established after immunization with Le(b)/Le(a) glycosphingolipid, a novel extended type 1 chain antigen.

Extended lacto-series type 1 chain antigens lacking type 2 chain core have recently been shown to comprise a new type of tumor-associated carbohydrate antigen. Examples are Le(a)/Le(a) (IV3Gal beta 1----3[Fuc alpha 1----4]Glc-NAcLc4Cer) and Le(b)/Le(a) (IV3Fuc alpha 1----2Gal beta 1----3[Fuc alpha 1----4]Glc-NAcLc4Cer) (M. R. Stroud, et al., J. Biol. Chem., 266: 8439-8446, 1991; Eur. J. Biochem., 203: 577-586, 1992). We have now established an IgG3 mouse monoclonal antibody (IMH2) after immunization of mice with Le(b)/Le(a) antigen; however, monoclonal antibody (MAb) IMH2 reacted not only with the immunogen used but also with Le(y)/Le(x) and to a lesser degree with short-chain Le(y) or Le(b) with hexasaccharide ceramide (i.e., IV2FucIII3FucnLc4Cer or IV2FucIII4FucLc4Cer). It showed a high incidence of staining and strong reactivity with carcinomas of colon, rectum, liver, pancreas, and endometrium, but no reactivity with normal colonic mucosa at various loci, and minimal reactivity with normal liver, pancreas, or uterine endometrium. On the other hand, it reacted with normal gastric mucosa, cecal mucosa, urothelium, adrenal glands, and thymus. Its expression in colorectal tumors and normal cecal tissue was independent of secretor status, whereas that in normal urothelium was dependent on secretor status. MAb IMH2 displayed strong lymphocyte-activated or complement-dependent killing of human colonic cancer Colo205 cells in vitro, and inhibition of Colo205 growth in vivo; this inhibition was comparable to that by MAb NCC-ST-421, which is directed to Le(a)/Le(a) epitope (M. Watanabe, et al., Cancer Res., 51:2199, 1991). These results indicate that a new extended type 1 chain structure, Le(b)/Le(a), is a useful tumor marker associated with carcinomas of colon, rectum, pancreas, liver, and endometrium and that MAb IMH2 has potential diagnostic or therapeutic applicability for these carcinomas.

Animals↗

PubMed1992

Nuclear volume and expression of T-antigen, sialosyl-Tn-antigen, and Tn-antigen in carcinoma of the human bladder. Relation to tumor recurrence and progression.

The T-antigen system and the mean nuclear volume have been proposed as risk variables in bladder tumors. This study includes 34 patients with initially noninvasive (Ta) transitional cell carcinomas who experienced different courses of disease. Tissue specimens of primary tumors were analyzed for the expression of T-antigen, Tn-antigen, and sialosyl-Tn-antigen using monoclonal antibodies (MoAb) and the lectin peanut agglutinin (PNA) in an indirect immunoperoxidase method. In addition, the mean nuclear volume was estimated by morphometry. Tissue from 7 of 13 patients (54%) who had invasive disease during a follow-up period of 5 years expressed T-antigen, as defined by MoAb HH8 in the primary tumor, whereas tissue of only 3 of 21 patients who did not have invasive disease expressed the antigen (P less than 0.02). No association was found between tumor progression to invasion and the expression of Tn-antigen or sialosyl-Tn-antigen. Tn-antigen expression was partially lost in invasive tumors (P less than 0.03) when compared with the expression in primary noninvasive tumors. A high mean nuclear volume in tissue specimens of primary tumors correlated with a progression to invasive disease (P less than 0.01). A significantly (P less than 0.003) higher mean nuclear volume was found in tumor areas that were positive for PNA compared with areas that were negative for PNA in primary tumors. A significantly lower mean nuclear volume was found in Tn-antigen-positive invasive Grade 3 tumor areas than in Tn-antigen-negative areas (P less than 0.005). The combined use of T-antigen expression and mean nuclear volume is of potential clinical interest for determining patients who are at high risk of disease progression.

Aged↗

PubMed1992

Localization and identification of T-(Gal beta 1-3GalNAc alpha 1-O-R) and T-like antigens in experimental rat bladder cancer.

A mouse monoclonal antibody and a rabbit polyclonal antibody against the T-antigen (Gal beta 1-3GalNAc alpha 1-O-R) were used to study the distribution of T-antigens in an experimental rat bladder cancer model. Neoplasia was induced in 28 rats by intravesical installation of N-nitroso-N-methylurea (NMU) dissolved in acetate buffer. Fifteen rats were installed with acetate buffer, and served as controls. Urothelial samples were taken from all animals, the atypia was graded and detailed data on the location of the antibody binding structures were obtained by immunohistochemical methods. In addition, Western Blots of glycoproteins and thin-layer-chromatography (TLC) immunostainings of glycolipids extracted from normal and malignant tissue were performed to characterize the molecules presenting T-antigens. Examination of the histologic distribution of T-antigens showed that both the monoclonal and the polyclonal reagents reacted with atypical cells in proportion to the grade of atypia, but showed no reaction in invasive cells. These results confirm previously obtained data on the T-antigen using peanut (arachis hypogaea) agglutinin (PNA), and support the structure identity as being the classical O-linked mucin type T-antigen. Western blots of tumor glycoproteins showed that the monoclonal and the polyclonal antibody reacted with epitopes different from that of PNA, but all the probes correlated with atypia. In addition PNA, as the only anti-T reagent, bound to glycolipid. By using well characterized and highly specific immunological reagents the present study shows that the T-antigen is a highly selective marker of urothelial atypia.

Animals↗

PubMed1992

Squamous cell metaplasia of the bladder urothelium. A retrospective study of 36 patients.

In a retrospective study of 28 women and eight men with squamous cell metaplasia in different parts of the bladder, including the trigone, no histopathological differences were observed among the regions. All the five (female) patients with parakeratosis had a concomitant invasive bladder tumour. Thirty-eight% of all the patients had a simultaneous neoplastic tumour. The metaplastic lesions were investigated for keratin in 13 patients, and all were positive. In seven out of eight patients, the urothelium adjacent to the squamous cell metaplasia was also positive for keratin, indicating a direct transformation of the urothelium to squamous cell epithelium. The metaplastic cells were investigated for oestrogen receptors in five men and five women, and all were negative, suggesting no relationship between estrogens and squamous cell metaplasia of the bladder. Squamous cell metaplasia in the bladder is not considered a premalignant condition. However, metaplasia and neoplastic tumours are often associated with chronic tissue damage, and the presence of metaplasia may give a warning of conditions that can also cause cancer.

Adult↗

PubMed1992

Carbohydrate changes in bladder carcinomas.

The cloning of glycosyltransferases is a major breakthrough in the study of glycosylation in normal an malignant tissue. In bladder tumours we need a molecular explanation for the changes observed in especially Gal alpha 1-3 and GalNac alpha 1-3 transferase activity. Studies of the events (transcription/translation) taking place in the different cell layers would also be of importance, especially if the cells could be sorted layer by layer. The urine is a source of malignant cells, and could be a target for diagnosis as well as monitoring. Studies of urinary tumour-associated glycoconjugates or tumour-associated rearrangements in glycosyltransferase genes in exfoliated cells would be of great interest.

ABO Blood-Group System↗

PubMed1991

Differential tissue expression of the Lewis blood group antigens: enzymatic, immunohistologic, and immunochemical evidence for Lewis a and b antigen expression in Le(a-b-) individuals.

The Lewis blood group system comprises two main carbohydrate antigens, Le(a) and Le(b). Lewis typing has traditionally been based on serologic determinations using erythrocytes and saliva. Several recent studies have demonstrated that erythrocyte Lewis phenotype may change during pregnancy or disease, and inappropriate Lewis antigens have been found in both normal and neoplastic tissue. To evaluate whether these observations are in conflict with the presently proposed genetic and biosynthetic basis of the Lewis blood group system, we performed a combined enzymatic, immunohistologic, and immunochemical study of Lewis antigen expression in normal and neoplastic tissues, as well as erythrocytes, plasma, and saliva of Le(a-b-)-typed individuals. Of six cancer-bearing patients typed Le(a-b-), three were identified as nongenuine owing to the presence of alpha 1----4fucosyltransferase activity (alpha 1----4FT) and Lewis antigens in saliva and three were identified as genuine (lacking alpha 1----4FT and Lewis antigens in saliva). These genuine Le(a-b-) individuals were shown to express significant alpha 1----4FT in tissues, and Lewis antigens were detected in tissues by immunohistology as well as immunochemistry. We conclude that the Lewis phenotype obtained by serologic determination of erythrocytes and saliva does not apply to all tissues. We discuss biosynthetic and genetic consequences of this finding.

Colon↗