The immunomodulator glatiramer acetate influences spinal motoneuron plasticity during the course of multiple sclerosis in an animal model

The immunomodulador glatiramer acetate (GA) has been shown to significantly reduce the severity of symptoms during the course of multiple sclerosis and in its animal model - experimental autoimmune encephalomyelitis (EAE). Since GA may influence the response of non-neuronal cells in the spinal cord, it is possible that, to some extent, this drug affects the synaptic changes induced during the exacerbation of EAE. In the present study, we investigated whether GA has a positive influence on the loss of inputs to the motoneurons during the course of EAE in rats. Lewis rats were subjected to EAE associated with GA or placebo treatment. The animals were sacrificed after 15 days of treatment and the spinal cords processed for immunohistochemical analysis and transmission electron microscopy. A correlation between the synaptic changes and glial activation was obtained by performing labeling of synaptophysin and glial fibrillary acidic protein using immunohistochemical analysis. Ultrastructural analysis of the terminals apposed to alpha motoneurons was also performed by electron transmission microscopy. Interestingly, although the GA treatment preserved synaptophysin labeling, it did not significantly reduce the glial reaction, indicating that inflammatory activity was still present. Also, ultrastructural analysis showed that GA treatment significantly prevented retraction of both F and S type terminals compared to placebo. The present results indicate that the immunomodulator GA has an influence on the stability of nerve terminals in the spinal cord, which in turn may contribute to its neuroprotective effects during the course of multiple sclerosis.

Protective mechanism of agmatine pretreatment on RGC-5 cells injured by oxidative stress

Agmatine has neuroprotective effects on retinal ganglion cells (RGCs) as well as cortical and spinal neurons. It protects RGCs from oxidative stress even when it is not present at the time of injury. As agmatine has high affinity for various cellular receptors, we assessed protective mechanisms of agmatine using transformed RGCs (RGC-5 cell line). Differentiated RGC-5 cells were pretreated with 100 μM agmatine and consecutively exposed to 1.0 mM hydrogen peroxide (H2O2). Cell viability was determined by measuring lactate dehydrogenase (LDH), and the effects of selective alpha 2-adrenergic receptor antagonist yohimbine (0-500 nM) and N-methyl-D-aspartic acid (NMDA) receptor agonist NMDA (0-100 µM) were evaluated. Agmatine’s protective effect was compared to a selective NMDA receptor antagonist MK-801. After a 16-h exposure to H2O2, the LDH assay showed cell loss greater than 50%, which was reduced to about 30% when agmatine was pretreated before injury. Yohimbine almost completely inhibited agmatine’s protective effect, but NMDA did not. In addition, MK-801 (0-100 µM) did not significantly attenuate the H2O2-induced cytotoxicity. Our results suggest that neuroprotective effects of agmatine on RGCs under oxidative stress may be mainly attributed to the alpha 2-adrenergic receptor signaling pathway.

Caspase dependence of the death of neonatal retinal ganglion cells induced by axon damage and induction of autophagy as a survival mechanism

We examined the degeneration of post-mitotic ganglion cells in ex-vivo neonatal retinal explants following axon damage. Ultrastructural features of both apoptosis and autophagy were detected. Degenerating cells reacted with antibodies specific for activated caspase-3 or -9, consistent with the presence of caspase activity. Furthermore, peptidic inhibitors of caspase-9, -6 or -3 prevented cell death (100 µM Ac-LEDH-CHO, 50 µM Ac-VEID-CHO and 10 µM Z-DEVD-fmk, respectively). Interestingly, inhibition of autophagy by 7-10 mM 3-methyl-adenine increased the rate of cell death. Immunohistochemistry data, caspase activation and caspase inhibition data suggest that axotomy of neonatal retinal ganglion cells triggers the intrinsic apoptotic pathway, which, in turn, is counteracted by a pro-survival autophagic response, demonstrated by electron microscopy profiles and pharmacological autophagy inhibitor.

Intravenous regional block is similar to sympathetic ganglion block for pain management in patients with complex regional pain syndrome type I

Sympathetic ganglion block (SGB) or intravenous regional block (IVRB) has been recommended for pain management in patients with complex regional pain syndrome type I (CRPS-I). Forty-five patients were initially selected but only 43 were accepted for the study. The present study evaluated the efficacy of IVRB produced by combining 70 mg lidocaine with 30 µg clonidine (14 patients, 1 male/13 females, age range: 27-50 years) versus SGB produced by the injection of 70 mg lidocaine alone (14 patients, 1 male/13 females, age range: 27-54 years) or combined with 30 µg clonidine (15 patients, 1 male/14 females, age range: 25-50 years) into the stellate ganglion for pain management in patients with upper extremity CRPS-I. Each procedure was repeated five times at 7-day intervals, and pain intensity and duration were measured using a visual analog scale immediately before each procedure. A progressive and significant reduction in pain scores and a significant increase in the duration of analgesia were observed in all groups following the first three blocks, but no further improvement was obtained following the last two blocks. Drowsiness, the most frequent side effect, and dry mouth occurred only in patients submitted to SGB with lidocaine combined with clonidine. The three methods were similar regarding changes in pain intensity and duration of analgesia. However, IVRB seems to be preferable to SGB due to its easier execution and lower risk of undesirable effects.

Transplantation of mononuclear cells from human umbilical cord blood promotes functional recovery after traumatic spinal cord injury in Wistar rats

Cell transplantation is a promising experimental treatment for spinal cord injury. The aim of the present study was to evaluate the efficacy of mononuclear cells from human umbilical cord blood in promoting functional recovery when transplanted after a contusion spinal cord injury. Female Wistar rats (12 weeks old) were submitted to spinal injury with a MASCIS impactor and divided into 4 groups: control, surgical control, spinal cord injury, and one cell-treated lesion group. Mononuclear cells from umbilical cord blood of human male neonates were transplanted in two experiments: a) 1 h after surgery, into the injury site at a concentration of 5 x 10(6) cells diluted in 10 µL 0.9% NaCl (N = 8-10 per group); b) into the cisterna magna, 9 days after lesion at a concentration of 5 x 10(6) cells diluted in 150 µL 0.9% NaCl (N = 12-14 per group). The transplanted animals were immunosuppressed with cyclosporin-A (10 mg/kg per day). The BBB scale was used to evaluate motor behavior and the injury site was analyzed with immunofluorescent markers to label human transplanted cells, oligodendrocytes, neurons, and astrocytes. Spinal cord injury rats had 25% loss of cord tissue and cell treatment did not affect lesion extension. Transplanted cells survived in the injured area for 6 weeks after the procedure and both transplanted groups showed better motor recovery than the untreated ones (P < 0.05). The transplantation of mononuclear cells from human umbilical cord blood promoted functional recovery with no evidence of cell differentiation.

Effect of melatonin on the functional recovery from experimental traumatic compression of the spinal cord

Spinal cord injury is an extremely severe condition with no available effective therapies. We examined the effect of melatonin on traumatic compression of the spinal cord. Sixty male adult Wistar rats were divided into three groups: sham-operated animals and animals with 35 and 50% spinal cord compression with a polycarbonate rod spacer. Each group was divided into two subgroups, each receiving an injection of vehicle or melatonin (2.5 mg/kg, intraperitoneal) 5 min prior to and 1, 2, 3, and 4 h after injury. Functional recovery was monitored weekly by the open-field test, the Basso, Beattie and Bresnahan locomotor scale and the inclined plane test. Histological changes of the spinal cord were examined 35 days after injury. Motor scores were progressively lower as spacer size increased according to the motor scale and inclined plane test evaluation at all times of assessment. The results of the two tests were correlated. The open-field test presented similar results with a less pronounced difference between the 35 and 50% compression groups. The injured groups presented functional recovery that was more evident in the first and second weeks. Animals receiving melatonin treatment presented more pronounced functional recovery than vehicle-treated animals as measured by the motor scale or inclined plane. NADPH-d histochemistry revealed integrity of the spinal cord thoracic segment in sham-operated animals and confirmed the severity of the lesion after spinal cord narrowing. The results obtained after experimental compression of the spinal cord support the hypothesis that melatonin may be considered for use in clinical practice because of its protective effect on the secondary wave of neuronal death following the primary wave after spinal cord injury.

Effects of sciatic nerve transection on ultrastructure, NADPH-diaphorase reaction and serotonin-, tyrosine hydroxylase-, c-Fos-, glucose transporter 1- and 3-like immunoreactivities in frog dorsal root ganglion

Frogs have been used as an alternative model to study pain mechanisms. Since we did not find any reports on the effects of sciatic nerve transection (SNT) on the ultrastructure and pattern of metabolic substances in frog dorsal root ganglion (DRG) cells, in the present study, 18 adult male frogs (Rana catesbeiana) were divided into three experimental groups: naive (frogs not subjected to surgical manipulation), sham (frogs in which all surgical procedures to expose the sciatic nerve were used except transection of the nerve), and SNT (frogs in which the sciatic nerve was exposed and transected). After 3 days, the bilateral DRG of the sciatic nerve was collected and used for transmission electron microscopy. Immunohistochemistry was used to detect reactivity for glucose transporter (Glut) types 1 and 3, tyrosine hydroxylase, serotonin and c-Fos, as well as nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase). SNT induced more mitochondria with vacuolation in neurons, satellite glial cells (SGCs) with more cytoplasmic extensions emerging from cell bodies, as well as more ribosomes, rough endoplasmic reticulum, intermediate filaments and mitochondria. c-Fos immunoreactivity was found in neuronal nuclei. More neurons and SGCs surrounded by tyrosine hydroxylase-like immunoreactivity were found. No change occurred in serotonin- and Glut1- and Glut3-like immunoreactivity. NADPH-diaphorase occurred in more neurons and SGCs. No sign of SGC proliferation was observed. Since the changes of frog DRG in response to nerve injury are similar to those of mammals, frogs should be a valid experimental model for the study of the effects of SNT, a condition that still has many unanswered questions.

Expression of MMP-2 and MMP-9 in the rat trigeminal ganglion during the development of temporomandibular joint inflammation

Orofacial pain is a prevalent symptom in modern society. Some musculoskeletal orofacial pain is caused by temporomandibular disorders (TMDs). This condition has a multi-factorial etiology, including emotional factors and alteration of the masticator muscle and temporomandibular joints (TMJs). TMJ inflammation is considered to be a cause of pain in patients with TMD. Extracellular proteolytic enzymes, specifically the matrix metalloproteinases (MMPs), have been shown to modulate inflammation and pain. The purpose of this investigation was to determine whether the expression and level of gelatinolytic activity of MMP-2 and MMP-9 in the trigeminal ganglion are altered during different stages of temporomandibular inflammation, as determined by gelatin zymography. This study also evaluated whether mechanical allodynia and orofacial hyperalgesia, induced by the injection of complete Freund's adjuvant into the TMJ capsule, were altered by an MMP inhibitor (doxycycline, DOX). TMJ inflammation was measured by plasma extravasation in the periarticular tissue (Evans blue test) and infiltration of polymorphonuclear neutrophils into the synovial fluid (myeloperoxidase enzyme quantification). MMP expression in the trigeminal ganglion was shown to vary during the phases of the inflammatory process. MMP-9 regulated the early phase and MMP-2 participated in the late phase of this process. Furthermore, increases in plasma extravasation in periarticular tissue and myeloperoxidase activity in the joint tissue, which occurred throughout the inflammation process, were diminished by treatment with DOX, a nonspecific MMP inhibitor. Additionally, the increases of mechanical allodynia and orofacial hyperalgesia were attenuated by the same treatment.

Assessment of the diagnostic value of diffusion tensor imaging in patients with spinal cord compression: a meta-analysis

We investigated the diagnostic value of the apparent diffusion coefficient (ADC) and fractional anisotropy (FA) of magnetic resonance diffusion tensor imaging (DTI) in patients with spinal cord compression (SCC) using a meta-analysis framework. Multiple scientific literature databases were exhaustively searched to identify articles relevant to this study. Mean values and standardized mean differences (SMDs) were calculated for the ADC and FA in normal and diseased tissues. The STATA version 12.0 software was used for statistical analysis. Of the 41 articles initially retrieved through database searches, 11 case-control studies were eligible for the meta-analysis and contained a combined total of 645 human subjects (394 patients with SCC and 251 healthy controls). All 11 studies reported data on FA, and 9 contained data related to the ADC. The combined SMDs of the ADC and FA showed that the ADC was significantly higher and the FA was lower in patients with SCC than in healthy controls. Subgroup analysis based on the b value showed higher ADCs in patients with SCC than in healthy controls at b values of both ≤500 and >500 s/mm2. In summary, the main findings of this meta-analysis revealed an increased ADC and decreased FA in patients with SCC, indicating that DTI is an important diagnostic imaging tool to assess patients suspected to have SCC.

Effect of selective β-adrenoceptor blockade and surgical resection of the celiac-superior mesenteric ganglion complex on delayed liquid gastric emptying induced by dipyrone, 4-aminoantipyrine, and antipyrine in rats

There is evidence for participation of peripheral β-adrenoceptors in delayed liquid gastric emptying (GE) induced in rats by dipyrone (Dp), 4-aminoantipyrine (AA), and antipyrine (At). The present study aimed to determine whether β-adrenoceptors are involved in delayed GE induced by phenylpyrazole derivatives and the role of the prevertebral sympathetic nervous system in this condition. Male Wistar rats weighing 220-280 g were used in the study. In the first experiment rats were intravenously pretreated with vehicle (V), atenolol 30 mg/kg (ATE, β1-adrenergic antagonist), or butoxamine 25 mg/kg (BUT, β2-adrenergic antagonist). In the second experiment, rats were pretreated with V or SR59230A 2 mg/kg (SRA, β3-adrenergic antagonist). In the third experiment, rats were subjected to surgical resection of the celiac-superior mesenteric ganglion complex or to sham surgery. The groups were intravenously treated with saline (S), 240 µmol/kg Dp, AA, or At, 15 min after pretreatment with the antagonists or V and nine days after surgery. GE was determined 10 min later by measuring the percentage of gastric retention (%GR) of saline labeled with phenol red 10 min after gavage. The %GR (means±SE, n=6) values indicated that BUT abolished the effect of Dp (BUT+Dp vs V+Dp: 35.0%±5.1% vs 56.4%±2.7%) and At (BUT+At vs V+At: 33.5%±4.7% vs 52.9%±2.6%) on GE, and significantly reduced (P<0.05) the effect of AA (BUT+AA vs V+AA: 48.0%±5.0% vs 65.2%±3.8%). ATE, SRA, and sympathectomy did not modify the effects of treatments. These results suggest that β2-adrenoceptor activation occurred in delayed liquid gastric emptying induced by the phenylpyrazole derivatives dipyrone, 4-aminoantipyrine, and antipyrine. Additionally, the released neurotransmitter did not originate in the celiac-superior mesenteric ganglion complex.

Rescuing axons from degeneration does not affect retinal ganglion cell death

After a traumatic injury to the central nervous system, the distal stumps of axons undergo Wallerian degeneration (WD), an event that comprises cytoskeleton and myelin breakdown, astrocytic gliosis, and overexpression of proteins that inhibit axonal regrowth. By contrast, injured neuronal cell bodies show features characteristic of attempts to initiate the regenerative process of elongating their axons. The main molecular event that leads to WD is an increase in the intracellular calcium concentration, which activates calpains, calcium-dependent proteases that degrade cytoskeleton proteins. The aim of our study was to investigate whether preventing axonal degeneration would impact the survival of retinal ganglion cells (RGCs) after crushing the optic nerve. We observed that male Wistar rats (weighing 200-400 g; n=18) treated with an exogenous calpain inhibitor (20 mM) administered via direct application of the inhibitor embedded within the copolymer resin Evlax immediately following optic nerve crush showed a delay in the onset of WD. This delayed onset was characterized by a decrease in the number of degenerated fibers (P<0.05) and an increase in the number of preserved fibers (P<0.05) 4 days after injury. Additionally, most preserved fibers showed a normal G-ratio. These results indicated that calpain inhibition prevented the degeneration of optic nerve fibers, rescuing axons from the process of axonal degeneration. However, analysis of retinal ganglion cell survival demonstrated no difference between the calpain inhibitor- and vehicle-treated groups, suggesting that although the calpain inhibitor prevented axonal degeneration, it had no effect on RGC survival after optic nerve damage.