Secretagogue stimulation of neurosecretory cells elicits filopodial extensions uncovering new functional release sites

Regulated exocytosis in neurosecretory cells relies on the timely fusion of secretory granules (SGs) with the plasma membrane. Secretagogue stimulation leads to an enlargement of the cell footprint (surface area in contact with the coverslip), an effect previously attributed to exocytic fusion of SGs with the plasma membrane. Using total internal reflection fluorescence microscopy, we reveal the formation of filopodia-like structures in bovine chromaffin and PC12 cells driving the footprint expansion, suggesting the involvement of cortical actin network remodeling in this process. Using exocytosis-incompetent PC12 cells, we demonstrate that footprint enlargement is largely independent of SG fusion, suggesting that vesicular exocytic fusion plays a relatively minor role in filopodial expansion. The footprint periphery, including filopodia, undergoes extensive F-actin remodeling, an effect abolished by the actomyosin inhibitors cytochalasin D and blebbistatin. Imaging of both Lifeact-GFP and the SG marker protein neuropeptide Y-mCherry reveals that SGs actively translocate along newly forming actin tracks before undergoing fusion. Together, these data demonstrate that neurosecretory cells regulate the number of SGs undergoing exocytosis during sustained stimulation by controlling vesicular mobilization and translocation to the plasma membrane through actin remodeling. Such remodeling facilitates the de novo formation of fusion sites

MHC class I-related antigen-processing machinery component defects in feline mammary carcinoma.

Defects in HLA class I antigen-processing machinery (APM) component expression and/or function are frequent in human tumors. These defects may provide tumor cells with a mechanism to escape from recognition and destruction by HLA class I antigen-restricted, tumor antigen-specific cytotoxic T cells. However, expression and functional properties of MHC class I antigens and APM components in malignant cells in other animal species have been investigated to a limited extent. However, this information can contribute to our understanding of the mechanisms underlying the association of MHC class I antigen and APM component defects with malignant transformation of cells and to identify animal models to validate targeted therapies to correct these defects. To overcome this limitation in the present study, we have investigated the expression of the catalytic subunits of proteasome (Y, X, and Z) and of immunoproteasome (LMP2, LMP7, and LMP10) as well as of MHC class I heavy chain (HC) in 25 primary feline mammary carcinomas (FMCs) and in 23 matched healthy mammary tissues. We found a reduced expression of MHC class I HC and of LMP2 and LMP7 in tumors compared with normal tissues. Concordantly, proteasomal cleavage specificities in extracts from FMCs were different from those in healthy tissues. In addition, correlation analysis showed that LMP2 and LMP7 were concordantly expressed in FMCs, and their expression was significantly correlated with that of MHC class I HC. The abnormalities we have found in the APM in FMCs may cause a defective processing of some tumor antigens

A circulating cell population showing both M1 and M2 monocyte/macrophage surface markers characterizes systemic sclerosis patients with lung involvement.

Background: Systemic sclerosis (SSc) is a disorder characterized by immune system alterations, vasculopathy and fibrosis. SSc-related interstitial lung disease (ILD) represents a common and early complication, being the leading cause of mortality. Monocytes/macrophages seem to have a key role in SSc-related ILD. Interestingly, the classically (M1) and alternatively (M2) activated monocyte/macrophage phenotype categorization is currently under revision. Our aim was to evaluate if circulating monocyte/macrophage phenotype could be used as biomarker for lung involvement in SSc. To this purpose we developed a wide phenotype characterization of circulating monocyte/macrophage subsets in SSc patients and we evaluated possible relations with lung involvement parameter values. Methods: A single centre cross-sectional study was performed in fifty-five consecutive SSc patients, during the year 2017. All clinical and instrumental tests requested for SSc follow up and in particular, lung computed tomography (CT) scan, pulmonary function tests (PFTs), Doppler echocardiography with systolic pulmonary artery pressure (sPAP) measurement, blood pro-hormone of brain natriuretic peptide (pro-BNP) evaluation, were performed in each patient in a maximum one-month period. Flow cytometry characterization of circulating cells belonging to the monocyte/macrophage lineage was performed using specific M1 (CD80, CD86, TLR2 and TLR4) and M2 surface markers (CD204, CD163 and CD206). Non-parametric tests were used for statistical analysis. Results: A higher percentage of circulating CD204 + CD163 + CD206 + TLR4 + CD80 + CD86 + and CD14 + CD206 + CD163 + CD204 + TLR4 + CD80 + CD86 + mixed M1/M2 monocyte/macrophage subsets, was identified to characterize patients affected by SSc-related ILD and higher systolic pulmonary artery pressure. Mixed M1/M2 monocyte/macrophage subset showed higher percentages in patients positive for anti-topoisomerase antibody, a known lung involvement predictor. Conclusions: The present study shows for the first time, through a wide flow cytometry surface marker analysis, that higher circulating mixed M1/M2 monocyte/macrophage cell percentages are associated with ILD, sPAP and anti-topoisomerase antibody positivity in SSc, opening the path for research on their possible role as pathogenic or biomarker elements for SSc lung involvement

Autonomous Navigation and Security: A 13000h/3000km Case Study

This paper presents the design of an autonomous mobile platform and its security system. The MB835 mobile platform has been adopted for RoboX, a fully autonomous tour guide robot. In 2002, 11 of these tour guides have served the Robotics exhibition at Expo.02 (Swiss National Exhibition) from May 15 to October 20. This project has been conjointly conducted by the Autonomous Systems Lab, Swiss Federal Institute of Technology Lausanne (EPFL) and BlueBotics SA the spin-off company of the lab, which has produced the robots. The goal was to maximize the autonomy and mobility of the platform while ensuring high performance, robustness and security. The paper presents the platform, its navigation and security, which resulted in the ANT product (Autonomous Navigation Technology) and the results of the Robotics exhibition as empirical validation of the whole system

Mechanistic relationship among mutagenicity, skin sensitization, and skin carcinogenicity.

Twenty organic Salmonella mutagens, seven of which (including benzo[a]pyrene) are established skin carcinogens, and one of which (2-chloroethanol) is a well-defined noncarcinogen to skin, have been evaluated for skin-sensitizing activity using the local lymph node assay. The relative mutagenicity of the agents to Salmonella was also established. Fourteen of the chemicals were positive in the local lymph node assay, including the seven skin carcinogens. 2-Chloroethanol was inactive as a sensitizing agent. We suggest that a variety of factors contributes to the lack of sensitizing activity of the remaining six bacterial mutagens: extremes of intrinsic chemical reactivity, high water solubility reducing dermal translocation, and inappropriate dermal metabolism. Two reference skin-sensitizing agents (an oxazolinone and fluorescein isothiocyanate) were established as in vitro clastogens after their recognition as nonmutagens to Salmonella. These data imply that mutagenicity, rather than simply activity in the Salmonella assay, is a primary stimulus for electrophilic sensitization and carcinogenic initiation in the skin. We conclude that genotoxicity data for an agent can provide indications of the agent's potential to induce skin sensitization and that genotoxins which are skin-sensitizing agents have an enhanced potential to initiate skin carcinogenesis. We suggest that common, albeit individually distinct, structure-activity relationships underpin genotoxicity, skin sensitization, and the initiation of skin carcinogenesis. These relationships should simplify the hazard evaluation of chemicals and contribute to a reduction in animal usage. Several predictions of skin carcinogenicity are made based on the data presented