(−)-DHMEQ, a newly designed NF-B inhibitor, inhibited RANKL-induced osteoclast differentiation in mouse BMMs through downregulation of the induction of NFATc1, an essential transcription factor of osteoclastogenesis.Introduction: Bone destruction is often observed in advanced case of rheumatoid arthritis and neoplastic diseases, including multiple myeloma. Effective and nontoxic chemotherapeutic agents are expected for the suppression of these bone destructions. RANKL induces activation of NF-B and osteoclastogenesis in bone marrow-derived monocyte/macrophage precursor cells (BMMs). Targeted disruption or pharmacological suppression of NF-B result in impaired osteoclastogenesis, but how NF-B is involved in the regulation of osteoclastogenesis is not known. Materials and Methods:The effect of (−)-dehydroxymethylepoxyquinomicin [(−)-DHMEQ] on osteoclast differentiation was studied using a culture system of mouse BMMs stimulated with RANKL and macrophage colony-stimulating factor. The mechanism of the inhibition was studied by biochemical analysis such as immunoblotting and retroviral transfer experiments. Results: (−)-DHMEQ strongly inhibited RANKL-induced NF-B activation in BMMs and inhibited RANKL-induced formation of TRACP+ multinucleated cells. Interestingly, (−)-DHMEQ specifically inhibited the RANKL-induced expression of NFATc1 but not the expressions of TRAF6 or c-fos. Inhibition of osteoclast differentiation by (−)-DHMEQ was rescued by overexpression of NFATc1, suggesting that the inhibition is not caused by a toxic effect. Moreover, pit formation assays showed that (−)-DHMEQ also inhibited the bone-resorbing activity of mature osteoclasts. Conclusion:The inhibition of NF-B suppresses osteoclastogenesis by downregulation of NFATc1, suggesting that NFATc1 expression is regulated by NF-B in RANKL-induced osteoclastogenesis. Our results also indicate the possibility of (−)-DHMEQ becoming a new therapeutic strategy against bone erosion.

The hippocampal dentate gyrus in adult animals is known to contain neural progenitors that proliferate and differentiate into neurons in response to brain injury. Little has been observed, however, on regeneration of the granule cell layer of the dentate gyrus that has been directly injured. Using trimethyltin (TMT)-treated mice as an in vivo model, we evaluated the ability of this layer to regenerate after injury. The administration of TMT induced neuronal death in the dentate gyrus selectively 2 days later, with recovery of granule neurons on day 14 and thereafter. At an early stage (days 2-5) after the damage by TMT treatment, 5-bromo-2'-deoxyuridine (BrdU) incorporation into at least two different types of cells was facilitated in the dentate gyrus: BrdU-positive/neuronal nuclear antigen (NeuN)-negative cells were found predominantly in the subgranular zone and granule cell layer, whereas BrdU-positive/NeuN-positive cells were numerous in the dentate molecular layer and hilus. In addition, expression of proliferating cell nuclear antigen, nestin, NeuroD3, and doublecortin, which are markers for proliferating cells and neural progenitors/neuronal precursors, was extremely enhanced in the dentate gyrus at the early stage after treatment. Double staining revealed that BrdU was colocalized with nestin and doublecortin in the subgranular zone. Behavioral analysis revealed that TMT-induced cognition impairment was ameliorated by day 14 after the treatment. Taken together, our data indicate that the hippocampal dentate gyrus itself is capable of regenerating the neuronal cell layer through rapid enhancement of neurogenesis after injury.

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Abstract. The organotin trimethyltin (TMT) is known to cause neuronal degeneration in the murine brain. Earlier studies indicate that TMT-induced neuronal degeneration is enhanced by adrenalectomy. However, no evaluation has been attempted to determine the mechanism underlying the enhancement of TMT neurotoxicity by adrenalectomy and its implications in neuronal degeneration. To assess the implications and determine the mechanism of adrenalectomyelicited enhancement of TMT neurotoxicity, we examined neuronal degeneration and associated signaling pathways in adrenalectomized mice. Adrenalectomy dramatically enhanced the TMTinduced neuronal damage in certain brain regions including the dentate gyrus, olfactory bulb, and anterior olfactory nucleus, in addition to exacerbating the behavioral abnormalities. TMTinduced activation of caspase-3 and calpain was also enhanced by adrenalectomy. The above events elicited by TMT were almost entirely prevented by treatment with dexamethasone. In addition to the above events, adrenalectomy clearly enhanced the activation of c-Jun-N-terminal kinases and the formation of 4-hydroxynonenal in the dentate gyrus following TMT treatment. The dentate granule cell damage induced by TMT was exacerbated by mifepristone, a glucocorticoid-receptor antagonist. Taken together, our data suggest that endogenous and exogenous glucocorticoids prevent neurodegeneration induced by TMT in the central nervous system by attenuating intensive oxidative stress and associated signaling pathways.

Abstract. The organotin trimethyltin (TMT) is known to cause neuronal degeneration in the central nervous system. A systemic injection of TMT produced neuronal damage in the cerebral frontal cortex of mice. To elucidate the mechanism(s) underlying the toxicity of TMT toward neurons, we prepared primary cultures of neurons from the cerebral cortex of mouse embryos for use in this study. Microscopic observations revealed that a continuous exposure to TMT produced neuronal damage with nuclear condensation in an incubation time-dependent manner up to 48 h. The neuronal damage induced by TMT was not blocked by N-methyl-D-aspartate receptor channel-blocker MK-801. The exposure to TMT produced an elevation of the phosphorylation level of c-Jun N-terminal kinase (JNK)p46 , but not JNK p54 , prior to neuronal death. Under the same conditions, a significant elevation was seen in the phosphorylation level of stress-activated protein kinase 1, which activates JNKs. Furthermore, TMT enhanced the expression and phosphorylation of c-Jun during a continuous exposure. The JNK inhibitor SP600125 was effective in significantly but only partially attenuating the TMT-induced nuclear condensation and accumulation of lactate dehydrogenase in the culture medium. Taken together, our data suggest that the neuronal damage induced by TMT was independent of excitotoxicity but that at least some of it was dependent on the JNK cascades in primary cultures of cortical neurons.

Abstract. Differential regulation of gene expression by transcription factors is widely viewed as one of the principal mechanisms guiding development. Although numerous DNA binding proteins have been identified in various tissues, the role of individual transcription factors in the differentiation of specific cell groups, such as those populating the inner ear, is just beginning to be elucidated. It is known that transcription factors are induced in response to many signals that lead to cell growth, differentiation, inflammatory responses, the regulation of apoptosis, and neoplastic transformation. There are various transcription factors in the cochlea of the inner ear. These include activator protein-1 and nuclear factor-kappa B, glucocorticoid receptor, and so on. Based on recent reports and our investigation, in this article we review possible functions and expression of these transcription factors.