Switching Rat Resident Macrophages from M1 to M2 Phenotype by Iba1 Silencing Has Analgesic Effects in SNL-Induced Neuropathic Pain.

Resident macrophages from dorsal root ganglia are important for the development of traumatic-induced neuropathic pain. In the first 5-7 days after a traumatic sciatic nerve injury (i.e., spinal nerve ligation (SNL), spared nerve injury (SNI), sciatic nerve transection or sciatic nerve ligation and transection), Ionized binding adapter protein 1 (Iba1) (+) resident macrophages cluster around dorsal root ganglia neurons, possibly contributing to nerve injury-induced hypersensitivity. Since infiltrating macrophages gradually recruited to the lesion site peak at about 7 days, the first few days post-lesion offer a window of opportunity when the contribution of Iba1 (+) resident macrophages to neuropathic pain pathogenesis could be investigated. Iba1 is an actin cross-linking cytoskeleton protein, specifically located only in macrophages and microglia. In this study, we explored the contribution of rat Iba1 (+) macrophages in SNL-induced neuropathic pain by using intra-ganglionic injections of naked Iba1-siRNA, delivered at the time the lesion occurred. The results show that 5 days after Iba1 silencing, Iba1 (+) resident macrophages are switched from an M1 (pro-inflammatory) phenotype to an M2 (anti-inflammatory) phenotype, which was confirmed by a significant decrease of M1 markers (CD32 and CD86), a significant increase of M2 markers (CD163 and Arginase-1), a reduced secretion of pro-inflammatory cytokines (IL-6, TNF-α and IL-1ß) and an increased release of pro-regenerative factors (BDNF, NGF and NT-3) which initiated the regrowth of adult DRG neurites and reduced SNL-induced neuropathic pain. Our data show for the first time, that it is possible to induce macrophages towards an anti-inflammatory phenotype by interacting with their cytoskeleton.

Effects of Venlafaxine, Risperidone and Febuxostat on Cuprizone-Induced Demyelination, Behavioral Deficits and Oxidative Stress.

Multiple sclerosis (MS) is a demyelinating, autoimmune disease that affects a large number of young adults. Novel therapies for MS are needed considering the efficiency and safety limitations of current treatments. In our study, we investigated the effects of venlafaxine (antidepressant, serotonin-norepinephrine reuptake inhibitor), risperidone (atypical antipsychotic) and febuxostat (gout medication, xanthine oxidase inhibitor) in the cuprizone mouse model of acute demyelination, hypothesizing an antagonistic effect on TRPA1 calcium channels. Cuprizone and drugs were administered to C57BL6/J mice for five weeks and locomotor activity, motor performance and cold sensitivity were assessed. Mice brains were harvested for histological staining and assessment of oxidative stress markers. Febuxostat and metabolites of venlafaxine (desvenlafaxine) and risperidone (paliperidone) were tested for TRPA1 antagonistic activity. Following treatment, venlafaxine and risperidone significantly improved motor performance and sensitivity to a cold stimulus. All administered drugs ameliorated the cuprizone-induced deficit of superoxide dismutase activity. Desvenlafaxine and paliperidone showed no activity on TRPA1, while febuxostat exhibited agonistic activity at high concentrations. Our findings indicated that all three drugs offered some protection against the effects of cuprizone-induced demyelination. The agonistic activity of febuxostat can be of potential use for discovering novel TRPA1 ligands.

Toxicity of plant extracts containing pyrrolizidine alkaloids using alternative invertebrate models.

Pyrrolizidine alkaloids (PAs) are a widespread class of hepatotoxic heterocyclic organic compounds found in approximately 3% of world flora. Some PAs have been shown to have genotoxic and carcinogenic effects. The present study focuses on the toxicity effects of four dry extracts obtained from medicinal plants (Senecio vernalis, Symphytum officinale, Petasites hybridus and Tussilago farfara), on two aquatic organisms, Artemia salina and Daphnia magna, and the correlation with their PAs content. A new GC­MS method, using a retention time (TR)­5MS type capillary column was developed. PAs Kovats retention indices, for this type of column were computed for the first time. The lethal dose 50% (LC50) values for the two invertebrate models were correlated (Pearson 's coefficient, >0.9) and the toxicity was PA concentration-dependent, for three of the four extracts. All tested extracts were found to be toxic in both aquatic organism models. The results can be used to develop a GC­MS validated method for the assay of PAs in medicinal plants with a further potential application in the risk assessment study of PAs toxicity in humans.

Therapeutic potential of certain drug combinations on paclitaxel-induced peripheral neuropathy in rats.

BACKGROUND AND AIMS: Experimental research and clinical data support the potential combination therapy for the treatment of neuropathic pain. We aimed to investigate the analgesic effect of the following associations: gabapentin + etifoxine; tramadol + etifoxine; gabapentin + tramadol, in an experimental model of peripheral neuropathy induced by paclitaxel. MATERIALS AND METHODS: Neuropathy was induced in male Wistar rats by the daily administration of 2 mg÷kg body weight (bw) paclitaxel intraperitoneally, four days in a row. Analgesics were given concomitantly with paclitaxel, in the following doses: tramadol 15 mg÷kg bw, etifoxine 100 mg÷kg bw, gabapentin 300 mg÷kg bw. Tactile allodynia and mechanical hyperalgesia were assessed using the Dynamic Plantar Aesthesiometer apparatus (Ugo Basile). After 18 days of treatment, the brain and liver tissue susceptibility to lipid peroxidation was evaluated and the sciatic nerve histological examination of the effect on myelin fibers was performed. RESULTS AND CONCLUSIONS: Experimental data have shown a strong analgesic effect of these three tested combinations expressed mainly by the statistically significant increased maximum response time, both in the assessment of allodynia and hyperalgesia. The gabapentin + tramadol combination lead to the maximum analgesic effect, immediately after the discontinuation of paclitaxel (44.94%, p<0.0001) and throughout the study. The treatment associated with tramadol caused a reduction in lipid peroxidation in the brain as compared to paclitaxel group. Combination therapy showed reduced damage to myelinated fiber density in the sciatic nerve. The drug combinations used in the experiment showed therapeutic potential in the fight against neuropathic pain induced by the administration of taxanes.

Pharmacotoxicological screening on new derivatives of beta-phenylethylamine, potential agonists of beta3-adrenergic receptors.

BACKGROUND AND AIMS: Beta3-adrenergic receptors (beta3-ARs) have been initially characterized in 1989. Afterwards, their tissue distribution was established: white and brown adipose tissue, central nervous system, myocardium (atrial and ventricular), blood vessels, smooth gastrointestinal muscles (stomach, small intestine, colon), gallbladder, urinary bladder, prostate, skeletal muscles. Non-clinical trials have demonstrated the major implication of beta3-ARs in glucose metabolism, implicitly, in insulin release, and also in obesity. Therefore, new compounds were synthesized starting from beta-phenylethylamine nucleus and substituted in various positions, for possible antidiabetic and÷or antiobesity action. MATERIALS AND METHODS: In the present research, the antidiabetic action of newly synthesized compounds was investigated on an experimental model of alloxan-induced diabetes, administered in dose of 130 mg÷kg body weight (bw), intraperitoneally (i.p.). After 14 days of treatment, glycemia and enzymes involved in homeostasis of glucose metabolism, glucose-6-phosphate dehydrogenase (G6PD), glucose-6-phosphatase (G6Pase) and hexokinase were determined. Animals were then euthanized and histopathology examinations were performed on harvested liver, kidney, spleen and brain in order to document pathological changes induced by alloxan-induced diabetes and÷or by tested compounds. RESULTS AND CONCLUSIONS: Glycemia in animals treated with the tested compounds decreased statistically significant for groups C2 and C3 (-42.13% and -37.2%, respectively), compared to diabetic control group. C2 was also the compound to favorably modify the dynamics of determined enzymes, together with the display of very good safety profile supported by minor, non-significant, histopathological changes.

Influence of hyperforin on the morphology of internal organs and biochemical parameters, in experimental model in mice.

BACKGROUND AND AIMS: Hyperforin (HY) is used to treat depression and skin irritation and has been shown a number of pharmacological activities. The literature does no cite data on changes that may occur in the body after HY intake (ethylene diammonium salt - EDS) in long-term administration. From this point of view, the present work is a key to determining the pharmacotoxicological profile of the HY-EDS, in long-term administration. MATERIALS AND METHODS: In present research, the influence of toxic doses of HY-EDS was investigated on the biochemical serum parameters and the histopathological changes in internal organs on the experimental mice model. For acute toxicity determination, the HY-EDS was tested in doses of 2000-5000 mg÷kg, administered once per day orally. For subacute toxicity, the HY-EDS was tested in three groups of mice, in doses of 50, 75 and 100 mg÷kg÷day, administered once daily, for 28 consecutive days. RESULTS AND CONCLUSIONS: As concern acute toxicity, a lethal effect has not occurred at any of the two tested doses and HY-EDS was classified as Class V toxic: median lethal dose (LD50) >5000 mg÷kg, p.o. After 14 days of follow-up in acute toxicity, the experimental results showed a statistically significant increase of aspartate transaminase (AST) and alanine transaminase (ALT), compared to the control group. There were no changes in creatinine and serum glucose compared to the control group. After the administration of repeated doses, it was observed an increase of serum transaminases and alkaline phosphatase. Histological examination revealed that the liver injuries were in an initial stage, making them reversible in case of HY-EDS treatment discontinuation. There was no evidence of kidney damage to any of the doses of HY-EDS.