The objective of this study was to investigate the potential role of the caspase protease family in meat tenderisation by examining if caspase 3 was capable of causing myofibril protein degradation. Full-length human recombinant caspase 3 (rC3) was expressed in Escherichiacoli and purified. The rC3 was active in the presence of myofibrils isolated from porcine longissimus dorsi muscle (LD) and retained activity in a buffer system closely mimicking post mortem conditions. The effect of increasing concentrations of rC3, incubation temperature, as well as incubation time on the degradation of isolated myofibril proteins were all investigated in this study. Myofibril protein degradation was determined by SDS-PAGE and Western blotting. There was a visible increase in myofibril degradation with a decrease in proteins identified as desmin and troponin I and the detection of protein degradation products at approximately 32, 28 and 18 kDa with increasing concentrations of rC3. These degradation products were analysed using MALDI-TOF mass spectrometry and identified to occur from the proteolysis of actin, troponin T and myosin light chain, respectively. The production of these degradation products was not inhibited by 5 mM EDTA or semi-purified calpastatin but was inhibited by the caspase-specific inhibitor Ac-DEVD-CHO. The temperature at which isolated myofibrils were incubated with rC3 was also found to affect degradation, with increasing incubation temperatures causing increased desmin degradation and cleavage of pro-caspase 3 into its active isoform. Incubation of isolated myofibrils at 4°C for 5 days with rC3 resulted in the visible degradation of a number of myofibril proteins including desmin and troponin I. This study has shown that rC3 is capable of causing myofibril degradation, hydrolysing myofibril proteins under conditions that are similar to those found in muscle in the post mortem conditioning period.

The present study was conducted to determine the effect of growth rate of calves on the activity of the calpain system post mortem, post mortem desmin degradation, myofibrillar fragmentation index and meat tenderness of m. longissimus lumborum (LL) and m. supraspinatus (SS). Twenty-four Friesian heifer calves were allocated to two treatment groups: MM and HH. The MM calves were given food to achieve a moderate growth rate (678 g/day from 5 days of age to 90 kg body weight (BW) (period I) and 770 g/day from 90 kg BW to slaughter at 250 kg BW (period II)) and the HH calves were given food to achieve a high growth rate (period I: 895 g/day and period II: 1204 g/day). The myofibril fragmentation index (MFI), desmin degradation and the activity of µ-calpain, m-calpain and calpastatin were measured in LL and SS at slaughter, and 1 and 7 days post mortem. Shear force was measured in LL and SS after 1 and 7 days of ageing and a sensory panel evaluated the eating quality of the loin aged 7 days. MFI (P < 0·01) and tenderness (P < 0·01) were higher and shear force was lower (P < 0·001) in meat from HH calves compared with meat from MM calves. However, growth rate did not affect the activity of µ-calpain, m-calpain or calpastatin or the degradation pattern of desmin at any time post mortem. Besides growth rate and MFI at day 7 post mortem, which both correlated positively with meat tenderness (P < 0·001 and P < 0·01, respectively), fibre type traits and DNA or RNA concentrations were not generally related to final tenderness. Thus, it was concluded that increased growth rate of calves before slaughter increased the ease of fragmentation (MFI) of the meat and improved the tenderness but these findings could not be related to any changes in the activity of the calpain system.

In this report, we describe the distribution of the platelet-derived growth factor receptor α (PDGFRα) by immunolocalization in the embryonic day 10.5 mouse heart and defects in heart development associated with the absence of this receptor in the Patch mouse. The Patch mouse is a naturally occurring mutant that has been accepted as a model for determining the role of the PDGFRα in early cardiac development. Even though other genetic defects exist in this naturally occurring mutant, most defects associated with cardiac development are believed to be a result of the absence of this receptor. Gross morphological defects including improper septation of the outflow tract, dysmorphic shape of the heart, and lack of trabecular development are similar to those that have been previously described. Many of these defects have been attributed to the failure of a subset of non-neuronal neural crest cells to properly migrate into the region of the developing outflow tract. In these studies, we have also used confocal scanning laser and transmission electron microscopy to describe and compare the organization and differentiation of the cytoskeletal proteins actin and myosin in littermate control and Patch mouse hearts. Cytoskeletal organization of the cardiac myocytes in Patch mouse hearts has not previously been described. In most cardiac myocytes of Patch mice, actin was found only on the periphery of the cells, and the organization of actin, myosin, and precursor Z-band material into distinct myofibrils was greatly reduced.

Fast-myosin in frozen histological sections of eight, 10, 11 and nine muscles of the upper forelimb, lower forelimb, upper hindlimb and lower hindlimb, respectively, of goats was quantified by an immunohistochemical micromethod based on the enzyme-linked immunosorbent assay. The structure of the muscles is well preserved during the immunohistochemical measurement. High fast-myosin levels (more than 201 mg/g total protein) were observed in the triceps brachii (lateral head), rectus femoris, vastus lateralis, semitendinosus, semimembranosus, gastrocnemius (lateral head) and long digital extensor muscles. In contrast, low fast-myosin levels (less than 50 mg/g) were found in the triceps brachii (medial head), superficial digital flexor, vastus intermedialis, and soleus muscles. Fast-myosin-positive fibres (type II or fast-twitch type) were distributed more in the superficial regions than in the deeper regions in the triceps brachii (lateral and long heads), biceps brachii, brachialis, biceps femoris, vastus lateralis, vastus medialis, semimembranosus and gastrocnemius (lateral and medial heads) muscles. In contrast, type IIfibres were distributed more in the deeper regions than in the superficial regions in the extensor carpi radialis, deep digital flexor, cranial tibial, deep digital flexor and superficial digital flexor muscles. When the results obtained by the immunohistochemical micromethod were compared with those obtained by biochemical techniques and by histomorphometrical analyses, high correlations were noted. This technique could be used in research projects to study the muscle characteristics that determine meat quality.

A study of the determinants of inter- and intra-litter variation in muscle fibre number was carried out on five litters of Large White piglets. Fresh frozen, whole mid-belly sections of m. semitendinosus were stained to demonstrate acid-stable myosin adenosine triphosphatase activity. From these sections it was possible to identify which fibres had developed as primary and which as secondary fibres. Estimations of total muscle fibre number, total primary fibre number and ratio of secondary fibres to primary fibres were made for each animal. Results demonstrated that primary fibre number varied between litters (P<0·01) and was responsible for the variation in total muscle fibre number (P < 005) between litters since there was no significant variation in secondary: primary ratio. Within-litter differences in total fibre number could be attributed to both the secondary: primary ratio and primary fibre number, in almost equal contributions. However, when only the largest and smallest extremes of the litters were compared, variation in fibre number was due to the significant difference in the secondary: primary ratio (P<0·01). Taken as a whole, the results appear to show that primary fibre number is responsible for all the variation in muscle fibre number between litters, and also makes a significant contribution, with secondary: primary fibre ratio, to the variation present within a litter. The factors responsible for variations in primary and secondary fibre numbers are discussed.