RAR/RXR and PPAR/RXR Signaling in Spinal Cord Injury

Abstract

The retinoid acid receptors (RAR) and peroxisome proliferator-activated receptors (PPAR) have been implicated in the regulation of inflammatory reactions. Both receptor families contain ligand-activated transcription factors which form heterodimers with retinoid X receptors (RXR). We review data that imply RAR/RXR and PPAR/RXR pathways in physiological reactions after spinal cord injury. Experiments show how RAR signaling may improve axonal regeneration and modulate reactions of glia cells. While anti-inflammatory properties of PPAR are well documented in the periphery, their possible roles in the central nervous system have only recently become evident. Due to its anti-inflammatory function this transcription factor family promises to be a useful target after spinal cord or brain lesions.

Figure 1

Retinoic acid synthesis and nuclear translocation of RAR/RXR after spinal cord injury. (a) Double immunostaining of rat spinal cord sections shows RALDH-2 immunoreactivity (red) in a subpopulation of NG2-positive glia (green) 7 days after contusion injury. RALDH2/NG2 expressing cells were only detected in the vicinity of the lesion site, many NG2 cells do not contain the RA-synthesizing enzyme (yellow arrow heads). White arrows point to DAPI-stained cell nuclei of RALDH2/NG2 cells in superimposed photographs. (b) At 7 dpo, ED1-positive macrophages at the lesion site express RXRβ in their cell nuclei. (c) SCI-induced transient translocation of RARβ and RXRα from the cytosol into the nuclei of activated macrophages/microglia. All scale bars are 20μ m (sources: [12, 14]).

Figure 1

Retinoic acid synthesis and nuclear translocation of RAR/RXR after spinal cord injury. (a) Double immunostaining of rat spinal cord sections shows RALDH-2 immunoreactivity (red) in a subpopulation of NG2-positive glia (green) 7 days after contusion injury. RALDH2/NG2 expressing cells were only detected in the vicinity of the lesion site, many NG2 cells do not contain the RA-synthesizing enzyme (yellow arrow heads). White arrows point to DAPI-stained cell nuclei of RALDH2/NG2 cells in superimposed photographs. (b) At 7 dpo, ED1-positive macrophages at the lesion site express RXRβ in their cell nuclei. (c) SCI-induced transient translocation of RARβ and RXRα from the cytosol into the nuclei of activated macrophages/microglia. All scale bars are 20μ m (sources: [12, 14]).

Figure 1

Retinoic acid synthesis and nuclear translocation of RAR/RXR after spinal cord injury. (a) Double immunostaining of rat spinal cord sections shows RALDH-2 immunoreactivity (red) in a subpopulation of NG2-positive glia (green) 7 days after contusion injury. RALDH2/NG2 expressing cells were only detected in the vicinity of the lesion site, many NG2 cells do not contain the RA-synthesizing enzyme (yellow arrow heads). White arrows point to DAPI-stained cell nuclei of RALDH2/NG2 cells in superimposed photographs. (b) At 7 dpo, ED1-positive macrophages at the lesion site express RXRβ in their cell nuclei. (c) SCI-induced transient translocation of RARβ and RXRα from the cytosol into the nuclei of activated macrophages/microglia. All scale bars are 20μ m (sources: [12, 14]).

Figure 2

Localization of PPARα and PPARβ/δ in astrocytes and oligodendrocytes in the spinal cord. (a) Western blotting shows PPARα and PPARβ but not PPARγ in spinal cord, telencephalon, and diencephalon. Identical expression patterns were detected with RT-PCR. (b) Detection of PPAR immunoreactive cells in the white matter of rat spinal cord. GFAP-positive/CNPase-negative astrocytes are immunoreactive for PPARα (marked a, upper four panels) while GFAP-negative/CNPase-positive oligodendrocytes express PPARβ (marked ol, lower panels) (scale bars: 25μ m, source: [38]). (c) Distribution of PPARβ/δ and PPARγ immunoreactive cells in the spinal cord of the adult rat (cervical level, coronal sections, scale bar: 100μ m, source: [41]).

Figure 3

Anti-inflammatory effects of PPAR and RAR agonists in astrocytes. Astrocyte primary cultures from cortices of newborn mice were treated with 9-cis RA (RA) and/or the PPARα agonist fenofibrate (F) for 1hour and then stimulated with 2μg/mL LPS for 24 hours. (a) Nitrite production as an indicator of NOS activity was reduced by 1μ M RA or 100μ M F, and completely suppressed with RA plus F. Pretreatment of RA and/or F also decreased the release of (b) the chemokine MCP-1 (RA: 2μ M, F: 200μ M), and cytokines (c) IL-1β (RA: 1μ M, F: 100μ M), (d) IL-6 (RA: 2μ M, F: 50μ M), and (e) TNFα (RA: 1μ M, F: 100μ M) in response to LPS. Cytokine production was measured with ELISA. Experiments with microglia cultures revealed similar effects, except for MCP-1, whose production was stimulated by 9-cis RA in microglia. Error bars indicate SEM, asterisks indicate significant differences compared to the LPS condition (sources: [54, 78]).

Figure 4

Transfection of RARβ induces regeneration of corticospinal axons in vivo. (a) Longitudinal sections of the adult rat spinal cord were traced with BDA to determine fibre regeneration in the corticospinal tract; overview of the lesion site in an animal injected with the RARβ2 expressing construct (EIAV-RARβ2, Rostral: left). (b) Animals with RARβ2-transfected cells displayed less fiber degeneration than control rats. (c) Labeled axons were detected up to the edge of the lesion, (d) at the distal edge of the lesion, and (e–g) at various distances caudal to the lesion. Growth cone-like endings were detected at the tips of some axons (white arrowheads in d). Axons appear to send collaterals (white arrowhead in g) from white matter to grey matter. (Scale bars in (a): 1mm, in (b–g): 100μ m.) (h–i) Quantification of BDA-labeled fibres in the corticospinal tract after spinal cord lesion. EIAV-RARβ2-treated animals displayed increased fiber numbers rostral and caudal to the lesion compared to control animals (P < .05, two-way ANOVA) (source: [100]).