Source and symptoms of COVID-19 among hospital workers in Milan.

BACKGROUND: Healthcare workers (HCWs) are commonly infected by SARS-CoV-2 and represent one of the most vulnerable groups. Adequate prevention strategies are necessary to guarantee HCWs' safety, as well as to prevent dissemination of the infection among patients. AIMS: To describe a case series of SARS-CoV-2-positive HCWs in a large public healthcare organization in Milan (Italy) during the most devastating weeks of the epidemic and analyse the sources, symptoms and duration of SARS-CoV-2 infection. METHODS: This study included 172 SARS-CoV-2-positive HCWs who were infected between the 25th of February and the 7th of April 2020. A nasopharyngeal swab (NPS) and RT-PCR were used to indicate. RESULTS: Initially, the most common sources of infection were other positive HCWs (49%). Medical doctors and nursing assistants were most frequently infected, with infection rates of 53/1000 and 50/1000, respectively. COVID-19 departments were less affected than internal medicine, surgery, intensive care, or emergency room. The most commonly reported symptom was mild cough, while loss of smell (anosmia) and loss of taste (ageusia) were reported as moderate and severe by 30-40% of HCWs. The time necessary for 50% of workers to recover from the infection was 23 days, while it took 41 days for 95% of HCWs to become virus-free. CONCLUSIONS: HCWs are commonly infected due to close contacts with other positive HCWs, and non-COVID departments were most affected. Most HCWs were asymptomatic or subclinical but contact tracing and testing of asymptomatic HCWs help identify and isolate infected workers.

Circulating innate immune markers and outcomes in treatment-naïve advanced non-small cell lung cancer patients.

INTRODUCTION: Innate immunity represents the first step of activation of the immune system and dictates the quality of adaptive immune responses. Studies have reported links between systemic inflammatory or innate immune markers and prognosis in patients with lung cancer. To our knowledge, the prospective and concomitant study of these systemic markers has never been performed. METHODS: Advanced treatment-naive non-small cell lung cancer (NSCLC) patients eligible for first-line platinum-based chemotherapy were prospectively included from December 2012 to July 2015 (N = 148). Blood samples of patients were collected before the first cycle for fresh NK cell phenotyping. Peripheral blood mononuclear cells were cryopreserved for natural cytotoxicity receptor (NCR) genotyping as well as sera for NCR's ligand quantification. Data on leukocytes, neutrophils and monocyte counts and lactate dehydrogenase (LDH) levels were extracted from electronic medical records. RESULTS: Among all studied markers, monocytosis, neutrophilia, leucocytosis, high LDH and sBAG6 levels and reduced levels of NCR3 transcripts were associated with poor overall survival (OS) in univariate analysis. The levels of NCR3 transcripts was linked to age, number of metastatic sites, monocyte counts, LDH and sBAG6 levels. Neutrophilia was associated to high sBAG6 levels. NCR3 was the unique innate immune parameter that remained as an independent factor associated with both OS (P = 0.003) and progression-free survival (P = 0.009) in the multivariate analysis. CONCLUSION: This study brought evidence that these biomarkers are entangled; parameters associated with an inflammatory process were related to reduced levels of NCR3 transcripts. Finally, the level of NCR3 transcripts was independently associated with outcomes in treatment-naive patients with advanced NSCLC.

Biological recognition of graphene nanoflakes.

The systematic study of nanoparticle-biological interactions requires particles to be reproducibly dispersed in relevant fluids along with further development in the identification of biologically relevant structural details at the materials-biology interface. Here, we develop a biocompatible long-term colloidally stable water dispersion of few-layered graphene nanoflakes in the biological exposure medium in which it will be studied. We also report the study of the orientation and functionality of key proteins of interest in the biolayer (corona) that are believed to mediate most of the early biological interactions. The evidence accumulated shows that graphene nanoflakes are rich in effective apolipoprotein A-I presentation, and we are able to map specific functional epitopes located in the C-terminal portion that are known to mediate the binding of high-density lipoprotein to binding sites in receptors that are abundant in the liver. This could suggest a way of connecting the materials' properties to the biological outcomes.

Association of free-living physical activity measures with metabolic phenotypes in type 2 diabetes at the time of diagnosis. The Verona Newly Diagnosed Type 2 Diabetes Study (VNDS).

BACKGROUND AND AIM: Lifestyle is considered a major determinant of risk of type 2 diabetes (T2D). We investigated whether daily physical activity (DPA) is associated with beta-cell function (BF) and/or insulin sensitivity (IS) in patients with T2D at the time of diagnosis. METHODS AND RESULTS: In 41 subjects enrolled in the Verona Newly-Diagnosed Type 2 Diabetes Study we assessed: (1) IS, by euglycaemic insulin clamp; (2) BF, estimated by prolonged-OGTT minimal modeling and expressed as derivative and proportional control; (3) DPA and energy expenditure (EE), assessed over 48-h monitoring by a validated wearable armband system. Study participants (median [IQR]; age: 62 [53-67] years, BMI: 30.8 [26.5-34.3] Kg m-2, HbA1c: 6.7 [6.3-7.3]%; 49.7 [45.4-56.3] mmol/mol) were moderately active (footsteps/day: 7773 [5748-10,927]; DPA≥3MET: 70 [38-125] min/day), but none of them exercised above 6 metabolic equivalents (MET). EE, expressed as EETOT (total daily-EE) and EE≥3MET (EE due to DPA≥3MET) were 2398 [2226-2801] and 364 [238-617] Kcal/day, respectively. IS (M-clamp 630 [371-878] µmol/min/m2) was positively associated with DPA and EE, independent of age, sex and BMI (p < 0.05). Among the DPA and EE parameters assessed, DPA≥3MET and EETOT were independent predictors of IS in multivariable regression analyses, adjusted for age, sex, BMI (R2 = 16%, R2 = 19%, respectively; p < 0.01). None of model-derived components of BF was significantly associated with DPA or accompanying EE. CONCLUSIONS: Our study highlighted moderate levels of DPA and total EE as potential determinants of IS, but not BF, in T2D at the time of diagnosis. Intervention studies are needed to conclusively elucidate the effect of DPA on these features. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. UNIQUE IDENTIFIER: NCT01526720.

Baseline gut microbiota predicts clinical response and colitis in metastatic melanoma patients treated with ipilimumab.

BACKGROUND: Ipilimumab, an immune checkpoint inhibitor targeting CTLA-4, prolongs survival in a subset of patients with metastatic melanoma (MM) but can induce immune-related adverse events, including enterocolitis. We hypothesized that baseline gut microbiota could predict ipilimumab anti-tumor response and/or intestinal toxicity. PATIENTS AND METHODS: Twenty-six patients with MM treated with ipilimumab were prospectively enrolled. Fecal microbiota composition was assessed using 16S rRNA gene sequencing at baseline and before each ipilimumab infusion. Patients were further clustered based on microbiota patterns. Peripheral blood lymphocytes immunophenotypes were studied in parallel. RESULTS: A distinct baseline gut microbiota composition was associated with both clinical response and colitis. Compared with patients whose baseline microbiota was driven by Bacteroides (cluster B, n = 10), patients whose baseline microbiota was enriched with Faecalibacterium genus and other Firmicutes (cluster A, n = 12) had longer progression-free survival (P = 0.0039) and overall survival (P = 0.051). Most of the baseline colitis-associated phylotypes were related to Firmicutes (e.g. relatives of Faecalibacterium prausnitzii and Gemmiger formicilis), whereas no colitis-related phylotypes were assigned to Bacteroidetes. A low proportion of peripheral blood regulatory T cells was associated with cluster A, long-term clinical benefit and colitis. Ipilimumab led to a higher inducible T-cell COStimulator induction on CD4+ T cells and to a higher increase in serum CD25 in patients who belonged to Faecalibacterium-driven cluster A. CONCLUSION: Baseline gut microbiota enriched with Faecalibacterium and other Firmicutes is associated with beneficial clinical response to ipilimumab and more frequent occurrence of ipilimumab-induced colitis.

Detecting the shape of anisotropic gold nanoparticles in dispersion with single particle extinction and scattering.

The shape and size of nanoparticles are important parameters affecting their biodistribution, bioactivity, and toxicity. The high-throughput characterisation of the nanoparticle shape in dispersion is a fundamental prerequisite for realistic in vitro and in vivo evaluation, however, with routinely available bench-top optical characterisation techniques, it remains a challenging task. Herein, we demonstrate the efficacy of a single particle extinction and scattering (SPES) technique for the in situ detection of the shape of nanoparticles in dispersion, applied to a small library of anisotropic gold particles, with a potential development for in-line detection. The use of SPES paves the way to the routine quantitative analysis of nanoparticles dispersed in biologically relevant fluids, which is of importance for the nanosafety assessment and any in vitro and in vivo administration of nanomaterials.

Towards a classification strategy for complex nanostructures.

The range of possible nanostructures is so large and continuously growing, that collating and unifying the knowledge connected to them, including their biological activity, is a major challenge. Here we discuss a concept that is based on the connection of microscopic features of the nanomaterials to their biological impacts. We also consider what would be necessary to identify the features that control their biological interactions, and make them resemble each other in a biological context.

Is common genetic variation at IRS1, ENPP1 and TRIB3 loci associated with cardiometabolic phenotypes in type 2 diabetes? An exploratory analysis of the Verona Newly Diagnosed Type 2 Diabetes Study (VNDS) 5.

BACKGROUND AND AIMS: Insulin resistance is a hallmark of type 2 diabetes (T2DM), it is often accompanied by defective beta-cell function (BF) and is involved in the pathophysiology of cardiovascular disease (CVD). Commonalities among these traits may recognize a genetic background, possibly involving the genetic variation of insulin signaling pathway genes. We conducted an exploratory analysis by testing whether common genetic variability at IRS1, ENPP1 and TRIB3 loci is associated with cardiovascular risk traits and metabolic phenotypes in T2DM. METHODS AND RESULTS: In 597 drug-naïve, GADA-negative, newly-diagnosed T2DM patients we performed: 1) genotyping of 10 independent single-nucleotide polymorphisms covering ∼ 90% of common variability at IRS1, ENPP1 and TRIB3 loci; 2) carotid artery ultrasound; 3) standard ECG (n = 450); 4) euglycaemic insulin clamp to assess insulin sensitivity; 5) 75 g-OGTT to estimate BF (derivative and proportional control) by mathematical modeling. False discovery rate of multiple comparisons was set at 0.20. After adjustment for age, sex and smoking status, rs4675095-T (IRS1) and rs4897549-A (ENPP1) were significantly associated with carotid atherosclerosis severity, whilst rs7265169-A (TRIB3) was associated with ECG abnormalities. Rs858340-G (ENPP1) was significantly associated with decreased insulin sensitivity, independently of age, sex and body-mass-index. No consistent relationships were found with BF. CONCLUSION: Some associations were found between intermediate phenotypes of CVD and common genetic variation of gatekeepers along the insulin signaling pathway. These results need be replicated to support the concept that in T2DM the CVD genetic risk clock may start ticking long before hyperglycemia appears. ClinicalTrials.gov Identifier: NCT01526720.

Type 2 diabetes mellitus: a disease of the governance of the glucose-insulin system: an experimental metabolic control analysis study.

BACKGROUND AND AIMS: The relatives role of each component of the glucose-insulin system in determining hyperglycemia in type 2 diabetes is still under debate. Metabolic Control Analysis (MCA) quantifies the control exerted by each component of a system on a variable of interest, by computing the relevant coefficients of control (CCs), which are systemic properties. We applied MCA to the intravenous glucose tolerance test (IVGTT) to quantify the CCs of the main components of the glucose-insulin system on intravenous glucose tolerance. METHODS AND RESULTS: We combined in vivo phenotyping (IVGTT/euglycaemic insulin clamp) and in silico modeling (GLUKINSLOOP.1) to compute the CCs of intravenous glucose tolerance in healthy insulin-sensitive (n = 9, NGR-IS), healthy insulin-resistant (n = 7, NGR-IR) and subdiabetic hyperglycemic (n = 8, PreT2DM) individuals and in patients with newly diagnosed type 2 diabetes (n = 7, T2DM). Altered insulin secretion and action were documented in NGR-IR and PreT2DM groups, but only 1st phase insulin secretion was significantly lower in T2DM than in PreT2DM (p < 0.05). The CCs changed little in the nondiabetic groups. However, several CCs were significantly altered in the patients (e.g. CCs of beta cell: -0.75 ± 0.10, -0.64 ± 0.15, -0.56 ± 0.09 and -0.19 ± 0.04 in NGR-IS, NGR-IR, PreT2DM and T2DM, respectively; p < 0.01 by MANOVA), and they could not be corrected by matching in silico nondiabetic and T2DM groups for 1st phase secretion. CONCLUSIONS: Type 2 diabetes is characterized not only by loss of function of the elements of the glucose-insulin system, but also by changes in systemic properties (CCs). As such, it could be considered a disease of the governance of the glucose-insulin system.

Variants and haplotypes of TCF7L2 are associated with ß-cell function in patients with newly diagnosed type 2 diabetes: the Verona Newly Diagnosed Type 2 Diabetes Study (VNDS) 1.

CONTEXT: Intronic variants of TCF7L2 are confirmed genetic risk factors for type 2 diabetes and are associated to alterations in beta cell function in nondiabetic individuals. OBJECTIVE: The objective of the study was to test whether TCF7L2 variability may affect ß-cell function also in patients with type 2 diabetes. DESIGN: This was a cross-sectional association study. SETTING: The study was conducted at a university hospital referral center for diabetes. PATIENTS: Patients included 464 (315 males and 149 females) glutamic acid decarboxylase-negative patients [age: median 59 yr (interquartile range: 52-65); body mass index: 29.3 kg/m(2) (26.5-32.9); fasting plasma glucose: 7.0 mmol/liter (6.1-8.0)] with newly diagnosed type 2 diabetes. INTERVENTION(S): Interventions included frequently sampled oral glucose tolerance test and euglycemic insulin clamp. MAIN OUTCOME MEASURE(S): ß-Cell function (derivative control and proportional control); insulin sensitivity; genotypes of the following TCF7L2 single-nucleotide polymorphisms: rs7901695, rs7903146, rs11196205, and rs12255372. RESULTS: Both rs7901695 and rs7903146 diabetes risk alleles were associated with reduced proportional control of ß-cell function (P = 0.019 and P = 0.022, respectively). Two low-frequency haplotypes were associated with extreme (best and worst) phenotypes of ß-cell function (P < 0.01). No associations between TCF7L2 genotypes and insulin sensitivity were detected. CONCLUSIONS: TCF7L2 diabetes risk variants, either as single-nucleotide polymorphisms or as haplotypes, detrimentally influence ß-cell function and might play a role in determining the metabolic phenotype of patients with newly diagnosed type 2 diabetes.

Prevalence of nosocomial infections in Italy: result from the Lombardy survey in 2000.

A one-day survey was carried out in 88 out of 113 public hospitals in Lombardy to obtain prevalence rates of hospital-acquired infections (HAIs) by hospital departments and to identify the pathogens more frequently involved. In total 18667 patients were surveyed, representing 72% of the average daily total of occupied beds in public hospitals in Lombardy. The overall prevalence of HAI was 4.9%. The highest prevalence was observed in intensive care units and in spinal units. The prevalence of bloodstream infections was 0.6%; pneumonia 1.1%; urinary tract infections 1.6% and gastrointestinal infections 0.4%. In surgical patients the prevalence of surgical site infections was 2.7%. The most frequently isolated pathogen from all sites of infections was Escherichia coli (16.8%), followed by Staphylococcus aureus (15.0%), Pseudomonas aeruginosa (13.2%) and Candida spp. (8.7%). Methicillin-resistant S. aureus accounted for 23% of all isolated S. aureus. The results provide baseline data for rational priorities in allocation of resources, for further studies and for infection control activities.

Altered blood pressure variability in patients with congestive heart failure.

OBJECTIVE: Congestive heart failure (CHF) is characterized by sympathetic overactivity but reduced variability of heart interval and sympathetic nerve activity; little information exists, however, about the alterations in blood pressure variability in this syndrome, especially during excitatory manoeuvres such as tilting or exercise. DESIGN AND METHODS: Nine patients with CHF (age 62+/-1 years, NYHA class II-III, ejection fraction 33+/-1%, peak VO2 14.1+/-3.2 ml/min per kg body weight [mean +/- SEM]) and eight healthy control subjects (age 58+/-1 years) with normal left ventricular function were studied. Blood pressure (Finapres), R-R interval (ECG) and respiration (nasal thermistor) were recorded during 15-min periods of supine rest, 70 degree head-up tilting, submaximal bicycling exercise and post-exercise recovery. Total variance and the power of the spectral components of blood pressure (HF, respiratory-related; LF, 0.03-0.14 Hz; and VLF, 0.02-0.003 Hz) were measured. RESULTS: Compared with control subjects, CHF patients have, first, a normal overall blood pressure variability during supine rest but a failure to increase this variability in response to head-up tilt and exercise; second, a suppressed LF spectral component of blood pressure at rest and in response to head-up tilt and exercise; and third, reappearance of LF blood pressure power during postexercise recovery. CONCLUSIONS: In CHF patients, overall blood pressure variability and its LF spectral component are altered at rest and during sympathoexcitatory manoeuvres. Somewhat paradoxically, however, the depressed LF blood pressure power is partially restored during a 15-min recovery period, indicating that at least part of the CHF-related alterations of blood pressure variability have the potential to revert back towards normal under appropriate physiological circumstances.

Prevention and treatment of vasospasm.

Cerebral vasospasm is a frequent and severe complication of SAH. Angiographic vasospasm may be seen in 70% of patients and delayed cerebral ischemic deficits are observed in 30% of patients. Since vasodilator drugs cannot reverse cerebral vasospasm, treatment is directed to prevent vasospasm and to prevent or reverse ischemic deficits. The mainstay of treatment of vasospasm is the hypertensive hypervolemia dilution (triple H therapy); the mainstay of prevention is the calcium channel blocker nimodipine. The efficacy of triple H therapy has not been demonstrated in randomized clinical trials, while several randomized trials have demonstrated that nimodipine reduces poor outcome due to vasospasm in all grades of patients. Some randomized, clinical trials are recently performed on the efficacy of rTPA (on the basis of the correlation between the amount of cisternal blood and the incidence and severity of vasospasm) and of tirilazed (on the basis of the role of lipidic peroxidation and free radical generation in the pathogenesis of spasm). Balloon angioplasty and/or super-selective intra-arterial infusion of papaverine can be considered when patient is refractory to medical and pharmacological treatment.

Contribution of the neuro-intensivist in the choice of treatment of patients with aneurysm.

During the last years embolization with Guglielmi detachable coils has provided a new alternative of treatment of intracranial aneurysms. Neuroanesthesiologists and neurointensivists have an important role in the selection of the patients to traditional surgical treatment or endovascular treatment. This selection must be a team decision on the basis of the patient's conditions. The main contribution of neuroanesthesiologists is to evaluate the patient's medical and neurologic conditions and to prospect the anesthesiological problems in each technique, especially when the patient is treated in acute phase after SAH.

[Anesthesia in early surgery and endovascular therapy for aneurysmic subarachnoid hemorrhage]. / Anestesia nella chirurgia precoce dell'ESA aneurismatica e nei trattamenti endovascolari.

Early surgery after SAH is frequently performed. The most important problems for anesthesiologists are the risk of rebleeding, the alteration of autoregulation and CO2 responsiveness, cardiac, respiratory and electrolytic alterations. In this phase the brain may be ischemic-edematous or haemorrhagic-compressive and the choice of anesthetic agent is made on the basis of cerebral conditions. The main goal is to control ICP and maintain adequate CPP. The endovascular treatment with Guglielmi detachable coils is usually performed in patients with poor neurologic and/or medical conditions. General anaesthesia under aggressive monitoring is advisable to control systemic pressure and to avoid movements.

Parotid gland tumors: a comparison of postoperative radiotherapy techniques using three dimensional (3D) dose distributions and dose-volume histograms (DVHS).

PURPOSE: To compare different treatment techniques for unilateral treatment of parotid gland tumors. METHODS AND MATERIALS: The CT-scans of a representative parotid patient were used. The field size was 9 x 11 cm, the separation was 15.5 cm, and the prescription depth was 4.5 cm. Using 3D dose distributions, tissue inhomogeneity corrections, scatter integration (for photons) and pencil beam (for electrons) algorithms and dose-volume histogram (DVH), nine treatment techniques were compared. [1] unilateral 6 MV photons [2] unilateral 12 MeV electrons [3] unilateral 16 MeV electrons [4] an ipsilateral wedge pair technique using 6 MV photons [5] a 3-field AP (wedged), PA (wedged) and lateral portal technique using 6 MV photons [6] a mixed beam technique using 6 MV photons and 12 MeV electrons (1:4 weighting) [7] a mixed beam technique using 6 MV photons and 16 MeV electrons (1:4 weighting) [8] a mixed beam technique using 18 MV photons and 20 MeV electrons (2:3 weighting) [9] a mixed beam technique using 18 MV photons and 20 MeV electrons (1:1 weighting). RESULTS: Using dose-volume histograms to evaluate the dose to the contralateral parotid gland, the percentage of contralateral parotid volume receiving > or = 30% of the prescribed dose was 100% for techniques [1], [8] and [9], and < 5% for techniques [2] through [7]. Evaluating the "hottest" 5 cc of the ipsilateral mandible and temporal lobes, the hot spots were: 152% and 150% for technique [2], 132% and 130% for technique [6]. Comparing the exit doses, techniques [1], [8] and [9] contributed to > or = 50% of the prescribed dose to the contralateral mandible and the temporal lobes. Only techniques [2] and [6] kept the highest point doses to both the brain stem and the spinal cord below 50% of the prescribed dose. CONCLUSION: The single photon lateral field [1] and the mixed electron-photon beams [8] and [9] are not recommended treatment techniques for unilateral parotid irradiation because of high doses delivered to the contralateral parotid gland and high exit doses which are associated with Xerostomia. The en face electron beam technique [2] and the mixed electron-photon beam technique [6] are unacceptable due to the excessive dose heterogeneity to the contiguous normal structures. In spite of optimal dose fall-off achieved using the en face technique [3], most patients cannot tolerate the resulting high skin doses. We conclude that the ipsilateral wedge pair [4], the 3-field [5], and the mixed electron-photon beam [7] techniques are optimal techniques in providing relatively homogeneous dose distributions within the target area and for minimizing dose to the relevant normal structures.

Propofol anaesthesia in spontaneously breathing paediatric patients during magnetic resonance imaging.

BACKGROUND: The aim of this study was to evaluate the use of propofol to induce and maintain anaesthesia in spontaneously breathing paediatric patients (age 2 weeks-11 years) during Magnetic Resonance Imaging (MRI) of the CNS. METHODS: All patients were spontaneously breathing, without intubation, and received supplemental O2. Pulse rate, blood pressure (BP), electrocardiogram and EtCO2 were recorded in all patients, and in 38 subjects SpO2 was also monitored. Patients were divided in 2 groups according to their body weights: Group A (n = 34, bwt < or = 10 kg), and Group B (n = 48, bwt > 10 kg). RESULTS: Dosage of propofol during the time of induction (from insertion of the i.v. cannula to positioning on the MRI table) was significantly higher in smaller children (Group A; 5.4 +/- 2.2 (SD) mg/kg) as compared to children with bwt above 10 kg (Group B; 3.7 +/- 1.6 mg/kg). Propofol dosage for maintenance of anaesthesia was significantly higher in smaller children (Group A: 10.1 +/- 5.7 vs Group B: 7.1 +/- 3.0 mgkg-1 h-1, P = 0.003). During the time of induction, transient episodes of reduced BP (< or = 20%) occurred in 6 patients in Group A and 2 patients in Group B. During anaesthesia in Group B there was 1 episode of oxygen desaturation (95%), and 3 episodes of short and mild increases of EtCO2(< or = 52 mmHg). No other side effects occurred in any patient. MRI studies were successfully completed, only 3 sequences (Group A) had to be restarted. CONCLUSION: Propofol can be safely used for total intravenous anaesthesia in children undergoing MRI.

Peri-operative prophylaxis with phenytoin: dosage and therapeutic plasma levels.

Early postoperative epilepsy is a frequent complication of supratentorial intracranial surgery. The lack of consensus on prophylaxis of early postoperative seizures with phenytoin (PHT) may be due to the different dosages used in several studies, owing to inadequate therapeutic plasma level. The aim of this study was to evaluate which dosage of PHT can maintain the therapeutic range in the early postoperative period. Twenty patients operated on for supratentorial neoplasms were randomly allocated to receive, during the last hour of the surgical procedure, loading doses of either 10 mg/kg (group A, n = 10) or 15 mg/kg (group B, n = 10) of PHT. PHT infusion rate never exceeded 30 mg/min. Six hours after the loading dose, PHT maintenance treatment (250 mg, i.v., every 8 hours) was started in all patients. PHT plasma levels were evaluated from the end of the intra-operative loading infusion up to 24 h. During the first six hours after the loading dose, phenytoin plasma levels fell below the therapeutic range (10-20 mg/l) in 7 out of the 10 patients receiving 10 mg/kg, while in the patients treated with 15 mg/kg, PHT plasma levels were always in the therapeutic range (P < or = 0.0001). PHT maintenance dose was sufficient to keep plasma levels within the therapeutic range in 8 patients in group A, and in all the patients in group B. It is concluded that a loading dose of 15 mg/kg, followed by postoperative treatment, is necessary to guarantee therapeutic plasma levels of phenytoin in the immediate postoperative period, when seizure risk is very high.

EGF stimulates lamellipod extension in metastatic mammary adenocarcinoma cells by an actin-dependent mechanism.

Changes in lamellipod extension and chemotaxis in response to EGF were analysed for MTLn3 cells (a metastatic cell line derived from the 13762NF rat mammary adenocarcinoma). Addition of EGF produced a cessation of ruffling followed by extension of hyaline lamellipods containing increased amounts of F-actin at the growing edge. A non-metastatic cell line (MTC) derived from the same tumor did not show such responses. Lamellipod extension was maximal within 5 min, followed by retraction and resumption of ruffling. Maximal area increases due to lamellipod extension occurred at about 5 nM EGF. Chemotactic and chemokinetic responses, measured using a microchemotaxis chamber, were also greatest at 5 nM. Cytochalasin D inhibited EGF-stimulated responses including lamellipod extension, increases in F-actin in lamellipods, and chemotaxis. Nocodazole affected chemotaxis at higher concentrations but not EGF-induced lamellipod extension. We conclude that polymerization of F-actin at the leading edges of lamellipods is necessary for extension of lamellipods and chemotaxis of MTLn3 cells in response to EGF. The motility and chemotaxis responses of this metastatic cell line have strong similarities to those seen in well-characterized chemotactic cells such as Dictyostelium and neutrophils.