mercaptopurine and Subarachnoid-Hemorrhage

Upregulation of high-level toll-like receptors (TLRs) is observed in the serum of animals following experimental subarachnoid hemorrhage (SAH) and is highly related to SAH-induced early brain injury (EBI). The present study was of interest to examine the effect of 6-mercaptopurine (6-MP) on alternation of TLR-2, -3, and -4 in this model.. A rodent SAH model was used. Administration with 6-MP (0.5/1/2 mg/kg/d) was initiated 1 h after the induction of SAH via an osmotic minipump. Cerebral cortex was harvested to measure TLRs messenger RNA and protein (reverse transcription polymerase chain reaction [rt-PCR] and Western blot). Cerebral cortex was harvested for activated caspases (rt-PCR) measurement.. Cellular evaluation revealed increased neuronal nuclei(+) neurons with vacuolated nuclear and glial fibrillary acidic protein(+) astrocytes in the SAH group, but absent in the 6-MP treatment and healthy controls. The TLR-3 levels were not significantly increased in animals subject to SAH, compared with the controls (no SAH). The levels of TLR-2 and -4 in the SAH only and SAH plus vehicle groups were significantly elevated (P < 0.01), and treatment with 6-MP reduced TLR-2, -3 (at 2 mg/kg), and -4 (dose-dependently) protein expression following SAH. Likewise, the TLR-4 messenger RNA levels were also significantly reduced in the 6-MP (at 1 mg/kg and 2 mg/kg) groups. Cleaved caspase-3 and caspase-9a were reduced at 2-mg/kg 6-MP treatment group.. These results show that 6-MP attenuates the expression of TLR-2, -4, especially TLR-4, which play an antiapoptotic effect on SAH-induced EBI. This finding supported that through modulating TLRs, 6-MP can attenuate SAH-induced EBI. Those results offer credit to the neuroprotective effect of 6-MP.

Topics: Animals; Antigens, Nuclear; Antimetabolites; Brain Injuries; Caspase 3; Caspase 9; Cytokines; Disease Models, Animal; Drug Evaluation, Preclinical; Male; Mercaptopurine; Nerve Tissue Proteins; Proto-Oncogene Proteins c-bcl-2; Random Allocation; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Toll-Like Receptors

Increased endothelin-1 (ET-1) production and diminished nitric oxide synthase (NOS) bioavailability has been observed in aneurysmal subarachnoid hemorrhage (SAH). The authors previously found that 6-mercaptopurine (6-mp) is effective in preventing and reversing arterial narrowing in a rodent SAH model. This present study is of interest to examine the effect of 6-mp on ET-1/endothelial nitric oxide synthase (eNOS) in this animal model.. A rodent double hemorrhage SAH model was employed. Animals were randomly assigned to six groups (sham, SAH only, vehicle, 0.5, 1.0 and 2 mg kg(-1) day(-1) 6-mp treatment). Monoclonal CD45 immunostaining was utilized to evaluate monocytes and microglia. The level of pro-inflammatory cytokines, such as IL-1, IL-6 and TNF-α(RT-PCR), and ET-1 (ELISA) was measured. The basilar arteries (BAs) were harvested and sliced, and their cross-sectional areas were determined. Radiolabeled NOS assay kit was applied to detect eNOS.. Morphologically, convolution of internal elastic lamina, endothelial cells distortion, and necrotic smooth muscle were prevalently present in the basilar artery of SAH groups, which was absent in the 1 and 2 mg kg(-1) day(-1) 6-mp plus SAH group or the healthy controls. Significant vasospasm was noted in the vehicle group (lumen patency, 54.6%, p ≤ 0.01 compared with the sham group), but it was less prominent in the 2 mg kg(-1) day(-1) 6-mp treatment group (lumen patency, 87.6%, p < 0.05). In addition, administration with 2 mg kg(-1) day(-1) 6-mp reduced cytokine levels by 11%, 47%, and 34% for IL-1, IL-6, and TNF-α, respectively, and increased ET-1 levels were found in all the animals subject to SAH (SAH only, SAH plus vehicle, SAH plus 0.5 and 1.0 mg kg(-1) day(-1) 6-mp) except in the 2 mg kg(-1) day(-1) 6-mp SAH group, when compared with the healthy controls (no SAH). Meanwhile, treatment with 6-mp did not induce the levels of expressed eNOS in BAs in the 6-mp groups (0.5, 1.0, and 2 mg kg(-1) day(-1) 6-mp plus SAH) when compared with that in the SAH groups (p > 0.1).. In summary, treatment with 6-mp decreased the release of pro-inflammatory cytokines and diminished experimental vasospasm. This study offered first evidence that 6-mp dose-dependently reduces the level of ET-1 in a NO-independent mechanism, which corresponds to its antivasospastic effect in the condition of chronic vasospasm.

Topics: Aneurysm, Ruptured; Animals; Chemotaxis; Connective Tissue; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelin-1; Immunosuppressive Agents; Inflammation Mediators; Intracranial Aneurysm; Male; Mercaptopurine; Microglia; Muscle, Smooth, Vascular; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Vasospasm, Intracranial

Adhesion molecules, including intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin, are important inflammatory mediators which are elevated in the serum of patients following aneurysmal subarachnoid hemorrhage (SAH). The authors previously found that 6-mercaptopurine (6-mp) was effective in preventing and reversing arterial narrowing in a rodent SAH model. The present study was to examine whether levels of adhesion molecules were altered after treatment with 6-mp in this animal model.. Animals were each injected with autologous blood into the cisterna magna, and intraperitoneal treatment with 6-mp (2 mg/kg) was initiated 1 h before (prevention) or later (treatment). The compound was subsequently administered at 24 and 48 h post-SAH. Blood samples were collected at 72 h post-SAH to measure ICAM-1, VCAM-1, and E-selectin levels. The basilar arteries were harvested and sliced, and their cross-sectional areas were measured. Morphologically, convolution of the internal elastic lamina, distorted endothelial wall, and myonecrosis of the smooth muscle were prominently observed in the SAH only and vehicle-treated SAH groups, but not in the 6-mp-treated SAH group or in healthy controls. No significant differences were found in the levels of VCAM-1 among all groups. However, the levels of E-selectin were increased in all animals subjected to SAH (SAH only and SAH plus vehicle groups) compared with healthy controls (no SAH), but not in the 6-mp group (SAH plus 6-mp treatment and preventive treatment with 6-mp).Likewise, the levels of ICAM-1 in the SAH only and SAH plus vehicle groups were significantly elevated (p < 0.001), and pretreatment and treatment with 6-mp reduced ICAM-1 to control levels.. These results show that ICAM-1 and E-selectin may play a role in mediating SAH-induced vasospasm and that a reduction of both adhesive molecules after SAH may partly contribute to the antispastic effect of 6-mp.

Topics: Animals; Antimetabolites; Basilar Artery; Cell Adhesion Molecules; Cerebral Arteries; Disease Models, Animal; E-Selectin; Inflammation Mediators; Intercellular Adhesion Molecule-1; Male; Mercaptopurine; Rats; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Treatment Outcome; Up-Regulation; Vascular Cell Adhesion Molecule-1; Vasodilation; Vasospasm, Intracranial

Topics: Adolescent; Child; Hematoma; Hematoma, Epidural, Cranial; Hematoma, Subdural; Humans; Hydrocephalus; Leucovorin; Leukemia; Leukemia, Lymphoid; Leukemia, Myeloid; Meninges; Mercaptopurine; Methotrexate; Pathology; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Prednisone; Radiography; Subarachnoid Hemorrhage; Vinblastine