A novel lupene derivative from Thymus capitatus possesses an apoptosis-inducing effect via Let-7 miRNA/Cyclin D1/VEGF cascade in the A549 cell line.

Non-small-cell lung carcinoma (NSCLC) is a type of epithelial lung cancer accounting for about 85% of all lung cancers. In our research, a novel lupene derivative namely acetoxy-lup-5(6), 20(29)-diene (ALUP), as well as two known triterpenes; lupeol (LUP) and betulinic acid (BA) were isolated through the chromatographic purification of the 95% ethanolic extract of Thymus capitatus. Identification of the compounds was carried out by physicochemical properties as well as spectral 1D and 2D NMR analysis. The anti-cancer activity of the three triterpenes was assessed on non-small cell lung cancer cell line; A549 using MTT assay and cell cycle analysis using annexin V/propidium iodide. The molecular mechanism underlying anti-apoptotic effects was determined by analyzing Let-7 miRNA and miRNA-21 expression, the mRNA gene expression level of Bax, CASP-8, CD95, Bcl2, KRAS, VEGF, Cyclin D1 using qRT-PCR. Our results revealed that the three isolated compounds ALUP, LUP, and BA caused cell cycle arrest at the G2/M phase with an increase in the apoptosis which may be attributed to their significant effect on raising Bax, CASP-8, and CD95 and reducing the mRNA expression levels of Bcl-2, KRAS, VEGF, and Cyclin D1 compared to control cells. RT-PCR results showed that the ALUP, LUP, and BA significantly downregulated miRNA-21 expression. Meanwhile, the three compounds caused significant overexpression of Let-7 miRNA. This is the first report on the anti-cancer activity of acetoxy-lup-5(6), 20(29)-diene (ALUP) in reducing the proliferation and differentiation of the A549 cell line through inducing apoptosis. Finally, by targeting the Let-7 miRNA/Cyclin D1/VEGF cascade, acetoxy-lup-5(6), 20(29)-diene could be a potential therapeutic agent for lung cancer.

Enhancing Anti-cancer Activity: Green Synthesis and Cytotoxicity Evaluation of Turmeric-Gold Nanocapsules on A549 Lung Cancer Cells.

Background Lung cancer remains a major global health concern, with a notable increase in new cases in recent years. This study aims to investigate the cytotoxic effects of polymeric turmeric-gold nanocapsules on A549 human lung cancer cells, utilizing green-synthesized gold nanoparticles from Curcuma longa L. and ethyl cellulose-based nanocapsules. Methods Gold nanoparticles were synthesized using the aqueous root extract of Curcuma longa L., and the resulting nanoparticles were characterized using UV-Vis, fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), and energy dispersive x-ray (EDX) techniques. Subsequently, polymeric nanocapsules of turmeric with encapsulated gold nanoparticles were prepared. The cytotoxicity of these nanocapsules was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay on both A549 lung cancer cell lines and normal cell lines. Results The turmeric-gold nanocapsules exhibited a half maximal inhibitory concentration (IC50) value of 40 µg/ml, while the gold nanoparticles alone showed an IC50 value of 60 µg/ml when tested on A549 cells. Furthermore, apoptosis was observed in A549 cells treated with turmeric-gold nanocapsules. The combination of gold nanoparticles and turmeric polymer (gold turmeric nanocapsules) demonstrated a more potent anti-cancer effect on the lung cancer cell line, with an IC50 value of 40 µg/ml compared to green-synthesized gold nanoparticles (IC50 of 60 µg/ml). Conclusion The utilization of polymeric nanocapsules of turmeric, with green-synthesized gold nanoparticles, presents a promising solution to overcome the limited water solubility of turmeric. The results suggest that the combination of gold nanoparticles and turmeric enhances the cytotoxic effects on A549 human lung cancer cells. These findings contribute to the potential application of turmeric-gold nanocapsules as a novel therapeutic approach in lung cancer research.

Quercitrin loaded cyclodextrin based nanosponge as a promising approach for management of lung cancer and COVID-19.

Lung cancer and pandemic acute respiratory disease, COVID-19, are examples of the most worldwide widespread diseases. The aim of the current study is to develop cyclodextrin based nanosponge (CD-NS) for loading the flavonoid drug, quercitrin (QCT). This is to improve its solubility in an attempt to enhance its activity against lung cancer as well as SARS-CoV-2 virus responsible for COVID-19. Preparation of CD-NS was performed by ultrasound-assisted synthesis method. Two CDs were employed, namely, ß cyclodextrin (ßCD) and 2-hydroxy propyl-ß-cyclodextrin (2-HPßCD) that were crosslinked with diphenyl carbonate, one at a time. QCT loaded CD-NS revealed entrapment efficiency and particle size ranged between 94.17 and 99.03% and 97.10-325.90 nm, respectively. QCT loaded 2-HPßCD-NS revealed smaller particle size compared with that of QCT loaded ßCD-NS. Zeta potential absolute values of the prepared formulations were >20 mV, indicating physically stable nanosystems. The selected formulations were investigated by Fourier transform infrared spectroscopy, X-ray powder diffraction and scanning electron microscopy which proved the formation of QCT loaded CD-NS exhibiting porous structure. QCT exhibited partial and complete amorphization in ßCD-NS and 2-HPßCD-NS, respectively. In vitro release revealed an improved release of QCT from CD-NS formulations. The biological activity of free QCT and QCT loaded CD-NS was investigated against lung cancer cell line A549 as well as SARS-CoV-2 virus. The results revealed that IC50 values of free QCT against lung cancer cell line A549 and SARS-CoV-2 were higher than those exhibited by QCT loaded CD-NS by 1.57-5.35 and 5.95-26.95 folds, respectively. QCT loaded 2-HPßCD-NS revealed enhanced in vitro release and superior biological activity compared with QCT loaded ßCD-NS.

Comparative Study of Docosahexaenoic Acid with Different Molecular Forms for Promoting Apoptosis of the 95D Non-Small-Cell Lung Cancer Cells in a PPARγ-Dependent Manner.

Cancer is a leading cause of death in worldwide. Growing evidence has shown that docosahexaenoic acid (DHA) has ameliorative effects on cancer. However, the effects of DHA-enriched phosphatidylcholine (DHA-PC) and efficacy differences between DHA-PC, DHA-triglyceride (DHA-TG), and DHA- ethyl esters (DHA-EE) on cancer cells had not been studied. In this study, 95D lung cancer cells in vitro were used to determine the effects and underlying mechanisms of DHA with different molecular forms. The results showed that DHA-PC and DHA-TG treatment significantly inhibited the growth of 95D cells by 53.7% and 33.8%, whereas DHA-EE had no significantly effect. Morphological analysis showed that DHA-PC and DHA-TG prompted promoted cell contraction, increased concentration of cell heterochromatin, vacuolization of cytoplasm, and edema of endoplasmic reticulum and mitochondria. TUNEL and AO/EB staining indicated that both DHA-PC and DHA-TG promoted cell apoptosis, in which DHA-PC performed better than DHA-TG. Mechanistically, DHA-PC and DHA-TG treatment up-regulated the PPARγ and RXRα signal, inhibited the expression of NF-κB and Bcl-2, and enhanced the expression of Bax and caspase-3, thereby promoting cell apoptosis. In conclusion, DHA-PC exerted superior effects to DHA-TG and DHA-EE in promoting apoptosis in 95D non-small-cell lung cancer cells. These data provide new evidence for the application of DHA in treatment of cancer.

PEG-4000 formed polymeric nanoparticles loaded with cetuximab downregulate p21 & stathmin-1 gene expression in cancer cell lines.

Cetuximab (CTX) is known to have cytotoxic effects on several human cancer cells in vitro; however, as CTX is poorly water soluble, there is a need for improved formulations can reach cancer cells at high concentrations with low side effects. We developed (PEG-4000) polymeric nanoparticles (PEGNPs) loaded with CTX and evaluated their in vitro cytotoxicity and anticancer properties against human lung (A549) and breast (MCF-7) cancer cells. CTX-PEGNPs were formulated using the solvent evaporation technique, and their morphological properties were evaluated. Further, the effects of CTX-PEGNPs on cell viability using the MTT assay and perform gene expression analysis, DNA fragmentation measurements, and the comet assay. CTX-PEGNP showed uniformly dispersed NPs of nano-size range (253.7 ± 0.3 nm), and low polydispersity index (0.16) indicating the stability and uniformity of NPs. Further, the zeta potential of the preparations was -17.0 ± 1.8 mv. DSC and FTIR confirmed the entrapping of CTX in NPs. The results showed IC50 values of 2.26 µg/mL and 1.83 µg/mL for free CTX and CTX-PEGNPs on the A549 cancer cell line, respectively. Moreover, CTX-PEGNPs had a lower IC50 of 1.12 µg/mL in MCF-7 cells than that of free CTX (2.28 µg/mL). The expression levels of p21 and stathmin-1 were significantly decreased in both cell lines treated with CTX-PEGNPs compared to CTX alone. The CTX-PEGNP-treated cells also showed increased DNA fragmentation rates in both cancer cell lines compared with CTX alone. The results indicated that CTX-PEGNP was an improved formulation than CTX alone to induce apoptosis and DNA damage and inhibit cell proliferation through the downregulation of P21 and stathmin-1 expression.

Organophosphate flame retardants induce oxidative stress and Chop/Caspase 3-related apoptosis via Sod1/p53/Map3k6/Fkbp5 in NCI-1975 cells.

Organophosphate flame retardants (OPFRs) have been ubiquitously detected in dust and air which could cause damage to human health through inhalation. Currently the understanding of their adverse effects and potential mechanisms on the lung are still limited. In this study, human non-small cell lung cancer cell line NCI-H1975 was used to investigate the cytotoxicity, oxidative stress, cellular apoptosis of 9 typical OPFRs with concentrations varied from 0 to 200 µM, and their toxic mechanism associated with molecular structure was compared. After 72 h, tris(1,3-dichloro-2-propyl) phosphate (TDCIPP) displayed the highest cytotoxicity, followed by 2-ethylhexyl diphenyl phosphate (EHDPP), tris(2-butoxyethyl) phosphate (TBOEP) and tris(2-chloroisopropyl) phosphate (TCIPP), while tris(2-chloroethyl) phosphate (TCEP) and tris(2-ethylhexyl) phosphate (TEHP) exhibited the least suppression on cell viability. These results indicated that the variation of cytotoxicity on OPFRs could only be partially explained by their ester linkage. Moreover, the overexpression of intracellular reactive oxygen species (ROS), free Ca2+ and cellular apoptosis suggested that exposure to OPFRs can lead to apoptosis related to oxidative stress. Six genes associated with oxidative stress and apoptosis were upregulated dramatically compared with the control, demonstrating OPFRs induced Chop/Caspase 3-related apoptosis by activating Sod1/p53/Map3k6/Fkbp5 expression in NCI-H1975 cells. This is the first study to investigate cytotoxicity and related mechanism on commonly-used OPFRs in NCI-H1975 cells.

In vitro cytotoxicity assay of Fucoidan extracted from Turbinaria conoides against cancer cell lines MCF7, A549, and normal cell line L929

Abstract The main aim of the study is to quantify the cytotoxic property of the Fucoidan extracted from the Turbinaria conoides using the MTT assay with the standard fucose. Fucoidan was extracted using the soaked water method and it was determined using the HPLC procedure the obtained Test sample Fucoidan extracted from the Turbinaria conoides and standard fucose was subjected to the cytotoxicity assay against the MCF7 Human breast cancer cell line, A549 lung cancer cell line, and L929 normal mouse fibroblast cell line. From the results it was found that the Test sample showed good IC50 value for MCF7 cell line then A549 with an increasing concentration 24 hours incubation at 37°C The IC50 for MCF7 was 115.21 µg/ml and A549 396.46µg/ml and the Fucoidan extract was checked for its cytotoxicity against the normal mouse fibroblast cell line L929, Fucoidan was found non-lethal to the L929 mouse fibroblast normal cell line. Standard fucose also gave a significant result towards MCF7 and against the L929. This indicates that the Fucoidan extracted from Tubinaria conoides shows better anticancer potential in it. Hence its application can be further extended in the pharmacological fields.

circ-PRKDC affects the proliferation, apoptosis and radiosensitivity of lung cancer cells by regulating miR-505-3p / 中华放射肿瘤学杂志

Objective:To investigate the effect of circ-PRKDC on lung cancer cell proliferation, apoptosis and radiosensitivity and its molecular mechanism.Methods:Normal lung epithelial cells BEAS-2B and lung cancer cell lines NCI-H1299, NCI-H2170, NCI-H1975 were cultured. NCI-H1299 cells were divided into the si-NC, si-PRKDC, pcDNA-NC, pcDNA-PRKDC, miR-NC, miR-505-3p, anti-miR-NC, anti-miR-505-3p, si-PRKDC+ anti-miR-NC and si-PRKDC+ anti-miR-505-3p groups. RT-qPCR was used to detect the expression levels of circ-PRKDC and miR-505-3p. Western blot was employed to measure the protein expression. MTT was used to detect cell proliferation. Flow cytometry was utilized to detect cell apoptosis. Plate clone formation assay was conducted to detect the cell radiosensitivity. Dual luciferase reporter assay was performed to analyze the targeting relationship between circ-PRKDC and miR-505-3p.Results:Compared with normal lung epithelial cells BEAS-2B, the expression levels of circ-PRKDC in the lung cancer cell lines NCI-H1299, NCI-H2170 and NCI-H1975 were significantly up-regulated (3.65, 3.10, 2.67 vs. 1.00, all P<0.05), whereas those of miR-505-3p were significantly down-regulated (0.42, 0.50, 0.54 vs. 1.02, all P<0.05). After low expression of circ-PRKDC, the expression level of CyclinD1 was significantly down-regulated (0.42 vs. 0.81, P<0.05), whereas those of Cleaved-caspase-3(0.71 vs. 0.33, P<0.05) and γ-H 2AX (0.89 vs. 0.46, P<0.05) were significantly up-regulated, the cell A value was significantly decreased (0.413 vs. 0.839, P<0.05), cell apoptosis rate was significantly increased (20.35 vs. 6.21, P<0.05), cell survival fraction was significantly decreased ( P<0.05), and β-catenin expression was significantly down-regulated (0.35 vs. 0.73, P<0.05). After high expression of miR-505-3p, the expression level of CyclinD1 was significantly down-regulated (0.34 vs. 0.83, P<0.05), those of Cleaved-caspase-3(0.65 vs. 0.32, P<0.05) and γ-H 2AX (0.96 vs. 0.45, P<0.05) were significantly up-regulated, the cell A value was significantly decreased (0.386 vs. 0.851, P<0.05), the apoptosis rate was significantly increased (16.38 vs. 6.20, P<0.05), and the cell survival fraction was significantly decreased ( P<0.05). Compared with miR-NC, the luciferase activity of miR-505-3p group transfected with circ-PRKDC wild-type reporter plasmid was significantly decreased (0.44 vs. 1.00, P<0.05). Down-regulation of miR-505-3p could reverse the effect of low expression of circ-PRKDC on the proliferation, apoptosis, radiosensitivity and β-catenin expression of NCI-H1299 cells. Conclusion:Low expression of circ-PRKDC may inhibit lung cancer cell proliferation, promote cell apoptosis and enhance cell radiosensitivity by up-regulating miR-505-3p, which is probably associated with the Wnt/β-catenin signaling pathway.

Effect and possible mechanism of Salvianolic acid B on radiosensitivity of non-small cell lung cancer / 中华放射肿瘤学杂志

Objective:To evaluate the effect of Salvianolic acid B on the radiosensitivity of human non-small cell lung cancer cells and investigate its possible mechanism.Methods:Non-small cell lung cancer cells A549 and H1299 were cultured in vitro. The toxicity of Salvianolic acid B on non-small cell lung cancer cells was detected by MTT assay. The effect of Salvianolic acid B on the radiosensitivity was assessed by clone formation assay. Transwell chamber assay was used to evaluate the effect of Salvianolic acid B on the migration of tumor cells. Western blot was employed to detect the expression levels of OTUD7B, MMP-2, MMP-9, E-cadherin, Akt and p-Akt regulated by Salvianolic acid B. Results:Salvianolic acid B (5 μmol/L) could inhibit the proliferation of A549 and H1299 cells. Clone formation assay showed that Salvianolic acid B increased the radiosensitivity of A549 and H1299 cells, with a radiosensitization ratio of 1.45 and 1.38, respectively. Transwell chamber assay indicated that the ability of cell migration was significantly inhibited by Salvianolic acid B ( P<0.05). Western blot revealed that the expression levels of OTUD7B in A549 and H1299 cells were induced by irradiation in a time-dependent manner. Salvianolic acid B could down-regulate the expression levels of MMP-2, MMP-9 and p-Akt, whereas up-regulate the expression level of E-cadherin by down-regulating the expression level of OTUD7B. Conclusions:Salvianolic acid B can enhance the radiosensitivity of A549 and H1299 cells. The possible mechanism is that Salvianolic acid B down-regulates the expression level of OTUD7B induced by irradiation and inhibits the epithelial-mesenchymal transition process of tumor cells.

Sesquiterpenoids isolated from the flower of Inula japonica as potential antitumor leads for intervention of paclitaxel-resistant non-small-cell lung cancer.

Three new sesquiterpene lactone dimers (1-3) were isolated from the flowers of Inula japonica together with twenty-two known sesquiterpene derivatives (4-25). Their structures were established on the basis of detailed spectroscopic analyses. All isolates were evaluated for their antiproliferative activities against paclitaxel-resistant human non-small-cell lung cancer cell line A549/PTX. The preliminary structure-activity relationship was discussed. Compound 24 exhibited the most potent effect with the IC50 value of 0.34 ± 0.10 µM, even more active than the clinically used drug paclitaxel (PTX, IC50 = 1.40 ± 0.52 µM). Compound 24 showed significant efficacy of arresting the cell cycle at the G2-M stage, inducing apoptosis through mitochondria-mediated pathway, and inhibiting cell migration and invasion. Furthermore, compound 24 could reverse multidrug resistance through suppressing the expression of ABC family proteins.

Efficaciousness of Low Affinity Compared to High Affinity TSPO Ligands in the Inhibition of Hypoxic Mitochondrial Cellular Damage Induced by Cobalt Chloride in Human Lung H1299 Cells.

The 18 kDa translocator protein (TSPO) plays an important role in apoptotic cell death, including apoptosis induced by the hypoxia mimicking agent cobalt chloride (CoCl2). In this study, the protective effects of a high (CB86; Ki= 1.6 nM) and a low (CB204; Ki= 117.7 nM) affinity TSPO ligands were investigated in H1299 lung cancer cell line exposed to CoCl2. The lung cell line H1299 was chosen in the present study since they express TSPO and able to undergo programmed cell death. The examined cell death markers included: ATP synthase reversal, reactive oxygen species (ROS) generation, mitochondrial membrane potential (Δψm) depolarization, cellular toxicity, and cellular viability. Pretreatment of the cells with the low affinity ligand CB204 at a concentration of 100 µM suppressed significantly (p < 0.05 for all) CoCl2-induced cellular cytotoxicity (100%), ATP synthase reversal (67%), ROS generation (82%), Δψm depolarization (100%), reduction in cellular density (97%), and also increased cell viability (85%). Furthermore, the low affinity TSPO ligand CB204, was harmless when given by itself at 100 µM. In contrast, the high affinity ligand (CB86) was significantly effective only in the prevention of CoCl2-induced ROS generation (39%, p < 0.001), and showed significant cytotoxic effects when given alone at 100 µM, as reflected in alterations in ADP/ATP ratio, oxidative stress, mitochondrial membrane potential depolarization and cell death. It appears that similar to previous studies on brain-derived cells, the relatively low affinity for the TSPO target enhances the potency of TSPO ligands in the protection from hypoxic cell death. Moreover, the high affinity TSPO ligand CB86, but not the low affinity ligand CB204, was lethal to the lung cells at high concentration (100 µM). The low affinity TSPO ligand CB204 may be a candidate for the treatment of pulmonary diseases related to hypoxia, such as pulmonary ischemia and chronic obstructive pulmonary disease COPD.

Preparation and characterisation of tetrandrine nanosuspensions and in vitro estimate antitumour activity on A549 lung cancer cell line.

Aim: The aim of this study was to improve solubility and antitumour ability in vitro of tetrandrine (Tet) via preparing nanosuspensions (NSs).Methods: The Tet-NSs were prepared by wet media milling. The Tet-CCS-NS was prepared with croscarmellose sodium (CCS) as single stabiliser. The Tet-HACC-TPGS-NS was manufactured with D-α-tocopheryl polyethylene glycol 1,000 succinate (TPGS) and hydroponically trimethyl ammonium chloride chitosan (HACC) as combined stabilisers. Physicochemical properties of the NSs such as particle size, surface morphologies, crystallinity and molecular interactions were investigated. In addition, the in vitro dissolution and antitumour activities using A549 human lung cancer cells were evaluated.Results: The mean particle sizes and Zeta potential of freshly prepared Tet-CCS-NS, Tet-HACC-TPGS-NS were 469.1 ± 14nm and 157.3 ± 5nm, -29.4 ± 0.26 mV and 23.3 ± 0.36 mV, respectively. In comparison to pure Tet, the cumulative dissolution of Tet-NSs were increased by 4 ∼ 5 times in 2 h. In vitro antitumour studies on Tet- NSs in A549 cells, the cell survival rate of the Tet-NSs at high concentration (30-50µg/ml) were less than 10% within 48 h. Meanwhile, Tet-NSs were revealed to induce A549 cells apoptosis and promote cell uptake.Conclusion: The present study has proved that the Tet-NSs can increase Tet solubility as well as improve Tet antitumour activity in vitro.

Microwave-Assisted versus Conventional Isolation of Glucosinolate Degradation Products from Lunaria annua L. and Their Cytotoxic Activity.

Glucosinolates (GSLs) from Lunaria annua L. seeds were analyzed qualitatively and quantitatively by their desulfo counterparts using UHPLC-DAD-MS/MS technique and by their volatile breakdown products, isothiocyanates (ITCs), using GC-MS technique. GSL breakdown products were obtained by conventional techniques (hydrodistillation in a Clevenger type apparatus (HD), CH2Cl2 extraction after myrosinase hydrolysis (EXT) for 24 h) as well as by modern techniques, microwave-assisted distillation (MAD) and microwave hydrodiffusion and gravity (MHG). Seven GSLs were identified as follows: isopropyl GSL (1), sec-butyl GSL (2), 5-(methylsulfinyl)pentyl GSL (3), 6-(methylsulfinyl)hexyl GSL (4), 5-(methylsulfanyl)pentyl GSL (5), 6-(methylsulfanyl)hexyl GSL (6), and benzyl GSL (7). Additionally, pent-4-enyl- and hex-5-enyl ITCs were detected in the volatile extracts. However, their corresponding GSLs were not detected using UHPLC-DAD-MS/MS. Thus, they are suggested to be formed during GC-MS analysis via thermolysis of 5-(methylsulfinyl)pentyl- and 6-(methylsulfinyl)hexyl ITCs, respectively. Volatile isolates were tested for their cytotoxic activity using MTT assay. EXT and MHG showed the best cytotoxic activity against human lung cancer cell line A549 during an incubation time of 72 h (IC50 18.8, and 33.5 µg/mL, respectively), and against breast cancer cell line MDA-MB-231 after 48 h (IC50 6.0 and 11.8 µg/mL, respectively). These activities can be attributed to the ITCs originating from 3 and 4.

Effects of microRNAs on invasion, migration and apoptosis of lung cancer cell lines / 解剖学报

Objective To investigate the effects of microRNAs(miRNAs) on invasion, migration and apoptosis of lung cancer cell lines. Methods Target cells were infected with hsa-mir-933, hsa-mir-4700-3p, hsa-mir-3144-3p, hsamir-3972, and hsa-mir-548a-5p. Cell apoptosis was measured using flow cytomertry, Transwell invasion assay, cell migration were analyzed by lineation,miRNA was quantified using Real-time PCR. Results Compared with the vector and control, in hsa-mir-933, hsa-mir-4700-3p, hsa-mir-3144-3p, hsa-mir-397 and hsa-mir-548a-5p group the apoptosis significantly increased,cell invasion ability significantly decreased,cell migration ability significantly decreased, miRNAs expression significantly increased. Conclusion Five microRNAs can promote the apoptosis of lung cancer cell lines, change the high invasiveness of lung cancer cell lines, inhibit the migration of lung cancer cell lines, and increase the expression of corresponding miRNAs.

[The Effect of Methylation Level of microRNA Promoter on the Expression of microRNAs and on the Proliferation, Migration and Invasion of Lung Cancer Cells].

OBJECTIVE: To study the effect of methylation level of microRNA promoter on the expression of microRNAs (miRNA34a, miRNA34b, miRNA148a, miRNA203a) and on the proliferation, migration and invasion of lung cancer A549 cells. METHODS: The proliferation of A549 cells treated with different concentrations of demethylated drug 5-aza-2'-deoxycytidine (5-Aza-CdR) was measured by CCK8 assay and calculated the inhibitory rate in 24 h, 48 h and 72 h, respectively. After 72 h of treatment with 20 µmol/L 5-Aza-CdR, methylation-specific PCR (MSP) was used to detect the methylation level of A549 cells in miRNAs gene promoter regions, and real-time quantitative PCR (real-time PCR) was used to test the expression of miRNAs. The migration abilities of A549 cells treated with 20 µmol/L 5-Aza-CdR in 24 h and 48 h were performed with wound healing assay, while the invasion abilities in 48 h were evaluated by Transwell assay, respectively. RESULTS: The proliferation inhibition rate of A549 cells gradually increased with the treatment concentration of 5-Aza-CdR increased and the treatment time prolonged. Compared with the control group, the methylated band of the experimental group was weaker and the unmethylated band was stronger, and the miRNAs gene promoter regions methylation level of the experimental group was lower than that of the control group. The expression level of miRNAs was significantly increased in the experimental group (P<0.05) . The migration and invasion of the experimental group of A549 cells were inhibited compared with the control group (P<0.05) . CONCLUSION: 5-Aza-CdR can reverse methylation levels of miRNAs promoter regions and upregulate the expression level of miRNA34a, miRNA34b, miRNA148a, miRNA203a, resulting in significantly inhibiting the proliferation, migration and invasion of lung cancer cells.

Study on Synthesis and Anti-tumor Activity of 6 Kinds of Tectorigenin Mannich Base Derivatives / 中国药房

OBJECTIVE： To conduct structural modification of tectorigenin to search for new compounds with anti-tumor activity. METHODS： Tectorigenin was used as a lead compound， and then added into amine reagents as ethanolamine， methylamine， ethylamine， dimethylamine， diethylamine， n-propylamine and formaldehyde solution. Tectorigenin Mannich base derivatives were synthesized by mannich reaction with as the lead compound. The structures of the derivatives were identified according to IR， UV， MS and NMR data. Solubility of tectorigenin and its derivatives were investigated by solubility test method. MTT assay was used to investigate the inhibitory effects of tectorigenin and its derivatives on the proliferation of human colon cancer cell line HCT116， human lung cancer cell line A549 and human hepatoma cell line HepG2， and half inhibitory concentration （IC50） was calculated. The inhibition rate of tectorigenin and its derivatives （100 mg/kg） on H22 hepatoma-bearing mice in vivo was studied. RESULTS： Totally of 6 kinds of tectorigenin mannich base derivatives were synthesized， such as 8-（N-hydroxyethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-methyl）-methyleneamino-5，7，4′-trihydroxy-6- methoxyisoflavone， 8-（N， N-diethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N， N-dimethyl）-methyleneamino- 5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-ethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-propyl）- methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone （compounds 1-6 in turn）. Compared with tectorigenin， the water solubility of six derivatives was significantly improved， and the solubility was 5-20 times higher than that of tectorigenin. IC50 of compounds 1， 3 and 5 to HCT116 cells were （34.82±3.27）， （16.21±4.13）， （33.12±3.25） μmol/L， which were stronger than that of tectorigenin [（45.23±5.74） μmol/L]； IC50 of compounds 1， 3 and 5 to A549 cells were （37.05±5.74）， （26.88±4.52）， （30.13±6.23） μmol/L， which were stronger than that of tectorigenin [（53.24±6.34） μmol/L]； IC50 of compounds 1， 3 and 5 to HepG2 cells were （23.74±1.45）， （18.96±2.34）， （30.95±2.87） μmol/L， which were stronger than that of tectorigenin [（48.98±2.58） μmol/L]. Compounds 1， 3 and 5 showed higher inhibition rates （55.51％， 57.20％ and 49.15％） than tectorigenin （33.05％） on H22 hepatoma-bearing mice， respectively. The other three compounds had no obvious advantage over tectorigenin in anti-tumor activity. CONCLUSIONS： In this study， compounds 1， 3 and 5 of six tectorigenin mannich base derivatives synthesized in this study have stronger antitumor activity than tectorigenin.

Silencing LncRNA HOXD-AS1 enhances the radiosensitivity of H460 cells by regulating miR-454-3p expression / 中华放射肿瘤学杂志

Objective To explore the effect of LncRNA HOXD-AS1 on the radiosensitivity of H460 cells by targeting the regulation of miR-454-3p expression. Methods H460 cells were irradiated with 0, 2, 4, 6 and 8 Gy, and the expression levels of HOXD-AS1 and miR-454-3p were detected by qRT-PCR. Cell apoptosis rate were measured by flow cytometry experiments. Cell cloning experiments were used to detect cell radiosensitivity. The protein expression levels of Caspase-3, Cyclin D1 and γ-H2AX were detected by Western blot. Results The expression of HOXD-AS1 in H460 cells was significantly increased after X-ray irradiation ( P<0.05) , while the expression of miR-454-3p was significantly decreased ( P<0.05) . Silencing HOXD-AS1 significantly promoted apoptosis, increased radiosensitivity of lung cancer cells, promoted Caspase-3, γ-H2AX protein expression, and inhibited Cyclin D1 expression. HOXD-AS1 could target the expression of miR-454-3p. Conclusion Silencing HOXD-AS1 can promote the apoptosis of lung cancer cells and inhibit cell survival by targeting miR-454-3p to increase the radiosensitivity of lung cancer cells.

Cytotoxic consequences of Halloysite nanotube/iron oxide nanocomposite and iron oxide nanoparticles upon interaction with bacterial, non-cancerous and cancerous cells.

Cytotoxic effects of iron oxide (Fe3O4) nanoparticles and Halloysite nanotube/iron oxide (HNT/Fe3O4) nanocomposite are compared based on their interaction with Gram-negative bacteria Escherichia coli and Gram-positive bacteria Bacillus subtilis. Similarly, the action of these two nanomaterials on non-cancerous Vero cell lines and human lung cancerous (A-549) cell lines are compared. The cytotoxicity studies on Fe3O4 nanoparticles and HNT/Fe3O4 nanocomposite showed difference in the rate of killing of bacterial cells. This is reflected in differential cell growth, cell membrane integrity loss, lactate dehydrogenase (LDH) release and reactive oxygen species (ROS) production. These factors are measured over a range of concentrations of Fe3O4 nanoparticles and HNT/Fe3O4 nanocomposite and at specified time intervals, to test if there is any statistically significant difference between the toxicity of the two nanomaterials. Between the two nanomaterials, HNT/Fe3O4 nanocomposite is found to be less toxic to bacterial cells than Fe3O4 nanoparticles. HNT, when attached to the Fe3O4 nanoparticles, changes their surface characteristics and suppresses their inherent toxicity on bacteria. In the study on the effect on cell lines, Fe3O4 nanoparticles and HNT/Fe3O4 nanocomposite are both seen to be biocompatible with Vero cell lines. However, HNT/Fe3O4 nanocomposite showed more cytotoxicity than Fe3O4 nanoparticles on A-549 cell lines.

Synthesis and characterization of kaempferol-based ruthenium (II) complex: A facile approach for superior anticancer application.

In this study, we synthesized a novel metal flavonoid complex and investigated its effects on the non-small cell lung cancer cell lines, A549 and toxicity on the human dermal fibroblast cell lines, HDFa. 1H, 13C NMR, single crystal X-ray diffraction and elemental micro analysis (C,H,N,S/O) were used to characterize the synthesized kaempferol-based Ru (II) complex. Cell toxicity was studied using MTT assay and electric cell substrate impedance sensing (ECIS). It was evident from the MTT results that no significant cytotoxicity in HDFa cells occurs with the synthesized complex, but in case of A549 cells, significant cytotoxicity was observed even at low concentrations (10-20 µm). In addition, the effect of the newly synthesized complex on the A549 cell line was studied by investigating the cellular damage via atomic force microscopy and DNA fragmentation assay. The obtained results revealed that the synthesized complex was able to inhibit the cancer cells and have shown moderate anticancer activity against A549 cancer cell lines. In addition, it was evident that the complex was more active than kaempferol and well tolerated by normal cell lines.