Opportunities to engage health system leaders in whole systems approaches to physical activity in England.

BACKGROUND: Physical activity plays an important role in maintaining good health and wellbeing, non-communicable disease prevention and can improve healthcare outcomes. Some progress is being made on incorporating physical activity into routine care, but less on engaging health system leaders in the 'whole systems' approaches which are increasingly recognised as important for addressing complex public health challenges such as physical inactivity. This commentary builds upon the findings of a recent study and aims to identify opportunities for engaging National Health Service (NHS) systems leaders in whole systems approaches to physical activity. OPPORTUNITIES FOR ACTION IN ENGLAND: Pockets of good practice exist from which lessons can be learned, but there are systemic issues that discourage and create barriers, and a need for meaningful engagement, leadership and action at national, regional and local levels. National and regional actors like Sport England, NHS England, health professional bodies, Active Partnerships, the Local Government Association and the Office for Health Improvement and Disparities can encourage and support government and the NHS to change policy drivers, culture and practices. Emerging opportunities include the 2021 White Paper Integration and Innovation, development of local integrated care systems, leadership from health charities and investment in non-clinical interventions ('social prescribing'). At local level, public health and physical activity specialists and other organisations have a key role as champions and facilitators of local whole systems approaches and engagement of local NHS leaderships. Finally, although whole systems action is about collaborative leadership, individual champions of physical activity can make a difference in influencing NHS leaders at every level towards whole systems working.

Practices of the best companies in the medical industry.

A report containing information about three of the top medical manufacturing companies in the world--Baxter Healthcare Corporation, Johnson & Johnson and Medtronic, Inc. Resources have been gathered and employees in key positions from each company have been interviewed to obtain the most current and accurate information. Interviewees were asked what factors contribute to their company's success. These factors were then explored more explicitly through a variety of publications, and the results are included in this report. In today's competitive market, companies must keep current with the market trends and be flexible and open to changing and/or improving their practices in order to remain successful. It has been found that although the main focuses of each company may be similar, the processes by which they strive to meet their goals are quite unique.

An autoantigenic T cell epitope forms unstable complexes with class II MHC: a novel route for escape from tolerance induction.

The peptide rAc1-11 represents the dominant T cell epitope of rat myelin basic protein (MBP) in mice of the H-2u haplotype. Residue 4 has been shown previously to govern binding of the peptide to the class II molecule, I-Au. We have constructed peptide analogues bearing amino acid substitutions at position 4 and have assessed their ability to stimulate an antigen-specific T cell hybridoma when presented by viable antigen presenting cells (APC). Complexes between I-Au and one such analogue, rAc1-11[4A], were rapidly lost from the surface of live APC displaying a half-life (t 1/2) of approximately 10 min. Neither shedding of intact complexes from the cell surface, nor their internalization and recycling through an acidic intracellular compartment were found to account for their loss. The possible dissociation of rAc1-11[4A] from the peptide binding cleft was therefore addressed by comparing the t 1/2 of complexes between I-Au and peptide analogues of higher affinity. The tyrosine-substituted analogue, rAc1-11[4Y], remained stably bound to I-Au for at least 4 h, thereby displaying a t 1/2 far in excess of that evident for rAc1-11[4A]. Significantly, the wild type peptide, rAc1-11, bound so transiently that functional complexes could not be detected on the surface of peptide-pulsed APC. The physiological relevance of these findings was confirmed by extending our studies to an analysis of the homologous epitope of murine MBP; evidence that this epitope likewise displays minimal affinity for I-Au suggests a novel strategy for the escape from tolerance induction by encephalitogenic T cells.

Synthesis and characterization of mu-conotoxin IIIa.

mu-Conotoxin IIIa, a voltage-dependent sodium channel neurotoxin, has been synthesised using solid-phase peptide synthesis employing 9-fluorenylmethoxycarbonyl chemistry. After cleavage from the resin, the peptide was isolated by reverse-phase HPLC and then the six acetamidomethyl groups were removed by treatment with mercuric acetate. The reduced product so formed was purified by reverse-phase HPLC. Protocols were developed to optimize the oxidation of the cysteine residues to form disulphide bonds. Protocols employed using air oxidation together with 2-mercaptoethanol were the most effective. As complete oxidation was never obtained the oxidised peptide was purified by reverse-phase HPLC. The activity of our products was monitored using [3H]saxitoxin binding to eel membranes. The oxidised product was able to completely block [3H]saxitoxin binding in a competitive manner. Lineweaver-Burke analysis of [3H]saxitoxin binding gave a Ki of 1.5 nM, IC50 was determined as 26.6 nM. It was also shown that the pure synthetic mu-conotoxin IIIa had the same retention time on reversephase HPLC as the natural conotoxin IIIa. Thus an active toxin has been synthesised that can be used to probe sodium channels.

Mapping the cysteine residues and actin-binding regions of villin by using antisera to the amino and carboxyl termini of the molecule.

Peptide antisera specific for either the amino- or carboxyl-terminal regions of villin were used to locate the position of cysteine residues in immunoblots of villin cleaved with 2-nitro-5-thiocyanobenzoic acid. Maps constructed from the cleavage pattern suggest that villin contains six cysteine residues, two located in its amino-terminal peptide of Mr 44,000, and four located in the carboxyl-terminal peptide of Mr 51,000. Gel overlays of the partial cleavage fragments with 125I-labeled actin identified a calcium-dependent actin-binding region located within the amino-terminal peptide of Mr 32,000 of villin. The peptide antibody method used, called cleavage mapping, should be a convenient technique for mapping residues and ligand binding sites in proteins.

Metabolism of c-myc gene products: c-myc mRNA and protein expression in the cell cycle.

The presence and synthesis of c-myc protein and mRNA in the cell cycle has been studied. We find that c-myc mRNA is present, at equivalent levels, at all times in the cell cycle with the possible exception of mitosis. Furthermore, we demonstrate that this mRNA is transcribed in both G1 and G2 phases. An analysis of the c-myc protein in vivo shows that de novo synthesis occurs in G1 and G2 and the protein turns over with a half-life of approximately 20-30 min in both phases. Furthermore, the level of c-myc protein rapidly increases in cell populations when they re-initiate the cell cycle, thereafter decreasing as the culture reaches quiescence. The results therefore suggest that expression of c-myc can be rapidly modulated and that it is activated during the G0 to G1 transition, but is expressed thereafter in the cell cycle.

Conotoxin GI: disulfide bridges, synthesis, and preparation of iodinated derivatives.

The 13 amino acid toxic peptide from the marine snail Conus geographus, conotoxin GI, blocks the acetylcholine receptor at the neuromuscular junction. In this report, we describe a method for analyzing disulfide bonding in nanomole amounts of small cystine-rich peptides. The procedure involves partial reduction and a double-label alkylation of cysteine residues. Using this method, we show that the natural conotoxin GI has a (2-7, 3-13) disulfide configuration. The structure of conotoxin GI has been confirmed by chemical synthesis. The preparation and purification of molecularly homogeneous, iodinated derivatives of this toxin are also described. All derivatives, including the [diiodohistidine,diiodotyrosine]conotoxin GI, retained at least half of the biological activity of unmodified toxin. Since the tetraiodinated toxin, which is greater than 25% by weight iodine, retains considerable toxicity, unmodified histidine and tyrosine residues in conotoxin GI are not crucial for biological activity.

4-Chloromethylphenoxyacetyl polystyrene and polyamide supports for solid-phase peptide synthesis.

Two functionalised supports for the solid-phase synthesis of peptides under mild reaction conditions were prepared: 4-chloromethylphenoxyacetamidomethyl-copoly (styrene-1%-divinylbenzene) and 4-chloromethylphenoxyacetyl-norleucyl-poly (dimethylacrylamide). They were devised in order to avoid the danger of racemization which exists during base-catalyzed esterification of the first protected amino acid to the 4-alkoxybenzyl alcohol resins formerly employed in combination with N alpha-9-fluorenylmethoxycarbonyl and tert.-butyl side-chain protecting groups. Esterification of N alpha-protected amino acids to the new resins can be achieved easily and without significant levels of racemization by means of their caesium salts, while cleavage from the supports is possible by treatment with trifluoroacetic acid. The 4-chloromethylphenoxyacetyl polystyrene resin was tested by the synthesis of Leu-enkephalin which was cleaved, at the end of the synthesis, from the solid support in 91% yield by 60% trifluoroacetic acid in methylene chloride, and was shown to be more than 99% pure by ion-exchange chromatography and reverse phase high pressure liquid chromatography.

Synthesis of a 21-residue fragment of human proinsulin by the polyamide solid phase method.

The synthesis of human proinsulin fragment residues 40-60 by a solid phase method based on polydimethylacrylamide is described. Nineteen of the amino-acids were introduced as N alpha-(9-fluorenylmethoxycarbonyl) derivatives; functional side chains were protected as t-butyl ethers and esters. The yield of highly purified 21-residue peptide was 56.5%.

Synthesis and some properties and antitumor effects of the actinomycin lactam analog, (di(1-L-alpha, beta-diaminopropionic))actinomycin D1.

A lactam analog of actinomycin D (AMD) has been synthesized as a potential antitumor chemotherapeutic agent. Both L-threonine residues were replaced by L-alpha,beta-diaminopropionic acid. Starting with Nalpha-benzyloxycarbonyl-Nbeta-tert-butyloxycarbonyl-L-alpha,beta-diaminopropionic acid methyl ester hydrochloride the linear intermediate Nalpha-benzyloxycarbonyl-Nbeta-(tert-butyloxycarbonylsarcosyl-L-N-methylvalyl)-L-alpha,beta-diaminopropionyl-D-valyl-L-proline p-nitrophenyl ester was prepared by conventional methods of peptide synthesis in solution. Selective cleavage of the Nbeta-tert-butyloxycarbonyl group and lactam formation afforded the desired cyclic pentapeptide derivative. The chromophore precursor, Nalpha-(2-nitro-3-benzyloxy-4-methylbenzoyl) substituent, was introduced via its symmetric anhydride. Catalytic reduction followed by ferricyanide-mediated phenoxazinone formation provided the lactam analog, [di(1'-L-alpha,beta-diaminopionic acid)]actinomycin D ([Dpr1]2-AMD). Its binding to natural and synthetic DNA and that of an analogous L-threo-alpha,beta-diaminobutyric acid containing lactam ([Dbu1]2-AMD) compared with the binding of AMD (in which the peptides are in lactone form) was studied by circular dichroic (CD) spectroscopy. The visible and uv CD spectra of free AMD differed from those of the free lactam analogs, indicating that the asymmetric environment of the pentapeptide rings in the region of the chromophore differs in free actinomycin lactone and lactams. In the presence of calf thymus DNA, PM2 DNA, and the synthetic d(A-T)-like copolymers containing 2,6-diaminopurine (DAP), poly[d(DAP-T)], and poly[d(DAP-A-T)], the rotational strengths of the optically active transitions in the visible region of the actinomycins increased, and the CD spectra in the presence of the various DNA duplexes were qualitatively similar. The CD spectra of bound actinomycin lactams resembled the spectrum of bound AMD. This suggests that the lactone and lactam actinomycins acquire a similar environment when bound to DNA. [Dpr1]2-AMD was less cytotoxic than AMD in antibacterial assays but exhibited somewhat higher toxicity in mice than AMD. At optimal dose levels the lactam analog had little or no antitumor activity in three murine tumor systems.