Medicinal Plant Artemisia Annua Research Articles

Differential Anti-Fibrotic and Remodeling Responses of Human Dermal Fibroblasts to Artemisia sp., Artemisinin, and Its Derivatives.

Fibrosis is a ubiquitous pathology, and prior studies have indicated that various artemisinin (ART) derivatives (including artesunate (AS), artemether (AM), and dihydroartemisinin (DHA)) can reduce fibrosis in vitro and in vivo. The medicinal plant Artemisia annua L. is the natural source of ART and is widely used, especially in underdeveloped countries, to treat a variety of diseases including malaria. A. afra contains no ART but is also antimalarial. Using human dermal fibroblasts (CRL-2097), we compared the effects of A. annua and A. afra tea infusions, ART, AS, AM, DHA, and a liver metabolite of ART, deoxyART (dART), on fibroblast viability and expression of key fibrotic marker genes after 1 and 4 days of treatment. AS, DHA, and Artemisia teas reduced fibroblast viability 4 d post-treatment in up to 80% of their respective controls. After 4 d of treatment, AS DHA and Artemisia teas downregulated ACTA2 up to 10 fold while ART had no significant effect, and AM increased viability by 10%. MMP1 and MMP3 were upregulated by AS, 17.5 and 32.6 fold, respectively, and by DHA, 8 and 51.8 fold, respectively. ART had no effect, but A. annua and A. afra teas increased MMP3 5 and 16-fold, respectively. Although A. afra tea increased COL3A1 5 fold, MMP1 decreased >7 fold with no change in either transcript by A. annua tea. Although A. annua contains ART, it had a significantly greater anti-fibrotic effect than ART alone but was less effective than A. afra. Immunofluorescent staining for smooth-muscle α-actin (α-SMA) correlated well with the transcriptional responses of drug-treated fibroblasts. Together, proliferation, qPCR, and immunofluorescence results show that treatment with ART, AS, DHA, and the two Artemisia teas yield differing responses, including those related to fibrosis, in human dermal fibroblasts, with evidence also of remodeling of fibrotic ECM.

Advanced metabolic engineering strategies for increasing artemisinin yield in Artemisia annua L.

Artemisinin, also known as 'Qinghaosu', is a chemically sesquiterpene lactone containing an endoperoxide bridge. Due to the high activity to kill Plasmodium parasites, artemisinin and its derivatives have continuously served as the foundation for antimalarial therapies. Natural artemisinin is unique to the traditional Chinese medicinal plant Artemisia annua L., and its content in this plant is low. This has motivated the synthesis of this bioactive compound using yeast, tobacco, and Physcomitrium patens systems. However, the artemisinin production in these heterologous hosts is low and cannot fulfil its increasing clinical demand. Therefore, A. annua plants remain the major source of this bioactive component. Recently, the transcriptional regulatory networks related to artemisinin biosynthesis and glandular trichome formation have been extensively studied in A. annua. Various strategies including (i) enhancing the metabolic flux in artemisinin biosynthetic pathway; (ii) blocking competition branch pathways; (iii) using transcription factors (TFs); (iv) increasing peltate glandular secretory trichome (GST) density; (v) applying exogenous factors; and (vi) phytohormones have been used to improve artemisinin yields. Here we summarize recent scientific advances and achievements in artemisinin metabolic engineering, and discuss prospects in the development of high-artemisinin yielding A. annua varieties. This review provides new insights into revealing the transcriptional regulatory networks of other high-value plant-derived natural compounds (e.g., taxol, vinblastine, and camptothecin), as well as glandular trichome formation. It is also helpful for the researchers who intend to promote natural compounds production in other plants species.

Phytochemical profile and anti-inflammatory effect of Artemisia annua L. essential oil

Plant-derived natural compounds are desired for their anti-inflammatory properties. We investigated the chemical composition and the anti-inflammatory effects of the essential oil obtained from leaves of the Chinese medicinal plant Artemisia annua L. Artemisia annua L. essential oil (AO) was obtained by hydrodistillation; the extraction yield was approximately 1.2%. Gas chromatography–mass spectrometry (GC–MS) revealed the presence of 13 compounds in AO; the major compounds were caryophyllene (32.16%), eucalyptol (28.62%), α-pinene (19.08%), and β-pinene (14.72%). Cell counting kit-8 (CCK8) assay revealed that AO did not show in vitro cytotoxicity toward RAW264.7 cells and suppressed the expression of nitric oxide (NO), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) in lipopolysaccharide (LPS)-induced RAW264.7 cells. Pretreatment with AO significantly reduced xylene-induced inflammation in the ears of Kunming mice. AO inhibited inflammatory cell infiltration, decreased epidermis thickness in the ear, and suppressed expression of TNF-α in the serum. The in vivo anti-inflammatory activity of AO was similar to that of the positive control compound Di Song Zhang Bo Cream. Overall, AO was observed to be an effective anti-inflammatory agent. These results provide an experimental basis for commercializing AO-derived external preparations for pharmacological and cosmetic industries.

Graphene enhances artemisinin production in the traditional medicinal plant Artemisia annua via dynamic physiological processes and miRNA regulation

We investigated the effects of graphene on the model herb Artemisia annua, which is renowned for producing artemisinin, a widely used pharmacological compound. Seedling growth and biomass were promoted when A. annua was cultivated with low concentrations of graphene, an effect which was attributed to a 1.4-fold increase in nitrogen uptake, a 15%–22% increase in chlorophyll fluorescence, and greater abundance of carbon cycling–related bacteria. Exposure to 10 or 20 mg/L graphene resulted in a ∼60% increase in H2O2, and graphene could act as a catalyst accelerator, leading to a 9-fold increase in catalase (CAT) activity in vitro and thereby maintaining reactive oxygen species (ROS) homeostasis. Importantly, graphene exposure led to an 80% increase in the density of glandular secreting trichomes (GSTs), in which artemisinin is biosynthesized and stored. This contributed to a 5% increase in artemisinin content in mature leaves. Interestingly, expression of miR828 was reduced by both graphene and H2O2 treatments, resulting in induction of its target gene AaMYB17, a positive regulator of GST initiation. Subsequent molecular and genetic assays showed that graphene-induced H2O2 inhibits micro-RNA (miRNA) biogenesis through Dicers and regulates the miR828–AaMYB17 module, thus affecting GST density. Our results suggest that graphene may contribute to yield improvement in A. annua via dynamic physiological processes together with miRNA regulation, and it may thus represent a new cultivation strategy for increasing yield capacity through nanobiotechnology.

Attenuation of quorum sensing regulated virulence functions and biofilm of pathogenic bacteria by medicinal plant Artemisia annua and its phytoconstituent 1, 8-cineole.

The emergence of multidrug resistance (MDR) in bacterial pathogens is a serious public health concern. A significant therapeutic target for MDR infections is the quorum sensing-regulated bacterial pathogenicity. Determining the anti-quorum sensing abilities of certain medicinal plants against bacterial pathogens as well as the in-silico interactions of particular bioactive phytocompounds with QS and biofilm-associated proteins were the objectives of the present study. In this study, 6 medicinal plants were selected based on their ethnopharmacological usage, screened for Anti-QS activity and Artemisia annua leaf extract (AALE) demonstrated pigment inhibitory activity against Chromobacterium violaceum CV12472. Further, the methanol active fraction significantly inhibited the virulence factors (pyocyanin, pyoverdine, rhamnolipid and swarming motility) of Pseudomonas aeruginosa PAO1 and Serratia marcescens MTCC 97 at respective sub-MICs. The inhibition of biofilm was determined using a microtiter plate test and scanning electron microscopy. Biofilm formation was impaired by 70%, 72% and 74% in P. aeruginosa, C. violaceum and S. marcescens, respectively at 0.5xMIC of the extract. The phytochemical content of the extract was studied using GC-MS and 1, 8-cineole was identified as major bioactive compound. Furthermore, 1, 8-cineole was docked with quorum sensing (QS) proteins (LasI, LasR, CviR, and rhlR) and biofilm proteins (PilY1 and PilT). In silico docking and dynamics simulations studies suggested interactions with QS-receptors CviR', LasI, LasR, and biofilm proteins PilY1, PilT for anti-QS activity. Further, 1, 8-cineole demonstrated 66% and 51% reduction in violacein production and biofilm formation, respectively to validate the findings of computational analysis. Findings of the present investigation suggests that 1, 8-cineole plays a crucial role in the QS and biofilm inhibitory activity demonstrated by Artemisia annua extract. RESEARCH HIGHLIGHTS: Artemisia annua leaf extract (AALE) methanol fraction demonstrated broad-spectrum QS and biofilm inhibition Scanning electron microscopy (SEM) confirmed biofilm inhibition Molecular docking and simulation studies suggested positive interactions of 1,8-cineol with QS-receptors and biofilm proteins.

Jasmonic acid responsive AaJRM1 transcription factor positively regulates the biosynthesis of anti-malarial drug artemisinin in Artemisia annua

The phytohormone jasmonic acid (JA) and environmental factor light have the ability to synergistically affect the biosynthesis of anti-malarial drug artemisinin in traditional Chinese medicinal plant Artemisia annua. Previous studies have screened a JA-responsive R2R3-MYB transcription factor, AaJRM1, as a candidate gene for regulating artemisinin biosynthesis by transcriptome sequencing after JA treatment in light signals. However, the function and regulatory mechanism of AaJRM1 on artemisinin biosynthesis have not been illus. In this study, transcriptional level analysis showed that the expression of AaJRM1 was the most highly expressed in trichomes and rapidly responded to JA treatment, which was similar to the expression profiles of the specific genes belonging to artemisinin biosynthesis. The regulatory potential of artemisinin biosynthesis using transgenic A. annua plants indicated that overexpression and suppression of AaJRM1 increased and decreased the accumulation of artemisinin, respectively. The expression analysis revealed that the transcriptions of four core biosynthetic genes were significantly increased in AaJRM1-OE plants and obviously declined in AaJRM1-EAR transgenic lines compared with the wild-type control. Dual-Luciferase assays demonstrated that AaJRM1 significantly activated the transcription of artemisinin biosynthetic genes. Therefore, this study elucidated the molecular mechanism of JA-responsive AaJRM1 transcription factor regulating artemisinin biosynthesis, and provided a theoretical basis for the use of biotechnology to cultivate new germplasm with high artemisinin content.

Genome-wide characterization of B-box gene family in Artemisia annua L. and its potential role in the regulation of artemisinin biosynthesis

Artemisinin is a sesquiterpene lactone synthesized by the Chinese medicinal plant Artemisia annua L., and is the principal component used for the treatment of malaria. Recent studies have demonstrated the potential role of B-box (BBX) transcription factors in regulating plant secondary metabolism. However, the roles and functions of BBX members in the biosynthesis of artemisinin in A. annua have not been reported yet. In this study, a total of 27 BBX members (AaBBXs) were identified and classified into five sub-groups (I-V). The AaBBXs in each sub-group shared similar gene structures, conserved domains and motifs. Additionally, multiple cis-acting elements which are responsible for light, hormones, and stress signals were found in the promoter regions of AaBBX members. Based on transcriptome data, 27 AaBBXs showed constitutive expression in different organs/tissues, while several BBX genes exhibited higher expression levels in glandular secretory trichomes (GSTs), the site of artemisinin biosynthesis. To explore the roles of AaBBX members in regulating the artemisinin biosynthesis, 7 AaBBXs (AaBBX5/6/8/15/22/23/24) were selected for further investigation. The expressions of all 7 selected AaBBXs were strongly induced by MeJA and ABA, and they were all found to be nuclear localized. Additionally, all seven proteins were found to have the ability to physically interact with AaJAZ8 in vitro. Dual-luciferase assays revealed that AaBBX5/6/8/15/22/23 can activate the promoter activity of the artemisinin biosynthetic pathway genes such as AaADS, AaCYP71AV1, AaDBR2 and AaALDH1 at different degrees. AaBBX22 showed highest expression level in A. annua GSTs, and can significantly activate the promoters of all four artemisinin biosynthetic pathway genes. Furthermore, transgenic assay results indicated that AaBBX22 acted as a positive regulator for the biosynthesis of artemisinin in A. annua. Taken together, this study provided a comprehensive genome-wide characterization of the BBX family and identified several significant BBX genes, including AaBBX22, which paves the way for further investigation into the regulatory role of the BBX gene family in artemisinin biosynthesis.

Anti-Mitochondrial and Insecticidal Effects of Artemisinin against Drosophila melanogaster.

Artemisinin (ART) is an endoperoxide molecule derived from the medicinal plant Artemisia annua L. and is clinically used as an antimalarial drug. As a secondary metabolite, the benefit of ART production to the host plant and the possible associated mechanism are not understood. It has previously been reported that Artemisia annua L. extract or ART can inhibit both insect feeding behaviors and growth; however, it is not known whether these effects are independent of each other, i.e., if growth inhibition is a direct outcome of the drug's antifeeding activity. Using the lab model organism Drosophila melanogaster, we demonstrated that ART repels the feeding of larvae. Nevertheless, feeding inhibition was insufficient to explain its toxicity on fly larval growth. We revealed that ART provoked a strong and instant depolarization when applied to isolated mitochondria from Drosophila while exerting little effect on mitochondria isolated from mice tissues. Thus, ART benefits its host plant through two distinct activities on the insect: a feeding-repelling action and a potent anti-mitochondrial action which may underlie its insect inhibitory activities.

Comparative assessment of hepatoprotective properties of Artesunate and flavonoids from Artemisia annua on acetaminophen and carbon tetrachloride-induced cytotoxicity in primary mice hepatocytes

BackgroundArtesunate (ART) is a semi-synthetized molecule from Artemisinin, an active compound isolated from the medicinal plant Artemisia annua, widely used for the treatment of malaria. Previous studies reported that ART may exert a dual effect on the liver. Accordingly, this study investigated the potential protective action of ART against Acetaminophen (APAP) and Carbon tetrachloride (CCl4)-induced hepatotoxicity in primary mice hepatocytes, in comparison to that of flavonoid extracted from A. annua (FAA). In addition, the antioxidant properties of FAA were also assessed. MethodsThe antioxidant activities of FAA and Ascorbic acid (ASC) (0.01–100 μg/mL) were assessed through inhibition of lipid peroxidation, reduction of ferric and phosphomolydenum, and hydroxyl and DPPH radicals scavenging assays. The hepatoprotective effects of FAA and ART (0.1–100 μg/mL) were evaluated against APAP (11 mM) or CCl4 (4 mM) induced oxidative damage in primary mouse hepatocytes. Biochemical parameters associated with hepatotoxicity assessed include cell viability, cell membrane integrity, cellular glutathione, and antioxidant enzyme activities. ResultsThe obtained finding revealed FAA displayed a remarkable antioxidant activities as evidenced by the low IC50/EC50 values (3.85–19.32 μg/mL), comparable to that of ASC (3.26–18.04 μg/mL). When tested at 10 μg/mL, both FAA and ART significantly (p˂0.05) preserved cell viability, inhibited alanine aminotransferase leakage and lipid membrane peroxidation, and restored superoxide dismutase and catalase activities and glutathione content induced by APAP or CCl4 in a similar way as Silymarin. However, ART showed a significant (p˂0.05) cytotoxic effect on hepatocytes at 100 and 1000 μg/mL and did not confer obvious protection at 100 μg/mL. ConclusionOverall, our data demonstrated that ART harms mice hepatocytes at high concentration while conferring relative protection against APAP and CCl4-hepatotoxicity at low concentration. In contrast, FAA effectively protects liver cells without cytotoxicity effect, event at 100 μg/mL. Accordingly, ART should be given to the patient only under a medical prescription.

Artemisia annua L. plants lacking Bornyl diPhosphate Synthase reallocate carbon from monoterpenes to sesquiterpenes except artemisinin.

The monoterpene camphor is produced in glandular secretory trichomes of the medicinal plant Artemisia annua, which also produces the antimalarial drug artemisinin. We have found that, depending on growth conditions, camphor can accumulate at levels ranging from 1- 10% leaf dry weight (LDW) in the Artemis F1 hybrid, which has been developed for commercial production of artemisinin at up to 1% LDW. We discovered that a camphor null (camphor-0) phenotype segregates in the progeny of self-pollinated Artemis material. Camphor-0 plants also show reduced levels of other less abundant monoterpenes and increased levels of the sesquiterpene precursor farnesyl pyrophosphate plus sesquiterpenes, including enzymatically derived artemisinin pathway intermediates but not artemisinin. One possible explanation for this is that high camphor concentrations in the glandular secretory trichomes play an important role in generating the hydrophobic conditions required for the non-enzymatic conversion of dihydroartemisinic acid tertiary hydroperoxide to artemisinin. We established that the camphor-0 phenotype associates with a genomic deletion that results in loss of a Bornyl diPhosphate Synthase (AaBPS) gene candidate. Functional characterization of the corresponding enzyme in vitro confirmed it can catalyze the first committed step in not only camphor biosynthesis but also in a number of other monoterpenes, accounting for over 60% of total volatiles in A. annua leaves. This in vitro analysis is consistent with loss of monoterpenes in camphor-0 plants. The AaBPS promoter drives high reporter gene expression in A. annua glandular secretory trichomes of juvenile leaves with expression shifting to non-glandular trichomes in mature leaves, which is consistent with AaBPS transcript abundance.

Evaluation of biological mechanisms of artemisinin on bovine mammary epithelial cells by integration of network pharmacology and TMT-based quantitative proteomics.

The sesquiterpene lactone, artemisinin, is a primary component of the medicinal plant Artemisia annua L., which has anti-inflammatory, antibacterial and antioxidant activities. However, the potential effects of artemisinin on the mammary gland of dairy cows and the underlying molecular mechanisms remain unclear. Here, we utilized systematic network pharmacology and proteomics to elucidate the mechanism by which artemisinin affects milk production and the proliferation of bovine mammary epithelial cells (BMECs). Nineteen bioactive compounds and 56 key targets were identified through database mining. To delineate the mechanism of artemisia’s activity, a protein-protein interaction network and integrated visual display were generated from bioinformatics assays to explore the relationships and interactions among the bioactive molecules and their targets. The gene ontology (GO) terms and kyoto encyclopedia of genes and genomes annotation suggested that the apoptotic process, cell division, p53 pathway, prolactin and PI3K-Akt pathways played vital roles in mammary gland development. Using proteomics analysis, we identified 122 up-regulated and 96 down-regulated differentially significant expressed proteins (DSEPs). The differentially significant expressed proteins had multiple biological functions associated with cell division, apoptosis, differentiation, and migration. Gene ontology enrichment analysis suggested that differentially significant expressed proteins may promote cell proliferation and regulate apoptosis in bovine mammary epithelial cells. Kyoto encyclopedia of genes and genomes pathway analysis indicated that several biological pathways, such as those involved in antigen processing and presentation, cell adhesion molecules and ribosomes, played significant roles in the effects of artemisinin on bovine mammary epithelial cells. These findings contribute to a comprehensive understanding of the mechanism by which artemisinin affects bovine mammary epithelial cells to improve mammary gland turnover by inducing cell proliferation and mammary gland development.

Artemisinins in Combating Viral Infections Like SARS-CoV-2, Inflammation and Cancers and Options to Meet Increased Global Demand.

Artemisinin is a natural bioactive sesquiterpene lactone containing an unusual endoperoxide 1, 2, 4-trioxane ring. It is derived from the herbal medicinal plant Artemisia annua and is best known for its use in treatment of malaria. However, recent studies also indicate the potential for artemisinin and related compounds, commonly referred to as artemisinins, in combating viral infections, inflammation and certain cancers. Moreover, the different potential modes of action of artemisinins make these compounds also potentially relevant to the challenges the world faces in the COVID-19 pandemic. Initial studies indicate positive effects of artemisinin or Artemisia spp. extracts to combat SARS-CoV-2 infection or COVID-19 related symptoms and WHO-supervised clinical studies on the potential of artemisinins to combat COVID-19 are now in progress. However, implementing multiple potential new uses of artemisinins will require effective solutions to boost production, either by enhancing synthesis in A. annua itself or through biotechnological engineering in alternative biosynthesis platforms. Because of this renewed interest in artemisinin and its derivatives, here we review its modes of action, its potential application in different diseases including COVID-19, its biosynthesis and future options to boost production.

Global assessment of the distribution and conservation status of a key medicinal plant (Artemisia annua L.): The roles of climate and anthropogenic activities

As a medicinal plant, Artemisia annua L. is the main source of artemisinin in malaria drugs, but the lack of understanding of its distribution, environmental conditions and protection status limits the mass acquisition of artemisinin. Therefore, we used the ensemble forecast method to model the current and future global distribution areas of A. annua, evaluated the changes in suitable distribution areas on each continent under impacts of human activities and climate change, and its protection status on each continent in the corresponding period. The results showed that the main distribution areas of A. annua were concentrated in mid-latitudes in western and central Europe, southeastern Asia, southeastern North America and southeastern South America. Under the current climate scenario, human modifications have greatly reduced the suitable distribution area of A. annua, which was projected to expand inland with climate change and human socioeconomic impacts of CMIP6 in the future, but the effects of increasing temperature were different in different periods. Among all continents, the suitable distribution area in Europe was the most affected. However, at present and in the future, A. annua needs high priority protection on all continents. Asia and Europe have slightly better protection status scores than other continents, but the protection status scores of all continents are still very low. Our findings can be useful to guide development of protective measures for medicinal plants such as A. annua to further support drug production and disease treatment.

The Potential Mechanisms by which Artemisinin and Its Derivatives Induce Ferroptosis in the Treatment of Cancer.

Artemisinin (ART) is a bioactive molecule derived from the Chinese medicinal plant Artemisia annua (Asteraceae). ART and artemisinin derivatives (ARTs) have been effectively used for antimalaria treatment. The structure of ART is composed of a sesquiterpene lactone, including a peroxide internal bridge that is essential for its activity. In addition to their well-known antimalarial effects, ARTs have been shown recently to resist a wide range of tumors. The antineoplastic mechanisms of ART mainly include cell cycle inhibition, inhibition of tumor angiogenesis, DNA damage, and ferroptosis. In particular, ferroptosis is a novel nonapoptotic type of programmed cell death. However, the antitumor mechanisms of ARTs by regulating ferroptosis remain unclear. Through this review, we focus on the potential antitumor function of ARTs by acting on ferroptosis, including the regulation of iron metabolism, generation of reactive oxygen species (ROS), and activation of endoplasmic reticulum stress (ERS). This article systematically reviews the recent progress in ferroptosis research and provides a basis for ARTs as an anticancer drug in clinical practice.

The Effect of Different Light Spectrum Ratios and Photosynthetic Photon Flux Density (PPFD) on Some Agronomic and Physiological Traits in Artemisia annua L.

In order to investigate the effect of the ratio of different light spectra on the growth and physiological traits of the medicinal plant Artemisia annua, an experiment was conducted based on a randomized complete block design with three replications. Experimental treatments were six levels of different light wavelengths including control (base light: full spectrum), base light + ultraviolet spectrum, base light + blue light spectrum, base light + green light spectrum, base light + red light spectrum and base light + far-red light spectrum. The results showed that different light ratios influenced all the measured traits. Accordingly, the application of all light treatments significantly increased the dry weight of the Artemisia plant. Also, the light treatments had significant (P≤0.01) effects on plants height, contents of chlorophyll a and b, total chlorophyll, carotenoids and anthocyanin. The percentage and yield of plant essential oils were impressed significantly (P≤0.01) under the application of different light wavelengths. Although UV increased the content of the essential oil, it reduced the yield of the essential oil due to the reduction in the dry weight of plants. According to the results of this study, it can be concluded that a combination of base light with blue light can increase the biomass yield as well as percentage and yield of essential oils of Artemisia, compared to control.

Jasmonate- and abscisic acid-activated AaGSW1-AaTCP15/AaORA transcriptional cascade promotes artemisinin biosynthesis in Artemisia annua.

SummaryArtemisinin, a sesquiterpene lactone widely used in malaria treatment, was discovered in the medicinal plant Artemisia annua. The biosynthesis of artemisinin is efficiently regulated by jasmonate (JA) and abscisic acid (ABA) via regulatory factors. However, the mechanisms linking JA and ABA signalling with artemisinin biosynthesis through an associated regulatory network of downstream transcription factors (TFs) remain enigmatic. Here we report AaTCP15, a JA and ABA dual‐responsive teosinte branched1/cycloidea/proliferating (TCP) TF, which is essential for JA and ABA‐induced artemisinin biosynthesis by directly binding to and activating the promoters of DBR2 and ALDH1, two genes encoding enzymes for artemisinin biosynthesis. Furthermore, AaORA, another positive regulator of artemisinin biosynthesis responds to JA and ABA, interacts with and enhances the transactivation activity of AaTCP15 and simultaneously activates AaTCP15 transcripts. Hence, they form an AaORA‐AaTCP15 module to synergistically activate DBR2, a crucial gene for artemisinin biosynthesis. More importantly, AaTCP15 expression is activated by the multiple reported JA and ABA‐responsive TFs that promote artemisinin biosynthesis. Among them, AaGSW1 acts at the nexus of JA and ABA signalling to activate the artemisinin biosynthetic pathway and directly binds to and activates the AaTCP15 promoter apart from the AaORA promoter, which further facilitates formation of the AaGSW1‐AaTCP15/AaORA regulatory module to integrate JA and ABA‐mediated artemisinin biosynthesis. Our results establish a multilayer regulatory network of the AaGSW1‐AaTCP15/AaORA module to regulate artemisinin biosynthesis through JA and ABA signalling, and provide an interesting avenue for future research exploring the special transcriptional regulation module of TCP genes associated with specialized metabolites in plants.

A metabolomic approach to target antimalarial metabolites in the Artemisia annua fungal endophytes

Fungal endophytes are a major source of anti-infective agents and other medically relevant compounds. However, their classical blinded-chemical investigation is a challenging process due to their highly complex chemical makeup. Thus, utilizing cheminformatics tools such as metabolomics and computer-aided modelling is of great help deal with such complexity and select the most probable bioactive candidates. In the present study, we have explored the fungal endophytes associated with the well-known antimalarial medicinal plant Artemisia annua for their production of further antimalarial agents. Based on the preliminary antimalarial screening of these endophytes and using LC-HRMS-based metabolomics and multivariate analyses, we suggested different potentially active metabolites (compounds 1–8). Further in silico investigation using the neural-network-based prediction software PASS led to the selection of a group of quinone derivatives (compounds 1–5) as the most possible active hits. Subsequent in vitro validation revealed emodin (1) and physcion (2) to be potent antimalarial candidates with IC50 values of 0.9 and 1.9 µM, respectively. Our approach in the present investigation therefore can be applied as a preliminary evaluation step in the natural products drug discovery, which in turn can facilitate the isolation of selected metabolites notably the biologically active ones.

Experimental infection with Toxoplasma gondii in broiler chickens (Gallus domesticus): seroconversion, tissue cyst distribution, and prophylaxis.

Toxoplasma gondii is a widespread zoonotic protozoan that infects most species of mammals and birds, including poultry. This study aimed to investigate the course of T. gondii infection and the efficacy of diclazuril and Artemisia annua in preventing infection in experimentally infected chickens. Seventy-five 1-month-old chickens, female and male, were randomly divided into five groups (n = 15 each) as follows: (1) uninfected untreated (negative control, NC); (2) infected with T. gondii genotype II/III isolated from a wild cat (group WC); (3) infected with T. gondii genotype II isolated from a domestic cat (group DC); (4) infected with T. gondii domestic cat strain and treated with the anticoccidial diclazuril (group DC-D); and (5) infected with T. gondii domestic cat strain and treated with the medicinal plant Artemisia annua (group DC-A). Clinical signs, body temperature, mortality rate, weight gain, feed conversion ratio, hematological parameters, and the presence of T. gondii-specific IgY antibodies were recorded in all groups. Five chickens per group were euthanized 28days post-infection (p.i.) and their brains, hearts, and breast muscle tested for T. gondii by mouse bioassay and polymerase chain reaction (PCR). No clinical signs related to the experimental infection were observed throughout the study period. T. gondii-specific antibodies were detected by day 28 p.i., but not in all infected chickens. Overall, T. gondii DNA was detected (bioassay or tissue digests) in all infected and untreated chickens (10/10), while viable parasite (bioassay) was isolated from 7 out of 10 chickens. The parasite was most frequently identified in the brain (7/10). There were no differences in the T. gondii strains regarding clinical infection and the rate of T. gondii detection in tissues. However, higher antibody titers were obtained in chickens infected with T. gondii WC strain (1:192) comparing with T. gondii DC strain (1:48). A. annua reduced replication of the parasite in 3 out of 5 chickens, while diclazuril did not. In conclusion, broiler chickens were resistant to clinical toxoplasmosis, irrespective of the strain (domestic or wild cat strain). The herb A. annua presented prophylactic efficacy by reduced parasite replication. However, further studies are required aiming at the efficacy of diclazuril and A. annua for the prevention of T. gondii infection in chickens using quantitative analysis methods.

Dietary enzymatically-treated Artemisia annua L. supplementation could alleviate oxidative injury and improve reproductive performance of sows reared under high ambient temperature.

The medicinal plant Artemisia annua L. is well known for its antimalarial compound artemisinin and the antioxidant capacity of its active ingredients. However, low bioavailability of Artemisia annua L. limits its therapeutic potential, fermentation of Artemisia annua L. can improve its bioavailability. This study was aimed to investigate the effects of dietary supplementation of enzymatically-treated Artemisia annua L. (EA) on reproductive performance, antioxidant status, milk composition of heat-stressed sows and intestinal barrier integrity of their preweaning offspring. 135 multiparous sows of average parity 4.65 (Landrace×large white) at day 85 of pregnancy were randomly distributed into 3 treatments. Sows in the control group were housed at control rooms (temperature: 27.12±0.18°C, temperature-humidity index (THI): 70.90±0.80) and fed the basal diet. Sows in the HS, HS+EA groups were fed the basal diet supplemented with 0 or 1.0g/kg EA respectively, and reared at heat stress rooms (temperature: 30.11±0.16°C, THI: 72.70±0.60). Heat stress increased the malondialdehyde (MDA) content, reduced the activities of total antioxidant capacity (T-AOC) and total superoxide dismutase (T-SOD) of sows and piglets, and seriously compromised the antioxidant capacity of the sows and the intestinal integrity of their offspring. However, dietary supplementation of 1.0g/kg EA reduced the MDA content, increased the activities of T-SOD and T-AOC in serum, colostrum, and milk of heat-stressed sows, and increased colostrum yield and 14-d milk fat content. EA supplementation also increased piglet weaning weight and the activities of T-SOD and T-AOC in serum. In addition, the abundances of intestinal tight junction proteins claudin-1 and occludin were up-regulated in piglets in EA-supplemented group. In conclusion, dietary EA supplementation at 1.0g/kg can alleviate the oxidative stress in heat-stressed sows, improve the antioxidant capacity in both sows and their offspring, and promote the intestinal barrier integrity in their offspring. EA may be a potent dietary supplement that ameliorates oxidative stress in livestock production by improving the antioxidant capacity.

Dihydroartemisinin as a potential drug candidate for cancer therapy: a structural-based virtual screening for multitarget profiling

Cancer is a rapidly growing non-communicable disease worldwide that is responsible for high mortality rates, which account for 9.6 million death in 2018. Dihydroartemisinin (DHA) is an active metabolite of artemisinin, an active principle present in the Chinese medicinal plant Artemisia annua used for malaria treatment. Dihydroartemisinin possesses remarkable and selective anticancer properties however the underlying mechanism of the antitumor effects of DHA from the structural point of view is still not yet elucidated. In the present study, we employed molecular docking simulation techniques using Autodock suits to access the binding properties of dihydroartemisinin to multiple protein targets implicated in cancer pathogenesis. Its potential targets with comprehensive pharmacophore were predicted using a PharmMapper database. The co-crystallised structures of the protein were obtained from a Protein Data Bank and prepared for molecular docking simulation. Out of the 24 selected protein targets, DHA has shown about 29% excellent binding to the targets compared to their co-crystallised ligand. Additionally, 75% of the targets identified for dihydroartemisinin binding are protein kinases, and 25% are non-protein kinases. Hydroxyl functional group of dihydroartemisinin contributed to 58.5% of the total hydrogen interactions, while pyran (12.2%), endoperoxide (9.8%), and oxepane (19.5%) contributed to the remaining hydrogen bonding. The present findings have elucidated the possible antitumor properties of dihydroartemisinin through the structural-based virtual studies, which provides a lead to a safe and effective anticancer agent useful for cancer therapy. Communicated by Ramaswamy H. Sarma