Abstract: Tendon injuries are an important problem in athletic horses and are probably caused by excessive loading of the tendons during demanding activities. As a first step towards understanding these injuries, the tendon loading was quantified during jump landings. Kinematics and ground reaction forces were collected from the leading and trailing forelimbs of 6 experienced jumping horses. Joint moments were calculated using inverse dynamic analysis. It was found that the variation of movement and loading patterns was small, both within and between horses. The peak flexor joint moments in the coffin and fetlock joints were larger in the trailing limb (-0.62 and -2.44 Nm/kg bwt, respectively) than in the leading limb (-0.44 and -1.93 Nm/kg bwt, respectively) and exceeded literature values for trot by 82 and 45%. Additionally, there was an extensor coffin joint moment in the first half of the stance phase of the leading limb (peak value 0.26+/-0.18 Nm/kg bwt). From these results, it was concluded that the loading of the flexor tendons during landing was higher in the trailing than in the leading limb and that there was an unexpected loading of the extensor tendon in the leading limb.

Abstract: An acid-destabilized form of apomyoglobin, the so-called E state, consists of a set of heterogeneous structures that are all characterized by a stable hydrophobic core composed of 30-40 residues at the intersection of the A, G, and H helices of the protein, with little other secondary structure and no other tertiary structure. Relaxation kinetics studies were carried out to characterize the dynamics of core melting and formation in this protein. The unfolding and/or refolding response is induced by a laser-induced temperature jump between the folded and unfolded forms of E, and structural changes are monitored using the infrared amide I' absorbance at 1648-1651 cm(-1) that reports on the formation of solvent-protected, native-like helix in the core and by fluorescence emission changes from apomyoglobin's Trp14, a measure of burial of the indole group of this residue. The fluorescence kinetics data are monoexponential with a relaxation time of 14 micros. However, infrared kinetics data are best fit to a biexponential function with relaxation times of 14 and 59 micros. These relaxation times are very fast, close to the limits placed on folding reactions by diffusion. The 14 micros relaxation time is weakly temperature dependent and thus represents a pathway that is energetically downhill. The appearance of this relaxation time in both the fluorescence and infrared measurements indicates that this folding event proceeds by a concomitant formation of compact secondary and tertiary structures. The 59 micros relaxation time is much more strongly temperature dependent and has no fluorescence counterpart, indicating an activated process with a large energy barrier wherein nonspecific hydrophobic interactions between helix A and the G and H helices cause some helix burial but Trp14 remains solvent exposed. These results are best fit by a multiple-pathway kinetic model when U collapses to form the various folded core structures of E. Thus, the results suggest very robust dynamics for core formation involving multiple folding pathways and provide significant insight into the primary processes of protein folding.

Abstract: Ultrafast, laser-induced pH jump with time-resolved photoacoustic detection has been used to investigate the early protonation steps leading to the formation of the compact acid intermediate (I) of apomyoglobin (ApoMb). When ApoMb is in its native state (N) at pH 7.0, rapid acidification induced by a laser pulse leads to two parallel protonation processes. One reaction can be attributed to the binding of protons to the imidazole rings of His24 and His119. Reaction with imidazole leads to an unusually large contraction of -82 +/- 3 ml/mol, an enthalpy change of 8 +/- 1 kcal/mol, and an apparent bimolecular rate constant of (0.77 +/- 0.03) x 10(10) M(-1) s(-1). Our experiments evidence a rate-limiting step for this process at high ApoMb concentrations, characterized by a value of (0. 60 +/- 0.07) x 10(6) s(-1). The second protonation reaction at pH 7. 0 can be attributed to neutralization of carboxylate groups and is accompanied by an apparent expansion of 3.4 +/- 0.2 ml/mol, occurring with an apparent bimolecular rate constant of (1.25 +/- 0.02) x 10(11) M(-1) s(-1), and a reaction enthalpy of about 2 kcal/mol. The activation energy for the processes associated with the protonation of His24 and His119 is 16.2 +/- 0.9 kcal/mol, whereas that for the neutralization of carboxylates is 9.2 +/- 0.9 kcal/mol. At pH 4.5 ApoMb is in a partially unfolded state (I) and rapid acidification experiments evidence only the process assigned to carboxylate protonation. The unusually large contraction and the high energetic barrier observed at pH 7.0 for the protonation of the His residues suggests that the formation of the compact acid intermediate involves a rate-limiting step after protonation.

Abstract: Injuries to the superficial digital flexor tendon are common in horses required to gallop and jump at speed. Partial rupture of this tendon usually occurs in the central core of the midmetacarpal region and may be preceded by localised degenerative changes. Post mortem examination of apparently normal equine flexor tendons has revealed an abnormal macroscopic appearance in the central core, characterised by a reddish discolouration. We have previously shown that there is also physical damage to the collagen fibres. In the present study we tested the hypothesis that the abnormal appearance is accompanied by changes in the composition of the extracellular matrix of the tendon. Biochemical analysis of the extracellular matrix demonstrated an increase in total sulphated glycosaminoglycan content, increase in the proportion of type III collagen and decrease in collagen linked fluorescence in the central core of 'degenerated' tendons relative to tissue from the peripheral region of the same tendon. Dry matter content and total collagen content were not significantly different between tendon zones or normal and 'degenerated' tendons. These changes suggest a change in cell metabolism and matrix turnover in the central core of the tendon and are likely to contribute to a decrease in mechanical properties in this part of the tendon, predisposing to the characteristic partial rupture of the tendon.

Abstract: The purpose of this study was to demonstrate the relationships between jumping technique and dorsoventral acceleration measured at the sternum. Eight saddle horses of various jumping abilities competed on a selective experimental show jumping course including 14 obstacles. An accelerometric belt fastened onto the thorax continuously measured the dorsoventral acceleration during the course. At each jump, 11 locomotor parameters (acceleration peaks, durations and stride frequency) were obtained from the dorsoventral acceleration-time curves. The type of obstacle significantly influenced the hindlimb acceleration peak at take-off and the landing acceleration peak (P<0.01). The poor jumpers exhibited a higher mean forelimb acceleration peak at take-off, a higher forelimb/hindlimb ratio between peaks of acceleration (F/H), and a lower approach stride frequency than good jumpers. Knocking over an obstacle was significantly associated with a low hindlimb acceleration peak at take-off and a high F/H ratio (P<0.01). In order to observe the continuous changes in the frequency domain of the dorsoventral acceleration during the approach and take-off phase, a Morlet's wavelet analysis was computed for each horse jumping over a series of 3 vertical obstacles. Different patterns of time-frequency images obtained by wavelet analysis were found when the horse either knocked over a vertical obstacle or cleared it. In the latter case, the image pattern showed an instantaneous increase in stride frequency at the end of the approach phase, and a marked energy content in the middle frequency range at take-off.