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italic>Salvia miltiorrhiza Bunge is a traditional Chinese medicinal herb widely used to treat cardiovascular and cerebrovascular diseases at clinic. Its main water-soluble components are rosmarinic acid (RA) and salvianolic acid B (SAB), which are produced by phenylpropanoid pathway. 4-Hydroxyphenylpyruvate reductase (HPPR) is a key enzyme in phenylpropanoid metabolism pathway. SmHPPR1 was cloned from S. miltiorrhiza and was constructed into plant expression vector pJR-SmHPPR1. On this basis, SmHPPR1 transgenic Arabidopsis plants were induced and the content of 4-hydroxyphenyllactic acid (pHPL) was determined. SmHPPR1-overexpressing (SmHPPR1-OE) hairy roots of S. miltiorrhiza were obtained and the concentration of active components and transcriptome analysis were performed. The results showed that the concentration of pHPL in SmHPPR1 transgenic Arabidopsis T1 was 0.594 mg·g-1 dry weight. The concentration of RA, SAB and total salvianolic acid in SmHPPR1-OE-3 hairy roots were 1.09, 1.29, 1.15 times of that in control-3, respectively, and the content of Danshensu was 36.26% of that in control-3. Transcriptomic analysis revealed that overexpression of SmHPPR1 caused the upregulation of other phenylpropanoid pathway genes like SmTAT2. Protein-protein interaction indicated CYT (TR74706_c0_g1), NADP+ (TR26565_c0_g1) and NADP+ (TR68771_c0_g1) is the central node of the network and participated in metabolic process and cellular process. The tracking work in this study proved that SmHPPR1 could catalyze the reduction of 4-hydroxyphenylpyruvic acid to 4-hydroxyphenyllactic acid in SmHPPR1 transgenic Arabidopsis, and SmHPPR1-overexpressing in hairy roots of S. miltiorrhiza could increase the concentration of salvianolic acids through synergistically regulating other pathway genes.
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Traditional Chinese medicines (TCMs), with a history of thousands of years, are widely used clinically with effective treatment. However, the drug delivery systems (DDSs) for TCMs remains major challenges due to the characteristics of multi-components including alkaloids, flavones, anthraquinones, glycosides, proteins, volatile oils and other types. Therefore, the novel preparations and technology of modern pharmaceutics is introduced to improve TCM therapeutic effects due to instability and low bioavailability of active ingredients. Salviae Miltiorrhizae Radix et Rhizoma, the radix and rhizomes of Salvia miltiorrhiza Bunge (Danshen in Chinese), is a well known Chinese herbal medicine for protecting the cardiovascular system, with active ingredients mainly including lipophilic tanshinones and hydrophilic salvianolic acids. In this review, this drug is taken as an example to present challenges and strategies in progress of DDSs for TCMs. This review would also summary the characteristics of active ingredients in it including physicochemical properties and pharmacological effects. The purpose of this review is to provide inspirations and ideas for the DDSs designed from TCMs by summarizing the advances on DDSs for both single- and multi-component from Danshen.
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Objective: To explore the mechanism of Salvia miltiorrhiza in treatment of microcirculatory disturbance based on network pharmacology. Methods: The targets of S. miltiorrhiza’s active components for treatment of microcirculatory disturbance were screened and predicted by utilizing TCMSP, PubChem Search, Genecards database and Swiss target prediction online tool. Cytoscape 3.3.0 software was adopted to construct an active component-microcirculatory disturbance target network. The protein-protein interaction (PPI) network was established by using STRING database. DAVID database was used to analyze metabolism pathway in target gene ontology (GO) biological process, Kyoto encyclopedia of genes and gnomes (KEGG). Results: Totally 65 active components of S. miltiorrhiza and nine related targets were screened. GO and KEGG pathway enrichment analysis revealed that active components of S. miltiorrhiza participated in oxidation-reduction process, cellular calcium ion homeostasis and other biological processes, and S. miltiorrhiza may regulate VEGF signaling pathway, cholinergic synapse signal transduction, oxytocin signaling pathway, aldosterone-regulated sodium reabsorption pathway and so on. Conclusion: This study reflects the characteristics of multi-components, multi-targets, and multi-pathways of S. miltiorrhiza in the treatment of microcirculation disturbance, which may provide new ideas and methodology for further research on the treatment of microcirculatory disturbance using S. miltiorrhiza.
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Objective: Nitrogen is an important element affecting the accumulation of effective components in Chinese medicinal materials. The purpose of this study was to investigate effects of different nitrogen sources on the growth and active components accumulation of hairy roots of Salvia miltiorrhiza and Salvia castanea f. tomentosa. Methods: The hairy roots of S. miltiorrhiza and S. castanea f. tomentosa were treated with ammonium nitrate, hydrolyzed milk protein, peptone, beef extract, casein and yeast extract, respectively. The growth of hairy roots and the accumulation of active components were analyzed. Results: Ammonium nitrate was the most beneficial to the growth of the two kinds of hairy roots. Hydrolyzed milk protein significantly promoted the accumulation of salvianolic acids, compared with ammonium nitrate, the contents of rosmarinic acid and salvianolic acid B in S. miltiorrhiza were respectively increased by 2.94 times and 3.27 times, and the contents of rosmarinic acid and salvianolic acid B in S. castanea f. tomentosa were respectively increased by 13.74 times and 2.01 times. Yeast extract had the most significant effect on the accumulation of dihydrotanshinone I and cryptotanshinone in two kinds of hairy roots. Hydrolyzed milk protein significantly promoted the accumulation of tanshinone IIA in the hairy roots of S. miltiorrhiza, while beef extract had the most significant effect on the accumulation of tanshinone IIA in S. castanea f. tomentosa. Conclusion: Ammonium nitrate was the best nitrogen source for the growth of two kinds of hairy roots, and hydrolyzed milk protein was the best nitrogen source for salvianolic acids accumulation. The effects of different nitrogen sources on four kinds of tanshinones were different, and the responses of S. miltiorrhiza and S. castanea f. tomentosa to different nitrogen sources were also different. This study not only has certain guiding significance for large-scale cultivation of hairy roots of S. miltiorrhiza and industrialized production of active components, but also provides a reference for the development and utilization of S. castanea f. tomentosa resources.
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Objective: To summarize the prescription rules of traditional Chinese medicine (TCM) for treating alcoholic liver disease (ALD) based on TCM inheritance support system (V2.50). Methods: The literatures about TCM prescriptions for treating ALD were collected from CNKI, Wanfang database, and VIP database. The TCM inheritance platform system was used to analyze the prescription rules of TCM in the treatment of alcoholic liver disease. Results: Statistics showed that the majority of prescriptions were used to treat alcoholic fatty liver, alcoholic hepatitis, and alcoholic cirrhosis. Through "frequency statistics" analysis, 107 prescriptions were found involving 149 flavors of TCM, with a cumulative frequency of 1 195 times. Twenty-three Chinese medicines with a frequency of ≥ 15 times were used, and the cumulative frequency was 737 times (62%). The most frequently used medicines were blood-activating and stasis-removing drugs, water-diffusing and damp-permeating drugs, tonics, heat-clearing drugs, antialcoholic poisons and qi-regulating drugs. The commonly used doses of Salvia miltiorrhiza, Poria cocos, Bupleurum chinense, Glycyrrhiza uralensis, Atractylodes macrocephala, Alismatis Rhizoma, and Curcumae Radix in the top 10 medicines ranked in the frequency of medication accorded with the prescribed doses in the Pharmacopoeia of the People's Republic of China (2015 edition), while Crataegi Fructus, Artemisiae Scopariae Herba, and Puerariae Lobatae Radix exceeded the prescribed doses. In the frequency analysis of drug pairs, the combination of S.miltiorrhiza and B. chinense was the most widely used. According to the association rules of drug combination, the correlation between Curcumae Radix and S. miltiorrhiza was the strongest, that was, the probability of S. miltiorrhiza appearing with the emergence of Curcumae Radix was 88%. From the network display chart, it was indicated that S. miltiorrhiza and P. cocos were the main herbs for treatment. Through unsupervised entropy hierarchical clustering algorithm, 14 core combinations for new clustering were extracted, and seven new prescriptions can be obtained by further clustering. Conclusion: The basic principles of TCM treatment of ALD include promoting blood circulation and removing blood stasis, removing dampness, tonifying, detoxifying alcohol, and promoting qi, and with "protecting spleen and stomach function" as its purpose, which accords with the theoretical basis of traditional Chinese medicine in treating alcoholic liver disease. Core combinations and new prescriptions provide references for clinical drug use and new drug research and development, but new prescriptions must be further evaluated with the combination of traditional Chinese medicine theory and clinical practice.
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Objective: To analyze the rules of coronary heart disease syndrome differentiation based on the data mining technology. Methods: First of all, the famous medical records and prescriptions were collected and normalized to construct the database of coronary heart disease prescriptions. Then, IBM SPSS Statistics 20.0, a statistical software, was used to conduct statistics on the distribution of drug frequency, disease site syndrome frequency and disease syndrome frequency. Finally, the Apriori algorithm was applied to carry out data mining and analyze the underlying law of drug compatibility. Results: A total of 145 prescriptions were selected and analyzed, including 216 Chinese medicines, eight syndromes of disease location and 12 syndromes of disease nature. Forty-six herbs with higher frequency were found, and the top five TCM herbs were Salvia miltiorrhiza, Glycyrrhiza uralensis, Trichosanthes kirilowii, Pinellia ternata and Ligusticum chuanxiong. The main syndromes of disease location were heart, kidney, spleen and liver, while the main syndromes of disease nature were blood stasis, qi deficiency, phlegm, deficiency of yang, qi stagnation, and deficiency of yin. When the minimum support was 15% and the minimum confidence coefficient was 70%, the association rule analysis results showed that a total of 15 association rules for two-herb-combinations and 26 association rules for the trigeminy medications were obtained. The most frequency two-herb-combinations were Allium macrostemon→T. kirilowii", "Schisandra chinensis→Ophiopogon japonicus", "Curcumae Radix→S. miltiorrhiza. The most frequently used trigeminy medications were A. macrostemon + P. ternata→T. kirilowii, S. miltiorrhiza + A. macrostemon→T. kirilowii and A. macrostemon + G. uralensis→T. kirilowii. Conclusion: The syndrome differentiation and medication of TCM in the treatment of coronary heart disease mainly focus on invigorating the circulation of blood and resolving stasis, moving qi, nourishing and replenishing qi, clearing up phlegms. It is consistent with the main syndromes of the disease nature, such as blood stasis and qi deficiency, and the herbs mostly return to the heart meridian, which is consistent with the main syndromes of the disease location. The rules of syndrome differentiation and medication of based on the data mining technique have great value for clinical medication guidance and application to analyze the prescription for coronary heart disease.
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Objective: Network pharmacology method was adopted in this study to explore the active compounds and mechanism of Salvia miltiorrhiza for cirrhosis. Methods: TCMSP database was utilized to obtain the active components of S. miltiorrhiza. Through GeneCards, OMIM and DRAR-CPI, the potential targets of S. miltiorrhiza for the treatment of cirrhosis were screened. Cytoscape 3.6.0 software was established to construct the active components-targets network of S. miltiorrhiza. STRING database and Generate style from statistics of Cytoscape 3.6.0 software were conducted to draw a graph of protein interaction network. Molecular docking was carried out through Systems Dock Web Site with the active components of S. miltiorrhiza. The GO classified enrichment analysis and the KEGG pathway enrichment analysis were performed by using DAVID database. Results: Selecting the OB ≥ 30% and DL ≥ 0.18 as filter condition, 65 active components and 75 targets of S. miltiorrhiza were involved. S. miltiorrhiza exerted its effects on treating cirrhosis mainly by regulating signaling pathways including MAPK, Toll-like receptor, Gap junction, PI3K/AKT, Natural killer cell mediated cytotoxicity signaling pathway and so on. Conclusion: This study preliminarily predicted the major targets and pathways of S. miltiorrhiza acting on cirrhosis, which provided new ideas and clues for its further research.
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Objective: Salvia miltiorrhiza is a valuable herbal medicine with tanshinone and phenolic acid as the main biological active ingredients. The biosynthetic regulation of these bioactive compounds is controlled by a set of transcription factors (TFs). The basic helix-loop-helix (bHLH) transcription factor plays an important role in various physiological and biochemical processes in plants. However, research on bHLH TFs regulating phenolic acid or tanshinone biosynthesis in S. miltiorrhiza is limited. Methods: qRT-PCR was used for gene expression analysis. The subcellular localization of SmbHLH92 was detected by SmbHLH92-GFP transient transformation into tobacco leaves, and its fluorescence was observed using a confocal laser scanning microscope. The transcriptional activity of SmbHLH92 was confirmed in the AH109 yeast strain. RNA interference hairy roots of SmbHLH92-RNAi transgenic lines were obtained through Agrobacterium-mediated genetic transformation. Ultra performance liquid chromatography (UPLC) was used to detect the changes of phenolic acids and tanshinones. Results: SmbHLH92 is a bHLH transcription factor that is highly expressed in the root and phloem of S. miltiorrhiza. The subcellular localization and transcriptional activity of SmbHLH92 indicated that SmbHLH92 was located in the nucleus and may be a transcription factor. RNA interference (RNAi) of SmbHLH92 in hairy roots of S. miltiorrhiza significantly increased the accumulation of phenolic acid and tanshinone. Quantitative RT-PCR (RT-qPCR) analysis showed the transcription level of genes encoding the key enzymes involved in the phenolic acid and tanshinone biosynthetic pathways was increased in the hairy roots of the SmbHLH92-RNAi transgenic line, comparing with the control line. Conclusion: These data indicate that SmbHLH92 is a negative regulator involved in the regulation of phenolic acid and tanshinone biosynthesis in S. miltiorrhiza.
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OBJECTIVE@#To investigate the antiproliferative activity of Salvia miltiorrhiza Bunge. (SM) on the castration-resistant prostate cancer (CRPC) cell line DU-145, in vitro and in vivo.@*METHODS@#Prostate cancer cell line (DU-145) and normal prostate cell line (RWPE-1) were treated with SM at different concentrations (3.125, 12.5, 25 and 50 μg/mL) to investigate the antiproliferative effects. DNA laddering analysis was performed to investigate the apoptosis of DU-145 cells. Molecular mechanism was investigated by Western blot analysis of p53, Bcl-2, prostate specific antigen (PSA), and androgen receptor (AR). Six-week-old male BALB/c nude mice were randomly divided into normal control group (n=101) and treated group (n=101) which administered 500 mg/kg SM for 2 weeks. Tumor volumes were measured.@*RESULTS@#Treatment with SM resulted in a dose-dependent decrease in cell number of DU-145 cells in comparison with RWPE-1. DNA laddering analysis indicated the apoptosis of DU-145 cells. Treatment with SM increased the expression of p53 and reduced the expression of Bcl-2 proteins. The levels of PSA were considerably reduced in SM-treated group compared to the controls, and a decrease in AR expression was observed when cells were treated with SM in the same pattern as a reduction in PSA. In the tumour xenograft study, SM given once a day for 2 weeks significantly inhibited tumour growth.@*CONCLUSION@#SM might contribute to the anticancer actions such as induction of apoptosis and inhibition of proliferation of prostate cancer cells.
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Objective: The exogenous gibberellin (GA) and ethylene (ET) treatment can improve the medicinal ingredients of Salvia miltiorrhiza. Interestingly, many reports pointed out that WRKY transcription factors played an important regulatory role in these treatment responses. However, whether the SmWRKY mediate these treatment signalings in S. miltiorrhiza remains largely elusive. Methods: qRT-PCR was used for SmWRKY42-like in response to exogenous GA and ethephon (Eth) treatment. The subcellular location of SmWRKY42-like was transiently transformed into onion epidermal cells by particle bombardment. The self-activating activity of SmWRKY42-like was verified in AH109 yeast strain. Results: SmWRKY42-like was a WRKY family gene in S. miltiorrhiza. The subcellular localization and transcriptional activity results of the SmWRKY42-like protein indicated that SmWRKY42-like mainly enriched in nucleus and might be a transcription factor in S. miltiorrhiza. In the meantime, the SmWRKY42-like gene significantly responded to exogenous GA and Eth treatment. Conclusion: These results collectively indicated the SmWRKY42-like gene functions, as an important hormone-responsive gene, might play a potentially role in ET and GA signaling pathways.
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Objective To provide a basis for further study and utilization of the functional gene family of Pht1 in Salviae miltiorrhizae, the Pht1 gene family members from S. miltiorrhizae genome should be explored, identified and functionally predicted. Methods By utilizing the bioinformatics methods, based on the genomic and NCBI database of S. miltiorrhiza, the sequences of Pht1 family candidate genes of S. miltiorrhiza were obtained firstly by screening, and then carried on the multiple sequence alignment, conserved structure lookup as well as phylogenetic tree construction. Finally, the characteristic analysis and biological function prediction were carried out. Results The results showed that 12 Pht1 candidate genes were extracted from the genome of Salvia miltiorrhiza, together with the reported Smpht1, there are 13 Pht1 members in total, their homology as high as 74.34%, and nine members of them could provide more reliable functional annotation. Among the gene family members, Smpht1 & Sm1, Sm4 & Sm6, Sm5 & Sm11, in which these three pairs may share similar function as well as coevolution relation between each other; Smpht1, Sm1, Sm3, Sm5, Sm7, and Sm11, all of the six members can be only expressed in the roots of S. miltiorrhiza. furthermore, except Sm7 induced by p-deficiency and Sm3 member unaffected by the phosphorus supply factor, the other four probably belong to the P-deficiency induced up-regulated expressive genotype, which play an important role in phosphorus absorption of S. miltiorrhiza; Moreover, both Sm7 and Sm16 are possible to expressed in the flower organs of S. miltiorrhiza and then participate in the regulation of phosphorus balance in the flowering stage of plants. In addition, Sm14 is likely to be a mycorrhizal-induced specific expressive gene with the function of phosphorus transfer in Pht1 family members. Conclusion The Pht1 gene family was screened and identified systematically from the genome of S. miltiorrhiza for the first time, in addition, the members with characteristic and biological function were found out, which will provide dependable background support and ideas reference for the further research on the role of Pht1 in the growth and development of S. miltiorrhiza.
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Objective: The potency of multi-source information fusion technology was explored to improve the model calibration and prediction performance of Chinese medicine extraction process. Methods: The ethanol extraction process for isolating fat-soluble components from Salvia miltiorrhiza was taken as the research carrier. S. miltiorrhiza from different sources were collected to simulate the fluctuation of materials. The changes of process parameters were simulated by design of experimental (DOE), and the process near infrared spectra (NIRS) were used as the process state variables. The contents of tanshinone IIA, cryptotanshinone, and tanshinone I were determined by HPLC. The raw material properties, process parameters and process state variables were combined as independent variables. The content of effective components in the extract was taken as the dependent variable. The partial least squares (PLS) algorithm was used to establish the quality prediction model of the extracts. Results: The modeling results respectively showed that the RMSECV was 0.172 8 mg/g, RMSEP was 0.031 7 mg/g, RPD was 6.91 (tanshinone IIA); RMSECV was 0.153 4 mg/g, RMSEP was 0.024 2 mg/g, RPD was 4.02 (cryptotanshinone); RMSECV was 0.117 1 mg/g, RMSEP was 0.043 2 mg/g, RPD was 4.76 (tanshinone I). Conclusion: The calibration and prediction performance of multi-source information fusion model are better than the conventional model, which can effectively improve the quality predictability and controllability of S. miltiorrhiza extract.
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Objective To explore the protective effects of Salvia miltiorrhiza and Carthamus tinctorius active ingredients in different compatibility on cerebral ischemia-reperfusion injury in rats. Methods Adult male Sprague-Dawley (SD) rats were selected to establish a cerebral ischemia-reperfusion (I/R) injury model by middle cerebral artery occlusion (MCAO) method. The rats were randomly divided into sham operation group, model group, and positive control group (Danhong Injection 2 mL/kg); And orthogonal test method L9(34) was adopted to compose nine compatibility groups from main active ingredients of S. miltiorrhiza and C. tinctorius (trashinol, salvianolic acid A, salvianolic acid B, and HYSA). Rats were tail iv administrated once daily for continuous 3 d. Score neurological deficit was evaluated after 3 d; qRT-PCR was used to detect mRNA expression of glucose regulated protein 78 (GRP-78), nuclear factor-κB (NF-κB), C/EBP homologous protein (CHOP), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6); HE staining was used to detect the pathological changes of cerebral cortex, and the expression of NF-κB p65 protein in cerebral cortex was detected by immunohistochemistry. Results Compared to the model group, Danhong Injection group and orthogonal compatibility nine groups significantly reduced the neurological deficit scores of rats with cerebral ischemia-reperfusion; increased the mRNA expression of GRP-78 in the cerebral cortex; decrease the mRNA expression of NF-κB, CHOP, TNF-α, and IL-6, and decrease the protein expression of NF-κB p65 in the cerebral cortex. The results also showed that the protective effects of the 4th group (danshensu 30 mg/kg, salvianolic acid A 2.5 mg/kg, salvianolic acid B 16 mg/kg, and HYSA 8 mg/kg), 6th group (trashinol 30 mg/kg, salvianolic acid A 10 mg/kg, salvianolic acid B 8 mg/kg, and HYSA 4 mg/kg) in the 9th group were more significant for cerebral ischemia reperfusion injury in rats. Conclusion The active components of S. miltiorrhiza and C. tinctorius can play a good protective role in endoplasmic reticulum stress and inflammation in rats with cerebral ischemia reperfusion injury.
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Objective To clone the R2R3 MYB transcription factor gene SmMYB87 in subgroup 14 from Salvia miltiorrhiza, and analyze the bioinformatics and expression of this gene. Methods Total RNA extracted from S. miltiorrhiza was used as cDNA synthesis template and the full length cDNA sequence was obtained through homology-based cloning and rapid amplification of cDNA ends (RACE) technology. The structure and physicochemical properties of SmMYB87 gene and its coded protein were analyzed with bioinformatics softwares. The expression of SmMYB87 in different organs was determined with qRT-PCR, and a GFP fusion expression vector was constructed to investigate the subcellular laicization of SmMYB87 protein. Results SmMYB87 gene contained two introns and an open reading frame (ORF) of 732 bp, encoding 243 amino acid polypeptides. It expressed in roots, stems, leaves and flowers with similar expression levels and the SmMYB87 protein located in nucleus and cytomembrane. Conclusion The analysis of sequence structure and expression pattern of SmMYB87 will be helpful to study the regulating roles of this gene in S. miltiorrhiza.
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Abstract One bioactive compound, identified as alternariol 9-methyl ether, was isolated from the crude extract of the endophytic fungus Alternaria sp. Samif01 residing in the roots of Salvia miltiorrhiza Bunge. Alternariol 9-methyl ether was active against bacteria with minimum inhibitory concentration values ranging from 25 to 75 µg/mL and median inhibitory concentration (IC50) values ranging from 16.00 to 38.27 µg/mL. The IC50 value of alternariol 9-methyl ether against spore germination of Magnaporthe oryzae was 87.18 µg/mL. Alternariol 9-methyl ether also showed antinematodal activity against Bursaphelenchus xylophilus and Caenorhabditis elegans with IC50 values of 98.17 µg/mL and 74.62 µg/mL, respectively. This work is the first report on alternariol 9-methyl ether and its biological activities from the endophytic fungus Alternaria sp. Samif01 derived from S. miltiorrhiza Bunge. The results indicate the potential of Alternaria sp. Samif01 as a source of alternariol 9-methyl ether and also support that alternariol 9-methyl ether is a natural compound with high potential bioactivity against microorganisms.
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Animals , Alternaria/chemistry , Anti-Infective Agents/isolation & purification , Endophytes/chemistry , Lactones/isolation & purification , Alternaria/isolation & purification , Anti-Infective Agents/metabolism , Bacteria/drug effects , Endophytes/isolation & purification , Lactones/metabolism , Microbial Sensitivity Tests , Magnaporthe/drug effects , Nematoda/drug effects , Plant Roots/microbiology , Salvia/microbiologyABSTRACT
In recent years, the Chinese medicine Salvia miltiorrhiza has been widely used in cardiovascular and cerebrovascular diseases. So, the interaction between S. miltiorrhiza and cytochrome P450s are focused increasingly. This review summarized the research progress on the cytochrome P450s-based metabolism and interaction of S. miltiorrhiza, including its extracts, major individuals, and S. miltiorrhiza-based formulas. It systematically summarized the information which were useful for the further applications and researches on S. miltiorrhiza and its active components.
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Objective: To investigate the effect of water-soluble components from Salvia miltiorrhiza on liver sinusoid endothelial cell function and angiogenesis. Methods: Different dosages of water-soluble components from S. miltiorrhiza were incubated for 24 h with HHSEC cell line, and the toxicity of HHSEC cells was assayed by CCK-8 method. The proliferation of HHSEC cells was induced by endothelial cell growth supplements (ECGS), with receptor tyrosine kinase inhibitor sorafenib as positive control, cell proliferation was analyzed by EdU DNA cell proliferation kit; Fluorescence probe method was used to assay the intracellular NO level and NOS activity; Immunofluorescence method was performed to observe the expression of CD31; The transgenic zebrafishes were incubated for 24 h with each component. The fluorescence vessels, the number of functional blood vessels, and intersegmental vessel changes were observed; Chicken chorioallantoic membranes were incubated for 24 h with each component, Image Analysis Software was used to analyze the vessel area changes. Results: Compared with control group, the proliferation of HHSEC cells induced by ECGS increased, the level of NO and NOS activity reduced and the expression of CD31 increased; Compared with the model group, salvianolic acid A, salvianolic acid B, salvianolic acid C, and sorafenib inhibited the proliferation of HHSEC cells induced by ECGS, reduced the level of NO, NOS activity, and expression of CD31. The vessel area of chicken chorioallantoic membranes was decreased in sorafenib, salvianolic acid A, salvianolic acid B and caffeic acid. Salvianolic acid C also significantly inhibited the intersegmental vessel changes of transgenic zebrafish. Conclusion: Salvianolic acid A, salvianolic acid B, and salvianolic acid C have the potential effects on function of endothelial cell proliferation and angiogenesis.
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Tanshinones and phenolic acids are two major classes of effective compounds in S.miltiorrhizae Radix et Rhizoma.Accumulative levels of these compounds directly determined its quality and efficacy.Presently,for the purpose of understanding the underlying the mechanism behind the biosynthesis and regulation of effective compounds in S.miltiorrhiza,a series of progresses have been pressed ahead.Benefiting from the boost and convenience of high-throughput genomic,transcriptomic,and metabolic analysis methods,remarkable achievements have been accessed in this field.In this paper,a summary for the recent progress have been made with the provision of a reference for better understanding over the quality of S.miltiorrhiza.
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To optimize the solvent system of high-speed counter-current chromatography (HSCCC) for diterpenoidtanshinone separation and define it's antitumor activity in vitro. Total diterpenoidtanshinone was made by CO2 supercritical fluid extraction (SFE), UPLC was applied to determining the peak area of tanshinone IIA and cryptotanshinone in different solvents up or under phase, and the partition coefficient K values of them were calculated. CKK-8 was used to observe the inhibitory effects of diterpenoidtanshinone on human liver cancer (QGY-7703), lung cancer (PC9, A549), gastric cancer (MKN-45, HGC-27), colon cancer (HCT116), myeloma (U266, RPMI8226), and breast cancer (MCF-7). The best solvent system for HSCCC was petroleum ether-ethyl acetate-methanol-aqua (12∶8∶ 13∶7). The yield of diterpenoidtanshinone was 8.65%. Diterpenoidtanshinone has good effect of antitumor in vitro especially on human PC9 cell. The selected solvent system is suitable for diterpenoidtanshinone separation by HSCCC, and the established HSCCC method is reliable and easy for operating. Diterpenoidtanshinone has good antitumor effect in vitro.
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Objective: To clone and characterize a late embryogenesis abundant protein SmLEA2 with its promoter region from Salvia miltiorrhiza, and to predict its probable function. Methods: SmLEA2 was cloned by PCR and RT-PCR from genomic DNA and cDNA. Protein structure and phylogenetic relationships were carried out by bioinformatic analysis. Gene expression in different organs and different development periods was detected by qPCR. Gene expression was also detected under different treatments. Results: By analyzing the cDNA library for S. miltiorrhiza with BLAST program, one of these sequences showed a high homology with late embryogenesis abundant (LEA) protein and was named as SmLEA2 (GenBank: HQ676610). We obtained 1 961 bp gene sequences of SmLEA2, which contained an intron and a single opening reading frame (ORF) of 960 bp encoding 319 amino acid peptides. Bioinformatic analysis showed that the putative SmLEA2 protein was a hydrophilic protein without signal peptides and transmembrane domains. The SmLEA2 protein was predicted with a molecular weight of 35 340 and a theoretical isoelectric point of 4.77. SmLEA2 was expressed in the roots, stems, and leaves of S. miltiorrhiza, and the most abundant in the stems. With the development of the flowers and seeds, its expression increased gradually. Expression of SmLEA2 could be induced by methyl jasmonate (MeJA) and abscisic acid (ABA). Conclusion: Thus, we speculate that SmLEA2 may be involved in seed development and plant defenses.