Linalool, a Fragrance Compound in Plants, Protects Dopaminergic Neurons and Improves Motor Function and Skeletal Muscle Strength in Experimental Models of Parkinson's Disease.

Parkinson's disease (PD) is a common neurodegenerative disorder characterized by the gradual loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc), resulting in reduced dopamine levels in the striatum and eventual onset of motor symptoms. Linalool (3,7-dimethyl-1,6-octadien-3-ol) is a monoterpene in aromatic plants exhibiting antioxidant, antidepressant, and anti-anxiety properties. The objective of this study is to evaluate the neuroprotective impacts of linalool on dopaminergic SH-SY5Y cells, primary mesencephalic and cortical neurons treated with 1-methyl-4-phenylpyridinium ion (MPP+), as well as in PD-like mice induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Cell viability, α-tubulin staining, western blotting, immunohistochemistry and behavioral experiments were performed. In MPP+-treated SH-SY5Y cells, linalool increased cell viability, reduced neurite retraction, enhanced antioxidant defense by downregulation of apoptosis signaling (B-cell lymphoma 2 (Bcl-2), cleaved caspase-3 and poly ADP-ribose polymerase (PARP)) and phagocyte NADPH oxidase (gp91phox), as well as upregulation of neurotrophic signaling (brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF)) and nuclear factor-erythroid 2 related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) pathway. In MPP+-treated primary mesencephalic neurons, linalool enhanced the expressions of tyrosine hydroxylase (TH), Sirtuin 1 (SirT1), and parkin. In MPP+-treated primary cortical neurons, linalool upregulated protein expression of SirT1, γ-Aminobutyric acid type A-α1 (GABAA-α1), and γ-Aminobutyric acid type B (GABAB). In PD-like mice, linalool attenuated the loss of dopamine neurons in SNpc. Linalool improved the motor and nonmotor behavioral deficits and muscle strength of PD-like mice. These findings suggest that linalool potentially protects dopaminergic neurons and improves the impairment symptoms of PD.

Guilu Erxian Jiao enhances protein synthesis, glucose homeostasis, mitochondrial biogenesis and slow-twitch fibers in the skeletal muscle.

Guilu Erxian Jiao (GEJ) is a commonly used nutritional supplement due to its rich content of amino acids. It is also a traditional herbal medicine for improving degenerative joint. This study aimed to investigate the effect and mechanism of GEJ water extract (GEJ-WE) on skeletal muscle in C2C12 myotubes and C57BL/6J mice. Analysis of GEJ-WE were performed by high-performance liquid chromatography fingerprinting with chemical standards. Protein expression, mRNA level, glycogen content, mitochondria activity and ATP level were evaluated by western blots, real-time PCR, PAS staining, MTT and ATP bioluminescence assay, respectively. Skeletal muscle strength was evaluated by grip strength. Skeletal muscle volume, mass and fiber types were evaluated by micro computed tomography, histological analysis and immunofluorescence staining, respectively. Motor function was evaluated by rotarod performance and locomotor activity. In C2C12 myotubes, GEJ-WE significantly enhanced myogenic differentiation and myotube growth, protein synthesis signaling IGF-1/IGF-1R/IRS-1/Akt, Glut4 translocation, glycogen content, mitochondrial biogenesis signaling PGC-1α/NRF1/TFAM, mitochondrial activity and ATP production. However, IGF-1R antagonist AG1024 and PI3K inhibitor wortmannin reduced GEJ-WE-induced protein expression of MyHC, p-Akt, p-mTOR and p-GSK-3ß, Glut4 translocation and glycogen content. In C57BL/6J mice, GEJ-WE not only upregulated protein synthesis and mitochondrial biogenesis signaling, but it also increased muscle volume, relative muscle weight, cross-sectional area of myofibers, glycogen content and transition of fast-to-slow type fibers of skeletal muscles. Moreover, GEJ-WE enhanced grip strength and motor activity of mice. In conclusion, the upregulation of protein synthesis, myogenic differentiation, glucose homeostasis, mitochondrial biogenesis and slow-twitch fibers contributes to the mechanisms of GEJ-WE on the enhancement of skeletal muscle mass and motor function.

An efficient cellular image-based platform for high-content screening of neuroprotective agents against chemotherapy-induced neuropathy.

Chemotherapy-induced peripheral neuropathy (CIPN) is a major dose-limiting side effect, with no approved therapy for prevention or treatment. Here, we aimed to establish a high-content image platform based on the neurite outgrowth of dorsal root ganglia (DRG)-derived neuron cells for the discovery of neuroprotective agents against paclitaxel-induced CIPN. ND7/23 cells, an immortalized hybrid DRG cell line, were maturely differentiated by induction with nerve growth factor and upregulation of intracellular cAMP levels. High-content image analyses of the neurofilament-stained neurite network showed that paclitaxel disrupted the neurite outgrowth of well-differentiated ND7/23 DRG neuron cells, recapitulating characteristic effects of paclitaxel on primary cultured DRG neurons. This process coincided with the upregulated activity of store-operated Ca2+ entry, similar to those found in rodent models of paclitaxel-induced CIPN. The previously identified neuroprotective agents, minoxidil and 8-Br-cyclic adenosine monophosphate ribose (8-Br-cADPR), attenuated the reduction in total neurite outgrowth in paclitaxel-damaged ND7/23 cells. Additionally, the total neurite outgrowth of well-differentiated ND7/23 cells was concentration-dependently reduced by the neurotoxic chemotherapeutic agents, oxaliplatin and bortezomib, but not the less neurotoxic 5-fluorouracil. We demonstrated that high-content analyses of neurite morphology in well-differentiated DRG neuron-derived cells provide an effective, reproducible, and high-throughput strategy for developing therapeutics against CIPN.

Prevalence and Outcomes of Major Psychiatric Disorders Preceding Index Surgery for Degenerative Thoracic/Lumbar Spine Disease.

The relationship between preexisting major psychiatric disorders and outcomes of spine surgery for degenerative thoracic/lumbar disease remains unclear. A 5% subset of inpatients was randomly selected from the Taiwan National Health Insurance Research Database. A total of 10,109 inpatients aged 18 years or over with degenerative thoracic/lumbar disease and underwent spine surgery met inclusion criteria. Major psychiatric disorders diagnosed by psychiatrists preceding index surgery, including anxiety disorder, depression disorder, bipolar disorder, schizophrenia and dementia, were identified. The prevalence of psychiatric disorders, and their differential risks on in-hospital and post-discharge outcomes were examined. 10.4% had major psychiatric disorders, of which depression (6.6%) and anxiety (4.9%) were most common. Logistic regression revealed increased risks of ventilator use in depression (OR = 1.62, 95% CI = 1.04-2.54, p < 0.05), extended hospitalization length in bipolar (OR = 1.77, 95% CI = 1.08-2.89, p < 0.05), and higher rehabilitation utilization in depression (OR = 1.25, 95% CI = 1.06-1.47, p < 0.01) and bipolar (OR = 1.69, 95% CI = 1.04-2.76, p < 0.05). Those patients with anxiety had a decreased risk of longer hospitalization duration (OR = 0.77, 95% CI = 0.60-0.98, p < 0.05), while those with dementia and schizophrenia had no change in risks. Preoperative recognition of major psychiatric disorders for risk and treatment assessment is suggested as people with preexisting depression or bipolar disorder have worse outcomes after spine surgery.

Triptolide prevents LPS-induced skeletal muscle atrophy via inhibiting NF-κB/TNF-α and regulating protein synthesis/degradation pathway.

BACKGROUND AND PURPOSE: Increasing evidence suggests systemic inflammation-caused skeletal muscle atrophy as a major clinical feature of cachexia. Triptolide obtained from Tripterygium wilfordii Hook F possesses potent anti-inflammatory and immunosuppressive effects. The present study aims to evaluate the protective effects and molecular mechanisms of triptolide on inflammation-induced skeletal muscle atrophy. EXPERIMENTAL APPROACH: The effects of triptolide on skeletal muscle atrophy were investigated in LPS-treated C2C12 myotubes and C57BL/6 mice. Protein expressions and mRNA levels were analysed by western blot and qPCR, respectively. Skeletal muscle mass, volume and strength were measured by histological analysis, micro-CT and grip strength, respectively. Locomotor activity was measured using the open field test. KEY RESULTS: Triptolide (10-100 fM) up-regulated protein synthesis signals (IGF-1/p-IGF-1R/IRS-1/p-Akt/p-mTOR) and down-regulated protein degradation signal atrogin-1 in C2C12 myotubes. In LPS (100 ng·ml-1 )-treated C2C12 myotubes, triptolide up-regulated MyHC, IGF-1, p-IGF-1R, IRS-1 and p-Akt. Triptolide also down-regulated ubiquitin-proteasome molecules (n-FoxO3a/atrogin-1/MuRF1), proteasome activity, autophagy-lysosomal molecules (LC3-II/LC3-I and Bnip3) and inflammatory mediators (NF-κB, Cox-2, NLRP3, IL-1ß and TNF-α). However, AG1024, an IGF-1R inhibitor, suppressed triptolide-mediated effects on MyHC, myotube diameter, MuRF1 and p62 in LPS-treated C2C12 myotubes. In LPS (1 mg·kg-1 , i.p.)-challenged mice, triptolide (5 and 20 µg·kg-1 ·day-1 , i.p.) decreased plasma TNF-α levels and it increased skeletal muscle volume, cross-sectional area of myofibers, weights of the gastrocnemius and tibialis anterior muscles, forelimb grip strength and locomotion. CONCLUSIONS AND IMPLICATIONS: These findings reveal that triptolide prevented LPS-induced inflammation and skeletal muscle atrophy and have implications for the discovery of novel agents for preventing muscle wasting.

Effective Activation by Kynurenic Acid and Its Aminoalkylated Derivatives on M-Type K+ Current.

Kynurenic acid (KYNA, 4-oxoquinoline-2-carboxylic acid), an intermediate of the tryptophan metabolism, has been recognized to exert different neuroactive actions; however, the need of how it or its aminoalkylated amide derivative N-(2-(dimethylamino)ethyl)-3-(morpholinomethyl)-4-oxo-1,4-dihydroquinoline-2-carboxamide (KYNA-A4) exerts any effects on ion currents in excitable cells remains largely unmet. In this study, the investigations of how KYNA and other structurally similar KYNA derivatives have any adjustments on different ionic currents in pituitary GH3 cells and hippocampal mHippoE-14 neurons were performed by patch-clamp technique. KYNA or KYNA-A4 increased the amplitude of M-type K+ current (IK(M)) and concomitantly enhanced the activation time course of the current. The EC50 value required for KYNA- or KYNA-A4 -stimulated IK(M) was yielded to be 18.1 or 6.4 µM, respectively. The presence of KYNA or KYNA-A4 shifted the relationship of normalized IK(M)-conductance versus membrane potential to more depolarized potential with no change in the gating charge of the current. The voltage-dependent hysteretic area of IK(M) elicited by long-lasting triangular ramp pulse was observed in GH3 cells and that was increased during exposure to KYNA or KYNA-A4. In cell-attached current recordings, addition of KYNA raised the open probability of M-type K+ channels, along with increased mean open time of the channel. Cell exposure to KYNA or KYNA-A4 mildly inhibited delayed-rectifying K+ current; however, neither erg-mediated K+ current, hyperpolarization-activated cation current, nor voltage-gated Na+ current in GH3 cells was changed by KYNA or KYNA-A4. Under whole-cell, current-clamp recordings, exposure to KYNA or KYNA-A4 diminished the frequency of spontaneous action potentials; moreover, their reduction in firing frequency was attenuated by linopirdine, yet not by iberiotoxin or apamin. In hippocampal mHippoE-14 neurons, the addition of KYNA also increased the IK(M) amplitude effectively. Taken together, the actions presented herein would be one of the noticeable mechanisms through which they modulate functional activities of excitable cells occurring in vivo.

Naringenin, a dietary flavanone, enhances insulin-like growth factor 1 receptor-mediated antioxidant defense and attenuates methylglyoxal-induced neurite damage and apoptotic death.

Objectives: Recent studies revealed the neuroprotective effects of naringenin (NGEN), a common dietary bioflavonoid contained in citrus fruits. However, there are limited data on its protection against methylglyoxal (MG), the most potent precursor of advanced glycation end-products. The present study was to investigate the protection of NGEN on MG-induced neurotoxicity and the involvement of insulin-like growth factor 1 receptor (IGF-1R) signaling. Methods: NSC34 motor neuron-like cells was used. Cell viability was measured by MTT assay. Protein expressions were analyzed by western blots. Morphological changes of neurites were observed by an inverted microscope. Reactive oxygen species (ROS) production and apoptotic cell numbers were measured by flow cytometer. Glutathione (GSH) level and superoxide dismutase (SOD) activity were measured by ELISA. Results: >NGEN attenuated ROS production and increased GSH level, SOD activity and nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear expression in MG-treated NSC34 cells. NGEN also increased neurite length and enhanced IGF-1R and p-Akt in MG-treated NSC34 cells. Furthermore, NGEN attenuated MG-induced apoptotic death accompanied with down-regulation of cleaved-poly (ADP-ribose) polymerase (PARP) and up-regulation of B-cell lymphoma-2 (Bcl-2). However, AG1024, an IGF-1R antagonist, attenuated the anti-oxidative and anti-apoptotic effects of NGEN in MG-treated cells. Discussion: The present results demonstrated that NGEN decreased neuronal apoptosis and improved antioxidant defense in MG-treated NSC34 cells. Moreover, IGF-1R-mediated antioxidant defense plays an important role in this protective mechanism. These findings suggest the potential benefits of NGEN on the prevention of MG-induced or diabetes/hyperglycemia-related neurotoxicity. In vivo studies are needed for further confirmation on NGEN-mediated neuroprotection.

The Antioxidant, Anti-Inflammatory, and Neuroprotective Properties of the Synthetic Chalcone Derivative AN07.

Chalcones belong to a class of biologically active polyphenolic natural products. As a result of their simple chemical nature, they are easily synthesized and show a variety of promising biological activities. 2-Hydroxy-4'-methoxychalcone (AN07) is a synthetic chalcone derivate with potential anti-atherosclerosis effects. In this study, we demonstrated the novel antioxidant, anti-inflammatory, and neuroprotective effects of AN07. In RAW 264.7 macrophages, AN07 attenuated lipopolysaccharide (LPS)-induced elevations in reactive oxygen species (ROS) level and oxidative stress via down-regulating gp91phox expression and stimulating the antioxidant system of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) pathways, which were accompanied by increased glutathione (GSH) levels. Additionally, AN07 attenuated LPS-induced inflammatory factors, including NO, inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and phosphorylated inhibitor of nuclear factor kappa B-alpha (p-IκBα) in RAW 264.7 macrophages. However, the effects of AN07 on promoting nuclear Nrf2 levels and decreasing COX-2 expressions were significantly abrogated by the peroxisome proliferator-activated receptor-γ (PPARγ) antagonist GW9662. In human dopaminergic SH-SY5Y cells treated with or without methylglyoxal (MG), a toxic endogenous by-product of glycolysis, AN07 up-regulated neurotrophic signals including insulin-like growth factor 1 receptor (IGF-1R), p-Akt, p-GSK3ß, glucagon-like peptide 1 receptor (GLP-1R), and brain-derived neurotrophic factor (BDNF). AN07 attenuated MG-induced apoptosis by up-regulating the B-cell lymphoma 2 (Bcl-2) protein and down-regulating the cytosolic expression of cytochrome c. AN07 also attenuated MG-induced neurite damage via down-regulating the Rho-associated protein kinase 2 (ROCK2)/phosphorylated LIM kinase 1 (p-LIMK1) pathway. Moreover, AN07 ameliorated the MG-induced down-regulation of neuroprotective Parkinsonism-associated proteins parkin, pink1, and DJ-1. These findings suggest that AN07 possesses the potentials to be an anti-inflammatory, antioxidant, and neuroprotective agent.

Characterization of Inhibitory Effectiveness in Hyperpolarization-Activated Cation Currents by a Group of ent-Kaurane-Type Diterpenoids from Croton tonkinensis.

Croton is an extensive flowering plant genus in the spurge family, Euphorbiaceae. Three croton compounds with the common ent-kaurane skeleton have been purified from Croton tonkinensis. METHODS: We examined any modifications of croton components (i.e., croton-01 [ent-18-acetoxy-7α-hydroxykaur-16-en-15-one], croton-02 [ent-7α,14ß-dihydroxykaur-16-en-15-one] and croton-03 [ent-1ß-acetoxy-7α,14ß-dihydroxykaur-16-en-15-one] on either hyperpolarization-activated cation current (Ih) or erg-mediated K+ current identified in pituitary tumor (GH3) cells and in rat insulin-secreting (INS-1) cells via patch-clamp methods. RESULTS: Addition of croton-01, croton-02, or croton-03 effectively and differentially depressed Ih amplitude. Croton-03 (3 µM) shifted the activation curve of Ih to a more negative potential by approximately 11 mV. The voltage-dependent hysteresis of Ih was also diminished by croton-03 administration. Croton-03-induced depression of Ih could not be attenuated by SQ-22536 (10 µM), an inhibitor of adenylate cyclase, but indeed reversed by oxaliplatin (10 µM). The Ih in INS-1 cells was also depressed effectively by croton-03. CONCLUSION: Our study highlights the evidence that these ent-kaurane diterpenoids might conceivably perturb these ionic currents through which they have high influence on the functional activities of endocrine or neuroendocrine cells.

Characterization of Convergent Suppression by UCL-2077 (3-(Triphenylmethylaminomethyl)pyridine), Known to Inhibit Slow Afterhyperpolarization, of erg-Mediated Potassium Currents and Intermediate-Conductance Calcium-Activated Potassium Channels.

UCL-2077 (triphenylmethylaminomethyl)pyridine) was previously reported to suppress slow afterhyperpolarization in neurons. However, the information with respect to the effects of UCL-2077 on ionic currents is quite scarce. The addition of UCL-2077 decreased the amplitude of erg-mediated K+ current (IK(erg)) together with an increased deactivation rate of the current in pituitary GH3 cells. The IC50 and KD values of UCL-2077-induced inhibition of IK(erg) were 4.7 and 5.1 µM, respectively. UCL-2077 (10 µM) distinctly shifted the midpoint in the activation curve of IK(erg) to less hyperpolarizing potentials by 17 mV. Its presence decreased the degree of voltage hysteresis for IK(erg) elicitation by long-lasting triangular ramp pulse. It also diminished the probability of the opening of intermediate-conductance Ca2+-activated K+ channels. In cell-attached current recordings, UCL-2077 raised the frequency of action currents. When KCNH2 mRNA was knocked down, a UCL-2077-mediated increase in AC firing was attenuated. Collectively, the actions elaborated herein conceivably contribute to the perturbating effects of this compound on electrical behaviors of excitable cells.

Characterization in Dual Activation by Oxaliplatin, a Platinum-Based Chemotherapeutic Agent of Hyperpolarization-Activated Cation and Electroporation-Induced Currents.

Oxaliplatin (OXAL) is regarded as a platinum-based anti-neoplastic agent. However, its perturbations on membrane ionic currents in neurons and neuroendocrine or endocrine cells are largely unclear, though peripheral neuropathy has been noted during its long-term administration. In this study, we investigated how the presence of OXAL and other related compounds can interact with two types of inward currents; namely, hyperpolarization-activated cation current (Ih) and membrane electroporation-induced current (IMEP). OXAL increased the amplitude or activation rate constant of Ih in a concentration-dependent manner with effective EC50 or KD values of 3.2 or 6.4 µM, respectively, in pituitary GH3 cells. The stimulation by this agent of Ih could be attenuated by subsequent addition of ivabradine, protopine, or dexmedetomidine. Cell exposure to OXAL (3 µM) resulted in an approximately 11 mV rightward shift in Ih activation along the voltage axis with minimal changes in the gating charge of the curve. The exposure to OXAL also effected an elevation in area of the voltage-dependent hysteresis elicited by long-lasting triangular ramp. Additionally, its application resulted in an increase in the amplitude of IMEP elicited by large hyperpolarization in GH3 cells with an EC50 value of 1.3 µM. However, in the continued presence of OXAL, further addition of ivabradine, protopine, or dexmedetomidine always resulted in failure to attenuate the OXAL-induced increase of IMEP amplitude effectively. Averaged current-voltage relation of membrane electroporation-induced current (IMEP) was altered in the presence of OXAL. In pituitary R1220 cells, OXAL-stimulated Ih remained effective. In Rolf B1.T olfactory sensory neurons, this agent was also observed to increase IMEP in a concentration-dependent manner. In light of the findings from this study, OXAL-mediated increases of Ih and IMEP may coincide and then synergistically act to increase the amplitude of inward currents, raising the membrane excitability of electrically excitable cells, if similar in vivo findings occur.

Characterization of the Inhibitory Effect of Gastrodigenin and Gastrodin on M-type K+ Currents in Pituitary Cells and Hippocampal Neurons.

Gastrodigenin (HBA) and gastrodin (GAS) are phenolic ingredients found in Gastrodia elata Blume (GEB), a traditional Chinese herbal medicine. These compounds have been previously used to treat cognitive dysfunction, convulsion, and dizziness. However, at present, there is no available information regarding their potential ionic effects in electrically excitable cells. In the current study, the possible effects of HBA and GAS on different ionic currents in pituitary GH3 cells and hippocampal mHippoE-14 neurons were investigated using the patch-clamp technique. The addition of HBA or GAS resulted in the differential inhibition of the M-type K+ current (IK(M)) density in a concentration-dependent manner in GH3 cells. HBA resulted in a slowing of the activation time course of IK(M), while GAS elevated it. HBA also mildly suppressed the density of erg-mediated or the delayed-rectifier K+ current in GH3 cells. Neither GAS nor HBA (10 µM) modified the voltage-gated Na+ current density, although they suppressed the L-type Ca2+ current density at the same concentration. In hippocampal mHippoE-14 neurons, HBA was effective at inhibiting IK(M) density as well as slowing the activation time course. Taken together, the present study provided the first evidence that HBA or GAS could act on cellular mechanisms, and could therefore potentially have a functional influence in various neurologic disorders.

Evidence for the Capability of Roxadustat (FG-4592), an Oral HIF Prolyl-Hydroxylase Inhibitor, to Perturb Membrane Ionic Currents: An Unidentified yet Important Action.

Roxadustat (FG-4592), an analog of 2-oxoglutarate, is an orally-administered, heterocyclic small molecule known to be an inhibitor of hypoxia inducible factor (HIF) prolyl hydroxylase. However, none of the studies have thus far thoroughly investigated its possible perturbations on membrane ion currents in endocrine or heart cells. In our studies, the whole-cell current recordings of the patch-clamp technique showed that the presence of roxadustat effectively and differentially suppressed the peak and late components of IK(DR) amplitude in response to membrane depolarization in pituitary tumor (GH3) cells with an IC50 value of 5.71 and 1.32 µM, respectively. The current inactivation of IK(DR) elicited by 10-sec membrane depolarization became raised in the presence of roxadustatt. When cells were exposed to either CoCl2 or deferoxamine (DFO), the IK(DR) elicited by membrane depolarization was not modified; however, nonactin, a K+-selective ionophore, in continued presence of roxadustat, attenuated roxadustat-mediated inhibition of the amplitude. The steady-state inactivation of IK(DR) could be constructed in the presence of roxadustat. Recovery of IK(DR) block by roxadustat (3 and 10 µM) could be fitted by a single exponential with 382 and 523 msec, respectively. The roxadustat addition slightly suppressed erg-mediated K+ or hyperpolarization-activated cation currents. This drug also decreased the peak amplitude of voltage-gated Na+ current with a slowing in inactivation rate of the current. Likewise, in H9c2 heart-derived cells, the addition of roxadustat suppressed IK(DR) amplitude in combination with the shortening in inactivation time course of the current. In high glucose-treated H9c2 cells, roxadustat-mediated inhibition of IK(DR) remained unchanged. Collectively, despite its suppression of HIF prolyl hydroxylase, inhibitory actions of roxadustat on different types of ionic currents possibly in a non-genomic fashion might provide another yet unidentified mechanism through which cellular functions are seriously perturbed, if similar findings occur in vivo.

Evidence for Effective Inhibitory Actions on Hyperpolarization-Activated Cation Current Caused by Ganoderma Triterpenoids, the Main Active Constitutents of Ganoderma Spores.

The triterpenoid fraction of Ganoderma (Ganoderma triterpenoids, GTs) has been increasingly demonstrated to provide effective antioxidant, neuroprotective or cardioprotective activities. However, whether GTs is capable of perturbing the transmembrane ionic currents existing in electrically excitable cells is not thoroughly investigated. In this study, an attempt was made to study whether GTs could modify hyperpolarization-activated cation currents (Ih) in pituitary tumor (GH3) cells and in HL-1 atrial cardiomyocytes. In whole-cell current recordings, the addition of GTs produced a dose-dependent reduction in the amplitude of Ih in GH3 cells with an IC50 value of 11.7 µg/mL, in combination with a lengthening in activation time constant of the current. GTs (10 µg/mL) also caused a conceivable shift in the steady-state activation curve of Ih along the voltage axis to a more negative potential by approximately 11 mV. Subsequent addition of neither 8-cyclopentyl-1,3-dipropylxanthine nor 8-(p-sulfophenyl)theophylline, still in the presence of GTs, could attenuate GTs-mediated inhibition of Ih. In current-clamp voltage recordings, GTs diminished the firing frequency of spontaneous action potentials in GH3 cells, and it also decreased the amplitude of sag potential in response to hyperpolarizing current stimuli. In murine HL-1 cardiomyocytes, the GTs addition also suppressed the amplitude of Ih effectively. In DPCPX (1 µM)-treated HL-1 cells, the inhibitory effect of GTs on Ih remained efficacious. Collectively, the inhibition of Ih caused by GTs is independent of its possible binding to adenosine receptors and it might have profound influence in electrical behaviors of different types of electrically excitable cells (e.g., pituitary and heart cells) if similar in vitro or in vivo findings occur.

2-Iodo-4'-Methoxychalcone Attenuates Methylglyoxal-Induced Neurotoxicity by Activation of GLP-1 Receptor and Enhancement of Neurotrophic Signal, Antioxidant Defense and Glyoxalase Pathway.

Methylglyoxal (MG) acts as a reactive precursor of advanced glycation end products (AGEs). This compound is often connected with pathologies such as diabetes, neurodegenerative processes and diseases of aging. 2-iodo-4'-methoxychalcone (CHA79), a synthetic halogen-containing chalcone derivative, has been reported its anti-diabetic activity. This study aims to investigate the potential protective capability of CHA79 against MG-mediated neurotoxicity in SH-SY5Y cells. Results indicated CHA79 increased viability of cells and attenuated the rate of apoptosis in MG-exposed SH-SY5Y. CHA79 up-regulated expression of anti-apoptotic protein (Bcl-2) and down-regulated apoptotic proteins (Bax, cytochrome c, caspase-3, caspase-9). Moreover, CHA79 significantly up-regulated expression of neurotrophic factors, including glucagon-like peptide-1 receptor (GLP-1R), brain derived neurotrophic factor (BDNF), p75NTR, p-TrkB, p-Akt, p-GK-3ß and p-CREB. CHA79 attenuated MG-induced ROS production and enhanced the antioxidant defense including nuclear factor erythroid 2-related factor 2 (Nrf2), HO-1, SOD and GSH. Furthermore, CHA79 attenuated MG-induced reduction of glyoxalase-1 (GLO-1), a vital enzyme on removing AGE precursors. In conclusion, CHA79 is the first novel synthetic chalcone possessing the GLP-1R and GLO-1 activating properties. CHA 79 also exhibits neuroprotective effects against MG toxicity by enhancing neurotrophic signal, antioxidant defense and anti-apoptosis pathway.

The novel protective effects of loganin against 1-methyl-4-phenylpyridinium-induced neurotoxicity: Enhancement of neurotrophic signaling, activation of IGF-1R/GLP-1R, and inhibition of RhoA/ROCK pathway.

Loganin, a major iridoid glycoside obtained from fruits of Cornus officinalis, possesses anti-inflammatory, antitumor, antidiabetic, and osteoporosis prevention effects. Loganin has been linked to neuroprotection in several models of neurodegeneration, including Parkinson's disease (PD). However, mechanisms underlying the neuroprotective effects of loganin are still mostly unknown. Here, we demonstrated the protective effects of loganin against PD mimetic toxin 1-methyl-4-phenylpyridinium (MPP+ ) and the important roles of insulin-like growth factor 1 receptor (IGF-1R) and glucagon-like peptide 1 receptor (GLP-1R) in the neuroprotective mechanisms of loganin. In primary mesencephalic neuronal cultures treated with or without MPP+ , loganin up-regulated expressions of neurotrophic signals including IGF-1R, GLP-1R, p-Akt, BDNF, and tyrosine hydroxylase. Loganin protected against MPP+ -induced apoptosis by up-regulating antiapoptotic protein and down-regulating proapoptotic protein. Moreover, loganin attenuated MPP+ -induced neurite damage via up-regulation of GAP43 and down-regulation of membrane-RhoA/ROCK2/p-LIMK/p-cofilin. Loganin also attenuated MPP+ -induced reactive oxygen species (ROS) production. However, both AG1024, an IGF-1R antagonist, and exendin 9-39, a GLP-1R antagonist, attenuated the protective effects of loganin on MPP+ -induced cytotoxicity, apoptosis, neurite length decrease, and ROS production. Our results suggest that loganin attenuates MPP+ -induced apoptotic death, neurite damage, and oxidative stress through enhancement of neurotrophic signaling, activation of IGF-1R/GLP-1R, and inhibition of RhoA/ROCK pathway, providing the evidence that loganin possesses novel neuroprotective effects.

Resveratrol prevents nanoparticles-induced inflammation and oxidative stress via downregulation of PKC-α and NADPH oxidase in lung epithelial A549 cells.

BACKGROUND: Exposure to carbon black nanoparticles (CBNPs), a well-known industrial production, promotes pulmonary toxicity through inflammation and oxidative stress. Recent studies show that some polyphenols exert their antioxidant properties through regulation of protein kinase C-α (PKC-α) and NADPH oxidase (Nox) signaling. Resveratrol, a dietary polyphenol in fruits, possesses various health beneficial effects including anti-inflammatory and antioxidative properties. In this study, we aimed to elucidate the involvement of PKC-α and Nox in CBNPs-induced inflammation and oxidative stress, and to investigate the protective effects of resveratrol on CBNP-induced inflammation and oxidative stress in human lung epithelial A549 cells. METHODS: The production of reactive oxygen species (ROS) and the change of mitochondrial membrane potential (ΔΨm) were measured by flow cytometry. Nitric oxide (NO) was measured using the Griess reagent, and prostaglandin E2 (PGE2) production was detected by ELISA, while protein expressions were measured by Western blotting analysis. RESULTS: In lung epithelial A549 cells, CBNPs significantly enhanced oxidative stress by upregulation of Nox2 and membrane expression of p67phox accompanied with increase of ROS production. CBNPs also increased inflammatory factors, including iNOS, COX-2, NO and PGE2. However, resveratrol attenuated the above effects induced by CBNPs in A549 cells; additionally, CBNPs-induced activation of PKC-α was observed. We found that PKC-α inhibitor (Gö6976) could attenuate CBNPs-induced inflammation by down-regulation of ROS, NO and PGE2 production in A549 cells, suggesting PKC-α might be involved in CBNPs-induced oxidative stress and inflammation. Our results also found resveratrol was able to inhibit protein expression of PKC-α induced by CBNPs. Moreover, ROS scavenger (NAC) and Nox inhibitor (DPI) attenuated CBNPs-induced expressions of iNOS and COX-2. DPI could also attenuate CBNPs-induced ROS, NO and PGE2 production. CONCLUSIONS: Resveratrol attenuated CBNPs-induced oxidative and inflammatory factors in lung epithelial A549 cells, at least in part via inhibiting PKC-α- and Nox-related signaling.

Multiple regulatory actions of 2-guanidine-4-methylquinazoline (GMQ), an agonist of acid-sensing ion channel type 3, on ionic currents in pituitary GH3 cells and in olfactory sensory (Rolf B1.T) neurons.

GMQ (2-guanidine-4-methylquinazoline or N-(4-methyl-2-quinazolinyl)-guanidine hydrochloride), an agonist of acid-sensing ion channel type 3, has been increasingly used for in vivo studies of alternations in nociceptic behavior. In this study, we tried to investigate whether GMQ has any possible effect on other types of ion channels. Addition of GMQ to pituitary GH3 cells raised the amplitude of Ca2+-activated K+ currents (IK(Ca)), which was reversed by verruculogen or PF1022A, but not by TRAM-39. Under inside-out current recordings, addition of GMQ into bath enhanced the probability of large-conductance Ca2+-activated K+ (BKCa) channels with an EC50 value of 0.95 µM. The activation curve of BKCa channels during exposure to GMQ shifted to a lower depolarized potential, with no change in the gating charge of the curve; however, there was a reduction of free energy for channel activation in its presence. As cells were exposed to GMQ, the amplitude of ion currents were suppressed, including delayed rectifying K+ current, voltage-gated Na+ and L-type Ca2+ currents. In Rolf B1.T olfactory sensory neuron, addition of GMQ was able to induce inward current and to suppress peak INa. Taken together, findings from these results indicated that in addition to the activation of ASIC3 channels, this compound might directly produce additional actions on various types of ion channels. Caution should be taken in the interpretation of in vivo experimental results when GMQ or other structurally similar compounds are used as targets to characterize the potential functions of ASIC3 channels.

Differential regulation of tefluthrin and telmisartan on the gating charges of INa activation and inactivation as well as on resurgent and persistent INa in a pituitary cell line (GH3).

Voltage-gated Na+ currents (INa), known to contain many components (e.g., transient, resurgent and persistent INa) with unique gating properties, are involved in the generation and propagation of action potentials in excitable cells. In this study, how tefluthrin (Tef), a synthetic pyrethoid, and telmisartan (TEL), blocker of angiotensin II receptors can perturb those components of INa was investigated. The presence of either Tef or TEL increased the values of the gating charges of INa involved in the activation (za) and inactivation (zi). Tef also increased the amplitude of resurgent INa (INa(R)) or persistent INa (INa(P)) in a pituitary cell line (GH3), while TEL produced minimal effects on them. Subsequent addition of either ranolazine (a blocker of late INa) or d-limonene (a monoterpene), could reverse the changes by TEL or Tef on za or zi. In SCN5A-expressing HEK293T cells, addition of Tef or TEL also increased the peak amplitude and the inactivation time constant of INa which was accompanied by the increased za value of INa. Taken together, the results indicated that Tef- or TEL-mediated changes in the gating kinetics of INa are linked to their actions on functional activity of neurons, neuroendocrine or endocrine cells.

Ginger Phytochemicals Inhibit Cell Growth and Modulate Drug Resistance Factors in Docetaxel Resistant Prostate Cancer Cell.

Ginger has many bioactive compounds with pharmacological activities. However, few studies are known about these bioactive compounds activity in chemoresistant cells. The aim of the present study was to investigate the anticancer properties of ginger phytochemicals in docetaxel-resistant human prostate cancer cells in vitro. In this study, we isolated 6-gingerol, 10-gingerol, 4-shogaol, 6-shogaol, 10-shogaol, and 6-dehydrogingerdione from ginger. Further, the antiproliferation activity of these compounds was examined in docetaxel-resistant (PC3R) and sensitive (PC3) human prostate cancer cell lines. 6-gingerol, 10-gingerol, 6-shogaol, and 10-shogaol at the concentration of 100 µM significantly inhibited the proliferation in PC3R but 6-gingerol, 6-shogaol, and 10-shogaol displayed similar activity in PC3. The protein expression of multidrug resistance associated protein 1 (MRP1) and glutathione-S-transferase (GSTπ) is higher in PC3R than in PC3. In summary, we isolated the bioactive compounds from ginger. Our results showed that 6-gingerol, 10-gingerol, 6-shogaol, and 10-shogaol inhibit the proliferation of PC3R cells through the downregulation of MRP1 and GSTπ protein expression.