Publications by authors named "Zhen-Hao Li"

12 Publications

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Spore Polysaccharide Inhibits the Growth of Hepatocellular Carcinoma Cells by Altering Macrophage Polarity and Induction of Apoptosis.

J Immunol Res 2021 5;2021:6696606. Epub 2021 Mar 5.

School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing 100029, China.

Background: has certain components with known pharmacological effects, including strengthening immunity and anti-inflammatory activity. seeds inherit all its biological characteristics. spore polysaccharide (GLSP) is the main active ingredient to enhance these effects. However, its specific biological mechanisms are not exact. Our research is aimed at revealing the specific biological mechanism of GLSP to enhance immunity and inhibit the growth of H22 hepatocellular carcinoma cells.

Methods: We extracted primary macrophages (M) from BALB/c mice and treated them with GLSP (800 g/mL, 400 g/mL, and 200 g/mL) to observe its effects on macrophage polarization and cytokine secretion. We used GLSP and GLSP-intervened macrophage supernatant to treat H22 tumor cells and observed their effects using MTT and flow cytometry. Moreover, real-time fluorescent quantitative PCR and western blotting were used to observe the effect of GLSP-intervened macrophage supernatant on the PI3K/AKT and mitochondrial apoptosis pathways.

Results: In this study, GLSP promoted the polarization of primary macrophages to M1 type and the upregulation of some cytokines such as TNF-, IL-1, IL-6, and TGF-1. The MTT assay revealed that GLSP+M at 400 g/mL and 800 g/mL significantly inhibited H22 cell proliferation in a dose-dependent manner. Flow cytometry analysis revealed that GLSP+Mø induced apoptosis and cell cycle arrest at the G2/M phase, associated with the expression of critical genes and proteins (PI3K, p-AKT, BCL-2, BAX, and caspase-9) that regulate the PI3K/AKT pathway and apoptosis. GLSP reshapes the tumor microenvironment by activating macrophages, promotes the polarization of primary macrophages to M1 type, and promotes the secretion of various inflammatory factors and cytokines.

Conclusion: Therefore, as a natural nutrient, GLSP is a potential agent in hepatocellular carcinoma cell treatment and induction of apoptosis.
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http://dx.doi.org/10.1155/2021/6696606DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7954632PMC
November 2021

Antitumor effects of different Ganoderma lucidum spore powder in cell- and zebrafish-based bioassays.

J Integr Med 2021 03 13;19(2):177-184. Epub 2021 Jan 13.

Zhejiang Shouxiangu Pharmaceutical Co., Ltd, Wuyi 321200, Zhejiang Province, China; Pharmaceutical Informatics Institute, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310012, Zhejiang Province, China. Electronic address:

Objective: Ganoderma lucidum spore (GLS) is gaining recognition as a medicinal part of G. lucidum and has been reported to possess various pharmacological properties, such as antitumor activity. In this work, wall-broken GLS powder (BGLSP) and wall-removed GLS powder (RGLSP), two kinds of GLS powder with different manufacturing techniques, were compared in terms of contents of active constituents and in vivo and in vitro antitumor effects.

Methods: The ultraviolet and visible spectrophotometry method was used to determine the contents of polysaccharides and total triterpenoids in BGLSP and RGLSP. Seventeen individual triterpenoids were further quantified using ultra-high-performance liquid chromatography and quantitative analysis of multi-components by single marker. The antitumor effects of BGLSP and RGLSP were evaluated using in vitro cell viability assay against human gastric carcinoma SGC-7901, lung carcinoma A549 and lymphoma Ramos and further validated by in vivo zebrafish xenograft models with transplanted SGC-7901, A549 and Ramos.

Results: The results showed that the contents of polysaccharides, total triterpenoids and individual triterpenoids of RGLSP were significantly higher than those of BGLSP. Although both BGLSP and RGLSP inhibited the three tumor cell lines in vitro in a dose-dependent manner, the inhibitory effects of RGLSP were much better than those of BGLSP. In the in vivo zebrafish assay, RGLSP exhibited more potent inhibitory activities against tumors transplanted into the zebrafish compared with BGLSP, and the inhibition rates of RGLSP reached approximately 78%, 31% and 83% on SGC-7901, A549 and Ramos, respectively.

Conclusion: The results indicated that the antitumor effects of GLS were positively correlated with the contents of the polysaccharides and triterpenoids and demonstrated that the wall-removing manufacturing technique could significantly improve the levels of active constituents, and thereby enhance the antitumor activity.
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http://dx.doi.org/10.1016/j.joim.2021.01.004DOI Listing
March 2021

Prophylactic effects of sporoderm-removed Ganoderma lucidum spores in a rat model of streptozotocin-induced sporadic Alzheimer's disease.

J Ethnopharmacol 2021 Apr 25;269:113725. Epub 2020 Dec 25.

Department of Pharmacology, School of Basic Medical Science, Peking University, 38 Xueyuan Road, Beijing, 100191, China. Electronic address:

Ethnopharmacological Relevance: Ganoderma lucidum (G. lucidum, Lingzhi), also known as "immortality mushroom" has been broadly used to improve health and longevity for thousands of years in Asia. G. lucidum and its spores have been used to promote health, based on its broad pharmacological and therapeutic activity. This species is recorded in Chinese traditional formula as a nootropic and has been suggested to improve cognitive dysfunction in Alzheimer's disease. However, little is known about the nootropic effects and molecular mechanism of action of G. lucidum spores.

Aim Of The Study: The present study investigated the protective effects of sporoderm-deficient Ganoderma lucidum spores (RGLS) against learning and memory impairments and its mechanism of action.

Materials And Methods: In the Morris water maze, the effects of RGLS on learning and memory impairments were evaluated in a rat model of sporadic Alzheimer's disease that was induced by an intracerebroventricular injection of streptozotocin (STZ). Changes in amyloid β (Aβ) expression, Tau expression and phosphorylation, brain-derived neurotrophic factor (BDNF), and the BDNF receptor tropomyosin-related kinase B (TrkB) in the hippocampus were evaluated by Western blot.

Results: Treatment with RGLS (360 and 720 mg/kg) significantly enhanced memory in the rat model of STZ-induced sporadic Alzheimer's disease and reversed the STZ-induced increases in Aβ expression and Tau protein expression and phosphorylation at Ser199, Ser202, and Ser396. The STZ-induced decreases in neurotrophic factors, including BDNF, TrkB and TrkB phosphorylation at Tyr816, were reversed by treatment with RGLS.

Conclusion: These findings indicate that RGLS prevented learning and memory impairments in the present rat model of STZ-induced sporadic Alzheimer's disease, and these effects depended on a decrease in Aβ expression and Tau hyperphosphorylation and the modulation of BDNF-TrkB signaling in the hippocampus.
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http://dx.doi.org/10.1016/j.jep.2020.113725DOI Listing
April 2021

[UPLC fingerprint combined with quantitative analysis of multi-components by single marker for quality assessment of Danshen Injection].

Zhongguo Zhong Yao Za Zhi 2019 Sep;44(17):3724-3731

Department of Chinese Medical Science and Engineering,College of Pharmaceutical Science,Zhejiang University Hangzhou 310058,China.

Testing and analysis of chemical markers is currently the prevailing approach for quality assessment of traditional Chinese medicines. However,several important issues remain to be addressed,including the trade-off between accuracy and coverage. In this study,in order to give full play to the advantages of their respective methods and provide technical support for more comprehensively and rapidly evaluate the quality of Danshen Injection products,a fingerprint method was coupled with quantitative analysis of multicomponents by single marker( QAMS),with Danshen Injection as the carrier. Ultra performance liquid chromatography( UPLC) was used to establish the quantitative fingerprint. The UPLC fingerprints contained 13 common peaks,11 of which were identified by using ultra performance liquid chromatography-quadrupole time-of-flight mass spectrometry( UPLC-QTOF-MS). Furthermore,with sodium danshensu as the internal reference substance,relative correction factors( RCFs) of protocatechuic aldehyde,caffeic acid,rosmarinic acid,lithospermic acid,and salvianolic acid B were calculated through slope analysis method,and the QAMS method was adopted to determine the contents of these 6 components. The UPLC fingerprint was employed to assess the consistency of 12 batches of Danshen Injection,which showed good batch-to-batch consistency with similarity higher than 0. 99. In the comparison of contents of the six constituents obtained by QAMS and external standard method( ESM),RSD was all less than 4. 3%,indicating the good accuracy of the QAMS method. The QAMS method developed in this study combined with UPLC fingerprint can comprehensively reflect the internal quality of Danshen Injection when only the reference substance sodium danshensu is consumed,with greatly reduced detection cost and time. It provides a technical basis for further improving the quality standard of Danshen Injection.
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http://dx.doi.org/10.19540/j.cnki.cjcmm.20190410.310DOI Listing
September 2019

A multiple biomarker assay for quality assessment of botanical drugs using a versatile microfluidic chip.

Sci Rep 2017 09 25;7(1):12243. Epub 2017 Sep 25.

Pharmaceutical Informatics Institute, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, China.

Quality control is critical for ensuring the safety and effectiveness of drugs. Current quality control method for botanical drugs is mainly based on chemical testing. However, chemical testing alone may not be sufficient as it may not capture all constituents of botanical drugs. Therefore, it is necessary to establish a bioassay correlating with the drug's known mechanism of action to ensure its potency and activity. Herein we developed a multiple biomarker assay to assess the quality of botanicals using microfluidics, where enzyme inhibition was employed to indicate the drug's activity and thereby evaluate biological consistency. This approach was exemplified on QiShenYiQi Pills using thrombin and angiotensin converting enzyme as "quality biomarkers". Our results demonstrated that there existed variations in potency across different batches of the intermediates and preparations. Compared with chromatographic fingerprinting, the bioassay provided better discrimination ability for some abnormal samples. Moreover, the chip could function as "affinity chromatography" to identify bioactive phytochemicals bound to the enzymes. This work proposed a multiple-biomarker strategy for quality assessment of botanical drugs, while demonstrating for the first time the feasibility of microfluidics in this field.
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http://dx.doi.org/10.1038/s41598-017-12453-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5612938PMC
September 2017

[Situation analysis and standard formulation of pesticide residues in traditional Chinese medicines].

Zhongguo Zhong Yao Za Zhi 2017 Jun;42(12):2284-2290

The State Key Laboratory Breeding Base of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China.

Chinese Pharmacopoeia provides nine pesticide Maximum Residual Limits(MRLs) of traditional Chinese medicines(TCMs), The number of pesticides used in production are far more than those listed in pharmacopoeia. The lack of the standards make it's hard to reflect the real situation of pesticide residues in TCMs correctly. The paper is aimed to analyze the data of pesticide residues in TCMs from 7 089 items in 140 reports, and judging the exceedance rate of pesticides in TCMs using the MRLs of European pharmacopoeia,which is widely accepted in many countries. The results show that:①Pesticide residues in 18 kinds of TCMs are higher than MRLs,while in 137 kinds are below MRLs, such as Atractylodis Macrocephalae Rhizoma, Menthae Haplocalycis Herba and Fritillariae Thunbergii Bulbus. The average exceedance rate of all TCMs is 1.72%. The average exceedance rates of organochlorine, organophosphorus and pyrethroid are 2.26%, 1.51%, 0.37%,respectively. ②The average exceedance rate of pesticides is 2.00%, and the exceedance rate is more than 5%, accounting for 8.33%, the exceedance rate is between 1%-5%, accounting for 18.75%. the exceedance rate is between 0%-1%, accounting for 18.75%. The remaining 29 kinds of pesticides were not exceeded, accounting for 60.42%.Some reports like Greenpeace's organization exaggerated the pesticide residues in TCMs.But the pesticide residue question is still worthy of attention, so we proposed to amend the Chinese Pharmacopoeia pesticide residues standards, to increase the pesticide species of traditional Chinese medicine in production on the basis of retaining the existing types of pesticide residues, to strengthen the system research of pesticide residues in TCMs, providing a basis for making standard and promoting import and export trade in TCMs.
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http://dx.doi.org/10.19540/j.cnki.cjcmm.2017.0108DOI Listing
June 2017

[The technological innovation strategy for quality control of Chinese medicine based on Big Data].

Zhongguo Zhong Yao Za Zhi 2015 Sep;40(17):3374-8

The evolution of the quality control concepts of medical products within the global context and the development of the quality control technology of Chinese medicine are briefly described. Aimed at the bottlenecks in the regulation and quality control of Chinese medicine, using Big Data technology to address the significant challenges in Chinese medicine industry is proposed. For quality standard refinements and internationalization of Chinese medicine, a technological innovation strategy encompassing its methodology, and the R&D direction of the subsequent core technology are also presented.
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September 2015

[Effects of different penetration enhancers on pharmacokinetics of active components in Xiangfu Siwu transdermal patches].

Zhongguo Zhong Yao Za Zhi 2016 Jan;41(2):294-302

China National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, Nanjing University of Chinese Medicine, Nanjing 210023, China.

To study the effect of different penetration enhancers on the pharmacokinetic characters of six active components in Xiangfu Siwu transdermal patch (XBW) and optimize the best penetration enhancers. During the experiment, the patches containing different penetration enhancers were stuck on the rat's skin, and then the blood samples were acquired at different time points. Six active components in plasma were determined by UPLC-MS/MS. The main pharmacokinetic parameters were calculated with DAS software package. The total factor scores (F) of the plasma concentrations of six components at every time point in different groups were calculated using principle component analysis, and the areas under F versus time curves (AUCF-t) were employed to be the indexes for selecting penetration enhancers. The results demonstrated that compared with the control group, the AUCF-t from other groups increased prominently and furthermore, 5% menthol manifested the best effect. In this research, 5% menthol could remarkably promote the percutaneous penetration effect of the six active compounds in XBW, and it could provide a scientific basis for the preparation research of XBW.
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http://dx.doi.org/10.4268/cjcmm20160221DOI Listing
January 2016

Comparisons of pharmacokinetic and tissue distribution profile of four major bioactive components after oral administration of Xiang-Fu-Si-Wu Decoction effective fraction in normal and dysmenorrheal symptom rats.

J Ethnopharmacol 2014 Jul 14;154(3):696-703. Epub 2014 May 14.

Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, Nanjing University of Chinese Medicine, Nanjing 210023, China; National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, Nanjing University of Chinese Medicine, Nanjing 210023, China. Electronic address:

Ethnopharmacological Relevance: Xiang-Fu-Si-Wu Decoction (XFSWD) has been widely used to treat primary dysmenorrhea in clinical practice for hundreds of years and shown great efficacy. One fraction of XFSWD, which was an elution product by macroporous adsorption resin from aqueous extract solution with 60% ethanol (XFSWE), showed great analgesic effect. The present study was conducted to investigate the possible pharmacokinetic and tissue distribution profiles of four major bioactive constituents (berberine, protopine, tetrahydrocoptisine and tetrahydropalmatine) after oral administration of XFSWE in dysmenorrheal symptom rats, and to compare the difference between normal and dysmenorrheal symptom rats.

Materials And Methods: Estradiol benzoate and oxytocin were used to produce dysmenorrheal symptom rat model. The experimental period was seven days. At the final day of experimental period, both normal and dysmenorrheal symptom rats were orally administrated with XFSWE, and then the blood and tissues samples were collected at different time points. Berberine, protopine, tetrahydrocoptisine and tetrahydropalmatine in blood and tissue samples were determined by LC-MS/MS. Pharmacokinetic parameters were calculated from the plasma concentration-time data using non-compartmental methods. The differences of pharmacokinetic parameters among groups were tested by one-way analysis of variance (ANOVA).

Results: There were statistically significant differences (P<0.05) in Cmax, Tmax, AUC(0-t), AUC(0-∞), MRT(0-t), MRT(0-∞) and CL/F between normal and dysmenorrheal symptom rats that orally administered with same dosage of XFSWE. In tissue distribution study, the results showed that the overall trend was C(Spleen)>C(Liver)>C(Kidney)>C(Uterus)>C(Heart)>C(Lung)>C(Ovary)>C(Brain)>C(Thymus), C(M-60 min)>C(M-120 min)>C(M-30 min)>C(C-60 min)>C(C-120 min)>C(C-30 min). The contents of protopine in liver, spleen and uterus were more than that in other tissues of dysmenorrheal symptom rats. Compared to normal rats, partial contents of the compounds in dysmenorrheal symptom rats׳ tissues at different time points had significant difference (P<0.05).

Conclusions: This study was the first report about pharmacokinetic and tissue distribution investigation in dysmenorrheal symptom animals. The results indicated that berberine, protopine, tetrahydrocoptisine and tetrahydropalmatine have higher uptake and slower elimination in the rats with dysmenorrheal syndrome, which suggests that the rate and extent of drug metabolism were altered in dysmenorrheal syndrome rats. And the results also demonstrated that berberine, protopine and tetrahydropalmatine in normal and dysmenorrheal symptom rats had obvious differences in some organs and time points, suggesting that the blood flow and perfusion rate of the organ were altered in dysmenorrheal symptom animals.
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http://dx.doi.org/10.1016/j.jep.2014.04.044DOI Listing
July 2014

[In vitro transdermal delivery of the active fraction of xiangfusiwu decoction based on principal component analysis].

Yao Xue Xue Bao 2013 Jun;48(6):933-9

Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing 210046, China.

The objective of the present study was to establish a method based on principal component analysis (PCA) for the study of transdermal delivery of multiple components in Chinese medicine, and to choose the best penetration enhancers for the active fraction of Xiangfusiwu decoction (BW) with this method. Improved Franz diffusion cells with isolated rat abdomen skins were carried out to experiment on the transdermal delivery of six active components, including ferulic acid, paeoniflorin, albiflorin, protopine, tetrahydropalmatine and tetrahydrocolumbamine. The concentrations of these components were determined by LC-MS/MS, then the total factor scores of the concentrations at different times were calculated using PCA and were employed instead of the concentrations to compute the cumulative amounts and steady fluxes, the latter of which were considered as the indexes for optimizing penetration enhancers. The results showed that compared to the control group, the steady fluxes of the other groups increased significantly and furthermore, 4% azone with 1% propylene glycol manifested the best effect. The six components could penetrate through skin well under the action of penetration enhancers. The method established in this study has been proved to be suitable for the study of transdermal delivery of multiple components, and it provided a scientific basis for preparation research of Xiangfusiwu decoction and moreover, it could be a reference for Chinese medicine research.
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June 2013

[Study on evaluation system for gynecological disease model characterized by Qi stagnation and blood stasis syndrome with female SD rats].

Zhongguo Zhong Yao Za Zhi 2013 Apr;38(8):1211-8

Jiangsu Key Laboratory for High Technology Research of Traditional Chinese Medicine Formulae, Nanjing University of Chinese Medicine, Nanjing 210046, China.

Objective: To establish an evaluation system for animal model with gynecological disease characterized by Qi stagnation and blood stasis syndrome, in order to disprove syndrome characteristics of the model by classic clinical prescriptions, and evaluate the specificity and reliability of the model with macroscopic biological signs and symptoms.

Method: The model characterized by Qi stagnation and blood stasis syndrome was established by injecting adrenaline into female SD rats and conducting unpredictable chronic stimulus such as reversal of day and night, swimming in cold water, thermal stimulation in oven, noise and tail suspension for two weeks. They were also orally administered with Xiangfu Siwu Tang, Shaofu Zhuyu Tang and positive control drug aspirin in groups. A comprehensive evaluation was conducted for the model on the basis of haemorheology, four blood coagulation indexes, four diagnostic information (digital imaging of tongue, paw and tail, temperature, weight, ingestion, electrocardiograph, and open filed test), and syndrome rating.

Result: Compared with the normal group, the model group showed obvious changes in haemorheology, four blood coagulation indexes, animal behavior, weight, ingestion, syndrome rating and heart rate. Their tongue and paw pictures were analyzed with Photoshop 7.0, showing significant difference in red, green and blue percentage composition from the normal group. Groups given aspirin and Xiangfu Siwu Tang showed notable changes in haemorheology, four blood coagulation indexes, animal behavior, weight, ingestion, heart rate, syndrome rating, and red, green and blue percentage composition in tongue and paw pictures, whereas the group given Shaofu Zhuyu Tang showed no remarkable improvement.

Conclusion: The evaluation system for the animal model with gynecological disease characterized by Qi stagnation and blood stasis syndrome is established to provide reference for studies on the evaluation system for qi stagnation and blood stasis syndrome models.
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April 2013

UPLC-Q-TOF/MS coupled with multivariate statistical analysis as a powerful technique for rapidly exploring potential chemical markers to differentiate between radix paeoniae alba and radix paeoniae rubra.

Nat Prod Commun 2013 Apr;8(4):487-91

Jiangsu Key Laboratory for High Technology of TCM Formulae Research, Nanjing University of Chinese Medicine, 210023 Nanjing, P R China.

To explore rapidly the potential chemical markers for differentiating Radix Paeoniae Alba and Radix Paeoniae Rubra, a method is proposed based on ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) coupled with multivariate statistical analysis. Batches of commercial samples were analyzed by UPLC-Q-TOF/MS. The datasets of t(R)-m/z pair, ion intensities and sample codes were further processed with orthogonal partial least squared discriminant analysis (OPLS-DA) to compare holistically the difference between these two kinds of samples. Then statistics were used to generate an S-plot, in which the variables (t(R)-m/z pair) contributing most to the difference were clearly depicted as points at the two ends of "S", and the components correlated to these ions should be regarded as the chemical markers. The identities of the most changed markers can be identified by comparing the mass/UV spectra and retention times with those of reference compounds and/or tentatively assigned by matching empirical molecular formulae with those of known compounds published in the literature. Using this proposed approach, albflorin, paeoniflorin, oxypaeoniflorin, benzoylpaeoniflorin, galloylalbiflorin and paeoniflorigenone were found to be the differentiating components for discrimination of Radix Paeoniae Alba and Radix Paeoniae Rubra. Moreover, paeoniflorin sulfonate and its isomer, isomaltopaeoniflorin sulfonate, were found to be the characteristic markers for all Radix Paeoniae Alba samples that were processed by sulfurdioxide gas fumigation. The results suggested that this newly established approach could be used to explore rapidly the potential chemical markers for herbs with similar chemical characteristics.
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April 2013
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