Effects of <i>AMPD1</i> C34T and MAD activity on the metabolic-chronotropic relationship.

<p>A Representative plots of the heart rate versus oxygen consumption according to MAD activity in three patients and in one control subject. For comparison, the four subjects have similar anthropometric data, i.e. similar predicted values for maximal exercise testing data. f, female. MCR, metabolic-chronotropic relationship. B Scatter plots reporting the metabolic-chronotropic relationship according to the <i>AMPD1</i> C34T genotype (left panel) and the MAD activity (right panel). Horizontal bars represent the mean. Rs, Spearman’s rank correlation coefficient along with associated two-tailed <i>p</i> value. *Significantly different from the C34T-genotype spearman correlation coefficient (Williams-Steiger’ t-test, <i>p</i> = 0.015). Intergroup comparison was performed by one way ANOVA followed by the Scheffe post hoc test. <b>†</b> significantly different from MAD Absent (<i>p</i> < 0.05). ‡ significantly different from MAD Absent (<i>p</i> < 0.001). § significantly different from MAD Decreased (<i>p</i> < 0.01).</p

Confusion matrix of MAD activity in skeletal muscle vs. the most common <i>AMPD1</i> mutation in patients.

<p>Confusion matrix of MAD activity in skeletal muscle vs. the most common <i>AMPD1</i> mutation in patients.</p

<i>AMPD1</i> rare variants carriers phenotype for the metabolic-chronotropic relationship and the predicted maximal O₂ pulse.

<p><i>AMPD1</i> rare variants carriers phenotype for the metabolic-chronotropic relationship and the predicted maximal O₂ pulse.</p

Effects of MAD activity on the % age-predicted O₂ pulse.

<p>Horizontal bars represent the mean.* MAD defect (Absent MAD activity + Decreased MAD activity) significantly different from normal (Normal MAD activity + Control) according to the Scheffé grouping post hoc test (<i>p</i> < 0.01). Inset, O₂ pulse is determined by stroke volume (SV), hemoglobin content (Hb) and arteriovenous oxygen saturation difference.</p

Flowchart of the study.

<p>Muscle biopsies were analysed using standard techniques including histoenzymology with Periodic acid-Schiff (PAS) and MAD staining (Fishbein’s method [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132972#pone.0132972.ref035" target="_blank">35</a>]).</p

Effects of incremental exercise on plasma metabolite levels according to muscle MAD activity.

<p>Patients performed an incremental exercise testing (<i>inset</i>). Blood was sampled before (Rest), during exercise (50% of Predicted Maximal Power and Peak exercise), and after exercise (2, 5, 10 and 15 min recovery). Filled symbols correspond to glycogenoses, open symbols correspond to the absence of glycogenoses. (A) Ammonia. (B) Ammonia/rest. (C) Lactate. (D) Lactate/rest. (E) Lactate/Pyruvate ratio. Data are represented as means ± standard error of mean (error bars not included for ammonia values in the subgroup with glycogenose and normal MAD in panel A).*: Absent vs. Normal MAD staining, †: Decreased vs. Normal MAD staining (<i>P</i> < 0.05, Games-Howell <i>post-hoc</i> test).</p

Diagnostic algorithm for glycogenoses and MAD deficiencies.

<p>(A) The algorithm classifies subjects referred for metabolic exercise testing into four groups. The figure combines the optimal cut-off values reported in decision tree nodes (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132972#pone.0132972.g004" target="_blank">Fig 4A and 4B</a>). L/P: lactate-to-pyruvate ratio, R: Recovery from exercise. (B) A contingency table was constructed on the basis of whether subjects have a metabolic myopathy or not, and corresponding common performance metrics with 95% confidence interval were calculated. TP: True positive, FP: False positive, FN: False negative, TN: True negative.</p

Anthropometric characteristics and maximal exercise test data.

<p>Data are means ± SD. MAD: Myoadenylate deaminase, f: Female, m: Male, BMI: Body Mass Index</p><p>*: Different from the Normal MAD activity group (<i>P</i> < 0.05, Games-Howell <i>post-hoc</i> test).</p><p>Anthropometric characteristics and maximal exercise test data.</p

Diagnostic performance of blood parameters to discriminate abnormal from normal MAD activity (A) and decreased from absent MAD activity (B).

<p>Diagnostic indices for predictors, with areas under receiver operating characteristic curves (AUC) ≥ 0.750 listed in the tables. The best AUC for each parameter is shown in the ROC space, with the corresponding 95% confidence interval. Circles denote cut-off points, corresponding to the highest concomitant sensitivity and specificity. (A) Performance of blood parameters to discriminate between Normal (n = 37) and Abnormal (n = 11) MAD activity. (B) Classifiers for the differentiation of Absent (n = 5) from Decreased (n = 6) MAD activity. 50% PMP: 50% from predicted maximal power (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132972#sec006" target="_blank">Methods</a>), R: Recovery from exercise, Se: Sensibility, Sp: Specificity, Acc: Accuracy, PPV: Positive predictive value, NPV: Negative predictive value, LR+: Positive likelihood ratio, LR-: Negative likelihood ratio, DOR: Diagnostic odds ratio, NC: Not calculable.</p

Histochemical MAD staining using <i>p</i>-nitro blue tetrazolium.

<p>(A) Relative <i>p</i>-NBT staining intensity was expressed as a percentage of the mean optical density in control muscle biopsies (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132972#sec006" target="_blank">Methods</a>). Horizontal bars represent the mean value of each group. *: Decreased vs. Normal MAD staining, †: Absent vs. Normal MAD staining, ‡: Absent vs. Decreased MAD staining (<i>P</i> < 0.05, Games-Howell <i>post-hoc</i> test). Representative serial cross sections of lateral vastus biopsies: (B) Normal, (C) Decreased, and (D) Absent MAD staining (original magnification: × 100).</p