N, P and C removal simultaneously and microbial population numbers in a cyclic activated sludge system treating village and township domestic wastewater by altering the cycle times.

It was necessary to research an efficient treatment process suitable for township domestic wastewater. In this paper, the performance of the cyclic activated sludge system (CASS) system for simultaneous carbon (C), nitrogen (N) and phosphorus (P) removal was investigated by changing the operation cycle of the CASS reactor. Four operating conditions were set up, T1, T2, T3 and T4, with cycle times of 6, 8, 12 and 8 h (with carbon source), respectively. The results showed that the CASS system had good simultaneous removal of C, N and P. The highest removal rates of COD, TN, NH4+ -N and TP were 87.69, 72.99, 98.60 and 98.38%, respectively, at a cycle time of 8 h. The TN removal rate could be increased to 82.51% after the addition of carbon source. Microbial community analysis showed that Proteobacteria, Bacteroidetes and Candidatus Saccharibacteria were the main phylum-level bacteria. Their presence facilitated the effectiveness of the CASS process for nitrogen removal and phosphorus removal. Functional analysis of genes revealed that the abundance values of genes associated with C, N and P metabolism were higher when the treatment was effective.

Sanguisorba officinalis L. enhances the 5-fluorouracil sensitivity and overcomes chemoresistance in 5-fluorouracil-resistant colorectal cancer cells via Ras/MEK/ERK and PI3K/Akt pathways.

Sanguisorba officinalis L., a traditional Chinese medicine (TCM) called DiYu (DY) in China, has a strong tradition of utilization as a scorching, blood-cooling, and hemostatic medication, and was used for cancer prevention and treatment due to its potential immune-enhancing and hematological toxicity-reducing effects. Previous studies have reported significant effects of DY on cancers including colorectal cancer (CRC), which is one of the most common malignancies worldwide. The first-line cure 5-fluorouracil (5-FU) plays decisive commerce in the sedative of CRC as a clinically available chemotherapeutic agent. One of the primary causes of cancer treatment failure is the acquisition of chemotherapy drug resistance. In order to successfully combat the emergence of chemoresistance, it is essential to identify herbs or traditional Chinese medicine that have adjuvant therapeutic effects on CRC. Therefore, this study aimed to determine whether DY could improve the sensitivity, conquer the chemoresistance of 5-FU-resistant CRC cells, and investigate its intrinsic mechanism. Materials and methods: MTT, Hoechst 33258 staining, and flow cytometry assays were used to determine the anticancer activity of DY alone or in combination with 5-FU against 5-FU-resistant CRC cells (RKO-R and HCT15-R) and wound healing assays were conducted to detect cell migration. Transcriptomic techniques were carried out to explore the effect and mechanism of DY on drug-resistant CRC cells. Western Blot and RT q-PCR assays were performed to validate the mechanism by which DY overcomes drug-resistant CRC cells. Results: These results indicated that DY alone or in combination with 5-FU significantly inhibited the proliferation and the migration of resistant CRC cells, and potentiated the susceptibility of 5-FU to drug-resistant CRC cells. GO and KEGG enrichment analysis showed that the mechanisms of drug resistance in CRC cells and DY against drug-resistant CRC cells highly overlapped, involved in the modulation of biological processes such as cell migration, positive regulation of protein binding and cytoskeleton, and MAPK (Ras-ERK-MEK), PI3K/Akt, and other signaling pathways. Moreover, DY can mediate the expression of p-R-Ras, p-ERK1/2, p-MEK1/2, p-PI3K, p-AKT, HIF-1A and VEGFA proteins. In addition, DY significantly suppressed the expression of AKT3, NEDD9, BMI-1, and CXCL1 genes in resistant CRC cells. Conclusion: In conclusion, DY could inhibit the proliferation and migration of 5-FU-resistant cells and strengthen the sensitivity of 5-FU to CRC-resistant cells. Furthermore, DY may prevail over chemoresistance through the Ras/MEK/ERK and PI3K/Akt pathways. These findings imply that DY may be a potential drug for clinical treatment or adjuvant treatment of drug-resistant CRC.

Sound induces analgesia through corticothalamic circuits.

Sound-including music and noise-can relieve pain in humans, but the underlying neural mechanisms remain unknown. We discovered that analgesic effects of sound depended on a low (5-decibel) signal-to-noise ratio (SNR) relative to ambient noise in mice. Viral tracing, microendoscopic calcium imaging, and multitetrode recordings in freely moving mice showed that low-SNR sounds inhibited glutamatergic inputs from the auditory cortex (ACxGlu) to the thalamic posterior (PO) and ventral posterior (VP) nuclei. Optogenetic or chemogenetic inhibition of the ACxGlu→PO and ACxGlu→VP circuits mimicked the low-SNR sound-induced analgesia in inflamed hindpaws and forepaws, respectively. Artificial activation of these two circuits abolished the sound-induced analgesia. Our study reveals the corticothalamic circuits underlying sound-promoted analgesia by deciphering the role of the auditory system in pain processing.

Ameliorative Potential of Resveratrol in Dry Eye Disease by Restoring Mitochondrial Function.

Methods: The mitochondrial dysfunction of HCE-2 human corneal epithelial cells was induced by high osmotic pressure exposure and treated with resveratrol (50 µM). Western blotting was used to detect the expression of the antioxidant proteins SOD2, GPx, and SIRT1, and flow cytometry was used to detect cell apoptosis and ROS production. The DED mouse model was induced by 0.2% benzalkonium chloride (BAC) and treated with resveratrol. The tear yield was measured by the phenol cotton thread test, the density of cup cells in the conjunctiva was measured by periodic acid-Schiff (PAS) staining, and the expression levels of SIRT1, GPx, and SOD2 in lacrimal glands were detected by Western blotting. Results: In hypertonic conditions, the apoptosis of HCE-2 cells increased, the expression of the antioxidant proteins SOD2 and GPx decreased, ROS production increased, and the expression of SIRT1 protein, an essential regulator of mitochondrial function, was downregulated. Treatment with resveratrol reversed the mitochondrial dysfunction mediated by high osmotic pressure. In the DED mouse model, resveratrol treatment promoted tear production and goblet cell number in DED mice, decreased corneal fluorescein staining, upregulated SIRT1 expression, and induced SOD2 and GPx expression in DED mice. Conclusion: Resveratrol alleviates mitochondrial dysfunction by promoting SIRT1 expression, thus reducing ocular surface injury in mice with dry eye. This study suggests a new path against DED.

In vitro anti-Helicobacter pylori activity of Syzygium aromaticum and the preliminary mechanism of action.

ETHNOPHARMACOLOGICAL RELEVANCE: The dried flower bud of Syzygium aromaticum (L.) Merr. & L.M Perry (S. aromaticum) (Myrtaceae), also known as clove, was used in Traditional Chinese Medicine (TCM) to aid gastrointestinal function and treat stomach disorders including vomiting, flatulence and nausea. And it is a food homology medicine which is a promising candidate for H. pylori treatment. H. pylori is a Gram-negative bacterium that infects approximately 50% of the human population worldwide, which is closely related to multiple gastric diseases, including gastric cancer. However, there are still no sufficient studies on the anti-H. pylori activity of S. aromaticum, especially for the mechanism of action. AIM OF STUDY: This study aimed to study the antibacterial activities of S. aromaticum extracts on both antibiotic-sensitive and -resistant H. pylori strains, and to explore the underlying mechanisms of action. MATERIALS AND METHODS: The S. aromaticum extracts were obtained by heat reflux extraction and lyophilized to powder form. The phytochemical analyses were performed by High-performance liquid chromatography (HPLC) and UPLC-electrospray ionization mass spectrometry (ESI-MS). In vitro anti-H. pylori activity was evaluated by broth microdilution method. Mechanism of action studies included morphological observation using electron microscopy, determination of expression of virulence genes by reverse transcription quantitative polymerase chain reaction (RT-qPCR), genes expression profile identification by transcriptomic analysis, and exploration of anti-H. pylori infection mechanisms by network pharmacology analysis and western blotting validation. RESULTS: The S. aromaticum extracts, aqueous extract (AE) and 75% hydroalcoholic extract (HE), exerted significant antibacterial activities against both antibiotic-sensitive and -resistant H. pylori strains with MICs of 160∼320 µg/ml, without developing drug resistance. Among them, AE was bactericide to all the tested strains with MBCs of less than 4MIC, while HE was merely bacteriostatic to most of the tested strains with MBCs of 2MIC∼16MIC. Besides, they showed no antagonistic effects in combination with clarithromycin, metronidazole, levofloxacin, and amoxicillin. Additionally, these extracts altered the morphology and ultrastructure and down-regulated the virulence genes expression of H. pylori. And transcriptomic analysis showed that they regulated genes expression of multiple H. pylori biological processes, including tricarboxylic acid cycle (TAC) and pyruvate metabolic pathways. Furthermore, these extracts combated the abnormal activation of PI3K-Akt and MAPK signaling pathways caused by H. pylori infection. CONCLUSIONS: Overall, the present study firstly analyzed the chemical compositions of S. aromaticum extracts, and then confirmed their activities on both antibiotic-sensitive and -resistant H. pylori strains. In addition, the mechanisms of action of S. aromaticum extracts against H. pylori were found to be destroying the bacterial structure, down-regulating the expression of virulence genes, and interfering TAC and pyruvate metabolic pathways. Finally, S. aromaticum extracts were found to combated the abnormal activation of PI3K-Akt and MAPK signaling pathways to treat H. pylori infection. This study should accelerate further research and application of S. aromaticum against H. pylori infection.

In-vitro anti-Helicobacter pylori activity and preliminary mechanism of action of Canarium album Raeusch. fruit extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Canarium album Raeusch. belongs to the Burseraceae family. Its ripe fruits, known as Qing Guo (QG) in Traditional Chinese Medicine (TCM), are used to treat sore throat, cough, and fish or crab poisoning. QG was reported to have antibacterial activity, and it exerted excellent anti-Helicobacter pylori (H. pylori) activity in our screening of abundant TCM. However, few studies have reported its anti-H. pylori activity and mechanism. AIM OF STUDY: The commonly used eradication therapies for H. pylori infection are antibiotic-based therapies. With the increasing antibiotic resistance of H. pylori, interest in finding alternative therapies has been aroused. This study investigated the phytochemistry profile, in vitro anti-H. pylori activity and possible anti-bacterial mechanism of QG extracts. MATERIALS AND METHODS: QG extracts were obtained by heat reflux extraction, ultrasonic extraction or liquid-liquid extraction with different solvents. The quantitative and qualitative phytochemical analyses were performed by colorimetric determination, high-performance liquid chromatography (HPLC), and UPLC-electrospray ionization mass spectrometry (ESI-MS). In vitro anti- H. pylori activity was assessed by broth micro-dilution method. Mechanism of action studies included morphological observation using electron microscopy, urease inhibition assay and determination of expression of virulence genes by RT-qPCR. RESULTS: All QG extracts especially ethyl acetate extract (QGEAE) were rich in phenolic components, with the minimum inhibitory concentrations (MICs) on H.pylori of 39-625 µg/ml and minimum bactericidal concentrations (MBCs) of 78-1250 µg/ml. Both aqueous extract (QGAE) and QGEAE could induce the morphological and structural changes of H. pylori, inhibit urease activity with IC50 of 1093 µg/ml and 332.90 µg/ml, respectively, and down-regulate the virulence genes, such as vacA and cagA. CONCLUSIONS: QG may exhibit in vitro anti-H. pylori activity by inhibiting growth, destroying the bacterial structure and down-regulating the expression of virulence factors. Moreover, QG is the homology of food and TCM, which can be considered as a safe and convenient agent against H. pylori infection.

Effects of chitooligosaccharide-functionalized graphene oxide on stability, simulated digestion, and antioxidant activity of blueberry anthocyanins.

In this study, we tested the in vitro efficacy of a graphene oxide-chitooligosaccharide (GO-COS) complex developed to protect blueberry anthocyanins (An) from degradation by various physicochemical factors and the digestive process. We prepared a GO-COS complex to adsorb An and protect them from the destructive effects of their ambient environment. The complex protected the An under various temperature, pH, light, oxidant, and reductant conditions. We evaluated An content and composition in a simulated digestive system using the pH differential method and the high performance liquid chromatography-mass spectrometry (HPLC-MS). The GO-COS carrier stabilized An in the intestine and protected their peroxyl radical-scavenging capacity. Additionally, we observed a dose-response relationship between An content and cellular antioxidant activity, and simultaneous improvement of An bioavailability when the An were encapsulated in the complex. The complex inhibited HepG2 cell proliferation at the tested dose range. This study provides valuable information for stability of An-rich products.

Sanguisorba officinalis L. suppresses 5-fluorouracil-sensitive and-resistant colorectal cancer growth and metastasis via inhibition of the Wnt/ß-catenin pathway.

BACKGROUND: Colorectal cancer (CRC) is a widespread cancer with high morbidity and mortality. Chemoresistance and metastasis are the current challenges for CRC treatment. Sanguisorba officinalis Linn. (called DiYu in Chinese, DY) is a traditional Chinese medicine (TCM) whose root is long used as medicinal part. In our previous study, the aqueous extract of DY could inhibit the Wnt/ß-catenin pathway and showed great antitumor effect against CRC. The Wnt/ß-catenin pathway is involved in CRC chemoresistance and metastasis. However, there is little study on the antitumor and antimetastatic effects of DY on resistant CRC cells. The aim of this study is to explore the effect of aqueous extract of DY on the growth and metastasis of 5-fluorouracil (5-FU) sensitive and resistant CRC, and to elucidate the underlying molecular mechanism. METHODOLOGY: In this study, cell viability, cell colony formation and apoptosis analyses were performed to verify the in vitro antitumor effect of DY on 5-FU-sensitive and -resistant CRC cells. Next, transwell assays were used to test the inhibition activity of DY on CRC migration and invasion. Western Blotting assays were carried out to identify the molecular mechanism underlying the efficacy of DY extract. Xenograft CRC nude mice model and tumor metastasis model were used to confirm the in vivo antitumor and antimetastatic effects of DY. RESULTS: DY inhibited cell proliferation and apoptosis via the upregulation of Bax, cleaved-caspase3 and cleaved-PARP proteins and downregulation of Bcl-2 protein. DY also inhibited cell migration and invasion via the downregulation of N-cadherin, vimentin and snail proteins and upregulation of E-cadherin protein, demonstrating that DY suppressed cell metastasis by reversing epithelial-to-mesenchymal transition (EMT) procession. Moreover, the protein expression levels of ß-catenin in whole cell, cytoplasm and nucleus were decreased after DY treatment. Taken together, DY suppressed CRC cell growth and metastasis via inhibition of the Wnt pathway. Additionally, DY also demonstrated effective antitumor and anti-metastasis activities in vivo. CONCLUSIONS: In conclusion, DY suppressed the growth and metastasis of 5-FU-sensitive and -resistant CRC via inhibition of the Wnt pathway, which indicated that DY could be a potential drug to treat CRC patients and improve clinic outcome.

A Bioactive Compound from Sanguisorba officinalis L. Inhibits Cell Proliferation and Induces Cell Death in 5-Fluorouracil-Sensitive/Resistant Colorectal Cancer Cells.

Colorectal cancer (CRC) is one of the most common cancer in the world. The first line chemotherapeutic agent, 5-fluorouracil (5-FU), plays a predominant role in the clinical treatment of CRC. However, with the wide use of 5-FU, more and more CRC patients have been obtaining drug resistance to 5-FU, which leads to a large amount of treatment failures. One of the effective strategies to overcome this obstacle is to find bioactive natural products from traditional medicine. In our previous work, Sanguisorba officinalis L. was found to exert a strong anti-proliferative activity against 5-FU-senstive/resistant CRC cells. Therefore, several compounds were isolated from this herb and screened for their anti-CRC effects to find promising compounds. Among them, a triterpenoid compound named 3ß-[(α-l-arabinopyranosyl) oxy]-urs-12,18(19)-dien-28-oic acid ß-d-glucopyranosyl ester (AGE), showed strong activity against both 5-FU-senstive and resistant CRC cells. In order to further study the mechanism of AGE on CRC cells, flow cytometer analysis, mitochondrial membrane potential (MMP) measurement, Western blotting, and RT-PCR assays were performed. Results demonstrated that AGE induced cell death by apoptosis pathway and autophagy, and inhibited cell proliferation via cell cycle arrest in G0-G1 phase mediated by Wnt signaling pathway. Therefore, AGE may be a potential bioactive compound for CRC treatment in clinic.

Isoangustone A induces autophagic cell death in colorectal cancer cells by activating AMPK signaling.

Phytochemicals, especially flavonoids, have been widely investigated for their diversified pharmacological activities including anticancer activities. Previously we identified isoangustone A from licorice-derived compounds as a potent inducer of cell death. In the present study, the exact mechanism by which isoangustone A induced cell death was further investigated, with autophagy as an indispensible part of this process. Isoangustone A treatment activated autophagic signaling and induced a complete autophagic flux in colorectal cancer cells. Knockdown of ATG5 or pre-treatment with autophagy inhibitors significantly reversed isoangustone A-induced apoptotic signaling and loss of cell viability, suggesting autophagy plays an important role in isoangustone A-induced cell death. Isoangustone A inhibited Akt/mTOR signaling, and overexpressing of a constitutively activated Akt mildly suppressed isoangustone A-induced cell death. More importantly, isoangustone A inhibited cellular ATP level and activated AMPK, and pre-treatment with AMPK inhibitor or overexpression of dominant negative AMPKα2 significantly reversed isoangustone A-induced autophagy and cell death. Further study shows isoangustone A dose-dependently inhibited mitochondrial respiration, which could be responsible for isoangustone A-induced activation of AMPK. Finally, isoangustone A at a dosage of 10 mg/kg potently activated AMPK and autophagic signaling in and inhibited the growth of SW480 human colorectal xenograft in vivo. Taken together, induction of autophagy through activation of AMPK is an important mechanism by which isoangustone A inhibits tumor growth, and isoangustone A deserves further investigation as a promising anti-cancer agent.

In vitro anti-bacterial activity and network pharmacology analysis of Sanguisorba officinalis L. against Helicobacter pylori infection.

BACKGROUND: Helicobacter pylori (H. pylori) infection has become an international public health problem, and antibiotic-based triple or quadruple therapy is currently the mainstay of treatment. However, the effectiveness of these therapies decreases due to resistance to multiple commonly used antibiotics. Sanguisorba officinalis L. (S. officinalis), a traditional Chinese medicine clinically used for hemostasis and treatment of diarrhea, has various pharmacological activities. In this study, in vitro antimicrobial activity was used for the preliminary evaluation of S. officinalis against H. pylori. And a pharmacology analysis approach was also utilized to elucidate its underlying mechanisms against H. pylori infection. METHODS: Micro-broth dilution method, agar dilution method, checkerboard assay, scanning electron microscopy (SEM), and transmission electron microscopy (TEM) were used for the assessment of anti-bacterial activity. Active ingredients screening, GO analysis, KEGG analysis, construction of PPI network, molecular docking, and RT-qPCR were used to elucidate the underlying pharmacological mechanisms of S. officinalis against H. pylori infection. RESULTS: The minimum inhibitory concentration (MIC) values of S. officinalis against multiple H. pylori strains including clinically isolated multi-drug resistant (MDR) strains were ranging from 160 to 320 µg/ml. These results showed that S. officinalis had additive interaction with four commonly used antibiotics and could exert antibacterial effect by changing the morphology of bacteria without developing drug resistance. Through network pharmacology analysis, 8 active ingredients in S. officinalis were screened out for subsequent studies. Among 222 putative targets of S. officinalis, 49 targets were identified as potential targets for treatment of H. pylori infection. And these 49 targets were significantly enriched in GO processes such as protein kinase B signaling, protein kinase activity, protein kinase binding, and KEGG pathways such as Pathways in cancer, MicroRNAs in cancer, and TNF signaling pathway. Protein-protein interaction analysis yielded 5 core targets (AKT1, VEGFA, EGFR, SRC, CCND1), which were validated by molecular docking and RT-qPCR. CONCLUSIONS: Overall, this study confirmed the in vitro inhibitory activity of S. officinalis against H. pylori and explored the possible pharmacological mechanisms, laying the foundation for further research and clinical application.

Effect of Blueberry Anthocyanin-Rich Extracts on Peripheral and Hippocampal Antioxidant Defensiveness: The Analysis of the Serum Fatty Acid Species and Gut Microbiota Profile.

The current study investigated the positive effects of blueberry anthocyanin-rich extracts (BAE) on either peripheral or hippocampal antioxidant defensiveness and established the connection of the improved antioxidant status with the altered fatty acid species and gut microbiota profile. High-fat diet-induced oxidative stress in C57BL/6 mice was attenuated by BAE administration, which was reflected by strengthened antioxidant enzymes, alleviated hepatic steatosis, and improved hippocampal neuronal status. Serum lipidomics analysis indicated that the fatty acid species were altered toward the elevated unsaturated/saturated ratio, along with phospholipid species toward enriched n-3 polyunsaturated fatty acid compositions. The modulated antioxidant pattern could be attributed to the increased bacteria diversity, stimulated probiotics (Bifidobacterium and Lactobacillus) and short-chain fatty acid (SCFA) producers (Roseburia, Faecalibaculum, and Parabacteroides) improved by anthocyanins and their metabolites, which improved the colon environment, characterized by promoted SCFAs, restored colonic mucosa, and reorganized microbial structure. Thus, anthocyanin-rich dietary intervention is a promising approach for the defensiveness in human oxidative damage and neurodegeneration.

Effects of α-casein and ß-casein on the stability, antioxidant activity and bioaccessibility of blueberry anthocyanins with an in vitro simulated digestion.

Blueberry anthocyanins are well-known for their diverse biological functions. However, the instability during digestion results in their weak bioavailability. The current study aimed to investigate the alteration in the stability, antioxidant capacity and bioaccessibility of blueberry anthocyanins with the addition of α-casein and ß-casein in a simulated digestion system using pH differential method, HPLC-MS analysis, peroxyl scavenging capacity (PSC) assay, cellular antioxidant activity (CAA) and penetration test. The results showed that both α-casein and ß-casein could increase the stability of blueberry anthocyanins during intestinal digestion and protect their antioxidant capacity. Moreover, the addition of α-casein or ß-casein would enhance the bioaccessibility of blueberry anthocyanins. In conclusion, our study highlights that the interaction between α-casein or ß-casein with blueberry anthocyanins can protect the compounds against influences associated with the simulated digestion.

Serum Ceramide Reduction by Blueberry Anthocyanin-Rich Extract Alleviates Insulin Resistance in Hyperlipidemia Mice.

Blueberry anthocyanin-rich extract (BAE) was supplemented to high-fat diet (HFD)-fed mice to investigate sphingolipid metabolism modulating factors involved in the attenuated hyperinsulinemia and hyperlipidemia. A BAE-containing diet effectively controlled food intake and liver weight and significantly attenuated insulin resistance triggered by a HFD. Higher BAE (200 mg/kg of body weight) administration performed more efficiently in the improvement of hepatic steatosis and adipocyte hypertrophy, together with distinct suppressions in serum triacylglycerol and cholesterol in total and species. Serum lipid compositions revealed 200 mg/kg of BAE supplementation remarkably suppressed ceramide accumulation. Consistently, genes encoding enzymes associated with sphingomyelin conversion and ceramide de novo synthesis were modulated toward a healthy direction for restrained sphingolipid accumulation. Further, the inhibited mRNA expressions of protein phosphatase 2A and protein kinase Cζ involved in blocking Akt phosphorylation connected the controlled ceramides with the restored insulin sensitivity.

Post-transcriptional regulation activity through alternative splicing involved in the effects of Aloe vera on the Wnt/ß-catenin and Notch pathways in colorectal cancer cells.

Aloe vera (L.) Burm.f. is widely used as laxative drugs, cosmetics, and functional food due to a variety of therapeutic effects. However, several studies indicated a colonic carcinogenic activity of Aloe vera. But the underline mechanism has not been well clarified. This study aimed to explore the potential mechanism at the post-transcriptional level. Identification of Differential Expressed Alternative Splicing (DEAS) genes and events and the corresponding functional enrichment analyses were conducted on RKO cells after treated with Aloe vera aqueous extract and its two active components, aloin and aloesin. And RT-qPCR was conducted for validation. Results indicated that they induced 2200, 2342 and 2133 DEAS events, respectively. The GO enrichment and the COG classification results of DEAS genes showed that they were associated with transcription, as well as functions like signal transduction mechanisms. Moreover, DEAS genes related to the two colorectal cancerous pathways, Wnt and Notch pathways, were annotated. In conclusion, aloe extract, aloin and aloesin significantly regulated the DEAS profile of RKO cells. The colonic carcinogenicity of Aloe vera may due to its post-transcriptional regulatory activity through Alternative Splicing (AS) on genes, especially on Wnt-related and Notch-related key genes.

Investigation into perturbed nucleoside metabolism and cell cycle for elucidating the cytotoxicity effect of resveratrol on human lung adenocarcinoma epithelial cells.

In an effort to understand the molecular events contributing to the cytotoxicity activity of resveratrol (RSV), we investigated its effects on human lung adenocarcinoma epithelial cell line A549 at different concentrations. Cellular nucleoside metabolic profiling was determined by an established liquid chromatography-mass spectrometry method in A549 cells. RSV resulted in significant decreases and imbalances of deoxyribonucleoside triphosphates (dNTPs) pools suppressing subsequent DNA synthesis. Meanwhile, RSV at high concentration caused significant cell cycle arrest at S phase, in which cells required the highest dNTPs supply than other phases for DNA replication. The inhibition of DNA synthesis thus blocked subsequent progression through S phase in A549 cells, which may partly contribute to the cytotoxicity effect of RSV. However, hydroxyurea (HU), an inhibitor of RNR activity, caused similar dNTPs perturbation but no S phase arrest, finally no cytotoxicity effect. Therefore, we believed that the dual effect of high concentration RSV, including S phase arrest and DNA synthesis inhibition, was required for its cytotoxicity effect on A549 cells. In summary, our results provided important clues to the molecular basis for the anticancer effect of RSV on epithelial cells.

Study of the aqueous extract of Aloe vera and its two active components on the Wnt/ß-catenin and Notch signaling pathways in colorectal cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Aloe vera (L.) Burm. f. (Aloe vera) is a common Traditional Chinese Medicine (TCM) recorded in Pharmacopoeia of the People's Republic of China (version 2015). It has been traditionally used for treatment of constipation. Aloe vera requires much attention for its safety evaluation because several studies have reported the association between oral consumption of Aloe vera and the development of colorectal cancer (CRC). However the material basis and molecular mechanism are.still less well elucidated. Although Wnt/ß-catenin and Notch signaling pathway have been known to be closely related to the initiation and development of CRC, the impacts of Aloe vera on these cancerous pathways have not been completely determined yet. AIM OF THIS STUDY: Hence, this study aimed to study the impacts of Aloe vera on the Wnt/ß-catenin and Notch signaling pathway, as well as proliferation of CRC cells. MATERIALS AND METHODS: Firstly, the effects of Aloe vera aqueous extract and its two active components (aloin and aloesin) on the Wnt/ß-catenin and Notch signaling pathway were studied by luciferase reporter, RT-qPCR, western blotting and immunofluorescence assays, respectively. Furthermore, RNA sequencing analysis (RNA-seq) was then performed to verify their regulatory activities on the Wnt-related and Notch-related genes expression. Finally, their impacts on RKO cell proliferation and cell cycle phase were also evaluated via MTT assay and cell cycle analysis. RESULTS: Our results indicate that the aqueous extract of Aloe vera and its active component aloin activated the Wnt/ß-catenin pathway and inhibited the Notch signaling pathway only in the presence of Wnt3a. While aloesin was characterized to directly activate the Wnt/ß-catenin pathway and inhibit the Notch pathway independent of Wnt3a. Within 24h, the Aloe vera extract and its two components were failed to affect the proliferation or cell cycle phase of RKO cells. Nevertheless, in the presence of Wnt3a, the aqueous extract of Aloe vera with the concentration of 33.3 µg/ml start to promote the cell proliferation of RKO cells after 48h incubation. CONCLUSION: In conclusion, this study showed that Aloe vera extract and its active component aloin activated the Wnt/ß-catenin pathway and inhibited the Notch pathway in the presence of Wnt3a. While another active component, aloesin, activated the Wnt/ß-catenin pathway and inhibited the Notch signaling pathway independent of Wnt3a. Given that Wnt/ß-catenin and Notch pathway are closely associated with the progression of CRC, these findings would be helpful to better understand the colonic carcinogenicity of Aloe vera.

Blueberry Malvidin-3-galactoside Suppresses Hepatocellular Carcinoma by Regulating Apoptosis, Proliferation, and Metastasis Pathways In Vivo and In Vitro.

Anthocyanin, a natural antioxidant, is reported to have cytotoxicity against cancer cells; however, the mechanism remains unclear. The aim of the present study was to investigate the mechanism by which malvidin-3-galactoside (M3G), the prominent anthocyanin in blueberry, suppresses the development of hepatocellular carcinoma. In vitro, M3G suppressed the proliferation, polarization, migration, and invasion activities of HepG2 cells by regulating the protein expression of cyclin D1, cyclin B, cyclin E, caspase-3, cleaved caspase-3, Bax, p-JNK, and p-p38, activating phosphatase and tensin homologue deleted on chromosome 10 (PTEN), accompanied by a decrease in the p-AKT level, and lowering the protein expression levels of MMP-2 and MMP-9. In vivo, M3G promoted the apoptosis of liver tumor cells, as determined by immunohistochemistry (cleaved caspase-3, Ki-67, PTEN, and p-AKT), a terminal deoxynucleotidyl transferase dUTP nick end labeling assay, and hematoxylin-eosin staining. Overall, these results suggest that M3G, as an adjuvant ingredient or nutritional supplement, may be beneficial for liver cancer prevention and the modulatory mechanism seems to be associated with inhibition of proliferation, apoptosis, migration, and invasion-related pathways.

Spatially constrained tandem bromodomain inhibition bolsters sustained repression of BRD4 transcriptional activity for TNBC cell growth.

The importance of BET protein BRD4 in gene transcription is well recognized through the study of chemical modulation of its characteristic tandem bromodomain (BrD) binding to lysine-acetylated histones and transcription factors. However, while monovalent inhibition of BRD4 by BET BrD inhibitors such as JQ1 blocks growth of hematopoietic cancers, it is much less effective generally in solid tumors. Here, we report a thienodiazepine-based bivalent BrD inhibitor, MS645, that affords spatially constrained tandem BrD inhibition and consequently sustained repression of BRD4 transcriptional activity in blocking proliferation of solid-tumor cells including a panel of triple-negative breast cancer (TNBC) cells. MS645 blocks BRD4 binding to transcription enhancer/mediator proteins MED1 and YY1 with potency superior to monovalent BET inhibitors, resulting in down-regulation of proinflammatory cytokines and genes for cell-cycle control and DNA damage repair that are largely unaffected by monovalent BrD inhibition. Our study suggests a therapeutic strategy to maximally control BRD4 activity for rapid growth of solid-tumor TNBC cells.

Multisensor-integrated organs-on-chips platform for automated and continual in situ monitoring of organoid behaviors.

Organ-on-a-chip systems are miniaturized microfluidic 3D human tissue and organ models designed to recapitulate the important biological and physiological parameters of their in vivo counterparts. They have recently emerged as a viable platform for personalized medicine and drug screening. These in vitro models, featuring biomimetic compositions, architectures, and functions, are expected to replace the conventional planar, static cell cultures and bridge the gap between the currently used preclinical animal models and the human body. Multiple organoid models may be further connected together through the microfluidics in a similar manner in which they are arranged in vivo, providing the capability to analyze multiorgan interactions. Although a wide variety of human organ-on-a-chip models have been created, there are limited efforts on the integration of multisensor systems. However, in situ continual measuring is critical in precise assessment of the microenvironment parameters and the dynamic responses of the organs to pharmaceutical compounds over extended periods of time. In addition, automated and noninvasive capability is strongly desired for long-term monitoring. Here, we report a fully integrated modular physical, biochemical, and optical sensing platform through a fluidics-routing breadboard, which operates organ-on-a-chip units in a continual, dynamic, and automated manner. We believe that this platform technology has paved a potential avenue to promote the performance of current organ-on-a-chip models in drug screening by integrating a multitude of real-time sensors to achieve automated in situ monitoring of biophysical and biochemical parameters.