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RETURN TO ISSUEPREVREVIEWNEXT Fluorescence Lifetime Measurements and Biological Imaging

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When a molecule absorbs a photon of appropriate energy, a chain of photophysical events ensues, such as internal conversion or vibrational relaxation (loss of energy in the absence of light emission), fluorescence, intersystem crossing (from singlet state to a triplet state) and phosphorescence, as shown in the Jablonski diagram for organic molecules (Fig. 1). Each of the processes occurs with a certain probability, characterized by decay rate constants (k). It can be shown that the average length of time τ for the set of molecules to decay from one state to another is reciprocally proportional to the rate of decay: τ = 1/k. This average length of time is called the mean lifetime, or simply lifetime. It can also be shown that the lifetime of a photophysical process is the time required by a population of N electronically excited molecules to be reduced by a factor of e. Correspondingly, the fluorescence lifetime is the time required by a population of excited fluorophores to decrease exponentially to N/e via the loss of energy through fluorescence and other non-radiative processes. The lifetime of photophycal processes vary significantly from tens of femotoseconds for internal conversion1,2 to nanoseconds for fluorescence and microseconds or seconds for phosphorescence.1

Mexican Households’ Food Shopping Patterns in 2015: Analysis Following Nonessential Food and Sugary Beverage Taxes

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Objective To examine patterns of taxed and untaxed food and beverage shopping across store types after Mexico’s sugary drink and non-essential food taxes, the nutritional quality of these patterns and the socio-economic characteristics associated with them. Design We performed k -means cluster analyses using households’ percentage of food and beverage purchases from each store type (i.e. convenience stores, traditional shops (e.g. bodegas, tiendas, mom-and-pop shops), supermarkets, wholesalers and others). We calculated adjusted mean proportions of taxed and untaxed products (ml or g/capita per d) purchased in each pattern. We studied the associations between households’ SES and shopping patterns using multinomial logistic regressions. Within shopping patterns, we obtained mean volumes and proportions of taxed and untaxed food and beverage subgroups and calculated the proportion of products purchased at each store type. Setting Mexico. Participants Urban Mexican households ( n 5493) from the Nielsen Mexico Consumer Panel Survey 2015. Results We found four beverage shopping patterns and three food shopping patterns, driven by the store type where most purchases were made. For beverages, 48 % of households were clustered in the Traditional pattern and purchased the highest proportion of taxed beverages. Low-SES households had the highest probability of clustering in the Traditional beverage shopping pattern. For foods, 35 % of households were clustered into the Supermarket pattern. High-SES households had the highest probability of clustering in the Supermarket food shopping pattern. Conclusions The combination of store types where Mexican households purchase packaged foods and beverages varies. However, households in all shopping patterns and SES purchase taxed beverages mainly at traditional stores. Store-level strategies should be developed to intervene on traditional stores to improve the healthfulness of purchases.

Biomimetic Microsystems for Cardiovascular Studies

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Traditional tissue culture platforms have been around for several decades and have enabled key findings in the cardiovascular field. However, these platforms failed to recreate the mechanical and dynamic features found within the body. Organs-on-chips (OOCs) are cellularized microfluidic-based devices that can mimic the basic structure, function, and responses of organs. These systems have been successfully utilized in disease, development, and drug studies. OOCs are designed to recapitulate the mechanical, electrical, chemical, and structural features of the in vivo microenvironment. Here, we review cardiovascular-themed OOC studies, design considerations, and techniques used to generate these cellularized devices. Furthermore, we will highlight the advantages of OOC models over traditional cell culture vessels, discuss implementation challenges, and provide perspectives on the state of the field.

Epigenetic regulator BMI1 promotes alveolar rhabdomyosarcoma proliferation and constitutes a novel therapeutic target

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Rhabdomyosarcoma (RMS) is an aggressive pediatric soft tissue sarcoma. There are two main subtypes of RMS, alveolar rhabdomyosarcoma (ARMS) and embryonal rhabdomyosarcoma. ARMS typically encompasses fusion-positive rhabdomyosarcoma, which expresses either PAX3-FOXO1 or PAX7-FOXO1 fusion proteins. There are no targeted therapies for ARMS; however, recent studies have begun to illustrate the cooperation between epigenetic proteins and the PAX3-FOXO1 fusion, indicating that epigenetic proteins may serve as targets in ARMS. Here, we investigate the contribution of BMI1, given the established role of this epigenetic regulator in sustaining aggression in cancer. We determined that BMI1 is expressed across ARMS tumors, patient-derived xenografts, and cell lines. We depleted BMI1 using RNAi and inhibitors (PTC-209 and PTC-028) and found that this leads to a decrease in cell growth/increase in apoptosis in vitro, and delays tumor growth in vivo. Our data suggest that BMI1 inhibition activates the Hippo pathway via phosphorylation of LATS1/2 and subsequent reduction in YAP levels and YAP/TAZ target genes. These results identify BMI1 as a potential therapeutic vulnerability in ARMS and warrant further investigation of BMI1 in ARMS and other sarcomas.

N Domain of the Lon AAA+ Protease Controls Assembly and Substrate Choice

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The protein quality control network (pQC) plays critical roles in maintaining protein and cellular homeostasis, especially during stress. Lon is a major pQC AAA+ protease, conserved from bacteria to human mitochondria. It is the principal enzyme that degrades most unfolded or damaged proteins. Degradation by Lon also controls cellular levels of several key regulatory proteins. Recently, our group determined that Escherichia coli Lon, previously thought to be an obligate homo-hexamer, also forms a dodecamer. This larger assembly has decreased ATPase activity and displays substrate-specific alterations in degradation compared with the hexamer. Here we experimentally probe the physical hexamer-hexamer interactions and the biological roles of the Lon dodecamer. Using structure prediction methods coupled with mutagenesis, we identified a key interface and specific residues within the Lon N domain that participates in an intermolecular coiled coil unique to the dodecamer. With this knowledge, we made a Lon variant (Lon^VQ ) that forms a dodecamer with increased stability, as determined by analytical ultracentrifugation and electron microscopy. Using this altered Lon, we characterize the Lon dodecamer’s activities using a panel of substrates. Lon dodecamers are clearly functional, and complement critical lon- phenotypes but also exhibit altered substrate specificity. For example, the small heat shock proteins IbpA and IbpB are only efficiently degraded well by the hexamer. Thus, by elucidating the intermolecular contacts connecting the hexamers, we are starting to illuminate how dodecamer formation versus disassembly can alter Lon function under conditions where controlling specific activities and substrate preferences of this key protease may be advantageous.

Candida Albicans Stimulates Streptococcus Mutans Microcolony Development Via Cross-Kingdom Biofilm-Derived Metabolites

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Candida albicans is frequently detected with heavy infection of Streptococcus mutans in plaque-biofilms from children affected with early-childhood caries, a prevalent and costly oral disease. The presence of C. albicans enhances S. mutans growth within biofilms, yet the chemical interactions associated with bacterial accumulation remain unclear. Thus, this study was conducted to investigate how microbial products from this cross-kingdom association modulate S. mutans build-up in biofilms. Our data revealed that bacterial-fungal derived conditioned medium (BF-CM) significantly increased the growth of S. mutans and altered biofilm 3D-architecture in a dose-dependent manner, resulting in enlarged and densely packed bacterial cell-clusters (microcolonies). Intriguingly, BF-CM induced S. mutans gtfBC expression (responsible for Gtf exoenzymes production), enhancing Gtf activity essential for microcolony development. Using a recently developed nanoculture system, the data demonstrated simultaneous microcolony growth and gtfB activation in situ by BF-CM. Further metabolites/chromatographic analyses of BF-CM revealed elevated amounts of formate and the presence of Candida-derived farnesol, which is commonly known to exhibit antibacterial activity. Unexpectedly, at the levels detected (25–50 μM), farnesol enhanced S. mutans-biofilm cell growth, microcolony development, and Gtf activity akin to BF-CM bioactivity. Altogether, the data provide new insights on how extracellular microbial products from cross-kingdom interactions stimulate the accumulation of a bacterial pathogen within biofilms.

Dendritic Cell Activation and T Cell Priming with Adjuvant- and Antigen-Loaded Oxidation-Sensitive Polymersomes

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Quantifying Colorimetric Assays in Paper-Based Microfluidic Devices by Measuring the Transmission of Light through Paper

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This article describes a point-of-care (POC) system—comprising a microfluidic, paper-based analytical device (μ-PAD) and a hand-held optical colorimeter—for quantifying the concentration of analytes in biological fluids. The μ-PAD runs colorimetric assays, and consists of paper that has been (i) patterned to expose isolated regions of hydrophilic zones and (ii) wet with an index-matching fluid (e.g., vegetable oil) that is applied using a disposable, plastic sleeve encasement. Measuring transmittance through paper represents a new method of quantitative detection that expands the potential functionality of μ-PADs. This prototype transmittance colorimeter is inexpensive, rugged, and fully self-contained, and thus potentially attractive for use in resource-limited environments and developing countries.

Untapped Resources: Black Parent Engagement That Contributes to Learning

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Conversations around the achievement gap are often centered on what parents and students can do to close it. Overall school leaders have not made the achievement of Black students a priority, but it continues to be a priority for Black parents. Despite the vast research on parental involvement, little is known about the specific contributions of Black families to student learning. This study surveyed 130 parents/guardians of Black K–12 students throughout the United States to identify the strategies and resources they use in engaging with their children’s education. The families exhibited high rates of supporting learning at home, communicating with schools, and providing educational experiences in the community. Participants reported using two types of resources: programs and organizations geared specifically to Black students and their parents and social interactions through friends, parents, and the Internet. School personnel may improve communication and collaboration with Black parents by revisiting policy and restructuring engagement programs to incorporate this information.

The Danger of an Untold Story: Excerpts of My Life as a Black Academic.

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People of color who choose careers in higher education may encounter microaggressions, marginalization, and other unpleasant situations. This narrative shares the experiences and perspectives of one African-American academic’s experiences with the intent of encouraging academic professionals from underrepresented populations to persevere.