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Biomedical subjects

G Mincione

Publications and source records attributed to G Mincione.

At least 19 recordsLinked to original sources
PubMed1998

Cyclin D1 and Cyclin E expression in malignant thyroid cells and in human thyroid carcinomas.

Evidence of the involvement of cyclin gene alterations in human cancer is growing. In this study, we sought to determine the pattern of expression of cyclin D1 and cyclin E in normal and malignant thyroid cells. Quiescent rat thyroid cells in culture, induced to synthesize DNA by thyrotropin (TSH), expressed cyclin D1 gene after 6 hr and cyclin E gene with a peak at 18 hr from the stimulus; K-ras-transformed rat thyroid cells, which grew without addition of hormones necessary for normal cell proliferation, expressed elevated levels of cyclin D1 and cyclin E, compared with normal differentiated thyroid cells. Human benign and malignant thyroid tumors and their relative normal tissues were then analyzed. Neither major genetic alterations nor amplifications for cyclin D1 and cyclin E genes were found by Southern blot analysis in genomic DNAs extracted from all types of thyroid tumors. Moreover, statistical analyses of densitometric values from Northern blots did not show increased levels of cyclin D1 and E mRNAs in the tumor samples, compared with normal thyroid. Immunohistochemical analyses of formalin-fixed paraffin-embedded sections of tissues with specific antibodies revealed a prevalent cytoplasmic cyclin E staining in the thyroid tissues analyzed. Cyclin D1, instead, was present in the cytoplasm of normal thyroids and adenomas, but in 31% of thyroid papillary carcinomas analysed, it was overexpressed, with a localization in the nucleus. Our in vivo observations suggest that unlike cyclin E, elevated nuclear cyclin D1 expression defines a subset of thyroid papillary carcinomas, and might be a contributory factor to thyroid tumorigenesis.

Animals↗
PubMed1998

Heregulin-dependent autocrine loop regulates growth of K-ras but not erbB-2 transformed rat thyroid epithelial cells.

The EGF-like family of proteins, such as epidermal growth factor (EGF), transforming growth factor alpha (TGFalpha), amphiregulin (AR), betacellulin (BTC), cripto-1 (CR-1), and heregulin (HRG), plays an important role in the pathogenesis of several human carcinomas as autocrine growth factors. Differentiation and proliferation of rat thyroid cells in culture (FRTL-5 cells) are regulated by thyrotropin (TSH); withdrawal of TSH from culture medium produces growth arrest, whereas its addition to quiescent cells stimulates cell entry into S phase. Instead, transformed thyroid cell lines as FRTL-5H2 cell line, overexpressing erbB-2, Kimol cells, transformed by the wild-type K-ras and A6 clone, transformed by a temperature sensitive K-ras mutant, can grow without addition of TSH to the culture medium. In order to identify whether EGF-like growth factors and corresponding receptors (erbB-2, erbB-3, and erbB-4) could be involved in the autonomous growth of these transformed rat thyroid epithelial cells, Northern blot for mRNA analysis and Western blot for protein expression were performed. In contrast to normal control FRTL-5 cells, both K-ras and erbB-2-transformed cells expressed elevated levels of erbB-2 receptor. Moreover, both K-ras transformed cells, Kimol and A6 cells, but no FRTL-5H2 cells, were found able to express also high levels of erbB-4 receptor and HRG/NDF ligand. Treatment of K-ras transformed thyroid cells with neutralizing antibody against HRG/NDF reduced by 50% cell proliferation. These data indicate that unlike the erbB-2 overexpressing FRTL-5 cells, in K-ras rat thyroid epithelial cells, the growth factor heregulin signals through the heterodimer erbB-2/erbB-4 receptors in an autocrine fashion.

Amphiregulin↗
PubMed1998

Carbonic anhydrase inhibitors: inhibition of isozymes I, II and IV with N-hydroxysulfonamides--a novel class of intraocular pressure lowering agents.

A series of N-hydroxy sulfonamides has been prepared by reaction of alkyl-, arylalkyl- and arylsulfonyl halides or sulfonic acid anhydrides with hydroxylamine. Structurally related inhibitors were also obtained from acyl chlorides and hydroxylamine, as well as by reaction of tosyl isocyanate with hydroxylamine, sulfamic acid and sulfamide. Inhibition of three carbonic anhydrase (CA) isozymes, hCA I, hCA II and bCA IV (h = human; b = bovine) with the prepared compounds has been investigated. Good inhibitors, as well as compounds with moderate activity against these isozymes were detected, depending on the R group to which the SO2NHOH or CONHOH moieties were attached. Susceptibility to inhibition was generally: hCA II > bCA IV >> hCA I. Some of the new inhibitors showed very good antiglaucoma action when administered directly into the eye in experimental animals, acting as more efficient intraocular pressure lowering agents as compared to the clinical drug dorzolamide. This constitutes an encouraging result for obtaining novel antiglaucoma drugs from this class of CA inhibitors.

Acetamides↗
PubMed1997

Human malignant thyroid tumors displayed reduced levels of transforming growth factor beta receptor type II messenger RNA and protein.

Transforming growth factor beta (TGF-beta) is a physiological regulator of thyroid epithelial cell growth and differentiation. This factor signals through a heteromeric complex composed of type I (TGF-beta receptor type I) and type II [TGF-beta receptor type II (TbetaRII)] receptors. Loss of TbetaRII expression has been related to resistance to TGF-beta inhibition of cell proliferation. In the present work, we analyzed the TbetaRII expression in a series of human thyroid tumors, from benign lesions (adenomas) to neoplastic lesions of increasing aggressiveness (papillary and follicular carcinomas) up to the extremely aggressive anaplastic tumors. Results obtained indicated a clear reduced expression of TbetaRII mRNA only in the group of thyroid carcinomas when compared with their relative normal tissues. Immunohistochemical analyses with specific anti-TbetaRII antibodies confirm these observations. These data indicate that loss of expression of TbetaRII can contribute to thyroid cancer progression, inducing cancer cells to escape the growth-inhibitory effect of TGF-beta.

Adenocarcinoma, Follicular↗
PubMed1997

Restored expression of transforming growth factor beta type II receptor in k-ras-transformed thyroid cells, TGF beta-resistant, reverts their malignant phenotype.

Transforming growth factor beta 1 (TGF beta 1) inhibits the growth of normal rat epithelial thyroid cells (FRTL-5 strain) by counteracting thyrotropin (TSH)-stimulated DNA synthesis and by slowing the cells in the G1 phase of the cell cycle. Here, we have studied two clones of FRTL-5 thyroid cell line transformed by the wild type (wt) v-k-ras oncogene (K.M.A1, K.M.A2) and one clone (A6) transformed by a temperature-sensitive (ts) v-k-ras mutant. Anchorage-dependent as well as anchorage-independent growth of these k-ras-transformed cells was not inhibited by TGF beta 1. TGF beta 1 resistance appeared to be dependent by a functional p21 k-ras, because A6 cell growth was partially inhibited at the nonpermissive temperature (39 degrees C). To determine the basis for TGF beta 1 resistance in k-ras-transformed thyroid cells, we looked for possible defects in the expression of type I (T beta R-I/ALK5) and type II TGF beta receptors (T beta R-II). Lower levels of type II receptors were present in all of the k-ras-transformed clones, as revealed by both Northern blot and cross-linking experiments. A partial reversion of the malignant phenotype of the wt k-ras-transformed clone was obtained in two clones isolated after transfection of the malignant thyroid cells (K.M.A1) with a T beta R-II expression vector. These two clones also showed restored levels of exogenous T beta R-II mRNA and protein, and both clones showed a partially reacquired sensitivity to TGF beta 1. Similarly, the reversion of the malignant phenotype of the A6 clone grown at the nonpermissive temperature was accompanied by a restored expression of the T beta R-II receptors. These data indicate that active k-ras oncogene can induce TGF beta 1 resistance in rat thyroid cells and suggest that one of the possible mechanisms of escape from TGF beta 1 growth control in k-ras-induced thyroid carcinogenesis involves a reduced expression of T beta R-II receptors.

Animals↗
PubMed1997

Increased transforming growth factor-beta production and gene expression by peripheral blood monocytes of hypertensive patients.

Cultured human peripheral blood monocytes are known to secrete and express transforming growth factor-beta (TGF-beta), a multifunctional cytokine that can be involved in myocardial and vascular remodeling. In addition, monocytes/macrophages have been demonstrated to be colocalized with fibrosis of hypertrophied heart and in the vascular wall of hypertensive vessels. In this study, we tested TGF-beta production and mRNA expression in peripheral blood monocytes from hypertensive patients with myocardial hypertrophy and increased carotid myointimal thickness with respect to healthy normotensive control subjects. We found an increased TGF-beta activity in the conditioned medium of monocytes from hypertensive patients compared with control subjects as evaluated by inhibition of [3H]thymidine incorporation by mink lung epithelial cells (-83% and -18% in hypertensive and normotensive subjects; P<.001). Western blot analysis confirmed a significant difference in the amount of TGF-beta protein secreted in the conditioned medium of hypertensive patients compared with that of normotensive subjects. Finally, we also observed a 4.2- and 5.5-fold increase in the amount of TGF-beta1 and TGF-beta2 transcripts, respectively. Our results indicate an upregulation of the TGF-beta system in the peripheral blood monocytes of hypertensive patients with cardiovascular structural changes, suggesting a possible role of TGF-beta monocyte production in hypertensive disease.

Biological Assay↗
PubMed1997

Oncogenes and antioncogenes involved in human thyroid carcinogenesis.

The development of cancer is due to the accumulation of multiple somatic mutations, in some cases following germline mutations, which occur in hereditary malignancies such as retinoblastomas or multiple endocrine neoplasia (MEN 2A and B). Genetic alterations or changes in the expression of growth regulatory genes can lead to the initiation of malignant transformation and to eventual tumor progression. Cells that have undergone these cumulative alterations in either the structure or expression of these regulatory genes generally possess a selective growth and/or metastatic advantage over other normal non-transformed cells. Thus, activation of dominantly transforming oncogenes by point mutations, gene amplification, chromosomal translocation or insertional mutagenesis can lead to uncontrolled cellular growth or to a disruption in normal differentiation or apoptosis. Equally contributory to the process of malignant progression is the inactivation of recessive tumor suppressor genes due to point mutations and/or loss of heterozygosity in one allele, which can ultimately lead to a reduction of homozygosity in both alleles. Thyroid tumors in humans represent a particularly suitable multistage model of epithelial tumorigenesis. In fact, even though most thyroid neoplasms originate from a single cell type, i.e. the thyroid follicular cell, they include a broad spectrum of tumors with different phenotypic characteristics and variable biological and clinical behaviour. Multiple degrees of malignancies have been defined: from the benign colloid adenomas through the slowly progressive differentiated papillary and follicular carcinomas to the invariably fatal anaplastic carcinomas, although these histological changes are not necessarily sequential. In this review an effort has been made to summarize and integrate new data published on genetic lesions and altered expression of genes involved in the tumorigenesis of the follicular type of thyroid cancer. We have focused our interest only on gene alterations inducing gain or loss of function, that have been studied in vivo in human thyroid tumor specimens by the use of different techniques, such as PCR mediated DNA analyses, sequencing, mRNA level evaluation and protein expression by immunohistochemical staining.

Genes, Tumor Suppressor↗
PubMed1996

Transforming growth factor beta regulates differentiation and proliferation of human neuroblastoma.

The effects of transforming growth factor-beta1 (TGFbeta) on two human neuroblastoma cell lines, LAN-5 and SK-N-AS, and one human glioblastoma cell line, GL15, were evaluated. Of the three cultures, only two, SK-N-AS and GL15, had a complete response to TGFbeta, with induction of the following effects: (i) inhibition of cell proliferation; (ii) up-regulation of the extracellular matrix glycoprotein fibronectin, together with down-regulation of the VLA5 integrin receptor; (iii) up-regulation of histotype-specific cytoskeletal intermediate filaments (neurofilaments for neuroblastoma and GFAP for glioblastoma); and (iv) increase in the glycoprotein CD44, only in SK-N-AS. In the third cell line, neuroblastoma LAN-5, the effects exerted by TGFbeta consisted only of (i) neurofilament increase and (ii) morphological differentiation. The TGFbeta receptor pattern was different in each culture: SK-N-AS expressed low rates of type I and type II receptors and high rates of type III receptor; LAN-5 expressed high rates of type I, low rates of type II, and no type III; GL15 expressed high rates of all three receptors. These data suggest that TGFbeta can induce a histotype-specific cell maturation and that the neuroblastoma expressing low type II and at the same time lacking type III receptor responds only partially to TGFbeta, with induction of neural differentiation but without inhibition of cell growth.

Cell Differentiation↗
PubMed1996

Enhanced expression of heregulin in c-erb B-2 and c-Ha-ras transformed mouse and human mammary epithelial cells.

Heregulin beta 1 was found to stimulate the anchorage-dependent, serum-free growth of nontransformed human MCF-10A mammary epithelial cells. Unlike epidermal growth factor, transforming growth factor alpha, or amphiregulin, heregulin beta 1 was also able to induce the anchorage-independent growth of MCF-10A cells. In contrast, the anchorage-independent, serum-free growth of c-Ha-ras or c-erb B-2 transformed MCF-10A cells was unaffected by heregulin beta 1, whereas heregulin beta 1 was able to stimulate the anchorage-independent growth of these cells. c-Ha-ras or c-erb B-2 (c-neu) transformed MCF-10A or mouse NOG-8 mammary epithelial cells express elevated levels of 2.5, 5.0, 6.5, 6.8, and 8.5 kb heregulin mRNA transcripts and/or synthesize cell-associated 25, 29, 50, and 115 kDa isoforms of heregulin. Since the MCF-10A cells and transformants also express c-erb B-3, these data suggest that endogenous heregulin might function as an autocrine growth factor for Ha-ras or erb B-2 transformed mammary epithelial cells.

Animals↗
PubMed1995

Cysteinyl-leukotriene D4 induced IL-1 beta expression and release in rat vascular smooth muscle cells.

Vascular cells, including smooth muscle cells (VSMC), may release interleukin 1 (IL-1) and transcribe its genes for both isoforms. Previous studies have shown that cysteinyl-leukotrienes can modulate cytokine production by monocytes and a cytokine-eicosanoid network has been suggested during atherosclerosis. In this study the effects of cysteinyl-leukotriene D4 (LTD4) on IL-1 beta production and IL-1 beta mRNA expression were tested on rat VSMC. LTD4 showed a significant dose-dependent (from basal production of 55 +/- 15 pg/ml to maximal production of 177 +/- 14 pg/ml) and time-dependent (peaking at 24 h 16 +/- 54 pg/ml) increase of IL-1 beta immunoreactivity in the supernatants of conditioned medium and cell lysates. Furthermore, LTD4 induced an increased mRNA expression which began at 1 h and peaked at 12 h incubation time. The production of IL-1 beta was inhibited by MK-571 (from 145 +/- 12 to 60 +/- 10 pg/ml), a specific receptor antagonist of LTD4 and partially reduced by IL-1 receptor antagonist (IL-1 ra) (from 160 +/- 12 to 85 +/- 5 pg/ml). These experiments suggest that cysteinyl-leukotrienes, potentially produced in the vascular wall by leukocytes or by transcellular metabolism, may be involved in local IL-1 production.

Analysis of Variance↗
PubMed1995

Epithelial rat thyroid cell clones, escaping from transforming growth factor beta negative growth control, are still inhibited by this factor in the ability to trap iodide.

Transforming growth factor beta (TGF beta) acts on epithelial thyroid cells, negatively controlling their proliferation and functions. The effects of TGF beta on epithelial rat thyroid cells (FRTL-5) and on two TGF beta-resistant rat thyroid cell clones (FRTL-5H2 and FRTL-R) were investigated. FRTL-5H2 represents a rat thyroid cell clone overexpressing active erbB-2 oncogene, recently obtained after FRTL-5 cell infection with a retrovirus vector carrying the erbB-2 human oncogene (G. Mincione et al., Cancer Res., 53: 5548-5553, 1993). FRTL-R is a FRTL-5 subclone spontaneously isolated after long term in culture. FRTL-5H2 and FRTL-R cell clones were stimulated by TGF beta at the same concentration of 5 ng/ml that induced 70% inhibition of [3H]thymidine incorporation in control FRTL-5 thyroid cells. Nuclear events regulated by TGF beta, such as cyclin and cyclin-dependent kinase gene expression, were then analyzed. In FRTL-5 cells, TGF beta was found to reduce the expression of cdk2 and cyclin A genes; the same treatment did not modify nuclear gene expression in the resistant cell clones. TGF beta is known to reduce iodide uptake in thyroid cells; in both FRTL-5H2 and FRTL-R cells, TGF beta was found to inhibit the thyrotropin-induced iodide uptake. Thus, thyroid cell clones, resistant to the growth-inhibitory activity of TGF beta, were sensitive to TGF beta inhibition of iodide incorporation, suggesting that TGF beta activates divergent signaling pathways in these cells, separately controlling cell proliferation and differentiation parameters. Studies on TGF beta receptors showed similar amounts of TGF beta-binding species on FRTL-5 cells and TGF beta-resistant clones, while 125I-labeled TGF beta cross-linking experiments revealed differences; thus, the TGF beta-resistant cells showed a 40% decrease in the amount of labeled type II TGF beta receptor on the cell surface. However, this different pattern of TGF beta receptors cannot totally account for the shown TGF beta resistance to growth inhibition that might also be due to perturbation in signaling pathways.

Animals↗
PubMed1995

Vesical dysfunction in systemic sclerosis (scleroderma)

There have been only a few reports on the involvement of the urinary tract in patients with systemic sclerosis, a disease of the connective tissue characterized by thickening and fibrosis of the skin, abnormality of the small arteries, and involvement of the gastrointestinal tract, heart, lung and kidney. We report the urodynamic assessment and histological examination of 9 women with scleroderma. Three patients voided less than 100 ml. with a significant residual volume and 4 presented with detrusor areflexia during a filling cystometrogram. Histopathological examination in all patients with detrusor areflexia demonstrated the presence of arterial lesions and derangement of the capillary bed of the detrusor musculature. Our data provide evidence for the functional and histological involvement of the bladder in patients with systemic sclerosis.

Female↗
PubMed1994

Antisense peptides to the 43-57 region of acylphosphatase and to the 46-60 region of two isoenzymes of a low-M(r) phosphotyrosine protein phosphatase do not interact with the corresponding proteins.

Three peptides complementary to exposed regions of two low-M(r) phosphotyrosine protein phosphatase isoenzymes and of the acylphosphatase muscle isoenzyme have been synthesized. Each peptide was synthesized on two different types of resins; the peptides were anchored to the resins by amide linkages. The peptide resins were checked by amino acid analysis and Edman degradation and directly used for enzyme purification. Despite our attempts, none of the resins was able to bind significant amounts of the corresponding protein, indicating the lack of interaction between the three proteins and the corresponding complementary peptides. This result agrees with many other reports, confirming that the molecular-recognition theory has no general validity.

Acid Anhydride Hydrolases↗
PubMed1993

Loss of thyrotropin regulation and transforming growth factor beta-induced growth arrest in erbB-2 overexpressing rat thyroid cells.

Amplification of erbB-2 gene and overexpression of gp185erbB-2 gene product is found in approximately one-third of primary human breast and ovarian cancer. Overexpression of gp185erbB-2 was recently found in human papillary thyroid carcinomas, but not in thyroid follicular carcinomas or adenomas. The erbB-2 gene encodes a cell surface growth factor receptor with intrinsic tyrosine kinase activity. Wild type human erbB-2 has been shown to act as a potent oncogene when overexpressed in mouse fibroblasts. To test whether overexpression of normal human erbB-2 gene can transform epithelial differentiated rat thyroid cells, these cells were infected with a recombinant retroviral expression vector containing the erbB-2 protooncogene. Rat thyroid cells expressing high levels of gp185erbB-2 do not display a fully transformed and tumorigenic phenotype. However, the isolated cell clones that overexpress gp185erbB-2, show changes in their growth properties if compared to normal thyroid cells, since they can grow in absence of thyrotropin, the main growth factor controlling thyroid cell proliferation in vitro, and do not respond to the growth inhibitory effect of transforming growth factor beta.

Animals↗
PubMed1992

TGF beta inhibits rat thyroid cell proliferation without alterations in the expression of TSH-induced cell cycle-related genes.

Transforming growth factor beta (TGF beta) is a secreted polypeptide factor that is thought to play a major role in the regulation of proliferation of many cell types and various differentiation processes. TGF beta acts on thyroid cells by inhibiting cell proliferation and expression of differentiation markers, such as thyroglobulin production and iodide uptake. Exponentially growing thyroid cells cultures accumulate mostly in G0/G1 after exposure to TGF beta for 48 hours. TGF beta inhibits the TSH induced transition of quiescent thyroid cell from the G0 to the S phase. These effects on the thyroid cell growth, however, are not mediated by changes in the TSH-induced cell cycle-related genes expression; both immediate early and progression genes expression is unaffected by the TGF beta treatment.

Animals↗