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Background: Autism spectrum disorder (ASD) diagnoses have rapidly increased globally. Both environmental and genetic factors appear to contribute to the development of ASD. Several studies have shown a potential association between prenatal or postnatal pesticide exposure and the risk of developing ASD. Methods: We reviewed the available literature concerning the relationship between early life exposure to pesticides used in agriculture, such as organochlorines, organophosphates and pyrethroids, and ASD onset in childhood. We searched on Medline and Scopus for cohort or case–control studies published in English from 1977 to 2020. Results: A total of seven articles were selected for the review. We found a remarkable association between the maternal exposure to pyrethroid, as well as the exposure to organophosphate during pregnancy or in the first years of childhood, and the risk of ASD onset. This association was found to be less evident with organochlorine pesticides. Pregnancy seems to be the time when pesticide exposure appears to have the greatest impact on the onset of ASD in children. Conclusions: Among the different environmental pollutants, pesticides should be considered as emerging risk factors for ASD. The potential association identified between the exposure to pesticides and ASD needs to be implemented and confirmed by further epidemiological studies based on individual assessment both in outdoor and indoor conditions, including multiple confounding factors, and using statistical models that take into account single and multiple pesticide residues.

Among therapeutic approaches that have been investigated, targeting of receptors implicated in managing neuroinflammation has been described. One such family of receptors comprises the formyl peptide receptors (FPRs) whose ligands could play a role in host defense. The murine FPR gene family includes at least six members while in humans there are only three. The two most important members are the Fpr1 and Fpr2. Fpr1encodes murine FPR1, which is considered the murine orthologue of human FPR. Resveratrol, a non-flavonoid polyphenol rich in red wine and grapes, apart from its beneficial health effects and anti-inflammatory properties, has been reported to reduce neuroinflammation in different neurodegenerative disease models. Resveratrol anti-inflammatory responses involve the activation of the protein deacetylase sirtuin 1 (SIRT1) gene. In this work we have investigated in an LPS-based murine model of neuroinflammation the role of FPR1, examining not only if this receptor undergoes a reduction of its expression during neuroinflammation, but also whether treatment with resveratrol was able to modulate its expression leading to an amelioration of neuroinflammatory picture in a murine model of neuroinflammation. Results of this work showed that FPR1 together with SIRT1 resulted upregulated by resveratrol treatment and that this increase is associated with an amelioration of the neuroinflammatory picture, as demonstrated by the induction of IL-10 and IL1-RA expression and the downregulation of proinflammatory mediators, such as TNF-α and IL-1β. The expression and the modulation of FPR1 by resveratrol may be evaluated in order to propose a novel anti-inflammatory and pro-resolving therapeutic approach for the reduction of the detrimental effects associated with neuro-inflammation based neurodegenerative diseases and also as a promising strategy to promote human health by a diet rich in antioxidative bioactive compounds.

Vitamin C (Vit C) is anutrient present in many foods, particularly citrus fruits, green vegetables, tomatoes, and potatoes. Vit C is studied for its applications in the prevention and management of different pathologies, including neurodegenerative diseases. Neuroinflammation is a defense mechanism activated by a stimulus or an insult that is aimed at the preservation of the brain by promoting tissue repair and removing cellular debris, however, persistent inflammatory responses are detrimental and may lead to the pathogenesis and progression of neurodegenerative diseases like Parkinson’s disease (PD) and Alzheimer’s disease. PD is one of the most common chronic progressive neurodegenerative disorders, and oxidative stress is one of the most important factors involved in its pathogenesis and progression.Due to this, research on antioxidant and anti-inflammatory compounds is an important target for counteracting neurodegenerative diseases, including PD. In the central nervous system, the presence of Vit C in the brain is higher than in other body districts, but why and how this occurs is still unknown. In this research, Vit C, with its anti-inflammatory and anti-oxidative properties, is studied to better understand its contribution to brain protection, in particular, we have investigated the neuroprotective effects of Vit C in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced animal model of PD and its role in the modulation of neuroinflammation. First, we observed that Vit C significantly decreased the MPTP-induced loss of tyrosine hydroxylase (TH)-positive dopaminergic neuronal cells in the substantia nigra, as well as microglial cell activation and astrogliosis. Furthermore, gait and spontaneous locomotor activity, evaluated by an automated treadmill and the Open Field test, respectively, were partially ameliorated by Vit C treatment in MPTP-intoxicated animals. In relation to neuroinflammation, results show that Vit C reduced the protein and mRNA expression of inflammatory cytokines such as IL-6, TLR4, TNF-α, iNOS, and CD40, while anti-inflammatory proteins such as IL-10, CD163, TGF-β, and IL-4 increased. Interestingly, we show for the first time that Vit C reduces neuroinflammation by modulating microglial polarization and astrocyte activation. Moreover, Vit C was able to reduce NLRP3 activation, which is linked to the pathogenesis of many inflammatory diseases, including neuroinflammatory disorders. In conclusion, our study provides evidence that Vit C may represent a new promising dietary supplement for the prevention and alleviation of the inflammatory cascade of PD, thus contributing to neuroprotection.

International audience; Several studies have identified a specific metabolic program that is associated with the process of epithelial-mesenchymal transition (EMT). Whereas much is known about the association between glucose metabolism and EMT, the contribution of lipid metabolism is not still completely understood. Here, we studied epithelial and mesenchymal breast cancer cells by proteomic and lipidomic approaches and identified significant differences that characterised these models concerning specific metabolic enzymes and metabolites including fatty acids and phospholipids. Higher levels of monounsaturated fatty acids together with increased expression of enzymes of de novo fatty acid synthesis is the distinct signature of epithelial with respect to mesenchymal cells that, on the contrary, show reduced lipogenesis, higher polyunsaturated fatty acids level and increased expression of genes involved in the triacylglycerol (TAG) synthesis and lipid droplets formation. In the mesenchymal model, the diacylglycerol acyltransferase (DGAT)-1 appears to be the major enzyme involved in TAG synthesis and inhibition of DGAT1, but not DGAT2, drastically reduces the incorporation of labeled palmitate into TAG. Moreover, knockdown of β-catenin demonstrated that this metabolic phenotype in under the control of a network of transcriptional factors and that β-catenin has a specific role in the regulation of lipid metabolism in mesenchymal cells.

In this study, the effects of Radio Electric Asymmetric Conveyer (REAC), a non-invasive physical treatment, on neuroinflammatory responses in a mouse model of parkinsonism induced by intoxication with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), were investigated in vivo. We found that the REAC tissue optimization treatment specific for neuro-regenerative purposes (REAC TO-RGN-N) attenuated the inflammatory picture evoked by MPTP-induced nigro-striatal damage in mice, decreasing the levels of pro-inflammatory molecules and increasing anti-inflammatory mediators. Besides, there was a significant reduction of both astrocyte and microglial activation in MPTP-treated mice exposed to REAC TO-RGN-N. These results indicated that REAC TO-RGN-N treatment modulates the pro-inflammatory responses and reduces neuronal damage in MPTP-induced parkinsonism.

This paper presents a step toward the development of a data-centric approach to prevention of Mild Cognitive Impairment and frailty in the elderly population. The scientific literature provides a large number of “indicators” for assessing the quality of behavior for aged individuals, in order to predict possible decaying. On the opposite side, a large variety of sensors and datasets today allows the effective collection of elementary data about actions performed by individuals. This paper proposes to build a bridge between these two sides. In a bottom-up vision, data from sensors and smart cities' datasets are aggregated and interpreted in a way that leads to reliable assessment of the indicators. In a top-down vision, indicators are translated into data analysis. The work described in this paper is part of City4age, a project partially funded by the EU within the H2020 Programme.

Activated microglia secrete an array of pro-inflammatory factors, such as prostaglandins, whose accumulation contributes to neuronal damages. Prostaglandin endoperoxide synthases or cyclooxygenases (COX-1 and COX-2), which play a critical role in the inflammation, are the pharmacological targets of non-steroidal anti-inflammatory drugs, used to treat pain and inflammation. Since it was reported that COX-1 is the major player in mediating the brain inflammatory response, the aim of this study was to evaluate the effects of highly selective COX-1 inhibitors, such as P6 and mofezolac, in neuroinflammation models. Lipopolysaccharide (LPS)-activated mouse BV-2 microglial cells and LPS intracerebroventricular-injected mice as in vitro and in vivo neuroinflammation models, respectively, were used to probe the antiinflammatory efficacy of P6 and mofezolac. Both P6 and mofezolac reduce COX-1 expression in LPS-activated BV-2 cells. This reduction was accompanied with PGE2 release reduction and NF-kB activation downregulation. Coextensively, in the in vivo model, both glial fibrillary acidic protein and ionized calcium-binding adapter molecule-1 expression, two markers of inflammation, were reduced by mofezolac to a rank depending on the encephalon area analyzed. The increase of COX-1 expression observed in all the brain sections of LPS-treated mice was selectively downregulated by the in vivo treatment with mofezolac as well as PGE2 release and Ikβα phosphorylation amount assayed in the brain areas tested. These results indicate the capability of P6 and mofezolac to modulate the NF-kB signaling pathway, emphasizing the neuroprotective effect and therapeutic potential of COX-1 inhibitors in the control of neuroinflammatory diseases.

The most common methods used by neurologist to evaluate Parkinson’s Disease (PD) patients are rating scales, that are affected by subjective and non-repeatable observations. Since several research studies have revealed that walking is a sensitive indicator

Microglia-mediated neuroinflammation has been described as a common hallmark of Parkinson's disease (PD) and is believed to further exacerbate the progressive degeneration of dopaminergic neurons. Current therapies are unable to prevent the disease progression. A significant association has been demonstrated between PD and low levels of vitamin D in patients serum, and vitamin D supplement appears to have a beneficial clinical effect. Herein, we investigated whether vitamin D administered orally in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced preclinical animal model of PD protects against glia-mediated inflammation and nigrostriatal neurodegeneration. Vitamin D significantly attenuated the MPTP-induced loss of tyrosine hydrlase (TH)-positive neuronal cells, microglial cell activation (Iba1-immunoreactive), inducible nitric oxide synthase (iNOS) and TLR-4 expression, typical hallmarks of the pro-inflammatory (M1) activation of microglia. Additionally, Vitamin D was able to decrease pro-inflammatory cytokines mRNA expression in distinct brain areas of the MPTP mouse. Importantly, we also assessed the anti-inflammatory property of vitamin D in the MPTP mouse, in which it upregulated the anti-inflammatory cytokines (IL-10, IL-4 and TGF-β) mRNA expression as well as increasing the expression of CD163, CD206 and CD204, typical hallmarks of alternative activation of microglia for anti-inflammatory signalling (M2). Collectively, these results demonstrate that vitamin D exhibits substantial neuroprotective effects in this PD animal model, by attenuating pro-inflammatory and up-regulating anti-inflammatory processes.

A fast and simple isocratic high-performance liquid chromatography method for the determination of 3,4-dihydroxyphenylacetic acid (DOPAC), norepinephrine (NE), dopamine (DA), and serotonin (5-HT) in homogenate samples of mouse striatum employing the direct fluorescence of the neurotransmitters is described. The method has been optimized and validated. The analytes were separated in 15min on a reversed-phase column (C18) with acetate buffer (pH 4.0, 12mM)-methanol (86:14, v/v) as mobile phase; the flow rate was 1ml/min. The fluorescence measurements were carried out at 320nm with excitation at 279nm. The calibration curve for DA was linear up to about 2.5μg/ml, with a coefficient of determination (r(2)) of 0.9995 with a lower limit of quantification of 0.031μg/ml. Since the procedure does not involve sample pre-purification or derivatisation, the recovery ranged from 97% to 102% and relative standard deviation (RSD) was better than 2.9%, the use of the internal standard is not mandatory, further simplifying the method. Similar performance was obtained for the other analytes. As a result, thanks to its simplicity, rapidity and adequate working range, the method can be used for the determination of 3,4-dihydroxyphenylacetic acid, dopamine, norepinephrine and serotonin in animal tissues. An experimental 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson-like disease has been used to demonstrate the method is fit-for-purpose.

The uterine myoma pseudocapsule is a neurovascular bundle surrounding fibroid, containing neuropeptides, probably involved in uterine scar healing. We studied neurotensin (NT), neuropeptide tyrosine (NPY), and protein gene product 9.5 (PGP 9.5) nerve fibres in the pseudocapsule neurovascular bundle of intramural uterine fibroids on 67 no pregnant women by intracapsular myomectomy sparing the neurovascular bundle, sampling full thickness specimens of the pseudocapsule of uterine fibroids (PUF) and normal myometrium (NM) obtained from the fundus uteri (FU) and the uterine body (UB). The samples were sent for histological and immunofluorescent analyses and compared by morphometrical quantification. The Conventional Unit (C.U.) difference of NT, NPY, and PGP 9.5 nerve fibres was statistically analyzed. Our results showed that NT, NPY, and PGP 9.5 neurofibers are almost equally present in PUF as in NM of a no pregnant uterus. As all of these neuropeptides are present in the uterine muscle and can affect muscle contractility, uterine peristalsis and muscular healing. A myomectomy respecting the pseudocapsule neurofibers should facilitate smooth muscle scarring and promote restoration of normal uterine peristalsis with a possible positive influence on fertility.

STUDY QUESTION Can uterine scar healing after laparoscopic intracapsular myomectomy (LIM) be adequately monitored by traditional two-dimensional (2D) ultrasound (US) and Doppler velocimetry? SUMMARY ANSWER The myometrial area of the scar after LIM can be followed by 2D US and Doppler velocimetry. WHAT IS KNOWN ALREADY Apart from post-surgical adhesions, the main concern linked to laparoscopic myomectomy is the quality of healing of the myometrial incision: it has been suggested that US could be useful for assessing uterine scars after myomectomy. However, no diagnostic method has yet been widely accepted to assess the healing process. STUDY DESIGN, SIZE, DURATION A cohort prospective study (level of evidence II-2), run in University-affiliated hospitals: 149 women with symptomatic uterine fibroids (UFs) underwent LIM, between January 2007 and October 2011. During follow up 13 patients withdrew from the study. PARTICIPANTS/MATERIALS, SETTING, METHODS After LIM, all patients were followed by traditional 2D US scanning and Doppler velocimetry on Days: 0, 1, 7, 30 and 45. Authors evaluated: number, size and location of UFs, scar diameter and Doppler velocimetry and resistance index (RI) of the uterine arteries, at their ascending branch. MAIN RESULTS AND THE ROLE OF CHANCE The uterine examination showed a significant (P < 0.05) progressive reduction of uterine scar area from 78% of the previous UF location on the first day, to 19% on 30th day, and

AIMS/HYPOTHESIS: Beta cell failure is caused by loss of cell mass, mostly by apoptosis, but also by simple dysfunction (decline of glucose-stimulated insulin secretion, downregulation of specific gene expression). Apoptosis and dysfunction are caused, at least in part, by lipoglucotoxicity. The mechanisms implicated are oxidative stress, increase in the hexosamine biosynthetic pathway (HBP) flux and endoplasmic reticulum (ER) stress. Oxidative stress plays a role in glucotoxicity-induced beta cell dedifferentiation, while glucotoxicity-induced ER stress has been mostly linked to beta cell apoptosis. We sought to clarify whether ER stress caused by increased HBP flux participates in a dedifferentiating response of beta cells, in the absence of relevant apoptosis. METHODS: We used INS-1E cells and murine islets. We analysed the unfolded protein response and the expression profile of beta cells by real-time RT-PCR and western blot. The signal transmission pathway elicited by ER stress was investigated by real-time RT-PCR and immunofluorescence. RESULTS: Glucosamine and high glucose induced ER stress, but did not decrease cell viability in INS-1E cells. ER stress caused dedifferentiation of beta cells, as shown by downregulation of beta cell markers and of the transcription factor, pancreatic and duodenal homeobox 1. Glucose-stimulated insulin secretion was inhibited. These effects were prevented by the chemical chaperone, 4-phenyl butyric acid. The extracellular signal-regulated kinase (ERK) signal transmission pathway was implicated, since its inhibition prevented the effects induced by glucosamine and high glucose. CONCLUSIONS/INTERPRETATION: Glucotoxic ER stress dedifferentiates beta cells, in the absence of apoptosis, through a transcriptional response. These effects are mediated by the activation of ERK1/2.

Obstructive sleep apnea syndrome (OSAS) is an independent and modifiable risk factor for cardiovascular diseases; however, the pathophysiological mechanisms underlying this association are not yet fully understood. Intermittent hypoxemia, one of the physiological markers of OSAS, is characterized by transient periods of oxygen desaturation followed by reoxygenation. The hypoxia-reoxygenation cycles are associated with oxidative stress that, in turn, triggers the activation of pathways that lead to cardiovascular damage. The results of several studies show that OSAS causes oxidative stress and that nasal continuous positive airway pressure therapy normalizes these biological abnormalities. In conclusion, treatment of OSAS with continuous positive airway pressure may lower cardiovascular risk by reducing sympathetic nerve activity, ambulatory blood pressure and arterial stiffness, and by increasing sensitivity of the arterial baroreflex. Newer modalities such as C-Flex and A-Flex also show promise as treatment options in the future. However, the evidence supporting the use of these alternative modalities remains scant, in particular with regards to long-term cardiovascular outcomes.

Systemic infections are often associated with neurodegenerative processes in many diseases of the central nervous system (CNS), including Parkinson’s disease. Toll-like receptor (TLR)4 and CD14 act as receptors for lipopolysaccharide (LPS) released by gram-negative bacteria. In this contest, CD14 functions as the main LPS ligand and TLR4 transmits the LPS signal into the cell. In this paper, we investigated the expression of TLR4 and CD14, in different anatomical areas of the CNS, in an experimental model of Parkinson’s-like disease, represented by MPTP-treated mouse. In particular, we analyzed the gene transcripts and proteins expression of CD14 and TLR4, in the substantia nigra and caudate-putamen nuclei of these animals. Results demonstrated an augmented expression of both CD14 and TLR4 in the substantia nigra of mice treated with MPTP in comparison to untreated animals, suggesting that the endotoxin receptors are over expressed in different manner in specific areas of the CNS during Parkinson’s-like disease.

Several types of K+ channels have been identified in epithelial cells. Among them high conductance Ca2+-activated K+ channels (BK channels) are of relevant importance

Authors investigated 75 patients with uterine myomas, appraising whether fibroid pseudocapsule (FP) thickness varies depending on fibroid location, by a prospective cohort trial (level of evidence II-2) settled in University-affiliated Hospitals. Uteri were scanned via bidimensional and power Doppler ultrasound (US) to map the fibroids and record the FP thickness, prior to hysterectomy for symptomatic uterine fibroids. After hysterectomy, FP specimens were sampled and analyzed by pathologists. Ultrasound and histology data were matched. Pseudocapsule thickness of 108 fibroids was measured: subserosal fibroids (SSFs), intramural fibroids (IMFs), and fibroids near the endometrial cavity (FEC). The FEC's pseudocapsules were considerably thicker than those of IMF and SSF measured by US and histology (P = .001). A clear cutoff existed between FEC pseudocapsule thickness and all other pseudocapsules, with significant differences observed at 2 mm (P = .001). Similarity between histological and US measurements was observed only with IMF pseudocapsules, whereas FEC or SSF showed significant differences. The pseudocapsule of fibroids is considerably thicker near the endometrial cavity when compared to those of both IMFs and SSFs. Since fibroids closest to the endometrial cavity are the most involved in fertility and infertility and FP is considerably thicker near the endometrial cavity, it is possible to hypothesize an involvement of FP of fibroid near the endometrium since FP contains many neuropeptides and neurotransmitters that are physiologically active, even if these data may take on a broader meaning in a study on a larger number of patients.

In the present study we used a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) mouse model to analyze resveratrol neuroprotective effects. The MPTP-induced PD model is characterized by chronic inflammation, oxidative stress and loss of the dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc). We observed that resveratrol treatment significantly reduced glial activation, decreasing the levels of IL-1β, IL-6 and TNF-α, as well as their respective receptors in the SNpc of MPTP-treated mice, as demonstrated by Western blotting, RT-PCR and quantitative PCR analysis. This reduction is related to possible neuroprotection as we also observed that resveratrol administration limited the decline of tyrosine hydroxylase-immunoreactivity induced in the striatum and SNpc by MPTP injection. Consistent with these data, resveratrol treatment up-regulated the expression of the suppressor of cytokine signaling-1 (SOCS-1), supporting the hypothesis that resveratrol protects DA neurons of the SNpc against MPTP-induced cell loss by regulating inflammatory reactions, possibly through SOCS-1 induction.

Newly synthesized thyroglobulin (Tg), the thyroid prohormone, forms detectable high molecular weight mixed disulfide adducts: until now, only Tg "adduct B" was identified as primarily engaging the endoplasmic reticulum oxidoreductases ERp57 and protein disulfide isomerase. Here, we demonstrate that the faster migrating Tg adduct C primarily engages the CaBP1/P5 oxidoreductase, whereas the slower migrating Tg adduct A primarily engages ERp72. Upon siRNA-mediated knockdown of CaBP1/P5 or ERp72, adducts C or A, respectively, are decreased. Within the three Tg adduct bands that do not exhibit a precursor-product relationship, Tg exhibits distinct oxidation patterns. We present evidence suggesting that disulfide maturation occurs within Tg monomers engaged in each of the adduct bands. Moreover, the same Tg substrate molecules can form simultaneous mixed disulfides with both CaBP1/P5 and protein disulfide isomerase, although these are generally viewed as components of distinct oxidoreductase-chaperone protein complexes. Such substrate-oxidoreductase combinations offer Tg the potential for simultaneous oxidative maturation along different parallel tracks leading to the native state.

Eel intestinal epithelium when bathed symmetrically with normal Ringer solution develops a net Cl– current (short circuit current, Isc) giving rise to a negative transepithelial potential (Vt) at the basolateral side of the epithelium, lower in fresh-water (FW)-acclimated animals with respect to sea-water (SW). The aim of the present work was to study the cell response to hypertonic stress of FW eel intestinal epithelium in relation to Cl– absorption. The hypertonicity of the external bathing solutions produced first a transient increase of Vt and Isc, then followed (after10-15min) by a gradual and sustained increase which reached the maximum value after 40-60 min. The morphometric analysis of the intestine revealed the shrinkage of the cells after 5 min hypertonicity exposure, and then a regulatory volume increase (RVI) response, which parallels the gradual and sustained increase in the electrophysiological parameters. This last phase is inhibited by drugs known to block Cl– absorption in eel intestine, such as luminal bumetanide (10 µM), specific inhibitor of Na+-K+-2Cl– cotransport, or basolateral NPPB (0.5 mM), dichloro-DPC (0.5 mM), inhibitors of basolateral Cl– conductance. Serosal dimethyl-amiloride (100 µM), specific inhibitor of the Na+/H+ antiport, was ineffective on the hyperosmotic response. Bicarbonate revealed a crucial role as a modulator of hypertonicity response, since in bicarbonate-free conditions or in the presence of serosal 0.25 mM SITS, blocker of HCO3– transport systems, the Isc response to hypertonicity was lost. In nominally Ca2+-free conditions the Isc response to hypertonicity was abolished. The same results were obtained by bilateral addition of 100 µM verapamil or 50 µM nifedipine or 1 mM lanthanum, known Ca2+ channel blockers, indicating that extracellular Ca2+ plays a key role for the activation of Cl– current in the response to hypertonic stress. The data show that in the eel intestinal epithelium the hypertonicity of the external medium affects cell volume which in turn might represent the signal to increase the rate of Cl– transport. This response is sustained by the activation of the luminal Na+-K+-2Cl– cotransporter and the functionality of basolateral Cl– channels.

Tumour infiltrating lymphocytes (TILs) implicated in immunologic cytotoxicity were evaluated by immunohistochemistry and digitally counted in serial sections from 90 breast cancers in order to assess their number, the relationships between them and to tumour histology. CD3 + , CD4 + , CD8 + , CD20 + , CD25 + and CD56 + lymphocytes were found in 58 (64.4%), 52 (57.7%), 50 (55.5%), 22 (24.4%), 11 (12.2%) and 21 (23.3%) tumours, respectively. There was no difference in the number of TILs between pure infiltrating ductal (NOS) and non-ductal carcinomas, and no relationship between TILs and histological grades was found. CD3 + TILs directly correlated to age, while lymph node negative patients had tumours infiltrated by fewer CD4 + TILs with respect to lymph node positive patients. In 25/90 patients, randomly chosen, the status of peripheral blood lymphocytes was evaluated but no differences with respect to the status found in healthy blood donors was obtained; nonetheless while in some patients CD8 + TILs outnumbered CD4 + TILs in situ, the CD4/CD8 ratio was normal in their peripheral blood. The results show a considerable diversity of TILs among breast tumours, their lack of relationship with the status of the peripheral blood cells, and their potential important relationship with age (CD3 + ) and lymph node status (CD4 + ).

Background: The aim of this study was to investigate relationships between the genotypic frequencies of genes F5-coagulation factor V (proaccelerin, labile factor), F2-coagulation factor II (thrombin), MTHFRmethylenetetrahydrofolate reductase (NADPH), PAI-1 - plasminogen activator inhibitor-1, the frequencies of HLA haplotypes DQ2/DR3, DQ2/DR7, DQ8/DR4, 5, DQ7/DR5 and clinical features and severity of OSAS in comparison to a control population of 163 patients without OSAS. Methods and results: Eighty-two consecutive outpatients with OSAS were enrolled in the respiratory, cardio, and genetic study. By means of cluster analysis it was shown that 40% of patients with PAI-1 5G/5G polymorphism had pulmonary artery hypertension, with a mean value of 46 mm Hg, and also 40% of OSAS patients with pulmonary artery hypertension had PAI-1 5G/5G polymorphism, 50% had 4G/5G and 10% had 4G/4G; as regards OSAS patients without PH, 8.95% only had PAI-1 5G/5G polymorphism, 22.38% had PAI-1 4G/4G and 67.16% had PAI- 1 4G/5G. PAI-1 polimorphisms were detected by RDB techniques. Moreover, these patients with OSAS showed HLA haplotypes polymorphism: DQ7 (40%), DQ2.5 (20%) and DQ2.2 (10%). Nevertheless, OSAS patients without PAH showed HLA polymorphism as follows: DQ7 (47, 76%); DQ2.5 (8.95%); DQ2.2 (16.41%). OSAS patients with pulmonary hypertension were 12.99 % of all OSAS patients. HLA polymorphism was analyzed by PCR amplification and gel electrophoresis. Conclusions: Our results suggested a genotypic heterogeneity among OSAS patients PAI-1 5G/5G polymorphism with PAH possibly in relation with severity of the disease.

Significant heterogeneity of clinical presentation and disease progression exists within chronic obstructive pulmonary disease (COPD). This article discusses and refines the concept of desaturator phenotypes in COPD with pulmonary hypertension (PH) obtained by cluster analysis and presents a pattern of phenotypic markers that could be used as a framework for future diagnosis and research. Nocturnal oxygen desaturation results in sleep disturbances which predispose to nocturnal cardiac dysrhythmias, PH and possibly nocturnal death, particularly during acute exacerbations. We assume that in patients with COPD at least two factors play a role in PH: the severity of pulmonary impairment, and the severity of systemic nocturnal hypoxaemia due to reduced pulmonary functions. Establishing a common language for future research will facilitate our understanding and management of such a disease. This knowledge could lead to different pharmacological treatments and other interventions directed at specific phenotypic groups.

Domenico Maurizio Toraldo1, Francesco De Nuccio2, Annarita Gaballo1, Giuseppe Nicolardi21A Galateo Lung Disease Hospital, Regional Service Puglia, San Cesario di Lecce, 2Laboratory of Human Anatomy, Department of Biological and Environmental Sciences and Technologies, University of Lecce, Lecce, ItalyBackground: Significant heterogeneity of clinical presentation and disease progression exists within chronic obstructive pulmonary disease (COPD). Although forced expiratory volume in 1 second (FEV1) inadequately describes this heterogeneity, a clear alternative has not emerged. This article discusses and refines the concept of phenotyping desaturators in COPD and shows a possible pattern which could be used as a framework for future research.Recent findings: COPD is a complex condition with pulmonary and extrapulmonary manifestations. We suggest that COPD phenotypes should be associated with clinically meaningful outcomes. The innovation of COPD phenotyping is defined as COPD desaturators. Sleep-related hypoxemia and hypercapnia are well recognized in COPD and the development of systemic inflammation during sleep. These sleep-related changes predispose to nocturnal cardiac arrhythmias, pulmonary hypertension, and possibly death, particularly during acute exacerbations.Conclusion: A more focused definition makes possible a classification of patients into two distinct subgroups for both clinical and research purposes. Establishing a common language for future research will facilitate our understanding and management of such diseases. Even if different treatment strategies have different outcomes for these groups, we will have confirmation, or otherwise, of the clinical value of cluster analysis. This knowledge could lead to pharmacological treatment and other interventions directed to specific phenotypic groups.Keywords: phenotypes, chronic obstructive pulmonary disease, desaturator, nocturnal hypoxemia, systemic inflammation, intermittent hypoxia

Objective The fibroid pseudocapsule is a structure which surrounds the uterine fibroid, separates it from the uterine tissue and contains a vascular network rich in neurotransmitters like a neurovascular bundle. The authors examined the composition of the fibroid pseudocapsule using electron microscopy. Study design Twenty non-pregnant patients were submitted to laparoscopic myomectomy by the intracapsular method and samples of the removed pseudocapsules were analyzed using transmission electron microscopy. Results At the ultrastructural level the pseudocapsule cells have the features of smooth muscle cells similar to the myometrium. So, the pseudocapsules are part of the myometrium which compresses the leiomyoma. Conclusion This ultrastructural feature suggests that when removing fibroids their pseudocapsules should be preserved. This study confirms preliminary evidence that pseudocapsules contain neuropeptides together with their related fibers, as a neurovascular bundle. The surgeon's behavior should be directed to carefully control and spare this muscular surrounding tissue during fibroid excision, in order to preserve the myometrium as much as possible.

Parkinson’s disease (PD) is a common neurodegenerative disease characterised by a slow and progressive degeneration of dopaminergic neurons in the substantia nigra (SN). Despite intensive research, the cause of neuronal loss in PD is poorly understood. Inflammatory mechanisms have been implicated in the pathophysiology of PD. In this study, conducted on an experimental 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model, we investigated the expression of interleukin (IL)-1β, tumor necrosis factor (TNF)-α, IL-6 and their receptors (IL-1RI, TNF-αRI, IL-6Rα) at the SN and caudate-putamen (CP) levels. In MPTP-treated animals we observed a significant increase in IL-1β, TNF-α and IL-6 mRNA expression levels both in the SN and CP in comparison with untreated mice. In addition, both mRNA and protein levels of IL-1RI, TNF-αRI and IL-6Rα were significantly enhanced in the SN of MPTP-treated mice in comparison to controls, whereas no significant differences were observed in the CP between treated and untreated mice. Overall, these results indicate a role of both pro-inflammatory cytokines and their receptors in the pathogenesis of PD.

Astrocytes have a key role in the pathogenesis of several diseases, including multiple sclerosis, and are proposed as a possible target for immunotherapy. Our earlier study reported that astrocytes treated with IFN-beta modified their biomechanical properties possibly due to changes in the expression of the proteins involved in cytoskeleton organization and other important physiological processes. To gain insight into the mechanism underlying IFN-beta action during inflammation, we stimulated astrocytes with LPS, a bacterial wall component used as a model for both in vitro and in vivo immunological stimulation of microglia and astrocytes. We showed that IFN-beta reverses the effects of LPS on the proteome of astrocytes. To better examine this result, we performed a proteomic analysis of astrocytes treated with LPS or LPS plus IFN-beta. Treatment with LPS caused increases both in a series of proteins mainly involved in cytoskeletal changes and in protein degradation, as well as protective enzymes like superoxide dismutase. IFN-beta reverses LPS effects on astrocyte proteome, supporting its protective role during inflammatory insults.

The aim of this study was to investigate the time course of body weight, daytime sleepiness, and functional cardiorespiratory parameters in patients with both chronic obstructive pulmonary disease (COPD) and obstructive sleep apnea syndrome (OSA), after institution of domiciliary nasal continuous positive airway pressure (nCPAP).Twelve consecutive obese outpatients (mean age = 61 +/- 11 years; four women) were evaluated before (baseline) and after 3, 12, and 24 months of nocturnal nCPAP (4 h per night).At baseline, all patients were hypercapnic and hypoxemic, suffering from night desaturation (T (90) is the percentage of total recording time (TRT) spent with SaO(2)or= 90% = 38 +/- 2%) and sleepy (Epworth sleepiness scale [ESS] = 16.58 +/- 0.86). Three months after the implementation of nCPAP, daytime PaCO(2) and PaO(2) improved up to 45.1 +/- 0.9 and 69.0 +/- 1 mmHg, respectively; mean pulmonary artery pressure (MPAP) decreased from 24.7 +/- 1.1 to 19.2 +/- 04 mmHg. All other variables showed progressive improvements up to 12 months. At 3 and 12 months, mean body mass index was slightly decreased (to 31.6 +/- 0.2 and 30.7 +/- 0.1 kg/m(2), respectively); daytime sleepiness, nocturnal O(2) desaturation, and maximal inspiratory pressure were also improved and thereafter remained stable.In conclusion, in our patients with both severe OSA and mild-to-moderate COPD, arterial blood gasses and MPAP improved and stabilized after 3 months of nCPAP therapy, with the greatest improvements being in ESS score, T (90), and maximal inspiratory force from 3 up to 12 months; these parameters remained stable over the following 12 months. Finally, our data support early treatment with nCPAP in such patients.

Introduction: Tuberculosis (TB) is a rare curable infective disease, caused mainly by Mycobacterium tuberculosis, which in abdominopelvic (AP) localisation, can mimic a disseminated carcinomatosis. Symptoms of AP-TB are non-specific, so diagnosis is difficult and elusive as the affected patients have normal chest X-ray and elevated levels of CA125. Female ultrasonographic features of AP-TB mimic peritoneal carcinomatosis, and the computed tomography has also been suggested to be helpful, but the final diagnosis was reached by histology and serology. Aim of the study: To propose the validity of the combination of laparoscopy (LPS), histopathology and enzyme-linked immuno-spot (ELIspot) in the diagnosis of AP-TB. Methods: In the last two years, we had six women with suspect of AP-TB, who, after the routine exams, were referred for a diagnostic LPS that revealed turbid-free fluid in pelvis or ascites (collected for serology), multiple peritoneal and/or bowel tubercles (randomly sampled), fibrous bands, adhesions, hyperaemic and oedematous bowel loops. Results: LPS diagnosis was confirmed by intra-LPS biopsy of nodules and histological examination of specimens: epithelioid granulomas with central caseous necrosis in five patients (83.3%) and a non-caseating granulomatous inflammation in the last one. An outer layer of epithelioid histiocytes and Langhans cells was present in all patients. Using the ELIspot technique performed on free fluids, the final diagnosis of TB was made in all patients. Conclusions: Even if gynaecological LPS appearance of the peritoneum can mimic other conditions, ELIspot and histopathological exam can confirm the suspect of AP-TB.

Several types of K(+) channels have been identified in epithelial cells. Among them high conductance Ca(2+)-activated K(+) channels (BK channels) are of relevant importance for their involvement in regulatory volume decrease (RVD) response following hypotonic stress. The aim of the present work was to investigate the functional and molecular expression of BK in the eel intestine, which is a useful experimental model for cell volume regulation research. In the present paper using rat BK channel-specific primer, a RT-PCR signal of 696 pb cDNA was detected in eel intestine, whole nucleotide sequence showed high similarity (83%) to the alpha subunit of BK channel family. BK channel protein expression was verified by immunoblotting and confocal microscopy, while the functional role of BK channels in epithelial ion transport mechanisms and cell volume regulation was examined by electrophysiological and morphometric analysis on the intact tissue. BK(Ca) channels appeared to be localized along all the plasma membrane of the enterocytes; the apical part of the villi showed the most intense immunostaining. These channels were silent in basal condition, but were activated on both membranes (apical and basolateral) by increasing intracellular Ca(2+) concentration with the Ca(2+) ionophore ionomycin (1 microM). BK(Ca) channels were also activated on both membranes by hypotonic swelling of the epithelium and their inhibition by 100 nM iberiotoxin (specific BK(Ca) inhibitor) abolished the Regulatory Volume Decrease (RVD) of the intestinal cells after hypotonic swelling. In conclusion, our results demonstrated the molecular and functional expression of high conductance Ca(2+) -activated K(+) channels in eel intestine; the physiological role of these channels is mainly related to the RVD response of the epithelial cells following hypotonic swelling.

Study objectives: The aims of this study were to define, by cluster analysis, a pattern of clinical variables that differentiate desaturator (D) from nondesaturator (ND) patients affected by COPD, and to identify daytime variables that are predictive of nocturnal desaturation. Patients: Fifty-one random, consecutive COPD outpatients (20 women; mean [± SD] age, 69.6 ± 4.0 years) with mild daytime hypoxemia (Pa o 2 , 60 to 70 mm Hg) were enrolled into the study. Obstructive sleep apnea syndrome patients were excluded. Measurements and results: Lung volumes, arterial blood gas levels, and mean pulmonary artery pressure (MPAP) were measured, and nocturnal desaturation was evaluated with nighttime polygraphy. With least squares simple linear regression, the percentage of total recording time was highly correlated with a total nocturnal recording time of arterial oxygen saturation of 90 ) and MPAP ( R = 0.84; R 2 = 71.20%); T 90 was also highly correlated with daytime Pa co 2 ( R = 0.70; R 2 = 48.96%). Multiple regression showed that T 90 was highly correlated with both MPAP and Pa co 2 ( R 2 = 97.75%). Hierarchical cluster analysis conducted with these three variables showed that D and ND patients differed in both nocturnal and daytime variables. The mean T 90 was 30 ± 3.5% in 19.2% and 8%, respectively, of the D and ND groups. Moreover, two D subgroups differing in MPAP and two ND subgroups differing in Pa co 2 were identified. Conclusions: D patients may be identified by a pattern of T 90 , MPAP, and Pa co 2 values, rather than by T 90 alone, with the latter two variables being predictors of nocturnal desaturation severity.

Astrocytes have a key role in the pathogenesis of several diseases including multiple sclerosis and were proposed as the designed target for immunotherapy. In this study we used atomic force microscopy (AFM) and proteomics methods to analyse and correlate the modifications induced in the viscoleastic properties of astrocytes to the changes induced in protein expression after interferon- beta (IFN-beta) treatment. Our results indicated that IFN-beta treatment resulted in a significant decrease in the Young's modulus, a measure of cell elasticity, in comparison with control cells. The molecular mechanisms that trigger these changes were investigated by 2DE (two-dimensional electrophoresis) and confocal analyses and confirmed by western blotting. Altered proteins were found to be involved in cytoskeleton organization and other important physiological processes.

With the aim of investigating some factors and mechanisms of the chicken brain development, the same thick sections of brain stems from twelve E13-to-E21-aged chick embryos were sequentially tested with a rabbit anti-Somatostatin antiserum, using a PAP-DAB technique, and with anti-tyrosine hydroxylase (-TH) monoclonal antibodies, using an indirect immuno-fluorescence technique. As regards the pons and mesencephalon, the following main results were obtained. At E21 almost the same distribution of the TH-like immunoreactivity (ir) as at E13 was observed. Neuroblasts in a central, relatively wide region of mesencephalic tegmentum and in the central portion of the pons showed TH-like ir. A co-localization of the 2 immunoreactivities was detected only at E18, within some neuroblasts of the mesencephalic and pontine regions with TH-like ir. It is possible that this transitory co-localization plays a role in the development of the pons and mesencephalon of this species.

The aim of the present research is to study the development of spinal ganglia implanted onto chorio-allantoic membrane (CAM), where they are supplied by 'foreign', extraembryonic vessels. Small blocks including a segment of the neural tube and 4 pairs of somites were isolated from the brachial or thoracic or lumbar region (neuromeres C13 - T1 or T3 - T6 or L1 - L4) of thirty 3-day donor chick embryos and grafted on the CAM of as many 10-day host chick embryos. At regular time intervals after grafting, the implants were fixed, embedded in paraffin, serially cross-sectioned and stained. The preliminary results are similar for the ganglia from the 3 regions: they seem to indicate that in the spinal ganglia grafted on the CAM the developing blood vessels do not exert any morphogenetic influence on the substratum; this corroborates results from previous researches (2) on the normally 'in situ' developing ganglia. It is now the author purpose to submit to a careful analysis a datum from the present research: namely, the earlier appearance of the endoganglionic vessels in the brachial and lumbar grafted ganglia than in the thoracic ones.

An immunocytochemical analysis according to the PAP-DAB procedure was performed on the chicken pontine and mesencephalic tegmentum, to study the distribution here of the neurons reacting to anti-cyclic-somatostatin-14 polyclonal rabbit antibodies. Immunoreactivity was not increased by any pretreatment; positive and negative controls of the immunoreaction were carried out. A positivity was chiefly detected: 1. in perikarya of the inferior half of the mesencephalic tegmentum, belonging probably to the n. mesencephalicus profundus cranially, and to the n. loci coerulei caudally; 2. in the perikarya of the cranial half of the mesencephalic tegmentum, belonging to the n. tegmentalis latero-dorsalis; 3. in perikarya of the cranial part of the n. interpeduncularis; 4. in long neuronal processes (a) of the pontine tegmentum – situated mainly in the dorsal half and running longitudinally -, (b) at the ponto-mesencephalic border – oriented almost transversally -, (c) of the mesencephalic tegmentum – placed mainly in its ventral half and running longitudinally; 5. in short neuronal processes within the periaqueductal grey.

Clinical, neuropathological, and immunohistochemical (PAP) findings are reported in a 7 month-old girl with Tuberous sclerosis (TS). Polycystic kidneys and massive cardiomegaly constituted the prominent pathological features. The neuropathology of the syndrome was characterized by Megalencephaly, Subependymal giant cell astrocytoma, Very large protoplasmic astrocytes in Golgi-rapid method preparations of cortical tubers and Glial fibrillary acidic protein (GFAP)-negative giant cells in the subependymal nodules as well as in the cortical tubers. The identity of these giant cells is discussed.

An immunocytochemical analysis was performed on the chicken pons and mesencephalon (except the optic tectum) according to the PAP-DAB procedure, to study the distribution here of the neurons reacting to anti-VIP antibodies and the gliocytes reacting to anti-GFAP antibodies. Positive and negative controls were carried out in both the immunoreactions. The VIP-immunoreactive neurons showed a distribution essentially corresponding to that observed in other species by various Authors. They appeared scattered mainly in 3 sites: (a) the subventricular grey between pons and mesencephalon; (b) the periaqueductal grey, up to the diencephalon; (c) the rostro-ventral portion of the mesencephalic tegmentum, up to the diencephalon. Furthermore, some perivascular VIP-immunoreactive neuronal processes were seen. No differences have so far been detected as regards the GFAP-like immunoreactivity distribution, in comparison with the data reported by the authors in the chicken medulla and by others in in the brain stem of some other species.

An immunocytochemical analysis following the PAP-DAB procedure was performed on the chick embryo midbrain tegmentum, to study through which developmental steps the nerve cell groups, showing positivity to anti-Somatostatin (SRIF) antibodies in adult animals, form. The SRIF-like immunoreactivity appears within neuroblasts at the 12th incubation day. From the 12th to the 18th incubation day the distribution of the SRIF-like immunoreactivity gradually changes. At the 18th incubation day the positivity detented within neuroblasts perikarya corresponds to that described in adult animals.