Genetic over-expression of cytokeratin 18 in bovine luteal cells: Effects on intermediate filament formation and Fas ligand-induced apoptosis

The receptor Fas and its ligand, FasL, are implicated in apoptosis of luteal cells during regression of the corpus luteum (CL). In some cell types, cytokeratin (CK) 8/18 filaments impair FasL-induced apoptosis. Here, we determined if genetic over-expression of CK18 filaments in luteal cells similarly prevents FasL-induced apoptosis. Luteal cell cultures were prepared from early and late stage bovine CL (day 5 and days 16-18 post-ovulation, respectively). The cells were transduced with media, mock vector containing GFP, or a CK18-containing vector, and then exposed to a cytokine cocktail containing FasL for 24 hours. Transduction of luteal cells with CK18 vector resulted in CK18 aggregates rather than filaments. The aggregates did not affect surface expression of Fas or progesterone production. Moreover, the CK18 aggregates did not prevent FasL-induced apoptosis. Thus, while successful in transducing primary cultures of bovine luteal cells with CK18, aggregation of CK failed to prevent FasL-induced apoptosis

Estrous cycle-dependent changes of Fas expression in the bovine corpus luteum: influence of keratin 8/18 intermediate filaments and cytokines

Background Fas expression and Fas-induced apoptosis are mechanisms attributed to the selective destruction of cells of the corpus luteum (CL) during luteal regression. In certain cell-types, sensitivity to these death-inducing mechanisms is due to the loss or cleavage of keratin-containing intermediate filaments. Specifically, keratin 8/18 (K8/K18) filaments are hypothesized to influence cell death in part by regulating Fas expression at the cell surface. Methods Here, Fas expression on bovine luteal cells was quantified by flow cytometry during the early (Day 5, postovulation) and late stages (Days 16–18, postovulation) of CL function, and the relationship between Fas expression, K8/K18 filament expression and cytokine-induced cell death in vitro was evaluated. Results Both total and cell surface expression of Fas on luteal cells was greater for early versus late stage bovine CL (89% vs. 44% of cells for total Fas; 65% vs.18% of cells for cell surface Fas; respectively, P0.05, n=4 CL/stage), despite evidence these conditions increased Fas expression on HepG2 cells (P0.05) or stage of CL (P\u3e0.05, n= 4 CL/stage) on this outcome. Conclusion In conclusion, we rejected our null hypothesis that the cell surface expression of Fas does not differ between luteal cells of early and late stage CL. The results also did not support the idea that K8/K18 filaments influence the expression of Fas on the surface of bovine luteal cells. Potential downstream effects of these filaments on death signaling, however, remain a possibility. Importantly, the elevated expression of Fas observed on cells of early stage bovine CL compared to late stage bovine CL raises a provocative question concerning the physiological role(s) of Fas in the corpus luteum, particularly during early luteal development

Markers for genetic change

Wildlife conservation has focused primarily on species for the last decades. Recently, popular perception and laws have begun to recognize the central importance of genetic diversity in the conservation of biodiversity. How to incorporate genetic diversity in ongoing monitoring and management of wildlife is still an open question.Peer reviewe

Estrous cycle-dependent changes of Fas expression in the bovine corpus luteum: influence of keratin 8/18 intermediate filaments and cytokines

Abstract Background Fas expression and Fas-induced apoptosis are mechanisms attributed to the selective destruction of cells of the corpus luteum (CL) during luteal regression. In certain cell-types, sensitivity to these death-inducing mechanisms is due to the loss or cleavage of keratin-containing intermediate filaments. Specifically, keratin 8/18 (K8/K18) filaments are hypothesized to influence cell death in part by regulating Fas expression at the cell surface. Methods Here, Fas expression on bovine luteal cells was quantified by flow cytometry during the early (Day 5, postovulation) and late stages (Days 16–18, postovulation) of CL function, and the relationship between Fas expression, K8/K18 filament expression and cytokine-induced cell death in vitro was evaluated. Results Both total and cell surface expression of Fas on luteal cells was greater for early versus late stage bovine CL (89% vs. 44% of cells for total Fas; 65% vs.18% of cells for cell surface Fas; respectively, P0.05, n=4 CL/stage), despite evidence these conditions increased Fas expression on HepG2 cells (P0.05) or stage of CL (P>0.05, n= 4 CL/stage) on this outcome. Conclusion In conclusion, we rejected our null hypothesis that the cell surface expression of Fas does not differ between luteal cells of early and late stage CL. The results also did not support the idea that K8/K18 filaments influence the expression of Fas on the surface of bovine luteal cells. Potential downstream effects of these filaments on death signaling, however, remain a possibility. Importantly, the elevated expression of Fas observed on cells of early stage bovine CL compared to late stage bovine CL raises a provocative question concerning the physiological role(s) of Fas in the corpus luteum, particularly during early luteal development.</p

Perceptions of and decision making about clinical trials in adolescent and young adults with Cancer: a qualitative analysis

Abstract Background Adolescent and young adults (AYA) enrolment rates into cancer clinical trials (CCT) are the lowest of any age group globally. As AYA have distinct biological, psychosocial and relational needs, we aimed to explore any unique factors influencing their CCT decision-making process, including AYA-specific perceptions or attitudes towards CCT. Methods Qualitative interpretive descriptive methodology was used to explore AYA perceptions and decision-making related to CCT. An analytic approach conducive to inductive imagining and exploratory questioning was used in order to generate insights and interpret data. Results A total of 21 AYA were interviewed (median age: 31 (18–39)). Twelve (57%) participants had previously been approached to participate in CCT. Major themes influencing trial enrolment decisions were: 1) severity of illness/urgency for new treatment 2) side effect profile of investigational drug in the short and long term (e.g., impact on future quality of life) 3) who approached patient for trial participation (oncologist vs. other) 4) additional information found on-line about the trial and investigators, and 5) family, friends and peer group opinion regarding the CCT. Conclusions Several psychosocial and relational factors were identified as influencing AYA CCT decisions, some of which are unique to this demographic. Specific strategies to address barriers to CCT and enable supportive decision-making include: 1) involving family in decision-making and 2) helping AYA appreciate short- and long-term implications of trial participation. Finally, exploring social networking and general education about CCT that AYA can independently access may increase participation

Perceptions of and decision making about clinical trials in adolescent and young adults with Cancer: a qualitative analysis

Abstract Background Adolescent and young adults (AYA) enrolment rates into cancer clinical trials (CCT) are the lowest of any age group globally. As AYA have distinct biological, psychosocial and relational needs, we aimed to explore any unique factors influencing their CCT decision-making process, including AYA-specific perceptions or attitudes towards CCT. Methods Qualitative interpretive descriptive methodology was used to explore AYA perceptions and decision-making related to CCT. An analytic approach conducive to inductive imagining and exploratory questioning was used in order to generate insights and interpret data. Results A total of 21 AYA were interviewed (median age: 31 (18–39)). Twelve (57%) participants had previously been approached to participate in CCT. Major themes influencing trial enrolment decisions were: 1) severity of illness/urgency for new treatment 2) side effect profile of investigational drug in the short and long term (e.g., impact on future quality of life) 3) who approached patient for trial participation (oncologist vs. other) 4) additional information found on-line about the trial and investigators, and 5) family, friends and peer group opinion regarding the CCT. Conclusions Several psychosocial and relational factors were identified as influencing AYA CCT decisions, some of which are unique to this demographic. Specific strategies to address barriers to CCT and enable supportive decision-making include: 1) involving family in decision-making and 2) helping AYA appreciate short- and long-term implications of trial participation. Finally, exploring social networking and general education about CCT that AYA can independently access may increase participation

Evolutionary diversification of the HAP2 membrane insertion motifs to drive gamete fusion across eukaryotes.

HAPLESS2 (HAP2) is a broadly conserved, gamete-expressed transmembrane protein that was shown recently to be structurally homologous to viral class II fusion proteins, which initiate fusion with host cells via insertion of fusion loops into the host membrane. However, the functional conformation of the HAP2 fusion loops has remained unknown, as the reported X-ray structure of Chlamydomonas reinhardtii HAP2 lacked this critical region. Here, we report a structure-guided alignment that reveals diversification of the proposed HAP2 fusion loops. Representative crystal structures show that in flowering plants, HAP2 has a single prominent fusion loop projecting an amphipathic helix at its apex, while in trypanosomes, three small nonpolar loops of HAP2 are poised to interact with the target membrane. A detailed structure-function analysis of the Arabidopsis HAP2 amphipathic fusion helix defines key residues that are essential for membrane insertion and for gamete fusion. Our study suggests that HAP2 may have evolved multiple modes of membrane insertion to accommodate the diversity of membrane environments it has encountered during eukaryotic evolution

Additional file 1: of Perceptions of and decision making about clinical trials in adolescent and young adults with Cancer: a qualitative analysis

Table S1. Interview guide is presented in the additional file. (DOCX 17 kb

Motivating factors, barriers and facilitators of participation in COVID-19 clinical research: A cross-sectional survey of Canadian community intensive care units.

Only a small proportion of COVID-19 patients in Canada have been recruited into clinical research studies. One reason is that few community intensive care units (ICUs) in Canada participate in research. The objective of this study was to examine the motivating factors, barriers and facilitators to research participation amongst Canadian community ICU stakeholders. A cross-sectional online survey was distributed between May and November 2020. The survey focused on 6 domains: participant demographics, ICU characteristics, ICU research infrastructure, motivating factors, perceived barriers, and perceived facilitators. Responses were received from 73 community ICU stakeholders, representing 18 ICUs. 7/18 ICUs had a clinical research program. Participants rated their interest in pandemic research at a mean of 5.2 (Standard Deviation [SD] = 1.9) on a 7-point Likert scale from 'not interested' to 'very interested'. The strongest motivating factor for research participation was the belief that research improves clinical care and outcomes. The most significant facilitators of research involvement were the availability of an experienced research coordinator and dedicated external funding to cover start-up costs, while the most significant barriers to research involvement were a lack of start-up funding for a research coordinator and a lack of ICU research experience. Canadian Community ICU stakeholders are interested in participating in pandemic research but lack basic infrastructure, research personnel, research experience and start-up funding. Evolution of a research support model at community hospitals, where most patients receive acute care, may increase research participation and improve the generalizability of funded research in Canada