Maturational and social factors contributing to relative age effects in school sports: Data from the London Youth Games.

Few studies have investigated whether relative age effects (RAEs) exist in school sport. None have sought to test the competing maturational and social-agent hypotheses proposed to explain the RAE. We aimed to determine the presence of RAEs in multiple school sports and examine the contribution of maturational and social factors in commonplace school sports. We analyzed birth dates of n=10645 competitors (11-18 years) in the 2013 London Youth Games annual inter-school multisport competition and calculated odds ratio (OR) for students competing based on their yearly birth quarter (Q1-Q4). Multivariate logistic regression was used to determine the relative contribution of constituent year (Grade) and relative age in netball and football which used multiyear age groupings. In girls, RAEs were present in the team sports including hockey, netball, rugby union, cricket and volleyball but not football. In boys, RAEs were stronger in common team sports (football, basketball cricket) as well as athletics and rowing. In netball and football teams with players from two constituent years, birth quarter better-predicted selection than did constituent year. Relatively older players (Q1) from lower constituent years were overrepresented compared with players from Q3 and Q4 of the upper constituent years. RAEs are present in the many sports commonplace in English schools. Selection of relatively older players ahead of chronologically older students born later in the selection year suggests social agents contribute to RAEs in school sports.

Variation in Responses to Sprint Training in Male Youth Athletes: A Meta-analysis.

The trainability of youths and the existence of periods of accelerated adaptation to training have become key subjects of debate in exercise science. The purpose of this meta-analysis was to characterise youth athletes' adaptability to sprint training across PRE-, MID-, and POST-peak height velocity (PHV) groups. Effect sizes were calculated as a measure of straight-line sprinting performance with studies qualifying based on the following criteria: (a) healthy male athletes who were engaged in organised sports; (b) groups of participants with a mean age between 10 and 18 years; (c) sprint training intervention duration between 4 and 16 weeks. Standardised mean differences showed sprint training to be moderately effective (Effect size=1.01, 95% confidence interval: 0.43-1.59) with adaptive responses being of large and moderate magnitude in the POST- (ES=1.39; 0.32-2.46) and MID- (ES=1.15; 0.40-1.9) PHV groups respectively. A negative effect size was found in the PRE group (ES=-0.18; -1.35-0.99). Youth training practitioners should prescribe sprint training modalities based on biological maturation status. Twice weekly training sessions should comprise up to 16 sprints of around 20 m with a work-to-rest ratio of 1:25 or greater than 90 s.

Athletic performance and birth month: is the relative age effect more than just selection bias?

The aim of this study was to determine if month of birth affects performance in 3 tests of physical function in children and adolescents. We measured cardiorespiratory fitness, handgrip strength and lower-body power expressed them relative to (whole year) age then compared scores between calendar year birth-months. We also expressed test performance as the likelihood of achieving criterion-referenced fitness standards. There were significant main effects of birth-month for cardiorespiratory fitness (F=4.54, p<0.001), strength (F=6.81, p<0.001) and power (F=3.67, p<0.001). Children born in November were fitter and more powerful than those born at other times, particularly the summer months (April, May and June). October-born children were stronger than those born in all months except September and November. This relationship was evident despite controlling for decimal age and despite no significant inter-month differences in anthropometric characteristics.There is a clear physical advantage for those born in the autumn and this may explain some of the bias in sports selection attributed to the relative age effect, particularly when the British school-year (September) cut-off is used.

Intermediate filament structure.

In the past year, several new developments concerning the structure of intermediate filament proteins and their assembly into intact intermediate filaments have been made: the coiled-coil structure of a rod domain has been elucidated; the basis of the chain interaction and its role in intermediate filament assembly has been specified; the organization of nearest-neighbour molecules in keratin intermediate filaments has been determined; and the glycine loop structures of the terminal domains of epidermal keratin chains have been defined. In addition, mutations in intermediate filament chains that promote pathology have been reported for the first time.

Hypomaturation amelogenesis imperfecta due to WDR72 mutations: a novel mutation and ultrastructural analyses of deciduous teeth.

BACKGROUND: Mutations in WDR72 have been identified in autosomal recessive hypomaturation amelogenesis imperfecta (AI). OBJECTIVE: to describe a novel WDR72 mutation and report the ultrastructural enamel phenotype associated with a different WDR72 mutation. METHODS: A family segregating autosomal recessive hypomaturation AI was recruited, genomic DNA obtained and WDR72 sequenced. Four deciduous teeth from one individual with a previously published WDR72 mutation, extracted as part of clinical care, were subjected to scanning electron microscopy, energy-dispersive X-ray analysis and transverse microradiography. RESULTS: A novel homozygous nonsense mutation, R897X, was identified in WDR72 in a family originating from Pakistan. Ultrastructural analysis of enamel from the deciduous teeth of an AI patient with the WDR72 mutation S783X revealed energy-dispersive X-ray analysis spectra with normal carbon and nitrogen peaks, excluding retention of enamel matrix protein. However, transverse microradiography values were significantly lower for affected teeth when compared to normal teeth, consistent with reduced mineralisation. On scanning electron microscopy the enamel rod form observed was normal, yet with inter-rod enamel more prominent than in controls. This appearance was unaltered following incubation with either α-chymotrypsin or lipase. CONCLUSIONS: The novel WDR72 mutation described brings the total reported WDR72 mutations to four. Analyses of deciduous tooth enamel in an individual with a homozygous WDR72 mutation identified changes consistent with a late failure of enamel maturation without retention of matrix proteins. The mechanisms by which intracellular WDR72 influences enamel maturation remain unknown.

Ultrastructural analyses of deciduous teeth affected by hypocalcified amelogenesis imperfecta from a family with a novel Y458X FAM83H nonsense mutation.

BACKGROUND: Nonsense mutations in FAM83H are a recently described underlying cause of autosomal dominant (AD) hypocalcified amelogenesis imperfecta (AI). OBJECTIVE: This study aims to report a novel c.1374C>A p.Y458X nonsense mutation and describe the associated ultrastructural phenotype of deciduous teeth. METHODS: A family of European origin from the Iberian Peninsula with AD-inherited AI was ascertained. Family members were assessed through clinical examination and supporting investigations. Naturally exfoliated deciduous teeth from 2 siblings were investigated by scanning electron microscopy (SEM), energy dispersive X-ray analysis (EDX) and transverse microradiography (TMR). RESULTS: On clinical and radiographic investigation the appearances of the affected deciduous and permanent teeth were consistent with hypocalcified AI with small focal areas of more normal looking enamel. DNA sequencing identified a novel c.1374C>A p.Y458X FAM83H nonsense mutation in affected, but not in either unaffected family members or unrelated controls. Exfoliated teeth were characterised by substantial post-eruptive enamel loss on gross examination. Irregular, poor quality enamel prisms were observed on SEM. These were coated in amorphous material. TMR and EDX confirmed reduced mineral and increased organic content in enamel, respectively. CONCLUSIONS: FAM83H nonsense mutations have recently been recognised as a cause of AD hypocalcified AI. We report a novel nonsense FAM83H mutation and describe the associated preliminary ultrastructural phenotype in deciduous teeth. This is characterised by poorly formed enamel rods with inappropriate retention of amorphous material, which is likely to represent retained organic matrix that contributes to the overall hypomineralised phenotype.

Periplakin, a novel component of cornified envelopes and desmosomes that belongs to the plakin family and forms complexes with envoplakin.

The cornified envelope is a layer of transglutaminase cross-linked protein that is assembled under the plasma membrane of keratinocytes in the outermost layers of the epidermis. We have determined the cDNA sequence of one of the proteins that becomes incorporated into the cornified envelope of cultured epidermal keratinocytes, a protein with an apparent molecular mass of 195 kD that is encoded by a mRNA with an estimated size of 6.3 kb. The protein is expressed in keratinizing and nonkeratinizing stratified squamous epithelia and in a number of other epithelia. Expression of the protein is upregulated during the terminal differentiation of epidermal keratinocytes in vivo and in culture. Immunogold electron microscopy was used to demonstrate an association of the 195-kD protein with the desmosomal plaque and with keratin filaments in the differentiated layers of the epidermis. Sequence analysis showed that the 195-kD protein is a member of the plakin family of proteins, to which envoplakin, desmoplakin, bullous pemphigoid antigen 1, and plectin belong. Envoplakin and the 195-kD protein coimmunoprecipitate. Analysis of their rod domain sequences suggests that the formation of both homodimers and heterodimers would be energetically favorable. Confocal immunofluorescent microscopy of cultured epidermal keratinocytes revealed that envoplakin and the 195-kD protein form a network radiating from desmosomes, and we speculate that the two proteins may provide a scaffolding onto which the cornified envelope is assembled. We propose to name the 195-kD protein periplakin.

In vitro assembly and structure of trichocyte keratin intermediate filaments: a novel role for stabilization by disulfide bonding.

Intermediate filaments (IF) have been recognized as ubiquitous components of the cytoskeletons of eukaryotic cells for 25 yr. Historically, the first IF proteins to be characterized were those from wool in the 1960s, when they were defined as low sulfur keratins derived from "microfibrils." These proteins are now known as the type Ia/type IIa trichocyte keratins that constitute keratin IF of several hardened epithelial cell types. However, to date, of the entire class of >40 IF proteins, the trichocyte keratins remain the only ones for which efficient in vitro assembly remains unavailable. In this paper, we describe the assembly of expressed mouse type Ia and type IIa trichocyte keratins into IF in high yield. In cross-linking experiments, we document that the alignments of molecules within reduced trichocyte IF are the same as in type Ib/IIb cytokeratins. However, when oxidized in vitro, several intermolecular disulfide bonds form and the molecular alignments rearrange into the pattern shown earlier by x-ray diffraction analyses of intact wool. We suggest the realignments occur because the disulfide bonds confer substantially increased stability to trichocyte keratin IF. Our data suggest a novel role for disulfide bond cross linking in stabilization of these IF and the tissues containing them.

Cingulin contains globular and coiled-coil domains and interacts with ZO-1, ZO-2, ZO-3, and myosin.

We characterized the sequence and protein interactions of cingulin, an M(r) 140-160-kD phosphoprotein localized on the cytoplasmic surface of epithelial tight junctions (TJ). The derived amino acid sequence of a full-length Xenopus laevis cingulin cDNA shows globular head (residues 1-439) and tail (1,326-1,368) domains and a central alpha-helical rod domain (440-1,325). Sequence analysis, electron microscopy, and pull-down assays indicate that the cingulin rod is responsible for the formation of coiled-coil parallel dimers, which can further aggregate through intermolecular interactions. Pull-down assays from epithelial, insect cell, and reticulocyte lysates show that an NH(2)-terminal fragment of cingulin (1-378) interacts in vitro with ZO-1 (K(d) approximately 5 nM), ZO-2, ZO-3, myosin, and AF-6, but not with symplekin, and a COOH-terminal fragment (377-1,368) interacts with myosin and ZO-3. ZO-1 and ZO-2 immunoprecipitates contain cingulin, suggesting in vivo interactions. Full-length cingulin, but not NH(2)-terminal and COOH-terminal fragments, colocalizes with endogenous cingulin in transfected MDCK cells, indicating that sequences within both head and rod domains are required for TJ localization. We propose that cingulin is a functionally important component of TJ, linking the submembrane plaque domain of TJ to the actomyosin cytoskeleton.

Communication between the transverse cervical nerve (C2,3) and marginal mandibular branch of the facial nerve: a cadaveric and clinical study.

Several branches of the facial nerve are known to anastomose with branches of the cervical plexus, other cranial nerves, and the trigeminal nerve. Communication between the sensory transverse cervical nerve (C2, 3) and marginal mandibular nerve is, however, less well known, and in a previous study of 86 neck dissections we reported a 2.3% incidence of anastomoses between them. In this prospective study, we meticulously searched for more examples using both formalin-fixed cadavers and neck dissections. A total of 102 necks were included (both sides of 36 cadavers (n=72 necks), and 30 patients who had neck dissection for the management of squamous cell carcinoma). We found communications between these nerves on one side of a cadaver and in one neck dissection. When combined with the numbers from our previous study, the overall incidence was 2.1% in 188 necks. The marginal mandibular nerve was inseparable from the anastomosis with the transverse cervical nerve, and the variant should not be forgotten if we are to reduce the chance of postoperative weakness of the lower lip, particularly when operative exposure is more limited (such as during removal of the submandibular gland).

Sleep: its importance and the effects of deprivation on surgeons and other healthcare professionals.

As clinicians, we sometimes fail to look after ourselves properly and do not regularly eat healthy foods or drink enough. Sleep is another factor that we often neglect. A lack of it can compromise our personal health and performance at work, and the "sleep debt" that results when this is chronic can take far longer to recover from than one might think. Now that junior doctors work more shift rotas and senior colleagues have onerous on-call responsibilities, we all need to be aware of the effects of sleep deprivation, which can lower the mood and motivation, weaken leadership, and result in more clinical errors. In this review we consider what might constitute enough sleep, the consequences of inadequate sleep, and how these might be addressed for surgeons.

Can dietary supplements improve a clinician's well-being and health?

Many people use dietary supplements to improve their physical and mental well-being and their general health, but do not know if they really have any benefit. To our knowledge, little has been published on their use in the clinical environment, so we evaluated the evidence for their benefits in people whose work is physically and mentally challenging. Studies on nutrition and supplementation in athletes and military personnel have clearly shown that several compounds improve cognition, mental well-being, and physical performance. Based on this evidence, and with the many pressures faced by healthcare workers, as well as the need for concentration and endurance, some dietary supplements might be beneficial. Supplementation of a balanced diet with omega-3 fatty acids, vitamin B3, vitamin C and associated antioxidants, vitamin D, and protein, may improve a clinician's physical and mental health and their performance at work. Specific research is, however, needed to evaluate this more fully.

Could exercise improve mental health and cognitive skills for surgeons and other healthcare professionals?

Workplace-related illness is common in the UK, and in healthcare more than five million working days over 10years have been lost as a result. Occupational stress is well known and can affect clinicians at any stage, yet many healthcare professionals continue to work with this or other psychological problems (including anxiety, chronic fatigue, and burnout) as they do not wish to let their colleagues down. Mental health issues might be dismissed, particularly in surgery, because there is a misconception that surgeons can cope better with stress than those working in other specialties, and are better protected from clinical burnout. The benefit of exercise on physical health is clear, but its role in the maintenance of good mental health and well-being should not be underestimated. As society adopts an increasingly sedentary lifestyle, exercise for many has a lower priority than other activities. In this article we give an overview of the mental health issues that might affect doctors and surgeons, and explore how exercise can benefit our well-being and clinical performance.

Developmentally regulated cytokeratin gene in Xenopus laevis.

We have determined the sequence of cloned cDNAs derived from a 1,665-nucleotide mRNA which transiently accumulates during Xenopus laevis embryogenesis. Computer analysis of the deduced amino acid sequence revealed that this mRNA encodes a 47-kilodalton type I intermediate filament subunit, i.e., a cytokeratin. As is common to all intermediate filament subunits so far examined, the predicted polypeptide, named XK70, contains N- and C-terminal domains flanking a central alpha-helical rod domain. The overall amino acid homology between XK70 and a human 50-kilodalton type I keratin is 47%; homology within the alpha-helical domain is 57%. The N-terminal domain, which is not completely contained in our cDNAs, is basic, contains 42% serine plus alanine, and includes five copies of a six-amino-acid repeating unit. The C-terminal domain has a high alpha-helical content and contains a region with sequence homology to the C-terminal domains of other type I and type III intermediate filament proteins. We suggest that different keratin filament subtypes may have different functional roles during amphibian oogenesis and embryogenesis.

Coiled-coil trigger motifs in the 1B and 2B rod domain segments are required for the stability of keratin intermediate filaments.

Many alpha-helical proteins that form two-chain coiled coils possess a 13-residue trigger motif that seems to be required for the stability of the coiled coil. However, as currently defined, the motif is absent from intermediate filament (IF) protein chains, which nevertheless form segmented two-chain coiled coils. In the present work, we have searched for and identified two regions in IF chains that are essential for the stability necessary for the formation of coiled-coil molecules and thus may function as trigger motifs. We made a series of point substitutions with the keratin 5/keratin 14 IF system. Combinations of the wild-type and mutant chains were assembled in vitro and in vivo, and the stabilities of two-chain (one-molecule) and two-molecule assemblies were examined with use of a urea disassembly assay. Our new data document that there is a region located between residues 100 and 113 of the 2B rod domain segment that is absolutely required for molecular stability and IF assembly. This potential trigger motif differs slightly from the consensus in having an Asp residue at position 4 (instead of a Glu) and a Thr residue at position 9 (instead of a charged residue), but there is an absolute requirement for a Glu residue at position 6. Because these 13 residues are highly conserved, it seems possible that this motif functions in all IF chains. Likewise, by testing keratin IF with substitutions in both chains, we identified a second potential trigger motif between residues 79 and 91 of the 1B rod domain segment, which may also be conserved in all IF chains. However, we were unable to find a trigger motif in the 1A rod domain segment. In addition, many other point substitutions had little detectable effect on IF assembly, except for the conserved Lys-23 residue of the 2B rod domain segment. Cross-linking and modeling studies revealed that Lys-23 may lie very close to Glu-106 when two molecules are aligned in the A(22) mode. Thus, the Glu-106 residue may have a dual role in IF structure: it may participate in trigger formation to afford special stability to the two-chain coiled-coil molecule, and it may participate in stabilization of the two-molecule hierarchical stage of IF structure.

Active cdk6 complexes are predominantly nuclear and represent only a minority of the cdk6 in T cells.

We have performed biochemical analyses of cdk6 complexes in T cells. By gel filtration chromatography we observed at least three cdk6 containing complexes in the cell, the most abundant eluting at 450 kDa and 50-70 kDa and a minor complex eluting at 170 kDa. Cyclin D was present in the minor 170 kDa complex which co-eluted with the peak of cdk associated in vitro Rb kinase activity. Analysis of proteins that co-immunoprecipitated with cdk6 showed that the 450 kDa complex contained both Hsp90 and CDC37. The 50-70 kDa complex was made up of two moieties, a 66 kDa complex containing cdk6 bound to p19INK4d and monomeric cdk6. The subcellular localisation of the cdk6 complexes was analysed by preparing cytoplasmic and nuclear extracts. The 450 kDa complex was shown to be predominantly cytoplasmic, whereas the 170 kDa cyclin D/cdk6 and the 50-70 kDa complexes were present in both nuclear and cytoplasmic compartments. When these same extracts were assayed for cdk6 associated kinase activity, only the nuclear cdk6 complexes were active. These data suggest that even though there are cdk6/cyclin D complexes detectable in both the cytoplasm and nucleus, only the cdk6 that is in the nucleus is active.

Ionic interactions in the coiled-coil domain of laminin determine the specificity of chain assembly.

Laminins are a family of large (800 to 900 kDa) multidomain glycoproteins specifically found in basement membranes. They consist of one heavy A chain and two light chains B1 and B2, and several tissue-specific laminin isoforms exist. Chain assembly is mediated through the formation of a rod-like triple-stranded alpha-helical coiled-coil domain about 75 nm long. The interacting edges of the chains are mostly formed by hydrophobic residues in positions a and d of an (abcdefg)n heptad sequence repeat and by a distinct pattern of charged residues in positions e and g. Here, we have analyzed the sequences of known laminin chains in an effort to relate them to interaction potential. Initially, those sequences localized in the long arm were arranged in an optimum heptad-repeating scheme. The interacting edges between chains were then analyzed for interchain hydrophobic and ionic interactions. The short heptad blocks were allowed to shift axially with respect to each other to maximize the number of interactions. The number of hydrophobic interactions was very high and similar for all chain combinations, but especially so for homodimers. As these were not observed experimentally, it seems that hydrophobic interactions probably represent only a prerequisite for coiled-coil formation. The number of ionic interactions, however, directly resembles the interaction potential observed in in vitro experiments. In particular, the number of interchain ionic interactions is high for parallel heterodimer configurations of A and B chains, but low for homodimer arrangements. When the laminin isoform chains, rat s-laminin (B1s) and human merosin (Am), are included in the analysis, they show rather low numbers of mutual interactions but high ionic interaction potentials between them and distinct mouse laminin chains are predicted. For mouse laminin the analysis was extended to a full three-stranded coiled-coil structure. The highest number of interchain ionic interactions occurs for an anti-clockwise chain arrangement of A-->B1-->B2 when viewed from the N terminus. None of the laminin chains appears to be designed for the formation of homodimers, although such conformations are frequently found in other alpha-fibrous proteins.

Keratin intermediate filament structure. Crosslinking studies yield quantitative information on molecular dimensions and mechanism of assembly.

One of the major obstacles to solving the full three-dimensional structure of keratin intermediate filaments (KIF) is the determination of the exact mode(s) of alignment of nearest-neighbor molecules; this in turn requires precise information of the lengths of the non-alpha-helical linker segments within the coiled-coil alpha-helical heterodimer molecule. In this study, we have induced lysine-lysine and cysteine-cysteine crosslinks between keratin intermediate filament molecules in small assembly-competent oligomers, isolated them and then characterized the natures and locations of the crosslinks. Of more than 100 found, 21 quantitatively major crosslinks were used to obtain the relative axial alignments of rod domain segments by least-squares fitting methods. Three dominant modes of alignment were found. In each case the molecules are antiparallel with the first involving molecules in approximate register (stagger = -0.2 nm), the second involving molecules staggered so as to bring the 1B segments into approximate alignment (stagger = -16.1 nm), and the third involving molecules staggered so as to bring the 2B segments into approximate alignment (stagger = 28.2 nm). In addition, the data enable quantitative estimates to be made for the first time of the lengths of the non-coiled-coil segments (L1 = 2.5 nm, L12 = 1.6 nm, L2 = 0.8 nm), and the total length of the rod domain (46.0 nm). Alignment of molecules according to these parameters permits construction of a two-dimensional surface lattice which displays a 1.6 nm (10 or 11 residue) overlap between similarly directed molecules. Together, the data predict six important overlapping sequence regions that recur about 16 times per 46 nm of filament length. Interestingly, synthetic peptides corresponding to these sequences, singly or in combination, significantly interfere with keratin filament structural integrity. These results thus represent the most significant set of structural constraints for KIF yet available and provide insights into how disease-causing mutations disrupt filaments and their organization in cells.

Crystalline fibril structure of type II collagen in lamprey notochord sheath.

We report here the existence of a crystalline molecular packing of type II collagen in the fibrils of the lamprey notochord sheath. This is the first finding of a crystalline structure in any collagen other than type I. The lamprey notochord sheath has a composition similar to that of cartilage, with type II collagen, a minor collagen component with 1 alpha, 2 alpha and 3 alpha chains, and cartilage-like proteoglycan. The high degree of orientation of fibrils in the notochord makes it possible to use X-ray diffraction to determine collagen fibril organization in this type II-containing tissue. The low angle equatorial scattering shows the fibrils are all about 17 nm in diameter and have an average center-to-center separation of 31 nm. These results are supported by electron microscope observations. A set of broad equatorial diffraction maxima at higher angles represents the sampling of the collagen molecular transform by a limited crystalline lattice, extending over a lateral dimension close to the diameter of one fibril. This indicates that each 17 nm fibril contains a crystalline array of molecules and, although a unit cell is difficult to determine because of the broad overlapping reflections, it is clear that the quasi-hexagonal triclinic unit cell of type I collagen in rat tail tendon is not consistent with the data. The meridional diffraction pattern showed 26 orders with the characteristic 67 nm periodicity found for tendon. However, the intensities of these reflections differ markedly from those found for tendon and cannot be explained by an unmodified gap/overlap model within each 67 nm period. Both X-ray diffraction and electron microscope data indicate a low degree of contrast along the fibril axis and are consistent with a periodic binding of a non-collagenous component in such a way as to obscure the gap region.