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Biomedical subjects

F Sesti

Publications and source records attributed to F Sesti.

At least 19 recordsLinked to original sources
PubMed2006

Isobaric gasless laparoscopic myomectomy for removal of large uterine leiomyomas.

BACKGROUND: This study aimed to evaluate the feasibility and safety of isobaric laparoscopic removal of large myomas (> or = 8 cm) using the Laparotenser, a subcutaneous abdominal wall-lifting system. METHODS: A series of 63 consecutive patients with at least one large symptomatic subserosal or intramural uterine myoma (> or = 8 cm) underwent an isobaric gasless laparoscopic myomectomy. Conventional laparotomy instruments were used. RESULTS: The procedure was successfully completed for all 63 consecutive patients. The average size of the dominant myoma was 11 cm. The mean number of myomas removed from each patient was 3.6. The mean blood loss was 143 ml, and the mean operating time was 72 min. No intraoperative complication occurred. CONCLUSIONS: Gasless laparoscopic myomectomy for the removal of large myomas using the Laparotenser is feasible and safe. It offers several advantages over laparoscopy with pneumoperitoneum.

Adult↗
PubMed2005

Silencing of endogenous IGFBP-5 by micro RNA interference affects proliferation, apoptosis and differentiation of neuroblastoma cells.

Signal transduction through the IGF axis is implicated in proliferation, differentiation and survival during development and adult life. The IGF axis includes the IGF binding proteins (IGFBPs) that bind IGFs with high affinity and modulate their activity. In neuroblastoma (NB), a malignant childhood tumor, we found that IGFBP-5 is frequently expressed. Since NB is an IGF2-sensitive tumor, we investigated the relevance and the function of endogenous IGFBP-5 in LAN-5 and in SY5Y(N) cell lines transfected with micro and small interfering RNAs directed to IGFBP-5 mRNA. Cells in which IGFBP-5 expression was suppressed were growth-inhibited and more prone to apoptosis than the parental cell line and controls. Apoptosis was further enhanced by X-ray irradiation. The ability of these cells to undergo neuronal differentiation was impaired after IGFBP-5 inhibition but the effect was reversed by exposure to recombinant IGFBP-5. Together, these data demonstrate the importance of IGFBP-5 for NB cell functions and suggest that IGFBP-5 might serve as a novel therapeutic target in NB.

Apoptosis↗
PubMed2004

Cellular retinol-binding protein-1 expression in endometrial stromal cells: physiopathological and diagnostic implications.

AIMS: Cellular retinol-binding protein-1 (CRBP-1) contributes to the maintenance of the differentiated state of the endometrium through retinol bioavailability regulation. The aim was to analyse CRBP-1 expression in endometrial stromal cells at eutopic and ectopic sites in different physiopathological conditions. METHODS AND RESULTS: Antibodies to CRBP-1, CD10 and alpha-smooth muscle actin were applied to proliferative (n = 10), secretory (n = 9) and atrophic (n = 7) endometrium, decidua (n = 4), adenomyosis (n = 5), endometriosis (n = 10), endometrial polyps (n = 9), simple endometrial hyperplasia (n = 6), well-differentiated endometrioid carcinoma (n = 6) and submucosal leiomyomas (n = 5). In some cases, Western blotting and reverse transcription-polymerase chain reaction were also applied. CRBP-1 was expressed by eutopic and ectopic endometrial stromal cells more markedly during the late secretory phase and in decidua of pregnancy. CRBP-1 expression was low in the stroma of atrophic endometrium and absent in myometrium, leiomyomas and cervical stroma. CD10 immunoreactivity was weak in atrophic endometrium and in decidua. CONCLUSIONS: CRBP-1 expression characterizes endometrial stromal cells at eutopic and ectopic sites and appears to be more specific than CD10. The level of CRBP-1 varies in intensity according to hormonal variations, reaching its maximum in predecidua and decidua. Thus, immunodetection of CRBP-1 may help to elucidate the physiopathological changes which occur in endometrial stroma and can also be applied as an adjuvant stromal marker.

Adult↗
PubMed2004

Human papillomavirus (HPV) testing in the management of women with abnormal Pap smears. Experience of a colposcopy referral clinic.

BACKGROUND AND OBJECTIVES: Several detailed algorithms for the appropriate use of human papillomavirus (HPV) testing in the management of women with abnormal Pap (Papanicolaou) smears have been launched, but their direct country-to-country adoption is difficult. This necessitates their testing in individual settings, which is ongoing in our colposcopy referral clinic. METHODS: A series of 224 consecutive women attending the clinic with the usual referral indications (ASC-US or higher in Pap) were examined by the conventional diagnostic tools (PAP smear, colposcopy, punch biopsy) and subjected to HPV testing and viral typing for both low-risk (L-R) and high-risk (H-R) types by nested PCR-based techniques. Predictors of the high-grade diagnostic categories were analysed using both univariate- and multivariate modelling, and the performance characteristics (sensitivity, specificity, NPV, PPV) of all tests in detecting high-grade CIN were calculated. RESULTS: In the PAP test, ASC-US smears were most common (37.9%), followed by low-grade squamous intraepithelial lesions (LSIL) (26.3%) and high-grade SIL (HSIL) (4.9%). Colposcopy was performed for 180 women, of whom 48.3% had a normal transformation zone (TZ), 40.6% had ATZ1 (abnormal TZ grade 1), and 5.6% had ATZ2. In biopsy (n = 71), 49.3% had CIN1, 5.6% CIN2, and 16.9% CIN3. The HPV test was positive in 64 (28.8%) women, more often in those aged < 35 years (p = 0.025). High-grade colposcopy (ATZ2) was significantly associated with HSIL in the Pap test (OR 20.5; 95% CI: 4.34-96.47), and with HPV test positivity (OR 6.37; 95% CI: 1.58-25.73). The most significant predictors of CIN3 were HSIL in the PAP, HPV test positivity, and high-grade colposcopy. HSIL and HPV test (for H-R types), but not colposcopy, retained their significance as independent predictors of CIN3 also in adjusted multivariate models: OR 88.27; 95% CI 4.17-1867.04, and OR 19.46; 95% CI 2.01-187.75, for the HSIL and H-R HPV test, respectively. Changing the cut-off level of the Pap test from ASC-US to HSIL increased the specificity of the test up to 96.4%, with the loss in sensitivity from 87.5% to 43.8%. Colposcopy (ATZ2) had 92% specificity, and NPV competing with that of the Pap test. The sensitivity of HPV test exceeds that of the Pap test at HSIL cut-off level, but the specificity of the PAP test is clearly superior. CONCLUSIONS: Accurate predictors of significant cervical pathology (CIN3) are well defined, but the problem is the different performance of the diagnostic tools in clinical practice. A proficient combination of the tests is likely to result in the most satisfactory clinical practice in the management of women with abnormal Pap tests (MAPS).

Adolescent↗
PubMed2004

Assessment of risk factors and human papillomavirus (HPV) related pathogenetic mechanisms of CIN in HIV-positive and HIV-negative women. Study design and baseline data of the HPV-PathogenISS study.

OBJECTIVES: In women with HIV-associated immunosuppression, HPV infections have an increased risk of progression to high-grade cervical intraepithelial neoplasia (CIN). With the HAART-induced prolonged survival and more protracted clinical course of AIDS, progression of CIN to cervical cancer (CC) has become a clinically relevant issue, and the mechanisms responsible for HIV-HPV interactions need further elucidation. The study design and analysis of the baseline data of our new project are presented. MATERIAL AND METHODS: This project is a combination of a prospective cohort study of HIV- and HIV+ women, and a retrospective analysis of CIN lesions and cervical cancer. Up to the present, 244 women have been enrolled (17 HIV+) and subjected to epidemiological interview, colposcopic examination, sampling for HPV testing and typing (PCR, InnoLiPA), and HPV serology. The retrospective series of biopsies were analysed for 13 biomarkers (monitoring key molecular events) using immunohistochemistry and tested for HPV by PCR and TaqMan. RESULTS: HIV- and HIV+ women differ in their exposure status to many of the key epidemiological risk factors of cervical cancer, the most significant ones being number of sexual partners (p = 0.0001), age at onset of sexual activity (p = 0.002), and contraception (yes-no) (p = 0.009). The differences in the baseline clinical observations are less dramatic; HIV-positive women had more frequent HSIL PAP tests (p = 0.040), CIN2 or higher in cervical biopsy (p = 0.049), and external genital warts (p = 0.019). The factors predicting intermediate endpoint markers of cervical cancer, i.e., HSIL PAP smear, ATZ2 in colposcopy, and high-grade CIN in biopsy were analysed in univariate and multivariate regression models. All factors significant in univariate analysis were entered in the multivariate model; HIV-status and Pap smear history maintained their independent predictive power of the HSIL Pap test. The most powerful predictor of ATZ2 colposcopy was HSIL in Pap test. Only the HSIL Pap test and ATZ2 colposcopy remained significant independent predictors of high-grade CIN (p = 0.0001 and p = 0.008, respectively) in the multivariate model. CONCLUSIONS: The three intermediate endpoint markers are closely interrelated, but predicted in part by different covariantes in the causal pathway to cervical cancer. To elucidate whether the increased risk of HIV-positive women to high-grade CIN is due a) to their different exposure status to the risk factors, b) to the direct effects of HIV, or c) to molecular interactions between HIV and HPV, we need to complete these analyses separately in HIV+ and HIV- women.

Adult↗
PubMed2001

Immunity to K1 killer toxin: internal TOK1 blockade.

K1 killer strains of Saccharomyces cerevisiae harbor RNA viruses that mediate secretion of K1, a protein toxin that kills virus-free cells. Recently, external K1 toxin was shown to directly activate TOK1 channels in the plasma membranes of sensitive yeast cells, leading to excess potassium flux and cell death. Here, a mechanism by which killer cells resist their own toxin is shown: internal toxin inhibits TOK1 channels and suppresses activation by external toxin.

Gene Expression Regulation, Fungal↗
PubMed2000

A common polymorphism associated with antibiotic-induced cardiac arrhythmia.

Drug-induced long QT syndrome (LQTS) is a prevalent disorder of uncertain etiology that predisposes to sudden death. KCNE2 encodes MinK-related peptide 1 (MiRP1), a subunit of the cardiac potassium channel I(Kr) that has been associated previously with inherited LQTS. Here, we examine KCNE2 in 98 patients with drug-induced LQTS, identifying three individuals with sporadic mutations and a patient with sulfamethoxazole-associated LQTS who carried a single-nucleotide polymorphism (SNP) found in approximately 1.6% of the general population. While mutant channels showed diminished potassium flux at baseline and wild-type drug sensitivity, channels with the SNP were normal at baseline but inhibited by sulfamethoxazole at therapeutic levels that did not affect wild-type channels. We conclude that allelic variants of MiRP1 contribute to a significant fraction of cases of drug-induced LQTS through multiple mechanisms and that common sequence variations that increase the risk of life-threatening drug reactions can be clinically silent before drug exposure.

Amino Acid Sequence↗
PubMed2000

MinK endows the I(Ks) potassium channel pore with sensitivity to internal tetraethylammonium.

I(Ks) channels are heteromeric complexes of pore-forming KvLQT1 subunits and pore-associated MinK subunits. Channels formed only of KvLQT1 subunits vary from I(Ks) channels in their gating kinetics, single-channel conductance, and ion selectivity. Here we show that I(Ks) channels are more sensitive to blockade by internal tetraethylammonium ion (TEA) than KvLQT1 channels. Inhibition by internal TEA is shown to proceed by a simple bimolecular interaction in the I(Ks) conduction pathway. Application of a noise-variance strategy suggests that MinK enhances blockade by increasing the dwell time of TEA on its pore site from approximately 70 to 370 micros. Mutation of consecutive residues across the single transmembrane segment of MinK identifies positions that alter TEA blockade of I(Ks) channels. MinK is seen to determine the pharmacology of I(Ks) channels in addition to establishing their biophysical attributes.

Amino Acid Sequence↗
PubMed1999

A molecular target for viral killer toxin: TOK1 potassium channels.

Killer strains of S. cerevisiae harbor double-stranded RNA viruses and secrete protein toxins that kill virus-free cells. The K1 killer toxin acts on sensitive yeast cells to perturb potassium homeostasis and cause cell death. Here, the toxin is shown to activate the plasma membrane potassium channel of S. cerevisiae, TOK1. Genetic deletion of TOK1 confers toxin resistance; overexpression increases susceptibility. Cells expressing TOK1 exhibit toxin-induced potassium flux; those without the gene do not. K1 toxin acts in the absence of other viral or yeast products: toxin synthesized from a cDNA increases open probability of single TOK1 channels (via reversible destabilization of closed states) whether channels are studied in yeast cells or X. laevis oocytes.

Animals↗
PubMed1999

The native rat olfactory cyclic nucleotide-gated channel is composed of three distinct subunits.

Cyclic nucleotide-gated (CNG) channels play central roles in visual and olfactory signal transduction. In the retina, rod photoreceptors express the subunits CNCalpha1 and CNCbeta1a. In cone photoreceptors, only CNCalpha2 expression has been demonstrated so far. Rat olfactory sensory neurons (OSNs) express two homologous subunits, here designated CNCalpha3 and CNCalpha4. This paper describes the characterization of CNCbeta1b, a third subunit expressed in OSNs and establishes it as a component of the native channel. CNCbeta1b is an alternate splice form of the rod photoreceptor CNCbeta1a subunit. Analysis of mRNA and protein expression together suggest co-expression of all three subunits in sensory cilia of OSNs. From single-channel analyses of native rat olfactory channels and of channels expressed heterologously from all possible combinations of the CNCalpha3, -alpha4, and -beta1b subunits, we conclude that the native CNG channel in OSNs is composed of all three subunits. Thus, CNG channels in both rod photoreceptors and olfactory sensory neurons result from coassembly of specific alpha subunits with various forms of an alternatively spliced beta subunit.

Alternative Splicing↗
PubMed1999

MiRP1 forms IKr potassium channels with HERG and is associated with cardiac arrhythmia.

A novel potassium channel gene has been cloned, characterized, and associated with cardiac arrhythmia. The gene encodes MinK-related peptide 1 (MiRP1), a small integral membrane subunit that assembles with HERG, a pore-forming protein, to alter its function. Unlike channels formed only with HERG, mixed complexes resemble native cardiac IKr channels in their gating, unitary conductance, regulation by potassium, and distinctive biphasic inhibition by the class III antiarrhythmic E-4031. Three missense mutations associated with long QT syndrome and ventricular fibrillation are identified in the gene for MiRP1. Mutants form channels that open slowly and close rapidly, thereby diminishing potassium currents. One variant, associated with clarithromycin-induced arrhythmia, increases channel blockade by the antibiotic. A mechanism for acquired arrhythmia is revealed: genetically based reduction in potassium currents that remains clinically silent until combined with additional stressors.

Amino Acid Sequence↗
PubMed1998

Phosphorylation of mammalian olfactory cyclic nucleotide-gated channels increases ligand sensitivity.

In vertebrate olfactory sensory neurons, odorant receptors couple the sensory signal to the synthesis of the second messenger cAMP. Cyclic nucleotide-gated (CNG) channels are activated by binding of cAMP and conduct a depolarizing receptor current that leads to electrical excitation of the neuron. The sensitivity of olfactory CNG channels for cAMP can be significantly reduced by binding of calmodulin to a regulatory domain that resides within the N-terminus of the alpha-subunit of the channel. This regulatory domain also contains a consensus phosphorylation sequence for protein kinase C (PKC). We have investigated the effect of channel phosphorylation by PKC and found that phosphorylation increases ligand sensitivity without counteracting modulation of the channel by calmodulin. We have identified the amino acid residue that is phosphorylated by PKC and have localized three isoforms of PKC in olfactory sensory cilia. The results of this study provide information about the control of ligand sensitivity in olfactory CNG channels by an intrinsic regulatory domain, representing both a calmodulin-binding site and a substrate for PKC.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1998

Single-channel characteristics of wild-type IKs channels and channels formed with two minK mutants that cause long QT syndrome.

IKs channels are voltage dependent and K+ selective. They influence cardiac action potential duration through their contribution to myocyte repolarization. Assembled from minK and KvLQT1 subunits, IKs channels are notable for a heteromeric ion conduction pathway in which both subunit types contribute to pore formation. This study was undertaken to assess the effects of minK on pore function. We first characterized the properties of wild-type human IKs channels and channels formed only of KvLQT1 subunits. Channels were expressed in Xenopus laevis oocytes or Chinese hamster ovary cells and currents recorded in excised membrane patches or whole-cell mode. Unitary conductance estimates were dependent on bandwidth due to rapid channel "flicker." At 25 kHz in symmetrical 100-mM KCl, the single-channel conductance of IKs channels was approximately 16 pS (corresponding to approximately 0.8 pA at 50 mV) as judged by noise-variance analysis; this was fourfold greater than the estimated conductance of homomeric KvLQT1 channels. Mutant IKs channels formed with D76N and S74L minK subunits are associated with long QT syndrome. When compared with wild type, mutant channels showed lower unitary currents and diminished open probabilities with only minor changes in ion permeabilities. Apparently, the mutations altered single-channel currents at a site in the pore distinct from the ion selectivity apparatus. Patients carrying these mutant minK genes are expected to manifest decreased K+ flux through IKs channels due to lowered single-channel conductance and altered gating.

Animals↗
PubMed1996

Time-dependent current decline in cyclic GMP-gated bovine channels caused by point mutations in the pore region expressed in Xenopus oocytes.

1. Amino acids with a charged or a polar residue in the putative pore region, between lysine 346 and glutamate 372 of the alpha-subunit of the cGMP-gated channel from bovine rods were mutated to a different amino acid. The mRNA encoding for the wild-type, i.e. the alpha-subunit, or mutant channels was injected in Xenopus laevis oocytes. 2. When glutamate 363 was mutated to asparagine, serine or alanine, the current activated by a steady cGMP concentration declined in mutant channels. No current decline was observed when glutamate 363 was mutated to aspartate, glutamine or glycine, when theronine 359, 360 and 364 were mutated to alanine or when other charged residues in the pore region were neutralized. 3. The amount of current decline and its time course were significantly voltage dependent. In mutant E363A the current decline developed within about 1.5 s at -100 mV, but in about 6 s at +100 mV. In the same mutant, the current declined to about 55% of its initial level at +100 mV and to about 10% at -100 mV. 4. The current decline in mutants E363A, E363S and E363N was only moderately dependent on the cGMP concentration (from 10 to 1000 microM) and was not caused by a reduced affinity of the mutant channels for cGMP. Analysis of current fluctuations at a single-channel level indicated that current decline was primarily caused by a decrease of the open probability. 5. The wild-type channel was not permeable to dimethylammonium. When glutamate 363 was replaced by a smaller residue such as serine, mutant channels became permeable to dimethylammonium. 6. The current decline observed in mutant channels is reminiscent of desensitization of ligand-gated channels and of inactivation of voltage-gated channels. These results suggest also that gating and permeation through the cGMP-gated channel from bovine rods are intrinsically coupled and that glutamate 363 is part of the molecular structure controlling both the gating and the narrowest region of the pore.

Amino Acid Sequence↗
PubMed1996

Effect of changing temperature on the ionic permeation through the cyclic GMP-gated channel from vertebrate photoreceptors.

Native cGMP-gated channels were studied in rod outer segments of the larval tiger salamander Ambystoma tigrinum. The alpha subunit of the cGMP-gated channel from bovine rods, here referred to as the wild type (w.t.), and mutant channels were heterologously expressed in Xenopus laevis oocytes. These channels were studied in excised membrane patches in the inside-out configuration and were activated by the addition of 100 or 500 microM cGMP. The effect of temperature on the ionic permeation was studied. The macroscopic current flowing through the native channel at +100 mV had an activation energy of 35.8, 30, 31.8, 34.5, 41.3, and 22.4 kJ mol-1 in the presence of Li+, Na+, K+, Rb+, Cs+, and NH4+, respectively. The macroscopic current flowing through the w.t. channel at +100 mV had an activation energy of 45.2, 38.2, 37.5, 47.3, 49.4, and 38.9 kJ mol-1 in the presence of Li+, Na+, K+, Rb+, Cs+, and NH4+, respectively. The activation energy of the macroscopic current flowing through the native and w.t. channels did not vary significantly when the ionic concentration of the permeant ion was changed between 2.5 and 110 mM. The activation energy of the single-channel current of the w.t. channel at +100 mV was 40.4 and 33 kJ mol-1 for Na+ and NH4+, respectively. The reversal potential of biionic solutions changed significantly with temperature. These results can be used to obtain an estimate of the enthalpic and entropic contributions to the barrier of the Gibbs free energy experienced by an ion during its permeation through the open channel. These estimates indicate that the ionic permeation and selectivity of the cGMP-gated channel are controlled both by enthalpic and entropic factors and that the selectivity of the native channel for Li+ over Na+ is primarily caused by entropic effects.

Ambystoma↗