Chaperone co‐inducer BGP‐15 mitigates early contractile dysfunction of the soleus muscle in a rat ICU model

Figures

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  Figure 1. Survival in control and BGP‐15‐treated animals. Kaplan‐Meier survival plot showing 100% survival of the 13 BGP‐15‐treated animals killed at 5, 8 and 10 d. The corresponding survivals in control animals (n = 30) were 67% (5 d), and 39% (8‐10 d)

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  Figure 2. Rats body weight, soleus muscle weight, Myosin: actin ratio, and single muscle fibre CSA. The change in rats body weight (A), soleus muscle weight (B) and the per cent of soleus to body weight change (C) in controls and rats exposed to the ICU condition for 5, 8 and 10 d and rats exposed to BGP‐15 treatment and the ICU condition for 5, 8 and 10 d. Myosin: actin ratios in the soleus (control n = 5, 5d n = 3, 5d + BGP‐15 n = 4, 8d n = 3, 8d + BGP‐15 n = 3, 10d n = 5, and 10d + BGP‐15 n = 3) (D), and in the diaphragm (control n = 4, 5d n = 5, 5d + BGP‐15 n = 2, 8d n = 2, 8d + BGP‐15 n = 2, 10d n = 4, and 10d + BGP‐15 n = 2) (E) determined by 12% SDS‐PAGE, with the dots on the figure represent the individual ratio from each rat. Single muscle fibre CSA in the soleus (F) and in the diaphragm (G) measured at fixed sarcomeric length. In all the figure panels, the p values represent statistical significance when compared with control except when stated otherwise in the figure. Values are represented as means + SEM. *P < .05; **P < .01; ***P < .001

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  Figure 3. Single muscle fibres absolute force, specific force and myosin in vitro motility speed and force. Single muscle fibre absolute force measured at fixed sarcomeric length (A), and specific force (B) in soleus fibres in response to exposure to the ICU condition and BGP‐15 treatment for 5, 8 and 10 d. Single muscle fibre absolute force measured at fixed sarcomeric length (C), and specific force (D) in diaphragm fibres in response to exposure to the ICU condition and BGP‐15 treatment for 5 d. In vitro motility speed (E), force index (F) and efficiency (G) of myosin extracted from soleus single muscle fibre segments expressing the type I MyHC in rats exposed to the ICU condition for 5 d with and without BGP‐15 treatment. Values are represented as means + SEM. *P < .05; **P < .01; ***P < .001

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  Figure 4. β/slow (type I) MyHC post‐translational modifications. The BGP‐15 treatment was found to rescue the myosin motor protein from two PTMs (labelled in blue), that is, a methylation in the 1372 position (arginine) and oxidation in the 1774 position (aspartic acid). At the 1720 position (asparagine) we found an oxidation and in the 1749 position (arginine) and a methylation independent on BGP‐15 in response to 5 d mechanical ventilation (labelled in red)

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  Figure 5. mRNA expression of E3 ligases, MyHCI isoform and actin in the soleus. mRNA expression of the E3 ligases MuRF1 (A), Atrogin‐1(B), SMART (C) and Fbxo31 (D), and MyHCI (E), and actin (F) in the soleus of rats exposed to the ICU condition and BGP‐15 treatment for 5 and 10 d. For each experimental group the values were normalized to the controls. Values are presented as means + SEM. Significant differences compared with controls are denoted as *P < .05

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  Figure 6. mRNA expression of E3 ligases, MyHCs isoforms and actin in the diaphragm. mRNA expression of the E3 ligases MuRF1 (A), Atrogin‐1 (B), SMART (C) and Fbxo31 (D); myosin heavy chain isoforms MyHCIIa (E), MyHCIIx (F); and actin (G) in the diaphragm muscles of rats exposed to the ICU condition and BGP‐15 treatment for 5 and 10 d. The values for the different experimental groups were normalized to the control group. Values are presented as means + SEM. Significant differences compared with controls are denoted as *P < .05

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  Figure 7. Intermyofibrillar (IMF) and subsarcolemmal (SS) mitochondria structural changes. Electron micrographs captured at 10 000× magnification showing IMF and SS mitochondria structure in the soleus muscles of control animals (A), animal exposed to the ICU condition for 5 d (B), and 10 d (D), and animals treated with BGP‐15 and exposed to the ICU condition for 5 (C) and 10 d (E). Mitochondria structure at 40 000× magnification is shown in control animals (F), and animals exposed to 10 d ICU treatment (G). Ratio of IMF mitochondria abnormal fraction for all animal groups (H). Ratio of SS mitochondria abnormal fraction for all animal groups (I). Examples of normal mitochondria structure, with electron dense matrix and organized cristae, are labelled with yellow arrows, while some abnormal mitochondria, with swollen or vesicular structure, are labelled with red arrows. Values are presented as means + SEM

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  Figure 8. Mitochondrial dynamic and biogenesis. Mitochondrial dynamic gene expression (A‐D); mitochondrial fusion gene expression MFN1 (A), MFN2 (B), and OPA1 (C), and mitochondrial fission gene expression DRP1 (D). Mitochondrial biogenesis evaluated as the gene expression of total PGC‐1α (E) for each experimental group normalized to control group. Values are presented as means + SEM. Significant differences compared with controls are denoted as *P < .05