Design considerations in a clinical trial of a cognitive behavioural intervention for the management of low back pain in primary care : Back Skills Training Trial

Background Low back pain (LBP) is a major public health problem. Risk factors for the development and persistence of LBP include physical and psychological factors. However, most research activity has focused on physical solutions including manipulation, exercise training and activity promotion. Methods/Design This randomised controlled trial will establish the clinical and cost-effectiveness of a group programme, based on cognitive behavioural principles, for the management of sub-acute and chronic LBP in primary care. Our primary outcomes are disease specific measures of pain and function. Secondary outcomes include back beliefs, generic health related quality of life and resource use. All outcomes are measured over 12 months. Participants randomised to the intervention arm are invited to attend up to six weekly sessions each of 90 minutes; each group has 6–8 participants. A parallel qualitative study will aid the evaluation of the intervention. Discussion In this paper we describe the rationale and design of a randomised evaluation of a group based cognitive behavioural intervention for low back pain

Altered H19/miR‐675 expression in skeletal muscle is associated with low muscle mass in community‐dwelling older adults

Background: Despite increasing knowledge of the pathogenesis of muscle ageing, the molecular mechanisms are poorly understood. Based on an expression analysis of muscle biopsies from older Caucasian men, we undertook an in-depth analysis of the expression of the long non-coding RNA, H19, to identify molecular mechanisms that may contribute to the loss of muscle mass with age. Methods: We carried out transcriptome analysis of vastus lateralis muscle biopsies from 40 healthy Caucasian men aged 68–76 years from the Hertfordshire Sarcopenia Study (HSS) with respect to appendicular lean mass adjusted for height (ALMi). Validation and replication was carried out using qRT-PCR in 130 independent male and female participants aged 73–83 years recruited into an extension of the HSS (HSSe). DNA methylation was assessed using pyrosequencing. Results: Lower ALMi was associated with higher muscle H19 expression (r2 = 0.177, P < 0.001). The microRNAs, miR-675-5p/3p encoded by exon 1 of H19, were positively correlated with H19 expression (Pearson r = 0.192 and 0.182, respectively, P < 0.03), and miR-675-5p expression negatively associated with ALMi (r2 = 0.629, P = 0.005). The methylation of CpGs within the H19 imprinting control region (ICR) were negatively correlated with H19 expression (Pearson r = −0.211 to −0.245, P ≤ 0.05). Moreover, RNA and protein levels of SMAD1 and 5, targets of miR-675-3p, were negatively associated with miR-675-3p (r2 = 0.792 and 0.760, respectively) and miR-675-5p (r2 = 0.584 and 0.723, respectively) expression, and SMAD1 and 5 RNA levels positively associated with greater type II fibre size (r2 = 0.184 and 0.246, respectively, P < 0.05). Conclusions: Increased expression profiles of H19/miR-675-5p/3p and lower expression of the anabolic SMAD1/5 effectors of bone morphogenetic protein (BMP) signalling are associated with low muscle mass in older individuals

Association between perinatal methylation of the neuronal differentiation regulator HES1 and later childhood neurocognitive function and behaviour

Background: Early life environments induce long-term changes in neurocognitive development and behaviour. In animal models, early environmental cues affect neuropsychological phenotypes via epigenetic processes but as yet there is little direct evidence for such mechanisms in humans. Method: We examined the relation between DNA methylation at birth and child neuropsychological outcomes in two culturally diverse populations using a genome-wide methylation analysis and validation by pyrosequencing. Results: Within the UK Southampton Women’s Survey (SWS) we first which identified 41 differentially methylated regions of interest (DMROI) at birth associated with child’s full-scale IQ at age 4-years. Associations between HES1 DMROI methylation and later cognitive function were confirmed by pyrosequencing in 175 SWS children. Consistent with these findings, higher HES1 methylation was associated with higher executive memory function in a second independent group of 200 SWS seven-year olds. Finally, we examined a pathway for this relationship within a Singaporean cohort (n=108). Here, HES1 DMROI methylation predicted differences in early infant behavior, known to be associated with academic success. In vitro, methylation of HES1 inhibited ETS transcription factor binding, suggesting a functional role of this site. Conclusions: Thus, our findings suggest that perinatal epigenetic processes mark later neuro-cognitive function and behavior, providing support for a role of epigenetic processes in mediating the long-term consequences of early life environment on cognitive development. <br/

Trigonelline is a novel NAD+ precursor that improves muscle function during ageing and is reduced in human sarcopenia

Mitochondrial dysfunction and low nicotinamide adenine dinucleotide (NAD+) levels are hallmarks of skeletal muscle aging and sarcopenia1-3, but it remains unclear whether these defects result from local changes or can be mediated by systemic or dietary cues. Here we reporta functional link between circulating levels of the natural alkaloid trigonelline, which is structurally related to nicotinic acid4, NAD+ levels and muscle health in multiple species. In humans, serum trigonelline levels are reduced with sarcopenia, and correlate positively with muscle strength and mitochondrial oxidative phosphorylation in skeletal muscle. Using naturally-occuring and isotopically-labelled trigonelline, we demonstrate that trigonelline incorporates into the NAD+ pool, and increases NAD+ levels in C. elegans, mice and primary myotubes from healthy and sarcopenic humans. Mechanistically, trigonelline does not activate GPR109A but is metabolized via the NAPRT/Preiss-Handler pathway5,6 across models. In C. elegans, trigonelline improves mitochondrial respiration and biogenesis, reduces age-related muscle wasting and increases lifespan and mobility through an NAD+-dependent mechanism requiring sirtuin. Dietary trigonelline supplementation in male mice enhances muscle strength and prevents fatigue during aging. Collectively, we identify nutritional supplementation of trigonelline as a novel NAD+-boosting strategy with therapeutic potential for age-associated muscle decline

Maternal protein restriction with or without folic acid supplementation during pregnancy alters the hepatic transcriptome in adult male rats

Feeding pregnant rats a protein-restricted (PR) diet induces altered expression of candidate genes in the liver of the adult offspring, which can be prevented by supplementation of the PR diet with folic acid (PRF). We investigated the effect of maternal nutrition during pregnancy on the liver transcriptome in their adult male offspring. Pregnant rats were fed control, PR or PRF diets. Male offspring were killed on day 84. The liver transcriptome was analysed by microarray (six livers per maternal dietary group) followed by post hoc analysis of relative mRNA levels and gene ontology. These results were confirmed for selected genes by real-time RT-PCR. There were 311 genes that differed significantly ( &gt;/= 1.5-fold change; P &lt; 0.05) between PR offspring (222 increased) and control offspring, while 191 genes differed significantly between PRF offspring (forty-five increased) compared with offspring of control dams. There were sixteen genes that were significantly altered in both PR and PRF offspring compared with controls. Ion transport, developmental process, and response to reactive oxygen species (RROS) and steroid hormone response (SHR) ontologies were altered in PR offspring. Folic acid supplementation prevented changes within RROS and SHR response pathways, but not in ion transport or developmental process. There was no effect of maternal PR on mRNA expression of imprinted genes. Insulin 1 and Pleckstrin homology-like domain family A member 2 were increased significantly in PRF compared with PR offspring. The present findings show that the pattern of induced changes in the adult liver transcriptome were dependent on maternal protein and folic acid intakes during pregnancy

The serum small non-coding RNA (SncRNA) landscape as a molecular biomarker of age associated muscle dysregulation and insulin resistance in older adults

Small noncoding RNAs (sncRNAs) are implicated in age-associated pathologies, including sarcopenia and insulin resistance (IR). As potential circulating biomarkers, most studies have focussed on microRNAs (miRNAs), one class of sncRNA. This study characterized the wider circulating sncRNA transcriptome of older individuals and associations with sarcopenia and IR. sncRNA expression including miRNAs, transfer RNAs (tRNAs), tRNA-associated fragments (tRFs), and piwi-interacting RNAs (piRNAs) was measured in serum from 21 healthy and 21 sarcopenic Hertfordshire Sarcopenia Study extension women matched for age (mean 78.9 years) and HOMA2-IR. Associations with age, sarcopenia and HOMA2-IR were examined and predicted gene targets and biological pathways characterized. Of the total sncRNA among healthy controls, piRNAs were most abundant (85.3%), followed by tRNAs (4.1%), miRNAs (2.7%), and tRFs (0.5%). Age was associated (FDR &lt; 0.05) with 2 miRNAs, 58 tRNAs, and 14 tRFs, with chromatin organization, WNT signaling, and response to stress enriched among gene targets. Sarcopenia was nominally associated (p &lt; .05) with 12 tRNAs, 3 tRFs, and 6 piRNAs, with target genes linked to cell proliferation and differentiation such as Notch Receptor 1 (NOTCH1), DISC1 scaffold protein (DISC1), and GLI family zinc finger-2 (GLI2). HOMA2-IR was nominally associated (p&lt;0.05) with 6 miRNAs, 9 tRNAs, 1 tRF, and 19 piRNAs, linked with lysine degradation, circadian rhythm, and fatty acid biosynthesis pathways. These findings identify changes in circulating sncRNA expression in human serum associated with chronological age, sarcopenia, and IR. These may have clinical utility as circulating biomarkers of ageing and age-associated pathologies and provide novel targets for therapeutic intervention.</p

Pyrosequencing analysis of the PPARα promoter in adipose tissue of saline or leptin treated adult female rats.

<p>A) Schematic diagram showing the location of the CpGs sequenced. All exon positions are indicated relative to the Ensembl transcription start site B) Pyrosequencing analysis of CpGs within the PPARα promoter. Values represent mean methylation levels ±SEM (n = 8/group).Only CpG sites where a significant difference in methylation between the saline and leptin treated groups are shown. Saline treated, black bars (C), leptin treated white bars (L). Significant differences in DNA methylation between saline and leptin treated groups was determined using a Students unpaired t-test where *p&lt;0.05.</p

Leptin activates the PPARα P2 Promoter in HepG2 Cells P1, P2 and P3 promoter constructs were transfected into HepG2 cells and treated for 24 hrs with a vehicle control or an increasing concentration of leptin (0,50, 500, 1000 ng/ml).

<p>B) P1 and P2 promoter constructs were transfected into HepG2 cells and treated for 24 hrs with leptin (1000 ng/ml) and an increasing concentration of the STAT3 inhibitor PpYLKTK-mts (0,1,10 nM). C) Mutation of the Sp1 site blocks leptin activation of P2 promoter activity. P2 (P2-pGL3) and P2 promoter construct containing the mutated Sp1 response element (SP1M EcoRI-pGL3) was transfected into HepG2 cells and treated with leptin (1000 ng/ml) for 24 hrs. All values represent the mean of 6 independent experiments ±SEM. Statistical comparisons of luciferase activity between treatments relative to the untreated control were determined by ANOVA followed by Bonferroni post hoc analysis. (* p&lt;0.05, ** p&lt;0.001, *** p&lt;0.001).</p

P1 and P2 promoters are active in HepG2 cells.

<p>A) Schematic diagram showing the relative locations of PPARα P1, P2 and P3 cloned promoter regions and their positioning relative to Ensembl transcription start site and 5′UTR exons. B) PPARα P1, P2 and P3 promoter constructs (P1, P2 and P3 Prom 1 µg) and an empty control vector (pGL3Basic 1 µg) were transfected into HepG2 cells and promoter activity assessed 24 hrs later. C and D) Regulation of PPARα promoter activity by clofibric acid and dexamethasone. P2 and P3 promoter constructs were transfected into HepG2 cells and treated for 24 hrs with vehicle control or an increasing concentration of dexamethasone (0, 0.1, 1 or 10 µM) or clofibric acid (60, 80, 100 µM). All values represent the mean of 6 independent experiments ±SEM.). Statistical comparisons of luciferase activity between treatments relative to the untreated control were determined by ANOVA followed by Bonferroni post hoc analysis. (* p&lt;0.05, ** p&lt;0.001, *** p&lt;0.001).</p

Alternative PPARα 5′UTR exon/intron junctions conform to the GT-AG splice site rule.

<p>5′ (donor) and 3′ (acceptor) intron splice site consensus sequences are indicated. The sequences at P1, P2 and P3 transcript exon/intron boundaries are shown. Splice site sequences within introns are shown in lowercase; Exons are shown in capitals, and sizes of exons are given. All transcript exon boundaries conform to the GT-AG splice site rule.</p