Increased Skeletal Muscle 11βHSD1 mRNA Is Associated with Lower Muscle Strength in Ageing

Background

Sarcopenia, the loss of muscle mass and function with age, is associated with increased morbidity and mortality. Current understanding of the underlying mechanisms is limited. Glucocorticoids (GC) in excess cause muscle weakness and atrophy. We hypothesized that GC may contribute to sarcopenia through elevated circulating levels or increased glucocorticoid receptor (GR) signaling by increased expression of either GR or the GC-amplifying enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11βHSD1) in muscle.

Methods

There were 82 participants; group 1 comprised 33 older men (mean age 70.2years, SD 4.4) and 19 younger men (22.2years, 1.7) and group 2 comprised 16 older men (79.1years, 3.4) and 14 older women (80.1years, 3.7). We measured muscle strength, mid-thigh cross-sectional area, fasting morning plasma cortisol, quadriceps muscle GR and 11βHSD1 mRNA, and urinary glucocorticoid metabolites. Data were analysed using multiple linear regression adjusting for age, gender and body size.

Results

Muscle strength and size were not associated with plasma cortisol, total urinary glucocorticoids or the ratio of urinary 5β-tetrahydrocortisol +5α-tetrahydrocortisol to tetrahydrocortisone (an index of systemic 11βHSD activity). Muscle strength was associated with 11βHSD1 mRNA levels (β -0.35, p = 0.04), but GR mRNA levels were not significantly associated with muscle strength or size.

Conclusion

Although circulating levels of GC are not associated with muscle strength or size in either gender, increased cortisol generation within muscle by 11βHSD1 may contribute to loss of muscle strength with age, a key component of sarcopenia. Inhibition of 11βHSD1 may have therapeutic potential in sarcopenia.     

Time course of expression and function of the serotonin transporter in the neonatal rat's primary somatosensory cortex

Immunocytochemical and autoradiographic techniques were employed to determine the time course of expression of the serotonin (5-HT) transporter (SERT) on thalamocortical afferents in the rat's primary somatosensory cortex (S-I), and to correlate this expression to the transient vibrissae-related patterning of 5-HT immunostaining previously described. In additional in vivo and in vitro experiments, 5-HT and 3 H-5-HT were applied directly to the cortices of untreated and 5,7-dihydroxytryptamine-treated (5,7-DHT) rats in order to determine the period during which SERT functions on thalamocortical axons to take up 5-HT. In postnatal rats, SERT immunohistochemistry revealed a somatotopic patterning in S-I that persisted until P-15, which is 6 days after the disappearance of the vibrissae-related 5-HT immunostaining. 3 H-citalopram autoradiography revealed a vibrissae-related pattern in layer IV of S-I until at least P-30. Following destruction of raphe-cortical afferents with 5,7-DHT on the day of birth, this binding pattern remained visible until at least P-25, indicating that SERT located on thalamocortical axons is responsible for the 3 H-citalopram patterning observed in S-I. Tissue from 5,7-DHT-treated rats that had 5-HT applied directly to their cortices revealed a normal vibrissae-related pattern of 5-HT immunostaining in S-I at P-7 and P-11 but only a faint pattern at P-13 and none at P-14. In addition, 3 H-5-HT injected directly into S-I labeled layer IV barrels at P-6 and P-12 but not at P-18. The results of these experiments demonstrate that SERT is expressed by thalamocortical afferents and remains functional long after the vibrissae-related 5-HT immunostaining in cortex disappears.     

Enantiomer‐Specific In Vitro Biotransformation of Select Pharmaceuticals in Rainbow Trout (Oncorhynchus mykiss)

The occurrence of pharmaceuticals in the environment represents a challenge of emerging concern. Many pharmaceuticals are chiral compounds; however, few studies have examined the relative toxicity of pharmaceutical enantiomers to wildlife. Further, our understanding of stereospecific pharmacokinetics remains largely informed by research on humans and a few well‐studied laboratory test animals, and not by studies conducted with environmentally relevant species, including fish. The objective of this study was to investigate whether rainbow trout display stereospecific in vitro metabolism of three common chiral pharmaceuticals. Metabolism by trout liver S9 fractions was evaluated using a substrate depletion approach, which provides an estimate of intrinsic hepatic clearance (CL_{IN VITRO,INT}). No biotransformation was observed for rac‐, R‐, or S‐fluoxetine. Ibuprofen, including both enantiomers and the racemic mixture, appeared to undergo slow metabolism, but the resulting substrate depletion curves did not differ significantly from those of inactive controls. Contrary to relative clearance rates in humans, S(?)‐propranolol was more rapidly cleared than the R(+)‐ enantiomer. This work demonstrates that relative clearance rates and the effects of racemic mixtures in trout could not have been predicted based on human data. Additional research describing species differences and exploring tools for species extrapolation in biomedical and environmental studies is needed. Chirality 25:763–767, 2013, © 2013 Wiley Periodicals, Inc.     

Song and Sperm in Crickets: A Trade‐off between Pre‐ and Post‐copulatory Traits or Phenotype‐Linked Fertility?

When females mate multiply (polyandry) both pre‐ and post‐copulatory sexual selection can occur. Sperm competition theory predicts there should be a trade‐off between investment in attracting mates and investment in ejaculate quality. In contrast, the phenotype‐linked fertility hypothesis predicts a positive relationship should exist between investment in attracting mates and investment in ejaculate quality. Given the need to understand how pre‐ and post‐copulatory sexual selection interacts, we investigated the relationship between secondary sexual traits and ejaculate quality using the European house cricket, Acheta domesticus. Although we found no direct relationship between cricket secondary sexual signals and ejaculate quality, variation in ejaculate quality was dependent on male body weight and mating latency: the lightest males produced twice as many sperm as the heaviest males but took longer to mate with females. Our findings are consistent with current theoretical models of sperm competition. Given light males may have lower mating success than heavy males because females take longer to mate with them in no‐choice tests, light males may be exhibiting an alternative reproductive tactic by providing females with more living sperm. Together, our findings suggest that the fitness of heavy males may depend on pre‐copulatory sexual selection, while the fitness of light males may depend on post‐copulatory fertilization success.     

Laforin, a dual-specificity phosphatase involved in Lafora disease, is phosphorylated at Ser25 by AMP-activated protein kinase

Lafora progressive myoclonus epilepsy [LD (Lafora disease)] is a fatal autosomal recessive neurodegenerative disorder caused by loss-of-function mutations in either the EPM2A gene, encoding the dual-specificity phosphatase laforin, or the EPM2B gene, encoding the E3-ubiquitin ligase malin. Previously, we and others showed that laforin and malin form a functional complex that regulates multiple aspects of glycogen metabolism, and that the interaction between laforin and malin is enhanced by conditions activating AMPK (AMP-activated protein kinase). In the present study, we demonstrate that laforin is a phosphoprotein, as indicated by two-dimensional electrophoresis, and we identify Ser(25) as the residue involved in this modification. We also show that Ser(25) is phosphorylated both in vitro and in vivo by AMPK. Lastly, we demonstrate that this residue plays a critical role for both the phosphatase activity and the ability of laforin to interact with itself and with previously established binding partners. The results of the present study suggest that phosphorylation of laforin-Ser(25) by AMPK provides a mechanism to modulate the interaction between laforin and malin. Regulation of this complex is necessary to maintain normal glycogen metabolism. Importantly, Ser(25) is mutated in some LD patients (S25P), and our results begin to elucidate the mechanism of disease in these patients.     

Renal sodium transporter/channel expression and sodium excretion in P2Y2 receptor knockout mice fed a high-NaCl diet with/without aldosterone infusion

The P2Y(2) receptor (P2Y2-R) antagonizes sodium reabsorption in the kidney. Apart from its effect in distal nephron, hypothetically, P2Y(2)-R may modulate activity/abundances of sodium transporters/channel subunits along the nephron via antagonism of aldosterone or vasopressin or interaction with mediators such as nitric oxide (NO), and prostaglandin E(2) (PGE(2)) or oxidative stress (OS). To determine the extent of the regulatory role of P2Y(2)-R in renal sodium reabsorption, in study 1, we fed P2Y(2)-R knockout (KO; n = 5) and wild-type (WT; n = 5) mice a high (3.15%)-sodium diet (HSD) for 14 days. Western blotting revealed significantly higher protein abundances for cortical and medullary bumetanide-sensitive Na-K-2Cl cotransporter (NKCC2), medullary α-1-subunit of Na-K-ATPase, and medullary α-subunit of the epithelial sodium channel (ENaC) in KO vs. WT mice. Molecular analysis of urine showed increased excretion of nitrates plus nitrites (NOx), PGE(2), and 8-isoprostane in the KO, relative to WT mice, supporting a putative role for these molecules in determining alterations of proteins involved in sodium transport along the nephron. To determine whether genotype differences in response to aldosterone might have played a role in these differences due to HSD, in study 2 aldosterone levels were clamped (by osmotic minipump infusion). Clamping aldosterone (with HSD) led to significantly impaired natriuresis with elevated Na/H exchanger isoform 3 in the cortex, and NKCC2 in the medulla, and modest but significantly lower levels of NKCC2, and α- and β-ENaC in the cortex of KO vs. WT mice. This was associated with significantly reduced urinary NOx in the KO, although PGE(2) and 8-isoprostane remained significantly elevated vs. WT mice. Taken together, our results suggest that P2Y(2)-R is an important regulator of sodium transporters along the nephron. Pre- or postreceptor differences in the response to aldosterone, perhaps mediated via prostaglandins or changes in NOS activity or OS, likely play a role.     

Impaired cardiac reserve and severely diminished skeletal muscle O₂ utilization mediate exercise intolerance in Barth syndrome

Barth syndrome (BTHS) is a mitochondrial myopathy characterized by reports of exercise intolerance. We sought to determine if 1) BTHS leads to abnormalities of skeletal muscle O(2) extraction/utilization and 2) exercise intolerance in BTHS is related to impaired O(2) extraction/utilization, impaired cardiac function, or both. Participants with BTHS (age: 17 ± 5 yr, n = 15) and control participants (age: 13 ± 4 yr, n = 9) underwent graded exercise testing on a cycle ergometer with continuous ECG and metabolic measurements. Echocardiography was performed at rest and at peak exercise. Near-infrared spectroscopy of the vastus lateralis muscle was continuously recorded for measurements of skeletal muscle O(2) extraction. Adjusting for age, peak O(2) consumption (16.5 ± 4.0 vs. 39.5 ± 12.3 ml·kg(-1)·min(-1), P < 0.001) and peak work rate (58 ± 19 vs. 166 ± 60 W, P < 0.001) were significantly lower in BTHS than control participants. The percent increase from rest to peak exercise in ejection fraction (BTHS: 3 ± 10 vs. control: 19 ± 4%, P < 0.01) was blunted in BTHS compared with control participants. The muscle tissue O(2) saturation change from rest to peak exercise was paradoxically opposite (BTHS: 8 ± 16 vs. control: -5 ± 9, P < 0.01), and the deoxyhemoglobin change was blunted (BTHS: 0 ± 12 vs. control: 10 ± 8, P < 0.09) in BTHS compared with control participants, indicating impaired skeletal muscle extraction in BTHS. In conclusion, severe exercise intolerance in BTHS is due to both cardiac and skeletal muscle impairments that are consistent with cardiac and skeletal mitochondrial myopathy. These findings provide further insight to the pathophysiology of BTHS.     

Hypoxic culture and in vivo inflammatory environments affect the assumption of pericyte characteristics by human adipose and bone marrow progenitor cells

Previous studies have shown that exposure to a hypoxic in vitro environment increases the secretion of pro-angiogenic growth factors by human adipose-derived stromal cells (hASCs) [Cao Y, et al., Biochem Biophys Res Commun 332: 370-379, 2005; Kokai LE, et al., Plast Reconstr Surg 116: 1453-1460, 2005; Park BS, et al., Biomed Res (Tokyo) 31: 27-34, 2010; Rasmussen JG, et al., Cytotherapy 13: 318-328, 2010; Rehman J, et al., Circulation 109: 1292-1298, 2004]. Previously, it has been demonstrated that hASCs can differentiate into pericytes and promote microvascular stability and maintenance during angiogenesis in vivo (Amos PJ, et al., Stem Cells 26: 2682-2690, 2008; Traktuev DO, et al., Circ Res 102: 77-85, 2008). In this study, we tested the hypotheses that angiogenic induction can be increased and pericyte differentiation decreased by pretreatment of hASCs with hypoxic culture and that hASCs are similar to human bone marrow-derived stromal cells (hBMSCs) in these regards. Our data confirms previous studies showing that hASCs: 1) secrete pro-angiogenic proteins, which are upregulated following culture in hypoxia, and 2) migrate up gradients of PDGF-BB in vitro, while showing for the first time that a rat mesenteric model of angiogenesis induced by 48/80 increases the propensity of both hASCs and hBMSCs to assume perivascular phenotypes following injection. Moreover, culture of both cell types in hypoxia before injection results in a biphasic vascular length density response in this model of inflammation-induced angiogenesis. The effects of hypoxia and inflammation on the phenotype of adult progenitor cells impacts both the therapeutic and the basic science applications of the cell types, as hypoxia and inflammation are common features of natural and pathological vascular compartments in vivo.     

Risk of injury associated with bodychecking experience among youth hockey players

Background:

In a previous prospective study, the risk of concussion and all injury was more than threefold higher among Pee Wee ice hockey players (ages 11–12 years) in a league that allows bodychecking than among those in a league that does not. We examined whether two years of bodychecking experience in Pee Wee influenced the risk of concussion and other injury among players in a Bantam league (ages 13–14) compared with Bantam players introduced to bodychecking for the first time at age 13.

Methods:

We conducted a prospective cohort study involving hockey players aged 13–14 years in the top 30% of divisions of play in their leagues. Sixty-eight teams from the province of Alberta (n = 995), whose players had two years of bodychecking experience in Pee Wee, and 62 teams from the province of Quebec (n = 976), whose players had no bodychecking experience in Pee Wee, participated. We estimated incidence rate ratios (IRRs) for injury and for concussion.

Results:

There were 272 injuries (51 concussions) among the Bantam hockey players who had bodychecking experience in Pee Wee and 244 injuries (49 concussions) among those without such experience. The adjusted IRRs for game-related injuries and concussion overall between players with bodychecking experience in Pee Wee and those without it were as follows: injury overall 0.85 (95% confidence interval [CI] 0.63 to 1.16); concussion overall 0.84 (95% CI 0.48 to 1.48); and injury resulting in more than seven days of time loss (i.e., time between injury and return to play) 0.67 (95% CI 0.46 to 0.99). The unadjusted IRR for concussion resulting in more than 10 days of time loss was 0.60 (95% CI 0.26 to 1.41).

Interpretation:

The risk of injury resulting in more than seven days of time loss from play was reduced by 33% among Bantam hockey players in a league where bodychecking was allowed two years earlier in Pee Wee compared with Bantam players introduced to bodychecking for the first time at age 13. In light of the increased risk of concussion and other injury among Pee Wee players in a league where bodychecking is permitted, policy regarding the age at which hockey players are introduced to bodychecking requires further consideration.Rates of participation in youth-level ice hockey are high in North America.^1,2 There is growing concern regarding the impact of concussion in this population.^3–5 Body-checking is the reported mechanism for 45%–86% of injuries in youth ice hockey.^5–11 Internationally, the age group at which bodychecking is introduced varies. In Canada, bodychecking is introduced in Pee Wee leagues (ages 11–12 years), except in the province of Quebec, where it is introduced in Bantam (ages 13–14).¹²The age at which bodychecking should be introduced is controversial. We recently reported that the risk of injury and concussion in a Pee Wee league that allows bodychecking was more than threefold higher than in a Pee Wee league that does not allow bodychecking.⁹ Findings from systematic reviews support these findings.^13,14Injury rates may increase when players begin to learn bodychecking, because it is a new skill. If so, injury rates would be expected to be higher among players without bodychecking experience in Pee Wee (i.e., those in Quebec) than among players introduced to body-checking two years earlier in Pee Wee (e.g., in Alberta). We examined whether the risk of concussion and other injury among hockey players in Bantam leagues differed between players with and those without bodychecking experience in Pee Wee.