Progesterone Receptor and Prostaglandins Mediate Luteinizing Hormone-Induced Changes in Messenger RNAs for ADAMTS Proteases in Theca Cells of Bovine Periovulatory Follicles

Little is known about the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family of extracellular proteases in ovarian follicles of non‐rodent species, particularly in theca cells. In the present study, temporal changes in the abundance of mRNA encoding four ADAMTS subtypes and hormonal regulation of mRNA encoding two subtypes were investigated in theca interna cells during the periovulatory period in cattle. Gonadotropin‐releasing hormone (GnRH) was injected into animals to induce a luteinizing hormone (LH)/follicle‐stimulating hormone (FSH) surge, and follicles were obtained at 0 hr post‐GnRH (preovulatory) or at 6, 12, 18, or 24 hr (periovulatory). ADAMTS1, ‐2, ‐7, and ‐9 transcript abundance was then determined in the isolated theca interna. ADAMTS1 and ‐9 mRNA levels were up‐regulated at 24 hr post‐GnRH, whereas ADAMTS2 mRNA was higher at 12–24 hr post‐GnRH and ADAMTS7 mRNA increased transiently at 12 hr post‐GnRH compared to other time points. Subsequent in vitro experiments using preovulatory theca interna (0 hr post‐GnRH) showed that application of LH in vitro can mimic the effects of the gonadotropin surge on mRNAs encoding ADAMTS1 and ‐9 and that progesterone/progesterone receptor and/or prostaglandins may regulate the levels of mRNA encoding ADAMTS1 and ‐9 in theca interna, downstream of the LH surge. Time‐ and subtype‐specific changes in ADAMTS mRNA abundance in vivo, and their regulation in vitro by hormones, indicate that ADAMTS family members produced by theca cells may play important roles in follicle rupture and the accompanying tissue remodeling in cattle

Increasing doses of fiber do not influence short-term satiety or food intake and are inconsistently linked to gut hormone levels

Background: People who eat more fiber often have a lower body weight than people who eat less fiber. The mechanism for this relationship has been explained, in part, by increased satiety, which may occur as a result of changes in appetite-suppressing gut hormone levels, and decreases in food intake at subsequent meals. Objective: We hypothesized that increasing doses of mixed fiber, consumed in muffins for breakfast, would proportionally influence satiety, gut hormone levels, and subsequent food intake. Design: This was a randomized, double-blind, crossover study. Healthy men (n=10) and women (n=10) with a BMI of 24±2 (mean±SEM) participated in this study. Fasting subjects consumed a muffin with 0, 4, 8, or 12 g of mixed fibers and approximately 500 kcal. Visual analog scales rated hunger and satiety for 3 h; blood was drawn to measure ghrelin, glucagon-like peptide-1 (GLP-1), and peptide YY3–36 (PYY3–36) at various intervals; and food intake was measured at an ad libitum lunch. Results: Responses to satiety-related questions did not differ among treatments. However, despite lack of differences in satiety, gut hormone levels differed among treatments. Ghrelin was higher after the 12 g fiber dose than after the 4 and 8 g fiber doses. GLP-1 was higher after the 0 g fiber dose than after the 12 and 4 g fiber doses, and PYY3–36 did not differ among fiber doses. Food intake was also indistinguishable among doses. Conclusion: Satiety, gut hormone response, and food intake did not change in a dose-dependent manner after subjects consumed 0, 4, 8, and 12 g of mixed fiber in muffins for breakfast

A Natural Plasmid Uniquely Encodes Two Biosynthetic Pathways Creating a Potent Anti-MRSA Antibiotic

Background Understanding how complex antibiotics are synthesised by their producer bacteria is essential for creation of new families of bioactive compounds. Thiomarinols, produced by marine bacteria belonging to the genus Pseudoalteromonas, are hybrids of two independently active species: the pseudomonic acid mixture, mupirocin, which is used clinically against MRSA, and the pyrrothine core of holomycin. Methodology/Principal Findings High throughput DNA sequencing of the complete genome of the producer bacterium revealed a novel 97 kb plasmid, pTML1, consisting almost entirely of two distinct gene clusters. Targeted gene knockouts confirmed the role of these clusters in biosynthesis of the two separate components, pseudomonic acid and the pyrrothine, and identified a putative amide synthetase that joins them together. Feeding mupirocin to a mutant unable to make the endogenous pseudomonic acid created a novel hybrid with the pyrrothine via “mutasynthesis” that allows inhibition of mupirocin-resistant isoleucyl-tRNA synthetase, the mupirocin target. A mutant defective in pyrrothine biosynthesis was also able to incorporate alternative amine substrates. Conclusions/Significance Plasmid pTML1 provides a paradigm for combining independent antibiotic biosynthetic pathways or using mutasynthesis to develop a new family of hybrid derivatives that may extend the effective use of mupirocin against MRSA

The challenges of changing national malaria drug policy to artemisinin-based combinations in Kenya

Backgound: Sulphadoxine/sulphalene-pyrimethamine (SP) was adopted in Kenya as first line therapeutic for uncomplicated malaria in 1998. By the second half of 2003, there was convincing evidence that SP was failing and had to be replaced. Despite several descriptive investigations of policy change and implementation when countries moved from chloroquine to SP, the different constraints of moving to artemisinin-based combination therapy (ACT) in Africa are less well documented. Methods: A narrative description of the process of anti-malarial drug policy change, financing and implementation in Kenya is assembled from discussions with stakeholders, reports, newspaper articles, minutes of meetings and email correspondence between actors in the policy change process. The narrative has been structured to capture the timing of events, the difficulties and hurdles faced and the resolutions reached to the final implementation of a new treatment policy. Results: Following a recognition that SP was failing there was a rapid technical appraisal of available data and replacement options resulting in a decision to adopt artemether-lumefantrine (AL) as the recommended first-line therapy in Kenya, announced in April 2004. Funding requirements were approved by the Global Fund to Fight AIDS, Tuberculosis and Malaria (GFATM) and over 60 million US$ were agreed in principle in July 2004 to procure AL and implement the policy change. AL arrived in Kenya in May 2006, distribution to health facilities began in July 2006 coincidental with cascade in-service training in the revised national guidelines. Both training and drug distribution were almost complete by the end of 2006. The article examines why it took over 32 months from announcing a drug policy change to completing early implementation. Reasons included: lack of clarity on sustainable financing of an expensive therapeutic for a common disease, a delay in release of funding, a lack of comparative efficacy data between AL and amodiaquine-based alternatives, a poor dialogue with pharmaceutical companies with a national interest in antimalarial drug supply versus the single sourcing of AL and complex drug ordering, tendering and procurement procedures. Conclusion: Decisions to abandon failing monotherapy in favour of ACT for the treatment of malaria can be achieved relatively quickly. Future policy changes in Africa should be carefully prepared for a myriad of financial, political and legislative issues that might limit the rapid translation of drug policy change into action

Engineered transient and stable overexpression of translation factors eIF3i and eIF3c in CHOK1 and HEK293 cells gives enhanced cell growth associated with increased c-Myc expression and increased recombinant protein synthesis

There is a desire to engineer mammalian host cell lines to improve cell growth/biomass accumulation and recombinant biopharmaceutical protein production in industrially relevant cell lines such as the CHOK1 and HEK293 cell lines. The over-expression of individual subunits of the eukaryotic translation factor eIF3 in mammalian cells has previously been shown to result in oncogenic properties being imparted on cells, including increased cell proliferation and growth and enhanced global protein synthesis rates. Here we report on the engineering of CHOK1 and HEK cells to over-express the eIF3i and eIF3c subunits of the eIF3 complex and the resultant impact on cell growth and a reporter of exogenous recombinant protein production. Transient over-expression of eIF3i in HEK293 and CHOK1 cells resulted in a modest increase in total eIF3i amounts (maximum 40% increase above control) and an approximate 10% increase in global protein synthesis rates in CHOK1 cells. Stable over-expression of eIF3i in CHOK1 cells was not achievable, most likely due to the already high levels of eIF3i in CHO cells compared to HEK293 cells, but was achieved in HEK293 cells. HEK293 cells engineered to over-express eIF3i had faster growth that was associated with increased c-Myc expression, achieved higher cell biomass and gave enhanced yields of a reporter of recombinant protein production. Whilst CHOK1 cells could not be engineered to over-express eIF3i directly, they could be engineered to over-express eIF3c, which resulted in a subsequent increase in eIF3i amounts and c-Myc expression. The CHOK1 eIF3c engineered cells grew to higher cell numbers and had enhanced cap- and IRES-dependent recombinant protein synthesis. Collectively these data show that engineering of subunits of the eIF3 complex can enhance cell growth and recombinant protein synthesis in mammalian cells in a cell specific manner that has implications for the engineering or selection of fast growing or high producing cells for production of recombinant proteins

A sociological exploration of the tensions related to interprofessional collaboration in acute-care discharge planning

Patient discharge is a key concern in hospitals, particularly in acute care, given the multifaceted and challenging nature of patients' healthcare needs. Policies on discharge have identified the importance of interprofessional collaboration, yet research has described its limitations in this clinical context. This study aimed to extend our understanding of interprofessional interactions related to discharge in a general internal medicine setting by using sociological theories to illuminate the existence of, and interplay between, structural factors and microlevel practices. An ethnographic approach was employed to obtain an in-depth insight into healthcare providers' perspectives, behaviours, and interactions regarding discharge. Data collection involved observations, interviews, and document analysis. Approximately 65 hours of observations were undertaken, 23 interviews were conducted with healthcare providers, and government and hospital discharge documents were collected. Data were analysed using a directed content approach. The findings indicate the existence of a medically dominated division of healthcare labour in patient discharge with opportunities for some interprofessional negotiations; the role of organizational routines in facilitating and challenging interprofessional negotiations in patient discharge; and tensions in organizational priorities that impact an interprofessional approach to discharge. The findings provide insight into the various levels at which interventions can be targeted to improve interprofessional collaboration in discharge while recognizing the organizational tensions that challenge an interprofessional approach

Cooling-induced SUMOylation of EXOSC10 down-regulates ribosome biogenesis.

The RNA exosome is essential for 3? processing of functional RNA species and degradation of aberrant RNAs in eukaryotic cells. Recent reports have defined the substrates of the exosome catalytic domains and solved the multimeric structure of the exosome complex. However, regulation of exosome activity remains poorly characterized, especially in response to physiological stress. Following the observation that cooling of mammalian cells results in a reduction in 40S:60S ribosomal subunit ratio, we uncover regulation of the nuclear exosome as a result of reduced temperature. Using human cells and an in vivo model system allowing whole-body cooling, we observe reduced EXOSC10 (hRrp6, Pm/Scl-100) expression in the cold. In parallel, both models of cooling increase global SUMOylation, leading to the identification of specific conjugation of SUMO1 to EXOSC10, a process that is increased by cooling. Furthermore, we define the major SUMOylation sites in EXOSC10 by mutagenesis and show that overexpression of SUMO1 alone is sufficient to suppress EXOSC10 abundance. Reducing EXOSC10 expression by RNAi in human cells correlates with the 3? preribosomal RNA processing defects seen in the cold as well as reducing the 40S:60S ratio, a previously uncharacterized consequence of EXOSC10 suppression. Together, this work illustrates that EXOSC10 can be modified by SUMOylation and identifies a physiological stress where this regulation is prevalent both in vitro and in vivo