Spinocerebellar ataxia (SCA) is an inherited neurodegenerative disorder that is characterized by cerebellar atrophy and progressive ataxia and is classified into 31 types by the genetic locus. Recently, missense mutations of PRKCG genes that code protein kinase Cγ (γPKC) have been identified as a causal gene of SCA14. To explore the molecular mechanism of SCA14 pathogenesis, we investigated how mutant γPKC causes the neurodegeneration of cerebellar Purkinje cells (PCs) by expressing mutant γPKC-GFP in cell lines and primary cultured PCs. Mutant γPKC was susceptible to aggregation in the cytoplasm, which led to an impairment of the ubiquitin–proteasome system and apoptosis. Furthermore, mutant γPKC induced improper dendritic development of cultured PCs in an aggregation-independent manner. Stimulation-induced translocation of mutant γPKC in PC dendrites was prominently attenuated by the reduced mobility of oligomerized mutant γPKC, which resulted in attenuated signal transduction and the improper morphology of PC dendrites. These findings suggested that the oligomerization and aggregation of mutant γPKC caused improper dendritic development and apoptosis of PCs, which led to cerebellar dysfunction and SCA14 pathogenesis. We screened the chemicals that improved these cellular dysfunctions and identified several compounds, including trehalose and Congo red, which could be novel therapeutics for SCA14.
The prevailing view is that L-glutamate (Glu) functions as an excitatory amino acid neurotransmitter through a number of molecular machineries required for the neurocrine signaling at synapses in the brain. These include Glu receptors for signal input, Glu transporters for signal termination, and vesicular Glu transporters for signal output through exocytotic release. Although relatively little attention has been paid to the functional expression of these molecules required for glutamatergic signaling in peripheral tissues, recent molecular biological analyses including ours give rise to a novel function for Glu as an extracellular signal mediator in the autocrine and/or paracrine system in several peripheral and non-neuronal tissues, including bone and cartilage. In particular, a drastic increase is demonstrated in the endogenous levels of both Glu and aspartate in the synovial fluid with intimate relevance to increased edema and sensitization to thermal hyperalgesia in experimental arthritis models. However, to date, there is only limited information about the physiological and pathological significance of glutamatergic signaling machineries expressed by articular synovial tissues. In this review, we have outlined the role of Glu in synovial fibroblasts in addition to the possible involvement of glutamatergic signaling machineries in the pathogenesis of joint diseases such as rheumatoid arthritis.
The present study was designed to explore the possible nitriergic influence and role of p38MAPK and NF-κB in the diminished anti-anxiety effect of diazepam in stressed mice, using the elevated plus maze and light/dark box to assess anxiety. Immobilization stress for 6 h enhanced an anxiety-like behavior and increased plasma nitrite levels in mice. Diazepam (2 mg/kg, i.p.) produced an anti-anxiety effect in unstressed mice, but could not produce any change in anxiety levels of stressed mice. SB-203580 (2 mg/kg, i.p.), a specific inhibitor of p38MAPK, per se produced a significant antianxiety-like activity in stressed mice. Administration of a combination of SB-203580 (2 mg/kg, i.p.) and diazepam (2 mg/kg) in stressed mice produced a significantly higher antianxiety-like activity than that produced by SB-203580 alone. Pyrrolidine dithiocarbamate (PDTC), an inhibitor of the activation of NF-κB, per se produced a significant antianxiety-like activity in stressed mice. Combination of PDTC and diazepam also served to produce a higher significant antianxiety-like activity in stressed mice than that produced by PDTC alone. Diazepam could not produce any change in plasma nitrite levels in both unstressed and stressed mice. Both SB-203580 (2 mg/kg, i.p.) and PDTC (100 mg/kg, i.p.) significantly decreased plasma nitrite levels in stressed mice. The observations indicate that the diminished anti-anxiety effect of diazepam in stressed mice may involve strong nitriergic influence and may further be p38MAPK- and NF-κB–dependent.
Stiffness and cytokine in blood levels show 24-h rhythms in rheumatoid arthritis (RA) patients. We previously revealed that higher therapeutic effects were obtained in RA patients and RA model animals when the dosing time of methotrexate was chosen according to the 24-h rhythms to cytokine. In this study, we examined whether a dosing time–dependency of the therapeutic effect of tacrolimus (TAC) could be detected in collagen-induced arthritis (CIA) and MRL/lpr mice. To measure the levels of cytokines and serum amyloid A (SAA), blood was collected from CIA mice at different times. TAC was administered at two different dosing times based on these findings and its effects on arthritis and toxicity were examined. Plasma tumor necrosis factor (TNF)-α, interleukin-6 (IL-6), and SAA concentrations showed obvious 24-h rhythms with higher levels during the light phase and lower levels during the dark phase after RA crisis. The arthritis score and leukocyte counts were significantly lower in the group treated at 2 h after the light was turned on (HALO) than in the control and 14 HALO–treated groups. Our findings suggest that choosing an optimal dosing time could lead to the effective treatment of RA by TAC.
We have reported that nicotine and the specific α7AChR agonist ameliorate indomethacin-induced intestinal lesions in mice by activating α7 nicotinic acetylcholine receptors (α7nAChR). Dopamine D2–receptor antagonists, such as domperidone and metoclopramide, enhance the release of ACh from vagal efferent nerves. The present study examined the effects of domperidone and metoclopramide on indomethacin-induced small intestinal ulceration in mice, focusing on the α7AChR. Male C57BL/6 mice were administered indomethacin (10 mg/kg, s.c.) and sacrificed 24 h later. Domperidone (0.1 – 10 mg/kg) and metoclopramide (0.03 – 0.3 mg/kg) were administered i.p. twice, at 0.5 h before and 8 h after indomethacin treatment, while methyllycaconitine (a selective antagonist of α7nAChR, 30 mg/kg) was administered twice, at 0.5 h before each domperidone treatment. Indomethacin caused severe hemorrhagic lesions in the small intestine, mostly to the jejunum and ileum, with a concomitant increase in myeloperoxidase (MPO) activity. Domperidone suppressed the severity of lesions and the increase in MPO activity at low doses (0.1 – 3 mg/kg), but not at a high dose (10 mg/kg). Similar effects were also observed by metoclopramide. The protective effects of domperidone and metoclopramide were totally abolished by prior administration of methyllycaconitine. Indomethacin treatment markedly enhanced inducible nitric oxide synthase and chemokine mRNA expression in the small intestine, but these responses were all significantly attenuated by either domperidone or metoclopramide. These findings suggest that dopamine D2–receptor antagonists ameliorate indomethacin-induced small intestinal ulceration through the activation of endogenous anti-inflammatory pathways mediated by α7nAChR.
Asperlin is a fungal metabolite isolated from Aspergillus sp. SF-5044. In the present study, we isolated asperlin from the marine-derived fungus Aspergillus sp. SF-5044 and demonstrated that it inhibited inducible nitric oxide synthase (iNOS) expression, reduced iNOS-derived NO, suppressed cyclooxygenase (COX)-2 expression, and reduced COX-derived prostaglandin (PG) E2 production in lipopolysaccharide (LPS)-stimulated RAW264.7 and murine peritoneal macrophages. Similarly, asperlin reduced the production of tumor necrosis factor (TNF)-α and interleukin (IL)-1β. In addition, asperlin inhibited the phosphorylation and degradation of IκB-α, as well as the nuclear translocation of p65 caused by the stimulation of LPS in RAW264.7 macrophages. Furthermore, asperlin induced heme oxygenase (HO)-1 expression through nuclear translocation of nuclear factor E2–related factor 2 and increased HO activity in RAW264.7 macrophages. The effects of asperlin on the LPS-induced expression of iNOS and COX-2 and production of NO, PGE2, TNF-α, and IL-1β were partially reversed by a HO-1 inhibitor, tin protoporphyrin. These findings suggest that asperlin-induced HO-1 expression plays a role in the anti-inflammatory effects of asperlin in macrophages.
Compelling evidence supports the notion that the majority of neurodegenerative diseases are associated with microglia-mediated neuroinflammation. Therefore, quelling of microglial activation may lead to neuronal cell survival. The present study investigated the effects of Kamebakaurin (KMBK), a kaurane diterpene isolated from Isodon japonicus HARA (Labiatae), on the production of pro-inflammatory mediators in lipopolysaccharide (LPS)-stimulated cytotoxicity in rat primary microglial cultures and the BV-2 cell line. KMBK significantly inhibited the LPS-induced production of nitric oxide (NO) in a concentration-dependent fashion in activated microglial cells. The mRNA and protein levels of inducible nitric oxide synthase (iNOS) and cyclooxycenase-2 (COX-2) were also decreased dose-dependently. Furthermore KMBK inhibited the JNK and p38 mitogen-activated protein kinases (MAPKs) in LPS-stimulated BV-2 microglial cells. Considering the results obtained, the present study authenticated the potential benefits of KMBK as a therapeutic target in ameliorating microglia-mediated neuroinflammatory diseases.
Irbesartan, an angiotensin-receptor blocker, is a known agonist of peroxisome proliferator-activated receptor (PPAR) γ. In this study, thirteen-week-old spontaneously hypertensive (SHR)/NDmcr-cp rats, representing a genetic model of metabolic syndrome, were treated daily with placebo, irbesartan (30 mg/kg), valsartan (10 mg/kg), or pioglitazone (10 mg/kg) for 4 weeks. Significant reductions in systolic blood pressure were seen in the irbesartan- and valsartan-treated groups, but not in the pioglitazone-treated group. Compared with the placebo group, plasma insulin, homeostasis model assessment of insulin resistance index, and plasma triglyceride levels were significantly lower while plasma adiponectin levels were significantly higher in the pioglitazone- and irbesartan-treated groups, but not in the valsartan-treated group. Significant increases in the gene expression of adiponectin and GLUT4 within adipose tissue were also observed in the pioglitazone- and irbesartan-treated groups, but not in the valsartan-treated group. These findings suggest that through PPARγ stimulation along with angiotensin II inhibition, irbesartan may be an optimal treatment option in the prevention of metabolic syndrome as well as hypertension.
In the present study, we investigated the effect of the Kampo medicine Yokukansan (YKS) on pentobarbital-induced sleep in group-housed and socially isolated mice. Socially isolated mice showed shorter sleeping time than the group-housed mice. YKS (300 mg/kg, p.o.) prolonged the pentobarbital-induced sleeping time in socially isolated mice without affecting pentobarbital sleep in group-housed mice. The prolongation of sleeping time by YKS was reversed by bicuculline (3 mg/kg, i.p.) and flumazenil (3 mg/kg, i.p.), but not WAY100635. These findings suggest that the GABAA – benzodiazepine receptor complex, but not 5-HT1A receptors, is involved in the reversal effect of YKS on the decrease of pentobarbital sleep by social isolation.