Probing a Complex of Cytochromecand Cardiolipin by Magnetic Circular Dichroism Spectroscopy: Implications for the Initial Events in Apoptosis

Oxidation of cardiolipin (CL) by its complex with cytochrome c (cyt c) plays a crucial role in triggering apoptosis. Through a combination of magnetic circular dichroism spectroscopy and potentiometric titrations, we show that both the ferric and ferrous forms of the heme group of a CL:cyt c complex exist as multiple conformers at a physiologically relevant pH of 7.4. For the ferric state, these conformers are His/Lys- and His/OH–-ligated. The ferrous state is predominantly high-spin and, most likely, His/–. Interconversion of the ferric and ferrous conformers is described by a single midpoint potential of -80 ± 9 mV vs SHE. These results suggest that CL oxidation in mitochondria could occur by the reaction of molecular oxygen with the ferrous CL:cyt c complex in addition to the well-described reaction of peroxides with the ferric form

How can we optimise inhaled beta2 agonist dose as ‘reliever’ medicine for wheezy pre-school children? Study protocol for a randomised controlled trial

Background: Asthma is a common problem in children and, if inadequately controlled, may seriously diminish their quality of life. Inhaled short-acting beta2 agonists such as salbutamol are usually prescribed as ‘reliever’ medication to help control day-to-day symptoms such as wheeze. As with many medications currently prescribed for younger children (defined as those aged 2 years 6 months to 6 years 11 months), there has been no pre-licensing age-specific pharmacological testing; consequently, the doses currently prescribed (200–1000 μg) may be ineffective or likely to induce unnecessary side effects. We plan to use the interrupter technique to measure airway resistance in this age group, allowing us for the first time to correlate inhaled salbutamol dose with changes in clinical response. We will measure urinary salbutamol levels 30 min after dosing as an estimate of salbutamol doses in the lungs, and also look for genetic polymorphisms linked to poor responses to inhaled salbutamol. Methods: This is a phase IV, randomised, controlled, observer-blinded, single-centre trial with four parallel groups (based on a sparse sampling approach) and a primary endpoint of the immediate bronchodilator response to salbutamol so that we can determine the most appropriate dose for an individual younger child. Simple randomisation will be used with a 1:1:1:1 allocation. Discussion: The proposed research will exploit simple, non-invasive and inexpensive tests that can mostly be performed in an outpatient setting in order to help develop the evidence for the correct dose of salbutamol in younger children with recurrent wheeze who have been prescribed salbutamol by their doctor. Trial registration: EudraCT2014-001978-33, ISRCTN15513131. Registered on 8 April 2015

T-Lymphocytes Enable Osteoblast Maturation via IL-17F during the Early Phase of Fracture Repair

While it is well known that the presence of lymphocytes and cytokines are important for fracture healing, the exact role of the various cytokines expressed by cells of the immune system on osteoblast biology remains unclear. To study the role of inflammatory cytokines in fracture repair, we studied tibial bone healing in wild-type and Rag1−/− mice. Histological analysis, µCT stereology, biomechanical testing, calcein staining and quantitative RNA gene expression studies were performed on healing tibial fractures. These data provide support for Rag1−/− mice as a model of impaired fracture healing compared to wild-type. Moreover, the pro-inflammatory cytokine, IL-17F, was found to be a key mediator in the cellular response of the immune system in osteogenesis. In vitro studies showed that IL-17F alone stimulated osteoblast maturation. We propose a model in which the Th17 subset of T-lymphocytes produces IL-17F to stimulate bone healing. This is a pivotal link in advancing our current understanding of the molecular and cellular basis of fracture healing, which in turn may aid in optimizing fracture management and in the treatment of impaired bone healing