bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sertoli cells in culture secrete paracrine factor(s) that inhibit peritubular myoid cell proliferation: Identification of heparinoids as likely candidates
Authors: Tung Pierre S. , Fritz Irving B. 
Published: February 2005

AbstractConditioned medium from Sertoli cells, prepared from testes of 20‐day‐old rats, contains component(s) that inhibit the incorporation of [3H]‐thymidine into DNA of peritubular myoid cells (PMC) and inhibit the proliferation of PMC. These components are trypsin‐resistant, heat‐stable compounds having a molecular weight less than 30,000. The active inhibitory components in Sertoli cell conditioned medium are inactivated by treatment with heparinase, but not by treatment with hyaluronidase or chondroitin sulfate lyases. Addition of heparin or heparan sulfate results in inhibition of DNA synthesis by PMC in a dose‐dependent manner, whereas other glycosaminoglycans (GAGs) examined (hyaluronic acid, keratan sulfate, and chondroitin sulfate) have no detectable effects. Heparin and heparan sulfate are unique among GAGs tested in inhibiting the characteristic multilayer growth pattern of PMC following the attainment of confluence in serum‐rich medium. On the basis of these and other data presented, it is concluded that heparin and other heparin‐like GAGs synthesized by Sertoli cells are implicated in the modulation of growth of PMC in vitro during co‐culture. It is postulated that heparin may play a similar role in maintaining the quiescent peritubular myoid cell phenotype in vivo.
Increased glutathione levels in quiescent, serum‐stimulated NRK‐49F cells are associated not with a response to growth factors but with nutrient repletion
Authors: Kang Yu‐Jian , Duane Enger M. 
Published: February 2005

AbstractTreatment of quiescent cells with serum results concomitantly in an increase in cellular glutathione (GSH) content and growth stimulation. A possible association between the GSH increase and the growth response was examined by studying separately the effects of nutrients and growth factors on the levels of cellular GSH and proliferation of quiescent NRK‐49F cells. The addition of fresh medium with 10% calf serum was found to result in both a twofold increase in cellular GSH and growth stimulation (DNA synthesis and cell proliferation). 10% calf serum alone, without fresh medium, stimulated cell growth but failed to cause a comparable increase in cellular GSH. The addition of fresh medium without 10% serum, and of 0.5 mM cysteine and glutamate, resulted in both instances in a marked increase in cellular GSH, but failed to stimulate cell growth, EGF, in contrast, induced a complete mitogenic response but did not increase cellular GSH. Finally, pretreatment with L‐buthionine‐(S, R)‐sulfoximine (BSO), a specific inhibitor of GSH synthesis, decreased cellular GSH and inhibited EGF‐induced DNA synthesis, but these two responses do not, in their dose dependency, correlate. The results obtained thus show that the increase in cellular GSH that occurs in quiescent, serum‐stimulated NRK‐49F cells is a result of nutrient repletion rather than mitogenic stimulation, and increased GSH levels do not necessarily precede DNA synthesis and mitosis.
Pathway of glutamate oxidation and its regulation in the HuH13 line of human hepatoma cells
Authors: Matsuno Tetsuya 
Published: February 2005

AbstractWell‐coupled mitochondria were isolated from a HuH13 line of human hepatoma cells and human liver tissue. The liver mitochondria showed a feeble glutamine oxidation activity in contrast to the hepatoma mitochondria, whereas they utilized glutamate well for the oxidative phosphorylation. In the liver mitochondria, glutamate was cxidized via the routes of transamination and deamination. On the other hand, glutamate oxidation was initiated preferentially via transamination pathway in the tumor mitochondria. In the liver mitochondria, bicarbonate nearly at a physiological concentration inhibited oxygen uptake with glutamate as substrate. The interaction of bicarbonate with the pathway of glutamate oxidation occured primarily at the level of succinate dehydrogenase, due to competitive inhibition of the enzyme by the compound. In contrast to the liver mitochondria, glutamate oxidation was not affected by bicarbonate in the tumor mitochondria. These results indicate that the aberrations in the glutamate metabolism and its regulation observed in the hepatoma mitochondria may be favorable to the respiration utilizing glutamine and/or glutamate as an energy source.
Continuous activation of primitive hematopoietic cells in long‐term human marrow cultures containing irradiated tumor cells
Authors: Otsuka Teruhisa , Keith Humphries R. , Hogge Donna E. , Eaves Allen C. , Eaves Connie J. 
Published: February 2005

AbstractHuman hematopoietic cells can be maintained in vitro for many weeks in the absence of exogenously provided hematopoietic growth factors if an adequate stromal cell containing adherent layer is present. We have now extended the use of this type of long‐term culture (LTC) system to create a model of perturbed hematopoiesis in which human tumor cells that constitutively produce a variety of factors are co‐cultured together with normal human marrow cells. In the present study, we used the human bladder carcinoma cell line (5637) because these cells were known to produce not only a variety of factors active directly on hematopoietic cells but also factors that can stimulate hematopoietic growth factor production by human marrow stromal cells. Analysis of mRNA extracted from the adherent layer and measurement of growth factor bioactivity in the medium of established LTC of human marrow containing irradiated 5637 cells, showed increased levels of interleukin‐1 and ‐6, as well as granulocyte and granulocyte‐macrophage colony‐stimulating factor production by comparison to control cultures. As in normal cultures, high proliferative potential clonogenic hematopoietic cells were found almost exclusively in the adherent layer of these co‐cultures, but these primitive cells were maintained in a state of continuous turnover, in contrast to control cultures where the same cell types showed the expected oscillation between a quiescent and a proliferating state following each weekly change of the medium. A similar perturbation of primitive progenitor cycling was achieved by adding medium conditioned by 5637 cells twice a week to otherwise normal LTC. The presence of irradiated 5637 cells in the LTC or the addition of 5637 conditioned medium also resulted in modest (2‐ to 3‐fold) but sustained increases in the total hematopoietic progenitor population, as well as in the final output of terminally differentiated granulocytes and macrophages. These findings indicate that primitive hematopoietic cells in LTC can be kept in a state of continuous activation for many weeks by appropriate endogenous or exogenous hematopoietic growth factor provision and that this does not necessarily lead either to their rapid exhaustion or to a large amplification in output of mature progeny.
Reduction of TGF‐beta activity abrogates growth promoting tumor cell‐cell interactions in vivo
Authors: Theodorescu D. , Caltabiano M. , Greig R. , Rieman D. , Kerbel R. S. 
Published: February 2005

AbstractWe have shown in previous studies that metastatically‐competent variant subpopulations (B5, C1) derived from a non‐metastatic murine mammary adenocarcinoma (SP1) have a pronounced growth advantage over their non‐metastatic tumor cell counterparts in primary tumors. As a result, primary tumors can be progressively overgrown by cells having the competence to spread elsewhere in the body. This occurs despite any evidence to indicate an intrinsic in vivo growth rate advantage of the metastatic cells when grown as isolated populations. This suggested that cell‐cell interactions between metastatic and non‐metastatic tumor populations may be involved in the metastatic cell growth dominance process. Evidence was therefore sought for growth factors released by SP1 cells which could preferentially stimulate the B5 or C1 variants and thereby mediate this cell‐cell interaction process. We found that cocultures of SP1 and C1 or B5 cells with irradiated C1, B5, or SP1 “feeder” cells showed significant stimulation of C1 and B5 by SP1 “feeder” cells. Cell growth stimulation in response to EGF, TGF‐α, TGF‐β1, bFGF, PDGF, NGF, IGF‐1, or IGF‐2 demonstrated that only TGF‐β1 could duplicate this effect. A repeat of the coculture experiment in the presence of specific neutralizing anti‐TGF‐β antibodies was therefore undertaken and this was found to markedly reduce the stimulation of C1 or B5 cells by irradiated SP1 cells. Conditioned media from the SP1 and C1 cell lines was quantitated for TGF‐β activity and contained 4.5 ng/ml and 2.0 ng/ml, respectively. However, the majority of the TGF‐β released by SP1 cells was found to be spontaneously active, whereas 70% of the TGF‐β released by C1 cells was in its latent form. Scatchard analysis revealed approximately four times the number of TGF‐β receptors, of similar type and affinity, present on C1 as compared with SP1 cells. The in vitro results support the hypothesis that active TGF‐β released by SP1 cells may stimulate the proliferation of metastatic variant cells in a paracrine like fashion. In vivo evidence for this was obtained by showing that coinjection of irradiated SP1 cells could selectively stimulate tumor growth of viable C1 cells and this effect was markedly diminished by neurtralizing polyclonal anti‐TGF‐β antibodies. Taken together, the results suggest a novel role for TGF‐β in clonal evolution of malignant tumor growth and as a molecular mediator of tumor cell‐tumor cell interactions involved in facilitating tumor progression.
Regulation of interleukin‐6 expression in the lymphoma cell line OCI‐LY3
Authors: Yee C. S. , Messner H. A. , Minden M. D. 
Published: February 2005

AbstractPreviously we described a cell line OCI‐LY3 derived from a patient with non‐Hodgkin's lymphoma. The cell line produced interleukin‐6 (IL‐6) mRNA and protein and demonstrated an autocrine pattern of growth for IL‐6. Southern blot analysis of the IL‐6 gene did not reveal any rearrangement. To determine whether the production of IL‐6 by OCI‐LY3 was due to subtle changes in the promoter of IL‐6 or due to the expression of trans‐acting factors chloramphenicol acetyltransferase (CAT) reporter constructs containing from ‐1, 180 to + 13 to ‐112 to + 13 of a normal IL‐6 gene were elecroporated into the cell line. When these constructs are transferred into unstimulated fibroblasts, no CAT activity is seen; however, CAT activity is induced when the cells are stimulated with either IL‐1α, lipopolysaccharide (LPS), or cyclic adenosine monophosphate (cAMP) analogues. When the cell line OCI‐LY3 was transfected with these constructs, CAT activity was observed; it was not necessary to stimulate the cells with exogenous factors to observe this activity. No CAT activity was observed in a second lymphoma cell line, OCI‐LY13.1, that does not produce IL‐6. These results suggest that the constitutive production of IL‐6 by the cell line OCI‐LY3 is due to the presence of trans‐acting factors that stimulate the expression of IL‐6 and not due to a cis‐acting mutation of the IL‐6 promoter.
Induction of heat‐shock response and alterations of protein phosphorylation by a novel topoisomerase ii inhibitor, withangulatin A, in 9L rat brain tumor cells
Authors: Lee Wen‐Chuan , Lin Kae‐Yuan , Chen Chiu‐Ming , Chen Zong‐Tsi , Liu Hon‐Ju , Lai Yiu‐Kay 
Published: February 2005

AbstractWithangulatin A is a newly identified in vitro topoisomerase II inhibitor isolated from the Chinese antitumor herb Physalis angulata. In vivo, it was found to be cytotoxic, capable of suppressing general protein synthesis and of inducing the synthesis of a small set of proteins including those generated by heat‐shock treatment. The 70 kDa protein generated by withangulatin A was unequivocally identified as the heat‐shock protein70 (HSP70) since both proteins migrated to the same position on two‐dimensional polyacrylamide gels, could be recognized by a monoclonal antibody to human HSP70, and exhibited identical peptide maps. The induction of protein synthesis by withangulatin A was regulated at the transcriptional level since it was aborted in cells pre‐treated with actinomycin D. However, the initiation of this process did not require de novo protein synthesis since it was not affected by cycloheximide. Other cellular effect of withangulatin A was alterations of protein phosphorylation including an enhancement of phosphorylation of a 32 kDa protein which was also detected in the heat‐shocked cells. Morevoer, this process was observed within 7.5 min after the initial heat treatment which is much faster than the onset of HSP synthesis. Therefore, increased phosphorylation of the 65 kDa protein may represent on of the earliest signals generated by both heat‐shock and withangulatin A and may be involved in the upstream regulation of heat‐shock response in cells.
Effect of tunicamycin on glycosylation of a 50 kDa protein and thermotolerance development
Authors: Lee Yong J. , Kim Dooha , Hou Zi‐Zheng , Corry Peter M. 
Published: February 2005

AbstractWe investigated whether or not a 50 kDa glycoprotein might play an important role in protein synthesis‐independent thermotolerance development in CHO cells. When cells were heated for 10 min at 45.5°C, they became thermotolerant to a heat treatment at 45.5°C administered 12 hr later. The thermotolerance ratio at 10−3 isosurvival was 4.4. The cellular heat shock response leads to enhanced glycosylation of a 50 kDa protein. The glycosylation of proteins including a 50 kDa glycoprotein was inhibited by treatment with various concentrations of tunicamycin (0.2–2 μg/ml). The development of thermotolerance was not affected by treatment with tunicamycin after the initial heat treatment, although 2 μg/ml tunicamycin inhibited glycosylation by 95%. However, inhibiting protein synthesis with cycloheximide (10 μg/ml) after the initial heat treatment partially inhibited the development of thermotolerance. Nevertheless, there was no further reduction of thermotolerance. development by treatment with a combination of 2 μg/ml tunicamycin and 10 μg/ml cycloheximide. These data suggest that development of thermotolerance, especially protein synthesis‐independent thermotolerance, is not correlated with increased glycosylation of the 50 kDa protein.
Recessive (mediator−) revertants from c‐h‐ras oncogene‐transformed NIH 3T3 cells: Tumorigenicity in nude mice and transient anchorage and serum independence of the recovered tumor cells in culture
Authors: Omata‐Yamada Toshiko , Yamada Hisafumi , Lengyel Peter 
Published: February 2005

AbstractWe have reported earlier the isolation of two recessive, serum‐ and anchoragedependent revertants (R116 and R260) from a c‐H‐ras oncogene‐transformed NIH 3T3 line. In both revertants, the oncogene was fully expressed and fusion of either revertant with (untransformed) NIH 3T3 cells, or of the two revertants with one another, resulted in transformed progeny. These, and other data, indicated that the transforming activity of the oncogene was impaired in the two revertants in consequence of defects in distinct genes needed to mediate this activity.We report here that neither revertant could be re‐transformed by the K‐ras or N‐ras oncogene (though they could be re‐transformed by several other oncogenes). The two revertants turned out to be tumorigenic in nude mice (though less so than the parental transformed cells). The tumor cells, as recovered, formed foci and had a transformed morphology and a greatly diminished serum and anchorage dependence. Growth of the cells in culture (for 20 passages) resulted in their regaining the characteristics (i.e., anchorage and serum dependence) of cultured R116 and R260 cells. Proliferation of the cells in nude mice was not accompanied by a change in the level of ras oncogene expression or in gene amplification, at least as manifested in the lack of appearance of double‐minute chromosomes. The addition of the growth factors TGF alpha and beta to the medium of either revertant did not support anchorage‐independent growth.
Multiple pathways for ligand internalization in rat hepatocytes I: Effects of anoxia, phenylarsine oxide and monensin
Authors: Moss Anita L. , Ward Walter F. 
Published: February 2005

AbstractIt has been suggested that there are multiple pathways for the cellular internalization of insulin. To investigate these pathways we have examined the effects of three perturbations of endocytosis on the insulin internalization process and have compared these effects with those obtained using an asialoglycoprotein, asialofetuin (Afet), and epidermal growth factor (EGF). Freshly isolated hepatocytes were incubated with radiolabeled ligands and internalization measured under conditions of anoxia to deplete cellular ATP, in the presence of phenylarsine oxide (PAO) to inhibit endocytosis, and in the presence of monensin to interfere with endosomal acidification. Afet internalization essentially was blocked by all three treatment processes, while insulin internalization was inhibited approximately 40% in the presence of anoxia, and 54% in the presence of PAO. Monensin exhibited differential effects on internalization of high and low insulin concentrations. The effects of the treatment processes on EGF internalization were intermediate to those seen with Afet and insulin. These results suggest that insulin and EGF utilize routes of internalization exhibiting different energy requirements that may correspond to coated pit, non‐coated pit, and fluid‐phase internalization pathways. The observations with Afet internalization remain consistent with utilization of the coated pit pathway.
Inhibition of protein synthesis and heat protection: Histidinol‐resistant mutant cell lines
Authors: Lee Yong J. , Hou Zi‐Zheng , Kim Dooha , Al‐Saadi Abdul , Corry Peter M. 
Published: February 2005

AbstractThe mechanism of histidinol (HST) ‐induced heat protection was investigated to test the hypothesis that the cessation of protein synthesis itself is one of the events involved in heat protection. For this study, we isolated three HST‐resistant mutant strains. HST (5 mM), which inhibited protein synthesis by 88% in the wild type, caused only 0, 9, and 25% inhibition in three mutants, respectively. The drug, which afforded heat protection, (i.e., a 125‐fold increase in survival from 4 x 10−3 to5 x 10−1 after 2 hr at 43°C in wild type), did not protect mutant cells from heat killing. In contrast, cycloheximide (10 μg/ml) which inhibited protein synthesis by 95% in both wild type and mutant cell types, protected both cell types from heat killing. Therefore, these results suggest that the cessation of protein synthesis, per se, preventing synthesis of nascent polypeptides, is a major event leading to heat protection.
An IGF binding protein is an inhibitor of fgf stimulation
Authors: Villaudy J. , Delbé J. , Blat C. , Desauty G. , Golde A. , Harel L. 
Published: February 2005

AbstractWe purified to homogeneity a growth inhibiting diffusible factor (IDF45) secreted by dense cultures of mouse 3T3 cells and which was able to inhibit 100% of DNA synthesis stimulated by serum in chick embryo fibroblasts (CEF) (Blat et al., 1989a). We then demonstrated that this factor was an IGF‐binding protein (Blat et al., 1989b). Indeed, its N‐terminal amino acid sequence was homologous to that of rat IGFBP‐3. Our present results show that basic fibroblast growth factor (bFGF) induced, respectively, a fivefold and threefold increase in DNA synthesis in mouse embryo fibroblasts (MEF) and CEF. IDF45 inhibited the stimulation induced by bFGF by about 65%, while stimulation induced by insulin, PDGF, or EGF was only weakly or not at all inhibited by IDF45. When bFGF stimulation was determined in the presence of a high concentration of insulin in conditions which minimize the effect of endogenous IGF‐I or ‐II, this stimulation was decreased by about 50% in the presence of IDF45. This result suggests that addition of bFGF stimulates IGF secretion, thereby resulting in partial loss of inhibition, by IDF45, of bFGF stimulation.
Na+/K+/Cl− cotransport is stimulated by a Ca++‐calmodulin–;mediated pathway in BALB/c 3T3 fibroblasts
Authors: Snyder David , Atlan Henri , Markus Miriam , Panet Rivka 
Published: February 2005

AbstractIn the present study, we investigated the role of intracellular Ca++ in the stimulation of the Na+/K+/Cl– cotransport in synchronized BALB/c 3T3 cells. The Na+/K+/Cl– cotransport was stimulated by the growth factors EGF, TGF‐α, IGF‐1, and IGF‐2, which do not activate protein kinase C, but do induce a transient increase in free cytoplasmic Ca++ In addition, direct activation of protein kinase C by the phorbol ester 12‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA) did not affect the Na+/K+/Cl– cotransport activity of quiescent cells. The Na+/K+/Cl– cotransport was also stimulated by the above mitogens in cells pretreated with the phorbol ester TPA. This treatment led to a progressive decline in the activity of cellular protein kinase C. This result implies that cells deficient in protein kinase C may still support stimulation of the Na+/K+/Cl– cotransport. Taken as a whole, these findings suggest that the Na+/K+/Cl– cotransport is stimulated predominantly by a protein kinase C‐independent mechanism in BALB/c 3T3 fibroblasts.Both the intracellular Ca++ antagonist 8‐(N,N‐diethylamino)octyl‐3,4,5‐trimethoxybenzoate (TMB‐8) and two potent calmodulin antagonists, trifluoperazine (TFP) and chloropromazine(CP), blocked serum‐ and mitogen‐stimulated Na+/K+/Cl– cotransport. These results suggest that the Na+/K+/Cl– cotransport is stimulated by an increase of intracellular Ca++ and subsequently by a Ca++‐calmodulin‐mediated pathway in the synchronized BALB/c 3T3 fibroblasts.
Biologically active synthetic fragments of human basic fibroblast growth factor (bFGF): Identification of two Asp‐Gly‐Arg‐Containing domains involved in the mitogenic activity of bFGF in endothelial cells
Authors: Presta Marco , Rusnati Marco , Urbinati Chiara , Sommer Andreas , Ragnotti Giovanni 
Published: February 2005

AbstractSynthetic peptides derived from the amino acid sequence of human basic fibroblast growth factor (bFGF) have been assayed for the capacity to exert bFGF agonist and antagonist activities in cultured endothelial cells. bFGF fragments A and C, which correspond to the sequences bFGF (38–61) and bFGF (82–101), induce a limited but statistically significant increase in cell number when administered to cultures of fetal bovine aortic endothelial GM 7373 cells and adult bovine aortic endothelial cells. The two peptides also exert a partial antagonist activity when GM 7373 cells are stimulated to proliferate by bfGF, but they do not affect cell proliferation induced by serum, epidermal growth factor (EGF), phorbol ester (TPA), or 1,2‐diacylglycerol (diC8). Moreover, antibodies raised against peptides A and C specifically quench the mitogenic activity of bFGF. Peptides A and C contain the amino acid sequence Asp‐Gly‐Arg (DGR), which is the inverse of the cell adhesion signal sequence RGD recognized by integrins. DGR‐ and RGD‐containing tetra‐ and heptapeptides inhibit the mito‐genic activity exerted by bFGF and by the two active bFGF fragments. They do not affect cell proliferation induced by acidic FGF, EGF, serum, TPA, and diC8. However, neither peptides A and C, their corresponding antibodies, nor DGR‐and RGD‐containing peptides inhibit the binding of 125I‐bFGF to its low and high affinity binding sites. The data suggest that amino acid residues 38–61 and 82–101, both containing a core DGR sequence, represent two “activation” domains of bFGF. Both domains are involved in the modulation of the mitogenic activity of bFGF without interacting directly with the bFGF receptor.