| Summary: | 碩士 === 國防醫學院 === 生理學研究所 === 101 === Intracerebral hemorrhage (ICH) induces a complex sequence of pathological cascades, including oxidative stress, neurogenic inflammation and apoptosis. These factors contribute to blood-brain barrier (BBB) breakdown, brain edema and lead to subsequent brain damage. Transient receptor potential vanilloid subtype 1 (TRPV1), a nonselective cation channel activated by the vanilloids, has been reported to trigger apoptotic cell death via increasing intracellular Ca2+ accumulation, causing mitochondrial disruption in cultured neurons. Activation of TRPV1 also initiated neurogenic inflammation by releasing inflammatory peptides. However, the contribution of TRPV1 to ICH has not yet been reported. This study hypothesized that TRPV1 would play a deleterious role via regulating neurogenic inflammation and apoptosis in ICH.
Primary cultured neuron or mixed glial cultures of C57BL/6 background were stimulated with hemin and exposed to capsaicin (CAP). The mouse ICH model was performed by intracerebral injection of bacterial collagenase (type VII-S, 0.05U) into the C57BL/6 mice striatum. Capsaicin (1 or 3M) or vehicle was intracerebrally injected 30 mins before ICH. TRPV1-knock out (KO) mice were also used to examine the effects of TRPV1 deficiency in ICH.
The results showed that the TRPV1 protein level was upregulated in the peri-hematomal area, predominantly localizing in neurons following mouse ICH. In hemin-stimulated cultured neurons, a brief exposure (10 min) of CAP led to a 12.12% reduction of cell viability by the MTT test. In contrast, after a longer exposure (1 h and 24 h), cell viability was increased by 15.72% and 25.88% compared with the vehicle control. These changes were associated with an increase of [Ca2+]i following a 10- min exposure of CAP and a reversed reduction of [Ca2+]i following a 24-h exposure using fluorescent Ca2+ images. However, CAP did not influence the increase in hemin-induced nitric oxide production in glial cells. After ICH, deficiency or desensitization of TRPV1 following 24-h exposure of CAP resulted in reduced tissue injury and neuronal degeneration at 1 day. TRPV1 KO mice also exhibited better motor performance up to 28 days. Furthermore, TRPV1 desensitization attenuated neuropeptide substance P expression, BBB permeability, brain edema and neutrophil infiltration at 1 day. Deficiency or desensitization of TRPV1 also reduced the cleaved caspase-3 protein level at 1 day following ICH. The protective effect of TRPV1 desensitization was associated with a significant decrease of cleaved caspase-3, cytochrome C and Smac release to the cytosol in cultured neurons. These data suggest that TRPV1 plays a deleterious role in ICH-induced brain injury. Induction of TRPV1 desensitization may translate into a novel therapy for hemorrhagic stroke.
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