bluuebunny/crossref_metadata_2025 huggingface dataset:
Biochemical Properties of Human Urinary Megakaryocyte Colony-Stimulating Factor and Erythropoietin: The Role of Sulfhydryl Groups and Disulfide Bonds
Authors: Shimizu Tomoe , Miyake Takaji , Pilch Ann M. , Mantel Charles , Murphy, Jr. Martin J.
Published: October 2008

The labeling of cystine residues with [l-14C]iodoacetic acid showed that urinary preparations from patients with aplastic anemia contained 3.06 × 10-9 mol of sulfhydryl groups and 2.90 × 10-7 mol of half-cystine as disulfide bonds in the native state, and 6.36 × 10-7 mol in the denatured state per absorbance unit of protein, respectively. Sulfhydryl reagent-treated proteins retained full activity of megakaryocyte colony-stimulating factor (Meg-CSF) and erythropoietin (Epo), except with DTNB-treated protein. Reduction-car-boxymethylation and reduction-mercuration resulted in complete loss of Meg-CSF and Epo activities, suggesting that one of the essential chemical groups of Meg-CSF and Epo is a disulfide bond. Reduction of disulfide bonds at neutral pH revealed that Meg-CSF is less susceptible to reduction than Epo. Reactivation occurred by spontaneous reoxidation in most of the reduced Meg-CSF (92.6%) and part of the reduced Epo (22.1 %). These molecular behaviors may reflect differences in the spatial configurations of Meg-CSF and Epo.
Retinoic Acid-Induced Differentiation of Human Neuroblastoma: A Cell Variant System Showing Two Distinct Responses
Authors: Sidell Neil , Sarafian Theodore , Kelly Michael , Tsuchida Tetsuya , Haussler Mark
Published: October 2008

Retinoic acid (RA) has been shown to induce the differentiation of human neuroblastoma cells in vitro. In this study, we describe two variants of the SK-N-SH human neuroblastoma cell line thai have dramatically different responses to RA. RA induces neuronal-like differentiation characterized by extensive neurite outgrowth, thick neurite bundles, and large cellular aggregates of SK-N-SH-N (SH-N) cells. In contrast, RA treatment of SK-N-SH-F (SH-F) cultures transforms the small neuroblast cells into large flattened, fibroblastic or epithelial-like cells. Karyotype analysis verified that the SH-N and SH-F cultures were derived from a common precursor cell. Confirmation of their markedly different responses to RA was obtained by metabolic labelling of glycol-proteins and SDS-PAGE analysis. While both sublines showed very similar Coomassie-labelled protein bands and glycol-protein profiles in control cultures, dramatic differences between the lines were revealed following RA treatment. In contrast to their similar protein profiles, untreated SH-N and SH-F cells had quite different patterns of ganglioside biosynthesis in that GM3 was detected in SH-F cells but not in SH-N, while GM1 was only detected in SH-N. Cellular RA binding protein (CRABP) was detected in both SH-F and SH-N cells and their RA-transformed derivatives. These results demonstrate heterogeneity in the response to RA of neuroblastoma cells derived from a common origin that cannot be accounted for by differences in CRABP content. The SH-N and SH-F neuroblastoma sublines should provide a useful system for further studies of the molecular processes through which RA exerts its differentiation-inducing activity on this type of tumor.
Embryogenesis of Stratified Squamous Epithelium
Authors: Shuler Charles F. , Schwartz Stephen A.
Published: October 2008

Although several types of modified stratified squamous epithelia are present in the adult mammal, most are presumably derived from similar embryonic progenitors. Mechanisms responsible for region-specific specialization are poorly understood. To correlate epithelial diversification with the expression of tissue-specific markers, we analyzed keratin protein composition in four representative types of rat squamous epithelia from early embryonic through adult phases of development. Keratin subsets synthesized in palatal mucosa were qualitatively similar, but differed dramatically in relative abundance. Tongue mucosa synthesized a different, but consistent subset of keratins which also changed quantitatively throughout morphogenesis. In contrast, different keratin genes were sequentially expressed during histogenesis of backskin and footpad epidermis. These data indicate that tissue-specific keratin biosynthesis is genetically predetermined early in embryogenesis.
Embryogenesis of Stratified Squamous Epithelium
Authors: Shuler Charles F. , Schwartz Stephen A.
Published: October 2008

Throughout embryogenesis of the rat palate, from the early fetal to adult stages, a consistent subset of keratin proteins is synthesized in the epithelial lining cells. Although the relative abundance of particular keratins has been shown to vary with ongoing palatogenesis, the expression of finite keratins appears to be genetically predetermined. In order to preliminarily ascertain whether conformational changes accompanied intermediate filament ‘maturation’ from monomeric to polymeric keratin formation, we screened cytokeratins with polyclonal and monoclonal antibodies generated against adult-type keratins. Until epithelial stratification occurred on the 16th day of gestation, the keratin proteins were weakly immunoreactive. On the other hand, subsequent to epithelial thickening, adult-type immunoreactivity was initiated and progressed concomitantly with ongoing palatal development. These findings suggest that the cytokeratin intermediate filaments may progress through conformational ‘maturation’ during polymerization, and play a role in the eventual acquisition of the adult-type epithelial structure and function.
Human Embryonic Liver Cell Differentiation
Authors: Hakim Anwar A. , Siraki Charles M. , Dube Volker E.
Published: October 2008

When incubated in preconditioned medium, i.e. spent media from the hepatocellular carcinoma (PLC/PRF/5) cell cultures, human embryonic liver (HEL) cells differentiate and acquire oncologic phenotypes. This is caused by transcriptional alterations in fetal gene expression. This occurs when hepatocytes are proliferating rapidly and secreting alpha fetoprotein (AFP), and later when the quiescent state is reached with secretion of albumin (ALB). The present studies examined the parameters signaling oncologic transformation during liver cell differentiation in the conditioned medium. mRNAs coding for AFP, ALB and HBsAg were isolated from HEL, adult liver cells (ADLC) and from PLC/PRF/5 cells, respectively. cDNA molecules complementary to these polysomal mRNA molecules were constructed and labeled with 32P. These tracers were used to quantitate changes in cellular mRNA • AFP, mRNA • ALB and mRNA·HBsAg directly by DNA molecular hybridization during HEL cells cultivation in the preconditioned medium. Under these conditions, the changes in cellular mRNA·HBsAg and mRNA • AFP correlated with an increased tumorigenicity in athymic Nu/Nu mice, membrane galactosyltransferase and phospho-tyrosine kinase activities.
Hypothesis: Persistent Expression of Fetal Phenotypic Characteristics by Fibroblasts Is Associated with an Increased Susceptibility to Neoplastic Disease
Authors: Schor S.L. , Schor A.M. , Howell A. , Crowther D.
Published: October 2008

Published data from our own and other laboratories have indicated that fibroblasts obtained from patients with different types of epithelial cancers commonly display aberrant (i.e. fetal-like or transformed) phenotypic characteristics. Previous interpretations of these results have tended to regard the study of fibroblasts as a convenient means to demonstrate genetic abnormalities also expressed in the target epithelial cell population and did not ascribe a particular causative role to fibroblast abnormalities in the genesis of neoplastic lesions. In this communication we present an alternative, but not mutally exclusive, hypothesis suggesting that aberrations expressed solely by fibroblasts may lead to the development of an epithelial tumour by virtue of a dysfunction in normal epithelial-mesenchymal interactions.
Transglutaminase Activity and Embryonal Carcinoma Cell Differentiation
Authors: Uhl Lynne , Schindler Joel
Published: October 2008

Murine embryonal carcinoma (EC) cells induced to differentiate by retinoic acid (RA) modulate transglutaminase (TGase) activity shortly after exposure to the inducer. Compounds that inhibit TGase enzyme activity in vitro can successfully block RA induced EC cell differentiation in culture. These observations suggest that TGase may play a role in mediating RA induced EC cell differentiation.
Alkaline Phosphatase Expression in Human Chorionic Villi
Authors: Novelli Giuseppe , Mannello Ferdinando , Cosmi Ermelando V. , Biagioni Stefano , Dallapiccola Bruno
Published: October 2008

The physicochemical properties and electrophoretic mobility of different isoforms of alkaline phosphatase were studied in chorionic villi. Based on selective inactivation and inhibition studies (thermal stability, inactivation by urea, EDTA and L(+)ascorbic acid and L-amino acid inhibition), evidence was obtained for the existence of two distinct types of alkaline phosphatase in trophoblast cells. One type is peculiar to chorionic villi while the other is also found in term placenta. Both show two isoforms. These two isoforms were observed with polyacrylamide gel electrophoresis, carried out at pH 6.0 and 9.5. It is suggested that the qualitative and quantitative methods of alkaline phosphatase analysis could be used for first trimester fetal diagnosis of severe infantile hypophosphatasia and for understanding genetic control during early fetal development.
Activity and Function of Hybrid Ribonuclease in Cells of Acute and Chronic Myelogenous Leukemia
Authors: Papaphilis Anastasios D. , Kamper Elli F. , Pangalis Gerassimos A. , Grammenou Stavroula , Kattamis Christos
Published: October 2008

The activity of hybrid ribonuclease (ribonuclease H) has been determined in mononuclear blood cells (lymphocytes plus monocytes) from 23 normal individuals and cells (pool of immature granulocytes, metamyelocytes and lymphocytes) from 35 untreated acute and chronic myelogenous leukemia cases. It was found that in 86% of the leukemic samples the activity of ribonuclease H was above two standard deviations from the mean activity level drawn for the group of normal samples along the 0–100% substrate hydrolysis scale. The activity of the enzyme in leukemic cells correlated linearly with the DNA-synthesizing activity of the cells in vitro and in the examined CML cases it paralleled the inverse relationship of the incorporation of tritiated thymidine into DNA to the size of the pool of immature granulocytes. In one CML patient who received chemotherapy with Myleran, the activity of ribonuclease H, high at the initiation of drug therapy, was reduced to a normal level at remission, but increased again at the stage of subsequent relapse. These findings indicate that the levels of ribonuclease H in leukemic cells reflect the proliferative activity of the population in the cases of untreated myelogenous leukemias.
Interaction of Retinoids and Tamoxifen on the Inhibition of Human Mammary Carcinoma Cell Proliferation
Authors: Fontana Joseph A.
Published: October 2008

The growth of chemically induced mammary tumors is inhibited by both hormone manipulation as well as by retinoids. Numerous mammary carcinoma cell lines are also inhibited by retinoids. Co-treatment of estrogen receptor (ER)-positive breast cancer cells resulted in an additive effect in terms of inhibition of cellular proliferation. The addition of varying concentrations of retinoic acid (RA) to varying concentrations of tamoxifen (TMX) resulted in an additive effect on the inhibition of proliferation of the ER-positive human carcinoma cell lines (MCF-7). Co-treatment of MCF-7 cells over time with RA and TMX resulted in enhanced inhibition of growth. A similar phenomenon was observed when other synthetic retinoids were combined with TMX. This enhanced inhibition by the combination of retinoids and TMX was also observed with other ER-positive cell lines (ZR-75, T47-D), while no effect was noted on the ER-negative cell lines (MDA-MB-231, Hs578T).
Autoneuralization in the Amphibian Ectoderm -A Species-Specific and Stage-Specific Phenomenon
Authors: Mattsson Mats-Olof , Nakatsuji Norio , Løvtrup Søren
Published: October 2008

Ectoderm from Ambystoma is especially prone to undergo ‘autoneuralization’. This assertion has led to the maxim that ectoderm from this species is unsuitable for studying cell differentiation. Here we report that the degree of neuralization in cultured explants is stage-dependent. Control explants from blastulae (stage 8-9) show no neuralization, while explants treated with LiCl (10 mM) give rise to neuralization in about 70% of the cases. This difference between control and experiment decreases during gastrulation, in late gastrulae (stage 12) it is more or less negligible. Ectoderm from Cynops pyrrhogaster reacts like that of Ambystoma when exposed to LiCl, but like Triturus ectoderm it is insensitive to cyclic nucleotides.
Initiation of Primary Cell Cultures from Human Intracranial Tumors on Extracellular Matrix from Bovine Corneal Endothelial Cells
Authors: Westphal Manfred , Hänsel Marianne , Brunken Martin , König Albrecht , Köppen J.A. , Herrmann Hans-Dietrich
Published: October 2008

Tissue specimens from 105 human gliomas and 57 human meningiomas were obtained at surgery, dissociated into single cells and small cell aggregates and then plated onto plain plastic tissue culture dishes and dishes which had been precoated with an extracellular matrix (ECM) derived from bovine corneal endothelium. In 80% of the glioma cases we observed a marked improvement in initial plating efficiency, colony formation and speed of attachment when cells were plated on ECM. In 5 cases cells attached only to the ECM-coated dishes but remained afloat in the untreated dishes. In addition it could be noted that over the first 2 days, those cells which had been initiated on ECM showed more signs of morphological differentiation, i.e., extension of cytoplasmic processes or formation of fiber networks between cell groups. If adaptation occurred and proliferation began in vitro, either immediately or after a several days’ lag phase, both the ECM-cultured cells as well as those which slowly had adapted to culture on plastic could be passed on to untreated culture ware and perpetuated thereon. In the case of well-differentiated low-grade gliomas where no growth in culture took place, the cultures on ECM could at least be used for initial experiments in the primary cultures (P0). Meningiomas usually attached well to both, plastic or ECM. In 50% of our cases the plating efficiency was higher on ECM but after successful initial culture, the delay until the cells on plastic reached confluence in comparison with those on ECM was 1 or 2 days. Again there were 2 cases in which the cells would not plate on plastic. Here the cells which after 1 day were still afloat plated to more than 80% within the first 2 h after transfer to ECM. In all cases the cells from plastic and ECM cultures were indistinguishable and could be passed onto untreated dishes henceforth. In later culture stages ECM offers several advantages: It is easier to shift cells to serum-free defined culture conditions, the cells will grow at a faster rate on ECM when in higher passages and the maximal number of passages possible is higher on ECM.
Polyadenylate Polymerase Activity in Stationary and Growing Cell Cultures
Authors: Kazazoglou Theodossia , Tsiapalis Chris M. , Havredaki Maria
Published: October 2008

Soluble polyadenylic acid (poly(A)) polymerase content of stationary and growing cell populations from a variety (6) of cell lines was determined. Cell populations from stationary cultures presented poly(A) polymerase values with a mean of 31 ± 12 enzyme units/mg protein. The mean value for growing cell populations were 62 ± 18 enzyme units per mg protein. A statistically significant difference was found between stationary and growing cell populations from the variety of cell lines examined (p < 0.1). The observed differences in poly(A) polymerase levels persisted after fractionation of the crude extracts and revealed two molecular forms of enzyme activity with a net charge difference in stationary and growing cell cultures.
Transfer of Granulomatous Inflammation with Nonviable Preparations of Schistosome Granulomas in Naive Mice
Authors: Okamoto Mitsuse , Epstein William L. , Suya Hideko , Kanazawa Kazunori , Fukuyama Kimie
Published: October 2008

Hepatic granulomas of euthymic (nu/+) mice infected with Schistosoma mansoni were freeze-dried or freeze-thawed 3 times and transplanted subcutaneously into naive nu/+ and athymic (nu/nu) mice. The grafted sites, studied histologically, showed formation of organized granulomas in nu/+ mice similar to donor granulomas as observed after grafting of freshly isolated granulomas. On the other hand, in nu/nu mice, the nonviable transplants elicited small and disorganized granulomas, like hepatic granulomas in nu/nu mice with schistosomiasis, but different from fresh nu/+ transplants in nu/nu skin. The findings indicate viable cells are not required for transfer of granulomatous reactions, but T cells are needed for full expression.
Morphometric Analysis of Collagen Fibrils in Idiopathic Carpal Tunnel Syndrome: Part 2
Authors: Weis Serge , Stransky Gregor , Dimitrov Leonid , Wenger Emanuel , Neumüller Josef , Hakimzadeh Ahmed , Firneis Fred , Partsch Gerald , Eberl Rudolf
Published: October 2008

Morphometric parameters were evaluated in order to analyze the relation between number and covered area of collagen fibrils in normal and carpal tunnel syndrome tissue. This analysis revealed that in normal tissue twice as many collagen fibrils as in pathological tissue occupy an equal area. Taking these facts into account, some hypotheses are advanced.
Proliferation of Human Carcinoma Cells in Calcium-Deficient Culture Medium May Depend on Autocrine Growth Factor(s)
Authors: Swierenga Sabine H.H. , Auersperg Nelly
Published: October 2008

The ability to proliferate in media with low calcium concentrations ( < 0.1 mM) at clonal seeding densities is a characteristic of malignant cells. Cells of the carcinoma line C-4I are an exception, and are unable to proliferate in medium with 0.02 mM calcium (LCM) when seeded at low densities. Conditioned medium, derived from a subline of C-4I cells that were adapted to grow in LCM, contained an acid-stable factor of > 8 kilodaltons molecular weight that mediated proliferation and colony formation of unadapted C-4I cells in LCM in a concentration-dependent manner. The effect was not related to enhanced cell attachment or spreading, and was maximal when the unadapted C-4I cells were seeded in aggregates of 2-20 cells rather than singly. Thus, calcium independence, a component of the neoplastic phenotype, may be mediated by autocrine tumor-cell-derived factor(s) and may involve long- and short-range cell interactions.
Differential Effects of Sodium Butyrate and Dimethylsulfoxide on Gamma-Glutamyl Transpeptidase and Alkaline Phosphatase Activities in MCF-7 Breast Cancer Cells
Authors: Wasserman Lina , Nordenberg Jardena , Beery Einat , Deutsch Alexander A. , Novogrodsky Abraham
Published: October 2008

Sodium butyrate and dimethylsulfoxide (DMSO), two known chemical inducers of cell differentiation, were examined on MCF-7 breast cancer cells. Both agents reduce the proliferative capacity of MCF-7 cells, as reflected by inhibition of colony formation in semi-solid agar. Sodium butyrate is shown to enhance markedly the activity of two plasma membrane-bound enzymes, alkaline phosphatase and γ-glutamyl transpeptidase. DMSO does not enhance the activity of these enzymes, but rather induces a small decrease in γ-glutamyl transpeptidase activity. The present results show that although both agents inhibit cell proliferation, they have a distinct effect on phenotypic expression.
Isolation and Morphologic Characterization of Human Ovarian Carcinoma Cell Clusters Present in Effusions
Authors: Allen Howard J. , Porter Carl , Gamarra Marie , Piver Steven , Johnson Edward A.Z.
Published: October 2008

Serous, mucinous, endometrioid and clear cell human ovarian carcinoma cells were isolated as multicellular aggregates from patient effusions by filtration on nylon mesh of defined porosity and examined by light microscopy. The cell clusters ranged from compact to loosely adherent groups of cells to spheroids with a central lumen surrounded by a cell monolayer. There was considerable variation in cluster morphology between effusions from different patients as well as within effusion from the same patient. Apparent budding of clusters was observed as well as different stages of cluster growth and development. This was observed for all histologic types studied. Electron microscopy of serous, mucinous and clear cell types showed that cells forming clusters were attached to each other by desmosomes, demonstrating that cluster formation did not result from a nonspecific stickiness of cells. Irregular micro villi were present on the external periphery of the various carcinoma cells and a prominent glycocalyx was present on the surface of mucinous carcinoma cells. Extensive interdigitation of cytoplasmic extensions and extended villi was present in mucinous and serous clusters which appeared to strengthen cluster cohesiveness. Nuclei were irregular with prominent nucleoli frequently present. The cell clusters usually remained intact and viable in culture but generally did not attach to glass or plastic substrata, whereas mesothelial cells and nonactivated histiocytes rapidly attached. When carcinoma cell clusters did attach, they were resistant to detachment by trypsin-EDTA treatment, in contrast to the nonmalignant cells.
Effects of Urinary Extracts from Patients with Idiopathic Thrombocytopenic Purpura or Aplastic Anemia on Rodent Platelet Production and Megakaryocytopoiesis
Authors: Kuriya Shin-ichiro , Kamamoto T. , Murphy, Jr. Martin J.
Published: October 2008

Urinary extracts from idiopathic thrombocytopenic purpura (ITP) patients, aplastic anemia (AA) patients and normal subjects were investigated for their effects on in vivo platelet production, and both in vitro and in vivo megakaryocytopoiesis in rodents. Daily intraperitoneal injection of 1.2 absorbance units (AU, A278) of urinary protein for three consecutive days induced statistically significant increases in rat blood platelet numbers. This increase was observed for 1 of 4 ITP urinary extracts and for all 3 AA urinary extracts, and occurred 24 h after the final injection. In vitro levels of megakaryocyte colony-stimulating factor (Meg-CSF) in ITP urinary extracts were similar to those of normal urinary extracts, and were in dramatic contrast to the markedly elevated levels of Meg-CSF in extracts from AA urine. A single intraperitoneal injection of 0.5 AU of AA urinary protein induced a significant increase in spleen-derived megakaryocyte colony-forming cells (CFU-meg) 48 h past injection. In the group injected with ITP urinary extract, CFU-meg levels remained within normal limits. These results provide evidence that urinary extracts of ITP patients do not contain increased levels of Meg-CSF and a factor which directly stimulates in vivo CFU-meg production, and that the decrease in circulating platelet numbers that is characteristic of ITP patients is not a primary in vivo determinant in the elaboration of these factors.
Human Bone Marrow CFU-GM and BFU-E Localized by Light Scatter Cell Sorting
Authors: Atzpodien Jens , Gulati Subhash C. , Kwon Jong H. , Wachter Maria , Fried Jerrold , Clarkson Bayard D.
Published: October 2008

Flow cytometric separation was performed on the normal human bone marrow (BM) by using the low-angle (0°) or high-angle (90°) light scatter. Four distinct subpopulations of cells can be enriched from normal human BM and these fractions were subsequently evaluated for their morphological properties as well as their clonogenic capacity in various progenitor cell assays. Our results indicate that human erythroid and granulocyte-macrophage progenitor cells can be separated from BM low-density cells by cell sorting, and these cells show similar 0° and 90° light scatter properties to those observed with murine bone marrow studies. Flow cytometric analysis also suggests that the majority of sorted BFU-E and CFU-GM resides in the blast cell subset of human BM mononuclear cells.
5-Azacytidine Inhibits Adipocytic Conversion of Human Skin Fibroblasts by Kirsten Murine Sarcoma Virus and Dexamethasone
Authors: Kopelovich Levy
Published: October 2008

Transformation of human skin fibroblasts (SF) by the Kirsten murine sarcoma virus (KiMSV) is associated with their neodifferentiation1 into preadipose cells. Hydrocortisone or dexamethasone (DX) promote the transformation/neodifferentiation of preadipocytes to fully differentiated adipocytes. 5-Azacytidine (5-Aza-CR) by itself, or together with DX, or immediately after inoculation with KiMSV did not cause adipocytic conversion of human SF. Addition of 5-Aza-CR either before or immediately after exposure of human SF to KiMSV and DX reduced the total number of foci and significantly inhibited expression of adipocyte-containing foci. The results suggest that adipocytic conversion of human SF by KiMSV and DX is specific. This conversion apparently does not involve mechanisms directly related to the effects of 5-Aza-CR on the adipocytic conversion of murine embryo cells.
Morphological Aspects of Interactions between Asbestos Fibers and Human Mesothelial Cell Cytoskeleton
Authors: Rüttner J.R. , Lang A.B. , Gut D.R. , Wydler M.U.
Published: October 2008

The interaction of chrysotile and amphibole asbestos fibers with the cytoskeleton of cultured human mesothelial cells from nontumoral pleural effusions was studied using scanning electron and immunofluorescence microscopy. Asbestos-exposed mesothelial cells show a massive annular condensation of cytokeratin filaments, forming a concentric ring enveloping the nucleus and the phagocytosed asbestos fibers. By detergent extraction of the cells it could be shown that the asbestos fibers are in close contact with the nuclear membrane and associated with the cytoskeletal framework of the cells. An association of cytokeratin filaments with the asbestos could be observed during phagocytosis of the fibers. The disturbance of the cell cytoskeleton and the close morphologic contact between asbestos fibers and the nuclear membrane may have some relevance in explaining the well-recognized carcinogenic effects of asbestos mineral fibers.
Influence of Albumin on Granulocyte and Macrophage Colony-Forming Efficiency
Authors: Iizuka Yoshikazu , Oshima Toshiteru , Murphy, Jr. Martin J.
Published: October 2008

Mouse granulocyte and macrophage precursors were assayed in plasma clot and fibrin clot cultures, and the effect of bovine serum albumin (BSA) on colony formation was investigated. The number of granulocyte colonies (CFU-g) and clusters increased as the albumin concentration was increased and the number of macrophage colonies (CFU-m) and clusters concomitantly decreased. The albumin-mediated suppression of macrophage colony formation was overcome by the addition of more than 10% fetal bovine serum (FBS) to the plasma clot culture. The effect of BSA and fatty-acid-free BSA on colony-forming efficiency was also tested in fibrin clot cultures containing 10% FBS. Both BSA and fatty-acid-free BSA at a final concentration of 0.5-2% enhanced CFU-g colony formation, while both forms of BSA reduced the number of CFU-m colonies. However, neither BSA nor fatty-acid-free BSA had any effect on colony formation in FBS-free fibrin clot cultures, and only BSA enhanced colony formation when transferrin, linoleic acid, α-thioglycerol and dextran were added to the culture. The number of CFU-g (15.6 ± 3.1) was higher in cultures containing BSA, transferrin, etc., than the number (9.8 ± 2.5) in cultures without BSA and including transferrin, etc. (p < 0.01). The number of CFU-m (32.0 ± 6.8) in cultures containing BSA and the other four factors was lower than the number (72.2 ± 5.6) in the culture without BSA (p > 0.001). These results indicated that although albumin was a factor regulating granulocyte and macrophage proliferation, by itself it did not stimulate colony formation. The effect of albumin was observed only in the presence of other factors.
Discrepancy between Flow Cytometric Analyses of CALLA Using Two Monoclonal Antibodies
Authors: Fried Jerrold , Perez Amaury G. , Shimazaki Chihiro , Clarkson Bayard D.
Published: October 2008

Both the J5 and BA-3 monoclonal antibodies are considered to be specific for epitopes on the common acute lymphoblastic leukemia antigen (CALLA). Flow-cytometric analyses of three cell lines and one normal bone marrow sample using these antibodies as CALLA markers demonstrated that J5-labeled cells were always brighter than those labeled with BA-3, and that the ratio of their fluorescence intensities varied widely in the different systems. Furthermore, one of the lines, RPMI 8226, while positive for J5, appeared to be negative when labeled with BA-3, except for a slight displacement of the fluorescence distribution relative to the control. A possible explanation for the observed results is that the BA-3 binding epitope or epitopes on CALLA may vary in their number and/or accessibility to the antibody. These observations suggest that the use of a single monoclonal antibody to detect a cell surface antigen may be misleading, particularly when a negative result is obtained.
Effects of Hyperinsulinism and of Diabetes on Proteoglycans of the Intervertebral Disc in Weanling Sand Rats
Authors: Silberberga Ruth , Adler Jonathan H. , Meier-Ruge William
Published: October 2008

Weanling sand rats (Psammomys obesus) develop hyperinsulinemia or diabetes or both, if fed a standard laboratory diet without a supplement of fiber rich salt bush. The annuli fibrosi of hyperinsulinemic or diabetic animals, which are still hyperinsulinemic, show a slight but statistically significant increase in chondroitin sulfate and a lesser, statistically nonsignificant increase in keratan sulfate. Possible causes of these changes are discussed and the likelihood of a role of hyperinsulinism in their production is pointed out.
Effect of Melphalan on Growth Curves and Cell Cycle Distribution of Four Human Small Cell Carcinomas of the Lung Grown in Nude Mice
Authors: Engelholm S.Aa. , Spang-Thomsen M. , Vindeløv L.L. , Brünner N.
Published: October 2008

Four human small cell carcinomas of the lung grown in nude mice were exposed to melphalan. Two of the tumors were derived from subpopulations isolated by in vitro cloning from the same tumor biopsy. The chemosensitivity of the tumors was determined by calculating the specific growth delay. Drug-induced changes in the cell cycle were detected by flow cytometric DNA analysis. The specific growth delay of the tumors was very different with the greatest differences between the two subpopulations originating from the same tumor. Melphalan induced a dose-related S phase accumulation in three sensitive tumors, whereas no changes were seen in a resistant tumor. Furthermore, the amount of S phase accumulation reflected the sensitivity to melphalan. The results suggest that heterogeneity in chemosensitivity is an important reason for chemotherapy failures.
Quantitative Evaluation of Anticancer Agents against Human Melanoma Cells Implanted in Nude Mice
Authors: Lockshin Arnold , Giovanella Beppino C. , Stehlin, Jr. John S.
Published: October 2008

BRO human melanoma cells, which are exceptionally tumorigenic and lethal for nude mice, were inoculated intraperitoneally or intracerebrally in varying numbers. An inverse linear correlation was observed between the logarithm of the number of cells inoculated and host survival. In mice bearing 107 cells intraperitoneally, 2.4–2.8 logio units of cell kill were obtained after a single intraperitoneal injection of vincristine, and some mice inoculated with 105 cells were cured by this treatment. Fewer cells were killed by L-phenylalanine mustard. Vincristine did not prolong survival of nude mice with intracerebral BRO tumors. Cell kill after administration of anticancer agents can be quantitated for BRO cells inoculated intraperitoneally or intracerebrally.
Antigen Presentation in Human Autoimmune Thyroid Disease
Authors: De Bernardo Erica , Davies Terry F.
Published: October 2008

Monocyte/macrophage function in patients with autoimmune thyroiditis was investigated by their presentation of two distinct antigens; sheep red blood cells (SRBC) and human thyroglobulin (hTg) using in vitro systems designed for antibody induction. Purified peripheral blood monocyte/macrophages were primed by prefeeding with antigen for 60 min at 37 °C, washed, and co-cultured with autologous lymphocytes under a variety of incubation conditions. The most successful system employed 5% monocyte/macrophages with autologous T-B cells in the presence of the mitogen Staphylococcus aureus and B-cell differentiating factors. Under such conditions the anti-SRBC plaque-forming cell (PFC) response was amplified equally (approximately 10-fold) by SRBC-fed monocyte/macrophages in normal controls and patients with autoimmune thyroiditis rendered euthyroid with thyroxine replacement. hTg-fed monocyte/macrophages induced a 4-fold increase in anti-hTg PFC in selected patients with autoimmune thyroiditis examined under similar conditions (mean 36 ± 3 PFC per 106 T-B cells). These data indicated that antigen processing by monocyte/macrophages was normal in patients with autoimmune thyroid disease.
Modified Distribution of Epidermal Glycoproteins in the Nude Mouse
Authors: Brysk Miriam M. , Miller Joanne , Chen Shu-Jen , Rajaraman Srinivasan
Published: October 2008

The nude mouse is an athymic mutant whose immunological deficiency has been exploited for transplantation of normal and diseased xenogeneic tissue. Histologically, its skin has no unusual features apart from the absence of hair. We report here a biochemical study of its epidermis, with comparison to the hairless mouse (which is devoid of hair but otherwise functionally normal). The epidermal glycoproteins were probed with the lectin, concanavalin A (Con A). Fluorescein isothyocyanate (FΙTC)-Con A overlays of cryostat skin sections gave a similar fluorescent pattern for both mouse strains: all the viable epidermal cell layers were labeled but not the stratum corneum. In contrast, when different populations of keratinocytes that were separated on Percoll gradients were analyzed by gel electrophoresis, and the gels then overlaid with iodinated Con A, all the epidermal layers, including the stratum corneum, were labeled. For all the epidermal cell layers there are substantial differences between the two mouse strains. We observe changes in the glycoprotein distribution with the stage of differentiation. Comparison with our earlier data for human epidermis indicates that the discrepancies between the nude mouse and the hairless mouse are much greater than those between the latter and man. The most striking difference is the absence in the stratum corneum of the nude mouse of a 40 K glycoprotein which is the dominant feature for the hairless mouse and for man. The gel patterns point to functional discrepancies in the epidermis of the nude mouse, particularly in the stratum corneum, not evident histologically or with FITC-Con A.
Mouse Peritoneal Macrophages Produce CFU-gm and CFU-meg Growth-Promoting Factors
Authors: Sugimoto Masakuni , Wakabayashi Yoshihisa , Hirose Shun-ichi , Murphy, Jr Martin J.
Published: October 2008

The effect of peritoneal macrophage-conditioned medium (macrophage CM) from lipopolysaccharide-injected mice on granulocyte-macrophage colony (CFU-gm) and megakaryocyte colony (CFU-meg) formation was examined using a plasma clot culture system. Macrophage CM stimulated mouse bone marrow cells to form CFU-gm and CFU-meg in the presence of pokeweed mitogen-stimulated spleen cell-conditioned medium, but it had no effect on colony formation in the absence of exogenous colony-stimulating factors (CSF). CFU-gm and CFU-meg colony formation was enhanced by low concentrations of exogenous GM-CSF or Meg-CSF alone. These data demonstrate that macrophage CM contains an activity that sensitized CFU-gm and CFU-meg to exogenous CSF.
Growth Factor Effects on Small Cell Lung Cancer Cells Using a Colorimetric Assay: Can a Transferrin-Like Factor Mediate Autocrine Growth?
Authors: Nakanishi Yoichi , Cuttitta Frank , Kasprzyk Philip G. , Avis Ingalill , Steinberg Seth M. , Gazdar Adi F. , Mulshine James L.
Published: October 2008

A semiautomated colorimetric assay (MTT assay), based on the ability of live cells to reduce a tetrazolium-based compound, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), to a purplish colored formazan product that can be measured spectrophotometrically, has recently been adapted for use in drug sensitivity analysis of cultured human tumor cell lines. We report the application of this assay for the evaluation of the growth factor requirements of human small cell lung cancer (SCLC) cell lines. Specifically, the growth stimulation of each constituent of a previously reported serum-free defined medium system for SCLC including various concentrations of hydrocortisone, insulin, transferrin, 17β-estradiol, and selenium (HITES) was evaluated. The optimal concentrations for insulin, transferrin, and selenium derived in the previously reported experiments with direct counting of viable cells were similar to optimal concentrations determined for the growth of three SCLC cell lines (NCI-H82, NCI-N417, NCI-H526) using the MTT assay. In contrast to the previous report, the growth-stimulating effects of hydrocortisone and 17β-estradiol were negligible. Using the MTT we have shown that a SCLC cell line, NCI-H345 (which has been previously reported to produce a transferrin-like molecule), was growth-inhibited by an anti-transferrin receptor antibody, when grown in transferrin-free media. The conditioned media from this cell line is stimulatory to other transferrin-sensitive cell lines, suggesting the possibility of an autocrine role for this transferrin-like molecule at least in that cell line. With carefully defined conditions for a given cell line in which cell density and other parameters are within a range of constant MTT metabolism, the assay is well suited for precise analysis of growth factor effects.
Cultured Liver Epithelial Cells Can Incorporate Monoclonal Antibodies Directed against Intracellular Cytokeratin Structures without Microinjection
Authors: Swierenga Sabine H.H.
Published: October 2008

T51B rat liver cells in the exponential phase of growth were arrested in late G1 by medium calcium deprivation, exposed to mouse monoclonal anti-cytokeratin (Mr = 55,000) by addition of the antibody to the medium, induced to enter the S phase by readdition of 1.5 mM calcium, and incubated for a further 18 h. After fixing and staining with FITC-conjugated anti-mouse IgG, the cells exhibited an intact intracellular cytokeratin-staining pattern. No effect of antibody was observed on entry of cells into the S phase. These results show that rat liver-derived epithelial cells can actively take up macromolecules like cytokeratin antibodies without microinjection.
Changes in the Polyamine and Nucleotide Metabolism of P388 Leukemia Cells Treated with DL-α-Difluoromethylornithine in Culture
Authors: Kremmer T. , Pályi I. , Holczinger L. , Lörincz I. , Boldizsár M. , Paulik E. , Pokorny É.
Published: October 2008

Influence of DL-α-difiuoromethylornithine (DFMO) treatment on the growth kinetics, labelling index, extra- and intracellular polyamine and nucleotide concentrations was monitored in cultured P388 leukemia cells. A substantial decrease of cell proliferation was observed when the cells were continuously treated with 1–5 mM DFMO. Depletion of cellular polyamines, mostly of putrescine and spermidine, was seen with a concomitant but delayed increase of spermidine and spermine levels in the culture medium. Changes of DNA content and of labelling index of untreated and treated cells seem to indicate that DFMO arrested cells in G1/S transition. The results presented here provide additional in vitro evidence on the characteristic changes in the metabolic imbalance of ornithine in tumor cells induced by DFMO via inhibition of ornithine decarboxylase and ornithine carbamoyl transferase activities.
Deficiency of a 42-Kilodalton Protein in Tumor-Derived Fibroblastic Cells in Neurofibromatosis
Authors: Maruyama E. , Hayashi A. , Arima M.
Published: October 2008

Cell proteins obtained from cultured normal appearing skin and neurofibromas of neurofibromatosis patients, and normal skin of normal donors were compared by SDS-PAGE and isoelectric focusing analysis. Essentially, identical protein patterns were obtained for the pellet fractions of all the strains. The lysate fraction binding patterns were also similar to each other, but a deficiency of a 42-kilodalton protein with pI 4.3 was observed in the four tumor-derived cell strains examined. These results raise the possibility that tumor-derived fibroblastic cells are of the same cell origin as skin fibroblasts, and that the deficiency of a 42-kilodalton protein could be related to the tumorigenicity in neurofibromatosis.
Studies on Thymocyte Subpopulations in Guinea Pigs
Authors: Sandberg G. , Stenvinkel C. , Kölare S.
Published: October 2008

Guinea pig thymocytes were incubated with normal rabbit serum, which resulted in the death of a great majority of the cells. The remaining rabbit serum-resistant cells, representing less than 10% of the thymocytes, contained euchromatic DNA and were of intermediate size and low density. Functional tests indicated that they were enriched in immunologically mature cells, which responded to the mitogenic lectins phytohemagglutinin and concanavalin A, and were depleted of immature, spontaneously proliferating cells and in cells responding to the thymocyte growth peptide. The described procedure for enrichment of immunologically mature thymus cells in guinea pigs may become useful since glucocorticoid treatment, used in mice for enrichment of mature thymocytes, cannot be used for this purpose in guinea pigs.
The Role of Antibodies in Natural and Acquired Resistance of Mice to Vesicular Stomatitis Virus
Authors: Gobet R. , Cerny A. , Rüedi E. , Hengartner H. , Zinkernagel R.M.
Published: October 2008

Mice infected with live vesicular stomatitis virus (VSV) produced primary antibody responses more efficiently (with doses > 102 pfu) than those injected with UV-inactivated VSV ( > 106 pfu) or purified VSV glycoprotein G (equivalent to 107 pfu) by producing neutralizing antibodies. Very low doses of live VSV ( < 102 pfu) failed to prime mice. Normal mouse serum had the capacity to inactivate VSV by heat-labile and immunoglobulin-mediated mechanisms in vitro independently of specifically induced antibodies. Studies using B cell-depleted agammaglobulineamic mice showed that their serum lacked VSV-neutralizing capacity in vitro and that naturally resistant mice became susceptible to VSV-induced paralytic disease which could be prevented by adoptive transfer of immune but also of normal serum.
Expression of T Cell Receptor γ-Chain in Murine Cytotoxic T Cells: Analysis of a Highly Transcribed Nonrearranged Gene Cluster
Authors: Schulz Manfred , Lienhard Gabi , Rupp Fabio , Zinkernagel Rolf M. , Hengartner Hans
Published: October 2008

The murine T cell receptor γ-gene family is organized in 4 gene clusters which encode variable, joining and constant gene segments. Because the function of the γ-gene products is still unclear, there is great interest in functional rearrangements of these genes. We searched for new variable γ-gene segments in the γ4-gene cluster. The mapping and sequencing of a supposed new variable segment of a functionally spliced transcript from a cDNA library of the cytotoxic T cell clone 3F9 was attempted. The results show that the supposed variable segment is a transcribed noncoding germline sequence 5’ adjacent to the γ4-joining segment. With the γ4-constant sequence as a probe, strong signals could be shown in Northern analysis of the original T cell clone 3F9 and in other virus-specific T cell clones. Since all the T cell clones investigated did not show any rearrangement in the γ4-gene cluster, the observed high transcriptional activity in this region could be involved functionally or as an intermediate in T cell receptor rearrangement.
Cytologic Presentation of Malignant Mesothelioma in Pleural Effusions
Authors: Pedio Gino , Landolt-Weber Ursula
Published: October 2008

1,601 pleural effusions were found to be malignant between 1976 and 1987. Among these were 26 (1.6% of the malignant effusions) mesothelioma. Only 2 cases showed pronounced cytologic features that made a definite diagnosis possible on cytologic criteria alone. In 20 cases diagnosis of mesothelioma was strongly suggested by the patients’ history and cytology of the effusion was compatible with mesothelioma. In the other 4 cases special examinations (histo- and immunohistochemistry, electron microscopy) led to the final diagnosis. The cytologic features of mesothelioma and other examination techniques, needed to resolve the differential diagnosis of mesothelioma versus other neoplasm in pleural effusions, are discussed.
The Vertebral Column of Diabetic Sand Rats (Psammomys obesus)
Authors: Silberberg Ruth
Published: October 2008

Intervertebral discs and vertebral spongiosa of diabetic and nondiabetic sand rats were investigated histologically and morphometrically. In diabetic animals, degeneration of the intervertebral discs was accelerated. Morphometrically, there was a consistent trend in the diabetics toward an attenuated bone turnover, which was, however, statistically significant only in regard to the length of the trabecular surface covered by osteoblasts. In the vertebral column of the sand rat, the tendency to osteoporosis – age-linked or diabetes-related – is reduced apparently due to the influence of local forces which promote osteogenesis.
Growth and Metastasis of Lewis Lung Carcinoma in the Footpad of Mice
Authors: Kitinya James N. , Kinjo Mitsuru , Tanaka Kenzo
Published: October 2008

When Lewis lung carcinoma cells were transplanted into the foopad of mice, exponential growth of tumour was seen from the 4th day. Intravasation of tumour cells was first seen at this time, then increased steadily up to the 10th day. Tumour cells entered the circulation by destructive degeneration of the vascular endothelium and also by transcellular passage, mainly through capillaries, venules and thin-walled tumour vascular channels. Multiple sites of entry of tumour cells into the circulation were frequently found. The rate of intravasation of tumour cells into the circulation was the same at the proximal, middle and distal portions of the tumour.
Megakaryocyte Colony-Stimulating Factor and Burst-Promoting Activity in LPS-Treated Mouse Spleen Cell-Conditioned Medium
Authors: Sugimoto Masakuni , Wakabayashi Yoshihisa , Hirose Shun-ichi , Murphy, Jr Martin J.
Published: October 2008

In order to clarify the mechanism(s) of increased splenic hematopoiesis noted in lipopolysaccharide (LPS)-injected mice, the effects of spleen cell-conditioned medium (SPCM) on megakaryocyte colony (CFU-meg) formation and early erythroid (BFU-e) differentiation were investigated. After spleen cells from LPS-injected mice were incubated for 3 days, the SPCM was assayed for megakaryocyte colony-stimulating factor (Meg-CSF) in CFU-meg assay and for burst-promoting activity (BPA) and erythropoietin (Epo) in erythroid colony assays (i.e., CFU-e, BFU-e). Colony formation of CFU-meg and BFU-e peaked with the addition of 30 and 10–15% SPCM, respectively. Spleen cells from LPS-injected mice produced Meg-CSF and BPA when compared with controls. However, conditioned medium from spleen cells depleted of phagocytic cells had low Meg-CSF and BPA. SPCM did not contain detectable quantities of Epo. It appears likely that local splenic production of Meg-CSF and BPA may affect proliferation of CFU-meg and erythroid progenitor cells in the spleen.