The mammalian DNA-damage response (DDR) has evolved to protect genome stability and maximize cell survival following DNA-damage. One of the key regulators of the DDR is p53, itself tightly regulated ...by MDM2. Following double-strand DNA breaks (DSBs), mediators including ATM are recruited to the site of DNA-damage. Subsequent phosphorylation of p53 by ATM and ATM-induced CHK2 results in p53 stabilization, ultimately intensifying transcription of p53-responsive genes involved in DNA repair, cell-cycle checkpoint control and apoptosis.
In the current study, we investigated the stabilization and activation of p53 and associated DDR proteins in response to treatment of human colorectal cancer cells (HCT116p53+/+) with the MDM2 antagonist, Nutlin-3.
Using immunoblotting, Nutlin-3 was observed to stabilize p53, and activate p53 target proteins. Unexpectedly, Nutlin-3 also mediated phosphorylation of p53 at key DNA-damage-specific serine residues (Ser15, 20 and 37). Furthermore, Nutlin-3 induced activation of CHK2 and ATM - proteins required for DNA-damage-dependent phosphorylation and activation of p53, and the phosphorylation of BRCA1 and H2AX - proteins known to be activated specifically in response to DNA damage. Indeed, using immunofluorescent labeling, Nutlin-3 was seen to induce formation of γH2AX foci, an early hallmark of the DDR. Moreover, Nutlin-3 induced phosphorylation of key DDR proteins, initiated cell cycle arrest and led to formation of γH2AX foci in cells lacking p53, whilst γH2AX foci were also noted in MDM2-deficient cells.
To our knowledge, this is the first solid evidence showing a secondary role for Nutlin-3 as a DDR triggering agent, independent of p53 status, and unrelated to its role as an MDM2 antagonist.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
ABSTRACT
Introduction: In this study we aimed to characterize muscle composition of the medial gastrocnemius in children with spastic cerebral palsy (SCP) using quantitative ultrasound. Methods: ...Forty children with SCP, aged 4–14 years, participated in this study. Children were grouped according to the gross motor function classification system (GMFCS I–V) and compared with a cohort of age‐ and gender‐matched, typically developing children (TD; n = 12). Ultrasound scans were taken of the medial gastrocnemius. Images were then characterized using grayscale statistics to determine mean echo intensity (EI) and the size and number of spatially connected homogeneous regions (i.e., blobs). Results: Significant differences in skeletal muscle composition were found between children with SCP and their TD peers. Children classified as GMFCS III consistently exhibited the highest EI and blob area. Conclusions: This study demonstrates altered tissue composition in children with SCP visualized using ultrasound. Further work is required to determine the pathophysiology contributing to these alterations in SCP. Muscle Nerve 52:397–403, 2015
ABSTRACT
Introduction: In this study we aimed to determine the lower limb morphological characteristics of skeletal muscle of ambulant children with spastic cerebral palsy (CP) and typically ...developing (TD) children. Methods: Seventeen children with spastic diplegic CP (10 boys and 7 girls, 5–12 years of age, Gross Motor Function Classification System GMFCS level I or II) and 19 TD children (8 boys and 11 girls, 5–11 years of age) underwent lower limb T1‐weighted MRI. Morphological characteristics of the triceps surae, including muscle volume, anatomical cross‐sectional area, muscle length, and subcutaneous adipose tissue, were digitally quantified, and the proportional distribution calculated. Results: Children with GMFCS II had significantly reduced muscle volume, cross‐sectional area, and muscle length, and increased subcutaneous fat compared with TD children. Children classified as GMFCS II consistently exhibited the greatest deficits in all morphology variables. Discussion: Morphological variables were significantly different between the groups. These alterations have the potential to influence the functional capabilities of the triceps surae muscle group. Muscle Nerve 58:818–823, 2018
The implementation of an intervention protocol aimed at increasing vocal complexity in three pre-linguistic children with cerebral palsy (two males, starting age 15 months, and one female, starting ...age 16 months) was evaluated utilising a repeated ABA case series design. The study progressed until the children were 36 months of age. Weekly probes with trained and untrained items were administered across each of three intervention blocks. Successive blocks targeted more advanced protophone production and speech movement patterns, individualised for each participant. Positive treatment effects were seen for all participants in terms of a greater rate of achievement of target protophone categories and speech movement patterns. Tau coefficients for trained items demonstrated overall moderate to large AB phase contrast effect sizes, with limited evidence of generalisation to untrained items. Control items featuring protophones and speech movements not targeted for intervention showed no change across phases for any participant. Our data suggest that emerging speech-production skills in prelinguistic infants with CP can be positively influenced through a multimodal intervention focused on capitalising on early periods of plasticity when language learning is most sensitive.
The aim of this study is to contribute to the knowledge base on the long-term outcomes of evidence-based medical interventions used to improve gross motor function in children and adolescents with ...Cerebral Palsy.
Prospective cohort study of children with Cerebral Palsy in the birth years 2000-2009 attending a tertiary level service for children with Cerebral Palsy who's first recorded Gross Motor Function Classification System level was II.
A total of 40 children were eligible for the study, of whom 28 (72.7%) enrolled. The Botulinum toxin A treatment for this cohort, (median and interquartile ranges) were: total number of lower limb Botulinum toxin A injections 11 (6.7, 5.5); total dose of Botulinum Toxin A per lower limb treatment 6.95 u/kg (4.5, 11); and dose of Botulinum Toxin u/kg/muscle 2.95 (2.2, 4). For all 28 subjects there was a median of 15 (8.5 to 22) Gross Motor Function Classification System level recordings: six of the 28 children (21.4%) improved from level II to level I, the remaining 22 children remained stable at level II (78.6%). In this highly treated population, the average 66 item Gross Motor Function Measure score for the 22 children in level II was 72.55, which is consistent with the mean of 68.5 reported in the original Ontario cohort.
This cohort study has confirmed that children with Cerebral Palsy, Gross Motor Function level II treated at a young age with repeated doses of Botulinum Toxin A within an integrated comprehensive service, maintain or improve their functional motor level at a later age.
IntroductionFor children with cerebral palsy (CP), who are marginally ambulant, gross motor capacity peaks between 6 and 7 years of age with a subsequent clinical decline, impacting their ability to ...engage in physical activity. Active Strides-CP is a novel package of physiotherapy targeting body functions, activity and participation outcomes for children with bilateral CP. This study will compare Active Strides-CP to usual care in a multisite randomised waitlist-controlled trial.Methods and analysis150 children with bilateral CP (5–15 years), classified in Gross Motor Function Classification System (GMFCS) levels III and IV will be stratified (GMFCS III vs IV, age 5–10 years; 11–15 years and trial site) and randomised to receive either (1) 8 weeks of Active Strides-CP two times/week for 1.5 hours in clinic and one time/week for 1 hour alternating home visits and telehealth (total dose=32 hours) or (2) usual care. Active Strides-CP comprises functional electrical stimulation cycling, partial body weight support treadmill training, overground walking, adapted community cycling and goal-directed training. Outcomes will be measured at baseline, immediately post-intervention at 9 weeks primary endpoint and at 26 weeks post-baseline for retention. The primary outcome is the Gross Motor Function Measure-66. Secondary outcomes include habitual physical activity, cardiorespiratory fitness, walking speed and distance, frequency/involvement of community participation, mobility, goal attainment and quality of life. Analyses will follow standard principles for randomised controlled trials using two-group comparisons on all participants on an intention-to-treat basis. Comparisons between groups for primary and secondary outcomes will be conducted using regression models. A within-trial cost utility analysis will be performed.Ethics and disseminationThe Children’s Health Queensland Hospital and Health Service, The University of Queensland, The University of Melbourne and Curtin University Human Research Ethics Committees have approved this study. Results will be disseminated as conference abstracts and presentations, peer-reviewed articles in scientific journals, and institution newsletters and media releases.Trial registration numberACTRN12621001133820.
ObjectivesTo determine the feasibility of an intensive interdisciplinary programme in improving goal and motor outcomes for preschool-aged children with non-progressive neurodisabilities. The primary ...hypothesis was that the intervention would be feasible.DesignA single group feasibility study.SettingAn Australian paediatric community therapy provider.ParticipantsForty children were recruited. Inclusion criteria were age 2–5 years with a non-progressive neurodisability, Gross Motor Function Classification System (GMFCS) levels III–V or equivalent, and goals relating to mobility, communication and upper limb function. Exclusion criteria included orthopaedic surgery in the past 6 months, unstable hip subluxation, uncontrolled seizure disorder or treadmill training in the past month.InterventionA goal-directed programme of three 2-hour sessions per week for 4 weeks (24 hours total). This consisted of treadmill and overground walking, communication practice, and upper limb tasks tailored by an interdisciplinary team.Primary and secondary outcome measuresLimited-efficacy measures from preintervention (T1) to postintervention (T2) and 4-week follow-up (T3) included the Goal Attainment Scaling (GAS), Canadian Occupational Performance Measure (COPM), Gross Motor Function Measure (GMFM-66) and 10-Metre Walk Test (10MWT). Acceptability, demand, implementation and practicality were also explored.ResultsThere were improvements at T2 compared with T1 for all limited-efficacy measures. The GAS improved at T2 (mean difference (MD) 27.7, 95% CI 25.8 to 29.5) as well as COPM performance (MD 3.2, 95% CI 2.8 to 3.6) and satisfaction (MD 3.3, 95% CI 2.8 to 3.8). The GMFM-66 (MD 2.3, 95% CI 1.0 to 3.5) and 10MWT (median difference −2.3, 95% CI −28.8 to 0.0) improved at T2. Almost all improvements were maintained at T3. Other feasibility components were also demonstrated. There were no adverse events.ConclusionsAn intensive interdisciplinary programme is feasible in improving goal and motor outcomes for preschool children with neurodisabilities (GMFCS III–V or equivalent). A randomised controlled trial is warranted to establish efficacy.Trial registration numberACTRN12619000064101.
Abstract
Purpose: To investigate the muscle size-strength relationship of the knee flexors and extensors in children with spastic cerebral palsy (CP) in relation to typically developing children ...(TD). Methods: Eighteen children with spastic Diplegia, Gross Motor Function Classification System I-III (mean 7 y 5 mo SD 1 y 7 mo) and 19 TD children (mean 7 y 6 mo SD 1 y 9 mo) participated. Muscle volume (MV) and anatomical cross-sectional area (aCSA) were assessed using MRI. Measures of peak torque (PT) and work of the knee flexors and extensors were assessed isometrically and isokinetically using a Biodex dynamometer, and normalised to bodymass (Bm). Results: Children with CP were weaker than their TD peers across all torque variables (p < 0.05). MV and aCSA of the knee flexors (MV: p = 0.002; aCSA: p = 0.000) and extensors (MV: p = 0.003; aCSA: p < 0.0001) were smaller in children with CP. The relationship between muscle size and strength in children with CP was weaker than the TD children. The strongest relationship was between MV and isometric PT/Bm for TD children (r = 0.77-0.84), and between MV and isokinetic work (r = 0.70-0.72) for children with CP. Conclusions: Children with CP have smaller, weaker muscles than their TD peers. However, muscle size may only partially explain their decreased torque capacity. MV appears to be a better predictor of muscle work in children with CP than aCSA. This is an important area of research particularly in regard to treatment(s) that target muscle and strength in children with CP.Implications for RehabilitationThis research adds to the evidence that children with CP have smaller, weaker knee flexor and extensor muscles than their TD peers. However, unlike their TD peers, muscle size does not necessarily relate to muscle strength.The weak correlation between MRI-derived muscle volume and isometric peak torque suggests children with CP are underpowered relative to their muscle size.For children with CP, muscle volume appears to be the best predictor of isokinetic muscle torque output. Therefore, when assessing the capacity of a muscle, it appears preferable to measure total muscle volume and torque development through a range of motion (isokinetic strength).
Introduction:
The purpose of this study was to establish the nature and stability of the strength‐size relationship for the knee flexors and extensors across a 6‐month period of childhood growth.
...Methods:
Nineteen typically developing children aged 5–11 years underwent lower limb magnetic resonance imaging (MRI) and dynamometry strength assessments on 2 occasions, 6 months apart. Muscle volume (MV) and maximum anatomical cross‐sectional area (aCSA) for the knee flexors and extensors were determined using MRI analysis software. Isokinetic dynamometry determined corresponding isometric and isokinetic strength.
Results:
Strong correlations were found between muscle size and strength for both the knee flexors and extensors (r = 0.84–0.90; P < 0.01). Furthermore, the ratio of strength to muscle size remained consistent across 6 months of prepubescent growth.
Conclusions:
Increases in thigh muscle strength were relative to those in muscle size, suggesting that muscle growth may play an important role in the development of strength during childhood. Muscle Nerve 46: 360–366, 2012
We compared early vocal development in children "at risk" for cerebral palsy (CP) with typically developing (TD) infants aged 6 to 15 months using the SAEVD-R, investigating potential pre-linguistic ...markers of communication impairment. Additionally, we sought to examine the agreement between the SAEVD-R and IMP, which uses parent report, in identifying departure from typical vocal development in at-risk infants.
Utilising a longitudinal cohort study, >10,000 vocalisations of 33 infants (15 at risk for CP and 18 TD) were assessed at 6, 9, 12, and 15 months using the SAEVD-R. Generalised linear mixed models (GLMMs) compared groups, and Spearman correlations explored IMP ceiling scores and SAEVD-R measures.
At 6 months, both TD and CP groups reached SAEVD-R vocalisation level 3 (expansion). By 9 months, 51% of TD infants progressed to advanced babbling (levels 4 and 5), while 80% of at-risk infants remained at level 3. At 12 and 15 months, over 90% of TD children advanced, compared to 67% at 12 months and 53% at 15 months for at-risk infants, who stayed at the pre-canonical stage. Strong correlations were found between IMP scores and vocalisation levels at 9 and 12 months. Remaining at the pre-canonical stage at 12 months correlated with delayed vocal development as per IMP scores.
TD infants achieved higher SAEVD-R levels than at-risk infants. At 12 months, IMP scores effectively identified infants with speech-like vocalisation difficulties, demonstrating its clinical utility in identifying atypical vocal development in infants at risk for CP.
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