Elsevier

Neuropharmacology

Volume 79, April 2014, Pages 275-281
Neuropharmacology

All-trans retinoic acid prevents epidural fibrosis through NF-κB signaling pathway in post-laminectomy rats

https://doi.org/10.1016/j.neuropharm.2013.11.010Get rights and content

Highlights

  • ATRA shows anti-fibrotic effects in vitro and vivo.

  • ATRA is able to prevent EF in laminectomized rats in a dose-dependent manner.

  • ATRA is able to suppress the expression of TGF-β1 and IL-6.

  • ATRA works through the regulation of NF-κB signaling pathway.

Abstract

Laminectomy is a widely accepted treatment for lumbar disorders, and epidural fibrosis (EF) is a common complication. EF is thought to cause post-operative pain recurrence after laminectomy or discectomy. All-trans retinoic acid (ATRA) has shown anti-fibrotic, anti-inflammatory, and anti-proliferative functions. The object of this study was to investigate the effects of ATRA on the prevention of EF in post-laminectomy rats. In vitro, the anti-fibrotic effect of ATRA was demonstrated with cultured fibroblasts count, which comprised of those that were cultured with/without ATRA. In vivo, rats underwent laminectomy at the L1-L2 levels. We first demonstrated the beneficial effects using 0.05% ATRA compared to vehicle (control group). We found that a higher concentration of ATRA (0.1%) achieved dose-dependent results. Hydroxyproline content, Rydell score, vimentin-positive cell density, fibroblast density, inflammatory cell density and inflammatory factor expression levels all suggested better outcomes in the 0.1% ATRA rats compared to the other three groups. Presumably, these effects involved ATRA's ability to suppress transforming growth factor (TGF-β1) and interleukin (IL)-6 which was confirmed with reverse-transcriptase polymerase chain reaction (RT-PCR). Finally we demonstrated that ATRA down-regulated nuclear factor (NF)-κB by immunohistochemistry and western blotting for p65 and inhibition of κB (IκBα), respectively. Our findings indicate that topical application of ATRA can inhibit fibroblast proliferation, decrease TGF-β1 and IL-6 expression level, and prevent epidural scar adhesion in rats. The highest concentration employed in this study (0.1%) was the most effective. ATRA suppressed EF through down-regulating NF-κB signaling, whose specific mechanism is suppression of IκB phosphorylation and proteolytic degradation.

Introduction

Lumbar laminectomy is widely performed to treat lumbar disc herniation and other lumbar disorders, and epidural fibrosis (EF) is a common complication. The literature reported that “failed back surgery syndrome” (FBSS) was common in patients suffering from continued pain in the lower posterior trunk and/or lower extremities following lumbar laminectomy (Guyer et al., 2006), and extensive EF may be a contributing factor for FBSS (Ross et al., 1996).

First described in 1948, EF frequently deposits in the epidural space following posterior spinal surgery (Guyer et al., 2006, Cauchoix et al., 1978, Key and Ford, 1948). No effective treatments exist for patients with established EF (Ceviz et al., 1997), and most reoperations for FBSS are unsuccessful (Cruccu et al., 2007). Thus, prevention of EF formation is believed to be the best approach (Henderson et al., 1993). A variety of agents or mechanical barriers have been studied to prevent EF both in animal models and humans, including autologous fat grafts, Adcon-L, polytetrafluoroethylene membrane, and fibrinolytic agents. (Gill et al., 1985, Sandoval and Hernandez-Vaquero, 2008, Xu et al., 2012, Barberá et al., 1978). However, none of these approaches have been completely successful.

All-trans retinoic acid (ATRA) is a physiological metabolite of vitamin A that has a wide range of biological activities; it affects cell differentiation, proliferation, embryogenesis, inflammation and apoptosis (Dong et al., 2012, Datta and Lianos, 1999). It has been reported that ATRA might ease bleomycin-induced pulmonary fibrosis via inhibition expressions of interleukin (IL)-6 and transforming growth factor (TGF)-β (Dong et al., 2012). Moreover, amelioration of EF was observed after inhibiting IL family and TGF family member expression (Yan et al., 2013, Zhang et al., 2013).

ATRA is able to attenuate the increased activation of nuclear factor (NF)-κB (Datta and Lianos, 1999). NF-κB was first identified in activated B cells. This transcription factor has been found to induce several biologically essential processes including innate and adaptive immunity, cell proliferation, differentiation, and inflammation (Ghosh and Karin, 2002). A previous study reported that ATRA was able to protect cardiomyocytes by inhibiting NF-κB signaling (Nizamutdinova et al., 2013).

In the present study, fibroblasts isolated from the tail skin of the rats and laminectomized rats were employed to explore the anti-fibrotic, anti-inflammatory, and anti-proliferative effects of ATRA. We also investigated ATRA's effects on EF at different concentrations, as well as the mechanism of action. Our results are helpful for planning future human trials and clinical applications for treating EF.

Section snippets

Animals

Eighty five healthy male Wistar rats (mean weight = 250 g) were purchased from the Radiation Study Institute-Animal Center, Tianjin, China. Experiments were carried out in compliance with the principles of International Laboratory Animal Care and with the European Communities Council Directive (86/809 /EEC) and were approved by the local ethical committee. Five rats formed the tail skin sample collection group. The remaining eighty rats were randomly divided into four groups (20 rats per

ATRA reduced EF formation

Initially, ATRA (0.05%) was employed and it showed signs of suppressing EF.

In order to identify the potential ability of ATRA to suppress fibroblast proliferation, we cultured and counted fibroblasts and found that proliferation was suppressed by ATRA (Fig. 1). We then used ATRA in the rat model setting. Pre- and post-operation comparisons of posture, weight support, and coordination according to the BBB locomotion test did not show significant changes (BBB score = 21). Pre and post-operative

Discussion

In approximately 24% of FBSS cases, epidural scar formation causes extensive adhesions around the dural mater and nerve roots (Burton et al., 1981). EF is characterized by extracellular matrix protein deposition, fibroblast accumulation, and distortion of normal tissue architecture due to inflammation (Ward and Hunninghake, 1998). In order to effectively prevent EF, purposeful efforts should be guided toward these effects.

In this study, ATRA was shown to effectively ameliorate the

Conclusions

ATRA was able to prevent EF in laminectomized rats in a dose-dependent manner. The highest concentration used in this study (0.1%) proved to be the most effective. ATRA suppressed EF by regulating the NF-κB signaling pathway; specifically, it suppressed the phosphorylation and proteolytic degradation of IκB.

Acknowledgments

This study was sponsored by the National Natural Science Foundation of China (81330042), (81070982), (81371957), (81171714), (81201400) and (81301544), and the Swedish Medical Research Council Grant Nr 2710, Stockholm, Sweden, and the Air Force Material Command, USAF, under grant number FA8655-05-1-3065.

References (37)

  • C.V. Burton et al.

    Causes of failure ofsurgery on the lumbar spine

    Clin. Orthop. Relat. Res.

    (1981)
  • J.M. Cabezudo et al.

    Symptomatic rootcompression by a free fat transplant after hemilaminectomy: casereport

    J. Neurosurg.

    (1985)
  • J. Cauchoix et al.

    Repeat surgery after disc excision

    J. Spine

    (1978)
  • G. Cruccu et al.

    EFNS guidelines on neurostimulation therapy for neuropathic pain

    Eur. J. Neurol.

    (2007)
  • Z. Dong et al.

    The role of all-trans retinoic acid in bleomycin-inducedpulmonary fibrosis in mice

    Exp. Lung Res.

    (2012)
  • N. Fukui et al.

    Adhesion formation can be reduced by the suppression of transforming growth factor-beta1activity

    J. Orthop. Res.

    (2000)
  • S. Ghosh et al.

    Missing pieces in the NF-κB puzzle

    Cell.

    (2002)
  • G.G. Gill et al.

    Pedicle fat grafts for the prevention of scar in low back surgery. A preliminary report on the first 92 cases

    Spine(Phila Pa 1986)

    (1985)
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