Evaluation of collagen fiber maturation and ordering in regenerating tendons employing H-1 double quantum filtered NMR spectroscopy

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Abstract
It is difficult to monitor the chronic stage of the healing process of ruptured tendons employing the present diagnostic modes. However, the results of this study have shown that 1H double quantum filtered (DQF) NMR spectroscopy is sensitive to the later stages of the healing process. Regenerated tendons of rabbits were dissected and measured at the end of the acute phase (three weeks), the subacute phase (nine weeks), and the chronic phase (13 and 18 weeks after tenotomy). Four parameters were determined by 1H DQF NMR spectroscopy: (a) the maximum signal intensity (h(max)) relative to the single quantum spectrum, (b) the creation time of the maximum signal intensity (tau(max)), (c) the decay time from the maximum signal intensity to a value half of that intensity (tau(1/2)) and (d) the residual dipolar splitting of water (delta), representing the order of the collagen fibers. The values of h(max), tau(max), tau(1/2), and delta of the intact Achilles tendons were 11.3+/-1.0%, 0.48+/-0.03 ms, 0.67+/-0.04 ms and 732+/-62 Hz (mean +/- SEM, n=6), respectively. In the regenerating tendon, h(max) increased from 0.41+/-0.12% at three weeks to 7.07+/-0.77% at 18 weeks, tau(max) decreased from 1.88+/-0.31 ms at three weeks to 0.72+/-0.04 ms at 18 weeks, tau(1/2) decreased from 11.6+/-1.8 ms at 3 weeks to 1.48+/-0.16 ms at 18 weeks, and delta increased from 129+/-8 Hz at three weeks to 414+/-29 Hz at 18 weeks. We have concluded that reordering of collagen fibers proceeds continuously even in the chronic stage of healing. Thus, the 1H DQF NMR spectroscopy is a useful non-invasive technique to evaluate the reconstruction and the order of collagen fibers in regenerating tendon. It is also suggested that tau(1/2) and h(max) are most useful for in vivo DQF NMR spectroscopy and imaging, respectively, in combination with tau(max).