Stretch of contracting cardiac muscle abruptly decreases the rate of phosphate release at high and low calcium

The contractile performance of the heart is linked to the energy that is available to it. Yet, the heart needs to respond quickly to changing demands. During diastole, the heart fills with blood and the heart chambers expand. Upon activation, contraction of cardiac muscle expels blood into the circulation. Early in systole, parts of the left ventricle are being stretched by incoming blood, before contraction causes shrinking of the ventricle. We explore here the effect of stretch of contracting permeabilized cardiac trabeculae of the rat on the rate of inorganic phosphate (P(i)) release resulting from ATP hydrolysis, using a fluorescent sensor for P(i) with millisecond time resolution. Stretch immediately reduces the rate of P(i) release, an effect observed both at full calcium activation (32 μmol/liter of Ca(2+)), and at a physiological activation level of 1 μmol/liter of Ca(2+). The results suggest that stretch redistributes the actomyosin cross-bridges toward their P(i)-containing state. The redistribution means that a greater fraction of cross-bridges will be poised to rapidly produce a force-generating transition and movement, compared with cross-bridges that have not been subjected to stretch. At the same time stretch modifies the P(i) balance in the cytoplasm, which may act as a cytoplasmic signal for energy turnover.     

Priming the engine of DNA synthesis

In this issue of Structure, Rymer and colleagues present the first crystal structures of a bacterial DnaG primase with bound substrate NTPs and alarmone inhibitors. A thoughtful comparative structural analysis provides important insights into the chemical mechanism of primase.     

Lactoferrin activates macrophages via TLR4-dependent and -independent signaling pathways

Lactoferrin (LF) is a component of innate immunity and is known to interact with accessory molecules involved in the TLR4 pathway, including CD14 and LPS binding protein, suggesting that LF may activate components of the TLR4 pathway. In the present study, we have asked whether bovine LF (bLF)-induced macrophage activation is TLR4-dependent. Both bLF and LPS stimulated IL-6 production and CD40 expression in RAW 264.7 macrophages and in BALB/cJ peritoneal exudate macrophages. However, in macrophages from congenic TLR4(-/-) C.C3-Tlr4(lps-d) mice, CD40 was not expressed while IL-6 secretion was increased relative to wild-type cells. The signaling components NF-kappaB, p38, ERK and JNK were activated in RAW 264.7 cells and BALB/cJ macrophages after bLF or LPS stimulation, demonstrating that the TLR4-dependent bLF activation pathway utilizes signaling components common to LPS activation. In TLR4 deficient macrophages, bLF-induced activation of NF-kappaB, p38, ERK and JNK whereas LPS-induced cell signaling was absent. We conclude from these studies that bLF induces limited and defined macrophage activation and cell signaling events via TLR4-dependent and -independent mechanisms. bLF-induced CD40 expression was TLR4-dependent whereas bLF-induced IL-6 secretion was TLR4-independent, indicating potentially separate pathways for bLF mediated macrophage activation events in innate immunity.     

Predicting the regenerative capacity of conifer somatic embryogenic cultures by metabolomics

Somatic embryogenesis in gymnosperms is an effective approach to clonally propagating germplasm. However, embryogenic cultures frequently lose regenerative capacity. The interactions between metabolic composition, physiological state, genotype and embryogenic capacity in Pinus taeda (loblolly pine) somatic embryogenic cultures were explored using metabolomics. A stepwise modelling procedure, using the Bayesian information criterion, generated a 47 metabolite predictive model that could explain culture productivity. The model performed extremely well in cross-validation, achieving a correlation coefficient of 0.98 between actual and predicted mature embryo production. The metabolic composition and structure of the model implied that variation in culture regenerative capacity was closely linked to the physiological transition of cultures from the proliferation phase to the maturation phase of development. The propensity of cultures to advance into this transition appears to relate to nutrient uptake and allocation in vivo , and to be associated with the tolerance and response of cultures to stress, during the proliferation phase.     

Non-Human Primate Model of Kaposi's Sarcoma-Associated Herpesvirus Infection

Since Kaposi''s sarcoma-associated herpesvirus (KSHV or human herpesvirus 8) was first identified in Kaposi''s sarcoma (KS) lesions of HIV-infected individuals with AIDS, the basic biological understanding of KSHV has progressed remarkably. However, the absence of a proper animal model for KSHV continues to impede direct in vivo studies of viral replication, persistence, and pathogenesis. In response to this need for an animal model of KSHV infection, we have explored whether common marmosets can be experimentally infected with human KSHV. Here, we report the successful zoonotic transmission of KSHV into common marmosets (Callithrix jacchus, Cj), a New World primate. Marmosets infected with recombinant KSHV rapidly seroconverted and maintained a vigorous anti-KSHV antibody response. KSHV DNA and latent nuclear antigen (LANA) were readily detected in the peripheral blood mononuclear cells (PBMCs) and various tissues of infected marmosets. Remarkably, one orally infected marmoset developed a KS-like skin lesion with the characteristic infiltration of leukocytes by spindle cells positive for KSHV DNA and proteins. These results demonstrate that human KSHV infects common marmosets, establishes an efficient persistent infection, and occasionally leads to a KS-like skin lesion. This is the first animal model to significantly elaborate the important aspects of KSHV infection in humans and will aid in the future design of vaccines against KSHV and anti-viral therapies targeting KSHV coinfected tumor cells.     

Information from pharmaceutical companies and the quality, quantity, and cost of physicians' prescribing: a systematic review

Background

Pharmaceutical companies spent $57.5 billion on pharmaceutical promotion in the United States in 2004. The industry claims that promotion provides scientific and educational information to physicians. While some evidence indicates that promotion may adversely influence prescribing, physicians hold a wide range of views about pharmaceutical promotion. The objective of this review is to examine the relationship between exposure to information from pharmaceutical companies and the quality, quantity, and cost of physicians'' prescribing.

Methods and Findings

We searched for studies of physicians with prescribing rights who were exposed to information from pharmaceutical companies (promotional or otherwise). Exposures included pharmaceutical sales representative visits, journal advertisements, attendance at pharmaceutical sponsored meetings, mailed information, prescribing software, and participation in sponsored clinical trials. The outcomes measured were quality, quantity, and cost of physicians'' prescribing. We searched Medline (1966 to February 2008), International Pharmaceutical Abstracts (1970 to February 2008), Embase (1997 to February 2008), Current Contents (2001 to 2008), and Central (The Cochrane Library Issue 3, 2007) using the search terms developed with an expert librarian. Additionally, we reviewed reference lists and contacted experts and pharmaceutical companies for information. Randomized and observational studies evaluating information from pharmaceutical companies and measures of physicians'' prescribing were independently appraised for methodological quality by two authors. Studies were excluded where insufficient study information precluded appraisal. The full text of 255 articles was retrieved from electronic databases (7,185 studies) and other sources (138 studies). Articles were then excluded because they did not fulfil inclusion criteria (179) or quality appraisal criteria (18), leaving 58 included studies with 87 distinct analyses. Data were extracted independently by two authors and a narrative synthesis performed following the MOOSE guidelines. Of the set of studies examining prescribing quality outcomes, five found associations between exposure to pharmaceutical company information and lower quality prescribing, four did not detect an association, and one found associations with lower and higher quality prescribing. 38 included studies found associations between exposure and higher frequency of prescribing and 13 did not detect an association. Five included studies found evidence for association with higher costs, four found no association, and one found an association with lower costs. The narrative synthesis finding of variable results was supported by a meta-analysis of studies of prescribing frequency that found significant heterogeneity. The observational nature of most included studies is the main limitation of this review.

Conclusions

With rare exceptions, studies of exposure to information provided directly by pharmaceutical companies have found associations with higher prescribing frequency, higher costs, or lower prescribing quality or have not found significant associations. We did not find evidence of net improvements in prescribing, but the available literature does not exclude the possibility that prescribing may sometimes be improved. Still, we recommend that practitioners follow the precautionary principle and thus avoid exposure to information from pharmaceutical companies. Please see later in the article for the Editors'' Summary     

Syk activation initiates downstream signaling events during human polymorphonuclear leukocyte phagocytosis

We investigated the requirement for Syk activation to initiate downstream signaling events during polymorphonuclear leukocyte (PMN) phagocytosis of Ab-coated erythrocytes (EIgG). When PMN were challenged with EIgG, Syk phosphorylation increased in a time-dependent manner, paralleling the response of PMN phagocytosis. Pretreatment of PMN with piceatannol, a Syk-selective inhibitor, blocked EIgG phagocytosis and Syk phosphorylation. We found that piceatannol inhibited protein kinase Cdelta (PKCdelta) and Raf-1 translocation from cytosol to plasma membrane by >90%. Extracellular signal-regulated protein kinase-1 and -2 (ERK1 and ERK2) phosphorylation was similarly blocked. We also investigated phosphatidylinositide 3-kinase (PI 3-kinase) activity and Syk phosphorylation using piceatannol, wortmannin, and LY294002, inhibitors of PI 3-kinase. The phosphorylation of Syk preceded the activation of PI 3-kinase. Both wortmannin and piceatannol inhibited PI 3-kinase, but only piceatannol inhibited Syk. In contrast to piceatannol, wortmannin did not inhibit PKCdelta and Raf-1 translocation. To elucidate signaling downstream of Syk activation, we assessed whether the cell-permeable diacylglycerol analogue didecanoylglycerol could normalize PMN phagocytosis, PKCdelta and Raf-1 translocation, and ERK1 and ERK2 phosphorylation inhibited by piceatannol. The addition of didecanoylglycerol to the Syk-inhibited phagocytosing PMN normalized all three without a concomitant effect on PI 3-kinase activity and Syk phosphorylation. We conclude that Syk activation following Fcgamma receptor engagement initiates downstream signaling events leading to mitogen-activated protein kinase activation independent of PI 3-kinase activation.