Endogenous cannabinoid receptor CB1 activation promotes vascular smooth-muscle cell proliferation and neointima formation.

Percutaneous transluminal angioplasty is frequently used in patients with severe arterial narrowing due to atherosclerosis. However, it induces severe arterial injury and an inflammatory response leading to restenosis. Here, we studied a potential activation of the endocannabinoid system and the effect of FA amide hydrolase (FAAH) deficiency, the major enzyme responsible for endocannabinoid anandamide degradation, in arterial injury. We performed carotid balloon injury in atherosclerosis-prone apoE knockout (apoE(-/-)) and apoE(-/-)FAAH(-/-) mice. Anandamide levels were systemically elevated in apoE(-/-) mice after balloon injury. ApoE(-/-)FAAH(-/-) mice had significantly higher baseline anandamide levels and enhanced neointima formation compared with apoE(-/-) controls. The latter effect was inhibited by treatment with CB1 antagonist AM281. Similarly, apoE(-/-) mice treated with AM281 had reduced neointimal areas, reduced lesional vascular smooth-muscle cell (SMC) content, and proliferating cell counts. The lesional macrophage content was unchanged. In vitro proliferation rates were significantly reduced in CB1(-/-) SMCs or when treating apoE(-/-) or apoE(-/-)FAAH(-/-) SMCs with AM281. Macrophage in vitro adhesion and migration were marginally affected by CB1 deficiency. Reendothelialization was not inhibited by treatment with AM281. In conclusion, endogenous CB1 activation contributes to vascular SMC proliferation and neointima formation in response to arterial injury.

Mini invasive skeletal muscle biopsy technique with a tri-axial end cut needle.

OBJECTIVE: Skeletal Muscle Biopsy is a minor surgical procedure for the diagnosis of different neuromuscular pathological conditions and has recently gained popularity also in the research field of age-related muscular modifications and sarcopenia. Few studies focused on the application of mini-invasive muscular biopsy in both normal and pathological conditions. The aim of our study was to describe a mini invasive ultrasound-guided skeletal muscular biopsy technique in complete spinal cord injured (SCI) patients and healthy controls with a tri-axial end-cut needle. PATIENTS AND METHODS: Skeletal muscle biopsies were collected from 6 chronic SCI patients and 3 healthy controls vastus lateralis muscle with a tri-axial end cut needle (Biopince© - Angiotech). Muscle samples were stained for ATPase to determine fibers composition, moreover, gene expression of cyclooxygenase-1 (COX-1) and prostaglandin E2 receptor has been analyzed by Real Time RT-PCR. RESULTS: All the procedures were perfomed easily without failures and complications. Control tissue was macroscopically thicker than SCI one. Control specimen displayed an equal distribution of type I and type II fibers, while SCI sample displayed a prevalence of type II fibers SCI specimen displayed a significant reduction in COX-1 gene expression. This mini-invasive approach was easy, accurate and with low complication rate in performing skeletal muscle biopsy in both SCI patients and controls. CONCLUSIONS: This technique could be useful in conditions in which the overall quantity of specimen required is small like for molecular biology analysis. For histological diagnostic purposes and/or conditions in which the original tissue is already pathologically modified, this technique should be integrated with more invasive techniques.

The exercise leader's gender and physique salience : effects on self-presentational concerns in an exercise context

Self-presentation is the process by which individuals attempt to monitor and control how others perceive and evaluate them (Leary, 1992; Leary & Kowalski, 1990). Self-presentational concerns have been shown to influence a number of exercise-related behaviours, cognitions, and affective responses to exercise (e.g., social anxiety). Social anxiety occurs when an individual wants to create a specific impression on others, but is unsure (s)he will be successful (Leary & Kowalski, 1995). Social physique anxiety (SPA) is a specific form of social anxiety related the evaluation of one's body (Hart, Leary, & Rejeski, 1989). Both social anxiety and SPA may act as deterrents to exercise (Lantz, Hardy, & Ainsworth, 1997; Leary, 1992), so it is important to examine factors that may influence social anxiety and SPA; one such factor is self-presentational efficacy (SPE). SPE is one's confidence in successfully making desired impressions on others (Leary & Atherton, 1986) and has been associated with social anxiety and SPA (Leary & Kowalski, 1995; Gammage, Martin Ginis, & Hall, 2004). Several aspects of the exercise environment, such as the presence of mirrors, clothing, and the exercise leader or other participant characteristics, may be manipulated to influence self-presentational concerns (e.g., Gammage, Martin Ginis et aI., 2004; Martin & Fox, 2001; Martin Ginis, Prapavessis, & Haase, 2005). Given that the exercise leader has been recognized as one of the most important influences in the group exercise context (Franklin, 1988), it is important to further examine how the leader may impact self-presentational concerns. The present study examined the impact of the exercise leader's gender and physique salience (i.e., the extent to which the body was emphasized) on SPE, state social anxiety (SSA), and state social physique anxiety (SPA-S) of women in a live exercise class. Eighty-seven college-aged female non- or infrequent exercisers (i.e., exercised 2 or fewer times per week) participated in a group exercise class led by one of four leaders: a female whose physique was salient; a female whose physique was non-salient; a male whose physique was salient; or a male whose physique was non-salient. Participants completed measures of SPE, SSA, and SPA-S prior to and following completion of a 30- minute group exercise class. In addition, a measure of social comparison to the exercise leader and other participants with respect to attractiveness, skill, and fitness was completed by participants following the exercise class. A MANOV A was conducted to examine differences between groups on postexercise variables. Results indicated that there were no significant differences between groups on measures ofSPE, SSA, or SPA-S (allp's > .05). However, when all participants were collapsed into one group, a MANOV A showed a significant time effect (F(3, 81) = 19.45,p < .05, 1')2= .419). Follow-up ANOVAs indicated that post-exercise SPE increased significantly, while SSA and SPA-S decreased significantly (SPE: F(I, 83) = 30.87,p < .001,1')2 = .27; SSA: F(I,83) = 11.09,p < .001, 1')2 = .12; SPA-S: F (1,83) = 42.79,p < .001, 1')2 = .34). Further, results of a MANOVA revealed that participants who believed they were less fit than other group members (i.e., made negative social comparisons) reported significantly more post-exercise SSA and SP A-S than those who believed they were more fit than the other participants (i.e., made positive comparisons; SSA: F(2, 84) = 3.46, p < .05, 1')2 = .08; SPA-S: F(2, 84) = 5.69, p < .05, 1')2 = .12). These results may indicate that successfully completing an exercise class may serve as a source of SPE and lead to reduced social anxiety and SPA-S in this population. Alternatively, characteristics of the exercise leader may be less important than characteristics of the other participants. These results also suggest that the types of social comparisons made may influence self-presentational concerns in this sample. Future research should examine how the type of social comparison (i.e., negative or positive) made to the other group members may either generate or reduce anxiety. Also, factors that contribute to the types of social comparisons made with other exercisers should be examined. Implications for practice and research are discussed.

Exploring the experiences of parents with children who have communication delays, as a population at risk for the development of behavioural concerns

Existing research identifies preschoolers with communication delays as a population at risk for the development of behavioural concerns. This risk increases when additional environmental factors such as parental stress and family conflict are also present. Research has also shown that behavioural concerns can be stable over time when they develop early. However, early intervention has been shown to be effective in addressing these concerns. The effectiveness of early intervention in addressing both child and family outcomes increases when interventions are delivered in a family-centred approach. This research project made use of data related to child behaviour and parenting, gathered through the Family Resource Project which explored the parenting experiences and resource access and allocation decisions of families who have preschool children with and without communication delays. Cluster analysis was used to explore whether there were identifiable clusters of children and families within each sample. Interview data fi"om each identified family cluster was then explored further, to identify how parents described their child's behaviour and their experience of parenting. Results show that, within this sample, parents of preschoolers with communication delays described their child's behaviour and their experience of parenting differently than did parents of children without communication delays. Results also showed that within this sample parents experiencing parental stress and/or family conflict described their child's behaviour and their experience of parenting differently than did parents from other clusters. Results suggest support for early intervention and the use of family-centred intervention, particularly for families of children with communication delays.

The influence of skeletal muscle cell volume on the regulation of carbohydrate uptake and muscle metabolism

This study investigated the regulation of carbohydrate metabolism and glucose uptake through changes in skeletal muscle cell volume. Using an established invitro isolated whole muscle model, soleus (SOL) and extensor digitorum longus (EDL) muscles were dissected from male rats and incubated in an organ bath containing Sigma medium-199 with 8 mM D-glucose altered to target osmolality (hypo-osmotic: HYPO, iso-osmotic: ISO, hyper-osmotic: HYPER; 190, 290, 400 mmol/kg). Muscles were divided into two groups; metabolite (MM) and uptake (MU). MM (N=48) were incubated for 60 minutes and were then immediately flash frozen. MU (N=24) were incubated for 30 minutes and then the extracellular fluid was exchanged for media containing ^H-glucose and ^'*C-mannitol and incubated for another 30 minutes. After the incubation, the muscles were freeze clamped. Results demonstrated a relative water decrease and increase in HYPER and HYPO, respectively. EDL and SOL glucose uptakes were found to be significantly greater in HYPER conditions. The HYPER condition resulted in significant alterations in muscle metabolite concentrations (lower glycogen, elevated lactate, and G-6-P) suggesting a catabolic cell state, and an increase in glycogen synthase transformation when compared to the HYPO group. In conclusion, skeletal muscle cell volume alters rates of glucose uptake with further alterations in muscle metabolites and glycogen synthase transformation.

The effects of R(+)-lipoic acid supplementation on regulation of human skeletal muscle pyruvate dehydrogenase

This thesis investigated whole body glucose disposal and the adaptive changes in skeletal muscle carbohydrate metabolism following 28 d of supplementation with 1000 mg R(+)-lipoic acid in young sedentary males (age, 22.1 ± 0.67 yr, body mass, 78.7 ± 10.3 kg, n=9). In certain individuals, lipoic acid decreased the 180-min area under the glucose concentration and insulin concentration curve during an oral glucose tolerance test (OGTT) (n=4). In the same individuals, lipoic acid supplementation decreased pyruvate dehydrogenase kinase activity (PDK) (0.09 ± 0.024 min"^ vs. 0.137 ± 0.023 min'\ n=4). The fasting levels of the activated form of pyruvate dehydrogenase (PDHa) were decreased following lipoic acid (0.42 ± 0.13 mmol-min'kg'^ vs. 0.82 ± 0.32 mmolrnin'^kg"\ n=4), yet increased to a greater extent during the OGTT (1.21 ± 0.34 mmol-min'kg"' vs. 0.81 ±0.13 mmolmin"'kg'\ n=4) following hpoic acid supplementation. No changes were demonstrated in the remaining subjects (n=5). It was concluded that improved glucose clearance during an OGTT following lipoic acid supplementation is assisted by increased muscle glucose oxidation through increased PDHa activation and decreased PDK activity in certain individuals.

The capillary supply of human skeletal muscle in health and disease

BACKGROUND: Capillaries function to provide a surface area for nutrient and waste exchange with cells. The capillary supply of skeletal muscle is highly organized, and therefore, represents an excellent choice to study factors regulating diffusion. Muscle is comprised of three specific fibre types, each with specific contractile and metabolic characteristics, which influence the capillary supply of a given muscle; in addition, both environmental and genetic factors influence the capillary supply, including aging, physical training, and various disease processes. OBJECTIVE: The present study was undertaken to develop and assess the functionality of a data base, from which virtual experiments can be conducted on the capillary supply of human muscle, and the adaptations of the capillary bed in muscle to various perturbations. METHODS: To create the database, an extensive search of the literature was conducted using various search engines, and the three key words - "capillary, muscle, and human". This search yielded 169 papers from which the data for the 46 variables on the capillary supply and fibre characteristics of muscle were extracted for inclusion in the database. A series of statistical analyses (ANOVA) were done on the capillary database to examine differences in skeletal muscle capillarization and fibre characteristics between young and old individuals, between healthy and diseased individuals, and between untrained, endurance trained, endurance welltrained, and resistance trained individuals, using SAS. RESULTS: There was a significantly higher capillarization in the young compared to the old individuals, in the healthy compared to the diseased individuals, and in the endurance-trained and endurance well-trained compared to the untrained individuals. CONCLUSIONS: The results of this study support the conclusion that the capillary supply of skeletal muscle is closely regulated by factors aimed at optimizing oxygen and nutrient supply and/or waste removal in response to changes in muscle mass and/or metabolic activity.

Adaptations of skeletal muscle pyruvate dehydrogenase kinase in response to food-restriction in mitochondrial subpopulations

University, 2006 Dr. Sandra J. Peters Pyruvate dehydrogenase (PDH) catalyses the decarboxylation of pyruvate, to form acetyl-CoA. PDH activity is down-regulated by intrinsic PDH kinases (predominantly PDK2 and PDK4 isoforms), but the understanding of the PDK isoform distribution and adaptation to nutritional stresses has been restricted to mixed mitochondrial populations, and not delineated between subsarcolemmal (SS) and intermyofibrillar (IMF) subpopulations. SS and IMF mitochondria exhibit distinct morphological and biochemical properties; however the functional differences are not well understood. This study investigated the effect of fed (FED) versus 48 h total foodrestriction (FR) on rat red gastrocnemius muscle PDK2 and 4 isoform content in SS and IMF mitochondria. PDK4 content was ~3-5 fold higher in SS mitochondria compared to IMF (p=0.001), and increased with FR -3-4- fold in both subpopulations (p<0.001). PDK2 was -2.5-4 fold higher in SS mitochondria compared to IMF (p=0.001), but PDK2 was unaltered with FR. Citrate synthase activity (|imol/min/mg mitochondrial protein) was not different between either subpopulation. As well there were no significant differences between mitochondrial subpopulations in PDH complex components in both fed and FR states. These results demonstrate that there is a markedly higher content of both PDK isofonns in SS compared to IMF mitochondria. Although PDK2 does not increase in either subpopulation in response to FR, PDK4 increases to a similar extent in both SS and IMF after 48 h food-restriction.

Skeletal muscle fibre-type comparison of whole tissue and subcellular membrane phospholipids and fatty acids

Membranes are dynamic structures that affect cell structure and function. Compositional changes ofmembranes have been shown with the application of a perturbation; however these are limited to whole tissue analysis. The purpose of this thesis was to compare the phospholipid (PL) fatty acid (FA) composition of rat whole muscle (Wm) to 1) purified and non-purified subsarcolemmal (SS) mitochondria in soleus, plantaris, and red gastrocnemius, and 2) sarcolemma, transverse-tubules, SS and intermyofibrillar (IMF) mitochondria fix)m whole hindlimb. The major findings were that 1) contamination significantly altered the PL FA composition of the SS mitochondrial membrane fraction, 2) Wm and SS mitochondria compositions differed between muscle types, and 3) Wm did not accurately reflect the PL FA composition of any isolated subcellular membranes, with each being unique from each other. As such, the relevancy of the trends reported in the literature of the effects of perturbations on Wm may be limited.

Glucose dynamics in rat skeletal muscle : recovery from osmotic stress

The purpose of this study was to examine cell glucose kinetics in rat skeletal muscle during iso-osmotic recovery from hyper- and hypo-osmotic stress. Rat EDL muscles were incubated for sixty minutes in either HYPO (190 mmol/kg), ISO (290 mmol/kg), or HYPER (400 mmol/kg) media (Sigma medium-199, 8 mM glucose) according to an established in vitro whole muscle model. In addition to sixty minute baseline measures in aniso-osmotic conditions, (HYPO-0 n=8; ISO- 0, n=S; HYPER-0, n=8), muscles were subjected to either one minute (HYPO-1 n=8; ISO-1, n=8; HYPER-1, n=8) or five minutes (HYPO-5 n=8; ISO-5, n=8; HYPER-5, n=8) of iso-osmotic recovery media and analyzed for metabolite content and glycogen synthase percent activation. To determine glucose uptake during iso-osmotic recovery, muscles (n=6 per group) were incubated for sixty minutes in either hypo-, iso-, or hyper-osmotic media immediately followed by five minutes of iso-osmotic media containing 3H-glucose and 14 C-mannitol. Increased relative water content/decreased [glucose] (observed in HYPO-0) and decreased water content/increased [glucose] (observed in HYPER-0) returned to ISO levels within 5 minutes of recovery. Glycogen synthase percent activation increased significantly in HYPO-5 over iso-osmotic controls. Glucose uptake measurements revealed no significant differences between groups. It was determined that [glucose] and muscle water content rapidly recovered from osmotic stress demonstrating skeletal muscle's resilience to osmotic stress.

The effect of elevated muscle fluid volume on indices of muscle damage following an acute bout of eccentric exercise

The primary purpose of the current investigation was to develop an elevated muscle fluid level using a human in-vivo model. The secondary purpose was to determine if an increased muscle fluid content could alter the acute muscle damage response following a bout of eccentric exercise. Eight healthy, recreationally active males participated in a cross-over design involving two randomly assigned trials. A hydration trial (HYD) consisting of a two hour infusion of a hypotonic (0.45%) saline at a rate of 20mL/minVl .73m"^ and a control trial (CON), separated by four weeks. Following the infusion (HYD) or rest period (CON), participants completed a single leg isokinetic eccentric exercise protocol of the quadriceps, consisting of 10 sets of 10 repetitions with a one minute rest between each set. Muscle biopsies were collected prior to the exercise, immediately following and at three hours post exercise. Muscle analysis included determination of wet-dry ratios and quantification of muscle damage using toluidine blue staining and light microscopy. Blood samples were collected prior to, immediately post, three and 24 hours post exercise to determine changes in creatine kinase (CK), lactate dehydrogenase (LD), interleukin-6 (IL-6) and Creactive protein (CRP) levels. Results demonstrated an increased muscle fluid volume in the HYD condition following the infusion when compared to the CON condition. Isometric peak torque was significantly reduced following the exercise in both the HYD and CON conditions. There were no significant differences in the number of areas of muscle damage at any of the time points in either condition, with no differences between conditions. CK levels were significantly greater 24hour post exercise compared to pre, immediately and three hours post similarly in both conditions. LD in the HYD condition followed a similar trend as CK with 24 hour levels higher than pre, immediately post and three hours post and LD levels were significantly greater 24 hours post compared to pre levels in the CON condition, with no differences between conditions. A significant main effect for time was observed for CRP (p<0.05) for time, such that CRP levels increased consistently at each subsequent time point. However, CRP and IL-6 levels were not different at any of the measured time points when comparing the two conditions. Although the current investigation was able to successfully increase muscle fluid volume and an increased CK, LD and CRP were observed, no muscle damage was observed following the eccentric exercise protocol in the CON or HYD conditions. Therefore, the hypotonic infusion used in the HYD condition proved to be a viable method to acutely increase muscle fluid content in in-vivo human skeletal muscle.

The effect of extracellular osmolality on cell volume and resting skeletal muscle metabolism

The purpose of the current investigation was to establish an in-l'itro skeletal muscle model to study acute alterations in resting skeletal muscle cell volume. Isolated. whole muscle (SOL and EDL) was dissected from Long Evans rats and incubated for 60 min in Sigma Medium-199 (resting tension (lg). bubbled with 95:5% 02:C02, 30 ± 2°C, and pH 7.4). Media osmolality was altered to simulate hypo-osmotic (190 ± 10 Osm) (HYPO) or hyper-osmotic conditions (400 ± 10 Osm) (HYPER) while an iso-osmotic condition (290± 1 0 Osm) (CON) served as a control (n= 17.19.17). Following incubation, relative muscle water content decreased with HYPER and increased with HYPO in both muscle types (p<0.05). The cross-sectional area of HYPO SOL type I and type II fibres increased (p<0.05) while the EDL type 11 fibre area decreased in HYPER and increascd from HYPO exposure. Furthermore, HYPER exposure in both muscles lead to decreased ATP and phosphocreatine (p<0.05) and increased creatine and lactate (p<0.05) compared to CON. This isolated skeletal muscle model proved viable and demonstrated that altering extracellular osmolality could cause acutc alterations in muscle water content and resting muscle metabolism.

The acute effects of differential dietary fatty acids on PDHa activity in human skeletal muscle at the onset of exercise

Pyruvate dehydrogenase (PDH) is an important regulator of carbohydrate oxidation during exercise and its activity can be down-regulated by an increase in dietary fat. The purpose of this study was to determine the acute metabolic effects of differential dietary fatty acids on the activation of PDH in its active form (PDHa) at rest and at the onset of moderate-intensity exercise. University-aged male subjects (n=7) underwent 2 fat loading trials spaced at least 2 weeks apart. Subjects consumed saturated (SFA) or polyunsaturated (PUFA) fat over the course of 5 hours. Following this, participants cycled at 65% VO2 max for 15 min. Muscle biopsies were taken prior to and following fat loading and at 1 min exercise. Plasma free fatty acids increased from 0.15 ± 0.07 to 0.54 ± 0.19 mM over 5 hours with SFA and from 0.1 1 ± 0.04 to 0.35 ±0.13 mM with PUFA. PDHa activity was unchanged following fat loading, but increased at the onset of exercise in the SFA trial, from 1 .4 ± 0.4 to 2.2 ± 0.4 /xmol/min/kg wet wt. This effect was negated in the PUFA trial (1 .2 ± 0.3 to 1 .3 ± 0.3 pimol/min/kg wet wt.). PDH kinase (PDK) was unchanged in both trials, suggesting that the attenuation of PDHa activity with PUFA was a result of changes in the concentrations of intramitochondrial effectors, more specifically intramitochondrial NADH or Ca^*. Our findings suggest that attenuated PDHa activity participates in the preferential oxidation of PUFA during moderateintensity exercise.