Lycorine, a natural alkaloid, promotes the degradation of alpha-synuclein via PKA-mediated UPS activation in transgenic Parkinson's disease models.

BACKGROUND: Parkinson's disease (PD) is one of the most common neurodegenerative motor disorders, and is characterized by the presence of Lewy bodies containing misfolded α-synuclein (α-syn) and by selective degeneration of midbrain dopamine neurons. Studies have shown that upregulation of ubiquitin-proteasome system (UPS) activity promotes the clearance of aggregation-prone proteins such as α-syn and Tau, so as to alleviate the neuropathology of neurodegenerative diseases. PURPOSE: To identify and investigate lycorine as a UPS enhancer able to decrease α-syn in transgenic PD models. METHODS: Dot blot was used to screen α-syn-lowering compounds in an inducible α-syn overexpression cell model. Inducible wild-type (WT) and mutant α-syn-overexpressing PC12 cells, WT α-syn-overexpressing N2a cells and primary cultured neurons from A53T transgenic mice were used to evaluate the effects of lycorine on α-syn degradation in vitro. Heterozygous A53T transgenic mice were used to evaluate the effects of lycorine on α-syn degradation in vivo. mCherry-GFP-LC3 reporter was used to detect autophagy-dependent degradation. Ub-R-GFP and Ub-G76V-GFP reporters were used to detect UPS-dependent degradation. Proteasome activity was detected by fluorogenic substrate Suc-Leu-Leu-Val-Tyr-AMC (Suc-LLVY-AMC). RESULTS: Lycorine significantly promoted clearance of over-expressed WT and mutant α-syn in neuronal cell lines and primary cultured neurons. More importantly, 15 days' intraperitoneal administration of lycorine effectively promoted the degradation of α-syn in the brains of A53T transgenic mice. Mechanistically, lycorine accelerated α-syn degradation by activating cAMP-dependent protein kinase (PKA) to promote proteasome activity. CONCLUSION: Lycorine is a novel α-syn-lowering compound that works through PKA-mediated UPS activation. This ability to lower α-syn implies that lycorine has the potential to be developed as a pharmaceutical for the treatment of neurodegenerative diseases, such as PD, associated with UPS impairment and protein aggregations.

Elevated GTP Cyclohydrolase I Pathway in Endothelial Progenitor Cells of Overweight Premenopausal Women.

Background/Aims. Sexual differences exist in endothelial progenitor cells (EPCs), and various cardiovascular risk factors are associated with the preservation of endothelial function in premenopausal women. However, it is unclear whether differences in endothelial function and circulating EPCs exist between overweight premenopausal women and age-matched men. Methods. We compared EPC counting and functions in normal-weight and overweight premenopausal women and men, evaluated endothelial function in each group, and detected the expression of the guanosine triphosphate cyclohydrolase I (GTPCH I) pathway. Results. The number of EPCs was lower in the male group than in the female group, regardless of normal-weight or overweight status, and there was no significant difference between the different weight groups among females or males. Endothelial function and EPC migration and proliferation were preserved in overweight premenopausal women compared with overweight men as were nitric oxide (NO) levels in plasma and secreted by EPCs. Endothelial function, the circulating EPC population, and NO levels were not different between normal-weight and overweight premenopausal women. Flow-mediated dilatation was significantly correlated with EPC function, plasma NO levels, and EPC-secreted NO. Conclusions. This investigation provides the first evidence for sex-based differences in EPC activity and endothelial function in overweight middle-aged individuals; these differences are associated with alterations in NO production and may partly occur through downregulation of the GTPCH I pathway. The present results provide new insights into the mechanism underlying the preserved endothelial function in overweight premenopausal women and may uncover a potential therapeutic target for endothelial repair in overweight population.

Pharmacological enhancement of TFEB-mediated autophagy alleviated neuronal death in oxidative stress-induced Parkinson's disease models.

Autophagy, a conserved cellular degradation and recycling process, can be enhanced by nutrient depletion, oxidative stress or other harmful conditions to maintain cell survival. 6-Hydroxydopamine/ascorbic acid (6-OHDA/AA) is commonly used to induce experimental Parkinson's disease (PD) lesions by causing oxidative damage to dopaminergic neurons. Activation of autophagy has been observed in the 6-OHDA-induced PD models. However, the mechanism and exact role of autophagy activation in 6-OHDA PD model remain inconclusive. In this study, we report that autophagy was triggered via mucolipin 1/calcium/calcineurin/TFEB (transcription factor EB) pathway upon oxidative stress induced by 6-OHDA/AA. Interestingly, overexpression of TFEB alleviated 6-OHDA/AA toxicity. Moreover, autophagy enhancers, Torin1 (an mTOR-dependent TFEB/autophagy enhancer) and curcumin analog C1 (a TFEB-dependent and mTOR-independent autophagy enhancer), significantly rescued 6-OHDA/AA-induced cell death in SH-SY5Y cells, iPSC-derived DA neurons and mice nigral DA neurons. The behavioral abnormality of 6-OHDA/AA-treated mice can also be rescued by Torin 1 or C1 administration. The protective effects of Torin 1 and C1 can be blocked by autophagy inhibitors like chloroquine (CQ) or by knocking down autophagy-related genes TFEB and ATG5. Taken together, this study supports that TFEB-mediated autophagy is a survival mechanism during oxidative stress and pharmacological enhancement of this process is a neuroprotective strategy against oxidative stress-associated PD lesions.

Neuroprotective effects of berberine in animal models of Alzheimer's disease: a systematic review of pre-clinical studies.

BACKGROUND: Berberine is an isoquinoline alkaloid extracted from various Berberis species which is widely used in East Asia for a wide range of symptoms. Recently, neuroprotective effects of berberine in Alzheimer's disease (AD) animal models are being extensively reported. So far, no clinical trial has been carried out on the neuroprotective effects of berberine. However, a review of the experimental data is needed before choosing berberine as a candidate drug for clinical experiments. We conducted a systematic review on AD rodent models to analyze the drug effects with minimal selection bias. METHODS: Five online literature databases were searched to find publications reporting studies of the effect of berberine treatment on animal models of AD. Up to March 2018, 15 papers were identified to describe the efficacy of berberine. RESULTS: The included 15 articles met our inclusion criteria with different quality ranging from 3 to 5. We analyzed data extracted from full texts with regard to pharmacological effects and potential anti-Alzheimer's properties. Our analysis revealed that in multiple memory defects animal models, berberine showed significant memory-improving activities with multiple mechanisms, such as anti-inflammation, anti-oxidative stress, cholinesterase (ChE) inhibition and anti-amyloid effects. CONCLUSION: AD is likely to be a complex disease driven by multiple factors. Yet, many therapeutic strategies based on lowering ß-amyloid have failed in clinical trials. This suggest that the threapy should not base on a single cause of Alzheimer's disease but rather a number of different pathways that lead to the disease. Overall we think that berberine can be a promising multipotent agent to combat Alzheimer's disease.

Natural alkaloid harmine promotes degradation of alpha-synuclein via PKA-mediated ubiquitin-proteasome system activation.

BACKGROUND: Parkinson's disease (PD) is an age-dependent progressive movement disorder characterized by a profound and selective loss of nigrostriatal dopaminergic neurons. Accumulation of -synuclein (-syn) positive protein aggregates in the substantia nigra is a pathological hallmark of PD, indicating that protein turnover defect is implicated in PD pathogenesis. PURPOSE: This study aims to identify neuroprotective compounds which can alleviate the accumulation of -syn in neuronal cells and dissect the underlying mechanisms. METHODS: High throughput screening was performed by dot blot assay. The degradation of different forms of -syn by candidate compounds were assessed by western blot. The autophagy lysosome pathway and ubiquitin-proteasome system were examined to dissect the degradation pathway. The UPS activity was assessed by cellular UPS substrates degradation assay and biochemical proteasome activity assay. Q-PCR was performed to test the mRNA level of different proteasome subunits. Furthermore, Neuroprotective effect of candidate compound was tested by LDH assay and PI staining. RESULTS: Through the high throughput screening, harmine was identified as a potent -syn lowering compound. The time-dependent and dose-dependent effects of harmine on the degradation of different forms of -syn were further confirmed. Harmine could dramatically promote the degradation of UPS substrates GFP-CL1, Ub-R-GFP and Ub-G76V-GFP, and activate cellular proteasome activity. Mechanistically, harmine dramatically enhanced PKA phosphorylation to enhance proteasome subunit PSMD1 expression. PKA inhibitor blocked the effects of harmine in activating UPS, up regulating PSMD1 and promoting -syn degradation, indicating that harmine enhances UPS function via PKA activation. Moreover, harmine efficiently rescued cell death induced by over-expression of -syn, via UPS-dependent manner. CONCLUSION: Harmine, as a new proteasome enhancer, may have potential to be developed into therapeutic agent against neurodegenerative diseases associated with UPS dysfunction and aberrant proteins accumulation.

Online comprehensive two-dimensional hydrophilic interaction chromatography×reversed-phase liquid chromatography coupled with hybrid linear ion trap Orbitrap mass spectrometry for the analysis of phenolic acids in Salvia miltiorrhiza.

In the present study, a system was developed for the analysis of phenolic acids in Salvia miltiorrhiza using online comprehensive two-dimensional hydrophilic interaction chromatography and reversed-phase liquid chromatography coupled to a DAD detector and hybrid linear ion trap-Orbitrap mass spectrometry (HILIC×RP-DAD-ESI/HRMS/MSn). The system was configured based on the combination of an XBridge Amide column (150mm×4.6mm, 3.5µm) and Accucore PFP column (50mm×4.6mm, 2.6µm) for the first and second dimensions, respectively. An additional LC pump was used to dilute the eluent from the first dimension to decrease its elution strength in the second dimension. A back-flush trap column was selected as an interface to make up for the loss of efficiency and resolution due to the online dilution. Under the optimized conditions, a total of 196 peaks of polar compounds were successfully separated and detected in Salvia miltiorrhiza using the developed online HILIC×RP system, which exhibited high orthogonality (73%). The online combination of HILIC and RP provides powerful separation capability for the analysis of polar compounds in samples with complex matrices.

Trace determination of carbamate pesticides in medicinal plants by a fluorescent technique.

The safety issue of using carbamate pesticides in medicinal plants (MPs) has been a global concern and hence attracted attention of many researchers to develop analytical tools for trace pesticides detection. Derived from the fluorescence-based techniques, a rapid, convenient and efficient method for the detection of three carbamate pesticides, including carbofuran, aldicarb and methomyl has been developed by using core-shell QDs. By optimizing experimental parameters, the system demonstrated high detection sensitivities for the investigated carbamates, with the lowest detectable concentrations less than 0.05 µM. The molecular docking study indicated that the selected carbamate pesticides bound to the catalytic active site of acetylcholinesterase via π-π or H-π interactions, which also revealed the potential mechanism of the differences in inhibition strength among the three pesticides on AChE. Moreover, in order to investigate the applicability and reliability of the proposed method for the pesticide analysis in real sample with complex matrix, the matrix effects of eight common MPs have been systematically explored. These findings suggested that this technique was a simple, sensitive and reliable method for rapid determination of carbamate pesticides in real samples, especially those with complex matrices like MPs, vegetables, fruits, and other agricultural crops.

Reactivated astrocytes as a possible source of oligodendrocyte precursors for remyelination in remitting phase of experimental autoimmune encephalomyelitis rats.

Multiple sclerosis (MS) is ademyelinating disease in the central nervous system (CNS). Majority of the MS patients show relapsing-remitting disease course. Evidences show that oligodendrocyte precursor cells (OPCs), which remain relatively quiescent in normal adult CNS, play a key role in the remitting phase by proliferation and remyelination. In the present study, we found that spinal cord astrocytesco-expressed progenitor cell marker and oligodendroglial lineage markers in the remittance phase in adult rat experimental autoimmune encephalomyelitis (EAE) model. We suggest that activated astrocyte could de-differentiate into OPCs and re-differentiate into mature oligodendrocytes, raising the possibility that astrocytes can be a potential source of OPCs in the adult demyelinated spinal cord.

[Current topics on cancer biology and research strategies for anti-cancer traditional Chinese medicine].

Cancer, an abnormal cell proliferation resulted from multi-factors,has the highest morbidity and mortality among all the serious diseases. Considerable progress has been made in cancer biology in recent years. Tumor immunology, cancer stem cells (CSCs), autophagy, and epithelial-mesenchymal transition (EMT) have become hot topics of interests in this area. Detailed dissection of these biological processes will provide novel directions, targets, and strategies for the pharmacological evaluation, mechanism elucidation, and new drug development of traditional Chinese medicine.

Application of two-dimensional chromatography in the analysis of Chinese herbal medicines.

For the purpose of better understanding the complex Chinese herbal medicines (CHMs) and controlling their quality, powerful analytical techniques are essential. Although conventional one-dimensional (1D) chromatographic approaches have been widely used for the analysis of multiple components in CHMs, the complexity of CHM samples often exceeds the maximal capacity of any single separation mode. Therefore, in past decades, many researchers have attempted to explore the coupling of independent separation techniques to improve the resolving power for complex CHM samples. Two-dimensional (2D) separation systems, based on two independent columns with different separation mechanisms, have proven to be more powerful than 1D techniques and have been used successfully to separate and analyze CHM samples with excellent performance. This article aims to review the most recent advances in the strategies for analyzing CHMs using 2D chromatography. For this purpose, some remarkable applications of the commonly used couplings, mainly including 2D-GC and 2D-LC for analysis of CHMs, are described. Moreover, their major advantages and shortcomings are discussed, which might be helpful to the researchers who focus on quality control of CHMs.

Xyloketal B exhibits its antioxidant activity through induction of HO-1 in vascular endothelial cells and zebrafish.

We previously reported that a novel marine compound, xyloketal B, has strong antioxidative actions in different models of cardiovascular diseases. Induction of heme oxygenase-1 (HO-1), an important endogenous antioxidant enzyme, has been considered as a potential therapeutic strategy for cardiovascular diseases. We here investigated whether xyloketal B exhibits its antioxidant activity through induction of HO-1. In human umbilical vein endothelial cells (HUVECs), xyloketal B significantly induced HO-1 gene expression and translocation of the nuclear factor-erythroid 2-related factor 2 (Nrf-2) in a concentration- and time-dependent manner. The protection of xyloketal B against angiotensin II-induced apoptosis and reactive oxygen species (ROS) production could be abrogated by the HO-1 specific inhibitor, tin protoporphyrin-IX (SnPP). Consistently, the suppressive effects of xyloketal B on NADPH oxidase activity could be reversed by SnPP in zebrafish embryos. In addition, xyloketal B induced Akt and Erk1/2 phosphorylation in a concentration- and time-dependent manner. Furthermore, PI3K inhibitor LY294002 and Erk1/2 inhibitor U0126 suppressed the induction of HO-1 and translocation of Nrf-2 by xyloketal B, whereas P38 inhibitor SB203580 did not. In conclusion, xyloketal B can induce HO-1 expression via PI3K/Akt/Nrf-2 pathways, and the induction of HO-1 is mainly responsible for the antioxidant and antiapoptotic actions of xyloketal B.

Elevated blood pressure aggravates intracerebral hemorrhage-induced brain injury.

Elevated blood pressure (BP) is commonly seen in patients with intracerebral hemorrhage (ICH), and is independently associated with poor functional outcomes. Little is known about how elevated BP influences ICH-related brain injury. In the present study, we investigated the physiological and brain histological changes, as well as functional recovery following ICH in renovascular hypertensive rats. Renovascular hypertension (RVHT) was achieved by applying a silver clip onto the left renal artery of adult Sprague-Dawley rats. ICH was induced by an intrastriatal injection of bacterial collagenase IV about 5-6 weeks after left renal artery clipping or the sham operation. Following induction of ICH, both the normotensive and RVHT rats demonstrated an ultra-acute elevation in BP. Elevated BP increased hematoma volume, brain swelling, and apoptosis in the perihematomal areas. Brain degeneration, including local atrophy and lateral ventricle enlargement, was greater in the RVHT rats. In addition, many proliferating cells were seen over the ipsilateral striatum in the RVHT rats after ICH. The modified limb placing tests were done weekly for 3 weeks. In line with the histological damage, elevated BP worsened neurological deficits. These results suggest that ICH in the hypertensive rats mimics the clinical scenario of hypertensive ICH and may provide a platform to study the mechanisms of ICH-induced brain injury and potential therapies for ICH.

Early detection of neurodegeneration in brain ischemia by manganese-enhanced MRI.

This study aims to employ in vivo manganese-enhanced MRI (MEMRI) to detect neurodegenerative changes in two models of brain ischemia, photothrombotic cortical injury (PCI) and transient middle cerebral artery occlusion (MCAO) in rodents. After systemic Mn(2+) injection to both ischemic models, a close pattern of T1-weighted hyperintensity was observed throughout different brain regions in comparison to the distribution of GFAP, MnSOD and GS immunoreactivities, whereby conventional MRI could hardly detect such. In addition, the infarct volumes in the posterior parts of the brain had significantly reduced after Mn(2+) injection to the MCAO model. It is suggested that exogenous Mn(2+) injection may provide enhanced MEMRI detection of oxidative stress and gliosis early after brain ischemia. Manganese may also mediate infarctions at remote brain regions in transient focal cerebral ischemia before delayed secondary damage takes place.