Rapid characterization of non-volatile phenolic compounds reveals the reliable chemical markers for authentication of traditional Chinese medicine Xiang-ru among confusing Elsholtzia species.

The aerial parts of Mosla chinensis Maxim. and Mosla chinensis cv. 'Jiangxiangru' (MCJ) are widely utilized in traditional Chinese medicine (TCM), known collectively as Xiang-ru. However, due to clinical effectiveness concerns and frequent misidentification, the original plants have increasingly been substituted by various species within the genera Elsholtzia and Mosla. The challenge in distinguishing between these genera arises from their similar morphological and metabolic profiles. To address this issue, our study introduced a rapid method for metabolic characterization, employing high-resolution mass spectrometry-based metabolomics. Through detailed biosynthetic and chemometric analyses, we pinpointed five phenolic compounds-salviaflaside, cynaroside, scutellarein-7-O-D-glucoside, rutin, and vicenin-2-among 203 identified compounds, as reliable chemical markers for distinguishing Xiang-ru from closely related Elsholtzia species. This methodology holds promise for broad application in the analysis of plant aerial parts, especially in verifying the authenticity of aromatic traditional medicinal plants. Our findings underscore the importance of non-volatile compounds as dependable chemical markers in the authentication process of aromatic traditional medicinal plants.

The effect of scutellaria baicalensis and its active ingredients on major depressive disorder: a systematic review and meta-analysis of literature in pre-clinical research.

Background: Scutellaria baicalensis, the dry root of scutellaria baicalensis georgi, is a traditional Chinese medicine with long. In clinic, scutellaria baicalensis is commonly used in prescription for the treatment of depression. Additionally, numerous pre-clinical studies have shown that Scutellaria baicalensis and its active constituents are effective for depression. In this study, we aims to systematically review the roles of scutellaria baicalensis in depression and summarize the possible mechanism. Methods: A systematic review and meta-analysis were conducted to analyze the existing studies on the effects of scutellaria baicalensis on depression in animal models. Briefly, we searched electronic databases including Pubmed and Embase for preclinical trial studies from inception to September 2023. The items in each study were evaluated by two independent reviewers, and meta-analyses were performed on scutellaria baicalensis-induced behavioral changes in the study. Finally, random effects model is used to collect data. Results: A total of 49 studies were identified, and 13 studies were included in the final analysis. They all reported the different antidepressant effects of scutellaria baicalensis and the underlying biological mechanisms. Among the included 13 studies, the results of eight articles SPT[SMD = -2.80, 95%CI(-4.03, -1.57), p < 0.01], the results of the nine articles OFT[SMD = -2.38, 95%CI(-3.53, -1.23), p < 0.01], and the results of two articles NSFT[SMD = -2.98, 95%CI(-3.94, -2.02), p < 0.01] were significantly different from the control group. The risk of bias was moderate in all studies, however, there was a significant heterogeneity among studies. Conclusion: These results preliminarily suggest that scutellaria baicalensis can alleviate depressive behaviors and modulate underlying mechanisms, which is expected to be a promising antidepressant.

The Efficacy of Chaihu-Guizhi-Ganjiang Decoction on Chronic Non-Atrophic Gastritis with Gallbladder Heat and Spleen Cold Syndrome and Its Metabolomic Analysis: An Observational Controlled Before-After Clinical Trial.

Purpose: The aim of this study was to verify the effectiveness and explore the mechanism of Chaihu-Guizhi-Ganjiang decoction (CGGD) in the treatment of chronic non-atrophic gastritis (CNAG) with gallbladder heat and spleen cold syndrome (GHSC) by metabolomics based on UHPLC-Q-TOF/MS. Patients and Methods: An observational controlled before-after study was conducted to verify the effectiveness of CGGD in the treatment of CNAG with GHSC from January to June 2023, enrolling 27 patients, who took CGGD for 28 days. 30 healthy volunteers were enrolled as the controls. The efficacy was evaluated by comparing the traditional Chinese medicine (TCM) syndrome and CNAG scores, and clinical parameters before and after treatment. The plasma levels of hormones related to gastrointestinal function were collected by ELISA. The mechanisms of CGGD in the treatment of CNAG with GHSC were explored using a metabolomic approach based on UHPLC-Q-TOF/MS. Results: Patients treated with CGGD experienced a statistically significant improvement in TCM syndrome and CNAG scores (p < 0.01). CGGD treatment evoked the concentration alteration of 15 biomarkers, which were enriched in the glycerophospholipid metabolism, and branched-chain amino acids biosynthesis pathways. Moreover, CGGD treatment attenuated the abnormalities of the gastrointestinal hormone levels and significantly increased the pepsinogen level. Conclusion: It was the first time that this clinical trial presented detailed data on the clinical parameters that demonstrated the effectiveness of CGGD in the treatment of CNAG with GHSC patients. This study also provided supportive evidence that CNAG with GHSC patients were associated with disturbed branched-chain amino acid metabolism and glycerophospholipid levels, suggesting that CNAG treatment based on TCM syndrome scores was reasonable and also provided a potential pharmacological mechanism of action of CGGD.

Versatile CYP98A enzymes catalyse meta-hydroxylation reveals diversity of salvianolic acids biosynthesis.

Salvianolic acids (SA), such as rosmarinic acid (RA), danshensu (DSS), and their derivative salvianolic acid B (SAB), etc. widely existed in Lamiaceae and Boraginaceae families, are of interest due to medicinal properties in the pharmaceutical industries. Hundreds of studies in past decades described that 4-coumaroyl-CoA and 4-hydroxyphenyllactic acid (4-HPL) are common substrates to biosynthesize SA with participation of rosmarinic acid synthase (RAS) and cytochrome P450 98A (CYP98A) subfamily enzymes in different plants. However, in our recent study, several acyl donors and acceptors included DSS as well as their ester-forming products all were determined in SA-rich plants, which indicated that previous recognition to SA biosynthesis is insufficient. Here, we used Salvia miltiorrhiza, a representative important medicinal plant rich in SA, to elucidate the diversity of SA biosynthesis. Various acyl donors as well as acceptors are catalysed by SmRAS to form precursors of RA and two SmCYP98A family members, SmCYP98A14 and SmCYP98A75, are responsible for different positions' meta-hydroxylation of these precursors. SmCYP98A75 preferentially catalyses C-3' hydroxylation, and SmCYP98A14 preferentially catalyses C-3 hydroxylation in RA generation. In addition, relative to C-3' hydroxylation of the acyl acceptor moiety in RA biosynthesis, SmCYP98A75 has been verified as the first enzyme that participates in DSS formation. Furthermore, SmCYP98A enzymes knockout resulted in the decrease and overexpression leaded to dramatic increase of SA accumlation. Our study provides new insights into SA biosynthesis diversity in SA-abundant species and versatility of CYP98A enzymes catalytic preference in meta-hydroxylation reactions. Moreover, CYP98A enzymes are ideal metabolic engineering targets to elevate SA content.

Functional divergence of CYP76AKs shapes the chemodiversity of abietane-type diterpenoids in genus Salvia.

The genus Salvia L. (Lamiaceae) comprises myriad distinct medicinal herbs, with terpenoids as one of their major active chemical groups. Abietane-type diterpenoids (ATDs), such as tanshinones and carnosic acids, are specific to Salvia and exhibit taxonomic chemical diversity among lineages. To elucidate how ATD chemical diversity evolved, we carried out large-scale metabolic and phylogenetic analyses of 71 Salvia species, combined with enzyme function, ancestral sequence and chemical trait reconstruction, and comparative genomics experiments. This integrated approach showed that the lineage-wide ATD diversities in Salvia were induced by differences in the oxidation of the terpenoid skeleton at C-20, which was caused by the functional divergence of the cytochrome P450 subfamily CYP76AK. These findings present a unique pattern of chemical diversity in plants that was shaped by the loss of enzyme activity and associated catalytic pathways.

SbWRKY75- and SbWRKY41-mediated jasmonic acid signaling regulates baicalin biosynthesis.

Introduction: Scutellaria baicalensis Georgi is a traditional Chinese medicinal plant with broad pharmacological activities whose main active ingredient is the flavonoid baicalin. Given its medicinal value and increasing market demand, it is essential to improve the plant's baicalin content. Flavonoid biosynthesis is regulated by several phytohormones, primarily jasmonic acid (JA). Methods: In this study, we conducted transcriptome deep sequencing analysis of S. baicalensis roots treated with methyl jasmonate for different durations (1, 3, or 7 hours). Leveraging weighted gene co-expression network analysis and transcriptome data, we identified candidate transcription factor genes involved in the regulation of baicalin biosynthesis. To validate the regulatory interactions, we performed functional assays such as yeast one-hybrid, electrophoretic mobility shift, and dual-luciferase assays. Results: Our findings demonstrated that SbWRKY75 directly regulates the expression of the flavonoid biosynthetic gene SbCLL-7, whereas SbWRKY41 directly regulates the expression of two other flavonoid biosynthetic genes, SbF6H and SbUGT, thus regulating baicalin biosynthesis. We also obtained transgenic S.baicalensis plants by somatic embryo induction and determined that overexpressing SbWRKY75 increased baicalin content by 14%, while RNAi reduced it by 22%. Notably, SbWRKY41 indirectly regulated baicalin biosynthesis by modulating the expression of SbMYC2.1, SbJAZ3 and SbWRKY75. Discussion: This study provides valuable insights into the molecular mechanisms underlying JA-mediated baicalin biosynthesis in S. baicalensis. Our results highlight the specific roles of transcription factors, namely SbWRKY75 and SbWRKY41, in the regulation of key biosynthetic genes. Understanding these regulatory mechanisms holds significant potential for developing targeted strategies to enhance baicalin content in S. baicalensis through genetic interventions.

Stepwise changes in flavonoids in spores/pollen contributed to terrestrial adaptation of plants.

Protecting haploid pollen and spores against UV-B light and high temperature, 2 major stresses inherent to the terrestrial environment, is critical for plant reproduction and dispersal. Here, we show flavonoids play an indispensable role in this process. First, we identified the flavanone naringenin, which serves to defend against UV-B damage, in the sporopollenin wall of all vascular plants tested. Second, we found that flavonols are present in the spore/pollen protoplasm of all euphyllophyte plants tested and that these flavonols scavenge reactive oxygen species to protect against environmental stresses, particularly heat. Genetic and biochemical analyses showed that these flavonoids are sequentially synthesized in both the tapetum and microspores during pollen ontogeny in Arabidopsis (Arabidopsis thaliana). We show that stepwise increases in the complexity of flavonoids in spores/pollen during plant evolution mirror their progressive adaptation to terrestrial environments. The close relationship between flavonoid complexity and phylogeny and its strong association with pollen survival phenotypes suggest that flavonoids played a central role in the progression of plants from aquatic environments into progressively dry land habitats.

The transcription factors SbMYB45 and SbMYB86.1 regulate flavone biosynthesis in Scutellaria baicalensis.

Scutellaria baicalensis Georgi is an important Chinese medicinal plant that is rich in the flavones baicalin, wogonoside, and wogonin, providing it with anti-cancer, anti-inflammatory, and antibacterial properties. However, although the biosynthetic pathways of baicalin and its derivates have been elucidated, the regulation of flavone biosynthesis in S. baicalensis is poorly understood. Here, we found that the contents of baicalin and its derivates increased and that baicalin biosynthetic pathway genes were induced in response to light, and baicalin and baicalein are not exclusively produced in the roots of S. baicalensis. Based on the fact that MYB transcription factors are known to play important roles in flavone biosynthesis, we identified SbMYB45 and SbMYB86.1 in S. baicalensis and determined that they bind to the promoter of the flavone biosynthesis gene SbCHI to enhance its transcription. Moreover, overexpressing SbMYB45 and SbMYB86.1 enhanced the accumulation of baicalin in S. baicalensis leaves. We demonstrate that SbMYB45 and SbMYB86.1 bind to the cis-acting element MBSII in the promoter of CHI to redundantly induce its expression upon light exposure. These findings indicate that SbMYB45 and SbMYB86.1 transcriptionally activate SbCHI in response to light and enhance flavone contents in S. baicalensis.

Ameliorative effects of mangiferin derivative TPX on insulin resistance via PI3K/AKT and AMPK signaling pathways in human HepG2 and HL-7702 hepatocytes.

BACKGROUND: As a multifaceted metabolic disorder, insulin resistance is accompanied by the preceding onset of type 2 diabetes mellitus, hyperinsulinemia, metabolic dysfunction-associated fatty liver disease (MAFLD) and other metabolic syndromes. Currently, the number of existing drugs and mechanism-based strategies is limited to alleviate insulin resistance in clinics. As a natural polyphenol product derivative, 1,3,6,7-tetrapropylene acyloxy-ketone (TPX) showed a significant hypoglycemic effect in our previous studies. However, whether TPX could improve hepatic insulin sensitivity was unknown. PURPOSE: To explore whether insulin sensitivity can be improved by the treatment with TPX and further investigate its mechanism(s) of activity. METHODS: To mimic hyperglycemia and insulin resistance in vitro, human HepG2 and HL-7702 hepatocytes were exposed to high glucose. Cellular glucose uptake, glucose consumption, glycogen synthesis, and glucose production were quantified after TPX treatment. The effects of TPX on AMP-activated protein kinase (AMPK) phosphorylation, glucose metabolism, and insulin signal transduction were evaluated by western blotting and network pharmacology analysis. The eGFP-membrane of glucose transporter type 4 (GLUT4) lentivirus transfected cells were constructed to investigate the effects of TPX on GLUT4 mobilization. Reactive oxygen species activity in high glucose-induced insulin-resistant cells was measured by DCFH-DA to show oxidative stress. RESULTS: Treatment with TPX improved glycogen synthesis and inhibited gluconeogenesis by regulating GSK3ß, G6Pase, and PEPCK. Furthermore, high glucose-induced inhibition of glucose consumption, glucose uptake, and GLUT4-mediated membrane translocation were reverted by TPX. Accordingly, mechanistic investigations revealed that TPX interacted with AMPK protein and activated the phosphorylation of AKT, thereby improving energy homeostasis and further ameliorating hepatic insulin resistance. Network pharmacology analysis and molecular docking further confirmed AMPK as an active target of TPX. Concordantly, the pharmacological activity of TPX was reversed by the AMPK inhibitor compound C when hepatocytes were exposed to high glucose stimulation. CONCLUSION: In summary, our study confirmed TPX contributions to insulin resistance improvements by targeting AMPK and PI3K/AKT to restore the insulin signaling pathway, which may be an important potential treatment strategy for insulin-resistance-related diseases, including MAFLD and diabetes.

Genome-wide identification and characterisation of bHLH transcription factors in Artemisia annua.

BACKGROUND: A. annua (also named Artemisia annua, sweet wormwood) is the main source of the anti-malarial drug artemisinin, which is synthesised and stored in its trichomes. Members of the basic Helix-Loop-Helix (bHLH) family of transcription factors (TFs) have been implicated in artemisinin biosynthesis in A. annua and in trichome development in other plant species. RESULTS: Here, we have systematically identified and characterised 226 putative bHLH TFs in A. annua. All of the proteins contain a HLH domain, 213 of which also contain the basic motif that mediates DNA binding of HLH dimers. Of these, 22 also contained a Myc domain that permits dimerisation with other families of TFs; only two proteins lacking the basic motif contained a Myc domain. Highly conserved GO annotations reflected the transcriptional regulatory role of the identified TFs, and suggested conserved roles in biological processes such as iron homeostasis, and guard cell and endosperm development. Expression analysis revealed that three genes (AabHLH80, AabHLH96, and AaMyc-bHLH3) exhibited spatiotemporal expression patterns similar to genes encoding key enzymes in artemisinin synthesis. CONCLUSIONS: This comprehensive analysis of bHLH TFs provides a new resource to direct further analysis into key molecular mechanisms underlying and regulating artemisinin biosynthesis and trichome development, as well as other biological processes, in the key medicinal plant A. annua.

Tanreqing injection protects against bleomycin-induced pulmonary fibrosis via inhibiting STING-mediated endoplasmic reticulum stress signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Idiopathic pulmonary fibrosis (IPF), characterized by excessive collagen deposition, is a progressive and typically fatal lung disease without effective therapeutic methods. Tanreqing injection (TRQ), a Traditional Chinese Patent Medicine, has been widely used to treat inflammatory respiratory diseases clinically. AIM OF THE STUDY: The present work aims to elucidate the therapeutic effects and the possible mechanism of TRQ against pulmonary fibrosis. METHODS: The pulmonary fibrosis murine model were constructed by the intratracheal injection of bleomycin (BLM). 7 days later, TRQ-L (2.6 ml/kg) and TRQ-H (5.2 ml/kg) were administered via intraperitoneal injection respectively for 21 days. The efficacy and underlying molecular mechanism of TRQ were investigated. RESULTS: Here, we showed that TRQ significantly inhibited BLM-induced lung edema and pulmonary function. TRQ markedly reduced BLM-promoted inflammatory cell infiltration in BALF and inflammatory cytokines release (TNF-α, IL-6, and IL-1ß) in serum and lung tissues. Meanwhile, TRQ also alleviated BLM-induced collagen synthesis and deposition. Simultaneously, TRQ attenuated BLM-induced pulmonary fibrosis through regulating the expression of fibrotic hallmarks, manifested by down-regulated α-SMA and up-regulated E-cadherin. Moreover, we found that TRQ significantly prevented STING, p-P65, BIP, p-PERK, p-eIF2α, and ATF4 expression in lung fibrosis mice. CONCLUSIONS: Taken together, our results indicated that TRQ positively affects inflammatory responses and lung fibrosis by regulating STING-mediated endoplasmic reticulum stress (ERS) signal pathway.

Chaihu-Guizhi-Ganjiang Decoction is more efficacious in treating irritable bowel syndrome than Dicetel according to metabolomics analysis.

BACKGROUND: Chaihu-Guizhi-Ganjiang Decoction (CGGD) is a traditional Chinese medicine (TCM) prescription used to treat viral influenza. There is evidence that CGGD can be used to treat irritable bowel syndrome (IBS) but the potential mechanism of action and metabolites produced upon CGGD treatment remains elusive. METHODS: Patients with IBS were treated with pinaverium bromide (Dicetel™) and then CGGD after a washout period of 1 week. Both treatments lasted for 30 days. The efficacy and changes of metabolites in plasma after the two treatments were compared. Plasma samples were acquired before and after each treatment, and untargeted metabolics analysis was performed. RESULTS: Efficacy was measured according to the Rome IV criteria and TCM theory. Our results indicated that CGGD showed significantly better efficacy than Dicetel in the treatment of IBS utilizing each criterion. CGGD exerted greater effects on plasma metabolism than Dicetel. Dicetel treatment led to increased tryptophan metabolism (increased levels of 5-Hydroxyindoleacetaldehyde) and increased protein metabolism (increased levels of L-arginine). CGGD treatment significantly (p < 0.05) increased carnitine metabolism, with elevated levels of L-carnitine and acylcarnitine in plasma. Such changes in these metabolites could exert effects against IBS by improving gastrointestinal motility and suppressing pain, depression, and inflammation. CONCLUSIONS: CGGD appeared to be more efficacious than Dicetel for treating patients with IBS. The findings provide a sound support for the underlying biomolecular mechanism of CGGD in the prevention and treatment of IBS.

Mitochondrial toxicants in Xian-Ling-Gu-Bao induce liver injury by regulating the PI3K/mTOR signaling pathway: an in vitro study.

BACKGROUND: Drug-induced mitochondrial toxicity is thought to be a common mechanism of drug hepatotoxicity. Xian-Ling-Gu-Bao (XLGB) oral preparation is a commonly used drug for osteoporosis in China. Classical safety evaluation studies have shown that the entire preparation and six Chinese herbal medicines have high safety, but the incidence of drug-induced liver damage due to XLGB remains high, the mechanism and toxic substances causing liver injury are still unclear. The purpose of this study is to identify compounds with potential mitochondrial liabilities in XLGB, and to clarify their underlying mechanisms and related pathways. METHODS: The mitochondrial function analysis was performed using an extracellular flux assay, which simultaneously monitored both oxygen consumption rate (OCR) and extracellular acidification rate (ECAR). Through network pharmacology and in vitro experimental verification, the potential protein targets, signaling pathways and molecular mechanism of mitochondrial toxicity have been studied. RESULTS: We observed a significant decrease in mitochondrial respiration of Psoraleae Fructus and its five compounds in fundamental bioenergetics parameters such as basal respiration, ATP-linked production and maximal respiration, indicating mitochondrial dysfunction. The network pharmacology results showed that the influence of XLGB on mitochondrial dysfunction was closely related to PI3K-Akt signaling pathway, mTOR signaling pathway and Apoptosis. Western blot showed that the levels of mTOR, p-mTOR (Ser2448), Raptor, PI3K (p110α), Beclin 1, ATG5 and Caspase-9 were up-regulated after treatment with psoralidin, psoralen and bavachin, and the expression of Bcl-2 was down-regulated after bavachinin treatment. CONCLUSIONS: The hepatotoxicity of XLGB is associated with mitochondrial dysfunction. Five compounds in Psoraleae Fructus showed mitochondrial damage, they are psoralidin, isobavachalcone, bavachinin, bavachin and psoralen, especially psoralidin showed significant reduction in reserve capacity and respiratory control ratios. The molecular mechanism is related to the activation of PI3K/mTOR signaling pathway to inhibit autophagy and induce mitochondrial apoptosis.

Dissection of the Active Ingredients and Potential Mechanism of Han-Shi-Yu-Fei-Decoction in Treating COVID-19 Based on In Vivo Substances Profiling and Clinical Symptom-Guided Network Pharmacology.

This work was aimed to elucidate the mechanism of action of Han-Shi-Yu-Fei-decoction (HSYFD) for treating patients with mild coronavirus disease 2019 (COVID-19) based on clinical symptom-guided network pharmacology. Experimentally, an ultra-high performance liquid chromatography technique coupled with quadrupole time-of-flight mass spectrometry method was used to profile the chemical components and the absorbed prototype constituents in rat serum after its oral administration, and 11 out of 108 compounds were identified. Calculatingly, the disease targets of Han-Shi-Yu-Fei symptoms of COVID-19 were constructed through the TCMIP V2.0 database. The subsequent network pharmacology and molecular docking analysis explored the molecular mechanism of the absorbed prototype constituents in the treatment of COVID-19. A total of 42 HSYFD targets oriented by COVID-19 clinical symptom were obtained, with EGFR, TP53, TNF, JAK2, NR3C1, TH, COMT, and DRD2 as the core targets. Enriched pathway analysis yielded multiple COVID-19-related signaling pathways, such as the PI3K/AKT signaling pathway and JAK-STAT pathway. Molecular docking showed that the key compounds, such as 6-gingerol, 10-gingerol, and scopoletin, had high binding activity to the core targets like COMT, JAK2, and NR3C1. Our work also verified the feasibility of clinical symptom-guided network pharmacology analysis of chemical compounds, and provided a possible agreement between the points of views of traditional Chinese medicine and western medicine on the disease.

Integrated Chemical Interpretation and Network Pharmacology Analysis to Reveal the Anti-Liver Fibrosis Effect of Penthorum chinense.

Liver fibrosis is a disease with complex pathological mechanisms. Penthorum chinense Pursh (P. chinense) is a traditional Chinese medicine (TCM) for liver injury treatment. However, the pharmacological mechanisms of P. chinense on liver fibrosis have not been investigated and clarified clearly. This study was designed to investigate the chemicals in P. chinense and explore its effect on liver fibrosis. First, we developed a highly efficient method, called DDA-assisted DIA, which can both broaden mass spectrometry (MS) coverage and MS2 quality. In DDA-assisted DIA, data-dependent acquisition (DDA) and data-independent acquisition (DIA) were merged to construct a molecular network, in which 1,094 mass features were retained in Penthorum chinense Pursh (P. chinense). Out of these, 169 compounds were identified based on both MS1 and MS2 analysis. After that, based on a network pharmacology study, 94 bioactive compounds and 440 targets of P. chinense associated with liver fibrosis were obtained, forming a tight compound-target network. Meanwhile, the network pharmacology experimental results showed that multiple pathways interacted with the HIF-1 pathway, which was first identified involved in P. chinense. It could be observed that some proteins, such as TNF-α, Timp1, and HO-1, were involved in the HIF-1 pathway. Furthermore, the pharmacological effects of P. chinense on these proteins were verified by CCl4-induced rat liver fibrosis, and P. chinense was found to improve liver functions through regulating TNF-α, Timp1, and HO-1 expressions. In summary, DDA-assisted DIA could provide more detailed compound information, which will help us to annotate the ingredients of TCM, and combination with computerized network pharmacology provided a theoretical basis for revealing the mechanism of P. chinense.

Identification of Abietane-Type Diterpenoids and Phenolic Acids Biosynthesis Genes in Salvia apiana Jepson Through Full-Length Transcriptomic and Metabolomic Profiling.

Salvia apiana (S. apiana) Jepson is a medicinal plant that is frequently used by the Chumash Indians in southern California as a diaphoretic, calmative, diuretic, or antimicrobial agent. Abietane-type diterpenoids (ATDs) and phenolic acids (PAs) are the main bioactive ingredients in S. apiana. However, few studies have looked into the biosynthesis of ATDs and PAs in S. apiana. In this study, using metabolic profiling focused on the ATDs and PAs in the roots and leaves of S. apiana, we found a distinctive metabolic feature with all-around accumulation of ATDs, but absence of salvianolic acid B. To identify the candidate genes involved in these biosynthesis pathways, full-length transcriptome was performed by PacBio single-molecule real-time (SMRT) sequencing. A total of 50 and 40 unigenes were predicted to be involved in ATDs and PAs biosynthesis, respectively. Further transcriptional profile using Illumina HiSeq sequencing showed that the transcriptional variations of these pathways were consistent with the accumulation patterns of corresponding metabolites. A plant kingdom-wide phylogenetic analysis of cytochromes (CYPs) identified two CYP76AK and two CYP76AH subfamily genes that might contribute for the specific ATDs biosynthesis in S. apiana. We also noticed that the clade VII laccase gene family was significantly expanded in Salvia miltiorrhiza compared with that of S. apiana, indicating their involvements in the formation of salvianolic acid B. In conclusion, our results will enable the further understanding of ATDs and PAs biosynthesis in S. apiana and Salvia genus.

Analgesic and Anti-Arthritic Activities of Polysaccharides in Chaenomeles speciosa.

Chaenomeles speciosa (Sweet) Nakai has been long used as a folk medicine for rheumatic diseases treatment. This study aimed to investigate the effects and underlying mechanism of polysaccharides in Chaenomeles speciosa (CSP) on the pro-inflammatory cytokines and MAPK pathway in complete Freund's adjuvant (CFA)-induced arthritis and LPS-induced NR8383 cells. We used acetic acid (HAc)-induced writhing and CFA induced paw edema to determine the analgesic activity and anti-inflammatory activity, respectively. CFA rats were administered CSP (12.5, 25.0, and 50.0 mg/kg) daily for 3 weeks via oral gavage. The analgesic test was done using three different doses of the extract (50, 100, and 200 mg/kg). The anti-arthritic evaluation involved testing for paw swelling, swelling inhibition, and histological analysis in CFA rats. Finally, ELISA, western blot, qRT-PCR were done to determine the effect of CSP on the activation of MAPK pathway, production of pro-inflammatory cytokines in lipopolysaccharide (LPS)-stimulated NR838 macrophage cells. In pain models, oral uptake of CSP greatly reduced pain perception. Furthermore, in CFA rats, CSP substantially decreased paw swelling as well as synovial tissue proliferation and inflammatory cell infiltration. In addition, CSP was shown to inhibit pro-inflammatory cytokines (TNF-α, IL-1ß, and COX-2) as well as JNK and ERK1/2 phosphorylation in LPS-stimulated NR8383 cells. Thus, pro-inflammatory cytokine secretion and MAPK signaling downregulation promoted the analgesic and anti-arthritic effects of CSP.

Choline deficiency-related multi-omics characteristics are susceptible factors for chemotherapy-induced thrombocytopenia.

The XELOX chemotherapy protocol that includes capecitabine and oxaliplatin is the routine treatment for colorectal cancer (CRC), but it can cause chemotherapy-related adverse events such as thrombocytopenia (TCP). To identify predictive biomarkers and clarify the mechanism of TCP susceptibility, we conducted integrative analysis using normal colorectal tissue (CRT), plasma, and urine samples collected before CRC patients received adjuvant XELOX chemotherapy. RNA-sequencing and DNA methylation arrays were performed on CRT samples, while liquid chromatography-mass spectrometry was performed on CRT, plasma, and urine samples. Differentially expressed features (DEFs) from each uni-omics analysis were then subjected to integrative analysis using Multi-Omics Factor Analysis (MOFA). Choline-deficiency in plasma and CRT was found as the most critical TCP-related feature. Based on bioinformatic analysis and literature research, we further concluded that choline-deficiency was the possible reason for most of the other TCP-related multi-omics DEFs, including metabolites representing reduced sphingolipid de novo synthesis and elevated solute carrier-mediated transmembrane transportation in CRT and plasma, DNA hypermethylation and elevated expression of genes involved in neuronal system genes. In terms of thrombocytopoiesis, these TCP-related DEFs may cause atypical maintenance and differentiation of megakaryocyte, resulting a suppressed ability of thrombocytopoiesis, making patients more susceptible to chemotherapy-induced TCP. At last, prediction models were developed and validated with reasonably good discrimination. The area under curves (AUCs) of training sets were all > 0.9, while validation sets had AUCs between 0.778 and 0.926. In conclusion, our results produced reliable marker systems for predicting TCP and promising target for developing precision treatment to prevent TCP.

Effect of hepar-protecting Wuzhi capsule on pharmacokinetics and dose-effect character of tacrolimus in healthy volunteers.

Wuzhi capsule (WZC), a preparation of Fructus Schisandra sphenanthera extract, has been used widely for the treatment of viral and drug-induced hepatitis in China. This study aimed to determine the pharmacokinetic parameters of tacrolimus (TAC) when co-administered with WZC and the dose-effect of WZC on tacrolimus in healthy volunteers. The effect of an increased dosage of WZC (1, 2, 6, and 8 capsules once daily) on the relative oral exposure of tacrolimus was assessed to explore the dose-response relationship between WZC and tacrolimus using bioanalysis, pharmacokinetic, and genotypical analyses. The influence of CYP3A5 and MDR1 genetic polymorphisms on the WZC dose was elucidated by maintaining the Ctrough of tacrolimus in Chinese healthy volunteers. When co-administered with WZC, the Tmax of tacrolimus was increased significantly while the apparent oral clearance was decreased. The plasma tacrolimus level in volunteers with high CYP3A5 expression was much lower than that in those with mutant CYP3A5. However, polymorphisms of MDR1 exon26 C3435T, exon21 G2677T/A, and exon12 C1236T were not associated with plasma tacrolimus levels. Our findings provide important information on interactions between modern medications and herbal products, thus facilitating a better usage of tacrolimus in patients receiving WZC.

Biosynthesis-based spatial metabolome of Salvia miltiorrhiza Bunge by combining metabolomics approaches with mass spectrometry-imaging.

Defining the spatial distributions of metabolites and their structures are the two key aspects for interpreting the complexities of biosynthesis pathways in plants. As a means of obtaining information on the spatial distribution of metabolites, a strategy is needed that has high sensitivity and allows visualization. Toward this goal, we carried an untargeted metabolomics to obtain detailed metabolic information on different plant parts of Salvia miltiorrhiza, the roots of which are widely used in traditional Chinese medicine. Systematic optimization of desorption electrospray ionization mass spectrometry imaging (DESI-MSI) including parameter selection and sample preparation were carried out to improve the sensitivity of the method for plant samples. Guided by the metabolomics data, the spatial distributions of diverse metabolites, including phenolic acids, flavonoids, tanshinones, carbohydrates, and lipids, were characterized and visualized for both the underground and aerial parts. To integrate the information pertaining to the spatial distribution of metabolites, the flavonoids and phenolic acids (phenylpropanoid metabolic pathway) were chosen as examples for in-depth study the biosynthesis pathways in S. miltiorrhiza. The complementary data obtained from the metabolomics study and mass spectrometry imaging enabled the identification of key reactions involved in flavonoid biosynthesis in flowers, which lead the changes in metabolite distribution. The analysis also identified the core precursor for phenolic acid biosynthesis in Salvia species. Therefore, the powerful combination of metabolomics and mass spectrometry imaging provides a basis for obtaining detailed information on spatial metabolome and constitutes a platform for deep understanding the biosynthesis of bioactive metabolites in plants.