calpain and Burns

Calpains are intracellular nonlysosomal Ca(2+-) regulated cysteine proteases, widely located in the tissues of most mammals. Skeletal muscle tissue mainly expresses m-calpain, µ-caplain, n-calpain, and their endogenous inhibitor calpastatin. They are closely related to the cell apoptosis, cytoskeleton formation, cell cycles, etc. Calpains are also considered to be participating in the protein degradation process. Severe burns are typically followed by hypermetabolic responses that are characterized by hyperdynamic circulatory responses with increased proteolysis and cell apoptosis. Recently, overloading of Ca(2+) in skeletal muscle cells, which activates the calpains is observed after a serious burn. This paper aims to review the current research of the relationship between calpains and post-burn skeletal muscle wasting from the perspectives of structure, function, and physiological activities.

Topics: Animals; Burns; Calpain; Muscle, Skeletal

Mitochondrial damage is an important cause of heart dysfunction after severe burn injury. However, the pathophysiological process remains unclear. This study aims to examine the mitochondrial dynamics in the heart and the role of μ-calpain, a cysteine protease, in this scenario. Rats were subjected to severe burn injury treatment, and the calpain inhibitor MDL28170 was administered intravenously 1 h before or after burn injury. Rats in the burn group displayed weakened heart performance and decreased mean arterial pressure, which was accompanied by a diminishment of mitochondrial function. The animals also exhibited higher levels of calpain in mitochondria, as reflected by immunofluorescence staining and activity tests. In contrast, treatment with MDL28170 before any severe burn diminished these responses to a severe burn. Burn injury decreased the abundance of mitochondria and resulted in a lower percentage of small mitochondria and a higher percentage of large mitochondria. Furthermore, burn injury caused an increase in the fission protein DRP1 in the mitochondria and a decrease in the inner membrane fusion protein OPA1. Similarly, these alterations were also blocked by MDL28170. Of note, inhibition of calpain yielded the emergence of more elongated mitochondria along with membrane invagination in the middle of the longitude, which is an indicator of the fission process. Finally, MDL28170, administered 1 h after burn injury, preserved mitochondrial function and heart performance, and increased the survival rate. Overall, these results provided the first evidence that mitochondrial recruitment of calpain confers heart dysfunction after severe burn injury, which involves aberrant mitochondrial dynamics.

Topics: Animals; Burns; Calpain; Mitochondria; Mitochondrial Dynamics; Rats

Hypertrophic scars, the most common complication of burn injuries, are characterized by excessive deposition of fibroblast-derived extracellular matrix proteins. Calpain, a calcium-dependent protease, is involved in the fibroblast proliferation and extracellular matrix production observed in certain fibrotic diseases. However, its role in the formation of post-burn hypertrophic skin scars remains largely unknown. Here, calpain expression and activity were assessed in skin fibroblasts obtained directly from patients with third-degree burns, who consequently developed post-burn hypertrophic scars. Furthermore, the antifibrotic effect of calpastatin, an endogenous calpain inhibitor, was evaluated in human fibroblasts and a murine burn model. The activity, mRNA levels, and protein levels of calpain were markedly higher in fibroblasts from the burn wounds of patients than in normal cells. Selective calpain inhibition by calpastatin markedly reduced not only the proliferation of burn-wound fibroblasts but also the mRNA and protein expression of calpain, transforming growth factor-beta 1, α-smooth muscle actin, type I and type III collagens, fibronectin, and vimentin in burn-wound fibroblasts. The anti-scarring effects of calpastatin were validated using a murine burn model by molecular, histological, and visual analyses. This study demonstrates the pathological role of calpain and the antifibrotic effect of calpastatin via calpain inhibition in post-burn hypertrophic scar formation.

Topics: Adult; Animals; Burns; Calcium-Binding Proteins; Calpain; Cell Proliferation; Cicatrix, Hypertrophic; Collagen Type III; Extracellular Matrix; Extracellular Matrix Proteins; Female; Fibroblasts; Fibronectins; Humans; Hypertrophy; Male; Mice; Middle Aged; RNA, Messenger; Skin; Transforming Growth Factor beta1; Young Adult

Severe burns are typically followed by hypermetabolism characterized by significant muscle wasting, which causes considerable morbidity and mortality. The aim of the present study was to explore the underlying mechanisms of skeletal muscle damage/wasting post-burn. Rats were randomized to the sham, sham+4-phenylbutyrate (4-PBA, a pharmacological chaperone promoting endoplasmic reticulum (ER) folding/trafficking, commonly considered as an inhibitor of ER), burn (30% total body surface area), and burn+4-PBA groups; and sacrificed at 1, 4, 7, 14 days after the burn injury. Tibial anterior muscle was harvested for transmission electron microscopy, calcium imaging, gene expression and protein analysis of ER stress / ubiquitin-proteasome system / autophagy, and calpain activity measurement. The results showed that ER stress markers were increased in the burn group compared with the sham group, especially at post-burn days 4 and 7, which might consequently elevate cytoplasmic calcium concentration, promote calpain production as well as activation, and cause skeletal muscle damage/wasting of TA muscle after severe burn injury. Interestingly, treatment with 4-PBA prevented burn-induced ER swelling and altered protein expression of ER stress markers and calcium release, attenuating calpain activation and skeletal muscle damage/wasting after severe burn injury. Atrogin-1 and LC3-II/LC3-I ratio were also increased in the burn group compared with the sham group, while MuRF-1 remained unchanged; 4-PBA decreased atrogin-1 in the burn group. Taken together, these findings suggested that severe burn injury induces ER stress, which in turns causes calpain activation. ER stress and subsequent activated calpain play a critical role in skeletal muscle damage/wasting in burned rats.

Topics: Animals; Burns; Calcium; Calpain; Endoplasmic Reticulum Stress; Enzyme Activation; Homeostasis; Male; Muscle, Skeletal; Phenylbutyrates; Proteolysis; Rats; Rats, Wistar

37 degrees' warm water for 12 s; Burn group: 94 degrees' boiling water for 12 s; Blast group: 5 g cyclonite explode in 75 cm distance from left chest wall of rat; Burn-blast group: burn group and blast group combined modeling method. At 6, 24, 48, 72 h observation points after injury, abdominal aorta blood samples and myocardial specimen were collected. Left ventricular ejection fraction (EF), left ventricular fractional shortening index (FS) were measured through color Doppler ultrasound instrument; Myocardial tissue was stained with hematoxylin-eosin (HE); serum cardiac troponin I (CTnI) and creatine kinase isoenzyme (CK-MB) were detected; detection of cell apoptosis in myocardial tissue was performed by terminal deoxynucleotidyl transferase-mediated dUTP notch labeling technique (Tunel). Expression levels of calpain mRNA level and protein were detected with Real-time fluorescent quantitative polymerase chain reaction (RT-PCR) and Western imprinting method analysis; calpain activity was detected by fluorescence spectrophotometry.

Topics: Animals; Blast Injuries; Burns; Calpain; Male; Myocardium; Rats; Rats, Sprague-Dawley

To explore the effect and mechanism of ryanodine receptor antagonist dantrolene on skeletal muscle of rats with severe scald injury.. A total of 56 Wistar rats were divided into control, scald and dantrolene treatment groups according to a random digital table. Rats in scald and dantrolene treatment groups were subject to 50% total body surface area (TBSA) full-thickness scald by a 12-second immersion of back and a 6-second immersion of abdomen in 94 °C water and then received an intraperitoneal injection of Ringer's solution. At the same time, the rats in scald group received 5% mannitol through caudal vein while those in dantrolene treatment group received dantrolene 2 mg/kg (dissolved in 5% mannitol). Rats in control group were sham-injured through an immersion of back and abdomen into 37 °C warm water. Tibialis anterior muscle samples were harvested at Days 1, 4 and 7 post-scalding. Changes of skeletal muscle ultrastructure were observed by transmission electron microscope, subcellular calcium ion (Ca(2+)) contents of skeletal muscle (including cytoplasm, mitochondria & sarcoplasm reticulum) were detected by electron probe X-ray microanalysis (EPMA) and the levels of calpain-1 and calpain-2 protein were determined by Western blot. And the activities of calpain were detected by enzyme-linked immunosorbent assay.. In scald group, assorted arrangement appeared immediately at Day 1 post-injury and partial disappearance of Z lines at Day 7 post-injury. There were no significant ultrastructure changes in dantrolene treatment group at Day 1 and 4 post-injury. Curled filament and mild fracture occurred merely in dantrolene treatment group at Day 7 post-injury. The cytoplasmic contents of Ca(2+) were significantly higher in scald group than those in control group at Day 1 and 4 ((0.964 ± 0.060), (0.639 ± 0.067) vs (0.266 ± 0.029) µmol/L respectively, all P < 0.05) while the contents of Ca(2+) within sarcoplasm reticulum were obviously lower in scald group than those in control group at Day 1 and 4 ((0.368 ± 0.060), (0.814 ± 0.089) vs (1.337 ± 0.112) µmol/L respectively, all P < 0.05). However, those subcellular regions in dantrolene treatment group ((0.310 ± 0.069), (0.490 ± 0.039) and (1.241 ± 0.073), (1.161 ± 0.094) µmol/L) had no significant difference with control group (all P > 0.05). Calpain-1 and calpain-2 protein levels in scald group increased significantly at Day 1 and 4 post-injury versus control group (1.371 ± 0.034, 1.214 ± 0.030 vs 0.838 ± 0.017 & 1.464 ± 0.015, 1.390 ± 0.023 vs 0.806 ± 0.026 respectively, all P < 0.05), whereas calpain-1 and calpain-2 protein levels in dantrolene treatment (0.984 ± 0.031, 0.935 ± 0.023 and 0.836 ± 0.014, 0.741 ± 0.020) obviously were lower than those in scald group respectively (all P < 0.05). The activities of calpain in scald and dantrolene treatment groups at Day 1, 4 and 7 post-injury were (8.33 ± 0.21), (9.33 ± 0.21), (10.59 ± 0.18) and (7.76 ± 0.28), (7.86 ± 0.20), (7.91 ± 0.22) µmol/L respectively while the activity of calpain in control group was (7.62 ± 0.19) µmol/L. The activities of calpain in scald group were significantly higher than those in dantrolene treatment and control groups (all P < 0.05) whereas the activities of calpain in dantrolene treatment group had no obvious change versus control group (all P > 0.05).. Dantrolene offers significant protection from skeletal muscle tissue damage and minimizes the ultrastructural change of tibialis anterior muscle induced by severe scald injury. The mechanism is probably through inhibiting an excessive release of Ca(2+) within sarcoplasm reticulum and down-regulated cytoplasmic expression and activity of calpain-1 and calpain-2.

Topics: Animals; Burns; Calcium; Calpain; Dantrolene; Disease Models, Animal; Male; Muscle, Skeletal; Rats; Rats, Wistar