[This corrects the article DOI: 10.3892/ol.2020.12321.].

Objective To investigate the mechanistic role of Sijunzi decoction (SJZD) in overcoming chemoresistance through the suppression of adaptive metabolic responses in non-small cell lung cancer (NSCLC). Methods Chemical profiling of SJZD-derived components in systemic circulation was conducted using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in Sprague-Dawley rats. Multiomics integration and network pharmacology were employed to identify convergent targets shared by the bioactive constituents of SJZD and genes associated with cisplatin resistance. In vitro functional assessments using cisplatin-resistant human lung adenocarcinoma (A549/DDP) cells included the following: quantification of cell viability via Cell Counting Kit-8 (CCK-8) assays; evaluation of mitochondrial bioenergetics through targeted metabolomic profiling; and ultrastructural characterization of ferroptotic morphology via transmission electron microscopy (TEM). Cellular redox homeostasis was dynamically monitored using fluorescent probes, including a DCFH-DA probe for reactive oxygen species (ROS) and a C11-BODIPY581/591 probe for lipid peroxidation. siRNA-mediated gene silencing and immunohistochemical analysis were performed to elucidate the functional hierarchy of the p62/Keap1/nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant axis. Complementary in vivo validation was performed using BALB/c nude mice bearing A549/DDP xenografts, with longitudinal monitoring of tumor progression under SJZD treatment regimens. Results Untargeted metabolomics of SJZD-medicated serum revealed 392 differentially abundant metabolites, with pathway enrichment revealing significant dysregulation of glutamine metabolism. Structural validation confirmed 55 bioactive components of SJZD in serum, including glycyrrhizin, ginsenoside Ro, liquiritigenin, and atractylenolide I. Integration of these components with disease targets yielded 355 overlapping genes associated with both SJZD activity and cisplatin-resistant NSCLC, with significant enrichment in oxidative stress response pathways. Experimental assays confirmed that SJZD induced ferroptosis in cisplatin-resistant A549/DDP cells, as evidenced by disrupted iron homeostasis, lipid peroxidation, and characteristic mitochondrial damage. These effects and subsequent cell death were specifically abrogated by the ferroptosis inhibitor ferrostatin-1 (Fer-1) but not by apoptosis inhibition, confirming that ferroptosis is the primary mechanism of cell death. Mechanistically, the inhibition of p62/Keap1/Nrf2 signaling was involved in the modulation of SJZD-induced ferroptosis both in vitro and in vivo. Conclusions SJZD counteracts metabolic adaptation through ferroptosis mediated by the inhibition of p62/Keap1/Nrf2 in cisplatin-resistant NSCLC.

This study aimed to investigate the molecular mechanism by which Buzhong Yiqi Decoction(BZYQD),a representative TCM formula for invigorating the spleen and replenishing Qi,regulates fatty acid metabolism,induces ferroptosis in cisplatin-resistant lung adenocarcinoma A549/DDP cells,and reverses cisplatin resistance in non-small cell lung cancer(NSCLC)via the AMP-activated protein kinase(AMPK)/acetyl-CoA carboxylase 1(ACC1)signaling pathway.The sensitivity of parental A549 cells and cisplatin-resistant A549/DDP cells to different concentrations of cisplatin(0-128 μmol·L-1)was compared,and the optimal cisplatin concentration for subsequent intervention was selected via the CCK-8 assay.The BZYQD-containing serum was prepared,and the effects of cisplatin,low-(L),medium-(M),and high-concentration(H)BZYQD,as well as the combinations of low-,medium-,and high-concentration BZYQD with cisplatin on the viability of A549/DDP cells were evaluated via the CCK-8 assay.RNA-seq was employed to analyze the effect of BZYQD-H+cisplatin treatment on the gene expression profile of A549/DDP cells.A549/DDP cells were randomly allocated into the control group,cisplatin group,and BZYQD-L+cisplatin,BZYQD-M+cisplatin,BZYQD-H+cisplatin groups.Western blot was performed to quantify the protein levels of AMPK,phosphorylated AMPK(p-AMPK),ACC1,fatty acid synthase(FASN),ATP-citrate lyase(ACLY),and carnitine palmitoyltransferase 1A(CPT1A).Immunofluorescence was used to observe lipid droplet synthesis in cells of each group under the intervention of BZYQD+cisplatin and AMPK inhibitor.The CCK-8 assay was adopted to examine the effect of the inhibitor on the viability of A549/DDP cells treated with BZYQD-H+cisplatin,and thus the underlying molecular mechanism was clarified.A ferrous ion assay kit was used to measure intracellular ferrous ion levels.The DCFH-DA fluorescent probe was employed to assess reactive oxygen species(ROS)levels.Western blot was conducted to determine the expression levels of ferroptosis-related proteins,including glutathione peroxidase 4(GPX4),acyl-CoA synthetase 4(ACSL4),and transferrin receptor 1(TFR1).The Liperfluo kit was used to assess intracellular lipid peroxidation levels,and the JC-1 probe was employed to quantify mitochondrial membrane potential.The results showed that cisplatin demonstrated the half-maximal inhibitory concentrations(IC50)of 12.2 μmol·L-1 and 41.2 μmol·L-1 against A549 cells and A549/DDP cells,respectively,with a resistance index(RI)of 3.38.Compared with the cisplatin group,BZYQD-L+cisplatin,BZYQD-M+cisplatin,and BZYQD-H+cisplatin significantly inhibited the viability of A549/DDP cells.RNA-seq results revealed that the differentially expressed genes in the BZYQD-H+cisplatin group were associated with signaling pathways such as fatty acid synthesis and catabolism,AMPK,and ferroptosis.Compared with the cisplatin group,the BZYQD-L+cisplatin,BZYQD-M+cisplatin,and BZYQD-H+cisplatin groups showed a significant upregulation of p-AMPK,a significant downregulation of fatty acid synthesis-related proteins(FASN,ACC1,and ACLY),and a significant upregulation of CPT1A.Immunofluorescence results indicated that BZYQD+cisplatin treatment inhibited lipid droplet formation by activating the AMPK/ACC1 pathway.Meanwhile,CCK-8 results showed that the AMPK inhibitor significantly reversed the decrease in viability of cells treated by BZYQD-H+cisplatin.Compared with the cisplatin group,the BZYQD-M+cisplatin and BZYQD-H+cisplatin groups exhibited significant increases in intracellular ferrous ion levels,along with significant elevations in intracellular ROS levels and lipid peroxidation levels and a significant reduction in mitochondrial membrane potential.The expression of the ferroptosis-related protein GPX4 was significantly downregulated,suggesting an imbalance in the cellular antioxidant system,while the expression levels of ACSL4 and TFR1 were significantly upregulated.These findings indicate that BZYQD+cisplatin can induce ferroptosis in A549/DDP cells and reverse cisplatin resistance.In summary,the results indicate that BZYQD synergizes with cisplatin to inhibit fatty acid synthesis and induce ferroptosis in A549/DDP cells through the AMPK/ACC1 pathway,thereby reversing cisplatin resistance in NSCLC.

Lung cancer is a leading cause of global cancer mortality. Clinical observations reveal that histological transformation from non-small cell lung cancer (NSCLC) to small cell lung cancer (SCLC) is accompanied by mutations in TP53 and RB1. By applying gradually increasing cisplatin concentrations to mimic the escalating drug pressure within the tumor microenvironment, this study investigated the link between phenotypic transformation to SCLC in cisplatin-resistant human lung adenocarcinoma cells and alterations in cellular energy production pathways. We established two cisplatin-resistant NSCLC cell lines with varying resistance levels. RNAseq analyses identified TP53 and RB1 gene mutations. Comprehensive functional assays were performed to characterize A549/DDP1 μg/mL and A549/DDP3 μg/mL cells, focusing on proliferation and migratory capabilities. Cellular bioenergetics were assessed through glycolysis and oxidative phosphorylation analyses. Western blotting was employed to examine epithelial-mesenchymal transition (EMT), glucose metabolism, and lipid metabolism markers. Cell cycle distribution was analyzed by flow cytometry. Additionally, a xenograft mouse model was developed for in vivo validation. TP53 and RB1 mutations were associated with cisplatin concentration-dependent phenotypic transformation, with A549/DDP cells acquiring a more aggressive SCLC-like phenotype (In the article we call the A549/DDPSCLC cells). Analysis of cell bioenergetics profiling and Western blot analyses revealed enhanced glucose metabolism in A549/DDP1 μg/mL cells, while A549/DDPSCLC cells exhibited predominant lipid metabolism. Compound3K and Etomoxir specifically inhibit the activity of PKM2 and CPT1A, respectively, with Etomoxir demonstrating substantially inhibited A549/DDPSCLC cells growth and more cell cycle arrest in the G0/G1 phase. Combinatorial of Compound3K and Etomoxir effectively induced cell death in A549/DDPSCLC phenotype cells in vitro. Etomoxir alone or combined with Compound3K significantly inhibited tumor growth in vivo, with enhanced efficacy in the combination group. This study provides the first evidence of cisplatin concentration-dependent metabolic reprogramming during NSCLC-to-SCLC transformation. We identified a phenotypic transition from NSCLC to SCLC accompanied by a metabolic shift from glucose to fatty acid metabolism, offering new insights into therapeutic strategies for treatmentresistant lung cancer.

Introduction: The effectiveness of therapy is strongly impacted by the resistance of lung cancer cells to cisplatin. The objective of this research is to characterize the impact of ailanthone on cisplatin resistance in non-small cell lung cancer (NSCLC) and examine any potential in vivo and in vitro molecular pathways. Methods: Following treatment of A549/DDP cells with ailanthone and cisplatin, cell survival and apoptosis were measured using flow cytometry and Cell Counting Kit-8 (CCK- 8) assays, respectively. mRFP-GFP-LC3 adenovirus transfection was used to track autophagy, and protein expression levels were analyzed by western blotting. Hematoxylin and eosin (H&E) staining was used after the organs of the mice were removed for in vivo investigations. In A549/DDP cells, ailanthone and cisplatin together induced autophagy and apoptosis in a dose and time-dependent manner (P< 0.05). After receiving combined treatment with ailanthone and cisplatin, the expression levels of cleaved caspase- 3, Bcl-2, cleaved PARP, Beclin1L, and C3B-II were considerably elevated by inhibiting the PI3K/AKT/mTOR signaling pathway. Result: Our findings show that ailanthone, without causing adverse effects in vivo, greatly suppressed the growth of A549/DDP-grafted tumors and improved the anti-tumor efficaciousness of cisplatin. Conclusion: According to this study, ailanthone may increase sensitivity to cisplatin and promote autophagy and death in NSCLC A549/DDP cells through the signaling pathway of PI3K/AKT/mTOR.

FOXO3 was decreased, while LA and FBXO22 were increased in NSCLC patients. LA promoted cisplatin resistance in NSCLC in vitro and in vivo. SQFZ inhibited LA-induced cisplatin resistance in NSCLC by regulating FOXO3. FBXO22 affected p53 ubiquitination to reverse the inhibitory effect of SQFZ. SQFZ inhibited cisplatin resistance in NSCLC by FOXO3/FBXO22/p53 axis. Non-small cell lung cancer (NSCLC) accounts for 80

OBJECTIVE:Utilizing transcriptome analysis to investigate the mechanisms and therapeutic approaches for cisplatin resistance in non-small cell lung cancer (NSCLC). METHODS:Firstly, the biological characters of A549 cells and A549/DDP cells were detected by RNA sequencing, CCK-8 and hippocampal energy analyzer. Then, the differential Genes were functionally enriched by GO and KEGG and the competitive endogenous RNA network map was constructed. Finally, the effects of the predicted biogenesis pathway on the biological functions of A549/DDP cells were verified by in vitro and in vivo experiments. RESULT:The differentially transcribed genes of A549 and A549/DDP cells were analyzed by enrichment analysis and cell biological characteristics detection. The results showed that A549/DDP cells showed significantly increased resistance to cisplatin, glucose metabolism signaling pathway and glycolysis levels compared with A549 cells. Among glycolysis-related transcription genes, PKM had the most significant difference Fold Change is 8. LncRNA PCIF1 is a new marker of A549/DDP cells and can be used as a molecular sponge to regulate the expression of PKM. LncRNA PCIF1 targets miR-326 to induce PKM expression, promote glycolysis level, and enhance the resistance of A549/DDP cells to cisplatin. CONCLUSION:LncRNA PCIF1 as biomarkers of A549/DDP cells, higher expression can induce the PKM, promote cell glycolysis, lead to the occurrence of cisplatin resistance. LncRNA PCIF1 can be considered as a potential target for treating cisplatin-resistant NSCLC.

Background: Cisplatin resistance is one of the major obstacles in non-small cell lung cancer (NSCLC) treatment. Intriguingly, elevated lactate levels were observed in cisplatin-resistant cells, which spurred further investigation into their underlying biological mechanisms. Methods: Lactate levels were measured by lactate detection kit. Cisplatin-resistance NSCLC cells were established using progressive concentration of cisplatin. Cell viability, proliferation, and apoptosis were detected by CCK-8, EdU, and flow cytometry, respectively. Cell proliferation in vivo was determined by immunohistochemistry of Ki67 and apoptotic cells were calculated by the TUNEL. MeRIP-PCR was used to measure FOXO3 m6A levels. The interactions of genes were analyzed via RIP, ChIP, Dual-luciferase reporter, and RNA pull-down, respectively. Results: Elevated lactate levels were observed in both NSCLC patients and cisplatin-resistance cells. Lactate treatment increased cisplatin-resistance cell viability in vitro and promoted tumor growth in vivo. Mechanistically, lactate downregulated FOXO3 by YTHDF2-mediated m6A modification. FOXO3 transcriptionally reduced MAGI1-IT1 expression. FOXO3 overexpression inhibited the lactate-induced promotion of cisplatin resistance in NSCLC, which were reversed by MAGI1-IT1 overexpression. MAGI1-IT1 and IL6R competitively bound miR-664b-3p. FOXO3 overexpression or MAGI1-IT1 knockdown repressed lactate-mediated cisplatin resistance in vivo. Conclusion: Lactate promoted NSCLC cisplatin resistance through regulating FOXO3/MAGI1-IT1/miR-664b-3p/IL6R axis in YTHDF2-mediated m6A modification.

BACKGROUND:Qingshen exhibits anti-inflammatory and immunoregulation effects to renal damage. Dendritic cells (DCs) play a critical role in regulating the pathologic inflammatory environment in renal fibrosis (RF). PURPOSE:To investigate the immune modulation mechanism of qingshen granule (QSG) in RF, particularly focusing on the role of DCs. METHODS/STUDY DESIGN:Adenine-induced RF animal models were used to study the pharmacological effects of QSG and the immune cells differentiation and function. Glucose uptake, non-esterified fatty acids secretion, mitochondrial membrane potential (MMP) detection, and qPCR were used to explore the effect of QSG to glucose and lipid metabolism in DCs and T cells. The effect of QSG to PI3K-AKT-mTOR axis and the modulation of mTOR to PD-L1 were explored by co-culture experiments, co-immunoprecipitation and western blot assays. The interaction of DCs/CD8+T cells and renal tubular epithelial cells (RTECs) was investigated to demonstrate the direct action and/or the immune-mediated regulation of QSG to RF. The components of QSG in the serum were determined by HPLC. And the effect of active ingredients and formula to DCs and T cells was analyzed by cell experiments in vitro. RESULTS:QSG reduced nephritic histopathological damage and suppressed the release of proinflammatory cytokines in adenine-induced RF mice. Of note, QSG decreased the levels of CD86, MHC-II, and CCR7 on DCs, while, increased PD-L1 expression on DCs in RF. The results demonstrated that QSG promoted the maturation and inhibited the migration of DCs, and QSG decreased the antigen presenting of DCs to T cells. Additionally, QSG reduced the MMP and glucose/lipid utilization ratio in DCs. QSG also down-regulated the level of targeted metabolic genes included glucose transporter 1 (Glut1), sterol-regulatory element-binding protein 1 (Srebp1), acetyl-CoA carboxylase alpha (Acaca), phosphomevalonate kinase (Pmvk), and up-regulated sirtuin2 (Sirt2) in DCs. In terms of mechanism, QSG inhibited the metabolism-related PI3K-AKT-mTOR pathway, followed by regulating the interaction of mTOR with PD-L1 to enhance the membrane stability of PD-L1. Besides, HPLC analysis identified five active ingredients in QSG. The specific anti-inflammatory and immunosuppressive actions of these ingredients were found to be weaker than QSG as a whole. Finally, inhibiting DC function by QSG disrupted the communication among DCs, T cells, and RTECs. This disruption was associated with low expression of α-smooth muscle actin (α-SMA) and collagen type I (Col-I) in the kidney. CONCLUSIONS:QSG inhibits DC metabolism and function via the PI3K-AKT-mTOR pathway to alleviate RF. The study highlights the importance of the specific composition of the formula in targeting DC-mediated immune regulation.

Objective Tunneling nanotubes are membrane-tubule-like structures that exist between cells and have a direct long-distance biological information exchange function. Because the structure of TNTs is easily destroyed, exists for a short period of time and is unstable after formation, it is difficult to observe its dynamic formation and function. This study used the high content analysis system (HCA) combined with laser scanning confocal microscopy (LSCM) to try to observe the dynamic process of TNTs formation and its function of vesicular material transport. Methods Human lung adenocarcinoma A549 and cisplatin-resistant A549/DDP cells were labeled using fluorescent probes. Subsequently, HCA was used to observe the formation process of TNTs, LSCM to observe the three-dimensional structure of TNTs, and HCA combined with LSCM to analyze the vesicular material transport function of TNTs, respectively. Results TNTs structures can be formed between tumor cells of the same types (A549 or A549/DDP) or between different subtypes of tumor cells (A549 and A549/DDP). TNTs formed between A549/DDP cells were longer, thicker and had a higher formation index compared to A549 cells (the length, diameter and formation index of TNTs for A549 and A549/DDP were 14.71 mu m, 2.27 mu m, 4 and 25.44 mu m, 2.59 mu m, 11, respectively). TNTs are formed in two steps: first, two cell cytoplasmic membranes first come into contact with each other and then fuse, and as the two cells translocate in opposite directions, the fused regions of the membranes are continuously elongated and narrowed, resulting in the formation of TNTs; second, two cells extend filopodia-like membrane projections and fuse when they come into contact with each other's membrane projections, resulting in the formation of TNTs or one cell extends filopodia-like membrane projections and fuses after contacting the other cell membrane, thus forming TNTs. The transport of vesicles by TNTs is bidirectional. During transport, the rate and amounts of vesicles being transported vary depending on the stage of transport and the donor cell. The vesicular translocation from A549/DDP cells to A549 cells showed a fast initial rate and a slow terminal rate. A549 cells, as donor cells, transit vesicles to recipient A549/DDP cells in a higher number and proportion than A549/DDP as donor cells in the opposite direction. Conclusion The dynamic formation process and vesicular substance transport function of TNTs could be observed and analyzed by HCA combined with LSCM effectively.

Glutamine, like glucose, is a major nutrient consumed by cancer cells, yet these cells undergo glutamine starvation in the cores of tumors, forcing them to evolve adaptive metabolic responses. Pharmacologically targeting glutamine metabolism or withdrawal has been exploited for therapeutic purposes, but does not always induce cancer cell death. The mechanism by which cancer cells adapt to resist glutamine starvation in cisplatin-resistant non-small-cell lung cancer (NSCLC) also remains uncertain. Here, we report the potential metabolic vulnerabilities of A549/DDP (drug-resistant human lung adenocarcinoma cell lines) cells, which were more easily killed by the iron chelator deferoxamine (DFO) during glutamine deprivation than their parental cisplatin-sensitive A549 cells. We demonstrate that phenotype resistance to cisplatin is accompanied by adaptive responses during glutamine deprivation partly via higher levels of autophagic activity and apoptosis resistance characteristics. Moreover, this adaptation could be explained by sustained glucose instead of glutamine-dominant complex II-dependent oxidative phosphorylation (OXPHOS). Further investigation revealed that cisplatin-resistant cells sustain OXPHOS partly via iron metabolism reprogramming during glutamine deprivation. This reprogramming might be responsible for mitochondrial iron-sulfur [Fe-S] cluster biogenesis, which has become an “Achilles’ heel,” rendering cancer cells vulnerable to DFO-induced autophagic cell death and apoptosis through c-Jun N-terminal kinase (JNK) signaling. Finally, in vivo studies using xenograft mouse models also confirmed the growth-slowing effect of DFO. In summary, we have elucidated the adaptive responses of cisplatin-resistant NSCLC cells, which balanced stability and plasticity to overcome metabolic reprogramming and permitted them to survive under stress induced by chemotherapy or glutamine starvation. In addition, for the first time, we show that suppressing the growth of cisplatin-resistant NSCLC cells via iron chelator-induced autophagic cell death and apoptosis was possible with DFO treatment. These findings provide a solid basis for targeting mitochondria iron metabolism in cisplatin-resistant NSCLC for therapeutic purposes, and it is plausible to consider that DFO facilitates in the improvement of treatment responses in cisplatin-resistant NSCLC patients.

Aim: Chemoresistance is the biggest obstacle in cancer treatment. Our previous study demonstrated that Shenmai injection (SMI), a Chinese herbal medicine, enhanced the antitumor effect of cisplatin via glucose metabolism reprogramming. This study aimed to further determine whether the SMI sensitizes the non-small cell lung cancer (NSCLC) cells to cisplatin through regulation mitochondrial dynamics. Methods: The Kaplan-Meier Plotter database was used to investigate the relationship between mRNA expression of mitofusin-2 (Mfn2) and the survival analysis of NSCLC patients. The protein expression of Mfn2 in a lung adenocarcinoma tissue chip was detected by immunohistochemistry staining. The expression of Mfn2 and ATAD3A were compared between cisplatin-sensitive A549 and cisplatin-resistant A549/DDP cells. Additionally, A549/DDP cells were co-treated with cisplatin and SMI to detect mitochondrial morphology by fluorescent staining, apoptosis-related protein expression with Western blotting, and mitochondrial membrane potential (ΔΨm) with flow cytometry analysis. Results: The mean survival time of the Mfn2low group was significantly lower than that of the Mfn2high group by Kaplan-Meier Plotter database analysis, and the Mfn2 protein expression level was lower in cancer tissues than in adjacent tissues. The combination of SMI and cisplatin induced dynamic changes in A549/DDP cells, with increased mitochondrial fusion followed by upregulation of Mfn2 and downregulation of ATAD3A and reduced mitochondrial mass and ΔΨm. Moreover, SMI significantly enhanced cisplatin-induced A549/DDP apoptosis, upregulated Bax and the active subunit of caspase-3, and downregulated Bcl-2 expression, as shown via Hoechst staining and flow cytometry analysis. Conclusion: Our findings suggest SMI enhances cisplatin-induced apoptosis through regulation of Mfn2-dependent mitochondrial dynamics in cisplatin-resistant lung adenocarcinoma cells.

Tumor cells, especially drug-resistant cells, predominately support growth by glycolysis even under the condition of adequate oxygen, which is known as the Warburg effect. Glucose metabolism reprogramming is one of the main factors causing tumor resistance. Previous studies on Shenmai injection (SMI), a Chinese herbal medicine, have shown enhanced efficacy in the treatment of tumors in combination with chemotherapy drugs, but the mechanism is not clear. In this study, we investigated the effect of SMI combined with cisplatin on cisplatin-resistant lung adenocarcinoma A549/DDP cells. Our results showed that cisplatin-resistant A549/DDP cells exhibited increased glucose consumption, lactate production, and expression levels of key glycolytic enzymes, including hexokinase 2 (HK2), pyruvate kinase M1/2 (PKM1/2), pyruvate kinase M2 (PKM2), glucose transporter 1 (GLUT1), and lactate dehydrogenase A (LDHA), compared with cisplatin-sensitive A549 cells. SMI combined with cisplatin in A549/DDP cells, led to significantly lower expression levels of key glycolytic enzymes, such as HK2, PKM1/2, GLUT1, and pyruvate dehydrogenase (PDH). In addition, we found that the combination of SMI and cisplatin could inhibit cell proliferation and promote apoptosis by reducing the expression levels of p-Akt, p-mTOR, and c-Myc, and then, it reduced the glycolysis level. These results suggest that SMI enhances the antitumor effect of cisplatin via glucose metabolism reprogramming. Therefore, the combination of SMI and cisplatin may be a potential therapeutic strategy to treat cisplatin-resistant nonsmall cell lung cancer.

Skin cancer is the deadliest type of malignant disease and causes primary mortality worldwide. Dioscin, which exists in medicinal plants, has potent anticancer effects. However, its effects on skin cancer remain unknown. In the present study, the activity and mechanism of dioscin on the human skin cancer A431 cell line were investigated, MTT, colony formation, Transwell, wound-healing, TUNEL, Comet, immunofluorescence and western blot assays were used to assess the effects of dioscin on A431 cells. The results of MTT, colony formation, Transwell and wound-healing assays revealed that dioscin suppressed proliferation, colony formation and invasion of the cancer cells. TUNEL and comet assays demonstrated that dioscin exhibited significant effects on cell apoptosis and DNA damage. Investigations into the mechanism revealed that the expression levels of phosphorylated Ataxia telangiectasia-mutated (ATM) were considerably activated by dioscin, which significantly upregulated the expression levels of p53 to activate mitochondrial apoptosis signaling. Furthermore, the expression levels of BAX, cleaved caspase-3/9 and cleaved poly (ADP-ribose) polymerase were upregulated, and the expression levels of BCL-2 were downregulated by dioscin. Additionally, dioscin markedly downregulated the expression levels of matrix metalloproteinase 2 (MMP2), MMP9, RHO and cdc42, which are all associated with tumor invasion. In addition, p53-small interfering RNA transfection experiments indicated that dioscin exhibited excellent activity against skin cancer in vitro by decreasing p53 expression. Overall, the present results suggested that dioscin inhibited skin cancer cell proliferation via adjusting ATM/p53-mediated cell apoptosis, migration and DNA damage, which should be considered as a potential option for future treatments of skin cancer.

Cisplatin is one of the most active cytotoxic agents for non-small cell lung cancer (NSCLC) treatment. However, the development of cisplatin resistance is common. Bu-Zhong-Yi-Qi decoction (BZYQD), a Chinese traditional herbal medicine, is widely used for the enhancement of antitumor effect in other medications. In this study, we evaluated the effect and drug-resistance reversal mechanism of BZYQD combined with cisplatin on cisplatin-resistant A549/DDP cells. Our results showed that BZYQD exhibited direct cytotoxic and chemosensitizing effects. Cotreatment with BZYQD and cisplatin induced intrinsic apoptotic pathways which were measured by condensed nuclear chromatin, Annexin V/PI apoptosis assay, and apoptosis related proteins expression. In addition, cotreatment with BZYQD and cisplatin also activated autophagy, as indicated by an increase in LC3 puncta, classical autophagosomes and/or autolysosomes, and an accumulation of LC3-II and ATG7 protein. Finally, cotreatment with BZYQD and cisplatin resulted in the generation of ROS and scavenging ROS by NAC almost completely suppressing cell death. These results suggest that cotreatment with BZYQD and cisplatin might reverse cisplatin resistance by inducing ROS accumulation, which activates apoptosis and autophagy by oxidative stress. The combination of BZYQD and cisplatin may represent a novel approach in treatment for NSCLC and thus offer a new target for chemotherapy.

Quinocetone (QCT), an antimicrobial growth promoter, is widely used in food-producing animals. However, information about pharmacokinetics (PK) of QCT in ducks still remains unavailable up to now. In this study, QCT and its major metabolites (1-desoxyquinocetone, di-desoxyquinocetone and 3-methyl-quinoxaline-2-carboxylic) in ducks were studied using a simple and sensitive UHPLC-MS/MS assay. Twenty ducks were divided into two groups. (n = 10/group). One group received QCT by oral administration at dose of 40 mg/kg while another group received QCT intravenously at 10 mg/kg. Plasma samples were collected at various time points from 0 to 96 hr. QCT and its major metabolites in duck plasma samples were extracted by 1 ml acetonitrile and detected by UHPLC-MS/MS, with the gradient mobile phase that consisted of 0.1% formic acid in water (A) and acetonitrile (B). A noncompartment analysis was used to calculate the PK parameters. The results showed that following oral dosing, the peak plasma concentration (Cmax ) of QCT was 32.14 ng/ml and the area under the curve (AUCINF_obs) was 233.63 (h ng)/ ml. Following intravenous dosing, the Cmax , AUCINF_obs and Vss_obs were 96.70 ng/ml, 152.34 (h ng)/ ml and 807.00 L/kg, respectively. These data indicated that the QCT was less absorbed in vivo following oral administration, with low bioavailability (38.43%). QCT and its major metabolites such as 1-desoxyquinocetone and 3-methyl-quinoxaline-2-carboxylic were detected at individual time points in individual ducks, while the di-desoxyquinocetone was not detected in all time points in all ducks. This study enriches basic scientific data about pharmacokinetics of QCT in ducks after oral and intravenous administration and will be beneficial for clinical application in ducks.

Intestinal mucositis is a serious toxic side effect of 5-fluorouracil (5-FU) treatment. Bu-Zhong-Yi-Qi decoction (BZYQD), a water extract of Chinese traditional herbal medicine, is widely used in chemotherapy in Asia as an alternative treatment to reduce the side effects of chemotherapy. However, the mechanism is unknown. To evaluate its mechanism, we investigated the effect of BZYQD on 5-FU-induced intestinal mucositis in mice, especially with regard to apoptosis in the intestinal mucosal epithelia. In the present study, mice were divided into three groups: control, 5-FU, and 5-FU + BZYQD. Mice in the 5-FU and 5-FU + BZYQD groups were administered 5-FU (100 mg/kg/day, intraperitoneally) for 6 days, and the mice in the latter group were given BZYQD (8 g/kg/day, intragastrically) beginning 4 days before 5-FU and continuing until the termination of the experiment. Loss in body weight and diarrhea during the 5-FU treatment were significantly attenuated by administration of BZYQD. The morphological signs of intestinal damage, including shortened villi height, crypt destruction, apoptosis, and necrosis, in intestinal mucosal epithelia were also reversed, accompanied by reduced neutrophil infiltration, nitrite levels, and inflammatory factors (tumor necrosis factor α and interleukin 1β) and increased levels of reduced glutathione. These results suggest that BZYQD inhibits 5-FU-induced intestinal mucositis, and this effect may be due to the reduction in apoptosis and necrosis in intestinal mucosal epithelia via the suppression of inflammatory cytokine upregulation. In conclusion, inhibiting cytokine-mediated apoptosis or necrosis can be the molecular mechanism by which BZYQD reduces the gastrointestinal side effects of cancer chemotherapy.