PubMed Health⌕ Search

Biomedical subjects

W H Fishman

Publications and source records attributed to W H Fishman.

At least 55 records · Page 3Linked to original sources
PubMed1977

Inhibition of human placental-type alkaline phosphatase variants by peptides containing L-leucine.

The inhibition of normal human placental alkaline phosphatase, its rare "D-variant" phenotype, and the "Nagao isoenzyme" found in an ovarian cancer fluid by peptides containing L-leucine were studied. The Nagao isoenzyme and the D-variant phenotype were distinct from the normal placental enzyme, but similar to each other in their inhibition by L-leucine and dipeptides containing L-leucine in their amino termini. On the other hand, L-leucinamide, L-Leu-L-Leu-L-Leu and L-Leu-Gly-Gly inhibitions did not readily discriminate between D-variant and normal phenotypes, but did so between D-variant and Nagao isoenzymes. The distinctions among these enzymes in their responses to peptide inhibitors may reflect differences in their primary structures.

Alkaline Phosphatase↗
PubMed1976

Carcinoplacental alkaline phosphatase. Base level and hormone-induced activity associated with events in the cell cycle.

Using HeLa TCRC-1, a cell line which is monophenotypic with respect to the Regan isoenzyme of alkaline phosphatase, we have examined the factors which influence its expression in relation to events of the cell cycle. DNA synthesis is not required for hormone induction of the Regan isoenzyme as in the presence of hydroxyurea, a specific inhibitor of DNA synthesis, we found induction to occur. Additionally, when partially synchronised cells were allowed to leave the S period prior to hormone treatment, and hydroxyurea was added to prevent cells from entering the next S period, hormone induction of the Regan isoenzyme was still observed. This indicates that initiation of expression of hormone-induced carcinoplacental alkaline phosphatase occur prior to the DNA synthetic phase of the cell cycle. We propose a hypothetical two-step mechanism of hormone induction to interpret the present findings in relation to previous results.

Alkaline Phosphatase↗
PubMed1976

Direct immunoperoxidase staining for Regan isoenzyme of alkaline phosphatase in human tumor tissues.

Immunoperoxidase staining for Regan isoenzyme of alkaline phosphatase was performed on cryostat sections of five human tumor tisssues. With a direct immunoperoxidase staining for the localization of Regan isoenzyme at the light and electron microscope levels, sections previously fixed with 0.05 M phosphate-buffered 4% paraformaldehyde were reacted with rabbit antisera to human placenta alkaline phosphatase conjugated to horseradish peroxidase. Comparison of conventional histochemistry and immunohistochemistry for Regan isoenzyme indicated that strong specific immunoperoxidase staining appeared on the cell membrane surface, and a diffuse one, in the cytoplasm of lung and colon cancer tissue cells showing L-phenylalanine-sensitive alkaline phosphatase. No immunoperoxidase reaction was obtained in tumor cells showing sensitivity to L-homoarginine or lacking aklaline phosphatase activity.

Adenocarcinoma↗
PubMed1976

Isoelectric focusing of alkaline phosphatase isoenzymes in polyacrylamide gels. Use of Triton X-100 and improved staining technic.

The examination of alkaline phosphatase isoenzymes by means of isoelectric focusing in polyacrylamide gel rods using the apparatus and focusing method of Righetti and Drysdale is discussed. A simultaneous coupling procedure using alpha-naphthyl phosphate and fast blue salt R in 2-amino-2-methyl-1,3-propanediol buffer, pH 9.68, containing MgCl2 and ZnSO4 proved sensitive for developing the enzyme bands. Also discussed are the effects seen with the incorporation of Triton X100 into the gel and sample mixtures. Enzyme, which remained at the top of the gel without using this detergent, entered the gel easily with the addition of Triton X-100 into the application solution. Incorporation of Triton into the gel matrix resulted in some enzyme band patterns that showed distinct differences from gels containing no Triton.

Acrylamides↗
PubMed1976

Developmental phase-specific alkaline phosphatase isoenzymes of human placenta and their occurrence in human cancer.

Alkaline phosphatase electrophoretic patterns characteristic of three phases in early human trophoblast development are described in this preliminary communication. Phase 1 (6 to 10 weeks) consists entirely of two heat-sensitive, L-homoarginine-inhibited bands, the slower one of which possesses antigenic determinants of live-bone-type alkaline phosphatase, whereas the fast band lacks any of the known alkaline phosphatase antigenic determinants. Phase 2 pattern (11 to 13 weeks) is that of a mixture of Phase 1 and Phase 3 isozyme components, the latter exhibiting two isozyme bands with the characteristics of term placental alkaline phosphatases correspond in order to non-Regan isoenzyme, a mixture of Regan and non-Regan isoenzymes and Regan isoenzyme in a variety of human cancer tissues. The biochemical profile characteristic of trophoblast developmental Phase 1 alkaline phosphatase is expressed as 78.5% heat-sensitive inhibition (5 min at 65 degrees), 66.3% L-homoarginine inhibition, and 17.3% L-phenylalanine inhibition where n = 12. It is hypothesized that the alkaline phosphatase of human tumor tissues reflects the expression of placental genes corresponding to one or more phases of trophoblastic development.

Alkaline Phosphatase↗
PubMed1976

Activation of developmental genes in neoplastic transformation.

The view is advanced that it is important to fix the stage of development corresponding to the pattern of expression of oncodevelopmental proteins in neoplastic transformation and neoplasia. From a map of development focused on events beginning with the zygote and ending with the establishment of fetus and placenta, it is possible to explain why certain developmental gene products are restricted to fetus or placenta or distributed in both. It also suggests which products can be expected to be expressed concordantly in neoplastic transformation and in neoplasia.

Alkaline Phosphatase↗
PubMed1976

Regulatory controls of oncotrophoblast proteins and developmental alkaline phosphatases in cancer cells.

The two oncotrophoblast proteins, Regan isoenzyme (placental-type alkaline phosphatase) and human chorionic gonadotrophin, are readily studied oncodevelopmantal gene products in human cancer patients and in three experimental model systems. The latter consists of (a) HeLa sublines TCRC-1 and TCRC-2, which produce Regan and non-Regan isoenzymes, (b) HEp-2 and FL amnion cell lines as models for the reciprocal expression of developmental genes, and (c) modulation in vivo of developmental gene expression in HeLa cells. In the case of the third model, for example, HeLa TCRC-1 cells grow in immunosuppressed rats to form a tumor nodule, which expresses a new oncoamnion (FL) isoenzyme, while the Regan isoenzyme disappears. Return of the tumor cells to cell culture medium results in a disappearance of the oncoamnion (FL) species and the reappearance of Regan isoenzyme. This interesting model is expected to bridge the interpretation of experiments done in cell culture with observations made on tumors of cancer patients. Most helpful in interpretation of all these studies has been a chronology of early development. It appears that the counterparts of a number of tumor proteins appear as early as gametogenesis and as late as 10 weeks of gestation.

Alkaline Phosphatase↗
PubMed1975

Purification and characterization of mouse kidney beta-glucuronidase.

Beta-Glucuronidase has been purified from mouse kidneys previously induced by gonadotrophin to a specific enzyme activity 15 times higher than the non-induced kidney. The purification procedure includes ultrasonication to solubilize the enzyme, acid and ammonium sulfate precipitations, gel filtration in Sephadex G-200, DEAE-ion exchange chromatography, and isoelectric focusing. The resulting product has a specific activity of 284,000 Fishman units/mg of protein, representing a 1,090-fold purification and is 17,000-fold higher than the level in the non-induced kidney. The purified beta-glucuronidase is apparently homogeneous by criteria of gel filtration, sodium dodecyl sulfate gel electrophoresis, and immunodiffusion. Characterization of the purified enzyme showed that it is identical with the lysosomal isoenzymic from electrophoretically, has subunit molecular weight of 74,000 (estimated by sodium dodecyl sulfate gel electrophoresis) and oligomer molecular weight of 300,000. The purified enzyme is stable at high temperature (up to 55 degrees) and at wide range of pH (from 4 to 11). It has a pH optimum for its activity at 4.7 and a Km of 1.18 times 10- minus 4 M. The purification and characterization of this enzyme from mouse kidney will have significance in the understanding of the molecular nature of the isoenzymes of beta-glucuronidase and will be useful in future studies on the mechanism of intracellular transport and distribution of this hydrolase.

Animals↗
PubMed1975

Demonstration of lysosomal and extralysosomal sites for acid phosphatase in mouse kidney tubule cells with p-nitrophenylphosphate lead-salt technique.

Dual localization of acid phosphatase in lysosomal and extralysosomal sites of the tubule epithelial cells of normal mouse kidney was observed at the light and electron microscope level using a modified Gomori lead-salt method with p-nitrophenylphosphate (pNPP) as substrate. Based on previous biochemical and cytochemical findings, we developed optimal conditions for the enzyme activity in extralysosomal sites. The conditions used for the light microscopic level consisted of 1.5 mM PNPP, 2.0 MM Pb(NO3)2 and 0.05 M acetate buffer (pH 5.8). Those for the electron microscopic study required 3.0 mM PNPP, 3.6 MM Pb(NO3)2 and 0.1 M acetate buffer (pH 5.8). This modified lead-salt technique was highly specific and provided a suitable method for the demonstration of nonlysosomal as well as lysosomal sites of acid phosphatase activity in the tubule epithelial cells of normal mouse kidney. As expected, the enzyme activity appeared in the lysosomes, but the prominent reaction in the brush border, the rough endoplasmic reticulum and basal infolding plasma membranes was not anticipated. We were able to demonstrate in situ organelle precursors of microsomal acid phosphatase such as endoplasmic reticulum, plasma membrane and basal infolding membranes showing the same substrate preference, which had been observed previously in biochemical studies in our laboratory. Since the possible participation of alkaline phosphatases, K+-pNPPase or Na+-K+-adenosine triphosphatase was ruled out by use of appropriate inhibitors, the enzyme-reactive sites can be interpreted as reflecting nonspecific acid phosphatase.

Acetates↗
PubMed1975

A simple radioimmunoassay of human placental alkaline phosphatase (Regan isoenzyme) using specific antibody polymers.

Rabbit antiserum against highly purified high-molecular-weight B-variant of human placental alkaline phosphatase (M.W. 200,000) was rendered monospecific by absorption with polymerized pooled male serum proteins; the absorbed antiserum was then polymerized with ethyl chloroformate and used in radioimmunoassay as a stable solid-phase immunoabsorbent. Homogeneous preparation of the enzyme, with a specific activity of 477 mumoles phenol per mg per min, was also obtained by absorbing the chromatographically purified enzyme with polymerized rabbit antiserum directed to whole human serum proteins; the pure enzyme was then labeled with 125-I as the tracer retaining at least 80% of its antigenicity. Only a minute quantity of the polymerized antibody particles is required for each assay in admixture with the labeled and unlabeled enzyme. By adding a small amount of starch-gel particles before low-speed centrifugation, complete phase separation was achieved. The radioimmunoassay could detect 0.4 to 0.8 ng enzyme protein per tube, which is comparable to the sensitivity achieved by enzymic assays. However, radioimmunoassay is advantageous over the enzymic assay in being direct, specific (no interference by the nonplacental-type alkaline posphatase), and capable of detecting both catalytically active and inactive forms of the enzyme. Native variants of placental-type alkaline phosphatase including Regan isoenzyme and Nagao isoenzyme (D-phenotype of normal placental alkaline phosphatase), could thus be directly determined by this procedure in the clinical specimens.

Alkaline Phosphatase↗
PubMed1975

Regan isoenzyme and human chorionic gonadotropin in ovarian cancer.

Among 833 cancer patients whose sera were investigated for Regan isoenzyme and among 1,319 cancer patients from a different population whose sera were assayed for human chorionic gonadotropin (HCG), those patients with neoplasms of the testis or ovary showed the highest frequency of both placental proteins. Among another 22 patients with ovarian cancer, for whom both placental proteins were measured, 59% showed Regan isoenzyme and 68% showed HCG in ascitic fluids, whereas the figures were 65% and 30%, respectively, for sera. In 55% of both fluids and sera, there was a positive correlation of Regan isoenzyme with HCG (positive or negative). Almost invariably, the ascitic fluid was richer in Regan isoenzyme and HCG than the serum when both were collected on the same day. Progressively increasing levels of each placental protein generally correlated with the spread of the disease, though there were instances when only one was expressed. Evidence indicated the existence of two forms of alkaline phosphatase in ovarian cancer, Regan and non-Regan; the latter was assumed to be of fetal origin. Ultrastructural studies of one ovarian cancer revealed a morphologic entity, i.e., mitochondria enveloped by inverted tubules of endoplasmic reticulum.

Aged↗
PubMed1975

Markers for ovarian cancer: regan isoenzyme and other glycoproteins.

Since embryonic genes are not generally active in normal adult subjects and because certain of these genes are activated in cancer leading to ectopic synthesis, it is the difference between the ectopic level and the normal adult concentrations of embryonic gene products which we seek in developing "markers" for ovarian cancer. The carcinoplacental alkaline phosphatases corresponding to the term gestational phenotypes correlate positively with ovarian cancer as does hCG. Other fetal and placental glycoproteins whose presence is noted in ovarian cancer include CEA, alpha-FP, and Björklund's antigen. Antigens of mucinous cystadenocarcinoma have not yet been examined for their possible fetal or placental origins. The degree of concordance of expression of Regan isoenzyme and hCG is variable. Profiles of glycoproteins would appear to offer an opportunity to inquire more deeply into the nature of ovarian cancer and from this inquiry, one can expect to develop a system of markers which can be of clinical use.

Alkaline Phosphatase↗