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Parkinson's disease (PD) is one of the most common neurodegenerative diseases. Mutation in the phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1) gene causes an autosomal recessive form of PD. However, the etiology related to PINK1 is still not clear. Here, we examined the effect of PINK1 on heme oxygenase (HO)-1 induction in SH-SY5Y neuronal cells following H2O2 or 1-methyl-4-phenylpyridinium (MPP+) treatment. The HO-1 induction in response to H2O2 and MPP+ treatment was impaired by the expression of recombinant PINK1 G309D mutant. PINK1 G309D mutation increased the apoptosis of SH-SY5Y cells following H2O2 treatment and cell survival was rescued by the over-expression of HO-1 using adenovirus (Ad) infection. In addition, knockdown of tumor necrosis factor receptor-associated protein-1 (TRAP1), which is the substrate of PINK1 kinase, in SH-SY5Y cells also inhibited the expression of HO-1 in response to oxidative stress. The up-regulation of TRAP1 expression following H2O2 treatment was inhibited by the expression of recombinant PINK1 G309D mutant. The H2O2-induced HO-1 induction was Akt- and ERKdependent. The phosphorylation of ERK and Akt but not p38 was inhibited in cells expressing the PINK1 G309D mutant and knockdown of TRAP1. These results indicate a novel pathway by which the defect of PINK1 inhibits the oxidative stress-induced HO-1 production. Impairment of HO-1 production following oxidative stress may accelerate the dopaminergic neurodegeneration in Parkinson patients with PINK1 defect. [ABSTRACT FROM AUTHOR]

Heme-oxygenase-1 (HO-1), an important enzyme involved in vascular disease, transplantation, and inflammation, catalyzes the degradation of heme into carbon monoxide and biliverdin. It has been reported that overexpression of HO-1 inhibits osteoclastogenesis. However, the effect of HO-1 on osteoblast differentiation is still not clear. We here used adenoviral vector expressing recombinant human HO-1 and HO-1 inducer hemin to study the effects of HO-1 in primary cultured osteoblasts. The results showed that induction of HO-1 inhibited the maturation of osteoblasts including mineralized bone nodule formation, alkaline phosphatase activity and decreased mRNA expression of several differentiation markers such as alkaline phosphatase, osteocalcin, and RUNX2. Furthermore, downstream products of HO-1, bilirubin, carbon monoxide, and iron, are involved in the inhibitory action of HO-1. HO-1 can be induced by H2O2, lipopolysaccharide and inflammatory cytokines such as TNF-α and IL-1β in osteoblasts and also in STZ-induced diabetic mice. In addition, endogenous PPARγ ligand, 15-deoxy-Δ12,14-prostaglandin-J2 (15d-PGJ2) markedly increased both mRNA and protein levels of HO-1 in osteoblasts via PI3K-Akt and MAPK pathways. Blockade of HO activity by ZnPP IX antagonized the inhibitory action on osteocalcin expression by hemin and 15d-PGJ2. Our results indicate that upregulation of HO-1 inhibits the maturation of osteoblasts and HO-1 may be involved in oxidative- or inflammation-induced bone loss. J. Cell. Physiol. 222: 757–768, 2010. © 2009 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Matrix metalloproteinase-13 (MMP-13) is involved in the degradation of extracellular matrix in many kinds of tissues. Here we found that hypoxia increased MMP-13 protein and mRNA levels in primary rat astrocyte cultures. Hypoxia stimulation also increased the secretion of MMP-13 from astrocytes, as shown by zymographic analysis. In addition, exposure to hypoxia up-regulated the expression of c-Fos and c-Jun time-dependently. Hypoxia-induced MMP-13 overexpression was antagonized by transfection with antisense oligodeoxynucleotides (AS-ODN) of c-Fos or c-Jun. Furthermore, hypoxic-conditioned medium (Hx-CM) collected from astrocytes exposed to hypoxia increased paracellular permeability of adult rat brain endothelial cells (ARBECs). Administration of MMP-13 neutralizing antibody antagonized Hx-CM-induced paracellular permeability of ARBECs. Furthermore, pre-transfection of astrocytes with AS-ODN of c-Fos, c-Jun or MMP-13-shRNA significantly decreased hyperpermeability of ARBECs induced by Hx-CM. The arrangement of tight junction protein (TJP) zonular occludens-1 (ZO-1) of ARBECs disorganized in response to Hx-CM. Administration of Hx-CM to ARBECs also resulted in the production of proteolytic fragments of ZO-1, which was antagonized by transfection of MMP-13-shRNA in primary astrocytes. Administration of MMP-13 recombinant protein to ARBECs led to the disorganization and fragmentation of ZO-1 protein and also increased paracellular permeability. These results suggest that hypoxia-induced MMP-13 expression in astrocytes is regulated by c-Fos and c-Jun. MMP-13 is an important factor leading to the disorganization of ZO-1 and hyperpermeablility of blood–brain barrier in response to hypoxia. J. Cell. Physiol. 220: 163–173, 2009. © 2009 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

The dentate gyrus (DG) is a gateway that regulates seizure activity in the hippocampus. We investigated the site of action of lamotrigine (LTG), an effective anticonvulsant, in the regulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and N-methyl-D-aspartic acid (NMDA) receptor-mediated excitatory synaptic transmission on DG. Evoked AMPA and NMDA receptor-mediated excitatory postsynaptic currents (eEPSCampa and eEPSCnmda ) were recorded by whole-cell patch-clamp recording from the granule cells of DG in brain slice preparation of young Wistar rats (60–120 g). Exogenously applied AMPA and NMDA-induced currents and AMPA receptor-mediated miniature EPSC (mEPSCampa ) were recorded in the presence of specific antagonists. LTG inhibited both eEPSCampa and eEPSCnmda , and had no effect on exogenously applied NMDA-induced current indicating LTG inhibited glutamate release. Previous studies demonstrated that alteration in glutamate concentration in synaptic cleft causes parallel changes of eEPSCampa and eEPSCnmda . Our results showed that LTG inhibited eEPSCampa significantly more than eEPSCnmda (p < 0.05), suggesting that LTG may also have blocked the postsynaptic AMPA receptor. The hypothesis is further supported by the facts that; (1) LTG (30–100 μM) inhibited direct exogenously applied AMPA-induced currents (to 90%), (2) LTG significantly reduced the amplitude, but not the frequency of mEPSCampa and asynchronous (EPSC), and (3) LTG-induced reduction of eEPSCampa was not associated with a modification of the paired-pulse ratio. To sum up, LTG exerts a postsynaptic inhibitory mechanism on the AMPA receptor. Our results demonstrate that LTG suppress es postsynaptic AMPA receptors and reduces glutamate release in granule cells of DG. The postsynaptic effect can be one of the underlying mechanisms of LTG's anticonvulsant action. [ABSTRACT FROM AUTHOR]

It has been shown that ultrasound (US) stimulation accelerates fracture healing, bone maturation, and remodeling in the animal models and in clinical studies. One of the major factor involves in remodeling process is matrix metalloproteinases (MMPs) such as MMP-13 that has been shown to degrade the native interstitial collagens in several tissues. Here we found that US stimulation increased the secretion of MMP-13 in cultured rat osteoblasts, as shown by zymographic analysis. US stimulation also increased the mRNA level of MMP-13, c-Fos, and c-Jun. Cycloheximide (an inhibitor of protein translocation) and actinomycin D (an inhibitor of gene transcription) did not inhibit the MMP-13, c-Fos, and c-Jun mRNA expression, suggesting that such expression does not require de novo protein synthesis and not change their stabilities. p38 inhibitor, SB203580 or JNK inhibitor, SP600125 but not ERK inhibitor, PD98059 attenuated the US-induced MMP-13, c-Fos, and c-Jun expression; these results were further substantiated by transfecting with the dominant negative mutants of p38 or JNK. The binding of c-Fos and c-Jun to the AP-1 element on the MMP-13 promoter and the enhancement of AP-1 luciferase activity was enhanced by US stimulation. Taken together, our results provide evidence that US stimulation increases MMP-13 expression through p38 and JNK signaling pathway to regulate bone remodeling. J. Cell. Physiol. 215: 356–365, 2008. © 2007 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Fibronectin (Fn) is involved in early stages of bone formation and basic fibroblast growth factor (bFGF) is an important factor regulating osteogenesis. bFGF increased Fn expression, which was attenuated by phosphatidylinositol phospholipase inhibitor (U73122), protein kinase C inhibitor (GF109203X), Src inhibitor (PP2), NF-κB inhibitor (PDTC), IκBα phosphorylation inhibitor (Bay 117082), or IκB protease inhibitor (TPCK). bFGF-induced increase of Fn-luciferase activity was antagonized by cells transfected with Fn construct without NF-κB regulatory site. Stimulation of osteoblasts with bFGF activated IκB kinase α/β (IKK α/β) and increased IκBα phosphorylation, IκBα degradation, p65 and p50 translocation from the cytosol to the nucleus, the formation of an NF-κB-specific DNA-protein complex and κB-luciferase activity. bFGF-mediated an increase of IKKα/β activity and DNA-binding activity was inhibited by U73122, GF109203X, or PP2. The binding of p65 to the NF-κB element, as well as the recruitment of p300 and the enhancement of p50 acetylation on the Fn promoter was enhanced by bFGF. Overexpression of constitutively active FGF receptor 2 (FGFR2) increased Fn-luciferase activity, which was inhibited by co-transfection with dominant negative (DN) mutants of PLCγ2, PKCα, c-Src, IKKα, or IKKβ. Our results suggest that bFGF increased Fn expression in rat osteoblasts via the FGFR2/PLCγ2/PKCα/c-Src/NF-κB signaling pathway. J. Cell. Physiol. 211: 45–55, 2007. © 2007 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

1 Although capsaicin analogs might be a potential strategy to manipulate inflammation, the mechanism is still unclear. In this study, the effects and action mechanisms of vanilloid analogs on iNOS and COX-2 expression were investigated in RAW264.7 macrophages. 2 Capsaicin and resiniferatoxin (RTX) can inhibit LPS- and IFN-γ-mediated NO production, and iNOS protein and m RNA expression with similar 1C[sub50] values of around 10 μM. 3 Capsaicin also transcriptionally inhibited LPS-and PMA-induced COX-2 expression and PGE[sub2] production. However, this effect exhibited a higher potency (1C[sub50] 0.2μM). and RTX failed to elicit such responses at 10μM. 4 Interestingly, we found that capsazepine, a competitive TRPVI antagonist, did not prevent the inhibition elicited by Capsaicin or RTX. Nevertheless, it mimicked vanilloids in inhibiting iNOS/NO and COX-2/PGE[sub2] induction with an IC[sub50] value of 3μM. RT-PCR and immunoblotting analysis excluded the expression of TRPVl in RAW264.7 macrophages. 5 The DNA binding assay demonstrated the abilities of vanilloids to inhibit LPS-elicited NF-[subk]B and AP-I activation and IFN-Y-elicited STAT1 activation. The reporter assay of AIM activity also supported this action. 6 The kinase assay indicated that ERK,JNK, and IKK activation by LPS were inhibited by vanilloids. 7 In conclusionvanilloids can modulate the expression of inflammatory iNOS and COX-2 genes in macrophages through interference with upstream signalling events of LPS and IFN-γ. These findings provide new insights into the potential benefits of the active ingredient in hot chilli peppers in inflammatory conditions. [ABSTRACT FROM AUTHOR]