Arsenic compounds activate MAPK and inhibit Akt pathways to induce apoptosis in MA-10 mouse Leydig tumor cells.

Arsenic compounds have been applied treating acute promyelocytic 1eukemia and solid tumors with brief mechanism investigations. In fact, we have demonstrated that sodium arsenite plus dimethylarsenic acid could activate apoptosis in MA-10 mouse Leydig tumor cells by inducing caspase pathways. However, detail underlying mechanisms how caspase cascade is regulated remains elusive. Therefore, the apoptotic mechanism of sodium arsenite plus dimethylarsenic acid were examined in MA-10 cells in this study. Our results reveal that Fas/FasL protein expressions were stimulated by sodium arsenite plus dimethylarsenic acid in MA-10 cells. In addition, reactive oxygen species (ROS) generation, cytochrome C release, Bid truncation, and Bax translocation were induced in MA-10 cells by arsenic compounds. Moreover, activation of p38, JNK and ERK1/2, MAPK pathways was stimulated while Akt phosphorylated levels and Akt expression were decreased by sodium arsenite plus dimethylarsenic in MA-10 cells. In conclusion, sodium arsenite and dimethylarsenic acid did activate MAPK pathway plus ROS generation, but suppress Akt pathway, to modulate caspase pathway and then induce MA-10 cell apoptosis.

The application of flow cytometry for evaluating biological aggressiveness of intracranial meningiomas.

BACKGROUND: Meningiomas have classically been considered to include benign and atypical/anaplastic tumors. Despite the availability of clinical and pathologic parameters for prognostic prediction prognosis, the behavior of each meningioma may be difficult to predict. Here, we used DNA flow-cytometric studies to predict biological tumor behaviors of intracranial meningiomas. METHODS: The specimens were obtained from fresh tumoral tissues of 43 microsurgically resected meningiomas as approved by the institutional review board. The presence of G2/M-phase and S+G2/M-phase fractions were analyzed and correlated with the proliferation index of Ki-67 and the World Health Organization grading. The check point of G2/M-phase fraction, cyclin B, and pCdk1 (Y15), were analyzed by Western blotting. RESULTS: Our results showed that there were significant differences in Ki-67, G2/M-phase, S+G2/M-phase fractions, and cyclin B between benign and atypical/anaplastic meningiomas. The optimal cutoff point of G2/M-phase and S+G2/M-phase fractions were 5.12 and 7.52%, respectively, and this can be used to discriminate those cases with benign or atypical/anaplastic meningiomas. Besides, both the G2/M-phase and S+G2/M-phase fractions were correlated well with Ki-67 and the histopathological features such as focal necrosis, infiltration of dura mater and mitotic activity. In addition, the occurrence of tumor recurrence and patient age were correlated to the G2/M-phase and S+G2/M-phase fractions, respectively. The G2/M-phase and S+G2/M-phase fractions, however, did not correlate well with histologic invasion to adjacent bone, sinus, or brain tissues. CONCLUSIONS: The use of flow cytometry facilitates additional information for G2/M-phase and S+G2/M-phase fractions represent tumoral grading and risk of recurrence in patients with meningiomas.

Melatonin improves neuroplasticity by upregulating the growth-associated protein-43 (GAP-43) and NMDAR postsynaptic density-95 (PSD-95) proteins in cultured neurons exposed to glutamate excitotoxicity and in rats subjected to transient focal cerebral ischemia even during a long-term recovery period.

Recent evidence shows that the NMDAR postsynaptic density-95 (PSD-95), growth-associated protein-43 (GAP-43), and matrix metalloproteinase-9 (MMP-9) protein enhance neuroplasticity at the subacute stage of stroke. Here, we evaluated whether melatonin would modulate the PSD-95, GAP-43, and MMP-9 proteins in cultured neurons exposed to glutamate excitotoxicity and in rats subjected to experimental stroke. Adult male Sprague-Dawley rats were treated with melatonin (5 mg/kg) or vehicle at reperfusion onset after transient occlusion of the right middle cerebral artery (tMCAO) for 90 min. Animals were euthanized for Western immunoblot analyses for the PSD-95 and GAP-43 proteins and gelatin zymography for the MMP-9 activity at 7 days postinsult. Another set of animals was sacrificed for histologic and Golgi-Cox-impregnated sections at 28 days postinsult. In cultured neurons exposed to glutamate excitotoxicity, melatonin significantly upregulated the GAP-43 and PSD-95 expressions and improved dendritic aborizations (P<0.05, respectively). Relative to controls, melatonin-treated stroke animals caused a significant improvement in GAP-43 and PSD-95 expressions as well as the MMP-9 activity in the ischemic brain (P<0.05). Consequently, melatonin also significantly promoted the dendritic spine density and reduced infarction in the ischemic brain, and improved neurobehaviors as well at 28 days postinsult (P<0.05, respectively). Together, melatonin upregulates GAP-43, PSD-95, and MMP-9 proteins, which likely accounts for its actions to improve neuroplasticity in cultured neurons exposed to glutamate excitotoxicity and to enhance long-term neuroprotection, neuroplasticity, and brain remodeling in stroke rats.

Melatonin inhibits matrix metalloproteinase-9 (MMP-9) activation in the lipopolysaccharide (LPS)-stimulated RAW 264.7 and BV2 cells and a mouse model of meningitis.

We explored anti-inflammatory potential of melatonin against the lipopolysaccharide (LPS)-induced inflammation in vivo and in vitro. RAW 264.7 and BV2 cells were stimulated by LPS, followed by the treatment with melatonin or vehicle at various time intervals. In a mouse model of meningitis induced by LPS, melatonin (5mg/kg) or vehicle was intravenously injected at 30min postinsult. The activity of matrix metalloproteinase-2 (MMP-2) and metalloproteinase-9 (MMP-9) was determined by gelatin zymography. Nuclear factor-kappa B (NFκB) translocation and binding activity were determined by immunocytochemistry and electrophoretic mobility shift assay (EMSA). Our results showed that either pretreatment or cotreatment with melatonin at 50-500 µm effectively inhibited the LPS-induced proMMP-9 activation in the RAW 264.7 and BV2 cells, respectively (P<0.05). This melatonin-induced proMMP-9 inhibition remained effective when treatment was delayed up to 2 and 6hr postinsult for RAW 264.7 and BV2 cells, respectively (P<0.05 for both groups). Additionally, melatonin significantly attenuated the rises of circulatory and cerebral MMP-9 activity, respectively (P<0.05) and reduced the loss of body weight (P<0.05) in mice with meningitis. Moreover, melatonin (50µm) effectively inhibited nuclear factor-kappa B (NFκB) translocation and binding activity in the LPS-treated RAW 264.7 and BV2 cells, respectively (P<0.05). These results demonstrate direct inhibitory actions of melatonin against postinflammatory NFκB translocation and MMP-9 activation and highlight its ability to inhibit systemic and cerebral MMP-9 activation following brain inflammation.

Optimal Percoll concentration facilitates flow cytometric analysis for annexin V/propidium iodine-stained ischemic brain tissues.

We sought to determine the optimal Percoll concentration for ischemic rat brain prepared for flow cytometric (FC) measurements. Animals were subjected to the right middle cerebral artery (MCA) occlusion, and were euthanized at 3, 12, 24, and 72 h after reperfusion onset. The brains were processed by different concentrations (unisolated, 20, 25, 30, or 40%) of Percoll and stained with annexin V/propidium iodine (PI). Ischemic brain damage was evaluated by FC analysis and image analysis for histologic sections. The relative susceptibility of different phenotypes of cells to necrotic and apoptotic damage were evaluated by the FC analyses for the immunohistochemistry, PI, and the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL)-processed brain tissues. Our results showed that FC analysis effectively detected the extent and maturation of apoptotic/necrotic brain damage, and the results were consistent with those determined from histologic brain sections. Neuron was more vulnerable to apoptosis than glia, whereas both cellular phenotypes were compatible in susceptibility for necrotic cell death. Percoll at a low concentration (20%) could effectively remove tissue debris without affecting membranous integrity of the injured neurons. Conversely, high percentages of Percoll (30-40%) substantially increased membranous damage for the injured cells. These results supported the application of FC to determine the extent and progression in time, as well as relative phenotypes of apoptotic/necrotic cell deaths following ischemic damage. We highlighted the use of Percoll at low percentages to facilitate the removal of tissue debris and to improve membrane integrity preservation for the injured neurons.

Multiple tenting techniques improve dead space obliteration in the surgical treatment for patients with giant calcified chronic subdural hematoma.

BACKGROUND: Calcified chronic subdural hematoma (CCSDH), or "armored brain," is a rare disease entity. The optimal surgical procedure for CCSDH has not been established because it is hard to obtain brain re-expansion after surgery. In particular, a large CCSDH is difficult to completely extirpate, and the residual rigid inner and outer membranes facilitates dead space retention and hematoma recurrence. METHODS: We introduce the use a multiple suturing technique to tent the residual outer and inner membranes onto the dura matter so as to obliterate dead space after surgical treatment for CCSDH. Neuroimaging and surgical reports with illustrative images from two cases are shown. RESULTS: Two patients were admitted to our intensive care unit more than 10 years apart from their ventriculoperitoneal (V-P) shunt placements. The first patient presented with clinical signs of increased intracranial pressure. The second patient had a large CCSDH as a concomitant finding with ruptured aneurysmal subarachnoid hemorrhage. Computerized cranial tomography demonstrated large hematoma cavities with thick calcified inner membranes. After neurosurgical intervention by craniotomy and optimal resection of calcified membranes and muddy blood clot, we tented the residual calcified inner and outer membranes onto the dura matter by multiple sutures to reduce dead space accumulation. Postoperatively, the two patients had improved clinical symptoms along with much reduced hematoma cavity in imaging examinations. CONCLUSIONS: We reported an alternative technique using multiple tenting procedures to improve dead space obliteration after surgical treatment for patients with a large CCSDH presenting as a late complication after V-P shunting.