Progesterone attenuates spinal glial activation and the activity of pro-inflammatory enzymes and prevents neurophatic pain alter spinal cord injury

GONZALEZ S; CORONEL F; LABOMBARDA, F; DE NICOLA A; VILLAR M

Milan

Congreso; 14th. World Congress on Pain; 2012

International Association for the Study of Pain

Aim of Investigation: Spinal cord injury (SCI) results in the development of chronic pain that severely compromises the quality of life in nearly 70% of patients. Glial cell activation and the consequent production of inflammatory cytokines contribute to the pathology of central neuropathic syndromes and represent a therapeutic target. We have previously reported that progesterone (PG), a neuroprotective steroid, may offer a promising perspective in pain modulation. In a model of central pain, PG prevents the early injury-induced up-regulation of proinflammatory cytokines and the enzymes cyclooxigenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), all key players in nociceptive processing at the spinal level. This inflammatory response involves the activation of different signaling pathways, including the transcription factor nuclear factor-κB (NF-κB).To further explore the effects of PG after central injury and neuropathic pain, in the present study we evaluate the impact of the steroid administration on the profile of glial cell activation, enzyme activities, pain behaviors and the expression of the IκB-α protein, an index of NF-κB transactivation.   Methods: Sprague-Dawley male rats subjected to spinal hemisection at T13 level received daily subcutaneous injections of PG (Hx+PG; 16 mg/kg) or vehicle (Hx). Uninjured rats were used as control animals (CTL). Mechanical and thermal allodynia of the hindpaws were assessed with the von Frey and Choi tests, respectively. Immunohistochemical analysis was employed to assess the number of cells exhibiting the astrocytic marker GFAP or the microglial marker OX-42 in the dorsal horn. NOS activity was analyzed by measuring nitric oxide stable end products nitrite and nitrate (NOx) with the Griess reagent. COX-2 activity was determined by means of a commercial assay kit. Real-time PCR was used to determine the relative mRNA levels of IκB-α and the housekeeping gene cyclophilin.   Results: One day after SCI, a significant increase in NOS activity (μmol/ mg protein) and COX-2 activity (nmol/min/mg protein) was observed in the dorsal spinal cord as compared to CTL (NOx: CTL: 0.108±0.025, Hx: 1.422±0.116, p<0.001; COX-2: CTL: 2.274±0.142; Hx: 9.727±0.794, p<0.001). At this time point, and coincident with enzyme activity increase, a significant up-regulation in the mRNAs levels for IκB-α was observed (results are expressed as fold-increase relative to CTL levels; Hx: 2.586±0.423, p<0.05 vs. CTL). Interestingly, Hx+PG animals showed reduced enzymes activities as compared to Hx, which resulted similar to those observed in CTL rats (NOx: 0.336±0.044; COX-2: 2.102±0.236; p>0.05 vs CTL, p<0.001 vs Hx for both enzymes). PG administration also attenuated the injury-induced increase in the number of GFAP- and OX-42 positive cells (p<0.01 vs Hx in all cases).  In addition, at this time point, Hx+PG animals presented lower levels of mRNAs for IκB-α as compared to injured animals (1.506±0.278, p<0.05 vs Hx), suggesting a decreased NF-κB transactivation during steroid administration. In line with our previous results, Hx+PG animals did not develop mechanical allodynia and showed reduced sensitivity to cold stimulation.   Conclusions: Our data indicate that PG, by modulating the inflammatory events triggered after SCI, may provide an interesting therapeutic strategy to prevent the development of central chronic pain.