Rosmarinic acid attenuates development and existing pain in a rat model of neuropathic pain: An evidence of anti-oxidative and anti-inflammatory effects.

We aimed to investigate the potential prophylactic and curative effects of rosmarinic acid, one of the main constituents of rosemary, on the neuropathic pain induced by chronic constriction injury (CCI) in rats. CCI was used to induce peripheral neuropathic pain. In prophylactic groups, rosmarinic acid (10, 20, and 40 mg/kg, i.p.) was administered from the day of surgery (day 0) for 14 days. In treatment group, rosmarinic acid (40 mg/kg) was given from day 5 (after the pain was established), for 7 days. The degree of mechanical allodynia, cold allodynia, and heat hyperalgesia were measured on days 0, 3, 5, 7, 10 and 14 post-surgery. The open field test was carried out to assess locomotor activity of animals. Lumbar spinal cord levels of astroglia activation marker, glial fibrillary acidic protein (GFAP), microglial activation marker, ionized calcium-binding adapter molecule 1 (Iba-1), toll-like receptor 4 (TLR-4), tumor necrosis factor alpha (TNF-α), inducible isoform of nitric oxide synthase enzyme (iNOS) and apoptotic factors were quantified via western blot on days 7 and 14. CCI rats showed a significant mechanical allodynia, cold allodynia and thermal hyperalgesia, compared to sham ones on day 3, persisted up to day 14 post-CCI. Rosmarinic acid was able to prevent and also attenuate CCI-induced behavioral features in prophylactic as well as treatment groups, respectively. A significant increase in the levels of TNF-α, iNOS, apoptotic factors (Bax, caspases 3, 9), Iba-1, TLR-4, and GFAP was observed on both days 7 and 14, which was suppressed by 14 days administration of rosmarinic acid. These findings further support the use of rosemary in traditional medicine to alleviate pain. Rosmarinic acid could be a promising compound in prophylaxis and treatment of neuropathic pain. Anti-apoptotic and anti-inflammatory effects of rosmarinic acid may have important roles in the observed antinociceptive properties.

Rahbardar MG ，Amin B ，Mehri S ，Mirnajafi-Zadeh SJ ，Hosseinzadeh H ... -  《-》

Effect of alcoholic extract of aerial parts of Rosmarinus officinalis L. on pain, inflammation and apoptosis induced by chronic constriction injury (CCI) model of neuropathic pain in rats.

According to traditional medicine, rosemary (Rosmarinus officinalis) has been used in many ailments such as dysmenorrhea, rheumatic pain and stomachache. This work was carried out to evaluate putative anti-allodynic and anti-hyperalgesic effects of Rosmarinus officinalis alcoholic extract and some spinal cord molecular changes on a neuropathic pain model in rats. Peripheral neuropathy was induced by chronic constriction injury (CCI) of sciatic nerve. Rats were treated daily with alcoholic extract of aerial parts of R. officinalis (100, 200, and 400mg/kg, i.p.), from the day of surgery (day 0) for 14 days. Mechanical allodynia, cold allodynia and heat hyperalgesia were measured on days 0, 3, 5, 7, 10 and 14. Investigations into mechanisms involved measurement of apoptotic factors (bcl-2-like protein (Bax)), cleaved caspases 3 and 9, anti-apoptotic mediator, Bcl2, inflammatory mediators including tumor necrosis factor alpha (TNF-α), inducible isoform of nitric oxide synthase (iNOS), toll like receptor 4 (TLR4), microglial activation marker, ionized calcium-binding adapter molecule 1 (Iba-1) and astroglia activation marker, glial fibrillary acidic protein (GFAP) were measured via western blot on days 7 and 14. CCI rats exhibited a marked mechanical allodynia, cold allodynia, and thermal hyperalgesia on days 3, 5, 7, 10, and 14 post-CCI. All three doses of rosemary alcoholic extract were able to attenuate neuropathic behavioral changes as compared with CCI animals that received vehicle. In the vehicle-treated CCI group, a significant increase in levels of Bax, cleaved caspases 3 and 9, Iba1, TNF-α, iNOS and TLR4 levels was detected on both days 7 and 14. Rosemary extract, 400mg/kg significantly decreased the amounts of mentioned apoptotic, inflammatory and glial markers as compared with vehicle-treated CCI animals. Anti-inflammatory and anti-apoptotic processes might contribute to the anti-allodynic and anti-hyperalgesic effects of rosemary following nerve injury. Our findings support the ethnopharmacological use of rosemary as a potential candidate in treating neuropathic pain and different neurological disorders that associate with the activation of apoptosis and inflammatory pathways.

Ghasemzadeh MR ，Amin B ，Mehri S ，Mirnajafi-Zadeh SJ ，Hosseinzadeh H ... -  《-》

Anti-inflammatory effects of ethanolic extract of Rosmarinus officinalis L. and rosmarinic acid in a rat model of neuropathic pain.

Rosemary, Rosmarinus (R.) officinalis L. is a well-known plant with several useful properties such as analgesic, anti-inflammatory and anti-neurodegenerative. It has been used in folk medicine to alleviate rheumatic pain, stomachache and dysmenorrhea. Rosemary has several constituents such as rosmarinic acid which can be responsible for therapeutic properties been noted with rosemary. The aim of this study was to investigate the potential anti-inflammatory effects of R. officinalis and rosmarinic acid in a rat model of sciatic nerve chronic constriction injury (CCI)-induced neuropathic pain to verify usage of rosemary in folk medicine. Rats underwent CCI, were treated with either normal saline, ethanolic extract of aerial parts of R. officinalis (400mg/kg, i.p.) or rosmarinic acid (40mg/kg, i.p.) from the day of surgery (day 0) for 14days. The anti-inflammatory effects of R. officinalis extract and rosmarinic acid were evaluated by assessing the levels of some spinal inflammatory markers including cyclooxygenase-2 (COX2), prostaglandin E2 (PGE-2), interleukin 1 beta (IL-1β), matrix metallopeptidase 2 (MMP2) through western blotting and nitric oxide (NO) production via Griess reaction on days 7 and 14 post-surgery. CCI rats exhibited a marked expression in the levels of inflammatory markers (COX2, PGE-2, IL-1β, MMP2 and NO) on both days 7 (p<0.001) and 14 (p<0.001). Rosmarinic acid and ethanolic extract of R. officinalis were able to decrease amounts of mentioned inflammatory markers on both days 7 (p<0.001) and 14 (p<0.001). Our data support the traditional use of R. officinalis as an effective remedy for pain relief and inflammatory disorders. It also suggests that the ethanolic extract of R. officinalis and rosmarinic acid through modulating neuro-inflammation might be potential candidates in treating neuropathic pain and different neurological disorders associated with inflammation.

Ghasemzadeh Rahbardar M ，Amin B ，Mehri S ，Mirnajafi-Zadeh SJ ，Hosseinzadeh H ... -  《-》

The Effect of Verbascoside in Neuropathic Pain Induced by Chronic Constriction Injury in Rats.

We examined the effects of verbascoside in rats subjected to chronic constriction injury (CCI). Verbascoside (50, 100, and 200 mg/kg, i.p.), was administered from the day of surgery for 14 days. Spinal cord levels of apoptotic factors and glia markers were quantified on days 3, 7, and 14 post-CCI. Oxidative stress markers were assessed on days 7 and 14. CCI rats exhibited a marked mechanical allodynia, cold allodynia, and thermal hyperalgesia on days 3, 5, 7, 10, and 14 post-CCI. A significant increase in the levels of Iba (a marker of microglia activation) and Bax (a proapoptotic factor) was observed on day 3. Iba remained high on day 7. In contrast, there were no differences in glial fibrillary acidic protein contents between sham and CCI animals. Malondialdehyde increased and reduced glutathione decreased on day 14. Verbascoside significantly attenuated behavioral changes associated with neuropathy. Bax decreased, while Bcl-2 was increased by verbascoside on day 3. Verbascoside also reduced Iba protein on days 3 and 7. The results support evidence that microglial activation, apoptotic factors, and oxidative stress may have a pivotal role in the neuropathic pain pathogenesis. It is suggested that antinociceptive effects elicited by verbascoside might be through the inhibition of microglia activation, apoptotic pathways, and antioxidant properties.

Amin B ，Poureshagh E ，Hosseinzadeh H 《-》

Effects of intraperitoneal thymoquinone on chronic neuropathic pain in rats.

In this study, we evaluated the protective effects of thymoquinone, the major constituent of Nigella sativa seeds on the neuropathic pain of rats with chronic constrictive injury of the sciatic nerve. Rats received repeated administration of thymoquinone (1.25, 2.5, and 5 mg/kg, i. p.) once a day for 14 days, beginning immediately after the nerve injury. Mechanical allodynia, cold allodynia, and thermal hyperalgesia were assessed with the von Frey filament, acetone drop, or radiant heat stimulus, respectively. Recent evidence points towards a role of oxidative stress, spinal glia activation, and cell death in the pathogenesis of neuropathic pain. Ionized calcium-binding adapter molecule 1 (a marker of microglia), glial fibrillary acidic protein (a marker of astroglia), Bcl2-associated X protein (a proapoptotic protein), and B-cell lymphoma protein 2 (an antiapoptotic protein) were measured using Western blot on days 3, 7, and 14 post chronic constrictive injury. The changes in the protein levels of malondialdehyde and glutathione, biomarkers of oxidative stress, were assessed by spectrophotometric assay on day 14 post chronic constrictive injury. Repeated treatment with thymoquinone (2.5 and 5 mg/kg) significantly alleviated behavioral signs of neuropathic pain. In the lumbar spinal cord of neuropathic rats, ionized calcium-binding adapter molecule 1 and Bcl2-associated X protein increased on day 3 post chronic constrictive injury, whereas B-cell lymphoma protein 2 did not significantly change. After repeated thymoquinone administration, the elevated Bcl2-associated X protein and ionized calcium-binding adapter molecule reduced on day 3, while the level of B-cell lymphoma protein 2 was even stimulated. Ionized calcium-binding adapter molecule and Bcl2-associated X protein/B-cell lymphoma protein 2 ratio declined by days 7 and 14; consequently, there were no significant differences among groups. No or little change was observed in the glial fibrillary acidic protein content during the study. Chronic constrictive injury produced a significant increase in the levels of malondialdehyde and decrease in the contents of glutathione on day 14. Thymoquinone treatment (2.5 and 5 mg/kg) restored the levels of malondialdehyde. High dose of thymoquinone (5 mg/kg) also reversed the decreased glutathione in the injured animals. Our results indicate that, microglia, apoptotic factors, and oxidative stress rather than astroglia contribute to the pathogenesis of chronic constrictive injury, and thymoquinone plays an anti-nociceptive role possibly by antioxidant effects and inhibition of microglia activity.

Amin B ，Taheri MM ，Hosseinzadeh H 《-》