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.

Hyperbaric Oxygen Therapy Improves Parkinson's Disease by Promoting Mitochondrial Biogenesis via the SIRT-1/PGC-1α Pathway.

Hyperbaric oxygen therapy (HBOT) has been suggested as a potential adjunctive therapy for Parkinson's disease (PD). PD is a neurodegenerative disease characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The aim of this study was to investigate the protective mechanisms of HBOT on neurons and motor function in a 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD and 1-methyl-4-phenylpyridinium (MPP+)-mediated neurotoxicity in SH-SY5Y cells on the potential protective capability. In vivo: male C57BL/6 mice were randomly divided into three groups: control, MPTP group and MPTP+HBOT group. The MPTP-treated mice were intraperitoneally received MPTP (20 mg/kg) four times at 2 h intervals within a day. The day after MPTP treatment, MPTP+HBOT mice were exposed to hyperbaric oxygen at 2.5 atmosphere absolute (ATA) with 100% oxygen for 1 h once daily for 7 consecutive days. In vitro: retinoic acid (RA)-differentiated SH-SY5Y cells were treated with MPP+ for 1 h followed by hyperbaric oxygen at 2.5 ATA with 100% oxygen for 1 h. The results showed that MPTP induced a significant loss in tyrosine hydroxylase (TH)-positive neurons in the SNpc of mice. HBOT treatment significantly increased the number of TH-positive neurons, with enhanced neurotrophic factor BDNF, decreased apoptotic signaling and attenuated inflammatory mediators in the midbrain of MPTP-treated mice. In addition, MPTP treatment decreased the locomotor activity and grip strength of mice, and these effects were shown to improve after HBOT treatment. Furthermore, MPTP decreased mitochondrial biogenesis signaling (SIRT-1, PGC-1α and TFAM), as well as mitochondrial marker VDAC expression, while HBOT treatment was shown to upregulate protein expression. In cell experiments, MPP+ reduced neurite length, while HBOT treatment attenuated neurite retraction. Conclusions: the effects of HBOT in MPTP-treated mice might come from promoting mitochondrial biogenesis, decreasing apoptotic signaling and attenuating inflammatory mediators in the midbrain, suggesting its potential benefits in PD treatment.

Effectiveness in the Block by Honokiol, a Dimerized Allylphenol from Magnolia Officinalis, of Hyperpolarization-Activated Cation Current and Delayed-Rectifier K+ Current.

Background: Honokiol (HNK), a dimer of allylphenol obtained from the bark of Magnolia officinalis was demonstrated to exert an array of biological actions in different excitable cell types. However, whether or how this compound can lead to any perturbations on surface-membrane ionic currents remains largely unknown. Methods: We used the patch clamp method and found that addition of HNK effectively depressed the density of macroscopic hyperpolarization-activated cation currents (Ih) in pituitary GH3 cells in a concentration-, time- and voltage-dependent manner. By the use of a two-step voltage protocol, the presence of HNK (10 µM) shifted the steady-state activation curve of Ih density along the voltage axis to a more negative potential by approximately 11 mV, together with no noteworthy modification in the gating charge of the current. Results: The voltage-dependent hysteresis of Ih density elicited by long-lasting triangular ramp pulse was attenuated by the presence of HNK. The HNK addition also diminished the magnitude of deactivating Ih density elicited by ramp-up depolarization with varying durations. The effective half-maximal concentration (IC50) value needed to inhibit the density of Ih or delayed rectifier K+ current identified in GH3 cells was estimated to be 2.1 or 6.8 µM, respectively. In cell-attached current recordings, HNK decreased the frequency of spontaneous action currents. In Rolf B1.T olfactory sensory neurons, HNK was also observed to decrease Ih density in a concentration-dependent manner. Conclusions: The present study highlights the evidence revealing that HNK has the propensity to perturb these ionic currents and that the hyperpolarization-activated cyclic nucleotide-gated (HCN) channel is proposed to be a potential target for the in vivo actions of HNK and its structurally similar compounds.

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 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.

Protective effects of Liuwei dihuang water extracts on diabetic muscle atrophy.

BACKGROUND: Liuwei dihuang (LWDH), a widely used traditional Chinese herbal medicine, has been noticed for its potential on the improvement of diabetic complications including diabetic nephropathy and diabetic encephalopathy. However, whether LWDH can improve the effects of diabetic skeletal muscle atrophy has not yet been reported. PURPOSE: The present study aimed to investigate the protective effects and mechanisms of the water extract of Liuwei dihuang (LWDH-WE) on skeletal muscle in cellular and animal models of diabetic muscle atrophy. STUDY DESIGN: The muscle protective effects of LWDH-WE on diabetic muscle atrophy and weakness were examined in methylglyoxal (MG)-treated C2C12 myotubes and streptozotocin (STZ)-treated C57BL/6 mice, respectively. METHODS: C2C12 myoblasts were differentiated by differentiation medium to form myotube structures. C2C12 myotubes were pre-treated with LWDH-WE 1 h before MG treatment. Diabetic mice were induced by single intraperitoneal injection of STZ (150 mg/kg) in C57BL/6 mice. Cell viability was determined by MTT and LDH assays. Protein expressions were detected by western blots. Morphological changes of cells were observed by an inverted microscope. Mitochondria membrane potential and reactive oxygen species (ROS) production were measured by flow cytometry. Muscle strength was evaluated by measuring grip strength of mice. RESULTS: In C2C12 myotubes, LWDH-WE attenuated MG-induced cellular death and oxidative damage accompanied with improving mitochondrial membrane potential, inhibiting NADPH oxidase (Nox) activation, and ROS production. Moreover, LWDH-WE could attenuate MG-induced atrophy of C2C12 myotubes accompanied with regulating protein synthesis (IGF-1R, Akt, mTOR) and protein degradation (FoxO3a, atrogin-1, MuRF-1) signals. In STZ-induced diabetic mice, LWDH-WE improved body weight and skeletal muscle mass of mice. LWDH-WE also enhanced muscle strength of STZ-induced diabetic mice. Furthermore, LWDH-WE enhanced the improvement of insulin on gastrocnemius muscle mass and grip strength in STZ-treated mice. CONCLUSION: LWDH-WE possesses skeletal muscle protection via reducing oxidative damage and regulating protein synthesis and degradation pathways in MG-induced atrophy of C2C12 myotubes. We also reveal the novel protection of LWDH-WE against STZ-induced reduction of muscle mass and muscle strength in mice.

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.

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.

The water extract of Liuwei dihuang possesses multi-protective properties on neurons and muscle tissue against deficiency of survival motor neuron protein.

BACKGROUND: Deficiency of survival motor neuron (SMN) protein, which is encoded by the SMN1 and SMN2 genes, induces widespread splicing defects mainly in spinal motor neurons, and leads to spinal muscular atrophy (SMA). Currently, there is no effective treatment for SMA. Liuwei dihuang (LWDH), a traditional Chinese herbal formula, possesses multiple therapeutic benefits against various diseases via modulation of the nervous, immune and endocrine systems. Previously, we demonstrated water extract of LWDH (LWDH-WE) protects dopaminergic neurons and improves motor activity in models of Parkinson's disease. PURPOSE: This study aimed to investigate the potential protection of LWDH-WE on SMN deficiency-induced neurodegeneration and muscle weakness. STUDY DESIGN: The effects of LWDH-WE on SMN deficiency-induced neurotoxicity and muscle atrophy were examined by using SMN-deficient NSC34 motor neuron-like cells and SMA-like mice, respectively. METHODS: Inducible SMN-knockdown NSC34 motor neuron-like cells were used to mimic SMN-deficient condition. Doxycycline (1 µg/ml) was used to induce SMN deficiency in stable NSC34 cell line carrying SMN-specific shRNA. SMAΔ7 mice were used as a severe type of SMA mouse model. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays. Apoptotic cells and neurite length were observed by inverted microscope. Protein expressions were examined by western blots. Muscle strength of animals was evaluated by hind-limb suspension test. RESULTS: LWDH-WE significantly increased SMN protein level, mitochondrial membrane potential and cell viability of SMN-deficient NSC34 cells. LWDH-WE attenuated SMN deficiency-induced down-regulation of B-cell lymphoma-2 (Bcl-2) and up-regulation of cytosolic cytochrome c and cleaved caspase-3. Moreover, LWDH-WE prevented SMN deficiency-induced inhibition of neurite outgrowth and activation of Ras homolog gene family, member A (RhoA)/ Rho-associated protein kinase (ROCK2)/ phospho-LIM kinase (p-LIMK)/ phospho-cofilin (p-cofilin) pathway. Furthermore, in SMA-like mice, LWDH-WE improved muscle strength and body weight accompanied with up-regulation of SMN protein in spinal cord, brain, and gastrocnemius muscle tissues. CONCLUSION: The present study demonstrated that LWDH-WE protects motor neurons against SMN deficiency-induced neurodegeneration, and it also improves the muscle strength of SMA-like mice, suggesting the potential benefits of LWDH-WE as a complementary prescription for SMN deficiency-related diseases.

The Chinese herbal formula Liuwei dihuang protects dopaminergic neurons against Parkinson's toxin through enhancing antioxidative defense and preventing apoptotic death.

Liuwei dihuang (LWDH), a widely used traditional Chinese medicine (TCM), has been employed as an anti-aging prescription to improve declined function. Parkinson's disease (PD) is a common adult-onset neurodegenerative disorder characterized by the degeneration of dopaminergic nigrostriatal neurons with complex pathological mechanisms, including oxidative stress. Increasing evidence indicate that TCM has the potential to be neuroprotective drugs because of their antioxidant characteristics. The aim of this study is to investigate the mechanisms of LWDH-mediated protection in Parkinson's toxin-induced dopaminergic neurodegeneration by evaluating water extract of LWDH (LWDH-WE) in 1-methyl-4-phenylpyridinium (MPP(+))-treated primary mesencephalic neurons and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated C57BL/6 mice. In the present study, chemical profiling and quantitative analysis of LWDH-WE were revealed using 3D-HPLC technique, and were confirmed by the data of three batches of LWDH-WE. In primary mesencephalic neuronal cultures, LWDH-WE decreased MPP(+)-induced loss of tyrosine hydroxylase (TH)-positive neurons and increase of Annexin V-positive neurons. LWDH-WE reduced MPP(+)-induced oxidative damage via increasing antioxidant defense (SOD, GSH), decreasing ROS production, and down-regulating NADPH oxidases (Nox2 and Nox4). Also, LWDH-WE inhibited neuronal apoptosis by improving mitochondrial membrane potential, increasing antiapoptotic protein Bcl-2 expression, and down-regulating apoptotic signaling (Bax, cytochrome c, cleaved-caspase-3) in MPP(+)-treated neurons. In MPTP-treated C57BL/6 mice, LWDH-WE attenuated TH-positive neuronal loss in substantia nigra pars compacta (SNpc), and improved locomotor activity of mice. In conclusion, the present results reveal that LWDH-WE possesses protection on dopaminergic neurons through enhancing antioxidant defense and decreasing apoptotic death, suggesting the potential benefits of LWDH-WE for PD treatment.

Evidence for inhibitory effects of flupirtine, a centrally acting analgesic, on delayed rectifier k(+) currents in motor neuron-like cells.

Flupirtine (Flu), a triaminopyridine derivative, is a centrally acting, non-opiate analgesic agent. In this study, effects of Flu on K(+) currents were explored in two types of motor neuron-like cells. Cell exposure to Flu decreased the amplitude of delayed rectifier K(+) current (I(K(DR))) with a concomitant raise in current inactivation in NSC-34 neuronal cells. The dissociation constant for Flu-mediated increase of I(K(DR)) inactivation rate was about 9.8 µM. Neither linopirdine (10 µM), NMDA (30 µM), nor gabazine (10 µM) reversed Flu-induced changes in I(K(DR)) inactivation. Addition of Flu shifted the inactivation curve of I(K(DR)) to a hyperpolarized potential. Cumulative inactivation for I(K(DR)) was elevated in the presence of this compound. Flu increased the amplitude of M-type K(+) current (I(K(M))) and produced a leftward shift in the activation curve of I(K(M)). In another neuronal cells (NG108-15), Flu reduced I(K(DR)) amplitude and enhanced the inactivation rate of I(K(DR)). The results suggest that Flu acts as an open-channel blocker of delayed-rectifier K(+) channels in motor neurons. Flu-induced block of I(K(DR)) is unlinked to binding to NMDA or GABA receptors and the effects of this agent on K(+) channels are not limited to its action on M-type K(+) channels.

Berberine activates Nrf2 nuclear translocation and protects against oxidative damage via a phosphatidylinositol 3-kinase/Akt-dependent mechanism in NSC34 motor neuron-like cells.

Berberine (BBR) is a well-known anti-diabetic herbal medicine in Asia due to its beneficial effects on insulin sensitivity, glucose metabolism and glycolysis. Here, we identified the critical role of phosphatidylinositol 3-kinase (PI3K)/Akt involved BBR cellular defense mechanisms and first revealed the novel effect of BBR on nuclear factor (erythroid-derived 2)-related factor-2 (Nrf2)/heme oxygenase (HO)-1 induction in NSC34 motor neuron-like cells. BBR (0.1-10 nM) led to increasing insulin receptor expression, Akt phosphorylation and enhanced oxidant-sensitive Nrf2/HO-1 induction, which were blocked by a PI3K inhibitor, LY294002. In H(2)O(2)-treated cells, BBR significantly attenuated ROS production and increased cell viability, antioxidant defense (GSH and SOD) and oxidant-sensitive proteins (HO-1 and Nrf2), which also were blocked by LY294002. Furthermore, BBR improved mitochondrial function by increasing mitochondrial membrane potential and decreasing the oxygen consumption rate. BBR-induced anti-apoptotic function was demonstrated by increasing anti-apoptotic protein Bcl-2 and survival of motor neuron protein (SMN) and by decreasing apoptotic proteins (cytochrome c, Bax and caspase). These results suggest that BBR, which is active at nanomolar concentration, is a potential neuroprotective agent via PI3K/Akt-dependent cytoprotective and antioxidant pathways.

Neuroprotective Effects of San-Huang-Xie-Xin-Tang in the MPP(+)/MPTP Models of Parkinson's Disease In Vitro and In Vivo.

San-Huang-Xie-Xin-Tang (SHXT), composed of Coptidis rhizoma, Scutellariae radix, and Rhei rhizoma, is a traditional Chinese medicine used for complementary and alternative therapy of cardiovascular and neurodegenerative diseases via its anti-inflammatory and antioxidative effects. The aim of this study is to investigate the protective effects of SHXT in the 1-methyl-4-phenylpyridinium (MPP(+))/1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) models of Parkinson's disease. Rat primary mesencephalic neurons and mouse Parkinson disease model were used in this study. Oxidative stress was induced by MPP(+) in vitro and MPTP in vivo. In MPP(+)-treated mesencephalic neuron cultures, SHXT significantly increased the numbers of TH-positive neurons. SHXT reduced apoptotic signals (cytochrome and caspase) and apoptotic death. MPP(+)-induced gp91(phox) activation and ROS production were attenuated by SHXT. In addition, SHXT increased the levels of GSH and SOD in MPP(+)-treated neurons. In MPTP animal model, SHXT markedly increased TH-positive neurons in the substantia nigra pars compacta (SNpc) and improved motor activity of mice. In conclusion, the present results reveal the evidence that SHXT possesses beneficial protection against MPTP-induced neurotoxicity in this model of Parkinson's disease via its antioxidative and antiapoptotic effects. SHXT might be a potentially alternative and complementary medicine for neuroprotection.

San-Huang-Xie-Xin-Tang Protects against Activated Microglia- and 6-OHDA-Induced Toxicity in Neuronal SH-SY5Y Cells.

San-Huang-Xie-Xin-Tang (SHXT), composed of Coptidis rhizoma, Scutellariae radix and Rhei rhizoma, is a traditional Chinese herbal medicine used to treat gastritis, gastric bleeding and peptic ulcers. This study investigated the neuroprotective effects of SHXT on microglia-mediated neurotoxicity using co-cultured lipopolysaccharide (LPS)-activated microglia-like BV-2 cells with neuroblastoma SH-SY5Y cells. Effects of SHXT on 6-hydroxydopamine (6-OHDA)-induced neurotoxicity were also examined in SH-SY5Y cells. Results indicated SHXT inhibited LPS-induced inflammation of BV-2 cells by downregulation of iNOS, NO, COX-2, PGE(2), gp91(phox), iROS, TNF-α, IL-1ß, inhibition of IκBα degradation and upregulation of HO-1. In addition, SHXT increased cell viability and down regulated nNOS, COX-2 and gp91(phox) of SH-SY5Y cells co-cultured with LPS activated BV-2 cells. SHXT treatment increased cell viability and mitochondria membrane potential (MMP), decreased expression of nNOS, COX-2, gp91(phox) and iROS, and inhibited IκBα degradation in 6-OHDA-treated SH-SY5Y cells. SHXT also attenuated LPS activated BV-2 cells- and 6-OHDA-induced cell death in differentiated SH-SY5Y cells with db-cAMP. Furthermore, SHXT-inhibited nuclear translocation of p65 subunit of NF-κB in LPS treated BV-2 cells and 6-OHDA treated SH-SY5Y cells. In conclusion, SHXT showed protection from activated microglia- and 6-OHDA-induced neurotoxicity by attenuating inflammation and oxidative stress.

6-Hydroxycleroda-3,13-dien-15,16-olide protects neuronal cells from lipopolysaccharide-induced neurotoxicity through the inhibition of microglia-mediated inflammation.

Polyalthia longifolia var. pendula is used as an antipyretic agent in indigenous systems of medicine. Microglia-mediated inflammation plays an important role in the pathway leading to neuronal cell death in a number of neurodegenerative diseases. The aim of this study was to investigate the effects of 6-hydroxycleroda-3,13-dien-15,16-olide (PL3) extracted from Polyalthia longifolia var. pendula on lipopolysaccharide(LPS)-induced inflammation in microglia-like HAPI cells and primary microglia cultures. In microglia-neuron co-cultures, LPS decreased the cell viability of neuroblastoma SH-SY5Y cells. LPS-induced cell death was attenuated by the NOS inhibitor, L-NAME, the COX-2 inhibitor, NS-398 or the NADPH oxidase inhibitor, DPI, respectively. In LPS-treated microglia cells, PL3 decreased the expression of iNOS, COX-2, gp91 (phox), and NF- kappaBp65, the degradation of I kappaB alpha, and the production of NO, PGE (2), iROS, and TNF- alpha. PL3 also enhanced the expression of HO-1, a cytoprotective and anti-inflammatory enzyme. Moreover, PL3 reduced LPS-activated microglia-induced cell death. The present results suggest that PL3 inhibits microglia-mediated inflammation and inflammation-related neuronal cell death. Therefore, PL3 has potential use for the treatment of inflammation-related neurodegenerative diseases.

Anticancer activity of isoobtusilactone A from Cinnamomum kotoense: involvement of apoptosis, cell-cycle dysregulation, mitochondria regulation, and reactive oxygen species.

In this study, we investigate the anticancer effect of isoobtusilactone A (IOA), a constituent isolated from the leaves of Cinnamomum kotoense, on human non-small cell lung cancer (NSCLC) A549 cells. IOA was found to induce the arrest of G2-M phase, induce apoptosis, increase sub-G1, and inhibit the growth of these cells. Further investigation revealed that IOA's blockade of the cell cycle was associated with increased levels of p21/WAF1, p27 (kip1), and p53. In addition, IOA triggered the mitochondrial apoptotic pathway, as indicated by an increase in Bax/Bcl-2 ratios, resulting in a loss of mitochondrial membrane potential, release of cytochrome c, activation of caspase-9 and caspase-3, and cleavage of PARP. We also found the generation of reactive oxygen species (ROS) to be a critical mediator in IOA-induced inhibition of A549 cell growth. In antioxidant and NO inhibitor studies, we found that by pretreating A549 cells with either N-acetylcystenine (NAC), catalase, mannitol, dexamethasone, trolox, or L-NAME we could significantly decrease IOA production of ROS. Moreover, using NAC to block ROS, we could significantly suppress IOA-induced antiproliferation, antimigration, and anti-invasion. Finally, we found that IOA inhibited the migration and invasion of A549 cell migration and invasion. Taken together, these results suggest that IOA has anticancer effects on A549 cells.

Effects of San-Huang-Xie-Xin-Tang on U46619-induced increase in pulmonary arterial blood pressure.

ETHNOPHARMACOLOGICAL RELEVANCE: San-Huang-Xie-Xin-Tang (SHXT), composed of Coptidis rhizoma, Scutellariae radix and Rhei rhizoma, is traditionally used to treat hypertension. AIM OF THE STUDY: Our aim was to investigate the pharmacology effect of SHXT on a thromboxane A(2) analogue U46619-induced increase in pulmonary hypertension and protein expression in primary pulmonary smooth muscle cells (PASMCs). MATERIALS AND METHODS: Arterial blood pressure and isometric tension in the aorta and pulmonary artery of rats were measured by pressure and force transducers, respectively. Protein expressions on PASMCs were detected by Western blotting. RESULTS: SHXT significantly attenuated U46619-induced increase in arterial blood pressure. The inhibitory effect of SHXT on pulmonary arterial pressure was greater than systemic arterial pressure in U46619 treated rats. Similarly, the inhibitory effect of SHXT on U46619-induced vasoconstriction in rat pulmonary arterial rings was greater than that in aortic rings. In U46619 treated PASMCs, SHXT down-regulated expression of phosphodiesterase type 5 (PDE5), Rho-kinase (ROCK) II, cyclooxygenase-2 (COX-2) and up-regulated expression of soluble guanylyl cyclase (sGC) alpha(1) and sGCbeta(1). CONCLUSIONS: SHXT attenuated U46619-induced increase in systemic and pulmonary arterial blood pressure. Inhibition of PDE5, ROCK-II, COX-2 and stimulation of sGC may play important roles in the cardiovascular effects of SHXT.

Synthesis and anti-platelet evaluation of 2-benzoylaminobenzoate analogs.

Fifty-two 2-benzoylaminobenzoate analogs were synthesized and subjected to anti-platelet aggregation assay using arachidonic acid (AA), collagen (Col), thrombin (Thr), and U46619 as inducers. The results revealed that most of 2-benzoylaminobenzoic acid derivatives showed a selectively inhibitory effect on AA-induced platelet aggregation. As a result of the 2-benzoylaminobenzoic acid derivatives (18, 44, and 46), there were no inhibitory effects on platelet aggregation induced by U46619, but these elicited an inhibitory effect on thromboxane B(2) formation at 1.0microM. These 2-benzoylaminobenzoate analogs were therefore proposed as cyclooxygenase inhibitors.

Neuronal effects of 4-t-Butylcatechol: a model for catechol-containing antioxidants.

Many herbal medicines and dietary supplements sold as aids to improve memory or treat neurodegenerative diseases or have other favorable effects on the CNS contain a catechol or similar 1,2-dihydroxy aromatic moiety in their structure. As an approach to isolate and examine the neuroprotective properties of catechols, a simple catechol 4-t-Butylcatechol (TBC) has been used as a model. In this study, we investigated the effects of TBC on lipopolysaccharide (LPS)-activated microglial-induced neurotoxicity by using the in vitro model of coculture murine microglial-like cell line HAPI with the neuronal-like human neuroblastoma cell line SH-SY5Y. We also examined the effects of TBC on 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in human dopaminergic neuroblastoma SH-SY5Y cells. TBC at concentrations from 0.1-10 microM had no toxic effect on HAPI cells and SH-SY5Y cells, and it inhibited LPS (100 ng/ml)-induced increases of superoxide, intracellular ROS, gp91(Phox), iNOS and a decrease of HO-1 in HAPI cells. Under coculture condition, TBC significantly reduced LPS-activated microglia-induced dopaminergic SH-SY5Y cells death. Moreover, TBC (0.1-10 microM) inhibited 6-OHDA-induced increases of intracellular ROS, iNOS, nNOS, and a decrease of mitochondria membrane potential, and cell death in SH-SY5Y cells. However, the neurotoxic effects of TBC (100 microM) on SH-SY5Y cells were also observed including the decrease in mitochondria membrane potential and the increase in COX-2 expression and cell death. TBC-induced SH-SY5Y cell death was attenuated by pretreatment with NS-398, a selective COX-2 inhibitor. In conclusion, this study suggests that TBC might possess protective effects on inflammation- and oxidative stress-related neurodegenerative disorders. However, the high concentration of TBC might be toxic, at least in part, for increasing COX-2 expression.