Resistance to osimertinib in advanced EGFR-mutated NSCLC: a prospective study of molecular genotyping on tissue and liquid biopsies.

BACKGROUND: Resistance to osimertinib in advanced EGFR-mutated non-small cell lung cancer (NSCLC) constitutes a significant challenge for clinicians either in terms of molecular diagnosis and subsequent therapeutic implications. METHODS: This is a prospective single-centre study with the primary objective of characterising resistance mechanisms to osimertinib in advanced EGFR-mutated NSCLC patients treated both in first- and in second-line. Next-Generation Sequencing analysis was conducted on paired tissue biopsies and plasma samples. A concordance analysis between tissue and plasma was performed. RESULTS: Sixty-five advanced EGFR-mutated NSCLC patients treated with osimertinib in first- (n = 56) or in second-line (n = 9) were included. We managed to perform tissue and liquid biopsies in 65.5% and 89.7% of patients who experienced osimertinib progression, respectively. Acquired resistance mechanisms were identified in 80% of 25 patients with post-progression samples, with MET amplification (n = 8), EGFR C797S (n = 3), and SCLC transformation (n = 2) the most frequently identified. The mean concordance rates between tissue and plasma for the EGFR activating mutation and for the molecular resistance mechanisms were 87.5% and 22.7%, respectively. CONCLUSIONS: Resistance to osimertinib demonstrated to be highly heterogeneous, with MET amplification the main mechanism. Plasma genotyping is a relevant complementary tool which might integrate tissue analysis for the study of resistance mechanisms.

PD-L1 overexpression induces STAT signaling and promotes the secretion of pro-angiogenic cytokines in non-small cell lung cancer (NSCLC).

BACKGROUND: Monoclonal antibodies (ICI) targeting the immune checkpoint PD-1/PD-L1 alone or in combination with chemotherapy have demonstrated relevant benefits and established new standards of care in first-line treatment for advanced non-oncogene addicted non-small cell lung cancer (NSCLC). However, a relevant percentage of NSCLC patients, even with high PD-L1 expression, did not respond to ICI, highlighting the presence of intracellular resistance mechanisms that could be dependent on high PD-L1 levels. The intracellular signaling induced by PD-L1 in tumor cells and their correlation with angiogenic signaling pathways are not yet fully elucidated. METHODS: The intrinsic role of PD-L1 was initially checked in two PD-L1 overexpressing NSCLC cells by transcriptome profile and kinase array. The correlation of PD-L1 with VEGF, PECAM-1, and angiogenesis was evaluated in a cohort of advanced NSCLC patients. The secreted cytokines involved in tumor angiogenesis were assessed by Luminex assay and their effect on Huvec migration by a non-contact co-culture system. RESULTS: PD-L1 overexpressing cells modulated pathways involved in tumor inflammation and JAK-STAT signaling. In NSCLC patients, PD-L1 expression was correlated with high tumor intra-vasculature. When challenged with PBMC, PD-L1 overexpressing cells produced higher levels of pro-angiogenic factors compared to parental cells, as a consequence of STAT signaling activation. This increased production of cytokines involved in tumor angiogenesis largely stimulated Huvec migration. Finally, the addition of the anti-antiangiogenic agent nintedanib significantly reduced the spread of Huvec cells when exposed to high levels of pro-angiogenic factors. CONCLUSIONS: In this study, we reported that high PD-L1 modulates STAT signaling in the presence of PBMC and induces pro-angiogenic factor secretion. This could enforce the role of PD-L1 as a crucial regulator of the tumor microenvironment stimulating tumor progression, both as an inhibitor of T-cell activity and as a promoter of tumor angiogenesis.

A comparison of node vaccination strategies to halt SIR epidemic spreading in real-world complex networks.

We compared seven node vaccination strategies in twelve real-world complex networks. The node vaccination strategies are modeled as node removal on networks. We performed node vaccination strategies both removing nodes according to the initial network structure, i.e., non-adaptive approach, and performing partial node rank recalculation after node removal, i.e., semi-adaptive approach. To quantify the efficacy of each vaccination strategy, we used three epidemic spread indicators: the size of the largest connected component, the total number of infected at the end of the epidemic, and the maximum number of simultaneously infected individuals. We show that the best vaccination strategies in the non-adaptive and semi-adaptive approaches are different and that the best strategy also depends on the number of available vaccines. Furthermore, a partial recalculation of the node centrality increases the efficacy of the vaccination strategies by up to 80%.

White Blood Cells in Patients Treated with Programmed Cell Death-1 Inhibitors for Non-small Cell Lung Cancer.

PURPOSE: To investigate whether eosinophils and other white blood cell subtypes could be used as response and prognostic markers to anti-Programmed cell Death-1 or anti-PD-Ligand-1 treatments in non-small cell lung cancer patients. METHODS: We retrospectively analyzed data from the NSCLC patients consecutively treated at our hospital with a PD-1/PD-L1 inhibitor in monotherapy for advanced disease. A total of 191 patients were evaluated at three time-points to investigate any relation between tumor response and WBC counts. RESULTS: Baseline WBC and subtypes did not differ according to the type of response seen under treatment. A higher relative eosinophil count (REC) correlated with more objective responses (p = 0.019 at t1 and p = 0.014 at t2; OR for progression = 0.54 and 0.53, respectively) independently of the smoking status, PD-L1 status, and immune-related toxicity (IRT). Higher REC was also associated with a longer duration of treatment (p = 0.0096). Baseline absolute neutrophil count was prognostic (p = 0.049). At t1 relative lymphocytes, absolute and relative neutrophils, and neutrophil-to-lymphocyte ratio were prognostic (p = 0.044, p = 0.014, p = 0.0033, and p = 0.029, respectively). CONCLUSION: Our results show that in NSCLC patients anti-PD-1/PD-L1 therapy induces an early increase only in blood eosinophils, more prominent in responding patients and independent of the smoking status, PD-L1 status, and IRT. Eosinophils are also associated with a longer duration of treatment. Furthermore, our data support a prognostic role of neutrophils, lymphocytes, and their ratio for NSCLC patients with advanced disease treated with PD(L)-1 blockade.

Patient-reported outcomes after oesophagectomy in the multicentre LASER study.

BACKGROUND: Data on the long-term symptom burden in patients surviving oesophageal cancer surgery are scarce. The aim of this study was to identify the most prevalent symptoms and their interactions with health-related quality of life. METHODS: This was a cross-sectional cohort study of patients who underwent oesophageal cancer surgery in 20 European centres between 2010 and 2016. Patients had to be disease-free for at least 1 year. They were asked to complete a 28-symptom questionnaire at a single time point, at least 1 year after surgery. Principal component analysis was used to assess for clustering and association of symptoms. Risk factors associated with the development of severe symptoms were identified by multivariable logistic regression models. RESULTS: Of 1081 invited patients, 876 (81.0 per cent) responded. Symptoms in the preceding 6 months associated with previous surgery were experienced by 586 patients (66.9 per cent). The most common severe symptoms included reduced energy or activity tolerance (30.7 per cent), feeling of early fullness after eating (30.0 per cent), tiredness (28.7 per cent), and heartburn/acid or bile regurgitation (19.6 per cent). Clustering analysis showed that symptoms clustered into six domains: lethargy, musculoskeletal pain, dumping, lower gastrointestinal symptoms, regurgitation/reflux, and swallowing/conduit problems; the latter two were the most closely associated. Surgical approach, neoadjuvant therapy, patient age, and sex were factors associated with severe symptoms. CONCLUSION: A long-term symptom burden is common after oesophageal cancer surgery.

A comparative analysis of link removal strategies in real complex weighted networks.

In this report we offer the widest comparison of links removal (attack) strategies efficacy in impairing the robustness of six real-world complex weighted networks. We test eleven different link removal strategies by computing their impact on network robustness by means of using three different measures, i.e. the largest connected cluster (LCC), the efficiency (Eff) and the total flow (TF). We find that, in most of cases, the removal strategy based on the binary betweenness centrality of the links is the most efficient to disrupt the LCC. The link removal strategies based on binary-topological network features are less efficient in decreasing the weighted measures of the network robustness (e.g. Eff and TF). Removing highest weight links first is the best strategy to decrease the efficiency (Eff) in most of the networks. Last, we found that the removal of a very small fraction of links connecting higher strength nodes or of highest weight does not affect the LCC but it determines a rapid collapse of the network efficiency Eff and the total flow TF. This last outcome raises the importance of both to adopt weighted measures of network robustness and to focus the analyses on network response to few link removals.

Randomized clinical trial of psychological support and sleep adjuvant measures for postoperative sleep disturbance in patients undergoing oesophagectomy.

BACKGROUND: Major surgery such as oesophagectomy requires a postoperative stay in intensive care. Painful stimuli lead to sleep disturbance and impairment in quality of life. The aim of this study was to evaluate the effect of psychological counselling and sleep adjuvant measures on postoperative quality of sleep and quality of life. METHODS: This RCT was performed between January 2013 and October 2015. Patients undergoing oesophagectomy for cancer were randomized into one of four groups receiving: psychological counselling plus sleep adjuvant measures during the ICU stay; psychological counselling alone; sleep adjuvant measures alone during the ICU stay; or standard care. The primary endpoint was impairment in quality of life measured using the European Organisation for Research and Treatment of Cancer C30-QL2 questionnaire between admission for surgery and discharge from hospital. The secondary endpoint was impairment in quality of sleep assessed by means of the Pittsburgh Sleep Quality Index between admission for surgery and hospital discharge. RESULTS: The local ethics committee approved the early termination of the study because of relevant changes in the ICU setting. Some 87 patients were randomized and 74 patients were evaluated in the analysis. Psychological counselling reduced the impairment in quality of life (odds ratio 0·23, 95 per cent c.i. 0·09 to 0·61) and in quality of sleep (odds ratio 0·27, 0·10 to 0·73). CONCLUSION: Perioperative psychological support reduces impairment in quality of life and quality of sleep after oesophagectomy. Registration number: NCT01738620 (http://www.clinicaltrials.gov).

Hybrid minimally invasive esophagectomy for cancer: impact on postoperative inflammatory and nutritional status.

The purpose of this case-control study was to evaluate the impact of hybrid minimally invasive esophagectomy for cancer on surgical stress response and nutritional status. All 34 consecutive patients undergoing hybrid minimally invasive esophagectomy for cancer at our surgical unit between 2008 and 2013 were retrospectively compared with 34 patients undergoing esophagectomy with open gastric tubulization (open), matched for neoadjuvant therapy, pathological stage, gender and age. Demographic data, tumor features and postoperative course (including quality of life and systemic inflammatory and nutritional status) were compared. Postoperative course was similar in terms of complication rate. Length of stay in intensive care unit was shorter in patients undergoing hybrid minimally invasive esophagectomy (P = 0.002). In the first postoperative day, patients undergoing hybrid minimally invasive esophagectomy had lower C-reactive protein levels (P = 0.001) and white cell blood count (P = 0.05), and higher albumin serum level (P = 0.001). In this group, albumin remained higher also at third (P = 0.06) and seventh (P = 0.008) postoperative day, and C-reactive protein resulted lower at third post day (P = 0.04). Hybrid minimally invasive esophagectomy significantly improved the systemic inflammatory and catabolic response to surgical trauma, contributing to a shorter length of stay in intensive care unit.

Esophagectomy in elderly patients: which is the best prognostic score?

Our study aimed to identify the best prognostic score for fitness for surgery and postoperative morbidity in elderly patients. A prospectively collected database of a consecutive series of patients with esophageal cancer evaluated for possible esophagectomy at our unit was analyzed. Fitness for surgery and postoperative morbidity were used as measures of outcome. The performances of the Physiological and Operative Severity Score for the enUmeration of Mortality and morbidity (POSSUM) score, the Charlson Comorbidity Index, the age-related Charlson Comorbidity Index (ACCI), the American Society of Anesthesiologists scale and the prognostic nutritional index (PNI) were evaluated in elderly patients. Discrimination was measured with receiver operating characteristics curve analysis; calibration was assessed by the Hosmer-Lemeshow goodness-of-fit test. Age did not result a significant predictor for postoperative complications. In elderly patients, ACCI predicted the judgment of the multidisciplinary team about fitness for surgery with the best discrimination (C-index = 0.94). PNI had the best discrimination for postoperative complications (C-index = 0.71) in the elderly group. ACCI best predicted the fitness for surgery in elderly patients. In elderly patients, the most discriminative prognostic score for postoperative complication was PNI, which could be used at admission for surgery to correctly inform patients about their risk and, possibly, to take extra precaution in case of high risk.

Impact of esophagectomy for cancer on patients' occupational status.

BACKGROUND: The aim of our study was to investigate the impact of esophagectomy for cancer on patients' occupational status. METHODS: All 109 consecutive patients presenting with esophageal cancer to the Surgical Oncology Unit of the Veneto Institute of Oncology Padua (Italy) between November 1, 2009 and March 15, 2012, were included in the study. Information on occupational status at diagnosis and at 1 year after esophagectomy was retrieved. Health-related quality of life was evaluated at discharge after surgery using selected aspects of the EORTC QLQ-C30 questionnaire. Non parametric statistics were used. RESULTS: Sixty-one patients (49.6%) were active workers at diagnosis and 50 of them (82.0%) underwent esophagectomy. Eighteen active workers (18/50, 36.0%) quit their job within one year from esophagectomy. They received jejunostomy more often than patients still working after surgery (50.0% vs. 18.8%, respectively; p = 0.03) and reported lower social functioning at discharge (mean ± SD 63.6 ± 16.4 vs. 80.2 ± 25.6 in others, p = 0.02). Multivariable analysis identified jejunostomy as independent predictor of job-quitting at 1 year after esophagectomy (p = 0.03; OR 4.75, 95% C.I. 1.11-20.39) but not social functioning at discharge (p = 0.21). CONCLUSIONS: Patients should be informed that they may experience social and work disability due to cancer treatment and adequate interventions of return-to-work support should be provided. Adequate welfare strategy should be implemented for esophageal cancer survivors, enhancing their role competences and contributing to precision care medicine.

Postoperative outcome after oesophagectomy for cancer: Nutritional status is the missing ring in the current prognostic scores.

BACKGROUND: Several prognostic scores were designed in order to estimate the risk of postoperative adverse events. None of them includes a component directly associated to the nutritional status. The aims of the study were the evaluation of performance of risk-adjusted models for early outcomes after oesophagectomy and to develop a score for severe complication prediction with special consideration regarding nutritional status. METHODS: A comparison of POSSUM and Charlson score and their derivates, ASA, Lagarde score and nutritional index (PNI) was performed on 167 patients undergoing oesophagectomy for cancer. A logistic regression model was also estimated to obtain a new prognostic score for severe morbidity prediction. RESULTS: Overall morbidity was 35.3% (59 cases), severe complications (grade III-V of Clavien-Dindo classification) occurred in 20 cases. Discrimination was poor for all the scores. Multivariable analysis identified pulse, connective tissue disease, PNI and potassium as independent predictors of severe morbidity. This model showed good discrimination and calibration. Internal validation using standard bootstrapping techniques confirmed the good performance. CONCLUSIONS: Nutrition could be an independent risk factor for major complications and a nutritional status coefficient could be included in current prognostic scores to improve risk estimation of major postoperative complications after oesophagectomy for cancer.

Complications after surgery for colorectal cancer affect quality of life and surgeon-patient relationship.

AIM: This multicentric prospective study aimed to investigate how postoperative complications after surgery for colorectal cancer affect patients' quality of life and satisfaction with care. METHOD: One hundred and sixteen patients operated on for colorectal cancer were enrolled in this study. Patients answered three questionnaires about generic (EORTC QLQ-C30) and disease-specific (EORTC QLQ-CR29) quality of life and treatment satisfaction (EORTC IN-PATSAT32) at the time of admission and at 1 and 6 months after surgery. Non-parametric tests and linear multiple regression models were used for statistical analysis. RESULTS: Twelve patients had complications requiring further surgery (anastomotic leakage, abdominal bleeding, abdominal wall sepsis, wound infection). Patients with complications that required surgery reported a worse score of physical function, emotional function and anxiety than patients without such complications 1 month after surgery. These patients judged their general satisfaction with the quality of care and doctors' interpersonal skills, technical skills, information provision and availability to be worse than in patients without such complications. The presence of postoperative psychiatric complications and anastomotic leakage were independent predictors of quality of life (ß = -0.30, P = 0.004, and ß = -0.42, P < 0.001). CONCLUSION: In patients undergoing surgery for colorectal cancer, complications requiring any kind of surgical management significantly affected patients' perception of all doctor-related items suggesting an impairment of the entire surgeon-patient relationship. Convincing patients that 'zero risk' cannot be achieved in surgical practice is therefore a priority.

Cytocompatibility and cellular internalization mechanisms of SiC/SiO2 nanowires.

First evidence of in vitro cytocompatibility of SiC/SiO2 core-shell nanowires is reported. Different internalization mechanisms by adenocarcinomic alveolar basal epithelial cells, monocytic cell line derived from an acute monocytic leukemia, breast cancer cells, and normal human dermal fibroblasts are shown. The internalization occurs mainly for macropinocytosis and sporadically by direct penetration in all cell models considered, whereas it occurred for phagocytosis only in monocytic leukemia cells. The cytocompatibility of the nanowires is proved by the analysis of cell proliferation, cell cycle progression, and oxidative stress on the cells treated with NWs as compared to controls. Reactive oxygen species generation was detected as an early event that then quickly run out with a rapid decrease only in adenocarcinomic alveolar basal epithelial and human dermal fibroblasts cells. In all the cell lines, the intracellular presence of NWs induce the same molecular events but to a different extent: peroxidation of membrane lipids and oxidation of proteins. The NWs do not elicit either midterm (72 h) or long-term (10 days) cytotoxic activity leading to irreversible cellular damages or death. Our results are important in view of a possible use of SiC/SiO2 core-shell structures acting as biomolecule-delivery vectors or intracellular electrodes.

Epidermal growth factor receptor irreversible inhibitors: chemical exploration of the cysteine-trap portion.

Covalent EGFR irreversible inhibitors showed promising potential for the treatment of gefitinib-resistant tumors and for imaging purposes. They contain a cysteine-reactive portion forming a covalent bond with the protein. Irreversible kinase inhibitors have been advanced to clinical studies, mostly characterized by an acrylamide or butynamide warhead. However, the clinical usefulness of these compounds has been hampered by resistances, toxicity and pharmacokinetic problems. Investigation on the structure-activity and structure-reactivity relationships may provide useful information for compounds with improved selectivity and pharmacokinetic properties. This review focuses on the exploration of the cysteine-trap portions able to irreversibly inhibit EGFR and other erbB receptors.

Modeling the cell cycle: from deterministic models to hybrid systems.

The cell cycle is a complex biological system frequently investigated from a mathematical perspective. In fact, over the past years a huge number of deterministic mathematical models describing the dynamics and the regulation of this process have been proposed. A crucial point concerning the cell cycle modeling is the combination of continuous and discrete dynamics in order to obtain results which are coherent with the biological context. To face with this problem we propose a novel approach to the mathematical modeling of biological processes based on the use of hybrid systems. This new methodology essentially consists in a model reduction (using the modified Prony's method) which allows to define the crucial features of the dynamical system. The final aim is to implement a corresponding hybrid system which preserves the properties of the starting deterministic model. Thus, we implemented a methodology which allows to describe the cellular system by combining continuous behavior with discrete events by using the hybrid automata technology. In this way we try to overcome some drawbacks of the deterministic approach, especially regarding the possibility to introduce new variables during simulation and the associated variation of parameters in a more efficient way than the continuous method can do. We applied this innovative methodology to the reconstruction of a simplified hybrid model concerning one of the crucial mammalian cell cycle control point. In particular, we investigated the role of the transcription factors E2F in the R-point transition. The resulting hybrid model preserve the properties of the deterministic one and it allows the identification of the parameter which controls the transition from the inactive (quiescent) to the active state (R-point transition) after the mitogenic stimulation. At the best of our knowledge no hybrid model for the R-point transition are available in literature.

Effect of inducible FHIT and p53 expression in the Calu-1 lung cancer cell line.

Loss of FHIT expression and p53 mutations are critical events in the early stages of lung carcinogenesis. The restoration of Fhit function in FHIT-negative cancer cells has been reported to cause tumour suppression by inhibition of cell proliferation and/or activation of apoptotic pathways. However, the studies designed to elucidate the biological role of Fhit and its potential interaction with p53 have produced conflicting results. We investigated here the effects of the simultaneous restoration of FHIT and p53 in Calu-1 cells by using a hormone-inducible gene expression system. We demonstrate that the restoration of FHIT expression reinforces the anti-proliferative effect associated with the simultaneous replacement of p53. Indeed, a more pronounced inhibition of cell proliferation associated with an earlier and higher induction of p21(waf1) mRNA and protein expression was observed in Fhit/p53-expressing cells compared with cells expressing p53 alone. This effect was not due to Fhit-mediated up-regulation of p53 expression; in fact p53 protein was expressed at the same level in both FHIT-positive and FHIT-negative cell clones. Consistent with this result, Fhit did not affect the expression of MDM2, a protein known to interact directly with p53 and target p53 for proteolytic degradation, thus down-regulating its activity.

Cell size reduction induced by inhibition of the mTOR/S6K-signaling pathway protects Jurkat cells from apoptosis.

In Jurkat cells, the decreased cell growth rate associated with a long-lasting deactivation of the mammalian target of rapamycin (mTOR)/p70 ribosomal S6 kinase (S6K)-signaling pathway generates a cell population of progressively reduced cellular mass and size. When promoted by rapamycin as prototype inhibitor, the mTOR deactivation-dependent cell size reduction was associated with slowed, but not suppressed, proliferation. Small-size cells were significantly protected from apoptosis induced by Fas/Apo-1 death-receptor activation (as shown by reduced procaspase cleavage and decreased catalytic activity of relevant caspases) or by stress signals-dependent mitochondrial perturbation (as shown by reduced cleavage of caspase-2, lower dissipation of mitochondrial membrane potential and decreased release of cytochorome c and apoptosis-inducing factor from mitochondria). Protection faded when reactivation of the mTOR/S6K pathway promoted the cell recovery to normal size. These results suggest that cells induced to reduce their mass by the mTOR deactivation-dependent inhibition of cell growth become more resilient to lethal assaults by curbing the cell's suicidal response.

Use of IgG Avidity test in case definitions of toxoplasmosis in pregnancy.

A survey network for congenital toxoplasmosis (TOXO-NET) was set up in December 1996 in Piedmont (Italy). Participants were asked to classify the infections in pregnant mothers and newborns by the criteria of the European Network on Congenital Toxoplasmosis published by Lebech in 1996. Because the IgG Avidity test is largely employed as a 2nd level test in toxoplasmosis diagnosis and it could be helpful to date infection, the co-ordinators of TOXO-NET suggested including it in the "case definition" of "probable" infection and "unlikely" infection. 117 cases of toxoplasmosis in pregnancy divided into the risk categories under Lebech's criteria were re-examined using the "new" case definitions. 77 out of 117 (65.8%) Toxoplasma gondii infections during pregnancy could be defined with only one serum sample using the IgG Avidity test. The IgG Avidity test proved a useful method to classify the Toxoplasma gondii infections in pregnancy, especially when we had only one serum sample.

[Progression of chronic renal failure in remnant rats: role of arginase inhibition]. / Progressione dell'insufficienza renale cronica nel ratto remnant: ruolo della inibizione dell'arginasi.

BACKGROUND: Oral administration of arginine to remnant (REM) rats (5/6 nx) slows the progression of chronic renal failure through a nitric-oxide(NO)-dependent mechanism. We have recently shown that inhibition of arginase, the main metabolic pathway of arginine, was able to induce similar results on renal dynamics (GIN: 2001, 18:285-290). Aim of the present study was to test whether these changes were mediated by increased availability of arginine-derived NO. Methods. Three Groups of REM rats were studied for 8 weeks after surgery: 1) untreated REM (Group REM); 2) REM rats treated with arginine (1%) in tap water (Group ARG); 3) REM rats administered a Mn++-free diet, to induce partial inhibition of arginase (Group MNF). Normal unmanipulated rats were used as controls (Group NOR). RESULTS: Liver arginase activity was significantly depressed only in MNF-rats (-35% vs. REM, p < 0.01). Blood pressure was significantly lower in Group MNF vs. ARG and REM after 6 weeks (p < 0.05). Proteinuria was significantly decreased in Group ARG (-42%, p < 0.05 vs. REM) and even more in Group MNF (-57%, p < 0.01). ARG plasma levels, decreased in REM rats (-41% vs. Group CON), were normalized in Group ARG (p < 0.01 vs. Group REM); arginase inhibition was able to increase such levels in Group MNF (+38% vs. REM) and this resulted in a proportional rise in urinary nitrite excretion (+33% vs. REM), grossly depressed in REM rats. Renal arginase activity was lower in all the Groups of remnant rats vs. Group NOR, but intrarenal concentrations of ARG were significantly lower only in rats of Group MNF (p < 0.05 vs. all the other Groups). Histological examination showed that MNF-rats had a glomerular sclerosis index lower than in the other Groups (p < 0.05 vs. Group REM and ARG). CONCLUSIONS: In conclusion, inhibition of arginase in remnant rats slows the progression of CRF and preserves renal histology through a direct and/or indirect NO-dependent mechanism.

Evaluation of circulating levels and renal clearance of natural amino acids in patients with Cushing's disease.

Although the hypercortisolism-induced impairment of protein homeostasis is object of several studies, a detailed evaluation of the complete amino acid profile of patients with Cushing's syndrome (CS) has never been performed. The aim of the current open transversal controlled study was to evaluate serum and urinary concentrations as well as renal clearance of the complete series of natural amino acids and their relationship with glucose tolerance in patients with Cushing's disease (CD). Twenty patients with CD (10 active and 10 cured) and 20 sex- and age-matched healthy controls entered the study. Measurement of serum and urinary levels of the complete series of natural amino acids was performed in all patients analyzed by cationic exchange high performance liquid cromatography (HPLC) after 2 weeks of a standardized protein intake regimen. The renal clearance (renal excretion rate) of each amino acid was calculated on the basis of the serum and urinary concentrations of creatinine and the specific amino acid. Fasting glucose and insulin levels, glucose and insulin response to standard glucose load, insulinogenic and homeostasis model insulin resistance (Homa-R) indexes were also evaluated and correlated to the circulating levels and renal clearances of each amino acid. Significantly higher serum (p<0.01) and urinary (p<0.05) levels of alanine and cystine, lower serum and higher urinary levels of leucine, isoleucine and valine (p<0.05) and higher renal excretion rates of leucine, isoleucine and valine (p<0.01) were found in patients with active CD than in patients cured from the disease and in controls. No difference was found between cured patients and controls. Creatinine clearance was similar in active and cured patients and in controls. In patients with active CD, urinary cortisol levels were significantly correlated to urinary cystine levels (r=0.85; p<0.01) and renal excretion rate of leucine (r=-0.76; p<0.05), isoleucine (r=-0.76; p<0.05) and valine (r=-0.66; p<0.05). Fasting blood glucose levels were significantly correlated to serum alanine levels (r=0.70; p<0.05). Although Homa-R was significantly correlated to BMI in active patients (r=0.74 p<0.05), it was not correlated to amino acid levels. In conclusion, the results of the current study demonstrate that patients with CD have significant changes in serum and urinary concentration of several amino acids and changes in renal clearance of some specific amino acids. Normalization of cortisol levels restored the amino acid profile.