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Biomedical subjects

J A Fernandez-Pol

Publications and source records attributed to J A Fernandez-Pol.

At least 19 recordsLinked to original sources
PubMed2001

Differential expression of metallopanstimulin/S27 ribosomal protein in hepatic regeneration and neoplasia.

We have previously shown that human metallopanstimulin-1 (MPS-1) is a ubiquitous 9.4-kd multifunctional ribosomal S27/nuclear "zinc finger" protein that is expressed at high levels in a wide variety of actively proliferating cells and tumor tissues. In this study, we examined the expression of MPS-1 in chronic hepatitis, cirrhosis, and hepatocellular carcinoma. Tissue samples were obtained at the time of tumor resection, needle biopsy, or liver transplantation. MPS- 1 was studied by immunohistochemistry by use of specific antibodies to the N-terminus of MPS-1 in a biotin/streptavidin-amplified system. In chronic hepatitis, hepatocytes had very weak MPS-1 immunostaining. In contrast, hepatocytes in regenerating cirrhotic nodules stained strongly for MPS-1. In well-differentiated hepatocellular carcinoma, MPS-1 presence was intense at the periphery of the malignant nodule. In poorly differentiated hepatocellular carcinoma, MPS-1 presence was notably intense in malignant hepatocytes invading the septal tissues, in close contact with neovascular structures. These results suggest that MPS-1 may be involved in both progression toward malignancy in regenerating cirrhotic nodules and in subsequent steps of hepatocarcinogenesis.

Antibodies↗
PubMed2000

Molecular interactions of cancer and age.

The authors believe that the aging process--the loss of youthful resilience--is caused by the decline of many hormones. By restoring these hormone levels, the decay associated with old age can be eliminated and in some cases, perhaps reversed. The hormones that naturally occur in the body should be replenished through a medically sound regimen. The hormones must work together. The data indicate that aging occurs at two levels: systemically and cellularly. Systemic controls regulate the rates of intracellular enzymatic processes that accelerate to protect against aging.

Adult↗
PubMed1997

Differential expression of metallopanstimulin/S27 ribosomal protein in melanocytic lesions of the skin.

We have previously shown that human metallopanstimulin (MPS-1) is a ubiquitous 9.4-kDa multifunctional ribosomal S27/nuclear "zinc finger" protein which is expressed at high levels in a wide variety of cultured proliferating cells and tumor tissues, including melanoma. In the present study, we have examined the expression of the MPS-1 protein in various types of human benign and malignant melanocytic lesions of the skin. The expression of the MPS-1 protein was studied by immunohistochemistry using specific anti-MPS-1 antibodies. We found that in benign nevi, the staining is weak and in a gradient; most often, only type A melanocytes stain positive. The B and particularly the C types are negative. Remarkably, congenital nevi show a similar gradient staining of regular benign nevi, but in addition one example showed intensely positive dermal nodules adjacent to areas of negative melanocytes. In melanomas, the staining patterns for MPS-1 are more complex. While some melanomas stain evenly and intensely positive, others have remarkably variable expression of MPS-1. The scattered melanocytes migrating to the upper layers of the epidermis are usually intensely positive. In summary, benign lesions stain in an orderly pattern with staining gradients that correlate with the cellular differentiation of the nevi. Malignant melanomas have an erratic, often intense staining that also correlates with the disorderly growth of these neoplasms. These differential results indicate that the MPS-1 antigen is a useful marker for melanocytic lesions at the immunohistochemical level.

Amino Acid Sequence↗
PubMed1994

Metallopanstimulin gene product produced in a baculovirus expression system is a nuclear phosphoprotein that binds to DNA.

The protein product of the human Metallopanstimulin (MPS-1) gene was produced in the insect cell line Spodoptera frugiperda (Sf9) using the baculovirus expression vector system Autographa californica nuclear polyhedrosis virus (AcNPV). When a cloned MPS-1 complementary DNA sequence was inserted into the AcNPV viral genome downstream from the promoter of the polyhedrin gene, a polypeptide with an apparent molecular weight of approximately 10,000 was observed in extracts of infected Sf9 cells. This protein was not detected in Sf9 cells infected with AcNPV-MPS-1-Del, a vector in which the MPS-1 gene was deleted. The MPS-1 protein was produced at high levels in this host-vector system (congruent to 12% of total labeled soluble protein). Characterization of the MPS-1 protein extracted from Sf9 infected cells showed that it: binds zinc ions specifically; is phosphorylated; accumulates in the nucleus; is tightly bound to the nucleus; and binds to calf thymus DNA-cellulose. The MPS-1 protein constitutes one of the major proteins in the nuclear fraction of Sf9 cells and can be purified from this source to near homogeneity by a two-step procedure composed of high-performance liquid chromatography and gel electrophoresis. Antibodies were raised against selected peptide sequences of the MPS-1 protein and used to detect MPS-1 in insect cells and in various cultured mammalian cell types. Western analysis demonstrated that the baculovirus-generated protein had electrophoretic and immunological properties equivalent to those of MPS-1 spontaneously expressed in various human mammalian cell lines. Finally, recombinant MPS-1 generated a specific protein-DNA complex with a duplex oligomer containing a cyclic AMP-responsive element, as assessed by gel mobility shift assays. These results support the hypothesis that the MPS-1 protein may act, at least in part, by interacting with genes possessing the cyclic AMP-responsive element sequence.

Animals↗
PubMed1993

A growth factor-inducible gene encodes a novel nuclear protein with zinc finger structure.

Growth factors rapidly induce numerous genes that encode regulatory proteins, many of which have been identified by cDNA cloning. In this study, differential hybridization was used to screen a cDNA library constructed from human mammary carcinoma MDA-468 cells that were stimulated with transforming growth factor beta-1 (TGF-beta 1) in the presence of cycloheximide. One of the cDNA clones that was induced by TGF-beta 1 was found to have a nucleotide sequence that predicts a 9,450-Da protein with homology to regulatory DNA-binding proteins. This clone was designated metallopan-stimulin-1 (MPS-1) because it encodes a metalloprotein whose mRNA is expressed in a wide variety of actively proliferating cells and tumor tissues. MPS-1 protein contains one "zinc finger" domain of the C4 type, similar to those present in proteins of the steroid/thyroid hormone receptor superfamily and other DNA-binding proteins. The mRNA for MPS-1 was induced in MDA-468 cells by fetal calf serum, TGF-beta 1, TGF-beta 2, and cAMP analogues. The MPS-1 gene is expressed at relatively high levels in several human carcinoma cell lines, particularly those derived from ectodermal layers, and at higher levels in melanomas (ontogenically of neural origin). In contrast, the MPS-1 mRNA is expressed at low levels in normal WI-38 human lung diploid fibroblasts in culture. We hypothesize that MPS-1 protein may play a role as a potentially important mediator of cellular proliferative responses to various growth factors and other environmental signals.

Amino Acid Sequence↗
PubMed1993

Epidermal growth factor stimulates thyrotropin secretion in the rat.

The present work studied the effects of epidermal growth factor (EGF) on the release of thyrotropin (TSH) and prolactin (PRL) from perifused pituitary glands of 200-gram male Wistar rats. Each pituitary gland, cut into halves, was placed in a chamber of a perifusion system connected to a peristaltic pump which conveyed the perifusion medium (Medium 199, pH 7.3, Gibco, USA) from a reservoir to a chamber at a flow rate of 100 microliters/min. Each tightly closed chamber contained one pituitary gland and 600 microliters medium and it was placed in a water bath at 37 degrees C throughout the experiment. One milliliter samples of effluent were collected every 10 min for 60 min to obtain baseline values of TSH and PRL. Thereafter, TSH-releasing hormone (TRH) 10(-8) M or EGF (10(-11), 10(-10), 10(-9) or 10(-8) M) were added to individual chambers and the 10-min sampling of effluent continued for 60 min. EGF 10(-11) M elicited no TSH response, but 10(-10) and 10(-9) M doses induced significant increases in TSH secretion (p < 0.01) with a peak at 10 min after addition of EGF. In another experiment, EGF 10(-8) M or TRH 10(-8) M significantly elevated TSH secretion (p < 0.01). However, TRH, but not EGF, stimulated PRL secretion (p < 0.01). In the in vivo studies, the intravenous administration of EGF 10(-5) M or TRH 10(-5) M both induced significant elevation of TSH release at 10 min after the injection (p < 0.02 for EGF and p < 0.01 for TRH).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Modulation of EGF receptor protooncogene expression by growth factors and hormones in human breast carcinoma cells.

In this review I summarize the experimental data in favor of the notion that control of epidermal growth factor (EGF) receptor (R) and/or c-erbB-2 protooncogene expression by specific autocrine growth factors and certain classical endocrine hormones serves as a transducer of extracellular signals that ultimately lead to growth responses in breast carcinoma cells. I summarize some new results on the role of epidermal growth factor (EGF), transforming growth factor (TGF) alpha, and TGF beta in the control of EGF-R protooncogene expression in human breast carcinoma cells. Furthermore, the data embracing the hypothesis that the growth actions of hormone receptors that are homologous to the v-erbA oncogene (estrogens, progesterone, thyroid hormones, retinoic acid, and vitamin D) are mediated, in part, by modulating EGF-R and/or c-erbB-2 protooncogene transcription are reviewed. Finally, I develop the theme that cooperation of certain c-erb-A-related, c-erbB-2 and/or EGF-R gene products contribute to the uncontrolled growth of human mammary carcinoma cells. From the evidence reviewed, one can infer that elucidation of the molecular control of EGF-R/c-erbB-2 gene expression by c-erbA-related gene products may lead to both a better understanding of breast carcinogenesis and a new therapeutic approach directed at controlling the transcriptional responses of EGF-R/c-erbB-2 genes.

Biomarkers, Tumor↗
PubMed1989

Transcriptional regulation of proto-oncogene expression by epidermal growth factor, transforming growth factor beta 1, and triiodothyronine in MDA-468 cells.

We have examined the epidermal growth factor (EGF) dependence of the transcription of different proto-oncogenes, using cultured human mammary carcinoma MDA-468 cells and a nuclear run-on transcription assay. We found that the stimulation of MDA-468 cells with EGF regulates moderately and to different extents the transcription of the EGF-receptor(R) and c-erbB-2 proto-oncogenes. In contrast, the transcription of other proto-oncogenes, including c-myc, c-H-ras, and c-fps, was unchanged. Furthermore, we provide evidence that transforming growth factor beta 1 (TGF-beta 1) selectively and to different degrees modulates the EGF-dependent transcription of EGF-R and c-erbB-2 genes. In this study, we also discovered that T3 (triiodothyronine) exerts synergistic control on the action of EGF alone or in association with TGF-beta 1, on EGF-R and c-erbB-2 gene transcription. Moreover, we established that within 6 h after the addition of EGF, cytoplasmic EGF-R mRNA levels are increased several-fold and that this accumulation is enhanced by the presence of TGF-beta 1 and/or T3. The results described here are consistent with the hypothesis that a complex program of cooperative interactions among EGF-, TGF-beta 1-, and T3-generated signals at the transcriptional level may mediate, at least in part, the combined actions of EGF, TGF-beta 1, and T3 in target cells.

Breast Neoplasms↗
PubMed1989

Modulation of transforming growth factor alpha-dependent expression of epidermal growth factor receptor gene by transforming growth factor beta, triiodothyronine, and retinoic acid.

We have investigated the actions of transforming growth factor (TGF) type alpha on epidermal growth factor (EGF) receptor mRNA expression in MDA-468 human mammary carcinoma cells in serum-free media. We found that exposure of MDA-468 cells to TGF alpha results in elevated levels of EGF receptor mRNA. This increase in mRNA accumulation showed time and dose dependence. Addition of TGF beta 1 enhanced the accumulation of EGF receptor mRNA induced by TGF alpha in a time- and dose-dependent manner. We also found that triiodothyronine at physiological concentrations exerts synergistic control on the action of TGF alpha alone, or in association with TGF beta 1, on EGF receptor mRNA expression. Similarly, retinoic acid treatment also enhanced in a time- and dose-dependent manner the TGF alpha-dependent response of EGF receptor mRNA and acted synergistically with TGF beta 1. The results described here suggest that optimum regulation of EGF receptor gene expression by TGF alpha is a complex process involving synergistic interactions with heterologous growth factors and hormones.

Breast Neoplasms↗
PubMed1987

Modulation of epidermal growth factor receptor gene expression by transforming growth factor-beta in a human breast carcinoma cell line.

Modulation of epidermal growth factor (EGF) receptor expression determines cellular responsiveness to EGF and might play an important role in growth inhibition. We have investigated the actions of EGF and/or transforming growth factor type beta (TGF beta) on EGF receptor gene expression in MDA-468 human breast carcinoma cell line, which responds to EGF and/or TGF beta with growth inhibition. Using the cDNA clone pE7, which encodes 2.4 kilobases of the human EGF receptor mRNA, as a hybridization probe, we have found that exposure of MDA-468 cells to EGF results in elevated levels of EGF receptor mRNA. This increase in mRNA accumulation showed time and dose dependence. Addition of TGF beta enhances the accumulation of EGF receptor mRNA induced by EGF. Under this condition, stimulation could be detected after 1 h exposure to TGF beta with a maximum at 6-8 h. A concentration of 10 pM TGF beta gave detectable stimulation with maximal stimulation occurring at 300 pM in the presence of EGF (50 ng/ml). In contrast, TGF beta alone had no significant effect on EGF receptor mRNA accumulation. In the presence of cycloheximide, the EGF receptor mRNA was super-induced in response to EGF. Treatment of the cells with TGF beta enhances the EGF-dependent superinduction of EGF receptor mRNA produced by cycloheximide, suggesting that the stimulatory action of TGF beta does not depend on continuous protein synthesis. The results described here are consistent with the hypothesis that the growth inhibitory action of TGF beta in MDA-468 cells may be mediated, at least in part, by modulation of EGF receptor gene expression.

Breast Neoplasms↗
PubMed1987

Suppression of the EGF-dependent induction of c-myc proto-oncogene expression by transforming growth factor beta in a human breast carcinoma cell line.

Alterations in c-myc proto-oncogene expression after treatment of human mammary carcinoma MDA-468 cells with epidermal growth factor (EGF) and/or transforming growth factor beta (TGF beta) have been investigated. A stimulation of c-myc messenger RNA was detected within 60 min after treatment with EGF. This induction persisted for at least 24 hr, albeit to a lower extent. The early and late increase in c-myc mRNA levels induced by EGF were inhibited by the presence of TGF beta. TGF beta alone induced little change in c-myc mRNA levels. The effect of TGF beta represents a novel action of this hormone at the level of gene expression.

Breast Neoplasms↗
PubMed1986

Purification and biological properties of type beta transforming growth factor from mouse transformed cells.

Transforming growth factor type beta (TGF beta) has been purified from serum-free culture fluids of transformed mouse L-929 cells which are capable of continual growth in serum-free medium in the absence of any exogenously added polypeptide growth factors. TGF beta has been purified to homogeneity as judged by NH2-terminal amino acid sequence analysis. Analysis of the purified polypeptide by gel electrophoresis indicates that TGF beta is composed of two polypeptide chains of Mr 12,500 cross-linked by disulfide bonds. TGF beta was characterized by its ability to induce anchorage-dependent normal rat kidney (NRK) cells to grow in soft agar in the presence of epidermal growth factor (EGF). TGF beta was also able to enhance both EGF-induced DNA synthesis and cell proliferation on growth-arrested NRK cells in monolayer cultures under serum-free conditions. We also show that in mouse melanoma B-16 cells under serum-free conditions TGF beta stimulates both DNA synthesis in monolayer cultures and anchorage-independent growth in soft agar. Paradoxically, the anchorage-independent growth in the presence of serum of many human cell lines, including melanomas, and mammary, prostatic, vulvar, and lung carcinomas is inhibited by TGF beta at saturating concentrations similar to those that stimulate colony formation of the rodent cell lines L-929 and B-16 under serum-free conditions. The peculiar action of TGF beta is further revealed by the observations that while EGF and TGF beta synergize to induce inhibition of anchorage-independent growth of A-431 human vulvar carcinoma cells, their effects on the anchorage-independent growth of one human lung carcinoma cell line (A-549) and two human prostatic carcinoma cell lines (PC-3 and DU-145) are antagonistic. Moreover, we show that in the rodent and human cell lines TGF beta interacts with specific cellular receptors which may mediate the actions of TGF beta. We conclude that the expression of both TGF beta and TGF beta receptors by L-929 cells and the stimulation of growth of L-929 cells in serum-free medium by TGF beta suggests that TGF beta may be important for maintaining the transformed state of this tumor cell line, and the way in which a cell responds to TGF beta is dependent on the presence or absence of growth factors contained in the serum.

Amino Acid Sequence↗
PubMed1985

Epidermal growth factor receptor of A431 cells. Characterization of a monoclonal anti-receptor antibody noncompetitive agonist of epidermal growth factor action.

A monoclonal antibody to the epidermal growth factor (EGF) receptor of A431 cells, denoted 2D1-IgM, was generated after fusion of immunized BALB/c mouse spleen cells with SP2/0-Ag14 myeloma cells. Specific binding of 2D1-IgM to the A431 cell-surface receptor for EGF was demonstrated by indirect immunofluorescence, immunoprecipitation, and immunoblot analysis. Scatchard analysis of 125I-EGF binding to A431 cells demonstrated that 2D1-IgM treatment did not change the number of EGF receptors, but caused an increase in the affinity of EGF receptors from a population of low affinity to a uniform population of high affinity. Like EGF, 2D1-IgM induced phosphorylation of EGF receptors and EGF receptor clustering. As in the case of EGF, a biphasic growth response with stimulation of DNA synthesis at low and inhibition at high concentrations of 2D1-IgM was evident in A431 cells. The intrinsic "EGF-like" bioactivity of 2D1-IgM was enhanced by the presence of EGF. These results suggest that the binding of 2D1-IgM to the EGF receptor at a different site from that to which EGF binds can initiate an effective EGF-like biological response; and the EGF-like biological effects of 2D1-IgM may be mediated by a population of high affinity EGF receptors which may be involved in the control of cellular growth.

Animals↗
PubMed1982

Modulation of protein phosphorylation in human colon adenocarcinoma cell membrane preparations by epidermal growth factor in vitro.

Incubation of membranes prepared from the human colon adenocarcinoma cell line LoVo in vitro with [gamma-32P]ATP demonstrated numerous components whose phosphorylation was stimulated several fold by epidermal growth factor (EGF). One major component of Mn 170 K, which was either undetectable or very weakly phosphorylated in the absence of EGF, was primarily affected by exposure to EGF. The phosphorylation of the 170 K Mr membrane component required the presence of Mn2+; Mg2+ was ineffective. Although the phosphorylation of many LoVo membrane components was significantly modified by cAMP or dibutyryl-cAMP, none of these cyclic nucleotides appeared to be involved in the phosphorylation of the 170 K membrane component in the presence or absence of EGF. The phosphorylation system of the LoVo membranes efficiently utilized [gamma-32P]GTP in both the basal and EGF-stimulated reactions. All the membrane components phosphorylated in the absence or presence of EGF, except a band comigrating with the tracking dye front, were digested by trypsin. The possible glycoprotein nature of the 170 K dalton phosphoprotein was indicated by the fact that the 170 K dalton band comigrated with a periodic acid-Schiff base-stained band. These findings suggest that one of the biochemical steps in the mechanism of action of EGF in LoVo cells is enhanced phosphorylation of several membrane proteins, especially that of a glycoprotein of Mr 170 K, by a membrane-bound cyclic nucleotide independent protein kinase.

Adenocarcinoma↗
PubMed1982

Correlation between the loss of the transformed phenotype and an increase in superoxide dismutase activity in a revertant subclone of sarcoma virus-infected mammalian cells.

We have studied the effects of paraquat (methyl viologen), a herbicide that increases intracellular production of superoxide radical, on the viability of virus-transformed and nontransformed normal rat kidney (NRK) cells in culture. We have shown that a low concentration of paraquat (12.5 microM) is cytotoxic toward virus-transformed cell lines, including Kirsten sarcoma virus- and SV40-transformed NRK cells. The corresponding untransformed NRK cells were resistant to the same and a 4-fold higher concentration of paraquat. There was a good correlation between the susceptibility of transformed and untransformed cells to paraquat cytotoxicity and their ability to increase the superoxide dismutase (SOD) enzymatic activity. We found that paraquat is cytotoxic toward Kirsten sarcoma virus-transformed and SV40-transformed NRK cells which showed low intracellular SOD activity. The relationship between SOD activity and paraquat cytoxicity was strengthened by the finding that the tolerance of NRK cells to the drug was associated with high intracellular SOD activity. This report also describes the isolation of a revertant (revertant RE8G3) cell line derived from Kirsten sarcoma virus-transformed NRK cells after paraquat treatment which contains SOD activity at levels much higher than those found in NRK cells. This revertant is undistinguishable from NRK cells with respect to its lack of transformed cell properties. Not only are these cells normal morphologically but also they do not grow in soft agar, an in vitro property that closely correlates with in vivo tumorigenicity. Several biological and biochemical properties of RE8G3 cells, including growth characteristics, surface receptors for both transferrin and epidermal growth factor (EGF), and the EGF-dependent 32P phosphorylation of specific membrane polypeptides have been studied. The most interesting conclusion that can be drawn from these studies is that there is a correlation between loss of the transformed phenotype and an increase in both EGF receptors and EGF-dependent 32P phosphorylation of a m.w. 170,000 membrane-associated protein.

Animals↗