Regular articleFK506 enhances reinnervation by regeneration and by collateral sprouting of peripheral nerve fibers
Introduction
FK506 is an immunosuppressant drug widely used to prevent rejection after solid organ transplantation. FK506 and its nonimmunosuppressant derivatives also exert neuroprotective and neuroregenerative activities (for reviews, see Gold 1999, Gold 2000a, Gold 2000b, Snyder et al 1998. Several experimental studies have shown that administration of FK506 increases the rate of axonal regeneration after axotomy induced by nerve crush Gold et al 1995, Gold et al 1994, Lee et al 2000, Wang et al 1997 or after nerve transection and suture or graft repair Büttemeyer et al 1995, Doolabh and Mackinnon 1999, Fansa et al 1999, Navarro et al 2001. Functional recovery of distal targets, measured by the walking track or by electrophysiological methods, starts earlier in rats and mice treated with FK506, although ultimate levels of recovery are close to those of untreated animals Doolabh and Mackinnon 1999, Fansa et al 1999, Navarro et al 2001. In patients with a hand allograft where FK506 was used to prevent rejection, recovery of sensorimotor functions was faster than expected, this being attributed to an enhancement of axonal regeneration by FK506 Dubernard et al 1999, Jones et al 2000. However, after severe nerve injuries in larger species, including humans, accelerating target reinnervation may be more important for reducing the consequences of denervation (muscle atrophy, loss of sensory receptors, denervation hypersensitivity) (Fu and Gordon, 1995). In this context, the ability of FK506 to increase nerve regeneration following a chronic axotomy (Sulaiman et al., 2002) may be of importance for improving the chances for functional recovery. However, studies on the effects of FK506 on target reinnervation are lacking using any of these models. Furthermore, while FK506 is presently being used in human hand transplantations Dubernard et al 1999, Jones et al 2000 and allograft nerve repairs (Mackinnon et al., 2001), the dose–response needs to be characterized in multiple models to determine its optimal dose for use in clinical practice.
We recently examined the dose dependency for FK506 on the rate of nerve regeneration in the mouse and found a novel bimodal response (Udina et al., 2002). The largest increase in regeneration rate was achieved at a dose of 5 mg/kg, but doses of 0.2 and 2 mg/kg yielded a similar regeneration course through the first week following sciatic nerve crush. In contrast, results with intermediate doses of 0.5 and 1 mg/kg were not different from those of controls. In the rat, the 5 mg/kg dose also maximally accelerates axonal regeneration (Wang et al., 1997), while the 1 mg/kg was less effective; lower dosages were not examined. The efficacy of lower dosages has obvious implications for clinical use for nerve regeneration since low doses (e.g., 0.2 mg/kg) would have less risk of toxicity and would presumably not exhibit immunosuppressant activity Spencer et al 1997, Undre et al 1999.
In the present study, we examined the effects of FK506 on both the speed of axonal elongation and the degree of target reinnervation. Reinnervation of target tissue is achieved by either axonal regeneration from injured nerves or collateral sprouting from intact neurons. In regard to collateral sprouting, one immunophilin ligand (V-10,367) has been shown to increase neurite branching in vitro (Constantini and Isacson, 2000). Therefore, we also investigated the capabilities of FK506 to promote collateral sprouting in vivo. Thus, two models were used to study the effect of FK506 on target reinnervation: nerve regeneration following sciatic nerve crush and collateral sprouting following partial denervation of the mouse hind paw. In both models, we assessed recovery of different targets over a 1-month period in mice receiving FK506 at either 0.2 or 5 mg/kg/day, the two doses previously found as most effective (Udina et al., 2002).
Section snippets
Surgical procedures
Operations were performed under pentobarbital anesthesia (60 mg/kg ip) in 3-month-old female OF1 mice. For nerve regeneration assessment, the sciatic nerve was exposed at the mid thigh and crushed three times in succession with a Dumont No. 5 forceps at a constant point, 45 mm from the tip of the third digit. The saphenous nerve was also cut in the femoral space and a long segment of the distal stump removed to prevent regeneration. The wound was then sutured by layers and disinfected with
Effects of FK506 on nerve regeneration after sciatic nerve crush
Functional tests performed at 7 dpo demonstrated that all nerve-mediated functions were abolished in the denervated hind paw of all animals. The first CMAPs evoked by sciatic nerve stimulation and recorded from plantar muscles reappeared at 15 dpo in most mice and at 18 dpo in two mice in group C. During the following weeks, the latency of the CMAP shortened to close to normal (data not shown) and the amplitude progressively increased to reach mean final values (relative to preoperative values)
Discussion
The present study reveals that FK506 administration enhances the reinnervation of target organs both by regenerating axons following a nerve crush and by collateral sprouts of an intact nerve induced to expand following denervation of its nearby territory. The recovery achieved by regeneration and by collateral sprouting was greater overall with the 5 mg/kg dose than with the dose of 0.2 mg/kg of FK506. Nevertheless, the 0.2 mg/kg dose still produced better results than those found in
Acknowledgements
This work was supported by grants from the Fondo de Investigación Sanitaria (FIS00-0031-02), the Ministerio de Ciencia y Tecnología (SAF2002-04016), and the Acadèmia de Ciències Mèdiques de Catalunya i Balears, Spain. We thank Fujisawa Pharmaceuticals Inc. (Osaka, Japan) for its generous gift of FK506 and the Kerr Company (Romulus, MI) for providing Elasticon. The technical help of Jessica Jaramillo is greatly acknowledged.
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2014, Progress in NeurobiologyCitation Excerpt :Although the administration of FK506 increases the extent of neurological recovery (Chunasuwankul et al., 2002; Farrag et al., 2007; Udina et al., 2004), increases the onset time to initiation of axon regeneration (Navarro et al., 2001), and increases the half time to achieve neurological recovery, it does not induce a significant increase in the extent of long-term neurological recovery (Lanzetta et al., 2003; Terzis and Konofaos, 2010). However, FK506 exerts an additional benefit for restoring neurological function, which is to increase the specificity of sensory and motor target reinnervation (Udina et al., 2003, 2004). Clinically FK506 administered at 5 and 8 ng/ml//kg/day for a total of 60 days induces more rapid axon regeneration than is seen in control patients (Phan and Schuind, 2012).