Chemical components and protective effects of Atractylodes japonica Koidz. ex Kitam against acetic acid-induced gastric ulcer in rats.

BACKGROUND: Atractylodes japonica Koidz. ex Kitam. (A. japonica, Chinese name: Guan-Cangzhu, Japanese name: Byaku-jutsu), a perennial herb, which is mainly distributed in northeast area of China, it's often used to treat digestive system diseases such as gastric ulcer (GU). However, the mechanism of its potential protective effects against GU remains unclear. AIM: To investigate the protective effects of A. japonica on acetic acid-induced GU rats. METHODS: The chemical constituents of A. japonica were determined by ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) analysis. The rat model of GU was simulated by acetic acid method. The pathological changes of gastric tissues were evaluated by hematoxylin-eosin stain, the levels of epidermal growth factor (EGF), EGF receptor (EGFR), nuclear factor kappa-B (NF-κB), interleukin-1ß (IL-1ß), IL-10, Na+-K+-ATPase (NKA) in serum and gastric tissues were determined by enzyme-linked immunosorbent assay, and the mRNA expressions of EGFR, NF-κBp65, IkappaBalpha (IκBα) and Zonula Occludens-1 (ZO-1) in gastric tissues were determined by real-time reverse transcription polymerase chain reaction, and the efficacy was observed. Then, plasma metabolomic analysis was performed by UPLC-MS/MS to screen the specific potential biomarkers, metabolic pathways and to explore the possible mechanisms. RESULTS: 48 chemical constituents were identified. Many of them have strong pharmacological activity, the results also revealed that A. japonica significantly improved the pathological damage of gastric tissues, increased the expression levels of IL-10, IκBα related to anti-inflammatory factors, decreased the expression levels of IL-1ß, NF-κB, NF-κBp65, related to proinflammatory factors, restored the levels of factors about EGF, EGFR, ZO-1 associated with ulcer healing and the levels of factors about NKA associated with energy metabolism. Metabolomic analysis identified 10 potential differential metabolites and enriched 7 related metabolic pathways. CONCLUSION: These findings contribute to the understanding of the potential mechanism of A. japonica to improve acetic acid-induced GU, and will be of great importance for the development and clinical application of natural drugs related to A. japonica.

Integrative analysis of transcriptome and metabolome reveals the sesquiterpenoids and polyacetylenes biosynthesis regulation in Atractylodes lancea (Thunb.) DC.

Atractylodes lancea (Thunb.) is a perennial medicinal herb, with its dry rhizomes are rich in various sesquiterpenoids and polyacetylenes components (including atractylodin, atractylon and ß-eudesmol). However, the contents of these compounds are various and germplasms specific, and the mechanisms of biosynthesis in A. lancea are still unknown. In this study, we identified the differentially expressed candidate genes and metabolites involved in the biosynthesis of sesquiterpenoids and polyacetylenes, and speculated the anabolic pathways of these pharmaceutical components by transcriptome and metabolomic analysis. In the sesquiterpenoids biosynthesis, a total of 28 differentially expressed genes (DEGs) and 6 differentially expressed metabolites (DEMs) were identified. The beta-Selinene is likely to play a role in the synthesis of atractylon and ß-eudesmol. Additionally, the polyacetylenes biosynthesis showed the presence of 3 DEGs and 4 DEMs. Notably, some fatty acid desaturase (FAB2 and FAD2) significantly down-regulated in polyacetylenes biosynthesis. The gamma-Linolenic acid is likely involved in the biosynthesis of polyacetylenes and thus further synthesis of atractylodin. Overall, these studies have investigated the biosynthetic pathways of atractylodin, atractylon and ß-eudesmol in A. lancea for the first time, and present potential new anchor points for further exploration of sesquiterpenoids and polyacetylenes compound biosynthesis pathways in A. lancea.

Green carbon dots derived from Atractylodes macrocephala: A potential nanodrug for treating alcoholic gastric ulcer.

Alcoholic gastric ulcer is a common acute gastric injury disease. The drugs currently used in clinical practice not only cannot fundamentally treat gastric injury, but also have serious side effects. There is an urgent demand for the discovery of a mild drug to treat alcoholic gastric ulcers. Herein, the green carbon dots derived from charred Atractylodes macrocephala (CAM-CDs) were acquired and have been proven to be safe and effective in alleviating alcoholic gastric ulcers at an inhibition rate up to 60%. CAM-CDs can markedly attenuate the gastric mucosa damage such as mucosal defect, bleeding and inflammatory cell infiltration by histopathological examination. Serum and tissue inflammatory cytokine measurements, as well as immunohistochemistry results, indicate that its mechanism of gastric mucosal protection may involve the reduction of IL-1ß and TNF-α by regulating inflammatory signaling pathway of the NF-κB/NLRP3 axis, as well as elevation of IL-10 levels. CAM-CDs also can reduce oxidative stress markers (MDA), increase PGE2 and mucin secretion (MUC5AC), and it simultaneously exerts slight inhibition of hydrogen potassium ATPase and pepsin activity to protect gastric mucosa, as well as increases the microbial diversity and regulates species composition of gut microbiota in rats with gastric ulcer. Our work provides a new perspective on utilizing carbon-based nanomaterials in the development of new mild drugs.

Modulatory Effects of Atractylodin and ß-Eudesmol on Human Cytochrome P450 Enzymes: Potential Drug-Drug Interactions.

Atractylodin and ß-eudesmol, the major bioactive compounds in Atractylodes lancea, are promising candidates for anti-cholangiocarcinoma. The inhibitory effects of both compounds on human rCYP1A2, rCYP2C9, rCYP2C19, rCYP2D6 and rCYP3A4 enzymes were investigated using luminogenic CYP450 kits. The modulatory effects were investigated in mouse livers following a daily oral dose of atractylodin or ß-eudesmol at 100 mg/kg body weight for 1, 7, 14, and 21 days. The inhibitory effects of both compounds on all rCYP450s were weak (IC50: 167 to >686 µM). ß-Eudesmol showed the most potent inhibitory effect on rCYP2C19 (IC50 = 172.7 µM) and rCYP3A4 (IC50 = 218.6 µM). Results of the ex vivo study showed that short exposure (1-7 days) of atractylodin and ß-eudesmol resulted in the upregulation of mRNA. Prolonged exposure to the daily oral dose for at least 14 days significantly downregulated the expressions of mRNA and proteins, which correlated with the decrease in the activities of mCYP1A2 and mCYP3A11. Based on the results of the ex vivo study, clinical uses of atractylodin or ß-eudesmol for the treatment of cholangiocarcinoma are of concern for the risk of toxicity due to hCYP3A4 inhibition following chronic dosing, as well as the metabolic interaction with the coadministered drugs that are metabolized by hCYP3A4.

Isolation, characterisation, and expression profiling of DXS and DXR genes in Atractylodes lancea.

1-Deoxy-d-xylulose-5-phosphate synthase and 1-deoxy-d-xylulose-5-phosphate reductoismerase are considered two key enzymes in the 2-C-methyl-d-erythritol-4-phosphate pathway of terpenoid biosynthesis and are related to the synthesis and accumulation of sesquiterpenoids. We cloned two DXS and DXR genes from Atractylodes lancea and analysed their expression in different tissues and in response to methyl jasmonate (MeJA). Subcellular localisation analysis revealed that the AlDXS and AlDXR1 proteins are located in the chloroplasts and cytoplasm, whereas AlDXR2 is only located in the chloroplasts. pET-AlDXS-28a and pGEX-AlDXR-4T-1 were expressed in Escherichia coli BL21(DE3) and BL21, respectively. Based on the abiotic stress analysis, the growth rate of the recombinant pGEX-AlDXR-4T-1 was higher than that of the control in HCl and NaOH. AlDXS exhibited the highest expression level in rhizomes of A. lancea from Hubei but was highest in leaves from Henan. In contrast, AlDXR showed maximum expression in the leaves of A. lancea from Hubei and Henan. Moreover, DXS and DXR gene expression, enzyme activities, and antioxidant enzyme activities oscillated in response to MeJA, with expression peaks appearing at different time points. Our findings indicated that the characterisation and function of AlDXS and AlDXR could be useful for further elucidating the functions of DXR and DXR genes in A. lancea.

Atractylodes-I Overcomes the Oxidative Stress-induced Colonic Mucosal Epithelial Cells Dysfunction to Prevent Irritable Bowel Syndrome Via Modulating the miR-34a-5p-LDHA Signaling Pathway.

BACKGROUND: Irritable bowel syndrome (IBS) is a known brain-gut disorder. Currently, the molecular and cellular mechanisms of IBS remain unclear. Atractylenolide-I (ATL-I) is a majorly bioactive component extracted from Rhizoma Atractylodes Macrocephalae. METHODS: Studies have revealed that ATL-I functioned as an anti-tumor drug in various cancers. However, the effects and molecular mechanisms of ATL-I on the pathological processes of colonic mucosal epithelial cells (CMECs) during IBS remain unclear. This study reports ATL-I effectively alleviated the oxidative stress-induced colonic mucosal epithelial cell dysfunction. In colonic mucosal tissues from IBS patients, we detected upregulated miR-34a-5p and suppressed glucose metabolism enzyme expressions. Under H2O2 treatment which mimics in vitro oxidative stress, miR-34a-5p was induced and glucose metabolism was inhibited in the colon mucosal epithelial cell line, NCM460. Meanwhile, ATL-I treatment effectively overcame the oxidative stress-induced miR-34a- 5p expression and glucose metabolism in NCM460 cells. RESULT: By bioinformatics analysis, Western blot and luciferase assay, we illustrated that miR-34a-5p directly targeted the 3'UTR region of glucose metabolism key enzyme, lactate dehydrogenase-A (LDHA) in colonic mucosal epithelial cells. Rescue experiments validated that miR-34a-5p inhibited glucose metabolism by targeting LDHA. Finally, we demonstrated that ATL-I treatment reversed the miR-34a-5p-inhibited glucose metabolism and -exacerbated colonic mucosal epithelial cell dysfunction under oxidative stress by modulating the miR-34a-5p-LDHA pathway. CONCLUSION: Summarily, our study reports the roles and mechanisms of ATL-I in the oxidative stress-induced colonic mucosal epithelial cell dysfunction during IBS through regulating the miR-34a-5p-LDHA-glucose metabolism axis.

The Anti-hyperuricemia and Anti-inflammatory Effects of Atractylodes Macrocephala in Hyperuricemia and Gouty Arthritis Rat Models.

AIMS: Atractylodes macrocephala is a traditional Chinese medicine with a variety of pharmacological activities. This study aimed to evaluate its anti-hyperuricemia and antiinflammatory effects on gout, and to preliminarily explore its mechanism. METHODS: The hyperuricemia rat model was established by intraperitoneal injection of oteracil potassium and intragastric gavage of yeast powder solution. And the acute gouty arthritis (GA) model was established by injecting monosodium urate (MSU) suspension. In the study of the antihyperuricemia effect of Atractylodes macrocephala, the healthy male Sprague-Dawley rats were randomly divided into the blank group, hyperuricemia group allopurinol group as well as low, moderate and high dose groups of Atractylodes macrocephala decoction (N=8 rats in each group). Serum, liver and kidney tissue samples were collected from each group. Serum uric acid (UA), adenosine deaminase (ADA) and xanthine oxidase (XOD) levels in each group were detected by enzyme-linked immunosorbent assay (ELISA). Protein levels of ADA and XOD in liver tissues were detected by Western blot, and renal histological changes were observed by Hematoxylin-eosin (H&E) and Masson staining. In order to investigate the anti-inflammatory effect of Atractylodes macrocephala, the healthy male Sprague-Dawley rats were randomly divided into the blank group, GA group, colchicine group, high, moderate and low dose groups of Atractylodes macrocephala decoction (N=8 rats in each group), and serum and synovial tissue of each group were collected. Then the level of serum interleukin (IL)-1ß and tumor necrosis factor (TNF)-α was observed by ELISA, and the histological changes of synovial tissue were observed by H&E staining. Besides, the expression of adenosine monophosphate- activated protein kinase (AMPK) /silent information regulator (SIRT) 1/ nuclear factor kappa B (NF-κB) protein in synovial tissue was observed by Western blot and immunohistochemistry. The markers of M1 and M2 macrophages, inducible nitric oxide synthase (iNOS) and arginase-1 (ARG1) were observed by Western blot and immunofluorescence. RESULTS: Atractylodes macrocephala could reduce the production of UA by inhibiting the level of ADA and XOD, and could improve renal injury and fibrosis. In addition, Atractylodes macrophages could reduce the levels of IL-1ß and TNF-α, activate AMPK/SIRT1 signaling pathway, and inhibit the activation of NF-κB and the polarization of macrophages to a pro-inflammatory phenotype. CONCLUSION: Atractylodes macrocephala shows good anti-hyperuricemic and anti-inflammatory effects, and its anti-inflammation pharmacological activity may be related to the inhibition of M1 macrophage polarization and NF-κB activation through activating AMPK/SIRT1.

Atractylodes lancea volatile oils target ADAR2-miR-181a-5p signaling to mesenchymal stem cell chondrogenic differentiation.

Atractylodeslancea Rhizoma (Rhizoma atractylodis [RA]) has long been recommended for the treatment of arthritis in traditional Chinese medicine, but its mechanism of action is still unclear. RA contains a large amount of Atractylodes lancea volatile oils (Atr). In this study, we investigated whether Atr can promote mesenchymal stem cells (MSCs) chondrogenic differentiation. The Atr were extracted from RA by steam distillation method, and the effect of Atr on MSCs was detected by the CCK8 assay. The optimal concentration of Atr for MSCs cultivation was 3 µg/ml. The differentially expressed miR-181a-5p was screened by miRNA microarray assay, and its mimics and inhibitors were transfected into MSCs. It was found that the inhibitor of miR-181a-5p could upregulate cartilage-specific genes such as SOX9, COL2A1, and ACAN. Meanwhile, we also found that the expression of gene editing enzyme ADAR2 was significantly increased in the chondrogenic differentiation of MSCs induced by Atr, and the bases of precursor sequence of miR-181a-5p were changed from A to G. After ADAR2 deletion, the expression of cartilage-specific genes was significantly down-regulated and the precursor sequence bases of miR-181a-5p were not changed. Bioinformatics analysis revealed that the predicted target gene of miR-181a-5p was yingyang1 (YY1), and the targeting relationship was verified by dual-luciferase reporter assay. After deleting YY1, the expression of cartilage-specific genes was significantly down-regulated. In conclusion, our study demonstrated that Atr can promote chondrogenic differentiation of MSC through regulation of the ADAR2-miR-181a-5p signaling pathway. This may provide a new insight into the possible mechanism of traditional Chinese medicine (Atr) in treating inflammatory joint diseases.

Mild shading promotes sesquiterpenoid synthesis and accumulation in Atractylodes lancea by regulating photosynthesis and phytohormones.

Atractylodes lancea rhizome (AR) has high medicinal and economic value. A previous study has reported that the accumulation of sesquiterpenoids in AR has obvious advantages under bamboo canopy. A concrete shade value to promote the cultivation of high-quality AR has not been established. In this study, 80% shading was screened at six different light intensities (100%, 80%, 60%, 40%, 20%, 7%), and the mechanism was explored in terms of photosynthetic efficiency and phytohormones levels. The results indicated that the total sesquiterpenoid content of 80% mild shading increased by 58%, 52%, and 35%, respectively, compared to 100% strong light in seedling, expansion, and harvest stages and increased by 144%, 178%, and 94%, respectively, compared with 7% low light. The sesquiterpenoids hinesol and ß-eudesmol contributed approximately 70% to the differential contribution ratio between mild shading and strong light (100%) or between mild shading and low light (7%). Furthermore, HMGR, DXR, and FPPS genes, which regulate sesquiterpenoid synthesis, were significantly upregulated in 80% mild shading. Transpiration rate; the intercellular CO2 concentration; net photosynthetic rate; and levels of jasmonic acid, abscisic acid, and gibberellin were strongly correlated (r > 0.85) with sesquiterpenoid accumulation. Cis-acting elements responding to light and phytohormones were present within the promoter regions of HMGR, DXR, and FPPS. Therefore, 80% shading promotes the synthesis and accumulation of sesquiterpenoids in AR by regulating photosynthetic efficiency and phytohormone production, thereby promoting transcriptional expression.

Chloroform extracts of Atractylodes chinensis inhibit the adhesion and invasion of Salmonella typhimurium.

There are 27 million cases of Salmonella Typhimurium (STM) reported worldwide annually, which have resulted in 217,000 deaths to date. Thus, there is an urgent requirement to develop novel antibacterial agents to target the multidrug-resistant strains of STM. We evaluated the inhibitory effect of the chloroform extracts of Atractylodes chinensis (Ac-CE) on the virulence of STM in vitro and develop it as a potential antibacterial agent. First, we determined the in vitro effects of Ac-CE on STM biofilm formation, and swimming, swarming, and adhesion to mucin. Further, we evaluated the effect of Ac-CE on the adhesion and invasion of STM at the gene level. Lastly, we evaluated the inhibitory effect of Ac-CE on STM infectivity at the cellular level. Ac-CE could attenuate both the adhesion and invasion abilities of STM in vitro. At the gene level, it could inhibit the expression of flagella, pilus, biofilm, SPI-1, and SPI-2 genes, which are related to the adhesion and invasion ability of STM in cells. Ac-CE significantly downregulated the expression of inflammatory cytokines and the TLR4/MyD88/NF-κB pathway in an STM infection cell model. It also significantly recovered the expression of intestinal barrier-related genes and proteins in intestinal cells that are damaged during STM infection. Ac-CE is effective as an antivirulence agent in alleviating STM infection. Although the main components of Ac-CE were analyzed.We have not demonstrated the antivirulence effect of the active ingredients in Ac-CE. And the antivirulence effect of Ac-CE and its active ingredients warrant further in vivo studies.

Apoptotic and Anti-metastatic Effects of Atractylodes lancea (Thunb.) DC. in a Hamster Model of Cholangiocarcinoma.

OBJECTIVES: Cholangiocarcinoma (CCA) is a highly aggressive tumor with a greater risk of distant metastasis. The promising anti-CCA activity and safety profile of Atractylodes lancea (AL) have previously been reported in a series of in vitro, in vivo and clinical studies. The present study investigated the effect of AL extract on apoptosis and metastasis signaling pathways in the Opisthorchis viverrini/dimethylnitrosamine (OV/DMN)-induced CCA hamster model. MATERIALS AND METHODS: Hamster liver tissues were obtained from the four groups (n = 5 per group), i.e., (i) 5-FU treated CCA (40 µg/mL); (ii) CCA; (iii) AL-treated CCA (5,000 mg/kg), and (iv) normal hamsters. Total RNA was isolated, and the expression levels of apoptosis-related and metastasis-related genes were determined by qRT-PCR analysis. RESULTS: The expression levels of p16, caspase-3, caspase-8, caspase-9, Apaf-1, p53 and Eef1a1 were downregulated, while that of the remaining genes were upregulated in CCA hamsters compared with normal hamsters. AL treatment increased the expression of p16, caspase-9, caspase-3, Apaf-1, p53 and E-cadherin and decreased the expression of cyclin D1, cdk4, Bax, Akt/PKB, Bcl-2, Mfge-8, Lass4, S100A6, TGF-ß, Smad-2, Smad-3, Smad-4, MMP-9, and N-cadherin. The expression of Eef1a1 was unchanged. CONCLUSION: The anti-CCA activity of AL in OV/DMN-induced CCA hamsters could be due to the induction of cell cycle arrest at the G1 phase and activation of the apoptosis pathway, resulting in cancer cell death. The activation of the apoptosis pathway mainly involved the intrinsic pathway (activation of caspase-3 and caspase-9 through p53 and Mfge-8 modulation and downregulation of anti-apoptotic genes Akt and Bcl-2). In addition, AL could also inhibit the canonical TGF-ß signaling pathway, MMP-9 and N-cadherin to suppress tumor metastasis.

Differences in gene expression and endophytic bacterial diversity in Atractylodes macrocephala Koidz. rhizomes from different growth years.

Atractylodes macrocephala Koidz. (AMK) is widely used in traditional Chinese medicine owing to its pharmacological activity. Here, we aimed to characterize the differentially expressed genes (DEGs) of one- and three-year growth (OYG and TYG) rhizomes of AMK, combined with endophytic bacterial diversity analysis using high-throughput RNA sequencing. A total of 114 572 unigenes were annotated using six public databases. In all, 3570 DEGs revealed a clear difference, of which 936 and 2634 genes were upregulated and downregulated, respectively. The results of KEGG pathway analysis indicated that DEGs corresponding to terpenoid synthesis gene were downregulated in TYG rhizomes. In addition, 414 424 sequences corresponding to the 16S rRNA gene were divided into 1267 operational taxonomic units (OTUs). Moreover, the diversity of endophytic bacteria changed with species in the OYG (773) and TYG (1201) rhizomes at the OTU level, and Proteobacteria, Actinobacteria, and Bacteroidetes were the dominant phyla. A comparison of species differences among different growth years revealed that some species were significantly different, such as Actinomycetes, Variovorax, and Cloacibacterium. Interestingly, the decrease in the function-related metabolism of terpenoids and polyketides was correlated with the low expression of terpene synthesis genes in TYG rhizomes, as assessed using PICRUSt2. These data provide a scientific basis for elucidating the mechanisms underlying metabolite accumulation and endophytic bacterial diversity in relation to the growth years in AMK.

Cloning, Prokaryotic Expression, and Purification of Acetyl-CoA C-Acetyltransferase from Atractylodes lancea.

BACKGROUND: Cangzhu (Atractylodes lancea), a valuable and common traditional Chinese medicinal herb, is primarily used as an effective medicine with various health-promoting effects. The main pharmacological bioactive ingredients in the rhizome of A. lancea are terpenoids. Acetyl-CoA C-acetyltransferase (AACT) is the first enzyme in the terpenoid synthesis pathway and catalyzes two units of acetyl-CoA into acetoacetyl-CoA. OBJECTIVE: The objective of the present work was to clone and identify function of AlAACT from Atractylodes lancea. METHODS: A full-length cDNA clone of AlAACT was isolated using PCR and expressed in Escherichia coli. The expressed protein was purified using Ni-NTA agarose column using standard protocols. AlAACT was transiently expressed in N. benthamiana leaves to determine their subcellular location. The difference in growth between recombinant bacteria and control bacteria under different stresses was observed using the droplet plate experiment. RESULTS: In this study, a full-length cDNA of AACT (AlAACT) was cloned from A. lancea, which contains a 1,227 bp open reading frame and encodes a protein with 409 amino acids. Bioinformatic and phylogenetic analysis clearly suggested that AlAACT shared high similarity with AACTs from other plants. The recombinant protein pET32a(+)/AlAACT was successfully expressed in Escherichia coli BL21 (DE3) cells induced with 0.4 mM IPTG at 30°C as the optimized condition. The recombinant enzyme pET-32a-AlAACT was purified using the Ni-NTA column based on the His-tag, and the molecular weight was determined to be 62 kDa through SDS-PAGE and Western Blot analysis. The recombinant protein was eluted with 100, 300, and 500 mM imidazole; most of the protein was eluted with 300 mM imidazole. Under mannitol stress, the recombinant pET-32a- AlAACT protein showed a substantial advantage in terms of growth rates compared to the control. However, this phenomenon was directly opposite under NaCl abiotic stress. Subcellular localization showed that AlAACT localizes to the nucleus and cytoplasm. CONCLUSION: The expression and purification of recombinant enzyme pET-32a-AlAACT were successful, and the recombinant strain pET-32a-AlAACT in showed better growth in a drought stress. The expression of AlAACT-EGFP fusion protein revealed its localization in both nuclear and cytoplasm compartments. This study provides an important foundation for further research into the effects of terpenoid biosynthesis in A. lancea.

Polysaccharides from Chrysanthemun indicum L. enhance the accumulation of polysaccharide and atractylenolide in Atractylodes macrocephala Koidz.

Atractylodes macrocephala Koidz. (AM), an herb of traditional Chinese medicine, is well-known for anti-oxidant, anti-tumor and immune regulation potential. However, it is low bioactive compound content that restricts the application of this species. Elicitation is considered as an effective method to enhance biomass and bioactive compound in plants. Our precious study found that polysaccharide of Chrysanthemun indicum L. could promote plant growth by triggering plant defense. In the present study, polysaccharide of Chrysanthemun indicum L. is used to stimulate the accumulation of biomass and bioactive compound with different concentration in Atractylodes macrocephala Koidz. during pot, plot and field experiments. The results suggested that polysaccharide of Chrysanthemun indicum L. could significantly enhance the accumulation of biomass, atractylenolides and polysacchrides. Moreover, 2 mg/mL is determined and verified to be the appropriate concentration during field experiments. In addition, RT-qPCR revealed that CIP-induced terpenoid synthesis in AM mainly depended on mevalonate (MVA) pathway. This is the first report on the discovery of polysaccharide of Chrysanthemun indicum L. for the enhanced accumulation of biaomass and bioactive compound and the use of its for agricultural production.

Atractylodes chinensis Water Extract Ameliorates Obesity via Promotion of the SIRT1/AMPK Expression in High-Fat Diet-Induced Obese Mice.

Obesity remains a continuing global health concern, as it is associated with an increased risk of many chronic diseases. Atractylodes chinensis Koidz. (Ac) is traditionally used in the treatment of inflammatory diseases, such as arthritis, hepatitis, and gastric ulcers. Despite the diverse pharmacological activities of Ac, scientific evidence for the use of Ac in obesity is still limited. Therefore, the present study aimed to determine the anti-obesity effects of Ac. C57BL/6N mice were divided into five groups as follows: chow diet group (CON), 45% HFD group, HFD + oral administration of orlistat group, and HFD + oral administration of Ac groups. RT-PCR and western blotting were used to examine the expression of molecules relating to obesity progression. Ac-administered mice showed dramatically decreased body weight and weight gain compared to the high-fat diet (HFD)-fed mice. In addition, Ac administration attenuated the protein expression levels of adipogenic transcription factors in the white adipose tissue (WAT) and livers of HFD-fed mice. Furthermore, Ac administration declined the expression levels of lipogenic genes, while enhancing those of the fatty acid oxidation genes in the WAT of HFD-fed mice. Importantly, Ac administration highly upregulated the AMP-activated kinase (AMPK) and sirtuin 1 (SIRT1) expression levels in WAT of the HFD-induced obese mouse model. Our results provide evidence that Ac can effectively ameliorate weight gain and adipose tissue expansion.

Comparative transcriptome analysis reveals sesquiterpenoid biosynthesis among 1-, 2- and 3-year old Atractylodes chinensis.

BACKGROUND: Atractylodes chinensis (DC.) Koidz is a well-known medicinal plant containing the major bioactive compound, atractylodin, a sesquiterpenoid. High-performance liquid chromatography (HPLC) analysis demonstrated that atractylodin was most abundant in 3-year old A. chinensis rhizome, compared with those from 1- and 2-year old rhizomes, however, the molecular mechanisms underlying accumulation of atractylodin in rhizomes are poorly understood. RESULTS: In this study, we characterized the transcriptomes from rhizomes of 1-, 2- and 3-year old (Y1, Y2 and Y3, respectively) A. chinensis, to identify differentially expressed genes (DEGs). We identified 240, 169 and 131 unigenes encoding the enzyme genes in the mevalonate (MVA), methylerythritol phosphate (MEP), sesquiterpenoid and triterpenoid biosynthetic pathways, respectively. To confirm the reliability of the RNA sequencing analysis, eleven key gene encoding factors involved in the sesquiterpenoid and triterpenoid biosynthetic pathway, as well as in pigment, amino acid, hormone and transcription factor functions, were selected for quantitative real time PCR (qRT-PCR) analysis. The results demonstrated similar expression patterns to those determined by RNA sequencing, with a Pearson's correlation coefficient of 0.9 between qRT-PCR and RNA-seq data. Differential gene expression analysis of rhizomes from different ages revealed 52 genes related to sesquiterpenoid and triterpenoid biosynthesis. Among these, seven DEGs were identified in Y1 vs Y2, Y1 vs Y3 and Y2 vs Y3, of which five encoded four key enzymes, squalene/phytoene synthase (SS), squalene-hopene cyclase (SHC), squalene epoxidase (SE) and dammarenediol II synthase (DS). These four enzymes directly related to squalene biosynthesis and subsequent catalytic action. To validate the result of these seven DEGs, qRT-PCR was performed and indicated most of them displayed lower relative expression in 3-year old rhizome, similar to transcriptomic analysis. CONCLUSION: The enzymes SS, SHC, SE and DS down-regulated expression in 3-year old rhizome. This data corresponded to the higher content of sesquiterpenoid in 3-year old rhizome, and confirmed by qRT-PCR. The results of comparative transcriptome analysis and identified key enzyme genes laid a solid foundation for investigation of production sesquiterpenoid in A. chinensis.

Effect of drought on photosynthesis, total antioxidant capacity, bioactive component accumulation, and the transcriptome of Atractylodes lancea.

BACKGROUND: Atractylodes lancea (Thunb.) DC, a medicinal herb belonging to the Asteraceae family, often faces severe drought stress during its growth. Until now, there has been no research on the effect of drought stress on the quality formation of A. lancea. Therefore, the present study aimed to study the effects of drought stress on A. lancea through physical and chemical analysis, and to reveal the related molecular mechanisms via transcriptome analysis. RESULTS: The photosynthesis was markedly inhibited under drought stress. There were alterations to photosynthetic parameters (Pn, Gs, Ci) and chlorophyll fluorescence (Fv/Fm, NPQ), and the chlorophyll content decreased. Twenty genes encoding important regulatory enzymes in light and dark reactions, including the Rubisco gene of the Calvin cycle, were significantly downregulated. After exposure to drought stress for more than 4 days, the activities of four antioxidative enzymes (SOD, POD CAT and APX) began to decrease and continued to decrease with longer stress exposure. Meanwhile, most of the genes encoding antioxidative enzymes were downregulated significantly. The downregulation of 21 genes related to the respiratory electron transport chain indicated that the blocked electron transfer accelerated excessive ROS. The MDA content was significantly elevated. The above data showed that 15 days of drought stress caused serious oxidative damage to A. lancea. Drought stress not only reduced the size and dry weight of A. lancea, but also lowered the amount of total volatile oil and the content of the main bioactive components. The total volatile oil and atractylodin content decreased slightly, whereas the content of atractylon and ß-eudesmol decreased significantly. Moreover, ten significantly downregulated genes encoding sesquiterpene synthase were mainly expressed in rhizomes. CONCLUSIONS: After exposed to drought stress, the process of assimilation was affected by the destruction of photosynthesis; stress tolerance was impaired because of the inhibition of the antioxidative enzyme system; and bioactive component biosynthesis was hindered by the downregulation of sesquiterpene synthase-related gene expression. All these had negative impacts on the quality formation of A. lancea under drought stress.

Therapeutic potential and pharmacological activities of ß-eudesmol.

Herbal medicines are attracting the attention of researchers worldwide. ß-Eudesmol is one of the most studied and major bioactive sesquiterpenes, mainly extracted from Atractylodes lancea (Thunb) DC. rhizomes. It has potential anti-tumor and anti-angiogenic activities and is an inhibitor of tumor growth by inhibiting angiogenesis by suppressing CREB activation of the growth factor signaling pathway. It also stimulates neurite outgrowth in rat pheochromocytoma cells with activation of mitogen-activated protein kinases. It may be a promising lead compound for enhancing neural function, and it may help to explain the underlying mechanisms of neural differentiation. In this review, we summarized the currently available clinical and preclinical studies describing the therapeutic applications of ß-eudesmol.

Newly Generated Atractylon Derivatives in Processed Rhizomes of Atractylodes macrocephala Koidz.

Thermally processed rhizomes of Atractylodes macrocephala (RAM) have a long history of use in traditional Chinese medicine (TCM) for treating various disorders, and have been an integral part of various traditional drugs and healthcare products. In TCM, herbal medicines are, in most cases, uniquely processed. Although it is thought that processing can alter the properties of herbal medicines so as to achieve desired functions, increase potency, and/or reduce side effects, the underlying chemical changes remain unclear for most thermally processed Chinese herbal medicines. In an attempt to shed some light on the scientific rationale behind the processes involved in traditional medicine, the RAM processed by stir-frying with wheat bran was investigated for the change of chemical composition. As a result, for the first time, five new chemical entities, along with ten known compounds, were isolated. Their chemical structures were determined by spectroscopic and spectrometric analyses. The possible synthetic pathway for the generation of such thermally-induced chemical entities was also proposed. Furthermore, biological activity evaluation showed that none of the compounds possessed cytotoxic effects against the tested mammalian cancer and noncancer cell lines. In addition, all compounds were ineffective at inhibiting the growth of the pathogenic microorganisms.

Alisma Shugan Decoction (ASD) Ameliorates Hepatotoxicity and Associated Liver Dysfunction by Inhibiting Oxidative Stress and p65/Nrf2/JunD Signaling Dysregulation In Vivo.

BACKGROUND Liver fibrosis, defined as the aberrant accumulation of extracellular matrix (ECM) proteins such as collagen in the liver, is a common feature of chronic liver disease, and often culminates in portal hypertension, liver cirrhosis, and hepatic failure. Though therapeutically manageable, fibrosis is not always successfully treated by conventional antifibrotic agents. While the traditional Chinese medicine (TCM) Alisma Shugan Decoction (ASD) has several health benefits, including anti-inflammation, anti-oxidation, and limitation of cardiovascular and respiratory disorders, it remains unclear if it has any hepato-protective potential. MATERIAL AND METHODS The present study examined the therapeutic effect of ASD in thioacetamide (TAA)-induced liver injury and fibrosis rat models. RESULTS We demonstrated that 50 mg/kg ASD significantly reversed TAA-induced elevation of alanine or aspartate transaminase levels, elicited no dyscrasia, and conferred a 40% (p<0.01) or 20% (p<0.05) survival advantage, compared to rats treated with TAA or TAA+ASD, respectively. Treatment with ASD reversed TAA-induced liver injury and fibrogenesis via repression of alpha-SMA protein and reduction of the collagen area and fibrosis score. Concurrently, ASD markedly suppressed the mRNA expression of fibrogenic procollagen, ICAM-1, MMP2, MMP9, and MMP13, and production of TIMP-1, ICAM-1, CXCL7, or CD62L cytokine in rat liver injury models. Interestingly, ASD-elicited reduction of liver injury and fibrogenesis was mediated by dysregulated p65/NrF-2/JunD signaling, with a resultant 3.18-fold (p<0.05) increase in GSH/GSSH ratio, and a 3.61-fold (p<0.01) or 1.51-fold (p<0.01) reduction in the 4-hydroxynonenal and malondialdehyde (MDA) levels, respectively, indicating reduced oxidative stress in the ASD-treated rats, and suggesting an hepato-protective role for ASD. CONCLUSIONS In conclusion, the present study provides supplementary evidence of the therapeutic benefit of ASD as an efficient treatment option in cases of liver injury and fibrosis. Further large-cohort validation of these findings is warranted.