Medial costal diaphragm remodeling after complete or partial ITTO.

<p>(<b>A</b>) CSA was significantly larger in type IIx/b fibers for ITTO, compared to ITTO-PAR and SHAM animals (‡ p<0.001). In addition, SHAM animals had significantly larger type IIx/b fibers than ITTO-PAR animals (* p<0.05). Type IIa fibers were significantly larger in ITTO animals, compared to ITTO-PAR (# p<0.01). (<b>B</b>) Although there were significantly fewer type IIx/b fibers in all groups (‡ p<0.001), training did not affect fiber type proportions. (<b>C</b>) Type IIx/b fibers occupied the largest A_A, regardless of group (‡ p<0.001).</p

Area fractions of remodeled tissue, including internally-nucleated cells, small/angular fibers, inflamed/necrotic fibers, and inflammatory cells.

<p>Area fractions of remodeled tissue, including internally-nucleated cells, small/angular fibers, inflamed/necrotic fibers, and inflammatory cells.</p

Demographic information in sham-occluded animals and animals treated with ITTO.

*<p>Values are median (IQR). ^† Values are mean ± SD. ^‡ Significant difference from ITTO, p<0.05. § Significant difference from ITTO, p<0.01.</p

Illustration of ITTO experimental setup.

<p>(<b>A</b>) ITTO was administered in a plethysmograph to permit limited animal movement. The tracheal cuff was inflated by injection of sterile saline into the actuating line. The timing and magnitude of cuff pressure was monitored with a polygraph. (<b>B</b>) Cut-out illustration of the tracheal cuff. The trachea was fully patent with the cuff deflated. With cuff inflated, the trachea was fully and reversibly occluded.</p

Area fraction (A_A) of remodeled cells in the medial costal diaphragm after 10-minute occlusion sessions (ITTO), 20-minute occlusion sessions (ITTO-20) or sham conditioning.

<p>Values are mean ± SD. *p<0.001Significant main effect for category: significantly different A_A from normal.</p

Area fraction (A_A) of remodeled cells in the costal and crural diaphragm.

<p>Values are mean ± SD. *p<0.001 Significant main effect for category: differences between A_A of normal, remodeled and connective tissue.</p

Remodeling of the medial costal diaphragm after 10- or 20-minute ITTO conditioning sessions.

<p>(<b>A</b>) CSA in type IIx/b fibers was significantly greater for ITTO (# p<0.01) and ITTO-20 (* p<0.05) animals, compared to SHAM animals. In all conditions, type IIx/b fibers were significantly larger than type I or IIa fibers (‡ p<0.001). (<b>B</b>) Type IIx/b fibers were significantly less prevalent than type I or type IIa (‡ p<0.001). The duration of ITTO sessions did not influence fiber type proportions. (<b>C</b>) Type IIx/b fibers occupied the largest A_A (‡ p<0.001), but there were no significant group variations in MHC A_A.</p

Regional remodeling of the diaphragm, following ITTO or SHAM training.

<p>(<b>A</b>) <b>CSA.</b> Type IIx/b CSA in the medial costal (#) and crural (*) regions was significantly greater in ITTO animals than SHAM animals. In the ITTO group, type IIx/b medial costal diaphragm fibers were significantly larger than dorsal, ventral and crural segments (†). In both groups, type IIx/b fibers were significantly larger than oxidative fibers (‡). (<b>B</b>) <b>Fiber proportions.</b> In the SHAM group, the ventral region contained significantly fewer type IIx/b fibers than the dorsal region (*). In all groups, type IIx/b fibers were significantly less prevalent than type IIa or I fibers (‡). (<b>C</b>) <b>MHC A_A.</b> The type I fiber A_A of the medial region was significantly larger in ITTO animals than SHAM animals (*). The A_A of type IIx/b fibers of the SHAM ventral diaphragm was significantly lower than the SHAM medial diaphragm (^#) and the ITTO ventral diaphragm (*). (Error bars are ± 1SD, *p<0.05; ^# p<0.01; ‡ p<0.001; † p<0.05 vs other regions).</p

¹³C NMR Metabolomics: Applications at Natural Abundance

¹³C NMR has many advantages for a metabolomics study, including a large spectral dispersion, narrow singlets at natural abundance, and a direct measure of the backbone structures of metabolites. However, it has not had widespread use because of its relatively low sensitivity compounded by low natural abundance. Here we demonstrate the utility of high-quality ¹³C NMR spectra obtained using a custom ¹³C-optimized probe on metabolomic mixtures. A workflow was developed to use statistical correlations between replicate 1D ¹³C and ¹H spectra, leading to composite spin systems that can be used to search publicly available databases for compound identification. This was developed using synthetic mixtures and then applied to two biological samples, <i>Drosophila melanogaster</i> extracts and mouse serum. Using the synthetic mixtures we were able to obtain useful ¹³C–¹³C statistical correlations from metabolites with as little as 60 nmol of material. The lower limit of ¹³C NMR detection under our experimental conditions is approximately 40 nmol, slightly lower than the requirement for statistical analysis. The ¹³C and ¹H data together led to 15 matches in the database compared to just 7 using ¹H alone, and the ¹³C correlated peak lists had far fewer false positives than the ¹H generated lists. In addition, the ¹³C 1D data provided improved metabolite identification and separation of biologically distinct groups using multivariate statistical analysis in the <i>D. melanogaster</i> extracts and mouse serum

T₁ weighted images of lower leg muscles.

<p>Representative T₁ weighted 3-D gradient echo images of A) control, and B) DMD subject. Outline of regions of interest (A) showing different muscles namely TA, tibialis anterior; EDL, extensor digitorum longus; Per, peroneal; MG, medial gastrocnemius; SOL, soleus; TP, tibilais posterior; FDL, flexor digitorum longus. (B) Showing areas of fatty tissue infiltration in leg muscles.</p