Elevation of fast but not slow troponin I in the circulation of patients with Becker and Duchenne muscular dystrophy.

INTRODUCTION: One of the hallmarks of injured skeletal muscle is the appearance of elevated skeletal muscle proteins in circulation. Human skeletal muscle generally consists of a mosaic of slow (type I) and fast (type IIa, IIx/d) fibers, defined by their myosin isoform expression. Recently, measurement of circulating fiber-type specific isoforms of troponin I has been used as a biomarker to suggest that muscle injury in healthy volunteers (HV) results in the appearance of muscle proteins from fast but not slow fibers. We sought to understand if this is also the case in severe myopathy patients with Becker and Duchenne muscular dystrophy (BMD, DMD). METHODS: An enzyme-linked immunosorbent assay (ELISA) that selectively measures fast and slow skeletal troponin I (TNNI2 and TNNI1) was used to measure a cross-section of patient plasma samples from HV (N = 50), BMD (N = 49), and DMD (N = 132) patients. Creatine kinase (CK) activity was also measured from the same samples for comparison. RESULTS: TNNI2 was elevated in BMD and DMD and correlated with the injury biomarker, CK. In contrast, TNNI1 levels were indistinguishable from levels in HV. There was an inverse relationship between CK and TNNI2 levels and age, but no relationship for TNNI1. DISCUSSION: We define a surprising discrepancy between TNNI1 and TNNI2 in patient plasma that may have implications for the interpretation of elevated muscle protein levels in dystrophinopathies.

Biomarker-focused multi-drug combination therapy and repurposing trial in mdx mice.

Duchenne muscular dystrophy is initiated by dystrophin deficiency, but downstream pathophysiological pathways such as membrane instability, NFĸB activation, mitochondrial dysfunction, and induction of TGFß fibrosis pathways are thought to drive the disability. Dystrophin replacement strategies are hopeful for addressing upstream dystrophin deficiency; however, all methods to date use semi-functional dystrophin proteins that are likely to trigger downstream pathways. Thus, combination therapies that can target multiple downstream pathways are important in treating DMD, even for dystrophin-replacement strategies. We sought to define blood pharmacodynamic biomarkers of drug response in the mdx mouse model of Duchenne muscular dystrophy using a series of repurposed drugs. Four-week-old mdx mice were treated for four weeks with four different drugs singly and in combination: vehicle, prednisolone, vamorolone, rituximab, ß-aminoisobutyric acid (BAIBA) (11 treatment groups; n = 6/group). Blood was collected via cardiac puncture at study termination, and proteomic profiling was carried out using SOMAscan aptamer panels (1,310 proteins assayed). Prednisolone was tested alone and in combination with other drugs. It was found to have a good concordance of prednisolone-responsive biomarkers (56 increased by prednisolone, 39 decreased) focused on NFκB and TGFß cascades. Vamorolone shared 45 (80%) of increased biomarkers and 13 (33%) of decreased biomarkers with prednisolone. Comparison of published human corticosteroid-responsive biomarkers to our mdx data showed 14% (3/22) concordance between mouse and human. Rituximab showed fewer drug-associated biomarkers, with the most significant being human IgG. On the other hand, BAIBA treatment (high and low dose) showed a drug-associated increase in 40 serum proteins and decreased 5 serum proteins. Our results suggest that a biomarker approach could be employed for assessing drug combinations in both mouse and human studies.

Serum biomarkers associated with baseline clinical severity in young steroid-naïve Duchenne muscular dystrophy boys.

Duchenne muscular dystrophy (DMD) is caused by loss of dystrophin in muscle, and while all patients share the primary gene and biochemical defect, there is considerable patient-patient variability in clinical symptoms. We sought to develop multivariate models of serum protein biomarkers that explained observed variation, using functional outcome measures as proxies for severity. Serum samples from 39 steroid-naïve DMD boys 4 to <7 years enrolled into a clinical trial of vamorolone were studied (NCT02760264). Four assessments of gross motor function were carried out for each participant over a 6-week interval, and their mean was used as response for biomarker models. Weighted correlation network analysis was used for unsupervised clustering of 1305 proteins quantified using SOMAscan® aptamer profiling to define highly representative and connected proteins. Multivariate models of biomarkers were obtained for time to stand performance (strength phenotype; 17 proteins) and 6 min walk performance (endurance phenotype; 17 proteins) including some shared proteins. Identified proteins were tested with associations of mRNA expression with histological severity of muscle from dystrophinopathy patients (n = 28) and normal controls (n = 6). Strong associations predictive of both clinical and histological severity were found for ERBB4 (reductions in both blood and muscle with increasing severity), SOD1 (reductions in muscle and increases in blood with increasing severity) and CNTF (decreased levels in blood and muscle with increasing severity). We show that performance of DMD boys was effectively modeled with serum proteins, proximal strength associated with growth and remodeling pathways and muscle endurance centered on TGFß and fibrosis pathways in muscle.

Exposure to cuticular bacteria can alter host behavior in a funnel-weaving spider.

Contact with environmental microbes are arguably the most common species interaction in which any animal participates. Studies have noted diverse relationships between hosts and resident microbes, which can have strong consequences for host development, physiology, and behavior. Many of these studies focus specifically on pathogens or beneficial microbes, while the benign microbes, of which the majority of bacteria could be described, are often ignored. Here, we explore the nature of the relationships between the grass spider Agelenopsis pennsylvanica and bacteria collected from their cuticles in situ. First, using culture-based methods, we identified a portion of the cuticular bacterial communities that are naturally associated with these spiders. Then, we topically exposed spiders to a subset of these bacterial monocultures to estimate how bacterial exposure may alter 3 host behavioral traits: boldness, aggressiveness, and activity level. We conducted these behavioral assays 3 times before and 3 times after topical application, and compared the changes observed in each trait with spiders that were exposed to a sterile control treatment. We identified 9 species of bacteria from the cuticles of 36 spiders and exposed groups of 20 spiders to 1 of 4 species of cuticular bacteria. We found that exposure to Dermacoccus nishinomiyaensis and Staphylococcus saprophyticus was associated with a 10-fold decrease in the foraging aggressiveness of spiders toward prey in their web. Since bacterial exposure did not have survival consequences for hosts, these data suggest that interactions with cuticular bacteria, even non-pathogenic bacteria, could alter host behavior.

Erratum: Exposure to cuticular bacteria can alter host behavior in a funnel-weaving spider.

[This corrects the article DOI: 10.1093/cz/zox064.]

The primary case is not enough: Variation among individuals, groups and social networks modify bacterial transmission dynamics.

The traits of the primary case of an infectious disease outbreak, and the circumstances for their aetiology, potentially influence the trajectory of transmission dynamics. However, these dynamics likely also depend on the traits of the individuals with whom the primary case interacts. We used the social spider Stegodyphus dumicola to test how the traits of the primary case, group phenotypic composition and group size interact to facilitate the transmission of a GFP-labelled cuticular bacterium. We also compared bacterial transmission across experimentally generated "daisy-chain" vs. "star" networks of social interactions. Finally, we compared social network structure across groups of different sizes. Groups of 10 spiders experienced more bacterial transmission events compared to groups of 30 spiders, regardless of groups' behavioural composition. Groups containing only one bold spider experienced the lowest levels of bacterial transmission regardless of group size. We found no evidence for the traits of the primary case influencing any transmission dynamics. In a second experiment, bacteria were transmitted to more individuals in experimentally induced star networks than in daisy-chains, on which transmission never exceeded three steps. In both experimental network types, transmission success depended jointly on the behavioural traits of the interacting individuals; however, the behavioural traits of the primary case were only important for transmission on star networks. Larger social groups exhibited lower interaction density (i.e. had a low ratio of observed to possible connections) and were more modular, i.e. they had more connections between nodes within a subgroup and fewer connections across subgroups. Thus, larger groups may restrict transmission by forming fewer interactions and by isolating subgroups that interacted with the primary case. These findings suggest that accounting for the traits of single exposed hosts has less power in predicting transmission dynamics compared to the larger scale factors of the social groups in which they reside. Factors like group size and phenotypic composition appear to alter social interaction patterns, which leads to differential transmission of microbes.

Individual differences in boldness influence patterns of social interactions and the transmission of cuticular bacteria among group-mates.

Despite the importance of host attributes for the likelihood of associated microbial transmission, individual variation is seldom considered in studies of wildlife disease. Here, we test the influence of host phenotypes on social network structure and the likelihood of cuticular bacterial transmission from exposed individuals to susceptible group-mates using female social spiders (Stegodyphus dumicola). Based on the interactions of resting individuals of known behavioural types, we assessed whether individuals assorted according to their behavioural traits. We found that individuals preferentially interacted with individuals of unlike behavioural phenotypes. We next applied a green fluorescent protein-transformed cuticular bacterium,Pantoeasp., to individuals and allowed them to interact with an unexposed colony-mate for 24 h. We found evidence for transmission of bacteria in 55% of cases. The likelihood of transmission was influenced jointly by the behavioural phenotypes of both the exposed and susceptible individuals: transmission was more likely when exposed spiders exhibited higher 'boldness' relative to their colony-mate, and when unexposed individuals were in better body condition. Indirect transmission via shared silk took place in only 15% of cases. Thus, bodily contact appears key to transmission in this system. These data represent a fundamental step towards understanding how individual traits influence larger-scale social and epidemiological dynamics.