Multispectral fingerprinting for improved in vivo cell dynamics analysis

Background: Tracing cell dynamics in the embryo becomes tremendously difficult when cell trajectories cross in space and time and tissue density obscure individual cell borders. Here, we used the chick neural crest (NC) as a model to test multicolor cell labeling and multispectral confocal imaging strategies to overcome these roadblocks. Results: We found that multicolor nuclear cell labeling and multispectral imaging led to improved resolution of in vivo NC cell identification by providing a unique spectral identity for each cell. NC cell spectral identity allowed for more accurate cell tracking and was consistent during short term time-lapse imaging sessions. Computer model simulations predicted significantly better object counting for increasing cell densities in 3-color compared to 1-color nuclear cell labeling. To better resolve cell contacts, we show that a combination of 2-color membrane and 1-color nuclear cell labeling dramatically improved the semi-automated analysis of NC cell interactions, yet preserved the ability to track cell movements. We also found channel versus lambda scanning of multicolor labeled embryos significantly reduced the time and effort of image acquisition and analysis of large 3D volume data sets. Conclusions: Our results reveal that multicolor cell labeling and multispectral imaging provide a cellular fingerprint that may uniquely determine a cell's position within the embryo. Together, these methods offer a spectral toolbox to resolve in vivo cell dynamics in unprecedented detail

Formation and removal of alkylthiolate self-assembled monolayers on gold in aqueous solutions

We report the development of novel reagents and approaches for generating recyclable biosensors. The use of aqueous media for the formation of protein binding alkylthiolate monolayers on Au surfaces results in accelerated alkylthiolate monolayer formation and improvement in monolayer integrity as visualized by fluorescence microscopy and CV techniques. We have also developed an electrocleaning protocol that is compatible with microfluidics devices, and this technique serves as an on-chip method for cleaning Au substrates both before and after monolayer formation. The techniques for the formation and dissociation of biotinylated SAMs from aqueous solvents reported here may be applied towards the development of Au-based sensor devices and microfluidics chips in the future. A potential use of these devices includes the specific capture and triggered release of target cells, proteins, or small molecules from liquid samples

The Future of Parenting Programs: II Implementation

This article examines the role that implementation science can play in evidence-based parenting programs. Although parenting programs can support parents in their caregiving roles, adapting and taking an evidence-based approach from one place to another without attending to implementation factors may contribute to poor impact in a new setting. Implementation science enables researchers to move beyond monitoring and evaluation of outcomes of a parenting program to understanding the process of putting the program into practice. Factors such as whether the program meets the needs of families and communities, how to secure buy-in from key stakeholders, what training and supervision are needed for the workforce, and ways that parenting programs can be integrated in existing infrastructure are all critical to successful implementation. Quality improvement can be built into the implementation process through feedback loops that inform rapid changes and testing cycles over time as a program is implemented. If researchers lead initial implementation of parenting programs, they must determine how the program can continue to work when using community workers and local systems rather than researchers. Open access components are especially important for the implementation of parenting programs in low- and middle-income countries to avoid prohibitive costs of proprietary programs and to benefit from flexibility in adapting components to meet the needs of particular local populations. Parenting programs benefit when policy makers, program leaders, and researchers attend not only to the what but also to the how of implementation.Objective.Design.Results.Conclusions

“They Aren’t Going to Do Jack Shit”: Text-Based Crisis Service Users’ Perceptions of Seeking Child Maltreatment-Related Support From Formal Systems

Many of the children reported to child protective services (CPS) exhibit signs and symptoms that allow others to recognize their abuse or neglect and intervene; others, especially adolescents, must disclose their experiences to be identified. Relatively little is known about young people’s disclosure experiences, but individual, interpersonal, and cultural factors appear to influence when and how young people disclose. Technology-facilitated approaches, such as text- or chat-based hotlines or crisis services, may be one way to help young people share their maltreatment experiences and seek help. The current study contributes to the small body of literature that includes nonsexual maltreatment disclosures and sheds some light on how to support young people during their disclosures. We conducted a qualitative content analysis of all conversations from a text-based crisis service that resulted in a report to CPS (n = 244). Many of the texters had previously sought support from their peers or parents, and some had engaged with more formal systems. Many young people were hesitant to reach out to formal systems in the future, in part because of negative experiences during past disclosure experiences. Young people may be more likely to seek support through their preferred communication medium, so providing text- and chat-based communication may be one way to encourage and facilitate disclosure. As these resources become increasingly available, determining best practices for receiving disclosures through technology-facilitated platforms will be critical

Severity and justness do not moderate the relation between corporal punishment and negative child outcomes: A multicultural and longitudinal study

There is strong evidence of a positive association between corporal punishment and negative child outcomes, but previous studies have suggested that the manner in which parents implement corporal punishment moderates the effects of its use. This study investigated whether severity and justness in the use of corporal punishment moderate the associations between frequency of corporal punishment and child externalizing and internalizing behaviors. This question was examined using a multicultural sample from eight countries and two waves of data collected one year apart. Interviews were conducted with 998 children aged 7–10 years, and their mothers and fathers, from China, Colombia, Italy, Jordan, Kenya, Philippines, Thailand, and the United States. Mothers and fathers responded to questions on the frequency, severity, and justness of their use of corporal punishment; they also reported on the externalizing and internalizing behavior of their child. Children reported on their aggression. Multigroup path models revealed that across cultural groups, and as reported by mothers and fathers, there is a positive relation between the frequency of corporal punishment and externalizing child behaviors. Mother-reported severity and father-reported justness were associated with child-reported aggression. Neither severity nor justness moderated the relation between frequency of corporal punishment and child problem behavior. The null result suggests that more use of corporal punishment is harmful to children regardless of how it is implemented, but requires further substantiation as the study is unable to definitively conclude that there is no true interaction effect

Estimating the lifecycle of pavement markings on primary and secondary roads in South Carolina

The absence of systematic procedures and standardized methods to quantitatively evaluate pavement marking materials on South Carolina's primary and secondary roads has made it difficult for the South Carolina Department of Transportation ) to track performance and determine lifecycle duration of pavement markings from installation to eventual restriping applications. This report summarizes findings of a three-year research project and includes a literature search, discussion of data collection and analysis, development of retroreflectivity degradation models, comparison of marking materials, and identification of recommended guidelines

Avian Primordial Germ Cells Contribute to and Interact With the Extracellular Matrix During Early Migration

During early avian development, primordial germ cells (PGC) are highly migratory, moving from the central area pellucida of the blastoderm to the anterior extra-embryonic germinal crescent. The PGCs soon move into the forming blood vessels by intravasation and travel in the circulatory system to the genital ridges where they participate in the organogenesis of the gonads. This complex cellular migration takes place in close association with a nascent extracellular matrix that matures in a precise spatio-temporal pattern. We first compiled a list of quail matrisome genes by bioinformatic screening of human matrisome orthologs. Next, we used single cell RNA-seq analysis (scRNAseq) to determine that PGCs express numerous ECM and ECM-associated genes in early embryos. The expression of select ECM transcripts and proteins in PGCs were verified by fluorescent in situ hybridization (FISH) and immunofluorescence (IF). Live imaging of transgenic quail embryos injected with fluorescent antibodies against fibronectin and laminin, showed that germinal crescent PGCs display rapid shape changes and morphological properties such as blebbing and filopodia while surrounded by, or in close contact with, an ECM fibril meshwork that is itself in constant motion. Injection of anti-β1 integrin CSAT antibodies resulted in a reduction of mature fibronectin and laminin fibril meshwork in the germinal crescent at HH4-5 but did not alter the active motility of the PGCs or their ability to populate the germinal crescent. These results suggest that integrin β1 receptors are important, but not required, for PGCs to successfully migrate during embryonic development, but instead play a vital role in ECM fibrillogenesis and assembly

Prometastatic GPCR CD97 Is a Direct Target of Tumor Suppressor microRNA-126

Tumor suppressor microRNA-126 (miR-126) is often down-regulated in cancer cells, and its overexpression is found to inhibit cancer metastasis. To elucidate the mechanism of tumor suppression by miR-126, we analyzed the proteomic response to miR-126 overexpression in the human metastatic breast cancer cell line MDA-MB-231. To acquire quantitative, time-resolved information, we combined two complementary proteomic methods, BONCAT and SILAC. We discovered a new direct target of miR-126: CD97, a pro-metastatic G-protein-coupled receptor (GPCR) that has been reported to promote tumor cell invasion, endothelial cell migration, and tumor angiogenesis. This discovery establishes a link between down-regulation of miR-126 and overexpression of CD97 in cancer and provides new mechanistic insight into the role of miR-126 in inhibiting both cell-autonomous and non-cell-autonomous cancer progression

FAPRI 1999 U.S. Agricultural Outlook

Crop Production/Industries, Livestock Production/Industries,