bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transcripts encoding protein kinase C‐α, ‐ϵ, ‐ζ, and ‐ζ are expressed in basal and differentiating mouse keratinocytes in vitro and exhibit quantitative changes in neoplastic cells
Authors: Dlugosz Andrzej A. , Mischak Harald , Mushinski J. Frederic , Yuspa Stuart H. 
Published: February 2007

AbstractThe protein kinase C (PKC) family of phospholipid‐dependent serine‐threonine kinases has been implicated in keratinocyte differentiation and neoplastic transformation. To determine if Ca2+‐mediated keratinocyte differentiation is associated with changes in PKC isozyme gene expression, RNA was isolated from primary mouse keratinocytes grown in medium with 0.05, 0.12, or 1.4 mM Ca2+. Based on northern blot analysis, primary keratinocytes expressed mRNA encoding PKC‐α, ‐ϵ, ‐ζ, and ‐neta but not PKC‐p or ‐y. Relatively little change was detected in the level of these transcripts in cells induced to differentiate by exposure to elevated extracellular Ca2+. Interestingly, the PKC‐ζ transcripts detected in RNA isolated from keratinocytes were approximately 200 nucleotides longer than those from mouse brain, suggesting the existence of an alternative form of this isozyme.An early change in benign neoplastic transformation of keratinocytes is the inability to differentiate in response to Ca2+ or the PKC activator 12‐O‐tetradecanoylphorbol‐13‐acetate, which is consistent with altered PKC function in these cells. The PKC isozyme mRNA profile was examined in two benign neoplastic keratinocyte cell lines, 308 and SP‐1, which contain an activating mutation of the c‐Ha‐ras gene. Like normal keratinocytes, 308 and SP‐1 cells expressed mRNA encoding PKC‐α, ‐ϵ, ‐ζ, and ‐η However, the abundance of PKC‐η transcripts in both cell lines was reduced by 74–89% when compared with normal keratinocytes at similar Ca2+ levels. In addition, SP‐1 but not 308 cells exhibited a sevenfold increase in PKC‐η mRNA when cultured in medium with 1.4 mM Ca2+. To address whether these changes were related to the presence of an activated ras gene, RNA was isolated from primary keratinocytes transduced to a benign neoplastic phenotype with the v‐Ha‐ras oncogene. As with normal, 308, and SP‐1 cells, v‐Ha‐ras keratinocytes expressed mRNA encoding PKC‐α, ‐ϵ, ‐ζ, and ‐neta. The level of PKC‐η transcripts was similar in normal and v‐Ha‐ras keratinocytes, indicating that reduction of this mRNA in both 308 and SP‐1 cells was not a direct result of ras activation. As in SP‐1 cells, PKC‐η in v‐Ha‐ras keratinocytes was responsive to extracellular Ca2+, with a four‐fold increase in transcript abundance in 0.12 mM Ca2+ medium relative to 0.05 mM Ca2+ medium. These findings illustrate the complexity of the PKC‐signal transduction pathway in cultured mouse keratinocytes and indicate that the level of certain PKC isozyme transcripts is modulated during the neoplastic process. © 1992 wiley‐Liss, Inc.
Homologous and heterologous Gap‐junctional intercellular communication in v‐raf‐, v‐myc‐, and v‐raf/v‐myc‐transduced rat liver epithelial cell lines
Authors: Kalimi Ghulam H. , Hampton Lori L. , Trosko James E. , Thorgeirsson Snorri S. , Huggett Anthony C. 
Published: February 2007

AbstractWe examined gap‐junctional intercellular communication (GJIC) in a series of normal and v‐raf‐, v‐myc‐, and v‐raf/v‐myc‐transduced rat liver epithelial (RLE) cell lines using the scrape loading‐dye transfer and fluorescence‐recovery‐after‐photobleaching (FRAP) assays. Whereas the normal RLE cell line, the control helper virus‐transduced cell line, and the v‐myc‐transduced cell line all showed excellent GJIC, the v‐raf‐transduced cell lines displayed decreasing levels of GJIC associated with their increasing tumorigenicity. The v‐raf/v‐myc‐transformed cell lines showed the lowest levels of GJIC and were also the most tumorigenic. Heterologous GJIC of these oncogenetransduced cell lines was also compared with that in the normal RLE cells. A modified FRAP assay, using fluorescent‐microbead labeling to identify the oncogene‐transduced cell from surrounding normal cells, was used to quantify the heterologous GJIC. The v‐raf/v‐myc‐transformed RLE cells had no heterologous communication with the normal RLE cells, whereas v‐raf‐ and v‐myc‐transduced cell lines maintained heterologous GJIC. Northern analysis showed that connexin 43 was the only gap‐junction protein message expressed in these cell lines; connexin 32 and connexin 26 were not expressed. The levels of connexin 43 mRNA expression were relatively unchanged in all cell lines, suggesting that the reduction in GJIC was primarily at the posttranslational level. These findings suggest that reduction of homologous GJIC in v‐raf‐ and v‐raf/v‐myc‐transformed RLE cells is linked to their tumorigenic potential. Furthermore, the loss of heterologous GJIC, which we observed only in the v‐raf/v‐myc‐transformed cells, might release such cells from the growth‐regulating effects of surrounding normal cells, possibly contributing to their enhanced tumorigenic potential. © 1992 Wiley‐Liss, Inc.
Differential induction of 12‐O‐tetradecanoylphorbol‐13‐acetate sequence gene expression in murine melanocytes and melanoma cells
Authors: Brooks Gavin , Goss Martin W. , Hart Ian R. 
Published: February 2007

AbstractWe previously showed that growth of the nontumorigenic, immortal murine melanocyte line Mel‐ab correlates with the depletion of protein kinase C (PKC), whereas quiescence is associated with elevated levels of this enzyme (Brooks G, et al., Cancer Res 51: 3281–3288, 1991). Here we report responses that occur in these cells downstream of PKC activation or downregulation. We examined induction of 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐inducible sequence (TIS) gene expression in Mel‐ab melanocytes and in their transformed counterparts, B16 melanoma cells. Exposure of quiescent Mel‐ab cells to the PKC‐activating phorbol esters TPA or sapintoxin A at 81 nM for 2 h increased levels of mRNA for six of seven TIS genes examined (twofold to 80‐fold increase in steady‐state RNA levels for TIS 1, 7, 8, 11, 21, and 28 (c‐fos); TIS 10 expression was not affected). No induction of 115 gene expression was observed either in growing Mel‐ab cells maintained in 324 nM phorbol 12,13‐dibutyrate or in B16 cells previously unexposed to phorbol esters, in which normal PKC levels were endogenously depressed. The cAMP‐elevating agents choleratoxin (10 nM) and dibutyryl cyclic AMP (2.5 mM) increased levels of TIS mRNA (with the exception of TIS 10) in both proliferating Mel‐ab and B16 cells, suggesting that downregulation of the PKC pathway is specific and not a consequence of a general inhibition of all signalling pathways. © 1992 Wiley‐List. Inc.
Additive effects of c‐erbB‐2, c‐Ha‐ras, and transforming growth factor‐α genes on in vitro transformation of human mammary epithelial cells
Authors: Ciardiello Fortunato , Gottardis Marco , Basolo Fulvio , Pepe Stefano , Normanno Nicola , Dickson Robert B. , Raffaele Bianco A. , Salomon David S. 
Published: February 2007

AbstractMCF‐10A cells are a spontaneously immortalized untransformed human mammary epithelial cell line. We have previously shown that overexpression of a human point‐mutated c‐Ha‐ras proto‐oncogene, the rat c‐neu (c‐erbB‐2) proto‐oncogene, or the human transforming growth factor‐α (TGF‐α) gene in MCF‐10A cells leads to in vitro transformation of such cells. To ascertain whether the introduction of two of these genes into MCF‐10A human mammary epithelial cells induces a completely tumorigenic phenotype, we infected MCF‐10A Ha‐ras and MCF‐10A TGF‐α cells with a recombinant retroviral vector containing the human c‐erbB‐2 proto‐oncogene and the hygromycin‐resistance gene. Ten MCF‐10A TGF‐α/c‐ethB‐2 (MCF‐10A TE) and 10 MCF‐10A Ha‐ras/c‐erbB‐2 (MCF‐10A HE) hygromycin‐resistant clones were randomly selected and expanded into cell lines. MCF‐10A TE and MCF‐10A HE clones expressed a 10‐fold to 40‐fold increase in p185 erbB‐2 protein levels compared with parental uninfected cells. These cells exhibited a fourfold increase in their growth rate in serum‐free medium and showed a strongly reduced mitogenic response to exogenous epidermal growth factor or TGF‐α compared with MCF‐10A cells. Moreover, both MCF‐10A TE and MCF‐10A HE clones exhibited a fivefold to 20‐fold higher cloning efficiency in soft agar than MCF‐10A Ha‐ras, MCF‐10A c‐erbB‐2, or MCF‐10A TGF‐α clones. However, neither MCF‐10A TE nor MCF‐10A HE cells were able to grow as tumors in vivo when they were injected into nude mice. These results suggest that c‐Ha‐ras, c‐erbB‐2, and TGF‐α genes have an additive effect on the in vitro transformation of an immortalized human mammary epithelial cell line, but that additional genetic changes such as activation of other proto‐oncogenes or inactivation of a tumor suppressor gene may be necessary to elicit a fully tumorigenic phenotype. © 1992 Wiley‐Liss, Inc.
Role of mutations at codon 61 of the c‐Ha‐ras gene during diethylnitrosamine‐induced hepatocarcinogenesis in C3H/He mice
Authors: Bauer‐Hofmann Richard , Klimek Fritz , Buchmann Albrecht , Müller Oliver , Bannasch Peter , Schwarz Michael 
Published: February 2007

AbstractLiver tumors of certain strains of mice frequently contain mutations at codon 61 of the c‐Ha‐ras gene. In our study, we investigated the significance of these mutations in the carcinogenic process. Male C3H/He mice received a single injection of diethylnitrosamine (DEN) on day 15 after birth, and groups of animals were killed at various time intervals between 11 and 52 wk after treatment. At the earlier time points (11–29 wk), we analyzed microdissected tissue from precancerous glucose‐6‐phosphatase‐deficient liver lesions larger than approximately 200 p.m in diameter, for the presence and pattern of c‐Ha‐ras codon 61 mutations. In parallel, the growth characteristics of these liver lesions were studied by pulse labeling with [3H]thymidine and by determining the size distribution of the lesions. At the later time points (42–52 wk after DEN treatment), liver tumors were dissected and also analyzed for the presence of c‐Ha‐ras mutations. We found mutations to be already present in some of the enzyme‐altered liver lesions at weeks 11–29, suggesting that the mutations occurred early in the carcinogenic process. Whereas about 10% of the precancerous focal liver lesions showed mutations in the c‐Ha‐ras gene, the mutation frequency was increased to about 50% in the later‐appearing hepatocellular adenomas and carcinomas, suggesting that c‐Ha‐ras codon 61 mutations may provide a selective advantage to the mutated cell clones. © 1992 Wiley‐Liss, Inc.
p53 mutations are absent from carcinogen‐induced mouse liver tumors but occur in cell lines established from these tumors
Authors: Kress Stefan , König Jorg , Schweizer Jürgen , Löhrke Heinz , Bauer‐Hofmann Richard , Schwarz Michael 
Published: February 2007

AbstractMutations in the p53 gene are frequent genetic alterations in human hepatocellular carcinomas We have examined, by single‐strand conformation polymorphism analysis of polymerase chain reaction products, a total of 93 carcinogen‐induced liver tumors from mice of three different strains (C3H/He, C57BL/6J, and B6C3F1) for the presence of p53 aberrations. Hepatoma lines, established from 12 liver tumors, were also included in the analysis. While structural aberrations of the p53 gene were not detected in any of the primary mouse liver tumors analyzed, single‐base substitutions occurred at different locations within the p53 gene in three of the cell lines during in vitro propagation. One hepatoma line carried two point mutations on separate alleles. All four mutations were either G:C→C:G or C:G→G:C transversions. Mutations at codon 61 of the c‐Ha‐ras gene, which were frequent in primary liver tumors from C3H/He and B6C3F1 mice, were not detected in the hepatoma lines. Our data indicate (i) that c‐Ha‐ras but not p53 mutations play an important role during the early stages of mouse hepatocarcinogenesis and (ii) that p53 mutations confer a selective growth advantage to the mutated hepatoma cells in vitro. © 1992 Wiley‐Liss, Inc.
Role of activator protein‐1 and methylation function in 12‐o‐tetradecanoylphorbol‐13‐acetate—mediated inhibition of differentiation of friend erythroleukemia cells
Authors: Bertrand Olivier , Giroldi Laurence , Nakazawa Hisayoshi , Sergeant Alain , Yamasaki Hiroshi 
Published: February 2007

AbstractFriend erythroleukemia cells (FELCs) differentiate after hexamethylene‐bis‐acetamide treatment. This differentiation is characterized by an increase in β‐globin gene expression that is followed by appearance of the hemoglobin. Phorbol‐ester tumor promoters, such as 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA), inhibit differentiation of TPA‐sensitive cells but not TPA‐resistant cells. We have shown that the increase in β‐globin expression is inhibited by TPA in a TPA‐sensitive clone but not in a TPA‐resistant clone. To study the molecular mechanisms of regulation of gene expression by TPA, we examined the possible involvement of gene methylation and the TPA‐responsive element (TRE). Both clones showed similar patterns of methylation around the β‐globin gene. Moreover, TPA‐induced TRE binding and TRE enhancer activity were similar in both variants. These results suggest that the TPA inhibition of induced differentiation may not be explained by regulation of the methylation state. The activator protein‐1 also does not play a crucial role in the sensitivity of FELCs to TPA. © 1992 Wiley‐Liss, Inc.
12‐O‐tetradecanoylphorbol‐13‐acetate—induced levels of ap‐1 proteins: a 46‐kda protein immunoprecipitated by anti‐fra‐1 and induced in promotion‐resistant but not promotion‐sensitive JB6 cells
Authors: Bernstein Lori R. , Bravo Rodrigo , Colburn Nancy H. 
Published: February 2007

AbstractNeoplastic transformation and transcriptional activation by activator protein‐1 (AP‐1) complex are stimulated by tumor‐promoting agents in promotion‐sensitive (P+) but not promotion‐resistant (P‐) mouse epidermal JB6 cells in culture. This implicates AP‐1 as a specific regulator of signal transduction pathways in the promotion phase of neoplastic transformation. We therefore hypothesized that the defective P‐ responsiveness may be due to limiting levels of AP‐1 protein components in those cells. In this investigation, steady‐state levels of AP‐1 protein components were measured by immunoprecipitating proteins from 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐treated P+ and P‐ cells to discern what may limit the AP‐1 response. Whereas the AP‐1 proteins junB, junD, and fosB did not show differential basal or TPA‐inducible levels in P+ and P‐ cells, a 46‐kDa species precipitated by anti‐fra‐1 antibody was TPA‐inducible in P‐ cells but not in P+ cells, and c‐jun protein was present at higher levels in TPA‐treated and untreated P+ cells than in P‐ cells. These data raise the possibility that the 46‐kDa fra‐1‐related protein may be a negative modulator of AP‐1 activity and suggest that elevated levels of this 46‐kDa species and limiting levels of c‐jun may significantly impair AP‐1 function or transformation response in P‐ cells or both. © 1992 Wiley‐Liss, Inc.
Suppression of radiation‐induced neoplastic transformation by overexpression of mitochondrial superoxide dismutase
Authors: St. Clair Daret K. , Steven Wan X. , Oberley Terry D. , Muse Kenneth E. , St. Clair William H. 
Published: February 2007

AbstractManganese superoxide dismutase (MnSOD) scavenges toxic superoxide radicals produced in the mitochondria. Transfection of the human MnSOD gene into mouse C3H 10T1/2 cells resulted in production of active MnSOD, which was properly transported into mitochondria. Overexpression of MnSOD protected cells from radiation‐, but not chemically‐induced neoplastic transformation. This finding demonstrates that oxidative stress that occurs in the mitochondria plays an important role in the development of neoplastic transformation. © 1992 Wiley‐Liss, Inc.
Keratin 14 protein in cultured nonparenchymal rat hepatic epithelial cells: Characterization of keratin 14 and keratin 19 as antigens for the commonly used mouse monoclonal antibody OV‐6
Authors: Bisgaard Hanne Cathrine , Parmelee David C. , Dunsford Harold A. , Sechi Salvatore , Thorgeirsson Snorri S. 
Published: February 2007

AbstractWe have recently reported that cell lines of nonparenchymal origin isolated from rat liver and pancreas, which have been suggested to be the progeny of a facultative stem cell compartment in vivo, express an unusual combination of keratins (K). These cell lines express K8 and K14 but not K18 and K5, their normal partners in filament formation (Bisgaard HC, Thorgeirsson SS, J Cell Physiol 147:333–343, 1991). However, upon spontaneous transformation and differentiation toward a hepatoblastlike progeny, K14 expression is abrogated and replaced by expression of K18 (Wirth et al., Electrophoresis 13:305–332, 1992). In the study presented here, we confirmed by protein sequence analysis that K14 was a major component of the intermediate filaments in a nonparenchymal cell line of hepatic origin. Immunocytochemical analysis of the cells in monolayer demonstrated that K8 as well as K14 were incorporated in the cellular cytoskeleton. Further analysis by immunoprecipitation showed that filament complexes were formed between K8 and K14 as atypical partners. Thus, we concluded that in some nonparenchymal cell lines isolated from rat liver, K8 and K14 form a major intermediate filament network. Finally, we showed that an antibody widely used in studies of the cell lineages of hepatic and pancreatic tissues and their neoplasms, the mouse monoclonal antibody OV‐6, recognizes a common epitope in K14 and K19.
Induction of epidermal hyperplasia, hyperkeratosis, and papillomas in transgenic mice by a targeted v‐Ha‐ras oncogene
Authors: Greenhalgh David A. , Rothnagel Joseph A. , Quintanilla Maria I. , Orengo Christine C. , Gagne Todd A. , Bundman Donnie S. , Longley Mary A. , Roop Dennis R. 
Published: February 2007

AbstractThe regulatory elements of the human keratin K1 gene have been used to target expression of the v‐Ha‐ras oncogene exclusively in the epidermis of transgenic mice. We developed 12 transgenic mouse lines that express the HK1. ras transgene, producing epidermal hyperplasia in neonates and hyperkeratosis in juveniles. Eventually this skin phenotype diminished but with time adult animals developed papillomas that could persist or regress. The rate and frequency of tumorigenesis appeared to be limited, which suggests that v‐Ha‐ras requires a second or even third event to elicit and maintain a benign phenotype in transgenic mice. Since in certain transgenic lines papillomas appeared at wound sites, it appears that the promotion stimulus from wounding may be a second event. We envision that such transgenic mice that express v‐Ha‐ras in the epidermis will become a powerful model for assessing how environmental and molecular factors affect the process of multistage skin carcinogenesis in vivo, as well as a model for evaluating novel therapeutic protocols.
Correlation between lack of bone Gla protein mRNA expression in rat transplantable osteosarcomas and expression of both c‐fos and c‐jun proto‐oncogenes
Authors: Honoki Kanya , Dohi Yoshiko , Tabata Shiro , Mii Yoshio , Miyauchi Yoshizumi , Tsutsumi Masahiro , Tsujiuchi Toshifumi , Morishita Toru , Miura Syuichi , Moriyama Tadashige , Tamai Susumu , Konishi Yoichi 
Published: February 2007

AbstractAlkaline phosphatase (AP) activity and expression of bone Gla protein (BGP), c‐fos, and c‐jun were compared in two transplantable osteosarcomas with high potentials for metastasis to the lung. The original spontaneous osteosarcoma (SOS) gradually became histologically undifferentiated, losing its osteogenic activity during serial transfer, whereas the chemical (4‐hydroxyaminoquinoline 1‐oxide)—induced osteosarcoma (COS) retained osteogenesis. The two osteosarcomas showed similar doubling times and levels of lung metastasis, and strong AP activity was detected on the cell membranes of both. Northern blot analysis revealed that lack of BGP mRNA expression was associated with expression of both c‐fos and c‐jun proto‐oncogenes in SOS. In contrast, neither c‐fos nor c‐jun mRNAs were detected but BGP mRNA was expressed in the case of COS. These results suggest that the c‐fos and c‐jun genes may suppress the expression of BGP mRNA relevant to differentiation and osteoid formation in rat osteosarcomas. However, this does not appear to be directly related to proliferative or metastatic biological behavior.
Induction of superoxide by 12‐O‐tetradecanoylphorbol‐13‐acetate and thapsigargin, a non‐phorbol‐ester—type tumor promoter, in peritoneal macrophages elicited from SENCAR and B6C3F1 mice: A permissive role for the arachidonic acid cascade in signal transduction
Authors: Yoon Hyunah L. , Marcus Craig B. , Pfeifer Richard W. 
Published: February 2007

AbstractLocal production of reactive oxygen intermediates, e.g., superoxide anion, by tumor promoter—stimulated inflammatory macrophages (MPs) may contribute significantly to tumor development in classical models of two‐stage chemical‐induced carcinogenesis in murine skin. In the studies reported herein, peritoneal MPs elicited from phorbol‐ester—sensitive SENCAR mice demonstrated a time‐ and dose‐dependent release of superoxide anion (4–6 nmol/106 cells) when stimulated by 200 nM 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) in vitro; MP superoxide response was significantly inhibited (50–70%) by preincubation with 40 μM 1‐(5‐isoquinolinyl‐sulfonyl)‐2‐methylpiperazine (H‐7), a protein‐kinase inhibitor. Alternatively, TPA‐stimulated MPs derived from relatively resistant B6C3F1 mice generated negligible superoxide under the same conditions. A similar strain‐dependent induction of superoxide was observed when MPs were stimulated with thapsigargin (TG), a tumor promoter previously shown to act independently of protein kinase C (PKC). TG‐stimulated SENCAR MPs released a significant amount of superoxide (2–3 nmol/106 cells) that was not inhibited by H‐7; MPs from B6C3F1 mice demonstrated negligible stimulation by TG. Preincubation of SENCAR MPs with 100 μM dibromoacetophenone, an inhibitor of phospholipase A2, completely suppressed the superoxide induced by TPA and TG stimulation. Like TPA, 50 μM 1‐oleoyl‐2‐acetylglycerol, a diacylglycerol analogue and PKC activator, also induced a significant amount of superoxide from SENCAR MPs only. In parallel with the superoxide findings, TPA and TG stimulated significantly greater [3H]arachidonic acid release from prelabeled SENCAR MPs (a 32% and 48% increase, respectively, over unstimulated controls) relative to MPs from B6C3F1 mice. Two‐dimensional gel‐electrophoretic analysis indicated that TPA‐induced phosphorylation of a 47‐kDa protein (a presumed substrate for PKC previously linked to NADPH oxidase activation in guinea pig and human polymorphonuclear leukocytes) occurred in MPs from both SENCAR and B6C3F1 mice. Therefore, arachidonic acid production may be a common biochemical pathway by which phorbol‐ester— and non‐phorbol‐ester—type tumor promoters activate MPs in SENCAR mice; such a response may be “permissive” for additive (or synergistic) interactions with PKC‐driven signal pathways.