Alkaloids extracted from Uncaria rhynchophylla demonstrate neuroprotective effects in MPTP-induced experimental parkinsonism by regulating the PI3K/Akt/mTOR signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Alkaloids isolated from Uncaria rhynchophylla (Miq.) Miq. ex Havil. (Rubiaceae), alkaloids (URA) have been used to treat diseases related to the central nervous system, such as Parkinson's disease. Nevertheless, the potential mechanisms underlying their neuroprotective effects are not well-understood. AIM OF THE STUDY: We investigated the neuroprotective effects of URAs in a mouse model of 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) and the possible involvement of a molecular signaling pathway. MATERIALS AND METHODS: Two typical experiments for animal behavior despair, the spontaneous motor activity and the rotarod experiments, were employed to evaluate the efficacy of URAs in mice with PD symptoms. Dopamine (DA) neurons and their metabolism were evaluated using high-performance liquid chromatography-tandem mass spectrometry. The mechanism of action of the alkaloids was investigated by analyzing their effects on the phosphoinositide 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway using western blotting. RESULTS: URA treatment effectively improved the behaviors of the mice during the "spontaneous motor activity and latency to fall off the rotarod test". Moreover, URAs demonstrated a protective role in dopaminergic neurons by increasing the expression of the dopamine transporter and tyrosine hydroxylase, which were supposed to be reduced by MPTP, inhibiting dopamine turnover, and changing dopamine and relevant metabolites. In addition to its association with the increase in the Bcl-2/Bad ratio, URA treatment also attenuated the cleaved caspase-3 level and enhanced the phosphorylation of Akt and mTOR. CONCLUSION: These findings provide evidence that URA can effectively protect neurons from the neurotoxicity caused by MPTP in mouse models of PD by up-regulating the PI3K/Akt/mTOR signaling pathway.

Protection by rhynchophylline against MPTP/MPP+-induced neurotoxicity via regulating PI3K/Akt pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Isolated from Uncaria rhynchophylla (U. rhynchophylla), rhynchophylline (Rhy) has been applied for treating diseases related to central nervous system such as Parkinson's disease. Nevertheless, the molecular mechanism of the neuroprotective effect has not been well interpreted. AIM OF THE STUDY: To investigate the effects of Rhy on MPTP/MPP + -induced neurotoxicity in C57BL/6 mice or PC12 cells and study the mechanisms involved. MATERIALS AND METHODS: The neuroprotective effect of Rhy on MPTP-induced neurotoxicity was evaluated by spontaneous motor activity test, as well as a test of rota-rod on a rat model of Parkinson's disease. The numbers of TH-positive neurons in the substantia nigra pars compacta (SNpc) was assessed by immunohistological. CCK-8, lactate dehydrogenase (LDH), reactive oxygen species (ROS), the concentration of intracellular calcium ([Ca2+]i) and flow cytometry analysis were performed to evaluate the pharmacological property of Rhy on 1-methyl-4-phenylpyridinium (MPP+) induced neurotoxicity in PC12 cells. Besides, LY294002, a PI3K inhibitor was employed to determine the underlying molecular signaling pathway revealing the effect of Rhy by western-blot analysis. RESULTS: The results showed that Rhy exhibited a protective effect against the MPTP-induced decrease in tyrosine hydroxylase (TH)-positive fibers in the substantia nigra at 30 mg/kg, demonstrated by the immunohistological and behavioral outcomes. Furthermore, it has been indicated that cell viability was improved and the MPP+-induced apoptosis was inhibited after the treatment of Rhy at 20 µM, which were severally analyzed by the CCK-8 and the Annexin V/propidium iodide staining method. In addition, Rhy treatment attenuated MPP+-induced up-regulation of LDH, ([Ca2+]i), and the levels of ROS. Besides, it can be revealed from the Western blot assay that LY294002, as a selective Phosphatidylinositol 3-Kinase (PI3K) inhibitor, effectively inhibited the Akt phosphorylation caused by Rhy, which suggested that Rhy showed its protective property through the activated the PI3K/Akt signaling pathway. Moreover, the Rhy-induced decreases of Bax and caspase-3 as the proapoptotic markers and the increase of Bcl-2 as the antiapoptotic marker, were blocked by LY294002 in the MPP+-treated PC12 cells. CONCLUSIONS: Rhy exerts a neuroprotective effect is partly mediated by activating the PI3K/Akt signaling pathway.

Over-Expression of LcPDS, LcZDS, and LcCRTISO, Genes From Wolfberry for Carotenoid Biosynthesis, Enhanced Carotenoid Accumulation, and Salt Tolerance in Tobacco.

It is of great importance to combine stress tolerance and plant quality for breeding research. In this study, the role of phytoene desaturase (PDS), ζ-carotene desaturase (ZDS) and carotene isomerase (CRTISO) in the carotenoid biosynthesis are correlated and compared. The three genes were derived from Lycium chinenses and involved in the desaturation of tetraterpene. Their over-expression significantly increased carotenoid accumulation and enhanced photosynthesis and salt tolerance in transgenic tobacco. Up-regulation of almost all the genes involved in the carotenoid biosynthesis pathway and only significant down-regulation of lycopene ε-cyclase (ε-LCY) gene were detected in those transgenic plants. Under salt stress, proline content, and activities of catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) were significantly increased, whereas malonaldehyde (MDA) and hydrogen peroxide (H2O2) accumulated less in the transgenic plants. The genes encoding ascorbate peroxidase (APX), CAT, POD, SOD, and pyrroline-5-carboxylate reductase (P5CR) were shown to responsive up-regulated significantly under the salt stress in the transgenic plants. This study indicated that LcPDS, LcZDS, and LcCRTISO have the potential to improve carotenoid content and salt tolerance in higher plant breeding.

Total glucosides of paeony (TGP) extracted from Radix Paeoniae Alba exerts neuroprotective effects in MPTP-induced experimental parkinsonism by regulating the cAMP/PKA/CREB signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The totally-amounted glucosides of paeony (TGP), which are made up of paeoniflorin, albiflorin, oxypaeoniflorin as well as benzoylpaeoniflorin, constitute the Baishao' actively-working component extracted from Radix Paeonia alba employed in conventional oriental medicine aiming to treat cerebrovascular disorders, such as Parkinson's disease. However, its pharmacologic mechanism is not clear. AIM OF THE STUDY: The initial investigation was made on TGP's neuroprotective effects on PD of the mouse model based on 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) as well as the identification of potential involvement of a molecular signaling pathway. MATERIALS AND METHODS: The evaluation of the behavioral damage as well as neurotoxicity in mice was made through MPTP. Spontaneous motor activity test, as well as a test of Rota-rod on mice was employed for the measurement of bradykinesia symptom. Additionally, liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS) works as the determiner of the main monoamine neurotransmitters dopamine (DA) along with its metabolites 3, 4-dihydroxyphenylacetic acid (DOPAC) as well as homovanillic acid (HVA) based on mouse hippocampus connected with the anti-Parkinson's disease like effect of TGP. Besides, the measurement of the effects of TGP treatment on the expressions level of TH, DAT, a-synuclein, p-CREBS133 as well as apoptosis influence was made with the help of western-blot assay with apoptosis-related markers such as Bax and Bcl-2. RESULTS: The results showed that TGP treatment lessened the behavior-based loss shown "in the spontaneous motor activity as well as the potential of falling to rotarod test". In addition, we found that pretreatment with TGP markedly improved motor coordination, striatal dopamine and its metabolite levels. Furthermore, pretreatment of TGP conducted the protection for dopaminergic neurons with the prevented MPTP-induced reductions within the tyrosine hydroxylase (TH), substantia nigra dopaminergic transporter (DAT), as well as increasing α-synuclein protein levels with transformed dopamine catabolism as well as inhibited dopamine turnover. Besides, TGP treatment helped reversed apoptosis signaling molecules Bcl-2/Bax' reduction; meanwhile improving p-CREBS133 the factor of growth signaling in the substantia nigra' decrease. CONCLUSION: These results suggested that TGP can enhance dopaminergic neuron's cell survival in the SNpc in virtue of the activated cAMP/PKA/CREB factor of growth on inhibiting the pathway of second messenger apoptosis as well. In conclusion, the current findings indicate TGP is expected to be a new cure for PD.

Neuroprotection by Paeoniflorin in the MPTP mouse model of Parkinson's disease.

Paeoniflorin (PF) is a major bioactive ingredient in Radix Paeonia alba roots that has low toxicity and has been shown to have neuroprotective effects. Our in vitro experiments suggested that PF affords a significant neuroprotective effect against MPP+-induced damage and apoptosis in PC12 cells through Bcl-2/Bax/caspase-3 pathway. The objectives of the present study were to explore the potential neuroprotective effect of PF in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-treated mouse model of Parkinson's disease (PD). Our results demonstrated that PF treatment ameliorated the behavioral deficits of "in spontaneous motor activity and latency to fall of the rotarod test", and reduced dopaminergic cell loss that were induced by MPTP in a dose-dependent manner in an in vivo model of PD. In addition, we found that treatment of PF protected dopaminergic neurons by preventing MPTP-induced decreases in striatal and substantia nigra dopaminergic transporter (DAT) and tyrosine hydroxylase (TH) protein levels, and by changing dopamine catabolism and inhibiting dopamine turnover. Furthermore, it was also associated with up-regulation of the Bcl-2/BAD ratio, and inhibition of the activation of caspase-9 and caspase-3. These results showed that PF promoted dopamine neuron survival in vivo due to the MAO-B inhibition, and the PI3K/Akt signaling pathway may have mediated the protection of PF against MPTP, suggesting that PF treatment might represent a neuroprotective treatment for PD.

Protective Effects of Paeoniflorin Against MPP(+)-induced Neurotoxicity in PC12 Cells.

In the present study, we investigated the protective mechanism of paeoniflorin (PF), a monoterpene glycoside extracted from Radix Paeoniae alba roots, on MPP(+)-induced neurotoxicity in cultured rat pheochromocytoma cells (PC12). Our work included examination of cell viability assessment, amounts of released lactic dehydrogenase (LDH), intracellular Ca(2+) concentration, cell apoptosis, mitochondrial membrane potential, caspase-3 activity, and expression profiling of two apoptosis-related genes (Bcl-2 and Bax). It was shown that, PF functioned as an MPP(+) antagonist, being able to suppress apoptosis, decrease LDH release and Ca(2+) concentration, attenuate membrane potential collapse and, inhibit caspase-3 activation, decrease in Bax/Bcl-2 ratio. These observations suggest that PF could protect PC12 cells against MPP(+)-induced injury and the mechanism PF's neuroprotective effect was closely associated with Bcl-2 up-regulation and Bax down-regulation. PF has neuroprotective effects on MPP(+)-induced apoptosis in PC12 cells via regulating mitochondrial membrane potential and Bcl-2/Bax/caspase-3 signaling pathways, and this new insight will help develop a PF-based therapeutic strategy for treatmenting neurodegenerative diseases and injury.

A simple and effective system for foreign gene expression in plants via root absorption of agrobacterial suspension.

Due to the laborious and scale-up limitation we have developed a simple system named "root absorption" to express foreign proteins in plants successfully. It has been shown that GFP was expressed in tobacco plants by root absorbing the Agrobacterium suspension containing TMV-based P35S-30B-GFP vector. Various factors influencing the gene expression were studied including Agrobacterium cell density, seedling age, plant materials and inoculation conditions. This system has the special advantages as simple and convenient work process, ease to scale-up and higher level of expression than leaf infiltration. Interestingly, GFP was expressed at 24h post-absorption. We assume that the root absorption system will facilitate the large-scale production of the recombinant pharmaceutical proteins in plants by means of transient expression.

A modified method of coracoid transposition for the treatment of complete dislocation of acromioclavicular joint.

OBJECTIVE: To report a new method of coracoid transposition for the treatment of complete dislocation of acromioclavicular joint and to evaluate its efficacy. METHODS: We modified Dewar's surgical method as follows: (1) Two small incisions, a transversal incision on the acromioclavicular joint and a longitudinal incision on the coracoid, were made instead of a conventional large arc incision from the acromion to coracoid. (2) The foreign body in the acromioclavicular joint was cleared out. The chondral surface at the lateral segment of clavicle was resected to form a pseudarthrosis and meanwhile the residual joint capsule and ligaments were repaired. (3) The coracoid was moved to the anteroinferior edge of the clavicle instead of the anterior margin and (4) the coracoid was moved to the lateral border of the clavicle instead of the superior border of the coracoclavicular ligament. RESULTS: The follow-up duration in 30 patients of the series was from 6 to 72 months (mean 41 months). Functional assessment was carried out by the criteria delineated previously by Karkson, in which Grade A was in 24 cases, Grade B in 4 cases, and C in 2. CONCLUSIONS: This modified technique, having less postoperative complications and less injuries to tissues and according well with the requirement of biomechanics, can achieve a stable reduction of acromioclavicular joint with a good functional and cosmetic result and therefore is preferable to use clinically on a large scale.