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The lung tissue is one of the main targets of oxidative stress due to external sources and respiratory activity. In our previous work, we have demonstrated in that O 3 exposure alters the Cl - current-voltage relationship, with the appearance of a large outward rectifier component mainly sustained by outward rectifier chloride channels (ORCCs) in human lung epithelial cells (A549 line). In the present study, we have performed patch clamp experiments, in order to identify which one of the O 3 byproducts (4hydroxynonenal (HNE) and/or H 2 O 2 ) was responsible for chloride current change. While 4HNE exposition (up to 25 μM for 30' before electrophysiological analysis) did not reproduce O 3 effect, H 2 O 2 produced by glucose oxidase 10 mU for 24 hr before electrophysiological analysis mimicked O 3 response. This result was confirmed treating the cell with catalase (CAT) before O 3 exposure (1,000 U/ml for 2 hr): CAT was able to rescue Cl - current alteration. Since CAT is regulated by Nrf2 transcription factor, we pre-treated the cells with the Nrf2 activators, resveratrol and tBHQ. Immunochemical and immunocytochemical results showed Nrf2 activation with both substances that lead to prevent OS effect on Cl - current. These data bring new insights into the mechanisms involved in OS-induced lung tissue damage, pointing out the role of H 2 O 2 in chloride current alteration and the ability of Nfr2 activation in preventing this effect., (© 2017 Wiley Periodicals, Inc.)

Epidemiological evidences have correlated airbone particulate matter (PM) to adverse health effects, mainly linking to pulmonary and cardiovascular disease. Nevertheless, only recently, some studies reported detrimental effects of PM on other organs such as skin. In a recent work, we have reported increased oxidative and inflammatory responses in Reconstituted Human Epidermis (RHE) exposed to ambient particles (CAPs) and we also demonstrated the ability of CAPs to penetrate the skin tissue. The present study was aimed to better understand the cellular mechanisms beyond the oxidative changes induced by CAPs (5-10-25μg/mL) in human immortalized keratinocytes (HaCaT). After 24h of treatment, CAPs were able to enter the cells leading to a decrease in viability, increased levels of 4-hydroxinonenal products (4-HNE) and IL-1α release. Overall these data, suggest lipid and protein oxidative damage, as well as an increase of inflammatory response after being challenged with CAPs. In addition, 3h after CAPs exposure we found a significant increase in NF-kB and Nrf2 translocation into the nucleus. In contrast, no differences in gene expression and enzymatic activity of Nrf2 target genes were detected. This last finding could be explained by the ability of CAPs to possibly alter the binding of Nrf2 to the ARE DNA sequence., (Published by Elsevier B.V.)

Background: Wound healing is a biological process that can get in a state of pathologic inflammation, requiring the use of specific medications able to promote proper tissue repair., Objective: The study describes the production and characterization of nanoparticle based gel for wound healing treatment designed to deliver hyaluronic acid and retinyl palmitate onto the skin., Methods: Tristearin solid lipid nanoparticles and nanostructured lipid carriers based on a tristearin and caprylic/capric triglyceride mixture were produced and characterized. Gel spreadability and viscosity were investigated. Drug diffusion and "in vitro" wound healing were assessed by Franz cell and scratch wound assay in keratinocytes., Results: Cryogenic transmission electron microscopy evidenced flat discoid nanoparticles. Photon correlation spectroscopy analysis indicated homogeneous dimensional distribution and mean diameter 132±46 nm. X-ray evidenced a lamellar inner structure of lipid nanoparticles. Nanostructured lipid carriers, being based on a heterogeneous solid/ liquid lipid mixture, could better solubilize retinyl palmitate and control its stability. The hyaluronic acid directly added into nanoparticles' dispersion enabled to obtain a shear-thinning gel suitable for cutaneous administration. Retynil palmitate diffusion was slower from the nanoparticulate gel with respect to the plain nanoparticle dispersion. The "wound healing" effect of nanoparticulate gel containing retinyl palmitate and hyaluronic acid, analyzed in HaCaT cells, showed significant differences in wounded areas between treated and control cells during the first 24 h postwounding suggesting a synergic effect of retinyl palmitate and hyaluronic acid in "in vitro" wound healing., Conclusions: This study suggests that a nanoparticle based hyaluronate gel containing retinyl palmitate can be efficiently used for wound healing., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Air pollution continues to be a major public health concern affecting 9 out of 10 individuals living in urban areas worldwide. Respiratory tract is the organ most exposed to gas pollution, and ozone has been shown to be one of the most noxious pollutants to which living organisms are exposed. In the present work, we have investigated the effects of 0.1 ppm of ozone on chloride currents in human lung epithelial cells (A549 line) and whether this effect could be modulated by vitamin E pre-treatment. Whole-cell patch clamp technique was applied to not excitable cells in order to obtain information about chloride currents behavior, important for epithelial lung cells homeostasis. Significant alteration of the I-V curve after ozone treatment was observed, with the appearance of a large outward rectifier component decreasing over time and returning to the basal state levels after 24 h. Statistical analysis indicated a modification of the amount of ions passing the membrane in the unit of time as a possible cause of this difference. RT-qPCR analysis showed an increase in ClC-2 and ORCC mRNA after ozone exposure. In addition, pre-treatment with vitamin E was able to suppress the outward rectifier component induced by ozone, bringing back the current values to the control level and preventing ozone induced chloride channels up regulation. Our data suggest that ozone exposure is able to modify chloride current density and the use of vitamin E can prevent the above-mentioned damage. J. Cell. Physiol. 232: 1817-1825, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Objective: The purpose of this study was to assess the prevalence, distribution and intensity of tooth wear in a sample of an ancient Italian population in order to explain the pattern in terms of dietary habits and/or non-dietary tooth-use behaviors during the Early Bronze Age, with a focus on possible age-group and sex differences., Design: Well-preserved permanent teeth of individuals from the Bronze Age site of Ballabio (Lecco) in northern Italy were examined for tooth wear by different methods. Eight 3D models of teeth at increasing severity of wear were created., Results: In total, 357 permanent teeth belonging to male and female individuals were included in the study. Dental wear was present in 96.6% of the total sample. Males showed significantly greater levels of wear than females in the mandibular teeth. Both sexes exhibited a significantly different wear direction between the anterior (oblique and flat) and posterior (oblique and concave) teeth. Significant age differences were observed in the direction and level of wear in the incisors, canines and premolars, with higher wear in the older group. Complete and rotatable virtual 3D images of different wear patterns are proposed., Conclusions: The findings of the present study confirm the data from archaeological studies on this site and on northern Italian habits during the Early Bronze Age suggesting a diet rich in vegetables. The observed wear patterns can be related both to the diet of this Bronze age population, based on hard and abrasive food requiring vigorous mastication, and to sex differences in cultural practices., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

For its critical location, the skin represents the major interface between the body and the environment, therefore is one of the major biological barriers against the outdoor environmental stressors. Among the several oxidative environmental stressors, cigarette smoke (CS) has been associated with the development and worsening of many skin pathologies such as acne, dermatitis, delayed wound healing, aging and skin cancer. In our previous work we have demonstrated that CS is able to affect genes involved in skin cholesterol trafficking, among which SRB1, a receptor involved in the uptake of cholesterol from HDL, seems to be very susceptible to the oxidative stress induced by CS. In the present work we wanted to investigate the presence of SRB1 in human sebocytes and whether CS can affect cholesterol cellular uptake via the redox modulation of SRB1. By using a co-culture system of keratinocytes/sebocytes, we found that CS exposure induced a SRB1 protein loss without affecting sebocytes viability. The decrease of SRB1 levels was a consequence of SRB1/HNE adducts formation that leads to SRB1 ubiquitination and degradation. Moreover, the CS-induced loss of SRB1 induced an alteration of sebocytes lipid content, also demonstrated by cholesterol quantification in SRB1 siRNA experiments. In conclusion, exposure to CS, induced SRB1 post-translational modifications in sebocytes and this might affect sebocytes/skin functionality., (Copyright © 2016 Elsevier Inc. All rights reserved.)

While surgery is the definitive treatment for early-stage melanoma, the current therapies against advanced melanoma do not yet provide an effective, long-lasting control of the lesions and a satisfactory impact on patient survival. Thus, research is also focused on novel treatments that could potentiate the current therapies. In the present study, we evaluated the effect of potassium ascorbate with ribose (PAR) treatment on the human melanoma cell line, A375, in 2D and 3D models. In the 2D model, in line with the current literature, the pharmacological treatment with PAR decreased cell proliferation and viability. In addition, an increase in Connexin 43 mRNA and protein was observed. This novel finding was confirmed in PAR-treated melanoma cells cultured in 3D, where an increase in functional gap junctions and a higher spheroid compactness were observed. Moreover, in the 3D model, a remarkable decrease in the size and volume of spheroids was observed, further supporting the treatment efficacy observed in the 2D model. In conclusion, our results suggest that PAR could be used as a safe adjuvant approach in support to conventional therapies for the treatment of melanoma.

Rett syndrome (RTT) is a pervasive developmental disorder, primarily affecting girls with a prevalence of 1 in every 10,000 births. A clear etiological factor present in more than 90% of classical RTT cases is the mutation of the gene encoding methyl-CpG-binding protein 2 (MECP2). Recent work from our group was able to shown a systemic oxidative stress (OxS) in these patients that correlates with the gravity of the clinical features. Using freshly isolated skin fibroblasts from RTT patients and healthy subjects, we have performed a two-dimensional gel electrophoresis in order to evidence the oxidative modifications of proteins with special focus on the formation of protein adducts with 4-hydroxynonenal (4-HNE PAs)-a major secondary product of lipid peroxidation- and Nitrotyrosine, a marker derived from the biochemical interaction of nitric oxide (NO) or nitric oxide-derived secondary products with reactive oxygen species (ROS). Then, oxidatively modified spots were identified by mass spectrometry, LC-ESI-CID-MS/MS. Our results showed that 15 protein spots presented 4-HNE PAs and/or nitrotyrosine adducts in fibroblasts proteome from RTT patients compared to healthy control cells. Post-translationally modified proteins were related to several functional categories, in particular to cytoskeleton structure and protein folding. In addition, clear upregulated expression of the inducible NO synthase (iNOS) with high nitrite levels were observed in RTT fibroblasts, justifying the increased nitrotyrosine protein modifications. The present work describes not only the proteomic profile in RTT fibroblasts, but also identifies the modified proteins by 4-HNE and nitrotyrosine. Of note, for the first time, it appears that a dysregulation of NO pathway can be associated to RTT pathophysiology. In conclusion, the evidence of a wide range of proteins able to forms adducts with 4-HNE, Nitrotyrosine or with both confirms the possible alteration of several aspects of cellular functions that well correlates to the complex clinical features of RTT patients., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The beneficial effect of low doses of ozone on wound healing has been well documented and attributed mainly to its bactericidal and pro-oxidant properties. Because ozone itself does not penetrate the cells but immediately reacts with polyunsaturated fatty acids, its effects are the results of oxidative mediators. Among the molecule produces by the interaction of ozone with biological systems, there are HNE and H2O2. At today, the cellular mechanisms accounting for the positive effects of mild ozonization on wound closure are still largely unexplored. The aim of the present study was to evaluate the effect of different non-toxic doses of ozonated saline ranging from 2 to 300 μM, in an in vitro wound scratch model by the use of human keratinocytes. The results showed that ozonated saline is able to improve in vitro wound healing by stimulating cell proliferation as measured by BrdU assay and PCNA protein levels. In order to better elucidate the molecules that play the main role in the beneficial effect of ozonated saline in wound healing, HNE and H2O2 were used alone or in combination to mimic ozonated saline effect. Surprisingly, keratinocytes treated with different doses of HNE and H2O2 did not significantly improve the wound closure, while the combination of the two compounds was able to improve wound closure. In addition, Nrf2 pathways were also activated as determined by its translocation to the nucleus and the increased HO1 gene expression. The present work suggests that ozonated saline effect on wound closure is the results of the combination of more molecules among which HNE and H2O2 play a key role.

Microtubules are composed by α- and β-tubulin polypeptides. α-tubulin undergoes a reversible posttranslational modification whereby the C-terminal tyrosine residue is removed (Glu-tubulin) and re-added (Tyrtubulin). Recent studies have shown that α-tubulin tyrosine residues can be nitrated and the incorporation of NO2Tyr into the C-terminus of Glu-tubulin forms a complex that blocks the tyrosination/detyrosination cycle, an event that can compromise protein/enzyme functions, such as cell division. Since many studies demonstrated that Glu-tubulin levels increase in cancer, the aim of the present study was to investigate the effect of new drugs, fluorazone derivatives (K1-K2-K9-K10-K11), on the proliferation of melanoma cells. Our results demonstrated that these drugs, except for K2, were able to inhibit cellular proliferation without exhibiting cytotoxicity. The anti-proliferative effect was accompanied by the decrease of Glu-tubulin levels and the increase of its nitration. This effect seems to be a consequence of NO2 induction and NO2Tyr ligation to Glu-tubulin. Collectively, these results, showing that the fluorazone derivatives, by promoting NO2Tyr incorporation into α-tubulin, are able to arrest the cycle of detyrosination/tyrosination and to inhibit cell proliferation, offer new perspectives for the possible usage of these drugs, alone or in combination, as non-toxic, anti-proliferative agents in melanoma.

A potential role for immune dysfunction in autism spectrum disorders (ASD) has been well established. However, immunological features of Rett syndrome (RTT), a genetic neurodevelopmental disorder closely related to autism, have not been well addressed yet. By using multiplex Luminex technology, a panel of 27 cytokines and chemokines was evaluated in serum from 10 RTT patients with confirmed diagnosis of MECP2 mutation (typical RTT), 12 children affected by classic autistic disorder and 8 control subjects. The cytokine/chemokine gene expression was assessed by real time PCR on mRNA of isolated peripheral blood mononuclear cells (PBMCs). Moreover, ultrastructural analysis of PBMCs was performed using transmission electron microscopy (TEM). Significantly higher serum levels of interleukin-8 (IL-8), IL-9, IL-13 were detected in RTT compared to control subjects, and IL-15 shows a trend toward the upregulation in RTT. In addition, IL-1β and VEGF were the only down-regulated cytokines in autistic patients with respect to RTT. No difference in cytokine/chemokine profile between autistic and control groups was detected. These data were also confirmed by ELISA real time PCR. At the ultrastructural level, the most severe morphological abnormalities were observed in mitochondria of both RTT and autistic PBMCs. In conclusion, our study shows a deregulated cytokine/chemokine profile together with morphologically altered immune cells in RTT. Such abnormalities were not quite as evident in autistic subjects. These findings indicate a possible role of immune dysfunction in RTT making the clinical features of this pathology related also to the immunology aspects, suggesting, therefore, novel possible therapeutic interventions for this disorder., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Epidemiological studies suggest a correlation between increased airborne particulate matter (PM) and adverse health effects. The mechanisms of PM-health effects are believed to involve oxidative stress and inflammation. To evaluate the ability of PM promoting skin tissue damage, one of the main organs exposed to outdoor pollutants, we analyzed the effect of concentrated ambient particles (CAPs) in a reconstructed human epidermis (RHE) model. RHE tissues were exposed to 25 or 100 µg/ml CAPs for 24 or 48 h. Data showed that RHE seems to be more susceptible to CAPs-induced toxicity after 48 h exposure than after 24 h. We found a local reactive O(2) species (ROS) production increase generated from metals present on the particle, which contributes to lipids oxidation. Furthermore, as a consequence of altered redox status, NFkB nucleus translocation was increase upon CAPs exposure, as well as cyclooxygenase 2 and cytochrome P450 levels, which may be involved in the inflammatory response initiated by PM. CAPs also triggered an apoptotic process in skin. Surprisingly, by transition electron microscopy analysis we showed that CAPs were able to penetrate skin tissues. These findings contribute to the understanding of the cutaneous pathophysiological mechanisms initiated by CAPs exposure, where oxidative stress and inflammation may play predominant roles., (© The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background: A recent study conducted in humans demonstrated for the first time that bone loss in the immediate period after implant insertion before loading did not significantly differ in organ transplant recipients with respect to normal subjects., Purpose: The purpose of this study is to evaluate bone and periodontal response and peri-implant microflora in a group of organ-transplanted patients 1 year after prosthetic loading., Materials and Methods: The study population included 13 consecutive organ-transplanted (11 hearts, two livers) patients and 13 normal (healthy) control subjects who received 29 and 28 submerged dental implants, respectively. Crestal bone level, peri-implant probing depth (PIPD), and bleeding on probing were evaluated at prosthetic loading and after 1 year. Samples for microbiological testing were taken from the subgingival microbiota of each implant 1 year after loading and analyzed with polymerase chain reaction. All samples were examined for the presence of five microorganisms: Treponema denticola, Porphyromonas gingivalis, Prevotella intermedia, Tannarella forsythensis, and Actinobacillus actinomycetemcomitans., Results: A mean bone loss of 0.17 ± 0.10 and 0.20 ± 0.10 mm at 1 year was observed in the group of transplant recipients and in controls, respectively (N.S.). Similar results were obtained considering PIPD changes at 1 year (respectively 0.06 ± 0.71 mm in transplants vs 0.11 ± 0.74 mm in controls). Detection frequencies of pathogens were not statistically different between normal and transplanted patients., Conclusions: The present pilot study seems to indicate that bone and periodontal response and microbiological status around submerged dental implants in immunocompromised organ-transplanted patients do not differ 1 year after loading from those observed in control patients and that this particular population of patients may be successfully rehabilitated with dental implants., (© 2014 Wiley Periodicals, Inc.)

A strong correlation between oxidative stress (OS) and Rett syndrome (RTT), a rare neurodevelopmental disorder affecting females in the 95% of the cases, has been well documented although the source of OS and the effect of a redox imbalance in this pathology has not been yet investigated. Using freshly isolated skin fibroblasts from RTT patients and healthy subjects, we have demonstrated in RTT cells high levels of H2O2 and HNE protein adducts. These findings correlated with the constitutive activation of NADPH-oxidase (NOX) and that was prevented by a NOX inhibitor and iron chelator pre-treatment, showing its direct involvement. In parallel, we demonstrated an increase in mitochondrial oxidant production, altered mitochondrial biogenesis and impaired proteasome activity in RTT samples. Further, we found that the key cellular defensive enzymes: glutathione peroxidase, superoxide dismutase and thioredoxin reductases activities were also significantly lower in RTT. Taken all together, our findings suggest that the systemic OS levels in RTT can be a consequence of both: increased endogenous oxidants as well as altered mitochondrial biogenesis with a decreased activity of defensive enzymes that leads to posttranslational oxidant protein modification and a proteasome activity impairment., (Copyright © 2015 Elsevier B.V. All rights reserved.)

CDKL5 mutation is associated with an atypical Rett syndrome (RTT) variant. Recently, cholesterol homeostasis perturbation and oxidative-mediated loss of the high-density lipoprotein receptor SRB1 in typical RTT have been suggested. Here, we demonstrate an altered lipid serum profile also in CDKL5 patients with decreased levels of SRB1 and impaired activation of the defensive system Nrf2. In addition, CDKL5 fibroblasts showed an increase in 4-hydroxy-2-nonenal- and nitrotyrosine-SRB1 adducts that lead to its ubiquitination and probable degradation. This study highlights a possible common denominator between two different RTT variants (MECP2 and CDKL5) and a possible common future therapeutic target., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Introduction: One of the main functions of cutaneous tissues is to protect our body from the outdoor insults. Ozone (O3) is among the most toxic stressors to which we are continuously exposed and because of its critical location, the skin is one of the most susceptible tissues to the oxidative damaging effect of O3. O3 is not able to penetrate the skin, and although it is not a radical per se, the damage is mainly a consequence of its ability to induce oxidative stress via the formation of lipid peroxidation products., Aim of Study: In this study we investigated the protective effect of defined "antioxidant" mixtures against O3 induced oxidative stress damage in human keratinocytes and understand their underlying mechanism of action., Results: Results showed that the mixtures tested were able to protect human keratinocytes from O3-induced cytotoxicity, inhibition of cellular proliferation, decrease the formation of HNE protein adducts, ROS, and carbonyls levels. Furthermore, we have observed the decreased activation of the redox sensitive transcription factor NF-kB, which is involved in transcribing pro-inflammatory cytokines and therefore constitutes one of the main players associated with O3 induced skin inflammation. Cells exposed to O3 demonstrated a dose dependent increase in p65 subunit nuclear expression as a marker of NF-kB activation, while pre-treatment with the mixtures abolished NF-kB nuclear translocation. In addition, a significant activation of Nrf2 in keratinocytes treated with the mixtures was also observed., Conclusion: Overall this study was able to demonstrate a protective effect of the tested compounds versus O3-induced cell damage in human keratinocytes. Pre-treatment with the tested compounds significantly reduced the oxidative damage induced by O3 exposure and this protective effect was correlated to the abolishment of NF-kB nuclear translocation, as well as activation of Nrf2 nuclear translocation activating the downstream defence enzymes involved in cellular detoxification process.

This study describes the production and characterization of nanostructured lipid dispersions (NLDs) containing curcumin (CUR) as new tools for curcumin topical delivery. Four types of NLDs based on monoolein in association with different emulsifiers were produced: Na cholate and poloxamer 407 (NLD1), poloxamer alone (NLD2), the mixture of Na cholate and Na caseinate (NLD3) and Na cholate alone (NLD4). Morphology and dimensional distribution of lipid dispersions were investigated by cryo-TEM and photon correlation spectroscopy (PCS). In vitro studies based on Franz cell, membrane nylon and stratum corneum-epidermis (SCE) were carried out to compare the four NLDs in terms of cytotoxicity in human keratinocytes and CUR diffusion. Our PCS studies showed differences in particles diameter among the different NLDs. In addition, cytotoxicity results in HaCaT cells evidenced that NLD1 and NLD2 were toxic at doses over 1 μm. Therefore, cryo-TEM was determined only for NLD3 and NLD4 showing that CUR did not affect their structure. Diffusion measurement in SCE and nylon membrane evidenced that CUR had a time-delayed release for NLD4. The 'wound healing' effect of NLD3 and NLD4 with and without CUR analysed keratinocytes in vitro, and a clear inhibition of cell proliferation/migration by CUR was observed. This effect was mediated by the inhibition of cyclin D1 expression as a consequence of the impaired NFkB activation. This study confirms the antiproliferative properties of CUR and evidenced a new possible model of CUR topical delivery for hyperproliferative cutaneous diseases such as psoriasis., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Chronic obstructive pulmonary disease (COPD) has been recognized as one of the major causes of morbidity and mortality in the United States; it is the third leading cause of deaths in the United States, with approximately 15 million Americans affected with COPD. Although exposure to cigarette smoke has been shown to be the main, if not the only, risk factor for COPD, the mechanisms underlying this association remain unclear. Most smokers do not develop COPD, suggesting that a combination of exposure and susceptibility (genetic background) is required. Several mechanisms contribute to the pathogenesis of COPD, such as influx of inflammatory cells into the lung, imbalance between proteolytic and antiproteolytic molecules, disruption of the balance between apoptosis and replenishment of structural cells in the lung, and disruption of oxidant/antioxidant balance. The scavenger receptor BI (SRB1) plays an important role in mediating the uptake of high-density lipoprotein (HDL)-derived cholesterol and cholesteryl ester in tissues. In addition to its role as the HDL receptor, SRB1 is also involved in pathogen recognition, identification of apoptotic cells, tissue antioxidant uptake (tocopherol and carotenoids), and lung surfactant composition, all factors involved in COPD pathogenesis. Therefore, it is possible that lung SRB1 levels are involved in the development of COPD., (© 2015 New York Academy of Sciences.)

Oral leukoplakia (OL) is the most common potentially malignant lesion of the oral cavity. Immunohistochemical analysis of p53 and Ki67 proteins is a simple and inexpensive method widely used in non-dysplastic OLs to reveal lesions predicted to develop oral cancer. The present longitudinal study evaluated the predictive role of p53 and Ki67 proteins alone or in combination in a group of OLs without dysplasia followed for many years. Seventy-seven OL patients referred to our Department between January 2006 and October 2013 underwent histochemical analysis of p53 and Ki67 expression. OLs were considered at high risk in the presence of either high p53 expression (>20%), or low/normal p53 expression associated with high Ki67 expression (Ki67/p53 ratio >3). Seven OLs evolved to OSCC during the follow-up period. Three cases had p53 overexpression, while four had a high Ki67/p53 ratio. Statistical significance was reached when samples with p53 overexpression were combined with samples with high Ki67/p53 ratio (Chi square 5.3; p<0.02). The combined immunohistochemical expression of p53 and Ki67 proteins could be a useful and simple molecular marker for early detection of non-dysplastic OLs at risk of developing oral cancer.

Cigarette smoke (CS) contains over 4700 compounds, many of which can affect cellular redox balance through free radicals production or through the modulation of antioxidant enzymes. The respiratory tract is one of the organs directly exposed to CS and it is known that CS can damage the integrity of lung epithelium by affecting cell junctions and increasing epithelium permeability. In this study, we have used a human lung epithelial cell line, Calu-3, to evaluate the effect of CS on lung epithelial cell junctions levels, with special focus on the expression of two proteins involved in intercellular communication: connexins (Cx) 40 and 43. CS exposure increased Cx40 gene expression but not of Cx43. CS also induced NFκB activation and the formation of 4HNE-Cxs adducts. Since corilagin, a natural polyphenol, is able to inhibit NFκB activation, we have determined whether corilagin could counteract the effect of CS on Cxs expression. Corilagin was able to diminish CS induced Cx40 gene expression, 4HNE-Cx40 adducts formation, and NFκB activation. The results of this study demonstrated that CS induced the loss of cellular junctions in lung epithelium, possibly as a consequence of Cx-4HNE adducts formation, and corilagin seems to be able to abolish these CS induced alterations.

The modulation of the HDL receptor scavenger receptor B1 (SRB1) was evaluated in skin fibroblasts isolated from Rett syndrome (RTT) patients, a rare neurodevelopmental disorder affecting almost exclusively females associated in up to 95% of cases to de novo loss-of-function mutations in the X-chromosome-linked gene encoding the methyl-CpG-binding protein 2 (MeCP2). Patients showed an altered plasma lipid profile, while their skin fibroblasts showed a dramatic reduction in SRB1 (immunogold, Western blot and immunohistochemistry). The decreased SRB1 levels were demonstrated to be the consequence of its binding with 4-hydroxy-2-nonenal (4HNE), a product of lipid peroxidation, and its increased ubiquitination. Therefore the loss of SRB1 in RTT cells is a consequence of the chronic oxidative stress status present in RTT. In addition RTT fibroblast presented high intracellular levels of H2O2 and 4HNE protein adducts. This finding was correlated with the constitutive activation of NADPH oxidase (NOX) and was reverted by DPI (NOX inhibitor) or Desferal (Iron chelator) pre-treatment. To confirm the alteration of status redox in RTT cells, the activity of several enzymes involved in protecting the cell from OS was also evaluated. Glutathione peroxidase (GPx), Supeoxide dismutase and Glucose-6-phosphate dehydrogenase (G6PDH) activity were decreased respect to control. These data paralleled with a constitutive activation of NRF2 and elevated gene expression of Heme oxigenase-1 (HO-1) and NAD(P)H dehydrogenase quinone 1 (NQO-1). Of note, when NRF2 pathway was stimulated via exogenous oxidants, RTT fibroblast did not respond as the control cells., (Copyright © 2014. Published by Elsevier Inc.)

The plant polyphenol, resveratrol (Resv, 3,4,5-trihydroxystilbene), naturally occurring in a number of fruits and other food products, has been extensively studied over the last two decades for its beneficial properties. Recently, its possible topical use in ameliorating skin conditions has also been proposed; however, its role in preventing cigarette smoke (CS)-induced keratinocyte damage has not been investigated yet. Because of its peculiar location, cutaneous tissue is constantly exposed to several environmental stressors, such as CS. Many compounds presented in CS, have been shown to induce, directly or indirectly, cellular oxidative stress (OS) and inflammation via the production of ROS and lipid peroxidation compounds, among which 4HNE has been shown to be one of the most reactive. In this study, we have shown that resveratrol (at a dose of 10 μM) can decrease CS-induced ROS and carbonyl formation in human keratinocytes. In addition, pre-treatment with resveratrol prevented the induction of TRPA1 expression (mRNA and protein levels), a known receptor involved in cellular differentiation and inflammation, which has been recently shown to be activated by 4HNE. Finally, in keratinocytes, resveratrol could increase the expression of MsrA, enzyme involved in cell defence against oxidative protein damage. The present study further confirms the idea that the topical use of resveratrol can provide a good defence against CS-induced skin damage.

The aim of the present study was to develop controlled release inhalable lipid microparticles (LMs) loaded with the antioxidant flavonoid, quercetin and to investigate the interaction of these microparticles with A549 pulmonary alveolar epithelial cells. The LMs were produced using different lipidic materials and surfactants, by melt emulsification followed by a sonication step. The most efficient modulation of the in vitro release of quercetin was achieved by the LMs prepared with tristearin and hydrogenated phosphatidylcholine, which were used for subsequent studies. These LMs exhibited a quercetin loading of 11.8±0.3%, and a volume median diameter, determined by laser diffraction, of 4.1±0.2μm. Moreover, their mass median aerodynamic diameter (4.82±0.15μm) and fine particle fraction (27.2±3.9%), as measured by multi-stage liquid impinger, were suitable for pulmonary delivery. Quercetin was found to be highly unstable (complete decomposition within 6-h incubation) in Ham's F-12 medium used for A549 cell culture. Degradation was markedly reduced (16.4% of the initial quercetin content still present after 24-h incubation) after encapsulation in the lipid particle system. Viability studies performed by lactate dehydrogenase assay, demonstrated that quercetin LMs showed no significant cytotoxicity on the A549 cells, over the concentration 0.1-5μM. The uptake of quercetin by the A549 lung alveolar cells was also investigated. After 4-h incubation, the accumulation of quercetin in the A549 cells was significantly higher (2.3-fold increase) for the microparticle entrapped flavonoid when compare to non-encapsulated quercetin. The enhanced intracellular delivery of quercetin achieved by the LMs is likely due to the flavonoid stabilization after encapsulation., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The modulation of the HDL receptor scavenger receptor B1 (SRB1) was evaluated in skin fibroblasts isolated from patients with Rett syndrome (RTT), a genetic form of infantile autism. Patients showed an altered plasma lipid profile, while their skin fibroblasts showed a dramatic reduction in SRB1 (immunogold, Western blot and immunohistochemistry). The decreased SRB1 levels were demonstrated to be the consequence of its binding with 4-hydroxy-2-nonenal (4HNE), a product of lipid peroxidation, and its increased ubiquitination. Our findings show for the first time a loss of SRB1 in RTT cells and its relationship with a chronic oxidative stress status., (Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Selective COX-2 inhibitors (COXib) belonging to the class of diaryl heterocycles (e.g., celecoxib, rofecoxib, etc.), are devoid of the undesirable effects due to their capacity to inhibit selectively inducible (COX-2), responsible for inflammatory effects but not constitutive cyclooxygenase-1 (COX-1)(COX); responsible for cytoprotective effects on gastric mucosa. In addition, several reports have identified an increased risk of cardiovascular events associated with the use of COXib. We have developed a new series of anti-inflammatory agents (1,5-diarylpyrrole-3-alkoxyethyl esters and ethers). To evaluate the effect of two 1,5-diarylpyrrole-3-alkoxyethyl ethers, VA441 and VA428 (up to 100 μM), respectively, in comparison with two well known COXib, celecoxib and rofecoxib, on HaCaT cell (keratinocytes) proliferation and toxicity. Crucial molecules in cell cycle progression, i.e. NFκB and ERK as targets/mediators and cyclin D1 and p21 Cip1/Kip as final effectors were evaluated by Western blot, immunohystochemistry and q-PCR analysis. Both compounds, VA441 and VA428, showed a strong inhibition of cell proliferation, and did not exhibit cytotoxicity. The anti-proliferative effect was accompanied by a strong activation of ERK and induction of the cell cycle inhibitor p21. In addition, there was a clear inhibition of the transcription factor NF-κB and downregulation of cyclin D1, with enforced inhibition of the HaCaT cell cycle progression. These data suggest that compounds VA441 and VA428, along with their role in inhibiting COX-2 and inflammation, could have a possible therapeutic (topical and systemic) use against skin proliferative disorders, such as psoriasis., (Copyright © 2013 Elsevier B.V. All rights reserved.)

We investigated the effect of high-frequency electromagnetic fields (HF-EMFs) and 17-β-estradiol on connexins (Cxs), integrins (Ints), and estrogen receptor (ER) expression, as well as on ultrastructure of trophoblast-derived HTR-8/SVneo cells. HF-EMF, 17-β-estradiol, and their combination induced an increase of Cx40 and Cx43 mRNA expression. HF-EMF decreased Int alpha1 and β 1 mRNA levels but enhanced Int alpha5 mRNA expression. All the Ints mRNA expressions were increased by 17-β-estradiol and exposure to both stimuli. ER-β mRNA was reduced by HF-EMF but augmented by 17-β-estradiol alone or with HF-EMF. ER-β immunofluorescence showed a cytoplasmic localization in sham and HF-EMF exposed cells which became nuclear after treatment with hormone or both stimuli. Electron microscopy evidenced a loss of cellular contact in exposed cells which appeared counteracted by 17-β-estradiol. We demonstrate that 17-β-estradiol modulates Cxs and Ints as well as ER-β expression induced by HF-EMF, suggesting an influence of both stimuli on trophoblast differentiation and migration.

Rett syndrome (RTT) is mainly caused by mutations in the X-linked methyl-CpG binding protein (MeCP2) gene. By binding to methylated promoters on CpG islands, MeCP2 protein is able to modulate several genes and important cellular pathways. Therefore, mutations in MeCP2 can seriously affect the cellular phenotype. Today, the pathways that MeCP2 mutations are able to affect in RTT are not clear yet. The aim of our study was to investigate the gene expression profiles in peripheral blood lymphomonocytes (PBMC) isolated from RTT patients to try to evidence new genes and new pathways that are involved in RTT pathophysiology. LIMMA (Linear Models for MicroArray) and SAM (Significance Analysis of Microarrays) analyses on microarray data from 12 RTT patients and 7 control subjects identified 482 genes modulated in RTT, of which 430 were upregulated and 52 were downregulated. Functional clustering of a total of 146 genes in RTT identified key biological pathways related to mitochondrial function and organization, cellular ubiquitination and proteosome degradation, RNA processing, and chromatin folding. Our microarray data reveal an overexpression of genes involved in ATP synthesis suggesting altered energy requirement that parallels with increased activities of protein degradation. In conclusion, these findings suggest that mitochondrial-ATP-proteasome functions are likely to be involved in RTT clinical features.

Purpose: To compare the short-term outcome of dental implant therapy in a group of organ transplant patients with that of a control group., Materials and Methods: The study population included consecutive organ transplant patients and consecutive normal (healthy) subjects as controls. Two films were taken of all patients: one at baseline (implant placement) and one after 3 months of healing. All radiographs were analyzed twice (15 days apart) blindly by two independent trained radiologists. Crestal bone level (CBL) was measured, defined as the perpendicular distance from the reference point on the implant to the first visible apical bone-to-implant contact., Results: The study population included 10 organ transplant patients (eight hearts, two livers) and 10 control patients, who received 20 and 12 submerged dental implants, respectively. At the 3-month follow-up visit, no implants showed any exposed cover screws. CBL increased in both groups, without any significant difference between the groups (CBL increased from 0.08±0.09 mm to 0.28±0.20 mm in transplant patients and from 0.11±0.16 mm to 0.42±0.32 mm in controls). Multiple analysis of variance showed that the mean bone loss of 0.21±0.18 mm observed in the group of transplant patients was not statistically different from that (0.32±0.25 mm) seen in the control group and was not influenced by any of the variables considered., Conclusions: The present pilot study seems to indicate that the bone response around submerged dental implants in immunocompromised organ transplant patients does not differ from that observed in control patients and that this particular population of patients may be successfully rehabilitated with dental implants.

Living organisms are continuously exposed to environmental pollutants. Because of its critical location, the skin is a major interface between the body and the environment and provides a biological barrier against an array of chemical and physical environmental pollutants. The skin can be defined as our first defense against the environment because of its constant exposure to oxidants, including ultraviolet (UV) radiation and other environmental pollutants such as diesel fuel exhaust, cigarette smoke (CS), halogenated hydrocarbons, heavy metals, and ozone (O₃). The exposure to environmental pro-oxidant agents leads to the formation of reactive oxygen species (ROS) and the generation of bioactive molecules that can damage skin cells. This short review provides an overview of the effects and mechanisms of action of CS, O₃, and UV on cutanous tissues., (© 2012 New York Academy of Sciences.)

Objective: This study aimed to see whether clinical healing after amalgam removal corresponds to histologic healing, i.e., a complete disappearance of any histologic sign of lichenoid lesion., Study Design: The study evaluated 64 patients with lichenoid lesions and at least one amalgam filling., Results: After amalgam removal, complete clinical healing was obtained in 14 patients (22%) and was significantly related to lesion topography (χ(2) 4.7; P < .05) and positive patch test (χ(2) 6.3; P < .01). Complete histologic healing was obtained in only 7 cases (50% of clinically healed patients), and was significantly related to the combination of positive patch test and strict contact with amalgams (Fisher's exact test P < .01)., Conclusions: Contact with amalgams and positive patch testing are good but not absolute indicators of the beneficial effect of amalgam replacement. In addition, complete clinical healing does not necessarily mean a disappearance of the histologic characteristics of OLL/OLP lesions., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Laser excision has been used with increasing success to treat oral leukoplakia. The aim of the present study was to evaluate whether clinical healing of a leukoplakia after laser surgery is associated with a normal functional status of the new epithelium and whether pathological alterations in these parameters are related to the risk of local recurrence. The study population consisted of 13 consecutive patients with oral leukoplakia in which clinical healing was achieved after laser therapy using an Nd:YAG laser (Model 6000, Laser Sonics, Cooper Laser Sonics). At the end of the therapy, all patients underwent a second biopsy of the clinically healthy tissue. Epithelial cell turnover was evaluated before and after laser surgery by Ki67 protein expression, and positive staining of more than 20% was considered abnormal. All patients were followed on a monthly basis. Eight patients had abnormally high Ki67 values before laser therapy (mean 27.4 ± 6.2%), but the levels decreased significantly after treatment (17.6 ± 8.5%; t = 2.6, p < 0.05). High Ki67 values persisted in three patients, and local recurrences in the new epithelium was observed in two of these patients. Kaplan-Meier statistics showed that the between-group difference was statistically significant (Chi square 7.3; p < 0.01). In conclusion, this is the first prospective study to show that clinical healing of leukoplakia treated by laser surgery may be accompanied by altered cell turnover in 20% of the cases. Ki67, as a marker of proliferative status, may be a prognostic indicator in the mucosa replacing the lesion.

This report describes a rare case of glycogenic acanthosis (GA) located on the tongue of a 72-year-old man. The patient presented with a white plaque that had been on the right side of the tongue for 5 years. Histologically, the lesion was characterised by thickened squamous epithelium with abundant intracellular glycogen deposits. No epithelial dysplasia was noted. This is the first report of GA involving the tongue. This benign condition should be added to the large number of lesions responsible for leukoplakia in this anatomical location.

Aim:   To examine a group of lesions that progressed to oral squamous cell carcinoma (OSCC) to determine whether p16(INK4A) expression is an early finding during malignant transformation, and whether immunohistochemical evaluation of p16(INK4A) is an appropriate prognostic marker., Methods and Results:   Twenty cases of OSCC were investigated. All cases had had a biopsy on the same site as OSCC performed at least 1year before OSCC (range 1-11years; mean 3.15±3.1years). Twenty specimens from normal oral mucosa served as controls. p16(INK4A) expression was evaluated by immunohistochemical analysis and cases showing >5% of stained cells were defined as 'positive'. All 20 control cases were negative for p16(INK4A) . Oral lesions were p16(INK4A) -positive in nine cases and negative in 11. No significant relationship was found between p16(INK4A) positivity and the presence/absence of dysplasia. Among OSCC, nine tumours showed p16(INK4A) positivity and 11 showed negativity. A significant relationship (χ(2)=7.1; P<0.01) was found between the presence/absence of p16(INK4A) staining in OSCC and the presence/absence of p16(INK4A) staining in lesions preceding OSCC., Conclusions:   p16(INK4A) immunohistochemistry has a potential role in detecting a subset of p16(INK4A) -positive lesions with malignant potential., (© 2010 Blackwell Publishing Limited.)

Connexins (Cx) are membrane proteins able to influence trophoblast functions. Here we investigated the effect of high-frequency electromagnetic fields (HF-EMF) on Cx expression and localization in extravillous trophoblast cell line HTR-8/SVneo. We also analysed cell ultrastructural changes induced by HF-EMF exposure. Samples were exposed to pulse-modulated 1817 MHz sinusoidal waves (GSM-217 Hz; 1h: SAR of 2 W/kg). Cx mRNA expression was assessed through semi-quantitative RT-PCR, protein expression by Western blotting, protein localization by indirect immunofluorescence, cell ultrastructure using electron microscopy. HF-EMF exposure significantly and selectively increased Cx40 and Cx43, without altering protein expression. Nevertheless, Cx40 and Cx43 lost their punctuate fluorescence within the cell membrane, becoming diffuse after HF-EMF exposure. Electron microscopy evidenced a sharp decrease in intercellular gap junction-like structures. This study is the first to indicate that exposure of extravillous trophoblast to GSM-217 Hz signals can modify Cx gene expression, Cx protein localization and cellular ultrastructure.

Neoangiogenesis is a response to retinal hypoxia that is inhibited by somatostatin (SRIF) through its subtype 2 receptor (sst2). Using a mouse model of hypoxia-induced retinopathy, we investigated whether inhibition of adenylyl cyclase (AC) is involved in SRIF anti-angiogenic actions. Hypoxia increased AC responsiveness in wild type (WT) retinas and in retinas lacking sst2, but not in sst2-overexpressing retinas. Hypoxia also altered AC isoform expression with different patterns depending on sst2 expression level. The AC VII isoform mRNA and protein resulted the most affected. Indeed, in hypoxia AC VII expression was enhanced in WT retinas and it was further increased in sst2-lacking retinas, whereas in sst2 overexpressing retinas the increase of AC VII was lower than in WT retinas. These data suggest an involvement of AC/cAMP in mediating both hypoxia-evoked retinal neoangiogenesis and SRIF protective actions. The AC VII isoform is a candidate to a main role in these mechanisms.

p53 over-expression has been proposed as a reliable marker associated to oral carcinogenesis, although only about 50% of oral carcinomas (OSCC) are associated with p53 over-expression and even p53-negative lesions can progress to OSCC. The aim of the study was to determine whether the combination of p53 over-expression and p53 low-expression associated with Ki67 over-expression (high Ki67/p53 ratio) could lead to a more sensitive parameter. Immunohistochemical expression of Ki67 and p53 was measured in 54 specimens from OSCC; 27 specimens from moderate/severe epithelial dysplasia; 32 specimens from oral leukoplakias without epithelial dysplasia, and 13 specimens with normal epithelium. p53 over-expression was found in 31 (53%) samples from OSCC, in 10 (37%) samples from severe dysplasias, and in 5 (15%) samples from non-dysplastic lesions, while the combination of high p53 values with high Ki67/p53 ratio was observed in 93% of OSCC, in 81% of dysplastic lesions, and in 50% of non-dysplastic lesions. This parameter may have a clinical implication to detect early lesions with an impairment of p53 pathway, and probably at risk of progress to OSCC.

Our understanding of endothelial physiology is overdue compared with other areas of study. For too many years the complex mysteries of this thin single-layered cellular lamina covering the whole of the vascular network, from the large conduction vessel to the smallest resistance and diffusion vessel, have been hidden by an organ-based science more focused on organ pathology than on ultrastructural physiopathology. We tried to follow chronologically the alteration stages of this system of membranes in relation to the development of the atherosclerotic plaque in human biopsy.

The role of G proteins and related second messenger system on the modulation of acetylcholine release from [3H]choline-preloaded guinea-pig superior cervical ganglion was investigated using the potent general activator NaF. The electrically evoked (1 Hz, 5 min) [3H] release was inhibited by "low" F- concentrations (1-2.5 mM), by the adenylyl cyclase blocker MDL 12330A (10 microM), alone and in combination with 1 mM NaF, and increased by 0.5 mM 8Br-cAMP, 100 microM forskolin and 0.5 mM 3-isobutyl-1-methylxantine. No effect of 1 mM F- was observed on spontaneous release. Fluoride-induced inhibition was counteracted by the G protein blocker sulmazole (1 mM), forskolin and alteration of calcium influx by increasing [Ca2+]out from 2.2 to 6 mM, raising the rate of stimulation (10 Hz, 30 s), or broadening the presynaptic action potential with 10 microM 4-aminopyridine and 50 microM tetraethylammonium chloride. Thus a NaF-sensitive G protein, linked to cAMP synthesis, is determinant for the inhibition of neurosecretion in this cholinergic synapse, involving Ca2+-dependent mechanisms., (Copyright 2004 Elsevier Ireland Ltd.)

The aim of the present paper was to study production of methacrylate microparticles for the delivery (administration) of ascorbic acid via the oral route. Vitamin C is an important antioxidant that may be involved in the reduction of the risk of certain types of cancer, such as colorectal cancer. As polymers different acrylic compounds were considered, namely Eudragit RL, L and RS. Spray-drying was used as preparation method of vitamin C/Eudragit microspheres. Microspheres were first characterized by size and morphology by scanning electron microscopy, then in vitro release kinetics by mean of dialysis method were studied. Although the produced microparticles were unable to slow down the release of the drug with respect to the free form of ascorbic acid, these microspheres showed a good morphology and size distribution that permit to propose them as candidate for the delivery of vitamin C as associated therapy in the treatment of colorectal cancer by oral route.

Purpose: To evaluate the effectiveness of a dentin bonding agent containing HEMA compared to a resin emulsion in the reduction of cervical dentin hypersensitivity (CDH)., Materials and Methods: The study was a randomized-within-subject, double blind design. Twenty patients who were suffering from CDH from at least two teeth (canines and/or premolars) were enrolled. The subjective perceptions of pain in response to tactile, air blasts and cold ethyl chloride stimuli were evaluated using a visual analog scale (VAS) at baseline and 10 minutes, 1 week and 4 weeks after the treatments. Scotchbond 1 (a dentin bonding agent similar to Single Bond in the USA) and MS Coat (a resin emulsion), both applied according to the manufacturers' directions, were selected for the study., Results: Multiple regression ANOVA for repeated measures showed that both treatments significantly reduced the CDS with respect the baseline values after just 10 minutes. There were no significant differences between the ability of the two treatments to reduce CDH for each of the three evaluation stimuli or for subjective evaluation.

Starting from a group of 736 renal biopsy patients, evaluated by ultrastructural studies over a period of 22 years, the authors present a rare case of immunotactoid glomerulopathy, suggesting that these forms, until a few years ago considered in the same group as fibrillary glomerulonephritis, are in fact a separate entity; moreover, they may represent a very early manifestation of plasmacellular dyscrasia still at the initial stage.

The aim of this study was to evaluate the influence of operating parameters on the characteristics of methacrylate microparticles prepared by spray-drying technique. Eudragit microparticles were prepared by a spray-drying method. The influence of different experimental parameters (i.e., solvent, feed rate, air flow rate, air-drying temperature, and aspiration flow rate) on microparticle morphology, size distribution, and recovery was studied. In addition, different Eudragit types and Eudragit RS concentrations were employed. Optical and electron microscopy were employed to analyze microparticle morphology and dimensional distribution. Finally, prednisolone as model drug was encapsulated in Eudragit RS microparticles. Low feed rate values yielded the best results in terms of microparticle morphology. Changes in nebulizing air flow did not result in a corresponding effect on microparticle characteristics. An increase of air-drying temperature resulted in a reduction of microparticle dimension and recovery. A low flow of drying air was preferable because this resulted in microparticles with an optimal morphology. The polymer concentration affected both morphology and dimensions of microparticles. The encapsulation of prednisolone led to good incorporation efficiencies without altering percentage of recovery, morphology, and mean dimension of the microparticles. The selection of appropriate parameters yielded spray-dried Eudragit RS microparticles characterized by good morphology and narrow dimensional distribution.

We investigated the ultrastructure of Birbeck granules which are found in some malignant histiocytoses such as histiocytosis X, Letterer-Siwe disease, Hand-Schüller-Christian disease, eosinophilic granuloma of the bone and self-healing reticulohistiocytosis. The research is based on the systemic study of Birbeck granules, from their formation to intracytoplasmic development, examining with the electron microscope at regular intervals ultrathin sections derived from biopsies of two cases of Langerhans cell histiocytosis.

Gastric metaplasia is a frequent epithelial change in gallbladder of lithiasic patients. Particles consistent with cholesterol-containing vesicles were searched in areas overlying gastric metaplastic cells, in gastric metaplastic cells, in areas overlying normal-looking epithelial cells and in normal-looking epithelial cells in 25 patients with radiolucent calculi and 10 control patients undergoing cholecystectomy for other reasons. Important trends were found confirming the peculiar role of mucous layer adherent to metaplastic gallbladder epithelium in beginning the nucleation process and interesting pictures concerning the formation of uni- and multilamellar vesicles in these "nucleating areas" were seen.

Xanthine oxidase increases the rate of actin polymerization. This occurs at oxidase concentrations as low as 40 nM provided the concentration of the polymerizing agent is low (0.5 mM MgCl2). In the presence of 0.1 M KCl plus 1 mM MgCl2 as the polymerizing agents, xanthine oxidase does not affect the rate of the polymerization but increases significantly the rate of the conversion of F(ATP)actin into F(ADP.Pi)actin and probably also the rate of the orthophosphate release.