Performance evaluation of a new prognostic-efficacy-combination design in the context of telemedical interventions.

AIMS: Telemedical interventions in heart failure patients intend to avoid unfavourable, indication-related events by an early, individualized care, which reacts to the current patients need. However, telemedical support is an expensive intervention, and usually only patients with high risk for unfavourable follow-up events will be able to profit from it. Möckel et al. therefore adapted a new design which we call 'prognostic-efficacy-combination design'. This design allows to define a biomarker cut-off and to perform a randomized controlled trial (RCT) in a biomarker-selected population within a single study. However, so far, it has not been evaluated if this double use of the control group for biomarker cut-off definition and efficacy assessment within the RCT leads to a bias in treatment effect estimation. In this methodological research work, we therefore want to evaluate whether the 'prognostic-efficacy-combination design' leads to biased treatment effect estimates and also compare it to alternative designs. If there is a bias, we further want to analyse its magnitude under different parameter settings. METHODS: We perform a systematic Monte Carlo simulation study to investigate among others potential bias, root mean square error and sensitivity, and specificity as well as the total treatment effect estimate in various realistic trial scenarios that mimic and vary the true data characteristics of the published TIM-HF2 Trial. In particular, we vary the event proportion, the sample size, the biomarker distribution, and the lower bound for the sensitivity. RESULTS: The results show that indeed the proposed design leads to some bias in the effect estimators, indicating an overestimation of the effect. However, this bias is relatively small in most scenarios. CONCLUSIONS: The 'prognostic-efficacy-combination design' can generally be recommended for clinical applications due to its efficiency compared to two separate trials. We recommend a sufficiently large sample size depending on the trial scenario. Our simulation code can be adapted to explore suitable sample sizes for other settings.

The weight of culture: Societal individualism and flexibility explain large global variations in obesity.

BACKGROUND: Obesity rates have been rising steeply across the globe in recent decades, posing a major threat to global human health. Despite this almost universal increase, differences between countries remain striking, even among equally developed societies. METHODS: We test if two cultural dimensions derived from a revised Hofstede model of culture from Minkov (2018), namely collectivism vs. individualism and monumentalism vs. flexibility, could help explain national variations in prevalence of obesity (BMI ≥ 30) among women and men around the world. We develop a theoretical framework that links these two cultural dimensions with obesity and then test their association empirically in analyses including 51 countries from all regions of the world as well as using imputed data for a total of 155 countries, representing 98% of the global population. RESULTS: In contrast to previous studies, we find that, adjusting for undernourishment and other potential confounds, individualism is associated with higher obesity prevalence in the male population, but not among the female population. We explain these findings by pointing to the different mechanisms through which individualism relates to health behavior, some of which are more gender-specific than others. A further novel finding is that flexibility, a national cultural trait that emphases humility, self-control, and restraint of desires, is a strong negative predictor of obesity in both genders beyond various potential confounds and is highly robust in specification curve analyses. CONCLUSIONS: Our findings suggest that taking national culture into account can enhance our understanding of the obesity pandemic and should thus be considered by policy-makers in their design of interventions.

Male Sexual Preference for Female Swimming Activity in the Guppy (Poecilia reticulata).

Mate choice that is based on behavioural traits is a common feature in the animal kingdom. Using the Trinidadian guppy, a species with mutual mate choice, we investigated whether males use female swimming activity-a behavioural trait known to differ consistently among individuals in many species-as a trait relevant for their mate choice. In the first experiment, we assessed male and female activity in an open field test alone (two repeated measures) and afterwards in heterosexual pairs (two repeated measures). In these pairs, we simultaneously assessed males' mating efforts by counting the number of sexual behaviours (courtship displays and copulations). Male and female guppies showed consistent individual differences in their swimming activity when tested both alone and in a pair, and these differences were maintained across both test situations. When controlling for male swimming behaviour and both male and female body size, males performed more courtship displays towards females with higher swimming activity. In a second experiment, we tested for a directional male preference for swimming activity by presenting males video animations of low- and high-active females in a dichotomous choice test. In congruence with experiment 1, we found males to spend significantly more time in association with the high-active female stimulus. Both experiments thus point towards a directional male preference for higher activity levels in females. We discuss the adaptive significance of this preference as activity patterns might indicate individual female quality, health or reproductive state while, mechanistically, females that are more active might be more detectable to males as well.

Banking on cooperation: an evolutionary analysis of microfinance loan repayment behaviour.

Microfinance is an economic development tool that provides loans to low-income borrowers to stimulate economic growth and reduce financial hardship. Lenders typically require joint liability, where multiple borrowers share the responsibility of repaying a group loan. We propose that this lending practice creates a cooperation dilemma similar to that faced by humans and other organisms in nature across many domains. This could offer a real-world test case for evolutionary theories of cooperation from the biological sciences. In turn, such theories could provide new insights into loan repayment behaviour. We first hypothesise how group loan repayment efficacy should be affected by mechanisms of assortment from the evolutionary literature on cooperation, i.e. common ancestry (kin selection), prior interaction (reciprocity), partner choice, similarity of tags, social learning, and ecology and demography. We then assess selected hypotheses by reviewing 41 studies from 32 countries on micro-borrowers' loan repayment, evaluating which characteristics of borrowers are associated with credit repayment behaviour. Surprisingly, we find that kinship is mostly negatively associated with repayment efficacy, but prior interaction and partner choice are both more positively associated. Our work highlights the scope of evolutionary theory to provide systematic insight into how humans respond to novel economic institutions and interventions.

Defective lysosome reformation during autophagy causes skeletal muscle disease.

The regulation of autophagy-dependent lysosome homeostasis in vivo is unclear. We showed that the inositol polyphosphate 5-phosphatase INPP5K regulates autophagic lysosome reformation (ALR), a lysosome recycling pathway, in muscle. INPP5K hydrolyzes phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] to phosphatidylinositol 4-phosphate [PI(4)P], and INPP5K mutations cause muscular dystrophy by unknown mechanisms. We report that loss of INPP5K in muscle caused severe disease, autophagy inhibition, and lysosome depletion. Reduced PI(4,5)P2 turnover on autolysosomes in Inpp5k-/- muscle suppressed autophagy and lysosome repopulation via ALR inhibition. Defective ALR in Inpp5k-/- myoblasts was characterized by enlarged autolysosomes and the persistence of hyperextended reformation tubules, structures that participate in membrane recycling to form lysosomes. Reduced disengagement of the PI(4,5)P2 effector clathrin was observed on reformation tubules, which we propose interfered with ALR completion. Inhibition of PI(4,5)P2 synthesis or expression of WT INPP5K but not INPP5K disease mutants in INPP5K-depleted myoblasts restored lysosomal homeostasis. Therefore, bidirectional interconversion of PI(4)P/PI(4,5)P2 on autolysosomes was integral to lysosome replenishment and autophagy function in muscle. Activation of TFEB-dependent de novo lysosome biogenesis did not compensate for loss of ALR in Inpp5k-/- muscle, revealing a dependence on this lysosome recycling pathway. Therefore, in muscle, ALR is indispensable for lysosome homeostasis during autophagy and when defective is associated with muscular dystrophy.

Metabolic remodeling of dystrophic skeletal muscle reveals biological roles for dystrophin and utrophin in adaptation and plasticity.

OBJECTIVES: Preferential damage to fast, glycolytic myofibers is common in many muscle-wasting diseases, including Duchenne muscular dystrophy (DMD). Promoting an oxidative phenotype could protect muscles from damage and ameliorate the dystrophic pathology with therapeutic relevance, but developing efficacious strategies requires understanding currently unknown biological roles for dystrophin and utrophin in dystrophic muscle adaptation and plasticity. METHODS: Combining whole transcriptome RNA sequencing and mitochondrial proteomics with assessments of metabolic and contractile function, we investigated the roles of dystrophin and utrophin in fast-to-slow muscle remodeling with low-frequency electrical stimulation (LFS, 10 Hz, 12 h/d, 7 d/wk, 28 d) in mdx (dystrophin null) and dko (dystrophin/utrophin null) mice, two established preclinical models of DMD. RESULTS: Novel biological roles in adaptation were demonstrated by impaired transcriptional activation of estrogen-related receptor alpha-responsive genes supporting oxidative phosphorylation in dystrophic muscles. Further, utrophin expression in dystrophic muscles was required for LFS-induced remodeling of mitochondrial respiratory chain complexes, enhanced fiber respiration, and conferred protection from eccentric contraction-mediated damage. CONCLUSIONS: These findings reveal novel roles for dystrophin and utrophin during LFS-induced metabolic remodeling of dystrophic muscle and highlight the therapeutic potential of LFS to ameliorate the dystrophic pathology and protect from contraction-induced injury with important implications for DMD and related muscle disorders.

Copeptin combined with either non-high sensitivity or high sensitivity cardiac troponin for instant rule-out of suspected non-ST segment elevation myocardial infarction.

BACKGROUND: Whether Copeptin combined with high sensitivity troponin below the respective decision cut-offs improves rule-out of NSTEMI and may predict all-cause death at 30-day is under debate. METHODS: Data on 10,329 patients from 5 trials were pooled to evaluate the diagnostic and prognostic performance of an initial Copeptin below decision cut-off in combination with a (hs)-cTn below the uppler limit of normal (ULN) compared to a) the initial (hs)-cTn alone in the standard serial sampling strategy based on the 99th percentile and b) a single marker strategy (SMS) based on hs-cTn < limit of detection. Endpoints were sensitivities and negative predictive values (NPV) for rule-out of NSTEMI, 30-day all-cause mortality and rates of eligibility for DMS or SMS. RESULTS: NPV for NSTEMI was higher for DMS than for the initial cTn, regardless assay sensitivity. The highest NPVs were observed with DMS vs. hs-cTn (99.4% [95% CI: 99.0%-99.6%] vs. 98.8% [98.4%-99.1%],) , and improved performance was consistent across all important subgroups including presentation <3 h, again irrespective of assay sensitivity. The point estimate of all NPVs for all-cause death exceeded 99.75%. In the label populations, DMS versus SMS demonstrated comparably high NPVs for rule-out of NSTEMI (99.4% [99.0%-99.6%] vs. 99.9% [99.2%-100.0%]), very low mortality after rule-out (0.1% [0.0%-0.4% vs. 0.0% [0.0%-1.2%]), but eligibility for rule-out was 2.4-fold higher (61.4% [59.9%-62.9%] vs. 25.3% [23.7%-26.9%]) with DMS than SMS. CONCLUSION: Findings from a large pooled cohort corroborate the safety of the dual marker strategy for instant rule-out of NSTEMI, extending evidence to hs-cTn. Copeptin below cut-off in combination with hs-cTn below ULN may be used in more than 2.4-times more patients presenting with suspected ACS than a single marker strategy based on very low hs-cTn, without the need to exclude very early presenters or other important subgroups.

Fair payments for effective environmental conservation.

Global efforts for biodiversity protection and land use-based greenhouse gas mitigation call for increases in the effectiveness and efficiency of environmental conservation. Incentive-based policy instruments are key tools for meeting these goals, yet their effectiveness might be undermined by such factors as social norms regarding whether payments are considered fair. We investigated the causal link between equity and conservation effort with a randomized real-effort experiment in forest conservation with 443 land users near a tropical forest national park in the Vietnamese Central Annamites, a global biodiversity hotspot. The experiment introduced unjustified payment inequality based on luck, in contradiction of local fairness norms that were measured through responses to vignettes. Payment inequality was perceived as less fair than payment equality. In agreement with our preregistered hypotheses, participants who were disadvantaged by unequal payments exerted significantly less conservation effort than other participants receiving the same payment under an equal distribution. No effect was observed for participants advantaged by inequality. Thus, equity effects on effort can have consequences for the effectiveness and efficiency of incentive-based conservation instruments. Furthermore, we show that women exerted substantially more conservation effort than men, and that increasing payment size unexpectedly reduced effort. This emphasizes the need to consider social comparisons, local equity norms, and gender in environmental policies using monetary incentives to motivate behavioral change.

Biomarker guidance allows a more personalized allocation of patients for remote patient management in heart failure: results from the TIM-HF2 trial.

AIMS: The TIM-HF2 study showed less days lost due to unplanned cardiovascular hospitalization or all-cause death and improved survival in patients randomly assigned to remote patient management (RPM) instead of standard of care. METHODS AND RESULTS: This substudy explored whether the biomarkers mid-regional pro-adrenomedullin (MR-proADM) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) could be used to identify low-risk patients unlikely to benefit from RPM, thereby allowing more efficient allocation of the intervention. For 1538 patients of the trial (median age 73 years, interquartile range 64-78 years, 30% female), baseline biomarkers were used to select subpopulations recommended for RPM with various safety endpoints (100%, 98%, 95% sensitivity), and efficacy of RPM was assessed. Both biomarkers were strongly associated with events. The primary endpoint of lost days increased from 1.0% (1.4%) in the lowest to 17.3% (17.6%) in the highest quintile of NT-proBNP (MR-proADM). After combining biomarkers to identify patients recommended for RPM with 95% sensitivity, in the most efficient scenario (excluding 27% of patients; NT-proBNP < 413.7 pg/mL and MR-proADM < 0.75 nmol/L), the effect of RPM on patients was highly similar to the original trial (ratio of lost days: 0.78, hazard ratio for all-cause death: 0.68). Number needed to treat for all-cause death was lowered from 28 to 21. Rates of emergencies and telemedical efforts were significantly lower among patients not recommended for RPM. Biomarker guidance would have saved about 150 h effort/year per 100 patients of the eligible population. CONCLUSIONS: The combined use of MR-proADM and NT-proBNP may allow safe, more precise, effective and cost-saving allocation of patients with heart failure to RPM and warrants further prospective studies.

Sociocultural heterogeneity in a common pool resource dilemma.

Collective action of resource users is essential for sustainability. Yet, often user groups are socioculturally heterogeneous, which requires cooperation to be established across salient group boundaries. We explore the effect of this type of heterogeneity on resource extraction in lab-in-the-field Common Pool Resource (CPR) experiments in Zanzibar, Tanzania. We create heterogeneous groups by mixing fishers from two neighbouring fishing villages which have distinct social identities, a history of conflict and diverging resource use practices and institutions. Additionally, we analyse between-village differences in extraction behaviour in the heterogeneous setting to assess if out-group cooperation in a CPR dilemma is associated with a community's institutional scope in the economic realm (e.g., degree of market integration). We find no aggregate effect of heterogeneity on extraction. However, this is because fishers from the two villages behave differently in the heterogeneity treatment. We find support for the hypothesis that cooperation under sociocultural heterogeneity is higher for fishers from the village with larger institutional scope. In line with this explanation, cooperation under heterogeneity also correlates with a survey measure of individual fishers' radius of trust. We discuss implications for resource governance and collective action research.

Discovery of a multispecies shark aggregation and parturition area in the Ba Estuary, Fiji Islands.

Population declines in shark species have been reported on local and global scales, with overfishing, habitat destruction and climate change posing severe threats. The lack of species-specific baseline data on ecology and distribution of many sharks, however, makes conservation measures challenging. Here, we present a fisheries-independent shark survey from the Fiji Islands, where scientific knowledge on locally occurring elasmobranchs is largely still lacking despite the location's role as a shark hotspot in the Pacific. Juvenile shark abundance in the fishing grounds of the Ba Estuary (north-western Viti Levu) was assessed with a gillnet- and longline-based survey from December 2015 to April 2016. A total of 103 juvenile sharks identified as blacktip Carcharhinus limbatus (n = 57), scalloped hammerhead Sphyrna lewini (n = 35), and great hammerhead Sphyrna mokarran (n = 11) sharks were captured, tagged, and released. The condition of umbilical scars (68% open or semihealed), mean sizes of individuals (±SD) (C. limbatus: 66.5 ± 3.8 cm, S. lewini: 51.8 ± 4.8 cm, S. mokarran 77.4 ± 2.8 cm), and the presence of these species over recent years (based on fishermen interviews), suggest that the Ba Estuary area is a critical habitat for multiple species that are classified as "Near Threatened" or "Endangered." Specifically, the area likely acts as a parturition ground over the studied period, and potentially as a subsequent nursery area. We identified subareas of high abundance and found that temperature, salinity and depth acted as small-scale environmental drivers of shark abundance. The data suggests a tendency for species-specific spatial use, both horizontally (i.e., between sampling areas) and vertically (i.e., across the water column). These results enhance the understanding of shark ecology in Fiji and provide a scientific basis for the implementation of local conservation strategies that contribute to the protection of these threatened species.

Scriptaid enhances skeletal muscle insulin action and cardiac function in obese mice.

AIM: To determine the effect of Scriptaid, a compound that can replicate aspects of the exercise adaptive response through disruption of the class IIa histone deacetylase (HDAC) corepressor complex, on muscle insulin action in obesity. MATERIALS AND METHODS: Diet-induced obese mice were administered Scriptaid (1 mg/kg) via daily intraperitoneal injection for 4 weeks. Whole-body and skeletal muscle metabolic phenotyping of mice was performed, in addition to echocardiography, to assess cardiac morphology and function. RESULTS: Scriptaid treatment had no effect on body weight or composition, but did increase energy expenditure, supported by increased lipid oxidation, while food intake was also increased. Scriptaid enhanced the expression of oxidative genes and proteins, increased fatty acid oxidation and reduced triglycerides and diacylglycerides in skeletal muscle. Furthermore, ex vivo insulin-stimulated glucose uptake by skeletal muscle was enhanced. Surprisingly, heart weight was reduced in Scriptaid-treated mice and was associated with enhanced expression of genes involved in oxidative metabolism in the heart. Scriptaid also improved indices of both diastolic and systolic cardiac function. CONCLUSION: These data show that pharmacological targeting of the class IIa HDAC corepressor complex with Scriptaid could be used to enhance muscle insulin action and cardiac function in obesity.

BGP-15 Improves Aspects of the Dystrophic Pathology in mdx and dko Mice with Differing Efficacies in Heart and Skeletal Muscle.

Duchenne muscular dystrophy is a severe and progressive striated muscle wasting disorder that leads to premature death from respiratory and/or cardiac failure. We have previously shown that treatment of young dystrophic mdx and dystrophin/utrophin null (dko) mice with BGP-15, a coinducer of heat shock protein 72, ameliorated the dystrophic pathology. We therefore tested the hypothesis that later-stage BGP-15 treatment would similarly benefit older mdx and dko mice when the dystrophic pathology was already well established. Later stage treatment of mdx or dko mice with BGP-15 did not improve maximal force of tibialis anterior (TA) muscles (in situ) or diaphragm muscle strips (in vitro). However, collagen deposition (fibrosis) was reduced in TA muscles of BGP-15-treated dko mice but unchanged in TA muscles of treated mdx mice and diaphragm of treated mdx and dko mice. We also examined whether BGP-15 treatment could ameliorate aspects of the cardiac pathology, and in young dko mice it reduced collagen deposition and improved both membrane integrity and systolic function. These results confirm BGP-15's ability to improve aspects of the dystrophic pathology but with differing efficacies in heart and skeletal muscles at different stages of the disease progression. These findings support a role for BGP-15 among a suite of pharmacological therapies for Duchenne muscular dystrophy and related disorders.

Skeletal muscle-specific overexpression of IGFBP-2 promotes a slower muscle phenotype in healthy but not dystrophic mdx mice and does not affect the dystrophic pathology.

OBJECTIVE: The insulin-like growth factor binding proteins (IGFBPs) are thought to modulate cell size and homeostasis via IGF-I-dependent and -independent pathways. There is a considerable dearth of information regarding the function of IGFBPs in skeletal muscle, particularly their role in the pathophysiology of Duchenne muscular dystrophy (DMD). In this study we tested the hypothesis that intramuscular IGFBP-2 overexpression would ameliorate the pathology in mdx dystrophic mice. DESIGN: 4week old male C57Bl/10 and mdx mice received a single intramuscular injection of AAV6-empty or AAV6-IGFBP-2 vector into the tibialis anterior muscle. At 8weeks post-injection the effect of IGFBP-2 overexpression on the structure and function of the injected muscle was assessed. RESULTS: AAV6-mediated IGFBP-2 overexpression in the tibialis anterior (TA) muscles of 4-week-old C57BL/10 and mdx mice reduced the mass of injected muscle after 8weeks, inducing a slower muscle phenotype in C57BL/10 but not mdx mice. Analysis of inflammatory and fibrotic gene expression revealed no changes between control and IGFBP-2 injected muscles in dystrophic (mdx) mice. CONCLUSIONS: Together these results indicate that the IGFBP-2-induced promotion of a slower muscle phenotype is impaired in muscles of dystrophin-deficient mdx mice, which contributes to the inability of IGFBP-2 to ameliorate the dystrophic pathology. The findings implicate the dystrophin-glycoprotein complex (DGC) in the signaling required for this adaptation.

FHL1 reduces dystrophy in transgenic mice overexpressing FSHD muscular dystrophy region gene 1 (FRG1).

Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal-dominant disease with no effective treatment. The genetic cause of FSHD is complex and the primary pathogenic insult underlying the muscle disease is unknown. Several disease candidate genes have been proposed including DUX4 and FRG1. Expression analysis studies of FSHD report the deregulation of genes which mediate myoblast differentiation and fusion. Transgenic mice overexpressing FRG1 recapitulate the FSHD muscular dystrophy phenotype. Our current study selectively examines how increased expression of FRG1 may contribute to myoblast differentiation defects. We generated stable C2C12 cell lines overexpressing FRG1, which exhibited a myoblast fusion defect upon differentiation. To determine if myoblast fusion defects contribute to the FRG1 mouse dystrophic phenotype, this strain was crossed with skeletal muscle specific FHL1-transgenic mice. We previously reported that FHL1 promotes myoblast fusion in vitro and FHL1-transgenic mice develop skeletal muscle hypertrophy. In the current study, FRG1 mice overexpressing FHL1 showed an improvement in the dystrophic phenotype, including a reduced spinal kyphosis, increased muscle mass and myofiber size, and decreased muscle fibrosis. FHL1 expression in FRG1 mice, did not alter satellite cell number or activation, but enhanced myoblast fusion. Primary myoblasts isolated from FRG1 mice showed a myoblast fusion defect that was rescued by FHL1 expression. Therefore, increased FRG1 expression may contribute to a muscular dystrophy phenotype resembling FSHD by impairing myoblast fusion, a defect that can be rescued by enhanced myoblast fusion via expression of FHL1.

Dysfunctional muscle and liver glycogen metabolism in mdx dystrophic mice.

BACKGROUND: Duchenne muscular dystrophy (DMD) is a severe, genetic muscle wasting disorder characterised by progressive muscle weakness. DMD is caused by mutations in the dystrophin (dmd) gene resulting in very low levels or a complete absence of the dystrophin protein, a key structural element of muscle fibres which is responsible for the proper transmission of force. In the absence of dystrophin, muscle fibres become damaged easily during contraction resulting in their degeneration. DMD patients and mdx mice (an animal model of DMD) exhibit altered metabolic disturbances that cannot be attributed to the loss of dystrophin directly. We tested the hypothesis that glycogen metabolism is defective in mdx dystrophic mice. RESULTS: Dystrophic mdx mice had increased skeletal muscle glycogen (79%, (P<0.01)). Skeletal muscle glycogen synthesis is initiated by glycogenin, the expression of which was increased by 50% in mdx mice (P<0.0001). Glycogen synthase activity was 12% higher (P<0.05) but glycogen branching enzyme activity was 70% lower (P<0.01) in mdx compared with wild-type mice. The rate-limiting enzyme for glycogen breakdown, glycogen phosphorylase, had 62% lower activity (P<0.01) in mdx mice resulting from a 24% reduction in PKA activity (P<0.01). In mdx mice glycogen debranching enzyme expression was 50% higher (P<0.001) together with starch-binding domain protein 1 (219% higher; P<0.01). In addition, mdx mice were glucose intolerant (P<0.01) and had 30% less liver glycogen (P<0.05) compared with control mice. Subsequent analysis of the enzymes dysregulated in skeletal muscle glycogen metabolism in mdx mice identified reduced glycogenin protein expression (46% less; P<0.05) as a possible cause of this phenotype. CONCLUSION: We identified that mdx mice were glucose intolerant, and had increased skeletal muscle glycogen but reduced amounts of liver glycogen.

Tranilast administration reduces fibrosis and improves fatigue resistance in muscles of mdx dystrophic mice.

BACKGROUND: Duchenne muscular dystrophy (DMD) is a severe and progressive muscle-wasting disorder caused by mutations in the dystrophin gene that result in the absence of the membrane-stabilising protein dystrophin. Dystrophic muscle fibres are susceptible to injury and degeneration, and impaired muscle regeneration is associated with fibrotic deposition that limits the efficacy of potential pharmacological, cell- and gene-based therapies. Novel treatments that can prevent or attenuate fibrosis have important clinical merit for DMD and related neuromuscular diseases. We investigated the therapeutic potential for tranilast, an orally bioavailable anti-allergic agent, to prevent fibrosis in skeletal muscles of mdx dystrophic mice. RESULTS: Three-week-old C57Bl/10 and mdx mice received tranilast (~300 mg/kg) in their food for 9 weeks, after which fibrosis was assessed through histological analyses, and functional properties of tibialis anterior muscles were assessed in situ and diaphragm muscle strips in vitro. Tranilast administration did not significantly alter the mass of any muscles in control or mdx mice, but it decreased fibrosis in the severely affected diaphragm muscle by 31% compared with untreated mdx mice (P < 0.05). A similar trend of decreased fibrosis was observed in the tibialis anterior muscles of mdx mice (P = 0.10). These reductions in fibrotic deposition were not associated with improvements in maximum force-producing capacity, but we did observe small but significant improvements in the resistance to fatigue in both the diaphragm and TA muscles of mdx mice treated with tranilast. CONCLUSION: Together these findings demonstrate that administration of potent antifibrotic compounds such as tranilast could help preserve skeletal muscle structure, which could ultimately increase the efficacy of pharmacological, cell and gene replacement/correction therapies for muscular dystrophy and related disorders.

Alterations in Notch signalling in skeletal muscles from mdx and dko dystrophic mice and patients with Duchenne muscular dystrophy.

New Findings What is the central question of this study? The Notch signalling pathway plays an important role in muscle regeneration, and activation of the pathway has been shown to enhance muscle regeneration in aged mice. It is unknown whether Notch activation will have a similarly beneficial effect on muscle regeneration in the context of Duchenne muscular dystrophy (DMD). What is the main finding and its importance? Although expression of Notch signalling components is altered in both mouse models of DMD and in human DMD patients, activation of the Notch signalling pathway does not confer any functional benefit on muscles from dystrophic mice, suggesting that other signalling pathways may be more fruitful targets for manipulation in treating DMD. Abstract In Duchenne muscular dystrophy (DMD), muscle damage and impaired regeneration lead to progressive muscle wasting, weakness and premature death. The Notch signalling pathway represents a central regulator of gene expression and is critical for cellular proliferation, differentiation and apoptotic signalling during all stages of embryonic muscle development. Notch activation improves muscle regeneration in aged mice, but its potential to restore regeneration and function in muscular dystrophy is unknown. We performed a comprehensive examination of several genes involved in Notch signalling in muscles from dystrophin-deficient mdx and dko (utrophin- and dystrophin-null) mice and DMD patients. A reduction of Notch1 and Hes1 mRNA in tibialis anterior muscles of dko mice and quadriceps muscles of DMD patients and a reduction of Hes1 mRNA in the diaphragm of the mdx mice were observed, with other targets being inconsistent across species. Activation and inhibition of Notch signalling, followed by measures of muscle regeneration and function, were performed in the mouse models of DMD. Notch activation had no effect on functional regeneration in C57BL/10, mdx or dko mice. Notch inhibition significantly depressed the frequency-force relationship in regenerating muscles of C57BL/10 and mdx mice after injury, indicating reduced force at each stimulation frequency, but enhanced the frequency-force relationship in muscles from dko mice. We conclude that while Notch inhibition produces slight functional defects in dystrophic muscle, Notch activation does not significantly improve muscle regeneration in murine models of muscular dystrophy. Furthermore, the inconsistent expression of Notch targets between murine models and DMD patients suggests caution when making interspecies comparisons.

Identification of FHL1 as a therapeutic target for Duchenne muscular dystrophy.

Utrophin is a potential therapeutic target for the fatal muscle disease, Duchenne muscular dystrophy (DMD). In adult skeletal muscle, utrophin is restricted to the neuromuscular and myotendinous junctions and can compensate for dystrophin loss in mdx mice, a mouse model of DMD, but requires sarcolemmal localization. NFATc1-mediated transcription regulates utrophin expression and the LIM protein, FHL1 which promotes muscle hypertrophy, is a transcriptional activator of NFATc1. By generating mdx/FHL1-transgenic mice, we demonstrate that FHL1 potentiates NFATc1 activation of utrophin to ameliorate the dystrophic pathology. Transgenic FHL1 expression increased sarcolemmal membrane stability, reduced muscle degeneration, decreased inflammation and conferred protection from contraction-induced injury in mdx mice. Significantly, FHL1 expression also reduced progressive muscle degeneration and fibrosis in the diaphragm of aged mdx mice. FHL1 enhanced NFATc1 activation of the utrophin promoter and increased sarcolemmal expression of utrophin in muscles of mdx mice, directing the assembly of a substitute utrophin-glycoprotein complex, and revealing a novel FHL1-NFATc1-utrophin signaling axis that can functionally compensate for dystrophin.

Intramuscular administration of PEGylated IGF-I improves skeletal muscle regeneration after myotoxic injury.

OBJECTIVE: Musculoskeletal injuries represent a major public health problem and drugs that can improve muscle repair and restore function are needed for patients with these conditions and other related muscular pathologies. Increasing insulin-like growth factor-I (IGF-I) levels in skeletal muscle improves regeneration after myotoxic injury and while administration of IGF-I has a potential for accelerating healing after trauma, optimizing its method of delivery and obviating potential side-effects currently associated with recombinant human (rh) IGF-I, remain a hurdle. DESIGN: We compared the treatment efficacy of rhIGF-I with a polyethylene glycol modified IGF-I (PEG-IGF-I) analog to improve functional repair of mouse tibialis anterior muscles after myotoxic injury, testing the hypothesis that PEG-IGF-I would exert greater beneficial effects on regenerating skeletal muscles than rhIGF-I due to improved pharmacokinetic properties. We also examined the relative efficacy of systemic versus local delivery of these IGF-I variants for improving functional muscle regeneration. RESULTS: Local delivery of PEG-IGF-I, but not rhIGF-I, at 4 days post-injury significantly improved early functional recovery as evident by a 27% increase in normalized force compared with saline control (P<0.05), whereas systemic application of either IGF-I variant was not effective. The improved function with intramuscular PEG-IGF-I administration was attributed to a greater and prolonged residence within the regenerating muscles, resulting in increased Akt activation and a 13% larger fiber cross-sectional area compared with rhIGF-I (P<0.05). CONCLUSIONS: These data support the hypothesis that PEG-IGF-I is more efficacious than rhIGF-I in hastening early fiber regeneration and improving muscle function after injury, highlighting its therapeutic potential for muscular pathologies.