The SmMYC2-SmMYB36 complex is involved in methyl jasmonate-mediated tanshinones biosynthesis in Salvia miltiorrhiza.

The jasmonic acid (JA) signaling pathway plays an important role in promoting the biosynthesis of tanshinones. While individual transcription factors have been extensively studied in the context of tanshinones biosynthesis regulation, the influence of methyl jasmonate (MeJA)-induced transcriptional complexes remains unexplored. This study elucidates the positive regulatory role of the basic helix-loop-helix protein SmMYC2 in tanshinones biosynthesis in Salvia miltiorrhiza. SmMYC2 not only binds to SmGGPPS1 promoters, activating their transcription, but also interacts with SmMYB36. This interaction enhances the transcriptional activity of SmMYC2 on SmGGPPS1, thereby promoting tanshinones biosynthesis. Furthermore, we identified three JA signaling repressors, SmJAZ3, SmJAZ4, and SmJAZ8, which interact with SmMYC2. These repressors hindered the transcriptional activity of SmMYC2 on SmGGPPS1 and disrupted the interaction between SmMYC2 and SmMYB36. MeJA treatment triggered the degradation of SmJAZ3 and SmJAZ4, allowing the SmMYC2-SmMYB36 complex to subsequently activate the expression of SmGGPPS1, whereas SmJAZ8 inhibited MeJA-mediated degradation due to the absence of the LPIARR motif. These results demonstrate that the SmJAZ-SmMYC2-SmMYB36 module dynamically regulates the JA-mediated accumulation of tanshinones. Our results reveal a new regulatory network for the biosynthesis of tanshinones. This study provides valuable insight for future research on MeJA-mediated modulation of tanshinones biosynthesis.

Synthetic consortia of four strains promote Schisandra chinensis growth by regulating soil microbial community and improving soil fertility.

MAIN CONCLUSION: Synthetic consortia performed better in promoting Schisandra chinensis growth than individual strains, and this result provides valuable information for the development of synthetic microbial fertilizers. Schisandra chinensis is an herbal medicine that can treat numerous diseases. However, the excessive reliance on chemical fertilizers during the plantation of S. chinensis has severely restricted the development of the S. chinensis planting industry. Plant growth-promoting rhizobacteria (PGPR) can promote the growth of a wide range of crops, and synthetic consortia of them are frequently superior to those of a single strain. In this study, we compared the effects of four PGPR and their synthetic consortia on S. chinensis growth. The pot experiment showed that compared with the control, synthetic consortia significantly increased the plant height, biomass, and total chlorophyll contents of S. chinensis, and their combined effects were better than those of individual strains. In addition, they improved the rhizosphere soil fertility (e.g., TC and TN contents) and enzyme activities (e.g., soil urease activity) and affected the composition and structure of soil microbial community significantly, including promoting the enrichment of beneficial microorganisms (e.g., Actinobacteria and Verrucomicrobiota) and increasing the relative abundance of Proteobacteria, a dominant bacterial phylum. They also enhanced the synergistic effect between the soil microorganisms. The correlation analysis between soil physicochemical properties and microbiome revealed that soil microorganisms participated in regulating soil fertility and promoting S. chinensis growth. This study may provide a theoretical basis for the development of synthetic microbial fertilizers for S. chinensis.

SmJAZs-SmbHLH37/SmERF73-SmSAP4 module mediates jasmonic acid signaling to balance biosynthesis of medicinal metabolites and salt tolerance in Salvia miltiorrhiza.

Salvia miltiorrhiza holds significant importance in traditional Chinese medicine. Stress-associated proteins (SAP), identified by A20/AN1 zinc finger structural domains, play crucial roles in regulating plant growth, development, resistance to biotic and abiotic stress, and hormone responses. Herein, we conducted a genome-wide identification of the SAP gene family in S. miltiorrhiza. The expression analysis revealed a significant upregulation of SmSAP4 under methyl jasmonate (MeJA) and salt stress. Overexpressing SmSAP4 in S. miltiorrhiza hairy roots increased tanshinones content while decreasing salvianolic acids content, while RNAi-silencing SmSAP4 had the opposite effect. SmSAP4 overexpression in both Arabidopsis thaliana and S. miltiorrhiza hairy roots decreased their salt stress tolerance, accompanied by increased activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), and a hindered ability to maintain the Na+ : K+ ratio. Further investigations demonstrated that MeJA alleviated the inhibitory effect of SmJAZ3 on SmSAP4 activation by SmbHLH37 and SmERF73. However, MeJA did not affect the inhibition of SmSAP4 activation by SmJAZ8 through SmbHLH37. In summary, our research reveals that SmSAP4 negatively regulates the accumulation of salvianic acid through the SmJAZs-SmbHLH37/SmERF73-SmSAP4 module and positively impacting the accumulation of tanshinones. Additionally, it functions as a negative regulator under salt stress.

Salicylic acid regulates phenolic acid biosynthesis via SmNPR1-SmTGA2/SmNPR4 modules in Salvia miltiorrhiza.

Phenolic acids are the main active ingredients in Salvia miltiorrhiza, which can be used for the treatment of many diseases, particularly cardiovascular diseases. It is known that salicylic acid (SA) can enhance phenolic acid content, but the molecular mechanism of its regulation is still unclear. Nonexpresser of PR genes 1 (NPR1) plays a positive role in the SA signaling pathway. In this study, we identified a SmNPR1 gene that responds to SA induction and systematically investigated its function. We found that SmNPR1 positively affected phenolic acid biosynthesis. Then, we identified a novel TGA transcription factor, SmTGA2, which interacts with SmNPR1. SmTGA2 positively regulates phenolic acid biosynthesis by directly up-regulating SmCYP98A14 expression. After double-gene transgenic analysis and other biochemical assays, it was found that SmNPR1 and SmTGA2 work synergistically to regulate phenolic acid biosynthesis. In addition, SmNPR4 forms a heterodimer with SmNPR1 to inhibit the function of SmNPR1, and SA can alleviate this effect. Collectively, these findings elucidate the molecular mechanism underlying the regulation of phenolic acid biosynthesis by SmNPR1-SmTGA2/SmNPR4 modules and provide novel insights into the SA signaling pathway regulating plant secondary metabolism.

Chlorogenic acid inhibits proliferation in human hepatoma cells by suppressing noncanonical NF-κB signaling pathway and triggering mitochondrial apoptosis.

Chlorogenic acid (CGA), a phenylpropanoid derived from Eucommia ulmoides Oliver, has been shown to exhibit potent cytotoxic and anti-proliferative activities against several human cancers. However, the effects of CGA on hepatocellular carcinoma (HCC) and the underlying mechanisms have not been intensively studied. In this study, the CGA treatment effects on the viability of human hepatoma cells were investigated by MTT assay. Our data showed that CGA could dose-dependently inhibit the activity of human hepatoma cells Hep-G2 and Huh-7, but did not affect the activity and growth of normal human hepatocyte QSG-7701. The genes and pathways influenced by CGA treatment were explored by RNA sequencing and bioinformatics analysis, which identified 323 differentially expressed genes (DEGs) involved in multiple pharmacological signaling pathways such as MAPK, NF-κB, apoptosis and TGF-ß signaling pathways. Further analyses by real-time quantitative PCR, Western blot and flow cytometry revealed that CGA effectually suppressed the noncanonical NF-κB signaling pathway, meanwhile it activated the mitochondrial apoptosis of HCC by upregulation of the BH3-only protein Bcl-2 binding component 3 (BBC3). Our findings demonstrated the potential of CGA in suppressing human hepatoma cells and provided a new insight into the anti-cancer mechanism of CGA.

Plasma Membrane H+-ATPase SmPHA4 Negatively Regulates the Biosynthesis of Tanshinones in Salvia miltiorrhiza.

Salvia miltiorrhiza Bunge has been widely used in the treatment of cardiovascular and cerebrovascular diseases, due to the pharmacological action of its active components such as the tanshinones. Plasma membrane (PM) H+-ATPase plays key roles in numerous physiological processes in plants. However, little is known about the PM H+-ATPase gene family in S. miltiorrhiza (Sm). Here, nine PM H+-ATPase isoforms were identified and named SmPHA1-SmPHA9. Phylogenetic tree analysis showed that the genetic distance of SmPHAs was relatively far in the S. miltiorrhiza PM H+-ATPase family. Moreover, the transmembrane structures were rich in SmPHA protein. In addition, SmPHA4 was found to be highly expressed in roots and flowers. HPLC revealed that accumulation of dihydrotanshinone (DT), cryptotanshinone (CT), and tanshinone I (TI) was significantly reduced in the SmPHA4-OE lines but was increased in the SmPHA4-RNAi lines, ranging from 2.54 to 3.52, 3.77 to 6.33, and 0.35 to 0.74 mg/g, respectively, suggesting that SmPHA4 is a candidate regulator of tanshinone metabolites. Moreover, qRT-PCR confirmed that the expression of tanshinone biosynthetic-related key enzymes was also upregulated in the SmPHA4-RNAi lines. In summary, this study highlighted PM H+-ATPase function and provided new insights into regulatory candidate genes for modulating secondary metabolism biosynthesis in S. miltiorrhiza.

Genome-wide characterization and expression profiling of MAPK cascade genes in Salvia miltiorrhiza reveals the function of SmMAPK3 and SmMAPK1 in secondary metabolism.

BACKGROUND: The contribution of mitogen-activated protein kinase (MAPK) cascades to plant growth and development has been widely studied, but this knowledge has not yet been extended to the medicinal plant Salvia miltiorrhiza, which produces a number of pharmacologically active secondary metabolites. RESULTS: In this study, we performed a genome-wide survey and identified six MAPKKK kinases (MAPKKKKs), 83 MAPKK kinases (MAPKKKs), nine MAPK kinases (MAPKKs) and 18 MAPKs in the S. miltiorrhiza genome. Within each class of genes, a small number of subfamilies were recognized. A transcriptional analysis revealed differences in the genes' behaviour with respect to both their site of transcription and their inducibility by elicitors and phytohormones. Two genes were identified as strong candidates for playing roles in phytohormone signalling. A gene-to-metabolite network was constructed based on correlation analysis, highlighting the likely involvement of two of the cascades in the synthesis of two key groups of pharmacologically active secondary metabolites: phenolic acids and tanshinones. CONCLUSION: The data provide insight into the functional diversification and conservation of MAPK cascades in S. miltiorrhiza.

Lipopolysaccharide Enhances Tanshinone Biosynthesis via a Ca2+-Dependent Manner in Salvia miltiorrhiza Hairy Roots.

Tanshinones, the major bioactive components in Salvia miltiorrhiza Bunge (Danshen), are synthesized via the mevalonic acid (MVA) pathway or the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway and the downstream biosynthesis pathway. In this study, the bacterial component lipopolysaccharide (LPS) was utilized as a novel elicitor to induce the wild type hairy roots of S. miltiorrhiza. HPLC analysis revealed that LPS treatment resulted in a significant accumulation of cryptotanshinone (CT) and dihydrotanshinone I (DTI). qRT-PCR analysis confirmed that biosynthesis genes such as SmAACT and SmHMGS from the MVA pathway, SmDXS and SmHDR from the MEP pathway, and SmCPS, SmKSL and SmCYP76AH1 from the downstream pathway were markedly upregulated by LPS in a time-dependent manner. Furthermore, transcription factors SmWRKY1 and SmWRKY2, which can activate the expression of SmDXR, SmDXS and SmCPS, were also increased by LPS. Since Ca2+ signaling is essential for the LPS-triggered immune response, Ca2+ channel blocker LaCl3 and CaM antagonist W-7 were used to investigate the role of Ca2+ signaling in tanshinone biosynthesis. HPLC analysis demonstrated that both LaCl3 and W-7 diminished LPS-induced tanshinone accumulation. The downstream biosynthesis genes including SmCPS and SmCYP76AH1 were especially regulated by Ca2+ signaling. To summarize, LPS enhances tanshinone biosynthesis through SmWRKY1- and SmWRKY2-regulated pathways relying on Ca2+ signaling. Ca2+ signal transduction plays a key role in regulating tanshinone biosynthesis in S. miltiorrhiza.

Physiological and transcriptome analysis reveal molecular mechanism in Salvia miltiorrhiza leaves of near-isogenic male fertile lines and male sterile lines.

BACKGROUND: Our previous study finds that male sterility in Salvia miltiorrhiza could result in stunted growth and reduced biomass, but their molecular mechanisms have not yet been revealed. In this article, we investigate the underlying mechanism of male sterility and its impact on plant growth and metabolic yield by using physiological analysis and mRNA sequencing (RNA-Seq). RESULTS: In this study, transcriptomic and physiological analysis were performed to identify the mechanism of male sterility in mutants and its impact on plant growth and metabolic yield. Through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, it is found that the pathways are mainly enriched in processes including organ development, primary metabolic process and secondary metabolic process. Physiological analysis show that the chloroplast structure of male sterile mutants of S. miltiorrhiza is abnormally developed, which could result in decrease in leaf gas exchange (A, E and gs), chlorophyll fluorescence (Fv, Fm and Fv/Fm), and the chlorophyll content. Expression level of 7 differentially expressed genes involved in photosynthesis-related pathways is downregulated in male sterile lines of S. miltiorrhiza, which could explain the corresponding phenotypic changes in chlorophyll fluorescence, chlorophyll content and leaf gas exchange. Transcriptomic analysis establishes the role of disproportionating enzyme 1 (DPE1) as catalyzing the degradation of starch, and the role of sucrose synthase 3 (SUS3) and cytosolic invertase 2 (CINV2) as catalyzing the degradation of sucrose in the S. miltiorrhiza mutants. The results also confirm that phenylalanine ammonialyase (PAL) is involved in the biosynthesis of rosmarinic acid and salvianolic acid B, and flavone synthase (FLS) is an important enzyme catalyzing steps of flavonoid biosynthesis. CONCLUSIONS: Our results from the physiological and transcriptome analysis reveal underlying mechanism of plant growth and metabolic yield in male sterile mutants, and provide insight into the crop yield of S. miltiorrhiza.

Combining transcriptomics and metabolomics to reveal the underlying molecular mechanism of ergosterol biosynthesis during the fruiting process of Flammulina velutipes.

BACKGROUND: Flammulina velutipes has been recognized as a useful basidiomycete with nutritional and medicinal values. Ergosterol, one of the main sterols of F. velutipes is an important precursor of novel anticancer and anti-HIV drugs. Therefore, many studies have focused on the biosynthesis of ergosterol and have attempted to upregulate its content in multiple organisms. Great progress has been made in understanding the regulation of ergosterol biosynthesis in Saccharomyces cerevisiae. However, this molecular mechanism in F. velutipes remains largely uncharacterized. RESULTS: In this study, nine cDNA libraries, prepared from mycelia, young fruiting bodies and mature fruiting bodies of F. velutipes (three replicate sets for each stage), were sequenced using the Illumina HiSeq™ 4000 platform, resulting in at least 6.63 Gb of clean reads from each library. We studied the changes in genes and metabolites in the ergosterol biosynthesis pathway of F. velutipes during the development of fruiting bodies. A total of 13 genes (6 upregulated and 7 downregulated) were differentially expressed during the development from mycelia to young fruiting bodies (T1), while only 1 gene (1 downregulated) was differentially expressed during the development from young fruiting bodies to mature fruiting bodies (T2). A total of 7 metabolites (3 increased and 4 reduced) were found to have changed in content during T1, and 4 metabolites (4 increased) were found to be different during T2. A conjoint analysis of the genome-wide connection network revealed that the metabolites that were more likely to be regulated were primarily in the post-squalene pathway. CONCLUSIONS: This study provides useful information for understanding the regulation of ergosterol biosynthesis and the regulatory relationship between metabolites and genes in the ergosterol biosynthesis pathway during the development of fruiting bodies in F. velutipes.

SmJAZ8 acts as a core repressor regulating JA-induced biosynthesis of salvianolic acids and tanshinones in Salvia miltiorrhiza hairy roots.

Jasmonates (JAs) are important plant hormones that regulate a variety of plant development and defense processes, including biosynthesis of secondary metabolites. The JASMONATE ZIM DOMAIN (JAZ) proteins act as negative regulators in the JA signaling pathways of plants. We first verified that methyl jasmonate (MeJA) enhanced the accumulation of both salvianolic acids and tanshinones in Salvia miltiorrhiza (Danshen) hairy roots by inducing the expression of their biosynthetic pathway genes. Nine JAZ genes were cloned from Danshen and their expression levels in hairy roots were all increased by treatment with MeJA. When analyzed in detail, however, SmJAZ8 showed the strongest expression in the induced hairy roots. Overexpression or RNAi of SmJAZ8 deregulated or up-regulated the yields of salvianolic acids and tanshinones in the MeJA-induced transgenic hairy roots, respectively, and transcription factors and biosynthetic pathway genes showed an expression pattern that mirrored the production of the compounds. Genetic transformation of SmJAZ8 altered the expression of other SmJAZ genes, suggesting evidence of crosstalk occurring in JAZ-regulated secondary metabolism. Furthermore, the transcriptome analysis revealed a primary-secondary metabolism balance regulated by SmJAZ8. Altogether, we propose a novel role for SmJAZ8 as a negative feedback loop controller in the JA-induced biosynthesis of salvianolic acids and tanshinones.

Metabolite Profiles, Bioactivity, and HPLC Fingerprint of Different Varieties of Eucommia ulmoides Oliv.: Towards the Utilization of Medicinal and Commercial Chinese Endemic Tree.

Eucommia ulmoides Oliv. is widely regarded in China as a precious medicinal and commercial endemic tree. Due to cross-breeding or natural variation of E. ulmoides, the metabolite composition may vary significantly, making control of the medical quality difficult. In order to improve the rational development and utilization, the quality of seven varieties of E. ulmoides were evaluated based on metabolite profiles (total phenolic, total flavonoid, gutta-percha, aucubin, geniposidic acid, chlorogenic acid, geniposide, pinoresinol diglucoside, rutin, hyperoside, and astragalin), bioactivities (in vitro, in vivo antioxidant activities, and antibacterial activities) and HPLC fingerprint combined with chemometrics analysis. On this basis, the differences of medicinal parts (leaf and bark) were further carried out. For the traditional use of bark, Purple-leaf E. ulmoides was the most suitable. For the use of leaf, Qinzhong 1 and Purple-leaf E. ulmoides were appropriate. HPLC fingerprint analysis showed that significant differences in metabolite profiles exist among seven varieties of E. ulmoides. Combined with chemometrics analysis, seven varieties of E. ulmoides were divided into three groups from the use of leaf and bark. The analysis not only evaluated quality of seven varieties of E. ulmoides, but also could distinguish different varieties and different regions of origin. The results can provide theoretical basis for E. ulmoides resources utilization and cultivation of fine varieties.

[Effect of different fertilization treatments on yield and secondary metabolites of Codonopsis pilosula].

The research studies the effect of different fertilization treatments on yield and accumulation of secondary metabolites of Codonopsis pilosula by using single factor randomized block design, in order to ensure reasonable harvesting time and fertilization ratio, and provide the basis for standardized cultivation of C. pilosula. According to the clustering results, the nitrogen fertilizer benefitted for the improvement of root diameter and biomass of C. pilosula. The phosphate fertilizer could promote the content of C. pilosula polysaccharide. The organic fertilizers could increase the content of lobetyolin. With the time going on, C. pilosula's yield, polysaccharide and ehanol-soluble extracts increased while the content of lobetyolin decreased. According to various factors, October is a more reasonable harvest period. Organic fertilizers are more helpful to the yield and accumulation of secondary metabolites of C. pilosula.

Antioxidative activities and active compounds of extracts from Catalpa plant leaves.

In order to screen the Catalpa plant with high antioxidant activity and confirm the corresponding active fractions from Catalpa ovata G. Don, C. fargesii Bur., and C. bungei C. A. Mey., total flavonoid contents and antioxidant activities of the extracts/fractions of Catalpa plant leaves were determined. The determined total flavonoid content and antioxidant activity were used as assessment criteria. Those compounds with antioxidant activity were isolated with silica gel column chromatography and ODS column chromatography. Our results showed that the total flavonoid content in C. bungei C. A. Mey. (30.07 mg/g · DW) was the highest, followed by those in C. fargesii Bur. (25.55 mg/g · DW) and C. ovata G. Don (24.96 mg/g · DW). According to the determination results of total flavonoid content and antioxidant activity in 3 clones of leaves of C. bungei C. A. Mey., the total flavonoid content and antioxidant activity in crude extracts from C. bungei C. A. Mey. 6 (CA6) leaves were the highest. Moreover, the results showed that the total flavonoid content and antioxidant activities of ethyl acetate (EA) fraction in ethanol crude extracts in CA6 leaves were the highest, followed by n-butanol, petroleum ether (PE), and water fractions. Two flavonoid compounds with antioxidant activity were firstly isolated based on EA fraction. The two compounds were luteolin (1) and apigenin (2), respectively.

Hydrogen peroxide is involved in salicylic acid-elicited rosmarinic acid production in Salvia miltiorrhiza cell cultures.

Salicylic acid (SA) is an elicitor to induce the biosynthesis of secondary metabolites in plant cells. Hydrogen peroxide (H2O2) plays an important role as a key signaling molecule in response to various stimuli and is involved in the accumulation of secondary metabolites. However, the relationship between them is unclear and their synergetic functions on accumulation of secondary metabolites are unknown. In this paper, the roles of SA and H2O2 in rosmarinic acid (RA) production in Salvia miltiorrhiza cell cultures were investigated. The results showed that SA significantly enhanced H2O2 production, phenylalanine ammonia-lyase (PAL) activity, and RA accumulation. Exogenous H2O2 could also promote PAL activity and enhance RA production. If H2O2 production was inhibited by NADPH oxidase inhibitor (IMD) or scavenged by quencher (DMTU), RA accumulation would be blocked. These results indicated that H2O2 is secondary messenger for signal transduction, which can be induced by SA, significantly and promotes RA accumulation.

Hydrogen peroxide and nitric oxide are involved in salicylic acid-induced salvianolic acid B production in Salvia miltiorrhiza cell cultures.

Hydrogen peroxide (H2O2) and nitric oxide (NO) are key signaling molecules in cells whose levels are increased in response to various stimuli and are involved in plant secondary metabolite synthesis. In this paper, the roles of H2O2 and NO on salvianolic acid B (Sal B) production in salicylic acid (SA)-induced Salvia miltiorrhiza cell cultures were investigated. The results showed that H2O2 could be significantly elicited by SA, even though IMD (an inhibitor of NADPH oxidase) or DMTU (a quencher of H2O2) were employed to inhibit or quench intracellular H2O2. These elicited H2O2 levels significantly increased NO production by 1.6- and 1.46 fold in IMD+SA and DMTU+SA treatments, respectively, and induced 4.58- and 4.85-fold Sal B accumulation, respectively. NO was also markedly elicited by SA, in which L-NNA (an inhibitor of NO synthase) and cPTIO (a quencher of NO) were used to inhibit or quench NO within cells, and the induced NO could significantly enhance H2O2 production by 1.92- and 1.37-fold in L-NNA+SA and cPTIO+SA treatments, respectively, and 3.27- and 1.50-fold for Sal B accumulation, respectively. These results indicate that elicitation of SA for either H2O2 or NO was independent, and the elicited H2O2 or NO could act independently or synergistically to induce Sal B accumulation in SA-elicited cells.

Lignan contents of Schisandra chinensis (Turcz.) Baill. from different origins-A new model for evaluating the content of prominent components of Chinese herbs.

BACKGROUND: As a traditional Chinese herbal medicine, Schisandra chinensis exhibits various effects such as liver protection, blood sugar regulation, blood lipid regulation, immune function regulation, antidepressant activity, etc. However, because of its intricate composition, diverse origins, and medicinal effects depending on complex compound groups, there are differences in the lignan composition of S. chinensis from different origins. Therefore, it is currently difficult to evaluate the quality of medicinal materials from plants of different origins using a single qualitative quality control index. PURPOSE: This paper aims to investigate the potential relationship between the lignan components of S. chinensis from different origins and to establish stable assessment indices for determining the lignan content of S. chinensis from multiple perspectives. METHODS: In this study, we collected S. chinensis samples of seven major origins in China, and randomly sampled 6-9 batches of each origin for a total of 60 batches. The lignan content was determined by HPLC, and its distribution law of the ratio of each lignan component of S. chinensis to Schisandrol A content was analyzed. Combining network pharmacology and differential analysis between samples, the stable and effective substances used as quality markers were determined. RESULTS: There were some correlations among the lignan contents of S. chinensis, some correlations between schisandrin A and other lignans of S. chinensis could be determined. The ratio of each component to the indicator component schisandrol A was evenly distributed and reflected the lignan content of S. chinensis to some extent. Four substances (schisandrol A, schisandrol B, schisantherin A, and schisandrin C) were determined by network pharmacology combined with the analysis results of HCA, PCA and PLS-DA to further optimize the model. They displayed a strong connection with the core target, a large contribution rate to the principal components, and a stable content in each batch of samples, suggesting that these components may be the main active substances of S. chinensis lignans. Therefore, they could be used as main indicators evaluating the advantages and disadvantages of S. chinensis by examining the consistency of component proportions. CONCLUSION: This method can intuitively evaluate the content of main lignans in S. chinensis. This quality assessment model is an exploration of the multi-component comprehensive evaluation system of S. chinensis, providing a new concept for the quality evaluation system of Chinese herbal medicines.

Raspberry Ketone Prevents LPS-Induced Depression-Like Behaviors in Mice by Inhibiting TLR-4/NF-κB Signaling Pathway via the Gut-Brain Axis.

SCOPE: Depression, a prevalent mental disorder, has significantly impacted the lives of 350 million people, yet it holds promise for amelioration through food-derived phenolics. Raspberries, renowned globally for their delectable flavor, harbor a phenolic compound known as raspberry ketone (RK). However, the impact of RK on depressive symptoms remains ambiguous. This study aims to investigate the impact of RK on lipopolysaccharide (LPS)-induced depressed mice and elucidates its potential mechanisms, focusing on the gut-brain axis. METHODS AND RESULTS: Through behavioral tests, RK exerts a notable preventive effect on LPS-induced depression-like behaviors in mice. RK proves capable of attenuating gut inflammation, repairing gut barrier impairment, modulating the composition of the gut microbiome (Muribaculaceae, Streptococcus, Lachnospiraceae, and Akkermansia), and promoting the production of short-chain fatty acids. Furthermore, RK alleviates neuroinflammation by suppressing the TLR-4/NF-κB pathway and bolsters synaptic function by elevating levels of neurotrophic factors and synapse-associated proteins. CONCLUSION: The current study provides compelling evidence that RK effectively inhibits the TLR-4/NF-κB pathway via the gut-brain axis, leading to the improvement of LPS-induced depression-like behaviors in mice. This study addresses the research gap in understanding the antidepressant effects of RK and illuminates the potential of utilizing RK as a functional food for preventing depression.

[Regulation effects of intracellular and extracellular Ca2+ on biosynthesis of rosmarinic acid induced by salicylic acid in young seedlings of Salvia miltiorrhiza].

OBJECTIVE: To investigate the effect of intracellular and extracellular Ca2+ on the biosynthesis of rosmarinic acid (RA) induced by salicylic acid in young seedlings of Salvia miltiorrhiza. METHOD: Young seedlings of S. miltiorrhiza were used to select an optimal concentration of salicylic acid (SA), and then use the optimal concentration of SA to investigate the effects of extracellular Ca2+ channel inhibitors Verapamil, LaCl3, intracelluar calmodulin antagonist TFP and intracelluar Ca2+ channel inhibitors LiCl on the biosynthesis of RA and related enzymes. RESULT: SA increased the accumulation of RA and the activities of PAL and TAT, especially the SA of 2 mmol x L(-1) after 24 h. SA improved the accumulation of RA to (40.51 +/- 2.16) mg x g(-1), which was 1.97 times than that of control, and the activities of PAL, TAT were 1.42 times and 1.29 times than those of the control. However, Vp, LaCl3, TFP, LiCl inhibited the effects of SA evidently. CONCLUSION: Ca2+ plays a key role in the regulation of the induction process.

The SmMYB36-SmERF6/SmERF115 module regulates the biosynthesis of tanshinones and phenolic acids in salvia miltiorrhiza hairy roots.

Tanshinone and phenolic acids are the most important active substances of Salvia miltiorrhiza, and the insight into their transcriptional regulatory mechanisms is an essential process to increase their content in vivo. SmMYB36 has been found to have important regulatory functions in the synthesis of tanshinone and phenolic acid; paradoxically, its mechanism of action in S. miltiorrhiza is not clear. Here, we demonstrated that SmMYB36 functions as a promoter of tanshinones accumulation and a suppressor of phenolic acids through the generation of SmMYB36 overexpressed and chimeric SmMYB36-SRDX (EAR repressive domain) repressor hairy roots in combination with transcriptomic-metabolomic analysis. SmMYB36 directly down-regulate the key enzyme gene of primary metabolism, SmGAPC, up-regulate the tanshinones biosynthesis branch genes SmDXS2, SmGGPPS1, SmCPS1 and down-regulate the phenolic acids biosynthesis branch enzyme gene, SmRAS. Meanwhile, SmERF6, a positive regulator of tanshinone synthesis activating SmCPS1, was up-regulated and SmERF115, a positive regulator of phenolic acid biosynthesis activating SmRAS, was down-regulated. Furthermore, the seven acidic amino acids at the C-terminus of SmMYB36 are required for both self-activating domain and activation of target gene expression. As a consequence, this study contributes to reveal the potential relevance of transcription factors synergistically regulating the biosynthesis of tanshinone and phenolic acid.