Laparoscopic Appendectomy Performed by junior SUrgeonS: impact of 3D visualization on surgical outcome. Randomized multicentre clinical trial. (LAPSUS TRIAL).

BACKGROUND: The purpose of this study was to evaluate the effect of 3D visualization applied to laparoscopic appendectomy (LA) performed by young surgeons (YS). We considered both operative features and clinical outcomes, aiming to highlight the benefits that this technology could bring to novice surgeons and their laparoscopic training. METHODS: All the surgical procedures were performed by residents who had performed less than 20 appendectomies prior to the beginning of the study under the supervision of an expert surgeon. At the time of enrolment patients were randomized into two arms: Experimental arm (EA): laparoscopic appendectomy performed with laparoscopic 3D vision technology. Control arm (CA): laparoscopic appendectomy performed with the "standard" 2D technology. The primary endpoint was to find any statistically significant difference in operative time between the two arms. Differences in conversion rate, intra-operative complications, post-operative complications and surgeons' operative comfort were considered as secondary endpoints. RESULTS: We randomized 135 patients into the two study arms. The two groups were homogeneous for demographic characteristics, BMI and ASA scores. The characteristics of clinical presentation and anatomical position showed no significant difference. The operative time was longer in the CA (57.5 vs. 49.6 min, p = 0.048, 95% CI). In the subgroup of complicated appendicitis, this trend toward inferior operative time was confirmed without reaching statistical significance (2D = 60 min, 3D = 49.5 min, p = 0.082 95% CI). No intra-operative complications were observed in either group. The conversion rate was 5.6% (4 patients) in the 2D group and 4.6% (3 patients) in 3D group. CONCLUSION: The utilization of 3D laparoscopy was associated with reduction in operative time without influencing other parameters, in particular without altering the safety profile of the procedure.

Everolimus is an active agent in medullary thyroid cancer: a clinical and in vitro study.

Everolimus, an mTOR inhibitor, which has been demonstrated to induce anti-tumour effects in different types of neuroendocrine tumours, has never been evaluated in patients with medullary thyroid cancer (MTC). The aim of this study was to evaluate the in vitro and in vivo effects of everolimus in combination with octreotide in MTC. Two patients with progressive metastatic MTC and high calcitonin levels were treated with everolimus 5-10 mg/day. Both patients were under treatment with octreotide LAR at the study entry. An in vitro study was also performed to assess everolimus effects on MTC cell lines (TT and MZ-CRC-1 cells). A tumour response was observed in both patients. Serum calcitonin decreased by 86% in patient 1 and by 42% in patient 2. In TT and MZ-CRC-1 cells, everolimus induced a significant dose-dependent inhibition in cell proliferation. This effect seems to be related to a cell cycle arrest in G(0) /G(1) phase in both cell lines and to the induction of cellular senescence in TT cells. Everolimus in combination with octreotide may be active as anti-tumour therapy in patients with progressive metastatic MTC, suggesting to further evaluate this agent in MTC patients in a large prospective study.

Perspectives and challenges associated with the determination of new psychoactive substances in urine and wastewater - A tutorial.

New psychoactive substances (NPS), often designed as (legal) substitutes to conventional illicit drugs, are constantly emerging in the drug market and being commercialized in different ways and forms. Their use continues to cause public health problems and is therefore of major concern in many countries. Monitoring NPS use, however, is arduous and different sources of information are required to get more insight of the prevalence and diffusion of NPS use. The determination of NPS in pooled urine and wastewater has shown great potential, adding a different and complementary light on this issue. However, it also presents analytical challenges and limitations that must be taken into account such as the complexity of the matrices, the high sensitivity and selectivity required in the analytical methods as a consequence of the low analyte concentrations as well as the rapid transience of NPS on the drug market creating a scenario with constantly moving analytical targets. Analytical investigation of NPS in pooled urine and wastewater is based on liquid chromatography hyphenated to mass spectrometry and can follow different strategies: target, suspect and non-target analysis. This work aims to discuss the advantages and disadvantages of the different data acquisition workflows and data exploration approaches in mass spectrometry, but also pays attention to new developments such as ion mobility and the use of in-silico prediction tools to improve the identification capabilities in high-complex samples. This tutorial gives an insight into this emerging topic of current concern, and describes the experience gathered within different collaborations and projects supported by key research articles and illustrative practical examples.

Transversus abdominis release (TAR) for ventral hernia repair: open or robotic? Short-term outcomes from a systematic review with meta-analysis.

PURPOSE: To compare early postoperative outcomes after transversus abdominis release (TAR) for ventral hernia repair with open (oTAR) and robotic (rTAR) approach. METHODS: A systematic search of PubMed/MEDLINE, EMBASE, SCOPUS and Web of Science databases was conducted to identify comparative studies until October 2020. A meta-analysis of postoperative short-term outcomes was performed including complications rate, operative time, length of stay, surgical site infection (SSI), surgical site occurrence (SSO), SSO requiring intervention (SSOPI), systemic complications, readmission, and reoperation rates as measure outcomes. RESULTS: Six retrospective studies were included in the analysis with a total of 831 patients who underwent rTAR (n = 237) and oTAR (n = 594). Robotic TAR was associated with lower risk of complications rate (9.3 vs 20.7%, OR 0.358, 95% CI 0.218-0.589, p < 0.001), lower risk of developing SSO (5.3 vs 11.5%, OR 0.669, 95% CI 0.307-1.458, p = 0.02), lower risk of developing systemic complications (6.3 vs 26.5%, OR 0.208, 95% CI 0.100-0.433, p < 0.001), shorter hospital stay (SMD - 4.409, 95% CI - 6.000 to - 2.818, p < 0.001) but longer operative time (SMD 53.115, 95% CI 30.236-75.993, p < 0.01) compared with oTAR. There was no statistically significant difference in terms of SSI, SSOPI, readmission, and reoperation rates. CONCLUSION: Robotic TAR improves recovery by adding the benefits of minimally invasive procedures when compared to open surgery. Although postoperative complications appear to decrease with a robotic approach, further studies are needed to support the real long-term and cost-effective advantages.

TRPM7 and MagT1 regulate the proliferation of osteoblast-like SaOS-2 cells through different mechanisms.

A correct magnesium (Mg2+) intake is essential for bone health. In particular, Mg2+ deficiency inhibits the proliferation of osteoblast-like SaOS-2 cells by increasing nitric oxide (NO) production through the upregulation of inducible NO synthase. At the moment, little is known about the expression and the role of TRPM7, a channel/enzyme involved in Mg2+ uptake, and MagT1, a Mg2+ selective transporter, in SaOS-2 cells. Here, we demonstrate that TRPM7 is not modulated by different extracellular concentrations of Mg2+ and its silencing exacerbates growth inhibition exerted by low Mg2+ through the activation of inducible NO synthase and consequent accumulation of NO. Moreover, MagT1 is upregulated in SaOS-2 cultured in high Mg2+ and its silencing inhibits the growth of SaOS-2 cultured in media containing physiological or high Mg2+, without any modulation of NO production. We propose that TRPM7 and MagT1 are both involved in regulating SaOS-2 proliferation through different mechanisms.

Monitoring psychoactive substance use at six European festivals through wastewater and pooled urine analysis.

The consumption of psychoactive substances is considered a growing problem in many communities. Moreover, new psychoactive substances (NPS) designed as (legal) substitutes to traditional illicit drugs are relatively easily available to the public through e-commerce and retail shops, but there is little knowledge regarding the extent and actual use of these substances. This study aims to gain new and complementary information on NPS and traditional illicit drug use at six music festivals across Europe by investigating wastewater and pooled urine. Samples were collected, between 2015 and 2018, at six music festivals across Europe with approximately 465.000 attendees. Wastewater samples were also collected during a period not coinciding with festivals. A wide-scope screening for 197 NPS, six illicit drugs and known metabolites was applied using different chromatography-mass spectrometric strategies. Several illicit drugs and in total 21 different NPS, mainly synthetic cathinones, phenethylamines and tryptamines, were identified in the samples. Ketamine and the traditional illicit drugs, such as amphetamine-type stimulants, cannabis and cocaine were most abundant and/or frequently detected in the samples collected, suggesting a higher use compared to NPS. The analyses of urine and wastewater is quick and a high number of attendees may be monitored anonymously by analysing only a few samples which allows identifying the local profiles of use of different drugs within a wide panel of psychoactive substances. This approach contributes to the development of an efficient surveillance system which can provide timely insight in the trends of NPS and illicit drugs use.

Frequency and clinical features of Lewy body dementia in Italian memory clinics.

BACKGROUND: The latest developments in Lewy Body Dementia (DLB) raise some controversies on clinical features, neuroimaging and therapy. The aim of our study is to determine clinical, neuropsychological, neuroimaging and EEG profile of DLB through retrospective and prospective data of 102 patients. METHODS: data were collected with an analytical form that was developed by an expertise of neurologists. RESULTS: DLB represented 4.8% of the dementia population, with no sex difference. Family history of dementia was common (24.5%), while familiarity for parkinsonism was rare (4.9%). Cognitive disturbances were the predominant clinical presentation at onset (49%), followed by behavioral symptoms (29.4%) and parkinsonism (21.6%). Clinical features at consultation were: memory disturbances (almost all cases), symmetrical (68.6%) or asymmetrical (18.6%) parkinsonism, cognitive fluctuations (49%), visuospatial deficits (53.9%), and visual hallucinations (44.1%). Autonomic signs were present in a third of the cases, while sleep disorders were present in 44.1%. Some clinical response to antiparkinsonian drugs was evident in half of the cases. MRI, SPET, EEG and Neuropsychiatric Inventory data were available in a subgroup of patients. CONCLUSIONS: Most of our data were in accordance with the previous literature. However, some data underline the relationship between DLB, Alzheimer's and Parkinson's disease.

Recent advances in analytical methods for the determination of 4-alkylphenols and bisphenol A in solid environmental matrices: A critical review.

The problem of endocrine disrupting compounds (EDCs) in the environment has become a worldwide concern in recent decades. Besides their toxicological effects at low concentrations and their widespread use in industrial and household applications, these pollutants pose a risk for non-target organisms and also for public safety. Analytical methods to determine these compounds at trace levels in different matrices are urgently needed. This review critically discusses trends in analytical methods for well-known EDCs like alkylphenols and bisphenol A in solid environmental matrices, including sediment and aquatic biological samples (from 2006 to 2018). Information about extraction, clean-up and determination is covered in detail, including analytical quality parameters (QA/QC). Conventional and novel analytical techniques are compared, with their advantages and drawbacks. Ultrasound assisted extraction followed by solid phase extraction clean-up is the most widely used procedure for sediment and aquatic biological samples, although softer extraction conditions have been employed for the latter. The use of liquid chromatography followed by tandem mass spectrometry has greatly increased in the last five years. The majority of these methods have been employed for the analysis of river sediments and bivalve molluscs because of their usefulness in aquatic ecosystem (bio)monitoring programs. Green, simple, fast analytical methods are now needed to determine these compounds in complex matrices.

Illicit drugs and pharmaceuticals in swimming pool waters.

The occurrence of illicit drugs (cocaine, opioids, amphetamines and cannabis derivatives), some of their metabolites and 48 pharmaceuticals, was investigated in pool and source waters in ten Italian indoor swimming pools. The samples were analyzed by highperformance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), after solid phase extraction (SPE). Cocaine and its metabolites were found in nine swimming pools, at concentrations from 0.3 to 4.2 ng/L for cocaine, 1.1 to 48.7 ng/L for norcocaine, 0.7 to 21.4 ng/L for benzoylecgonine and 0.1 to 7.3 ng/L for norbenzoylecgonine. Opioids, amphetamines and cannabis derivatives were never detected. The most frequent pharmaceuticals were anti-inflammatory drugs: ibuprofen was found in all the pool waters, with a maximum 197 ng/L and ketoprofen was detected in 9/10 samples (maximum 127 ng/L). Among anticonvulsants, carbamazepine and its metabolite, 10,11-dihydro-10,11dihydroxycarbamazepine, were frequent in swimming pool water (8/10 samples) at concentrations up to 62 ng/L. The cardiovascular drug valsartan was also found frequently (8/10 samples), but at lower concentrations (up to 9 ng/L). Other pharmaceuticals were detected occasionally and at lower concentrations (atenolol, enalapril, paracetamol, hydroclorothiazide, irbesartan and dehydro-erythromycin). Carbamazepine, irbesartan and dehydroerythromycin were detected at very low levels (up to 5 ng/L) in only one of the four source water samples. A quantitative risk assessment showed that the health risk for humans to these substance in swimming pool waters was generally negligible, even for vulnerable subpopulations such as children and adolescents.

Investigation of agreement between wastewater-based epidemiology and survey data on alcohol and nicotine use in a community.

BACKGROUND/AIMS: Alcohol and nicotine are the two most used substances world-wide and associated with increased burden of disease. Since surveys on substance use may be difficult due to response biases, wastewater-based epidemiology (WBE) was developed as a more objective measure of nicotine and alcohol use. This study compares estimates of nicotine and alcohol use from a wastewater sampling campaign in a medium-sized Belgian city with a concurrently executed population survey. METHODS: 29,083 letters about participation in an online survey study on weekly alcohol and tobacco use were sent to the inhabitants of Lier, Belgium. Wastewater samples were collected from the associated treatment plant in four bi-weekly periods. Samples were analyzed using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Ethylsulfate was used as alcohol biomarker and cotinine and trans-3'-hydroxycotinine as nicotine biomarker. RESULTS: 263 (1%) surveys were filled out on average per week. According to survey data, alcohol and nicotine were used less than in the rest of Belgium and this was matched by the wastewater data. Nicotine use, but not alcohol use, showed a significant variation over the sampling periods. Both nicotine and alcohol showed increase use during the weekend while only alcohol showed a different use pattern throughout the week. CONCLUSION: No correlation between WBE and survey data could be demonstrated, possibly due to small sample sizes. However, this study shows that weekly trends in alcohol and nicotine use can be quickly detected from wastewater analysis and the occurrence of major events such as festivals can be identified.

3D-conformal radiation therapy in prostate cancer. Technical considerations after 5 years of experience and 334 patients treated at the Istituto Europeo di Oncologia of Milan, Italy.

AIMS AND BACKGROUND: To report the technique of 3D-conformal radiation therapy (3D-CRT) currently used at our Institute for the treatment of prostate cancer with a curative intent. A critical review of the technical aspects of the technique is provided. METHODS AND STUDY DESIGN: Between December 1995 and October 2000, 334 patients with biopsy-proven adenocarcinoma of the prostate were treated with 3D-CRT. All patients were treated in a prone position with 15 MV X-ray beams and a 6-field technique for all but 20 patients, who were treated with a 3-field technique. Patients were simulated with the rectum and bladder empty. To ensure reproducible positioning, custom-made polyurethane foam or thermoplastic casts were produced for each patient. Subsequently, consecutive CT scan slices were obtained. The clinical target volume and critical organs (rectum and bladder) were identified on each CT slice. The beam's eye view technique was used to spatially display these structures, and the treatment portals were manually shaped based on the images obtained. The beam apertures were initially realized by conventional Cerrobend blocks (48 patients), which were replaced in October 1997 by a computer-driven multi-leaf collimator. The total target dose prescribed at the ICRU point is 76 Gy, delivered in 38 fractions and 54 days. The seminal vesicles are excluded at 70 Gy. Dose-volume histograms were obtained for all patients. If more than 30% of the bladder and/or more than 20% of the rectum receive >95% of the prescribed total dose, the treatment plan is judged as unsatisfactory and is adjusted. The dose-volume histogram can be improved by changing the beam's arrangement and/or weights or by introducing or modifying the wedge filters. CONCLUSIONS: 3D-CRT in prostate cancer patients is a highly sophisticated and time-consuming method of dose delivery. Important technical issues remain to be clarified.

Cigarette smoke condensate affects monocyte interaction with endothelium.

OBJECTIVE: Circulating monocytes adhere to the endothelium and migrate into the intima contributing to atherosclerotic plaque growth. Cigarette smoke is a risk factor for atherosclerosis, but it is not completely known how it affects monocyte behavior in atherogenesis. METHODS: We studied the effects of cigarette smoke condensate (CSC) on human monocytes (HM) chemotaxis and transmigration through an endothelial cell (EC) monolayer. RESULTS: Pre-treatment with CSC caused a decrease in HM chemotaxis and transmigration (-55% and -18% vs control, p < 0.05, respectively), paralleled by a reduced expression of Rac 1 GTPase. On the contrary, direct exposure of both HM and EC to CSC increased (+23% vs control, p < 0.05) HM transmigration, paralleled by a strong stimulation of VCAM1 and ICAM1 expression by ECs, and by a slight increase in monocyte integrin expression. An enhancement of monocyte transmigration was obtained after the exposure of both HM and EC to medium conditioned by HM previously incubated with CSC (+265% vs control, p < 0.001). CSC showed a stimulatory effect on the expression by HM of TLR4, MCP1, IL8, IL1beta, and TNFalfa, which was ablated by pre treatment with PDTC. Incubation with neutralizing antibodies against both MCP1 or IL8 completely abolished the CSC-conditioned medium induced HM transmigration. CONCLUSIONS: CSC induces HM to release chemotactic factor(s), which amplify the recruitment and transmigration of inflammatory cells through EC, but CSC may also reduce HM migratory capacity. Therefore, exposure to CSC affects monocyte behavior and interaction with the endothelium, thus potentially facilitating and/or further aggravating the atherogenic process.

Second interlaboratory exercise on non-steroidal anti-inflammatory drug analysis in environmental aqueous samples.

Several interlaboratory exercises were organised within the framework of European FP6 project NORMAN. Among others, non-steroidal anti-inflammatory drugs were investigated in different aqueous samples in two sequential ring studies. The aim of both studies was to evaluate the state-of-art in Europe and to determine possible sources of variation, while also attempting to diminish them. In the present paper we discuss the results of the 2nd Interlaboratory study, while the results of 1st round were presented before. The main scope of the 1st exercise organised within NORMAN project was to assess the laboratory proficiency regardless of the analytical method applied, to evaluate the stability of the target compounds during sample storage, and to define possible sources of variation during sample shipment, storage and analysis. In the 2nd round we primarily aimed to diminish these sources of variation by applying two predetermined analytical protocols based on liquid chromatography-mass spectrometry or gas chromatography-mass spectrometry. The two analytical protocols were compared in terms of their ability to determine individual analytes in matrices of different complexity, i.e. tap water, river water and wastewater. Furthermore, the 2nd exercise addressed also the filtration and compared the influence of different filter material categories on the analysis of non-steroidal anti-inflammatory drugs. Results presented herein evaluate laboratory performance using z-score, bias, proximity and Youden plots. Overall, the laboratory performances were found to be satisfactory for determining NSAIDs in aqueous samples. The two analytical protocols, LC-MS and GC-MS, are assessed according to their sensitivity and measurement uncertainty, where the GC-MS proved superior for the analysis of Ibuprofen, Ketoprofen and Naproxen in matrices with higher complexity. Finally, neither the filtration itself, nor the filter materials were shown to significantly affect the determination of NSAIDs.

The contribution of voxel-based morphometry in staging patients with mild cognitive impairment.

OBJECTIVE: To assess whether different patterns of regional gray matter loss in patients with mild cognitive impairment (MCI) are associated with different risks of conversion to Alzheimer disease (AD), using MRI and voxel-based morphometry (VBM). METHODS: The authors recruited 22 patients with MCI, 22 patients with probable AD, and 20 healthy subjects (HS). T1 volumes from each subject were postprocessed according to an optimized VBM protocol. All patients were clinically followed up (mean [SD] time = 28.7 [5.7] months), and patients with MCI were reclassified into two groups (converters and nonconverters to AD). RESULTS: When comparing patients with AD to HS, widespread areas of reduced gray matter density were found predominantly in temporal, frontal, and parietal lobes and in the insula. Comparing MCI converters and nonconverters with HS, the converters showed more widespread areas of reduced gray matter density than nonconverters, with a pattern of abnormalities similar to that seen in patients with AD. Conversely, when comparing the same groups with patients with AD, MCI nonconverters showed a pattern of gray matter density similar to that of HS. Areas of decreased gray matter density were also found in MCI converters compared with nonconverters. CONCLUSIONS: Different patterns of gray matter density distribution in patients with mild cognitive impairment may be associated to different rates of conversion to Alzheimer disease.

Low-tech electrophoresis, small but beautiful, and effective: electrophoretic titration curves of proteins.

Migration across a stationary pH gradient results in the electrophoretic titration of a protein's dissociable groups. From the resulting curves, some properties of the protein may be derived, including overall amino acid composition and type of mutation between polymorphic variants, as well as range of stability or, for enzymes, of catalytic activity. Analysis with this technique is a stringent purity criterion; other applications allow the study of interacting systems and the planning of chromatographic fractionations based on differences in surface charge.

Interactions between carbonic anhydrase and its inhibitors revealed by gel electrophoresis and circular dichroism.

Structural properties, and especially the differential stability, of complexes between carbonic anhydrase (CA) and three sulfonamide inhibitors, acetazolamide, dorzolamide and methazolamide, were investigated by spectroscopic and electrophoretic techniques. These included denaturant gradient gel electrophoresis either across a urea or a steady-state transverse sodium dodecyl sulfate (SDS) gradient. Acetazolamide, the smallest and most hydrophilic of the sulfonamides, forms the most stable complex in the presence of urea, whereas dorzolamide, with a bulky and hydrophobic structure, is most stable against the effects of SDS. At pH 7.4, complexes with dorzolamide show minimal changes in mobility across the SDS gradient, as if unaffected by the detergent, both in the presence and in the absence of excess ligand in the gel. When bound to both acetazolamide and methazolamide, on the other hand, CA displays an increase in mobility above 0.05% SDS, lower in the presence than in the absence of excess ligand. The finding of a distinct pattern for the unliganded enzyme, however, suggests the complexes can still retain the ligand, although binding of the surfactant changes their charge density. Under saturating conditions and in the presence of SDS, the surface charge of all complexes is much lower than for unliganded, denatured CA. Circular dichroism (CD) spectra clearly indicate that the increase in secondary structure and the decrease in tertiary structure brought about in CA by the presence of low concentrations of SDS are largely prevented by complexing with the inhibitors. These observations point out peculiar properties of each CA inhibitor, of potential value in the definition of their biological activities and also in the potential development of novel antagonist molecules.

Effects of hypothermic perfusion on pulmonary circulation in isolated rabbit lung.

The effects of hypothermic perfusion have been studied by using different perfusates in 24 isolated rabbit lung preparations, divided into three groups: G1, perfused with blood (hematocrit of 10%) and G2 and G3, perfused with erythrocyte-free plasma plus 6% protein in saline. In both G1 and G2 groups left atrial pressures were kept below airway pressure (Zone II conditions), and in G3 it was higher than airway pressure (Zone III conditions). Perfusate flow, pulmonary artery pressure, pulmonary vascular resistance, left atrial pressure, fluid filtration rate, colloid osmotic pressure and temperature were not different (p > 0.1) between G1 and G2 at the beginning of the experiments. Lowering perfusate temperature from 38 degrees C to 28 degrees C produced a significant increase in pulmonary artery pressure and pulmonary vascular resistance in G1 but they decreased in G2 lungs (p < 0.05). Fluid filtration rate increased in both groups during hypothermia. These responses were not inhibited by an alpha-adrenergic receptor blocker or a pulmonary vasodilator. In G3 lungs no changes were observed. The differences in the hemodynamic effects of hypothermia observed in G1 and G2, both in Zone II conditions, could result from the differences in the vessel distention state obtained by each of the perfusate before initiating hypothermia. As perfusate viscosity increase with cold, a greater possibility of vessel distention in G2 lungs occurs. This explains the decrease in pulmonary artery pressure and pulmonary vascular resistance with cold in this group. The increase in fluid filtration rate observed with hypothermia in G1 and G2 may be due to increases in fluid exchange area.