A three-gene signature marks the time to locoregional recurrence in luminal-like breast cancer.

BACKGROUND: Gene expression profiling (GEP)-based prognostic signatures are being rapidly integrated into clinical decision making for systemic management of breast cancer patients. However, GEP remains relatively underdeveloped for locoregional risk assessment. Yet, locoregional recurrence (LRR), especially early after surgery, is associated with poor survival. PATIENTS AND METHODS: GEP was carried out on two independent luminal-like breast cancer cohorts of patients developing early (≤5 years after surgery) or late (>5 years) LRR and used, by a training and testing approach, to build a gene signature able to intercept women at risk of developing early LRR. The GEP data of two in silico datasets and of a third independent cohort were used to explore its prognostic value. RESULTS: Analysis of the first two cohorts led to the identification of three genes, CSTB, CCDC91 and ITGB1, whose expression, derived by principal component analysis, generated a three-gene signature significantly associated with early LRR in both cohorts (P value <0.001 and 0.005, respectively), overcoming the discriminatory capability of age, hormone receptor status and therapy. Remarkably, the integration of the signature with these clinical variables led to an area under the curve of 0.878 [95% confidence interval (CI) 0.810-0.945]. In in silico datasets we found that the three-gene signature retained its association, showing higher values in the early relapsed patients. Moreover, in the third additional cohort, the signature significantly associated with relapse-free survival (hazard ratio 1.56, 95% CI 1.04-2.35). CONCLUSIONS: Our three-gene signature represents a new exploitable tool to aid treatment choice in patients with luminal-like breast cancer at risk of developing early recurrence.

Hydrogen-peroxide and silicone-hydrogel contact lenses: Worsening of external eye condition and tear film instability.

PURPOSE: The aim is discussing the origins of worsening of external eye condition (EEC) and of tear film (TF) instability after wear of silicone-hydrogel contact lenses (CLs) with hydrogen-peroxide (H2O2) care system. METHODS: EEC and TF stability were evaluated before and after 15days of wear combined with different care systems: (1) H2O2, (2) detergent solution and H2O2, (3) multipurpose solution (MPS), (4) H2O2 and artificial tears. In-vitro cell mortality tests were performed after 24h cell incubation with CLs treated with H2O2. Photon correlation spectroscopy (PCS) was carried out on tears of non-wearers and CL wearers who used MPS or H2O2 solution. RESULTS: Worsening of EEC was observed only for the group using H2O2 (group 1). In-vitro, cell mortality was found higher for worn CL than for unworn CLs. Worsening of TF stability was observed regardless of care system and also PCS results on tears of CL wearers were found different compared to non-wearers regardless of care system. The only observed remedy for tear instability of CL wearers was found to be the administration of artificial tears. CONCLUSIONS: Worsening of EEC of CL wearers using H2O2 is attributed to H2O2 scarce cleaning efficacy, which can be solved by adding a CL detergent solution. The origin of TF instability is found to be different. A remedy was found to be the administration of artificial tears, whose effect could be attributed either to the role of specific components or to rinsing and replacement of TF during wear.

Photon correlation spectroscopy applied to tear analysis.

This study aims to deepen the knowledge on tear film properties by the development of a protocol for analyses of Photon Correlation Spectroscopy (PCS) on human tears and by the comparison between PCS results obtained on tears of contact lens wearers and non-wearers. Tears (5µL) were collected by a glass capillary. The analyses provide the hydrodynamic diameter of tear components by analyzing intensity fluctuations in time of scattered light. PCS appears a promising technique for studying tear features and for shedding light on specific eye conditions, such as on the clinical effects of CL wear. In fact, statistical difference (p<0.001) was found between the measured mean hydrodynamic diameter of tear components of wearers and non-wearers, the resulting value significantly higher for CL wearers. The scenario does not substantially change after (25±5)min from the CL removal. The difference is attributed to changes in the interactions between tear constituents due to CL wear. In order to get deeper insights on the influence of CL wear on aggregation and structure of tear components, a preliminary Electron Spin Resonance (ESR) investigation was performed, monitoring Fe3+ species. ESR spectra on tears of both CL wearers and non-wearers showed the presence of intense signals, probably associated to iron (III) centers in proteins such as lactoferrin, and a weaker resonance attributable to Fe3+ species interacting with S-S bridges of lysozyme. Differences in ESR spectra between CL wearers and non-wearers were detected and tentatively ascribed to changes in coordination or in local environment of Fe3+ centers connected to aggregation phenomena induced by CL wear, which promote their interaction with other neighboring iron species.

Comparative analysis of full-time, half-time, and quarter-time myocardial ECG-gated SPECT quantification in normal-weight and overweight patients.

BACKGROUND: The introduction of a camera-based dose-reduction strategy in myocardial perfusion imaging (MPI) clinical setting entails the definition of objective and reproducible criteria for establishing the amount of activity to be injected. AIM: The aim is to evaluate the impact of count statistics on the estimation of summed-scores (SS), end-diastolic volume (EDV), end-systolic volume (ESV), and ejection fraction (EF). METHODS: Data rest/stress ECG-gated SPECT (2-day protocol and 8 MBq·kg-1) were acquired with Bright View gamma camera and Astonish algorithm for 40 normal-weight and 40 overweight patients. Assuming that count statistics of shorter acquisition time may simulate that of lower injected activity, three simultaneous scans (full-time, half-time, and quarter-time scans) were started at the same time but with different acquisition time/projection (30, 15 and 8 seconds). RESULTS: A significant difference between SS values of half-time and quarter-time stress scans was found for overweight group (P = .006). Post hoc test showed significant differences for ESV (P < .05), EDV (P < .01) and EF (P < .05) between half-time and quarter-time scans for both patient groups. CONCLUSIONS: The reduction of the count-statistics to a quarter of the MPI reference influenced negatively the quantification in overweight patients. The decrease of radiopharmaceutical activity to 25% of the reference seems practicable for normal-weight patients, while it is more appropriate an activity reduction limited to 50% for overweight and obese patients.

Myocardial perfusion scintigraphy dosimetry: optimal use of SPECT and SPECT/CT technologies in stress-first imaging protocol.

PURPOSE: Over the past decade, nuclear medicine experts have been seeking to minimize patient exposure to radiation in myocardial perfusion scintigraphy (MPS). This review describes the latest technological innovations in MPS, particularly with regard to dose reduction. METHODS: We searched in PubMed for original clinical papers in English, published after 2008, using the following research criteria: (dose) and ((reduction) or (reducing)) and ((myocardial) or (cardiac) or (heart)) and ((nuclear medicine) or (nuclear imaging) or (radionuclide) or (scintigraphy) or (SPET) or (SPECT)). Thereafter, recent reviews on the topic were considered and other relevant clinical papers were added to the results. RESULTS: Of 202 non-duplicate articles, 17 were included. To these, another eight papers cited in recent reviews were added. By optimizing the features of software, i.e., through algorithms for iterative reconstruction with resolution recovery (IRRs), and hardware, i.e., scanners and collimators, and by preferring, unless otherwise indicated, the use of stress-first imaging protocols, it has become possible to reduce the effective dose by at least 50% in stress/rest protocols, and by up to 89% in patients undergoing a diagnostic stress-only study with new technology. With today's SPECT/CT systems, the use of a stress-first protocol can conveniently be performed, resulting in an overall dose reduction of about 35% if two-thirds of stress-first examinations were considered definitively normal. CONCLUSION: Using innovative gamma cameras, collimators and software, as well as, unless otherwise indicated, stress-first imaging protocols, it has become possible to reduce significantly the effective dose in a high percentage of patients, even when X-ray CT scanning is performed for attenuation correction.

Observational prospective study comparing mucoactive and antibiotic treament in the management of acute cough from upper respiratory tract infections.

AIM: Since the efficacy of antibiotics in the management of cough from upper respiratory tract infections (URTI) is unclear, we conducted an observational study to evaluate the efficacy of antibiotic compared to symptomatic treatment in adults with productive cough from URTI in a real life setting. METHODS: One-hundred and 50 subjects presenting to five general practice ambulatories with productive cough due to URTI were included in the study. According to the physician judgment, patients were treated with either oral antibiotics (group A, 50 patients), a mucoactive syrup (sobrerol: group S, 50 patients) as symptomatic treatment, or both sobrerol and oral antibiotics (group SA, 50 patients). Cough severity was assessed daily with a diary, by means of a validated verbal category-descriptive (VCD) Scale. RESULTS: In every treatment group cough severity decreased over time (P<0.001); however, during the first week of treatment patients in groups S and SA experienced significantly lower VCD score values in comparison to Group A (P=0.045 and P=0.010, respectively). On the other hand, the VCD score was never significantly different between the two groups that received symptomatic treatment at any of the 4 weeks of the study, suggesting that the effect observed was due to the action of the mucoactive syrup. CONCLUSION: Our data confirm the lack of efficacy of antibiotics in the management of productive cough in URTI and suggest that symptomatic treatment should be preferred to antibiotics in acute cough from URTI.

Clinical evaluation and cellular electrophysiology of a recessive CLCN1 patient.

Here we present the case of a 32-year-old female patient with myotonia congenita. She carried two mutations in the CLCN1 gene that encodes the chloride channel ClC-1: p.Phe167Leu, which was previously identified in several families, and p.Val536Leu, which has been previously reported but not yet characterized by electrophysiological investigations. The patient's symptoms included generalized stiffness, myotonia, and muscle cramps mostly localized in the lower limbs. These symptoms started during childhood and worsened over the following years. The symptoms were exacerbated by low outside temperature, rest, stress, and fasting and were improved by mild exercise, suggesting a warm-up phenomenon. The mutation p.Phe167Leu has previously been associated with a slight shift in the overall open probability. Here we further analysed this mutation to extrapolate the voltage-dependence of the fast and slow gates. In our experimental conditions, p.Phe167Leu exclusively affected the slow gate, increasing the minimum open probability and displacing the voltage-dependence toward depolarized potentials. p.Val536Leu showed more severe effects, dramatically influencing the slow gate as well as modifying properties of the fast gate. Co-expression of the mutants in a human cell line to reproduce the compound heterozygous condition of the patient produced channels with altered voltage-dependence of the slow gate but a restored fast gate. The alteration of the slow mechanism was reflected by the relative open probability, reducing the contribution of ClC-1 channels in maintaining the resting membrane potential of skeletal muscles and thus explaining the myotonic phenotype of the patient.

Exploring dopaminergic activity in ring chromosome 20 syndrome: a SPECT study.

AIM: Several lines of evidence indicate that the dopaminergic system may play a role in the propagation of epileptic seizures and, indeed, DOPA metabolism impairment has recently been demonstrated in PET studies of ring chromosome 20 [r(20)] patients. We conducted a study looking for correlations between r(20) mosaicism, other clinical variables and both pre-synaptic dopamine transporter (DAT) expression and post-synaptic D2 receptor density. METHODS: Five patients with r(20) and epilepsy were enrolled in the study. DAT expression and D2 density were measured by single photon emission tomography (SPECT) imaging with 185 MBq of [¹²³I]ioflupane and [¹²³I]IBZM, respectively, on different days. Linear correlations between r(20) mosaicism, clinical variables and binding of [¹²³I]ioflupane or [¹²³I]IBZM were examined. RESULTS: A significant correlation between seizure frequency and r(20) mosaicism was detected (r=0.903, P<0.05), along with a negative correlation between r(20) mosaicism and binding of [¹²³I]ioflupane in the putamen and in the caudate nucleus (r=-0.692 and r=-807; P<0.05). Seizure frequency was positively correlated with post-synaptic D2 density (r=0.925, P<0.05). CONCLUSION: Striatal neurons are involved in r(20) epilepsy; the relationship found between r(20) mosaicism and DAT expression suggests that drugs acting on the dopaminergic system could have a place in the treatment of this rare form of epilepsy.

Exploring dopaminergic activity in ring chromosome 20 syndrome: a SPECT study.

AIM: Several lines of evidence indicate that the dopaminergic system may play a role in the propagation of epileptic seizures and, indeed, DOPA metabolism impairment has recently been demonstrated in PET studies of ring chromosome 20 [r(20)] patients. We conducted a study looking for correlations between r(20) mosaicism, other clinical variables and both pre-synaptic dopamine transporter (DAT) expression and post-synaptic D2 receptor density. METHODS: Five patients with r(20) and epilepsy were enrolled in the study. DAT expression and D2 density were measured by single photon emission tomography (SPECT) imaging with 185 MBq of [123I]ioflupane and [123I]IBZM, respectively, on different days. Linear correlations between r(20) mosaicism, clinical variables and binding of [123I]ioflupane or [123I]IBZM were examined. RESULTS: A significant correlation between seizure frequency and r(20) mosaicism was detected (r=0.903, P<0.05), along with a negative correlation between r(20) mosaicism and binding of [123I]ioflupane in the putamen and in the caudate nucleus (r=-0.692 and r=-807; P<0.05). Seizure frequency was positively correlated with post-synaptic D2 density (r=0.925, P<0.05). CONCLUSION: Striatal neurons are involved in r(20) epilepsy; the relationship found between r(20) mosaicism and DAT expression suggests that drugs acting on the dopaminergic system could have a place in the treatment of this rare form of epilepsy.

Differences in hippocampal metabolism between amnestic and non-amnestic MCI subjects: automated FDG-PET image analysis.

AIM: The aim of this study was to assess whether 18F-fluorodeoxyglucose positron emission tomography differentiates amnestic (aMCI) from single-non-amnestic mild cognitive impairment (snaMCI) with executive dysfunction. METHODS: Sixteen aMCI subjects (62% females, age 75+/-8 years) and 14 snaMCI subjects (71% females, age 74+/-6 years) underwent [18F]FDG-PET and clinical follow-up. Comparisons between MCI subgroups and with seven cognitively normal elderly subjects were performed using SPM2. RESULTS: At baseline aMCI and snaMCI exhibited a similar pattern of hypometabolism, mostly in the posterior cingulate gyrus, as compared with controls. In the comparison between the MCI subtypes, the aMCI subjects showed reduced metabolism in the medial temporal lobes (MTL) (hippocampus, fusiform gyrus and amygdala). At follow-up 12 aMCI developed Alzheimer's disease (AD), while snaMCI had a heterogeneous course, including five subjects who developed Lewy body dementia. CONCLUSIONS: The patterns of altered brain metabolism in aMCI and snaMCI subjects compared to controls are similar and do not provide evidence for making clinical distinctions between them. Comparison between the two MCI subtypes showed MTL hypometabolism in aMCI subjects, possibly reflecting the fact that most had prodromal AD.

Quinolinic acid induced neurodegeneration in the striatum: a combined in vivo and in vitro analysis of receptor changes and microglia activation.

PURPOSE: Huntington's disease (HD) is a progressive neurodegenerative disorder, which is characterised by prominent neuronal cell loss in the basal ganglia with motor and cognitive disturbances. One of the most well-studied pharmacological models of HD is produced by local injection in the rat brain striatum of the excitotoxin quinolinic acid (QA), which produces many of the distinctive features of this human neurodegenerative disorder. Here, we report a detailed analysis, obtained both in vivo and in vitro of this pharmacological model of HD. MATERIALS AND METHODS: By combining emission tomography (PET) with autoradiographic and immunocytochemical confocal laser techniques, we quantified in the QA-injected striatum the temporal behavior (from 1 to 60 days from the excitotoxic insult) of neuronal cell density and receptor availability (adenosine A(2A) and dopamine D(2) receptors) together with the degree of microglia activation. RESULTS: Both approaches showed a loss of adenosine A(2A) and dopamine D(2) receptors paralleled by an increase of microglial activation. CONCLUSION: This combined longitudinal analysis of the disease progression, which suggested an impairment of neurotransmission, neuronal integrity and a reversible activation of brain inflammatory processes, might represent a more quantitative approach to compare the differential effects of treatments in slowing down or reversing HD in rodent models with potential applications to human patients.

Instrumentation and probes for molecular and cellular imaging.

Molecular and cellular imaging is a branch of biomedical sciences that combines the use of imaging instrumentation and biotechnology to characterize molecular and cellular processes in living organisms in normal and pathologic conditions. The two merging areas of research behind molecular and cellular imaging are detection technology, i.e. scanners and imaging devices, and development of tracers, contrast agents and reporter probes that make imaging with scanners and devices possible. Several in vivo imaging instruments currently used in human studies, such as computer tomography, ultrasound, magnetic resonance, positron emission tomography and single photon emission computed tomography, have been rescaled for small animal studies, while other methods initially used for in vitro evaluation, such as bioluminescence and fluorescence, have been refined for in vivo imaging. Conventional imaging relies on the use of non specific contrast agents and classical probes; however, newly developed targeted contrast agents and activable ''smart'' imaging probes for so-called ''targeted imaging'' have demonstrated high specificity and high signal to noise ratio in small animal studies. This review focuses on basic recent findings in the technical aspects of molecular and cellular imaging modalities (equipment, targeted probe and contrast agents and applied combinations of instrumentation and probe) with particular attention to the choice of the future: the multimodal imaging approach.

Two-dimensional vs three-dimensional imaging in whole body oncologic PET/CT: a Discovery-STE phantom and patient study.

AIM: To evaluate the performance of the positron emission tomography (PET)/computed tomography (CT) Discovery-STE (D-STE) scanner for lesion detectability in two-dimensional (2D) and three-dimensional (3D) acquisition. METHODS: A NEMA 2001 Image-Quality phantom with 11 lesions (7-37 mm in diameter) filled with a solution of 18F (lesion/background concentration ratio: 4.4) was studied. 2D and 3D PET scans were sequentially acquired (10 min each) in list mode (LM). Each scan was unlisted into 4, 3 and 2-min scans. Ten [18F]FDG PET oncological patient studies were also evaluated. Each patient underwent a 3D PET/CT whole body scan, followed by a 2D PET scan (4 min LM) and a 3D PET scan (4 min LM) over a single field of view. Both 2D and 3D scans were unlisted in 3 and 2-min scans. Data were evaluated quantitatively by calculating quality measurements and qualitatively by two physicians who judged lesion detectability compared to statistical variations in background activity. RESULTS: Quantitative and qualitative evaluations showed the superiority of 3D over 2D across all measures of quality. In particular, lesion detectability was better in 3D than in 2D at equal scan times and 3D acquisition provided images comparable in quality to 2D in approximately half the time. Interobserver variability was lower in evaluation of 3D scans and lesion shape and volume were better depicted. CONCLUSION: In oncological applications, the D-STE system demonstrated good performance in 2D and 3D acquisition, while 3D exhibited better image quality, data accuracy and consistency of lesion detectability, resulting in shorter scan times and higher patient throughput.

Subretinal electrode implantation in the P23H rat for chronic stimulations.

BACKGROUND: In age related macular degeneration and inherited dystrophies, preservation of retinal ganglion cells has been demonstrated. This finding has led to the development of various models of subretinal or epiretinal implant in order to restore vision. This study addresses the development of a polyimide subretinal electrode platform in the dystrophic P23H rat in vivo. METHODS: A technique was developed for implanting a subretinal electrode into the subretinal space and stabilising the distal extremity of the cabling on the rat cranium in order to allow future electrical stimulations of the retina. RESULTS: In vivo imaging of the retina with the scanning laser ophthalmoscope demonstrated reabsorption of the surgically induced retinal detachment and the absence of major tissue reactions. These in vivo observations were confirmed by retinal histology. The extraocular fixation system on the rat cranium was effective in stabilising the distal connector for in vivo stimulation. CONCLUSION: This study demonstrates that a retinal implant can be introduced into the subretinal space of a dystrophic rat with a stable external connection for repeatable electrical measurements and stimulation. This in vivo model should therefore allow us to evaluate the safety and efficacy of electrical stimulations on dystrophic retina.

Ganglion cells from chick retina display multiple functional nAChR subtypes.

We have examined the properties of nicotinic acetylcholine receptors in embryonic chick retinal ganglion cells. Ganglion cells, identified according to morphological and physiological criteria, displayed spontaneous or induced action potentials. In 94/99 cells acetylcholine pulses evoked responses. In current clamp mode, acetylcholine provoked membrane depolarization and triggered action potentials. Under voltage clamp conditions, acetylcholine evoked inward currents that were readily blocked by d-tubocurarine. Antagonists specific for homomeric (alpha-bungarotoxin) and heteromeric (dihydro-beta-erythroidine) receptors revealed that ganglion cells express multiple functional receptor subtypes. These findings demonstrate that ACh modulates the electrical activity of these cells and is likely to mediate synaptic transmission. The presence of multiple receptor subtypes may contribute to processing and transmission of information in the retina.

Resveratrol derivatives and their role as potassium channels modulators.

A series of stilbenoid analogues of resveratrol (trans-3,4',5-trihydroxystilbene) with a stilbenic or a bibenzylic skeleton have been prepared by partial synthesis from resveratrol and dihydroresveratrol. The synthesized compounds have been evaluated for their ability to modulate voltage-gated channels.

Evaluation of the clinical performances of a large NaI(Tl) crystal 3D PET scanner.

AIM: This study was aimed at assessing the clinical performances of a NaI(Tl) crystal 3D PET scanner, C-PET (ADAC-UGM), using a multi-ring 2D BGO PET scanner (multi-ring PET), as a reference. METHODS: Thirty-seven oncological patients were studied in sequence with multi-ring PET and C-PET, within 30 days of a CT study. In order to assess the behaviour of C-PET in relation to acquisition count rate, patients were divided into 3 groups according to the count rate at the time of the C-PET scan acquisition. Group A (n=21): 3000-5000 kcounts/sec (recommended count rate range); Group B (n=8): <3000 Kcounts/sec and Group C (n=8): >5000 Kcounts/sec. RESULTS: The number of lesions detected by multi-ring PET and C-PET, classified according to size, was compared. For Group A and Group B there was a good agreement between C-PET and multi-ring PET in terms of lesion detectability (relative sensitivity: 99.9% and 96.0%, respectively), while for Group C the relative sensitivity of C-PET was 61.9%. CONCLUSION: Optimal performances of the C-PET scanner can thus be obtained at a count rate within or below the recommended range. Despite a lower lesion/background contrast resulting from a high scatter and random noise, the sensitivity of C-PET in detecting hypermetabolic lesions is comparable to that of multi-ring PET. These findings are discussed in relation to the physical performance of the two scanners and particularly in relation to the 3D vs 2D acquisition modality.

Glucose- and arginine-induced insulin secretion by human pancreatic beta-cells: the role of HERG K(+) channels in firing and release.

The human ether-a-go-go-related genes (herg) are expressed in tissues other than heart and brain where the HERG K(+) channels are known to regulate the repolarization of the heart action potential and the neuronal spike-frequency accommodation. We provide evidence that herg1 transcripts are present in human pancreatic islets that were used to study both insulin secretion and electrical activity with radioimmunoassay and single cell perforated patch-clamp techniques, respectively. Glucose- and arginine-induced islets insulin secretion data suggested a net increase of release under perfusion with antiarrhythmic drugs known to selectively block HERG channels. Indeed we could routinely isolate a K(+) current that was recognized as biophysically and pharmacologically similar to the HERG current. An analysis of the glucose- and arginine-induced electrical activity (several applications during 30 min) in terms of firing frequency and putative insulin release was done in control and in the presence of selective blockers of HERG channels: the firing frequency and the release increased by 32% and 77%, respectively. It is concluded that HERG channels have a crucial role in regulating insulin secretion and firing of human beta-cells. This raises the possibility that some genetically characterized hyperinsulinemic diseases of unknown origin might involve mutations in the HERG channels.