The caspase proteolytic system in callipyge and normal lambs in longissimus, semimembranosus, and infraspinatus muscles during postmortem storage1

Abstract

The objective of this experiment was to determine whether the caspase proteolytic system has a role in postmortem tenderization. Six ewes and 6 wethers that were noncarriers and 6 ewes and 6 wethers that were expressing the callipyge gene were used for this study. Caspase activities were determined in LM at 7 different time points during the postmortem storage period: 0 h, 4 h, 8 h, 24 h, 2 ci, 7 ci, and 21 cl and in semimembranosus (SM) and infraspinatus (IS) muscles at 0 h, 8 h. 24 h, and 7 d from callipyge and noncallipyge (normal) lambs. Calpastatin activity was determined at 0 h, 2 d. 7 d, and 21 d and slice shear force measured at 2, 7, and 21 d in the LM. Calpastatin activity and slice shear force were greater in LM from callipyge lambs than normal lambs at each time point (P < 0.001 and P < 0.0001, respectively). Caspases 3 and 7 are executioner caspases, and their combined activity was found to decrease during the postmortem storage period in LM, SM, and IS muscles from calhpyge and normal lambs. Similarly, activity of the initiator caspase (caspase 9) decreased (P < 0.05) in all 3 muscles across the postmortem storage period in callipyge and normal lambs, and its decrease in activity preceded that of the executioner caspases 3/7. A positive relationship also was detected between caspase 9 and caspase 3/7 in LM, SM. and IS muscles (P < 0.0001, r = 0.85, r = 0.86, r = 0.84, respectively), which is consistent with caspa.se 9 being responsible for the cleavage and activation of the executioner caspases (caspase 3/7) downstream. Caspase 3/7 and caspase 9 activities at 8 h in SM were greater in normal lamb than callipyge lamb (P < 0.05), with a trend for caspase 3/7 activity to be greater at 24 h postmortem (P = 0.0841). There also was a trend for caspase 3/7 activity to be greater in LM at 21 cliii nornial lamb than in callipyge lamb (P = 0.053), although there were no differences detected in caspase activities between genotypes in the IS muscle, which is not affected by the callipyge gene. A negative relationship also was detected between peak caspa.se 3/7 activity at 8 Ii in LM from normal lambs and calpastatin activity at 0 and 2 d (r =-0.65, r =-0.68, respectively, P < 0.05). This relationship was not observed in LM from callipyge lambs, suggesting that caspase 3/7 may be cleaving calpastatin in normal lambs but the level of calpastatin in callipyge lambs is such that caspase 3/7 cannot degrade it sufficiently to overcome the increased content of caipastatin, and thus, calpastatin activity is the overriding factor in postmortem proteolysis in these animals. There was no direct evidence from this study that ca.spases have a significant role in postmortem tenderization, but they may have some role through calpastatin degradation.

Autophagy during beef aging

“…Caspase activity changes across the postmortem conditioning period of pork, chicken, and beef, with the highest activities always detected in the early postmortem phase. 8,[28][29][30] Changes in CASP3/caspase 3 and CASP7, as well as CASP9 activity during the early phase of conditioning of porcine LD muscle have been identified to have a negative relationship with shear force; thus, the more caspase activation, the greater the change and the lower the shear force value, and therefore the higher the level of meat tenderness. 8 However, other authors 31 when examining changes in CASP3 activity and protein expression in beef muscle samples during postmortem aging, only detected the inactive isoform of CASP3, not the active one, and no association between caspase isoforms and shear force values was found.…”

Influence of ultrasound and proteolytic enzyme inhibitors on muscle degradation, tenderness, and cooking loss of hens during aging

“…A negative relationship was also detected between the peak caspase-3/7 activity at 8 h in longissimus muscles from normal lambs and calpastatin activity at 0 day and 2 days (r = -0.65, r = -0.68, respectively, P < 0.05). Again, in our experiment, calpastatin activity decreased with increased time, as found by Kemp et al (2009). This is therefore evidence for the indirect involvement of caspase in post mortem proteolysis and meat tenderisation, although other studies suggest a direct role for caspases and apoptosis in post mortem proteolysis.…”

“…Determination of Caspase 3/7 activity. Caspase 3/7 activity was determined in breast muscle samples according to the method of Wagner et al (2003) and Kemp et al (2009). Frozen samples were crushed in liquid nitrogen, and 1 g was homogenised in 3 volumes of extraction buffer [25 mmol/l HEPES (pH = 7.5), 1 mmol/l EGTA, 1 mmol/l EDTA, 0.1% (v/v) Triton X-100, 5 mmol/l MgCl 2 , 2 mmol/l DTT(1,4-dithiothreitol), (ethylene glycol tetra-acetic acid), 74 μmol/l antipain, 20 μmol/l leupeptin, 15 μmol/l pepstatin, 0.15 μmol/l aprotinin] for 2 × 20 seconds.…”

Aging/Tenderization Mechanisms

“…There is relatively little data, however, on the interaction between caspases and the calpain enzyme system in skeletal muscle as it relates to postmortem proteolysis. A negative relationship between peak caspase 3/7 activity at 8 hours postmortem and calpastatin activity at 0 and 2 days postmortem has been observed in the muscles of normal lambs but not in callipyge lambs (Kemp et al 2009 ). Thus, while there is no direct evidence that caspases contribute signifi cantly to postmortem tenderization, data suggest that they may play an indirect role by degrading calpastatin.…”

“…In this study, pro -caspase 3 was not activated during postmortem storage and caspase 3 activity was not correlated with Warner -Bratzler shear force in beef longissimus . The data from one study using muscle from callipyge and normal lambs indicated that caspase 3/7 and caspase 9 activities decreased between 1 and 21 days postmortem but did not directly support or reject the involvement of the caspase system in meat tenderization (Kemp et al 2009 ).…”