Evaluation of Anti-tumor and Chemoresistance-lowering Effects of Pectolinarigenin from Cirsium japonicum Fisch ex DC in Breast Cancer

Purpose: To investigate the antitumor and chemoresistance-lowering effects of pectolinarigenin on breast cancer cells.Methods: Pectolinarigenin was purified by a combination of silica gel and Sephadex LH-20 column chromatography from ethanol extracts of the aerial parts of C. japonicum DC. Breast cancer selfrenewal properties were tested by colony formation and tumor sphere formation assays. Thereafter, real-time polymerase chain reaction (PCR) was used to detect breast cancer stem cell markers. Furthermore, the effect of pectolinarigenin on breast cancer cell was evaluated by chemoresistance using 3-(4,5-dimethyl-2 thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Finally, tumor formation in nude mice was used to test the effect of pectolinarigenin on tumorigenicity of breast cancer cells in vivo.Results: The results showed that pectolinarigenin, extracted from Cirsium japonicum Fisch. ex DC., inhibited tumor cell self-renewal in MCF-7 breast cancer cells. Pectolinarigenin (25 μM) caused significant inhibition of colony formation (61.23 %, p < 0.001) and tumor sphere formation (59.49 %, p < 0.01) in MCF-7. The inhibitory effects were associated with changes in breast cancer stem cell markers. Treatment of breast cancer cells with pectolinarigenin reduced the chemoresistance of the cells to doxorubicin. At the same time, mRNA expression of chemoresistance genes (ATP binding cassette subfamily G member 2, ABCG2 and ATP binding cassette subfamily B member 1, MDR1) was repressed by pectolinarigenin. The inhibition efficiency of MDR1 and ABCG2 by 10 μM pectolinarigenin treatment was about 59.29 (p < 0.01) and 46.48 % (p < 0.01), respectively. Furthermore, pectolinarigenin reduced tumor mass in nude mice xenograft model.Conclusion: Pectolinarigenin inhibits breast cancer stem cell-like properties and lowers the chemoresistance of the cancer cells to chemotherapy. The results provide an insight into the mechanism of the anti-breast tumor effects and an experimental basis for the use of pectolinarigenin to enhance treatment of patients with breast cancer.Keywords: Pectolinarigenin, Cancer stem cells, Breast cancer, Chemoresistance, Cirsium japonicum Fisch. ex D

Strand-specific PCR of UV radiation-damaged genomic DNA revealed an essential role of DNA-PKcs in the transcription-coupled repair

<p>Abstract</p> <p>Background</p> <p>In eukaryotic cells, there are two sub-pathways of nucleotide excision repair (NER), the global genome (gg) NER and the transcription-coupled repair (TCR). TCR can preferentially remove the bulky DNA lesions located at the transcribed strand of a transcriptional active gene more rapidly than those at the untranscribed strand or overall genomic DNA. This strand-specific repair in a suitable restriction fragment is usually determined by alkaline gel electrophoresis followed by Southern blotting transfer and hybridization with an indirect end-labeled single-stranded probe. Here we describe a new method of TCR assay based on strand-specific-PCR (SS-PCR). Using this method, we have investigated the role of DNA-dependent protein kinase catalytic subunit (DNA-PKcs), a member of the phosphatidylinositol 3-kinase-related protein kinases (PIKK) family, in the TCR pathway of UV-induced DNA damage.</p> <p>Results</p> <p>Although depletion of DNA-PKcs sensitized HeLa cells to UV radiation, it did not affect the ggNER efficiency of UV-induced cyclobutane pyrimidine dimers (CPD) damage. We postulated that DNA-PKcs may involve in the TCR process. To test this hypothesis, we have firstly developed a novel method of TCR assay based on the strand-specific PCR technology with a set of smart primers, which allows the strand-specific amplification of a restricted gene fragment of UV radiation-damaged genomic DNA in mammalian cells. Using this new method, we confirmed that siRNA-mediated downregulation of Cockayne syndrome B resulted in a deficiency of TCR of the UV-damaged dihydrofolate reductase (<it>DHFR</it>) gene. In addition, DMSO-induced silencing of the c-myc gene led to a decreased TCR efficiency of UV radiation-damaged c-myc gene in HL60 cells. On the basis of the above methodology verification, we found that the depletion of DNA-PKcs mediated by siRNA significantly decreased the TCR capacity of repairing the UV-induced CPDs damage in <it>DHFR </it>gene in HeLa cells, indicating that DNA-PKcs may also be involved in the TCR pathway of DNA damage repair. By means of immunoprecipitation and MALDI-TOF-Mass spectrometric analysis, we have revealed the interaction of DNA-PKcs and cyclin T2, which is a subunit of the human transcription elongation factor (P-TEFb). While the P-TEFb complex can phosphorylate the serine 2 of the carboxyl-terminal domain (CTD) of RNA polymerase II and promote transcription elongation.</p> <p>Conclusion</p> <p>A new method of TCR assay was developed based the strand-specific-PCR (SS-PCR). Our data suggest that DNA-PKcs plays a role in the TCR pathway of UV-damaged DNA. One possible mechanistic hypothesis is that DNA-PKcs may function through associating with CyclinT2/CDK9 (P-TEFb) to modulate the activity of RNA Pol II, which has already been identified as a key molecule recognizing and initializing TCR.</p

Concept for a Future Super Proton-Proton Collider

Following the discovery of the Higgs boson at LHC, new large colliders are being studied by the international high-energy community to explore Higgs physics in detail and new physics beyond the Standard Model. In China, a two-stage circular collider project CEPC-SPPC is proposed, with the first stage CEPC (Circular Electron Positron Collier, a so-called Higgs factory) focused on Higgs physics, and the second stage SPPC (Super Proton-Proton Collider) focused on new physics beyond the Standard Model. This paper discusses this second stage.Comment: 34 pages, 8 figures, 5 table

Tumor Mutation Burden Predicts Relapse in Papillary Thyroid Carcinoma With Changes in Genes and Immune Microenvironment

BackgroundThe risk factors of papillary thyroid carcinoma (PTC) recurrence are meaningful for patients and clinicians. Tumor mutation burden (TMB) has been a biomarker for the effectiveness of immune checkpoint inhibitor (ICI) and prognosis in cancer. However, the role of TMB and its latent significance with immune cell infiltration in PTC are still unclear. Herein, we aimed to explore the effect of TMB on PTC prognosis.Material and MethodsRNA-seq and DNA-seq datasets of PTC patients were downloaded from The Cancer Genome Atlas (TCGA) database. The Gene Ontology (GO) and gene set enrichment analysis (GSEA 4.0.1) were applied further to explore potential differences in PTC patients’ biological functions. The differentially expressed genes (DEGs) and immune microenvironment between the high and low TMB groups were determined.ResultsTMB had the highest AUC score than other clinical indicators in ROC analysis on recurrence-free survival, and a higher TMB score was related to a worse prognosis. Further, GSEA showed a higher level of oxidative phosphorylation (OXPHOS) in the high TMB group, and four genes correlated with recurrence-free survival rate were identified. The abundance of CD8+ T cells and M1 macrophages in the high TMB group was significantly lower than that in the low TMB group.ConclusionsOur study found that TMB was a better predictor variable at evaluating the risk of PTC recurrence. Moreover, TMB-related genes conferred dramatically correlated prognosis, which was worth exploring in guiding postoperative follow-up and predicting recurrence for PTC patients

Sirtuin 6 maintains epithelial STAT6 activity to support intestinal tuft cell development and type 2 immunity

Dynamic regulation of intestinal epithelial cell (IEC) differentiation is crucial for both homeostasis and the response to helminth infection. SIRT6 belongs to the NAD+-dependent deacetylases and has established diverse roles in aging, metabolism and disease. Here, we report that IEC Sirt6 deletion leads to impaired tuft cell development and type 2 immunity in response to helminth infection, thereby resulting in compromised worm expulsion. Conversely, after helminth infection, IEC SIRT6 transgenic mice exhibit enhanced epithelial remodeling process and more efficient worm clearance. Mechanistically, Sirt6 ablation causes elevated Socs3 expression, and subsequently attenuated tyrosine 641 phosphorylation of STAT6 in IECs. Notably, intestinal epithelial overexpression of constitutively activated STAT6 (STAT6vt) in mice is sufficient to induce the expansion of tuft and goblet cell linage. Furthermore, epithelial STAT6vt overexpression remarkedly reverses the defects in intestinal epithelial remodeling caused by Sirt6 ablation. Our results reveal a novel function of SIRT6 in regulating intestinal epithelial remodeling and mucosal type 2 immunity in response to helminth infection

Common Phenolic Metabolites of Flavonoids, but Not Their Unmetabolized Precursors, Reduce the Secretion of Vascular Cellular Adhesion Molecules by Human Endothelial Cells

Background: Flavonoids have been implicated in the prevention of cardiovascular disease; however, their mechanisms of action have yet to be elucidated, possibly because most previous in vitro studies have used supraphysiological concentrations of unmetabolized flavonoids, overlooking their more bioavailable phenolic metabolites. Objective: We aimed to explore the effects of phenolic metabolites and their precursor flavonoids at physiologically achievable concentrations, in isolation and combination, on soluble vascular cellular adhesion molecule-1 (sVCAM-1). Method: Fourteen phenolic acid metabolites and 6 flavonoids were screened at 1 μM for their relative effects on sVCAM-1 secretion by human umbilical vein endothelial cells stimulated with tumor necrosis factor alpha (TNF-α). The active metabolites were further studied for their response at different concentrations (0.01 μM–100 μM), structure-activity relationships, and effect on vascular cellular adhesion molecule (VCAM)-1 mRNA expression. In addition, the additive activity of the metabolites and flavonoids was investigated by screening 25 unique mixtures at cumulative equimolar concentrations of 1 μM. Results: Of the 20 compounds screened at 1 μM, inhibition of sVCAM-1 secretion was elicited by 4 phenolic metabolites, of which protocatechuic acid (PCA) was the most active (−17.2%, P = 0.05). Investigations into their responses at different concentrations showed that PCA significantly reduced sVCAM-1 15.2–36.5% between 1 and 100 μM, protocatechuic acid-3-sulfate and isovanillic acid reduced sVCAM-1 levels 12.2–54.7% between 10 and 100 μM, and protocatechuic acid-4-sulfate and isovanillic acid-3-glucuronide reduced sVCAM-1 secretion 27.6% and 42.8%, respectively, only at 100 μM. PCA demonstrated the strongest protein response and was therefore explored for its effect on VCAM-1 mRNA, where 78.4% inhibition was observed only after treatment with 100 μM PCA. Mixtures of the metabolites showed no activity toward sVCAM-1, suggesting no additive activity at 1 μM. Conclusions: The present findings suggest that metabolism of flavonoids increases their vascular efficacy, resulting in a diversity of structures of varying bioactivity in human endothelial cells

Development of nanosecond spike pulse power supply for electrochemical micromachining

Abstract The use of pulse voltage can greatly improve the precision of electrochemical microfabrication, and the narrower the pulse width of the applied pulse voltage signal, the higher the machining precision. However, the commonly used chopper circuit topology of pulse power supplies is limited by the maximum switching frequency of the field-effect transistor. To address this problem, this paper proposes a nanosecond pulse electrochemical micromachining power supply based on a differential circuit. The power supply uses the STM32F103C8T6 microcontroller as the control core to output high-performance rectangular waves through a DDS device. After differential, rectification, filtering, and power amplification processing, stable, frequency, amplitude, and pulse width adjustable spike pulse voltage signals are obtained. By establishing a system mathematical model and optimizing the time constant of the differential circuit, theoretically, the sub-nanosecond pulse width can be obtained. Prototype performance tests show that the power supply has a maximum frequency of 20 MHz, a minimum pulse width of 1.8 ns, and a maximum peak voltage of 10 V. By using this power supply for microhole electrochemical machining experiments, nanometer-level machining precision has been achieved

Study on visible-light photocatalytic activity of g-C3N4 modified by trace Ag3PO4

Novel polymeric Ag3PO4/g-C3N4 photocatalysts modified by trace Ag3PO4 are prepared via a facile method. The obtained Ag3PO4/g-C3N4 composite products are characterized by X-ray diffraction (XRD), Fourier Transform Infrared Spectroscopy (FT-IR), transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), UV-vis diffuse reflection spectroscopy (DRS), and photoluminescence spectroscopy (PL). The photocatalytic activities of Ag3PO4/g-C3N4 samples are investigated by evaluating the decomposition of methyl orange (MO) under visible light irradiation. The results reveal that the polymeric material is doped by Ag3PO4, and the Ag3PO4 doping can hardly affect the crystallinity of the as-prepared g-C3N4 materials, but it improves greatly their visible light absorption and separation efficiency of photogenerated electron/hole pairs. The Ag3PO4/g-C3N4 photocatalysts exhibits significantly enhanced photocatalytic performance