| Alzheimer's disease (AD) is a progressive neurodegenerative disorder that affects the elderly. Neuropathological hallmarks of the disease include profound brain atrophy, intracellular neurofibrillary tangles and extracellular amyloid plaques. Memory decline and cognitive impairment are the main clinical manifestations of AD. Considering the still growing number of the elderly, it has become an increasing burden for the society.The etiology of AD is still unknown. However, a converging lines of evidence indicate that brain inflammation induced by B-amyloid (AB) deposition is closely associated with the pathogenesis of this disease. The hallmark of brain inflammation is the activation of microglia and astrocytes. Upon activation, these glial cells can produce a variety of cytotoxic mediators, including nitric oxide, proinflammatory cytokines, excitatory amino acid, reactive oxygen intermediates, which are toxic to neurons, Thus, drugs with the capability to inhibit microglia or astrocyte activation would provide neuroprotection. In addition, reduced expression of neurotrophic factors can lead to neuronal death and contribute to the pathogenesis of AD. Therefore, the application of neurotrophic factors and inducers of neurotrophic factor biosynthesis would have enormous therapeutic potential for this disease.Triptolide (named as T10) is an extract from the traditional Chinese herb Tripterygium Wilfordii Hook F (TWHF). It has been identified as the major component responsible for the immunosuppressive and anti-inflammatory effects of TWHE Our previous work reported that T10 could exert neuroprotective effects on dopaminergic neurons in vitro and in vivo via its anti-inflammatory potential and neurotrophic effect, indicating that it is a promising drug in the treatment of Parkinson's disease (PD). To investigate if T10 also provides neuroprotection for AD and trying to explore its underlying mechanisms, in the present study, using rat microglia and astrocyte cultures, we compared the effects of different assembly forms of AB on microglia/astrocyte activation, established AB-activated glial cell model and investigated the anti-inflammatory effects of T10. In addition, we explored the neurotrophic property of T10 on astrocyte by detecting the effect of T10 treatment on synthesis and release of NGF, BDNF and GDNF in rat astrocyte cultures. We also explored the neuroprotective effect and mechanism of T10-treated astrocyte conditioned medium on AB-induced hippocampal neuronal apoptosis. The main findings are as follows:1. Establishment of AB1-42-activated microglia cell model and anti-inflammatory effect of T10Rat microglia cultures were treated with monomeric, oligomeric, fibrillar Aβ1-42 or Aβ42-1 and the effects of different Aβpeptide species on release of proinflammatory cytokines-tumor necrosis factor-a (TNF-a) and interleukin-1β(IL-1β) were detected. We also explored the effects of different concentrations T10 on TNF-a and IL-1βrelease from Aβ-activated microglia.1.1. Treatment with 5-20μM either assembly form of Aβ1-42 increased TNF-a and IL-1βrelease from microglia, however, the effect of oligomeric Aβ1-42 is much stronger than the other two species. After treatment with 10μM oligomeric Aβ1-42, the release level of TNF-a and IL-1βreached its highest at 12h treatment. Neither forms of Aβ42-1 had effect on the release of TNF-a and IL-1βfrom microglia.1.2. 10(-11)-10(-8)M T10 pretreatment significantly inhibited oligomeric Aβ1-42-induced TNF-a and IL-1βrelease from microglia.1.3. 10(-11)-10(-8)M T10 treatment did not alter microglia cell viability, suggesting that these concentrations of T10 did not have toxic effect on microglia.2. Establishment of Aβ1-42-activated astrocyte cell model and anti-inflammatory effect of T10 Rat astrocyte cultures were treated with monomeric, oligomeric, fibrillar Aβ1-42 and the effects of different Aβpeptide species on the release of TNF-a and IL-1βfrom astrocyte were detected. We also explored the anti-inflammatory effects of T10 on Aβ-activated astrocytes.2.1. Treatment with 1.25-20μM either forms of Aβ1-42 increased TNF-a and IL-1βrelease from astrocytes, the effect of oligomeric Aβ1-42 is the strongest. After treatment with 10μM oligomeric Aβ1-42, the release level of TNF-a and IL-1βreached its highest at 12h treatment.2.2. 10-10-10-7M T10 pretreatment significantly inhibited oligomeric Aβ1-42-induced TNF-a and IL-1βrelease from astrocytes. 10-10-10-7M T10 treatment did not alter astrocyte viability.3. Neurotrophic effect of T10 on astrocytes and neuroprotective effect of T10 on primary hippocampal neuronsCultured rat astrocytes were treated with different concentration of T10 and effects of T10 on synthesis and release of NGF, BDNF and GDNF from astrocytes were detected. We also investigated the protective effect of T10-treated astrocyte conditioned medium on oligomeric Aβ1-42-induced hippocampal neuronal damage.3.1. 10-10-10-7M T10 treatment had a potent and specific effect on enhancing the expression and release of NGF from astrocytes, with the maximum effect occurring at the concentration of 10-8M. However, T10 had no detectable effect on either BDNF or GDNF synthesis at both mRNA and protein levels.3.2. 10-11-10-7M T10-treated astrocyte conditioned medium significantly inhibited Aβ1-42-induced cell viability decrease and apoptotic cell death of primary hippocampal neurons. The T10-treated astrocyte conditioned medium also significantly inhibited Aβ1-42-induced Bax up-regulation and Bc1-2 down-regulation in hippocampal neurons.The above results suggest that oligomeric Aβ-induced glial cell activation plays an important role in mediating neuroinflammation in AD. T10 significantly inhibits oligomeric Aβ1-42-induced microglia/astrocyte activation and release of proinflammatory cytokines-TNF-a and IL-1β. T10 has a potent and specific effect on up-regulating the synthesis and release of NGF in cultured rat astrocytes. T10 may inhibit Aβ1-42-induced hippocampal neuron apoptosis by enhancing neurotrophic factor release from astrocytes.
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