Membrane-binding and activation of LKB1 by phosphatidic acid is essential for development and tumour suppression.

The serine/threonine kinase LKB1 regulates various cellular processes such as cell proliferation, energy homeostasis and cell polarity and is frequently downregulated in various tumours. Many downstream pathways controlled by LKB1 have been described but little is known about the upstream regulatory mechanisms. Here we show that targeting of the kinase to the membrane by a direct binding of LKB1 to phosphatidic acid is essential to fully activate its kinase activity. Consequently, LKB1 mutants that are deficient for membrane binding fail to activate the downstream target AMPK to control mTOR signalling. Furthermore, the in vivo function of LKB1 during development of Drosophila depends on its capacity to associate with membranes. Strikingly, we find LKB1 to be downregulated in malignant melanoma, which exhibit aberrant activation of Akt and overexpress phosphatidic acid generating Phospholipase D. These results provide evidence for a fundamental mechanism of LKB1 activation and its implication in vivo and during carcinogenesis.

Inactivation of the LKB1-AMPK signaling pathway does not contribute to salivary gland tumor development - a short report.

PURPOSE: Activation of AMPK by the tumor suppressor LKB1 represents an essential gatekeeping step for cells under energetic stress to prevent their growth and proliferation by inhibiting mTOR activation, until the energy supply normalizes. The LKB1/AMPK pathway is frequently downregulated in various types of cancer, thereby uncoupling tumor cell growth and proliferation from energy supply. As yet, little information is available on the role of the LKB1/AMPK pathway in tumors derived from salivary gland tissues. METHODS: We performed LKB1 protein expression and AMPK and mTOR activation analyses in several salivary gland tumor types and their respective healthy control tissues using immunohistochemistry. RESULTS: No significant downregulation of LKB1 expression or decreased activation of AMPK or mTOR were observed in any of the salivary gland tumors tested. In contrast, we found that the salivary gland tumors exhibited an increased rather than a decreased AMPK activation. Although the PI3K/Akt pathway was found to be activated in most of the analyzed tumor samples, the unchanged robust activity of LKB1/AMPK likely prevents (over)activation of mTOR. CONCLUSION: In contrast to many other types of cancer, inactivation or downregulation of the LKB1/AMPK pathway does not substantially contribute to the pathogenesis of salivary gland tumors.

Interleukin-15 and soluble interleukin-15 receptor α in coronary artery disease patients: association with epicardial fat and indices of adipose tissue distribution.

Interleukin-15 (IL-15) is a pro-inflammatory cytokine which signals via a specific alpha receptor subunit (IL-15Rα). Increased IL-15 level has been observed in cardiovascular patients and IL-15 immunoreactivity has been detected at vulnerable atherosclerotic plaques. Due to the association between adipose tissue distribution, inflammation and coronary artery disease (CAD), we quantified IL-15 and IL-15Rα in CAD patients with different adiposity and adipose tissue distribution and we evaluated whether epicardial adipose tissue (EAT), a visceral fat depot surrounding and infiltrating myocardium, may be a source of both molecules. IL-15 and IL-15Rα proteins were quantified by enzyme-linked immunosorbent assays. Gene expression of IL-15 and IL-15Rα in EAT depots was evaluated by one colour microarray platform. EAT thickness was measured by echocardiography. Plasmatic IL-15 and IL-15Rα levels were higher in CAD than non-CAD patients. After classification according to adipose tissue distribution, IL-15 was higher in CAD patients with increased abdominal adiposity. Increased level of IL-15Rα was observed both in CAD and non-CAD patients with increased abdominal fat. EAT was a source of IL-15 and IL-15Rα and their expression was higher in CAD patients with increased EAT thickness. In conclusion, our data suggest that circulating levels of IL-15 and IL-15Rα seem to reflect visceral distribution of adipose tissue and that EAT may be a potential source of both IL-15 and IL-15Rα. Future studies on the relationship between IL-15, visceral fat and characteristics of atherosclerotic plaques could help to better understand the complex biology of this cytokine.

Low heart-type fatty acid binding protein level during aging may protect down syndrome people against atherosclerosis.

BACKGROUND: Aging is considered an important independent risk factor for atherosclerosis. Down syndrome people (DS) display an accelerated aging process compared to healthy subjects, anyway they are relatively resistant to developing atherosclerosis. The mechanisms involved in such protective effect are not well known. Since heart-type fatty acid binding protein (H-FABP) is a protein involved in the transport of fatty acids and it has been recently correlated with the presence of atherosclerosis, we aimed to measure H-FABP level both in DS and in healthy subjects during aging to evaluate the association between this molecule, aging and atherosclerosis. FINDINGS: We quantified plasmatic H-FABP level in three groups of male DS and age-matched healthy subjects (children, age 2-14 years; adults, age 20-50 years; elderly, > 60 years) using a biochip array analyzer. We observed that aging is associated with increased H-FABP level in healthy subjects but not in DS which display both the same protein level in the different ages of life and have also lower level compared to their age-matched healthy subjects. CONCLUSION: Reduced H-FABP level during aging in DS may play a protective role against atherosclerosis. The potential involvement of H-FABP in the relationship between aging, atherosclerosis and development of coronary artery disease needs further investigations.

Bone remodelling biomarkers after whole body cryotherapy (WBC) in elite rugby players.

Whole body cryotherapy (WBC) consists of a brief exposure to extreme cold air (-110°C) in a controlled chamber and it is applied in sports medicine to improve recovery from musculoskeletal trauma. The aim of this study is to better define the beneficial effect of WCB on the musculoskeletal system of athletes, in particular on bone remodelling. Remodelling osteoimmunological biomarkers OPG, RANKL and RANK were measured after WBC treatment in 10 male rugby players randomly selected from the Italian National team. OPG levels were increased significantly, supporting the view that WBC induces an osteogenic effect. Further studies evaluating the effect of WBC on bone metabolism are desirable.

Adipokines, hormonal parameters, and cardiovascular risk factors: similarities and differences between patients with erectile dysfunction of arteriogenic and nonarteriogenic origin.

INTRODUCTION: Erectile dysfunction (ED) is often associated with metabolic disorders. Leptin and adiponectin are adipose tissue-derived hormones involved in the regulation of metabolic homeostasis and considered important players in the relationship among obesity and cardiovascular diseases. AIM: Leptin, adiponectin, leptin to adiponectin ratio (L/A), and their correlation with hormonal and metabolic parameters were examined in male with arteriogenic- (A-ED) and nonarteriogenic-ED (NA-ED). MAIN OUTCOME MEASURES: Biochemical, metabolic, and hormonal parameters of men with A-ED were compared with those of male with NA-ED. METHODS: Diagnosis of ED was based on the International Index of Erectile Function Score. Its etiology was classified with penile echo-color Doppler at baseline and after intracavernous injection of prostaglandin E1. Leptin and adiponectin were measured by enzyme-linked immunosorbent assay. RESULTS: In A-ED subjects, increased levels of insulin, glycated hemoglobin, homeostasis model assessment of insulin resistance (HOMA-IR) index, body mass index (BMI), leptin, and L/A and decreased levels of total, free, and bioavailable testosterone were observed compared with NA-ED subjects. A trend toward lower estradiol level was also present in A-ED patients, even if not statistically significant. Reduced levels of adiponectin have been observed in both groups compared with patients without ED. Leptin and L/A correlated similarly with several parameters (negatively with testosterone/estradiol ratio and positively with BMI, insulin, HOMA-IR, and 17-beta estradiol). L/A resulted further correlated negatively with high-density lipoprotein and positively with triglycerides. CONCLUSIONS: Not all ED cases are similar. In fact, A-ED patients display a more complicated metabolic status characterized by overweight and obesity and associated to sexual hormone alteration. Whether changes in body composition and modulation of adipokine levels can improve local endothelial function need further investigation.

Natural zeolites chabazite/phillipsite/analcime increase blood levels of antioxidant enzymes.

Imbalance between reactive oxygen species generation and antioxidant capacity induces a condition known as oxidative stress which is implicated in numerous pathological processes. In this study we evaluated whether natural zeolites chabazite/phillipsite/analcime may affect the levels of different antioxidant enzymes (gluthatione peroxidase, superoxide dismutase, gluthatione reductase), total antioxidant status and oxidative stress in 25 clinically healthy men, both non-smokers and smokers. Measurements were performed on whole blood or on plasma samples before (T0) and after 4-weeks zeolites intake (T1). At T1, gluthatione peroxidase, superoxide dismutase and gluthatione reductase increased compared to T0 levels, both considering all subjects as joint and after subdivision in non-smokers and smokers. Differently, a reduction in total antioxidant status was observed at T1. Anyway, total antioxidant status resulted higher than the reference values in both groups at each time point. A decrease in lipid peroxidation, a major indicator of oxidative stress assessed by monitoring thiobarbituric acid reactive substances, was also observed in all subjects at T1. Our results suggested that chabazite/phillipsite/analcime may help to counteract oxidative stress in apparently healthy subjects exposed to different oxidative stress risk factors, such as smoking, thus representing a particular kind of food with potential antioxidant properties.

Serum amyloid a and C-reactive protein independently predict the recurrences of atrial fibrillation after cardioversion in patients with preserved left ventricular function.

BACKGROUND: Subclinical inflammation and atrial stretch have been recognized as important contributors to atrial fibrillation (AF) onset and perpetuation. The aim of the study was to compare the predictive role of serum inflammatory markers (serum amyloid A [SAA], and C-reactive protein [CRP]) and N-terminal pro brain natriuretic peptide (NT-proBNP) an indice of atrial strain in relation to subacute arrhythmic recurrence rate in patients with persistent AF and normal left ventricular ejection fraction (LVEF). METHODS: We studied 57 patients with a mean LVEF of 58.7 ± 6%. NT-proBNP, SAA and CRP levels were determined few hours before electrical cardioversion and 3 weeks after cardioversion. RESULTS: Subacute AF recurrences were documented in 19 (33 %) patients. Whereas NT-proBNP levels did not predict arrhythmic outcome, higher SAA (> 6.16-6.19 mg/L) and CRP levels (> 2.99-3.10 mg/L) were significantly associated with AF recurrences (odds ratio [OR], 5.39; 95% confidence interval [CI], 1.59-18.26; P = 0.007 and OR, 14.93; 95% CI, 3.90-57.19; P < 0.001). Both SAA (OR, 18.29; 95% CI, 2.07-161.46; P = 0.009) and high sensitivity CRP (OR, 42.03; 95% CI, 4.83-365.45; P = 0.001) through the multivariate logistic regression analysis show an independent role in predicting the AF recurrence with a sensitivity of 100% (38/38) and a specificity of 52.6% (10/19). CONCLUSIONS: The present study demonstrates that in patients with persistent AF and preserved LVEF, SAA and CRP levels are independent predictors of AF subacute recurrence rate, whereas NT-proBNP, not associated with arrhythmic outcome, reflects the hemodynamic alterations secondary to arrhythmia presence. The simultaneous determination of SAA and high sensitivity CRP has a very high sensitivity (100%) in predicting the AF recurrence.

Italian air force acrobatic pilots are protected against flight-induced oxidative stress.

BACKGROUND/AIM: The purpose of this study was to assess the oxidative stress of aircraft pilots by evaluating different markers of oxidative stress and any imbalance between free radicals and antioxidants in plasma. PATIENTS AND METHODS: A group of 13 supersonic aircraft pilots, following regular exercise and personalized diet, were compared with a group of 40 healthy controls. Oxidative stress indicators, such as reactive oxidative metabolites, carbonyl proteins, 8-hydroxy-2-deoxyguanosine and total antioxidant status, were evaluated after three months of intense flight. RESULTS: Reactive oxygen metabolites, carbonyl protein and 8-hydroxy-2-deoxyguanosine plasma levels did not differ in supersonic aircraft pilots and healthy controls. The two groups also had similar total antioxidant status levels. CONCLUSION: We suggest that supersonic aircraft pilots working at high altitude, even if exposed to physiological stresses, can, with proper diet, regular exercise and periodical medical examinations, maintain a healthy balance between oxidant and antioxidant status.

Carbon dioxide-enriched water inhalation in patients with allergic rhinitis and its relationship with nasal fluid cytokine/chemokine release.

BACKGROUND AND AIMS: Allergic rhinitis is characterized by eosinophil infiltration and accumulation in the nasal mucosa mainly due to IL-3, IL-5, and eotaxin activities. We undertook this study to investigate a possible in vivo effect of carbon dioxide-enriched water inhalation in patients with allergic rhinitis. METHODS: Twenty five consecutive patients inhaled carbon dioxide-enriched water at Fonti di Rabbi Spa Centre (Trento, Italy). Symptom scores for nasal obstruction, itching and sneezing were obtained before and after treatment. Nasal lavage was collected, and IL-3, IL-5, and eotaxin levels were assessed using the quantitative sandwich enzyme immunoassay technique. Cytometric analysis was performed on samples to measure total cell count, CD45+ cells, and percentages of polymorphonucleates and lymphocytes. RESULTS: There were statistically significant differences in chemokine levels and in cell populations between patients and healthy controls before treatment. After carbon dioxide-enriched water inhalation, we observed statistically significant improvements in symptom scores, chemokine levels, and percentages of cell populations. CONCLUSIONS: Our results seem to confirm the role of IL-3, IL-5, and eotaxin in the pathophysiology of allergy and the beneficial effect of carbon dioxide-enriched water inhalation in patients affected by allergic rhinitis.

Reduced plasma levels of P-selectin and L-selectin in a pilot study from Alzheimer disease: relationship with neuro-degeneration.

Neurodegenerative processes associated with Alzheimer's disease (AD) are accompanied by reactive astrogliosis and microglia activation and a role for chronic inflammation in the brain degeneration of these patients has been suggested. Moreover impaired immune functions in AD brains might also influence the disease's progression. Therefore, it is of interest to further characterized inflammatory molecules in the peripheral blood of patients with AD and its relationship with cognitive decline. A complex picture emerged in this pilot study and IL-8, IFN-gamma, MCP-1 and VEGF levels were increased in AD. Levels of P-selectin and L-selectin were decreased in AD and lowest in AD patients with highest cognitive decline. Our findings suggest that these molecules may induce alterations of endothelial regulation and influence neurodegenerative processes of AD.

Plasma concentrations of angiogenetic factors and angiogenetic inhibitors in patients with ductal pancreatic neoplasms. A pilot study.

BACKGROUND: The aim of the study was to evaluate the circulating concentrations of vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor-2 (VEGFR-2), vascular endothelial growth factor-D (VEGF-D) and endostatin in patients with intraductal papillary mucinous neoplasm (IPMN), and in those with ductal adenocarcinomas. METHODS: Sixty patients (32 males, 28 females, mean age 69.3±11.3 years) were enrolled: 31 (51.7%) had IPMNs and 29 (48.3%) had histologically confirmed pancreatic adenocarcinomas. Thirty blood donors were also studied as controls. In all study subjects, the concentrations of VEGF, VEGF-D, VEGFR-2, and endostatin were determined using enzyme-linked immunosorbent assays. RESULTS: Serum concentrations of VEGF, VEGF-D, and VEGFR-2 were significantly higher in patients with pancreatic ductal adenocarcinoma and those with IPMNs compared with healthy subjects, while endostatin was significantly higher only in patients with pancreatic ductal adenocarcinoma compared with healthy subjects. Within the group of patients, VEGFR-2 was significantly higher in patients with ductal adenocarcinoma compared to those with IPMNs. The sensitivity and the specificity of VEGFR-2 in differentiating patients with ductal adenocarcinomas from those with IPMN at a cut-off range of 4003-4034 pg/mL was 86.2% and 54.8%, respectively. CONCLUSIONS: IPMNs have serum VEGFR-2 concentrations different from those in patients with ductal adenocarcinomas. However, serum VEGFR-2 cannot be routinely utilized to differentiate IPMNs from pancreatic ductal adenocarcinomas.

Plasma and drainage fluid levels of soluble receptor activator of nuclear factor-kB (sRANK), soluble receptor activator of nuclear factor-kB ligand (sRANKL) and osteoprotegerin (OPG) during proximal humerus fracture healing.

Fracture healing is an ordered process that restores the structural integrity of the bone. Soluble receptor activator of nuclear factor-kB (sRANK), its ligand (sRANKL) and osteoprotegerin (OPG) are involved in bone remodelling, thus they may play a role in fracture repair. OPG, soluble RANK and RANKL levels were measured in plasma and in drainage fluid, collected from pre-surgery phase to healing in ten patients of both genders (age range 26-65 years) with proximal humerus fracture needing osteosynthesis. All patients showed fracture healing. No significant modifications in the concentrations of sRANKL and OPG were observed, while sRANK showed a significant increase in drainage fluid 24 hours post-surgery compared with intra-surgery time. OPG levels were higher in plasma and drainage fluid than sRANK and sRANKL at each time point. Since there are no published data about sRANK involvement in fracture healing, our study represents the first preliminary indication about a local increase of this marker concentration immediately after surgery.

Serum neutrophil gelatinase-B associated lipocalin (NGAL) levels in Down's syndrome patients.

Neutrophil gelatinase-associated lipocalin (NGAL) is a group of proteins with different functions.NGAL is released by different cell types such as epithelial cell, hepatocytes and renal tubular cells during inflammation and after cell injury. Expression of NGAL is induced under various pathophysiological conditions such as infection, cancer, inflammation, kidney injury, cardiovascular disease, burn injury, and intoxication, which has an important anti-apoptotic and anti-inflammatory role.Subjects with Down's syndrome (DS) are affected by many pathological age related conditions such as mental retardation, Alzheimer's disease, immune defects and increased susceptibility to infections. The aim of this study is to evaluate possible use of NGAL as a marker of inflammatory status for allow an early diagnosis of inflammatory disease such as autoimmune disease in DS patients, that are more susceptible to these pathologies, especially in elderly subjects.In this study were recruited 3 groups of DS subjects (children, adults and elderly) and compared them to healthy control group.The molecules of interest was determinated by immuno-enzymatic assay (ELISA).Our results show that NGAL plasmatic level was significantly higher in DS patients compared to healthy controls. Moreover NGAL levels increase in correlation with the age, and showed a significantly correlation between the increase with the severity of disease.DS is characterized by an enhancement of gene production such as GART, SOD-1 and CBS that encode specific protein and enzyme involved in hydrogen peroxide and superoxide production, species highly cytotoxic implicated in inflammation and ageing.NGAL may have the potential application to ameliorate the toxicity induced by oxidative stress conditions such as Alzheimer's disease, thalassemia, cardiovascular disease, burn injury, transplantation, diabetes, and aging.

The role of inflammation in patients with intraductal mucinous neoplasm of the pancreas and in those with pancreatic adenocarcinoma.

BACKGROUND: There are very few data regarding inflammation in patients with intraductal papillary mucinous neoplasms (IPMNs) of the pancreas. AIM: To evaluate the circulating concentrations of placental growth factor (PlGF), transforming growth factor-alpha (TGF-α), transforming growth factor-beta1 (TGF-ß1), tumour necrosis factor receptor 1 (TNF-R1) and matrix metalloproteinase-2 (MMP-2) in patients with IPMNs and in those with pancreatic adenocarcinomas. PATIENTS AND METHODS: Sixty-nine patients were enrolled: 23 (33.3%) had IPMNs and 46 (66.7%) had histologically confirmed pancreatic adenocarcinomas. Thirteen healthy subjects were also studied. PlGF, TGF-α, TGF-ß1, TNF-R1 and MMP-2 were determined using commercially available kits. RESULTS: TNF-R1 (p=0.003) was the only protein significantly different among the three groups. CONCLUSION: Serum TNF-R1 was elevated in patients with IPMNs and in those with pancreatic adenocarcinomas, suggesting a high apoptotic activity in both groups of patients studied.

The natural antioxidant alpha-lipoic acid induces p27(Kip1)-dependent cell cycle arrest and apoptosis in MCF-7 human breast cancer cells.

Unlike normal cells, tumor cells survive in a specific redox environment where the elevated reactive oxygen species contribute to enhance cell proliferation and to suppress apoptosis. Alpha-lipoic acid, a naturally occurring reactive oxygen species scavenger, has been shown to possess anticancer activity, due to its ability to suppress proliferation and to induce apoptosis in different cancer cell lines. Since at the moment little information is available regarding the potential effects of alpha-lipoic acid on breast cancer, in the present study we addressed the question whether alpha-lipoic acid induces cell cycle arrest and apoptosis in the human breast cancer cell line MCF-7. Moreover, we investigated some molecular mechanisms which mediate alpha-lipoic acid actions, focusing on the role of the PI3-K/Akt signalling pathway. We observed that alpha-lipoic acid is able to scavenge reactive oxygen species in MCF-7 cells and that the reduction of reactive oxygen species is followed by cell growth arrest in the G1 phase of the cell cycle, via the specific inhibition of Akt pathway and the up-regulation of the cyclin-dependent kinase inhibitor p27(kip1), and by apoptosis, via changes of the ratio of the apoptotic-related protein Bax/Bcl-2. Thus, the anti-tumor activity of alpha-lipoic acid observed in MCF-7 cells further stresses the role of redox state in regulating cancer initiation and progression.