Sheath fluid impacts the depletion of cellular metabolites in cells afflicted by sorting induced cellular stress (SICS).

Flow cytometrists have long observed a spectrum of cell-type-specific changes ranging from minor functional defects to outright cell destruction after purification of cells using conventional droplet cell sorters. We have described this spectrum of cell perturbations as sorter induced cellular stress, or SICS (Lopez and Hulspas, Cytometry, 2020, 97, 105-106). Despite the potential impact of this issue and ubiquitous anecdotes, little has been reported about this phenomenon in the literature, and the underlying mechanism has been elusive. Inspired by others' observations (Llufrio et al., Redox Biology, 2018, 16, 381-387 and Binek et al., Journal of Proteome Research, 2019, 18, 169-181), we set out to examine SICS at the metabolic level and use this information to propose a working model. Using representative suspension (Jurkat) and adherent (NIH/3T3) cell lines we observed broad and consistent metabolic perturbations after sorting using a high-speed droplet cell sorter. Our results suggest that the SICS metabolic phenotype is a common cell-type-independent manifestation and may be the harbinger of a wide-range of functional defects either directly related to metabolism, or cell stress response pathways. We further demonstrate a proof of concept that a modification to the fluidic environment (complete media used as sheath fluid) in a droplet cell sorter can largely rescue the intracellular markers of SICS, and that this rescue is not due to a contribution of metabolites found in media. Future studies will focus on characterizing the potential electro-physical mechanisms inherent to the droplet cell sorting process to determine the major contributors to the SICS mechanism.

Survey on Scientific Shared Resource Rigor and Reproducibility.

Shared scientific resources, also known as core facilities, support a significant portion of the research conducted at biomolecular research institutions. The Association of Biomolecular Resource Facilities (ABRF) established the Committee on Core Rigor and Reproducibility (CCoRRe) to further its mission of integrating advanced technologies, education, and communication in the operations of shared scientific resources in support of reproducible research. In order to first assess the needs of the scientific shared resource community, the CCoRRe solicited feedback from ABRF members via a survey. The purpose of the survey was to gain information on how U.S. National Institutes of Health (NIH) initiatives on advancing scientific rigor and reproducibility influenced current services and new technology development. In addition, the survey aimed to identify the challenges and opportunities related to implementation of new reporting requirements and to identify new practices and resources needed to ensure rigorous research. The results revealed a surprising unfamiliarity with the NIH guidelines. Many of the perceived challenges to the effective implementation of best practices (i.e., those designed to ensure rigor and reproducibility) were similarly noted as a challenge to effective provision of support services in a core setting. Further, most cores routinely use best practices and offer services that support rigor and reproducibility. These services include access to well-maintained instrumentation and training on experimental design and data analysis as well as data management. Feedback from this survey will enable the ABRF to build better educational resources and share critical best-practice guidelines. These resources will become important tools to the core community and the researchers they serve to impact rigor and transparency across the range of science and technology.

Do hostile sexual beliefs affect men's perceptions of sexual-interest messages?

This study tested the distorted cognitions component of Hall and Hirschman's (1991) quadripartite model of sexual aggression. Men (N = 107) with and without hostile sexual beliefs viewed erotic slides with a female partner who provided one of four patterns of feedback: clear disinterest, token disinterest, compliant interest, and clear interest. Hostile men reported fewer differences between women, were unresponsive to their partner's perceived embarrassment, and reported a consistent positive mood regardless of her feedback. Conversely, nonhostile men were more responsive to feedback, mirrored the partner's embarrassment, and experienced a less positive mood when she communicated clear disinterest in the erotica. These findings support the distorted cognitions component of the quadripartite model of sexual aggression. The authors also discuss the strengths and limitations of this study's methodology.

Postdrinking sexual perceptions and behaviors toward another person: alcohol expectancy set and gender differences.

Based on research showing that alcohol expectancy and gender both play a role in sexual perceptions, we factorially crossed the apparent drinking status of yoked pairs consisting of a participant and a target person (a confederate posing as co-participant). Alcohol expectancy interacted with gender in complex ways to influence sexual perceptions. We also found behavioral effects: Men showed more erotic material to both male and female targets than women did. Men perceived the target as more sexually aroused by erotic material than women did. Men also showed more erotic material to drinking targets than to non-drinking targets. Sexual perceptions and erotica showing behavior were correlated significantly and positively. These findings are discussed in terms of implications for postdrinking heterosexual encounters.

The gamma-secretase-generated intracellular domain of beta-amyloid precursor protein binds Numb and inhibits Notch signaling.

The beta-amyloid precursor protein (APP) and the Notch receptor undergo intramembranous proteolysis by the Presenilin-dependent gamma-secretase. The cleavage of APP by gamma-secretase releases amyloid-beta peptides, which have been implicated in the pathogenesis of Alzheimer's disease, and the APP intracellular domain (AID), for which the function is not yet well understood. A similar gamma-secretase-mediated cleavage of the Notch receptor liberates the Notch intracellular domain (NICD). NICD translocates to the nucleus and activates the transcription of genes that regulate the generation, differentiation, and survival of neuronal cells. Hence, some of the effects of APP signaling and Alzheimer's disease pathology may be mediated by the interaction of APP and Notch. Here, we show that membrane-tethered APP binds to the cytosolic Notch inhibitors Numb and Numb-like in mouse brain lysates. AID also binds Numb and Numb-like, and represses Notch activity when released by APP. Thus, gamma-secretase may have opposing effects on Notch signaling; positive by cleaving Notch and generating NICD, and negative by processing APP and generating AID, which inhibits the function of NICD.

Jun NH2-terminal kinase (JNK) interacting protein 1 (JIP1) binds the cytoplasmic domain of the Alzheimer's beta-amyloid precursor protein (APP).

The familial Alzheimer's disease gene product amyloid beta precursor protein (APP) is sequentially processed by beta- and gamma-secretases to generate the Abeta peptide. The biochemical pathway leading to Abeta formation has been extensively studied since extracellular aggregates of Abeta peptides are considered the culprit of Alzheimer's disease. Aside from its pathological relevance, the biological role of APP processing is unknown. Cleavage of APP by gamma-secretase releases, together with Abeta, a COOH-terminal APP intracellular domain, termed AID. This peptide has recently been identified in brain tissue of normal control and patients with sporadic Alzheimer's disease. We have previously shown that AID acts as a positive regulator of apoptosis. Nevertheless, the molecular mechanism by which AID regulates this process remains unknown. Hoping to gain clues about the function of APP, we used the yeast two-hybrid system to identify interaction between the AID region of APP and JNK-interacting protein-1 (JIP1). This molecular interaction is confirmed in vitro, in vivo by fluorescence resonance energy transfer (FRET), and in mouse brain lysates. These data provide a link between APP and its processing by gamma-secretase, and stress kinase signaling pathways. These pathways are known regulators of apoptosis and may be involved in the pathogenesis of Alzheimer's disease.