Nitric oxide binds to the proximal heme coordination site of the ferrocytochrome c/cardiolipin complex: formation mechanism and dynamics.

Mammalian mitochondrial cytochrome c interacts with cardiolipin to form a complex (cyt. c/CL) important in apoptosis. Here we show that this interaction leads to structural changes in ferrocytochrome c that leads to an open coordinate site on the central iron, resulting from the dissociation of the intrinsic methionine residue, where NO can rapidly bind (k = 1.2 x 10(7) m(-1) s(-1)). Accompanying NO binding, the proximal histidine dissociates leaving the heme pentacoordinate, in contrast to the hexacoordinate nitrosyl adducts of native ferrocytochrome c or of the protein in which the coordinating methionine is removed by chemical modification or mutation. We present the results of stopped-flow and photolysis experiments that show that following initial NO binding to the heme, there ensues an unusually complex set of kinetic steps. The spectral changes associated with these kinetic transitions, together with their dependence on NO concentration, have been determined and lead us to conclude that NO binding to cyt. c/CL takes place via an overall scheme comparable to that described for cytochrome c' and guanylate cyclase, the final product being one in which NO resides on the proximal side of the heme. In addition, novel features not observed before in other heme proteins forming pentacoordinate nitrosyl species, include a high yield of NO escape after dissociation, rapid (<1 ms) dissociation of proximal histidine upon NO binding and its very fast binding (60 ps) after NO dissociation, and the formation of a hexacoordinate intermediate. These features all point at a remarkable mobility of the proximal heme environment induced by cardiolipin

Ultrafast photochemistry of the bc₁ complex

We present a full investigation of ultrafast light-induced events in the membraneous cytochrome bc 1 complex by transient absorption spectroscopy. This energy-transducing complex harbors four redox-active components per monomer: heme c 1 , two 6-coordinate b-hemes and a [2Fe-2S] cluster. Using excitation of these components in different ratios under various excitation conditions, probing in the full visible range and under three well-defined redox conditions, we demonstrate that for all ferrous hemes of the complex photodissociation of axial ligands takes place and that they rebind in 5-7 ps, as in other 6-coordinate heme proteins, including cytoglobin, which is included as a reference in this study. By contrast, the signals are not consistent with photooxidation of the b hemes. This conclusion contrasts with a recent assessment based on a more limited data set. The binding kinetics of internal and external ligands are indicative of a rigid heme environment, consistent with the electron transfer function. We also report, for the first time, photoactivity of the very weakly absorbing iron-sulfur center. This yields the unexpected perspective of studying photochemistry, initiated by excitation of iron-sulfur clusters, in a range of protein complexes

Do not attempt resuscitation decisions in a cancer centre: addressing difficult ethical and communication issues

Talking to patients about ‘Do Not Attempt Resuscitation’ decisions is difficult for many doctors. Communication about ‘Do Not Attempt Resuscitation’ decisions should occur as part of a wider discussion of treatment goals at an earlier stage in the patient's illness. A doctor should not initiate any treatment, including cardio-pulmonary resuscitation if he/she does not believe it will benefit the patient. An ethical framework is offered which may be of practical help in clarifying decision-making

The native architecture of a photosynthetic membrane

In photosynthesis, the harvesting of solar energy and its subsequent conversion into a stable charge separation are dependent upon an interconnected macromolecular network of membrane-associated chlorophyll–protein complexes. Although the detailed structure of each complex has been determined, the size and organization of this network are unknown. Here we show the use of atomic force microscopy to directly reveal a native bacterial photosynthetic membrane. This first view of any multi-component membrane shows the relative positions and associations of the photosynthetic complexes and reveals crucial new features of the organization of the network: we found that the membrane is divided into specialized domains each with a different network organization and in which one type of complex predominates. Two types of organization were found for the peripheral light-harvesting LH2 complex. In the first, groups of 10–20 molecules of LH2 form light-capture domains that interconnect linear arrays of dimers of core reaction centre (RC)–light-harvesting 1 (RC–LH1–PufX) complexes; in the second they were found outside these arrays in larger clusters. The LH1 complex is ideally positioned to function as an energy collection hub, temporarily storing it before transfer to the RC where photochemistry occurs: the elegant economy of the photosynthetic membrane is demonstrated by the close packing of these linear arrays, which are often only separated by narrow 'energy conduits' of LH2 just two or three complexes wide

Terahertz underdamped vibrational motion governs protein-ligand binding in solution

Low-frequency collective vibrational modes in proteins have been proposed as being responsible for efficiently directing biochemical reactions and biological energy transport. However, evidence of the existence of delocalized vibrational modes is scarce and proof of their involvement in biological function absent. Here we apply extremely sensitive femtosecond optical Kerr-effect spectroscopy to study the depolarized Raman spectra of lysozyme and its complex with the inhibitor triacetylchitotriose in solution. Underdamped delocalized vibrational modes in the terahertz frequency domain are identified and shown to blue-shift and strengthen upon inhibitor binding. This demonstrates that the ligand-binding coordinate in proteins is underdamped and not simply solvent-controlled as previously assumed. The presence of such underdamped delocalized modes in proteins may have significant implications for the understanding of the efficiency of ligand binding and protein–molecule interactions, and has wider implications for biochemical reactivity and biological function

HGF Mediates the Anti-inflammatory Effects of PRP on Injured Tendons

Platelet-rich plasma (PRP) containing hepatocyte growth factor (HGF) and other growth factors are widely used in orthopaedic/sports medicine to repair injured tendons. While PRP treatment is reported to decrease pain in patients with tendon injury, the mechanism of this effect is not clear. Tendon pain is often associated with tendon inflammation, and HGF is known to protect tissues from inflammatory damages. Therefore, we hypothesized that HGF in PRP causes the anti-inflammatory effects. To test this hypothesis, we performed in vitro experiments on rabbit tendon cells and in vivo experiments on a mouse Achilles tendon injury model. We found that addition of PRP or HGF decreased gene expression of COX-1, COX-2, and mPGES-1, induced by the treatment of tendon cells in vitro with IL-1β. Further, the treatment of tendon cell cultures with HGF antibodies reduced the suppressive effects of PRP or HGF on IL-1β-induced COX-1, COX-2, and mPGES-1 gene expressions. Treatment with PRP or HGF almost completely blocked the cellular production of PGE2 and the expression of COX proteins. Finally, injection of PRP or HGF into wounded mouse Achilles tendons in vivo decreased PGE2 production in the tendinous tissues. Injection of platelet-poor plasma (PPP) however, did not reduce PGE2 levels in the wounded tendons, but the injection of HGF antibody inhibited the effects of PRP and HGF. Further, injection of PRP or HGF also decreased COX-1 and COX-2 proteins. These results indicate that PRP exerts anti-inflammatory effects on injured tendons through HGF. This study provides basic scientific evidence to support the use of PRP to treat injured tendons because PRP can reduce inflammation and thereby reduce the associated pain caused by high levels of PGE2. © 2013 Zhang et al

Epidemiological trends in nosocomial candidemia in intensive care

BACKGROUND: Infection represents a frequent complication among patients in Intensive Care Units (ICUs) and mortality is high. In particular, the incidence of fungal infections, especially due to Candida spp., has been increasing during the last years. METHODS: In a retrospective study we studied the etiology of candidemia in critically ill patients over a five-year period (1999–2003) in the ICU of the San Martino University Hospital in Genoa, Italy. RESULTS: In total, 182 episodes of candidaemia were identified, with an average incidence of 2.22 episodes/10 000 patient-days/year (range 1.25–3.06 episodes). Incidence of candidemia increased during the study period from 1.25 in 1999 to 3.06/10 000 patient-days/year in 2003. Overall, 40% of the fungemia episodes (74/182) were due to C.albicans, followed by C. parapsilosis(23%), C.glabrata (15%), C.tropicalis (9%) and other species (13%). Candidemia due to non-albicans species increased and this was apparently correlated with an increasing use of azoles for prophylaxis or empirical treatment. CONCLUSION: The study demonstrates a shift in the species of Candida causing fungemia in a medical and surgical ICU population during a 5 year period. The knowledge of the local epidemiological trends in Candida species isolated in blood cultures is important to guide therapeutic choices

Ectopic opening of the common bile duct and duodenal stenosis: an overlooked association

<p>Abstract</p> <p>Background</p> <p>Ectopic opening of the common bile duct into the duodenal bulb (EO-CBD-DB) is a rare disease that may be complicated by duodenal ulcer, deformity, stenosis and biliary stones. The aim of this study is to report clinical presentations, endoscopic diagnosis and treatment of this entity as well as to investigate its association with duodenal stenosis.</p> <p>Methods</p> <p>Gastroduodenoscopic findings and radiological imaging were evaluated for ectopic papilla and duodenal stenosis. Diagnostic methods, endoscopic procedures and long-term outcomes of the endoscopic treatment were presented.</p> <p>Results</p> <p>EO-CBD-DB was found in 74 (77.1%) of the 96 patients with duodenal deformity/stenosis (79 male, 17 female, mean age: 58.5, range: 30-87 years). The papilla with normal appearance was retracted to the bulb in 11 while it was at its usual location in the remaining 11. The history of biliodigestive surgery was more common in patients with EO-CBD-DB who were frequently presented with the common bile duct stone-related symptoms than the other patients. Thirteen (17.6%) of the patients with EO-CBD-DB were referred to surgery. Endoscopic treatment was completed in 60 (81.1%) patients after an average of 1.7 (range: 1-6) procedures. These patients were on follow-up for 24.8 (range: 2-46) months. Endoscopic intervention was required in 12 (20%) of them because of recurrent biliary problems. Treatment of the patient who had stricture due to biliary injury during laparoscopic cholecystectomy is still continued.</p> <p>Conclusions</p> <p>The presence of EO-CBD-DB should be considered particularly in middle-aged male patients who have duodenal deformity/stenosis. Endoscopic treatment is feasible in these patients. The long-term outcomes of endoscopic therapy need to be compared with surgical treatment.</p

Ethnic diversity outpatient clinic in paediatrics

<p>Abstract</p> <p>Background</p> <p>The health status of chronic sick ethnic minority children in the Netherlands is unequal compared with indigenous Dutch children. In order to optimize the health care for these children a specific patient-oriented clinic in ethnic-cultural diversity: the Mosaic Outpatient Clinic (MOC) was integrated in the general Paediatric Outpatient Departments (POPD) of three hospitals in Amsterdam.</p> <p>Methods</p> <p>Feasibility of the MOC, factors influencing the health care process and encountered bottlenecks in health care were studied in ethnic minority children with asthma, diabetes type 1 or metabolic disease originating from Morocco, Turkey and Surinam. Feasibility was determined by the number of patients attended, support from the paediatric medical staff and willingness of the patients to participate. Influences on the health care process comprised parents' level of knowledge of disease, sense of disease severity, level of effort, linguistic skills, health literacy, adherence to treatment and encountered bottlenecks in the health care process. Moreover, the number of admissions and visits to the POPD in the years before, during and after the MOC were analysed.</p> <p>Results</p> <p>In 2006 a total of 189 ethnic minority children were seen. Integration of the MOC within the general POPD of the hospital is feasible. The ability of the parents to speak and understand Dutch was found to be 58%, functional health literacy was 88%; sufficient knowledge of disease and sense of disease severity were 59% and 67%, respectively.</p> <p>The main bottlenecks in the healthcare process: poor knowledge of disease, limited sense of disease severity and low health literacy in the parents proved to be the best predictors for decreased adherence. After attending the MOC there was a decrease in the number of admissions and visits to the POPD for asthma while the number of visits increased in patients with diabetes and the amount of no-shows decreased in patients with a metabolic disease.</p> <p>Conclusion</p> <p>Integration of a MOC in the general POPD is feasible and appreciated by the parents, provides more insight in the problems ethnic minority children and their parents face and shows promising directions for optimizing adherence in these children.</p