Their orientation and distance dependence is fully consistent with the model, suggesting that torsional gene coupling may constitute a widespread mechanism of (co)regulation in eukaryotes.DNA torsional stress is generated by virtually all biomolecular processes involving the double helix, in particular transcription where a significant level of stress propagates over several kilobases. If another promoter is located in this range, this stress may strongly modify its opening properties, and hence facilitate or hinder its transcription. This mechanism implies that transcribed genes distant of a few kilobases are not independent, but coupled by torsional stress, an effect for which we propose the first quantitative and systematic model. In contrast to previously proposed mechanisms of transcriptional interference, the suggested coupling is not mediated by the transcription machineries, but results from the universal mechanical features of the double-helix. The model shows that the effect likely affects prokaryotes as well as eukaryotes, but with different consequences owing to their different basal levels of torsion. It also depends crucially on the relative orientation of the genes, enhancing the expression of eukaryotic divergent pairs while reducing that of prokaryotic convergent ones. To test the  in vivo expression levels in the drosophila genome. This observation suggests that torsional coupling may play an important role in genetic regulation, and might favor the orientation-dependent co-localization of genes involved in similar functions, which need to be expressed together.During the transcription process, the genetic sequence encoded in the DNA molecule is expressed by an enzymatic complex. This process is often considered as independent for each gene, despite numerous reported cases of one transcribed gene perturbing a neighbor gene's expression, which is then regarded as a side-effect. Here, we suggest in the contrary that such interactions are a widespread feature, resulting from the propagation along the DNA molecule of mechanical stress generated during gene transcription. This torsional stress modifies the facility with which the transcription machinery separates the two strands of the double-helix in order to access the bases, and thus the expression level of any gene located nearby. We develop a quantitative model of this effect, showing that it depends strongly on the orientation of the genes, which is confirmed by the analysis of  Transcription involves the separation of the two DNA strands by the RNA Polymerase complex during the initiation phase. The formation of this “transcription bubble” In vitro experiments have shown the influence of supercoiling in both prokaryotic and eukaryotic transcription, as shown in pelE, inserted on a plasmid, is expressed by bacterial polymerase only when the DNA is underwound at a level similar to the in vivo average level of −0.06 (B) CUP1 promoter on a torsionally relaxed plasmid, but only in the presence of a minimal set of in vivo relevant transcription factors, and in particular TFIIH which contains an ATP-consuming helicase subunit ensuring the formation of the transcription bubble in absence of any transcription factorin vivo, it could very well play a role for those genes located in underwound domains. Interestingly, for these two very different systems, the expression rate is proportional to σ2, i.e. precisely the expected dependence of the promoter opening free energy, arising from the elastic cost of unwinding double-helical DNA in vivo is transcription itself In vivo measurements suggested that this transcription-induced supercoiling is probably a major determinant of “topological domains” in eukaryotic as well as bacterial chromatin Conversely, an important source of supercoiling combination immediately suggests that the transcription of adjacent genes could be coupled by the propagation of torsional stress along the DNA. This mechanism has already been suggested and experimentally demonstrated in specific examples of both prokaryotic These two reciprocal aspects of transcription-supercoiling coupling have been known for decades. Their In this paper, we propose a simple theoretical framework for this interaction, which allows exploring the role of different parameters  on the time-averaged co-expression of neighbor genes. The model focuses on the most generic features of the interaction since they prevail at the genome-wide level. It voluntarily leaves aside several specific aspects of promoters response to supercoiling. The proposed mean-field description is derived from the knowledge-based physical properties of the double-helix, and requires only few adjustable parameters to quantitatively reproduce the behavior of model experimental systems in very different (prokaryotic or eukaryotic) organisms. By extrapolating this behavior to different parameters, it allows to predict the differential effect of the torsional coupling in a broad range of conditions and organisms.global role of supercoiling in prokaryotic and eukaryotic regulation, the local torsional perturbation is predicted to affect the regulation of nearby genes in both types of organisms , in particular in the case of symmetrically oriented (divergent or convergent) genes, as already observed even without any molecular contact between the machineries expressing distant genes, the propagation of torsional stress along the DNA could significantly couple (positively or negatively) their expression. Finally, we show that the predictions of the model are supported by published expression data of Drosophila melanogaster, where the expression of isolated gene pairs significantly depends on their orientation and distance.Interestingly, despite the very different The destabilization of the double-helix by torsional stress is a well-known phenomenon  , with thin vitro experimental data  data see . In thesdata see . In the The superhelical stress involved in transcriptional regulation can have different origins. In prokaryotes, this level is controlled at the global scale by ATP-consuming enzymes The transcribing polymerase acts as a torsional motor that generates positive superhelical stress ahead of the complex, and negative stress behind it in vivo with different protocols, involving either the intercalation of a psolaren-based agent in underwound DNA in vivo experiments reported remarkably consistent propagation distances of around 1000 bases. Surprisingly, this value was observed not only in prokaryotic, but also in eukaryotic organisms, suggesting that nucleosomes do not modify significantly the propagation distance , but this number is strongly affected by a minority of very long genes (up to 2 Mb for humans). In contrast, the median length, which reflects the majority of the genes, is closer to 1 kb (e.g. 1.75 kb for D. melanogaster). Our illustrations are therefore relevant to most eukaryotic genes, but not to very long genes where the elongation kinetics probably plays an important role.If another promoter is located within a few kilobases of the transcribed gene, the curves of orientation of the two genes, as shown on divergent promoters (A), each promoter favors the expression of the neighbor, which in turn increases the former's activity. The diagram of the dynamic system shows that the effect is predicted to be stronger for eukaryotic organisms, and decays sharply at a cutoff distance that decreases with the basal superhelical level. Note that the diagram shows the relative change in expression level; in general, for a given promoter sequence, the absolute basal level is higher for lower values of σ0 . This type of interaction in well-known in biochemical networks, and can lead either to a global reduction of both expression levels, or to the selective extinction of one of the two genes. This repressive effect is predicted to affect bacteria more strongly than eukaryotic organisms. Finally, for genes in tandem (C), the interaction is more subtle, with an asymmetrical influence leading to a limited increase of the upstream gene and repression of the downstream gene, especially at intermediate basal levels of supercoiling where both effects may coexist.In the case of es of σ0 . In contwithout any interaction of their transcription machineries, simply by propagation of the DNA mechanical deformations.The presence of an interaction between the transcription of neighbor genes is often referred to as “transcriptional interference” in the biological literature, and has been reported in many studies divergent genes were found to be the most frequent as well as more expressed and highly correlated over-expression of the divergent genes.The proposed torsional coupling implies that neighbor genes are not independent, but coupled in an orientation-dependent way. Genome-wide expression analysis studies have demonstrated the co-expression of adjacent genes in yeast Drosophila melanogasteri.e. pairs where (i) the genes are closeby, with the transcription units (start or end sites) less than 5 kb from the other gene's promoter and (ii) the two promoters are more than 3 kb away from any gene outside the pair, and therefore likely unperturbed by their transcription-induced torsion. This situation is rare in yeast where the genome is dense (and even more so in prokaryotes), and where short-range torsional interactions may form long chains of coupled genes, making it difficult to distinguish the proposed effect . But another, likely stronger, mechanism is the competition between the opening of the polymerase binding site and structural transitions at distal sites, which involves specific DNA-binding regulatory proteins in vitro model promoters of pelE  An important point to notice is that our model only describes the time-averaged properties of gene expression. How these properties relate to the dynamic, Drosophila. It does not mean however that other species are not affected by the interaction, but the small number of these pairs in denser genomes makes it more difficult to test the predictions. This is true in particular for prokaryotes, where the predicted effects are different, but where most promoters are expected to be simultaneously coupled to several other genes, often with different orientations Drosophila, many genes were disregarded because their promoter was within torsional influence of more than a single gene. This situation is probably also frequent in the less compact mammalian genomes, where many genes were found to be densely clustered In the analysis of RNA-Seq data, we focused on the “torsionally isolated” pairs of genes, where the mutual interaction could be most clearly identified. Only in eukaryotes could we find a sufficiently large number of these genes, and we therefore restricted the analysis to Importantly, these relations extend not only to coding genes, but also to promoters controlling non-coding transcripts. These promoters have attracted considerable attention recently for their possibly widespread role in transcriptional regulation. Interestingly, while short non-coding RNAs have widely recognized functional roles, that of long ones is less clear, and in particular a subclass of long antisense transcripts In this paper, we model the time-averaged effect of transcription-induced superhelicity on gene expression. The model is the combination of three ingredients, which are developed in the following paragraphs:the spatial distribution of superhelicity generated by a transcribed gene, as described by a mean-field approachthe supercoiling-dependent opening free energy of a promoter sequence the thermodynamic model of transcription, which takes Consistently with the time-average approximation of gene expression, the distribution of superhelicity σ generated by a transcribed gene is described with a mean-field approach.a at either end of the transcription unit is assumed proportional to the promoter strength k and transcript length l, consistent with experimental observations x): x0 and in vivo experiments involving very different organisms and protocols The average superhelical density ±σThe supercoiling-dependent opening free energy of DNA is estimated from a recent efficient implementation m, v, q are adjustable parameters , respectively.For the curves of Note that for extreme positive torsions  best reproducing the experimental curve. Interestingly, we found that the in vitro expression data of the eukaryotic promoter CUP1  the thermal opening of underwound promoters and (ii) the assisted opening of relaxed promoters (about 4 times less frequent than the former). Note that because of the relatively large error bars in both experiments, these values are not very precise, but even large modifications would not change the qualitative predictions of the model. For the simulations of in vivo expression data of The formation of the transcription bubble is not a purely thermal process, but is facilitated by conformational changes within the RNA polymerase complex. This contribution is difficult to estimate precisely, and probably depends on the type of RNA polymerase. In particular, we expect it to differ between bacterial polymerase which requires no external source of energy to initiate transcription, and eukaryotic RNA PolII which contains an ATP-hydrolysis-dependent helicase subunit ter CUP1  are bestk of the gene is then given by: The probability to form a transcript, and hence the average transcription rate 0 the basal supercoiling level of the organism.Note that within this framework, the transcription rate fold-change due to supercoiling , and based on communication The RNA-Seq expression data from 24 The two genes of a pair were considered as correlated if (i) they were simultaneously expressed in at least 6 of the 24 experiments; (ii) Pearson's correlation coefficient between the 24 pairs of expression levels is larger than 0.5. A modification of these threshold values changed the absolute number of “accepted” pairs, but not significantly the relative number of divergent vs. tandem or convergent correlated pairs.Figure S1In vitro expression profiles of prokaryotic and eukaryotic promoters. Same data as in 2, suggesting that the rate is governed by the promoter melting energy.(EPS)Click here for additional data file.FigureS2Melting profile of the CMV promoter  employed in the  and .(EPS)Click here for additional data file.Figure S3Expression properties of downstream genes in Drosophila melanogaster tandem isolated pairs. The quantities shown in the different rows are identical to those of (EPS)Click here for additional data file.Figure S4In vivo expression data from , for genes inserted in tandem in different loci and orientations in the mouse genome. (A) The expression is always stronger for divergent promoters, consistent with a torsional coupling between the genes. (B) For tandem genes, the expression level depends strongly on the insertion site, consistent with a less systematic effect of supercoiling. On the other hand, the expression level is sometimes (third panel) even larger than in the divergent construction, suggesting other mechanisms not described by our model. This is also the case when a single gene was inserted instead of a pair (EPS)Click here for additional data file.


Clinically effective and cost-effective methods for managing problematic sexual behaviour in adolescents are urgently needed. Adolescents who show problematic sexual behaviour have a range of negative psychosocial outcomes, and they and their parents can experience stigma, hostility and rejection from their community. Multisystemic therapy (MST) shows some evidence for helping to reduce adolescent sexual reoffending and is one of the few promising interventions available to young people who show problematic sexual behaviour. This paper describes the protocol for Services for Teens Engaging in Problem Sexual Behaviour (STEPS-B), a feasibility trial of MST for problem sexual behaviour (MST-PSB) in antisocial adolescents at high risk of out-of-home placement due to problematic sexual behaviour.Eighty participants and their families recruited from five London boroughs will be randomized to MST-PSB or management as usual with follow-up to 20 months post-randomization. The primary outcome is out-of-home placement at 20 months. Secondary outcomes include sexual and non-sexual offending rates and antisocial behaviours, participant well-being, educational outcomes and total service and criminal justice sector costs. Feasibility outcomes include mapping the clinical service pathways needed to recruit adolescents displaying problematic sexual behaviour, acceptability of a randomized controlled trial to the key systems involved in managing these adolescents, and acceptability of the research protocol to young people and their families. Data will be gathered from police computer records, the National Pupil Database and interviews and self-report measures administered to adolescents and parents and will be analysed on an intention-to-treat basis.The STEPS-B feasibility trial aims to inform policymakers, commissioners of services and professionals about the potential for implementing MST-PSB as an intervention for adolescents showing problem sexual behaviour. Should MST-PSB show potential, STEPS-B will determine what would be necessary to implement the programme more fully and at a scale that would warrant a full trial.ISRCTN28441235 (registered 25 January 2012). Systematic reviews of the prevalence of sexual abuse in young people indicate that 18 to 20 % of females and 6 to 8 % of males in the general population report having been abused before the age of 18 . VictimsSexual offenders are heterogeneous , 7 and iDespite the evolving knowledge concerning juvenile sex offenders, much less progress has been made in developing effective interventions. In a recent systematic review of interventions designed to prevent reoffending among known sex offenders and individuals at risk of sexually abusing children, Langstrom and colleagues  were ablMST is an intervention that has been developed in the United States. It has an established track record for successfully treating young people who show serious and persistent antisocial behaviour . More reMultisystemic Therapy for Youth with Problem Sexual Behaviours (MST-PSB) is an adaptation of MST aimed at adolescents who have committed sexual offences and demonstrated other problem behaviours. It is an intensive family- and home-based intervention uniquely developed to address the multiple determinants of problematic sexual behaviour in adolescents. MST-PSB is designed to reduce problematic sexual behaviours, antisocial behaviours and out-of-home placements. Problematic sexual behaviour in adolescents is sexual behaviour that is developmentally inappropriate or potentially harmful to the self or others and includes behaviour of a sexual nature that infringes on the rights of others (such as the use of coercion or force) .Recent reviews of the sexual offending outcome literature , 30, 31 MST-PSB is therefore the only evidence-based intervention currently available for adolescents showing problem sexual behaviour. However, despite some initial positive findings, there are several unresolved issues relating to transporting MST-PSB to other health and social care systems and jurisdictions outside the United States. First, the effectiveness of MST-PSB needs to be carefully assessed in the UK mental health, juvenile justice and social care systems. The pattern of results found in the transportability of RCT evaluations of standard MST in Canada and Europe suggest that the effectiveness of MST-PSB needs to be demonstrated by independent investigators and outside the United States. Specifically, the magnitude of the associations between standard MST and treatment outcomes are substantially higher in trials that involved the developers of the intervention (effect size = 0.81) than in studies conducted without their close involvement (effect size = 0.27) , suggestThe two promising MST-PSB trials , 32 wereIn the United Kingdom, juvenile sex offenders are identified and receive mental health support and treatment through youth offending services and social care, where they are identified as being high risk, and in need of intervention and potentially out-of-home placement for safety reasons. Given the different ways in which young people displaying sexually harmful behaviour are referred in the United Kingdom, it is possible that participants in the UK trial will differ from their counterparts in the United States in the nature and severity of their problem sexual behaviour, as well as their co-occurring mental health disorders and problems. For example, mental health systems for identifying and treating young people showing problematic sexual behaviours are underdeveloped , and theThis paper describes the protocol for the Services for Teens Engaging in Problem Sexual Behaviour (STEPS-B) trial, a UK feasibility evaluation of MST-PSB. The MST-PSB intervention uses the term ‘problem sexual behaviour’ to describe young people who display serious non-normative sexual behaviours that victimize others, which can be aggressive or non-aggressive sexual acts . GerardiThe STEPS-B trial aims to assess the feasibility of implementing MST-PSB and also the barriers to implementing it: young people who display problem sexual behaviour suffer intense stigma and blame . There aThe MST-PSB feasibility trial is being conducted at the Brandon Centre, London, a charitable organization serving the mental health needs of young people. The Brandon Centre is participating in the RCT as part of its funding agreement with the UK Department of Health for having a clinical team. In the run-up to the trial, clinicians at the Brandon Centre will have been working with MST-PSB families for at least 18 months. This represents a bedding-in period to help raise awareness of the Centre’s specialized service among referrers , as well as ensuring that the therapists are applying the therapeutic model of MST-PSB (see below) in accordance with MST principles and guidelines. The Brandon Centre developed expertise in delivering MST for general antisocial behaviour (‘standard’ MST) in the first RCT of MST in the United Kingdom  and has There are specific feasibility issues in implementing an RCT for problem sexual behaviour in the United Kingdom. These include the challenge of recruiting an adequate sample of adolescents displaying sexually problematic behaviour; the acceptability of an RCT to the key agencies and systems that are involved in managing these young people and in providing them with mental health services; the ability to identify comparable interventions specifically designed to target problem sexual behaviour ; and the acceptability of the research protocol to young people and their families, given the stigma associated with individuals who display problem sexual behaviour and high levels of anxiety within families and professional systems regarding the management and treatment of this behaviour.While the social and fiscal costs of adolescent sexual offending are substantive and warrant the development of effective and evidence-based services, European epidemiological studies of adolescent sexual offending in Oxfordshire, UK  and SwedThe available data on prevalence and the difficulty gaining accurate estimates of incidence and service use suggest that the size of the pool of available participants to recruit into an RCT for problem sexual behaviour is uncertain. Therefore, the success of the trial is likely to depend on efficient identification of potential participants. It will also rely on established systems for providing specialist treatments for adolescent sexual offenders and for managing their mental health needs, given that judicial processes can determine the offenders’ availability for mental health interventions or delay timely provision of treatment. Two recent reviews of the provisions for juvenile sexual offenders in the United Kingdom noted that, despite examples of good practice, services for identifying and intervening with young people displaying problem sexual behaviour are underdeveloped . For exaFinally, the feasibility of the MST-PSB trial will undoubtedly depend on the acceptability of the research protocol to young people and their families. The research protocol for the MST-PSB trial, including the majority of the measures that will be administered to young people and their carers, has been successfully implemented with a diverse and large sample of young people showing antisocial behaviour and their carers in the START trial of standard MST . HoweverThere are also several unresolved issues related to transporting MST-PSB to health and social care systems and jurisdictions outside the United States. In relation to a UK feasibility trial, the following issues require systematic monitoring: the clinical competence of therapists to practise MST-PSB in light of the curriculum differences for psychologists and social workers in the United Kingdom and the United States; the absence of data concerning the evidence base of usual services  in the United Kingdom, given there are no prior RCTs of interventions for problem sexual behaviour in this country; differences in the motivation of MST therapists and the research team who were not involved the development of MST-PSB; and contextual issues such as differences in national standards with regard to sentencing policy and practice. In addition, while the sample sizes of studies with sexual offenders in general, and RCTs in particular, are relatively small, the larger MST-PSB effectiveness trial  providedThe aim of STEPS-B is to carry out a feasibility trial that will inform policymakers, commissioners of services and professionals about the potential for implementing MST-PSB as a programme for intervening with young people showing problem sexual behaviour. Should MST-PSB show potential in the United Kingdom context, the trial will determine what would be necessary to implement the programme more fully and at a scale that would warrant a full trial.The trial will investigate whether MST is associated with the elimination or reduction of levels of sexual and nonsexual offending; reduction or elimination of problem sexual behaviours; reduction in nonsexual antisocial behaviour; less time spent in custodial institutions; and improved educational outcomes. It will also aim to establish the cost of MST-PSB relative to MAU, and the cost-effectiveness of providing this intensive form of intervention against the background of costs incurred in the 20-month period following randomization.Finally, the trial will investigate specific feasibility issues that pertain to effective implementation of MST-PSB in the United Kingdom. These include: specific characteristics of young people’s problem sexual behaviour, where they are referred from, and what services have been or are being provided to them whether or not they are randomized into the trial; the beliefs and attitudes held by professionals around family intervention for this population, given that the young person will remain in the family home despite having committed sexually harmful behaviour towards other young people, including, in some cases, a sibling living in the same home; the degree to which professionals are supportive of the procedures and demands associated with an RCT research evaluation, including the randomization and its consequences; and the degree to which the institutions and agencies that manage young people showing problem sexual behaviour, such as the courts and child protection services, are able to support an RCT evaluation. It will also investigate the acceptability of the trial protocol to young people and their carers, and will gather information on their experience of MST-PSB and MAU interventions, through qualitative interviews following completion of treatment.The STEPS-B trial is a single-site feasibility trial comparing MST-PSB with carefully documented MAU for adolescents who meet criteria for being at ‘high risk’ of requiring out-of-home placement, specifically when this risk is associated with problem sexual behaviour.The study protocol was approved by the London-South East Research Ethics Committee (reference number 11/LO/0772). Research and development approval has also been sought and given for the Brandon Centre by Camden and Islington Research Ethics Committee (11/LO/1707).The study will be conducted by the research team based at University College London (UCL). The team will be responsible for collecting data from cases referred to the Brandon Centre, London. Eighty families will be recruited to the study through referrals made to the Brandon Centre from at least five London boroughs. Referrals are anticipated from social care, CAMHS and youth offending services.Participants will be recruited over 3 years. Recruitment will commence about 18 months after the therapists at the Brandon Centre have started working on MST-PSB cases; this initial period will allow MST-PSB therapists sufficient time to reach fidelity to the MST-PSB model before randomization begins. The team of three MST-PSB therapists is guided by a supervisor - a doctoral-level  therapist with sound knowledge of the theories and therapeutic practices underpinning MST-PSB and experience in providing MST clinical supervision. The MST-PSB supervisor conducts weekly group supervision, and one-to-one supervision of the MST-PSB therapists as needed, to ensure therapists are adhering to MST-PSB principles in their delivery of the intervention.Eighty participants will be recruited and randomly allocated, with half of the consecutive qualifying cases being assigned to MST-PSB and the other half to MAU.It is estimated that the Brandon Centre will have approximately two eligible cases per month, where referrals are obtained from a minimum of four boroughs, which would result in 72 cases randomized into the trial over the 3-year recruitment period. Five more boroughs are set to join the trial, resulting in nine boroughs in total; it is predicted that this will result in 34 eligible referrals per year, to give a total of approximately 102 cases randomized over 3 years.P < 0.001, Fisher’s exact test) is 0.32 to 0.49 . In the current study, non-offender status is defined as no arrest leading to conviction over the follow-up period, based on police computer records. We expect to achieve the lower end of the effect size range, given that the trial is mounted independently  of the development team and involves the transportation of the clinical methodology across national boundaries, healthcare systems and cultures. A sample size estimate to give 80 % power based on the lower estimated effect size of 0.32 suggests 56 participants in a two-group design. Figure Effect size has been estimated on the basis of the available data from a recent MST intervention study of sexual offenders in the United States . As onlyParticipants will be adolescents who meet the criteria for ‘high risk’ of requiring out-of-home care, specifically when this risk is associated with problem sexual behaviour.Young people aged 10 to 17 years oldSufficient family involvement for MST-PSB to be applied: either living in the parental home, in long-term foster care, or in short-term out-of-home placement but family reunification is imminent (within 30 to 60 days)The young person and family are not already engaged in a problem sexual behaviour-specific treatmentProblem sexual behaviour within the past yearRisk of custody or out-of-home placementRisk of reoffendingDisplaying aggression at home, at school or in the community.Inclusion criteria are as described below:Young person is living independently or a primary caregiver cannot be identifiedThe sexual offence is under police investigation at the time of referralThe primary reason for referral is related to suicidal, homicidal or psychotic behavioursCaregiver persistently denies that the young person has engaged in problematic sexual behaviourYoung person has pervasive developmental delaysYoung person has generalized learning problems  as indicated by IQ <65Young person has severe substance misuse as the primary presenting problem.Exclusion criteria are as follows:The authors have developed a multiple gating procedure for recruitment, based partly on the experience of the largest adolescent mental health trial to date evaluating standard MST for youth displaying conduct problems , which, In order to maximize recruitment and to ensure that our planned sample size is achieved, research personnel are involved in several activities with different parts of the system that may influence recruitment. Specifically, a member of the research team attends service team meetings and multi-agency panels that review potential referrals in the different boroughs, and there is regular contact with service leads and managers, which will influence the visibility and importance of the trial within services. Finally, ongoing service and agency awareness is complemented by ongoing presentations , a quarterly newsletter sent to all potential referrers, and an active website, which provides information for service teams about how to make a referral, a referral leaflet, a description of MST-PSB and the STEPS-B trial, and a ‘frequently asked questions’ section for young people, parents/carers and professionals.Referrals are anticipated from youth offending services, CAMHS, social care and education services see Fig. . Prior tBefore a referral form is submitted, the potential referrer will complete a telephone case consultation with the MST-PSB supervisor. As the cohort is not homogeneous, individual cases will often require different levels of support. During the telephone consultation, the MST-PSB supervisor will screen the case against the eligibility criteria and begin to plan in parallel with the referrer another potential sexually harmful behaviour intervention, if the case and family are not randomly allocated to MST-PSB.Once a referral has been accepted, the clinical team sends a standard letter to the parents (and separately to the young person if aged 16 or 17), informing them of the trial. In addition, the MST-PSB supervisor contacts the family by telephone to discuss the trial and arrange an introductory meeting.The introductory meeting takes place at the family home, with the MST-PSB supervisor, senior RA, young person and primary carer(s) present. At this meeting, the nature of and concerns related to the young person’s problem sexual behaviour are reviewed and a credible MAU path , should the young person not be randomized to receive MST-PSB, is agreed upon by the young person, primary carer(s) and the trial team. Further information is also gathered regarding eligibility, such as agency involvement incompatible with participation in the trial , potential risk of harm to workers, or whether the young person has generalized learning problems. In the event that the primary carer steadfastly denies that the young person has engaged in problem sexual behaviour, the MST-PSB supervisor will offer a maximum of three sessions to explore the allegations with the primary carer(s), to see whether their position shifts. At the end of the introductory visit, the young person and primary carer(s) are given specifically tailored information outlining the trial, in age-appropriate and culturally appropriate language. The family is given contact details for the RA so that they can contact the RA if they have any further questions.Unless the family expresses a decision not to participate at the time of the introductory visit, they are given a minimum of 3 days to consider whether they would like to join the clinical trial. If the RA does not hear from the family within the agreed time period, the RA telephones the family, answers any additional questions they may have, and arranges an appointment for a research assessment.At the research assessment, each young person and primary carer(s) sign informed consent forms agreeing to join the clinical trial, including permission to access the police, correctional and educational databases, which remains in effect for 20 months. For families who sign the consent forms, the RA administers pre-randomization questionnaires and measures are completed during this contact .Once the young person and family’s eligibility have been confirmed and the instruments completed, the RA contacts the trial centre and relays the family’s randomization details. The clinical team informs the family and referrer of the outcome of randomization within 48 hours of the consent and pre-randomization visit being completed.Once the family has completed the compulsory instruments to be able to join the trial, the young person is asked whether they would like to take part in the optional Child Attachment Interview . If the young person and primary carer(s) agree, an additional meeting is arranged within 1 week of the date of randomization. At this additional visit, consent forms are signed by the primary carer(s) and young person and the Child Attachment Interview is conducted.Following consent and the baseline (pre-randomization) assessment, a trial identification number is assigned to the participant. Eligible consenting participants are randomized to MST-PSB or MAU on a 1:1 basis by the Trials Unit at UCL through the use of a secure randomization service that ensures allocation concealment. A computer-generated adaptive minimization algorithm that incorporates a random element is used with the following stratification factors: gender, age , conduct problems presenting with problem sexual behaviour (yes or no) and age differential between the perpetrator and victim (<4 years or ≥4 years). These strata were selected because previous research has shown that a greater age differential between perpetrator and victim, and whether the young person is also displaying antisocial behaviour, are associated with more severe maladjustment and poorer long-term outcomes , 47, 48.To minimize the possibility that RAs will learn of the treatment allocation for any individual participant, the RAs are blind to treatment allocation at randomization, and there is no direct communication between RAs and therapists. This procedure has been used in the START trial of standard MST  and has MST is based on the premise that behaviour is multi-causal; therefore, to change an individual’s behaviour successfully, multiple drivers need to be addressed. Consequently, the approach works with the young person’s ecology , as well as the young person themself; places a high premium on approaching each young person/family as unique; and has a specific focus on parental/carer involvement in treatment delivery, due to carer(s) being viewed as the key agents of change. MST has a track record of working with hard-to-reach, complex families with multiple needs. To increase engagement and formulation, a family is allocated one therapist who employs interventions that are strongly supported and informed by research. The sessions take place in the family home, and outreach work is undertaken when necessary. The family also has access to 24-hour/7 days a week telephone support with a therapist who is fully briefed on the family’s difficulties, safety plans and stage of therapy, so that support is readily available in times of crisis.MST was originally developed to work with youth who engage in antisocial behaviour. In recent years, the research and implementation base of standard MST has been built upon to produce interventions to treat young people with a range of other difficulties, such as substance abuse, diabetes management, psychiatric care, child abuse and neglect, as well as problem sexual behaviour.Problem sexual behaviour is an exclusion criterion for standard MST. MST-PSB, the adaptation of standard MST specifically for working with youths showing PSB, aims to reduce the level of both sexual and non-sexual offending in the target population. Supplementary to MST, MST-PSB has a greater focus on safety planning, individual factors , and interventions specific to problem sexual behaviour, such as offence clarification sessions aimed at increasing accountability and safety and the promotion of normative sexual behaviour. Furthermore, family therapy techniques, such as structural and strategic family therapy interventions, are utilized to a greater extent than in standard MST, as well as peer interaction skills. In addition, where this is a factor, the impact of the young person’s own victimization and experience of abuse is assessed.The intervention procedure consists of regular visits to the family home to meet with the young person and/or parent(s) for approximately 1 hour. Visits take place approximately three times per week at the beginning of the intervention and the frequency is reduced as the intervention progresses. Overall, the intervention time is 5 to 7 months.In the STEPS-B trial, MST-PSB will be delivered by a team of at least three specially trained clinicians under the supervision of an MST supervisor, with weekly 1-hour conference calls for consultation with an MST Services staff member. It is expected that MST-PSB therapists will also have the support of consultation with local mental health professionals. To further support the therapists’ supervision and delivery of the intervention, a Therapist Adherence Measure-Revised (TAM-R) will be completed by the participants’ carers with an independent researcher every 3 weeks to review the therapist’s adherence to the nine principles of the MST model . In addiThe MST-PSB team at the Brandon Centre is centrally funded by the Department of Health and will have been operational for 18 months by the start of the clinical trial; during this period the intervention has been offered to referrers free of charge . This bedding-in period has established the team in the referring boroughs, which has led to relationships being built with referring agencies and other intervention services addressing harmful sexual behaviour and to referrers increasing their awareness and knowledge of the referral criteria and intervention. Furthermore, as the MST-PSB team has been funded on condition that it participates in the research trial, greater collaboration is achieved between the MST-PSB team and research team by working together to meet joint targets such as recruitment numbers. Finally, the initial 18-month period will have given the MST-PSB team experience of implementing and addressing challenges to delivering the intervention in a UK context, therefore increasing treatment fidelity prior to the start of the transportability evaluation. To further ensure treatment fidelity, the MST developers have developed an extensive quality assurance system  and, in MAU constitutes the way in which a case would be managed if MST-PSB was not available. The authors aim to compare MST-PSB with specialist interventions for adolescents displaying problem sexual behaviour provided through social services, youth offending services and CAMHS. It is not assumed that MAU will be uniform across the referring boroughs. Instead, it is expected that the length, intensity and models of intervention will vary. Furthermore, it is envisaged that some young people assigned to MAU may not receive any interventions, owing to the limited service provision available where they live. The trial will map the specific profiles of services delivered to youth randomized to the MAU arm of the trial, as well as the approaches used within MST-PSB.The service to which a young person may be referred for MAU is likely to depend on the severity of the case, the interventions available in the borough’s directorate and the provision for the borough to spot purchase. Potential options for MAU may include: AIM (Assessment Intervention and Moving on), an assessment and intervention programme that can be delivered by CAMHS, youth offending teams and social care practitioners; Youth In Need, a group CBT programme with an emphasis on the developmental perspectives underpinning offence-related issues; The Portman Clinic, which provides individual therapy using a psychoanalytical model; SWAAY Child and Adolescent Services Ltd or the Bracton Centre, both of which offer residential placements for young people where they can access a variety of individual and group programmes specifically targeting problem sexual behaviour as well as attending to the young person’s educational needs; and practitioners in CAMHS, social care and youth offending services who have developed expertise in working with this client group. Depending on the seriousness of the problem sexual behaviour, the young person may attend the NSPCC’s National Clinical Assessment and Treatment Service (the NSPCC is a UK children’s charity with a major focus on abuse prevention and intervention), which offers assessment, a multidisciplinary risk assessment report and treatment. The project’s intervention models include CBT, psychotherapy, individual psychological therapy and group therapy.To maximize the clinical validity of the outcome evaluations, assessments are being made using multiple methods  completed by different informants  and teacher(s)) who are significant in the multiple domains that characterize young people’s functioning .The feasibility outcomes measure chosen variables that the investigators hypothesize will be related to the effective implementation of MST-PSB in the United Kingdom.First, it is predicted that specific characteristics of young people referred for displaying problematic sexual behaviour will be more likely to result in these young people being randomized into the trial. The specific characteristics that will be examined include the age of the young person, severity of the young person’s problematic sexual behaviour, source of the referral, and whether or not the young person was involved in judicial proceedings at the time of randomization.Second, the beliefs and attitudes held by professionals working in the agencies and services that will refer into the trial will be evaluated. The data gathered concerning their beliefs and attitudes will include the degree to which they are supportive of a family-based intervention for a young person displaying problematic sexual behaviour; the degree to which they are in accord with the procedures and demands associated with an RCT research evaluation, including the randomization and its consequences; and the degree to which they view the institutions and agencies that manage young people showing problematic sexual behaviour as able to support an RCT evaluation.The trial team has developed a bespoke 15-item questionnaire (the Feasibility Questionnaire) to assess beliefs and attitudes in the following areas: family treatment as an appropriate intervention for young people showing problematic sexual behaviour ; support for RCT evaluations of young people showing problematic sexual behaviour ’); and perceptions of the systemic context (agencies and institutions) as facilitative of RCT evaluations ’). These three areas have been chosen on the basis of preliminary discussions with referrers and agencies that manage young people showing problematic sexual behaviour in the localities where recruitment will take place, as well as our previous experience liaising with referrers and the major agencies involved in providing CAMHS while conducting two earlier RCTs of standard MST, including a large multi-site trial . The expThird, the acceptability of the trial protocol to young people and their carers will be investigated through qualitative interviews following the completion of treatment. The qualitative interviews concerning participants’ experience and views of the trial protocol will be part of a larger semi-structured interview, which will include discussion of their experiences of the MST-PSB and MAU interventions (see ‘Interviews’ below). The part of the interview that elicits information regarding the trial protocol will focus on several domains: the process of identifying and discussing the young person’s problematic sexual behaviour as part of the family visit at which consent is obtained; the explanation of randomization and their understanding of the two treatment options available to them; and their perceptions regarding the appropriateness and challenges of completing the study’s standardized measures.The primary outcome is the proportion of documented cases of offending within 20 months of randomization, based on police records supplemented by information obtained from the young person, parents/caregivers and social care services. While this may be only a partial or even an inaccurate reflection of the psychological conditions and social behaviours of the participants, it is the indicator of primary concern for the funders of the programme as, in most cases, this record triggers costly juvenile justice and social interventions. A linked outcome, of similarly high concern to funders of services, is assignment to long-term (3 months or longer) out-of-home placements in specialist residential provision when social services consider that the safety of the participant’s family or community is compromised by the individual, even in the absence of arrest and conviction. This means all participants who are placed into local authority care, are incarcerated, long-term hospitalized or offered residential schooling at any time in the 20 months following randomization will not be considered to have moved from the offending to the non-offending group. This information will be obtained from caregivers and cross-referenced with documented information from the national social care services database to maximize accuracy. The investigators expect this trial to give information on how many young people assigned to MAU and to MST-PSB require specialist residential provision either immediately after intervention or during the follow-up period.Secondary outcomes are limited in number to reduce the measurement burden on young people and their primary carer(s), as the investigators’ previous experience has shown that having too many measures to complete is a disincentive to continued participation , 28. TheObjective outcomes will be collected from reports of sexual and non-sexual offending behaviour based on police computer records, including details of custodial sentences. These measures will be taken at 6-month intervals, for the 6 months before randomization, the 8 months covering the intervention period (adjusted as MST-PSB lasts on average 5 to 7 months), and 6-monthly until the 20-month follow-up point. The number of records of offending behaviour (count data) will be obtained and 6-month periods free of any offending behaviour will also be recorded (binary data). Records will be obtained from the Police National Computer as well as from the Young Offender Information System database; these records detail information on offences, court appearances, criminal orders, police custody records and arrest rates. Records of school attendance and exclusions will be retrieved from both the schools themselves and the National Pupil Database to assess educational outcomes.The RA will administer pre-testing questionnaires during the initial contact with the young person and family after they have consented and before group assignment. Post-testing by the RA will take place 8 months after entry into the study; it is intended that this will be a minimum of 2 weeks after the family completes the intervention (MST-PSB or MAU). Follow-up assessments will be made at 14 and 20 months post-randomization. Self-report measures of well-being and adjustment, antisocial behaviour, inappropriate sexual interests and difficulties, parenting skills and family functioning, as well as parental mental health and adjustment, will be collected.A general assessment of well-being will be obtained using the Strengths and Difficulties Questionnaire . DepressThe Multidimensional Inventory of Development, Sex, and Aggression (MIDSA)  and the The prevalence and incidence of delinquent behaviour  and of peer delinquency will be assessed using the Self-Report Delinquency measure . Non-comThe quality of the relationship between adolescent and carer(s), family functioning and parenting practices will be assessed using the Family Adaptability and Cohesion Scales , the InvParental mental health will be briefly assessed using the General Health Questionnaire-28 , a commoA bespoke interview (Demographic Interview for Parents) covering general family information, including parental forensic history, schooling and economic information, constructed by the investigators, will be administered to all parents.Child psychometrics will be obtained by using the Wechsler Abbreviated Scale of Intelligence . PsychiaAfter the intervention, young people and their carer(s) will be interviewed to elicit their views regarding their experience of MST and factors that they perceive had facilitated or inhibited therapeutic change, based on qualitative research carried out during the Brandon Centre trial  and the Health economic analysis will be conducted by the Centre for the Economics of Mental Health at the Institute of Psychiatry, London. This analysis will explore the relative costs and cost-effectiveness of MST-PSB and MAU. The evaluation will take a broad perspective and will include all health, social services, education and voluntary sector services, as well as costs to the criminal justice sector, costs resulting from crimes committed, and out-of-pocket expenses to the young people and their families.Data on MST contacts will be collected directly from the Brandon Centre to avoid participants revealing their group allocation to the RAs. Data on the use of all other services will be collected in interviews using the Child and Adolescent Service Use Schedule (CA-SUS) and EQ-5D questionnaire, which have previously been used with young people with complex mental health and social care needs –73. The A bespoke fidelity measure, the Children and Young People - Resources, Evaluation and Systems Schedule (CYPRESS) , will be used to enable accurate characterization of the programme components of MST-PSB and MAU interventions. This approach is based on the initiative of the MST developers , who undTo obtain additional information about participants’ experience of MST-PSB and being in an RCT of this treatment, the semi-structured interview developed by Tighe and colleagues  in the BTranscripts will be analysed thematically using Interpretative Phenomenological Analysis (IPA). The primary objective of IPA research is to understand how individuals make sense of their experiences . The intFollow-up assessments will be conducted at 8, 14 and 20 months post-randomization. Table All analysis will be according to the intention-to-treat principle. The characteristics of the treatment groups will be described at baseline. Preliminary analysis will investigate the pattern of missing follow-up data.The primary outcome of the proportion of cases where a sexual offence has been incurred during the follow-up period will be described by a Kaplan-Meier graph and summarized by the proportions in each group with a new sexual offence by 20 months. The linked outcome of out-of-home placement will also be described by a Kaplan-Meier graph and summarized by the proportions in each group with out-of-home placement by 20 months. The number of events will be estimated in order to inform power calculations for a fully powered efficacy trial.For time-to-event offending outcomes, the event rate will be estimated. For continuous secondary outcomes, such as questionnaire data, linear mixed effects models will be fitted in order to estimate the individual-level residual variance.The feasibility outcome evaluating individual and systemic factors associated with recruitment into the trial will be analysed using logistic regression. The total score and subscale scores on the Feasibility Questionnaire will undergo descriptive analyses to characterize the direction and range of attitudes by referrers involved in the MST-PSB trial. As noted above, qualitative interviews eliciting the views of young people and their carer(s) on the underlying principles and demands of participating in an RCT will be analysed using IPA.There is very little research that has evaluated therapeutic interventions for young people displaying problem sexual behaviour, and to date, no RCTs have been conducted in the United Kingdom. Although the results from two RCTs of MST-PSB , 32 haveEvidence to support MST-PSB remains limited. Both prior evaluations of MST-PSB were conducted in the United States by the developers of the intervention and recruited participants solely through the juvenile court system. Moreover, in one of these two trials, measures of critical sexual and non-sexual reoffending outcomes were not reported and, as noted by the authors, the sample of young sexual offenders displayed very low levels of general psychopathology . This fiThe primary aim of this study is therefore to conduct a rigorous, community-based trial in which MST-PSB is compared with the range of services that are typically provided to adolescent sexual offenders in the United Kingdom . Whereas there is no good evidence that MAU for these young people and their families works, there is some evidence available to suggest that MST-PSB is of benefit. Therefore, this evaluation of both treatment options will enable closer observations of MAU services to provide an evaluation of interventions currently available in the localities, as well as MST-PSB. This pragmatic trial will inform policymakers, commissioners of services and professionals about the potential of MST-PSB in the UK context. Specifically, it will provide evidence to determine whether the provision of MST-PSB could reduce the incidence of out-of-home placements for young people at risk of being removed from their homes primarily because of problem sexual behaviour.To ensure that the study achieves the fairest and least biased assessment of the potential benefits of MST-PSB, the investigators have designed the study with the support of MST Services but will carry out the RCT independently from the developers; however, the developers provide regular consultation to the clinical service at the Brandon Centre to ensure that MST-PSB is delivered to the highest levels of fidelity. As part of the feasibility evaluation, individual and systemic factors will be assessed that may be associated with young people being recruited into the trial, the beliefs and attitudes held by practitioners about the usefulness of empirically evaluating services for problematic sexual behaviour in the United Kingdom, and the perceptions of young people and their carers of having participated in an RCT for problematic sexual behaviour and of the intervention that they received.absence of out-of-home placement) as the main measure of potential benefit. Other possible benefits of MST-PSB will also be examined, such as the impact on the young person’s mental health and well-being, and that of the carer(s).The research is being conducted by a team based at UCL, with data collection from cases referred to the Brandon Centre. MST-PSB aims to reduce levels of both sexual and non-sexual offending in the target population, and so offending will be used as a key measure to determine whether the intervention is effective. In addition, as out-of-home placement  represents an unhelpful outcome for antisocial adolescents in the majority of cases, the investigators have chosen to use preservation of the family , to be in line with changes in the purported mediating processes .To assess what the benefits of MST-PSB and MAU might be, care will be taken to gather accurate descriptions of the interventions delivered to both groups in the study. In addition, the investigators will attempt to chart the subjective experience of all stakeholders in the project . The data from the trial will be analysed to determine whether the expected benefit of MST-PSB is achieved and will provide initial data on whether MST-PSB represents an economically viable option for intervening with young people displaying problem sexual behaviour.We anticipate that the trial will also have positive effects on practice with this group of young people, irrespective of the trial outcomes. As noted, there is a lack of coherence within and across the systems that manage these young people; there are challenges to effectively identifying, assessing and intervening with this clinical population; and there has been very limited development of evidence-based interventions . As we dFinally, the qualitative aspects of the study, which will elicit the views of this stigmatized and marginalized group of young people, will provide valuable information about their lived experiences as sexual offenders, while the assessment of practitioners’ views specifically regarding provision of family and evidence-based treatments for this population will help us to understand the mental health context within which evidence-based services will need to develop. The qualitative data will complement the quantitative results about the treatment effectiveness of both MST-PSB and currently available services to enrich our understanding of the strengths and challenges facing mental health services for young sexual offenders in the United Kingdom.The trial is in its third year and recruitment is ongoing.


Stapled jejunocecal anastomoses are commonly performed in equine abdominal surgery. They carry higher complication rates compared to handsewn techniques. In human surgery various causes likely to lead to failure of stapled techniques have been evaluated, including staple line failure. Recently Freeman proposed a technique to perform a stapled jejunocecal anastomosis in horses while avoiding blind pouch formation. The aim of this study is to describe a method for stapled side-to-side jejunocecal anastomosis in horses and to compare it with other techniques with computed tomography to assess stomal area, shape and blind pouch size.Intestinal specimens comprising the cecum, ileum and jejunum from 18 horses were collected and were divided into three groups. In Group S a standard stapled side-to-side jejunocecal anastomosis was performed. In Group F the anastomosis was performed using a modified technique proposed by Freeman. In Group G the anastomosis was performed with a modified technique proposed by the authors. Inflated bowel segments were CT scanned to obtain a MultiPlanar Reconstruction of the stoma and afferent small intestine before calculating the cross-sectional area of each of these regions. The ratio of the measured areas was compared between the three techniques. The volume of the blind-end pouch was measured and its ratio with the intestinal area compared between techniques. The cecum was opened and the length of the stoma measured with a caliper and compared to the intended initial length.The stomal/intestinal area ratio was not significantly different between techniques.No statistically significant difference was found in the stomal ideal/real perimeter ratio.There was no statistically significant difference in the intended/real stomal length ratio, and all techniques featured an increase in stomal length ranging from 2 to 12 %. Blind pouch formation was a consistent finding in Group S and was virtually absent in Groups F and G.Both the Freeman and the new (G) technique were comparable to the standard technique in terms of stomal area, stomal shape and difference in stomal elongation. They consistently produced a smaller blind pouch and allowed easier placement of the staplers. Side-to-side jejunocecal anastomoses are commonly performed in equine abdominal surgery. Both handsewn and stapled techniques have been fully described , and havRecently Freeman proposed a new technique to perform a stapled jejunocecal anastomosis in horses while avoiding blind pouch formation, although the main focus of his work was to avoid complications caused by the stapler insertion sites . The sitThe purpose of the present study is to anatomically compare the standard stapled technique (S) with the technique described by Freeman (F) and a proposed modified technique (G) for stapled side-to-side jejunocecal anastomosis in terms of stomal area, stomal shape and blind pouch size.Intestinal specimens comprising the cecum, ileum and three meters of jejunum from 18 horses  without clinically evident intestinal disease were collected immediately after death at the Didactical Abattoir, Department of Veterinary Sciences, University of Turin, and maintained in warm Ringer lactate solution until testing, that was completed within 6 hours from collection. Bowel segments were divided into three groups of 6 specimens each: Group S underwent a standard stapled jejunocecal anastomosis [All anastomoses were performed by the same surgeon (MG) and the same assistant (GG) in order to reduce variability between the techniques. All anastomoses were placed between the dorsal and the medial band of the cecum, level with the cecocolic fold , approxiIn the modified technique (Group G) the jejunum was not cut. It was opposed to the cecal body wall with one stay suture placed 5 mm deep  to the aBowel segments were then connected to a manometer  as previously described  and inflThe images were acquired using a single slice Computed Tomography (CT) unit in axial mode using a slice thickness of 1mm, a matrix size of 512x512, a medium smooth reconstruction algorithm (defined as CHST by the CT software), 120kVp and 130mAs. The images were transferred to the visualization workstation and viewed with standardized windowing parameters . The Osirix software was then used to calculate the volume and area of the blind pouch. Three-dimensional MultiPlanar Reconstructions (MPR) of the stoma were exported as 16-bit Monochrome DICOM  lossless images, for use in the ImageJ measurement software. Four stomas were not monoplanar and required the use of thick-slice MPR reconstructions using volume rendering . Using the wand tool  the stomal area and perimeter were calculated on the MPR images, while the proximal jejunal area and perimeter were calculated on the original images.For each specimen, a ratio was obtained by dividing the stomal area by the jejunal area.We calculated the perimeter of an ideal circle of the same area as the stoma and then obtained a ratio by dividing this ideal value by the real one. This value gives an indication of the shape of the stoma. A ratio of 1 indicates a perfectly circular stoma.After the scan was completed, the cecum was opened at a site distant from the anastomosis and the effective length of the stoma was measured with a caliper and compared to the intended initial length. The intended initial stomal length is the length of the stoma that the procedure should have formed based on the choice of the length of the stapler. A percentage of variation was obtained by comparing the length of the stoma measured directly with a caliper on the specimen at the time of incision and after deflation following image acquisition.The volume of the blind pouch was calculated in Osirix by 2D region growing followed by ROI volume calculation using a PowerCrust algorithm. The resulting value was then divided by the area of the afferent jejunum.Using the R software (version 2.15.1 32-bit) we measured the normality of each group using a Shapiro-Wilk normality test and obtained means, standard deviation, median and interquartile ranges.Our work aims to ascertain the presence of statistically significant differences between various parameters of the anastomoses performed using these three techniques. The considered parameters were blind pouch volume/area ratio, stomal/jejunal area ratio, ideal/real perimeter ratio and percentage of stomal length variation.We used Pairwise Student's T-Test on normally distributed data  and Pairwise Wilcoxon's Rank Sum Test on non-normally distributed data  with significance set for p< 0.05.Results are summarized in Table The stomal/intestinal area ratio was not significantly different between techniques. No statistically significant difference was found in the stomal ideal/real perimeter ratio, although there was a trend towards a more circular shape in Group S. The technique in Group F resulted in the formation, upon inflation, of two folds in the jejunal wall placed internally to each row of staples Fig. . There wJejunocecal anastomoses in horses carry a higher risk of complications than other anastomotic techniques and, amongst them, stapled anastomoses carry higher complication rates compared to handsewn techniques . For thiin vitro to two other previously described techniques from an anatomical point of view. When compared to the handsewn technique, the modified version proposed by Freeman [in vivo cannot be easily established. The stomal/intestinal area ratio was higher in Group F than with the other two techniques, demonstrating that an 80 mm long stapler is able to produce a stoma as wide as the proximal intestine feeding into it in 400 kg horses. Also, there was no statistically significant difference in the stomal ideal/real perimeter ratio, although there was a trend to a more “circular” shape in Group S.In our study we proposed a modified stapled technique and compared it  Freeman  is repor Freeman , In our  Freeman , althougBy producing the two folds axially to the staple line, the F technique had a lower ideal/real perimeter ratio when compared to the other techniques. Although the difference was not statistically significant, the formation of these folds could decrease stomal compliance to the passage of ingesta, possibly participating in increasing the complication rate of this technique.All techniques exhibited an increase in the final stomal length, ranging from 2 to 12 % longer than the intended initial stomal length, and this datum is in accordance with other studies . This fiBlind pouch formation is a common complication of side-to-side anastomoses, both handsewn or stapled ,8-18, thIn the G technique, cutting the jejunum and closing the stump after stapling also removed the problem of placing the enterotomies in the correct position for stapler insertion. This is a crucial point in performing a functional anastomosis. The same goal can certainly be accomplished by a second application and firing of the linear cutting stapler or using a linear stapler (e.g. Ta-90) transversely on the jejunal stump . We prefWe performed the modified Parker-Kerr suture while the stapler was still in place because it avoided straining the staple line by holding the two bowel segments together. Removing the stapler before performing the second suture could further reduce the formation of the blind end, but we think modifying the stapler’s anvil tip accomplished the same goal. If the anvil tip is not removed then the removal of the stapler soon after firing will be necessary in order to reduce the blind pouch. We had the impression that closing the distal end of the jejunum without the stapler supporting the staple line would put sufficient strain on the staple line to disrupt it, so we preferred to avoid this risk by leaving the stapler in place until completion of the Parker-Kerr suture.The main advantage of the G technique versus the F technique is the possibility of placing the stapler in a proximo-distal direction. In fact, relative placement of the stoma onto the cecal body could be another issue that influences the performance of jejunocecal anastomoses. In the Freeman technique performed in vivo, having the surgeon insert the stapler disto-proximal might be a problem, possibly leading to an excessively apical placement of the stoma compared to the other techniques since the abdominal incision is in the way, forcing the stoma to be made too close to the apex. With the G and S techniques this is not an issue, although in our study this aspect might not have been fully evaluated because all the stoma were placed at the same distance from the ileocecal valve.The oversewing of staple lines has been proposed in stapled anastomoses, although this operation could remove the advantage offered by this type of technique in terms of reduced surgical time ,20, and in vivo compared to the former because of the proximo-distal insertion of the stapler. For these reasons the F and G techniques should be preferred to the standard technique, although further in vivo studies are needed.Both the Freeman and the G techniques were comparable to the standard technique in terms of stomal area, shape and difference in stomal elongation. They consistently produced a significantly smaller blind pouch and made proper placement of the staplers easier. The latter might prove easier to perform The authors declare that they have no competing interestsMG and GG developed the technique, studied the experimental protocol, performed the surgeries, participated in the CT acquisition, collected data, wrote and reviewed the paper.BI participated in the CT acquisition, performed all the image data analysis and statistics, wrote and reviewed the paper.FS and AV performed the CT acquisitions, participated in reviewing the paper.


It is increasingly clear that we extract patterns of temporal regularity between events to optimize information processing. The ability to extract temporal patterns and regularity of events is referred as temporal expectation. Temporal expectation activates the same cerebral network usually engaged in action selection, comprising cerebellum. However, it is unclear whether the cerebellum is directly involved in temporal expectation, when timing information is processed to make predictions on the outcome of a motor act. Healthy volunteers received one session of either active  or sham repetitive transcranial magnetic stimulation covering the right lateral cerebellum prior the execution of a temporal expectation task. Subjects were asked to predict the end of a visually perceived human body motion (right hand handwriting) and of an inanimate object motion (a moving circle reaching a target). Videos representing movements were shown in full; the actual tasks consisted of watching the same videos, but interrupted after a variable interval from its onset by a dark interval of variable duration. During the ‘dark’ interval, subjects were asked to indicate when the movement represented in the video reached its end by clicking on the spacebar of the keyboard. Performance on the timing task was analyzed measuring the absolute value of timing error, the coefficient of variability and the percentage of anticipation responses. The active group exhibited greater absolute timing error compared with the sham group only in the human body motion task. Our findings suggest that the cerebellum is engaged in cognitive and perceptual domains that are strictly connected to motor control. The traditional view of the cerebellum as a structure exclusively engaged in motor control has been largely modified in recent years, with increasing evidence pointing towards an involvement of cerebellar circuits in several domains pertaining to cognition and emotion –4. AmongHowever, it is still an open question whether the cerebellum might be involved in temporal expectation, when timing information is processed to make predictions on the outcome of a motor act, in the absence of self-action. By applying a conditioning rTMS train on cerebellar regions, it is possible to induce a transient modulation of the activity of the cerebellar cortex, hence to investigate whether changes in cerebellar neural activity would interfere with a specific task. Inhibitory rTMS (1Hz-rTMS) over lateral cerebellum has been used to explore the role of cerebellum in motor timing –17.As in our previous study  healthy Twenty-six subjects without any history of neurological or psychiatric disease  were recruited for the study and divided in two age- and gender-matched groups. The first group  performed the perceptual temporal expectation task immediately after receiving an active inhibitory rTMS train of stimuli on the right lateral cerebellum, whereas the second group  performed the task immediately after receiving sham rTMS stimulation on the same area. Importantly, a between-subjects design was preferred to a within-subjects design, in order to avoid a learning effect on the task performance. All subjects were right-handed. All participants gave their written informed consent prior to their inclusion in this study. The experimental protocol was approved by the ethics committee of the University of Genoa and was carried out in agreement with legal requirements and international norms .The experimental paradigm consisted of one session assessing temporal expectation of human body segment versus inanimate object motion. This task has been described elsewhere . Brieflytarget and control task), each condition  was administered six times in a randomized order, for 36 trials per experimental session. Handwriting (target task) and inanimate object motion (control task) were presented in separate blocks; the order of presentation of the blocks was random across subjects. The experimental design is summarised in Thus, participants used their knowledge of velocity of motion to extrapolate duration of the movement of the body segment (or object) represented in the video. For the sake of the analysis, we refer to the period spanning from the beginning of the dark interval to the subject’s response as ‘reproduced interval’, whereas we refer to the real duration of the dark interval as ‘target interval’. Participants did not receive any feedback on interval duration or their performance throughout the whole session. Three different dark intervals  were used. Separately for each task  connected to a figure-of-eight coil (wing diameters of 90 mm) positioned over the right lateral cerebellum (3 cm right and 1 cm inferior to the inion). The coil was positioned tangentially to the scalp with the handle pointing up. The current in the coil was directed upwards, which induced a downward current in the cerebellar cortex . The scaA single train  of 1Hz-rTMS was delivered at 90% of the motor threshold (MT) on right cerebellum. For sham rTMS the coil was positioned over the same scalp site, but angled away (90-degrees) so that no current was induced in the brain.The MT was determined as the minimum stimulus intensity capable of eliciting a motor evoked potential (MEP) of at least 50 μV in the contralateral first dorsal interosseus (FDI) muscle in five or more of ten consecutive stimulations. EMG was recorded with silver disc surface electrodes placed in a tendon belly arrangement over the FDI muscle. The ground electrode was placed at the wrist. EMG signals were amplified and filtered (20 Hz to 1 kHz) with a D360 amplifier . The signals were sampled at 5000 Hz, digitized using a laboratory interface  and stored on a personal computer for display and later off-line data analysis.Performance on the temporal expectation task was analysed measuring the timing error, the normalized absolute value of timing error, the coefficient of variability and the percentage of anticipation responses. These parameters allowed us to describe the ability in time processing determining the magnitude  and the variability (coefficient of variability) of the timing error, as well as the strategy adopted by subjects during time prediction (timing error and percentage of anticipation responses).The timing error was defined as the time elapsing between the subject’s response and the real end of the video, and had a negative value when the response preceded the real end of the video, and positive when the response was delayed in respect to the real end of the video. The absolute value of this error was then normalized with respect to the corresponding target interval and was expressed as a percentage. This parameter provides a measure of the accuracy of subjects in estimating the corresponding target interval: the greater the absolute error, the greater the error in temporal judgement regardless of its direction.The coefficient of variability was calculated as Standard Deviation (SD)/mean of the reproduced intervals, representing an index of performance variability in temporal judgement. Finally, the percentage of anticipation responses  represents the number of responses in which subjects pressed the space button before the real end of the movement (negative error). Together with the timing error, this parameter gives information on the tendency to under- or over-estimate the target interval.Timing parameters  were analysed by means of a repeated measures ANOVA with group (1Hz-rTMS and Sham) as between subject factor and task  and target interval  as within- subjects factors. Post hoc analysis of significant interactions was performed by means of t-tests applying the Bonferroni correction for multiple comparisons where necessary. P-values of < 0.05 were considered as threshold for statistical significance. Statistical analysis was performed with SPSS 13.0.The performance of the temporal expectation task was influenced by the active 1Hz-rTMS on right lateral cerebellum, as evidenced by significant changes in the absolute timing error parameter.In the presence of 1Hz-rTMS on the lateral right cerebellum, participants exhibited a greater normalized absolute timing error, at all target intervals in the human body motion task (handwriting) with respect to sham . AccordiVariability data are reported in Timing error data are reported in The results on percentage of responses in anticipation paralleled those on timing error. The percentages of response in anticipation for both target and control tasks are reported in In the present study we showed that the inhibition of lateral cerebellum activity, by means of 1Hz-rTMS, induced a worsening in timing performance when participants were engaged in a perceptual temporal expectation in which they had to make a deliberate estimate of the duration of a perceived common movement of a body segment (i.e. handwriting). Differently, we did not observe any effect of 1Hz-rTMS when subjects had to predict the outcome of a movement involving an inanimate object .Confirming our previous observations, apart from a specific effect of inhibitory 1Hz-rTMS, the ability to estimate the time point at which the visually perceived movement ended was influenced by the duration of the time interval in which subjects had to rely only on their mental representation of movement  and by the type of motion perceived.Shorter dark intervals were associated with higher tendency to shift the end of the perceived movement ahead in time , with greater variability of prediction response and greater normalized absolute timing error. These results, in accordance with what observed in a previous work of our group , lend suFurther, higher frequency of anticipated responses and a more negative timing error were observed on the inanimate object motion task compared with the human body motion task. Again, this finding is in accordance with what observed in a previous work of our group , and is Temporal expectations make use of timing information in order to optimise motor or perceptual performance . For exaIn agreement with the role of cerebellum in processing and adapting motor related information, it has been shown that the cerebellum is needed to optimize self-action by recalibrating predictions capturing the sensory consequences of one’s actions –32. HoweIn the present study, we wanted to expand this topic by exploring the role of cerebellum in predictions related to a visually perceived motor act . We found that subjects’ precision in in temporally predicting the end of a human movement  was influenced by 1Hz-rTMS disruption, whereas the variability in temporal expectation performance and the direction of the timing error (expressed by both the timing error and the percentage of responses in advance) were not influenced by 1Hz-rTMS disruption.In self-action it is well known that cerebellum is involved in predictive control of movement . The terHere, we demonstrated for the first time that cerebellum is involved in predictive movement control in absence of self-action; i.e., during observation of a motor act, thus confirming recent theories supporting a cognitive-perceptual role of the cerebellum . In accoThe perceptual predictive movement control operated by the cerebellum might be implicated in motor planning and its disruption might contribute to some movement abnormalities exhibited by patients with cerebellar damage, like lack of coordination, and poor accuracy . HoweverIntriguingly, when cerebellar activity was disrupted by inhibitory 1Hz-rTMS we did not observe any modification in time estimation when the perceived movement was that of a moving object . Although this does not exclude a cerebellar involvement in predicting the outcome of a moving object , it neveIn addition, there are some methodological characteristics of the study conducted by O’Reilly and co-workers that could partially justify this discrepancy, like the presence of an occluder, the type of task , the type of participants response  .Another noteworthy aspect of our results is that cerebellar disruption led to an impact on supra-second intervals. Most studies available in the literature suggest that the cerebellum is involved in sub-second timing, although some lesion studies suggest that the cerebellum is also involved in supra-second intervals . NotablyIt’s worthy to note that although we demonstrated that lateral cerebellum is directly involved in a temporal expectation task whose goal is to predict the temporal outcome of a motor act, we can only do some speculation about the cortical areas that may be recruited in this perceptual-motor ability. Indeed, given the cerebellar connectivity with cortical regions engaged in higher-order perceptual processing and spatiotemporal awareness  ,49, we mOne might argue that changes in motor performance, induced by cerebellar 1Hz-rTMS might have influenced the performance on the temporal expectation task. When applied to lateral cerebellum, 1Hz-rTMS is known to induce a disruption of cerebellar outputs to the contralateral motor cortex (M1). Low-frequency right cerebellar stimulation has been demonstrated to induce a decrease in the excitability of intracortical facilitatory circuits in left M1, likely due to the reduction of excitatory drive of cerebellar nuclei to the motor cortex, thus reflecting a suppressive effect of low-frequency rTMS on cerebellar Purkinje cells . FurtherWe propose that the cerebellum is directly implicated in temporal expectation, when timing information is processed in order to make accurate predictions on the outcome of an observed motor act. This result enlarges the current view of the cerebellum as a “non-motor” cerebral structure, engaged in cognitive and perceptual domains. However, the higher susceptibility of cerebellar activity to 1Hz-rTMS disruption for time estimation of a human motor act, rather than that of a moving object , suggests that cognitive and perceptual functions of cerebellum are likely to be strictly connected to motor control.


The Local Field Potential (LFP) is typically assumed to mainly stem from synaptic inputs and their subthreshold dendritic processing. The role of active dendritic conductances in shaping the LFP has often been ignored, even though several ion channels are active in the subthreshold voltage regime. Here we use biophysical modeling to investigate the impact of active conductances and their spatial distributions on the LFP in the subthreshold voltage regime.We used models of neocortical pyramidal neurons with realistic morphologies and systematically varied the spatial distributions of active conductances and synaptic inputs. We linearized the active conductances -- yielding the so-called quasi-active description -- thereby reducing the number of model parameters and highlighting that active currents can be divided into two classes: regenerative and restorative. In terms of the filtering properties of the cell membrane, regenerative conductances amplify low frequencies of the synaptic input current, whereas restorative conductances dampen them.The low-frequency components of the synaptic input current give the largest contributions to the LFP, due to their larger distance between synaptic currents and associated return currents. Since active conductances can enhance or diminish a neuron's response to low frequencies, they are also expected to affect the LFP.We indeed found that the effect of active conductances on the single-neuron LFP is strongest for the lowest frequencies (< 10-30 Hz). The impact was most pronounced when 1) the synaptic input is asymmetric , 2) the active conductances are distributed non-uniformly with the highest channel densities near the synaptic input, and 3) when the LFP is measured at the opposite pole of the cell relative to the synaptic input. Restorative conductances are typically expressed in the LFP as resonance peaks. However, these are considerably smaller than the corresponding peaks for the membrane potentials and transmembrane currents.H, can have a sizable impact on the shape of the LFP, since it is a restorative current, strong in the apical dendrite, and active at subthreshold voltages.We compared the findings from our quasi-active models with published, experimentally constrained pyramidal cell models that display various active conductances. We found that in particular the hyperpolarization-activated inward current, IWe conclude that subthreshold contributions of active currents to the LFP of detailed pyramidal cell models can be very well captured by quasi-active models containing one or two voltage-dependent currents, and that their contribution can be a major factor in shaping the LFP for the lowest frequencies.


A series of five cases were presented in which similar fractures of the shaft of the humerus occurred during the hand grenade throwing activity during the military education. All the fractures were in the 1/3 distal humeral shaft, and butterfly fragments were accompanying in two soldiers. All the fractures healed without any clinical complications with conservative treatment. The mechanism of the fracture is discussed with reference to the recent literature. Humerus fractures are generally secondary to the direct trauma , 2. Frac In the present study, spiral humeral shaft fractures are secondary to the hand grenade throwing in five military recruits. The causes of these fractures and the literature related to the hand grenade throwing were also reviewed.Between August 2008 and January 2009, 5 male military recruits were admitted to the Emergency Department of Ağrı Military Hospital with the right humerus shaft fractures during hand grenade throwing training period. Average patient age was 20.2 years (range 19–22). All the patients were right-handed, and none of them had an experience in throwing sports before their military obligation. The recruits reported that they used the maximum strength when throwing the hand grenade. According to their history, all the fractures occurred just before the hand grenade release.All fractures were closed and extra-articular. All the fractures were at the junction of the middle and distal third of the humeral shaft . Two of All patients were admitted to the clinic on the day of injury. Initial fracture stabilization was achieved with U-splint and the Velpeau bandage for all patients. Patients were systematically examined for accompanying any musculoskeletal disease. Because pathologic fracture was not suspected on plain radiographs, any further imaging techniques were not performed. On radiographs, average varus-valgus angulation was 12′ –15, and There are several cases of humerus fracture during the throwing activity. Several theories exist for the cause of this type of fracture, including uncoordinated muscular antagonism, lack of a regular exercise program, inadequate throwing technique, and muscle fatigue , 9. And, Allen reported one humeral shaft fracture that resulted from pitching. He described this as the “window of vulnerability” during the early stages of bone remodeling . Branch et al. reported 12 humeral shaft fractures in 12 baseball players, and they thought that the lack of exercise period and prolonged layoff periods are the main causes for this type of fractures . Ogawa and Yoshida suggested that this type of humerus fractures is secondary to the practice limitations, and they presented this as an external rotation fracture in 90 baseball players [ Throwing humeral shaft fractures can be secondary to the stress loading as called stress fracture. At this situation, there must be pain at the rest, and, also, there must be a history of repeating exercise program. Throwers who had fracture with no previous experience of throwing generally had no prodromal pain as in this study. The fracture site and type support the suggestions of Ogawa and Yoshida that these fractures occur due to mainly external rotation force on the distal humerus at the acceleration throw phase, as the proximal end internally rotates . In military education of hand grenade throwing, recruits are told to extend the elbow during the entire acceleration phase. This style does not create an external rotation force on the distal humerus, and only the proximal end internally rotates. In a faulty throwing style, if the elbow is flexed at the early acceleration throw phase, the distal humerus is exposed to external rotation force as the elbow is extended at the late acceleration phase. This antagonism of rotational torques, because of faulty throwing style, is the main cause of throwing fractures in military recruits . Malignant metastatic tumours are the most neoplasms of bone. These lesions whether metastatic or primary or malignant or benign can be the reason for humeral pathological fractures. As humerus is the second most frequently involving metatstatic lesions, patients ages generally are over 50 years old . Benign  In extra-articular humerus shaft fracture treatment modalities, the first choice is nonsurgical treatment methods; we used to begin the treatment with U-splint and the Velpeau bandage for 3 weeks, and after then, we applied functional brace as the Sarminento brace. This treatment method has low morbidity and high success rate. Rotational deformities decrease at the fracture site by contraction of the flexors and extensors . Angulat Whatever the condition of the humeral shaft before an overt fracture, the true cause of throwing fracture of the humerus is a rotational force at the acceleration phase of the throw. Almost all the throwing fractures, as in this present study, are spiral fractures, with or without butterfly fragment, and the course of the fracture line shows that they are external rotation fractures. The fracture site usually is at the junction of the middle and distal third of the humerus –10. Cons