Objective To establish the method for simultaneous determination of six index components in the water extract of Weile Prescription;To optimize the water extraction process.Methods UPLC-MS/MS was used with Waters CORTECS C18 column(2.1 mm×100 mm,1.6 μm)as the chromatographic conditions;the mobile phase was 0.1%formic acid water-acetonitrile with gradient elution;the flow rate was 0.25 mL/min;the column temperature was 40℃;the sample volume was 2 μL.Electrospray negative ion source,positive and negative ion switching multi-reaction monitoring(MRM)mode were detected.Taking the content of six index components(gallic acid,vitexin,paeoniflorin,naringin,hesperidin and glycyrrhizic acid)and extraction rate as evaluation indexes,the weight coefficient of each index was determined by G1-entropy weight method,and the optimum parameters of extraction process were determined by orthogonal experiment design with the amount of water,extraction time and extraction times as investigation factors.Results There was a good linear relationship of the six components in the water extract of Weile Prescription in the concentration range(r>0.999),and the average recovery rate was 96.83%-102.56%,RSD<4.0%.The best technological parameters were as follows:Chinese decoction pieces were soaked in 12 times of water for 2 h,and extracted twice,each time for 1.5 h.Conclusion The UPLC-MS/MS method established in the study for simultaneous determination of six components in Weile Prescription is rapid,simple and sensitive,and the optimized extraction process is stable and feasible,which provides experimental basis for the development and research of the preparation.

Objective To explore the application low of stimulation parameters of transcutaneous electrical acupoint stimulation(TEAS)and transcranial direct current stimulation(tDCS)for post-stroke movement disorders based on data mining.Methods The relevant clinical research literature was retrieved from CNKI,Wanfang Data,VIP,CBM,PubMed and Web of Science from January 2000 to May 2023.A database was set up after quality assessment.Frequency analysis,association rules and complex network analysis were used to explore the application law of core acupoints and electrical stimulation parameters.Results A total of 79 articles were included and 128 groups of data were contained.For TEAS,the core acupoints included Waiguan(TE5),Shousanli(LI10),Zusanli(ST36),Hegu(LI4),Neiguan(PC6),Yanglingquan(GB34),etc.,while the most commonly used acupoint combinations of upper limb and lower limb were Shousanli(LI10)-Waiguan(TE5)and Yanglingquan(GB34)-Zusanli(ST36).Among the electrical stimulation parameters of TEAS,the frequencies used vary widely,and 100 Hz was most commonly used,while 2 Hz TEAS was also mainly used for stimulating acupoints located on upper limbs in the treatment of flaccid paralysis.The application of other electrical stimulation parameters was relatively consistent.The bidirectional symmetrical square-wave with 200-250 μs pulse-width was used in majority of studies.The stimulus intensity was mostly determined by patient tolerance.For tDCS,stimulation electrodes were often positioned on the projection of the primary M1,and the safe stimulus intensity was mostly set as 1 to 2 mA.Conclusion In the treatment of post-stroke movement disorders,appropriate acupoints and electrical stimulation parameters of TEAS should be determined on the muscle strength and muscle tension of stroke patients at different stages after stroke,particularly the selection of electric stimulating frequency.

Bladder cancer remains the 10th most common cancer worldwide. In recent years, metformin has been found to have potential anti-bladder cancer activ-ity while high concentration of IC50 at millimolar level is needed, which could not be reached by regular oral administration route. Thus, higher efficient agent is urgently demanded for clinically treating bladder cancer. Here, by conjugating artesunate to metformin, a novel artesunate-metformin dimer triazine derivative AM2 was designed and synthesized. The inhibitory effect of AM2 on bladder cancer cell line T24 and the mechanism underlying was determined. Anti-tumor activity of AM2 was assessed by MTT, cloning formation and wound healing assays. Decreasing effect of AM2 on lipogenesis was determined by oil red O staining. The protein expressions of Clusterin, SREBP1 and FASN in T24 cells were evaluated by Western blotting. The results show that AM2 significantly inhibited cell proliferation and migration at micromolar level, much higher than parental metformin. AM2 reduced lipogenesis and down-regulated the expressions of Clusterin, SREBP1 and FASN. These results suggest that AM2 inhibits the growth of bladder cancer cells T24 by inhibiting cellular lipogenesis associated with the Clusterin/SREBP1/FASN signaling pathway.

Functionalized liposomes can improve the in vivo process of drugs to achieve high-efficiency delivery by enhancing drug absorption， changing drug distribution and reducing the elimination， which is one of the hotspots in nanomedicine research with broad application prospects. However， the drug information published by official websites of National Medical Products Administration （NMPA）， the United States Food and Drug Administration （FDA）， and the European Medicines Agency （EMA） shows that there are few liposomal products on the market， and the domestic varieties are mainly generic drugs. Excepting for polyethylene glycolized （PEGylated） liposomes， no other functionalized liposomes have been approved for marketing， which indicates that the clinical translation of functionalized liposomes remains at a low level. Therefore， the relevant reports of functionalized liposomes in recent years were reviewed in this paper， their application advantages and main challenges in preparation research and development were discussed based on the in vivo process， and their low clinical translation mainly because of the insufficient clinical thinking， safety and efficacy of functional materials， inaccurate in vitro and in vivo analysis methods and difficulty in scaling up production. Meanwhile， the possible strategies such as introducing the concept of clinical multi-function to improve clinical acuity， focusing on examining the modification density of functional materials and the interaction between the modified materials， evaluating the drug delivery performance of functionalized liposomes from multiple perspectives and scenarios， and conducting cost and convenience-oriented formulation composition and preparation process optimization were proposed in order to provide a reference for the development of functionalized liposomes and other carrier-based nanomedicines.

Epidemiological and animal studies indicate that pre-existing diabetes increases the risk of Parkinson's disease(PD).However,the mechanisms underlying this association remain unclear.In the present study,we found that high glucose(HG)levels in the cerebrospinal fluid(CSF)of diabetic rats might enhance the effect of a subthreshold dose of the neurotoxin 6-hydroxydopamine(6-OHDA)on the development of motor disorders,and the damage to the nigrostriatal dopaminergic neuronal pathway.In vitro,HG promoted the 6-OHDA-induced apoptosis in PC12 cells differentiated to neurons with nerve growth factor(NGF)(NGF-PC12).Metabolomics showed that HG promoted hyperglycolysis in neurons and impaired tricarboxylic acid cycle(TCA cycle)activity,which was closely related to abnormal mito-chondrial fusion,thus resulting in mitochondrial loss.Interestingly,HG-induced upregulation of pyruvate kinase M2(PKM2)combined with 6-OHDA exposure not only mediated glycolysis but also promoted abnormal mitochondrial fusion by upregulating the expression of MFN2 in NGF-PC12 cells.In addition,we found that PKM2 knockdown rescued the abnormal mitochondrial fusion and cell apoptosis induced by HG+6-OHDA.Furthermore,we found that shikonin(SK),an inhibitor of PKM2,restored the mito-chondrial number,promoted TCA cycle activity,reversed hyperglycolysis,enhanced the tolerance of cultured neurons to 6-OHDA,and reduced the risk of PD in diabetic rats.Overall,our results indicate that diabetes promotes hyperglycolysis and abnormal mitochondrial fusion in neurons through the upre-gulation of PKM2,leading to an increase in the vulnerability of dopaminergic neurons to 6-OHDA.Thus,the inhibition of PKM2 and restoration of mitochondrial metabolic homeostasis/pathways may prevent the occurrence and development of diabetic PD.

Diabetic neuropathic pain (DNP) is the most common disabling complication of diabetes. Emerging evidence has linked the pathogenesis of DNP to the aberrant sprouting of sensory axons into the epidermal area; however, the underlying molecular events remain poorly understood. Here we found that an axon guidance molecule, Netrin-3 (Ntn-3), was expressed in the sensory neurons of mouse dorsal root ganglia (DRGs), and downregulation of Ntn-3 expression was highly correlated with the severity of DNP in a diabetic mouse model. Genetic ablation of Ntn-3 increased the intra-epidermal sprouting of sensory axons and worsened the DNP in diabetic mice. In contrast, the elevation of Ntn-3 levels in DRGs significantly inhibited the intra-epidermal axon sprouting and alleviated DNP in diabetic mice. In conclusion, our studies identified Ntn-3 as an important regulator of DNP pathogenesis by gating the aberrant sprouting of sensory axons, indicating that Ntn-3 is a potential druggable target for DNP treatment.
Mice ; Animals ; Diabetes Mellitus, Experimental/metabolism* ; Axons/physiology* ; Diabetic Neuropathies ; Sensory Receptor Cells/metabolism* ; Neuralgia/metabolism*

Objective: To investigate the level of mental health literacy among residents in Jiaxing City, Zhejiang Province, so as to provide insights into implementation of mental health education and improvements of the quality of mental health services. Methods: A total of 2 248 permanent residents at ages of 18 years and older were sampled using a multi-stage stratified random sampling method from 3 streets (townships) in Jiaxing City. Residents' demographics were collected using self-designed questionnaires, and the mental health literacy was investigated using the Mental Health Literacy Questionnaire. Factors affecting the achievement of the target of mental health literacy were identified among residents using a multivariable logistic regression model. Results: A total of 2 248 questionnaires were allocated and 2 172 valid questionnaires were recovered, with an effective recovery rate of 96.62%. The respondents included 1 075 men (49.49%) and 1 097 women (50.51%). There were 623 respondents that met the target of the mental health literacy (28.68%). Multivariable logistic regression analysis showed a higher possibility of achieving the target of mental health literacy levels among women than among men (OR=1.282, 95%CI: 1.047-1.570), among unmarried residents (OR=1.685, 95%CI: 1.018-2.788) than among married residents, among residents with educational levels of junior high school (OR=1.689, 95%CI: 1.168-2.441), high school/vocational high school/technical secondary school (OR=2.420, 95%CI: 1.601-3.658) and college or above (OR=3.543, 95%CI: 2.252-5.574) than among residents with an educational level of primary and below, among students (OR=2.572, 95%CI: 1.013-6.527), medical personnel (OR=3.330, 95%CI: 2.029-5.467), teachers (OR=2.909, 95%CI: 1.202-7.040), freelance/self-employed staff (OR=1.519, 95%CI: 1.100-2.098) and other professional technical personnel (OR=1.529, 95%CI: 1.012-2.310) than among workers. Conclusions The proportion of mental health literacy levels meeting the target is high among residents in Jiaxing City, and gender, educational level, occupation and marital status are factors affecting mental health literacy levels.

OBJECTIVE: To investigate the effects of high dose vitamin C (VC) on proliferation of breast cancer cells and to explore its mechanisms. METHODS: Human breast cancer cells Bcap37 and MDA-MB-453 were treated with VC at low dose (0.01 mmol/L), medium dose (0.10 mmol/L) and high dose (2.00 mmol/L). Cell proliferation was determined with CCK-8 assay, protein expression was evaluated by Western blot, and the secretion of lactic acid in tumor cells was detected by colorimetric method. Bcap37 cells were inoculated in nude mice, and tumor baring nude mice were intraperitoneally injected with high VC(4 g/kg, VC group, =5)or normal saline (control group, =5) for 24 d. Tumor weight and body weight were calculated. RESULTS: experiments demonstrated that high dose VC significantly inhibited cell proliferation in Bcap37 and MDA-MB-453 cells (all <0.01); the expressions of Glut1 and mTOR signaling pathway-related proteins were decreased (all <0.05); and the secretion of lactic acid was also markedly reduced (all <0.05). experiment showed that the tumor weight was decreased in mice treated with high-dose VC as compared with control group (<0.05), but no difference in body weights between two groups was observed. CONCLUSIONS High dose VC may inhibit proliferation of breast cancer cells both and through reducing glycolysis and protein synthesis.
Animals ; Ascorbic Acid ; pharmacology ; Breast Neoplasms ; drug therapy ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Glycolysis ; drug effects ; Humans ; Mice ; Mice, Nude ; Protein Biosynthesis ; drug effects

OBJECTIVE: To investigate the effect of rapamycin on Parkinson's disease (PD) and its underlying mechanism in mice. METHODS: Sixty SPF adult male C57BL/6 mice were randomly divided into control group, model group and treatment group. 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine(MPTP) was used to induce Parkinson's disease in model group and treatment group. All mice were trained to cross the runway and were subjected to computer-assisted CatWalk. The numbers of tyrosine hydroxylase positive (TH) neurons in the substantia nigra (SN) were assessed by unbiased stereology using the optical fractionator method; protein expression was detected by Western blot analysis; and glutathione peroxidase (GSH-Px), malondialdehyde (MDA) and superoxide dismutase (SOD) were detected by spectrophotometry. RESULTS: In the model group, a decrease in stride rate and an increase in variation of stance and swing were noted by CatWalk system (<0.05 or <0.01); the numbers of TH neurons decreased (<0.01); expression of p-Akt, p-S6K, p-S6 and p-ULK increased (all <0.01); LC3-Ⅱ/Ⅰ ratio decreased (<0.01); MDA level was elevated while the levels of SOD and GSH-PX were reduced (all <0.01). Compared with the model group, after treated with rapamycin, the abnormal behavior including the stride length, variation of stance and swing and step patterns induced by MPTP were all improved (<0.05 or <0.01); the numbers of TH neurons increased (<0.05); the expression of p-Akt, p-S6K, p-S6 and p-ULK was suppressed (all <0.01); the LC3-Ⅱ/Ⅰ ratio was upregulated (<0.05); MDA level decreased while the levels of GSH-Px and SOD increased (all <0.01). CONCLUSIONS Rapamycin inhibits the activation of mTOR pathway, which contributes to protect against the loss of dopaminergic neurons and provide behavioral improvements in mice with Parkinson's disease. These results are partially related to the ability of rapamycin in inducing autophagy and reducing oxidative stress.
Animals ; Behavior, Animal ; drug effects ; Disease Models, Animal ; Male ; Mice ; Mice, Inbred C57BL ; Neuroprotective Agents ; pharmacology ; therapeutic use ; Oxidative Stress ; drug effects ; Parkinson Disease ; drug therapy ; Random Allocation ; Sirolimus ; pharmacology ; therapeutic use ; Substantia Nigra ; drug effects

OBJECTIVE: To investigate the effects of Honokiol on cognitive function in mice with epilepsy. METHODS: Kainic acid (38 mg/kg) was intraperitoneally injected in 5 weeks old male ICR mice to induce epilepsy. Honokiol at dose of 3, 10, 30 mg/kg was given to epilepic mice by intraperitoneal injection for 10 days. Fluoro-Jade B staining was used to assess neuronal death; Morris water maze and Y maze tests were used to measure cognitive function such as learning and memory; Western blot was performed to detect the expression of acetylated superoxide dismutase (SOD), microtubule associated protein 1 light chain 3-Ⅱ (LC3-Ⅱ) and P62 in hippocampus tissue; thiobarbituric acid and WST-1 methods were used to detect malondialdehyde (MDA) and SOD. RESULTS: Compared with control group, the levels of acetylated-SOD, MDA, LC3-Ⅱ, P62 and neuronal death increased, cognitive function and SOD decreased in model group (<0.05 or <0.01). Honokiol at the dose of 10 mg/kg and 30 mg/kg decreased SOD acetylation, MDA content, expression of LC3-Ⅱ and P62, as well as neuronal death, and the cognitive function was improved (<0.05 or <0.01), especially in 30 mg/kg Honokiol group. CONCLUSIONS Honokiol alleviates oxidative stress and autophagy degradation disorder, decreases neuronal death, and therefore improves cognitive function in epilepsy mice.
Animals ; Biphenyl Compounds ; pharmacology ; Cognition ; drug effects ; Epilepsy ; chemically induced ; Gene Expression Regulation ; drug effects ; Hippocampus ; drug effects ; Kainic Acid ; Lignans ; pharmacology ; Male ; Malondialdehyde ; Maze Learning ; drug effects ; Mice ; Mice, Inbred ICR ; Neurons ; drug effects ; Superoxide Dismutase ; genetics