Dihydroartemisinin Group Research Articles

Acidic Environment-Responsive Metal Organic Framework-Mediated Dihydroartemisinin Delivery for Triggering Production of Reactive Oxygen Species in Drug-Resistant Lung Cancer.

Dihydroartemisinin (DHA) has emerged as a promising candidate for anticancer therapy. However, the application of DHA in clinics has been hampered by several limitations including poor bioavailability, short circulation life, and low solubility, significantly restricting its therapeutic efficacy and leading to notable side effects during the treatment. We present DHA-loaded zeolitic imidazolate framework-8 (D-ZIF) with controllable and targeted DHA release properties, leading to enhanced antitumor effects while reducing potential side effects. D-ZIF was prepared by one-pot synthesis method using methylimidazole (MIM), Zn(NO3)2•6H2O and DHA. We characterized the physical and chemical properties of D-ZIF by TEM, DLS, XRD, FT-IR, and TG. We measured the drug loading efficiency and the cumulative release of DHA in different pH conditions. We evaluated the cytotoxicity of D-ZIF on renal cell carcinoma (RCC786-O), glioma cells (U251), TAX-resistant human lung adenocarcinoma (A549-TAX) cells by CCK8 in vitro. We explored the possible antitumor mechanism of D-ZIF by Western blot. We evaluated the biocompatibility and hemolysis of D-ZIF and explored the in vivo antitumor efficiency in mice model by TUNEL testing and blood biomarker evaluations. D-ZIF showed rhombic dodecahedral morphology with size of 129±7.2 nm and possessed a noticeable DHA encapsulation efficiency (72.9%). After 48 hours, D-ZIF released a cumulative 70.0% of the loaded DHA at pH 6.5, and only 42.1% at pH 7.4. The pH-triggered programmed release behavior of D-ZIF could enhance anticancer effect of DHA while minimizing side effects under normal physiological conditions. Compared with the free DHA group with 31.75% of A549-TAX cell apoptosis, the percentage of apoptotic cells was approximately 76.67% in the D-ZIF group. D-ZIF inhibited tumor growth by inducing tumor cell apoptosis through the mechanism of ROS production and regulation of Nrf2/HO-1 and P38 MAPK signaling pathways. D-ZIF showed potent effects in treating tumors with high safety in vivo. This pH-responsive release mechanism enhanced the targeting efficiency of DHA towards tumor cells, thereby increasing drug concentration in tumor sites with negligible side effects. Herein, D-ZIF holds great promise for curing cancers with minimal adverse effects.

Interventional effect and inflammatory regulation mechanism of dihydroartemisinin combined with pregabalin on neuropathic pain mice

This study aims to clarify the effects of dihydroartemisinin(DHA) combined with pregabalin(PGB) on neuropathic pain(NP) in mice and explore the neuroinflammatory regulatory mechanism. NP mice model was established using spinal nerve ligation, whereas the sham group exposed the spinal nerve without ligation. The mice were randomly divided into sham group, model group, PGB groups of low, medium, and high doses(PGB-L, PGB-M, and PGB-H, with 22, 45, and 91 mg·kg~(-1)), DHA group(16 mg·kg~(-1)), and DHA combined with PGB groups of low, medium, and high doses(DHA + PGB-L, DHA + PGB-M, and DHA + PGB-H). Administration by gavage 18 days after modeling. Von Frey and cold plate were used to detect mechanical pain threshold and cold pain sensitivity in mice. The tail suspension test and forced swimming test were used to investigate depressive behavior, and the open field test was used to estimate anxiety behavior. The Morris water maze was used to evaluate cognitive function. Liquid suspension chip technology was used to quantitatively analyze immune inflammation-related factors. Immunofluorescence was used to detect the expression of CC chemokine ligand 3(CCL3) and transmembrane protein 119(TMEM119). The results showed that compared with the sham group, the mechanical pain and cold pain sensitivity thresholds of the model group were significantly reduced, and the struggle time was significantly increased in the tail suspension test and forced swimming test. The activity time in the central area was significantly reduced in the open field test. The residence time in the second/fourth quadrant was significantly longer than that in other quadrants, and the latency time of platform climbing significantly increased after platform withdrawal in the Morris water maze experiment. The expression of CCL3 was significantly increased; the number of TMEM119 positive cells and the cell body area were significantly increased. Compared with the model group, the DHA + PGB-M group showed a significant increase in mechanical pain and cold pain sensitivity thresholds, as well as a significant increase in struggle time in the tail suspension test and forced swimming test. The activity time in the central area of the open field test was significantly reduced. The residence time in the second/fourth quadrant was significantly shorter than that in other quadrants, and the latency time of platform climbing after platform withdrawal was significantly reduced. Compared with the PGB-M group, the mechanical pain threshold of D14-17 in the DHA + PGB-M group was significantly increased, and the struggle time during forced swimming was significantly increased. The residence time in the second/fourth quadrant of the Morris water maze was significantly shorter than that in other quadrants. Compared with the model group, the expression of CCL3, the number of TMEM119 positive cells, and the cell body area in the DHA + PGB-M group were significantly decreased. This study indicates that DHA + PGB can enhance the analgesic effect of PGB on NP mice, break through the limitations of PGB tolerance, and make up for the shortcomings of PGB in antidepressant and cognitive improvement. Its mechanism may be related to regulating neuroinflammation by inhibiting the activation of microglial cells and expression of CCL3.

Therapeutic potential of dihydroartemisinin in mitigating radiation-induced lung injury: Inhibition of ferroptosis through Nrf2/HO-1 pathways in mice.

Radiation-induced lung injury (RILI) is a common consequence of thoracic radiation therapy that lacks effective preventative and treatment strategies. Dihydroartemisinin (DHA), a derivative of artemisinin, affects oxidative stress, immunomodulation, and inflammation. It is uncertain whether DHA reduces RILI. In this work, we investigated the specific mechanisms of action of DHA in RILI. Twenty-four C57BL/6J mice were randomly divided into four groups of six mice each: Control group, irradiation (IR) group, IR + DHA group, and IR + DHA + Brusatol group. The IR group received no interventions along with radiation treatment. Mice were killed 30 days after the irradiation. Morphologic and pathologic changes in lung tissue were observed with hematoxylin and eosinstaining. Detection of hydroxyproline levels for assessing the extent of pulmonary fibrosis. Tumor necrosis factor α (TNF-α), transforming growth factor-β (TGF-β), glutathione peroxidase (GPX4), Nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) expression in lung tissues were detected. In addition, mitochondrial ultrastructural changes in lung tissues were also observed, and the glutathione (GSH) content in lung tissues was assessed. DHA attenuated radiation-induced pathological lung injury and hydroxyproline levels. Additionally, it decreased TNF-α and TGF-β after irradiation. DHA may additionally stimulate the Nrf2/HO-1 pathway. DHA upregulated GPX4 and GSH levels and inhibited cellular ferroptosis. Brusatol reversed the inhibitory effect of DHA on ferroptosis and its protective effect on RILI. DHA modulated the Nrf2/HO-1 pathway to prevent cellular ferroptosis, which reduced RILI. Therefore, DHA could be a potential drug for the treatment of RILI.

Dihydroartemisinin alleviates mitochondrial damage and improves cardiomyocyte pyroptosis and ferroptosis through Sirt1-mediated PGC-1α

This study assesses the effect of dihydroartemisinin on pyroptosis and ferroptosis. Rat H9C2 cardiomyocytes were intervened with 35 mmol/L high glucose through assigned blank control, dihydro artemisinin, and dihydroartemisinin+Sirt1 groups. Confocal microscopy was used to observe the ROS levels, while proliferation ability was detected by CCK-8 method, and apoptosis was assessed by flow cytometry, and migration ability by Transwell transfer method. Moreover, analysis of pyroptosis-related factors expression and content of lipid peroxide were done using laser confocal microscopy. The average fluorescence intensity of dihydro artemisinin group and dihydroartemisinin+SIRT 1 group decreased significantly (P <0.05), among which the dihydroartemisinin+SIRT 1 group had lowest average fluorescence intensity (P <0.05). SIRT 1 level in the dihydroartemisinin and dihydroartemisinin+SIRT 1 groups was higher than blank control (P <0.05), with highest level in the dihydroartemisinin+SIRT 1 group (P <0.05). Cell proliferation in the dihydroartemisinin and dihydroartemisinin+SIRT 1 group was reduced (P <0.05), with lowest proliferation in combination group (P < 0.05). Cell migration in the dihydroartemisinin and dihydroartemisinin+SIRT 1 groups was reduced (P <0.05), with lowest number of migratory cells in the dihydroartemisinin+SIRT 1 group (P <0.05). Cell apoptosis in the dihydroartemisinin and dihydroartemisinin+SIRT 1 groups was increased (P <0.05), with lowest apoptosis in the dihydroartemisinin+SIRT 1 group (P <0.05). There was upregulation of SIRT 1 and PGC-1α mRNA expression in the dihydroartemisinin and dihydroartemisinin+SIRT 1 groups was elevated (P <0.05). The expression of NLRP3, GSDMD, and Caspase-1 were all decreased (P <0.05), while that of GPX4 was increased (P <0.05). Dihydroartemisinin inhibits the function of H9C2 cardiomyocytes, pyroptosis and ferroptosis, playing a positive role in ameliorating Diabetic cardiomyopathy (DCM).

Effects of dihydroartemisinin on cardiac fibroblast activity induced by angiotensin II

Dihydroartemisinin is currently now being studied for the treatment of diseases. The mechanism of action of dihydroartemisinin on cardiovascular diseases is unclear. We explored the effect of dihydroartemisinin on the activity of cardiac fibroblasts and the underlying mechanism. The cardiac fibroblasts were divided into blank control group, dihydroartemisinin group, vascular angiotensin II (AngII) group and angiotensin II+dihydroartemisinin group followed by analysis of cell proliferation, Col I and FN levels, and the levels of TGF-β1, α-SMA and IGF-I. The morphology of myocardial fibroblasts was spindle-shaped. Further immunofluorescence staining assessed positive expression of Vimentin of the myocardial fibroblasts and no expression of α-SMA, which confirmed the successful culture of the myocardial fibroblasts. Overexpression of AngII significantly promoted the proliferation ability of mouse cardiac fibroblasts, which was significantly reduced after dihydroartemisinin treatment. As shown by RT-qPCR, TGF-β1, α-SMA and IGF-I levels in AngII mice increased, while their levels reduced after dihydroartemisinin intervention. After overexpression of AngII in cardiomyocytes, cell proliferation and colonies formation increased and decreased after dihydroartemisinin treatment, suggesting that dihydroartemisinin inhibited the growth of cardiomyocytes. It showed that dihydroartemisinin could alleviate the expression of AngII-induced activation markers of mouse cardiac fibroblasts. Therefore, our research confirms that dihydroartemisinin plays an important role in anti-myocardial fibrosis, mainly through regulating the expression of TGF-β1.

Transcriptome analysis of Tetrahymena thermophila response to exposure with dihydroartemisinin

Dihydroartemisinin (DHA) is a derivative of artemisinin and is toxic to parasites. We used the Tetrahymena thermophila (T. thermophila) as a model to explore DHA toxicity. Results showed that low concentration of DHA (20 μmol/L) promoted cell proliferation, whereas high concentrations of DHA (40–1280 μmol/L) inhibited that. Appearance of nucleus was pycnosis by laser scanning confocal microscope. DHA significantly elevated activities of SOD and GSH-Px (P < 0.01) and MDA was markedly increased at high level but decreased at low level (P < 0.01). Further results of transcriptome in T. thermophila treated with different concentration DHA group (0, 20, 160 μmol/L) showed that differentially expressed genes (DEGs) were involved in oxidation-reduction and metabolism of exogenous substances indicated oxidative stress stimulation. Kyoto Encyclopedia of Genes and Genomes showed that DEGs were involved in the cytochrome P450-mediated metabolism of exogenous substances, glutathione metabolism and ABC transport. Remarkably, DNA replication was significantly enriched in low concentration DHA, energy metabolism related pathways and necrotic process were considerably enriched in high concentration DHA. The results of RT-qPCR of 13 DEGs were the same as that of transcriptome, in which the expression of GST and GPx family genes were significantly altered after exposed to high-DHA group. DHA induced oxidative stress damage through disturbing with energy. However, detoxification pathways in T. thermophila to resist oxidative damage and cell alleviated low concentration DHA stress by regulating antioxidant enzyme. This study provides good practice on pharmacological mechanism of artemisinin-based drugs in antiparasitic.

Effect of a primer containing Artemisinin on dentin endogenous MMPs and resin bond durability

ObjectivesThis study aims to investigate the potential effect of a primer containing Artemisinin (ART) or its derivative—dihydroartemisinin (DHA) on the dentin endogenous MMPs activity and resin bonding durability under etch-and-rinse mode. Materials and methodsMolecular docking was applied for evaluating the potential interactions of MMP-2 and MMP-9 with ART and DHA. Then human third molars were randomly assigned to six groups, after etch-and-rinse treatment, pretreated with the primers containing 0.25, 0.5 mg/mL ART, 0.25, 0.5 mg/mL DHA and 2% chlorhexidine (CHX) for 2 min, respectively, while one untreated group was served as the control group. The endogenous MMPs activity of demineralized dentin was tested with in situ zymography test and zymographic assay. Following the etch-and-rinse bonding process with a universal resin bond, the specimens of the microtensile bond strength (μTBS) and nanoleakage were tested with or without 10,000 °C thermocycling and 6-month storage in artificial saliva at 37 °C. ResultsMolecular docking showed that ART and DHA combined with MMP-2 and MMP-9 through forming covalent bonds with the zinc ion of MMPs. Compared with the control group, the ART, DHA and CHX groups showed less endogenous MMPs activity, and the MMPs activity of 0.5 mg/mL DHA group had no significant difference from that of 2% CHX group. Ageing condition reduced the μTBS values dramatically and also aggravated the nanoleakage of all the groups, specifically, the control group showed lower μTBS value and a broader, thicker and more consecutive silver deposition than other groups. ConclusionsThe primer containing ART or DHA could preserve bond durability and slow down the nanoleakage of resin-dentin interface over time through inhibiting the endogenous MMPs activity.

Dihydro-artemisinin inhibits angiotensin II-induced cardiac fibroblasts proliferation and induces apoptosis through phosphoinositide 3-kinase/akt pathway

Cardiac fibroblasts (CFs) abnormal proliferation can trigger myocardial fibrosis, which is closely related to a variety of cardiovascular diseases. This study investigated the effect of dihydroartemisinin (DHA) on CF growth through PI3K/AKT with angiotensin II (AngII). CCK-8 method and EdU were used detect cell proliferation, and flow cytometry assessed apoptosis of CFs. The PI3K and AKT expressions were determined, whilst immunofluorescence staining and RT-qPCR measured α-smooth muscle actin (α-SMA) level. With rat CFs exhibiting typical fibroblast morphology and nuclei in spindle, oval or polygonal shape, these cells were positive for the fibroblast marker vimentin. Treatment with AngII resulted in increased proliferation of CFs, but the proliferation declined in the presence of DHA (p &lt; 0.05). Meanwhile, the AngII group had lower apoptosis of CFs than blank and DHA groups with highest apoptosis observed in the DHA group. α-SMA increased at the mRNA and protein levels after Ang II treatment and decreased after DHA treatment (p &lt; 0.05). Importantly, the Ang II upregulated PI3K and AKT in CFs (p &lt; 0.05) and the increase was abrogated by DHA administration with lowest levels (p &lt; 0.05). Ang II activates PI3K/Akt in normal CFs, as administration of DHA can inhibit the effect of Ang II, thereby inhibiting proliferation of CFs induced by AngII and inducing apoptosis.

Retardant effect of dihydroartemisinin on ulcerative colitis in a JAK2/STAT3-dependent manner.

Dihydroartemisinin (DHA) is a semi-synthetic derivative and the main active metabolite of artemisinin. The purpose of this study was to investigate the effect of DHA on the ulcerative colitis (UC) in both in vivo and in vitro models. Weight, survival rate, colon length, and Disease Activity Index score were used to evaluate the severity of colitis. Reverse transcription quantitative polymerase chain reaction and enzyme-linked immunosorbent assay were used to detect the expressions of cytokines interleukin (IL)-1, IL-1β, IL-4, IL-6, IL-10, IL-12, and tumor necrosis factor-α (TNF-α). The expressions of Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3), and the phosphorylation of JAK2 (p-JAK2) and STAT3 (p-STAT3), were measured by western blot analysis. Western blot analysis and immunohistochemistry were used to detect the expressions of tight junction proteins. We found that the weights and colon lengths of mice in dextran sodium sulfate (DSS)+DHA group were significantly lower and longer than those in the DSS group, respectively. Compared with those in the DSS group, the expressions of IL-1β, IL-6, IL-17, and TNF-α in the DSS+DHA and DSS+5-aminosalicylic acid (5-ASA) groups were decreased, while the expressions of IL-4 and IL-10 were significantly upregulated. DHA largely increased the expressions of zonula occludens-1 and occludin. Western blot analysis and/or immunohistochemical staining analysis showed that the expressions of JAK2, STAT3, p-JAK2, and p-STAT3 in DSS+DHA and DSS+5-ASA groups were significantly lower than those in DSS group. DHA has a specific therapeutic effect on UC. The anti-inflammatory mechanism of DHA is related to the blockage of the JAK2/STAT3 signaling pathway. These findings provide evidence that DHA may be a useful drug and is expected to become a promising new treatment for human UC.

The Evaluation of 68Ga-Citrate PET/CT Imaging for Dihydroartemisinin in the Treatment of Rheumatoid Arthritis.

We aimed to use 68Ga-citrate, a labeled product of gallium (iron analog), combined with positron emission tomography/computed tomography (PET/CT) to non-invasively evaluate the potential of the iron-responsive product dihydroartemisinin (DHA) in the treatment of rheumatoid arthritis. From the establishment of chicken II collagen-induced arthritis (CIA) rat model over 40days, 20 rats with one-to-one corresponding arthritis index (AI) scores were randomly divided into two groups. One group received oral DHA (at a dose of 1.5ml/(kgday), containing 20mg DHA per 1ml) for 15days; the other group received stroke-physiological saline solution (SSS, 1.5ml/(kgday) for 15days. 68Ga-citrate micro-PET/CT imaging was performed on day 0 (D0), day 5 (D5), day 10 (D10), and day 15 (D15) of oral administration. After data reconstruction, the cross-sectional length "d" of the ankle joint of each rat was measured on the transverse CT, and the SUVmax of the ankle joint and muscle background was measured for statistical analysis. After micro-PET/CT collection, the ankle joint tissue was observed by HE staining. The ankle joint swelling in the DHA group was significantly suppressed, but the SSS group showed no significant suppression. 68Ga-citrate micro-PET/CT imaging results and microscope observation confirmed this finding. Statistical analysis indicated that the time tendency of AI score (Binteraction = 0.495, P < 0.001) and T/NT (Binteraction = 1.345, P < 0.001) were discrepant between DHA and SSS groups. The AI score and T/NT of the DHA group gradually increased with time, while the SSS group score gradually decreased. Furthermore, the Spearman correlation coefficient was used to describe the relationship between "d" and T/NT, which was positively correlated (r = 0.855, P < 0.001). This study showed that the anti-inflammatory effect of the iron-responsive product DHA in arthritis can be monitored by an iron-like radioactive tracer (68Ga-citrate).

Effects of dihydroartemisinin on the gut microbiome of mice.

Dihydroartemisinin (DHA) is a semisynthetic derivative of artemisinin, which has been found to exhibit a broad range of biological activities, excluding antimalarial effects; however its effects on the gut microbiota remain poorly understood. The present study aimed to investigate the effects of DHA on the gut microbiome in mice and to determine its potential biological and pharmaceutical activities through its alteration of the gut microbiota. Serum glucose, triglyceride (TG), total cholesterol, lipopolysaccharide, high density lipoprotein-cholesterol, low density lipoprotein-cholesterol, alanine aminotransferase and aspartate aminotransferase levels in mice treated with DHA were analyzed using the corresponding detection kits. In addition, hematoxylin and eosin staining was performed to determine the pathological effects of DHA on the liver, kidney and intestinal tissues of mice, and the effects of DHA on the gut microbiome were analyzed using 16S ribosomal (r)DNA gene analysis. The results demonstrated that the TG serum levels of mice treated with DHA were significantly decreased compared with the control group. Furthermore, 16S rDNA gene analysis demonstrated that the bacterial diversity of mice treated with DHA was enriched compared with the control group. The DHA group exhibited increased numbers of Firmicutes and Saccharibacteria, and decreased Deferribacteres and Actinobacteria compared with the control group at the phylum level. Kyoto Encyclopedia of Genes and Genomes signaling pathway enrichment analysis also revealed that the signaling pathways associated with ‘Energy metabolism’ and ‘Nucleotide metabolism’ were upregulated, whereas the signaling pathways associated with ‘Infectious diseases and ‘Neurodegenerative diseases’ were downregulated in the DHA group compared with the control group. In conclusion, the findings of the present study indicated that DHA may significantly decrease the serum TG levels and alter the gut microbiota, which suggested its potential to be used for the treatment of hyperlipidemia, inflammatory and neurodegenerative disorders.

Preliminary study of dihydroartemisin in inhibiting invasion and metastasis of hepatoma cells

It is reported that dihydroartemisinin could reduce the expression of phosphorylated adhesion kinase and matrix metalloproteinase-2, inhibit the growth, migration and invasion of ovarian cancer cells, promote the formation of Treg cells through TGF-beta/Smad signaling pathway, and play an immunosuppressive role; dihydroartemisinin could also inhibit the growth of lung cancer cells by inhibiting the expression of vascular endothelial growth factor(VEGF) receptor KDR. However, there are few studies on dihydroartemisinin in hepatocellular carcinoma cells. In order to preliminarily explore the effect of dihydroartemisinin on invasion and metastasis of hepatocellular carcinoma cells, CCK-8 method and crystal violet staining were used to detect the effect of dihydroartemisinin on the growth of hepatocellular carcinoma cell 7402 and highly metastatic hepatocellular carcinoma cell MHCC97 H. The effects of dihydroartemisinin on the invasion and metastasis of hepatocellular carcinoma cell 7402 and highly metastatic hepatocellular carcinoma cell MHCC97 H were studied by using cell wound healing and Transwell. Western blot was used to detect the protein expression of epidermal growth factor receptor(EGFR) and its downstream signaling pathway in cells treated with dihydroartemisinin for 48 hours. The results showed that dihydroartemisinin could inhibit the growth of hepatocellular carcinoma cell 7402 and highly metastatic hepatocellular carcinoma cell MHCC97 H at 25 μmol·L~(-1). As compared with the control group, the number of cell clones was significantly reduced, and the ability of cell migration and invasion was weakened. Western blot results showed that as compared with the control group, dihydroartemisinin group could down-regulate the protein expression of EGFR and its downstream signaling pathways p-AKT, p-ERK, N-cadherin, Snail and Slug, and up-regulate the expression of E-cadherin protein, thus affecting the migration, invasion and metastasis of hepatocellular carcinoma cells 7402 and MHCC97 H.

Dihydroartemisinin Ameliorated Ovalbumin-Induced Asthma in Mice via Regulation of MiR-183C.

BackgroundThe purpose of the present study was to investigate the function and mechanism of dihydroartemisinin (DHA) in treating ovalbumin-induced asthma in BALB/c mice.Material/MethodsThirty female BALB/c mice were randomly separated into 3 groups: the control group, the asthma model group stimulated by ovalbumin (OVA group), and the DHA treatment group (DHA group). The therapeutic effects and potential pharmacological mechanisms of DHA were specifically clarified by examining its effects on asthma-related phenomena, such as body weight, lung function, cell counts in bronchoalveolar lavage fluid (BALF), and hemotoxin and eosin staining. In addition, the expression of inflammatory factors was checked by enzyme-linked immunosorbent assay kits, and fractions of Th17 cells were detected by FACS analysis. Moreover, the downstream molecular pathway of IL-6/Stat3 (interleukin-6/signal transducer and activator of transcription 3) and expression of miR-183C was investigated by western blot and/or quantitative real-time polymerase chain reaction. Luciferase assay was used to reveal the function of miR-183C on the transcriptional regulation of Foxo1 (forkhead box O).ResultsDHA administration significantly relieved the severity of the asthma through its effect on body weight, survival rate, and airway pressure. DHA was able to ameliorate lung damage in terms of pathological morphology and it reduced the percentage of helper T 17 (Th17) cells and the secretion of cytokines. As a result, the activity of the IL-6/Stat3 pathway was inhibited by DHA. In addition, the adoption of DHA decreased the expression of miR-183C but increased the expression of the transcription factor Foxo1.ConclusionsOur results suggest that the therapeutic effects of DHA on asthma are partially realized via the regulation of miR-183C and IL-6/Stat3 pathway.

Dihydroartemisinin, an antimalarial drug, induces absent in melanoma 2 inflammasome activation and autophagy in human hepatocellular carcinoma HepG2215 cells.

As an effective antimalarial drug, Dihydroartemisinin (DHA) is readily isolated from the traditional Chinese medicine of Artemisia annua. DHA is not only an autophagy promoter but also a substance with strong antitumor efficiency. The relationship between autophagy and inflammasomes has been suggested in hepatocellular carcinoma (HCC). However, there are few reports describing relationships between inflammasomes and autophagy in HCC therapy. The present study demonstrated that DHA suppressed cell proliferation in HepG2215 cells in a dose- and time-dependent manner. The inhibitory activity is mediated by autophagy, in which reactive oxygen species (ROS) production induced nuclear and mitochondrial DNA damage. Then, DHA were first shown to promote AIM2/caspase-1 inflammasome. Compared with the DHA group, the autophagy inhibitor 3-MA significantly inhibited the expressions of activated Caspase-1, a pyroptotic marker proteins. Meanwhile, repression of mTOR by rapamycin promoted autophagy and AIM2/caspase-1 activation. The caspase-1 inhibitor Z-YVAD-FMK also notably blocked autophagy cell death characterized by the downexpression of Beclin-1 and LC3-II. Additionally, the study demonstrated that DHA suppressed pseudopodium formation and cell mobility. Therefore, we first reveal a novel mechanism that DHA promotes AIM2/caspase-1 inflammasome, which contributes to autophagy in HepG2215 cells. Moreover, nuclear and mitochondrial DNA damage was also involved in this process via ROS production.

Dihydroartemisinin Increases the Sensitivity of Photodynamic Therapy Via NF-κB/HIF-1α/VEGF Pathway in Esophageal Cancer Cell in vitro and in vivo

Background/Aims: Although photodynamic therapy (PDT) can relieve esophageal obstruction and prolong survival time of patients with esophageal cancer, it can induce nuclear factor-kappa B (NF-κB) activation in many cancers, which plays a negative role in PDT. Dihydroartemisinin (DHA), the most potent artemisinin derivative, can enhance the effect of PDT on esophageal cancer cells. However, the mechanism is still unclear. Methods: We generated stable cell lines expressing the super-repressor form of the NF-κB inhibitor IκBα and cell lines with lentivirus vector-mediated silencing of the HIF-1α gene. Esophageal xenograft tumors were created by subcutaneous injection of Eca109 cells into BALB/c nude mice. Four treatment groups were analyzed: a control group, photosensitizer alone group, light alone group, and PDT group. NF-κB expression was detected by an electrophoretic mobility shift assay, hypoxia-inducible factor α (HIF-1α) and vascular endothelial growth factor (VEGF) by real-time PCR, NF-κB, HIF-1α, and VEGF protein by western blot, and Ki-67, HIF-1α, VEGF, and NF-κB protein by immunohistochemistry. Results: PDT increased NF-κB activity and the gene expression of HIF-1α and VEGF in vitro and in vivo. In contrast, the DHA groups, particularly the combined DHA and PDT treatment group, abolished the effect. The combined treatment significantly inhibited tumor growth in vitro and in vivo. NF-κB activity and HIF-1α expression were also reduced in the stable IκBα expression group, whereas the former showed no change in HIF-1α-silenced cells. Conclusion: DHA might increase the sensitivity of esophageal cancer cells to PDT by inhibiting the NF-κB/HIF-1α/VEGF pathway.

Effects of dihydroartemisinin on migration and cell viability of human giant cell tumor cells of bone

Objective To research the effects of dihydroartemisinin (DHA) on migration and cell viability of human giant cell tumor cells of bone. Methods The human giant cell tumor cells of bone were treated with 0, 10, 20, 40 μmol/L DHA for 24 h. After treated, the cells were stained with Annexin V-fluoresceine isothiocyanate (FITC)/propidium iodide (PI) and the apoptosis rate was measured by flow cytometry. At the same time the mitochondrial membrane potential change was measured. Results After treated with DHA for 24 h, the apoptosis rates of human giant cell tumor cells of bone (0, 10, 20 μmol/L DHA) were respectively 0.16%, 18.57% and 34.28%. Compared with control group, the apoptosis rates of human giant cell tumor cells of bone in DHA groups were significantly decreased (P=0.000, 0.000). The relative cell viability were respectively (60.63±2.45)% and (45.93±9.36)% in 10 and 20 μmol/L DHA group, which were significantly decreased than that in control group (P=0.024, 0.000). The migration [(69.27±4.82)% and (46.43±3.96)%] and invasion [(61.98±9.37)% and (43.55±2.45)%] of human giant cell tumor cells of bone in 10 and 20 μmol/L DHA group, which were significantly decreased than those in control group. Compared with control group, the expression levels of matrix metalloproteinase (MMP)-2 and MMP-9 were decreased. Conclusion DHA can significantly inhibit the cell viability and increase cell apoptosis. At the same time, DHA can inhibit the migration and invasion of human giant cell tumor cells of bone through decreasing the expression levels of MMP-2 and MMP-9. Key words: Dihydroartemisinin; Human giant cell tumor cells of bone; Cell viability; Apoptosis; Migration; Invasion; Matrix metalloproteinase-2; Matrix metalloproteinase-9

The in vivo study of dihydroartemisinin inhibits the growth of human cholangiocarcinoma

Objective To investigate the effect and the potential mechanisms of dihydroartemisinin on the treatment of human cholangiocarcinoma cell line QBC939 xenograft in nude mouse.Methods A total of 18 animal models of nude mouse xenograft by human cholangiocarcinoma cell line QBC939 were established and randomly divided into three groups:the dihydroartemisinin group (DHA),the control group (saline),and the solvent control group (vegetable oil).The weight of tumor,the tumor inhibition rate,the expression of hypoxia-inducible factor (HIF)-1α,the microvessel density (MVD) and the apoptotic index (AI) in each group were compared and analyzed.Results The volume and the weight of xenograft tumor in DHA group [(1 979.63 ± 357.29) mm3 and (1.77 ± 0.23) g] were significantly lower than those in the control group [(2 827.69 ± 549.00) mm3 and (2.50 ± 0.65) g,P ＜ 0.05].The expression of HIF-1α in the experimental group (0.228 0 ± 0.043 9)was less than that in the control group (0.336 7 ± 0.089 6,P ＜ 0.05).MVD in the experimental group (11.06 ± 1.44) was less than that in the control group (17.44 ± 2.24,P ＜ 0.05).The AI in the experimental group [(13.18 ± 2.42) ％] was significantly greater than that in the control group [(3.28 ± 1.04) ％,P ＜ 0.05].Conclusion Dihydroartemisinin has an inhibitory effect on the tumor growth of human cholangiocarcinoma in nude mice,and the mechanism may be related to the induction of the apoptosis of tumor cells,and the inhibition of tumor angiogenesis. Key words: Cholangiocarcinoma; Dihydroartemisinin ; Hypoxia-inducible factor-1 α ; In vivo study

Intervention effect of dihydroartemisinin on pulmonary fibrosis in rats

Objective To investigate the effect of dihydroartemisinin on pulmonary fibrosis and its mechanism in rats. Methods Seventy-five SD rats were randomly divided into 5 groups: the control group, the model group, the high dihydroartemisinin group, the low dihydroartemisinin group, and the prednisone group(15 rats in each group). Saline(9 g/L, 0.3 mL )was injected into the rat trachea of the control group, and the rats of the rest 4 groups were injected bleomycin(3 mg/kg). The rats in the control group and the model group received saline lavage ( 9 g/L, 3 mL), while the rats in the high dihydroartemi-sinin group and the low dihydroartemisinin group received dihydroartemisinin lavage (100, 50 mg/kg), and the rats in the prednisone group received prednisone lavage each day. Five rats in each group were sacrificed and lung tissues were removed on the 8th, 15th, and 29th day of lavage, and the degree of inflammation in lung tissue was observed by HE staining. The deposit of collagen in lung tissue was observed by Masson staining. The expressions of transforming growth factor β1(TGF-β1), collagen Ⅰand collagen Ⅲ in pneumonic tissues were detected by immunohistochemistry. Results The degree of inflammation in pneumonic tissues in the high dihydroartemisinin group, the low dihydroartemisinin group, and the prednisone group were less significantly than that in the model group(all P 0.05). TGF-β1, collagen Ⅰand collagen Ⅲ in lung tissues in the high dihydroartemisinin group and the prednisone group were lower than those in the model group(P 0.05), except that collagen Ⅲ expression in lung tissues was lower than that of the prednisone group on the 29th day(P>0.05). Conclusions Dihydroartemisinin can lessen the extent of pulmonary fibrosis in rats and its anti-fibrosis effect is dose-dependent. The anti-fibrosis effect of dihydroartemisinin may decrease the deposit of collagens in the lung tissues by inhibiting TGF-β1 expression in the lung tissues and have an effect of antifibrosis. Key words: Dihydroartemisinin; Pulmonary fibrosis; Transforming growth factor β1; Collagen Ⅰ; Collagen Ⅲ

Exploring the autophagy of glioma cells induced by dihydroartemisinin in vitro

Objective To investigate the anti -glioma efficacy of dihydroartemisinin (DHA) and combined with temozolomide (TMZ) on human glioma cell line in vitro.Methods The growth inhibition of DHA on 10 glioma cell lines induced by DHA treatment for 72 h was analyzed by MTT method.TMZ combined with 0.5 μg/ml DHA,and the IC5o value was measured in SKMG -4 cells.The autofluorescent drug monodansylcadaverine (MDC) was used to mark autophagicvacuoles.The autophagy related protein LC3 - B and Beclin - 1,and the apoptosis protein caspase - 3 were analyzed by western blot (WB).Results The IC50 of DHA was different in ten glioma cell lines [ from ( 1.17 ± 0.078 ) μg/ml to (23.568±0.796) μg/ml].In SKMG -4 cells,the autophagicvacuoles were detected in the DHA group,the IC50 of TMZ had significant difference with 0.5 μg/ml DHA contrast to the control group ( P ＜ 0.01 ),and the levels of LC3 - B and Beclin - 1 protein were increased gradually with the increase of DHA concentration,and caspase -3 protein has no difference.Conclusion DHA can inhibit proliferation of glioma cell lines and increase the efficacy of TMZ,and the autophagy may be the mechanism. Key words: Glioma ; Autophagy ; Dihydroartemisinin ; Temozolomide ;

Topical dihydroartemisinin inhibits suture-induced neovascularization in rat corneas through ERK1/2 and p38 pathways.

To determine if topical instillation of dihydroartemisinin (DHA) inhibits corneal neovascularization (NV) in rats and to investigate the role of the extracellular regulated kinases (ERK) 1/2 and p38 pathways in this process. Suture-induced corneal NV was produced in rats and the eyes were topically treated with different concentrations of DHA (20mg/L, 10mg/L or 5mg/L) or normal saline 4 times a day for 7 days. The corneal NV was quantified as the proportion of NV area to the whole cornea. Western blot was used to determine the expressions of vascular endothelial growth factor (VEGF) and the phosphorylation status of VEGF receptor-2, ERK1/2 and p38 in the corneas. Immunofluorescent staining was used to determine the expressions of phospho-ERK1/2 and phospho-p38 in the corneal tissues from the eyes treated with 20 mg/L DHA (DHA group) or normal saline (control group). The proportion of corneal NV area in the eyes treated with normal saline or DHA at dosages of 20mg/L, 10mg/L or 5mg/L was (23.74±3.00)%, (15.73±2.88)%, (19.53±2.42)%, and (23.38±2.79)%, respectively. In the eyes treated with 20mg/L or 10mg/L DHA, the corneal NV area was significantly reduced when compared to that in eyes with normal saline (P<0.05). Western blot analyses revealed that 20mg/L DHA significantly inhibited the expressions of VEGF and phospho-VEGFR-2. Both 20mg/L and 10mg/L DHA inhibited the expressions of phospho-ERK1/2 and phospho-p38. Immunofluorescent staining further demonstrated that 20mg/L DHA lowered the expression levels of phospho-ERK1/2 and phospho-p38 in the corneas with suture-induced NV. Suture-induced NV in rat corneas was significantly inhibited by topical treatment with 20mg/L and 10mg/L DHA. The results suggest that the effects could be partially dependent on the DHA-mediated inhibitions of the ERK1/2 and p38 pathways.