Biomedical subjects

F Perini

Publications and source records attributed to F Perini.

At least 37 records · Page 2Linked to original sources

PubMed1995

A novel adhesion factor produced by hamster pancreatic cancer cell line is effective on normal and carcinoma cell lines of different species.

An adhesion factor, produced by the hamster pancreatic cancer cell line PC-1.0, was tested for its efficiency in promoting the in vitro adhesion of normal and tumor cells (pancreas, lung, kidney, colon, breast, skin, prostate, neuroblast, melanocyte) derived from human, monkey, bovine, hamster, and rat sources. Using a modification of the dimethylthyazol diphenyl tetrazolium (MTT) assay, the factor was found to induce adhesion in all cell lines in a dose-dependent manner. Although the effect was variously expressed, there was a statistically significant difference between the MTT absorbance of cells incubated in the presence or absence of the factor. Conditioned medium of each cell line tested showed significantly less adhesion effect than that produced by PC-1.0 cells. Because our previous study indicated that the adhesion factor produced by PC-1.0 cells differed from known growth factors and adhesion molecules including fibronectin, vibronectin, laminin, and collagen, it appears that PC-1.0 cells produce a novel adhesion factor that enhances adherence of normal and malignant cells of different species.

Animals↗

PubMed1995

The synthesis and compositional analysis of phosphopeptides.

Protein phosphorylation plays a critical role in the regulation of cell growth and differentiation. There is considerable interest, therefore, in the facile synthesis of peptides that possess selectively phosphorylated residues for use as molecular probes in mechanistic studies of the biological consequences of phosphorylation. This work will review the various synthetic protocols used in the generation of phosphopeptides and will discuss their characterization by amino acid compositional analysis.

Amino Acids↗

PubMed1995

Platelet and plasma levels of glutamate and glutamine in migraine with and without aura.

We evaluated plasma and platelet glutamate and glutamine levels in migraine with and without aura during headache-free periods and compared the results with those of normal controls. The plasma and platelet levels of glutamine in migraine with and without aura were normal. Migraine without aura patients had higher glutamate levels in plasma, and normal platelet levels. In migraine with aura patients, glutamate levels were high in platelets, but not in plasma. This suggests different profiles of excitatory amino acid metabolism in migraine with and without aura.

Adolescent↗

PubMed1995

Platelet models and their possible usefulness in the study of migraine pathogenesis.

Platelets may be linked to migraine. On the one hand they are activated during the migraine attack and thus may participate in the pathogenesis of the disorder (the nature of this activation is still unknown). In order to understand this platelet anomaly, we discuss the data available in the literature. In particular, we review recent in vitro studies of alpha-granules and dense bodies secretion, and aggregation induced by collagen and PAF. On the other hand, platelets share many metabolic characteristics with serotonergic neurons and endothelial cells. Accordingly, platelets have been used to investigate the possible role of serotonin turnover and nitric oxide function in migraine. In both cases, the data obtained have shown peculiar abnormalities that may explain pathogenetic and clinical aspects of primary headache.

Blood Platelets↗

PubMed1995

Efficacy of transdermal clonidine in short-term treatment of cluster headache: a pilot study.

Some clinical as well as pharmacological indications seem to suggest that a reduction of the noradrenergic tone occurs in cluster headache (CH), during both the active and remission periods. But sharp fluctuations of the sympathetic system may trigger the attacks. Clonidine, an alpha-2-adrenergic presynaptic agonist, regulates the sympathetic tone in the central nervous system. Therefore, a continuous administration of low-dose clonidine could be potentially beneficial in the active phase of CH by antagonizing the variations in noradrenergic tone. After a run-in week, we administered transdermal clonidine (5 - 7.5 mg) for one week to 13 patients suffering from CH, either episodic (8 cases) or chronic (5 cases). During clonidine treatment, the mean weekly frequency of attacks dropped from 17.7 +/- 7.0 to 8.7 +/- 6.6 (p = 0.0005), the pain intensity of attacks measured on the visual analogue scale from 98.0 +/- 7.2 to 41.1 +/- 36.1 mm (p = 0.001), and the duration from 59.3 +/- 21.9 to 34.3 +/- 24.6 min (p = 0.02). This open pilot study strongly suggests that transdermal clonidine may be an effective drug in the preventive treatment of CH. Its efficacy is possibly due to its central sympatho-inhibition, which reduces or prevents the occurrence of fluctuations of noradrenaline release that may induce the attacks.

Administration, Cutaneous↗

PubMed1995

A linear 23-residue peptide reveals a propensity to form an unusual native-like conformation.

To gain insight into the earliest events of protein folding, a 23-residue peptide with a sequence corresponding to the 38-60 fragment of the beta-subunit of human chorionic gonadotropin (hCG beta) was studied by NMR. In aqueous solution the majority of the peptide residues adopted an extended polyproline II (PII) conformation similar to those in mature, fully folded hCG beta. The finding that the isolated protein fragment may acquire native-like structural motifs, even without alpha-helices or beta-structures, extends the possibility of using free peptides as model systems to better understand the protein folding mechanisms. It was shown that the PII-rich structural motif can be determined efficiently by NMR spectroscopy. The observation that in the absence of extensive medium- and long-range interactions the majority of amino acid residues may adopt the PII conformation suggests that the PII-rich structural motifs may play an important role in early events of protein folding.

Base Sequence↗

PubMed1995

Metabolism and menstrual cycle rhythmicity of serotonin in primary headaches.

We investigated the platelet and plasma levels of serotonin and its metabolite, 5-hydroxyindoleacetic acid, in patients suffering from episodic tension-type headache and migraine with and without aura, during a headache-free period. In female subjects, blood samples were drawn during the follicular, ovulatory, and late luteal phases of the menstrual cycle. In tension headache and migraine with aura, the group mean values of serotonin and 5-hydroxyindoleacetic acid in platelets and plasma were significantly increased, but migraine without aura patients' levels were normal. The pattern of menstrual cycle-related fluctuations in platelet serotonin levels were similar in female patients with tension headache and in controls, with a maximum value in the follicular phase. In both migraine groups, in contrast, the peak occurred in the ovulatory phase. The results are discussed in view of whether these aberrations in peripheral markers of the metabolism and menstrual cycle-related rhythmicity of serotonin may reflect similar alterations in the central nervous system.

Adolescent↗

PubMed1995

[Combined treatment of stage I and II gastric lymphoma].

From 1981 to 1991, fifty-three patients with primary gastric lymphoma were referred to our Institute and submitted to a complete pathologic staging followed by gastric resection, i.e., total or partial gastrectomy. According to the Working Formulation criteria, 14 patients (26.4%) were affected with high grade lymphomas, 22 (41.5%) with intermediate lymphomas and 17 (32.1%) with low grade lymphomas. MALT (mucosa associated lymphoid tissue) lymphomas were observed in > 50% of cases. At the pathologic examination of the surgical specimens, infiltration depth was assessed, according to TNM criteria, in the patients whose disease was limited to the muscular gastric wall (T1-T2) and in those whose disease spread to the serosa or beyond it (T3-T4). Twenty-five patients were classified as stage I and 28 as stage II (9 of them in stage II E1 and 19 in stage II E2 according to Musshoff's classification). The treatment protocol of these patients was as follows: stage I patients (T1-T2) with normal surgical resection margins underwent no adjuvant treatment (10 patients); stage I patients (T1-T2) with resection margins infiltrated by the disease were submitted to local irradiation; stage I (T3-T4) and stage II E1 patients underwent large-field postoperative irradiation (14 patients); stage II E2 patients (n = 19) received conventional chemotherapy (CHOP, F-CVP, N-CVP): the ones who failed to reach complete remission or presented with bulky disease at diagnosis completed the treatment with large-field irradiation (10 patients). The disease-free survival (86.5%) and the overall survival (96%) rates of stage I patients exhibited no significant difference relative to stage II patients (DFS: 87% and OS: 90%). The analysis of relapses relative to disease extent demonstrated that this parameter is more significant to prognosis than nodal involvement (T1-T2: 2/39 relapses, 5.1%; T3-T4: 4/14 relapses, 28.6%).

Adult↗

PubMed1994

CEP regimen (CCNU, etoposide, prednimustine) for relapsed/refractory Hodgkin's disease.

AIMS AND BACKGROUND: Although initial treatment of Hodgkin's disease induces a complete remission in most patients, approximately 50% of patients with advanced disease will not achieve a complete remission or will relapse following the first complete remission. PATIENTS AND METHODS: Twenty-three patients with relapsed/resistant Hodgkin's disease, observed between January 1991 and October 1993, underwent CEP combination chemotherapy (CCNU, etoposide, prednimustine). All patients had previously received MOPP and ABVD regimens, in combination at diagnosis or sequentially (at diagnosis and at the first relapse). RESULTS: Thirteen (56%) patients achieved complete responses and 4 (18%) had partial responses. Two partial responders obtained a complete remission after a successive autologous bone marrow transplantation. The complete remission was not influenced by the timing of MOPP and ABVD treatments, presence of extranodal involvement or presence of bulky disease, but was affected by the presence of a primary disease refractory to the first standard programs. All the complete responders but 2 were alive and relapse-free at a median follow-up of 15 months; no major toxic effects were recorded. CONCLUSIONS: These data suggest, as did those of other studies, that CEP is an effective regimen in patients with Hodgkin's disease in first or second relapse, also to reduce the tumor burden and to determine chemosensitivity before contingent bone marrow or peripheral blood stem cell support.

Adult↗

PubMed1994

N-acetylgalactosamine glycosylation of MUC1 tandem repeat peptides by pancreatic tumor cell extracts.

Synthetic peptides corresponding to the human mucin MUC1 tandem repeat domain (20 residues) were glycosylated in vitro by using UDP-N-[3H]acetyl-D-galactosamine (GalNAc) and lysates of pancreatic tumor cell lines. Results obtained with peptides of different lengths (from one to five repeats) suggest that increasing the number of tandem repeats has neither a positive nor a negative effect on the density of glycosylation along the MUC1 tandem repeat protein backbone. Purified glycopeptides were sequenced on a gas-phase sequencer, and glycosylated positions were determined by measuring the incorporated radioactivity in fractions collected following each round of Edman degradation. The results showed that two of three threonine residues on the MUC1 tandem repeat peptides were glycosylated by pancreatic tumor cell lysates at the following positons: GVTSAPDTRPAPGSTAPPAH (underlined T indicates position of GalNAc attachment). None of the serine residues were glycosylated. Determination of the mass of the glycopeptides by mass spectrometry confirmed that a maximum of two molecules of GalNAc were covalently linked to each 20-residue repeat unit in the peptides. The data presented here show that acceptor substrate specificity of the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase detected in lysates of pancreatic and breast tumor cell lines is identical and is limited to some but not all threonines in the MUC1 tandem repeat peptide sequence. The influence of primary amino acid sequence on acceptor substrate activity was evaluated by using several peptides that contain single or double amino acid substitutions (relative to the native human MUC1 sequence). These included substitutions in the residues that were glycosylated and substitutions of the surrounding primary amino acid sequence. The results of these studies suggest that primary amino acid sequence, length, and relative position of the residue to be glycosylated dramatically affect the ability of peptides to serve as acceptor substrates for the UDP-GalNAc:polypeptide N-acetylgalatosaminyltransferase.

Acetylgalactosamine↗

PubMed1994

Influence of acceptor substrate primary amino acid sequence on the activity of human UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyltransferase. Studies with the MUC1 tandem repeat.

Synthetic peptides (30 and 20 residues long) corresponding to the native MUC1 tandem repeat sequence (20 residues long) were glycosylated in vitro using UDP-[3H]GalNAc and lysates from the human breast tumor cell line MCF7. Purified glycopeptides were sequenced on a gas-phase sequenator, and glycosylated positions were determined by measuring the incorporated radioactivity in fractions collected following each round of Edman degradation. The results showed that 2 of 3 threonines on the MUC1 tandem repeat peptides were glycosylated at the following positions: GVTSAPDTRPAPGSTAPPAH (underlined Thr residues indicate positions of GalNAc attachment); no glycosylation of serine residues was detected. Determination of the mass of the glycopeptides by mass spectrometry showed that a maximum of two molecules of GalNAc were covalently linked to each 20-residue repeat unit in the peptides. The influence of substrate primary amino acid sequence in determining the substrate specificity of UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyl-transferase activity was evaluated using as acceptor substrates a series of overlapping 9-residue peptides that represent a moving set through the tandem repeat of the MUC1 mucin. In addition, the influence of primary amino acid sequence on acceptor substrate activity was evaluated using several peptides that contained single or double amino acid substitutions (relative to the native human MUC1 sequence). These included substitutions in the residues that were glycosylated and substitutions in the surrounding primary amino acid sequence. This study demonstrates that primary amino acid sequence, length, and relative position of the residue to be glycosylated dramatically affect the ability of peptides to serve as acceptor substrates for UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyltransferase.

Amino Acid Sequence↗

PubMed1994

Decreased collagen-induced platelet aggregation and increased platelet arginine levels in migraine: a possible link with the NO pathway.

We studied whole blood platelet aggregation induced by collagen, platelet activating factor (PAF) and measured basal platelet L-arginine (L-arg) levels, as an indirect index of the nitric oxide (NO) pathway in migraine. Migraine, both with and without aura groups, showed a reduced aggregation to collagen, but not to PAF, compared with control subjects. Platelet L-arg levels were significantly increased in migraine with aura sufferers, whereas the plasma levels were in the same range in migraineurs and controls. Platelet hyperesponsiveness to collagen stimulation in migraine may be linked to an increased availability of the amino acid precursor and an abnormal NO synthesis.

Adolescent↗

PubMed1994

Platelet secretion from dense and alpha-granules in vitro in migraine with or without aura.

Several studies in vivo indicate platelet activation in migraine, as reflected by increased plasma concentrations of platelet secretory products. In vitro data on platelet secretion are scant, which prompted an investigation into agonist-induced platelet aggregation and secretion in platelets from patients with migraine. Sixty two patients with migraine with aura (MA) and 41 with migraine without aura (MwA) were studied during a headache-free phase, together with 26 healthy controls. Platelet aggregation and secretion in platelet-rich plasma were induced by collagen and platelet activating factor (PAF). Serotonin was measured by high performance liquid chromatography and platelet factor 4 (PF4) with an enzyme immunoassay kit. There were no significant aberrations in platelet aggregation in those with migraine compared with healthy controls. The platelet PF4 secretion induced by PAF (1.0 and 0.1 microM) was increased in MwA (p < 0.05, p < 0.0001) compared with controls, and there was a similar trend in MA (NS, p < 0.01). By contrast, the PF4 secretion induced by collagen (0.5 and 2.0 micrograms/ml) was reduced in MA (p < 0.01 and p < 0.05). Further, the MA group exhibited increased basal intraplatelet serotonin concentrations (p < 0.0001) and increased serotonin secretion induced by both concentrations of collagen (p < 0.0001) and PAF (p < 0.001). The data indicate an abnormal platelet a-granule secretion in those with migraine, and focus attention on PAF as a possible factor contributing to the platelet activation associated with migraine. The increased platelet content and secretion of serotonin was specific to MA, and may reflect different serotonin turnover in the two clinical migraine types.

Adolescent↗

PubMed1993

Protein folding and assembly in vitro parallel intracellular folding and assembly. Catalysis of folding and assembly of the human chorionic gonadotropin alpha beta dimer by protein disulfide isomerase.

Although purified proteins will refold and assemble in vitro, it is not known if cellular factors change the mechanisms of these processes. Based on the gel migration of folding intermediates, the kinetic relationships between these intermediates, and on the order of formation of six disulfide bonds, we have found that the in vitro folding pathway of the human chorionic gonadotropin beta subunit (hCG-beta) is indistinguishable from the intracellular folding pathway. The same rate-limiting event was found in both folding environments; however, the t1/2 for this step in a cell is 4 min, whereas in vitro the t1/2 was > or = 80 min. Protein disulfide isomerase (PDI) increased the in vitro rate of this event (t1/2 = 25 min) without changing the order of disulfide bond formation. PDI also catalyzed the in vitro rate of assembly of hCG subunits. In intact cells, assembly of the alpha beta heterodimer occurs before all of the intramolecular disulfide bonds of beta are formed. In vitro, assembly was increased after reduction of two of the carboxyl-terminal disulfide bonds of hCG-beta by PDI. These results strongly suggest that both in intact cells and in vitro, partially unfolded hCG-beta is more assembly-competent than is fully folded hCG-beta. The comparison of in vitro and intracellular hCG-beta folding and hCG subunit assembly which is shown in this report indicates that the assisted folding and assembly pathway that occurs in cells, where proteins such as PDI play a role, differs only in rate but not in the order of disulfide bond formation or in the precursor-product relationships among the folding intermediates.

Animals↗

PubMed1993

Increased platelet serotonin content and hypersecretion from dense and alpha-granules in vitro in tension-type headache.

We investigated platelet aggregation and secretion from dense and alpha-granules in vitro in 28 tension-type headache (TH) patients and 26 healthy controls. We also measured basal platelet serotonin levels. Platelet aggregation was normal in TH, but the secretion of serotonin and platelet factor 4 (PF4) was significantly increased in response to 0.5 and 2.0 micrograms/ml collagen and to 1.0 mumol/l PAF. The basal platelet serotonin levels were also higher in patients than in controls. The mechanisms of platelet hypersecretion remain to be determined, but the increased secretion of serotonin is probably in part related to the increased basal levels. The increased platelet serotonin in TH patients may reflect an enhanced serotonin turnover.

Adolescent↗

PubMed1992

Domain-dependent protein folding is indicated by the intracellular kinetics of disulfide bond formation of human chorionic gonadotropin beta subunit.

We have measured the intracellular rates of formation of the six disulfide bonds in the human chorionic gonadotropin beta subunit (hCG-beta) to determine whether the folding pathway of this molecule can be described by a simple sequential model. If such a model is correct, the formation of disulfide bonds, which is indicative of tertiary structural changes during protein folding, should occur in a discrete order. The individual rates of disulfide bridging were determined by identifying the extent of disulfide bond formation in hCG-beta intermediates purified from choriocarcinoma cells that had been metabolically labeled for 40 to 120 s and chased for 0 to 25 min. The results of these kinetic studies describe a folding pathway in which the disulfide bonds between cysteines 34-88, 38-57, 9-90 and 23-72 stabilize, in a discrete order, the putative domain(s) involving amino acids 1-90 of hCG-beta. However, the S-S bonds 93-100 and 26-110 begin to form before the complete formation of the disulfide bonds that stabilize the amino acid 1-90 domain(s), and continue to form after complete formation of these disulfide bonds, suggesting that hCG-beta does not fold by a simple sequential pathway. The order of completion of each of the six disulfide bonds of hCG-beta is: 34-88 (t1/2 = 1-2 min), 38-57 (t1/2 = 2-3 min), 9-90 and 23-72, 93-100, and 26-110. Moreover, 60-100% of each of the six disulfide bonds form posttranslationally, and nonnative disulfide bonds do not form in detectable amounts during intracellular folding of hCG-beta.

Choriocarcinoma↗

PubMed1992

Intracellular folding pathway of human chorionic gonadotropin beta subunit.

Few experimental models have been used to investigate how proteins fold inside a cell. Using the formation of disulfide bonds as an index of conformational changes during protein folding, we have developed a unique system to determine the intracellular folding pathway of the beta subunit of human chorionic gonadotropin (hCG). Three folding intermediates of the beta subunit were purified from [35S]cysteine-labeled JAR choriocarcinoma cells by immunoprecipitation and by reverse-phase high performance liquid chromatography (HPLC). To identify unformed disulfide bonds, nonreduced folding intermediates were treated with trypsin to liberate non-disulfide-bound, [35S]cysteine-containing peptides from the disulfide-linked peptides. Released peptides were purified by HPLC and identified by amino acid sequencing. The amount of a peptide that was released indicated the extent of disulfide bond formation involving the cysteine in that peptide. Of the six disulfide bonds in hCG-beta, bonds 34-88 and 38-57 form first. The rate-limiting event of folding involves the formation of the S-S bonds between cysteines 23 and 72 and cysteines 9 and 90. Disulfide bond 93-100, the formation of which appears to be necessary for assembly with the alpha subunit of the hCG heterodimer, forms next. Finally, disulfide bond 26-110 forms after assembly with the alpha subunit, suggesting that completion of folding of the COOH terminus in the beta subunit occurs after assembly with the alpha subunit.

Amino Acid Sequence↗

PubMed1991

Biosynthesis, primary structure and molecular cloning of snowdrop (Galanthus nivalis L.) lectin.

Poly(A)-rich RNA isolated from ripening ovaries of snowdrop (Galanthus nivalis L.) yielded a single 17-kDa lectin polypeptide upon translation in a wheat-germ cell-free system. This lectin was purified by affinity chromatography. Translation of the same RNA in Xenopus leavis oocytes revealed a lectin polypeptide which was about 2 kDa smaller than the in vitro synthesized precursor, suggesting that the oocyte system had removed a 2-kDa signal peptide. A second post-translational processing step was likely to be involved since both the in vivo precursor and the Xenopus translation products were about 2 kDa larger than the mature lectin polypeptide. This hypothesis was confirmed by the structural analysis of the amino acid sequence of the mature protein and the cloned mRNA. Edman degradation and carboxypeptidase Y digestion of the mature protein, and structural analysis of the peptides obtained after chemical cleavage and modification, allowed determination of the complete 105 amino acid sequence of the snowdrop lectin polypeptide. Comparison of this sequence with the deduced amino acid sequence of a lectin cDNA clone revealed that besides the mature lectin polypeptide, the lectin mRNA also encoded a 23 amino acid signal-sequence and a C-terminal extension of 29 amino acids, which confirms the results from in vitro translation experiments.

Amino Acid Sequence↗