The LINC complex is required for endothelial cell adhesion and adaptation to shear stress and cyclic stretch

The Linker of Nucleoskeleton and Cytoskeleton (LINC) complex is a structure consisting of nesprin, SUN, and lamin proteins. A principal function of the LINC complex is anchoring the nucleus to the actin, microtubule, and intermediate filament cytoskeletons. The LINC complex is present in nearly all cell types, including endothelial cells. Endothelial cells line the innermost surfaces of blood vessels and are critical for blood vessel barrier function. In addition, endothelial cells have specialized functions, including adaptation to the mechanical forces of blood flow. Previous studies have shown that depletion of individual nesprin isoforms results in impaired endothelial cell function. To further investigate the role of the LINC complex in endothelial cells we utilized dominant negative KASH (DN-KASH), a dominant negative protein that displaces endogenous nesprins from the nuclear envelope and disrupts nuclear–cytoskeletal connections. Endothelial cells expressing DN-KASH had altered cell–cell adhesion and barrier function, as well as altered cell–matrix adhesion and focal adhesion dynamics. In addition, cells expressing DN-KASH failed to properly adapt to shear stress or cyclic stretch. DN-KASH–expressing cells exhibited impaired collective cell migration in wound healing and angiogenesis assays. Our results demonstrate the importance of an intact LINC complex in endothelial cell function and homeostasis.     

Methods to Measure the Impact of Home,Social, and Sexual Neighborhoods of Urban Gay,Bisexual, and Other Men Who Have Sex with Men

Men who have sex with men (MSM) accounted for 61% of new HIV diagnoses in the United States in 2010. Recent analyses indicate that socio-structural factors are important correlates of HIV infection. NYCM2M was a cross-sectional study designed to identify neighborhood-level characteristics within the urban environment that influence sexual risk behaviors, substance use and depression among MSM living in New York City. The sample was recruited using a modified venue-based time-space sampling methodology and through select websites and mobile applications. This paper describes novel methodological approaches used to improve the quality of data collected for analysis of the impact of neighborhoods on MSM health. Previous research has focused predominately on residential neighborhoods and used pre-determined administrative boundaries (e.g., census tracts) that often do not reflect authentic and meaningful neighborhoods. This study included the definition and assessment of multiple neighborhoods of influence including where men live (home neighborhood), socialize (social neighborhood) and have sex (sexual neighborhood). Furthermore, making use of technological advances in mapping, we collected geo-points of reference for each type of neighborhood and identified and constructed self-identified neighborhood boundary definitions. Finally, this study collected both perceived neighborhood characteristics and objective neighborhood conditions to create a comprehensive, flexible and rich neighborhood-level set of covariates. This research revealed that men perceived their home, social and sexual neighborhoods in different ways. Few men (15%) had the same home, social and sexual neighborhoods; for 31%, none of the neighborhoods was the same. Of the three types of neighborhoods, the number of unique social neighborhoods was the lowest; the size of sexual neighborhoods was the smallest. The resultant dataset offers the opportunity to conduct analyses that will yield context-specific and nuanced understandings of the relations among neighborhood space, and the well-being and health of urban MSM.     

CATP-6, a C. elegans Ortholog of ATP13A2 PARK9, Positively Regulates GEM-1, an SLC16A Transporter

In previous work, we found that gain-of-function mutations that hyperactivate GEM-1 (an SLC16A transporter protein) can bypass the requirement for GON-2 (a TRPM channel protein) during the initiation of gonadogenesis in C. elegans. Consequently, we proposed that GEM-1 might function as part of a Mg²⁺ uptake pathway that functions in parallel to GON-2. In this study, we report that CATP-6, a C. elegans ortholog of the P5B ATPase, ATP13A2 (PARK9), is necessary for gem-1 gain-of-function mutations to suppress the effects of gon-2 inactivation. One possible explanation for this observation is that GEM-1 serves to activate CATP-6, which then functions as a Mg²⁺ transporter. However, we found that overexpression of GEM-1 can alleviate the requirement for CATP-6 activity, suggesting that CATP-6 probably acts as a non-essential upstream positive regulator of GEM-1. Our results are consistent with the notion that P5B ATPases govern intracellular levels of Mg²⁺ and/or Mn²⁺ by regulating the trafficking of transporters and other proteins associated with the plasma membrane.     

Spatial and Developmental Variation in Seed Dormancy Characteristics in the Fire-responsive Species Anigozanthos manglesii(Haemodoraceae) from Western Australia

Anigozanthos manglesii(Haemodoraceae) is a colourful, herbaceousperennial exhibiting intra- and inter-populational variationin germination in response to smoke and heat. This study investigatedthe extent and nature of this variation in A. manglesii populationscollected along a 550 km latitudinal cline from kwongan scrub(30°S) to more mesic jarrah forest habitats (34°S) insouthwestern Australia. Variation in seed germination alonga maturing inflorescence was also investigated. Germinationof this species is known to be prolific following fire, andtwo germination treatments, aerosol smoke and heat, were usedas germination cues. There was a trend of increasing responsivenessof A. manglesii to smoke with increasing latitude along a clinalrange, but there were no differences in germination of seedsalong the cline in response to control or heat treatments. Therewas no significant difference in intra-population seed germinationin response to any treatment. Lastly, a significant and increasingresponse to smoke for seeds from the apex to base of the inflorescencewas detected. This latter trend may be attributed to higherresource allocation and an accelerated after-ripening of basipetalcompared to acropetal seeds. Possible reasons for the clinalvariation are discussed.Copyright 2001 Annals of Botany Company Anigozanthos manglesii, populations, geographical cline, inflorescence level, germination, smoke, heat, fire, seed development     

Regulatory Considerations for the Classification of Video Monitoring in Dissolution Testing

The goal of this article is to discuss the classification of video recordings and images when applied to dissolution testing in USP apparatus 1 and 2. Three use cases are presented. The first case presents the use and classification of video as RBE (review by exception) data under GAMP 5. The second case presents the use of video in formulation development in a research and development environment. The third case presents a feasibility study using readily available computer vision software to recognize and measure objects in the dissolution vessel, setting the groundwork for the use of image analysis as a quantitative tool. The classification of video as “electronic data”, requiring 21 CFR part 11 compliance, versus its classification as a RBE data under GAMP 5, likely depends upon its use case. Another goal of this article is to establish a position on the use of video monitoring technology as a tool for dissolution testing that is fit for purpose and compliant with regulations regarding video data management and information.Key words: computer vision, dissolution, GAMP, USP, video monitoring     

Post-genomic Pseudomonas

A report on the 19th Whitehead Institute Symposium, Cambridge, USA, 14-16 October 2001.

     

Post-genomic Pseudomonas

A report on the Pseudomonas 2001 Meeting, Brussels, Belgium, 17-21 September 2001.