Biomedical subjects

D Fabbro

Publications and source records attributed to D Fabbro.

At least 37 records · Page 2Linked to original sources

PubMed1998

Selective ceramide binding to protein kinase C-alpha and -delta isoenzymes in renal mesangial cells.

Ceramide is an important lipid second messenger produced by sphingolipid metabolism in cells exposed to a limited number of agonists and in turn triggers several cell responses in a protein kinase C (PKC)-dependent manner. Stimulation of mesangial cells with a radioiodinated photoaffinity labeling analogue of ceramide, (N-[3-[[[2-(125I)iodo-4-[3-(trifluoromethyl)-3H-diazirin-3-yl]benz yl] oxy]carbonyl]propanoyl]-D-erythro-sphingosine) ([125I]-TID-ceramide), defines PKC-alpha and PKC-delta as direct targets of ceramide. No binding of ceramide to PKC-epsilon and PKC-zeta could be detected. Moreover, TID-ceramide selectively binds to recombinant PKC-alpha and -delta but not to PKC-epsilon and -zeta isoenzymes. In vitro kinase activity assays reveal that only the binding of ceramide to PKC-alpha is accompanied by an increase in kinase activity. In contrast, there is no change in in vitro kinase activity of the other isoforms tested, i.e., PKC-delta, -epsilon, and -zeta, toward any of the conventional substrates tested. However, it is noteworthy that PKC-delta shows a decreased autophosphorylation upon ceramide binding. In vivo, activation of PKC-alpha by ceramide is monitored by a delayed translocation of the isoform from the cytosol to the membrane fraction, detectable after 1 h of stimulation. In contrast, neither PKC-delta, nor -epsilon nor -zeta is redistributed by ceramide. One functional cell response mediated by PKC-alpha in mesangial cells is a negative feedback regulation of ligand-stimulated phosphoinositide hydrolysis. When cells are pretreated with ceramide, ATP-induced inositol trisphosphate formation is time-dependently reduced. A maximal inhibition is observed after 2 h of ceramide exposure. In summary, these results suggest that ceramide selectively interacts with the alpha- and delta-isoforms of PKC in mesangial cells. Whereas PKC-alpha is activated with pronounced inhibition of hormone-stimulated phosphoinositide signaling, PKC-delta displays a decrease in its autophosphorylation, suggesting a negative role of ceramide binding on PKC-delta activity.

Amino Acid Sequence↗

PubMed1998

Syntheses and biological evaluation of indolocarbazoles, analogues of rebeccamycin, modified at the imide heterocycle.

A series of 10 indolocarbazole derivatives, analogues to the antitumor antibiotic rebeccamycin, bearing modifications at the imide heterocycle were synthesized. They bear an N-methyl imide, N-methyl amide, or anhydride function instead of the original imide. Their inhibitory potencies toward topoisomerase I were examined using a DNA relaxation assay and by analyzing the drug-induced cleavage of 32P-labeled DNA. Protein kinase C (PKC) inhibition and interaction with DNA were also studied together with the in vitro antiproliferative activities against B16 melanoma and P388 leukemia cells. The antimicrobial activities against two Gram-positive bacteria (Bacillus cereus and Streptomyces chartreusis), a Gram-negative bacterium (Escherichia coli), and a yeast (Candida albicans) were tested as well as their antiviral activities toward HIV-1. The efficiency of the anhydride compounds was compared to that of the parent compound rebeccamycin and its dechlorinated analogue. All the compounds studied were inactive against PKC. The structural requirements for PKC and topoisomerase I inhibition are markedly different. In sharp contrast with the structure-PKC inhibition relationships, we found that an anhydride function does not affect topoisomerase I inhibition, whereas a methyl group on the indole nitrogen prevents the poisoning of topoisomerase I. The compounds exhibiting a marked toxicity to P388 leukemia cells had little or no effect on the growth of P388CPT5 cells which are resistant to the topoisomerase I inhibitor camptothecin. This study reinforces the conclusion that the DNA-topoisomerase I cleavable complex is the primary cellular target of the indolocarbazoles and significantly contributes to their cytotoxicity and possibly to their weak but noticeable anti-HIV-1 activities. The structure-activity relationships are also discussed.

Aminoglycosides↗

PubMed1998

The binding properties and biological activities of Bcl-2 and Bax in cells exposed to apoptotic stimuli.

The oncogene product Bcl-2 protects cells from apoptosis whereas its homolog Bax functions to kill cells. Several binding partners of Bcl-2 and Bax have been isolated, but none of them has yet provided clues as to exactly how Bcl-2 and Bax work. According to one view, Bcl-2 and Bax interact with survival and death effector molecules, respectively, and neutralize each other through heterodimerization. Alternatively, Bcl-2 requires Bax for death protection, and additional proteins bind to the heterodimer to regulate its activity. Here we used a co-immunoprecipitation strategy to distinguish between these two possibilities. We show that the Bcl-2-Bax heterodimer is maintained, and no other protein associates stably in detectable amounts with Bcl-2, Bax, or the heterodimer in anti-Bcl-2 and anti-Bax immunoprecipitates from normal cells and cells exposed to apoptotic stimuli. Analysis of cells expressing various levels of Bcl-2 and Bax, however, revealed that the degree of protection against apoptosis does not correlate with the number of Bcl-2-Bax heterodimers but the amount of Bcl-2 that is free of Bax. In addition, the survival activity of Bcl-2 is unaffected when Bax expression is ablated by an antisense strategy. Our findings suggest that the Bcl-2-Bax heterodimer is a negative regulator of death protection, and that Bcl-2 requires neither Bax nor major, stable interactions with other cellular proteins to exert its survival function. We therefore propose that Bcl-2 acts as an enzyme (capturing substrates in a transient way), as a homodi- or multimer, or through the interaction with non-proteaceous targets (lipids, ions).

Animals↗

PubMed1998

Structural and functional properties of the N transcriptional activation domain of thyroid transcription factor-1: similarities with the acidic activation domains.

The thyroid transcription factor 1 (TTF-1) is a tissue-specific transcription factor involved in the development of thyroid and lung. TTF-1 contains two transcriptional activation domains (N and C domain). The primary amino acid sequence of the N domain does not show any typical characteristic of known transcriptional activation domains. In aqueous solution the N domain exists in a random-coil conformation. The increase of the milieu hydrophobicity, by the addition of trifluoroethanol, induces a considerable gain of alpha-helical structure. Acidic transcriptional activation domains are largely unstructured in solution, but, under hydrophobic conditions, folding into alpha-helices or beta-strands can be induced. Therefore our data indicate that the inducibility of alpha-helix by hydrophobic conditions is a property not restricted to acidic domains. Co-transfections experiments indicate that the acidic domain of herpes simplex virus protein VP16 (VP16) and the TTF-1 N domain are interchangeable and that a chimaeric protein, which combines VP16 linked to the DNA-binding domain of TTF-1, undergoes the same regulatory constraints that operate for the wild-type TTF-1. In addition, we demonstrate that the TTF-1 N domain possesses two typical properties of acidic activation domains: TBP (TATA-binding protein) binding and ability to activate transcription in yeast. Accordingly, the TTF-1 N domain is able to squelch the activity of the p65 acidic domain. Altogether, these structural and functional data suggest that a non-acidic transcriptional activation domain (TTF-1 N domain) activates transcription by using molecular mechanisms similar to those used by acidic domains. TTF-1 N domain and acidic domains define a family of proteins whose common property is to activate transcription through the use of mechanisms largely conserved during evolutionary development.

Amino Acid Sequence↗

PubMed1998

Effect of intensive glycaemic control on fibrinogen plasma concentrations in patients with Type II diabetes mellitus. Relation with beta-fibrinogen genotype.

Recent studies show that in diabetic subjects an increase of plasma fibrinogen concentration is associated with a high risk of cardiovascular complications. Environmental and genetic factors contribute to the plasma fibrinogen concentration. Several studies indicate a relation between the polymorphism in the 5' region of the beta-fibrinogen gene and plasma protein concentrations and in diabetes the possible influence of hyperglycaemia on fibrinogen is still debated. In this study we investigated these relations. Hind III polymorphism was evaluated by a polymerase chain reaction-technique. On the basis of the observed allelic combination of fibrinogen beta-gene polymorphism and the existence of poor metabolic control (glycated haemoglobin > or = 7.5%), 50 Type II diabetic patients were selected. They were divided into three groups according to their beta-gene polymorphism (alpha1alpha1: n = 20, alpha1alpha2: n = 15, alpha2alpha2: n = 15) and then intensive insulin therapy was started. After 3 months of intensive treatment, the improvement in glycaemic control was equivalent, in terms of glycated haemoglobin, in all the three groups. A fibrinogen reduction was observed in alpha1alpha2 and alpha2alpha2 but not in alpha1alpha1 subjects. These results underline a possible relation between fibrinogen genotypes and glycaemic control in determining plasma fibrinogen concentrations in diabetic patients.

Blood Glucose↗

PubMed1998

Syntheses, biochemical and biological evaluation of staurosporine analogues from the microbial metabolite rebeccamycin.

The indolocarbazole antibiotics staurosporine and rebeccamycin (1) are potent antitumor drugs targeting protein kinase C and topoisomerase I, respectively. To obtain staurosporine analogues from rebeccamycin, different structural modifications were performed: coupling of the sugar moiety to the second indole nitrogen, dechlorination and then reduction of the imide function to amide. The newly synthesized compounds (3-6) were tested for their abilities to bind to DNA and to inhibit topoisomerase I and protein kinase C. Their antiproliferative effects in vitro against B16 melanoma and P388 leukemia (including the related P388CPT cell line resistant to camptothecin) as well as their anti-HIV-1 and antimicrobial activities against various strains of microorganisms were determined. The cytotoxicity of the dechlorinated imide analogue 5 correlates well with its DNA binding and anti-topoisomerase I activities. These findings provide guidance for the development of new topoisomerase I-targeted antitumor indolocarbazoles equipped with a carbohydrate attached to the two indole nitrogens.

Aminoglycosides↗

PubMed1998

Pax-8 protein levels regulate thyroglobulin gene expression.

Pax proteins are transcription factors that control differentiation of several cell types. In adult organisms Pax-8 is expressed in the follicular thyroid cell where it interacts with sequences of thyroglobulin and thyroperoxidase promoters. In this study, we provide evidence indicating that Pax-8 protein levels regulate thyroglobulin gene transcription. The most critical approach consisted in increasing Pax-8 protein levels by transfecting thyroid cells with a Pax-8 expression vector. In this situation the thyroglobulin promoter transcriptional activity was significantly increased with respect to untransfected cells. In contrast, the transfection of thyroid transcription factor-1 (TTF-1) expression vector causes a modest decrease of thyroglobulin promoter activity, rather than an increase. Northern blots of human papillary cancers reveal a significant correlation between Pax-8 and thyroglobulin mRNAs. Gel-retardation assays suggest that the mechanism by which the Pax-8 protein levels modulate thyroglobulin promoter activity may occur through competition with TTF-1 for a common binding site. Since we also demonstrate that Pax-8 expression is subjected to TSH control, our data strongly suggest that Pax-8 protein levels could represent an important determinant for the regulation of thyroid cells.

Base Sequence↗

PubMed1998

Antitumor activity of a PKC-alpha antisense oligonucleotide in combination with standard chemotherapeutic agents against various human tumors transplanted into nude mice.

A 20-mer phosphorothioate oligodeoxynucleotide (ODN) directed against human protein kinase C-alpha (CGP 64128A = ISIS 3521) was analyzed for its antitumor activity either alone or in combination therapy. Combination studies with CGP 64128A and standard chemotherapeutic agents (cisplatin, mitomycin-C, vinblastine, estracyt and adriamycin) were performed in nude mice that had been transplanted s.c. with a variety of human tumors (breast, prostate, large cell lung and small cell lung carcinomas, and melanomas). Additive antitumor effects with CGP 64128A and the cytotoxins were found for half of the combinations studied. The combination of CGP 64128A with vinblastine or cisplatin showed superadditive antitumor activities against MCF-7 human breast carcinomas and PC3 prostate carcinomas with complete responses. CGP 64128A in combination with adriamycin resulted in superadditive antitumor effects against BT20 human breast carcinomas with complete tumor responses, and in combination with mitomycin-C superadditive antitumor effects with cures were observed against NCI-H460 human large cell carcinomas. The antitumor activity of CGP 64128A appeared to be due to a sequence-dependent mechanism of action as two 20-mer control ODNs were completely inactive as single agents against A549 and NCI-H69 human lung carcinomas. The antitumor activity of cisplatin against NCI-H69 human small cell lung carcinomas was slightly inhibited by one of the control ODNs, indicating that the superadditive antitumor activities of CGP 64128A in combination with cisplatin are the result of a sequence-dependent mechanism of action.

Animals↗

PubMed1997

Syntheses and biological activities (topoisomerase inhibition and antitumor and antimicrobial properties) of rebeccamycin analogues bearing modified sugar moieties and substituted on the imide nitrogen with a methyl group.

As a part of studies on structure-activity relationships, several potential topoisomerase I inhibitors were prepared. Different analogues of the antitumor antibiotic rebeccamycin substituted on the imide nitrogen with a methyl group were synthesized. These compounds bore either the sugar residue of rebeccamycin, with or without the chlorine atoms on the indole moieties, or modified sugar residues (galactopyranosyl, glucopyranosyl, or fucopyranosyl) linked to the aglycone via a beta- or alpha-N-glycosidic bond. Their inhibitory properties toward protein kinase C, topoisomerase I, and topoisomerase II were examined, and their DNA-binding properties were investigated. Their in vitro antitumor activities against murine B16 melanoma and P388 leukemia cells were determined. Their antimicrobial activities were tested against Gram-positive bacteria Bacillus cereus and Streptomyces chartreusis, Gram-negative bacterium Escherichia coli, and yeast Candida albicans. These compounds are inactive toward topoisomerase II but inhibit topoisomerase I. A substitution with a methyl group on the imide nitrogen led to a loss of proteine kinase C inhibition in the maleimide indolocarbazole series but did not prevent topoisomerase I inhibition. Compounds possessing a beta-N-glycosidic bond, which fully intercalated into DNA, were more efficient at inhibiting topoisomerase I than their analogues with an alpha-N-glycosidic bond; however, both were equally toxic toward P388 leukemia cells. Dechlorinated rebeccamycin possessing a methyl group on the imide nitrogen was about 10 times more efficient in terms of cytotoxicity and inhibition of topoisomerase I than the natural metabolite.

Aminoglycosides↗

PubMed1997

Induction of apoptosis in small-cell lung cancer cells by an antisense oligodeoxynucleotide targeting the Bcl-2 coding sequence.

BACKGROUND: The emergence of resistance to chemotherapy remains a major problem in the treatment of patients with small-cell lung cancer. Elevated expression of Bcl-2, a protein that inhibits programmed cell death or apoptosis, has been associated with radiation and drug resistance and has been observed in the majority of small-cell lung cancer specimens and cell lines. PURPOSE: To test the hypothesis that Bcl-2 expression levels are critical for inhibiting apoptosis in small-cell lung cancer cells, we used an antisense strategy to reduce Bcl-2 expression in these cells in an attempt to restore the natural occurrence of apoptosis. METHODS: Thirteen antisense oligodeoxynucleotides (ODNs) targeting various regions of the bcl-2 messenger RNA and a control scrambled-sequence ODN were tested to identify the most effective sequence(s) for reducing Bcl-2 protein levels. Northern and western blot analyses were used to examine basal bcl-2 messenger RNA and protein levels, respectively, in four human small-cell lung cancer cell lines (SW2, NCI-H69, NCI-H82, and NCI-N417). SW2 cells were treated with the antisense ODNs in the presence of cationic lipids (to facilitate uptake), and cytotoxic effects were measured by use of a cell viability assay. Flow cytometric analysis of DNA fragmentation and cell morphology was also performed. The cytotoxic effect of the most potent antisense ODN was also tested on the three other cell lines. RESULTS: The viability of SW2 cells was effectively reduced by ODNs that targeted the translation initiation and termination sites of the bcl-2 messenger RNA, but ODN 2009 that targeted the coding region was the most cytotoxic. Treatment of SW2 cells with 0.15 microM ODN 2009 for 96 hours reduced their viability by 91% (95% confidence interval [CI] = 88%-94%) and caused a dose-dependent reduction in Bcl-2 levels that became detectable 24 hours after treatment and persisted up to 96 hours; analysis of cellular morphology demonstrated that viability was reduced through apoptosis. Moreover, ODN 2009 at 0.15 microM was cytotoxic to NCI-H69, NCI-H82, and NCI-N417 cells, resulting in decreases in cell viability of 82% (95% CI = 78%-86%), 100%, and 100%, respectively, after 96 hours of treatment. The cytotoxic effects were inversely correlated with the basal Bcl-2 levels in the cell lines (r = -9964). A control scrambled-sequence oligodeoxynucleotide had no statistically significant effect on the cell lines (P values ranging from .38 to .89). CONCLUSION: We have identified a novel antisense ODN sequence (ODN 2009) that effectively reduces the viability of small-cell lung cancer cells by reducing Bcl-2 levels and facilitating apoptosis.

Antineoplastic Agents↗

PubMed1997

Functional interference between contacting amino acids of homeodomains.

In a protein, the function of an amino acid at some position depends on the amino acids at other positions. Here we demonstrate a functional interference between base-contacting amino acids (at positions 50 and 54) of homeodomains. When, in the context of Antennapedia or Goosecoid homeodomains, Lys50 is paired to Tyr54 or Ala54 and Gln50 is paired to Met54, the resulting proteins efficiently discriminate among different DNA sequences. In contrast, in the presence of the pair Lys50-Met54, both homeodomains show a reduced capability to discriminate among different DNA sequences. Sequence selection experiments performed in the context of the Goosecoid homeodomain suggest that the presence of Met54 precludes the base-discriminating function of Lys50. These results may explain why the pair Lys50-Met54 is never found in natural homeodomains.

Amino Acid Sequence↗

PubMed1997

Activation of protein kinase C subtypes alpha, gamma, delta, epsilon, zeta, and eta by tumor-promoting and nontumor-promoting agents.

Protein kinase C (PKC) subtypes alpha, gamma, delta, epsilon, zeta, and eta have been expressed using the baculovirus expression system. The partially purified PKC subtypes have been studied for their substrate specificities and phospholipid-independent activation by various chemically different nontumor- and tumor-promoting agents, as well as their inhibition of kinase activity by staurosporine and two related compounds. An endogenous PKC-like kinase activity of Sf9 cells was detected and analyzed for cofactor requirements and inhibition. Protamine sulfate was most efficiently phosphorylated by all of the PKC subtypes tested, although this phosphorylation was independent of phosphatidylserine (PS) and diacylglycerol (DAG) or 12-O-tetradecanoylphorbol 13-acetate (TPA). Except for PKC-zeta, all subtypes tested phosphorylated myelin basic protein (MBP), histone, or a peptide derived from the pseudosubstrate region of PKC-alpha in a PS/DAG-dependent manner but to varying extents. Among the various agents tested, TPA most efficiently stimulated the kinase activities of the PKC subtypes in a phospholipid-dependent manner. Phorbol 12,13-dibutyrate (PDBu) was less effective than TPA but displayed no major difference among the subtypes. Activation of PKC-alpha by bryostatin-1 reached only half of the TPA response whereas the other subtypes were activated more effectively. The weak tumor promoter resiniferonol 9,13,14-orthophenyl acetate (ROPA) mainly stimulated PKC-alpha and PKC-gamma at 1 microM concentration, whereas PKC-epsilon and PKC-eta were much less activated. Sapintoxin D, mezerein, indolactam V, and resiniferatoxin at concentrations of 1-100 nM preferentially activated PKC-alpha in a DAG-like manner, whereas at 1 microM other subtypes were activated as well. Preferential activation of PKC-alpha was also noted for tinyatoxin and thapsigargin, but their mode of activation is unclear because these two compounds did not compete for the phorbol ester binding of the PKC subtypes as the other agents did. Of the three PKC inhibitors tested, staurosporine most efficiently inhibited kinase activity of the PKC subtypes, whereas K252a and CGP 41251 were at least 10 times less effective. However, K252a showed certain specificity for inhibition of PKC-alpha, and CGP 41251 failed to inhibit PKC-epsilon and PKC-zeta. Given the different substrate specificities and modes of activation by various tumor-promoting and nontumor-promoting agents, as well as the different sensitivities towards different inhibitors, our results indicate a divergence of individual PKC subtypes in signal transduction.

Animals↗

PubMed1997

Simplified determination of combined residues of prochloraz and its metabolites in vegetable, fruit and wheat samples by gas chromatography.

This paper describes a gas chromatographic method for the determination of combined residues of prochloraz and its metabolites in various plant materials, including vegetables, fruits, seeds, grains and roots. The method is much simpler than previous ones as it does not require steam distillation; it consists of preliminary extraction of the sample, hydrolysis with pyridine hydrochloride in a test tube, extraction of the 2,4,6-trichlorophenol without the aid of steam distillation/continuous extraction, derivatization with diazomethane and gas chromatographic determination using electron-capture detection. The method is very rapid and can be done in a test tube; detection limit is from 0.01 to 0.50 mg/kg; recoveries are dependent on concentration of analyte and sample matrix; the determined values were found in the region of 72-102%.

Chromatography, Gas↗

PubMed1997

Feedback regulation of extracellular ATP-stimulated phosphoinositide hydrolysis by protein kinase C-alpha in bovine glomerular endothelial cells.

In glomerular endothelial cells, extracellular ATP stimulates a phospholipase C with subsequent hydrolysis of polyphosphoinositides and an increase in cytosolic free Ca2+ concentration ([Ca2+]i). Short-term (30 min) pretreatment of endothelial cells with 12-O-tetradecanoylphorbol 13-acetate (TPA), a potent activator of protein kinase C (PKC), decreases the ATP-stimulated phosphoinositide degradation and Ca2+ mobilization. However, this inhibition was lost after incubating the cells for four hours with TPA. Longer-term pretreatment (10 to 48 hr) even potentiated ATP-induced phosphoinositide breakdown and Ca2+ mobilization. In addition, pretreating the cells for 30 minutes with the specific PKC inhibitor Ro 31-8220 dose-dependently increased ATP-stimulated phosphoinositide hydrolysis, thus clearly indicating a regulatory role for PKC in the inositol lipid signaling pathway in glomerular endothelial cells. By using specific antibodies recognizing the different PKC isoenzymes, it is observed that glomerular endothelial cells express five isoenzymes: PKC-alpha, -delta, -epsilon, -zeta and -theta. No PKC-beta, -gamma, -eta and -mu isoenzymes were detected. On exposure to TPA, a complete depletion of PKC-alpha is observed within four hours. In contrast, PKC-epsilon was more resistant to phorbol ester, and even after 48 hours of TPA treatment, only 60% of PKC-epsilon was down-regulated. PKC-theta decreased very slowly from the cytosol (47% left after 24 hr of phorbol ester treatment) and translocated to the Triton X100-insoluble fraction. Moreover, PKC-delta and PKC-zeta were not significantly affected by 48 hours of phorbol ester incubation. Thus, only PKC-alpha is depleted with a kinetic that corresponds to the loss of feedback inhibition of ATP-stimulated phosphoinositide turnover. In the next step, [Ca2+]i changes were measured in single cells loaded with Fura-2 after microinjection of neutralizing PKC isoenzyme-specific antibodies. Injection of antibodies specific for PKC-alpha potently increased Ca2+ mobilization in response to ATP stimulation when compared to cells injected with buffer only or antibodies specific for PKC-epsilon. These results provide evidence that PKC-alpha mediates feedback inhibition of ATP-stimulated phosphoinositide hydrolysis in glomerular endothelial cells.

Adenosine Triphosphate↗

PubMed1997

Expression of differentiation markers in cultured cells from various thyroid diseases.

The use of cell cultures as a model system for studying thyroid diseases requires establishment of appropriate culture conditions that allow in vitro propagation of populations that correspond to in vivo ones. We have defined these conditions and verified functional parameters such as thyrotropin-dependent cyclic adenosine monophosphate (cAMP) production and thymidine incorporation, and molecular markers such as thyroglobulin (by radioimmunoassay [RIA] and Northern blot), thyroperoxidase (by Northern blot), thyroid-specific transcription factor 1 (by immunohistochemistry and Northern blot) and PAX-8 (by Northern blot). The "in vitro profile" (functional parameters and molecular markers) was found to correlate with the degree of differentiation of the starting specimens and the pathological diagnosis. The data presented suggest that our culture technique allows in vitro growth of cell populations that may be used to perform functional assays and may facilitate the molecular characterization of pathological samples. This approach could be especially useful to define prognosis and also help to develop innovative therapies.

Biomarkers↗

PubMed1997

Immunocytochemical expression of tissue specific transcription factor-1 in lung carcinoma.

AIMS: To investigate the immunocytochemical expression of the tissue specific transcription factor-1 (TTF-1) on cytological specimens of small cell lung carcinoma (SCLC) and to establish its value in the cytological diagnosis of lung cancer. METHODS: For each case, the diagnosis was made on cytological specimens and confirmed on subsequent bronchial biopsy specimens. TTF-1 was detected immunocytochemically using the avidinbiotin complex technique with a rabbit antiserum. Expression of TTF-1 was evaluated in 41 cases of SCLC and 17 cases of non-small cell carcinoma (NSCC). The latter were subdivided into eight cases of adenocarcinomas and nine cases of squamous cell carcinomas (SCC). RESULTS: Positive nuclear immunoreactivity to TTF-1 was identified in 38 (92.7%) of the 41 cases of SCLC, in five (62.5%) of eight cases of adenocarcinoma, and one (11%) of nine cases of SCC. A significant difference was observed between the two main groups, SCLC and NSCC. A comparison between SCLC and adenocarcinoma and SCC showed that TTF-1 expression was significantly different. TTF-1 immunoreactivity was not detected in the inflammatory cells of the same cases. CONCLUSIONS: TTF-1 is strictly associated with SCLC; it was weakly expressed in the various subtypes of NSCC. Although TTF-1 is not specific for SCLC, it can be used to highlight neoplastic cells to good effect when a large inflammatory component is present, and to differentiate SCLC from lymphoid infiltrates.

Adenocarcinoma↗

PubMed1997

Synthesis and biological evaluation of monoindolyl and indolocarbazolyl oxazolones and imidazolones.

Eight compounds structurally related to protein kinase C inhibitor MDL 27032 and substituted with indole moieties were synthesized. Their activities towards protein kinase C (PKC) and protein kinase A (PKA) were determined. Their effect on PKC-mediated contraction of rat tracheal smooth muscle, their antiproliferative activity on two murine tumor cell lines, melanoma B16 and leukemia P388 and their antimicrobial activity on a gram-positive bacterium Bacillus cereus were also examined. The mammalian and bacterial cell antiproliferative activity, as well as vasorelaxant effect, observed for some of them could not be correlated to PKC or PKA inhibition. Only bulky bis-indolyl compounds exhibited biological activity in these experiments. Rigid indolocarbazoles had the strongest antiproliferative activity.

Animals↗

PubMed1997

In vitro cultures of pathological thyroid cells: a powerful diagnostic technique.

In order to get a correct predictivity from molecular and functional markers in neoplastic disease, cell cultures, correspondent to in vivo existing populations, should be available. The difficulty, as yet, to correlate in vivo conditions with in vitro molecular and functional markers, represents a hurdle for a better prognosis in several neoplastic diseases. We tackled the problem establishing cultures from surgical samples of human thyroid glands bearing various pathologies (pathological diagnosis were obtained for all samples). Cells were frozen after 2 passages, and molecular markers (thyroglobulin, TPO, TTF-1 and PAX-8) and functional parameters (TSH-dependent cAMP production and thymidine incorporation) were investigated after thawing. The "in vitro profile" (functional parameters and molecular markers) was found to correlate with the pathological diagnosis and the degree of differentiation of the starting specimens. The data presented suggest that our culture technique allows in vitro growth of cell populations that may be used to perform functional assays and may make the molecular characterization of pathological samples easier. These findings could be especially useful to better define prognosis and also help to develop innovative therapies.

Journal Article↗